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April 21, 2020  |  

Genomic investigation of Staphylococcus aureus recovered from Gambian women and newborns following an oral dose of intra-partum azithromycin.

Oral azithromycin given during labour reduces carriage of bacteria responsible for neonatal sepsis, including Staphylococcus aureus. However, there is concern that this may promote drug resistance.Here, we combine genomic and epidemiological data on S. aureus isolated from mothers and babies in a randomized intra-partum azithromycin trial (PregnAnZI) to describe bacterial population dynamics and resistance mechanisms.Participants from both arms of the trial, who carried S. aureus in day 3 and day 28 samples post-intervention, were included. Sixty-six S. aureus isolates (from 7 mothers and 10 babies) underwent comparative genome analyses and the data were then combined with epidemiological data. Trial registration (main trial): ClinicalTrials.gov Identifier NCT01800942.Seven S. aureus STs were identified, with ST5 dominant (n?=?40, 61.0%), followed by ST15 (n?=?11, 17.0%). ST5 predominated in the placebo arm (73.0% versus 49.0%, P?=?0.039) and ST15 in the azithromycin arm (27.0% versus 6.0%, P?=?0.022). In azithromycin-resistant isolates, msr(A) was the main macrolide resistance gene (n?=?36, 80%). Ten study participants, from both trial arms, acquired azithromycin-resistant S. aureus after initially harbouring a susceptible isolate. In nine (90%) of these cases, the acquired clone was an msr(A)-containing ST5 S. aureus. Long-read sequencing demonstrated that in ST5, msr(A) was found on an MDR plasmid.Our data reveal in this Gambian population the presence of a dominant clone of S. aureus harbouring plasmid-encoded azithromycin resistance, which was acquired by participants in both arms of the study. Understanding these resistance dynamics is crucial to defining the public health drug resistance impacts of azithromycin prophylaxis given during labour in Africa. © The Author(s) 2019. Published by Oxford University Press on behalf of the British Society for Antimicrobial Chemotherapy.


April 21, 2020  |  

One Aeromonas salmonicida subsp. salmonicida isolate with a pAsa5 variant bearing antibiotic resistance and a pRAS3 variant making a link with a swine pathogen.

The Gram-negative bacterium Aeromonas salmonicida subsp. salmonicida is an aquatic pathogen which causes furunculosis to salmonids, especially in fish farms. The emergence of strains of this bacterium exhibiting antibiotic resistance is increasing, limiting the effectiveness of antibiotherapy as a treatment against this worldwide disease. In the present study, we discovered an isolate of A. salmonicida subsp. salmonicida that harbors two novel plasmids variants carrying antibiotic resistance genes. The use of long-read sequencing (PacBio) allowed us to fully characterize those variants, named pAsa5-3432 and pRAS3-3432, which both differ from their classic counterpart through their content in mobile genetic elements. The plasmid pAsa5-3432 carries a new multidrug region composed of multiple mobile genetic elements, including a Class 1 integron similar to an integrated element of Salmonella enterica. With this new region, probably acquired through plasmid recombination, pAsa5-3432 is the first reported plasmid of this bacterium that bears both an essential virulence factor (the type three secretion system) and multiple antibiotic resistance genes. As for pRAS3-3432, compared to the classic pRAS3, it carries a new mobile element that has only been identified in Chlamydia suis. Hence, with the identification of those two novel plasmids harboring mobile genetic elements that are normally encountered in other bacterial species, the present study puts emphasis on the important impact of mobile genetic elements in the genomic plasticity of A. salmonicida subsp. salmonicida and suggests that this aquatic bacterium could be an important reservoir of antibiotic resistance genes that can be exchanged with other bacteria, including human and animal pathogens. Copyright © 2019 Elsevier B.V. All rights reserved.


April 21, 2020  |  

Oenococcus sicerae sp. nov., isolated from French cider.

Two Gram-stain-positive, small ellipsoidal cocci, non-motile, oxidase- and catalase-negative, and facultative anaerobic strains (UCMA15228T and UCMA17102) were isolated in France, from fermented apple juices (ciders). The 16S rRNA gene sequence was identical between the two isolates and showed 97 % similarity with respect to the closest related species Oenococcus oeni and O. kitaharae. Therefore, the two isolates were classified within the genus Oenococcus. The phylogeny based on the pheS gene sequences also confirmed the position of the new taxon. DNA-DNA hybridizations based on in silico genome-to-genome comparisons (GGDC) and Average Nucleotide Identity (ANI) values, as well as species-specific PCR, validated the novelty of the taxon. Various phenotypic characteristics such as the optimum temperature and pH for growth, the ability to metabolise sugars, the aptitude to perform the malolactic fermentation, and the resistance to ethanol and NaCl, revealed that the two strains are distinguishable from the other members of the Oenococcus genus. The combined genotypic and phenotypic data support the classification of strains UCMA15228T and UCMA17102 into a novel species of Oenococcus, for which the name O. sicerae sp. nov. is proposed. The type strain is UCMA15228T (=DSM107163T=CIRM-BIA2288T).Copyright © 2018 Elsevier GmbH. All rights reserved.


April 21, 2020  |  

MSC: a metagenomic sequence classification algorithm.

Metagenomics is the study of genetic materials directly sampled from natural habitats. It has the potential to reveal previously hidden diversity of microscopic life largely due to the existence of highly parallel and low-cost next-generation sequencing technology. Conventional approaches align metagenomic reads onto known reference genomes to identify microbes in the sample. Since such a collection of reference genomes is very large, the approach often needs high-end computing machines with large memory which is not often available to researchers. Alternative approaches follow an alignment-free methodology where the presence of a microbe is predicted using the information about the unique k-mers present in the microbial genomes. However, such approaches suffer from high false positives due to trading off the value of k with the computational resources. In this article, we propose a highly efficient metagenomic sequence classification (MSC) algorithm that is a hybrid of both approaches. Instead of aligning reads to the full genomes, MSC aligns reads onto a set of carefully chosen, shorter and highly discriminating model sequences built from the unique k-mers of each of the reference sequences.Microbiome researchers are generally interested in two objectives of a taxonomic classifier: (i) to detect prevalence, i.e. the taxa present in a sample, and (ii) to estimate their relative abundances. MSC is primarily designed to detect prevalence and experimental results show that MSC is indeed a more effective and efficient algorithm compared to the other state-of-the-art algorithms in terms of accuracy, memory and runtime. Moreover, MSC outputs an approximate estimate of the abundances.The implementations are freely available for non-commercial purposes. They can be downloaded from https://drive.google.com/open?id=1XirkAamkQ3ltWvI1W1igYQFusp9DHtVl. © The Author(s) 2019. Published by Oxford University Press. All rights reserved. For Permissions, please e-mail: journals.permissions@oup.com.


April 21, 2020  |  

Carbohydrate catabolic capability of a Flavobacteriia bacterium isolated from hadal water.

Flavobacteriia are abundant in many marine environments including hadal waters, as demonstrated recently. However, it is unclear how this flavobacterial population adapts to hadal conditions. In this study, extensive comparative genomic analyses were performed for the flavobacterial strain Euzebyella marina RN62 isolated from the Mariana Trench hadal water in low abundance. The complete genome of RN62 possessed a considerable number of carbohydrate-active enzymes with a different composition. There was a predominance of GH family 13 proteins compared to closely related relatives, suggesting that RN62 has preserved a certain capacity for carbohydrate utilization and that the hadal ocean may hold an organic matter reservoir distinct from the surface ocean. Additionally, RN62 possessed potential intracellular cycling of the glycogen/starch pathway, which may serve as a strategy for carbon storage and consumption in response to nutrient pulse and starvation. Moreover, the discovery of higher glycoside hydrolase dissimilarities among Flavobacteriia, compared to peptidases and transporters, suggested variation in polysaccharide utilization related traits as an important ecophysiological factor in response to environmental alterations, such as decreased labile organic carbon in hadal waters. The presence of abundant toxin exporting, transcription and signal transduction related genes in RN62 may further help to survive in hadal conditions, including high pressure/low temperature.Copyright © 2019 Elsevier GmbH. All rights reserved.


April 21, 2020  |  

Iron-associated protein interaction networks reveal the key functional modules related to survival and virulence of Pasteurella multocida.

Pasteurella multocida causes respiratory infectious diseases in a multitude of birds and mammals. A number of virulence-associated genes were reported across different strains of P. multocida, including those involved in the iron transport and metabolism. Comparative iron-associated genes of P. multocida among different animal hosts towards their interaction networks have not been fully revealed. Therefore, this study aimed to identify the iron-associated genes from core- and pan-genomes of fourteen P. multocida strains and to construct iron-associated protein interaction networks using genome-scale network analysis which might be associated with the virulence. Results showed that these fourteen strains had 1587 genes in the core-genome and 3400 genes constituting their pan-genome. Out of these, 2651 genes associated with iron transport and metabolism were selected to construct the protein interaction networks and 361 genes were incorporated into the iron-associated protein interaction network (iPIN) consisting of nine different iron-associated functional modules. After comparing with the virulence factor database (VFDB), 21 virulence-associated proteins were determined and 11 of these belonged to the heme biosynthesis module. From this study, the core heme biosynthesis module and the core outer membrane hemoglobin receptor HgbA were proposed as candidate targets to design novel antibiotics and vaccines for preventing pasteurellosis across the serotypes or animal hosts for enhanced precision agriculture to ensure sustainability in food security. Copyright © 2018. Published by Elsevier Ltd.


April 21, 2020  |  

Deciphering bacterial epigenomes using modern sequencing technologies.

Prokaryotic DNA contains three types of methylation: N6-methyladenine, N4-methylcytosine and 5-methylcytosine. The lack of tools to analyse the frequency and distribution of methylated residues in bacterial genomes has prevented a full understanding of their functions. Now, advances in DNA sequencing technology, including single-molecule, real-time sequencing and nanopore-based sequencing, have provided new opportunities for systematic detection of all three forms of methylated DNA at a genome-wide scale and offer unprecedented opportunities for achieving a more complete understanding of bacterial epigenomes. Indeed, as the number of mapped bacterial methylomes approaches 2,000, increasing evidence supports roles for methylation in regulation of gene expression, virulence and pathogen-host interactions.


April 21, 2020  |  

Global-level population genomics reveals differential effects of geography and phylogeny on horizontal gene transfer in soil bacteria.

Although microorganisms are known to dominate Earth’s biospheres and drive biogeochemical cycling, little is known about the geographic distributions of microbial populations or the environmental factors that pattern those distributions. We used a global-level hierarchical sampling scheme to comprehensively characterize the evolutionary relationships and distributional limitations of the nitrogen-fixing bacterial symbionts of the crop chickpea, generating 1,027 draft whole-genome sequences at the level of bacterial populations, including 14 high-quality PacBio genomes from a phylogenetically representative subset. We find that diverse Mesorhizobium taxa perform symbiosis with chickpea and have largely overlapping global distributions. However, sampled locations cluster based on the phylogenetic diversity of Mesorhizobium populations, and diversity clusters correspond to edaphic and environmental factors, primarily soil type and latitude. Despite long-standing evolutionary divergence and geographic isolation, the diverse taxa observed to nodulate chickpea share a set of integrative conjugative elements (ICEs) that encode the major functions of the symbiosis. This symbiosis ICE takes 2 forms in the bacterial chromosome-tripartite and monopartite-with tripartite ICEs confined to a broadly distributed superspecies clade. The pairwise evolutionary relatedness of these elements is controlled as much by geographic distance as by the evolutionary relatedness of the background genome. In contrast, diversity in the broader gene content of Mesorhizobium genomes follows a tight linear relationship with core genome phylogenetic distance, with little detectable effect of geography. These results illustrate how geography and demography can operate differentially on the evolution of bacterial genomes and offer useful insights for the development of improved technologies for sustainable agriculture.


April 21, 2020  |  

Antimicrobial resistance-encoding plasmid clusters with heterogeneous MDR regions driven by IS26 in a single Escherichia coli isolate.

IS26-flanked transposons played an increasingly important part in the mobilization and development of resistance determinants. Heterogeneous resistance-encoding plasmid clusters with polymorphic MDR regions (MRRs) conferred by IS26 in an individual Escherichia coli isolate have not yet been detected.To characterize the complete sequence of a novel blaCTX-M-65- and fosA3-carrying IncZ-7 plasmid with dynamic MRRs from an E. coli isolate, and to depict the mechanism underlying the spread of resistance determinants and genetic polymorphisms.The molecular characterization of a strain carrying blaCTX-M-65 and fosA3 was analysed by antimicrobial susceptibility testing and MLST. The transferability of a plasmid bearing blaCTX-M-65 and fosA3 was determined by conjugation assays, and the complete structure of the plasmid was obtained by Illumina, PacBio and conventional PCR mapping, respectively. The circular forms derived from IS26-flanked transposons were detected by reverse PCR and sequencing.A novel IncZ-7 plasmid pEC013 (~118kb) harbouring the blaCTX-M-65 and fosA3 genes was recovered from E. coli isolate EC013 belonging to D-ST117. The plasmid was found to have heterogeneous and dynamic MRRs in an individual strain and the IS26-flanked composite transposon-derived circular intermediates were identified and characterized in pEC013.The heterogeneous MRRs suggested that a single plasmid may actually be a cluster of plasmids with the same backbone but varied MRRs, reflecting the plasmid’s heterogeneity and the survival benefits of having a response to antimicrobial-related threatening conditions in an individual strain. © The Author(s) 2019. Published by Oxford University Press on behalf of the British Society for Antimicrobial Chemotherapy. All rights reserved. For permissions, please email: journals.permissions@oup.com.


April 21, 2020  |  

Assessment of the microbial diversity of Chinese Tianshan tibicos by single molecule, real-time sequencing technology.

Chinese Tianshan tibico grains were collected from the rural area of Tianshan in Xinjiang province, China. Typical tibico grains are known to consist of polysaccharide matrix that embeds a variety of bacteria and yeasts. These grains are widely used in some rural regions to produce a beneficial sugary beverage that is slightly acidic and contains low level of alcohol. This work aimed to characterize the microbiota composition of Chinese Tianshan tibicos using the single molecule, real-time sequencing technology, which is advantageous in generating long reads. Our results revealed that the microbiota mainly comprised of the bacterial species of Lactobacillus hilgardii, Lactococcus raffinolactis, Leuconostoc mesenteroides, Zymomonas mobilis, together with a Guehomyces pullulans-dominating fungal community. The data generated in this work helps identify beneficial microbes in Chinese Tianshan tibico grains.


April 21, 2020  |  

Epidemiologic and genomic insights on mcr-1-harbouring Salmonella from diarrhoeal outpatients in Shanghai, China, 2006-2016.

Colistin resistance mediated by mcr-1-harbouring plasmids is an emerging threat in Enterobacteriaceae, like Salmonella. Based on its major contribution to the diarrhoea burden, the epidemic state and threat of mcr-1-harbouring Salmonella in community-acquired infections should be estimated.This retrospective study analysed the mcr-1 gene incidence in Salmonella strains collected from a surveillance on diarrhoeal outpatients in Shanghai Municipality, China, 2006-2016. Molecular characteristics of the mcr-1-positive strains and their plasmids were determined by genome sequencing. The transfer abilities of these plasmids were measured with various conjugation strains, species, and serotypes.Among the 12,053 Salmonella isolates, 37 mcr-1-harbouring strains, in which 35 were serovar Typhimurium, were detected first in 2012 and with increasing frequency after 2015. Most patients infected with mcr-1-harbouring strains were aged <5?years. All strains, including fluoroquinolone-resistant and/or extended-spectrum ß-lactamase-producing strains, were multi-drug resistant. S. Typhimurium had higher mcr-1 plasmid acquisition ability compared with other common serovars. Phylogeny based on the genomes combined with complete plasmid sequences revealed some clusters, suggesting the presence of mcr-1-harbouring Salmonella outbreaks in the community. Most mcr-1-positive strains were clustered together with the pork strains, strongly suggesting pork consumption as a main infection source.The mcr-1-harbouring Salmonella prevalence in community-acquired diarrhoea displays a rapid increase trend, and the ESBL-mcr-1-harbouring Salmonella poses a threat for children. These findings highlight the necessary and significance of prohibiting colistin use in animals and continuous monitoring of mcr-1-harbouring Salmonella.Copyright © 2019. Published by Elsevier B.V.


April 21, 2020  |  

A novel plasmid carrying carbapenem-resistant gene blaKPC-2 in Pseudomonas aeruginosa.

A carbapenem-resistant Pseudomonas aeruginosa strain PA1011 (ST463) was isolated from a patient in a surgical intensive care unit. PCR detection showed that PA1011 carried the blaKPC-2 gene. A plasmid was isolated and sequenced using the Illumina NextSeq 500 and PacBio RSII sequencing platforms. The plasmid was named pPA1011 and carried the carbapenem-resistant gene blaKPC-2. pPA1011 was a 62,793 bp in length with an average G+C content of 58.8%. It was identified as a novel plasmid and encoded a novel genetic environment of blaKPC-2 gene (?IS6-Tn3-ISKpn8-blaKPC-2-ISKpn6-IS26).


April 21, 2020  |  

PacBio sequencing reveals bacterial community diversity in cheeses collected from different regions.

Cheese is a fermented dairy product that is popular for its unique flavor and nutritional value. Recent studies have shown that microorganisms in cheese play an important role in the fermentation process and determine the quality of the cheese. We collected 12 cheese samples from different regions and studied the composition of their bacterial communities using PacBio small-molecule real-time sequencing (Pacific Biosciences, Menlo Park, CA). Our data revealed 144 bacterial genera (including Lactobacillus, Streptococcus, Lactococcus, and Staphylococcus) and 217 bacterial species (including Lactococcus lactis, Streptococcus thermophilus, Staphylococcus equorum, and Streptococcus uberis). We investigated the flavor quality of the cheese samples using an electronic nose system and we found differences in flavor-quality indices among samples from different regions. We found a clustering tendency based on flavor quality using principal component analysis. We found correlations between lactic acid bacteria and the flavor quality of the cheese samples. Biodegradation and metabolism of xenobiotics, and lipid-metabolism-related pathways, were predicted to contribute to differences in cheese flavor using Phylogenetic Investigation of Communities by Reconstruction of Unobserved States (PICRUSt). This preliminary study explored the bacterial communities in cheeses collected from different regions and their potential genome functions from the perspective of flavor quality.Copyright © 2020 American Dairy Science Association. Published by Elsevier Inc. All rights reserved.


April 21, 2020  |  

Metagenomic assembly through the lens of validation: recent advances in assessing and improving the quality of genomes assembled from metagenomes.

Metagenomic samples are snapshots of complex ecosystems at work. They comprise hundreds of known and unknown species, contain multiple strain variants and vary greatly within and across environments. Many microbes found in microbial communities are not easily grown in culture making their DNA sequence our only clue into their evolutionary history and biological function. Metagenomic assembly is a computational process aimed at reconstructing genes and genomes from metagenomic mixtures. Current methods have made significant strides in reconstructing DNA segments comprising operons, tandem gene arrays and syntenic blocks. Shorter, higher-throughput sequencing technologies have become the de facto standard in the field. Sequencers are now able to generate billions of short reads in only a few days. Multiple metagenomic assembly strategies, pipelines and assemblers have appeared in recent years. Owing to the inherent complexity of metagenome assembly, regardless of the assembly algorithm and sequencing method, metagenome assemblies contain errors. Recent developments in assembly validation tools have played a pivotal role in improving metagenomics assemblers. Here, we survey recent progress in the field of metagenomic assembly, provide an overview of key approaches for genomic and metagenomic assembly validation and demonstrate the insights that can be derived from assemblies through the use of assembly validation strategies. We also discuss the potential for impact of long-read technologies in metagenomics. We conclude with a discussion of future challenges and opportunities in the field of metagenomic assembly and validation. © The Author 2017. Published by Oxford University Press.


April 21, 2020  |  

Dysbiosis and Variation in Predicted Functions of the Granulation Tissue Microbiome in HPV Positive and Negative Severe Chronic Periodontitis.

Retrospective analysis has already shown correlation between severe Chronic Periodontitis (CP) cases with human papiloma virus (HPV). Hence, we aimed to explore deep-seated infected granulation tissue removed during periodontal flap surgery procedures for residential bacterial species between HPV+ and HVP- CP cases, which may serve as good predisposition marker for oral cancer. All CP-granulation samples showed the prominence of Firmicutes, Proteobacteria, and Bacteroidetes phyla with an abundance of gram negative anaerobes, except Streptococcus. In Beta diversity nonmetric multidimensional scaling plot, the random distribution of species was observed between HPV+ and HPV- CP granulation-samples. However, an abundance of Capnocytophaga ochracea was observed in HPV+ CP samples (p<0.05), while Porphyromonas endodontalis, Macellibacteroides fermentas, Treponema phagedenis, and Campylobacter rectus species were highly abundant in HPV- CP samples (p<0.05). The differential species richness leads altered functions related to mismatch-repair and nucleotide excision-repair and cytoskeleton-proteins. Hence, differential abundance of gram negative bacterial species between HPV+ and HPV- granulation-samples under anaerobic conditions may release virulence factors which may alter pathways favouring carcinogenesis. Hence, these species may serve as good predisposition marker for oral-cancer.


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