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Edwardsiella ictaluri is a highly destructive pathogen in cultured yellow catfish, thus it was very necessary to study the immune response of yellow catfish against bacterial infection. In this study, RNA-Seq technology was used to study the immune response in two distinct tissues of yellow catfish at eight different time points (h) after E. ictaluri infection. The number of differentially expressed genes (DEGs) in the spleen and liver was low at 3 h and 6 h post-infection, respectively. Afterwards, the most number of DEGs in the spleen was detected at 72 h, while the number of DEGs in the liver maintained a high level from 24 h to 120 h. The GO and KEGG enrichment analyses of DEGs at different time points uncovered that cytokines were continuously transcribed at 6 h to 120 h; whereas the liver is the main organ that secretes the components of the complement system, and metabolic regulation was activated from 12 h to 120 h. Moreover, an overview of the inflammation response of yellow catfish was exhibited including pattern-recognition receptors, inflammatory cytokines, chemokines, complements, and inflammation-related signal pathways. The similar expression tendency of nine genes by qRT-PCR validated the accuracy of transcriptome analyses. The different transcriptomic profiles obtained from the spleen and liver will help to better understand the dynamic immune response of fish against bacterial infection, and will provide basic information for establishing effective measures to prevent and control diseases in fish.

In mammals, Interleukin-17 cytokine family plays critical roles in both acute and chronic inflammatory responses. In fish species, three Interleukin-17A/F (IL-17A/F) genes have been identified to be homologous to mammalian IL-17A and IL-17F, but little is known about their functional activity. In this study, Pf _IL-17A/F1, 2 and 3 genes were cloned from yellow catfish ( Pelteobagrus fulvidraco ) and they differed in protein structure and exon length, implying that they may have divergent bioactivity. Real-time quantitative PCR analyses revealed that three Pf _IL-17A/F genes were highly expressed in blood and mucosal tissues (skin+mucus and gill) from healthy adult fish. The mRNA expressions of Pf _IL-17A/F1, 2 and 3 genes were significantly up-regulated in the gill, skin+mucus, head kidney and spleen after challenge with Edwardsiella ictaluri and in the isolated peripheral blood leucocytes (PBLs) of yellow catfish after stimulation with phytohaemagglutinin (PHA), lipopolysaccharides (LPS), peptidoglycan (PGN) and polyinosinic-polycytidylic acid (Poly I:C). These results indicate that Pf _IL-17A/F1, 2 and 3 genes may play a vital role in the regulation of immune against pathogens. Additionally, the recombinant (r) Pf _IL-17A/F1, 2 and 3 proteins significantly induced the mRNA expressions of proinflammatory cytokines, chemokines and antibacterial peptides genes, and the r Pf _IL-17A/F 2 and 3 proteins promoted phagocytosis of PBLs more powerfully than the r Pf _IL-17A/F1. Furthermore, the r Pf _IL-17A/F1, 2 and 3 proteins might activate the NF-κB and MAPK signal pathways by IL-17RA, ACT1, TRAF6, TRAF2, TRAF5 and TAK1, indicating that the three Pf _IL-17A/F proteins may play different roles in promoting inflammatory response.

The yellow catfish (Pelteobagrus fulvidraco) is an economic fish with a large breeding scale, and diseases have led to huge economic losses. Tumor necrosis factor receptor-associated factors (TRAFs) are a class of intracellular signal transduction proteins that play an important role in innate and adaptive immune responses by mediating NF-κB, JNK and MAPK signaling pathways. However, there are few studies on the TRAF gene family in yellow catfish. In this study, the open reading frame (ORF) sequences of TRAF1, TRAF2a, TRAF2b, TRAF3, TRAF4a, TRAF4b, TRAF5, TRAF6 and TRAF7 genes were cloned and identified in yellow catfish. The ORF sequences of the nine TRAF genes of yellow catfish (Pf_TRAF1-7) were 1413–2025 bp in length and encoded 470–674 amino acids. The predicted protein structures of Pf_TRAFs have typically conserved domains compared to mammals. The phylogenetic relationships showed that TRAF genes are conserved during evolution. Gene structure, motifs and syntenic analyses of TRAF genes showed that the exon–intron structure and conserved motifs of TRAF genes are diverse among seven vertebrate species, and the TRAF gene family is relatively conserved evolutionarily. Among them, TRAF1 is more closely related to TRAF2a and TRAF2b, and they may have evolved from a common ancestor. TRAF7 is quite different and distantly related to other TRAFs. Real-time quantitative PCR (qRT-PCR) results showed that all nine Pf_TRAF genes were constitutively expressed in 12 tissues of healthy yellow catfish, with higher mRNA expression levels in the gonad, spleen, brain and gill. After infection with Edwardsiella ictaluri, the expression levels of nine Pf_TRAF mRNAs were significantly changed in the head kidney, spleen, gill and brain tissues of yellow catfish, of which four genes were down-regulated and one gene was up-regulated in the head kidney; four genes were up-regulated and four genes were down-regulated in the spleen; two genes were down-regulated, one gene was up-regulated, and one gene was up-regulated and then down-regulated in the gill; one gene was up-regulated, one gene was down-regulated, and four genes were down-regulated and then up-regulated in the brain. These results indicate that Pf_TRAF genes might be involved in the immune response against bacterial infection. Subcellular localization results showed that all nine Pf_TRAFs were found localized in the cytoplasm, and Pf_TRAF2a, Pf_TRAF3 and Pf_TRAF4a could also be localized in the nucleus, uncovering that the subcellular localization of TRAF protein may be closely related to its structure and function in cellular mechanism. The results of this study suggest that the Pf_TRAF gene family plays important roles in the immune response against pathogen invasion and will provide basic information to further understand the roles of TRAF gene against bacterial infection in yellow catfish.

The phylogeography of Schizothorax waltoni , an endemic and endangered tetraploid schizothoracine fish in the Yarlung Tsangpo River (YLTR) on southern margin of Qinghai-Tibet Plateau (QTP), was investigated using two mitochondrial DNA regions and eleven microsatellite loci. Analyses of concatenated sequences of cytochrome b (1141 bp) and the control region (712 bp) revealed high haplotype diversity and moderate nucleotide diversity. High genetic diversity was observed based on microsatellite variation. Both mtDNA and microsatellite analyses revealed significant genetic differentiation between the eastern population (Mainling) and the other four populations to the west, and non-significant genetic differentiation amongst the three central populations in the west. Significant genetic differentiation was observed between the western population (Shigatse) and the three central populations based on microsatellite analyses alone. Bayesian skyline plot analyses showed that S. waltoni experienced a pronounced population expansion 0.05 to 0.10 Ma. Hierarchical structure analyses of microsatellite data indicated that S. waltoni could be split into three groups (western, central and eastern YLTR). The results indicate that three management units should be considered for S. waltoni . Our findings highlight the need for the conservation and effective management of S. waltoni , which is a key member of the endemic and highly threatened fishes of the QTP.

The blunt snout bream (Megalobrama amblycephala) is an important freshwater aquaculture fish throughout China. Because of widespread introductions of this species to many regions, the genetic diversity of wild and natural populations is now threatened. In the present study, SRAP (sequence-related amplified polymorphism) markers were used to assess genetic diversity of blunt snout bream. Three natural populations (Liangzi Lake, Poyang Lake and Yuni Lake, one cultured population (Nanxian) and one genetic strain ('Pujiang No. 1') of blunt snout bream were screened with 88 SRAP primer combinations, of which 13 primer pairs produced stable and reproducible amplification patterns. In total, 172 bands were produced, of which 132 bands were polymorphic. Nei's gene diversity (h) and Shannon's information index (I) values provided evidence of differences in genetic diversity among the five populations (Poyang Lake>Liangzi Lake>Nanxian>'Pujiang No. 1'>Yuni Lake). Based on cluster analysis conducted on genetic distance values, the five blunt snout bream populations were divided into three groups, Poyang Lake and Liangzi Lake (natural populations), Nanxian and 'Pujiang No. 1' (cultured population and genetically selected strain), and Yuni Lake (natural population). Significant genetic differentiation was found among the five populations using analysis of molecular variance (AMOVA), with more genetic divergence existing among populations (55.49%), than within populations (44.51%). This molecular marker technique is a simple and efficient method to quantify genetic diversity within and among fish populations, and is employed here to help manage and conserve germplasm variability of blunt snout bream and to support the ongoing selective breeding programme for this fish.

Major threats to freshwater fish diversity now include loss of native genetic diversity as a consequence of translocations of fishes between sites and from hatcheries to sites, and small effective population sizes resulting from overfishing and/or habitat loss. Ten polymorphic microsatellite markers were employed to evaluate genetic diversity, population genetic structure and gene flow amongst nine populations of the ecologically and economically important fish, the northern snakehead (Channa argus), in three river systems in central China. Multiple analyses revealed evidence of high genetic diversity and pronounced subdivision based on both regional separation and on river systems. A lack of evidence of genetic bottleneck over recent generations was consistent with the long-term stability of population size and contemporary distribution. The effective population sizes for most C. argus populations were small, suggesting the need for future conservation efforts focusing on these populations. Different lines of evidence point to the local enhancement of stocks by both aquaculture-reared fish and the transfer of wild fish. This study illustrates how human activities may affect genetic diversity and population genetic structure of C. argus populations, and highlights the need for new management regimes to protect native freshwater fish genetic diversity.

Channa argus is an important commercial fish species in the aquaculture industry in China, however, it is declining in numbers and population size due to overfishing and environmental change. Based on the transcriptome database of C. argus from the Illumina Hiseq 2000 platform, 32 polymorphic single nucleotide polymorphism (SNP) markers were developed and characterized in 36 individuals using high resolution melting analysis. The observed and expected heterozygosities ranged from 0 to 0.694 and from 0.106 to 0.507, respectively, and the minor allele frequency ranged from 0.056 to 0.486. Only two loci showed significant deviation from Hardy–Weinberg equilibrium after Bonferroni correction. These SNP markers will serve as a valuable tool for population genetic analysis, natural resource conservation and selective breeding of this species.

The Qinghai-Tibet Plateau (QTP) is a biodiversity hotspot, resulting from its geological history, contemporary environment and isolation. Uplift of the QTP and Quaternary climatic oscillations are hypothesised to have influenced the genetic diversity, population structure and dynamics of all QTP endemic species. In this study, we tested this hypothesis by assaying variation at two mitochondrial DNA regions (cytochrome b and control region) and at 12 microsatellite loci of seven populations of the endemic fish, Schizothorax o’connori from the Yarlung Tsangpo River (YLTR) on the QTP. Analyses revealed one group of six populations to the west, above the Yarlung Tsangpo Grand Canyon (YTGC), and a second group to the east below the YTGC. Estimates of the timing of this east-west split indicate that these groups represent evolutionarily significant units that have evolved separately and rapidly in the middle Pleistocene, at the time of the Kunlun-Huanghe Movement A Phase and the Naynayxungla glaciation. Population dynamic analyses indicate that S. o’connori experienced a pronounced late Pleistocene expansion during the last interglacial period. The results of this study support the hypotheses that the QTP uplift and Quaternary climatic oscillations have played important roles in shaping the population genetics and dynamics of this endemic fish.

Odontobutis sinensis is an indigenous freshwater fish in China. The wild population size of this species has declined sharply in inland waters in recent years. For the purpose of the conservation of natural resources of O. sinensis, 14 polymorphic microsatellite loci were isolated and characterized in 30 individuals collected from Liangzi Lake in Hubei Province. The number of alleles per locus ranged from 2 to 14 with an average of 5.07. The expected and observed heterozygosities ranged from 0.291 to 0.887 and from 0.067 to 0.700, respectively. These newly isolated loci are currently being used for population genetic diversity analysis and will be valuable for the conservation of O. sinensis.

Cholecystokinin (CCK) is a multi-functional brain–gut peptide in fish and mammals. To investigate the role of CCK in appetite regulation in fish, a 770-bp full-length cDNA sequence of CCK gene was obtained by RT-PCR and rapid amplification of cDNA ends methods in grass carp Ctenopharyngodon idellus . Homology analysis showed that the CCK cDNA sequence of grass carp had the highest similarity (90 %) to that of goldfish Carassius auratus and a higher similarity (>70 %) to those of other teleosts than to mammals. The PCR amplification using genomic DNA identified that the CCK gene of grass carp was comprised of three exons and two introns. Real-time quantitative PCR was used to detect CCK mRNA expression in adult tissues. High levels of gene expression were found in the hypothalamus and pituitary; moderate levels in the intestine, muscle and white adipose tissue; and low levels in other tissues. During early development (i.e., fertilized eggs to 35-day post-hatching larvae) the levels of CCK mRNA expression were higher during embryonic developmental stages than during post-hatch larval stages. Fasting decreased CCK mRNA expression levels in the brain and intestine, whereas refeeding resulted in an increase of expression. The results suggest that CCK mRNA expression has obvious tissue specificity and may have a role in feed intake regulation in grass carp.