Males often perform mate choice with the aim of maximizing reproductive success. To identify profitable mates, the males of some animals are known to use visual and chemical cues derived from females. In this study, we aimed to examine mate discrimination by male guppies (Poecilia reticulata) using chemical cues received from females under different reproductive statuses, i.e., virgin females, pregnant females, females after copulation with another male, and post-partum females. We conducted a dichotomous choice experiment for each combination of chemical stimuli from females under different reproductive statuses. In experiment 1, in which females were removed from water that was subsequently used as a chemical stimulus, male preferences did not differ significantly in all combinations of chemical stimuli from females under different reproductive statuses. However, in experiment 2, in which females remained within bottles containing the water used as a chemical stimulus, with the exception of one combination of chemical stimuli, significant differences in male preferences were detected for chemical stimuli derived from females under different reproductive statuses. Overall, males generally showed a preference for chemical stimulus received from females after copulation with other males. The findings of this study indicate that male guppies can discriminate the reproductive status of females based solely on chemical cues that may disappear or degenerate within a short period of time, thereby facilitating profitable mate choice.

In most vertebrates, the oviducts and sperm ducts are derived from the Müllerian ducts and Wolffian ducts, respectively. However, in teleosts, the genital ducts are formed by the posterior extension of gonads in both sexes. Whether the genital ducts of teleosts are newly evolved organs or variants of Müllerian ducts is an important question for understanding evolutionary mechanisms of morphogenesis. One of the genes essential for Müllerian duct formation in mice is Wnt4, which is expressed in the mesenchyme and induces invagination of the coelomic epithelium and its posterior elongation. Here, we addressed the above question by examining genital duct development in mutants of two Wnt4 genes in the medaka (wnt4a is orthologous to mouse Wnt4, and wnt4b is paralogous). The wnt4b mutants had a short body but were fertile with normal genital ducts. In contrast, both male and female wnt4a mutants had their posterior elongation of the gonads stopped within or just outside the coelom. The mutants retained the posterior parts of ovarian cavities or sperm duct primordia, which are potential target tissues of Wnt4a. The gonads of female scl mutants (unable to synthesize sex steroids) lacked these tissues and did not develop genital ducts. Medaka wnt4a was expressed in the mesenchyme ventral to the genital ducts in both sexes. Taken together, the data strongly suggest that the mouse Müllerian ducts and the medaka genital ducts share homologous developmental processes. Additionally, the wnt4a or wnt4b single mutants and the double mutants did not show sex-reversal, implying that both genes are dispensable for gonadal sex differentiation in the medaka.

Salmincola californiensis is a parasitic copepod of freshwater salmonids in the North Pacific rim countries. Sixteen adult females of the species were found alive on the gills of an ocean-age 4, maturing steelhead trout, Oncorhynchus mykiss, caught in offshore waters (50°30′N, 179°30′W) of the North Pacific Ocean in July 1997. This is the first evidence of live individuals of S. californiensis from ocean-migrating salmonids. When found, copepods were attached to the distal ends of gill filaments, and their bodies were observed to be slowly moving in Petri dishes with seawater. Ocean-migrating steelhead trout comprise individuals originating from western Kamchatka (Russia) and western North America. Based on the date and catch location of the infected fish, it is inferred that it originated from western North America, where it acquired S. californiensis infection in fresh water. As this fish spent about 4 years in the ocean, the copepods likely survived the same period at sea. However, if the fish was a kelt, the survival period of the copepods in the ocean may be shorter than four years. To confirm identification of the copepods, adult females of S. californiensis are briefly described using the specimens collected from the fish.

Rhizocephalan barnacles (Thecostraca: Cirripedia) are parasitic crustaceans that lack appendages, segmentation, and a digestive system in adults, while instead infiltrating their hosts with a nutrient-absorbing system of rootlets. Sacculinids, belonging to the Rhizocephala order, are known for their various parasitization-induced effects on their decapod hosts, such as parasitic castration, reduction in the growth of secondary sexual characteristics, feminization of male crabs, and alteration of host behavior. In this study, we conducted field surveys in Japan at Manazuru Town (Kanagawa) on the Pacific coast, and on Sado Island and Noto Peninsula on the Sea of Japan side, and found that sacculinid-parasite-ratios on the grapsid crab Pachygrapsus crassipes were particularly high on the Sea of Japan coast. Molecular phylogenetic analysis revealed that the Manazuru population forms a single clade with Sacculina yatsui, and both Sado and Noto populations form a single clade with S. confragosa. We further demonstrated that external morphologies of male P. crassipes parasitized by sacculinids were changed to female phenotypes. This host-parasite interaction will be a useful model for understanding molecular mechanisms underlying rhizocephalan-driven morphological and behavioral feminization and castration.

The pond snail Lymnaea stagnalis is capable of learning by both classical conditioning and operant conditioning. Although operant conditioning related to escape behavior with punishment has been examined by some research groups, the molecular mechanisms are not known. In the present study, we examined changes in the expression levels of cAMP-response element binding protein 1 (CREB1), CREB2, CREB-binding protein (CBP), and monoamine oxidase (MAO) in the Lymnaea central nervous system (CNS) using real-time PCR following operant conditioning of escape behavior. CREB1 and CREB2 are transcription factors involved in long-term memory in Lymnaea; CBP is a coactivator with CREB1; and MAO is a degrading enzyme for monoamines (e.g., serotonin) with important roles in learning and memory in Lymnaea. In operant conditioning, the punishment cohort, in which snails escaping from the container encountered aversive KCl, exhibited significantly fewer escape attempts than the control cohort, in which snails escaping from the container encountered distilled water, during both the training and memory test periods. After the operant conditioning, CREB1 and CREB2 were upregulated, and the ratio of CREB1/CREB2 was also increased, suggesting that the operant conditioning of escape behavior involves these factors. MAO was also upregulated, suggesting that the content of monoamines such as serotonin in the CNS decreased. The upregulated genes identified in the present study will help to further elucidate learning and memory mechanisms in Lymnaea.

In the pond snail Lymnaea stagnalis, serotonin (5-HT) plays an important role in feeding behavior and its associated learning (e.g., conditioned taste aversion: CTA). The 5-HT content in the central nervous system (CNS) fluctuates with changes in the nutritional status, but it is also expected to be influenced by changes in the serotonin transporter (SERT) expression level. In the present study, we identified SERT in Lymnaea and observed its localization in 5-HTergic neurons, including the cerebral giant cells (CGCs) in the cerebral ganglia and the pedal A cluster neurons and right and left pedal dorsal 1 neurons in the pedal ganglia by in situ hybridization. Real-time PCR revealed that the SERT mRNA expression level was lower under severe food deprivation than under mild food deprivation in the whole CNS as well as in a single CGC. These results inversely correlated with previous data that the 5-HT content in the CNS was higher in the severely food-deprived state than in the mildly food-deprived state. Furthermore, in single CGCs, we observed that the 5-HT level was significantly increased in the severely food-deprived state compared with the mildly food-deprived state. Our present findings suggest that changes in the SERT expression level associated with food deprivation may affect 5-HT signaling, probably contributing to learning and memory mechanisms in Lymnaea.

Variations of the radula and shell microstructures in 33 species of Japanese chiton were investigated along with molecular phylogenetic trees. The molecular phylogenetic trees indicated that Chitonida was composed of four clades, of which two clades formed Acanthochitonina and corresponded to Mopalioidea and Cryptoplacoidea, respectively, and the other clades formed Chitonina. In the radula, the shapes of the central and centro-lateral teeth and the petaloid process varied greatly among species or genera and were useful for the identification of particular species or genera. The presence of accessory and petaloid processes and the cusp shape were relatively conserved and useful for recognizing particular genera or even suborders. In the valves, four to six shell layers were found at the section, but the ventral mesostracum was not observed in Acanthochitonina. The shell microstructures in the ventral sublayer of the tegmentum varied at suborder, but those in the other layers were almost constant. The megalaesthete chamber type varied at superfamily and was helpful to identify particular families or superfamilies. The characteristics of the shell layers and shell microstructures appear to be a synapomorphy shared by the members of Acanthochitonina. The classification within Chitonina needs to be reexamined because the variations of the cusp shape and megalaesthete chamber type were relatively large and did not correspond to the current classification. Callochiton formed a sister group with Chitonida and would be equally closely related to Chitonina and Acanthochitonina because of possessing a mosaic of characteristics from both.

High stocking densities have negative effects on fish. However, the mechanism mediating density perception and growth inhibition is still unknown. This study was conducted to confirm the occurrence of growth inhibition and evaluate changes in growth-related factors in fish reared under high-stocking-density conditions and to determine the role of vision in density perception of medaka. In the graduated-stocking experiment, growth inhibition was clearly observed in fish reared at higher densities, although environmental factors, such as water quality, dissolved oxygen, and feeding conditions, were the same in each experimental group. Differences in growth were observed between the 6-fish and 8-fish groups, indicating that medaka have a superior sense that allows them to accurately perceive the number of individuals in their surroundings. In the pseudo-high stocking experiment, the inner 2-L tank in both groups contained six fish; however, the outer 3-L tank in the pseudo group contained several fish, while that of the control group contained only water. Growth inhibition was observed among the fish in the inner tank of the pseudo group despite having similar spatial density with the control group. These findings suggest that vision is important for density perception. The gene expression of growth-related and metabolic-regulatory hormones decreased in the high-density group. Furthermore, neuropeptide Y expression increased, while pro-opiomelanocortin expression decreased in the high-density group. This study is the first to report that fish can visually perceive density and the resulting growth inhibition, and concluded that medaka is a suitable model for studying density effects and perception in fish.

A new species of Hua, Hua qiannanensis sp. nov., is described from Guizhou Province, China, based on morphological and molecular evidence. The new species can be distinguished from its congeners by the following combination of characters: the smooth shell, only three smaller cusps of lateral teeth on the inner side, outer marginal teeth with eight flattened and rounded denticles, an ovipositor pore in females, and BW/H ≥ 80%, B/H = 76.8–82.3%. Molecular analysis based on partial mitochondrial COI and 16S rDNA also supports the systematic position of the new taxon.