Combined use of Nassarius reticulatus imposex and statolith age determination for tracking temporal evolution of TBT pollution in the NW Portuguese continental shelf

Spatial and temporal trends of tributyltin (TBT) pollution on the northern continental shelf of Portugal (5-34 m depth) were assessed using Nassarius reticulatus as a bioindicator. Imposex levels were determined in 2006 and 2010, and integrated with age readings from statoliths for a better chronological analysis of the data. Females affected with imposex were detected in 84% of the sites surveyed in 2006, with levels higher than the Ecological Quality Objective (EcoQO) set by the OSPAR Commission for N. reticulatus (VDSI < 0.3) in approximately half of those sites. In 2010, imposex was only detected in 20% of the surveyed sites and, when detected, the VDSI were lower than the EcoQO, except for one location. The comparison between the current results and the levels previously reported for 2004 and 2005 in the same offshore area evidences a clear decline of imposex in 2010 that confirms the efficacy of Regulation (EC) No. 782/2003 and the IMO AFS-Convention (entered into force in 2003 and 2008, respectively). Moreover, the age of the whelks used in the monitoring is estimated in 3-5 years, indicating that actual TBT water contamination in 2010 may in fact be lower than that extrapolated from the imposex levels. We conclude that the legislation was indeed very effective in reducing TBT pollution in the NW Portuguese continental shelf, not only in terms of the magnitude but also in terms of fast field repercussion.     

An innovative statistical approach for analysing non-continuous variables in environmental monitoring: assessing temporal trends of TBT pollution

The current work presents an innovative statistical approach to model ordinal variables in environmental monitoring studies. An ordinal variable has values that can only be compared as "less", "equal" or "greater" and it is not possible to have information about the size of the difference between two particular values. The example of ordinal variable under this study is the vas deferens sequence (VDS) used in imposex (superimposition of male sexual characters onto prosobranch females) field assessment programmes for monitoring tributyltin (TBT) pollution. The statistical methodology presented here is the ordered logit regression model. It assumes that the VDS is an ordinal variable whose values match up a process of imposex development that can be considered continuous in both biological and statistical senses and can be described by a latent non-observable continuous variable. This model was applied to the case study of Nucella lapillus imposex monitoring surveys conducted in the Portuguese coast between 2003 and 2008 to evaluate the temporal evolution of TBT pollution in this country. In order to produce more reliable conclusions, the proposed model includes covariates that may influence the imposex response besides TBT (e.g. the shell size). The model also provides an analysis of the environmental risk associated to TBT pollution by estimating the probability of the occurrence of females with VDS ≥ 2 in each year, according to OSPAR criteria. We consider that the proposed application of this statistical methodology has a great potential in environmental monitoring whenever there is the need to model variables that can only be assessed through an ordinal scale of values.     

Factors affecting RPSI in imposex monitoring studies using Nucella lapillus (L.) as bioindicator

Nucella lapillus (L.) is a marine gastropod mollusc widely used as a bioindicator of TBT pollution in the North Atlantic coastlines. The species reproductive cycle and the male penis length seasonal/spatial variation were studied at a single site at Aveiro seashore (NW Portugal) between December 2005 and June 2007. The main objective of this work is to assess if the "Relative Penis Size Index" (RPSI)--an important imposex assessment index--varies seasonally and spatially in the same sampling site and how this can affect results obtained in imposex monitoring programmes. Animals able to reproduce were found every month but a seasonal pattern in N. lapillus reproductive cycle was evident. Female gametogenic maturation varied seasonally and a decrease in capsule gland volume and condition factor occurred in late summer/early autumn. The gametogenesis in males did not show a significant seasonal variation as in females but the condition factor, penis length, amount of sperm and prostate volume also diminished in late summer/early autumn. On the other hand, males that were close to egg capsules clusters had larger penises than those far away from clusters. The temporal and spatial male penis length variation introduces a bias on imposex assessment results when using RPSI and the magnitude of the error involved is evaluated for different TBT pollution levels scenarios. We consider that RPSI provides interesting and complementary information that should not be excluded from monitoring programmes, but temporal or spatial comparisons of imposex should be based on other more reliable imposex indices like the VDSI.     

The effects of organotin on female gastropods

Female masculinisation (FM) in gastropods (imposex, intersex and ovo-testis) has been documented in > 260 species globally. Biomonitoring studies use FM to indirectly assess the presence of tributyltin (TBT), a once popular biocide in antifouling paint, which has since been regulated. Laboratory studies confirm that FM in gastropods can be induced not only by TBT, but other tin (Sn) compounds, Sn "cocktails", steroids, and other contaminants such as Aroclor 1260. Even though FM in gastropods in the field has been attributed to TBT from boating activity, there is evidence that imposex occurred prior to the advent use of TBT in the 1960s, and for some species imposex occurs naturally. There are roughly 42 species that do not elicit a FM response even if they are exposed to TBT under laboratory conditions, collected sympatrically with other species displaying FM, and/or have TBT in their tissues. A geographic analysis of field studies in four regions indicates that the bulk of studies were conducted in Europe > Asia and Oceania > North America > Latin America. More data are needed regarding FM in Africa, the Middle East, and Latin America since data regarding the masculinisation of female gastropods in these areas are lacking. The most studied gastropod species used in TBT biomonitoring studies is the dogwhelk, Nucella lapillus. This review summarizes the progression of the literature from 1962 to 2009 outlining how the terminology, science, and theories have evolved over time.     

A temporal and spatial monitoring of organotin pollution in a harborside region of Brazil by imposex and ecological quality ratio using <Emphasis Type="Italic">Leucozonia nassa</Emphasis>

This study represents the first use of Leucozonia nassa (Mollusca-Gastropoda) in a program of long-term monitoring. Specimens of L. nassa were collected between 2006 and 2014 and analyzed for the occurrence of imposex in two areas in Southeastern Brazil, one control area and another characterized by the presence of many ports and marinas. Imposex was evaluated in nine sites based on the percentage of females exhibiting imposex (I%), relative penis length index and vas deferens sequence index, classical indices to evaluate tributyltin (TBT) impacts. Moreover, this study uses for the first time in a tropical environment the ecological quality ratio (EQR), an approach to verify the imposex levels as an indicator of the impact by organotin compounds according to the EU Water Framework Directive. The results confirm the occurrence of high levels of imposex in L. nassa with all the indices evaluated. Furthermore, the results also indicate that, despite the international ban, antifouling paints based on TBT are still possibly used in Brazil. In addition, this study shows that L. nassa has the characteristics that make it a suitable bioindicator to monitor TBT pollution, and that the use of the EQR may create important data regarding this problem.     

Imposex and butyltin concentrations in Bolinus brandaris (Gastropoda: Muricidae) from the northern Tunisian coast

The imposex incidence and butyltin concentration i.e. tributyltin with its di- and mono-substituted metabolites were investigated in the muricid Bolinus brandaris sampled from two sites on the northern Tunisian coast (the Lagoon of Bizerta and the small Gulf of Tunis). Both populations had imposex, with stages of imposex development varying between VDS 1 and VDS 4.3. All imposex indices (imposex frequency (I %), female penis length, female vas deferens length, vas deferens sequence index, relative penis length index, and vas deferens length index) were significantly higher in snails from the Bizerta lagoon. Butyltins were detected in the whole tissues of both sexes from the two sites. TBT levels were higher in gastropods collected from the lagoon of Bizerta (12.65 ±1.48 ng Sn g^???1 dw in female and 15.21 ±1.13 ng Sn g^???1 dw in male) than in individuals from the Gulf of Tunis (10.71 ±1.26 ng Sn g^???1 dw in female and 11.65 ±1.63 ng Sn g^???1 dw in male), corroborating the data of imposex analysis. These results confirmed that B. brandaris could be used as a bioindicator species of butyltin pollution in the studied areas. In addition, this study provided baseline data that could serve for long-term monitoring of TBT pollution in Tunisia, since legislation to reduce the use of TBT-based antifouling paints has not been introduced yet.     

Organotin contamination in South American coastal areas

Organotin compounds (OTs) were used in antifouling paints for more than four decades. However, due to their widespread intensive use and high toxicity, undesirable effects in non-target marine organisms have been detected since the early 1980s. Consequently, the International Maritime Organization banned new maritime applications of these products on January 1, 2003 and their presence on ship hulls from January 1, 2008. Although extensively studied in Europe, North America, Oceania, and Asia, environmental levels and effects of organotin contamination are still poorly known for South America. Thus, the current review aimed to present the actual status of this problem in South America by summarizing and comparing the available data in the literature. An overview of the OTs concentrations in sediment and biota and their effects, mainly imposex in marine gastropods, are presented. This work showed that in Atlantic coastal areas of South America there are “hot spots” of OTs contamination, similar to that observed in industrialized countries of Northern Hemisphere. On the other hand, the number of accomplished studies in the Pacific coast is extremely low. Despite the limitation on studies about OTs environmental levels and their related effects, the available data pointed out for a widespread TBT contamination along the South American coastal areas. Therefore, the establishment of baselines of organotin contamination in the Pacific coast and the implementation of temporal trend studies in the South American coastal areas is crucial to verify the effectiveness of local regulations and OTs global ban, and to map the most sensitive areas related to present and future antifouling impacts.     

Effect of freezing on the length of the penis in Nucella lapillus (L.)

When quantifying imposex in Nucella lapillus, two indices are used: the Vas Deferens Sequence Index (VDSI) and the Relative Penis Size Index (RPSI). Freezing and thawing increase the length of the penis in both male and female Nucella lapillus. In the population studied, this had no significant effect on the RPSI, but was potentially an important source of additional variance in the estimation of the mean penis length.     

Butyltin levels in several Portuguese coastal areas

This work aimed to report present levels (2007–2008 sampling) of tri- (TBT), di- (DBT), and monobutyltin (MBT) in surface sediments from 11 Portuguese coastal sites and discuss the evolution of BTs contamination in the last two decades. All the samples revealed quantifiable values of TBT, DBT, and MBT with total butyltin concentrations between 1 and 565 ng/g (of Sn in dry sediment). Maximum level of TBT, 66 ng/g, was observed in Sado estuary, at Lisnave site, in the proximity of a big shipyard. MBT decreased site by site by the same order as DBT and TBT did, but its concentrations were much higher in many cases, denoting that TBT contamination was much higher in the past. A comparison with the available previous data confirmed a marked decrease of TBT contamination all over the last years, indicating that the main sources of TBT in Portuguese coastwise stopped effectively.     

Description and evaluation of imposex in Strombus canarium Linnaeus, 1758 (Gastropoda, Strombidae): a potential bio-indicator of tributyltin pollution

Strombus canarium Linnaeus, 1758 is an important gastropod species within the study area and was traditionally collected for food by the locals. The objective of the present study is to assess the incidence of imposex and its severity in this species. Adult conchs were sampled during their main reproductive period, from October 2005 to January 2006, at Sungai Pulai estuary, Johor Straits, Malaysia. A total of 32.81% of adult females showed imposex characteristics, with varying degrees of severity though. The relative penis size (RPS) index ranged from 1.74 to 33.29 (mean = 13.40 ± 2.27, n?=?21), while the relative penis length (RPL) index ranged from 6.28 to 55.19 (mean = 25.83 ± 3.33, n?=?21). The use of vas deferens sequence (VDS) index was however cannot be applied as the presence of egg groove obscured any vas deferens development in affected females. Sequence of imposex (male penis) development in female conch, from merely a small stump to an advance male penis homologous was therefore carefully analyzed and described, and an alternative imposex classification scheme was proposed. S. canarium can be a good indicator for monitoring of organotin pollution within the study area. However, more studies are needed in order to further develop and test its validity and application, such as its correlation with levels of pollutant within the tissues and the environment, as well as its application on other Strombus species.     

Macrofaunal recovery following TBT ban

In the United Kingdom, the use of TBT-based anti-fouling paints on small vessels was banned in 1987, and a biological study of the Crouch Estuary, a yachting centre on the south-eastern coast of the UK, was conducted in order to determine the ecological improvements resulting from this legislation. We present the changes in the macro-infaunal communities along the estuary in relation to declining TBT concentrations between 1987 and 2005. Although the major changes in response to the ban were observed within the first 3 years (primarily an increase in the number of crustacean taxa and a shift in community structure), with changes still apparent between three and 5 years, the temporal duration of this study allowed the rapidity of the response to be truly determined.     

Effects of the fuel oil spilled by the Prestige tanker on reproduction parameters of wild mussel populations

The aim of this work was to assess possible effects of the Prestige oil spill on reproduction parameters of mussels along the Galician and Bay of Biscay coast. Studied endpoints included sex ratio, gonad histology and vitellogenin-like proteins using the alkali-labile phosphate (ALP) method. A high prevalence of haemocytic infiltration of follicles and severe oocyte atresia was found in most localities in April 2003. Spawning gonads were observed in most impacted populations in the same sampling. In April 2004 mature small sized follicles were observed. No histopathological changes were observed in April 2005 and 2006, except a high prevalence of necrotic gametes in 6 out of 22 localities in April 2006. Female ALP levels showed high interindividual variability in April 2004, which was reduced in April 2005 and 2006. No xenoestrogenic effects were observed in male mussels. Overall, gamete alterations were detected during 2003-2004 and a recovery trend was observed afterwards.     

QUASIMEME laboratory performance study of the biological effects of tributyltin (imposex and intersex) on two marine gastropod molluscs

The disruption of the endocrine systems of gastropod molluscs and consequential physiological changes (imposex and intersex) are used as biomarkers for environmental contamination by tributyltin compounds. The first international laboratory performance study on the determination of imposex and intersex in neogastropod molluscs, Nucella lapillus and Littorina littorea has been undertaken by the QUASIMEME (Quality Assurance of Information for Marine Environmental Monitoring in Europe) project. Samples of live gastropods were distributed and participants were asked to record shell height and sex, together with penis length and vas deferens sequence stage (VDS) in Nucella or the intersex stage (IS) and prostate length in Littorina. Calculations were made of vas deferens sequence index (VDSI) and the relative penis size index (RPSI) in Nucella and of intersex stage index (ISI) and the average female prostate length (FPrL) in Littorina. Thirteen (87%) of the 15 participating laboratories returned data. The remaining two laboratories asked to participate in later exercises. For Nucella, seven laboratories reported sex ratios significantly different from the reference laboratory data. Differences in penis length measurements between laboratories were largely random, although there were indications of systematic errors affecting the data from three laboratories. Seven laboratories reported satisfactory data (Z-score magnitude of Z < 2) for VDSI. The inclusion of a high proportion of sub-adults in the Nucella samples may have made separation of the sexes more difficult than in mature adults. The sub-adults will have had smaller pene than mature adults in the same population, and therefore any errors (random or systematic) in the measurement of penis length or observation of reproductive organs would have a potentially greater impact on the final reported values of the summary imposex indices. The Littorina sample did not show a high degree of intersex (ISI = 0.41). The laboratories could determine the sex of Littorina reliably and only one laboratory reported data significantly different from the reference laboratory. All except two laboratories reported satisfactory data for ISI.     

Application of a battery of biomarkers in mussel digestive gland to assess long-term effects of the Prestige oil spill in Galicia and Bay of Biscay: tissue-level biomarkers and histopathology

In order to assess the biological effects of the Prestige oil spill (POS), mussels, Mytilus galloprovincialis, were collected in 22 localities along the North coast of the Iberian Peninsula over 3 years (April 2003-April 2006). Different tissue-level biomarkers including cell type composition (volume density of basophilic cells, Vv(BAS)) in digestive gland epithelium, structural changes of digestive alveoli (mean luminal radius to mean epithelial thickness, MLR/MET) and histopathological alterations (prevalence and intensity) of the digestive gland were analysed. Severe alterations in the general condition of the digestive gland tissue were observed all over the study area up to 2004-2005. High Vv(BAS) values were recorded mainly in Galicia but also to a lesser extent in the Bay of Biscay in 2003-2004. Atrophy of the digestive alveoli, measured in terms of MLR/MET, was detected all along the studied area up to 2006. Inflammatory responses cannot be related to pollution due to the POS: (a) prevalence and intensity of focal hemocytic infiltration were higher in the Bay of Biscay than in Galicia but they did not show a clear temporal trend; (b) high intensities of brown cell aggregates were only sporadically recorded; and (c) granulocytomas were more frequently recorded in the Bay of Biscay than in Galicia and especially in localities (i.e. Arrigunaga) subjected to chronic pollution. Likewise, Marteilia, trematodes, intracellular ciliates, unidentified eosinophilic bodies, R/CLO and Mytilicola did not follow any recognisable pattern that could be associated to the POS. In contrast, high Nematopsis intensities recorded in several localities in 2003 might suggest some response of local interest after the POS (i.e., in combination with particular factors/conditions). More data at a regional scale are needed before histopathology may provide a reliable ecosystem health assessment but the present results suggest that the approach is worthwhile. Overall, although Vv(BAS) returned to reference values by 2004-2005, MLR/MET values indicated that the mussel health condition was affected during the whole study period up to April 2006.     

TBT in California coastal waters: Monitoring and assessment

Butyltins were monitored at over eighty sites, primarily marinas, in surface waters and sediments of the California coast. Values of tributyltin (TBT) in marina waters ranged from 20 to 600 ppt while lower values were usually found in harbors and on the coast. The origin of the tributyltin is in its use as an antifouling agent in marine paints. In those marinas where the concentrations are greater than about 100 ppt, there is usually a conspicuous absence of native organisms, especially molluscs, which are among the most sensitive animals to the highly toxic TBT. The impact of TBT upon not-target organisms recalls the DDT episode of the 1970s. Increasing uses of TBT-containing antifouling paints in the future may have even more drastic effects on coastal organisms than those observed today. The conclusion is inescapable that TBT should be banned for use in commercially available marine paints.     

Assessment of pollution profile in Buyukcekmece Watershed, Turkey

Buyukcekmece Lake located north of the Marmara Sea coast of Turkey is the third largest water resources among the six main reservoirs of a megacity Istanbul, providing 17% water demand. This study aims to investigate the water quality changes in Buyukcekmece Lake and to classify it in accordance to Turkish Water Pollution Control Regulation for inland surface waters, and assess the impacts of the land-use practices, population density, and settlement in the Buyukcekmece watershed. Physical, chemical, and biological quality of Büyükçekmece Lake has been investigated between December 2007 and June 2008 by collecting samples from the Lake and four streams that feed Büyükçekmece Lake. Land use profile maps of the water basin were created by using ARC-GIS to ensure the quality of drinking water. It was determined that the water quality of Büyükçekmece Lake was still of overall second class, implying that it can be used as a safe drinking water resource following a conventional water treatment scheme; however, it is moving toward the third class. Remarkable portion of the pollution in water basin was attributed to domestic wastewater discharges. Agricultural areas that occupy more than half of the water basin have also adversely affected the water quality.     

Distribution of polychlorinated biphenyls and organochlorine pesticides in wild mussels from two different sites in central Croatian Adriatic coast

Levels of 24 organochlorine compounds were investigated in wild mussels collected at two locations (Krka estuary and Ka?tela Bay) on the Croatian Adriatic coast in 2003, 2005, 2006, 2007, and 2008. PCB and OCP ranges found at the two locations overlapped and followed similar profiles despite the differences between the two locations indicating that they share a common pollutants source. Among organochlorine pesticides, the dominant compound was DDT. Among indicator PCBs, the dominant compound was PCB-153, while PCB-118 was dominant among the remaining 11 congeners. Generally, the sum of six indicator PCBs was constantly greater than the sum of 11 congeners at both locations. α-HCH/γ-HCH and DDE/DDT ratios were below 1, indicating recent input of γ-HCH and DDT into the marine environment. In the investigated period, almost all organochlorine compounds reached the highest values in 2006. The levels of PCBs and OCPs in this study were considerably below the Croatian maximum permissible levels, confirming that they do not pose any threat for human health.     

Copper, zinc, and cadmium body concentrations in Hexaplex trunculus collected from the Tunisian coast

Copper, Zn, and Cd were analyzed in the soft tissues of the edible gastropod Hexaplex trunculus collected from seven localities along the Tunisian coast. The body concentration of these metals was significantly different depending on the gender and site. Copper and Zn varied, respectively, from 47.70 to 343.64 μg/g dry weight (dw) and 149.46 to 530.44 μg/g dw, being higher in males. Cadmium varied from 0.22 to 18.95 μg/g dw and was always higher in females. Overall, the highest concentrations of the three metals were all recorded in Gabès fishing harbor. Comparison of metal concentrations with the European standards compiled by the Food and Agriculture Organization showed that values exceeded standards in several localities. Imposex had been previously recorded in H. trunculus along the Tunisian coast; however, it does not seem to be related with Cu, Zn, and Cd pollution because the body concentration of these metals is not correlated with imposex degree. Nevertheless, interactive effects resulting from the combination of different pollutants and other stressors cannot be disregarded.     

官厅水库入库断面水质多指标评价与演变特征分析

水体质量状况及其污染特征是流域水污染防治规划和治理措施制定的前提和基础。采用主成分分析和层次聚类分析等多元统计分析方法,选取溶解氧、高锰酸盐指数、化学需氧量等10个不同类型的监测指标,综合评价了官厅水库入库断面八号桥2006—2017年丰水期和枯水期水质年际变化特征,识别了不同阶段关键污染指标。结果表明:各年份丰水期(9月)大多数水质指标均优于枯水期(5月),特别是粪大肠菌群和氨氮,但总磷和高锰酸盐指数的差异较小。根据水质指标年际变化情况,可将研究期分为污染严重阶段(2006—2007年)、污染改善阶段(2008—2015年)和污染全面好转阶段(2016—2017年)。大部分水质指标呈现逐年好转的趋势,特别是粪大肠菌群、氨氮和五日生化需氧量,但总磷仍是官厅水库入库河流的重要污染指标。     

Statistical distributions of trace metal concentrations in the northwestern Mediterranean atmospheric aerosol

The concentrations of 11 crustal and anthropogenic trace metals (Li, Al, V, Mn, Fe, Co, Ni, Cu, Zn, Cd, Pb) were measured from 2006 to 2008 in the atmospheric aerosol at a northwestern Mediterranean coast (station of Cap Ferrat, situated on the southeastern coast of France). Statistical models (lognormal, Weibull, and gamma) that best represented the trace metal distribution for this environment are described. The lognormal model was selected for the distributions of (in decreasing strength of the fit) Al, Co, Li, Zn, Mn, Cu, Pb, and Cd, i.e., metals that are introduced into the atmospheric aerosol by pulses inducing temporal variability in their concentrations. The gamma model was associated with Fe, i.e., metals that exhibit less inter-annual variability than the former trace metals. The third mode (Weibull) represented the distribution of the concentrations of V and Ni. The statistical approach presented in this study contributed to better define and constrain the distribution of the 11 trace metals of the atmospheric aerosol from the northwestern Mediterranean coast. In a close future, knowledge of these statistical distributions will allow using convolution models to separate their natural and anthropogenic contributions, therefore increasing our ability to study anthropogenic emissions of trace metals and their impact on the environment.