Transcriptome analysis of reproductive tract tissues of male river prawn Macrobrachium americanum.

BACKGROUND: The river prawn, Macrobrachium americanum (M. americanum), is one of the largest prawns of the genus in Latin America and is an amphidromous species distributed along the Pacific coast of America. This prawn has commercial value due to its size and taste, making it a good option for aquaculture production. Its culture has been attempted in ponds and concrete tanks, but no successful technique can still support commercial production. Understanding the mechanisms that regulate reproduction at the molecular level is very important. This knowledge can provide tools for manipulating transcripts, which could increase the number or size of animals in the culture. Our understanding of the mechanism that regulates the reproduction of M. americanum at the molecular level is limited. AIM: Perform and analyze the transcriptome assembly of the testes, vas deferens, and terminal ampulla of M. americanum. to provide new molecular information about its reproduction. METHODS AND RESULTS: The cDNA library was constructed and sequenced for each tissue to identify novel transcripts. A combined transcriptome with the three tissues was assembled using Trinity software. Unigenes were annotated using BLASTx and BLAST2GO. The transcriptome assembly generated 1,059,447 unigenes, of which 7222 genes had significant hits (e-value < 1 × 10-5) when compared against the Swiss-Prot database. Around 75 genes were related to sex determination, testis development, spermatogenesis, spermiogenesis, fertilization, maturation of testicular cells, neuropeptides, hormones, hormone receptors, and/or embryogenesis. CONCLUSIONS: These results provide new molecular information about M. americanum reproduction, representing a reference point for further genetic studies of this species.

Distinct development trajectories and symbiosis modes in vent shrimps.

Most animal species have a singular developmental pathway and adult ecology, but developmental plasticity is well-known in some such as honeybees where castes display profoundly different morphology and ecology. An intriguing case is the Atlantic deep-sea hydrothermal vent shrimp pair Rimicaris hybisae and R. chacei that share dominant COI haplotypes and could represent very recently diverging lineages or even morphs of the same species. Rimicaris hybisae is symbiont-reliant with a hypertrophied head chamber (in the Mid-Cayman Spreading Centre), while R. chacei is mixotrophic with a narrow head chamber (on the Mid-Atlantic Ridge). Here, we use X-ray micro-computed tomography and fluorescence in situ hybridization to show that key anatomical shifts in both occur during the juvenile-subadult transition, when R. hybisae has fully established symbiosis but not R. chacei. On the Mid-Atlantic Ridge, the diet of R. chacei has been hypothetically linked to competition with the obligatorily symbiotic congener R. exoculata, and we find anatomical evidence that R. exoculata is indeed better adapted for symbiosis. We speculate the possibility that the distinct development trajectories in R. hybisae and R. chacei may be determined by symbiont colonization at a "critical period" before subadulthood, though further genetic studies are warranted to test this hypothesis along with the true relationship between R. hybisae and R. chacei.

Molecular phylogeny of deep-sea blind lobsters of the family Polychelidae (Decapoda: Polychelida), with implications for the origin and evolution of these "living fossils".

A comprehensive molecular analysis of the deep-sea blind lobsters of the family Polychelidae, often referred to as "living fossils", is conducted based on all six modern genera and 27 of the 38 extant species. Using six genetic markers from both mitochondrial and nuclear genomes, the molecular phylogenetic results differ considerably from previous morphological analyses and reveal the genera Polycheles and Pentacheles to be para- or polyphyletic. As the splitting of Polycheles has strong support from both molecular and morphological data, two new genera, Dianecheles and Neopolycheles, are erected for those species excluded from the clade containing the type species of Polycheles. The pattern of polyphyly of Pentacheles, however, is not robustly resolved, so it is retained as a single genus. Fossil evidence suggests that fossil polychelids inhabited deep-sea environments as early as the Early to Middle Jurassic, demonstrating the enduring adaptation of extant polychelid species to the deep-sea. Time-calibrated phylogeny suggested that modern polychelids probably had an Atlantic origin during the Jurassic period. Since their emergence, this ancient lobster group has continued to diversify, particularly in the West Pacific, and has colonized the abyssal zone, with the deepest genus, Willemoesia, representing the more 'derived' members among extant polychelids. Differences in eye reduction among extant polychelid genera highlight the necessity for ongoing investigations to ascertain the relative degree of functionality of their eyes, if they indeed retain any function.

Morphotype induced changes in the life history and population dynamics of an hippolytid shrimp.

One of the most exceptional, loud paradigm of shape polymorphisms constitutes the "rostral loss" condition in hippolytid shrimps. The intertidal shrimp Hippolyte sapphica includes two conspecific morphotypes, one of which demonstrates a neotenic rostrum (morph-B). Morphs' rostral elongation is controlled by a single genetic locus, with long rostra (morph-A) representing the recessive state and short, larval-like ones the completely dominant state. Geometrics morphometry on the species morphotypes revealed also the homozygous/heterozygous state of the gene site along with some induced body' adaptations, which compete the micro-evolutionary disadvantage of the "rostral loss". We found recently that females' viability and maternal energy investment selectively favors morph-A. The present contribution detects and discuss comparatively demographic and reproductive traits in species mixed (both morphs) and unmixed populations. Our results show that this sharp dimorphic rostral condition is a sex-related marker and that the species is gonochoric. Presence of morph-B results to (a) lower egg production (b) higher seasonally males' percentage (c) morph-A females' earlier maturation and (d) higher fecundity in morph-A mixed populations. It seems that the "rostral loss" state induces complex adaptations between the two morphotypes through sex ratios equilibria, morphotypes' growth rates, and morphs' fecundity differentiate inputs throughout the seasons.

Comparative Analysis of Transposable Elements Reveals the Diversity of Transposable Elements in Decapoda and Their Effects on Genomic Evolution.

Transposable elements (TEs) are mobile genetic elements that exist in the host genome and exert considerable influence on the evolution of the host genome. Since crustaceans, including decapoda, are considered ideal models for studying the relationship between adaptive evolution and TEs, TEs were identified and classified in the genomes of eight decapoda species and one diplostraca species (as the outgroup) using two strategies, namely homology-based annotation and de novo annotation. The statistics and classification of TEs showed that their proportion in the genome and their taxonomic composition in decapoda were different. Moreover, correlation analysis and transcriptome data demonstrated that there were more PIF-Harbinger TEs in the genomes of Eriocheir sinensis and Scylla paramamosain, and the expression patterns of PIF-Harbingers were significantly altered under air exposure stress conditions. These results signaled that PIF-Harbingers expanded in the genome of E. sinensis and S. paramamosain and might be related to their air exposure tolerance levels. Meanwhile, sequence alignment revealed that some Jockey-like sequences (JLSs) with high similarity to specific regions of the White spot syndrome virus (WSSV) genome existed in all eight decapod species. At the same time, phylogenetic comparison exposed that the phylogenetic tree constructed by JLSs was not in agreement with that of the species tree, and the distribution of each branch was significantly different. The abovementioned results signaled that these WSSV-specific JLSs might transfer horizontally and contribute to the emergence of WSSV. This study accumulated data for expanding research on TEs in decapod species and also provided new insights and future direction for the breeding of stress-resistant and disease-resistant crab breeds.

Penaeid Shrimp Chromosome Studies Entering the Post-Genomic Era.

Chromosome studies provide the foundation for comprehending inheritance, variation, systematics, and evolution. Penaeid shrimps are a group of crustaceans with great economic importance. Basic cytogenetic information obtained from these shrimps can be used to study their genome structure, chromosome relationships, chromosome variation, polyploidy manipulation, and breeding. The study of shrimp chromosomes experienced significant growth in the 1990s and has been closely linked to the progress of genome research since the application of next-generation sequencing technology. To date, the genome sequences of five penaeid shrimp species have been published. The availability of these genomes has ushered the study of shrimp chromosomes into the post-genomic era. Currently, research on shrimp cytogenetics not only involves chromosome counting and karyotyping, but also extends to investigating submicroscopic changes; exploring genome structure and regulation during various cell divisions; and contributing to the understanding of mechanisms related to growth, sexual control, stress resistance, and genome evolution. In this article, we provide an overview of the progress made in chromosome research on penaeid shrimp. We emphasize the mutual promotion between studies on chromosome structure and genome research and highlight the impact of chromosome-level assembly on studies of genome structure and function. Additionally, we summarize the emerging trends in post-genomic-era shrimp chromosome research.

Exploring the mitochondrial genome of Caridina pseudogracilirostris: a comparative analysis within the Atyidae Family.

BACKGROUND: Caridina pseudogracilirostris is a highly adaptive estuarine species found in brackish waters and marshes along the southwestern and southern coastal regions of India. METHODS AND RESULTS: The whole mitochondrial genome of C. pseudogracilirostris is 15,451 bp in length with 59.3% AT content and encodes 37 genes, including 22 tRNAs, 13 protein-coding genes, and two rRNAs, which are arranged in a distinctive pattern similar to most crustaceans. ML and BI methods were used for phylogenetic analysis of C. pseudogracilirostris clustered with other Caridina species, supporting the monophyly of the Caridina genus within the Atyidae family. The fully annotated mitochondrial genome of C. pseudogracilirostris was submitted to GenBank under accession number OQ534868.1. CONCLUSIONS: We are the first to report on the C. pseudogracilirostris whole mitochondrial genome, which provides a valuable resource for future research on genetics, evolution, phylogenetics, etc., among Caridina species and other species. The phylogenetic investigation supports the monophyly of the Caridina genus within the Atyidae family and emphasizes the value of mitochondrial genome data in determining the evolutionary relationships among crustaceans.

A new species of the mud shrimp genus Naushonia Kingsley, 1897 (Decapoda: Gebiidea: Laomediidae) from the Ryukyu Islands, southwestern Japan, inhabiting burrows of an axiidean shrimp.

A new species of the laomediid mud shrimp genus Naushonia Kingsley, 1897 is described and illustrated based on seven specimens collected from Okinawa Island, Ryukyu Islands, southwestern Japan. All specimens of Naushonia karashimai n. sp. were collected from burrows of a large axiidean shrimp, Neaxius acanthus (A. Milne-Edwards, 1879), which inhabits seagrass beds in inner reef lagoons. The new species is easily distinguished from all 16 known congeners by its tridentate rostrum and relatively narrow pereopod 1 palm with a proximally located pollex. Sequences of the mitochondrial 16S rRNA and COI genes were newly generated for five species and two species of Naushonia, including the new species, respectively. Preliminary phylogenetic analysis using sequences of the 16S rRNA gene was performed.

Comparison of Mitochondrial Genome Sequences between Two Palaemon Species of the Family Palaemonidae (Decapoda: Caridea): Gene Rearrangement and Phylogenetic Implications.

To further understand the origin and evolution of Palaemonidae (Decapoda: Caridea), we determined the mitochondrial genome sequence of Palaemon macrodactylus and Palaemon tenuidactylus. The entire mitochondrial genome sequences of these two Palaemon species encompassed 37 typical genes, including 13 protein-coding genes (PCGs), 2 ribosomal RNA genes (rRNAs), and 22 transfer RNA genes (tRNAs), and a control region (CR). The lengths of their mitochondrial genomes were 15,744 bp (P. macrodactylus) and 15,735 bp (P. tenuidactylus), respectively. We analyzed their genomic features and structural functions. In comparison with the ancestral Decapoda, these two newly sequenced Palaemon species exhibited a translocation event, where the gene order was trnK-trnD instead of trnD-trnK. Based on phylogenetic analysis constructed from 13 PCGs, the 12 families from Caridea can be divided into four major clades. Furthermore, it was revealed that Alpheidae and Palaemonidae formed sister groups, supporting the monophyly of various families within Caridea. These findings highlight the significant gene rearrangements within Palaemonidae and provide valuable evidence for the phylogenetic relationships within Caridea.

Assessment of genetic diversity and population structure of Neocaridina denticulata sinensis in the Baiyangdian drainage area, China, using microsatellite markers and mitochondrial cox1 gene sequences.

Neocaridina denticulata sinensis is a crustacean of major economic significance in the Baiyangdian drainage area. In this study, the first assessment of N. denticulata sinensis genetic diversity and population structure was performed based on sequence analysis of nine polymorphic microsatellite loci and the mitochondrial cytochrome oxidase subunit I (cox1) gene. Samples (n = 192) were collected from four different regions in the Baiyangdian drainage area i.e., Baiyangdian Lake, Jumahe River, Xidayang Reservoir, and Fuhe River. Microsatellite loci analysis identified high levels of genetic diversity represented by observed heterozygosity (Ho) of 0.6865 âˆ¼ 0.9583, expected heterozygosity (He) of 0.7151 âˆ¼ 0.8723, and polymorphism information content (PIC) of 0.6676 âˆ¼ 0.8585. Based on the analysis of cox1 sequences, haplotype diversity (Hd) ranged from 0.568 to 0.853 while nucleotide diversity (π) ranged from 0.0029 to 0.2236. Furthermore, there was no evidence of expansion events in the N. denticulata sinensis populations. Pairwise FST revealed pronounced genetic differentiation, and clustering analyses showed defined genetic structures within the N. denticulata sinensis population. Three groups were identified from four sampled stocks, with Xidayang Reservoir, and Fuhe River populations clustered in the same group. This work identified novel molecular markers and provided an important reference to guide management strategies to assist conservation of N. denticulata sinensis resources.

Immune responses and histopathological analyses of giant river prawn (Macrobrachium rosenbergii, De Man 1879) challenged with a sub-lethal dose of decapod iridescent virus 1 (DIV1) and chemical control investigation.

Decapod iridescent virus 1 (DIV1) is a lethal virus that has a significant influence on the shrimp and prawn culture industries. The mechanism through which infected prawns respond to the DIV1 virus is currently unknown. Here, we examined in detail the clinical signs, histopathology, and humoral, cellular, and immune-related gene responses after a sub-lethal dose of DIV1 during the acute infection period of 0-120 h post infection (hpi). Interestingly, at the end of the experiment, DIV1-infected prawns had black lesions on several external regions. The DIV1-infected prawns also exhibited few karyopyknotic nuclei in the gills and intestine tissues and exhibited increasing immunological responses, as revealed by significant increases in all examined parameters, including total hemocytes, phagocytosis, lysozyme, and overall bactericidal activity, from 6 to 48 hpi. In addition, between 72 and 120 hpi, all immune response activities of DIV1-infected prawn were impaired compared with those of normal prawns, indicating negative impacts on immunological parameters. A viral load analysis of various tissues by qPCR indicated that hemocytes were the dominant initial viral target tissues, followed by the gills and hepatopancreas. An expression analysis of crucial immune-related genes by qRT‒PCR revealed various expression patterns in response to DIV1 infection; in particular, fold changes in the relative expression of anti-lipopolysaccharide factors (ALFs), prophenoloxidase (proPO), lipopolysaccharide and ß-1,3-glucan binding protein (LGBP) were observed. Additionally, five common chemicals, calcium hypochlorite [Ca(OCl)2] at 16.25-130 ppm, hydrogen peroxide (H2O2) at 8.75-70 ppm, povidone iodine (PVP-I) at 3-24 ppm, benzalkonium chloride (BKC) at 20-160 ppm, and formalin at 25-200 ppm, had a significant effect on the killing of DIV1 particles in vitro within 24 h after exposure. These data will be helpful for determining the health status and immune defense mechanisms of giant river prawns during DIV1 infection periods. The study performed the first application of very common disinfectants, and the obtained information will be useful for implementing effective strategies to prevent and control DIV1 infection in both hatchery and grow-out ponds.

Occurrence of Deep-sea shrimp, Hadropenaeus lucasii (Spence Bate, 1881) from the southwest coast of India.

The present study provides an integrative report combining morphological and molecular analysis of the deep sea shrimp species Hadropenaeus lucasii (Spence Bate, 1881) from the southwest coast of India. The present specimen was obtained from the depths of 200-300m from the commercial bottom trawlers operated off Sakthikulangara fishing harbour off Kollam, Kerala. A phylogenetic analysis was used to explore the relationships of the genus. DNA barcoding and phylogenetic analysis were used to explore the relationship of the genus Hadropenaeus based on mitochondrial gene (16S: OK571387, OK571388; COI: OK569849, OK569850) sequences of the present specimen with the sequences retrieved from NCBI GenBank revealed an interspecies genetic divergence of 0.0% to 0.6%.

Genetic variation and cryptic lineage among the sergestid shrimp Acetes americanus (Decapoda).

The taxonomic status of the sergestid shrimp, Acetes americanus, has been questioned for several decades. No specific study has been performed thus far to resolve the incongruences. This species has a wide geographical range in the western Atlantic and is represented by two formally accepted subspecies: Acetes americanus carolinae, distributed in North America, and Acetes americanus americanus, present in South America. However, there are regions where the coexistence of both subspecies has been reported, such as Central America. This study aimed to genetically compare specimens of A. a. americanus collected in South America with A. a. carolinae sampled in North America to check for possible differences and the existence of more than one subspecies of A. americanus on the Brazilian coast. Based on the sequences of two informative markers, the cytochrome oxidase I region (COI) and 16S rRNA, phylogenetic reconstruction demonstrated well-defined clades with high support values, reinforcing the idea that A. a. americanus is genetically different from A. a. carolinae. Our hypothesis was corroborated as the specimens collected in Brazil were divided into two distinct lineages: the first composed of A. a. americanus sensu stricto (Brazil 1) and the second by Acetes americanus (Brazil 2). The three groups evidenced in the haplotype network were the same as those observed in the phylogenetic tree. The morphometric character (height/length of the thelycum) was effective in distinguishing A. a. Brazil 1 from A. a. carolinae. However, more detailed and conclusive studies comprising other characteristics to propose and describe a possible new entity are necessary. To the best of our knowledge, for the first time, the results of this study provide some insights into the taxonomic status of the sergestid shrimp A. americanus in the western Atlantic.

The Coding Mitogenome of the Freshwater Crayfish Pontastacus leptodactylus (Decapoda:Astacidea:Astacidae) from Lake Vegoritida, Greece and Its Taxonomic Classification.

Pontastacus leptodactylus (Eschscholtz, 1823) (Decapoda:Astacidea:Astacidae) constitutes an ecologically and economically highly important species. In the present study, the mitochondrial genome of the freshwater crayfish P. leptodactylus from Greece is analyzed for the first time, using 15 newly designed primer pairs based on available sequences of closely related species. The analyzed coding part of the mitochondrial genome of P. leptodactylus consists of 15,050 base pairs including 13 protein-coding genes (PCGs), 2 ribosomal RNA gene (rRNAs), and 22 transfer RNA genes (tRNAs). These newly designed primers may be particularly useful in future studies for analyzing different mitochondrial DNA segments. Based on the entire mitochondrial genome sequence, compared to other haplotypes from related species belonging in the same family (Astacidae) available in the GenBank database, a phylogenetic tree was constructed depicting the phylogenetic relationships of P. leptodactylus. Based on the results, the genetic distance between Astacus astacus and P. leptodactylus is smaller than the genetic distance between Austropotamobius pallipes and Austropotamobius torrentium, despite the fact that the latter two are classified within the same genus, questioning the phylogenetic position of A. astacus as a different genus than P. leptodactylus. In addition, the sample from Greece seems genetically distant compared with a conspecific haplotype available in the GenBank database, possibly implying a genetic distinction of P. leptodactylus from Greece.

Genome-wide identification and expression profiling of Wnt gene family in Neocaridina denticulata sinensis.

Wnt proteins are a class of hydrophobic secreted glycoproteins involved in diverse important biological processes, such as tissue formation and regeneration, embryonic development and innate immunity. The Wnt gene family has an early origin and is present in all deuterostomes. In the process of evolution, the phenomenon of gene expansion, contraction and adaptive evolution occurs in the Wnt gene family. In the current study, eleven Wnt genes (NdWnt1-2, NdWnt4-7, NdWnt9-11, NdWnt16, and NdWntA) belonging to different subfamilies were obtained based on the genomic and transcriptomic data of Neocaridina denticulata sinensis. Then the expression patterns of all NdWnts were analyzed in various tissues, at different developmental stages and under different stresses. The expression profiles of NdWnts at different developmental stages showed that most NdWnt genes were initially expressed at gastrula stage, and the expression of NdWnt5 and NdWnt16 throughout all developmental stages. The spatial expression of NdWnt genes presented tissue specificity. They were mainly expressed in four tissues, namely gill, intestines, ovary and eyestalk. After Vibrio parahemolyticus infection and under copper exposure, the expression levels of five NdWnts (NdWnt1, NdWnt5, NdWnt10, NdWnt16 and NdWntA) were variable. Our findings enrich the research on the Wnt gene family of N. denticulata sinensis and provide valuable insights into relationship between structure and function of Wnt genes in crustaceans.

Molecular cloning and expression patterns of a sex-biased transcriptional factor Foxl2 in the giant freshwater prawn (Macrobrachium rosenbergii).

BACKGROUD: Macrobrachium rosenbergii is an economically important species that is widely cultivated in some Asian nations. Foxl2 is a transcriptional regulator of ovarian differentiation and development. The aim of this study was to study the bioinformatics features and expression patterns of M. rosenbergii Foxl2 (MrFoxl2). METHODS: In this study, all experimental animals were mature M. rosenbergii (9-12 cm) individuals. The foxl2 gene was identified and characterized in the genome of M. rosenbergii using molecular cloning, bioinformatic analysis, in situ hybridization, and quantitative analysis. RESULTS: The identified cDNA encoded a putative 489-amino-acid MrFoxl2 protein. Bioinformatics analysis revealed a low identity of MrFoxl2 to other crustacean orthologues. The closest phylogenetic relationship was to Foxl2 of Eriocheir sinensis. The result of in situ hybridization demonstrated that transcripts of MrFoxl2 in M. rosenbergii were identified in spermatocytes, oocytes, and secretory epithelial cells of the vas deferens. The result of q-PCR suggested that a high expression of MrFoxl2 was identified in the testis, vas deferens, and ovaries. During ovarian development, MrFoxl2 expression was the highest in stage I. CONCLUSION: Our findings suggest that MrFoxl2 may play a role in gonadal development in both female and male M. rosenbergii.

Genomic analysis reveals strong population structure in the Giant Sydney Crayfish (Euastacus spinifer (Heller, 1865)).

Australia is home to over 140 species of freshwater crayfish (Decapoda: Parastacidae), representing a centre of diversity for this group in the Southern Hemisphere. Species delimitation in freshwater crayfish is difficult because many species show significant variation in colouration and morphology. This is particularly evident in the genus Euastacus, which exhibits large variations in colour and spination throughout its putative range. To understand this variation, we investigated the genetic diversity, population structure, phylogeny, and evolutionary timescale of the Giant Sydney Crayfish (Euastacus spinifer (Heller, 1865)). Our data set is sampled from over 70 individuals from across the ∼600 km range of the species, and includes a combination of two mitochondrial markers and more than 7000 single-nucleotide polymorphisms (SNPs) from the nuclear genome. Data were also obtained for representatives of the close relative, Euastacus vesper McCormack and Ahyong, 2017. Genomic SNP analyses revealed strong population structure, with multiple distinct populations showing little evidence of gene flow or migration. Phylogenetic analyses of mitochondrial data revealed similar structure between populations. Taken together, our analyses suggest that E. spinifer, as currently understood, represents a species complex, of which E. vesper is a member. Molecular clock estimates place the divergences within this group during the Pleistocene. The isolated and highly fragmented populations identified in our analyses probably represent relict populations of a previously widespread ancestral species. Periodic flooding events during the Pleistocene are likely to have facilitated the movement of these otherwise restricted freshwater crayfish within and between drainage basins, including the Murray-Darling and South East Coast Drainages. We present evidence supporting the recognition of populations in the southern parts of the range of E. spinifer as one or two separate species, which would raise the number of species within the E. spinifer complex to at least three. Our results add to the growing body of evidence that many freshwater crayfish exhibit highly fragmented, range-restricted distributions. In combination with the life-history traits of these species, the restricted distributions exacerbate the threats already placed on freshwater crayfish, which are among the five most endangered animal groups globally.

The marine palaemonid shrimps (Crustacea, Deapoda, Caridea) of the Dutch Caribbean.

Species of the decapod family Palaemonidae are common components of tropical coastal waters and coral reefs. The majority of these species are symbionts of various invertebrate phyla. Despite a long history of research on their species diversity in the Dutch Caribbean, recent field expeditions have yielded much new information. Combined with examinations of specimens housed in Naturalis Biodiversity Center and information from literature, a comprehensive list of Dutch Carribean palaemonids is provided. Newly collected material was primarily identified via morphological analyses. Additional molecular phylogenetic analyses based on mitochondrial COI and 16S and nuclear Histone 3 (H3) genes were conducted in search of cryptic species on the one hand and to check conspecifity in species that were found on multiple host species on the other hand. In total, 46 species are here listed for the Dutch Caribbean of which 24 are here recorded for the first time for one of the islands. One species new to science was discovered and is herein described. Sixty new host associations are recorded. In light of biodiversity loss and increasing anthropogenic pressure on declining coral reefs, documenting the diversity of palaemonids and other coral reef species to provide baseline data takes on a new urgency.

Characterization of the Cambaroides wladiwostokiensis Birstein & Vinogradov, 1934 (Decapoda: Astacidea) Mitochondrial Genome Using Genome Skimming and the Phylogenetic Implications within the Astacidea Infraorder.

BACKGROUND: The mitochondrial genome is a powerful tool for exploring and confirming species identity and understanding evolutionary trajectories. The genus Cambaroides, which consists of freshwater crayfish, is recognized for its evolutionary and morphological complexities. However, comprehensive genetic and mitogenomic data on species within this genus, such as C. wladiwostokiensis, remain scarce, thereby necessitating an in-depth mitogenomic exploration to decipher its evolutionary position and validate its species identity. METHODS: The mitochondrial genome of C. wladiwostokiensis was obtained through shallow Illumina paired-end sequencing of total DNA, followed by hybrid assembly using both de novo and reference-based techniques. Comparative analysis was performed using available Cambaroides mitochondrial genomes obtained from National Center for Biotechnology Information (NCBI). Additionally, phylogenetic analyses of 23 representatives from three families within the Astacidea infraorder were employed using the PhyloSuite platform for sequence management and phylogenetic preparation, to elucidate phylogenetic relationships via Bayesian Inference (BI), based on concatenated mitochondrial fragments. RESULTS: The resulting genome, which spans 16,391 base pairs was investigated, revealing 13 protein-coding genes, rRNAs (12S and 16S), 19 tRNAs, and a putative control region. Comparative analysis together with five other Cambaroides mitogenomes retrieved from GenBank unveiled regions that remained unread due to challenges associated with the genome skimming technique. Protein-coding genes varied in size and typically exhibited common start (ATG) and stop (TAA) codons. However, exceptions were noted in ND5 (start codon: GTG) and ND1 (stop codon: TAG). Landscape analysis was used to explore sequence variation across the five available mitochondrial genomes of Cambaroides. CONCLUSIONS: Collectively, these findings reveal variable sites and contribute to a deeper understanding of the genetic diversity in this genus alongside the further development of species-specific primers for noninvasive monitoring techniques. The partitioned phylogenetic analysis of Astacidea revealed a paraphyletic origin of Asian cambarids, which confirms the data in recent studies based on both multilocus analyses and integrative approaches.

Identification of potential functions of polo-like kinase 1 in male reproductive development of the oriental river prawn (Macrobrachium nipponense) by RNA interference analysis.

Polo-like kinase 1 (Plk1) has multiple functions in the cell cycle, including in the maturation of centrosomes during the G2/M transition, the separation of centrosomes, and the activation of cyclin-dependent kinase 1 expression and spindle assembly. In this study, we investigated the potential regulatory roles of Plk1 in the reproductive development of the male oriental river prawn (Machrobrachium nipponense). The full cDNA sequence of Mn-Plk1 was 2360 base pairs long, with an open reading frame of 1836 base pairs encoding 611 amino acids. Protein sequence alignment identified a conserved serine/threonine kinase domain and two Polo-boxes. Phylogenetic tree analysis revealed that Mn-Plk1 had the closest evolutionary distance with Plk1s of freshwater prawns and then with those of crustacean species, whereas the evolutionary distance with mollusks was much more distant. Quantitative PCR analysis predicted that Mn-Plk1 plays essential roles in the regulation of gonad development. RNA interference analysis and histological observations showed that expression of insulin-like androgenic gland hormone decreased as the expression of Mn-Plk1 decreased, and fewer than 5% of cells were sperm cells at day 14 in the dsPlk1 injected prawns. This result indicated that Plk1 positively regulated testis development in M. nipponense by affecting the expression of this hormone. Our results highlight the functions of Plk1 in M. nipponense and provide valuable information that can be applied to establish artificial techniques to regulate testis development in this species.