Susumu Chiba

Expression Profiling without Genome Sequence Information in a Non-Model Species, Pandalid Shrimp ( Pandalus latirostris ), by Next-Generation Sequencing

While the study of phenotypic variation is a central theme in evolutionary biology, the genetic approaches available to understanding this variation are usually limited because of a lack of genomic information in non-model organisms. This study explored the utility of next-generation sequencing (NGS) technologies for studying phenotypic variations between 2 populations of a non-model species, the Hokkai shrimp (Pandalus latirostris; Decapoda, Pandalidae). Before we performed transcriptome analyses using NGS, we examined the genetic and phenotypic differentiation between the populations. Analyses using microsatellite DNA markers suggested that these populations genetically differed from one another and that gene flow is restricted between them. Moreover, the results of our 4-year field observations indicated that the egg traits varied genetically between the populations. Using mRNA extracted from the ovaries of 5 females in each population of Hokkai shrimp, we then performed a transcriptome analysis of the 2 populations. A total of 13.66 gigabases (Gb) of 75-bp reads was obtained. Further, 58,804 and 33,548 contigs for the first and second population, respectively, and 47,467 contigs for both populations were produced by de novo assembly. We detected 552 sequences with the former approach and 702 sequences with the later one; both sets of sequences showed greater than twofold differences in the expression levels between the 2 populations. Twenty-nine sequences were found in both approaches and were considered to be differentially expressed genes. Among them, 9 sequences showed significant similarity to functional genes. The present study showed a de novo assembly approach for the transcriptome of a non-model species using only short-read sequence data, and provides a strategy for identifying sequences showing significantly different expression levels between populations.

Maladaptive sex ratio adjustment by a sex‐changing shrimp in selective‐fishing environments

1. Selective harvesting is acknowledged as a serious concern in efforts to conserve wild animal populations. In fisheries, most studies have focused on gradual and directional changes in the life-history traits of target species. While such changes represent the ultimate response of harvested animals, it is also well known that the life history of target species plastically alters with harvesting. However, research on the adaptive significance of these types of condition-dependent changes has been limited. 2. We explored the adaptive significance of annual changes in the age at sex-change of the protandrous (male-first) hermaphroditic shrimp and examined how selective harvesting affects life-history variation, by conducting field observations across 13 years and a controlled laboratory experiment. In addition, we considered whether plastic responses by the shrimp would be favourable, negligible or negative with respect to the conservation of fishery resources. 3. The age at sex-change and the population structure of the shrimp fluctuated between years during the study period. The results of the field observations and laboratory experiment both indicated that the shrimp could plastically change the timing of sex-change in accordance with the age structure of the population. These findings provide the first concrete evidence of adult sex ratio adjustment by pandalid shrimp, a group that has been treated as a model in the sex allocation theory. 4. The sex ratio adjustment by the shrimp did not always seem to be sufficient, however, as the supplement of females is restricted by their annual somatic growth rate. In addition, adjusted sex ratios are further skewed by the unintentional female-selectivity of fishing activity prior to the breeding season, indicating that the occurrence of males that have postponed sex-change causes sex ratio adjustment to become unfavourable. 5. We conclude that the plastic responses of harvested animals in selective fishing environments must be considered in efforts to conserve wild animal resources, because such responses can become maladaptive.

Mitochondrial and nuclear DNA analysis revealed a cryptic species and genetic introgression in Littorina sitkana (Mollusca, Gastropoda)

We investigated mitochondrial and nuclear DNA genotypes in nominal Littorina sitkana samples from 2 localities in Eastern Hokkaido, northern Japan. Our results indicated the existence of cryptic species. In the analysis of partial mitochondrial Cytchrome b gene sequences, haplotypes of L. sitkana samples were monophyletic in a phylogenetic tree with orthologous sequences from other Littorina species, but were apparently separated in 2 clades. One included typical L. sitkana (CBa clade) samples, which formed a clade with an allopatric species, L. horikawai. The other, CBb, was independent from CBa and L. horikawai. Haplotypes of the mitochondrial 16S rRNA gene also separated into 2 clades. We additionally examined intron sequence of the heat shock cognate 70 (HSC70) nuclear gene and identified 17 haplotypes. These were also separated into 2 clades, HSCa and HSCb. Among the examined Hokkaido samples, 60% of individuals were heterozygotes. However, each heterozygote consisted of haplotypes from the same clade, HSCa or HSCb, and no admixture of HSCa and HSCb haplotypes was observed. These results indicate reproductive isolation between the 2 clades. Among the genotyped Hokkaido samples, 93% of individuals had CBaxa0+xa0HSCa or CBbxa0+xa0HSCb genotypes, and 7% had CBbxa0+xa0HSCa genotypes. The discrepancy between the mtDNA and nuclear DNA haplotypes in a few individuals may have been caused by genetic introgression due to past hybridization.

Day–night change in fish community structure in a seagrass bed in subarctic waters

The day–night change in fish community structure over a year was examined in a seagrass bed in Lake Notoro, Hokkaido, northern Japan, to examine nocturnal increases in predation risk. This seagrass bed has previously been considered a predation refuge for juvenile and small-sized fishes. Species richness, abundance and biomass of piscivorous fishes during nighttime were higher than those during daytime on all sample dates surveyed (May, August and November 2013), indicating an increase in predation risk for juvenile and small-sized fishes during nighttime. The mean biomass-weighted trophic level of fish communities in the seagrass bed was also higher at night than in the day. The piscivorous fishes collected in the seagrass bed during the night included important fishery species. These may obtain energy through nocturnal feeding in the seagrass bed. Therefore, the function of seagrass beds as fish habitats should be re-evaluated by considering two possible characteristics contributing to fishery production: as a daytime predation refuge for juvenile and small-sized fishes, and as a nighttime foraging ground for piscivorous fishes.

Isolation and characterization of 13 polymorphic microsatellites for the Hokkai Shrimp, Pandalus latirostris

We have developed and characterized 13 novel polymorphic microsatellite markers for the Hokkai Shrimp, Pandalus latirostris, to provide an effective tool for conducting genetic studies on this species. No linkage disequilibria and no deviation from HWE were detected in these markers. In 32 individuals from Lake Notoro in Hokkaido, Japan, the number of alleles and expected heterozygosities ranged from 7 to 22 and from 0.84 to 0.95, respectively, suggesting the availability of these markers for ecological studies and conservation genetics in this species.

Spatial–temporal variations in the composition of two Zostera species in a seagrass bed: implications for population management of a commercially exploited grass shrimp

We examined spatial and temporal variations in the species composition of two seagrass species, Zostera marina and Z. caespitosa, in a lagoon facing the Sea of Okhotsk. We also considered how those variations affected habitat quality for motile epifauna, especially for a commercial shrimp. A long-interval comparison between 1996 and 2013 showed that seagrass species composition in the lagoon did not vary, while their relative abundances did. A survey in 2012 revealed that the abundance of Z. caespitosa was affected by the abundance of Z. marina, water depth, and location in the lagoon. Although these seagrass species have similar aboveground morphology, differences in their fine structures were detected. Diversities of motile animals inhabiting the seagrass species were the same. Differences in seagrass utilization were observed when we focused on a commercial shrimp, Pandalus latirostris. This shrimp always preferred higher densities of shoots irrespective of the species and seasonally changed their preference for the number of leaves per shoot. They were therefore more abundant in Z. caespitosa, which had those structural characteristics. The results suggest that the management of shrimp resources could be improved by flexibly changing protected areas in accordance with the dynamics of seagrass distribution in the lagoon.

Phylogeography of Littorina sitkana in the northwestern Pacific Ocean: evidence of eastward trans-Pacific colonization after the Last Glacial Maximum

We investigated genetic diversity and population structure of the Sitka periwinkle Littorina sitkana along the coastlines of the northwestern Pacific (NWP) to evaluate the possibility of trans-Pacific colonization of this species from the NWP to the northeastern Pacific (NEP) after the Last Glacial Maximum. We sampled L. sitkana from 32 populations in the NWP, and sequenced a region of the mitochondrial cytochrome b oxidase gene for population genetic analyses. The results were compared with those of previous reports from the NEP. The genetic diversity of L. sitkana was much higher in the NWP than in the NEP. Genetic connectivity between the NWP and NEP populations was indicated by an extremely abundant haplotype in the NEP that was also present in eastern Hokkaido and the Kuril Islands. To confirm these results, we compared sequences of the longest intron of the aminopeptidase N gene (APN54) in the nuclear genome in four populations of L. sitkana in the NWP with previous results from the NEP. Again, much higher genetic diversity was found in the NWP than in the NEP and genetic connectivity was supported between the Kuril Islands and the NEP. These results imply postglacial colonization of this species from the NWP to the NEP, probably along the Kuril and Aleutian Island chains. This study is the first report of possible trans-Pacific postglacial colonization of a direct-developing gastropod, inferred from genetic data.

Contributing to Restoration of Tidal Flats in Miyagi Prefecture’s Moune Bay Following the Great East Japan Earthquake and Tsunami

The Great East Japan Earthquake destroyed many artificial structures on the coast of northeastern Japan, and as a result it restored wetlands in many places. To conserve these wetlands, we started to estimate the ecological and economic values of a restored mudflat in Moune Bay, Miyagi Prefecture, Japan. Our tasks could be classified into three main categories: (1) establish the distribution of commercially important clams; (2) investigate the dynamics of the benthic community inhabiting the tidal flats and the floor of the bay; and (3) create a system so that monitoring of the tidal flat environment—including the two tasks mentioned—could be performed by nonspecialists. We hope that our actions will help to stimulate discussion about how Japan’s coastal areas should be used.