Different levels of mussel (Mytilus edulis) DNA strand breaks following chronic field and acute laboratory exposure to polycyclic aromatic hydrocarbons

Levels of polycyclic aromatic hydrocarbons (PAHs) including benzo[a]pyrene (B[a]P) were at least seven-fold higher in mussels sampled from a polluted site (Loch Leven, in Scotland, UK) compared to a nearby clean reference site (Loch Etive) throughout the year 2000. Levels of DNA strand breaks (alkaline COMET assay) using both gill and digestive gland nuclei were similar at both sites despite the difference in contaminant load (total PAH). In contrast, mussels collected from a reference site (Port Quin, Cornwall, UK) had an increase in DNA strand breaks in digestive gland cells following laboratory exposure to B[a]P-dosed Isochrysis galbana. However, after 14 days high dose (20 ppb-exposed diet) animals had returned to levels similar to the controls. There was no evidence of increased necrosis or apoptosis after treatments. The results from these two studies suggest that an adaptive response may prevent ongoing DNA damage in mussels exposed to high levels of B[a]P and PAH contamination.     

32P-postlabelling analysis of DNA adducts and EROD induction as biomarkers of genotoxin exposure in dab (Limanda limanda) from British coastal waters.

Dab (Limanda limanda) were sampled from a number of polluted and unpolluted areas in British coastal waters. The 32P-postlabelling assay was used to analyse the level of aromatic/hydrophobic DNA adducts in pooled samples of liver tissue. The mean levels of DNA adducts detected from areas known to receive anthropogenic pollutants ranged from 4.0 to 26.8 adducts per 10(8) nucleotides, with all sites containing samples displaying DNA adduct profiles consisting of diagonal radioactive zones. In contrast no DNA adducts were detectable in samples from an unpolluted reference site. The ranking of polluted sites based on DNA adduct levels did not correspond with the ranking of sites based on sediment associated polycyclic aromatic hydrocarbon levels, highlighting the problem of linking the presence of contamination with detectable biological responses. No correlation could be found in this study between EROD activity and the level of DNA adducts.     

Polycyclic aromatic hydrocarbons in surface seawater and in indigenous mussels (Mytilus galloprovincialis) from coastal areas of the Saronikos Gulf (Greece)

Polycyclic aromatic hydrocarbons (PAHs) were identified and measured in surface seawater and in the tissues (gills and mantle) of indigenous black mussels, Mytilus galloprovincialis, collected from three coastal sites of Saronikos Gulf (Greece), a gulf that exhibits high levels of pollution. The total PAHs measured by spectrofluorometry in the surface seawater were found in the range of 425–459 ng L⁻¹ at the most polluted sites 1 and 2 (Elefsis Bay–Salamis Island) and in the range of 103–124 ng L⁻¹ at site 3 (Aegina Island). PAHs' sources in seawater were identified by application of specific PAH ratios, such as phenanthrene/anthracene and fluoranthene/pyrene. Levels of PAHs in soft tissues (gills and mantle) of indigenous mussels were much higher than those reported for seawater. Total PAH concentrations in mantle tissues were in the range of 1300–1800 ng g⁻¹ dry weight (dw) tissue at sites 1 and 2 and approximately 380 ng g⁻¹ dw at site 3. In gill tissues total PAH concentrations were in the range of 1480–2400 ng g⁻¹ dw at sites 1 and 2 and approximately 430 ng g⁻¹ dw at site 3. PAHs composition was dominated by two-, three- and four-ring compounds in seawater, where 17 different PAH compounds were identified and measured in mussel tissues. Mussels can be used as sentinel organisms to monitoring PAHs' contamination, since they concentrate PAHs from the surrounding water media and therefore making the chemical analysis simpler and less prone to error than that for water. In surface seawater possible weathering and photodegradation due to hot climates contribute to reduced PAHs concentrations.     

Evaluation of neutral red retention assay,micronucleus test,acetylcholinesterase activity and a signal transduction molecule (cAMP) in tissues of Mytilus galloprovincialis (L.), in pollution monitoring

The neutral red lysosomal retention assay (NRR) of the haemocytes, and the acetylcholinesterase activity (AChE) in the haemolymph, the digestive gland, the gills and the mantle/gonad complex have been evaluated on mussels Mytilus galloprovincialis collected from Thermaikos and Strymonikos gulfs (northern Greece) in June and October 2001. The validity of performing the above core biomarkers is supported, firstly by their ability to respond to different pollution levels and, secondly, by the significant linear correlation among them. The evaluation of the micronuclei frequency (MN) has been performed in gill tissue and haemocytes of the same mussels and, according to the results, it needs more research in order its use as stress indices to be validated. In addition, the first results on cAMP levels in the gills, the mantle/gonad complex and the digestive gland, whose concentrations correlated to both, NRR and AChE introduce this signal transduction molecule as a new, promising biomarker.     

Immuno-localisations (GSSP) of subcellular accumulation sites of phenanthrene, aroclor 1254 and lead (Pb) in relation to cytopathologies in the gills and digestive gland of the mussel Mytilus edulis

Cell and tissue pathology of both, gill and digestive tissue, has been the subject of many studies for the elucidation of contaminant-induced biological effects. In the present study, cellular pathological alterations were linked to subcellular sites of chemical accumulation in gills and digestive gland tissues. For this purpose, mussels (Mytilus edulis) were exposed to the organic contaminants aroclor 1254 (PCB) (20 microg/L), phenanthrene (PAH) (150 microg/L) or the metal lead (Pb) (2.5mg/L). The localization of chemicals at the subcellular level was analysed by an antibody-based detection system (GSSP) by the use of commercially available antibodies specifically directed against the chemicals. Pathological changes were analysed in parallel in identical samples by transmission electron microscopy. After exposure to the different contaminants, cell organelles such as mitochondria, the endo-lysosomal system as well as endoplasmic reticulum showed clear evidence of chemically-induced alterations. Large numbers of crystalloid inclusions were found in mitochondria and in autophagic lysosomes as well as multi-lamellated whorls after PAH and aroclor exposure. Immunocytochemical detection of the chemicals showed their accumulation inside of various cell organelles such as lysosomes, mitochondria, and nuclei. Additionally, chemicals were localized in association to membranes, cilia and microvilli of gill and digestive gland cells. Furthermore, the chitinous rod and mucus secretions of gill epithelial cells were positively labelled for contaminants indicating their role in protection. Localization of contaminants by immuno-detection in combination with pathological diagnosis gives insights into the cellular targets of chemical attack.     

Phase I and II biotransformation enzymes and polycyclic aromatic hydrocarbons in the Mediterranean mussel (Mytilus galloprovincialis,Lamarck, 1819) collected in front of an oil refinery

The aim of the present study was to investigate the responses of phase I and II biotransformation enzymes and levels of PAHs in the Mediterranean mussel (Mytilus galloprovincialis, Lamarck, 1819) collected from three sites at different distance from an oil refinery. Phase I enzyme activities as NAD(P)H-cyt c red, NADH ferry red, B(a)PMO and phase II as UDPGT, GST were measured in digestive gland while 16 PAHs (US-EPA) in whole soft tissue. An added value to the data obtained in the present study rely on the RDA analysis which showed close correlations between PAHs levels and phase I enzyme activities in mussels collected in front of the refinery. And again a significant spatial correlation between B(a)P levels and NADPH-cyt c red activities was observed using linear models. No differences among sites for B(a)PMO and phase II GST activities were observed, while the application of UDPGT as biomarkers requires further investigation.     

In vitro mechanistic differences in benzo[a]pyrene-DNA adduct formation using fish liver and mussel digestive gland microsomal activating systems

Turbot (Scophthalmus maximus) and mussel (Mytilus edulis) microsomes were incubated with DNA to examine if microsomal in vitro metabolism of BaP could result in DNA adducts detected by 32P-postlabelling. Turbot DNA was incubated with benzo[a]pyrene (BaP), NADPH and microsomal activating systems prepared from either livers of unexposed turbot, turbot exposed to BaP or beta-naphthoflavone (beta-NF) or digestive glands from mussels. The beta-NF activating system generated the highest levels of DNA adducts detected in this study (451.7 adducts per 10(8) nucleotides) and were distributed in three discrete adduct TLC spots, one of which (97% of the total adducts) co-migrated with the 32P-postlabelled BaP 7,8-diol, 9,10-epoxide-N2-guanine adduct. Fewer adducts (P < 0.05) were generated by BaP-induced microsomes (9.4-30.6 adducts per 108 nucleotides) but levels were higher (P <0.05) than those generated from untreated fish (3.5 adducts per 10(8) nucleotides). Co-incubation with 500 microM alpha-naphthoflavone (alpha-NF) resulted in 97-99% inhibition in adduct formation implicating cytochrome P450-dependent (CYP) bioactivation however there was some evidence for carry over of BaP in the liver microsomal preparations from BaP injected fish. In contrast to the fish activating systems, no DNA adducts were observed when mussel microsomes were incubated with BaP, DNA and NADPH.     

Enhancement of polycyclic aromatic hydrocarbons in estuarine invertebrates by surface runoff at a decommissioned military fuel depot

Accumulation of polycyclic aromatic hydrocarbons (PAHs) was determined in blue mussels (Mytilus spp.) and shore crabs (Hemigrapsus sp.) at a recently closed military fuel depot in central San Francisco Bay, California. In April 1996, during a period of above average precipitation, specimens were collected at the depot, near the depot, and at sites 10 and 20 km south of the depot. Four weeks after the rains ended, blue mussels were again collected at the depot, and at two additional sites in the central Bay region. In April, total PAHs in mussels from the depot were significantly higher only than that in mussels collected 20 km from the depot; however, seven specific, substituted PAHs were higher at the depot than at all other sites. In June, only two of the 38 PAHs common in mussels in April were detected at the depot; these concentrations were comparable to ambient concentrations in mussels at the Bay. It seemed that bioavailability of PAHs at the depot was enhanced by rainfall, probably due to the mobilization of PAHs via groundwater into the Bay. Concentrations in mussels from chronically contaminated sites were about five times higher than mussels collected from the depot. Low PAH concentrations were detected in shore crabs near the depot, and the highest levels were not associated with the depot. Observed PAH concentrations are discussed in relation to upper trophic organisms.     

Evidence of uptake, biotransformation and DNA binding of polyaromatic hydrocarbons in Atlantic cod and corkwing wrasse caught in the vicinity of an aluminium works

Feral Atlantic cod (Gadus morhua) and corkwing wrasse (Symphodus melops) were investigated for polyaromatic hydrocarbon (PAH) contamination in the Karmsund strait, western Norway. This strait is highly contaminated with PAHs, and a main source is the chronic release of gas-scrubbing effluents from a local aluminium works. In both species, the level of biliary PAH metabolites and hepatic DNA adducts were higher in fish collected near the aluminium works. Interestingly, a significantly higher level of both biliary PAH metabolites and hepatic DNA adducts was found in corkwing wrasse as compared to cod, indicating a higher potential for genotoxic effects in this species. Hepatic cytochrome P4501A (CYP1A) in cod estimated by ethoxyresorufin-O-deethylase and an immunoassay technique (ELISA), seemed to be weakly induced at the contaminated sites. At the most contaminated site, skin ulcers and fin erosion were detected in about 70 and 45% of the cods, respectively. The data demonstrated that both cod and corkwing wrasse may be suitable target species for PAH pollution monitoring.     

Polycyclic aromatic hydrocarbon (PAH) burden of mussels (Mytilus sp.) in different marine environments in relation with sediment PAH contamination, and bioavailability

Sediments and mussels (Mytilus edulis, Mytilus galloprovincialis) were sampled in different European coastal environments (Germany, France, Spain) and analysed for their polycyclic aromatic hydrocarbon (PAH) content by Gas Chromatography/Mass Spectrometry (GC-MS). Bioaccumulation factors of individual compounds from the sediment were calculated and discussed according to the compound solubility. The mussels showed different accumulation patterns according to the pollution source they were exposed to (dissolved fraction of PAHs, particulate fraction, petroleum present in the water column). The exposure source also depends on the geographical location of the mussels. In the Mediterranean Sea, the bivalves were mainly exposed to the dissolved fraction of PAHs, while in the Baltic Sea and in the Atlantic Ocean, the PAHs associated to the particles were significant sources.     

In vitro mechanistic differences in benzo[a]pyrene–DNA adduct formation using fish liver and mussel digestive gland microsomal activating systems

Turbot (Scophthalmus maximus) and mussel (Mytilus edulis) microsomes were incubated with DNA to examine if microsomal in vitro metabolism of BaP could result in DNA adducts detected by ³²P-postlabelling. Turbot DNA was incubated with benzo[a]pyrene (BaP), NADPH and microsomal activating systems prepared from either livers of unexposed turbot, turbot exposed to BaP or β-naphthoflavone (ß-NF) or digestive glands from mussels. The β-NF activating system generated the highest levels of DNA adducts detected in this study (451.7 adducts per 10⁸ nucleotides) and were distributed in three discrete adduct TLC spots, one of which (97% of the total adducts) co-migrated with the ³²P-postlabelled BaP 7,8-diol, 9,10-epoxide-N²-guanine adduct. Fewer adducts (P <0.05) were generated by BaP-induced microsomes (9.4–30.6 adducts per 10⁸ nucleotides) but levels were higher (P <0.05) than those generated from untreated fish (3.5 adducts per 10⁸ nucleotides). Co-incubation with 500 μM α-naphthoflavone (α-NF) resulted in 97–99% inhibition in adduct formation implicating cytochrome P450-dependent (CYP) bioactivation however there was some evidence for carry over of BaP in the liver microsomal preparations from BaP injected fish. In contrast to the fish activating systems, no DNA adducts were observed when mussel microsomes were incubated with BaP, DNA and NADPH.     

多环芳烃化合物对栉孔扇贝组织芳烃羟化酶活力的影响

研究了苯并(a)芘(B[a]P)、苯并(k)荧蒽(B[k]F)以及两者的混合物对栉孔扇贝(Chlamys farreri)鳃丝和消化盲囊芳烃羟化酶(AHH)活力的影响。结果表明,多环芳烃化合物(PAHs)对栉孔扇贝鳃丝和消化盲囊AHH活力具有显著的诱导作用(P<0.05),并且呈现了明显的时间、剂量效应特征。B[a]P和B[k]F混合物处理组鳃丝和消化盲囊的AHH活力在实验期间呈现先升高后稳定趋势,B[a]P处理组则表现峰值变化,然后呈下降趋势。消化盲囊各处理组AHH活力显著高于鳃丝。作者认为可以采用消化盲囊的AHH活力来反映周围环境PAHs的污染,并应以最大饱和浓度为标准。3种PAHs中B[a]P的毒性最强,PAHs对栉孔扇贝的毒性大小与PAHs的组成有关,不能以不同种类单一PAHs毒性的总和来代表PAHs混合物的毒性。     

Measurement of stress effects (scope for growth) and contaminant levels in mussels (Mytilus edulis) collected from the Irish Sea

The objective of this research was to quantify the impact of pollution along the coastlines of the Irish Sea. Pollution assessment was based on the combined measurement of scope for growth (SFG), and chemical contaminants in the tissues of mussels (Mytilus edulis) collected from 38 coastal sites around the Irish Sea during June-July in 1996 and 1997. On the UK mainland coast, the SFG showed a general trend with a significant decline in water quality in the Liverpool and Morecambe Bay region. High water quality was recorded along the west coast of Wales, as well as southwest England and northwest Scotland (clean reference sites outside the Irish Sea). Along the coast of Ireland there was a similar trend with reduced SFG within the Irish Sea region. SFG was generally low north of Duncannon and then improved north of Belfast. The poor water quality on both sides of the Irish Sea is consistent with the prevailing hydrodynamics and the spatial distribution of contaminants associated with urban/ industrial development. The decline in SFG of mussels on both sides of the Irish Sea was associated with a general increase in contaminant levels in the mussels. Certain contaminants, including PAHs, TBT, sigmaDDT, Dieldrin, gamma-HCH, PCBs, and a few of the metals (Cd, Se, Ag, Pb), showed elevated concentrations. Many of these contaminants were particularly elevated in the coastal margins of Liverpool Bay, Morecambe Bay and Dublin Bay. A quantitative toxicological interpretation (QTI) of the combined tissue residue chemistry and SFG measurements indicated that at the majority of coastal sites, c. 50 to > 80% of the observed decline in SFG was due to PAHs as a result of fossil fuel combustion and oil spills. TBT levels were highest at major ports and harbours, but these concentrations only made a minor contribution to the overall reduction in SFG. At no sites were individual metals accumulated to concentrations that could cause a significant effect on SFG. The study identified many sites where the observed reduction in SFG was far greater than predicted from the limited number of chemical contaminants analysed, thus indicating the presence of additional 'unknown toxicants'. Sewage (containing domestic, agricultural and industrial components) appears to be an important contributor to reduced SFG and linear alkylbenzenes (LABs) and As may provide suitable 'sewage markers'. There was a highly significant positive correlation between SFG and As (P < 0.001). This relationship may be due to reduced As uptake by algal food material and mussels at sites with elevated P04 concentrations (e.g. at sites with sewage inputs). Phosphate is a known competitive inhibitor of As accumulation, at least in algae. The results highlight that further research is required on 'sewage markers' in mussels. The SFG approach therefore provides a rapid, cost-effective and quantitative measure of pollution impact, as well as a means of identifying the causes through a QTI of tissue contaminants levels. It also serves to identify the presence of unidentified toxicants and areas that require further study.     

Seasonal variation of histopathological and histochemical markers of PAH exposure in blue mussel (Mytilus edulis L.)

The aim of this work was to study seasonal variation of histopathological and histochemical markers in blue mussels (Mytilus edulis L.) exposed to pyrogenic PAH contaminants. Mussels were collected in January, June, September and October from a sampling site in the vicinity of the discharge from an aluminium smelter and from a clean reference site. Histopathological analysis was carried out on the digestive gland (DG) and the gonads, lipofuscin and neutral lipids were analysed in the DG. Clear responses in lipofuscin and neutral lipids were detected in the DG of mussels collected from the polluted site at some sampling times. Moreover, these mussels presented atrophy in digestive tubules and haemocytic aggregates in the gonad and DG. However, in all parameters studied, the magnitude of the response showed clear seasonal variation.     

Effects of environmental pollution in caged mussels (Mytilus galloprovincialis)

Biological effects of environmental pollution, mainly related to presence of PAHs, were assessed in mussels Mytilus galloprovincialis caged in Priolo, an anthropogenically-impacted area, and Vendicari, a reference site, both located along the eastern coastline of Sicily (Italy). PAHs concentration and histopathological changes were measured in digestive gland tissues. Expression of cytochrome P4504Y1 (CYP4Y1) and glutathione S-transferase (GST), indicative of xenobiotic detoxification, and activity of catalase (CAT) as oxidative stress index, were evaluated.The results show a direct correlation between the high concentrations of PAHs in digestive glands of mussels from Priolo and the significantly altered activity of phase I (P < 0.001) and phase II (P < 0.0001) biotransformation enzymes, along with increased levels of CAT activity (P < 0.05). These findings show the enhancement of the detoxification and antioxidant defense systems. The mussel caging approach and selected biomarkers demonstrated to be reliable for the assessment of environmental pollution effects on aquatic organisms.     

Use of the Comet assay to assess DNA damage in hemocytes and digestive gland cells of mussels and clams exposed to water contaminated with petroleum hydrocarbons

Oil spills can result in the deposition of large quantities of petroleum hydrocarbons into intertidal and shallow waters seriously impacting bivalve populations. Petroleum hydrocarbons are enriched in polycyclic aromatic hydrocarbons (PAH) and PAH analogs many of which may have potential to damage DNA. The Comet assay is useful for assessing DNA damage and has been used to a limited degree with aquatic organisms, but mostly with studies in vitro. We have carried out studies with the Comet assay to assess the DNA damaging potential of complex mixtures of petroleum hydrocarbons for bivalves. Experiments were carried out with mussels (Mytilus edulis) and clams (Mya arenaria) with dispersions and water soluble fractions of an Arabian crude oil which was also chemically characterized in detail by GC-MS. Pilot studies were first conducted to evaluate test performance and reproducibility. An interindividual coefficient of variation ranging from 17 to 30% was established for the assay with hemocytes and digestive gland cells of both species. Exposure to hydrocarbon fractions had no significant impact on clams. However, an increase in DNA damage was observed at P < 0.1 with digestive gland cells of mussels exposed to aqueous fractions of a light crude oil. These studies have demonstrated a potential for DNA damage in bivalves exposed to oil spills in inshore waters as well as potential for interspecies sensitivity.     

DNA strand breakage in mussels (Mytilus edulis L.) deployed in intertidal and subtidal zone in Reykjavík harbour

DNA single-strand breaks were analysed in the blue mussel (Mytilus edulis L.) deployed in intertidal and subtidal zones in the PAH contaminated Reykjavík harbour and at a reference site, Hvalfj?rethur, Iceland. DNA strand breaks were analysed by Comet assay in isolated gill and haemocyte cells from six mussels from each site and depth. Increased DNA damage in both gill cells and haemocytes were observed in mussels deployed in Reykjavík harbour compared to the reference site. Intertidal mussels from Reykjavík harbour had higher DNA damage in haemocytes compared to subtidal mussels. The Comet assay seems to be useful for measuring genotoxic exposure in mussels from the field, and that DNA damage might be higher in the intertidal zone either due to higher exposure to contaminants or because of physiological and biochemical responses to variations in oxygen availability.     

Anthropogenic pollution stimulates oxidative stress in soft tissues of mussel <Emphasis Type="Italic">Crenomytilus grayanus</Emphasis> (Dunker1853)

The digestive gland and gills of the mussel Crenomytilus grayanus extracted from three locations — (i) sampled from a clean and (ii) polluted site and (iii) transplanted from the nonpolluted to polluted site - were analysed for antioxidant enzymes (superoxide dismutase, catalase, glutathione reductase), total oxyradical scavenging capacity and levels of lipid peroxidation products (malondialdehyde, conjugated dienes and lipofuscin). Perturbation of redox status was found in both digestive gland and gill tissues of mussels living in the polluted area. As the activities of superoxide dismutase and catalase were 1.2–3 times higher, the total oxyradical scavenging capacity was lower by 20–35% and the levels of lipid peroxidation products were 2–7 times higher compared to mussels from the reference site. In transplanted mussels, the lipid peroxidation process in both tissues was significantly stimulated (the level of conjugated dienes was increased 1.7–2.5-fold; malondialdehyde and lipofuscin contents were increased 3.5–5-fold) and the total oxyradical scavenging capacity fell by 50–70%. In addition, the transplantation generally resulted in transient and variable responses of antioxidant enzymes for both tissues. Complex response-behaviour of the antioxidant enzymes strongly points to the necessity of employing a combined approach that takes into account activities of antioxidant enzymes and the total oxyradical scavenging capacity, as well as measurement of oxidative damage (e.g., lipid peroxidation) to evaluate the physiological health of molluscs.     

The PAH composition of surface sediments from Stagnone coastal lagoon, Marsala (Italy)

This paper examines the presence, distribution, nature and sources of 22 Aromatic Hydrocarbons (PAHs), which are important environmentally and toxicologically, in sediments from the Stagnone coastal lagoon at Marsala (Italy). Analysis was performed by gas chromatography/mass spectrometry (GC/MS) with selected ion monitoring (SIM). The total concentration of polycyclic aromatic hydrocarbons ranged from 72 to 18381 μg/kg of dry matrix. The relative standard deviation (RSD) of the replicates on the concentrations of individual compounds ranged from 5% to 20%. The accuracy of method was estimated by analyzing “blank” samples added of known quantities of analytes and the recover percentage was 88 ± 9%. The detection limit (LOD) of analytical procedure was less than 0.2 μg/kg d.w. for all analytes. The quantification limit (LOQ) of analytical procedure was less than 0.7 μg/kg d.w.The resulting distributions and weight ratios of specific compounds are discussed in terms of sampling location and origin of organic matter. A comparison with other studies of total PAHs suggests that the levels are within the concentration ranges already reported by other authors. From an eco-toxicological point of view, total PAH concentrations at seven out of the eight sites studied represent a relatively clean environment when compared to other sites.Organic matter content and PAH concentrations were found to be correlated and the compounds present in Stagnone sediments were shown to be mainly of pyrolitic origin, while a negligible quantity of PAHs may derive from biogenic sources since all the sediments contain perylene traces. There is no evidence of coal-tar contamination.Cluster analysis was carried out in order to discriminate between different PAH origins.     

Histopathology of mussels (Mytilus sp.) from the Tamar estuary, UK

This study assessed numerous histological parameters as markers of health status in mussels (Mytilus spp.) collected from several locations along a contamination gradient on the River Tamar and two coastal sites on the north and south coast of Cornwall, UK. Twenty-seven health parameters were assessed including the presence of pathogens, inflammatory lesions, non-specific pathologies and reproductive condition. Logistic regression showed that reproductive condition and inflammatory lesions showed changes indicative of a contamination gradient response. Mussel populations further upstream within the Tamar were relatively less developed compared to mussels situated downstream in the estuary. Compared to coastal locations, mussels sampled within the Tamar also exhibited a higher prevalence of inflammatory lesions that increased at locations situated furthest upstream. Similarly, levels of lipofuscin within the kidney epithelium were highest in mussels sampled within the estuary and Whitsand Bay compared to Trebarwith Strand. The most prevalent pathogens observed during this survey included Rickettsia/Chlamydia-like organisms (R/CLO), gill ciliates including Ancistrum mytili, the copepod Mytilicola intestinalis, Ciliophora-like organisms and Marteilia sp. Marteilia sp. was observed in 31% of the mussels sampled from the Cremyll Ferry site. Generally, the range and prevalence of pathogens was highest from the two estuarine locations at Cremyll Ferry and Wilcove. No significant differences were observed between estuarine sampling sites and Whitsand Bay with respect to median epithelial cell height of digestive tubules, however mussels from Trebarwith Strand did exhibit a reduced median epithelial cell height compared to mussels sampled from other sampling sites. Species genotyping was also undertaken during this study in order to consider any histological differences in relation to species. Results are discussed in relation to histopathology health parameters and biological effects monitoring.