Short Note on the Effects of Cadmium on the Gill of the Glass Eel (Anguilla anguilla)

Freshwater-adapted glass eels have been exposed to cadmium (5 mg/l) for 3 days and then transferred in clean water for 28 days. Perturbations occurred very early (after 2 hrs only of exposure to cadmium) and concerned mainly the respiratory epithelium: the epithelial cells were very swollen and they showed an important dilation of the endoplasmic reticulum and nuclear membrane. After the decontamination period, signs of recovery appear. Perturbations of osmo-regulatory and respiratory processes are suggested.     

Variation in Population and Life History Traits of the American Eel, Anguilla rostrata, in Four Rivers in Maine

We examined population traits of yellow American eels from nine sites with similar habitat characteristics in each of four rivers in Maine, U.S.A. Migrating silver eels were also collected to compare sex ratio, age and size at migration among the four rivers. Population density and biomass were not significantly different among rivers with mean ranges of 8.4–21.8 eels 100m^–2and 380–1485gm^–2. Pairwise comparisons of the slopes of weight–length relationships of log transformed data (pooled data: intercept = –6.007, slope = 3.094, r²= 0.99, and n = 3116) revealed no significant differences among rivers. Length–age relationships (pooled data: intercept = 87.826, slope = 23.444, r²= 0.76, and n = 2325) also showed no statistically significant pairwise differences in slopes among rivers. In all rivers, sexual differentiation was complete by 270mm total length and age eleven. The sex ratios of migrating silver eels were not correlated with yellow eel sex ratios among the four rivers. Mean age at migration among the four rivers was significantly different for males only, with a range of 1.3years. Both sexes had some significant differences in size at migration among rivers, but the biological importance of the differences is tenuous (male range: 15mm, female range: 36mm). The yellow and silver eel population traits from these four rivers showed little variation when riverine habitat was isolated. Variations in traits appeared to be greater when eels from non-riverine habitats may have been present.     

Temperature behavioral responses of the American eel,Anguilla rostrata (Lesueur), from Maryland

Acute temperature preference tests were conducted with American eels, Anguilla rostrata, collected from Maryland's eastern shore. Eels were acclimated to temperatures of 6, 12, 18, 24 and 30°C. Final temperature preferendum was 16.7°C. Data differ from the temperature responses of the majority of fishes tested to date in that acclimation temperature did not influence selected temperatures. Similar results were obtained for various other fishes (Oncorhynchus, Salmo, Salvelinus) by other investigators. Behavioral responses at various acclimation temperatures were observed.     

Genomic footprints of speciation in Atlantic eels (Anguilla anguilla and A. rostrata)

The importance of speciation‐with‐geneflow scenarios is increasingly appreciated. However, the specific processes and the resulting genomic footprints of selection are subject to much discussion. We studied the genomics of speciation between the two panmictic, sympatrically spawning sister species; European (Anguilla anguilla) and American eel (A. rostrata). Divergence is assumed to have initiated more than 3 Ma, and although low gene flow still occurs, strong postzygotic barriers are present. Restriction‐site‐associated DNA (RAD) sequencing identified 328 300 SNPs for subsequent analysis. However, despite the presence of 3757 strongly differentiated SNPs (F_ST > 0.8), sliding window analyses of F_ST showed no larger genomic regions (i.e. hundreds of thousands to millions of bases) of elevated differentiation. Overall F_ST was 0.041, and linkage disequilibrium was virtually absent for SNPs separated by more than 1000 bp. We suggest this to reflect a case of genomic hitchhiking, where multiple regions are under directional selection between the species. However, low but biologically significant gene flow and high effective population sizes leading to very low genetic drift preclude accumulation of strong background differentiation. Genes containing candidate SNPs for positive selection showed significant enrichment for gene ontology (GO) terms relating to developmental processes and phosphorylation, which seems consistent with assumptions that differences in larval phase duration and migratory distances underlie speciation. Most SNPs under putative selection were found outside coding regions, lending support to emerging views that noncoding regions may be more functionally important than previously assumed. In total, the results demonstrate the necessity of interpreting genomic footprints of selection in the context of demographic parameters and life‐history features of the studied species.     

Fecundity of the American eel Anguilla rostrata at 45° N in Maine, U.S.A.

The northern portion of the geographic range of the American eel Anguilla rostrata may contribute a great proportion of the reproductive potential to this panmictic species because of apparent increases in average female size and female percentage with latitude. The regressions of fecundity on body length and on body weight of 63 female eels captured at about 45° N latitude on their spawning migration to the sea were log F= 1·2601 + 2·9642 log L and log F= 4·1646+0·9153 log W , where F is fecundity, L is total length (cm), and W is total weight (g). Length and weight each explained about 90% of the variation in fecundity. Estimates of fecundity from counts of aliquots of eggs ranged from 1·84 million to 19·92 million eggs for eels ranging in length from 45 to 113 cm, nearly the range of sizes of migrating females reported in the literature. Fecundities of the American eel were greater than reported in one study at about 37° N and greater than reported for the European eel, A. anguilla , shortfin eel, A. australis , and longfin eel, A. dieffenbachii . If a geographic cline in fecundity does exist in American eels, it is established anew each generation because the species forms a single panmictic population.     

The first complete mitochondrial genome of the Indian Tent Turtle,Pangshura tentoria (Testudines: Geoemydidae): Characterization and comparative analysis

The characterization of a complete mitogenome is widely used in genomics studies for systematics and evolutionary research. However, the sequences and structural motifs contained within the mitogenome of Testudines taxa have rarely been examined. The present study decodes the first complete mitochondrial genome of the Indian Tent Turtle, Pangshura tentoria (16,657 bp) by using next‐generation sequencing. This denovo assembly encodes 37 genes: 13 protein‐coding genes (PCGs), 22 transfer RNA (tRNAs), two ribosomal RNA, and one control region (CR). Most of the genes were encoded on majority strand, except for one PCG (NADH dehydrogenase subunit 6) and eight tRNAs. Most of the PCGs were started with an ATG initiation codon, except for Cytochrome oxidase subunit 1 with “GTG” and NADH dehydrogenase subunit 5 with “ATA.” The termination codons, “TAA” and “AGA” were observed in two subunits of NADH dehydrogenase gene. The relative synonymous codon usage analysis revealed the maximum abundance of alanine, isoleucine, leucine, and threonine. The nonsynonymous/synonymous ratios were <1 in all PCGs, which indicates strong negative selection among all Geoemydid species. The study also found the typical cloverleaf secondary structure in most of the tRNA genes, except for serine with the lack of the conventional DHU arm. The comparative study of Geoemydid mitogenomes revealed the occurrence of tandem repeats was frequent in the 3′ end of CR. Further, two copies of a unique tandem repeat “TTCTCTTT” were identified in P. tentoria. The Bayesian and maximum‐likelihood phylogenetic trees using concatenation of 13 PCGs revealed the close relationships of P. tentoria with Batagur trivittata in the studied dataset. All the Geoemydid species showed distinct clustering with high bootstrap support congruent with previous evolutionary hypotheses. We suggest that the generations of more mitogenomes of Geoemydid species are required, to improve our understanding of their in‐depth phylogenetic and evolutionary relationships.     

Comments on a study of the thermal behaviour of the American eel (Anguilla rostrata) and some statistical suggestions for temperature preference studies

Some suggestions are made concerning methods and statistical evaluation of results in temperature preference studies. The use of regression analysis is advocated rather than graphical methods for evaluating results from acute temperature preference studies and a method of assessing the final preferendum and its confidence interval is demonstrated. This method is applied to a previously published study of the acutely preferred temperature of the American eel (Barila & Stauffer, 1980). The final preferendum and its 95% confidence interval is shown to be 17.4° ± 2.0 °C. In direct contrast to conclusions there drawn, the preferred temperature is shown to depend on the acclimated temperature. Some critical comments are also made concerning other parts of the study.     

Chromosome‐level genome assembly of the predator Propylea japonica to understand its tolerance to insecticides and high temperatures

The ladybird beetle Propylea japonica is an important natural enemy in agro‐ecological systems. Studies on the strong tolerance of P. japonica to high temperatures and insecticides, and its population and phenotype diversity have recently increased. However, abundant genome resources for obtaining insights into stress‐resistance mechanisms and genetic intra‐species diversity for P. japonica are lacking. Here, we constructed the P. japonica genome maps using Pacific Bioscience (PacBio) and Illumina sequencing technologies. The genome size was 850.90 Mb with a contig N50 of 813.13 kb. The Hi‐C sequence data were used to upgrade draft genome assemblies; 4,777 contigs were assembled to 10 chromosomes; and the final draft genome assembly was 803.93 Mb with a contig N50 of 813.98 kb and a scaffold N50 of 100.34 Mb. Approximately 495.38 Mb of repeated sequences was annotated. The 18,018 protein‐coding genes were predicted, of which 95.78% were functionally annotated, and 1,407 genes were species‐specific. The phylogenetic analysis showed that P. japonica diverged from the ancestor of Anoplophora glabripennis and Tribolium castaneum ~ 236.21 million years ago. We detected that some important gene families involved in detoxification of pesticides and tolerance to heat stress were expanded in P. japonica, especially cytochrome P450 and Hsp70 genes. Overall, the high‐quality draft genome sequence of P. japonica will provide invaluable resource for understanding the molecular mechanisms of stress resistance and will facilitate the research on population genetics, evolution and phylogeny of Coccinellidae. This genome will also provide new avenues for conserving the diversity of predator insects.     

Cloning and Sequencing of Cytochrome P450 1A Complementary DNA in Eel (Anguilla japonica)

Cytochrome P450 1A (CYP1A) complementary DNA was isolated from eel (Anguilla japonica) liver treated with 3-methylcholanthrene. The cDNA contained a 5′ untranslated region of 163 bp, an open reading flame of 1560 bp coding for 519 amino acids and a stop codon, and a 3′ untranslated region of 1730 bp. The predicted molecular weight was approximately 58.4 kDa. The deduced amino acid sequence exhibited identities with reported CYP1A sequences of 80% for rainbow trout, 79% for scup, 76% for plaice and butterfly fish, and 74% for toadfish. When compared with mammalian CYP proteins, the eel CYP1A was more similar to CYP1A1 (54%–56%) than to CYP1A2 (49%–52%). Northern and Southern blot analyses showed two distinct bands, suggesting the existence of another 3-methylcholanthrene-inducible CYP1A gene in eel. Received December 19, 1998; accepted February 18, 1999     

Population genetics of the American eel (Anguilla rostrata): FST = 0 and North Atlantic Oscillation effects on demographic fluctuations of a panmictic species

We performed population genetic analyses on the American eel (Anguilla rostrata) with three main objectives. First, we conducted the most comprehensive analysis of neutral genetic population structure to date to revisit the null hypothesis of panmixia in this species. Second, we used this data to provide the first estimates of contemporary effective population size (N_e) and to document temporal variation in effective number of breeders (N_b) in American eel. Third, we tested for statistical associations between temporal variation in the North Atlantic Oscillation (NAO), the effective number of breeders and two indices of recruit abundance. A total of 2142 eels from 32 sampling locations were genotyped with 18 microsatellite loci. All measures of differentiation were essentially zero, and no evidence for significant spatial or temporal genetic differentiation was found. The panmixia hypothesis should thus be accepted for this species. N_b estimates varied by a factor of 23 among 12 cohorts, from 473 to 10 999. The effective population size N_e was estimated at 10 532 (95% CI, 9312–11 752). This study also showed that genetically based demographic indices, namely N_b and allelic richness (Ar), can be used as surrogates for the abundance of breeders and recruits, which were both shown to be positively influenced by variation during high (positive) NAO phases. Thus, long‐term genetic monitoring of American glass eels at several sites along the North American Atlantic coast would represent a powerful and efficient complement to census monitoring to track demographic fluctuations and better understand their causes.     

Diel and seasonal movements of silver eels,Anguilla dieffenbachii,emigrating from a lake subject to hydro‐electric control

To investigate the behaviour and fate of silver female longfin eels Anguilla dieffenbachii in relation to flow variations from New Zealand's largest (700 MW) hydro‐electric station, 210 A. dieffenbachii were tagged with acoustic transmitters during summer and autumn over 6 years from 2005 to 2010. A shoreline array of receivers (2005–2008) was supplemented by positioning systems near the power station and natural outlet (2009–2010) that enabled continuous tracking of A. dieffenbachii within these areas. Tagged A. dieffenbachii (total length 785–1372 mm) emigrated from the lake over an 8 month period, either in the same season they were tagged (75%) or the following spring after over‐wintering for up to 7 months (25%). Most A. dieffenbachii (67%) emigrated via the natural outlet. Swimming activity was related to water temperature and was greatest during the summer months. Activity was strongly diel, although less so at the outlet where A. dieffenbachii migrating at depths greater than 20 m often did so during daylight. To analyse searching behaviour, sustained periods of continuous swimming activity (termed forays) were examined in the vicinity of the outlet. Most successful forays (where the A. dieffenbachii exited the lake via the lower Waiau River) were associated with outflows; such outflows were sometimes <10 m³ s^?1 although most successful forays took place at flows >50 m³ s^?1_. The timing of successful forays was related to outflow, the day of year and whether the A. dieffenbachii had delayed migration or not, but not lunar phase or rainfall. Passage time from the lake to the lower Waiau River 81 km downstream decreased from an average of 58 days early in the season to <1 day in late autumn. The wide range of behaviours and responses displayed by female silver A. dieffenbachii were interpreted as an inherent form of bet‐hedging to spread the risks associated with silvering and seaward migration in this slow‐growing species.     

Haplotype‐based genotyping‐by‐sequencing in oat genome research

In a de novo genotyping‐by‐sequencing (GBS) analysis of short, 64‐base tag‐level haplotypes in 4657 accessions of cultivated oat, we discovered 164741 tag‐level (TL) genetic variants containing 241224 SNPs. From this, the marker density of an oat consensus map was increased by the addition of more than 70000 loci. The mapped TL genotypes of a 635‐line diversity panel were used to infer chromosome‐level (CL) haplotype maps. These maps revealed differences in the number and size of haplotype blocks, as well as differences in haplotype diversity between chromosomes and subsets of the diversity panel. We then explored potential benefits of SNP vs. TL vs. CL GBS variants for mapping, high‐resolution genome analysis and genomic selection in oats. A combined genome‐wide association study (GWAS) of heading date from multiple locations using both TL haplotypes and individual SNP markers identified 184 significant associations. A comparative GWAS using TL haplotypes, CL haplotype blocks and their combinations demonstrated the superiority of using TL haplotype markers. Using a principal component‐based genome‐wide scan, genomic regions containing signatures of selection were identified. These regions may contain genes that are responsible for the local adaptation of oats to Northern American conditions. Genomic selection for heading date using TL haplotypes or SNP markers gave comparable and promising prediction accuracies of up to r = 0.74. Genomic selection carried out in an independent calibration and test population for heading date gave promising prediction accuracies that ranged between r = 0.42 and 0.67. In conclusion, TL haplotype GBS‐derived markers facilitate genome analysis and genomic selection in oat.     

Helping decision making for reliable and cost‐effective 2b‐RAD sequencing and genotyping analyses in non‐model species

High‐throughput sequencing has revolutionized population and conservation genetics. RAD sequencing methods, such as 2b‐RAD, can be used on species lacking a reference genome. However, transferring protocols across taxa can potentially lead to poor results. We tested two different IIB enzymes (AlfI and CspCI) on two species with different genome sizes (the loggerhead turtle Caretta caretta and the sharpsnout seabream Diplodus puntazzo) to build a set of guidelines to improve 2b‐RAD protocols on non‐model organisms while optimising costs. Good results were obtained even with degraded samples, showing the value of 2b‐RAD in studies with poor DNA quality. However, library quality was found to be a critical parameter on the number of reads and loci obtained for genotyping. Resampling analyses with different number of reads per individual showed a trade‐off between number of loci and number of reads per sample. The resulting accumulation curves can be used as a tool to calculate the number of sequences per individual needed to reach a mean depth ≥20 reads to acquire good genotyping results. Finally, we demonstrated that selective‐base ligation does not affect genomic differentiation between individuals, indicating that this technique can be used in species with large genome sizes to adjust the number of loci to the study scope, to reduce sequencing costs and to maintain suitable sequencing depth for a reliable genotyping without compromising the results. Here, we provide a set of guidelines to improve 2b‐RAD protocols on non‐model organisms with different genome sizes, helping decision‐making for a reliable and cost‐effective genotyping.     

Isolation and characterization of expressed sequence tag-linked microsatellite loci for the European eel (Anguilla anguilla)

A European eel (Anguilla anguilla) expressed sequence tag database consisting of 795 contigs and 4008 singletons was screened for microsatellites sequences. Primers were designed to amplify 96 repeats, of which 86 gave good quality amplification products. Twenty-eight microsatellites were selected for further microsatellite genotyping. Only two loci were found to be monomorphic; out of the 26 polymorphic loci, number of alleles per locus ranged from two to 14, while the observed and expected heterozygosities ranged from 0.05 to 0.93, and from 0.05 to 0.95, respectively. All 28 primer sets tested revealed positive amplification in American eel (Anguilla rostrata).