Sexual Disruption in the Freshwater Crab (Geothelphusa dehaani)1

SYNOPSIS. To clarify the effect of environmental endocrine disruptors on the induction of secondary sexual characteristics in wild animals, we observed 1,527 male and 1,847 female specimens of sexually mature Japanese freshwater crabs, Geothelphusa dehaani, collected at 12 sample points from 10 rivers in Sasebo City, Nagasaki Prefecture, Japan. The results obtained were as follows:

1. Abnormal males, with external appearance of a complete male, but having female genital openings, were observed at frequencies of 8-32% of males collected at all sample points. In contrast to this, all females collected were normal.

2. The frequency of abnormal males increased significantly with the increase in individual carapace width.

This species inhabits mountain streams in Japan, which are free from tributyltin that is known to induce the sexual mosaic phenomenon in mollusks. The streams are most likely polluted by some agricultural chemicals and chemicals present in waste gases from cars and especially particulate materials from diesel engines. The clear relationship of the frequencies of abnormal males to size (or age) suggests that the sexual mosaic phenomenon is induced not only at early stages of male development but after sexual maturation, in response to long-term exposure to chemical(s) during the whole life cycle of males. The definitive causal factors and mechanisms inducing sexual mosaics in this species are still unclear.

INTRODUCTION

Recently, a number of studies have been performed worldwide that have examined endocrine disrupters (EDs) and their interaction with the development and function of various systems in animals and humans (Colborn et al, 1993, 1996; Jobling et al, 1998). As invertebrates account for roughly 95% of the known species of animals on our planet (DeFur et al., 1999), evaluation of the adverse effects of ED exposure on invertebrates is important, in order to develop systems to test the health of any given environment. Field studies on imposex and organotin (butyltin and phenyltin) contamination in Japanese molluscs (the rock shell, Thais clavigera, and the Pacific oyster, Crassostrea gigas) conducted along the coast of both the Seto Inland Sea and in the Sanriku region, Japan, suggested that the percentage occurrence of imposex in the rock shell, T. clavigera, was 100% at all sites surveyed (Horiguchi et al., 1998). During a one-year survey, beginning in September 1990, six abnormal Geothelphusa dehaani were found out of 263 individuals collected in the Taniyama river, a small stream in north Kyushu, Fukuoka, Japan (Araki and Matsuura, 1995). The imposex phenomenon has been found to occur in crabs which inhabit tributyltin-uncontaminated mountain streams in Japan (Takahashi et ai, 2000).

To clarify the effect of EDs on the induction of secondary sexual characteristics of wild animals, we observed 1,527 males and 1,847 females of sexually mature Japanese freshwater crabs, G. dehaani, collected at 12 sample points from 10 rivers in Sasebo City, Nagasaki Prefecture, Japan. Crabs with the female genital opening gonopore, with incomplete male genitals were categorized as masculinized females; in addition, crabs possessing complete male genitals with the female genital opening gonopore were categorized as feminized males.

MATERIALS AND METHODS

We observed 1,527 males and 1,847 females of sexually mature Japanese freshwater crabs, G. dehaani, collected at 12 sample points (Point A-L) from 10 rivers (Sasebo City, Nagasaki Prefecture, Japan) between August 2000 and August 2001. A map depicting the research sites is shown in Figure 1. Among the collection sites, Yunoki Bridge on the main stream of the Ainoura River (Point D), Takao River, a tributary of the Ainoura River (Point F), and the Hiji River, a tributary of the Komori River (Point J) are located on the outskirts of Sasebo city where the rivers run through a residential area. In particular, samples collected from points D and F were found to be foul smelling, which indicated that these sites were eutrophic. The other 8 points are located in the mountains, in the upper reaches of rivers. Paddies and farms are located upstream of every river. A fruit farm (tangerine) is located upstream of the Horito River (Point K), a tributary of the Miyamura River and also located upstream of the Eshimo River (Point L). Golf courses are located in the foothills of Mt. Iimori, close to the Ogawachi River (Point A), a tributary of the Ainoura River, and at an area several hundred meters away to the northwest of the Komori River (Point I and Point J). An industrial waste disposal facility is located upstream of the Kubonita River (Point E). Points B and C are located in the upper reaches of the Ainoura River and points G and H are located in the upper reaches of the Sasebo River. A few private houses are dotted within the inter-mountain agricultural community area (Points B, C, G and H).

Observation of abnormality in crabs: The carapace sizes of collected crabs were larger than 10.9 mm and thus considered to be sexually mature. The external genital organs of crabs were checked for abnormality under a microscope. Crabs with the female genital opening gonopore, with incomplete male genitals were categorized as masculinized females. Crabs possessing complete male genitals with the female genital opening gonopore were categorized as feminized males. The individual crabs were released back into their original territory to avoid destruction of natural conditions as a result of this field study.