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Native to North America, largemouth bass Micropterus salmoides (Lacepède, 1802) were introduced in South Africa in 1928. Florida bass Micropterus floridanus (Lesueur, 1822) were introduced to enhance existing largemouth bass fisheries in 1980. While largemouth bass and Florida bass readily hybridise and produce offspring that are difficult to identify morphologically, confirmation of hybridisation requires genetic analysis. This study sought to understand: (1) the extent to which Florida bass spread within a catchment once introduced; and (2) whether a subset of samples taken from within a catchment accurately characterises hybridisation throughout the catchment. Samples were collected from the Breede River and Kowie River catchments and screened using 38 species-diagnostic single nucleotide polymorphisms to assess hybridisation. Collections from the mainstem of both rivers represented hybrid swarms, and neither pure largemouth bass nor Florida bass were observed. The absence of genetic differentiation among sampling sites suggests that hybridisation will occur throughout systems where both species are present. Hybridisation levels in dams located off the mainstem rivers were significantly variable and represented potential sources for Florida bass alleles observed within rivers. This finding, in conjunction with our limited knowledge of the distributions of the two species, suggests that applying independent management strategies to control and monitor the spread of both species may prove difficult. [ABSTRACT FROM AUTHOR]

Fish‐derived antimicrobial peptides are an important part of the innate immune system due to their potent antimicrobial properties. Piscidins are a class of antimicrobial peptides first described in hybrid striped bass (Morone chrysops x Morone saxatilis) but have also been identified in many other fish species. Previous work demonstrated the broad antimicrobial activity of piscidins against Gram‐negative and Gram‐positive bacterial species. This study sought to determine the extent to which class I (striped bass piscidin 1, white bass piscidin 1 and striped bass/white bass piscidin 3) and class II (striped bass piscidin 4 and white bass piscidin 5) piscidins inhibit biofilm formation of different Gram‐negative bacteria. In general, the class I and II piscidins demonstrate potent activity against Escherichia coli and Flavobacterium columnare biofilms. The class II piscidins showed more activity against E. coli and F. columnare isolates than did the class I piscidins. The piscidins in general were much less effective against inhibiting Aeromonas hydrophila and A. veronii biofilm growth. Only the class I piscidins showed significant growth inhibition among the Aeromonas spp. examined. [ABSTRACT FROM AUTHOR]

Hybrid zones between diverged lineages offer a unique opportunity to study evolutionary processes related to speciation. Natural and anthropogenic hybridization in the black basses (Micropterus spp.) is well documented, including an extensive intergrade zone between the widespread northern Largemouth Bass (M. salmoides) and the Florida Bass (M. floridanus). Phenotypic surveys have identified an estuarine population of Largemouth Bass (M. salmoides) in the Mobile‐Tensaw Delta, with larger relative weight and smaller adult size compared to inland populations, suggesting a potential third lineage of largemouth bass. To determine the evolutionary relationships among these Mobile Delta bass populations, M. salmoides and M. floridanus, putative pure and intergrade populations of all three groups were sampled across the eastern United States. Phylogenetic analyses of 8582 nuclear SNPs derived from genotype‐by‐sequencing and the ND2 mitochondrial gene determined that Delta bass populations stem from a recently diverged lineage of Largemouth Bass. Using a novel quantitative pipeline, a panel of 73 diagnostic SNPs was developed for the three lineages, evaluated for accuracy, and then used to screen 881 samples from 52 sites for genetic integrity and hybridization on the Agena MassARRAY platform. These results strongly support a redrawing of native ranges for both the intergrade zone and M. floridanus, which has significant implications for current fisheries management. Furthermore, Delta bass ancestry was shown to contribute significantly to the previously described intergrade zone between northern Largemouth Bass and Florida Bass, suggesting a more complex pattern of secondary contact and introgression among these diverged Micropterus lineages. [ABSTRACT FROM AUTHOR]

We report genetic and morphological evidence for the presence of Redeye Bass Micropterus coosae, in the Verde River of Arizona, previously thought to be Smallmouth Bass Micropterus dolomieu. We performed meristic measurements on 15 individuals sampled from the Upper Verde River Wildlife Area, Yavapai County, Arizona. Meristic data for lateral line scales, scales above lateral line, and scales below lateral line were all consistent with Redeye Bass and not Smallmouth Bass. We analyzed mitochondrial and nuclear genetic data to determine whether one of the black bass (Genus Micropterus) species historically introduced to the Verde River was Redeye Bass and whether they persist in the system. We extracted DNA from fin clips of five individuals for phylogenetic analysis of the nicotinamide adenine dinucleotide þ hydrogen (NADH) dehydrogenase subunit 2 (ND2) mitochondrial gene and for analysis of nuclear DNA using a diagnostic Single Nucleotide Polymorphism (SNP) panel. Results of the ND2 genetic sequencing and phylogenetic analysis indicated that these fish likely originated from native Redeye Bass stock from the Coosa River system of Alabama, Georgia, and Tennessee. Similarly, nuclear SNP data from the five individuals collected from the Verde River aligned with Redeye Bass reference genotypes based on STRUCTURE analysis. These results support the hypothesis that at least one of the introductions of black bass in Arizona's Verde River founded a previously unrecognized population of Redeye Bass. Further work is needed to determine the extent of the Redeye Bass presence in Arizona, whether Smallmouth Bass are also present in the Verde River system, and if hybridization of Redeye Bass and other black basses is occurring. [ABSTRACT FROM AUTHOR]

The Gram-negative bacterium, Aeromonas hydrophila , has been responsible for extensive losses in the catfish industry for over a decade. Due to this impact, there are ongoing efforts to understand the basic mechanisms that contribute to virulent A. hydrophila (vAh) outbreaks. Recent challenge models demonstrated that vAh cultured in the presence of the iron chelating agent deferoxamine mesylate (DFO) were more virulent to channel catfish (Ictalurus punctatus). Interestingly, differential gene expression of select iron acquisition genes was unremarkable between DFO and non-DFO cultures, posing the question: why the increased virulence? The current work sought to evaluate growth characteristics and protein expression of vAh after the addition of DFO. A comparative proteome analysis revealed differentially expressed proteins among tryptic soy broth (TSB) and TSB + DFO treatments. Upregulated proteins identified among the TSB + DFO treatment were enriched for gene ontology groups including iron ion transport, siderophore transport and siderophore uptake transport, all iron acquisition pathways. Protein-protein interactions were also evaluated among the differentially expressed proteins and predicted that many of the upregulated iron acquisition proteins likely form functional physiological networks. The proteome analysis of the vAh reveals valuable information about the basic biological processes likely leading to increased virulence during iron restriction in this organism. [ABSTRACT FROM AUTHOR]

The article discusses Walleye (Sander vitreus) is a popular sportfish threatened by overexploitation, habitat destruction, and loss of genetic integrity due to non-native walleye stocking. Topics include Walleye is an ecologically important and economically valuable freshwater fish species in family Percidae; and single nucleotide polymorphisms (SNPs) are rapidly becoming the preferred markers for routine genetic identification and hybridization tests in a variety of aquatic species.

Black basses (genus Micropterus) are apex predators in North American streams, rivers, and lakes and important game fishes. Translocation and introductions for angling, accompanied by intrinsically weak genetic barriers, have led to widespread introgressive hybridization and genetic swamping. Species-diagnostic (fixed allele) SNP markers have been utilized successfully in salmonids to monitor hybridization and genetic integrity. Here, we developed similar resources for black basses through initial genotyping-by-sequencing, followed by validation in additional samples using two panels of 64 SNPs. Results from > 1300 genotyped bass indicated that the developed panels robustly and clearly delineate fifteen species and their hybrids among black basses. The panels represent a flexible, rapid turnaround (~ 1 day), and cost-effective tool that should augment ongoing efforts toward black bass conservation and management. [ABSTRACT FROM AUTHOR]

The software programs STRUCTURE and NEWHYBRIDS are widely used population genetic programs useful in addressing questions related to genetic structure, admixture, and hybridization. These programs usually require a large number of independent runs with many iterations to provide robust data for downstream analyses, thus significantly increasing computation time. Programs such as Structure_threader and parallelnewhybrid were previously developed to address this problem by processing tasks in parallel on a multi-threaded processor; however some programming knowledge (e.g., R, Bash) is required to run these programs. We developed EasyParallel as a community resource to facilitate practical and routine population structure and hybridization analyses. The multi-threaded parallelization of EasyParallel allows processing of large genetic datasets in a very efficient way, with its point-and-click GUI providing ready access to users who have little experience in script programming. Performance evaluation of EasyParallel using simulated datasets showed similar speed-up and parallel execution time when compared to Structure_threader and Parallelnewhybrid. EasyParallel is written in Python 3 and freely available on the GitHub site https://github.com/hzz0024/EasyParallel. [ABSTRACT FROM AUTHOR]

BACKGROUND Genomes are vital to the study of genomics, population genetics, and evolution of species. To date, only one genome (Echinochloa crus‐galli) for C4 annual weedy grass species has been sequenced. Research was conducted to develop a draft genome of goosegrass (Eleusine indica; 2n = 2x = 18), one of the most common and troublesome weeds in the world. RESULTS: A draft assembly of an approximately 492 Mb whole‐genome sequence of goosegrass was obtained by de novo assembly of paired‐end and mate‐paired reads generated by Illumina sequencing of total genomic DNA. The genome was assembled into 24,072 scaffolds with N50 = 233,459 bp. More than 99% of transcriptome sequences were mapped to the goosegrass draft genome, and 95% of the commonly conserved plant genes were present. The assembled genome contains 25,467 unique protein‐coding genes. Genes associated with herbicide resistance were obtained and variant calling allowed the detection of 754,409 single nucleotide polymorphisms. In addition, we also report 115,417 simple sequence repeats which can be deployed in population genetics and phylogenetic analysis. CONCLUSION: This is the first report of genome sequence of goosegrass. Our assembly was able to identify all major herbicide‐resistance related genes and develop a useful tool for other genomic and evolutionary analysis. © 2019 Society of Chemical Industry [ABSTRACT FROM AUTHOR]

We evaluated a Saccharomyces cerevisiae fermentation product (Diamond V Original XPC) in hybrid catfish (Ictalurus furcatus x I. punctatus) for its potential effects on growth, blood parameters, and disease resistance. The trial featured four levels of inclusion that were added to a commercial 32% protein floating catfish ration. Following six weeks of feeding, we observed marginally heightened resistance to columnaris disease and saw significant changes in the levels of immune effectors in the serum, including lysozyme, complement, and immunoglobulin. [ABSTRACT FROM AUTHOR]

The widespread introduction of Largemouth Bass (Micropterus salmoides) and Florida Bass (M. floridanus) to establish sport fisheries represents a significant conservation concern given their role as apex predators and their ability to alter community diversity and species abundance. In regions like southern Africa, which has both high levels of aquatic endemism and imperilment, limiting the spread of invasive predators is a primary goal of current alien species legislation. Here, we applied two panels of species-diagnostic SNPs for a total of 60 markers to map the distribution of Largemouth Bass, Florida Bass, and their hybrids in 13 southern African water bodies. Using Bayesian clustering algorithms we documented the introgression of Florida Bass alleles across a broad geographic range, from the Cape Floristic region of South Africa to Mozambique. Several populations previously considered pure Largemouth Bass based on mitochondrial DNA sequences were found to consist exclusively of hybrids. Samples collected from Lake Chicamba, which was initially established with pure Largemouth Bass, are now almost exclusively comprised of Florida Bass alleles (89.3 % Florida Bass). The estimated hybrid class of sampled fish (e.g., F1 hybrid, pure Largemouth Bass, pure Florida Bass) showed that with few exceptions, populations were dominated by a single hybrid class. The present work provides enhanced resolution of the distribution and dynamics of Florida Bass, Largemouth Bass, and their hybrids in southern Africa. [ABSTRACT FROM AUTHOR]

The antimicrobial activity and mode of action of chitosan were evaluated against Streptococcus iniae, a pathogenic Gram‐positive bacterium of fish worldwide. Cell proliferation kinetics were examined following exposure to varying concentrations of chitosan. The action of chitosan on S. iniae was also investigated by measuring agglutination activity, conductivity, and extracellular and intracellular bacterial adenosine triphosphate (ATP) levels. Chitosan exhibited antibacterial activity against S. iniae at concentrations of 0.1% and above and was lethal at a concentration of 0.4% and higher. The mechanism of antibacterial activity of chitosan at the inhibitory level of bacterial growth appears to hinge upon the interaction between chitosan and the oppositely charged bacterial surface. This interplay causes agglutination, which was readily observed grossly and microscopically. After interacting with the cell surface via adsorption, an efflux of intracellular ATP was documented, which suggests that chitosan disrupts the bacterial cell causing leakage of cytosolic contents and ultimately cell death. Results suggest chitosan may be worth evaluating as a natural alternative to antibiotic against S. iniae infection of fish. [ABSTRACT FROM AUTHOR]

Following the Deepwater Horizon oil spill, eastern oyster (Crassostrea virginica) reefs in the northern Gulf of Mexico were exposed to oil and various associated clean-up activities that may have compromised oyster reef health. Included in the exposure was oil, dispersant, and in some locales, atypical salinity regimes. Oil and dispersants can be detrimental to oysters and the effects of salinity depend on the level. In addition to these extrinsic factors, genetic diversity of oyster populations may help the oysters respond to stressors, as demonstrated in other systems. We used a 3×3×2 factorial design to experimentally examine the effects of oil/dispersed oil, intraspecific genetic diversity, and salinity on juvenile (ca. 25 mm shell height) oyster survivorship and growth during a 21-d exposure in a closed, recirculating system. The genetic effect was weak overall, oil and dispersed oil negatively affected juvenile oyster survivorship, and low salinity mitigated mortality in oil and dispersed oil treatments. Survivorship was about 40% greater in low-salinity than in mesohaline water for both oil and dispersed oil treatments, bringing survivorship in low salinity oil-only treatments to a similar level with low salinity controls (no oil). Oyster growth was minimal after 21 d but appeared to be negatively affected by oil and dispersed oil, and had a significant interaction with salinity. Our results may be informative for future decisions regarding oil spill response activities and suggest that a pulse of low salinity water may be a viable short-term mitigation option for oysters if filtration characteristics, exposure time, and water temperatures are all considered, in addition to weighing the costs and benefits of this type of response on other organisms and habitats. [ABSTRACT FROM AUTHOR]

Abstract: Unusual persistent natural mortality occurred in a floating in‐pond raceway system intensively stocked with channel and hybrid catfish beginning in early November 2016 up until March 2017. The temperature during the period of outbreak ranged from 7.2 to 23.7°C. Gross examination of freshly dead and moribund fish revealed pale gills, slight abdominal distension and swollen inflamed vents. Comprehensive necropsy of 20 fish demonstrated vast amounts of bloody ascitic fluid in the coelomic cavity, visceral congestion, splenomegaly and pale friable livers but macroscopically normal kidneys, suggesting systemic bacterial infection. Bacterial cultures were initiated from skin, gills and major internal organs. Following incubation, a mixture of three bacterial colony phenotypes was observed on agar plates. Presumptive biochemical characterization of the isolates followed by 16S‐rRNA sequence analysis resulted in the identification of Aeromonas veronii, Streptococcus parauberis and Shewanella putrefaciens. Channel catfish juveniles were experimentally infected with the recovered isolates to fulfil Koch's postulates. Moreover, an antibiogram was used to evaluate the susceptibility of the isolates to antimicrobial drugs approved for use in aquaculture. Aquaflor was used successfully for treatment. Here, we report bacterial coinfection lead by A. veronii and the first identification of S. parauberis and S. putrefaciens from cultured catfish in North America. [ABSTRACT FROM AUTHOR]

Abstract: Flavobacterium columnare causes columnaris disease of farmed and wild freshwater fish. Skin mucus is an important factor in early stages of columnaris pathogenesis, albeit little studied. Our objectives were to (a) characterize the terminal glycosylation pattern (TGP) of catfish mucus, (b) determine the growth of F. columnare in formulated water (FW)‐containing channel catfish (Ictalurus punctatus) or hybrid catfish (Ictalurus punctatus X Ictalurus furcatus) mucus and (c) examine extracellular protease activity of two F. columnare isolates differing in virulence. The TGP of catfish mucus by lectin binding was as follows: alpha‐D‐mannose/alpha‐D‐glucose >N‐acetyl‐beta‐D‐glucosamine >N‐acetyl‐beta‐D‐glucosamine/N‐acetylneuraminic acid >N‐acetyl‐D‐galactosamine >alpha‐D‐galactose/N‐acetyl‐alpha‐D‐galactosamine >beta‐D‐galactose = alpha‐L‐fucose. Virulence studies demonstrated isolate AL‐02‐36 was highly virulent in channel catfish fry (0.1 g) with cumulative mortality of 90%‐100% versus 60% for isolate ALG‐00‐530 at equivalent doses (~3 × 106 CFU/ml); a similar result was observed in larger (0.7 g) catfish. In multiple experiments, F. columnare replicated (2‐3 logs) and survived (28 days) in formulated water‐containing catfish mucus. Highly virulent isolate AL‐02‐36 possessed at least 2.5‐ to fivefold higher protease activity following growth in mucus than the less virulent ALG‐00‐530. Flavobacterium columnare utilized catfish mucus as a nutrient source and mucus presence modulated extracellular protease production. [ABSTRACT FROM AUTHOR]

Culture of the eastern oyster (Crassostrea virginica) is rapidly expanding. Combined with their continuing role as an environmental sentinel species and ecological model, this trend necessitates improved molecular tools for breeding and selection, as well as population assessment and genetic conservation. Here, we describe the development and validation of two panels of 58 single nucleotide polymorphism markers (SNPs) for the species. Population analyses revealed three distinct populations, based on FST values and STRUCTURE, among wild oysters sampled from Delaware Bay (1), northwest Florida (2), Alabama (2), Louisiana (2), and the Texas Gulf Coast (3), consistent with previous microsatellite and mtDNA analyses. In addition, utilizing the developed panels for parentage assignment in cultured oysters (Rutgers, New Jersey) resulted in a highly accurate identification of parent pairs (99.37%). The SNP markers could, furthermore, clearly discriminate between hatchery stocks and wild-sourced individuals. The developed SNP panels may serve as an important tool for more rapid and affordable genetic analyses in eastern oyster. [ABSTRACT FROM AUTHOR]

Our aim was to transplant blue catfish germ line stem cells into blastulae of triploid channel catfish embryos to produce interspecific xenogenic catfish. The morphological structure of the gonads of blue catfish (Ictalurus furcatus) in ~ 90- to 100-day-old juveniles, two-year-old juveniles, and mature adults was studied histologically. Both oogonia (12-15 μm, diameter with distinct nucleus 7-8 μm diameter) and spermatogonia (12-15 μm, with distinct nucleus 6-7.5 μm diameter) were found in all ages of fish. The percentage of germ line stem cells was higher in younger blue catfish of both sexes. After the testicular tissue was trypsinized, a discontinuous density gradient centrifugation was performed using 70, 45, and 35% Percoll to enrich the percentage of spermatogonial stem cells (SSCs). Four distinct cell bands were generated after the centrifugation. It was estimated that 50% of the total cells in the top band were type A spermatogonia (diameter 12-15 μm) and type B spermatogonia (diameter 10-11 μm). Germ cells were confirmed with expression of vasa. Blastula-stage embryos of channel catfish (I. punctatus) were injected with freshly dissociated blue catfish testicular germ cells as donor cells for transplantation. Seventeen days after the transplantation, 33.3% of the triploid channel catfish fry were determined to be xenogenic catfish. This transplantation technique was efficient, and these xenogenic channel catfish need to be grown to maturity to verify their reproductive capacity and to verify that for the first time SSCs injected into blastulae were able to migrate to the genital ridge and colonize. These results open the possibility of artificially producing xenogenic channel catfish males that can produce blue catfish sperm and mate with normal channel catfish females naturally. The progeny would be all C × B hybrid catfish, and the efficiency of hybrid catfish production could be improved tremendously in the catfish industry. [ABSTRACT FROM AUTHOR]

Hypoxic condition is common in aquaculture, leading to major economic losses. Genetic analysis of hypoxia tolerance, therefore, is not only scientifically significant, but also economically important. Catfish is generally regarded as being highly tolerant to low dissolved oxygen, but variations exist among various populations, strains, and species. In this study, we conducted a genome-wide association study (GWAS) using the catfish 250 K SNP array to identify quantitative trait locus (QTL) associated with tolerance to low dissolved oxygen in the channel catfish × blue catfish interspecific system. Four linkage groups (LG2, LG4, LG23, and LG29) were found to be associated with low oxygen tolerance in hybrid catfish. Multiple significant SNPs were found to be physically linked in genomic regions containing significant QTL for low oxygen tolerance on LG2 and LG23, and in those regions containing suggestively significant QTL on LG2, LG4, LG23, and LG29, suggesting that the physically linked SNPs were genuinely segregating and related with low oxygen tolerance. Analysis of genes within the associated genomic regions suggested that many of these genes were involved in VEGF, MAPK, mTOR, PI3K-Akt, P53-mediated apoptosis, and DNA damage checkpoint pathways. Comparative analysis indicated that most of the QTL at the species level, as analyzed by using the interspecific system, did not overlap with those identified from six strains of channel catfish, confirming the complexity of the genetic architecture of hypoxia tolerance in catfish. [ABSTRACT FROM AUTHOR]

Interspecific hybrids provide a rich source for the analysis of allele-specific expression (ASE). In this work, we analyzed ASE in F1 hybrid catfish using RNA-Seq datasets. While the vast majority of genes were expressed with both alleles, 7-8 % SNPs exhibited significant differences in allele ratios of expression. Of the 66,251 and 177,841 SNPs identified from the datasets of the liver and gill, 5420 (8.2 %) and 13,390 (7.5 %) SNPs were identified as significant ASE-SNPs, respectively. With these SNPs, a total of 1519 and 3075 ASE-genes were identified. Gene Ontology analysis revealed that genes encoding cytoplasmic ribosomal proteins (RP) were highly enriched among ASE genes. Parent-of-origin was determined for 27 and 30 ASE RP genes in the liver and gill, respectively. The results indicated that genes from both channel catfish and blue catfish were involved in ASE. However, each RP gene appeared to be almost exclusively expressed from only one parent, indicating that ribosomes in the hybrid catfish were in the 'hybrid' form. Overall representation of RP transcripts among the transcriptome appeared lower in the F1 hybrid catfish than in channel catfish or blue catfish, suggesting that the 'hybrid' ribosomes may work more efficiently for translation in the F1 hybrid catfish. [ABSTRACT FROM AUTHOR]

Channel catfish ( Ictalurus punctatus) embryos were electroporated with sterilization constructs targeting primordial germ cell proteins or with buffer. Some embryos then were treated with repressor compounds, cadmium chloride, copper sulfate, sodium chloride or doxycycline, to prevent expression of the transgene constructs. Promoters included channel catfish nanos and vasa, salmon transferrin (TF), modified yeast Saccharomyces cerevisiae copper transport protein (MCTR) and zebrafish racemase (RM). Knock-down systems were the Tet-off (nanos and vasa constructs), MCTR, RM and TF systems. Knock-down genes included shRNAi targeting 5′ nanos (N1), 3′ nanos (N2) or dead end (DND), or double-stranded nanos RNA (dsRNA) for overexpression of nanos mRNA. These constructs previously were demonstrated to knock down nanos, vasa and dead end, with the repressors having variable success. Exogenous DNA affected percentage hatch (% hatch), as all 14 constructs, except for the TF dsRNA, TF N1 (T), RM DND (C), vasa DND (C), vasa N1 (C) and vasa N2 (C), had lower % hatch than the control electroporated with buffer. The MCTR and RM DND (T) constructs resulted in delayed hatch, and the vasa and nanos constructs had minimal effects on time of hatch ( P < 0.05). Cadmium chloride appeared to counteract the slow development caused by the TF constructs in two TF treatments ( P < 0.05). The 4 ppt sodium chloride treatment for the RM system decreased % hatch ( P < 0.05) and slowed development. In the case of nanos constructs, doxycycline greatly delayed hatch ( P < 0.05). Adverse effects of the transgenes and repressors continued for several treatments for the first 6 days after hatch, but only in a few treatments during the next 10 days. Repressors and gene expression impacted the yield of putative transgenic channel catfish fry, and need to be considered and accounted for in the hatchery phase of producing transgenically sterilized catfish fry and their fertile counterparts. This fry output should be considered to ensure that sufficient numbers of transgenic fish are produced for future applications and for defining repressor systems that are the most successful. [ABSTRACT FROM AUTHOR]

Efforts to improve recreational fisheries have included widespread stocking of Micropterus floridanus outside its native range of peninsular Florida. Hybridization of Florida bass ( M. floridanus) with largemouth bass ( Micropterus salmoides) has now dramatically expanded beyond a naturally occurring intergrade zone in the southeast U. S. In recent years, there has been growing interest in protecting the genetic integrity of native basses and assessing the impact and nature of M. salmoides/ M. floridanus introgression from the standpoint of hatchery and sport-fishery managers, fish biologists, ecologists and evolutionary biologists. Here, we conducted RNA-seq-based sequencing of the transcriptomes of M. salmoides, M. floridanus and their F1 hybrid and identified a set of 3674 SNP markers with fixed-allelic differences from 2112 unique genes. We then developed a subset of 25 of these markers into a single diagnostic multiplex assay and validated its capacity for assessing integrity and hybridization in hatchery and wild populations of largemouth and Florida bass. The availability of this resource, high-quality transcriptomes and a large set of gene-linked SNPs, should greatly facilitate functional and population genomics studies in these key species and allow the identification of traits and processes under selection during introgressive hybridization. [ABSTRACT FROM AUTHOR]

Blue catfish, Ictalurus furcatus, are valued in the United States as a trophy fishery for their capacity to reach large sizes, sometimes exceeding 45 kg. Additionally, blue catfish × channel catfish ( I. punctatus) hybrid food fish production has recently increased the demand for blue catfish broodstock. However, there has been little study of the genetic impacts and interaction of farmed, introduced and stocked populations of blue catfish. We utilized genotyping-by-sequencing ( GBS) to capture and genotype SNP markers on 190 individuals from five wild and domesticated populations ( Mississippi River, Missouri, D& B, Rio Grande and Texas). Stringent filtering of SNP-calling parameters resulted in 4275 SNP loci represented across all five populations. Population genetics and structure analyses revealed potential shared ancestry and admixture between populations. We utilized the Sequenom Mass ARRAY to validate two multiplex panels of SNPs selected from the GBS data. Selection criteria included SNPs shared between populations, SNPs specific to populations, number of reads per individual and number of individuals genotyped by GBS. Putative SNPs were validated in the discovery population and in two additional populations not used in the GBS analysis. A total of 64 SNPs were genotyped successfully in 191 individuals from nine populations. Our results should guide the development of highly informative, flexible genotyping multiplexes for blue catfish from the larger GBS SNP set as well as provide an example of a rapid, low-cost approach to generate and genotype informative marker loci in aquatic species with minimal previous genetic information. [ABSTRACT FROM AUTHOR]

The southeastern US has experienced recurrent drought during recent decades. Increasing demand for water, as precipitation decreases, exacerbates stress on the aquatic biota of the Southeast: a global hotspot for freshwater mussel, crayfish, and fish diversity. Freshwater unionid mussels are ideal candidates to study linkages between ecophysiological and behavioral responses to drought. Previous work on co-occurring mussel species suggests a coupling of physiology and behavior along a gradient ranging from intolerant species such as Pyganodon grandis (giant floater) that track receding waters and rarely burrow in the substrates to tolerant species such as Uniomerus tetralasmus (pondhorn) that rarely track receding waters, but readily burrow into the drying sediments. We utilized a next-generation sequencing-based RNA-Seq approach to examine heat/desiccation-induced transcriptomic profiles of these two species in order to identify linkages between patterns of gene expression, physiology and behavior. Sequencing produced over 425 million 100 bp reads. Using the de novo assembly package Trinity, we assembled the short reads into 321,250 contigs from giant floater (average length 835 bp) and 385,735 contigs from pondhorn (average length 929 bp). BLAST-based annotation and gene expression analysis revealed 2,832 differentially expressed genes in giant floater and 2,758 differentially expressed genes in pondhorn. Trancriptomic responses included changes in molecular chaperones, oxidative stress profiles, cell cycling, energy metabolism, immunity, and cytoskeletal rearrangements. Comparative analyses between species indicated significantly higher induction of molecular chaperones and cytoskeletal elements in the intolerant P. grandis as well as important differences in genes regulating apoptosis and immunity. [ABSTRACT FROM AUTHOR]

Lab-on-a-chip based Polymerase Chain Reaction-Restriction Fragment Length Polymorphism (PCR-RFLP) technology was developed for the identification of seven catfish species including an Ictalurid hybrid. RFLP profiles of mitochondrial cytochrome b fragments digested by three different restriction endonucleases (DdeI,HaeIII, andNlaIII) were constructed from morphologically verified catfish samples on an Agilent 2100 Bioanalyzer (Agilent Technologies, Inc., Santa Clara, CA, USA). High sensitivity and ease of handling through end-point microchip-based capillary electrophoresis increased resolution and accuracy of DNA fragment sizing. RFLP profiles derived from a combination of all three enzymes produced consistent species-specific identification profiles. Observed restriction fragment patterns forClarias batrachusandIctalurus punctatuswere similar, but both of these species could be consistently differentiated using a single band of theHaeIII restriction site. Assay advantages due to shorter assay times, assay ease, and minimum usage of harmful solvents and chemicals when compared to traditional DNA barcoding were counterbalanced by the need to develop and optimize specific restriction digest profiles for all potential species of interest. With further development, this method may be utilized in testing of catfish products to ensure the enforcement of seafood labeling regulations. [ABSTRACT FROM PUBLISHER]

Short-term feed deprivation (or fasting) is a common occurrence in aquacultured fish species whether due to season, production strategies, or disease. In channel catfish (Ictalurus punctatus) fasting impacts susceptibility to several bacterial pathogens including Flavobacterium columnare, the causative agent of columnaris disease. As columnaris gains entry through the gills and skin of fish, we examined here changes in transcriptional regulation induced in these surface mucosal tissues due to short-term (7 day) fasting. RNA-seq expression analysis revealed a total of 1,545 genes perturbed by fasting. Fasting significantly altered expression of critical innate immune factors in a manner consistent with lower immune fitness as well as dysregulating key genes involved in energy metabolism and cell cycling/proliferation. Downregulation of innate immune actors such as iNOS2b, Lysozyme C, and peptidoglycan recognition protein 6 is predicted to impact the delicate recognition/tolerance balance for commensal and pathogenic bacteria on the skin and gill. The highlighted expression profiles reveal potential mechanistic similarities between gut and surface mucosa and underscore the complex interrelationships between nutrition, mucosal integrity, and immunity in teleost fish. [ABSTRACT FROM AUTHOR]

Temperature is one of the most prominent abiotic factors affecting ectotherms. Most fish species, as ectotherms, have extraordinary ability to deal with a wide range of temperature changes. While the molecular mechanism underlying temperature adaptation has long been of interest, it is still largely unexplored with fish. Understanding of the fundamental mechanisms conferring tolerance to temperature fluctuations is a topic of increasing interest as temperature may continue to rise as a result of global climate change. Catfish have a wide natural habitat and possess great plasticity in dealing with environmental variations in temperature. However, no studies have been conducted at the transcriptomic level to determine heat stress-induced gene expression. In the present study, we conducted an RNA-Seq analysis to identify heat stress-induced genes in catfish at the transcriptome level. Expression analysis identified a total of 2,260 differentially expressed genes with a cutoff of twofold change. qRT-PCR validation suggested the high reliability of the RNA-Seq results. Gene ontology, enrichment, and pathway analyses were conducted to gain insight into physiological and gene pathways. Specifically, genes involved in oxygen transport, protein folding and degradation, and metabolic process were highly induced, while general protein synthesis was dramatically repressed in response to the lethal temperature stress. This is the first RNA-Seq-based expression study in catfish in response to heat stress. The candidate genes identified should be valuable for further targeted studies on heat tolerance, thereby assisting the development of heat-tolerant catfish lines for aquaculture. [ABSTRACT FROM AUTHOR]

Background: Single nucleotide polymorphisms (SNPs) have become the marker of choice for genome-wide association studies. In order to provide the best genome coverage for the analysis of performance and production traits, a large number of relatively evenly distributed SNPs are needed. Gene-associated SNPs may fulfill these requirements of large numbers and genome wide distribution. In addition, gene-associated SNPs could themselves be causative SNPs for traits. The objective of this project was to identify large numbers of gene-associated SNPs using high-throughput next generation sequencing. Results: Transcriptome sequencing was conducted for channel catfish and blue catfish using Illumina next generation sequencing technology. Approximately 220 million reads (15.6 Gb) for channel catfish and 280 million reads (19.6 Gb) for blue catfish were obtained by sequencing gene transcripts derived from various tissues of multiple individuals from a diverse genetic background. A total of over 35 billion base pairs of expressed short read sequences were generated. Over two million putative SNPs were identified from channel catfish and almost 2.5 million putative SNPs were identified from blue catfish. Of these putative SNPs, a set of filtered SNPs were identified including 342,104 intra-specific SNPs for channel catfish, 366,269 intra-specific SNPs for blue catfish, and 420,727 inter-specific SNPs between channel catfish and blue catfish. These filtered SNPs are distributed within 16,562 unique genes in channel catfish and 17,423 unique genes in blue catfish. Conclusions: For aquaculture species, transcriptome analysis of pooled RNA samples from multiple individuals using Illumina sequencing technology is both technically efficient and cost-effective for generating expressed sequences. Such an approach is most effective when coupled to existing EST resources generated using traditional sequencing approaches because the reference ESTs facilitate effective assembly of the expressed short reads. When multiple individuals with different genetic backgrounds are used, RNA-Seq is very effective for the identification of SNPs. The SNPs identified in this report will provide a much needed resource for genetic studies in catfish and will contribute to the development of a high-density SNP array. Validation and testing of these SNPs using SNP arrays will form the material basis for genome association studies and whole genome-based selection in catfish. [ABSTRACT FROM AUTHOR]

The article focuses on the study concerning gene expression and molecular characterization after iron treatment and bacterial infection of the channel catfish ferritin H subunit. The intention of the study of is to focus on the channel catfish on its ferritin H gene and to settle its copy numbers and genomic organization, to establish defense response after bacterial infection and to find its pattern of tissue expression. The study shows that catfish's ferritin H gene is highly conserved throughout the evolution.

Background: Genome annotation projects, gene functional studies, and phylogenetic analyses for a given organism all greatly benefit from access to a validated full-length cDNA resource. While increasingly common in model species, fulllength cDNA resources in aquaculture species are scarce. Methodology and Principal Findings: Through in silico analysis of catfish (Ictalurus spp.) ESTs, a total of 10,037 channel catfish and 7,382 blue catfish cDNA clones were identified as potentially encoding full-length cDNAs. Of this set, a total of 1,169 channel catfish and 933 blue catfish full-length cDNA clones were selected for re-sequencing to provide additional coverage and ensure sequence accuracy. A total of 1,745 unique gene transcripts were identified from the full-length cDNA set, including 1,064 gene transcripts from channel catfish and 681gene transcripts from blue catfish, with 416 transcripts shared between the two closely related species. Full-length sequence characteristics (ortholog conservation, UTR length, Kozak sequence, and conserved motifs) of the channel and blue catfish were examined in detail. Comparison of gene ontology composition between full-length cDNAs and all catfish ESTs revealed that the full-length cDNA set is representative of the gene diversity encoded in the catfish transcriptome. Conclusions: This study describes the first catfish full-length cDNA set constructed from several cDNA libraries. The catfish full-length cDNA sequences, and data gleaned from sequence characteristics analysis, will be a valuable resource for ongoing catfish whole-genome sequencing and future gene-based studies of function and evolution in teleost fishes. [ABSTRACT FROM AUTHOR]

Alternative splicing (AS) is a mechanism by which the coding diversity of the genome can be greatly increased. Rates of AS are known to vary according to the complexity of eukaryotic species potentially explaining the tremendous phenotypic diversity among species with similar numbers of coding genes. Little is known, however, about the nature or rate of AS in teleost fish. Here, we report the characteristics of AS in teleost fish and classification and frequency of five canonical AS types. We conducted both same-species and cross-species analysis utilizing the Genome Mapping and Alignment Program (GMAP) and an AS pipeline (ASpipe) to study AS in four genome-enabled species ( Danio rerio, Oryzias latipes, Gasterosteus aculeatus, and Takifugu rubripes) and one species lacking a complete genome sequence, Ictalurus punctatus. AS frequency was lowest in the highly duplicated genome of zebrafish (17% of mapped genes). The compact genome of the pufferfish showed the highest occurrence of AS (~43% of mapped genes). An inverse correlation between AS frequency and genome size was consistent across all analyzed species. Cross-species comparisons utilizing zebrafish as the reference genome allowed the identification of additional putative AS genes not revealed by zebrafish transcripts. Approximately, 50% of AS genes identified by same-species comparisons were shared among two or more species. A searchable website, the Teleost Alternative Splicing Database, was created to allow easy identification and visualization of AS transcripts in the studied teleost genomes. Our results and associated database should further our understanding of alternative splicing as an important functional and evolutionary mechanism in the genomes of teleost fish. [ABSTRACT FROM AUTHOR]

Background: SNPs are abundant, codominantly inherited, and sequence-tagged markers. They are highly adaptable to large-scale automated genotyping, and therefore, are most suitable for association studies and applicable to comparative genome analysis. However, discovery of SNPs requires genome sequencing efforts through whole genome sequencing or deep sequencing of reduced representation libraries. Such genome resources are not yet available for many species including catfish. A large resource of ESTs is to become available in catfish allowing identification of large number of SNPs, but reliability of EST-derived SNPs are relatively low because of sequencing errors. This project was designed to answer some of the questions relevant to quality assessment of EST-derived SNPs. Results: wo factors were found to be most significant for validation of EST-derived SNPs: the contig size (number of sequences in the contig) and the minor allele sequence frequency. The larger the contigs were, the greater the validation rate although the validation rate was reasonably high when the contigs contain four or more EST sequences with the minor allele sequence being represented at least twice in the contigs. Sequence quality surrounding the SNP under test is also crucially important. PCR extension appeared to be limited to a very short distance, prohibiting successful genotyping when an intron was present, a surprising finding. Conclusion: Stringent quality assessment measures should be used when working with ESTderived SNPs. In particular, contigs containing four or more ESTs should be used and the minor allele sequence should be represented at least twice. Genotyping primers should be designed from a single exon, completely avoiding introns. Application of such quality assessment measures, along with large resources of ESTs, should provide effective means for SNP identification in species where genome sequence resources are lacking. [ABSTRACT FROM AUTHOR]

Chemokines are a superfamily of cytokines responsible for regulating cell migration under both inflammatory and physiological conditions. CC chemokines are the largest subfamily of chemokines, with 28 members in humans. A subject of intense study in mammalian species, the known functional roles of CC chemokines ligands in both developmental and disease conditions continue to expand. They are also an important family for the study of gene copy number variation and tandem duplication in mammalian species. However, little is known regarding the evolutionary origin and status of these ligands in primitive vertebrates such as teleost fish. In this paper, we review the evolution of the teleost fish CC chemokine gene family, noting evidence of widespread tandem gene duplications and examining the implications of this phenomenon on immune diversity. Through extensive phylogenetic analysis of the CC chemokine sets of four teleost species, zebrafish, catfish, rainbow trout, and Atlantic salmon, we identified seven large groups of CC chemokines. It appeared that several major groups of CC chemokines are highly related including the CCL19/21/25 group, the CCL20 group, CCL27/28 group, and the fish-specific group. In the three remaining groups that contained the largest number of members, the CCL17/22 group, the MIP group, and the MCP group, similarities among species members were obscured by rapid, tandem duplications that may contribute to immune diversity. [ABSTRACT FROM AUTHOR]

Toll-like receptors (TLRs) are a family of transmembrane proteins that recognize specific pathogen-associated molecular patterns and use conserved signaling pathways to activate proinflammatory cytokines and type-1 interferons to fight infection. TLR3 in mammals is best known for its recognition of dsRNA as ligand and its MyD88-independent signaling. TLR3, upon recognition of dsRNA, recruits and binds its adaptor protein TIR domain-containing adapter molecule (TICAM) 1. Here we report the genomic sequences and structures of TLR3 and a TICAM adaptor from channel catfish ( Ictalurus punctatus). Whereas a partial TLR3 cDNA sequence has been reported from channel catfish, and complete TLR3 genes are known from other teleost fish species, a complete TICAM sequence has not been previously reported from a nonmammalian species. Analysis of catfish TLR3 and TICAM expression after infection with Edwardsiella ictaluri, the causative agent of enteric septicemia of catfish (ESC), suggested a conserved TLR3-TICAM receptor–adaptor relation in catfish. Comparison of TLR3 and TICAM expression profiles in channel catfish with those from the closely related blue catfish species ( Ictalurus furcatus), which exhibits strong resistance to ESC, revealed a striking pattern of species-specific expression. A dramatic downregulation of TLR3 and TICAM gene expression was observed in blue catfish head kidney and spleen, which we speculate may be the result of maturation and migration of different cell types to and from the lymphoid tissues following infection. [ABSTRACT FROM AUTHOR]

Chemokines, a superfamily of chemotactic cytokines involved in recruitment, activation, and adhesion of a variety of leukocyte types to inflammatory foci, are a crucial component of the immune system of Sarcopterygiian vertebrates. Although all mammalian chemokines are believed to have been found, the status of these molecules in Actinopterygii was unknown until recently. The identification of chemokines in fish species has been complicated by low sequence conservation and confusion over expected numbers. Earlier discoveries of single fish chemokines coupled with rapidly expanding genetic resources in these species have recently provided a foundation for large-scale in silico discoveries of these important immune regulators. We report here the identification and expression analysis of 12 new CC chemokine sequences from catfish. When added to our previous report of 14 catfish CC chemokines, the number of CC chemokines in catfish now stands at 26, two more than known from humans. Establishing orthologous relationships among the majority of catfish CC chemokines, a newly available set of chicken CC chemokines, and their mammalian counterparts remain difficult, suggesting high levels of duplication and divergence within individual species. [ABSTRACT FROM AUTHOR]

Chemokines represent a superfamily of chemotactic cytokines involved in recruitment, activation and adhesion of a variety of leukocyte types to inflammatory foci, as well as in the organization and maintenance of lymphoid organ architecture and in normal developmental processes. Nearly all chemokines have been identified in human and mouse, but only a handful of fish chemokines have been identified. Here we describe 14 distinct chemokines from channel catfish and blue catfish identified by analysis of 30,000 expressed sequence tags. Based on sequence analysis, sequence similarity, and the arrangement of the conserved cysteine residues, all 14 chemokines were identified as members of the CC subfamily. Phylogenetic analysis did not reveal clear evidence of orthology of the catfish and human or mouse chemokines. Similarity analysis indicated that nine of the 14 CC chemokines were identified for the first time in fish. The availability of this pool of catfish CC chemokines should facilitate rapid identification and phylogenetic analysis of CC chemokines from other fish and related species. [ABSTRACT FROM AUTHOR]

Striped bass ( Morone saxatilis) and white bass ( Morone chrysops) are the parental species of the hybrid striped bass, a major U.S. aquaculture species. Currently, genomic resources for striped bass, white bass, and their hybrid lag behind those of other aquaculture species. Current resources consist of a medium-density genetic linkage map and a well-annotated ovarian transcriptome. A well-annotated transcriptome from across striped bass and white bass tissues is needed to advance both broad-based RNA-seq studies of gene expression as well as aid in more targeted studies of important genes and pathways critical for reproductive physiology and immunity. Here, we carried out Illumina-based transcriptome sequencing and annotation in both species utilizing the trinity and trinotate packages. The assembled Moronid reference transcriptomes and identified SSRs and SNPs should advance ongoing studies of reproduction, physiology, and immunology in these species and provide markers for broodstock management and selection. [ABSTRACT FROM AUTHOR]

The impact of cortisol on Flavobacterium columnare biofilm formation was explored. Firstly, the dynamics of biofilm formation by one highly (HV) and one low virulent (LV) F. columnare isolate with and without the stress hormone cortisol under microfluidic flow conditions was characterized. This to confirm that F. columnare cells could form biofilm under cortisol supplementation, and to compare the temporal and structural differences between different treatment groups. One trial revealed that in both isolates cell aggregates resembling biofilms occurred within 7-h post-inoculation. Consequently, cell clusters were sloughed away, followed by a rebuilding of bacterial cell aggregates, suggestive for a high spreading capacity. While the HV isolate revealed cell aggregates formed upstream at all time-points, for the LV isolate this was only seen upon cortisol supplementation. Secondly, the transcriptional effect of genes (gldK, gldL, gldM, gldN, sprA, sprE, sprT, and porV) belonging to the Type IX secretion system involved in gliding motility was investigated in planktonic and biofilm cells of a HV and LV isolate to which no, a low (LD) or high (HD) dose of cortisol was added. Significantly lower expression of gliding genes gldK, gldL, gldM and gldN, and of protein secretion regulator porV was seen in the LV isolate planktonic cells supplemented with a HD-cortisol. The LV isolate biofilm cells treated with the HD-cortisol showed a significant upregulation of sprT, encoding mobile surface adhesion important in bacterial colonization. This is the first evidence for the co-regulatory effect of cortisol on biofilm formation and F. columnare gliding gene expression. [ABSTRACT FROM AUTHOR]

The current study was conducted to assess the effects of microinjection of different dosages of guide RNA (gRNA)/Cas9 protein on the mutation rate, embryo survival, embryonic development, hatchability and early fry survival in channel catfish, Ictalurus punctatus. Guide RNAs targeting two of the channel catfish immune-related genes, toll/interleukin 1 receptor domain-containing adapter molecule (TICAM 1) and rhamnose binding lectin (RBL) genes, were designed and prepared. Three dosages of gRNA/Cas9 protein (low, 2.5 ng gRNA/7.5 ng Cas9, medium, 5 ng gRNA/15 ng Cas9 and high, 7.5 ng gRNA/22.5 ng Cas9) were microinjected into the yolk of one-cell embryos. Mutation rate increased with higher dosages (p < 0.05). Higher dosages increased the mutation frequency in individual embryos where biallelic mutations were detected. For both genes, microinjection procedures increased the embryo mortality (p < 0.05). Increasing the dosage of gRNA/Cas9 protein increased the embryo mortality and reduced the hatching percent (p < 0.05). Embryonic development was delayed when gRNAs targeting RBL gene were injected. Means of fry survival time were similar for different dosages (p > 0.05). The current results lay the foundations for designing gene editing experiments in channel catfish and can be used as a guide for other fish species. [ABSTRACT FROM AUTHOR]