System for monitoring sea water quality
using toxicology data on cetaceans
Act. 4.4 MSP Pilot Project - Abruzzo Region Final Report
15 January 2014
External Expert:
Workgroup: Giovanna Lanciani, Nicola Celli, Luana Dragani, Barbara Mariani, Carmen Verri,
Andria D’Orazio, Lorena Salvatore, Cristina Ingarao, Tommaso Pagliani
Index
1
Summary ................................................................................................................ 3
2
Project objectives ................................................................................................... 6
3
Area of interest ....................................................................................................... 8
4
Project activities...................................................................................................... 9
4.1
5
Materials and methods ......................................................................................... 11
4.1.1
Phase I .................................................................................................. 11
4.1.2
Phases II and III .................................................................................... 13
Pilot project’s results............................................................................................. 20
5.1
Collection of literature on eco-toxicological studies ............................................. 20
5.1.1
Heavy metals ........................................................................................ 22
5.1.2
Organic contaminants ........................................................................... 25
5.2
The Adriatic Stranding Events (ASE) Database................................................... 29
5.3
The Preserved Organs and Tissues of Adriatic Cetaceans database
(POTAC) ............................................................................................................... 32
5.4
Stranding events data elaboration........................................................................ 33
5.5
Heavy metals data elaboration............................................................................. 40
5.6
Organic contaminants data elaboration................................................................ 52
5.7
Biomarkers data elaboration ................................................................................ 57
5.8
6
5.7.1
8-OHdG ................................................................................................. 57
5.7.2
Micronuclei assay.................................................................................. 60
The listening station at sea................................................................................... 63
Implication for MSP and ICZM ............................................................................. 64
6.1
Implication for MPS .............................................................................................. 64
6.2
Implication for ICZM ............................................................................................. 64
6.3
Implication for MSFD ............................................................................................ 65
6.4
Adriatic added valued of the project ..................................................................... 65
6.5
Innovative aspects of the projects ........................................................................ 66
7
Conclusions .......................................................................................................... 67
8
References ........................................................................................................... 70
Appendix – Complete results of inorganic and organic compounds analysis
Annex – Operating project oa a listening station at sea for Odontocetes populations and
acoustic marine pollution monitoring
pag. 2/73
1
Summary
The principal project purposes were three:
1.
Settlement reconstruction of the eco-toxicological trend on the Adriatic Cetaceans
populations;
2.
Evaluation of different environmental contaminants levels in tissues of cetaceans;
3.
Evaluation of the eco-toxicological stress linked to the different contaminants.
The activities were divided into 5 phases:
I.
Reconstruction of historical evolution and trends on eco-toxicological researches of
Adriatic cetaceans
It was carried out: an analysis of the scientific literature regarding cetaceans’ ecology, stranding and eco-toxicological studies in Mediterranean and Adriatic seas; a
geo-referenced archive (Geographical Information System) on stranding events recorded in Adriatic Sea during a period ranging from 1902 to November 2013 and on
biological materials collected from stranded animals. The possible presence of temporal trends, regional variation, differences in species, sex and body size of stranding animals were evaluated.
II.
Determination of different environmental contaminants levels in tissues of Adriatic
Cetaceans
Frozen tissues sampled from cetaceans stranded on the Adriatic sea coastline were
analysed to determine the concentration of 3 contaminants groups: heavy metals, by
Inductively Coupled Plasma Mass Spectrometry (ICP-MS); Organochlorines (OCs:
DDTs and HCB) and Polycyclic Aromatic Hydrocarbons (PAHs) by gas chromatography coupled to mass spectrometry (GC-MS).
III.
Evaluation of the eco-toxicological stress in Adriatic cetaceans due to different environmental contaminants
The cumulative effects of contaminants on the cetaceans were estimated by means
of diagnostic and prognostic instruments, such as biomarkers, that allow evaluating
the exposition and the effects of xenobiotic substances on the organism. Damages
induced to DNA are evaluated by: measuring the formation of one of the major products of DNA oxidation, 8-hydroxy-2’-deoxyguanosine (8-OhdG), by ELISA methodology; micronuclei counting, to highlight chromosomal mutations induced by mutagenic substances.
IV.
Construction of theoretical statistical models for Potential Hazard assessment
It was based on the start hypothesis that contaminants levels in stranding specimens
are significant higher than in free-ranging ones. The application of multivariate statis-
pag. 3/73
tical methods as the canonical discriminant analysis (ADC) would have to allow defining the Potential Hazard of the bottlenose dolphin for certain categories of pollutants. By the analysis of the limited studies existing in literature regarding the Potential Hazard (for a total of no. 4) it was not possible to develop this phase, due to the
inability to obtain tissue samples from live animals (by biopsy).
V.
Achievement of a “listening station” at sea
The executive project of a listening station at sea and a monitoring system of the anthropic noise impact on cetaceans’ population in the Adriatic Sea. It consists of a
permanent multi-hydrophonical omni-directional station installed at the Posidonia
tower, offshore of Francavilla (CH), equipped with specific acoustic instrumentation
connected via GSM to an ashore workstation.
The results can be summarized as follows.
Phase I
Geographical distribution of stranding is homogeneous along the Adriatic coast. Stranding appears quite regularly (with polynomial distribution) over the years, except for 1991, particularly
in the Central Adriatic, in which there is a strong increment probably caused by the measles
outbreak that attacked marine mammals populations, particularly dolphins, in the period 19901991.
The time period analysed (1982-2013) is too short to identify the existence of a cyclical nature
of the phenomenon, as shown instead in other contexts (Evans et al., 2005).
It is noticed an increase in the number of stranding during the summer months, which could be
partly due to the number of sightings during the bathing season; however, there is a peak that
moves along a north-south gradient: in the northern regions in July, in the central regions between July and August and in the southern regions in August-September. There is also a difference in the frequency peak of stranding among species during the summer period: in June
for Risso's dolphin (Grampus griseus); in July for the bottlenose dolphin (Tursiops truncatus)
and between August and September for the striped dolphin (Stenella coeruleoalba).
Statistical analysis didn’t highlight both the existence of a different probability to strand between males and females of the total species and between the species and body dimensions
of stranding specimens by species.
Phase II
Our results confirm the high concentrations of trace metals (mainly Hg, Cd, Zn, Fe and Se) in
various tissues. There is a relation between the tissue and the different concentrations of the
various compounds: the liver appears to be the preferential organ for mercury accumulation;
the kidney and the muscle are the next organs in terms of mercury uptake followed to skin
blubber and the melon; Cd tends to accumulate preferentially in the kidney, with lower concentrations found in the other tissues.
pag. 4/73
However there are no significant statistical differences on the concentration of metals among:
different species, different tissue and different tissue within the same species (except for G.
griseus and P. macrocephalus in kidney metal concentration). The values of metal concentrations do not differ significantly with those already published by scientific literature about Adriatic Sea.
As for the organic contaminants, most of Polycyclic Aromatic Hydrocarbons resulted below the
corresponding limits of quantitation in all the samples. Organochlorine compounds, as expected showed the highest concentration in skin-blubber and melon tissues of all the analysed
specimens, confirming their high affinity for lipophylic matrices. The distribution analysis of the
various DDT compounds shows 4,4’-DDE as the major constituent of the group, accounting for
at least 80% of the components sum.
Phase III
The results of biomarkers (OHdG and micronuclei) analysis indicate a consistent exposition to
genotoxic substances, as established by literature studies on different aquatic species. Considering the values of 8-OHdG found in literature studies, we can assume that the results we
obtained indicate a high level of oxidative stress. T. truncatus is the species characterized by
the presence of a higher frequency of adducts and micronuclei than S. coeruleoalba, in line
with the analytical data on heavy metals.
It was confirmed a positive correlation between the percentage of oxidation products with four
transition heavy metals. There is also a correlation between the 8-OHdG and Pb known for its
cytotoxic properties in soft tissues and Hg, responsible for inhibition of antioxidant molecules
that contain sulphur, resulting in increased oxidation. Among the organic substances only the
chlorinated fungicide hexachlorobenzene is positively correlated with the biomarker. The statistical analysis showed a correlation between the percentage of 8-OHdG and some metals,
such as Fe, Mo, Pb, Hg, and an organic compound, hexachlorobenzene, providing valuable
information for the marine eco-toxicology. The liver appears the most liable to both biological
damages (MN and adducts) according to toxicological literature data.
pag. 5/73
2
Project objectives
The aim of the project was the evaluation of pollution effects in the Adriatic ecosystem by
means of a biological monitoring of cetaceans, predators at the top of the food chain, defined
“guard species” of the marine environment health status.
Various studies on Mediterranean cetaceans have revealed bioaccumulation of different categories of contaminants. The susceptibility of these animals to inorganic and organic pollutants
and the relationship between bioaccumulation and some population decline (as in the case of
Delphinus delphis) are unexplored fields. Using stranded specimens found in a good state of
conservation and/or sampling free-ranging dolphins by non-destructive methods, it was possible to carry out several studies on the health status of Mediterranean Marine Mammals from
the toxicological point of view. By measuring levels of contaminants in organs and tissues of
cetaceans, it was possible to establish to what extent these substances are accumulated by
these mammals and their enzymatic response to contamination.
Nevertheless, eco-toxicological studies on cetaceans focused mainly on the Western side of
the Mediterranean basin. For the Adriatic side, something more has been done with regard to
heavy metals compared to Persistent Organic Pollutants (POPs) bioaccumulation. The environmental contaminants, object of the study were: heavy metals and two groups of organic
pollutants: Organochlorine compounds (OCs: DDTs and HCB) and Polycyclic Aromatic Hydrocarbon (PAHs).
Some geographic areas are potentially more threatened than others by these contaminants:
one of these is the Mediterranean Sea. In this peculiar environment, top predators, such as
large pelagic fish and marine mammals, tend to accumulate large quantities of OCs, PAHs
and toxic metals (Corsolini et al., 1995; Marsili, 2000).
A high concentration of heavy metals (mainly mercury, methyl-mercury, cadmium, zinc and
selenium) has been found in various tissues of stranded specimens in Mediterranean basin.
The high concentration is the main result of the trophic level of those species in the food chain,
the type of diet and the age of the specimen. No doubt that the main metal contaminant for the
Mediterranean Sea is mercury (Hg). The source of mercury is believed to be natural deposits
in the Mediterranean basin. Mercury levels in dolphins from the Mediterranean are generally
higher than those found in the same species from the Atlantic (André et al., 1991). This has
been explained by the existence of natural mercury sources in the Mediterranean Sea and
mercury rich inputs via rivers flowing through the Mt. Amiata area (one of the richest natural
reserves of cinnabar). In fact, very high mercury concentrations have been found in fish tissues from the Tyrrhenian Sea (NW Mediterranean; Bernhard, 1988).
Further, this basin has limited exchange of water with the Atlantic Ocean, and is surrounded
by some of the most heavily populated and industrialized countries in the world. Levels of
some xenobiotics are much higher here than in other seas and oceans (Aguilar et al., 2002).
pag. 6/73
The toxicological effect of these contaminants is a few studied fields that could drastically interfere with the stability of populations of Mediterranean top predators.
It is possible to evaluate the cumulative effects of contaminants on the cetaceans by means of
biomarkers, that allow evaluating the exposition and the effects of xenobiotic substances on
the organism.
Genotoxic effects interfere with the mechanisms of spontaneous genetic recombination, increasing the frequency of DNA mutations, with possible serious consequences, in case the
mutations affect genes involved in the growth regulation, in the cell cycle or in carcinogenesis
processes.
Therefore, the principal project purposes were three:
1.
Settlement reconstruction of the eco-toxicological trend on the Adriatic Cetaceans
populations;
2.
Evaluation of different environmental contaminants levels in tissues of cetaceans;
3.
Evaluation of the eco-toxicological stress linked to the different contaminants.
pag. 7/73
3
Area of interest
The entire basin of the Adriatic Sea with particular reference to Italian coast.
Figure 3-1 The interest area: the entire Adriatic Sea.
pag. 8/73
4
Project activities
The activities were divided into 5 phases:
I - Reconstruction of historical evolution and trends on eco-toxicological researches of
Adriatic cetaceans
The first phase of the service included:
•
Analysis of the scientific literature regarding cetaceans’ ecology, stranding and ecotoxicological studies in Mediterranean and Adriatic seas.
•
Creating and managing of a geo-referenced archive (Geographical Information System) on stranding events recorded in Adriatic Sea during a period ranging from 1902
to November 2013 and biological materials collected from stranded animals.
•
Stranding events data processing aimed at highlighting the possible existence of:
o
Temporal trends;
o
Regional variation and/or correlation;
o
Differences in species, sex and body size of stranding animals.
II - Determination of different environmental contaminants levels in tissues of Adriatic
Cetaceans
This action included: analysis of frozen tissues sampled from cetaceans stranded on the Adriatic sea coastline to estimate the concentration of 3 contaminants groups: heavy metals, by
Inductively Coupled Plasma Mass Spectrometry (ICP-MS); Organochlorine compounds (OCs:
DDTs and HCB) and Polycyclic Aromatic Hydrocarbon (PAHs) by gas chromatography coupled to mass spectrometry (GC-MS). Results are then used for standard statistical analysis to
obtain a variables description, correlations and differences among them. The following steps
were carried out:
•
Analysis of the scientific literature regarding the relationships between heavy metals
and/or organic contaminants with bio-ecological parameters and analytical methodologies for extracting samples and measuring the heavy metals and organic contaminants concentration, their extraction and determination in tissue samples.
•
Selection of the tissue samples available from the Tissue bank (BTMM).
•
Analysis of no. 33 elements (heavy metals) in the selected tissues.
•
Analysis of no. 7 Organochlorine Compounds (OCs) and no. 19 Polycyclic Aromatic
Hydrocarbons (PAHs) in the selected tissues.
•
Data elaboration of elements.
•
Data elaboration of OCs and PAHs.
pag. 9/73
III - Evaluation of the eco-toxicological stress in Adriatic cetaceans due to different environmental contaminants
The cumulative effects of contaminants on the cetaceans was estimated by means of diagnostic and prognostic instruments, such as biomarkers, that allow to evaluate the exposition and
the effects of xenobiotic substances on the organism.
In particular, damages induced to DNA are evaluated:
•
By measuring the formation in different tissues of one of the major products of DNA
oxidation, 8-hydroxy-2’-deoxyguanosine (8-OhdG), by ELISA methodology.
•
By micronuclei observation, that allows to highlight chromosomal mutations (broken or
badly distribution of chromosomes) induced by mutagenic substances, resulting in expulsion from the nucleus fragments or whole chromosomes, which are called micronuclei.
We carried out the following points:
•
Analysis of the scientific literature on biomarkers used for the evaluation of ecotoxicological stress on aquatic organisms, with particular reference to marine mammals (for a total of no. 10 studies);
•
Analysis of the DNA oxidation product: 8-hydroxy-2'-deoxyguanosine (dG-8OH).
•
Micronuclei assay on different cetaceans tissues.
•
Application of statistical tests to the results to correlate the observed DNA damage
degree to the contaminants concentrations in tissue samples.
IV - Construction of theoretical statistical models for Potential Hazard assessment
It was based on the start hypothesis that contaminants levels in stranding specimens are significant higher than in free-ranging ones. The application of multivariate statistical methods as
the canonical discriminant analysis (ADC) would have to allow defining the Potential Hazard of
the bottlenose dolphin for certain categories of pollutants. By the analysis of the limited studies
existing in literature regarding the Potential Hazard (for a total of no. 4) it was not possible to
develop this phase, due to the inability to obtain tissue samples from live animals (by biopsy).
V - Achievement of a “listening station” at sea
The executive project of a listening station at sea and a monitoring system of the anthropic
noise impact on cetaceans’ population in the Adriatic Sea is included as Annex to this report.
The project concerns a permanent multi-hydrophonical omni-directional station installed at the
Posidonia tower, offshore of Francavilla (CH), equipped with specific acoustic instrumentation
connected via GSM to an ashore workstation. Fulfilment of a digital sound archive, geared by
suitable numeric conversion devices of analogical signals coming from the sea station (Digital
Signal Processing Workstation), will allow the acoustic data processing by means of specific
library and software (discrimination of spectral components in the frequency dominion and of
dynamical ones in the time dominion) and the identification of transient species.
pag. 10/73
4.1
Materials and methods
®
®
All the statistical analyses were done by IBM SPSS Software vers. 22.
4.1.1
Phase I
The step was carried out by:
a)
Collection of literature studies and report about: the biology and ecology of Mediterranean cetaceans, stranding events along the Italian coasts, collection, studies and
biopsies of tissue /organs of stranded animals, eco-toxicological studies on heavy
metals, organic contaminants and biomarkers in Adriatic and Mediterranean cetaceans populations.
b)
Collection of historical data of stranding events on Adriatic coasts and of biological
materials (organs, tissues) from Adriatic specimens, obtained by existing data banks.
Data from point b) were organized in 3 groups, based on their localization, according to the
geographical sectors subdivision of the Mediterranean-Adriatic Sea, made by the Italian Marine Biology Society (SBM; Figure 4-1):
•
Northern Adriatic (sector 9) – from the border Italy-Slovenia to Riccione (RI, Italy);
•
Central Adriatic (sector 8) – from Riccione to “Testa del Gargano” (FG);
•
Southern Adriatic (sector 7): from Testa del Gargano (FG) to Capo Santa Maria di
Leuca (LE).
Figure 4-1 Partition areas of the Adriatic basin (according to SBM).
pag. 11/73
These data were then organized in two different databases regarding the stranding events and
the available preserved organs and tissues, respectively.
The Adriatic Stranding Events (ASE) Database
ASE collects two kinds of data regarding cetaceans stranding events, deriving from two database sources:
•
The Italian database of the "Coordination Centre for the data collection on stranded
animals" (CISBRA, University of Pavia). Thanks to the access grant of CISBRA, it has
been possible to obtain all the stranding events occurred along the Italian coasts of
Adriatic Sea from 1902 to 2012 registered by the national database. It is realized by a
monitoring network of several institutions and public bodies like Harbour offices,
Health Local Services and non-governmental institutions thanks to government funding. Data are constantly updated by the new events notifications arriving to us via email from Pavia University.
•
The Mediterranean area database “MEDACES” of the SPA-RAC, managed by the Valencia University (Spain). All the stranding events registered by MEDACES in the
available period 1996-2010 for the Adriatic basin and divided per country (Eastern Italy, Slovenia, Croatia and Albania; no information are available for BosniaHerzegovina, whilst Montenegro Republic is not included in the database) were collected and archived in ASE.
Preserved Organs and Tissues database of Adriatic Cetaceans (POTAC)
This database collects the information on stranding specimens (or those accidentally caught
by fish net), which tissues or/and organs are preserved at the National Bank of Preserved Organs and Tissues of Marine Mammals (BTMM) instituted at Padova University. No information
were obtained for the other Adriatic countries and, on the basis of information obtained by
SHAPE partners, this kind of database and/or tissues bank does not exist among the Adriatic
Balkan countries. No data were obtained from Shape Adriatic partners, because they do not
collect this kind of information. Moreover, the various institution/public entities of the Balkan
countries bordering on the Adriatic Sea, do not have or manage databases on stranding
events and/or tissue samples collection from stranding specimens. The two databases have
been geo-referenced on a GIS archive (ArcGIS, ver. 8.3; ESRI), constantly updated with the
new stranding events information). Stranding data were analysed by statistical methods to assess:
•
Possible patterns in spatial (by region) and temporal (yearly, seasonally and monthly)
stranding frequency distribution;
•
The number of events per: species, sex, time, region and localities and for a combination of two or three of these parameters;
•
The principal statistics regarding body dimensions of stranding specimens;
•
Possible differences in stranding frequency distribution between sex on the total species and between sexes for each species.
pag. 12/73
4.1.2
Phases II and III
BTMM of Padova University provided us tissue samples from specimens of Adriatic cetaceans
stored frozen at -20° and/or in formaldehyde. Tissues were obtained for no. 4 different species: Sperm whale (Physeter macrocephalus), Bottlenose dolphin (Tursiops truncates), Striped
dolphin (Stenella coeruleoalba), Risso’s dolphin (Grampus griseus) for a total of no. 17 specimens (Table 4-1). The selected tissues and storage methods were those which, according to
literature, fitted better for the specific analysis.
Table 4-1 – Specimens by species and tissue samples analysed.
ID
Species
Locality
Date
Length
(cm)
Weight
(kg)
Sex
Tissue frozen
162
T. truncatus
Comacchio
(FE)
15/07/
2009
119
20,3
M
skin-blubber, muscle,
liver,
kidney, melon
163
T. truncatus
Cavallino
(VE)
20/07/
2009
300
86
M
skin-blubber, muscle,
liver,
kidney, melon
164
T. truncatus
Comacchio
(FE)
27/07/
2009
250
125,5
M
skin-blubber, muscle,
liver, kidney
165
T. truncatus
Lido Adriano (RA)
28/07/
2009
280
167,5
M
skin-blubber, muscle,
liver,
kidney, melon
166
T. truncatus
Pellestrina
(VE)
29/07/
2009
260
75
M
skin-blubber, muscle,
liver
172
P. macrocephalus
Foggia
10/12/
2009
1.120
15,7
M
skin-blubber, muscle,
liver,
kidney, spermaceti
180
T. truncatus
Pellestrina
(VE)
27/04/
2010
209
99
M
skin-blubber, muscle,
liver, kidney
185
T. truncatus
Caorle
(VE)
08/06/
2010
292
177
M
skin-blubber, muscle,
liver, kidney
192
T. truncatus
Rosolina
(RO)
n.a.
240
178,5
F
skin-blubber, muscle,
liver, kidney
193
T. truncatus
Porto Tolle
(RO)
n.a.
280
n.a.
M
skin-blubber, muscle,
kidney, melon
196
T. truncatus
Cervia
(RA)
15/05/
2011
300
219
M
skin-blubber, muscle,
liver,
kidney, melon
203
T. truncatus
Rimini
06/07/
2011
284
288
M
skin-blubber, muscle,
kidney
214
S. coeruleoalba
Comacchio
(FE)
08/01/
2012
199
75
F
skin-blubber, muscle,
liver,
kidney, melon
Tissue in
formaldehyde
skin-blubber,
liver, spleen
pag. 13/73
ID
Species
Locality
Date
Length
(cm)
Weight
(kg)
Sex
Tissue frozen
215
G. griseus
Bibione
(VE)
11/01/
2012
300
n.a.
F
skin-blubber, muscle,
liver,
kidney, melon
218
S. coeruleoalba
Lido di
Classe
(RA)
20/01/
2012
198
81
M
skin-blubber, muscle,
liver, kidney, melon
220
S. coeruleoalba
Lido Volano (FE)
n.a.
208
100
N.I.
skin-blubber, muscle,
liver, kidney
221
S. coeruleoalba
Cattolica
(RN)
n.a.
200
78
M
skin-blubber, muscle,
liver, kidney
Tissue in
formaldehyde
skin-blubber,
muscle,
liver, kidney,
melon, spleen
*n.a.= not available
Heavy metals analysis
The tissues used were: skin-blubber, melon/spermaceti, liver, kidney and striated muscle. The
list of analysed elements is reported in Table 4-2.
Tissue samples (about 0,50 g of frozen sample, exactly weighted) were digested with 7 ml
HNO3 (67-69%), 1 ml HCl (32-35%) and 1 ml H2O2 (30-32%) in an Ethos One closed microwave system (Milestone). Digested samples were then diluted to 50 ml with ultrapure water in
a conical polypropylene tube and 10 ml were filtered through a PES (polyethersulfone) 0.45
µm syringe filter (Sartorius) and then analysed by ICP-MS (Inductively Coupled Plasma Mass
Spectrometry, Figure 4-2). A blank sample (0,50 g of ultrapure water treated as tissue samples) was performed during each digestion session. Standard curves were prepared in the
range 0,1-100 ppb (except for Hg, 0,1-5 ppb) from certified stock solution (Merck and Carlo
Erba Reagenti).
pag. 14/73
Figure 4-2 ICP-MS 7500cx System, with auto-sampler ASX-500 (Agilent Technologies).
Table 4-2 – Elements analysed by ICP-MS.
Beryllium (Be)
Aluminium (Al)
Titanium (Ti)*
Vanadium (V)*
Chromium (Cr)*
Manganese (Mn)
Nickel (Ni)*
Cobalt (Co)
Copper (Cu)*
Zinc (Zn)*
Gallium (Ga)
Arsenic (As)*
Selenium (Se)*
Rubidium (Rb)
Strontium (Sr)
Molybdenum (Mo)*
Rhodium (Rh)
Palladium (Pd)
Silver (Ag)
Cadmium (Cd)*
Indium (in)
Tin (Sn)*
Antimony (Sb)*
Tellurium (Te)
Cesium (Cs)
Mercury (Hg)
Thallium (Tl)
Lead (Pb)
Bismuth (Bi)
Uranium (U)
* elements analysed in ORS (Octapole Reaction System) mode.
Organochlorine compounds (OCs) analysis
The tissues used were: skin-blubber, melon/spermaceti, liver and kidney. The analysed compounds are reported in Table 4-2.
One (1) gram of the finely minced wet tissue was weighed in an extraction vessel and deuterated standard compounds and sodium sulphate were added to the sample. Microwave assisted extraction (MAE) of the sample, using n-hexane as solvent, was then performed with an
Ethos One closed microwave system (Milestone).
The extract was subsequently transferred in a polypropylene tube and centrifuged and the
hexanic solution was evaporated to dryness, redissolved in acetonitrile, shaking for at least 15
pag. 15/73
minutes, and kept in a freezer at about -20 °C for 24 hours. At this temperature, lipids are frozen out, having low solubility in acetonitrile, and the extracted organic solvent can be separated by filtration. This freezing-lipid filtration was repeated twice and the total organic extract
was evaporated to dryness. The dried extract was then redissolved in n-hexane and further
purified by solid-phase extraction clean-up on Florisil. The purified extract was evaporated to
dryness and finally redissolved in n-hexane in order to be analysed by GC-MS gas chromatography-mass spectrometry (Figure 4-3) in Selected Ion Monitoring (SIM) mode.
A blank sample (1 g of ultrapure water treated as tissue samples) was performed during each
extraction session. Standard curves were prepared in the range 1-1000 ppb from certified
stock solution (Sigma-Aldrich and Ultrascientific).
Table 4-3 - Organochlorine compounds analysed by GC-MS.
HCB Hexachlorobenzene
4,4’-DDT (dichlorodiphenyltrichloroethane)
2,4’-DDT
4,4’-DDD (dichlorodiphenyldichloroethane)
2,4’-DDD
4,4’-DDE (dichlorodiphenyldichloroethylene)
2,4’-DDE
Polycyclic aromatic hydrocarbons (PAHs) analysis
The tissues used were: skin-blubber, melon/spermaceti, liver and kidney. The analysed compounds are reported in Table 4-3.
One (1) gram of the finely minced wet tissue was weighed in an extraction vessel and deuterated standard compounds were added to the sample. Microwave assisted extraction and saponification (MAES) of the sample, using n-hexane as solvent and a saturated solution of potassium hydroxide (KOH), was then performed with an Ethos One closed microwave system
(Milestone).
The extract was subsequently transferred in a polypropylene tube and centrifuged and the
hexanic solution was evaporated to a volume of about 0.5 ml. The extract was then purified by
solid-phase extraction clean-up on Silica gel.
The purified extract was evaporated to dryness and finally redissolved in n-hexane in order to
be analysed by GC-MS gas chromatography-mass spectrometry (Figure 4-3) in Selected Ion
Monitoring (SIM) mode.
pag. 16/73
A blank sample (1 g of ultrapure water treated as tissue samples) was performed during each
extraction session. Standard curves were prepared in the range 1-1000 ppb from certified
stock solution (Ultrascientific).
Table 4-4 - Polycyclic Aromatic Hydrocarbons analysed by GC-MS.
Fluorene
Benzo(a)pyrene
Phenanthrene
Perylene
Anthracene
Indeno(1,2,3-cd)pyrene
Fluoranthene
Dibenz(a,h)anthracene
Pyrene
Benzo(g,h,i)perylene
Benz(a)anthracene
Dibenzo(a,l)pyrene
Chrysene
Dibenzo(a,e)pyrene
Benzo(b)fluoranthene
Dibenzo(a,i)pyrene
Benzo(k)fluoranthene
Dibenzo(a,h)pyrene
Benzo(e)pyrene
Figure 4-3 GC-MS 6890-5973N, with injector 7683 series (Agilent Technologies).
pag. 17/73
DNA oxidative products: 8-OHdG analysis
The tissues used were: liver and kidney. The tissue samples were extracted with the "Wizard
Genomic DNA Purification Kit” (PROMEGA) according to the manufacturer's protocol.
Briefly, few mg of the samples were treated with "Nuclei Lysis Solution"/EDTA and digested
overnight with Proteinase K at 55 °C, followed by digestion with RNase for 15-30 minutes at
37 °C. Then, the proteins were precipitated with the "Protein Precipitation Solution" and, after
centrifugation, the supernatant was recovered and the DNA was precipitated by adding isopropyl alcohol.
The DNA obtained was washed with 70% ethanol, dried and resuspended in the "DNA Rehydratation Solution". The concentration and the purity of DNA were analysed by spectrophotometric readings at 260 and 280 nm, and the quality of the DNA was determined by horizontal
agarose gel electrophoresis.
8-hydroxy-2'-deoxyguanosineis (8-OHdG) was determined on 300 ng/sample of purified DNA
by ELISA using the "EpiQuick™ 8-OHdG Direct DNA damage quantification kit (colorimetric)"
(EPIGENETEK) according to the manufacturer's protocol. Results were expressed as weight
percentage of 8-OHdG on the total analysed DNA. The total specimens tested were 12: no. 8
Tursiops truncatus, no. 3 Stenella coeruleoalba and no. 1 Physeter macrocephalus.
Micronuclei (MN) assay
The tissues used were: liver, kidney, muscle, skin-blubber and melon. The micronucleus test
allows to highlight chromosomal aberrations (breaks or misdistribution of chromosomes) resulting in expulsion from the nucleus fragments or whole chromosomes, which are called micronuclei; it may occur spontaneously or be induced by mutagens. On prepared slides for
viewing under a microscope free intracellular corpuscles appear as micronuclei, they have
round shape, with dimensions much smaller than those of the principal nucleus. Tissues were
fixed in Formalin (10% pH7.4). Small pieces were cut from each tissue, then dehydrated and
paraffin-embedded.
Protocol for dehydration
2 changes in 70% ethanol 30 min. each;
2 changes in 80% ethanol 30 min. each;
2 changes in 96% ethanol 30 min. each;
2 changes in 100% ethanol 30 min. each;
2 changes in 70% ethanol 30 min. each;
2 changes in xylene 30 min. each.
2 changes in liquid paraffin 60 min. each in the oven at 60 degrees.
Tissues were then embedded in the mold.
For each paraffin-embedded tissue were cut 5 sections (5 µm each) at microtome.
pag. 18/73
Protocol for micronuclei labelling
5 µm thick sections of formalin- fixed, paraffin-embedded tissues are rehydrated by
2 changes in xylene (10 min /each),
1 change in 100% ethanol (5 min),
1 change in 95% ethanol (5 min),
and by washing samples in 0.01M PBS under stirring (10 min).
Then sections are incubated for 30 min. at room temperature with Hoecst stain solution
(SIGMA-ALDRICH code no.H6024) a fluorescent DNA stain (1/10000 in 0.01 M PBS). Tissues
were washed 3 times in 0.01M PBS pH7.4, then 10 min under stirring and the coverslips
mounted with MowiolTM and evaluated under Zeiss-Axiophot microscope.
For each section no. 5 images were taken and the number of MN evaluated using Image J
software (imagej.en.softonic.com) and expressed as the percentage of MN on the total image
area. For each tissue the average of all the values obtained has been calculated.
pag. 19/73
5
Pilot project’s results
5.1
Collection of literature on eco-toxicological studies
Available literature regarding both stranding events and eco-toxicological investigation on
Mediterranean cetaceans, mainly those from Adriatic Basin, were collected in order to reconstruct historical and eco-toxicological researches and trends (Table 5-1).
Table 5-1 Literature data collected.
Topic
Number
Biology and ecology of Mediterranean cetaceans
21 (and 20 CSC reports)
Stranding events along the Italian coasts
18
Collection, studies and biopsies of tissue / organs of
stranded animals
15
Eco-toxicological studies on heavy metals
11 (no. 5 referred to Adriatic sea)
Eco-toxicological studies on OCs/PAHs
29 (no. 2 referred to Adriatic sea)
Biomarker studies
10 (no one referred to Adriatic
sea)
The Mediterranean Sea is home to approximately 20 different cetacean species (UNEP/
IUCN, 1994). The most abundant dolphin species are Tursiops truncatus (bottlenose dolphin),
Grampus griseus (Risso’s dolphin) and Stenella coeruleoalba (striped dolphin) and Delphinus
delphis (common dolphin).
Adriatic cetaceans population have been very poorly studied, both for ecological and ecotoxicological aspects. In fact, researches have been concentrated on Tirrenic side and mainly
regarded the most common Delphinidae present in the western Mediterranean side: Stenella
coreuloelba. Researchers are carried out principally by three organizations: Cetaceans Studies Centre of Milan’s Civic Museum of Natural History, for that concerning the stranding phenomenon; University of Pavia for bioacoustics researches and University of Siena for ecotoxicological facets.
Cetaceans of big dimensions are not habitual species in Adriatic sea, because of the little
deep bottoms, while species belonging to Delphinidae family are easier observable. The bottlenose dolphin is the only regularly encountered species in Adriatic waters. It is distributed all
across the Adriatic and is being studied in a few locations. In Italian waters, the only regularly
observed species in the Gulf of Venice during the last 15 years is the bottlenose dolphin (Tur-
pag. 20/73
siops truncatus). It is currently being studied in three locations of the western Adriatic sea:
around the island of Vis, in the Cres-Losinj archipelago and off the coast of Slovenia.
Striped dolphins and fin whales are sighted occasionally, while other species can be considered rare. Recent data show the occasional presence of the D. delphis in Dalmatia and Quarnero (Notarbartolo di Sciara and Demma, 1994) and the existence of a resident population in
Eastern Ionian Sea (Politi et al., 1999). Risso's dolphin (Grampus griseus) is still present principally in South Adriatic, where waters are deeper. It can be possible to sight occasionally the
fin whale (Balenoptera physalus) and the sperm whale (Physeter macrocephalus). Nevertheless, sperm whale needs deep waters for its hunt dives (in fact, it can go over the deep of
2,000 m). Among dolphins, two species can be considered proper to the Adriatic sea: the bottlenose dolphin (Tursiops truncatus) and the common dolphin (Delphinus delphis); nevertheless, in the last years the population of striped dolphins (Stenella coeruleoalba) seem to be increased in Adriatic basin (Table 5-2).
Table 5-2 The three dolphin species usually living in the Adriatic sea.
Species
Morphological characteristics
Ecology
Dimensions: 2-3 m
arched and curved dorsal
fin
rather dark back
body and head stout
Diet: fishes and cuttlefish
no. per groups: 1-10
Threats: pollution,
fishing, habitat alteration, sea human
activity
Dimensions: 2,5-2,6 m
dark back
arched and small dorsal
fin
dark back
Blue, white and pink
womb
Diet: squids, crustaceans, little fishes
No. per groups: 1030
Threats: pollution,
net, fishing
Dimensions: 2-3 m
dark back
colour hourglass on flanks
light womb
dark fins
Diet: fishes and cuttlefish
no. per groups: 1030
Threats: net and
fishing, pollution
Tursiops truncatus
Stenella coeruleoalba
Delphinus delphis
pag. 21/73
Among cetaceans, bottlenose dolphin is the most known, due to its ability to adapt at captivity
life allowing scientists to easily study it. Nevertheless, it is not easily recognizable on sea.
There are two ecotypes: a pelagic one, with bigger dimensions, who lives in numerous groups;
the inshore ecotype is of smaller dimensions and usually solitary. Common dolphin, as its
name indicates, was common in the past but now it is very rare in the Mediterranean waters.
Currently the number is considerably reduced: every sighting is exceptional. Groups are often
numerous; in frightening situations single specimens tends to groups in compact cluster, for
better facing dangers.
Stranding of dolphins in the Mediterranean Sea has been reported in literature (André et al.,
1991; Augier et al., 1993; Monaci et al., 1998, among others).
Interest in cetaceans has mainly been related to their economic value. They are actively marketed live for the tourist industry and dead for the food, cosmetic and chemical industries. Cetaceans are not hunted in the Mediterranean Sea and there has never been a tradition of
whale hunting in Italy. Trawling for tuna, swordfish and albacore causes the death by suffocation of many cetaceans, especially dolphins. Cetacean populations living in this environment
are also jeopardized by the rapid increase in levels of man-made chemicals in the marine environment in the second half of this century. The case of heavy metals and chlorinated hydrocarbons (HCB, DDTs and PCBs) are some examples.
5.1.1
Heavy metals
Extremely high concentrations of mercury have been reported in the livers of striped dolphin
from the Mediterranean coast of France and Italy (André et al. 1991; Storelli et al. 1999). Anyway, mercury levels in dolphins stranded on the south eastern Mediterranean coasts (Adriatic
coast) of Italy (Cardellicchio et al., 2000) and of Croatian coasts are generally lower than those
found in dolphins from the French Mediterranean coasts (André et al., 1991; Augier et al.,
1993), from the northern Mediterranean coasts of Italy (Ligurian Sea; Capelli et al., 2000),
from Corsican coast of the Mediterranean (Frodello et al. 2000) and from the Israeli coast of
the Mediterranean (Shoham-Frider et al. 2002).
The main studies on heavy metals in Adriatic cetaceans are on stranded specimens (Table
5-3) along the Apulia coasts (Southern Italy) by Storelli et al. (1999) and Cardellicchio et al.
(2000, 2002a, 2002b for Adriatic and Ionian seas) and by Bilandžić et al. (2012) in the Croatian side.
pag. 22/73
Table 5-3 Literature studies on heavy metals concentrations in Adriatic cetaceans.
Authors
Area
Species
Heavy metals
analysed
Organs/tissues
analysed
Storelli et al.
(1999)
Southern Adriatic
Grampus griseus
Ziphius cavirostris
Hg, Se, Cd, Pb,
Cr, MeHg
muscle, liver,
kidney
Cardellicchio et al.
(2000, 2002a,
2002b)
Apulia coasts
(Adriatic and Ionian seas)
Stenella coeruleoalba
Hg, Pb, Cd, Cr,
Zn, Cu, Fe, Mn
MeHg, Se
Liver, lung, kidney, muscle,
brain, melon
Bilandžić et al.
(2012)
Eastern Adriatic
(Croatian waters)
Tursiops truncatus
Stenella coeruleoalba
Grampus griseus
Cd, As, Hg, Pb
muscle, liver,
kidney
The distribution of different metals (mercury, lead, cadmium, chromium, zinc, copper, iron,
manganese, methyl mercury and selenium) were investigated in various tissues and organs
obtained from striped dolphins (Stenella coeruleoalba) stranded along the Apulian coasts
(Southern Italy) during 1987 (Cardellicchio et al., 2000) and 1991 (Cardellicchio et al., 2002a,
2002b) by atomic absorption spectrophotometry. The results obtained were:
1.
Metal levels were higher than those found in dolphins living in the Atlantic, but lower
than those recorded in the same species from the French Mediterranean coasts.
2.
Analysis showed considerable variations in the mercury concentration in the examined organs and tissues.
3.
The liver accumulated the highest concentrations of metals (especially mercury and
selenium), except for cadmium and chromium. The highest concentrations of mer-1
cury were found in the liver (from 2.27 to 374.50 µg g wet wt.).
4.
After the liver, lung, kidney, muscle and brain were the most contaminated, while the
lowest mercury contamination was found in the melon. Metal concentrations were
generally low in brain and melon.
5.
Some metals showed organ-specific accumulations: copper, tin and zinc exhibited
high concentrations in liver, the highest cadmium concentration was observed in kidney.
6.
Pathological, microbiological and parasitological surveys were performed on the animals. By necroscopic surveys it was found that some specimens were affected by
hemorrhagic gastritis, but the cause was not clear.
7.
It was not possible to relate the death of dolphins to a specific cause or to contaminants, but the accumulation of metals was likely to contribute to the health of the organism and represents a risk factor for dolphins.
pag. 23/73
For the Mediterranean Sea, unlike the case for species such as striped, bottlenose and common dolphins, few studies are focused on the Risso's dolphin (G. griseus) and Cuvier’s
beaked whale (Z. cavirostris), probably due to the relatively scantiness of stranding specimens
of both species. The few studies regarding the species above mentioned were conducted instead right in the Adriatic Sea (Storelli et al. 1999).
Trace metals concentrations (Hg, Se, Cd, Pb and Cr) and methylmercury have been studied in
the muscle, lung, liver and kidney of three specimens-samples of these two species stranded
along South Italy coast (South Adriatic Sea). Total mercury highest concentration in Z. cavirostris (one specimen) were found in the liver, followed by kidney, lung, and muscle. In G. griseus (two specimens) the highest total mercury level were found in liver, followed by lung, kidney, and muscle (Table 8). In addition to inter-tissue variations, inter-species differences in
metal concentrations were observed. The concentration of metals analysed in all tissues of Z.
cavirostris were lower than those of G. griseus, except for cadmium which showed higher levels in kidney and liver in Z. cavirostris compared to G. griseus.
Data regarding heavy metal concentrations in bottlenose (Tursiops truncatus), striped (Stenella coeruleoalba) and Risso’s (Grampus griseus) dolphins inhabiting the Adriatic Sea off the
coast of Croatia were recently obtained by Bilandžić et al. (2012). They are important results
which can also give an indication of the environmental condition with regard to the content of
toxic metals along the eastern coast of the Adriatic Sea (Croatian waters). Concentrations of
cadmium (Cd), arsenic (As), mercury (Hg) and lead (Pb) were measured in muscle, liver and
kidney of the three cetacean species.
In all three dolphin species Cd levels decreased in tissues in the order: kidney > liver > muscle, while As and Pb decreased in the order: liver > kidney > muscle for striped and Risso’s
dolphins, but with the order reversed for liver and kidney in the bottlenose dolphin for Pb. Levels of Hg consistently followed the order: liver > muscle > kidney, with mean concentrations in
the liver being 11–34 times higher than in the other tissues. The highest mean concentrations
of trace elements were measured in Risso’s dolphins.
Statistically significant differences between the three dolphin species were determined for Cd,
Hg and Pb in liver tissues, for As in muscle and for Cd in kidney. Significant correlations of
metals between tissues were determined in all three species.
The mean liver Hg levels determined in Risso’s dolphin were more than 5-times higher than
those in bottlenose and striped dolphins. It is known that Risso’s dolphin mainly feed on squids
(85%) and bottlenose dolphins mainly feed on fish (70 %), while the diet of striped dolphins is
more varied and is comprised of 35% squid and 55% fish (Shoham-Frider et al. 2002). Composition and characteristics of cephalopod prey in stomach contents of striped dolphins
showed a mixed diet of muscular and gelatinous-bodied squids, mainly consisting of oceanic
and pelagic or bathypelagic species (Blanco et al. 1995). Furthermore, comparisons between
bottlenose and striped dolphins do not reveal any difference in Hg accumulation in the Atlantic.
In Bilandžić et al. (2012) levels determined were comparable to those from the southern Adriatic Sea (Storelli et al. 1999) but higher than those from the Corsican coast and the Ligurian
Sea of the Mediterranean (Frodello et al. 2000; Capelli et al. 2008).
pag. 24/73
5.1.2
Organic contaminants
Endocrine disrupting chemicals (EDCs) are a structurally diverse group of compounds that
may adversely affect the health of humans, wildlife, and fisheries, or their progenies, by interaction with the endocrine system. They include chemicals used heavily in the past, in industry,
and agriculture, such as polychlorinated biphenyls and organochlorine pesticides, and currently used, such as plasticizers and surfactants.
Many of the known EDCs are estrogenic, affecting in particular reproductive functions. Because of the lipophilic and persistent nature of most xenobiotic estrogens and their metabolites, many bioaccumulate and biomagnify. Some geographic areas are potentially more
threatened than others by EDCs: one of these is the Mediterranean Sea. This basin has limited exchange of water with the Atlantic Ocean, and is surrounded by some of the most heavily
populated and industrialized countries in the world. Levels of some xenobiotics are much
higher here than in other seas and oceans. Mediterranean marine fauna could therefore be a
target of EDCs. In this peculiar environment, top predators (such as large pelagic fish and marine mammals) tend to accumulate high quantities of polyhalogenated aromatic hydrocarbons
and toxic metals and are potentially “at risk” due to EDCs contamination.
The first warning about toxicological risk to large Mediterranean pelagic fish due to Endocrine
Chemical Disrupters (EDCs) was pinpointed by the results of Fossi et al. (2001) in swordfish
and Fossi et al. (2002) in bluefin tuna. Levels of OCs in free-ranging striped dolphins are 10 to
20 times higher than in swordfish.
Four types of organochlorine endocrine disruptors (Adami et al., 1995; Hilscherova et al.,
2000; Fossi and Marsili, 2003) are commonly found in Mediterranean cetaceans (Aguilar et al.,
2002; Marsili, 2000; Fossi et al., 2003): 1) environmental estrogens, 2) environmental androgens, 3) antiestrogens and 4) anti-androgens.
The relative estrogenic power of these chemicals, identified by in vitro and in vivo screening
methods (Safe, 1995, 2000) is rather weak (10−3 or less) compared with the reference of 17estradiol or DES (Miyamoto and Klein, 1998). However, the high levels of organochlorine
compounds detected in marine mammals, particularly in pinnipeds and odontocetes, and consequently, the high levels of organochlorines with ED capacity, cannot be ignored. Some general considerations on potential hazard to these Mediterranean species can be drawn from
comparison of the data commonly detected in Mediterranean cetaceans and that of other cetacean species with known reproductive impairment (Fossi and Marsili, 2003). Several examples suggest that exposure to OC insecticides and PCBs has affected endocrine function and
reproduction in marine mammals. Here, it is worth noting that levels of PCBs found in Mediterranean free ranging Odontocetes (Fossi et al., 2003) are similar to those detected in the population of beluga whales of the St. Lawrence estuary where a hermaphrodite specimen was detected (PCBs mean value=78,900 ng/g lipid mass (l.w.)) (Muir et al., 1996); levels of PCBs detected in Mediterranean free ranging fin whales in the same period (mean value=7,331 ng/g
l.w.) (Fossi et al., 2003) are approximately 10 times higher than those found in the population
of bowhead whales (Balaena mysticetus) where pseudo-hermaphroditism and other reproduc-
pag. 25/73
tive dysfunctions have been detected (PCBs mean value=610 ng/g l.w.) (Tarpley et al., 1995).
This observation suggests the potential hazard that these species are exposed to in the Mediterranean Sea.
Organochlorine compounds (OCs)
The levels of OCs in a top predator of the Mediterranean, the striped dolphin (Stenella coeruleoalba), are 1–2 orders of magnitude higher than in Atlantic and Pacific dolphins of the
same species (Marsili, 2000). This suggests the hypothesis that Mediterranean top predator
species are potentially “at risk” due to EDs contamination.
Marsili et al. (1997) analysed the concentrations of OCs (HCB, DDTs and PCBs) in the tissues
and organs of the following cetaceans: Stenella coeruleoalba, Tursiops truncatus, Balaenoptera physalus, Steno bredanensis, Grampus griseus and Globicephala melaena stranded
along the Italian coasts in the period 1987–1993 (results for DDTs and PCBs in Figure 4). The
values are compared between species and between specimens of the same species. Chlorinated hydrocarbon (CH) levels were found to increase in relation to the quantity and type of lipids in each tissue and organ. Differences in accumulation encountered in the different species
are principally due to different feeding habitats. Remarkable differences found between males
and females of each species confirm that during gestation and lactation, females undergo detoxification by passing much of their total burden of CHs to their young. The study concluded
that Mediterranean cetaceans are an extraordinary example of organochlorine bioaccumulators. There are marked differences between species and in a given species, storage is related
to many parameters such as sex and age. Differences between males and females were almost always significant in most of the organs and tissues analysed, especially after sexual
maturity. The main storage tissue in all species was confirmed to be blubber. In view of its
contribution to total body weight (about 20%) its importance in the total burden of contaminants is evident.
pp’DDE accounted for more than 65% of total DDTs in all samples analysed. This refers to
contamination occurring considerable time ago and indicates the good metabolic capacity of
these marine mammals to transform DDT. The values of the PCBs/DDTs ratio show that most
CH input in these animals has been due to PCBs. The levels in relation to sampling years,
show an evident ratio shifting in favour of PCBs. Hexachlorobenezene was detectable in most
samples, but its concentrations were enough low not to be determinant in total CH burden. Although epidemics such as that of Morbillivirus can influence xenobiotic storage, it isn’t known
whether the disease is the cause or effect of high concentrations of organochlorines in the tissues. There were large epidemics of this virus in the Mediterranean in 1990 and 1991. The
levels of CHs found in the organs and tissues of cetaceans stranded in this period were significantly higher than in prior and successive years.
The results of these studies are synthesized in Figure 5. It is evident that the species with the
highest concentrations of both compounds is the Risso’s dolphin (Grampus griseus) followed
by the striped dolphin (Stenella coreuloelba).
pag. 26/73
Figure 5-1 DDTs and PCBs concentrations in Tyrrhenian cetaceans’ populations
(Marsili et al., 1997).
Fossi et al. (2000) used a non-destructive approach, the skin biopsy (subcutaneous tissue
consisting of skin and blubber of about 1-2 cm) to explore OCs bioaccumulation processes
and mixed function oxidase activity (BPMO) in four species of cetaceans: striped dolphin
(Stenella coeruleoalba), bottlenose dolphin (Tursiops truncatus), common dolphin (Delphinus
delphis) and fin whale (Balaenoptera physalus). Sampling was performed in the western Ligurian Sea (1992 until 1998), between Corsica and the French-Italian coast, and Ionian using
biopsy darts. Significant differences in BPMO induction and OC levels were found between
Odontocetes and Mysticetes, the former having mixed-function oxidase activities four times
higher than the latter, binding with levels of OCs one order of magnitude higher in Odontocetes. A significant correlation (P<0.05) between BPMO activities and OC levels was found in
B. physalus. Fibroblast cultures have been used as an alternative in vitro method of evaluating
interspecies susceptibility to contaminants such as OCs and Polycyclic Aromatic Hydrocarbons (PAHs). These results suggest that cetacean skin biopsies are a powerful non-invasive
tool for assessing eco-toxicological risk to Mediterranean marine mammals species.
Polycyclic Aromatic Hydrocarbons (PAHs)
Marsili et al. (2001) represents the only study available on the influence of PAHs accumulation
on Mediterranean cetaceans tissue by biopsy on free ranging specimens (subcutaneous blubber samples). The study involved two species: the fin whale (Balaenoptera physalus) from the
Ligurian sea and the striped dolphin (Stenella coeruleoalba) from Ligurian and Ionian seas.
Because these animals feed at different levels of the pelagic food chain, the aim was to determine whether a different PAH accumulation would follow.
pag. 27/73
Dolphins from two different sections of the Mediterranean sea were collected to detect correlation between PAH levels and habitat; they also investigated any relation between PAH levels
and gender in fin whales and analysed the three years trend of PAHs from 1993-1996.
The conclusions of the work were:
•
PAHs were present in both species. Therefore, Mediterranean cetaceans are exposed to PAHs.
•
The comparison between the total and carcinogenic PAH concentrations on whales
and dolphins showed a greater accumulation in subcutaneous blubber of dolphins
than in whales indicating differences in accumulating or metabolizing these compounds related to their different positions in the marine food chain. The same result
was found comparing whales and striped dolphins sampled only in the Ligurian sea.
•
No significant differences were found between Ligurian and Ionian dolphins population and between sex in fin whales, but only comparing samples of the three years
(much higher amount in 1993 compared to other years) probably due to the incident
of the tanker Haven in 1991.
Organic contaminants
Storelli and Marcotrigiano (2000) determined the levels of polychlorinated biphenyls (PCBs)
DDT and related compounds, and hexachlorobenzene (HCB) in tissues (blubber, heart, lung,
liver, spleen, kidney, stomach and muscle) of G. griseus beached along the Apulian coast.
The organochlorine concentrations decreased in the order DDTs>PCBs>HCB in all tissues
except for heart and liver, which exhibited higher PCB levels. Concentrations of DDT and related compounds (DDTs) were the highest in the blubber, followed in decreasing order by the:
heart, spleen, liver, muscle, lung, kidney and stomach. The major constituent of DDTs was
p,p’-DDE, accounting for at least 80% of DDTs in all tissues. HCB was present in all tissues
with similar concentrations, with the exception of blubber where the level was higher, with
comparable values found in the blubber of the same species collected from the west coast of
North America.
Storelli et al. (2012) measured the concentrations of polychlorinated biphenylthe (PCBs) including dioxin-like PCBs in different organs and tissues (melon, blubber, liver, kidney, lung,
heart, and muscle tissue) of striped dolphins (Stenella coeruleoalba) from the Adriatic Sea.
The mean highest levels were in blubber and melon, followed by liver, kidney, lung, heart, and
muscle tissue. PCB profiles were similar in all tissues and organs being dominated by the
higher chlorinated homologues.
pag. 28/73
Table 5-4 Available studies on organic contaminants in Adriatic sea cetaceans.
Authors
Area
Storelli and Marcotrigiano (2000)
Apulia coasts
(Southern
Adriatic sea)
Storelli et al
(2012)
5.2
Adriatic Sea
Species
POPs analysed
Organs/tissues
analysed
Grampus
griseus
Polychlorinated biphenyls
(PCBs) DDT and related
compounds, hexachlorobenzene (HCB)
blubber, heart,
lung, liver,
spleen, kidney,
stomach,
muscle
Stenella coeruleoalba
Polychlorinated biphenyls
(PCBs) including dioxin-like
PCBs (non-ortho, PCB
77, PCB 126, and PCB 169
and mono-ortho, PCB
105, PCB 118, and PCB 156)
melon, blubber,
liver, kidney,
lung, heart,
muscle
The Adriatic Stranding Events (ASE) Database
ASE is actually composed by:
•
no. 970 records overall the Italian side of the Adriatic Sea (about the 23% of the
whole stranding events along the Italian coasts) and no. 1 record from Croatia (Tables 9 and 10 show the number of stranding events registered from Italian coast per
region and species respectively) deriving from the Coordination Centre of CISBRA.
•
no. 567 records overall the Adriatic basin, considering all the country bordering the
Adriatic sea except Montenegro Republic (data missing) as reported in Table 5-7.
The real total number of stranding events recorded for the Adriatic basin area is no. 1,187.
This is because from a cross-comparison between the two databases, the no. 351 stranding
events registered by Medaces for Eastern Italy coast (Table 11 and Figure 6) are already included in the Italian database source.
For each event the following data are reported on ASE: date, location, competent authority
which issued the alert, geographic coordinates (UTM system), species (if identified), size
(measured or estimated), sex (if identified) and any information about the animal status (if
stranding alive or dead, the decomposition state and the method of disposal), as well as additional useful notes if available (if it will have made inquiries necropsy or biopsies, if any organs/tissues were collected for storage in the BTMM, photos available, etc.).
pag. 29/73
Table 5-5 Number of specimens stranding (period January 1902 - November 2013) in
each sector of the Adriatic basin (Italian side) recorded by the Italian national database.
Sector
Number
Northern Adriatic
328
Central Adriatic
276
Southern Adriatic
366
Total
970
Table 5-6 Number of stranding specimens per species along the Italian Adriatic coasts
recorded by Italian national database (period January 1902 - November 2013).
Species
Suborder
Family
Mysticetes Balaenopteridae
Physeteridae
Common English name
Balaenoptera physalus
Fin Whale
(Linnaeus,1758)
Delphinidae
Ziphiidae
%
5
0,52
Sperm whale
Physeter macrocephalus
(Linnaeus,1758)
9
0,93
Common dolphin
Delphinus delphis
(Linnaeus, 1758)
6
0,62
Striped dolphin
Odontocetes
No.
Scientific name
Stenella coeruleoalba
(Meyen, 1833)
182 18,76
Bottlenose dolphin
Tursiops truncates
(Montagu, 1821)
Risso's Dolphin
Grampus griseus
(Cuvier, 1812)
23
2,37
Long-finned
pilot whale
Globicephala melas
(Traill, 1808)
1
0,10
Cuvier's
beaked whale
Zifius cavirostris
(Cuvier, 1823)
6
0,62
539 55,57
Undetermined
199 20,36
Total
970 100,00
pag. 30/73
Table 5-7 Number of stranding specimens per species and country along overall the
Adriatic basin coasts recorded by MEDACES (period: 1996 - 2010).
Species
Eastern
Italy
Slovenia
BosniaHerzegovina
T. truncatus
192
4
0
152
2
350
S. coeruleoalba
53
0
0
19
0
72
G. griseus
12
0
0
9
0
21
G. melas
1
0
0
0
0
1
Z. cavirostris
8
0
0
4
0
12
P. macrocephalus
1
0
0
0
0
1
B. physalus
2
0
0
2
0
4
Undetermined
82
0
0
24
0
106
Total x country
351
4
0
210
2
567
Croatia Albania
Total x species
Eastern Italy
Slovenia
Croatia
0,7%
Albania
37,0%
61,9%
0,4%
Figure 5-2 Number and percentage of stranding events recorded by MEDACES for the
Adriatic basin (1996-2010).
pag. 31/73
Figure 5-3 Screen graphic representation of the thematic "strandings in the central
Adriatic” (period: 1902 – 2013) of the developed GIS (ArcGIS, 8.3).
5.3
The Preserved Organs and Tissues of Adriatic Cetaceans
database (POTAC)
POTAC is composed, at the moment, by no. 63 records (time period: 2000-2013) regarding
specimens from overall the Italian side of the Adriatic sea (Table 5-7). For each record are indicated: belonging species, date, localization, sex, cause of death, available organs and tissues and their modality of preservations (if freezing or preserved in aldehydes).
pag. 32/73
Table 5-8 Number of found specimens in Adriatic Italian side per species whose tissues
are preserved at BTMM.
Sector
Species
Total
no.
Northern
Adriatic
Central
Adriatic
Southern
Adriatic
Not
known
Grampus griseus
3
1
-
1
5
Physeter macrocephalus
-
-
7
-
7
Stenella coeruleoalba
7
3
-
1
11
Ziphius cavirostris
-
-
-
1
1
Tursiops truncatus
32
1
-
6
39
Total
42
5
7
9
63
5.4
Stranding events data elaboration
Stranding data were analysed to assess:
•
Possible patterns in spatial (by region) and temporal (yearly, seasonally and monthly) stranding frequency distribution;
•
The number of events per: species, sex, time, region and localities and for a combination of two or three of these parameters;
•
The principal statistics regarding body dimensions of stranding specimens;
•
Possible difference in stranding frequency distribution between sex on the total species and between sexes for each species.
Geographical distribution of stranding is homogeneous along the Adriatic coast (Figure 5-4).
pag. 33/73
Figure 5-4 Cartographic stranding events along the Italian Adriatic coast (1902 - 2013;
ArcGis 8.3).
Strandings appear quite regularly, with a polynomial distribution (Figure 5-5) over the years,
except for 1991, particularly in the Central Adriatic (Figure 5-6) in which there is a strong increment probably caused by the measles outbreak that attacked marine mammals populations
(particularly of Stenella coeruleoalba Figure 5-7), in the period 1990-1991. Anyway, the time
period analysed (substantially from 1982 to 2013 because there is only one report dating back
pag. 34/73
to 1902) is too short to identify the existence of a cyclical nature of the phenomenon, as shown
instead in other contexts (Evans et al., 2005). In some studies it has been demonstrated the
presence of a 10-11 years cycle in relation to climatic events and the large-scale solar tempest, which cover a cycle of the same period of time.
Figure 5-5 Annual distribution of stranding events.
pag. 35/73
Figure 5-6 Annual distribution of stranding events by region (period 1987-2013).
Figure 5-7 Annual distribution of stranding events by species.
pag. 36/73
It is noticed an increase in the number of stranding during the summer months, which could
be partly due to the number of sightings during the bathing season; however, there is a peak
that moves along a north-south gradient: in the northern regions in July, in the central regions
between July and August and in the southern regions in August-September (Figure 5-8).
There is also a difference in the frequency peak of stranding among species (Figure 5-9) during the summer period: in June for Risso's dolphin (Grampus griseus); in July for the bottlenose dolphin (Tursiops truncatus) and between August and September for the striped dolphin
(Stenella coeruleoalba).
Figure 5-8 Monthly distribution of stranding events by region (period 1902-2013).
pag. 37/73
Figure 5-9 Monthly distribution of stranding events by species (period 1902-2013).
Figure 5-10 shows sex distribution of strandings for each species. Anyhow, there doesn’t
seem to be a different probability to strand between sexes of the different species: there are
not significant differences between total males and females and between males and females
per species (no-parametric tests: Kolmogorov-Smirnov and Kruskal-Wallis H, respectively).
Statistical analysis didn’t highlight difference in body dimension of stranding specimens by
species. From the mean length values of the body dimension of undetermined specimens
(Figure 5-10), it is possible to say that they are mainly dolphins (probably and principally bottlenose and striped dolphins); however, considering the maximum value of body length (10 m)
there are probably also some specimens belonging to species of larger dimensions, as for example the Sperm whale.
pag. 38/73
Figure 5-10 Distribution of stranding events by sex per species.
Table 5-9 Mean body length of the strandings specimens per species.
Species
No.
Mean
Std. Dev.
Median
Min.
Max.
(m)
Tursiops truncatus
463
2.49
1.69
2.32
0.56
20.00
Stenella coeruleoalba
143
2.16
1.46
2.00
0.60
16.00
Delphinus delphis
6
2.34
0.31
2.40
1.87
2.68
Grampus griseus
20
2.51
0.57
2.65
1.65
3.50
Globicephala melas
1
3.00
3.00
3.00
3.00
Ziphius cavirostris
5
2.03
0.72
2.00
1.26
3.20
Physeter macrocephalus
9
3.05
2.00
2.70
1.65
8.25
Balanoeptera physalus
5
5.61
5.23
2.80
1.93
14.20
Undetermined
143
2.31
1.06
2.22
0.65
11.80
pag. 39/73
5.5
Heavy metals data elaboration
The complete results for elements for each specimens per tissue are reported in Table I of the
Appendix to this report.
The results in the selected tissues are reported by species in the Table 5-10 and Table 5-13
and graphically visualized in the corresponding figures. Elements that resulted always below
the quantification limits were not considered for statistical analysis.
The heavy metals resulting in the highest concentrations were: mercury (Hg), zinc (Zn) and
selenium (Se) in all the analysed tissues although with high variation in order of magnitude.
Heavy metals tend to accumulate mainly in liver, kidney and muscle, rather than in fat tissue,
like blubber or melon; the highest concentrations found were: mercury in liver, muscle and
kidney, zinc and selenium in all the analysed tissues for all the 4 species. Hg is principally
concentrated in the liver; the kidney and the muscle are the next organs in terms of mercury
uptake followed to skin blubber and the melon; Cd tends to accumulate preferentially in the
kidney, with lower concentrations found in the other tissues.
As it can be seen in Tables (Table 5-10 and Table 5-13) and correspondent figures (Figure
5-11 and Figure 5-14), there is a large variation in the metals concentration among tissues and
species. Even within the same species, as shown by the standard deviations, sometimes there
is a rather wide variation within the same tissue among specimens: for example for mercury
(Hg), selenium (Se) and zinc (Zn) in liver and/or muscle of bottlenose dolphins (Tursiops truncatus) and striped dolphins (Stenella coreuleoalba).
Anyway, the ANOVA didn’t highlight significant statistical differences in mean concentration of
the various elements among species and tissue. Particularly, there were no significant statistical differences in the same tissue of the four species and also in the concentration of the elements in different tissues of the same species, with the exception of both kidney in G. griseus
and P. macrocephalus (p<0,5).
pag. 40/73
Table 5-10 Mean concentration (mg/kg ww) of elements in Bottlenose dolphin (Tursiops
truncates) by tissue (number of available samples = 11).
Element
Skin-blubber
Muscle
Liver
Kidney
Melon
Mean
S.D.
Mean
S.D.
Mean
S.D.
Mean
S.D.
Mean
S.D.
Cr
0,154
0,316
0,01
0,012
0,01
0,009
0,01
0,010
0,06
0,080
Ni
0,277
0,502
0,56
0,329
2,00
1,003
0,38
0,222
0,08
0,114
Cu
0,173
0,148
1,01
0,435
11,14
12,035
3,12
1,581
0,08
0,138
Zn
24,890
10,709
18,39
10,564
44,17
47,396
16,34
7,759
3,46
0,706
As
0,543
0,322
0,19
0,098
0,30
0,322
0,33
0,272
0,19
0,053
Se
1,142
0,881
6,91
15,562
39,47
69,643
3,26
2,688
1,13
0,964
Rb
0,394
0,146
1,22
0,332
1,04
0,258
1,00
0,169
0,09
0,027
Cd
0,002
0,005
0,01
0,024
0,32
0,853
1,05
1,906
0,01
0,022
Sn
< 0,10
< 0,10
0,09
0,198
0,34
0,302
< 0,10
< 0,10
0,03
0,057
Sb
0,004
0,011
0,004
0,014
0,004
0,011
< 0,01
< 0,01
0,01
0,016
Hg
2,386
1,805
28,63
60,962
226,91
457,835
10,89
12,295
3,45
3,073
Pb
0,024
0,051
< 0,01
< 0,01
0,04
0,040
0,01
0,010
0,01
0,009
Figure 5-11 Mean concentration (mg/kg ww) of elements in Bottlenose dolphin
(Tursiops truncates) by tissue.
pag. 41/73
Table 5-11 Mean concentration (mg/kg ww) of elements in Striped dolphin (Stenella
coreuleoalba) by tissue (number of available samples = 4).
Element
Skin-blubber
Muscle
Liver
Kidney
Melon
Mean
S.D.
Mean
S.D.
Mean
S.D.
Mean
S.D.
Mean
S.D.
Cr
0,27
0,382
0,02
0,019
0,16
0,213
0,14
0,280
0,01
0,011
Ni
0,27
0,265
0,94
0,676
1,02
0,285
0,48
0,411
0,03
0,040
Cu
0,87
1,258
2,49
3,275
4,86
2,914
2,01
1,386
0,18
0,001
Zn
46,37
33,996
18,19
20,431
20,57
12,470
34,44
33,791
25,47
30,617
As
0,78
0,062
0,46
0,228
0,75
0,458
0,75
0,481
0,36
0,288
Se
7,42
5,454
8,56
14,581
19,45
13,025
6,92
6,564
7,25
8,653
Rb
0,70
0,391
1,54
0,585
1,23
0,198
0,93
0,333
0,29
0,130
Cd
1,24
2,450
0,86
1,700
1,59
1,444
3,43
2,296
0,03
0,020
Sn
< 0,10
< 0,10
0,08
0,155
0,08
0,109
< 0,10
< 0,10
< 0,10
< 0,10
Sb
< 0,01
< 0,01
< 0,01
< 0,01
< 0,01
< 0,01
< 0,01
< 0,01
< 0,01
< 0,01
Hg
4,07
3,510
40,32
67,089
14,80
57,720
8,59
5,145
2,65
0,243
Pb
< 0,01
< 0,01
0,004
0,009
0,004
0,005
< 0,01
< 0,01
< 0,01
< 0,01
Figure 5-12 Mean concentration (mg/kg ww) of elements in Striped dolphin (Stenella
coreuleoalba) by tissue.
pag. 42/73
Table 5-12 Concentration of elements (mg/kg ww) in Risso’s dolphin (Grampus griseus)
by tissue (number of available samples = 1).
Element
Skin-blubber
Muscle
Liver
Kidney
Melon
Cr
0,25
0,05
0,01
0,01
0,09
Ni
< 0,02
0,95
3,16
1,38
< 0,02
Cu
0,27
1,11
2,49
1,77
0,08
Zn
71,29
12,15
16,75
16,17
2,13
As
0,67
0,90
2,02
1,01
0,14
Se
15,28
13,45
165,37
8,80
1,48
Rb
0,53
2,22
1,73
1,67
0,05
Cd
0,03
0,05
4,94
6,83
0,06
Sn
< 0,10
< 0,10
< 0,10
< 0,10
< 0,10
Sb
< 0,01
< 0,01
< 0,01
< 0,01
< 0,01
Hg
4,20
42,36
562,20
17,86
3,00
Pb
< 0,01
< 0,01
0,11
0,03
0,01
Figure 5-13 Concentration of elements (mg/kg ww) in Risso’s dolphin (Grampus
griseus) by tissue.
pag. 43/73
Table 5-13 Concentration of elements (mg/kg ww) in Sperm whale (Physeter
macrocephalus) by tissue (number of available samples = 1).
Element
Skin-blubber
Muscle
Liver
Kidney
Spermaceti
Cr
0,14
0,08
0,01
0,05
<0,01
Ni
0,14
0,43
1,76
0,81
0,13
Cu
0,42
0,40
3,42
1,21
0,15
Zn
28,59
57,77
23,51
11,06
5,70
As
0,37
0,89
0,63
0,76
0,25
Se
4,12
1,31
25,38
2,62
0,40
Rb
0,43
1,42
0,73
0,70
0,23
Cd
0,01
0,01
0,77
2,33
0,02
Sn
<0,01
<0,01
0,23
<0,01
<0,01
Sb
0,01
<0,01
<0,01
<0,01
<0,01
Hg
0,99
4,69
95,01
5,14
0,69
Pb
<0,01
<0,01
0,01
<0,01
<0,01
Figure 5-14 Concentration of elements (mg/kg ww) in Sperm whale (Physeter
macrocephalus) by tissue.
pag. 44/73
Table 5-14 Concentration (mg/kg ww) of the main/most heavy metals in kidney and liver
per species.
Kidney
Element
Liver
Sperm Bottlenose Striped Risso's Sperm Bottlenose Striped Risso's
whale
dolphin
dolphin dolphin whale
dolphin
dolphin dolphin
Cr
0,05
0,01
0,14
0,01
0,01
0,01
0,16
0,01
Cd
2,33
1,05
3,43
6,83
0,77
0,32
1,59
4,94
Hg
5,14
10,89
8,59
17,86
95,01
226,91
14,80
562,20
Pb
<0,01
0,01
< 0,01
0,03
0,01
0,04
0,004
< 0,01
pag. 45/73
Figure 5-15 Concentration of heavy metals in liver and kidney per species.
pag. 46/73
Figure 5-16 Concentration of
melon/spermaceti per species.
heavy
metals
in
skin-blubber,
muscle
and
pag. 47/73
As shown in
Table 5-15 and Table 5-17, the results obtained in the present study substantially agree with
data reported by other studies on cetaceans stranded on the Adriatic Sea coastline (statistical
analysis (T Test) did not highlight significant statistical difference among them), with mercury
concentrations often below the levels found by other Authors. It is not possible to establish if
these concentrations may be related to cetacean pathologies, since no data concerning the
concentrations of heavy metals in healthy animals are available.
Table 5-15 Concentration (mg/kg, w/w) of metals in Tursiops truncatus tissues compared with literature data for the Adriatic Sea.
Element
Present Study
Storelli et al., 2000
Bilandzic et al., 2012
Muscle
n = 10
-
n = 14
Cr
0,15
-
-
Cu
0,17
-
-
Zn
24,89
-
-
As
0,54
-
0,26
Se
1,14
-
-
Cd
< 0,01
-
0,01
Hg
2,39
-
28,6
Pb
0,02
-
0,02
n=3
n = 14
Liver
n=9
Cr
0,01
Cu
11,14
36,09
-
Zn
44,17
59,95
-
As
0,3
Se
39,47
Cd
0,32
Hg
226,9
Pb
0,04
-
1,36
45,49
0,63
135,52
450
0,14
Kidney
n = 10
Cr
0,01
Cu
3,12
n=3
n = 14
-
5,01
-
pag. 48/73
Element
Present Study
Storelli et al., 2000
Bilandzic et al., 2012
Muscle
Zn
16,34
As
0,33
Se
3,26
Cd
1,05
Hg
10,89
Pb
0,01
28,47
0,63
5,78
2,85
12,47
31,4
0,21
Table 5-16 Concentration (mg/kg, w/w) of metals in Grampus griseus tissues compared
with literature data for the Adriatic Sea.
Element
Present Study
Storelli et al., 1999
Bilandzic et al., 2012
Muscle
n=1
n=2
n=4
Cr
0,05
0,675
-
Cu
1,11
-
-
Zn
12,15
-
-
As
0,9
-
1,21
Se
13,45
11,87
-
Cd
0,05
0,15
0,09
Hg
42,36
28,695
42,7
Pb
< 0,01
0,05
0,01
n=1
n=2
n=4
Cr
0,01
0,345
-
Cu
2,49
-
-
Zn
16,75
-
-
As
2,02
-
2,41
Se
165,4
189,8
-
Cd
4,94
7,21
5,16
Hg
562,2
740,15
1115
Pb
0,11
0,13
0,63
Liver
Kidney
pag. 49/73
Element
Present Study
Storelli et al., 1999
Bilandzic et al., 2012
Muscle
n=1
n=2
n=4
Cr
0,01
0,29
-
Cu
1,77
-
-
Zn
16,17
-
-
As
1,01
-
1,79
Se
8,8
16,515
-
Cd
6,83
11,61
14,9
Hg
17,86
54,39
33,1
Pb
0,03
0,11
0,1
Table 5-17 Concentration (mg/kg, w/w) of metals in Stenella coreuleoalba tissues
compared with literature data for the Adriatic Sea.
Element
Present Study
Cardellicchio et al., 2002 (a, b)
Bilandzic et al., 2012
Skin-Blubber
n=4
n = 10
-
Cr
0,36
0,88
-
Cu
1,17
1,21
-
Zn
46,37
17,03
-
As
0,78
-
-
Se
7,42
2,61
-
Cd
1,24
0,11
-
Hg
4,07
0,78
-
Pb
< 0,01
0,15
-
Muscle
n=4
n = 10
n=5
Cr
0,02
0,31
-
Cu
2,49
1,78
-
Zn
18,19
12,88
-
As
0,46
-
0,33
Se
8,56
4,43
-
Cd
0,86
0,04
0,02
pag. 50/73
Element
Present Study
Cardellicchio et al., 2002 (a, b)
Bilandzic et al., 2012
Skin-Blubber
Hg
40,32
8,61
16,50
Pb
0,02
0,12
0,01
Liver
n=4
n = 10
n=5
Cr
0,31
0,15
-
Cu
4,86
9,99
-
Zn
20,57
55,22
-
As
0,75
-
1,87
Se
19,45
63,18
-
Cd
1,60
1,50
2,19
Hg
87,65
170,76
182,00
Pb
0,01
0,22
0,07
Kidney
n=4
n = 10
n=5
Cr
0,14
0,23
-
Cu
2,01
4,06
-
Zn
34,44
27,71
-
As
0,75
-
0,66
Se
6,92
7,68
-
Cd
3,43
6,35
6,10
Hg
8,59
8,99
12,60
Pb
< 0,01
0,07
0,04
Melon
n=2
n = 10
-
Cr
0,01
0,34
-
Cu
0,18
1,07
-
Zn
25,47
8,33
-
As
0,36
-
-
Se
7,25
0,16
-
Cd
0,03
0,06
-
Hg
2,65
0,22
-
Pb
< 0,01
0,16
-
pag. 51/73
5.6
Organic contaminants data elaboration
Due to properties such as high fat solubility, high chemical stability and low volatility, Organochlorine Compounds are subject to biomagnification in the marine food chain. Organisms
at the top of the food chain, such as fish-eating cetaceans, suffer severe consequences. The
thick layer of blubber protecting cetaceans, the only warm-blooded animals that live their entire life in the sea, is an important energy reserve for them but also makes them the target of
fat soluble and apolar contaminants. As a result, their concentrations in this compartment are
very high. Cetaceans are thus excellent accumulators of persistent environmental contaminants but have a poor detoxifying potential. This makes them very sensitive to toxic effects,
increasing their vulnerability to various pathogens.
PAHs are toxic compounds which have attracted scientific interest due to their genotoxicity.
Their lipophilic nature enable them to cross biological membranes and accumulate in organisms. They are released into the environment by natural (pyrolysis etc.) and man-made processes (industrial processes, combustion of wood and fossil fuels, motor vehicles, incinerators,
oil plants and refineries, oil spills). These molecules are highly photosensitive and thermolabile, so in the presence of light and oxygen they are quickly degraded. As evident in Table II
included in the Appendix, most of PAHs resulted below their corresponding Limits of Quantitation in all the analysed samples. So, data were not graphically reported. These results were
not compared with those of the few data published in literature about PAH levels in Mediterranean cetaceans (Marsili et al, 2001), since these were obtained from samples collected
from live specimens, and not from stranded animals for which sampling was done after an undefined time from their death.
Table II reported in the Appendix to this report shows Organochlorine compounds (OCs) results obtained for those stranded specimens used in the present study for which the tissue
was enough to perform the analytical determination. The total DDT (Σ DDT) was calculated as
the sum of all the six DDT compounds. The Table also shows the values of the ratios 4,4’DDE/Σ DDT. Since 4,4’-DDE is the main metabolite of DDT, this ratio is useful since it may reflect the efficiency of metabolic processes in a particular species. It is also used to estimate
the time and intensity of exposure to DDT. The level of 4,4’-DDE generally increases with the
interval since DDT exposure began and with the quantity to which the animal was exposed.
Mean concentrations of OCs in each tissue of different species are summarised in Table 5-18
and graphically visualized in the corresponding Figure 5-17.
pag. 52/73
Table 5-18 Mean concentration of organochlorine compounds (mg/kg, ww).
Physeter macrocephalus
(sperm whale)
Physeter macrocephalus
(sperm whale)
Tursiops
truncatus
(bottlenose
dolphin)
Tursiops
truncatus
(bottlenose
dolphin)
Tursiops truncatus (bottlenose dolphin)
Tursiops
truncatus
(bottlenose
dolphin)
Stenella coeruleoalba
(striped dolphin)
Tissue
kidney
spermaceti
kidney
liver
melon
skin-blubber
skin-blubber
n
1
1
2
1
3
7
3
HCB
0,0037
0,0192
0,0062
0,0005
0,0432
0,0443
0,0174
2,4’-DDE
0,0126
0,0913
0,0177
0,0059
0,0761
0,0875
0,0831
4,4’-DDE
0,8856
6,2046
2,4432
0,6639
13,4222
15,8132
8,8805
2,4’-DDD
0,0104
0,0382
0,0176
0,0072
0,1674
0,1320
0,0671
4,4’-DDD
0,0309
0,1761
0,1181
0,0170
0,5199
0,9082
0,2798
2,4’-DDT
0,0194
0,2889
0,0284
0,0070
0,1016
0,2795
0,1980
4,4’-DDT
0,0245
0,5061
0,0365
0,0034
0,0797
0,2856
0,2228
∑ DDT
0,9833
7,3051
2,6615
0,7043
14,3668
17,5060
9,7313
4,4’-DDE/ ∑ DDT
0,8493
0,8885
0,9006
0,9304
0,9097
0,9426
0,8523
Figure 5-17 Mean concentration of organochlorine compounds (mg/kg, ww).
pag. 53/73
It was not possible to evaluate the results of the different tissues according to sex, since only
three on a total of seventeen specimens were female, and only one of these female samples
was a Tursiops truncatus, representing the most of the analysed samples.
As expected, the two tissues with the highest levels of OCs were melon and skin-blubber
(Figure 5-18). In fact, the level of these highly lipophylic contaminants, is known to be related
to the lipid content of the various tissues.
Σ DDT values for skin-blubber in T. truncatus (n=7) resulted in the concentration range 3.1 –
43 mg/kg on wet weight (see Table II in the Appendix I) with a medium value of 17.5 mg/kg
ww (Table 5-18).
Data obtained in this study were not compared directly with those reported in the overview of
Storelli et al (1997) about cetaceans stranded along the Italian coasts in the period 1987-1993,
since the data in the cited reference are reported on dry weight basis and not wet weight as for
our results. Nevertheless, Shoham-Frider et al (2009) converted data of Storelli (1997) to wet
weight basis obtaining in blubber of T. truncatus (n=8, the great part of them stranded along
Tyrrhenian coasts) a mean Σ DDT value of 6.03 mg/kg and 0.67 for the ratio 4,4’-DDE/Σ DDT.
The same authors analysed DDTs levels in blubber of T. truncatus (n=6) found along the Israeli Mediterranean coast and reported a range of concentration of 0.92 – 142 mg/kg on wet
weight and the values of 0.85-0.96 for the ratio 4,4’-DDE/Σ DDT.
Figure 5-18 Mean concentration of organochlorine compounds (mg/kg, ww) in Tursiops
truncatus (bottlenose dolphin) tissues.
pag. 54/73
Figure 5-19 and Figure 5-20 show the distribution of the various components of DDTs in skinblubber of Tursiops truncatus and Stenella coeruleoalba, respectively. The major constituent
of DDTs was 4,4’-DDE in all tissues, as confirmed by most studies of organochlorine compounds in dolphins and other cetacean species (O’Shea and Tanabe, 2003). It accounted for
at least 80% of Σ DDT in all the examined matrices. The relatively high fraction of 4,4’-DDE
indicates the metabolic capacity for conversion of DDTs to DDE and the absence of metabolic
enzymes to detoxify 4,4’-DDE, as has been shown in cetaceans in general (Colborn and Smolen, 2003).
As shown in Table 5-18, the ratio 4,4’-DDE/Σ DDT in the analysed sample was in the range
0,85 – 0,94. These values fit the general trend of increase in the last 20 years in the Mediterranean sea, indicating the progressive degradation of the remnant DDT and the absence of
new inputs (Aguilar and Borrell, 2005).
Figure 5-19 Percentage composition of DDTs (DDT and metabolites) in Tursiops
truncatus (bottlenose dolphin) skin-blubber tissues.
pag. 55/73
Figure 5-20 Percentage composition of DDTs (DDT and metabolites) in Stenella
coeruleoalba (striped dolphin) skin-blubber tissues.
pag. 56/73
5.7
Biomarkers data elaboration
5.7.1
8-OHdG
The obtained results (expressed in weight percentage of the DNA oxidation product 8-OhdG
on the total DNA analysed) range from the total absence of oxidation products (no. 14 samples), to a minimum of 0.00014% and a maximum of 0.0173% (Table 5-19).
Scientific literature studies use different units of measurement to express the 8-OhdG concentrations in animal tissues, due to different analytical techniques employed. Therefore, to make
a comparison it is necessary to convert data in the same unit of measurement. It follows that it
is useful to convert the number of adducts for dG nucleotide in number of adducts for total nucleotides.
We did it in two steps:
1. The molecular weight of the oxidation product and the different nucleotides were approximated to their average weight (according to scientific practices), useful for converting "percent by weight" in "number per nucleotide".
2. The frequency of deoxyguanosine (dG) per total nucleotides in T. truncatus was identified at 20% (Zhou X. et al, 2013) and we extended this value to the other species of
cetaceans. There is a high homology between cetaceans chromosomal sequences
and the human ones, in which the frequency of GC base pairs is 41%, as showed in
the Figure 5-21.
The table below (23) shows the levels of 8-OHdG in our study in the two different units of
6
measurement ‘8-OHdG % in weight’ and ‘number of 8-OHdG/10 nucleotides’.
Table 5-19 8-OHdG contents (% in weight) in the analysed samples.
6
ID specimen
Species
Sample
8-OHdG % in weight
8-OHdG (No/10
nucleotides)
mean
S.D.
mean
S.D.
ID166
T. truncatus
liver
0,01733
0,00271
173,25
27,08
ID163
T. truncatus
liver
0,01532
0,00392
153,2
39,17
ID165
T. truncatus
liver
0,01403
0,00317
140,25
31,75
ID185
T. truncatus
liver
0,01403
0,00125
140,25
12,52
ID192
T. truncatus
liver
0,01320
0,00052
132
5,23
ID214
S. coeruleoalba
liver
0,00407
0,00005
40,65
0,49
ID196
T. truncatus
liver
0,00342
0,00042
34,15
4,17
ID203
T. truncatus
kidney
0,00296
0,00109
29,6
10,89
pag. 57/73
6
ID specimen
Species
Sample
8-OHdG % in weight
8-OHdG (No/10
nucleotides)
mean
S.D.
mean
S.D.
ID172
P. macrocephalus
kidney
0,00285
0,00090
28,25
8,98
ID214
S. coeruleoalba
kidney
0,00073
0,00034
7,3
3,39
ID218
S. coeruleoalba
liver
0,00157
0,00069
15,65
6,86
ID162
T. truncatus
liver
0,00062
0,00021
6,15
2,05
ID180
T. truncatus
liver
0,00048
0,00066
4,75
6,58
ID193
T. truncatus
kidney
0,00014
0,00021
1,40
2,12
ID220
S. coeruleoalba
liver
0,00138
0,00070
13,75
7,00
ID196
T. truncatus
kidney
absent
/
absent
/
ID215
G. griseus
kidney
absent
/
absent
/
ID162
T. truncatus
kidney
absent
/
absent
/
ID163
T. truncatus
kidney
absent
/
absent
/
ID164
T. truncatus
liver
absent
/
absent
/
ID164
T. truncatus
kidney
absent
/
absent
/
ID165
T. truncatus
kidney
absent
/
absent
/
ID180
T. truncatus
kidney
absent
/
absent
/
ID185
T. truncatus
kidney
absent
/
absent
/
ID192
T. truncatus
kidney
absent
/
absent
/
ID218
S. coeruleoalba
kidney
absent
/
absent
/
ID220
S. coeruleoalba
kidney
absent
/
absent
/
ID221
S. coeruleoalba
liver
absent
/
absent
/
ID221
S. coeruleoalba
kidney
absent
/
absent
/
pag. 58/73
Figure 5-21 Percentage of guanine-cytosine pairs in the DNA of various species (from:
Zhou X. et al, 2013).
Overall, liver samples have a greater number of oxidation products compared to kidney ones,
in which they are often absent (no. 11 samples); in fact, the liver appears the most susceptible
to induction of 8-OHdG, in agreement with literature data.
In particular, no. 5 specimens belonging to Tursiops truncatus species exhibit the greater percentages of 8OH-dG; those with intermediate values belonging to Stenella coeruleoalba in
three cases and to T. truncatus in one case; finally, two T. truncatus specimens showed the
lower percentages. One specimen of T. truncatus and one of S. coeruleoalba didn’t present
oxidation products.
For kidney samples, a specimen of Physeter macrocephalus and one of T. truncatus showed
the greater levels of biomarkers.
The species T. truncatus seems to present the greatest variability in the appearance of oxidation products, and at the same time to show the highest level, while S. coeruleoalba is characterized by intermediate values. However, the variability present in T. truncatus is probably attributable to the greater number of samples analysed in relation to S. coeruleoalba.
Grampeus griseus presents the highest concentration for 5 metals (Zn, As, Se, Cd, Hg) compared with the remaining species, not justifying the anomalous peak of the species T. truncatus.
Looking for the reasons of the results, it should be taken into account the heavy metals concentrations in the corresponding tissues. T. truncatus specimens show in average higher values for 7 of the 12 analysed metals (Ni, Cu, Zn, Se, Sn, Sb, Hg) then others species and S.
coeruleoalba specimens for 3 metals (Cr, As, Cd); two elements (Pb, Rb) are similar, in
pag. 59/73
agreement with the biomarker concentration range. In addition, the bioaccumulation of metals
is greater in liver tissue than in the kidney one, except for Cd (lower) and As (in the same
range).
DNA oxidation products formation represents the extreme oxidative cell damage, which is responsible for some reactive compounds produced by several mechanisms, included the Fenton reaction, in presence of transition heavy metals and hydrogen peroxide [Novo and Word,
2008].
It was confirmed a positive correlation between the percentage of oxidation products with four
transition heavy metals: Fe (p= 0,0000008) and Mo (p=0,0000092), Ni (p= 0,0000012) and Ag
(P= P= 0,0000012). There is also a correlation between the biomarker and Pb (p=0,000035),
known for its cytotoxic properties in soft tissues and Hg (p=0,0021), heavy metal responsible
for inhibition of antioxidant molecules that contain sulphur, resulting in increased oxidative
stress [Houston MC, 2011].
Among the organic substances only the chlorinated fungicide hexachlorobenzene is positively
correlated with the biomarker in question. It is a probable carcinogen for humans and responsible for an increased incidence of tumors of the liver, kidneys, and thyroid in animals.
6
Li et al (2005) obtained a number of 90/10 nucleotides in cetaceans liver from Taiwan coast,
in the same range of our values. In both studies, it can be assumed the fundamental role of
the chemical contamination of sea water on the oxidative stress biomarkers levels.
Li et al (2005) highlighted a correlation between 8-OHdG and PCB concentrations, whilst no
correlation where found between 8-OHdG and the health status of specimens.
Other available studies regarding fish species subjected to chemical substances show 8OHdG values both lower (in terms of unit fractions) and higher (in the order of hundreds) then
our data [Ploch S.A. et al, 1999].
6
The values of 8-OHdG present in human lymphocytes range to 0.2-2/10 nucleotides [Valavanidis A. et al. 2009]. Therefore, we can assume that the results we obtained indicate an av6
erage high level of oxidative stress, with the presence of some peaks range from 132/10 and
6
173.25 /10 nucleotides.
DNA oxidation products formation represents the extreme oxidative cell damage, which is responsible for some reactive compounds produced by several mechanisms, included the Fenton reaction, in presence of transition heavy metals and hydrogen peroxide [Novo and Word,
2008].
5.7.2
Micronuclei assay
It was carried out the micronuclei (MN) analysis in different tissues (liver, kidney, skin-blubber,
melon and muscle) preserved in formalin of three specimens belonging to the following species: G. griseus, T. truncatus and S. coeruleoalba (Figure 5-22).
The MN percentage values for total cells analysed ranged in the order of unit, except for a sin-
pag. 60/73
gle value, much higher than the other (the skin-blubber belonging to T. truncatus; Table 5-20
and Figure 5-23).
A.
B.
Figure 5-22A Slides of liver tissues adapted for the counting of Micronuclei (MN): A. - T.
truncatus; B. - S. coreuleoalba (the white arrows indicate some of the counted MN);
photo by: Lorena Salvatore.
pag. 61/73
Figure 5-23B Slides of liver tissues adapted for the counting of Micronuclei (MN): G.
griseus (the white arrows indicate some of the counted MN); photo by: Lorena
Salvatore.
Table 5-20 Mean MN percentage per tissue and species.
ID
Species
Tissue
% MN
203
T. truncatus
Liver
7,8
203
T. truncatus
Skin-blubber
15,2
215
G. griseus
Liver
8,4
215
G. griseus
Skin-blubber
6
215
G. griseus
Kidney
6,2
215
G. griseus
Muscle
3,5
215
G. griseus
Melon
2,6
218
S. coeruleoalba
Liver
6,2
218
S. coeruleoalba
Skin-blubber
5,6
218
S. coeruleoalba
Kidney
5,8
218
S. coeruleoalba
Muscle
8,2
218
S. coeruleoalba
Melon
7,3
pag. 62/73
Figure 5-24 % of MN in different tissues per species.
The higher percentages of MN were obtained in descending order for: skin-blubber, liver, kidney and muscle, melon. Samples from T. truncatus have higher concentrations of MN than S.
coeruleoalba in three (liver, skin-blubber, kidney) of the five organs investigated. Grampeus
griseus shows a higher level for 5 metals (Zn, As, Se, Cd, Hg) compared with the other species.
It was not observed a corresponding change between metals concentration and MN, which
therefore seems to be a cellular response to other factors.
The observed MN values can be compared with those found in fish specimens exposed to an
organophosphorus insecticide; the percentage of genotoxic damage detected in lymphocytes
of the fish are comparable with that we found in cetaceans tissues [Sepici-Dincel A. et al,
2011]. From an artificial exposure to an environmental pollutant a high cellular response is expected; therefore, our results on micronuclei frequency show a significant genotoxic damage.
5.8
The listening station at sea
The executive project of a listening station at sea and a monitoring system of the anthropic
noise impact on cetaceans’ population in the Adriatic sea is included as attached document to
this report (see Annex).
pag. 63/73
6
Implication for MSP and ICZM
6.1
Implication for MPS
Adriatic cetaceans population have been poorly studied, both for ecological and ecotoxicological aspects. The project could be a contribute to a deepen knowledge on their ecology, to verify the role of Adriatic geographic area for this species and compare it with other
marine areas.
In fact, one of the principal causes of Adriatic disappearance of common dolphins is the habitat contamination. Understand the eco-toxicological state of the Adriatic populations may represent a starting point to plan safeguard action plans of the local population, also by the creation and management of specific marine Sites of Community Importance (SCI), still in very limited numbers in the Adriatic basin and by the creation and safeguard of biological corridors
along the routes (maritime route of global significance and migration corridors) usually followed by the Adriatic population. These areas should be planned in a broader context of marine spatial planning strategies.
6.2
Implication for ICZM
Our pilot subproject aimed at defining the state of the art of the contamination phenomena of
the Adriatic cetaceans; therefore, it can be assimilated, in general terms, to an environmental
monitoring.
The adoption of a holistic strategic approach in the coastal areas planning and management,
must takes also into account indirect and cumulative causes and effects. One of these effect is
the biomagnification phenomenon at the top of food chain, caused by the discharge into the
sea and by the dispersion of pollutants coming from the terrestrial environment, with all its
consequences for the health status of marine populations.
The implementation of this biomonitoring system could lead to the creation of a Adriatic network for monitoring and surveillance of the ecological status of the Adriatic food chain.
pag. 64/73
6.3
Implication for MSFD
The results obtained will be useful to contribute to the determination of the Adriatic environmental status by:
•
Improving the organization of the competent authorities of stranding phenomena
monitoring and indirectly assessing the ecological status of Adriatic marine mammals’ population (descriptors no. 1 and 4 of MSFD).
•
The detection of eco-toxicological parameters in cetaceans specimens, indicating
possible environmental contamination (descriptor no. 8 and 9 of MSFD).
•
The detection of the eco-toxicological stress to which cetacean populations are subjected as a result of anthropogenic pollution and the risk for their long-term abundance and the retention of their full reproductive capacity (descriptors no. 4).
•
The realization of the hydroacoustic station, which will allow the project to lay the
foundation for the anthropogenic noise assessment of Adriatic environment (descriptor no. 11) and the monitoring of the noise effects on marine mammals.
6.4
Adriatic added valued of the project
The project could give a contribution and highlight the following points:
•
Provision of tools of global significance in the ecological management of the interest
species.
•
Provision of field studies concerning Italian seas. They have been concentrated on
the Tirrenic side.
•
The project will allow to enlarge studies to the Adriatic side, to deepen and integrate
at the same time ecological, bio-acoustical and biochemical-toxicological researches
(stranding phenomenon, bioaccumulation and bio-acoustical studies) within a single
research group, which get all the required competences and professional skills for
every treated field.
•
Collected and processed data and analytic results will merge into a digital database/archive, which will feed the first GIS collecting this type of information for the
Adriatic basin. The GIS will permit the application of standard and spatial multiparametric statistical methodologies, by means of several overlapped georeferenced informative layers (thematic field/parameters). That will allow to underline
spatial correlation among different origins variables (physical, biological and anthropic) and emphasize possible relations at different scales among phenomena (of
cause-effect too), otherwise not identifiable.
pag. 65/73
•
Increasing the sparse database of contaminant data available for cetacean species
and allowing the comparison of contaminants concentrations in various tissues of cetaceans species from Adriatic with those found in other populations from the Mediterranean or non-Mediterranean area.
•
Understanding causes which have harmed to the rarefaction of cetaceans species in
Adriatic sea and giving a comparison with the status of Mediterranean populations
and assessing the different cetaceans species vulnerability to waters contamination.
•
Better use and interpretation of environmental data (number and placement of
stranding, eco-toxicological measures by bioaccumulation studies), with a view to increase their knowledge on the Adriatic basin scale, based on best practices present
on the Italian territory;
•
Production of new knowledge in the environmental monitoring and conservation of
nature fields, in terms of possible implementation of the Natura 2000 Network in the
Adriatic environment.
The project will grant in the future the construction of a theoretical model to estimate the Potential Hazard for some Adriatic species, in relation to the three categories of principal pollutants (PAHs, OCs and heavy metals). In fact, at now only models for striped dolphin in the Tirrenic sea have been structured. A task for future research will be to obtain models speciesspecific valid for each species in general, no matter where they live (in Tirrenic, Adriatic or
other seas).
6.5
Innovative aspects of the projects
Valuating the spatial and temporal distribution of Adriatic pollutants concentration by means of
biomonitoring,
Further innovative aim of the project is to acquire a different key for interpreting data on OCs,
PAHs and heavy metals accumulation in tissue (especially blubber) with applied statistics. The
information that can be obtained from this multidisciplinary approach is a theoretical evaluation
of stress caused to Adriatic cetaceans by the exposure to certain types of xenobiotics.
In a tightly analytical point of view, carrying out chemical and biological analysis on tissues deriving from stranding dolphins, that will offer the great opportunity of debug specific methodologies to obtain experimental data on species classified “at risk of extinction”. The physiological and cells damages picked out by biomarkers could be rendered in reference to
effective pollution impact on cetaceans’ health.
pag. 66/73
7
Conclusions
The focal points emerged from the project are the following:
•
Stranding events along the west Adriatic coast are homogeneous in time, space and
in biological terms (species, dimension, specimens etc.).
•
They appear quite regularly over the years, except for 1991, particularly in the Central Adriatic, probably due to the measles outbreak that attacked marine mammals
populations, particularly dolphins. The time period analysed (1982-2013) is too short
to identify the existence of a cyclical nature of the phenomenon, as shown instead in
other contexts (Evans et al., 2005). It is noticed an increase in the number of strandings during the summer months and there is a peak that moves along a north-south
gradient: in the northern regions in July, in the central regions between July and August and in the southern regions in August-September. There is also a difference in
the frequency peak of strandings among species during the summer period. Statistical analysis didn’t highlight the existence of a different probability to strand among
species, between males and females of the total species and per species, and any
difference in body dimension by species.
•
Our results confirm the few literature data regarding the rather high level of inorganic
and organic contaminants present in different tissues of the Adriatic cetaceans and
the values of metal concentrations we have found do not differ significantly from
those already published.
•
High concentrations of trace metals, principally Hg, Cd, Zn, Fe and Se, were found in
all analysed tissues, with a relation (not significant statistically) between the kind of
tissues and the different elements.
•
Most of PAHs resulted below the corresponding limits of quantitation in all the samples. Organochlorine compounds showed the highest concentration in skin-blubber
and melon tissues of all the analysed specimens, confirming their high affinity for lipophylic matrices. The distribution analysis of the various DDT compounds shows
4,4’-DDE as the major constituent of the group, accounting for at least 80% of the
components sum.
•
The biomarkers results indicate a consistent exposition to genotoxic substances, as
established by literature studies on different aquatic species. Our results on 8-OHdG
levels indicate a high level of oxidative stress. The statistical analysis showed a correlation between the percentage of 8-OHdG and some metals, such as Fe, Mo, Pb,
Hg, and an organic compound, hexachlorobenzene.
•
Liver is most liable to both biological damages (MN and adducts) according to toxicological literature data.
pag. 67/73
Unfortunately, a several lack of information in the available national databases didn’t allow to
relate the stranding events to eco-toxicological data. Reasons are the following:
•
It was not possible to correlate the Adriatic stranding events from CIBRA database
with the BTMM one, except for a few cases of which, however, the biological material was not available for the analysis. Further, in CIBRA database the cause of
death of stranding specimens is almost never reported, as a result of the advanced
state of decomposition in which the specimens are found at the time of reporting.
•
it was not possible to verify any relation between pollutants concentrations with the
body length or the animal age, because of the lack of this information in our data collected. BTMM indicates a body length and weight of the specimens at the biopsy
moment, often when the animal was in an advanced decomposition stage.
•
Considered the few samples available for the analysis, it was not possible to value
the existence of any variation of organic and inorganic pollutants concentration
among the species examined, the populations of the three regions (Northern Central
and Southern Adriatic) and among the same region (taking into consideration the
length, the sex, and the age of the specimens).
•
It is not possible to point out a direct correlation among the different groups of contaminants in the analysed samples and the diseases / causes of death of specimens
to which the samples belong. In any case, there are no literature data able to link the
contaminants and the diseases of cetaceans, except for extremely high values (but
extremely rare).
Conclusive findings of the pilot project are summarised through the 4-pillars matrix common to
all pilot projects. The matrix intends to highlights: (i) main outcome and deliverables of the project, (ii) improved skills, (iii) possible future uses of the project outcome, (iv) future opportunities and conflicts related to the evolution of pilot project contents in an MSP perspective.
These are also discussed in the following text.
pag. 68/73
Output
What we have done
A complete database on strandings events
occurred in the western Adriatic coast from
1902
How can we use output in the future?
Analysis of inorganic and organic pollutants
accumulated in different tissues of Adriatic
cetaceans specimens
Replicate the biomonitoring analysis to future
data
Creating a GIS archive on strandings events
and tissue bank of Adriatic cetaceans
An executive project of a listening station at sea
and a monitoring system of the antropic noise
impact on cetaceans population in the Adriatic
sea
Using the results for the Potential Hazard
determination of single species
Opportunities
Involvement of other actors and entity to
constitute a biomonitoring network for the
Adriatic sea
What we have learned/Skills improved
Creation of a work ing group with specific sk ills
on bioaccumulation and matabolic stress
consequences in the Adriatic cetaceans
A more in-depth k nowledge in technical and
regulation terms on European strategies for
planning in coastal areas
Looking Forward
Capitalization
First evaluation on Adriatic sea of cetaceans
stress caused by exposure to water
contaminants by biomak er analysis
the realization of the project on “listening station
at sea and a monitoring system of the antropic
noise impact” could fill the gap that is present
to a Adriatic basin scale about the marine noise
pollution currently significantly undervalued
Criticalities
Lack of information and samples coming from
national database/tissue bank
Skills
Figure 7-1 Conclusive findings of the pilot project.
pag. 69/73
8
References
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to 2002, as shown by levels in striped dolphins (Stenella coeruleoalba). Mar. Environ. Res.,
59(4):391-404.
André J., Boudou A., Ribeyre F., Bernhard M. (1991). Comparative study of mercury accumulation in dolphins (Stenella coeruleoalba) from French Atlantic and Mediterranean coasts. Sci.
Total Environ., 104; pp. 191–209.
Augier, H., Park, W. K., & Ronneau, C. (1993). Mercury contamination of the striped dolphin
Stenella coeruleoalba Meyen from French Mediterranean coasts. Mar. Pollut. Bull., 26(6): 306311.
Bernhard M. (1988). Mercury in the Mediterranean, Regional Seas Reports and Studies, No
98.
Bilandžić N, Sedak M, Ðokić M, Ðuras Gomerčić M, Gomerčić T, Zadravec M, Benić M, Prevendar Crnić (2012). A Toxic Element Concentrations in the Bottlenose (Tursiops truncatus),
Striped (Stenella coeruleoalba) and Risso’s (Grampus griseus) Dolphins Stranded in Eastern
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Appendix Complete results of inorganic and organic compounds analysis
Appendix - 1
Table I – Complete results of elemental analysis of cetacean tissues by ICP-MS after sample mineralization by closed vessel microwave
acid digestion.
Sample Name
Sample ID
Tissue
Specie
Sample Weight (g)
Element / Atomic weight
LLoQ(2)
[analytical mode(1)]
Be / 9 [#1]
0,01
Al / 27 [#1]
0,10
Ti / 48 [#2]
0,05
V / 51 [#2]
0,05
Cr / 52 [#2]
0,01
Mn / 55 [#1]
0,50
Fe / 56 [#2]
0,50
Ni / 58 [#2]
0,02
Co / 59 [#1]
0,01
Cu / 63 [#2]
0,05
Zn / 66 [#2]
0,05
Ga / 69 [#1]
0,01
As / 75 [#2]
0,01
Se / 78 [#2]
0,05
Rb / 85 [#1]
0,01
Sr / 88 [#1]
0,05
Mo / 98 [#2]
0,05
Rh / 103 [#1]
0,01
Pd / 105 [#1]
0,02
Ag / 107 [#2]
0,01
Cd / 111 [#2]
0,01
SHAPE-01
(ID172)
CUTE-ADIPE
172
Skin-Blubber
Physeter macrocephalus
0,65
SHAPE-02
(ID172)
MUSCOLO
172
Muscle
Physeter macrocephalus
0,53
SHAPE-03
(ID172)
FEGATO
172
Liver
Physeter macrocephalus
0,50
< 0,01
10,85
0,94
< 0,05
0,14
1,48
26,38
0,14
0,01
0,42
28,59
0,01
0,37
4,12
0,43
1,21
< 0,05
< 0,01
< 0,02
< 0,01
0,01
< 0,01
8,92
1,48
0,06
0,08
3,48
126,13
0,43
0,02
0,40
57,77
0,01
0,89
1,31
1,42
0,98
< 0,05
< 0,01
< 0,02
< 0,01
0,01
< 0,01
2,60
0,30
< 0,05
0,01
0,66
540,20
1,76
0,06
3,42
23,51
< 0,01
0,63
25,38
0,73
0,36
0,45
< 0,01
< 0,02
0,61
0,77
SHAPE-04
SHAPE-05
(ID172)
(ID172)
RENE
SPERMACETE
172
172
Kidney
Spermaceti
Physeter macPhyseter macrocephalus
rocephalus
0,51
0,46
Concentration
(mg/kg)
< 0,01
< 0,01
0,13
3,41
0,20
0,51
< 0,05
< 0,05
0,05
< 0,01
< 0,50
1,14
245,98
34,12
0,81
0,13
0,02
0,01
1,21
0,15
11,06
5,70
< 0,01
< 0,01
0,76
0,25
2,62
0,40
0,70
0,23
0,29
0,46
< 0,05
< 0,05
< 0,01
< 0,01
< 0,02
< 0,02
0,01
< 0,01
2,33
0,02
SHAPE-06
(ID196)
CUTE-ADIPE
196
Skin-Blubber
Tursiops
truncatus
0,56
SHAPE-07
(ID196)
MUSCOLO
196
Muscle
Tursiops truncatus
0,51
SHAPE-08
(ID196)
FEGATO
196
Liver
Tursiops truncatus
0,58
< 0,01
< 0,10
0,21
< 0,05
0,01
< 0,50
5,71
0,03
< 0,01
0,51
36,20
< 0,01
0,41
1,48
0,31
0,18
< 0,05
< 0,01
< 0,02
< 0,01
< 0,01
< 0,01
< 0,10
0,19
< 0,05
0,04
< 0,50
149,80
0,49
< 0,01
0,78
14,59
< 0,01
0,08
0,61
0,85
0,14
< 0,05
< 0,01
< 0,02
< 0,01
< 0,01
< 0,01
< 0,10
0,15
0,06
< 0,01
2,18
836,98
2,67
< 0,01
2,41
13,77
< 0,01
0,13
9,70
0,89
0,12
0,43
< 0,01
< 0,02
0,38
0,03
Appendix - 2
Sample Name
Sample ID
Tissue
Specie
Sample Weight (g)
Element / Atomic weight
LLoQ(2)
[analytical mode(1)]
In / 115 [#2]
0,01
Sn / 118 [#2]
0,10
Sb / 121 [#2]
0,01
Te / 130 [#1]
0,01
Cs / 133 [#1]
0,01
Ir / 193 [#1]
0,01
Pt / 195 [#1]
0,01
Hg / 202 [#1]
0,01
Tl / 205 [#1]
0,01
Pb / 208 [#1]
0,01
Bi / 209 [#1]
0,01
U / 238 [#1]
0,01
(1) [#1] without ORS, [#2] with ORS;
SHAPE-01
(ID172)
CUTE-ADIPE
172
Skin-Blubber
Physeter macrocephalus
0,65
SHAPE-02
(ID172)
MUSCOLO
172
Muscle
Physeter macrocephalus
0,53
SHAPE-03
(ID172)
FEGATO
172
Liver
Physeter macrocephalus
0,50
< 0,01
< 0,10
0,01
< 0,01
< 0,01
< 0,01
< 0,01
0,99
< 0,01
< 0,01
< 0,01
< 0,01
< 0,01
< 0,10
< 0,01
< 0,01
< 0,01
< 0,01
< 0,01
4,69
< 0,01
< 0,01
< 0,01
< 0,01
< 0,01
0,23
< 0,01
< 0,01
< 0,01
< 0,01
< 0,01
95,01
< 0,01
0,01
< 0,01
< 0,01
SHAPE-04
SHAPE-05
(ID172)
(ID172)
RENE
SPERMACETE
172
172
Kidney
Spermaceti
Physeter macPhyseter macrocephalus
rocephalus
0,51
0,46
Concentration
(mg/kg)
< 0,01
< 0,01
< 0,10
< 0,10
< 0,01
< 0,01
< 0,01
< 0,01
< 0,01
< 0,01
< 0,01
< 0,01
< 0,01
< 0,01
5,14
0,69
< 0,01
< 0,01
< 0,01
< 0,01
< 0,01
< 0,01
< 0,01
< 0,01
SHAPE-06
(ID196)
CUTE-ADIPE
196
Skin-Blubber
Tursiops
truncatus
0,56
SHAPE-07
(ID196)
MUSCOLO
196
Muscle
Tursiops truncatus
0,51
SHAPE-08
(ID196)
FEGATO
196
Liver
Tursiops truncatus
0,58
< 0,01
< 0,10
< 0,01
< 0,01
< 0,01
< 0,01
< 0,01
1,71
< 0,01
< 0,01
< 0,01
< 0,01
< 0,01
< 0,10
< 0,01
< 0,01
0,02
< 0,01
< 0,01
3,11
< 0,01
< 0,01
< 0,01
< 0,01
< 0,01
0,12
< 0,01
0,02
0,01
< 0,01
< 0,01
36,98
< 0,01
0,01
0,01
< 0,01
(2) Lower Limit of Quantification
Appendix - 3
Sample Name
Sample ID
Tissue
Specie
Sample Weight (g)
Element / Atomic weight
LLoQ(2)
[analytical mode(1)]
Be / 9 [#1]
0,01
Al / 27 [#1]
0,10
Ti / 48 [#2]
0,05
V / 51 [#2]
0,05
Cr / 52 [#2]
0,01
Mn / 55 [#1]
0,50
Fe / 56 [#2]
0,50
Ni / 58 [#2]
0,02
Co / 59 [#1]
0,01
Cu / 63 [#2]
0,05
Zn / 66 [#2]
0,05
Ga / 69 [#1]
0,01
As / 75 [#2]
0,01
Se / 78 [#2]
0,05
Rb / 85 [#1]
0,01
Sr / 88 [#1]
0,05
Mo / 98 [#2]
0,05
Rh / 103 [#1]
0,01
Pd / 105 [#1]
0,02
Ag / 107 [#2]
0,01
Cd / 111 [#2]
0,01
In / 115 [#2]
0,01
Sn / 118 [#2]
0,10
Sb / 121 [#2]
0,01
SHAPE-09
(ID196)
RENE
196
Kidney
Tursiops truncatus
0,54
SHAPE-10
(ID196)
MELONE
196
Melon
Tursiops truncatus
0,53
SHAPE-11
(ID214)
CUTE-ADIPE
214
Skin-Blubber
Stenella coeruleoalba
0,61
< 0,01
< 0,10
0,36
< 0,05
< 0,01
< 0,50
131,48
0,44
0,02
1,79
11,11
< 0,01
0,11
1,66
0,90
0,27
< 0,05
< 0,01
< 0,02
0,01
0,20
< 0,01
< 0,10
< 0,01
< 0,01
0,47
0,32
< 0,05
0,02
< 0,50
23,00
0,11
0,01
0,32
3,67
< 0,01
0,24
0,98
0,09
0,26
< 0,05
< 0,01
< 0,02
< 0,01
0,01
< 0,01
0,13
< 0,01
< 0,01
2,69
0,99
< 0,05
0,27
< 0,50
11,75
0,07
0,01
0,50
60,24
< 0,01
0,77
10,69
0,85
1,15
< 0,05
< 0,01
< 0,02
< 0,01
0,02
< 0,01
< 0,10
< 0,01
SHAPE-12
SHAPE-13
(ID214)
(ID214)
MUSCOLO
FEGATO
214
214
Muscle
Liver
Stenella coeStenella coeruleoalba
ruleoalba
0,50
0,54
Concentration
(mg/kg)
< 0,01
< 0,01
0,83
< 0,10
0,21
0,13
< 0,05
0,08
0,02
0,45
< 0,50
3,54
291,20
334,17
0,95
1,09
0,01
0,01
1,22
6,36
9,40
35,23
< 0,01
< 0,01
0,36
0,64
2,00
24,25
2,39
1,36
0,10
0,12
< 0,05
0,63
< 0,01
< 0,01
< 0,02
< 0,02
< 0,01
0,61
0,01
1,35
< 0,01
< 0,01
< 0,10
0,23
< 0,01
< 0,01
SHAPE-14
(ID214)
RENE
214
Kidney
Stenella coeruleoalba
0,53
SHAPE-15
(ID214)
MELONE
214
Melon
Stenella coeruleoalba
0,56
SHAPE-16
(ID215)
CUTE-ADIPE
215
Skin-Blubber
Grampus griseus
0,58
< 0,01
< 0,10
0,67
< 0,05
< 0,01
0,56
284,62
0,91
0,02
3,19
22,28
< 0,01
0,29
3,58
1,30
0,30
< 0,05
< 0,01
< 0,02
0,02
4,74
< 0,01
< 0,10
< 0,01
< 0,01
0,18
0,10
< 0,05
< 0,01
< 0,50
12,58
0,06
0,01
0,18
3,82
< 0,01
0,16
1,13
0,19
0,11
< 0,05
< 0,01
< 0,02
< 0,01
0,04
< 0,01
< 0,10
< 0,01
< 0,01
1,35
3,04
< 0,05
0,25
< 0,50
8,80
< 0,02
0,01
0,27
71,29
< 0,01
0,67
15,28
0,53
0,29
< 0,05
< 0,01
< 0,02
< 0,01
0,03
< 0,01
< 0,10
< 0,01
Appendix - 4
Sample Name
Sample ID
Tissue
Specie
Sample Weight (g)
Element / Atomic weight
LLoQ(2)
[analytical mode(1)]
Te / 130 [#1]
0,01
Cs / 133 [#1]
0,01
Ir / 193 [#1]
0,01
Pt / 195 [#1]
0,01
Hg / 202 [#1]
0,01
Tl / 205 [#1]
0,01
Pb / 208 [#1]
0,01
Bi / 209 [#1]
0,01
U / 238 [#1]
0,01
(1) [#1] without ORS, [#2] with ORS;
SHAPE-09
(ID196)
RENE
196
Kidney
Tursiops truncatus
0,54
SHAPE-10
(ID196)
MELONE
196
Melon
Tursiops truncatus
0,53
SHAPE-11
(ID214)
CUTE-ADIPE
214
Skin-Blubber
Stenella coeruleoalba
0,61
< 0,01
0,01
< 0,01
< 0,01
4,35
< 0,01
< 0,01
0,01
< 0,01
< 0,01
< 0,01
< 0,01
< 0,01
1,74
< 0,01
< 0,01
< 0,01
< 0,01
< 0,01
0,01
< 0,01
< 0,01
3,39
< 0,01
< 0,01
< 0,01
< 0,01
SHAPE-12
SHAPE-13
(ID214)
(ID214)
MUSCOLO
FEGATO
214
214
Muscle
Liver
Stenella coeStenella coeruleoalba
ruleoalba
0,50
0,54
Concentration
(mg/kg)
< 0,01
< 0,01
0,05
0,02
< 0,01
< 0,01
< 0,01
< 0,01
8,56
107,04
< 0,01
< 0,01
< 0,01
0,01
< 0,01
< 0,01
< 0,01
< 0,01
SHAPE-14
(ID214)
RENE
214
Kidney
Stenella coeruleoalba
0,53
SHAPE-15
(ID214)
MELONE
214
Melon
Stenella coeruleoalba
0,56
SHAPE-16
(ID215)
CUTE-ADIPE
215
Skin-Blubber
Grampus griseus
0,58
< 0,01
0,02
< 0,01
< 0,01
11,11
< 0,01
0,01
< 0,01
< 0,01
< 0,01
< 0,01
< 0,01
< 0,01
2,82
< 0,01
< 0,01
< 0,01
< 0,01
< 0,01
< 0,01
< 0,01
< 0,01
4,20
< 0,01
< 0,01
< 0,01
< 0,01
(2) Lower Limit of Quantification
Appendix - 5
Sample Name
Sample ID
Tissue
Specie
Sample Weight (g)
Element / Atomic weight
LLoQ(2)
[analytical mode(1)]
Be / 9 [#1]
0,01
Al / 27 [#1]
0,10
Ti / 48 [#2]
0,05
V / 51 [#2]
0,05
Cr / 52 [#2]
0,01
Mn / 55 [#1]
0,50
Fe / 56 [#2]
0,50
Ni / 58 [#2]
0,02
Co / 59 [#1]
0,01
Cu / 63 [#2]
0,05
Zn / 66 [#2]
0,05
Ga / 69 [#1]
0,01
As / 75 [#2]
0,01
Se / 78 [#2]
0,05
Rb / 85 [#1]
0,01
Sr / 88 [#1]
0,05
Mo / 98 [#2]
0,05
Rh / 103 [#1]
0,01
Pd / 105 [#1]
0,02
Ag / 107 [#2]
0,01
Cd / 111 [#2]
0,01
In / 115 [#2]
0,01
Sn / 118 [#2]
0,10
Sb / 121 [#2]
0,01
Te / 130 [#1]
0,01
SHAPE-17
(ID215)
MUSCOLO
215
Muscle
Grampus griseus
0,53
SHAPE-18
(ID215)
FEGATO
215
Liver
Grampus griseus
0,63
SHAPE-19
(ID215)
RENE
215
Kidney
Grampus griseus
0,54
< 0,01
2,09
2,18
0,06
0,05
< 0,50
349,24
0,95
< 0,01
1,11
12,15
< 0,01
0,90
13,45
2,22
0,10
< 0,05
< 0,01
< 0,02
< 0,01
0,05
< 0,01
< 0,10
< 0,01
< 0,01
< 0,01
0,93
1,95
< 0,05
0,01
2,02
1080,03
3,16
0,01
2,49
16,75
< 0,01
2,02
165,37
1,73
0,26
0,35
< 0,01
< 0,02
1,70
4,94
< 0,01
< 0,10
< 0,01
< 0,01
< 0,01
0,45
2,28
< 0,05
0,01
0,79
541,54
1,38
0,01
1,77
16,17
< 0,01
1,01
8,80
1,67
0,32
< 0,05
< 0,01
< 0,02
0,03
6,83
< 0,01
< 0,10
< 0,01
< 0,01
SHAPE-20
SHAPE-21
(ID215)
(ID162)
MELONE
CUTE-ADIPE
215
162
Melon
Skin-Blubber
Grampus griTursiops trunseus
catus
0,59
0,54
Concentration
(mg/kg)
< 0,01
< 0,01
1,14
< 0,10
0,97
1,27
< 0,05
< 0,05
0,09
< 0,01
< 0,50
< 0,50
2,83
11,63
< 0,02
< 0,02
< 0,01
< 0,01
0,08
0,12
2,13
28,26
< 0,01
< 0,01
0,14
0,14
1,48
0,24
0,05
0,41
0,09
0,15
< 0,05
0,01
< 0,01
< 0,01
< 0,02
< 0,02
< 0,01
< 0,01
0,06
< 0,01
< 0,01
< 0,01
< 0,10
< 0,10
< 0,01
0,04
< 0,01
< 0,01
SHAPE-22
(ID162)
MUSCOLO
162
Muscle
Tursiops truncatus
0,48
SHAPE-23
(ID162)
FEGATO
162
Liver
Tursiops truncatus
0,53
SHAPE-24
(ID162)
RENE
162
Kidney
Tursiops truncatus
0,49
< 0,01
< 0,10
1,66
0,08
< 0,01
< 0,50
59,06
0,17
< 0,01
1,27
12,67
< 0,01
0,24
0,14
1,18
0,12
0,01
< 0,01
< 0,02
< 0,01
< 0,01
< 0,01
< 0,10
0,04
< 0,01
< 0,01
< 0,10
2,29
< 0,05
0,02
1,29
404,53
1,19
< 0,01
42,08
32,24
< 0,01
0,11
0,98
0,88
0,14
0,10
< 0,01
0,02
0,19
< 0,01
< 0,01
< 0,10
0,03
< 0,01
< 0,01
< 0,10
2,12
0,06
< 0,01
< 0,50
70,50
0,22
0,01
3,02
13,94
< 0,01
0,21
0,73
1,06
0,17
0,03
< 0,01
< 0,02
< 0,01
< 0,01
< 0,01
< 0,10
0,04
< 0,01
Appendix - 6
Sample Name
Sample ID
Tissue
Specie
Sample Weight (g)
Element / Atomic weight
LLoQ(2)
[analytical mode(1)]
Cs / 133 [#1]
0,01
Ir / 193 [#1]
0,01
Pt / 195 [#1]
0,01
Hg / 202 [#1]
0,01
Tl / 205 [#1]
0,01
Pb / 208 [#1]
0,01
Bi / 209 [#1]
0,01
U / 238 [#1]
0,01
(1) [#1] without ORS, [#2] with ORS;
SHAPE-17
(ID215)
MUSCOLO
215
Muscle
Grampus griseus
0,53
SHAPE-18
(ID215)
FEGATO
215
Liver
Grampus griseus
0,63
SHAPE-19
(ID215)
RENE
215
Kidney
Grampus griseus
0,54
0,02
< 0,01
< 0,01
42,36
< 0,01
< 0,01
< 0,01
< 0,01
< 0,01
< 0,01
< 0,01
562,20
< 0,01
0,11
< 0,01
< 0,01
0,01
< 0,01
< 0,01
17,86
< 0,01
0,03
< 0,01
< 0,01
SHAPE-20
SHAPE-21
(ID215)
(ID162)
MELONE
CUTE-ADIPE
215
162
Melon
Skin-Blubber
Grampus griTursiops trunseus
catus
0,59
0,54
Concentration
(mg/kg)
< 0,01
< 0,01
< 0,01
< 0,01
< 0,01
< 0,01
3,00
0,16
< 0,01
< 0,01
0,01
< 0,01
< 0,01
< 0,01
< 0,01
< 0,01
SHAPE-22
(ID162)
MUSCOLO
162
Muscle
Tursiops truncatus
0,48
SHAPE-23
(ID162)
FEGATO
162
Liver
Tursiops truncatus
0,53
SHAPE-24
(ID162)
RENE
162
Kidney
Tursiops truncatus
0,49
0,02
< 0,01
< 0,01
0,44
< 0,01
< 0,01
< 0,01
< 0,01
0,01
< 0,01
< 0,01
1,14
< 0,01
< 0,01
0,01
< 0,01
0,01
< 0,01
< 0,01
0,35
< 0,01
< 0,01
0,01
< 0,01
(2) Lower Limit of Quantification
Appendix - 7
Sample Name
Sample ID
Tissue
Specie
Sample Weight (g)
Element / Atomic weight
LLoQ(2)
[analytical mode(1)]
Be / 9 [#1]
0,01
Al / 27 [#1]
0,10
Ti / 48 [#2]
0,05
V / 51 [#2]
0,05
Cr / 52 [#2]
0,01
Mn / 55 [#1]
0,50
Fe / 56 [#2]
0,50
Ni / 58 [#2]
0,02
Co / 59 [#1]
0,01
Cu / 63 [#2]
0,05
Zn / 66 [#2]
0,05
Ga / 69 [#1]
0,01
As / 75 [#2]
0,01
Se / 78 [#2]
0,05
Rb / 85 [#1]
0,01
Sr / 88 [#1]
0,05
Mo / 98 [#2]
0,05
Rh / 103 [#1]
0,01
Pd / 105 [#1]
0,02
Ag / 107 [#2]
0,01
Cd / 111 [#2]
0,01
In / 115 [#2]
0,01
Sn / 118 [#2]
0,10
Sb / 121 [#2]
0,01
SHAPE-25
(ID162)
MELONE
162
Melon
Tursiops truncatus
0,50
SHAPE-26
(ID163)
CUTE-ADIPE
163
Skin-Blubber
Tursiops truncatus
0,50
SHAPE-27
(ID163)
MUSCOLO
163
Muscle
Tursiops truncatus
0,52
< 0,01
1,51
0,77
< 0,05
0,01
< 0,50
11,29
< 0,02
< 0,01
< 0,05
2,35
< 0,01
0,11
0,17
0,13
0,09
< 0,01
< 0,01
< 0,02
< 0,01
< 0,01
< 0,01
< 0,10
0,04
< 0,01
103,00
66,36
0,22
0,19
1,62
128,10
0,41
0,06
0,08
8,73
0,03
0,24
0,56
0,27
16,90
0,04
0,01
0,11
< 0,01
< 0,01
< 0,01
< 0,10
< 0,01
< 0,01
367,02
6,74
< 0,05
0,02
1,04
386,83
1,17
0,01
1,90
30,98
0,01
0,11
48,29
1,05
0,39
0,02
< 0,01
< 0,02
< 0,01
0,07
< 0,01
0,57
< 0,01
SHAPE-28
SHAPE-29
(ID163)
(ID163)
FEGATO
RENE
163
163
Liver
Kidney
Tursiops trunTursiops truncatus
catus
0,54
0,49
Concentration
(mg/kg)
< 0,01
< 0,01
2,09
18,94
6,46
47,42
0,07
0,11
0,02
0,03
8,88
0,82
1242,59
105,61
3,60
0,33
0,01
0,09
8,01
7,22
165,09
35,91
< 0,01
< 0,01
0,08
0,14
218,06
10,02
0,74
0,98
0,40
3,28
2,05
0,08
< 0,01
< 0,01
< 0,02
0,03
3,61
0,15
2,46
5,32
< 0,01
< 0,01
0,96
< 0,10
< 0,01
< 0,01
SHAPE-30
(ID163)
MELONE
163
Melon
Tursiops truncatus
0,52
SHAPE-31
(ID164)
CUTE-ADIPE
164
Skin-Blubber
Tursiops truncatus
0,50
SHAPE-32
(ID164)
MUSCOLO
164
Muscle
Tursiops truncatus
0,51
< 0,01
49,74
42,25
0,19
0,20
1,55
71,14
0,27
0,05
0,09
4,30
0,01
0,18
1,31
0,09
3,91
0,02
< 0,01
0,04
< 0,01
0,01
< 0,01
< 0,10
< 0,01
0,01
138,30
127,40
0,36
1,07
7,27
310,00
1,60
0,10
0,09
12,30
0,05
0,18
0,14
0,28
3,60
0,02
< 0,01
0,04
< 0,01
< 0,01
< 0,01
< 0,10
< 0,01
< 0,01
2,39
57,91
0,07
< 0,01
< 0,50
96,57
0,31
0,05
0,67
13,37
< 0,01
0,25
0,24
1,37
10,97
0,01
< 0,01
0,07
< 0,01
< 0,01
< 0,01
< 0,10
< 0,01
Appendix - 8
Sample Name
Sample ID
Tissue
Specie
Sample Weight (g)
Element / Atomic weight
LLoQ(2)
[analytical mode(1)]
Te / 130 [#1]
0,01
Cs / 133 [#1]
0,01
Ir / 193 [#1]
0,01
Pt / 195 [#1]
0,01
Hg / 202 [#1]
0,01
Tl / 205 [#1]
0,01
Pb / 208 [#1]
0,01
Bi / 209 [#1]
0,01
U / 238 [#1]
0,01
(1) [#1] without ORS, [#2] with ORS;
SHAPE-25
(ID162)
MELONE
162
Melon
Tursiops truncatus
0,50
SHAPE-26
(ID163)
CUTE-ADIPE
163
Skin-Blubber
Tursiops truncatus
0,50
SHAPE-27
(ID163)
MUSCOLO
163
Muscle
Tursiops truncatus
0,52
< 0,01
< 0,01
< 0,01
< 0,01
0,06
< 0,01
< 0,01
< 0,01
< 0,01
0,01
0,01
< 0,01
< 0,01
2,31
< 0,01
0,02
< 0,01
0,02
< 0,01
0,03
< 0,01
< 0,01
189,33
< 0,01
< 0,01
< 0,01
< 0,01
SHAPE-28
SHAPE-29
(ID163)
(ID163)
FEGATO
RENE
163
163
Liver
Kidney
Tursiops trunTursiops truncatus
catus
0,54
0,49
Concentration
(mg/kg)
0,10
0,02
0,02
0,02
< 0,01
< 0,01
< 0,01
< 0,01
1425,93
41,90
0,02
< 0,01
0,11
0,02
0,11
0,05
< 0,01
0,01
SHAPE-30
(ID163)
MELONE
163
Melon
Tursiops truncatus
0,52
SHAPE-31
(ID164)
CUTE-ADIPE
164
Skin-Blubber
Tursiops truncatus
0,50
SHAPE-32
(ID164)
MUSCOLO
164
Muscle
Tursiops truncatus
0,51
< 0,01
< 0,01
< 0,01
< 0,01
7,17
< 0,01
0,01
< 0,01
0,02
0,01
0,02
0,01
< 0,01
1,74
< 0,01
0,17
< 0,01
0,04
< 0,01
0,03
< 0,01
< 0,01
1,60
< 0,01
< 0,01
< 0,01
< 0,01
(2) Lower Limit of Quantification
Appendix - 9
Sample Name
Sample ID
Tissue
Specie
Sample Weight (g)
Element / Atomic weight
LLoQ(2)
[analytical mode(1)]
Be / 9 [#1]
0,01
Al / 27 [#1]
0,10
Ti / 48 [#2]
0,05
V / 51 [#2]
0,05
Cr / 52 [#2]
0,01
Mn / 55 [#1]
0,50
Fe / 56 [#2]
0,50
Ni / 58 [#2]
0,02
Co / 59 [#1]
0,01
Cu / 63 [#2]
0,05
Zn / 66 [#2]
0,05
Ga / 69 [#1]
0,01
As / 75 [#2]
0,01
Se / 78 [#2]
0,05
Rb / 85 [#1]
0,01
Sr / 88 [#1]
0,05
Mo / 98 [#2]
0,05
Rh / 103 [#1]
0,01
Pd / 105 [#1]
0,02
Ag / 107 [#2]
0,01
Cd / 111 [#2]
0,01
In / 115 [#2]
0,01
Sn / 118 [#2]
0,10
Sb / 121 [#2]
0,01
Te / 130 [#1]
0,01
SHAPE-33
(ID164)
FEGATO
164
Liver
Tursiops truncatus
0,52
SHAPE-34
(ID164)
RENE
164
Kidney
Tursiops truncatus
0,52
SHAPE-36
(ID165)
CUTE-ADIPE
165
Skin-Blubber
Tursiops truncatus
0,49
< 0,01
0,21
4,84
< 0,05
0,01
1,41
233,27
0,76
0,01
7,61
33,98
< 0,01
0,24
1,54
1,30
0,38
0,72
< 0,01
< 0,02
0,27
0,01
< 0,01
0,25
< 0,01
< 0,01
< 0,01
< 0,10
1,44
0,05
< 0,01
0,58
63,79
0,18
0,04
3,29
15,69
< 0,01
0,24
1,77
1,18
0,17
0,06
< 0,01
< 0,02
0,01
0,06
< 0,01
< 0,10
< 0,01
< 0,01
< 0,01
< 0,10
1,87
< 0,05
< 0,01
< 0,50
5,43
< 0,02
0,01
< 0,05
22,87
< 0,01
0,57
1,74
0,47
0,29
0,01
< 0,01
< 0,02
< 0,01
< 0,01
< 0,01
< 0,10
< 0,01
< 0,01
SHAPE-37
SHAPE-38
(ID165)
(ID165)
MUSCOLO
FEGATO
165
165
Muscle
Liver
Tursiops trunTursiops truncatus
catus
0,49
0,48
Concentration
(mg/kg)
< 0,01
< 0,01
< 0,10
< 0,10
1,38
1,51
< 0,05
0,19
< 0,01
< 0,01
< 0,50
1,93
278,06
740,73
0,82
2,20
0,01
0,01
0,54
2,80
12,66
14,22
< 0,01
< 0,01
0,08
0,11
0,43
35,89
1,69
1,08
0,12
0,20
0,01
0,76
< 0,01
< 0,01
< 0,02
< 0,02
< 0,01
1,81
< 0,01
0,01
< 0,01
< 0,01
< 0,10
0,53
< 0,01
< 0,01
< 0,01
0,09
SHAPE-39
(ID165)
RENE
165
Kidney
Tursiops truncatus
0,52
SHAPE-40
(ID165)
MELONE
165
Melon
Tursiops truncatus
0,49
SHAPE-41
(ID166)
CUTE-ADIPE
166
Skin-Blubber
Tursiops truncatus
0,54
< 0,01
< 0,10
7,69
0,05
< 0,01
< 0,50
280,96
0,82
0,06
1,90
15,78
< 0,01
0,09
3,70
1,15
0,67
0,03
< 0,01
< 0,02
0,02
0,12
< 0,01
< 0,10
< 0,01
0,01
< 0,01
0,29
2,10
< 0,05
0,05
< 0,50
16,69
0,02
0,01
< 0,05
3,43
< 0,01
0,23
0,52
0,06
0,18
0,01
< 0,01
< 0,02
< 0,01
< 0,01
< 0,01
< 0,10
< 0,01
< 0,01
< 0,01
74,94
101,20
0,27
0,27
2,36
208,43
0,78
0,09
0,20
10,94
< 0,01
0,29
0,38
0,47
31,29
0,05
0,01
< 0,02
< 0,01
< 0,01
< 0,01
< 0,10
< 0,01
< 0,01
Appendix - 10
Sample Name
Sample ID
Tissue
Specie
Sample Weight (g)
Element / Atomic weight
LLoQ(2)
[analytical mode(1)]
Cs / 133 [#1]
0,01
Ir / 193 [#1]
0,01
Pt / 195 [#1]
0,01
Hg / 202 [#1]
0,01
Tl / 205 [#1]
0,01
Pb / 208 [#1]
0,01
Bi / 209 [#1]
0,01
U / 238 [#1]
0,01
(1) [#1] without ORS, [#2] with ORS;
SHAPE-33
(ID164)
FEGATO
164
Liver
Tursiops truncatus
0,52
SHAPE-34
(ID164)
RENE
164
Kidney
Tursiops truncatus
0,52
SHAPE-36
(ID165)
CUTE-ADIPE
165
Skin-Blubber
Tursiops truncatus
0,49
0,02
< 0,01
< 0,01
5,80
< 0,01
0,02
0,01
< 0,01
0,02
< 0,01
< 0,01
3,27
< 0,01
0,02
0,01
< 0,01
0,01
< 0,01
< 0,01
2,87
< 0,01
< 0,01
< 0,01
< 0,01
SHAPE-37
SHAPE-38
(ID165)
(ID165)
MUSCOLO
FEGATO
165
165
Muscle
Liver
Tursiops trunTursiops truncatus
catus
0,49
0,48
Concentration
(mg/kg)
0,04
0,01
< 0,01
< 0,01
< 0,01
< 0,01
4,24
158,65
< 0,01
< 0,01
< 0,01
0,08
< 0,01
0,04
< 0,01
< 0,01
SHAPE-39
(ID165)
RENE
165
Kidney
Tursiops truncatus
0,52
SHAPE-40
(ID165)
MELONE
165
Melon
Tursiops truncatus
0,49
SHAPE-41
(ID166)
CUTE-ADIPE
166
Skin-Blubber
Tursiops truncatus
0,54
0,02
< 0,01
< 0,01
13,26
< 0,01
< 0,01
0,03
< 0,01
< 0,01
< 0,01
< 0,01
2,08
< 0,01
0,02
< 0,01
0,01
0,02
0,01
< 0,01
2,51
< 0,01
0,07
< 0,01
0,04
(2) Lower Limit of Quantification
Appendix - 11
Sample Name
Sample ID
Tissue
Specie
Sample Weight (g)
Element / Atomic weight
LLoQ(2)
[analytical mode(1)]
Be / 9 [#1]
0,01
Al / 27 [#1]
0,10
Ti / 48 [#2]
0,05
V / 51 [#2]
0,05
Cr / 52 [#2]
0,01
Mn / 55 [#1]
0,50
Fe / 56 [#2]
0,50
Ni / 58 [#2]
0,02
Co / 59 [#1]
0,01
Cu / 63 [#2]
0,05
Zn / 66 [#2]
0,05
Ga / 69 [#1]
0,01
As / 75 [#2]
0,01
Se / 78 [#2]
0,05
Rb / 85 [#1]
0,01
Sr / 88 [#1]
0,05
Mo / 98 [#2]
0,05
Rh / 103 [#1]
0,01
Pd / 105 [#1]
0,02
Ag / 107 [#2]
0,01
Cd / 111 [#2]
0,01
In / 115 [#2]
0,01
Sn / 118 [#2]
0,10
Sb / 121 [#2]
0,01
SHAPE-43
(ID166)
FEGATO
166
Liver
Tursiops truncatus
0,50
SHAPE-46
(ID180)
CUTE-ADIPE
180
Skin-Blubber
Tursiops truncatus
0,47
SHAPE-47
(ID180)
MUSCOLO
180
Muscle
Tursiops truncatus
0,49
< 0,01
1,46
3,74
0,05
< 0,01
3,50
1037,00
3,05
0,01
10,50
57,63
< 0,01
0,40
25,99
0,97
0,26
1,23
< 0,01
< 0,02
1,95
0,05
< 0,01
0,43
< 0,01
< 0,01
0,51
5,29
< 0,05
0,06
< 0,50
14,17
0,02
< 0,01
0,38
26,59
< 0,01
0,64
1,47
0,53
0,25
0,01
< 0,01
< 0,02
< 0,01
< 0,01
< 0,01
< 0,10
< 0,01
< 0,01
< 0,10
1,12
< 0,05
< 0,01
< 0,50
276,33
0,81
< 0,01
1,16
6,91
< 0,01
0,38
0,36
1,20
0,09
0,01
< 0,01
< 0,02
< 0,01
< 0,01
< 0,01
< 0,10
< 0,01
SHAPE-48
SHAPE-49
(ID180)
(ID180)
FEGATO
RENE
180
180
Liver
Kidney
Tursiops trunTursiops truncatus
catus
0,50
0,48
Concentration
(mg/kg)
< 0,01
< 0,01
< 0,10
< 0,10
2,71
7,99
0,09
0,10
0,02
< 0,01
3,78
0,60
445,90
40,47
1,32
0,10
0,01
0,03
9,94
3,45
27,61
15,58
< 0,01
< 0,01
1,09
0,59
3,84
1,61
1,07
0,77
0,36
0,66
1,09
0,03
< 0,01
< 0,01
< 0,02
< 0,02
1,09
0,01
0,11
0,27
< 0,01
< 0,01
0,11
< 0,10
< 0,01
< 0,01
SHAPE-51
(ID185)
CUTE-ADIPE
185
Skin-Blubber
Tursiops truncatus
0,48
SHAPE-52
(ID185)
MUSCOLO
185
Muscle
Tursiops truncatus
0,50
SHAPE-53
(ID185)
FEGATO
185
Liver
Tursiops truncatus
0,53
< 0,01
1,26
8,49
< 0,05
0,02
< 0,50
19,16
0,02
0,01
0,11
26,36
< 0,01
0,65
0,90
0,21
2,59
0,01
< 0,01
< 0,02
< 0,01
< 0,01
< 0,01
< 0,10
< 0,01
< 0,01
< 0,10
0,83
< 0,05
0,01
< 0,50
168,90
0,49
< 0,01
1,01
10,06
< 0,01
0,11
0,39
1,09
0,08
0,01
< 0,01
< 0,02
< 0,01
< 0,01
< 0,01
< 0,10
< 0,01
< 0,01
< 0,10
2,16
0,13
< 0,01
1,37
769,72
2,28
0,01
11,44
36,47
< 0,01
0,10
56,51
0,87
0,14
1,71
< 0,01
< 0,02
5,03
0,20
< 0,01
0,52
< 0,01
Appendix - 12
Sample Name
Sample ID
Tissue
Specie
Sample Weight (g)
Element / Atomic weight
LLoQ(2)
[analytical mode(1)]
Te / 130 [#1]
0,01
Cs / 133 [#1]
0,01
Ir / 193 [#1]
0,01
Pt / 195 [#1]
0,01
Hg / 202 [#1]
0,01
Tl / 205 [#1]
0,01
Pb / 208 [#1]
0,01
Bi / 209 [#1]
0,01
U / 238 [#1]
0,01
(1) [#1] without ORS, [#2] with ORS;
SHAPE-43
(ID166)
FEGATO
166
Liver
Tursiops truncatus
0,50
SHAPE-46
(ID180)
CUTE-ADIPE
180
Skin-Blubber
Tursiops truncatus
0,47
SHAPE-47
(ID180)
MUSCOLO
180
Muscle
Tursiops truncatus
0,49
0,05
0,02
< 0,01
< 0,01
139,90
< 0,01
0,04
0,12
0,00
< 0,01
0,02
< 0,01
< 0,01
1,52
< 0,01
< 0,01
< 0,01
< 0,01
< 0,01
0,08
< 0,01
< 0,01
2,34
< 0,01
< 0,01
< 0,01
< 0,01
SHAPE-48
SHAPE-49
(ID180)
(ID180)
FEGATO
RENE
180
180
Liver
Kidney
Tursiops trunTursiops truncatus
catus
0,50
0,48
Concentration
(mg/kg)
< 0,01
< 0,01
0,04
0,03
< 0,01
< 0,01
< 0,01
< 0,01
16,33
3,28
< 0,01
< 0,01
< 0,01
< 0,01
0,02
< 0,01
< 0,01
< 0,01
SHAPE-51
(ID185)
CUTE-ADIPE
185
Skin-Blubber
Tursiops truncatus
0,48
SHAPE-52
(ID185)
MUSCOLO
185
Muscle
Tursiops truncatus
0,50
SHAPE-53
(ID185)
FEGATO
185
Liver
Tursiops truncatus
0,53
< 0,01
< 0,01
< 0,01
< 0,01
4,90
< 0,01
< 0,01
< 0,01
0,01
< 0,01
0,03
< 0,01
< 0,01
5,72
< 0,01
< 0,01
< 0,01
< 0,01
0,08
0,01
< 0,01
< 0,01
245,94
< 0,01
0,07
0,10
< 0,01
(2) Lower Limit of Quantification
Appendix - 13
Sample Name
Sample ID
Tissue
Specie
Sample Weight (g)
Element / Atomic weight
LLoQ(2)
[analytical mode(1)]
Be / 9 [#1]
0,01
Al / 27 [#1]
0,10
Ti / 48 [#2]
0,05
V / 51 [#2]
0,05
Cr / 52 [#2]
0,01
Mn / 55 [#1]
0,50
Fe / 56 [#2]
0,50
Ni / 58 [#2]
0,02
Co / 59 [#1]
0,01
Cu / 63 [#2]
0,05
Zn / 66 [#2]
0,05
Ga / 69 [#1]
0,01
As / 75 [#2]
0,01
Se / 78 [#2]
0,05
Rb / 85 [#1]
0,01
Sr / 88 [#1]
0,05
Mo / 98 [#2]
0,05
Rh / 103 [#1]
0,01
Pd / 105 [#1]
0,02
Ag / 107 [#2]
0,01
Cd / 111 [#2]
0,01
In / 115 [#2]
0,01
Sn / 118 [#2]
0,10
Sb / 121 [#2]
0,01
SHAPE-54
(ID185)
RENE
185
Kidney
Tursiops truncatus
0,49
SHAPE-56
(ID192)
CUTE-ADIPE
192
Skin-Blubber
Tursiops truncatus
0,53
SHAPE-57
(ID192)
MUSCOLO
192
Muscle
Tursiops truncatus
0,49
< 0,01
0,10
4,51
< 0,05
< 0,01
< 0,50
163,27
0,47
0,02
2,95
17,30
< 0,01
0,14
3,85
0,96
0,27
0,04
< 0,01
< 0,02
0,03
0,27
< 0,01
< 0,10
< 0,01
< 0,01
< 0,10
2,09
< 0,05
0,01
< 0,50
31,77
0,06
< 0,01
0,09
32,39
< 0,01
0,85
0,95
0,69
0,17
< 0,05
< 0,01
< 0,02
< 0,01
< 0,01
< 0,01
< 0,10
< 0,01
< 0,01
< 0,10
1,54
0,12
< 0,01
< 0,50
86,10
0,26
< 0,01
0,71
41,37
< 0,01
0,27
0,20
1,82
0,07
< 0,05
< 0,01
< 0,02
< 0,01
< 0,01
< 0,01
< 0,10
< 0,01
SHAPE-58
SHAPE-59
(ID192)
(ID192)
FEGATO
RENE
192
192
Liver
Kidney
Tursiops trunTursiops truncatus
catus
0,49
0,52
Concentration
(mg/kg)
< 0,01
< 0,01
< 0,10
< 0,10
1,59
2,12
0,05
0,06
< 0,01
0,02
0,83
< 0,50
324,29
117,21
0,92
0,30
< 0,01
0,02
5,50
1,88
16,48
7,66
< 0,01
< 0,01
0,41
0,88
2,77
1,30
1,57
0,99
0,14
0,17
0,60
< 0,05
< 0,01
< 0,01
< 0,02
< 0,02
0,72
< 0,01
0,02
0,04
< 0,01
< 0,01
0,16
< 0,10
< 0,01
< 0,01
SHAPE-61
(ID193)
CUTE-ADIPE
193
Skin-Blubber
Tursiops truncatus
0,50
SHAPE-62
(ID193)
MUSCOLO
193
Muscle
Tursiops truncatus
0,51
SHAPE-64
(ID193)
RENE
193
Kidney
Tursiops truncatus
0,52
< 0,01
21,93
22,38
< 0,05
0,06
< 0,50
28,97
0,10
0,02
0,21
26,22
0,01
0,87
1,50
0,25
3,73
< 0,05
< 0,01
0,03
< 0,01
0,02
< 0,01
< 0,10
< 0,01
< 0,01
< 0,10
1,77
< 0,05
0,01
< 0,50
287,35
0,84
< 0,01
1,44
19,82
< 0,01
0,20
18,00
1,22
0,17
< 0,05
< 0,01
< 0,02
< 0,01
0,04
< 0,01
0,33
< 0,01
< 0,01
< 0,10
23,95
0,05
< 0,01
< 0,50
233,46
0,66
0,04
3,35
20,01
< 0,01
0,30
3,73
1,28
2,40
0,07
< 0,01
0,02
0,01
3,89
< 0,01
0,02
< 0,01
Appendix - 14
Sample Name
Sample ID
Tissue
Specie
Sample Weight (g)
Element / Atomic weight
LLoQ(2)
[analytical mode(1)]
Te / 130 [#1]
0,01
Cs / 133 [#1]
0,01
Ir / 193 [#1]
0,01
Pt / 195 [#1]
0,01
Hg / 202 [#1]
0,01
Tl / 205 [#1]
0,01
Pb / 208 [#1]
0,01
Bi / 209 [#1]
0,01
U / 238 [#1]
0,01
(1) [#1] without ORS, [#2] with ORS;
SHAPE-54
(ID185)
RENE
185
Kidney
Tursiops truncatus
0,49
SHAPE-56
(ID192)
CUTE-ADIPE
192
Skin-Blubber
Tursiops truncatus
0,53
SHAPE-57
(ID192)
MUSCOLO
192
Muscle
Tursiops truncatus
0,49
< 0,01
0,02
< 0,01
< 0,01
15,45
< 0,01
< 0,01
0,02
< 0,01
< 0,01
< 0,01
< 0,01
< 0,01
0,39
< 0,01
< 0,01
< 0,01
< 0,01
< 0,01
0,02
< 0,01
< 0,01
1,54
< 0,01
< 0,01
< 0,01
< 0,01
SHAPE-58
SHAPE-59
(ID192)
(ID192)
FEGATO
RENE
192
192
Liver
Kidney
Tursiops trunTursiops truncatus
catus
0,49
0,52
Concentration
(mg/kg)
0,01
0,01
0,01
0,01
< 0,01
< 0,01
< 0,01
< 0,01
11,52
1,90
< 0,01
< 0,01
< 0,01
< 0,01
0,02
0,01
< 0,01
< 0,01
SHAPE-61
(ID193)
CUTE-ADIPE
193
Skin-Blubber
Tursiops truncatus
0,50
SHAPE-62
(ID193)
MUSCOLO
193
Muscle
Tursiops truncatus
0,51
SHAPE-64
(ID193)
RENE
193
Kidney
Tursiops truncatus
0,52
< 0,01
0,01
< 0,01
< 0,01
6,28
< 0,01
0,01
< 0,01
0,03
< 0,01
0,03
< 0,01
< 0,01
75,65
< 0,01
< 0,01
< 0,01
< 0,01
< 0,01
0,03
< 0,01
< 0,01
15,41
< 0,01
0,02
0,03
< 0,01
(2) Lower Limit of Quantification
Appendix - 15
Sample Name
Sample ID
Tissue
Specie
Sample Weight (g)
Element / Atomic weight
LLoQ(2)
[analytical mode(1)]
Be / 9 [#1]
0,01
Al / 27 [#1]
0,10
Ti / 48 [#2]
0,05
V / 51 [#2]
0,05
Cr / 52 [#2]
0,01
Mn / 55 [#1]
0,50
Fe / 56 [#2]
0,50
Ni / 58 [#2]
0,02
Co / 59 [#1]
0,01
Cu / 63 [#2]
0,05
Zn / 66 [#2]
0,05
Ga / 69 [#1]
0,01
As / 75 [#2]
0,01
Se / 78 [#2]
0,05
Rb / 85 [#1]
0,01
Sr / 88 [#1]
0,05
Mo / 98 [#2]
0,05
Rh / 103 [#1]
0,01
Pd / 105 [#1]
0,02
Ag / 107 [#2]
0,01
Cd / 111 [#2]
0,01
In / 115 [#2]
0,01
Sn / 118 [#2]
0,10
Sb / 121 [#2]
0,01
Te / 130 [#1]
0,01
SHAPE-65
(ID193)
MELONE
193
Melon
Tursiops truncatus
0,52
SHAPE-66
(ID203)
CUTE-ADIPE
203
Skin-Blubber
Tursiops truncatus
0,49
SHAPE-67
(ID203)
MUSCOLO
203
Muscle
Tursiops truncatus
0,48
< 0,01
0,71
3,53
0,07
0,02
< 0,50
4,74
< 0,02
0,02
< 0,05
3,57
< 0,01
0,19
2,67
0,09
0,34
< 0,05
< 0,01
0,02
< 0,01
0,05
< 0,01
< 0,10
< 0,01
< 0,01
< 0,01
0,20
2,17
< 0,05
< 0,01
< 0,50
20,62
0,04
< 0,01
0,11
42,95
< 0,01
1,13
3,19
0,45
0,15
< 0,05
< 0,01
< 0,02
< 0,01
< 0,01
< 0,01
< 0,10
< 0,01
< 0,01
< 0,01
< 0,10
2,39
0,07
< 0,01
< 0,50
92,36
0,27
< 0,01
0,62
21,46
< 0,01
0,15
0,43
0,77
0,10
< 0,05
< 0,01
< 0,02
< 0,01
< 0,01
< 0,01
< 0,10
< 0,01
< 0,01
SHAPE-69
SHAPE-71
(ID203)
(ID218)
RENE
CUTE-ADIPE
203
218
Kidney
Skin-Blubber
Stenella coeStenella coeruleoalba
ruleoalba
0,51
0,54
Concentration
(mg/kg)
< 0,01
< 0,01
< 0,10
0,68
2,98
3,09
< 0,05
< 0,05
0,01
< 0,01
< 0,50
< 0,50
135,20
20,19
0,35
0,02
0,02
0,01
2,32
0,26
10,44
87,65
< 0,01
< 0,01
0,63
0,82
4,21
13,36
0,76
0,69
0,23
0,41
< 0,05
< 0,05
< 0,01
< 0,01
< 0,02
< 0,02
0,04
< 0,01
0,36
0,01
< 0,01
< 0,01
< 0,10
< 0,10
< 0,01
< 0,01
< 0,01
< 0,01
SHAPE-72
(ID218)
MUSCOLO
218
Muscle
Stenella coeruleoalba
0,50
SHAPE-73
(ID218)
FEGATO
218
Liver
Stenella coeruleoalba
0,48
SHAPE-74
(ID218)
RENE
218
Kidney
Stenella coeruleoalba
0,54
< 0,01
0,05
1,46
0,05
0,01
< 0,50
90,44
0,45
< 0,01
0,64
7,69
< 0,01
0,25
0,92
1,49
0,15
< 0,05
< 0,01
< 0,02
< 0,01
0,01
< 0,01
< 0,10
< 0,01
< 0,01
< 0,01
< 0,10
1,26
0,10
< 0,01
1,32
412,81
1,19
0,01
6,94
23,67
< 0,01
1,35
30,66
1,42
0,21
1,23
< 0,01
< 0,02
1,51
3,50
< 0,01
0,11
< 0,01
0,01
< 0,01
< 0,10
3,28
< 0,05
< 0,01
< 0,50
122,69
0,30
0,03
2,89
18,44
< 0,01
0,51
5,05
1,06
0,54
0,07
< 0,01
< 0,02
0,05
4,83
< 0,01
< 0,10
< 0,01
< 0,01
Appendix - 16
Sample Name
Sample ID
Tissue
Specie
Sample Weight (g)
Element / Atomic weight
LLoQ(2)
[analytical mode(1)]
Cs / 133 [#1]
0,01
Ir / 193 [#1]
0,01
Pt / 195 [#1]
0,01
Hg / 202 [#1]
0,01
Tl / 205 [#1]
0,01
Pb / 208 [#1]
0,01
Bi / 209 [#1]
0,01
U / 238 [#1]
0,01
(1) [#1] without ORS, [#2] with ORS;
SHAPE-65
(ID193)
MELONE
193
Melon
Tursiops truncatus
0,52
SHAPE-66
(ID203)
CUTE-ADIPE
203
Skin-Blubber
Tursiops truncatus
0,49
SHAPE-67
(ID203)
MUSCOLO
203
Muscle
Tursiops truncatus
0,48
< 0,01
< 0,01
< 0,01
6,22
< 0,01
< 0,01
< 0,01
< 0,01
0,01
< 0,01
< 0,01
1,86
< 0,01
< 0,01
< 0,01
< 0,01
0,02
< 0,01
< 0,01
2,33
< 0,01
< 0,01
< 0,01
< 0,01
SHAPE-69
SHAPE-71
(ID203)
(ID218)
RENE
CUTE-ADIPE
203
218
Kidney
Skin-Blubber
Stenella coeStenella coeruleoalba
ruleoalba
0,51
0,54
Concentration
(mg/kg)
0,01
0,01
< 0,01
< 0,01
< 0,01
< 0,01
9,69
2,35
< 0,01
< 0,01
< 0,01
< 0,01
0,03
< 0,01
< 0,01
< 0,01
SHAPE-72
(ID218)
MUSCOLO
218
Muscle
Stenella coeruleoalba
0,50
SHAPE-73
(ID218)
FEGATO
218
Liver
Stenella coeruleoalba
0,48
SHAPE-74
(ID218)
RENE
218
Kidney
Stenella coeruleoalba
0,54
0,02
< 0,01
< 0,01
6,75
< 0,01
< 0,01
< 0,01
< 0,01
0,01
< 0,01
< 0,01
138,44
< 0,01
< 0,01
< 0,01
< 0,01
0,01
< 0,01
< 0,01
14,06
< 0,01
< 0,01
< 0,01
< 0,01
(2) Lower Limit of Quantification
Appendix - 17
Sample Name
Sample ID
Tissue
Specie
Sample Weight (g)
Element / Atomic weight
LLoQ(2)
[analytical mode(1)]
Be / 9 [#1]
0,01
Al / 27 [#1]
0,10
Ti / 48 [#2]
0,05
V / 51 [#2]
0,05
Cr / 52 [#2]
0,01
Mn / 55 [#1]
0,50
Fe / 56 [#2]
0,50
Ni / 58 [#2]
0,02
Co / 59 [#1]
0,01
Cu / 63 [#2]
0,05
Zn / 66 [#2]
0,05
Ga / 69 [#1]
0,01
As / 75 [#2]
0,01
Se / 78 [#2]
0,05
Rb / 85 [#1]
0,01
Sr / 88 [#1]
0,05
Mo / 98 [#2]
0,05
Rh / 103 [#1]
0,01
Pd / 105 [#1]
0,02
Ag / 107 [#2]
0,01
Cd / 111 [#2]
0,01
In / 115 [#2]
0,01
Sn / 118 [#2]
0,10
Sb / 121 [#2]
0,01
SHAPE-75
(ID218)
MELONE
218
Melon
Stenella coeruleoalba
0,50
SHAPE-76
(ID220)
CUTE-ADIPE
220
Skin-Blubber
Stenella coeruleoalba
0,49
SHAPE-77
(ID220)
MUSCOLO
220
Muscle
Stenella coeruleoalba
0,48
< 0,01
0,21
3,05
< 0,05
0,02
< 0,50
13,75
< 0,02
0,01
0,18
47,12
< 0,01
0,56
13,37
0,38
0,25
< 0,05
< 0,01
< 0,02
< 0,01
0,01
< 0,01
< 0,10
< 0,01
< 0,01
90,48
176,63
0,31
0,81
4,78
159,80
0,58
0,04
< 0,05
13,81
0,02
0,69
2,12
0,17
0,85
< 0,05
< 0,01
< 0,02
< 0,01
0,01
< 0,01
< 0,10
< 0,01
< 0,01
2,17
5,91
0,18
0,05
< 0,50
170,10
0,47
0,01
0,71
6,87
< 0,01
0,45
0,90
1,19
0,18
< 0,05
< 0,01
< 0,02
< 0,01
0,01
< 0,01
< 0,10
< 0,01
SHAPE-78
SHAPE-79
(ID220)
(ID220)
FEGATO
RENE
220
220
Liver
Kidney
Stenella coeStenella coeruleoalba
ruleoalba
0,50
0,49
Concentration
(mg/kg)
< 0,01
< 0,01
17,89
31,36
13,37
79,28
0,09
0,10
0,17
0,56
2,05
2,19
370,60
185,41
1,20
0,72
0,02
0,07
5,57
1,86
18,17
12,30
< 0,01
0,01
0,77
0,79
22,23
2,42
0,99
0,86
0,85
8,01
0,73
< 0,05
< 0,01
< 0,01
< 0,02
0,05
1,31
< 0,01
1,52
4,15
< 0,01
< 0,01
< 0,10
< 0,10
< 0,01
< 0,01
SHAPE-81
(ID221)
CUTE-ADIPE
221
Skin-Blubber
Stenella coeruleoalba
0,49
SHAPE-82
(ID221)
MUSCOLO
221
Muscle
Stenella coeruleoalba
0,54
SHAPE-83
(ID221)
FEGATO
221
Liver
Stenella coeruleoalba
0,55
< 0,01
< 0,10
6,23
0,06
0,01
< 0,50
101,08
0,39
0,02
2,74
23,78
< 0,01
0,83
3,53
1,09
0,44
0,06
< 0,01
< 0,02
0,01
4,91
< 0,01
< 0,10
< 0,01
< 0,01
< 0,10
1,53
0,05
0,01
2,26
411,02
1,89
0,01
7,39
48,80
< 0,01
0,78
30,42
1,11
0,10
0,85
< 0,01
< 0,02
1,86
3,41
< 0,01
0,31
< 0,01
< 0,01
< 0,10
0,90
< 0,05
< 0,01
< 0,50
211,55
0,60
0,01
0,57
5,20
< 0,01
0,24
0,67
1,14
0,05
< 0,05
< 0,01
< 0,02
< 0,01
0,02
< 0,01
< 0,10
< 0,01
Appendix - 18
Sample Name
Sample ID
Tissue
Specie
Sample Weight (g)
Element / Atomic weight
LLoQ(2)
[analytical mode(1)]
Te / 130 [#1]
0,01
Cs / 133 [#1]
0,01
Ir / 193 [#1]
0,01
Pt / 195 [#1]
0,01
Hg / 202 [#1]
0,01
Tl / 205 [#1]
0,01
Pb / 208 [#1]
0,01
Bi / 209 [#1]
0,01
U / 238 [#1]
0,01
(1) [#1] without ORS, [#2] with ORS;
SHAPE-75
(ID218)
MELONE
218
Melon
Stenella coeruleoalba
0,50
SHAPE-76
(ID220)
CUTE-ADIPE
220
Skin-Blubber
Stenella coeruleoalba
0,49
SHAPE-77
(ID220)
MUSCOLO
220
Muscle
Stenella coeruleoalba
0,48
< 0,01
< 0,01
< 0,01
< 0,01
2,48
< 0,01
< 0,01
< 0,01
< 0,01
< 0,01
0,01
< 0,01
< 0,01
1,35
< 0,01
< 0,01
< 0,01
0,01
< 0,01
0,02
< 0,01
< 0,01
5,03
< 0,01
< 0,01
< 0,01
< 0,01
SHAPE-78
SHAPE-79
(ID220)
(ID220)
FEGATO
RENE
220
220
Liver
Kidney
Stenella coeStenella coeruleoalba
ruleoalba
0,50
0,49
Concentration
(mg/kg)
< 0,01
< 0,01
0,02
0,01
< 0,01
< 0,01
< 0,01
< 0,01
100,40
6,93
< 0,01
< 0,01
0,01
0,04
0,01
0,01
< 0,01
< 0,01
SHAPE-81
(ID221)
CUTE-ADIPE
221
Skin-Blubber
Stenella coeruleoalba
0,49
SHAPE-82
(ID221)
MUSCOLO
221
Muscle
Stenella coeruleoalba
0,54
SHAPE-83
(ID221)
FEGATO
221
Liver
Stenella coeruleoalba
0,55
< 0,01
0,03
< 0,01
< 0,01
9,18
< 0,01
< 0,01
< 0,01
0,01
0,01
0,02
< 0,01
< 0,01
140,93
0,01
0,02
< 0,01
< 0,01
< 0,01
0,03
< 0,01
< 0,01
4,72
< 0,01
< 0,01
< 0,01
< 0,01
(2) Lower Limit of Quantification
Appendix - 19
Sample Name
Sample ID
Tissue
Specie
Sample Weight (g)
Element / Atomic weight
LLoQ(2)
[analytical mode(1)]
Be / 9 [#1]
0,01
Al / 27 [#1]
0,10
Ti / 48 [#2]
0,05
V / 51 [#2]
0,05
Cr / 52 [#2]
0,01
Mn / 55 [#1]
0,50
Fe / 56 [#2]
0,50
Ni / 58 [#2]
0,02
Co / 59 [#1]
0,01
Cu / 63 [#2]
0,05
Zn / 66 [#2]
0,05
Ga / 69 [#1]
0,01
As / 75 [#2]
0,01
Se / 78 [#2]
0,05
Rb / 85 [#1]
0,01
Sr / 88 [#1]
0,05
Mo / 98 [#2]
0,05
Rh / 103 [#1]
0,01
Pd / 105 [#1]
0,02
Ag / 107 [#2]
0,01
Cd / 111 [#2]
0,01
In / 115 [#2]
0,01
Sn / 118 [#2]
0,10
Sb / 121 [#2]
0,01
Te / 130 [#1]
0,01
SHAPE-84
(ID221)
RENE
221
Kidney
Stenella coeruleoalba
0,48
Concentration
(mg/kg)
< 0,01
0,30
1,75
< 0,05
< 0,01
< 0,50
6,86
< 0,02
< 0,01
0,12
84,75
< 0,01
1,40
16,64
0,51
0,11
< 0,05
< 0,01
< 0,02
< 0,01
0,02
< 0,01
< 0,10
< 0,01
< 0,01
Appendix - 20
Sample Name
Sample ID
Tissue
Specie
Sample Weight (g)
Element / Atomic weight
LLoQ(2)
[analytical mode(1)]
Cs / 133 [#1]
0,01
Ir / 193 [#1]
0,01
Pt / 195 [#1]
0,01
Hg / 202 [#1]
0,01
Tl / 205 [#1]
0,01
Pb / 208 [#1]
0,01
Bi / 209 [#1]
0,01
U / 238 [#1]
0,01
(1) [#1] without ORS, [#2] with ORS;
SHAPE-84
(ID221)
RENE
221
Kidney
Stenella coeruleoalba
0,48
Concentration
(mg/kg)
0,01
< 0,01
< 0,01
2,24
< 0,01
< 0,01
< 0,01
< 0,01
(2) Lower Limit of Quantification
Appendix - 21
Table II – Complete results of Polycyclic Aromatic Hydrocarbons (PAHs) and Organochlorine Compounds (OCs) concentrations (mg/kg on
wet weight, ww) in tissues of the cetaceans examined in the present study. For a number of samples it was not possible to perform the
analytical determination of both PAHs and OCs (*) or only OCs (**) since the tissue was not enough.
Sample Name
Sample ID
Tissue
Species
SHAPE-01
172
SHAPE-03
172
SHAPE-04
172
SHAPE-05
172
Skin-Blubber
Liver
Kidney
Spermaceti
Physeter macrocephalus
**
Physeter macrocephalus
*
Physeter macrocephalus
Physeter macrocephalus
SHAPE-06
196
SkinBlubber
Tursiops
truncatus
SHAPE-08
196
SHAPE-09
196
SHAPE-10
196
Liver
Kidney
Melon
Tursiops
truncatus
Tursiops truncatus
Tursiops truncatus
**
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0024
< 0,0049
< 0,0049
< 0,0049
< 0,0004
< 0,0004
< 0,0004
< 0,0004
< 0,0004
< 0,0004
< 0,0004
< 0,0004
< 0,0004
< 0,0004
< 0,0004
< 0,0004
< 0,0004
< 0,0004
< 0,0004
< 0,0022
< 0,0045
< 0,0045
< 0,0045
< 0,0008
< 0,0008
< 0,0008
< 0,0008
< 0,0008
< 0,0008
< 0,0008
< 0,0008
< 0,0008
< 0,0008
< 0,0008
< 0,0008
< 0,0008
< 0,0008
< 0,0008
< 0,0042
< 0,0085
< 0,0085
< 0,0085
0,0005
0,0007
POLYCYCLIC AROMATIC HYDROCARBONS (mg/kg, ww)
Fluorene
Phenanthrene
Anthracene
Fluoranthene
Pyrene
Benz(a)anthracene
Chrysene
Benzo(b)fluoranthene
Benzo(k)fluoranthene
Benzo(e)pyrene
Benzo(a)pyrene
Perylene
Indeno(1,2,3-cd)pyrene
Dibenz(a,h)anthracene
Benzo(g,h,i)perylene
Dibenzo(a,l)pyrene
Dibenzo(a,e)pyrene
Dibenzo(a,i)pyrene
Dibenzo(a,h)pyrene
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0024
< 0,0048
< 0,0048
< 0,0048
< 0,0005
< 0,0005
< 0,0005
0,0005
0,0007
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0023
< 0,0046
< 0,0046
< 0,0046
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0023
< 0,0047
< 0,0047
< 0,0047
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0024
< 0,0049
< 0,0049
< 0,0049
ORGANOCHLORINE COMPOUNDS (mg/kg, ww)
Hexachlorobenzene
0,0037
0,0192
0,0268
Appendix - 22
Sample Name
Sample ID
Skin-Blubber
Liver
Kidney
Spermaceti
Physeter macrocephalus
**
Physeter macrocephalus
*
Physeter macrocephalus
Physeter macrocephalus
SHAPE-06
196
SkinBlubber
Tursiops
truncatus
2,4’-DDE
4,4’-DDE
2,4’-DDD
4,4’-DDD
2,4’-DDT
4,4’-DDT
0,0126
0,8856
0,0104
0,0309
0,0194
0,0245
0,0913
6,2046
0,0382
0,1761
0,2889
0,5061
0,1898
39,8335
0,2396
1,2111
1,0047
0,4047
0,0059
0,6639
0,0072
0,0170
0,0070
0,0034
0,0074
1,1454
0,0094
0,0202
0,0148
0,0047
∑ DDT
4,4’-DDE/∑ DDT
0,9833
0,9006
7,3051
0,8493
42,8833
0,9289
0,7043
0,9426
1,2018
0,9530
Tissue
Species
SHAPE-01
172
SHAPE-03
172
SHAPE-04
172
SHAPE-05
172
SHAPE-08
196
SHAPE-09
196
SHAPE-10
196
Liver
Kidney
Melon
Tursiops
truncatus
Tursiops truncatus
Tursiops truncatus
**
Appendix - 23
Sample Name
Sample ID
Tissue
Species
SHAPE-11
214
Skin-Blubber
Stenella coeruleoalba
*
SHAPE-13
214
Liver
Stenella coeruleoalba
**
SHAPE-14
214
Kidney
Stenella coeruleoalba
**
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0023
< 0,0046
< 0,0046
< 0,0046
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0023
< 0,0045
< 0,0045
< 0,0045
SHAPE-15
214
Melon
Stenella coeruleoalba
**
SHAPE-16
215
Skin-Blubber
Grampus griseus
**
SHAPE-18
215
Liver
Grampus griseus
**
SHAPE-19
215
Kidney
Grampus griseus
**
SHAPE-20
215
Melon
Grampus griseus
**
< 0,0005
< 0,0005
< 0,0005
< 0,0005
0,0160
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
0,0259
< 0,0005
< 0,0026
< 0,0051
< 0,0051
< 0,0051
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0023
< 0,0046
< 0,0046
< 0,0046
POLYCYCLIC AROMATIC HYDROCARBONS (mg/kg, ww)
Fluorene
Phenanthrene
Anthracene
Fluoranthene
Pyrene
Benz(a)anthracene
Chrysene
Benzo(b)fluoranthene
Benzo(k)fluoranthene
Benzo(e)pyrene
Benzo(a)pyrene
Perylene
Indeno(1,2,3-cd)pyrene
Dibenz(a,h)anthracene
Benzo(g,h,i)perylene
Dibenzo(a,l)pyrene
Dibenzo(a,e)pyrene
Dibenzo(a,i)pyrene
Dibenzo(a,h)pyrene
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0025
< 0,0051
< 0,0051
< 0,0051
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
0,0018
0,0024
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
0,0048
0,0035
< 0,0005
< 0,0023
< 0,0045
< 0,0045
< 0,0045
< 0,0005
< 0,0005
< 0,0005
< 0,0005
0,0494
0,0380
0,0439
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0026
< 0,0051
< 0,0051
< 0,0051
ORGANOCHLORINE COMPOUNDS (mg/kg, ww)
Hexachlorobenzene
2,4’-DDE
4,4’-DDE
2,4’-DDD
4,4’-DDD
Appendix - 24
Sample Name
Sample ID
Tissue
Species
SHAPE-11
214
Skin-Blubber
Stenella coeruleoalba
*
SHAPE-13
214
Liver
Stenella coeruleoalba
**
SHAPE-14
214
Kidney
Stenella coeruleoalba
**
SHAPE-15
214
Melon
Stenella coeruleoalba
**
SHAPE-16
215
Skin-Blubber
Grampus griseus
**
SHAPE-18
215
Liver
Grampus griseus
**
SHAPE-19
215
Kidney
Grampus griseus
**
SHAPE-20
215
Melon
Grampus griseus
**
2,4’-DDT
4,4’-DDT
∑ DDT
4,4’-DDE/∑ DDT
Appendix - 25
Sample Name
Sample ID
Tissue
Species
SHAPE-21
162
Skin-Blubber
Tursiops truncatus
SHAPE-23
162
Liver
Tursiops truncatus
**
SHAPE-24
162
Kidney
Tursiops truncatus
**
SHAPE-25
162
Melon
Tursiops truncatus
SHAPE-26
163
Skin-Blubber
Tursiops truncatus
*
SHAPE-28
163
Liver
Tursiops truncatus
**
SHAPE-29
163
Kidney
Tursiops truncatus
**
SHAPE-30
163
Melon
Tursiops truncatus
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0025
< 0,0049
< 0,0049
< 0,0049
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0026
< 0,0052
< 0,0052
< 0,0052
POLYCYCLIC AROMATIC HYDROCARBONS (mg/kg, ww)
Fluorene
Phenanthrene
Anthracene
Fluoranthene
Pyrene
Benz(a)anthracene
Chrysene
Benzo(b)fluoranthene
Benzo(k)fluoranthene
Benzo(e)pyrene
Benzo(a)pyrene
Perylene
Indeno(1,2,3-cd)pyrene
Dibenz(a,h)anthracene
Benzo(g,h,i)perylene
Dibenzo(a,l)pyrene
Dibenzo(a,e)pyrene
Dibenzo(a,i)pyrene
Dibenzo(a,h)pyrene
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
0,0046
0,0046
0,0048
0,0050
< 0,0005
< 0,0005
< 0,0005
0,0092
0,0073
0,0099
< 0,0024
< 0,0048
< 0,0048
< 0,0048
< 0,0006
< 0,0006
< 0,0006
< 0,0006
< 0,0006
< 0,0006
< 0,0006
< 0,0006
< 0,0006
< 0,0006
< 0,0006
< 0,0006
< 0,0006
< 0,0006
< 0,0006
< 0,0032
< 0,0064
< 0,0064
< 0,0064
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0025
< 0,0050
< 0,0050
< 0,0050
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0025
< 0,0049
< 0,0049
< 0,0049
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0024
< 0,0049
< 0,0049
< 0,0049
ORGANOCHLORINE COMPOUNDS (mg/kg, ww)
Hexachlorobenzene
2,4’-DDE
4,4’-DDE
2,4’-DDD
4,4’-DDD
2,4’-DDT
0,0463
0,0328
2,3687
0,0579
0,4711
0,0243
0,0746
0,0356
2,2318
0,0561
0,3758
0,0424
0,0201
0,1101
24,7986
0,3609
0,5835
0,0367
Appendix - 26
Sample Name
Sample ID
Tissue
Species
SHAPE-21
162
Skin-Blubber
Tursiops truncatus
SHAPE-23
162
Liver
Tursiops truncatus
**
SHAPE-24
162
Kidney
Tursiops truncatus
**
SHAPE-25
162
Melon
Tursiops truncatus
SHAPE-26
163
Skin-Blubber
Tursiops truncatus
*
SHAPE-28
163
Liver
Tursiops truncatus
**
SHAPE-29
163
Kidney
Tursiops truncatus
**
SHAPE-30
163
Melon
Tursiops truncatus
4,4’-DDT
0,1383
0,0957
0,0065
∑ DDT
4,4’-DDE/∑ DDT
3,0932
0,7658
2,8374
0,7866
25,8962
0,9576
Appendix - 27
Sample Name
Sample ID
Tissue
Species
SHAPE-31
164
Skin-Blubber
Tursiops truncatus
**
SHAPE-33
164
Liver
Tursiops truncatus
**
SHAPE-34
164
Kidney
Tursiops truncatus
**
SHAPE-36
165
Skin-Blubber
Tursiops truncatus
SHAPE-38
165
Liver
Tursiops truncatus
**
SHAPE-39
165
Kidney
Tursiops truncatus
**
SHAPE-40
165
Melon
Tursiops truncatus
SHAPE-41
166
Skin-Blubber
Tursiops truncatus
**
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0024
< 0,0048
< 0,0048
< 0,0048
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
0,0063
0,0064
0,0091
0,0086
0,0067
0,0078
0,0089
0,0103
0,0088
0,0114
0,0063
0,0138
0,0115
0,0100
POLYCYCLIC AROMATIC HYDROCARBONS (mg/kg, ww)
Fluorene
Phenanthrene
Anthracene
Fluoranthene
Pyrene
Benz(a)anthracene
Chrysene
Benzo(b)fluoranthene
Benzo(k)fluoranthene
Benzo(e)pyrene
Benzo(a)pyrene
Perylene
Indeno(1,2,3-cd)pyrene
Dibenz(a,h)anthracene
Benzo(g,h,i)perylene
Dibenzo(a,l)pyrene
Dibenzo(a,e)pyrene
Dibenzo(a,i)pyrene
Dibenzo(a,h)pyrene
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
0,0027
0,0031
< 0,0005
< 0,0025
< 0,0051
< 0,0051
< 0,0051
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0026
< 0,0051
< 0,0051
< 0,0051
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0025
< 0,0050
< 0,0050
< 0,0050
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0023
< 0,0046
< 0,0046
< 0,0046
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0025
< 0,0050
< 0,0050
< 0,0050
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0025
< 0,0050
< 0,0050
< 0,0050
ORGANOCHLORINE COMPOUNDS (mg/kg, ww)
Hexachlorobenzene
2,4’-DDE
4,4’-DDE
2,4’-DDD
4,4’-DDD
2,4’-DDT
0,0542
0,2064
45,0004
0,3023
1,9130
0,6390
0,0348
0,0827
13,2362
0,0851
0,6003
0,2256
Appendix - 28
Sample Name
Sample ID
Tissue
Species
SHAPE-31
164
Skin-Blubber
Tursiops truncatus
**
SHAPE-33
164
Liver
Tursiops truncatus
**
SHAPE-34
164
Kidney
Tursiops truncatus
**
SHAPE-36
165
Skin-Blubber
Tursiops truncatus
SHAPE-38
165
Liver
Tursiops truncatus
**
SHAPE-39
165
Kidney
Tursiops truncatus
**
SHAPE-40
165
Melon
Tursiops truncatus
4,4’-DDT
0,4331
0,1369
∑ DDT
4,4’-DDE/∑ DDT
48,4942
0,9280
14,3667
0,9213
SHAPE-41
166
Skin-Blubber
Tursiops truncatus
**
Appendix - 29
Sample Name
Sample ID
Tissue
Species
SHAPE-43
166
Liver
Tursiops truncatus
**
SHAPE-46
180
Skin-Blubber
Tursiops truncatus
SHAPE-48
180
Liver
Tursiops truncatus
**
SHAPE-49
180
Kidney
Tursiops truncatus
**
SHAPE-51
185
Skin-Blubber
Tursiops truncatus
SHAPE-53
185
Liver
Tursiops truncatus
**
SHAPE-54
185
Kidney
Tursiops truncatus
**
SHAPE-56
192
Skin-Blubber
Tursiops truncatus
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0024
< 0,0048
< 0,0048
< 0,0048
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0024
< 0,0048
< 0,0048
< 0,0048
POLYCYCLIC AROMATIC HYDROCARBONS (mg/kg, ww)
Fluorene
Phenanthrene
Anthracene
Fluoranthene
Pyrene
Benz(a)anthracene
Chrysene
Benzo(b)fluoranthene
Benzo(k)fluoranthene
Benzo(e)pyrene
Benzo(a)pyrene
Perylene
Indeno(1,2,3-cd)pyrene
Dibenz(a,h)anthracene
Benzo(g,h,i)perylene
Dibenzo(a,l)pyrene
Dibenzo(a,e)pyrene
Dibenzo(a,i)pyrene
Dibenzo(a,h)pyrene
< 0,0005
< 0,0005
< 0,0005
< 0,0005
0,0021
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0025
< 0,0051
< 0,0051
< 0,0051
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0024
< 0,0048
< 0,0048
< 0,0048
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0023
< 0,0046
< 0,0046
< 0,0046
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0025
< 0,0050
< 0,0050
< 0,0050
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0023
< 0,0047
< 0,0047
< 0,0047
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0025
< 0,0050
< 0,0050
< 0,0050
ORGANOCHLORINE COMPOUNDS (mg/kg, ww)
Hexachlorobenzene
2,4’-DDE
4,4’-DDE
2,4’-DDD
4,4’-DDD
2,4’-DDT
0,0751
0,0741
9,0642
0,1367
1,1334
0,1356
0,0337
0,0313
5,0613
0,0696
0,6145
0,0387
0,0549
0,0385
3,6743
0,0787
0,6597
0,0350
Appendix - 30
Sample Name
Sample ID
Tissue
Species
SHAPE-43
166
Liver
Tursiops truncatus
**
SHAPE-46
180
Skin-Blubber
Tursiops truncatus
SHAPE-48
180
Liver
Tursiops truncatus
**
SHAPE-49
180
Kidney
Tursiops truncatus
**
SHAPE-51
185
Skin-Blubber
Tursiops truncatus
SHAPE-53
185
Liver
Tursiops truncatus
**
SHAPE-54
185
Kidney
Tursiops truncatus
**
SHAPE-56
192
Skin-Blubber
Tursiops truncatus
4,4’-DDT
0,4767
0,1881
0,2347
∑ DDT
4,4’-DDE/∑ DDT
11,0208
0,8225
6,0035
0,8431
4,7209
0,7783
Appendix - 31
Sample Name
Sample ID
Tissue
Species
SHAPE-58
192
Liver
Tursiops truncatus
**
SHAPE-59
192
Kidney
Tursiops truncatus
**
SHAPE-61
193
Skin-Blubber
Tursiops truncatus
**
SHAPE-64
193
Kidney
Tursiops truncatus
**
SHAPE-65
193
Melon
Tursiops truncatus
**
SHAPE-66
203
Skin-Blubber
Tursiops truncatus
SHAPE-69
203
Kidney
Tursiops truncatus
SHAPE-71
218
Skin-Blubber
Stenella coeruleoalba
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0024
< 0,0048
< 0,0048
< 0,0048
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0023
< 0,0046
< 0,0046
< 0,0046
0,0117
0,0280
3,7411
0,0258
0,2159
0,0420
0,0078
0,0431
6,0821
0,0447
0,1375
0,1201
POLYCYCLIC AROMATIC HYDROCARBONS (mg/kg, ww)
Fluorene
Phenanthrene
Anthracene
Fluoranthene
Pyrene
Benz(a)anthracene
Chrysene
Benzo(b)fluoranthene
Benzo(k)fluoranthene
Benzo(e)pyrene
Benzo(a)pyrene
Perylene
Indeno(1,2,3-cd)pyrene
Dibenz(a,h)anthracene
Benzo(g,h,i)perylene
Dibenzo(a,l)pyrene
Dibenzo(a,e)pyrene
Dibenzo(a,i)pyrene
Dibenzo(a,h)pyrene
< 0,0005
< 0,0005
< 0,0005
< 0,0005
0,0009
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0024
< 0,0048
< 0,0048
< 0,0048
< 0,0005
< 0,0005
< 0,0005
< 0,0005
0,0019
0,0017
0,0019
0,0016
0,0014
< 0,0005
< 0,0005
< 0,0005
0,0023
0,0022
0,0021
< 0,0024
< 0,0048
< 0,0048
< 0,0048
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0025
< 0,0050
< 0,0050
< 0,0050
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0024
< 0,0048
< 0,0048
< 0,0048
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0025
< 0,0050
< 0,0050
< 0,0050
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0024
< 0,0049
< 0,0049
< 0,0049
ORGANOCHLORINE COMPOUNDS (mg/kg, ww)
Hexachlorobenzene
2,4’-DDE
4,4’-DDE
2,4’-DDD
4,4’-DDD
2,4’-DDT
0,0192
0,0394
5,6898
0,0389
0,3548
0,0794
Appendix - 32
Sample Name
Sample ID
Tissue
SHAPE-66
203
Skin-Blubber
Tursiops truncatus
SHAPE-69
203
Kidney
Tursiops truncatus
SHAPE-71
218
Skin-Blubber
Stenella coeruleoalba
4,4’-DDT
0,1239
0,0684
0,1065
∑ DDT
4,4’-DDE/∑ DDT
6,3263
0,8994
4,1212
0,9078
6,5340
0,9308
Species
SHAPE-58
192
Liver
Tursiops truncatus
**
SHAPE-59
192
Kidney
Tursiops truncatus
**
SHAPE-61
193
Skin-Blubber
Tursiops truncatus
**
SHAPE-64
193
Kidney
Tursiops truncatus
**
SHAPE-65
193
Melon
Tursiops truncatus
**
Appendix - 33
Sample Name
Sample ID
Tissue
Species
SHAPE-73
218
Liver
Stenella coeruleoalba
**
SHAPE-74
218
Kidney
Stenella coeruleoalba
**
SHAPE-75
218
Melon
Stenella coeruleoalba
**
SHAPE-76
220
Skin-Blubber
Stenella coeruleoalba
SHAPE-78
220
Liver
Stenella coeruleoalba
**
SHAPE-79
220
Kidney
Stenella coeruleoalba
**
SHAPE-81
221
Skin-Blubber
Stenella coeruleoalba
**
SHAPE-83
221
Liver
Stenella coeruleoalba
< 0,0007
< 0,0007
< 0,0007
< 0,0007
< 0,0007
< 0,0007
< 0,0007
< 0,0007
< 0,0007
< 0,0007
< 0,0007
< 0,0007
< 0,0007
< 0,0007
< 0,0007
< 0,0036
< 0,0071
< 0,0071
< 0,0071
< 0,0005
< 0,0005
< 0,0005
< 0,0005
0,0019
0,0016
0,0018
0,0013
0,0014
< 0,0005
< 0,0005
< 0,0005
< 0,0005
0,0018
0,0020
< 0,0023
< 0,0047
< 0,0047
< 0,0047
POLYCYCLIC AROMATIC HYDROCARBONS (mg/kg, ww)
Fluorene
Phenanthrene
Anthracene
Fluoranthene
Pyrene
Benz(a)anthracene
Chrysene
Benzo(b)fluoranthene
Benzo(k)fluoranthene
Benzo(e)pyrene
Benzo(a)pyrene
Perylene
Indeno(1,2,3-cd)pyrene
Dibenz(a,h)anthracene
Benzo(g,h,i)perylene
Dibenzo(a,l)pyrene
Dibenzo(a,e)pyrene
Dibenzo(a,i)pyrene
Dibenzo(a,h)pyrene
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0026
< 0,0051
< 0,0051
< 0,0051
< 0,0005
< 0,0005
< 0,0005
< 0,0005
0,0012
0,0013
0,0006
0,0007
0,0007
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0023
< 0,0046
< 0,0046
< 0,0046
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0023
< 0,0046
< 0,0046
< 0,0046
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0025
< 0,0050
< 0,0050
< 0,0050
< 0,0005
< 0,0005
< 0,0005
< 0,0005
0,0014
0,0014
0,0016
0,0013
0,0013
< 0,0005
< 0,0005
< 0,0005
< 0,0005
0,0022
< 0,0005
< 0,0023
< 0,0047
< 0,0047
< 0,0047
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0024
< 0,0047
< 0,0047
< 0,0047
ORGANOCHLORINE COMPOUNDS (mg/kg, ww)
Hexachlorobenzene
2,4’-DDE
4,4’-DDE
2,4’-DDD
4,4’-DDD
2,4’-DDT
0,0178
0,0459
5,4249
0,0389
0,2909
0,1264
0,0265
0,1602
15,1346
0,1178
0,4109
0,3475
Appendix - 34
Sample Name
Sample ID
Tissue
Species
SHAPE-73
218
Liver
Stenella coeruleoalba
**
SHAPE-74
218
Kidney
Stenella coeruleoalba
**
SHAPE-75
218
Melon
Stenella coeruleoalba
**
SHAPE-76
220
Skin-Blubber
Stenella coeruleoalba
SHAPE-78
220
Liver
Stenella coeruleoalba
**
SHAPE-79
220
Kidney
Stenella coeruleoalba
**
SHAPE-81
221
Skin-Blubber
Stenella coeruleoalba
**
SHAPE-83
221
Liver
Stenella coeruleoalba
4,4’-DDT
0,2296
0,3324
∑ DDT
4,4’-DDE/∑ DDT
6,1566
0,8812
16,5034
0,9171
Appendix - 35
Sample Name
Sample ID
Tissue
Species
SHAPE-84
221
Kidney
Stenella coeruleoalba
**
POLYCYCLIC AROMATIC HYDROCARBONS (mg/kg, ww)
Fluorene
Phenanthrene
Anthracene
Fluoranthene
Pyrene
Benz(a)anthracene
Chrysene
Benzo(b)fluoranthene
Benzo(k)fluoranthene
Benzo(e)pyrene
Benzo(a)pyrene
Perylene
Indeno(1,2,3-cd)pyrene
Dibenz(a,h)anthracene
Benzo(g,h,i)perylene
Dibenzo(a,l)pyrene
Dibenzo(a,e)pyrene
Dibenzo(a,i)pyrene
Dibenzo(a,h)pyrene
< 0,0005
< 0,0005
< 0,0005
< 0,0005
0,0007
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0005
< 0,0025
< 0,0050
< 0,0050
< 0,0050
ORGANOCHLORINE COMPOUNDS (mg/kg, ww)
Hexachlorobenzene
2,4’-DDE
4,4’-DDE
2,4’-DDD
4,4’-DDD
2,4’-DDT
Appendix - 36
Sample Name
Sample ID
Tissue
Species
SHAPE-84
221
Kidney
Stenella coeruleoalba
**
4,4’-DDT
∑ DDT
4,4’-DDE/∑ DDT
Appendix - 37
Annex - Operating project of a listening station at sea for Odontocetes populations and acoustic marine pollution monitoring
Introduzione
L'ambiente marino ha le sue peculiarità acustiche e i cetacei sono straordinariamente ben adattati a questo ambiente. La comunicazione acustica ha acquisito un ruolo privilegiato rispetto ad altri canali di comunicazione in questi animali: essi si affidano infatti al suono per comunicare tra loro, analizzare l'ambiente, per trovare prede ed evitare ostacoli. I cetacei hanno
dunque sviluppato una particolare sensibilità al suono, essendo l’udito il loro senso principale.
Scopi e obiettivi del progetto
Il progetto si propone di realizzare una “stazione di ascolto di Cetacei e di monitoraggio
dell’inquinamento acustico marino”. Il sistema consisterà nella realizzazione di una postazione
acustica fissa, installata presso la torre Posidonia a largo di Francavilla (CH) e di tre postazioni mobili, tutte dotate di specifica strumentazione acustica e collegate via GSM a una workstation a terra, situata presso la Fondazione Mario Negri Sud.
Gli obiettivi generali del progetto sono:
1) identificare la presenza di Cetacei in transito nell’area di interesse e di discriminare le
specie, attraverso l’elaborazione dei dati acustici in arrivo dalla stazione a mare.
2) monitorare l’inquinamento acustico marino generato da attività antropiche di vario tipo e,
quindi, degli effetti di quest’ultimo sui mammiferi marini.
Il progetto nasce dall’esigenza di sopperire ad una carenza di informazioni esistente sul nostro
territorio relativamente a:
•
le popolazioni di cetacei (specie e abbondanza) che vivono lungo la costa centrooccidentale del Mar Adriatico in generale e, nello specifico, lungo la costa abruzzesemolisana;
•
alla valutazione del clima acustico esistente lungo la costa abruzzese e agli eventuali impatti generati dalle attività umane sulle comunità marine, in modo particolare quelli prodotti
dalle attività di petrolizzazione che vedono sempre più coinvolto il nostro territorio negli ultimi anni.
I Mammiferi Marini e, nello specifico i Cetacei, sono inclusi in Appendice I delle liste CITES
(per cui è richiesta massima protezione) e nell’All. II della Direttiva Habitat (92/43/CE) e richiedono pertanto la massima tutela da parte dei paesi aderenti alla Convenzione di Washington e
Annex - 1
membri della Comunità Europea. La Direttiva quadro sulla strategia per l’ambiente marino
(Marine Strategy Framework Directive MSFD) 2008/56/CE, in particolare, fa esplicito riferimento alla presenza di rumore antropogenico in termini di qualità dell’ambiente marino. Gli effetti ambientali generati dal rumore subacqueo sono divenuti, di conseguenza, di rilevanza europea, e gli impatti relativi necessitano di essere accuratamente valutati.
Tutti gli Stati Membri dell’Unione Europea sono obbligati a condurre valutazioni di ciascuno
degli 11 descrittori ambientali elencati nell’Annesso I della Direttiva, tra cui il livello di rumore
(descrittore n. 11), per definire il “buono stato ambientale” dei loro mari entro il 2020.
Da qui l’esigenza di conoscere le popolazioni locali per tutelarle…
I Cetacei dell’Adriatico
Le specie riscontrate in Adriatico sono riportate in Tabella 1.
Tabella I – Specie di cetacei osservati/avvistati nel Mar Adriatico.
Famiglia
Delphinidae
Physeteridae
Balaenopteridae
Nome scientifico
Nome comune
Tipologia di
presenza
Tursiops truncatus
common bottlenose
dolphin
Stanziale
Delphinus delphis
short-beaked common
dolphin
Stanziale
Stenella coeruleoalba
striped dolphin
Stanziale
Grampus griseus
Risso's dolphin
Stanziale
Globicephala melas
long-finned pilot whale
Accidentale
Pseudorca crassidens
false killer whale
Accidentale
Physeter macrocephalus
sperm whale
Stanziale
Balaenoptera physalus
fin whale
Stanziale
Megaptera novaeangliae
humpback whale
Accidentale
Il tursiope è l'unica specie regolarmente presente nelle acque dell'Adriatico. È distribuito in tutto il bacino anche se è stato accuratamente studiato solo in alcune località. Nelle acque italiane è l'unica specie regolarmente osservata, durante gli ultimi 15 anni, nel Golfo di Venezia. E'
attualmente studiato in tre località del mare Adriatico occidentale: attorno all'isola di Vis, in tutto l'arcipelago di Cres-Losinj e al largo della costa della Slovenia.
Annex - 2
Tra i cetacei il tursiope è sicuramente la specie più conosciuta e studiata, grazie alla sua capacità di adattarsi alla vita cattività. Esistono due ecotipi: uno pelagico di dimensioni più grandi, che vive in gruppi numerosi e l'ecotipo costiero, di dimensioni inferiori e di solito solitario.
Le stenelle e balenottere sono avvistate di tanto in tanto, mentre le altre specie possono essere considerate rare. Dati recenti mostrano la presenza occasionale del delfino comune in Damalzia e nel Quarnero (Notarbartolo di Sciara e Demma, 1997) e l'esistenza di una popolazione residente nel Mar Ionio orientale (Politi et al., 1998). Il Grampo (Grampus griseus) è presente principalmente nel Nord Adriatico, dove le acque sono più profonde. Può essere vista
occasionalmente la balenottera comune (Balaenoptera physalus) e il capodoglio (Physeter
macrocephalus). Tuttavia il capodoglio ha bisogno di acque profonde per le sue immersioni
caccia.
Attualmente il Delfino comune è notevolmente raro in Adriatico; vive in gruppi composti da 1020 individui.
Gli impatti del rumore antropico sui Cetacei
Gli effetti generati dall’inquinamento acustico sull’ambiente marino e, in particolare, sui cetacei, sono a livello quantitativo ancora poco conosciuti, ma è senza dubbio un problema sempre più sentito negli ultimi anni. Innumerevoli esperimenti scientifici e diversi incidenti, anche
mortali, come ampiamente riportato dalla bibliografia scientifica, hanno evidenziato una relazione conflittuale fra attività umane rumorose e i cetacei. Il rumore antropico può causare nei
cetacei una perdita della capacità di navigazione e di orientamento, del trovare prede, di riposare, di riprodursi e di svolgere tutte le altre attività essenziali per la sopravvivenza di questi
mammiferi marini. Questi ultimi possono subire effetti negativi variabili da semplici disturbi nelle proprie attività a manifestazioni di stress vero e proprio che si ripercuotono sul sistema endocrino, fino a traumi di carattere fisico con danni a diversi organi in aggiunta al sistema uditivo.
La comunità scientifica è ormai unanime nel ritenere che il rumore prodotto dalle attività umane può avere un pesante impatto sulla qualità di un ambiente naturale, e in taluni casi provocare danni fisici o la morte degli animali nelle vicinanze di sorgenti acustiche di elevata potenza. Questo è particolarmente vero per l’ambiente subacqueo (si guardi in particolare la Direttiva quadro sulla strategia per l’ambiente marino: 2008/56/CE.
In acqua, infatti, il suono si propaga molto meglio che in aria, sia in termini di intensità che di
distanza; il rumore prodotto da alcune attività umane avrà quindi un forte impatto in una vasta
area circostante. A questo scopo si rende necessaria l’applicazione di una serie di azioni volte
ad eliminare o minimizzare il rischio di disturbo arrecato alla fauna marina e, sicuramente, ad
evitare danni fisici diretti. Ogni volta ci si appresti a svolgere attività che producono rumore in
mare, è necessario attivare una serie di procedure che includano lo studio preliminare
dell’ambiente interessato e dei mammiferi marini presenti, il monitoraggio della loro presenza
Annex - 3
e comportamento durante l’emissione di rumore, e le conseguenze a lungo termine sulla popolazione anche dopo la cessazione dell’inquinamento acustico.
Durante lo svolgimento delle attività che causano inquinamento acustico, è fondamentale monitorare l’area interessata da livelli di rumore ritenuti dannosi in modo da bloccarne l’emissione
in caso uno o più animali entrino in tale raggio.
Il limite di pericolosità è stabilito in 180 db re 1 µPa per i cetacei e 190db re 1 µPa per i Pinnipedi, sulla base delle norme già riconosciute ed applicate dal National Marine Fishery Service
del Governo Americano (NMFS, 2000); norme più restrittive (160 db re 1 µPa) possono essere di volta in volta richieste in casi specifici (criticità relative a situazioni, habitat o specie particolarmente vulnerabili). In linea generale è necessario basare procedure e protocolli su un
approccio conservativo che rifletta i livelli di incertezza (best practices, Richardson, 1995): agire applicando il principio di precauzione.
Figura I - Schema dei livelli di sovrapposizione ed interferenza del range di frequenza
generato da varie fonti di rumore antropico con quello emesso dai mammiferi marini.
Annex - 4
Le domande cui questo progetto vorrebbe rispondere sono le seguenti:
•
Qual è il livello di “rumore” attuale lungo la nostra costa?
•
Attività legate all’estrazione di petrolio in mare o altri tipi di attività di quanto aumenterebbero questo livello?
•
Il nuovo range sarebbe compatibile con la vita delle popolazioni naturali di mammiferi marini?
•
Cosa potrebbe essere fatto per ridurre gli impatti?
Fasi del progetto
Le ricerche bioacustiche sui cetacei in mare possono far uso di due tipi di postazioni:
-
mobili, costituite da sistemi di rilevazione acustica passiva “Array” (serie di idrofoni), generalmente a 2 o più canali trainati da un’imbarcazione;
-
fisse, sistemi di registrazione acustica autonomi “Bottom recorder” posizionati sul fondo
marino o su strutture fisse.
Nello specifico, il progetto farà uso di un sistema di registrazione Bottom recorder su struttura
fissa presso la Posidonia srl per quanto riguarda l’obiettivo 1 e su fondo marino per quanto riguarda l’obiettivo 2.
Le fasi del progetto:
1. realizzazione di una stazione fissa multi-idrofonica omnidirezionale (MHODS). Il MHODS
è un'apparecchiatura basata su una specifica strumentazione acustica (sistema con funzioni di idrofono, amplificatore/trasduttore e registratore) collegata ad un sistema di acquisizione dati autonomo, dotato di idrofoni collegati ad un sistema di conversione ad alta risoluzione analogico-digitale con una memoria di massa allo stato solido. Esso assicura un
grado di robustezza adatto per l'ambiente in cui deve operare. Questi dati verranno memorizzati all’occorrenza in una workstation.
2. realizzazione di un sistema di trasmissione dei dati a terra attraverso dispositivi di telecomunicazione mediante telefonia mobile;
3. analisi dei suoni provenienti dagli idrofoni e loro archiviazione in una fonoteca digitale a
terra;
4. descrizione e rappresentazione grafica bidimensionale dei segnali (spettrogrammi: frequenza-tempo) registrati;
5. distribuzione spaziale (tridimensionale) dei dati acquisiti relativamente al clima acustico
(obiettivo 2);
6. analisi della struttura spettrografica dei segnali discretizzati attraverso unità di conversione
ed elaborazione digitale del segnale (Digital Signal Processing, Workstation situata presso
Annex - 5
il FMNS), equipaggiata con idonei dispositivi di conversione numerica dei segnali analogici (ADC e DAC converters ad alta risoluzione: 24 bit). Il sistema di elaborazione dati così
composto sarà in grado di eseguire analisi in tempo reale e la discriminazione delle componenti spettrali, nel dominio della frequenza, e della dinamica nel dominio del tempo, attraverso specifici software e librerie per l'elaborazione;
7. elaborazione dei dati acustici registrati per la valutazione del livello di rumore (obiettivo 2)
e per la comparazione attraverso algoritmi statistici dei dati con quelli di fonoteche di altri
enti di ricerca (obiettivo 1).
8. stima qualitativa della presenza delle varie specie (obiettivo 1) e valutazione degli impatti
del clima acustico sui cetacei (obiettivo 2).
La struttura fissa
La Torre a mare Posidonia è la sede operativa della Società Posidonia a r.l., con sede legale
a Pescara, fondata nel 1992 con l’intento di ideare, progettare, realizzare ed avviare a produzione un innovativo sistema di allevamento ittico in mare.
La torre è situata in mare aperto, a 3 miglia (5,5 km) al largo di Francavilla al Mare (CH), a 4
miglia di distanza dai due porti di Pescara e a 9 miglia dal porto di Ortona. La sede è costituita
da una struttura fissa rappresentata da una grande torre in acciaio e vetroresina (Figura 1),
alta complessivamente 23 m sul livello del mare, adibita a centro di gestione e controllo di un
impianto di acquicoltura, costituito da numerose gabbie e strutture accessorie, sommerse e di
2
superficie, destinate ad ospitare pesci e molluschi. Con un totale di 632 m coperti e protetti
2
distribuiti su 4 piani, e oltre 500 m di terrazzature poste a vari livelli, la torre è in grado di ospitare in modo confortevole per molti giorni numerose persone, anche nel pieno dell’inverno.
Annex - 6
Figura II - La Torre Posidonia.
La struttura (Figura II), dal basso verso l’alto, è così organizzata:
•
due banchine di attracco per imbarcazioni;
•
un primo livello a terrazzo scoperto di circa 50 m posto a m 2,78 sul livello medio del mare (s.l.m.m.);
•
un secondo livello a terrazzo scoperto di circa 90 m posto a m 6,96 s.l.m.m.;
•
un primo piano chiuso ed abitabile di circa 72 m con ampia terrazzatura sui 4 lati, posto a
m 10,10 s.l.m.m.;
•
un secondo piano chiuso ed abitabile di 234 m posto a m 12,75 s.l.m.m.;
•
un terzo piano chiuso ed abitabile di 234 m posto a m 15,40 s.l.m.m.;
•
un quarto piano chiuso ed abitabile di 92 m con ampia terrazzatura sui 4 lati, posto a m
18,05 s.l.m.m.;
•
un ultimo terrazzo di circa 110 m posto a m 20,70 s.l.m.m., con gli attacchi per
un’eventuale piazzola d’atterraggio per elicotteri.
2
2
2
2
2
2
2
La planimetria completa e particolari della struttura sono riportati in Allegato al documento.
Annex - 7
Poiché la struttura è impiantata nel mare territoriale, la sua gestione comporta il pagamento
allo Stato di un canone demaniale annuo di circa 6.400 €. La concessione terminerà nel 2014
e sarà, come da attuali disposizioni di legge, prorogabile a richiesta.
La torre si affaccia su un fondale di 20 metri costituito da un misto di sabbia e fango, in acque
particolarmente ricche di biodiversità. La zona è classificata, da molti anni e ininterrottamente,
come “Zona A” (D.L.vo 30 dicembre 1992, n. 530) dalla Regione Abruzzo, come attestato dalle analisi che l’Autorità Veterinaria Pubblica (A.S.L.) preposta effettua ogni 15 giorni sui campioni di acqua e di molluschi direttamente raccolti sul posto.
L’area di mare è oggi circondata da un allevamento di cozze esteso 110 ha, a sua volta attorniato da una fascia di 1.000 m interdetta alla pesca ed alla navigazione profonda. In totale,
l’area riservata e vietata alla pesca ed alla navigazione a forma quadrata, con lato di km 3,050
con superficie di 930 ha. Le gabbie, appoggiate sul fondo tutt’intorno alla torre, sono state colonizzate da molteplici forme di vita. Infatti, ad una semplice osservazione dalla superficie, è
visibile la massiccia presenza di varie specie ittiche in prossimità delle strutture sommerse. Le
molte tonnellate di cozze in allevamento forniscono all’ambiente circostante una base di avvio
della catena alimentare in quanto ospitano, nutrono e proteggono le piccole forme di vita che
ne costituiscono i primi anelli. Il personale addetto all’allevamento presta una costante sorveglianza a protezione di tutta l’area e tiene lontani curiosi e pescatori.
La struttura attualmente è sotto-utilizzata rispetto alle sue potenzialità, e potrebbe essere ulteriormente dotata di strumentazione per consentire una più vasta gamma di analisi, soprattutto
quelle di immediato intervento e quelle da effettuare su forme di vita le cui caratteristiche biologiche non sopportino il trasporto a terra fino ad un laboratorio lontano. Sarà fondamentale in
questo senso, la collaborazione con il Centro di Scienze Ambientali (CSA) della Fondazione
Mario Negri Sud (CMNS), struttura che possiede la strumentazione, l’esperienza e le competenze necessarie per avviare nuove attività di indagine.
Le caratteristiche possedute dalla struttura la rendono particolarmente qualificata per essere
destinata a scopi scientifici. Disporre di una struttura operativa dislocata direttamente in mare,
apporta notevoli vantaggi sia economici che produttivi, poiché è in grado di accorpare la funzione di indagine e di rilevamento dei dati, con quella di elaborazione e di studio.
E’ sicuramente unica nel contesto mediteranno ed europeo: è sita a 4 miglia dai porti di Pescara, distanza percorsa in soli 10 minuti da qualunque piccola imbarcazione a motore; tali
porti, inoltre, distano poche centinaia di metri dallo snodo autostradale e dalla stazione ferroviaria e 7 km dall’aeroporto. Ne consegue quindi una grande facilità e comodità di comunicazione con il resto d’Italia e d’Europa.
La strumentazione
Il sistema si comporrà di una stazione di ascolto fissa sulla Torre Posidonia e di tre idrofoni
smart mobili (collocabili secondo l’area da monitorare) che permetteranno l’identificazione,
nello spazio, dell’intensità dei segnali rilevati. La distribuzione nello spazio dei segnali acustici
Annex - 8
avverrà attraverso tecniche di triangolazione e l’analisi dell’intensità dei segnali riflessi. Gli idrofoni mobili saranno collocati su boe e la loro alimentazione elettrica sarà garantita da sistemi fotovoltaici.
La strumentazione acustica (Ocean Sonics, Canada) si comporrà dei seguenti elementi:
1.
n. 4 icListen HF 24-bit Broadband Smart Hydrophone (1 fisso + 3 mobili). Caratteristiche tecniche: frequenza 10 Hz - 200 kHz; profondità tra 0 m e 200 m, interfaccia
standard Ethernet.
2.
n. 1 software di pre-elaborazione dati e settaggio parametrico (Lucy software) per
la visualizzazione in tempo reale e la registrazione dei dati in arrivo all’idrofono smart.
Si tratta di una strumentazione specifica utilizzata in vari settori tra cui, appunto, nel monitoraggio acustico ambientale marino in generale (sia di tipo naturale che provocato dalle attività
antropiche) e, nello specifico, nel monitoraggio bioacustico.
Il sistema icListen Smart Hydrophone è un sistema “tutto in uno”, in quanto sostituisce preamplificatore, filtri, convertitori di dati e unità di collegamento con un unico strumento compatto, collegabile direttamente ad un pc (Figura III).
Figura III - Schema del sistema icListen Hydrophone rispetto all’approccio tradizionale.
L’idrofono smart può essere utilizzato semplicemente come idrofono digitale, in tethering (collegamento WI-FI) per la raccolta di dati in tempo reale oppure come data logger in standalone. Può gestire modelli matematici legati al fenomeno acustico discriminale, al fine di ridurre la quantità di dati da trasmettere ed elaborare, attraverso la scelta di uno specifico intervallo
di confidenza. I dati possono essere elaborati in formato acustico (wav) ed in formato spettrale
(trasformazione tramite idoneo software); opzione, quest’ultima, che permette di alleggerire
notevolmente la trasmissione dei dati. La trasmissione continua del formato wav richiede infatti un volume di 15 Mb al secondo, con un segnale di frequenza massima di 200 kHz; la trasmissione di dati spettrali richiede invece solo 20 Kb al secondo, facilmente gestibili via radio
Annex - 9
o via GPRS, laddove la banda a disposizione è più stretta. L’idrofono è raggiungibile anche
attraverso smartphone.
La manutenzione è poi notevolmente agevolata in quanto è in grado di effettuare misurazioni
di temperatura e di umidità al suo interno, allo scopo di informare su eventuali infiltrazioni
d’acqua.
Il software Lucy gestisce i dati raccolti e/o trasmessi dall’idrofono con filtrazione del rumore. In
particolare: individua e misura eventi in real time con un determinato livello di precisione, riproduce i dati registrati, riconosce in automatico l’idrofono collegato, ascolta i dati in formato
wav attraverso la scheda audio del computer, permette la gestione di file wav registrati anche
da altra strumentazione acustica. Il software in modalità realtime registra tutti i dati raccolti e
visualizzati e permette la modifica delle informazioni visualizzate senza modificare le registrazioni originali.
Monitoraggio dell’impatto acustico sui mammiferi marini
Il sistema di monitoraggio si proporrà di:
1. misurare il livello di rumore generato da differenti fonti di inquinamento acustico in mare (attività antropiche);
2. misurare l’impatto generato da tali attività sui cetacei dell’Adriatico;
3. individuare misure necessarie atte a garantire un accettabile livello di protezione.
Essendo queste, in special modo l’acustica, attività estremamente specializzate, si dovrà aver
cura che tali operazioni vengano svolte da personale con chiara competenza nel settore, e dotato di adeguati mezzi tecnici. La serietà e completezza dell’output scientifico di ogni operazione di mitigazione è altresì importante per incrementare il set di dati necessario per meglio
definire ed evolvere le tecniche stesse e i livelli di protezione. A tale scopo i protocolli di raccolta dati ed i formati dei file devono essere il più standardizzati possibile e facilmente consultabili ed esportabili.
Le fonti di inquinamento acustico e, quindi, le attività da monitorare saranno le seguenti:
-
Piattaforme off-shore;
-
Lavori di costruzione costieri e offshore;
-
Navigazione mercantile;
-
Utilizzo di sonar ad alta potenza (militari e civili);
-
Surveys sismici (Oil and Gas, Mineral and Geophysical exploration) e uso di airguns;
-
Esperimenti di playback ed esposizione a sorgenti acustiche;
Annex - 10
-
Altre fonti di impatto quali: navigazione da diporto, whale watching, operazioni di brillamento di residuati bellici, uso di esplosivi per test militari o per lo smantellamento di strutture, sistemi acustici attivi subacquei.
Le azioni previste sono le seguenti:
-
Determinare la presenza/assenza dei cetacei nell’area in prossimità del cantiere (sorgente
del rumore). Le tecniche di monitoraggio dei cetacei dovranno prevedere sia
l’avvistamento che la detezione acustica.
-
Comprendere se le emissioni sonore nelle immediate vicinanze sono compatibili con le
esigenze di protezione di queste specie;
-
Determinare lo stato acustico dell’ambiente durante il periodo di attività (definizione di
“paesaggi acustici” in relazione alle diverse fasi).
-
Attuare eventuali misure di mitigazione.
Il sistema di monitoraggio seguirà le seguenti linee guida redatte dal CIBRA (2011):
1.
Il monitoraggio e la riduzione del rumore subacqueo.
2.
La riduzione dell’impatto del rumore di origine antropica sull’ambiente marino e
sui mammiferi marini, che segue i protocolli specifici in base alle fonti di inquinamento
già implementati dal National Marine Fishery Service (NMFS) del Governo Americano.
I protocolli di monitoraggio acustico standardizzato prevedono misurazioni in determinate posizioni rispetto alla fonte di rumore (varie distanze dal cantiere) al fine di ridurre l’intensità del
rumore da lì proveniente per una più puntuale (eventuale) localizzazione acustica dei cetacei
potenzialmente presenti, al fine di:
•
integrare il monitoraggio visivo della presenza dei cetacei nell’area;
•
arricchire il monitoraggio dei rumori di cantiere con l’acquisizione di informazioni acustiche
specifiche sulla possibile (concomitante) presenza di animali nella zona.
Annex - 11
Costi
Voce
Strumentazione
Quantità
Costo totale
€
icListen HF Smart Hydrophone
4
12.000
58.000
Software pre-elaborazione e settaggio
1
2.000
2.000
PC
1
1.000
1.000
Boe
4
250
1.000
Pannelli fotovoltaici
4
1.000
4.000
Accumulatore amb. marino
4
300
1.200
GSM
4
400
1.600
Noleggio imbarcazione
1
5.000/anno
1
7.000/anno
Tecnico in bioacustica
1
3.500/anno
Segreteria, coordinamento
2
3.500/anno
Ricercatore senior in bioacustica
Personale
Costo
singolo €
Annex - 12

Settore I Sistemi Informatici Servizio AH Sistema informativo U.O.

Sistemi di ancoraggio e legatura UNI EN 12195

Settore I Sistemi Informatici Servizio AH Sistema informativo U.O.

Circolare esplicativa - Confesercenti Grosseto

(c.600-1453) - The British School at Rome

Cirque Du Soleil - feder cral italia

reperti parassitari in cetacei spiaggiati lungo la costa ligure

conto corrente di base isee

Adriatic Sea Forum 2015

MarCal. Calibri