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July 7, 2019

The characterization of goat genetic diversity: Towards a genomic approach

The investigation of genetic diversity at molecular level has been proposed as a valuable complement and sometimes proxy to phenotypic diversity of local breeds and is presently considered as one of the FAO priorities for breed characterization. By recommending a set of selected molecular markers for each of the main livestock species, FAO has promoted the meta-analysis of local datasets, to achieve a global view of molecular genetic diversity. Analysis within the EU Globaldiv project of two large goat microsatellite datasets produced by the Econogene Consortium and the IAEA CRP–Asia Consortium, respectively, has generated a picture of goat diversity across continents. This indicates a gradient of decreasing diversity from the domestication centre towards Europe and Asia, a clear phylogeographic structure at the continental and regional levels, and in Asia a limited genetic differentiation among local breeds. The development of SNP panels that assay thousands of markers and the whole genome sequencing of livestock permit an affordable use of genomic technologies in all livestock species, goats included. Preliminary data from the Italian Goat Consortium indicate that the SNP panel developed for this species is highly informative. The existing panel can be improved by integrating additional SNPs identified from the whole genome sequence alignment of goats adapted to extreme climates. Part of this effort is being achieved by international projects (e.g. EU FP7 NextGen and 3SR projects), but a fair representation of the global diversity in goats requires a large panel of samples (i.e. as in the recently launched 1000 cattle genomes initiative). Genomic technologies offer new strategies to investigate complex traits difficult to measure. For example, the comparison of patterns of diversity among the genomes in selected groups of animals (e.g. adapted to different environments) and the integration of genome-wide diversity with new GIScience-based methods are able to identify molecular markers associated with genomic regions of putative importance in adaptation and thus pave the way for the identification of causative genes. Goat breeds adapted to different production systems in extreme and harsh environments will play an important role in this process. The new sequencing technologies also permit the analysis of the entire mitochondrial genome at maximum resolution. The complete mtDNA sequence is now the common standard format for the investigation of human maternal lineages. A preliminary analysis of the complete goat mtDNA genome supports a single Neolithic origin of domestic goats rather than multiple domestication events in different geographic areas.

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July 7, 2019

Genome sequencing of an extended series of NDM-producing Klebsiella pneumoniae isolates from Neonatal infections in a Nepali hospital characterizes the extent of community- versus hospital-associated transmission in an endemic setting.

NDM-producing Klebsiella pneumoniae strains represent major clinical and infection control challenges, particularly in resource-limited settings with high rates of antimicrobial resistance. Determining whether transmission occurs at a gene, plasmid, or bacterial strain level and within hospital and/or the community has implications for monitoring and controlling spread. Whole-genome sequencing (WGS) is the highest-resolution typing method available for transmission epidemiology. We sequenced carbapenem-resistant K. pneumoniae isolates from 26 individuals involved in several infection case clusters in a Nepali neonatal unit and 68 other clinical Gram-negative isolates from a similar time frame, using Illumina and PacBio technologies. Within-outbreak chromosomal and closed-plasmid structures were generated and used as data set-specific references. Three temporally separated case clusters were caused by a single NDM K. pneumoniae strain with a conserved set of four plasmids, one being a 304,526-bp plasmid carrying blaNDM-1. The plasmids contained a large number of antimicrobial/heavy metal resistance and plasmid maintenance genes, which may have explained their persistence. No obvious environmental/human reservoir was found. There was no evidence of transmission of outbreak plasmids to other Gram-negative clinical isolates, although blaNDM variants were present in other isolates in different genetic contexts. WGS can effectively define complex antimicrobial resistance epidemiology. Wider sampling frames are required to contextualize outbreaks. Infection control may be effective in terminating outbreaks caused by particular strains, even in areas with widespread resistance, although this study could not demonstrate evidence supporting specific interventions. Larger, detailed studies are needed to characterize resistance genes, vectors, and host strains involved in disease, to enable effective intervention. Copyright © 2014 Stoesser et al.

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July 7, 2019

A precise chloroplast genome of Nelumbo nucifera (Nelumbonaceae) evaluated with Sanger, Illumina MiSeq, and PacBio RS II sequencing platforms: insight into the plastid evolution of basal eudicots.

BackgroundThe chloroplast genome is important for plant development and plant evolution. Nelumbo nucifera is one member of relict plants surviving from the late Cretaceous. Recently, a new sequencing platform PacBio RS II, known as `SMRT (Single Molecule, Real-Time) sequencing¿, has been developed. Using the SMRT sequencing to investigate the chloroplast genome of N. nucifera will help to elucidate the plastid evolution of basal eudicots.ResultsThe sizes of the de novo assembled complete chloroplast genome of N. nucifera were 163,307 bp, 163,747 bp and 163,600 bp with average depths of coverage of 7×, 712× and 105× sequenced by Sanger, Illumina MiSeq and PacBio RS II, respectively. The precise chloroplast genome of N. nucifera was obtained from PacBio RS II data proofread by Illumina MiSeq reads, with a quadripartite structure containing a large single copy region (91,846 bp) and a small single copy region (19,626 bp) separated by two inverted repeat regions (26,064 bp). The genome contains 113 different genes, including four distinct rRNAs, 30 distinct tRNAs and 79 distinct peptide-coding genes. A phylogenetic analysis of 133 taxa from 56 orders indicated that Nelumbo with an age of 177 million years is a sister clade to Platanus, which belongs to the basal eudicots. Basal eudicots began to emerge during the early Jurassic with estimated divergence times at 197 million years using MCMCTree. IR expansions/contractions within the basal eudicots seem to have occurred independently.ConclusionsBecause of long reads and lack of bias in coverage of AT-rich regions, PacBio RS II showed a great promise for highly accurate `finished¿ genomes, especially for a de novo assembly of genomes. N. nucifera is one member of basal eudicots, however, evolutionary analyses of IR structural variations of N. nucifera and other basal eudicots suggested that IR expansions/contractions occurred independently in these basal eudicots or were caused by independent insertions and deletions. The precise chloroplast genome of N. nucifera will present new information for structural variation of chloroplast genomes and provide new insight into the evolution of basal eudicots at the primary sequence and structural level.

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July 7, 2019

Co-option of Sox3 as the male-determining factor on the Y chromosome in the fish Oryzias dancena.

Sex chromosomes harbour a primary sex-determining signal that triggers sexual development of the organism. However, diverse sex chromosome systems have been evolved in vertebrates. Here we use positional cloning to identify the sex-determining locus of a medaka-related fish, Oryzias dancena, and find that the locus on the Y chromosome contains a cis-regulatory element that upregulates neighbouring Sox3 expression in developing gonad. Sex-reversed phenotypes in Sox3(Y) transgenic fish, and Sox3(Y) loss-of-function mutants all point to its critical role in sex determination. Furthermore, we demonstrate that Sox3 initiates testicular differentiation by upregulating expression of downstream Gsdf, which is highly conserved in fish sex differentiation pathways. Our results not only provide strong evidence for the independent recruitment of Sox3 to male determination in distantly related vertebrates, but also provide direct evidence that a novel sex determination pathway has evolved through co-option of a transcriptional regulator potentially interacted with a conserved downstream component.

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July 7, 2019

Complete genome sequence of Pseudomonas rhizosphaerae IH5(T) (=DSM 16299(T)), a phosphate-solubilizing rhizobacterium for bacterial biofertilizer.

Pseudomonas rhizosphaerae IH5(T) (=DSM 16299(T)), isolated from the rhizospheric soil of grass growing in Spain, has been reported as a novel species of the genus Pseudomonas harboring insoluble phosphorus solubilizing activity. To understanding the multifunctional biofertilizer better, we report the complete genome sequence of P. rhizosphaerae IH5(T). Copyright © 2014 Elsevier B.V. All rights reserved.

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July 7, 2019

Complete genome sequence for the shellfish pathogen Vibrio coralliilyticus RE98 isolated from a shellfish hatchery.

Vibrio coralliilyticus is a pathogen of corals and larval shellfish. Publications on strain RE98 list it as a Vibrio tubiashii; however, whole genome sequencing confirms RE98 as V. coralliilyticus containing a total of 6,037,824 bp consisting of two chromosomes (3,420,228 and 1,917,482 bp) and two megaplasmids (380,714 and 319,400 bp). Copyright © 2014 Richards et al.

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July 7, 2019

Genome sequence of the dark pink pigmented Listia bainesii microsymbiont Methylobacterium sp. WSM2598.

Strains of a pink-pigmented Methylobacterium sp. are effective nitrogen- (N2) fixing microsymbionts of species of the African crotalarioid genus Listia. Strain WSM2598 is an aerobic, motile, Gram-negative, non-spore-forming rod isolated in 2002 from a Listia bainesii root nodule collected at Estcourt Research Station in South Africa. Here we describe the features of Methylobacterium sp. WSM2598, together with information and annotation of a high-quality draft genome sequence. The 7,669,765 bp draft genome is arranged in 5 scaffolds of 83 contigs, contains 7,236 protein-coding genes and 18 RNA-only encoding genes. This rhizobial genome is one of 100 sequenced as part of the DOE Joint Genome Institute 2010 G enomic E ncyclopedia for B acteria and A rchaea- R oot N odule B acteria (GEBA-RNB) project.

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July 7, 2019

Complete genome determination and analysis of Acholeplasma oculi strain 19L, highlighting the loss of basic genetic features in the Acholeplasmataceae.

BACKGROUND: Acholeplasma oculi belongs to the Acholeplasmataceae family, comprising the genera Acholeplasma and ‘Candidatus Phytoplasma’. Acholeplasmas are ubiquitous saprophytic bacteria. Several isolates are derived from plants or animals, whereas phytoplasmas are characterised as intracellular parasitic pathogens of plant phloem and depend on insect vectors for their spread. The complete genome sequences for eight strains of this family have been resolved so far, all of which were determined depending on clone-based sequencing. RESULTS:The A. oculi strain 19L chromosome was sequenced using two independent approaches. The first approach comprised sequencing by synthesis (Illumina) in combination with Sanger sequencing, while single molecule real time sequencing (PacBio) was used in the second. The genome was determined to be 1,587,120bp in size. Sequencing by synthesis resulted in six large genome fragments, while the single molecule real time sequencing approach yielded one circular chromosome sequence. High-quality sequences were obtained by both strategies differing in six positions, which are interpreted as reliable variations present in the culture population. Our genome analysis revealed 1,471 protein-coding genes and highlighted the absence of the F1FO-type Na+ ATPase system and GroEL/ES chaperone. Comparison of the four available Acholeplasma sequences revealed a core-genome encoding 703 proteins and a pan-genome of 2,867 proteins. CONCLUSIONS:The application of two state-of-the-art sequencing technologies highlights the potential of single molecule real time sequencing for complete genome determination. Comparative genome analyses revealed that the process of losing particular basic genetic features during genome reduction occurs in both genera, as indicated for several phytoplasma strains and at least A. oculi. The loss of the F1FO-type Na+ ATPase system may separate Acholeplasmataceae from other Mollicutes, while the loss of those genes encoding the chaperone GroEL/ES is not a rare exception in this bacterial class.

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July 7, 2019

Potential impact on kidney infection: a whole-genome analysis of Leptospira santarosai serovar Shermani.

Leptospira santarosai serovar Shermani is the most frequently encountered serovar, and it causes leptospirosis and tubulointerstitial nephritis in Taiwan. This study aims to complete the genome sequence of L. santarosai serovar Shermani and analyze the transcriptional responses of L. santarosai serovar Shermani to renal tubular cells. To assemble this highly repetitive genome, we combined reads that were generated from four next-generation sequencing platforms by using hybrid assembly approaches to finish two-chromosome contiguous sequences without gaps by validating the data with optical restriction maps and Sanger sequencing. Whole-genome comparison studies revealed a 28-kb region containing genes that encode transposases and hypothetical proteins in L. santarosai serovar Shermani, but this region is absent in other pathogenic Leptospira spp. We found that lipoprotein gene expression in both L. santarosai serovar Shermani and L. interrogans serovar Copenhageni were upregulated upon interaction with renal tubular cells, and LSS19962, a L. santarosai serovar Shermani-specific gene within a 28-kb region that encodes hypothetical proteins, was upregulated in L. santarosai serovar Shermani-infected renal tubular cells. Lipoprotein expression during leptospiral infection might facilitate the interactions of leptospires within kidneys. The availability of the whole-genome sequence of L. santarosai serovar Shermani would make it the first completed sequence of this species, and its comparison with that of other Leptospira spp. may provide invaluable information for further studies in leptospiral pathogenesis.

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July 7, 2019

Ferrets exclusively synthesize Neu5Ac and express naturally humanized influenza A virus receptors.

Mammals express the sialic acids N-acetylneuraminic acid (Neu5Ac) and N-glycolylneuraminic acid (Neu5Gc) on cell surfaces, where they act as receptors for pathogens, including influenza A virus (IAV). Neu5Gc is synthesized from Neu5Ac by the enzyme cytidine monophosphate-N-acetylneuraminic acid hydroxylase (CMAH). In humans, this enzyme is inactive and only Neu5Ac is produced. Ferrets are susceptible to human-adapted IAV strains and have been the dominant animal model for IAV studies. Here we show that ferrets, like humans, do not synthesize Neu5Gc. Genomic analysis reveals an ancient, nine-exon deletion in the ferret CMAH gene that is shared by the Pinnipedia and Musteloidia members of the Carnivora. Interactions between two human strains of IAV with the sialyllactose receptor (sialic acid-a2,6Gal) confirm that the type of terminal sialic acid contributes significantly to IAV receptor specificity. Our results indicate that exclusive expression of Neu5Ac contributes to the susceptibility of ferrets to human-adapted IAV strains.

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July 7, 2019

Dissemination of cephalosporin resistance genes between Escherichia coli strains from farm animals and humans by specific plasmid lineages.

Third-generation cephalosporins are a class of ß-lactam antibiotics that are often used for the treatment of human infections caused by Gram-negative bacteria, especially Escherichia coli. Worryingly, the incidence of human infections caused by third-generation cephalosporin-resistant E. coli is increasing worldwide. Recent studies have suggested that these E. coli strains, and their antibiotic resistance genes, can spread from food-producing animals, via the food-chain, to humans. However, these studies used traditional typing methods, which may not have provided sufficient resolution to reliably assess the relatedness of these strains. We therefore used whole-genome sequencing (WGS) to study the relatedness of cephalosporin-resistant E. coli from humans, chicken meat, poultry and pigs. One strain collection included pairs of human and poultry-associated strains that had previously been considered to be identical based on Multi-Locus Sequence Typing, plasmid typing and antibiotic resistance gene sequencing. The second collection included isolates from farmers and their pigs. WGS analysis revealed considerable heterogeneity between human and poultry-associated isolates. The most closely related pairs of strains from both sources carried 1263 Single-Nucleotide Polymorphisms (SNPs) per Mbp core genome. In contrast, epidemiologically linked strains from humans and pigs differed by only 1.8 SNPs per Mbp core genome. WGS-based plasmid reconstructions revealed three distinct plasmid lineages (IncI1- and IncK-type) that carried cephalosporin resistance genes of the Extended-Spectrum Beta-Lactamase (ESBL)- and AmpC-types. The plasmid backbones within each lineage were virtually identical and were shared by genetically unrelated human and animal isolates. Plasmid reconstructions from short-read sequencing data were validated by long-read DNA sequencing for two strains. Our findings failed to demonstrate evidence for recent clonal transmission of cephalosporin-resistant E. coli strains from poultry to humans, as has been suggested based on traditional, low-resolution typing methods. Instead, our data suggest that cephalosporin resistance genes are mainly disseminated in animals and humans via distinct plasmids.

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July 7, 2019

Inconsistency of phenotypic and genomic characteristics of Campylobacter fetus subspecies requires reevaluation of current diagnostics.

Classifications of the Campylobacter fetus subspecies fetus and venerealis were first described in 1959 and were based on the source of isolation (intestinal versus genital) and the ability of the strains to proliferate in the genital tract of cows. Two phenotypic assays (1% glycine tolerance and H2S production) were described to differentiate the subspecies. Multiple molecular assays have been applied to differentiate the C. fetus subspecies, but none of these tests is consistent with the phenotypic identification methods. In this study, we defined the core genome and accessory genes of C. fetus, which are based on the closed genomes of five C. fetus strains. Phylogenetic analysis of the core genomes of 23 C. fetus strains of the two subspecies showed a division into two clusters. The phylogenetic core genome clusters were not consistent with the phenotypic classifications of the C. fetus subspecies. However, they were consistent with the molecular characteristics of the strains, which were determined by multilocus sequence typing, sap typing, and the presence/absence of insertion sequences and a type I restriction modification system. The similarity of the genome characteristics of three of the phenotypically defined C. fetus subsp. fetus strains to C. fetus subsp. venerealis strains, when considering the core genome and accessory genes, requires a critical evaluation of the clinical relevance of C. fetus subspecies identification by phenotypic assays. Copyright © 2014, American Society for Microbiology. All Rights Reserved.

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