Dataset Information

Diversity and relative abundance of the bacterial pathogen, Flavobacterium spp., infecting reproductive ecotypes of kokanee salmon.

ABSTRACT: BACKGROUND: Understanding the distribution and abundance of pathogens can provide insight into the evolution and ecology of their host species. Previous research in kokanee, the freshwater form of sockeye salmon (Oncorhynchus nerka), found evidence that populations spawning in streams may experience a greater pathogen load compared with populations that spawn on beaches. In this study we tested for differences in the abundance and diversity of the gram-negative bacteria, Flavobacterium spp., infecting tissues of kokanee in both of these spawning habitats (streams and beaches). Molecular assays were carried out using primers designed to amplify a ~200 nucleotide region of the gene encoding the ATP synthase alpha subunit (AtpA) within the genus Flavobacterium. Using a combination of DNA sequencing and quantitative PCR (qPCR) we compared the diversity and relative abundance of Flavobacterium AtpA amplicons present in DNA extracted from tissue samples of kokanee collected from each spawning habitat. RESULTS: We identified 10 Flavobacterium AtpA haplotypes among the tissues of stream-spawning kokanee and seven haplotypes among the tissues of beach-spawning kokanee, with only two haplotypes shared between spawning habitats. Haplotypes occurring in the same clade as F. psychrophilum were the most prevalent (92% of all reads, 60% of all haplotypes), and occurred in kokanee from both spawning habitats (streams and beaches). Subsequent qPCR assays did not find any significant difference in the relative abundance of Flavobacterium AtpA amplicons between samples from the different spawning habitats. CONCLUSIONS: We confirmed the presence of Flavobacterium spp. in both spawning habitats and found weak evidence for increased Flavobacterium diversity in kokanee sampled from stream-spawning sites. However, the quantity of Flavobacterium DNA did not differ between spawning habitats. We recommend further study aimed at quantifying pathogen diversity and abundance in population-level samples of kokanee combined with environmental sampling to better understand the ecology of pathogen infection in this species.

SUBMITTER: Lemay MA

PROVIDER: S-EPMC4228061 | biostudies-literature | 2014

REPOSITORIES: biostudies-literature

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Diversity and relative abundance of the bacterial pathogen, Flavobacterium spp., infecting reproductive ecotypes of kokanee salmon.

Lemay Matthew A MA Russello Michael A MA

BMC research notes 20141104

<h4>Background</h4>Understanding the distribution and abundance of pathogens can provide insight into the evolution and ecology of their host species. Previous research in kokanee, the freshwater form of sockeye salmon (Oncorhynchus nerka), found evidence that populations spawning in streams may experience a greater pathogen load compared with populations that spawn on beaches. In this study we tested for differences in the abundance and diversity of the gram-negative bacteria, Flavobacterium sp ...[more]

PMID: 25367228

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Project description:BACKGROUND:High throughput next-generation sequencing technology has enabled the collection of genome-wide sequence data and revolutionized single nucleotide polymorphism (SNP) discovery in a broad range of species. When analyzed within a population genomics framework, SNP-based genotypic data may be used to investigate questions of evolutionary, ecological, and conservation significance in natural populations of non-model organisms. Kokanee salmon are recently diverged freshwater populations of sockeye salmon (Oncorhynchus nerka) that exhibit reproductive ecotypes (stream-spawning and shore-spawning) in lakes throughout western North America and northeast Asia. Current conservation and management strategies may treat these ecotypes as discrete stocks, however their recent divergence and low levels of gene flow make in-season genetic stock identification a challenge. The development of genome-wide SNP markers is an essential step towards fine-scale stock identification, and may enable a direct investigation of the genetic basis of ecotype divergence. RESULTS:We used pooled cDNA samples from both ecotypes of kokanee to generate 750 million base pairs of transcriptome sequence data. These raw data were assembled into 11,074 high coverage contigs from which we identified 32,699 novel single nucleotide polymorphisms. A subset of these putative SNPs was validated using high-resolution melt analysis and Sanger resequencing to genotype independent samples of kokanee and anadromous sockeye salmon. We also identified a number of contigs that were composed entirely of reads from a single ecotype, which may indicate regions of differential gene expression between the two reproductive ecotypes. In addition, we found some evidence for greater pathogen load among the kokanee sampled in stream-spawning habitats, suggesting a possible evolutionary advantage to shore-spawning that warrants further study. CONCLUSIONS:This study provides novel genomic resources to support population genetic and genomic studies of both kokanee and anadromous sockeye salmon, and has the potential to produce markers capable of fine-scale stock assessment. While this RNAseq approach was successful at identifying a large number of new SNP loci, we found that the frequency of alleles present in the pooled transcriptome data was not an accurate predictor of population allele frequencies.

Project description:Flavobacterium spartansii strain T16T was isolated from a disease outbreak in hatchery-reared Chinook salmon (Oncorhynchus tshawytscha) fingerlings. To gain insight into its genomic content, structure and virulence pathogenesis factors, comparative genome analyses were performed using genomes from environmental and virulent Flavobacterium strains. F. spartansii shared low average nucleotide identity (ANI) to well-known fish-pathogenic flavobacteria (e.g., F. columnare, F. psychrophilum, and F. branchiophilum), indicating that it is a new and emerging fish pathogen. The genome in T16T had a length of 5,359,952 bp, a GC-content 35.7%, and 4,422 predicted protein-coding sequences. Flavobacterium core genome analysis showed that the number of shared genes decreased with the addition of input genomes and converged at 1182 genes. At least 8 genomic islands and 5 prophages were predicted in T16T. At least 133 virulence factors associated with virulence in pathogenic bacteria were highly conserved in F. spartansii T16T. Furthermore, genes linked to virulence in other bacterial species (e.g., those encoding for a type IX secretion system, collagenase and hemolysin) were found in the genome of F. spartansii T16T and were conserved in most of the analyzed pathogenic Flavobacterium. F. spartansii was resistant to ampicillin and penicillin, consistent with the presence of multiple genes encoding diverse lactamases and the penicillin-binding protein in the genome. To allow for future investigations into F. spartansii virulence in vivo, a transposon-based random mutagenesis strategy was attempted in F. spartansii T16T using pHimarEm1. Four putative gliding motility deficient mutants were obtained and the insertion sites of pHimarEm1 in the genome of these mutants were characterized. In total, study results clarify some of the mechanisms by which emerging flavobacterial fish pathogens may cause disease and also provide direly needed tools to investigate their pathogenesis.

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Dataset Information

Diversity and relative abundance of the bacterial pathogen, Flavobacterium spp., infecting reproductive ecotypes of kokanee salmon.

Publications

Diversity and relative abundance of the bacterial pathogen, Flavobacterium spp., infecting reproductive ecotypes of kokanee salmon.

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