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Über dieses Buch

This book provides an overview of the induction mechanism of imposex caused by organotin compounds in gastropods, as well as fundamental information on the physiology and biochemistry of reproduction in mollusks. Are the sex hormones of gastropod mollusks vertebrate-type steroids, or neuropeptides? What about lipid disturbance and membrane toxicity due to organotin compounds? The book also discusses the latest findings on the role of nuclear receptors, such as retinoid X receptor (RXR), retinoic acid receptor (RAR) and peroxisome proliferator-activated receptor (PPAR), in the development of imposex in gastropods.

Further, it describes the current state of contamination by organotins in the marine environment and gastropod imposex, especially focusing on Europe and Asia, introduces readers to analytical techniques for organotin compounds, and assesses the contamination and adverse effects of alternatives to organotin-based antifouling paints.

Imposex, a superimposition of male genital tracts, such as penis and vas deferens, on female gastropod mollusks, is known as a typical phenomenon or consequence of endocrine disruption in wildlife. Imposex is typically induced by very low concentrations of organotin compounds, such as tributyltin (TBT) and triphenyltin (TPhT) from antifouling paints on ships and fishing nets. Reproductive failure may be brought about in severely affected stages of imposex, resulting in population decline and/or mass extinction. Thus, gastropod imposex has been recognized as a critical environmental pollution issue. Although gastropod imposex is also highly interesting for the biological sciences because of its acquired pseudohermaphroditism and/or sex change by certain chemicals, such as TBT and TPhT, the mechanism that induces the development of imposex remains unclear, possibly due to our limited understanding of the endocrinology of gastropod mollusks. This book offers a useful guide for professionals and students interested in the fields of aquatic biology, invertebrate physiology, ecotoxicology and environmental science.



Analytical Techniques for Trace Levels of Organotin Compounds and Contamination by Organotin and Alternative Antifouling Paints in the Marine Environment


Chapter 1. Analytical Techniques for Trace Levels of Organotin Compounds in the Marine Environment

Organotins still remain a major concern for the safety of the marine environment, and their determination is covered under legislation in quite a number of nations. Because their usage is totally banned, the demand for determining organotins at sub-nanogram concentrations is ever increasing, which is achieved by elimination of matrix interferences, reduction of sample volume, and analyte enrichment. Organotin speciation is a complex technique involving a long and laborious sample treatment procedure that is prone to various uncertainties. To overcome the shortfalls in extraction and pre-treatment, newer microextraction techniques were developed with reduction in sample and solvent volume, extraction time, and enrichment procedures. Moreover, the recent techniques are developed with a major focus on green analytical chemistry to reduce the impact of anthropogenic (laboratory) activities on the environment. Decreasing the detection limit of methods without greatly compromising their sensitivity was a profound topic of environmental research for organotin analysis. In the case of analytical technique, from the late 1970s, the usage of titrometric and spectrophotometric methods were substituted with more sensitive and lower-cost detectors at nanogram level. Furthermore, detection at femtogram levels was achieved by a mass spectrometer coupled to either gas chromatography (GC) or liquid chromatography (LC) systems. One of the significant developments in instrumentation is the application of the isotope dilution technique to detect the transformation/degradation of organotin species during extraction analysis steps. This chapter discusses the methods available for measuring organotins and their metabolites in seawater, sediment, and biota such as fish and oysters and compares the performance of the various analytical methods available.
Babu Rajendran Ramaswamy

Chapter 2. Continuing Issues of Contamination by Organotins in the Marine Environment After Domestic and International Legislation

Organotin compounds, which are used for painting on bottoms of ship’s hull, have high residual property in the environment, and examples of their detection in the environment have been reported even after the introduction of international regulations on their use. To better understand the residual concentrations of organotins in marine environments, studies have been conducted on regional and small-scale fishing harbors, rather than in areas with integrated, large-scale commercial harbors and dockyards and areas where industrial facilities unrelated to the fishing industry exist. This approach of using small fishing harbors has advantages because there are many sites available for analysis, and the analytical noise is low. It is found that the organotin compound concentrations in sediments of fishing harbors with repainting facilities are clearly higher than those of harbors without such facilities. The organotin contamination depends on the presence of the repainting facilities and not on the scale of the fishing harbor. It is proposed that paint flakes produced during the repainting of hull bottoms are discharged into the sea, and these paint flakes are a high concentration source of contamination. Since the difference in organotin values between harbors with and without repainting facilities is less than ten times at the median values, it is considered that organotin compounds are released intermittently into the environment during periods that a ship is in harbor and that the quantities of organotins in the environment are not as low as expected.
Yuji Takao

Chapter 3. Emerging Issues on Contamination and Adverse Effects by Alternative Antifouling Paints in the Marine Environments

The current status of antifouling biocides contaminations was reviewed in water, sediment, and biological samples, and the effect of antifouling biocides for aquatic organisms was evaluated.
Irgarol 1051 (3-methylthio-4-tetrabutylamino-6-cyclopropylamino-s-triazine), diuron (3-(3,4-dichlorophenyl)-1,1-dimethylurea), sea nine 211 (2-n-octyl-4,5-dichloro-2-methyl-4-isothiazolin-3-one), chlorothalonil (2,4,5,6-tetrachloro-isophthalonitrile), dichrofluanid (N′-dimethyl-N-phenylsulphamide), metal pyrithions (metal complex of 2-mercaptopyridine-1-oxide), and PTPB (pyridine triphenylborane) in water were in the range of 0.5–2,430 ng/l, <0.7–6,742 ng/l, <0.3–3,700 ng/l, <1–1,380 ng/l, <1–55 ng/l, <80 ng/l, and 0.0036–0.021 ng/l, respectively. The concentrations of Irgarol 1051, diuron, sea nine 211, chlorothalonil, dichrofluanid, and pythiones in sediment were in the range of <0.02–816 μg/kg dry, <0.02–1,350 μg/kg dry, <0.04–150 μg/kg dry, <0.01–46.5 μg/kg dry, <0.1–688.2 μg/kg dry, <8–420 μg/kg dry, respectively. Irgarol 1051 was detected in the range of <0.1–35 μg/kg in clam, mussel, and oyster from Vietnam, Thailand, and Japan. The concentrations of diuron and sea nine 211 in bivalves were <0.1–9.6 μg/kg and <0.1–0.3 μg/kg, respectively.
EC50 and LC50 of Irgarol 1051 were in the range of 0.09–50,800 μg/l and 0.38 to >40,000 μg/l, respectively. EC50 of sea nine 211 were in the range of 0.42–12 μg/l. EC50 and LC50 of diuron were in the range of 4.3–43,000 μg/l and 5.9 to >127,000 μg/l, respectively. EC50 and LC50 of chlorothalonile were in the range of 4.4–390 μg/l and 12–110 μg/l, respectively. EC50 of dichlofluanid was in the range of 87–1,050 μg/l. EC50 of tolylfluanid was in the range of 9.9–405 μg/l. EC50 and LC50 of PTPB were in the range of 2.2–140 μg/l and 54 μg/l, respectively. EC50 of TCMTB ((2-thiocyanomethylthio) benzothiazole) was in the range of 46–433 μg/l.
Judging from toxicity data, most of these alternative biocides concentrations which were detected in the aquatic environment were below the level that causes an adverse effect in aquatic organisms.
Hiroya Harino

Contamination by Organotins and Organotin-Induced Imposex in Gastropod Mollusks


Chapter 4. Contamination by Organotins and Its Population-Level Effects Involved by Imposex in Prosobranch Gastropods

A history of the production and use of organotin compounds is briefly introduced. The worldwide use of tributyltin (TBT)- or triphenyltin (TPhT)-based antifouling paints since the mid-1960s has caused extensive contamination in the aquatic environment, especially in the marine environment, which led to contamination of aquatic organisms by these compounds and became a concern in terms of both seafood safety and ecotoxicology. Legislation of TBT- and TPhT-based antifouling paints began in Europe and the U.S.A in the 1980s and in Japan in 1990. An International Convention on the Control of Harmful Anti-fouling Systems on Ships (AFS Convention) for the worldwide ban of TBT- and TPhT-based antifouling paints came into force on 17 September 2008. Organotins have various toxicities to vertebrates and invertebrates; imposex is known to be induced in many gastropod species by TBT and also by TPhT released from antifouling paints on ships and fishing nets. Reproductive failure may be brought about in severely affected stages of imposex, resulting in population decline or mass extinction. Population-level effects involved by imposex and similar phenomena are described in the rock shell (Thais clavigera), the ivory shell (Babylonia japonica), and the giant abalone (Haliotis madaka), with special reference to tissue distributions of TBT, TPhT and their metabolites.
Toshihiro Horiguchi

Chapter 5. Organotins and Imposex in Europe: A Pre-ban and Post-ban Perspective

The use of tributyltin (TBT) and, to a lesser extent, triphenyltin (TPT) as biocides in antifouling paint formulations during the last decades of the twentieth century was responsible for the global contamination of the coastal and offshore areas of the European aquatic environment. This widespread contamination was responsible for extensive deleterious effects in nontarget organisms particularly in gastropods, which, as a consequence of TBT exposure, developed imposex in a dose-dependent manner. Over the last decades, more than 20 gastropod species registering the occurrence of imposex were used by European researchers to ascertain the degree of TBT pollution in the European coastal and marine environment. In this chapter, we will evaluate the status of organotin (OT) contamination in European countries, using imposex in gastropods as a surrogate. The temporal trends uncovered are evaluated under the light of the 2003 European ban (Directive 2002/62/EC and Regulation 782/2003) and the AFS Convention (entered into force in 2008). The reduction of TBT pollution is addressed under the same light. Overall, we conclude that significant reductions in imposex levels started to emerge after the European ban (2003), though those reductions were more pronounced from 2008 onward. The most recent surveys disclose that the recovery of the ecosystems from the “TBT nightmare” is ongoing.
Ana Catarina A. Sousa, M. Ramiro Pastorinho

Chapter 6. Current Status of Organotin Contamination and Imposex in Neogastropods Along Coastal Marine Environments of Southeast Asia and China

Organotin compounds (OTs), in particular tributyltin and triphenyltin, have been contaminating various coastal marine environments around the world since their first application back in the 1920s. These compounds have been proven to adversely affect a wide range of marine organisms from microalgae to marine mammals, and they have a great potential of bioaccumulation via the food web. Some OTs such as triphenyltin can even be biomagnified through marine food chains. In Southeast Asia, OT contamination has been a widespread problem because most countries or regions do not implement local regulations to restrict the use of OT-based antifouling paints on seagoing vessels and fish farming facilities (e.g., open-sea cages), although some of them are members of the International Maritime Organization, which has enforced a global ban of such paints since September 2008. Contamination by OTs was the most severe in coastal waters where intensive shipping or mariculture activities could be found. To rectify the problem and safeguard the marine ecosystem and human health, long-term monitoring of OT contamination and enforcement of more stringent regulations on controlling the use and release of these pollutants in Southeast Asian countries are urgently needed.
Kevin King Yan Ho, Kenneth Mei Yee Leung

Chapter 7. Current Status of Contamination by Organotins and Imposex in Prosobranch Gastropods in Korea

Organotin compounds are considered to be dangerous chemicals because of their deleterious effects on non-target marine organisms. In 2003, the use of TBT-based antifouling paints was totally banned in Korea, and the International Maritime Organization (IMO) proposed to extend the ban to almost all ocean-going vessels. In this study, the concentrations of organotins in the coastal environment are analyzed to illustrate the differences of these analyzed items in the periods before and after the IMO and Korean regulations, focusing on organotins concentrations in molluscan soft tissues and the imposex phenomenon in the rock shell (Thais clavigera), a gastropod species sensitive to organotin compounds, collected from the Korean coasts from 1995 to 1997 and 2002, and from 2005 to 2009. TBT and TPhT were dominant organotins. Higher organotin concentrations were observed in areas with frequent shipping activities, including regions adjacent to harbors or shipyards, than in areas away from shipping activities. Concentrations of TBT, TPhT, and their metabolites in tissue, imposex frequency, and relative penis length index and sterility ratio of rock shell specimens collected from the southern coast after the regulations for TBT- and/or TPhT-based antifouling paints were in place showed lower values than those before the regulations. Because of the continued occurrence of imposex in the rock shell populations after the regulations were established, it is necessary to carry out further studies to monitor organotin concentrations in tissues and imposex frequency of rock shell specimens along with evaluations of the organotins residues in seawater and sediment in Korean coastal areas.
Hyeon-Seo Cho, Toshihiro Horiguchi

Fundamental Knowledge of Physiology and Mode of Action of Organotins to Induce the Development of Imposex in Gastropod Mollusks


Chapter 8. Neuropeptides and Their Physiological Functions in Mollusks

Neuropeptides have essential functions in the neural regulation of physiological functions of various tissues and organs, as well as of animal behaviors. Many neuropeptides have been identified in mollusks, and investigation of their functions is currently proceeding. In this review, I attempt to give an overview of the neuropeptides in mollusks. Then, regulatory actions of neuropeptides are described with a special reference to reproduction. I chose three neuropeptides: egg-laying hormone (ELH) and caudodorsal cell hormone (CDCH), gonadotropin-releasing hormone (GnRH), and APGWamide. ELH and CDCH are well-investigated peptide hormones that trigger complex egg-laying behaviors in Aplysia and Lymnaea. GnRH, which is a key peptide that induces gonadal maturation and ovulation in mammals, also regulates gonadal maturation in bivalves and cephalopods. However, evidence suggests that GnRH also mediates other activities such as feeding and locomotion in mollusks. APGWamide, which regulates the male copulatory activity in freshwater snails, seems to have pheromonal actions in bivalves and cephalopods. These facts collectively emphasize the diverse actions of neuropeptides and peptide hormones on the regulation of reproduction in mollusks.
Fumihiro Morishita

Chapter 9. Mode of Action of Organotins to Induce the Development of Imposex in Gastropods, Focusing on Steroid and the Retinoid X Receptor Activation Hypotheses

Basic knowledge of endocrinology or reproductive physiology of prosobranch gastropods is reviewed, focusing on vertebrate-type steroids as possible sex hormones in gastropods. Major points of the view for criticism are steroid-producing cells, enzymes to synthesize and/or metabolize steroids, and functional receptors for steroids. Mechanism of induction and promotion of the development of imposex is also reviewed, regarding six hypotheses proposed as the mechanism by which organotins, such as TBT and TPhT, induce the development of imposex in gastropods: (1) an increase in androgen (e.g., testosterone) levels as a result of TBT-mediated inhibition of aromatase; (2) an increase in testosterone levels owing to the inhibition of acyl CoA-steroid acyltransferase; (3) TBT-mediated inhibition of the excretion of androgen sulfate conjugates, with a consequent increase in androgen levels; (4) TBT interference with the release of penis morphogenetic/retrogressive factor from the pedal/cerebropleural ganglia; (5) an increase in the level of an alanine-proline-glycine-tryptophan amide (APGWamide) neuropeptide in response to TBT; and (6) activation of the retinoid X receptor (RXR). The latest information about nuclear receptors other than RXR in gastropods, namely, retinoic acid receptor (RAR) and peroxisome proliferator-activated receptor (PPAR), is also described.
Toshihiro Horiguchi

Chapter 10. Effects of Organotins in Mollusk’s Lipids

Organotin compounds have been shown to alter lipid homeostasis and trigger adipocyte differentiation and a predisposition to obesity in vertebrates by binding to nuclear receptors (e.g., the retionid X receptor, RXR). RXR is highly conserved in evolution, and RXR homologues with high ligand affinity for tributyltin and triphenyltin have been cloned from gastropods. Thus, significant alteration of lipids as a consequence of exposure to organotin compounds is likely to occur also in mollusks. This chapter reviews the still fragmentary knowledge on the induction of lipid disturbance and membrane toxicity by organotin compounds and the potential link between those lipid alterations and the occurrence of imposex and/or altered levels of esterified steroids. Finally, the chapter emphasizes the need to characterize the richness of mollusk’s lipids (e.g., cardiolipin, plasmalogens, and many others) with the new available technologies to better understand the toxicity of organotin compounds but also other pollutants/stressors, in this very diverse animal group.
Denise Fernandes, Cinta Porte

Chapter 11. Reproductive Organ Development in the Ivory Shell Babylonia japonica and the Rock Shell Thais clavigera

Results of histological examination of normal differentiation and development of the genital tract and gonad in the ivory shell Babylonia japonica (Buccinidae) are described. The formation of male-type genitalia (penis and vas deferens) in imposex-exhibiting females seems to mimic the normal development of male genitalia in prosobranch gastropods, on the basis of observations using a wild-caught 2-year-old specimen and laboratory-reared juveniles aged 0–24 months. Gonad differentiation was unclear before age 14 months but progressed after 16 months. Both sexes had a complete genital tract and mature gonads at 20 months. However, differentiation and development occurred earlier in females than in males. Development of the genital tract preceded gonad differentiation. Vas deferens morphogenesis in males resembled that in imposex-exhibiting females. Histological examination of the development of male genitalia in imposex-exhibiting female rock shells, Thais clavigera (Muricidae), using specimens from a wild population and tributyltin (TBT)-exposed females in the laboratory, allowed observation of a variety of vas deferens morphogenesis patterns. Taking into consideration observed results both from wild female specimens and from TBT-exposed females in the laboratory, the vas deferens sequence (VDS) index for T. clavigera has been proposed as VDS 1–6, which is a little different from that for Nucella lapillus. Comparison of the differentiation and development of male genitalia in normal males and imposex-exhibiting females among gastropod species implies it does not seem to be strictly regulated: relatively large variation in the differentiation and development of genitalia could occur among individuals, as well as among species of prosobranch gastropods.
Toshihiro Horiguchi
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