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April 21, 2020  |  

Biphasic cellular adaptations and ecological implications of Alteromonas macleodii degrading a mixture of algal polysaccharides.

Algal polysaccharides are an important bacterial nutrient source and central component of marine food webs. However, cellular and ecological aspects concerning the bacterial degradation of polysaccharide mixtures, as presumably abundant in natural habitats, are poorly understood. Here, we contextualize marine polysaccharide mixtures and their bacterial utilization in several ways using the model bacterium Alteromonas macleodii 83-1, which can degrade multiple algal polysaccharides and contributes to polysaccharide degradation in the oceans. Transcriptomic, proteomic and exometabolomic profiling revealed cellular adaptations of A. macleodii 83-1 when degrading a mix of laminarin, alginate and pectin. Strain 83-1 exhibited substrate prioritization driven by catabolite repression, with initial laminarin utilization followed by simultaneous alginate/pectin utilization. This biphasic phenotype coincided with pronounced shifts in gene expression, protein abundance and metabolite secretion, mainly involving CAZymes/polysaccharide utilization loci but also other functional traits. Distinct temporal changes in exometabolome composition, including the alginate/pectin-specific secretion of pyrroloquinoline quinone, suggest that substrate-dependent adaptations influence chemical interactions within the community. The ecological relevance of cellular adaptations was underlined by molecular evidence that common marine macroalgae, in particular Saccharina and Fucus, release mixtures of alginate and pectin-like rhamnogalacturonan. Moreover, CAZyme microdiversity and the genomic predisposition towards polysaccharide mixtures among Alteromonas spp. suggest polysaccharide-related traits as an ecophysiological factor, potentially relating to distinct ‘carbohydrate utilization types’ with different ecological strategies. Considering the substantial primary productivity of algae on global scales, these insights contribute to the understanding of bacteria-algae interactions and the remineralization of chemically diverse polysaccharide pools, a key step in marine carbon cycling.


April 21, 2020  |  

Emergence of a ST2570 Klebsiella pneumoniae isolate carrying mcr-1 and blaCTX-M-14 recovered from a bloodstream infection in China.

The worldwide emergence of the plasmid-borne colistin resistance mediated by mcr-1 gene not only extended our knowledge on colistin resistance, but also poses a serious threat to clinical and public health [1, 2]. Since its first discovery, mcr-1-carrying Enterobacteriaceae from human, animal, food, and environmental origins have been widely identified, but few mcr-1-positive clinical strains of Klebsiella pneumoniae have been reported so far, especially when associated with community-acquired infections [3, 4]. Here, we report the emergence of a colistin-resistant K. pneumoniae isolate, which belonged to a rare sporadic clone, co-carrying mcr-1 and blaCTX-M-14 genes simultaneous recovered from a community-acquired bloodstream infection in China. Whole-genome sequencing and microbiological analysis were performed to elucidate its antimicrobial resistance mechanisms.


April 21, 2020  |  

Whole genome sequencing of NDM-1-producing serotype K1 ST23 hypervirulent Klebsiella pneumoniae in China.

The emergence and spread of carbapenem-resistant hypervirulent Klebsiella pneumoniae (CR-hvKP) is causing worldwide concern, whereas NDM-producing hvKP is still rare. Here we report the complete genome sequence characteristics of an NDM-1-producing ST23 type clinical hvKP in PR China.Capsular polysaccharide serotyping was performed by PCR. The complete genome sequence of isolate 3214 was obtained using both the Illumina Hiseq platform and Pacbio RS platform. Multilocus sequence type was identified by submitting the genome sequence to mlst 2.0 and the antimicrobial resistance genes and plasmid replicons were identified using ResFinder and PlasmidFinder, respectively. Transferability of the blaNDM-1-bearing plasmid was determined by conjugation experiment, S1 pulsed-field gel electrophoresis and Southern hybridization.Isolate 3214 was classified to ST23 and belonged to the K1 capsular serotype. The isolate’s total genome size was 6 171 644?bp with a G+C content of 56.39 %, consisting of a 5 448 209?bp chromosome and seven plasmids. The resistome included 18 types of antibiotic resistance genes. Fourteen resistance genes including blaNDM-1 and blaCTX-M-14 were located on plasmids and five also including blaCTX-M-14 were in the chromosome. Plasmid pNDM_3214 carrying blaNDM-1 harboured six types of resistance genes surrounded by insertion sequences and was conjugative. The worldwide pLVPK-like virulence plasmid harbouring rmpA2 and rmpA was also found in this isolate.This study provides basic information of phenotypic and genomic features of ST23 CR-hvKP isolate 3214. Our data highlights the potential risk of spread of NDM-1-producing ST23 hvKP.


April 21, 2020  |  

Genomic Survey of Bordetella pertussis Diversity, United States, 2000-2013.

We characterized 170 complete genome assemblies from clinical Bordetella pertussis isolates representing geographic and temporal diversity in the United States. These data capture genotypic shifts, including increased pertactin deficiency, occurring amid the current pertussis disease resurgence and provide a foundation for needed research to direct future public health control strategies.


April 21, 2020  |  

Potential KPC-2 carbapenemase reservoir of environmental Aeromonas hydrophila and Aeromonas caviae isolates from the effluent of an urban wastewater treatment plant in Japan.

Aeromonas hydrophila and Aeromonas caviae adapt to saline water environments and are the most predominant Aeromonas species isolated from estuaries. Here, we isolated antimicrobial-resistant (AMR) Aeromonas strains (A. hydrophila GSH8-2 and A. caviae GSH8M-1) carrying the carabapenemase blaKPC-2 gene from a wastewater treatment plant (WWTP) effluent in Tokyo Bay (Japan) and determined their complete genome sequences. GSH8-2 and GSH8M-1 were classified as newly assigned sequence types ST558 and ST13, suggesting no supportive evidence of clonal dissemination. The strains appear to have acquired blaKPC-2 -positive IncP-6-relative plasmids (pGSH8-2 and pGSH8M-1-2) that share a common backbone with plasmids in Aeromonas sp. ASNIH3 isolated from hospital wastewater in the United States, A. hydrophila WCHAH045096 isolated from sewage in China, other clinical isolates (Klebsiella, Enterobacter and Escherichia coli), and wastewater isolates (Citrobacter, Pseudomonas and other Aeromonas spp.). In addition to blaKPC-2 , pGSH8M-1-2 carries an IS26-mediated composite transposon including a macrolide resistance gene, mph(A). Although Aeromonas species are opportunistic pathogens, they could serve as potential environmental reservoir bacteria for carbapenemase and AMR genes. AMR monitoring from WWTP effluents will contribute to the detection of ongoing AMR dissemination in the environment and might provide an early warning of potential dissemination in clinical settings and communities. © 2019 The Authors. Environmental Microbiology Reports published by Society for Applied Microbiology and John Wiley & Sons Ltd.


April 21, 2020  |  

Genomic analysis provides insights into the transmission and pathogenicity of Talaromyces marneffei.

Talaromyces marneffei (T. marneffei) is a medically important opportunistic dimorphic fungus that infects both humans and bamboo rats. However, the mechanisms of transmission and pathogenicity of T. marneffei are poorly understood. In our study, we combined Illumina and PacBio sequencing technologies to sequence and assemble a complete genome of T. marneffei. To elucidate the transmission route and source, we sequenced three additional T. marneffei isolates using Illumina sequencing technology. Variations among isolates were used to develop a multilocus sequence typing (MLST) system comprising five housekeeping genes that can be used to discriminate between isolates derived from different sources. Our analysis revealed that human and bamboo rat share identical genotypes in these five loci. Thus, we hypothesized that T. marneffei is transmitted to humans through inhalation of spores in the surrounding environment into the lungs and that the bamboo rat can serve as an important natural reservoir for pathogens. Furthermore, we also identified temperature-dependent polyketide synthases, non-ribosomal peptide synthetases and secreted proteins as putative pathogenicity-related factors. In addition, we identified antifungal drug targets that can be investigated in future studies to elucidate the mechanisms underlying drug resistance. In summary, our study presents the basic features of the T. marneffei genome and provides insights into the transmission and pathogenicity of T. marneffei, which warrant fundamental experimental research.Copyright © 2019 Elsevier Inc. All rights reserved.


April 21, 2020  |  

Transmission of ESBL-producing Escherichia coli between broilers and humans on broiler farms.

ESBL and AmpC ß-lactamases are an increasing concern for public health. Studies suggest that ESBL/pAmpC-producing Escherichia coli and their plasmids carrying antibiotic resistance genes can spread from broilers to humans working or living on broiler farms. These studies used traditional typing methods, which may not have provided sufficient resolution to reliably assess the relatedness of these isolates.Eleven suspected transmission events among broilers and humans living/working on eight broiler farms were investigated using whole-genome short-read (Illumina) and long-read sequencing (PacBio). Core genome MLST (cgMLST) was performed to investigate the occurrence of strain transmission. Horizontal plasmid and gene transfer were analysed using BLAST.Of eight suspected strain transmission events, six were confirmed. The isolate pairs had identical ESBL/AmpC genes and fewer than eight allelic differences according to the cgMLST, and five had an almost identical plasmid composition. On one of the farms, cgMLST revealed that the isolate pairs belonging to ST10 from a broiler and a household member of the farmer had 475 different alleles, but that the plasmids were identical, indicating horizontal transfer of mobile elements rather than strain transfer. Of three suspected horizontal plasmid transmission events, one was confirmed. In addition, gene transfer between plasmids was found.The present study confirms transmission of strains as well as horizontal plasmid and gene transfer between broilers and farmers and household members on the same farm. WGS is an important tool to confirm suspected zoonotic strain and resistance gene transmission. © 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  |  

Diversity of phytobeneficial traits revealed by whole-genome analysis of worldwide-isolated phenazine-producing Pseudomonas spp.

Plant-beneficial Pseudomonas spp. competitively colonize the rhizosphere and display plant-growth promotion and/or disease-suppression activities. Some strains within the P. fluorescens species complex produce phenazine derivatives, such as phenazine-1-carboxylic acid. These antimicrobial compounds are broadly inhibitory to numerous soil-dwelling plant pathogens and play a role in the ecological competence of phenazine-producing Pseudomonas spp. We assembled a collection encompassing 63 strains representative of the worldwide diversity of plant-beneficial phenazine-producing Pseudomonas spp. In this study, we report the sequencing of 58 complete genomes using PacBio RS II sequencing technology. Distributed among four subgroups within the P. fluorescens species complex, the diversity of our collection is reflected by the large pangenome which accounts for 25 413 protein-coding genes. We identified genes and clusters encoding for numerous phytobeneficial traits, including antibiotics, siderophores and cyclic lipopeptides biosynthesis, some of which were previously unknown in these microorganisms. Finally, we gained insight into the evolutionary history of the phenazine biosynthetic operon. Given its diverse genomic context, it is likely that this operon was relocated several times during Pseudomonas evolution. Our findings acknowledge the tremendous diversity of plant-beneficial phenazine-producing Pseudomonas spp., paving the way for comparative analyses to identify new genetic determinants involved in biocontrol, plant-growth promotion and rhizosphere competence. © 2018 Society for Applied Microbiology and John Wiley & Sons Ltd.


April 21, 2020  |  

Investigating the bacterial microbiota of traditional fermented dairy products using propidium monoazide with single-molecule real-time sequencing.

Traditional fermented dairy foods have been the major components of the Mongolian diet for millennia. In this study, we used propidium monoazide (PMA; binds to DNA of nonviable cells so that only viable cells are enumerated) and single-molecule real-time sequencing (SMRT) technology to investigate the total and viable bacterial compositions of 19 traditional fermented dairy foods, including koumiss from Inner Mongolia (KIM), koumiss from Mongolia (KM), and fermented cow milk from Mongolia (CM); sample groups treated with PMA were designated PKIM, PKM, and PCM. Full-length 16S rRNA sequencing identified 195 bacterial species in 121 genera and 13 phyla in PMA-treated and untreated samples. The PMA-treated and untreated samples differed significantly in their bacterial community composition and a-diversity values. The predominant species in KM, KIM, and CM were Lactobacillus helveticus, Streptococcus parauberis, and Lactobacillus delbrueckii, whereas the predominant species in PKM, PKIM, and PCM were Enterobacter xiangfangensis, Lactobacillus helveticus, and E. xiangfangensis, respectively. Weighted and unweighted principal coordinate analyses showed a clear clustering pattern with good separation and only minor overlapping. In addition, a pure culture method was performed to obtain lactic acid bacteria resources in dairy samples according to the results of SMRT sequencing. A total of 102 LAB strains were identified and Lb. helveticus (68.63%) was the most abundant, in agreement with SMRT sequencing results. Our results revealed that the bacterial communities of traditional dairy foods are complex and vary by type of fermented dairy product. The PMA treatment induced significant changes in bacterial community structure.Copyright © 2019 American Dairy Science Association. Published by Elsevier Inc. 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  |  

A systematic review of the Trypanosoma cruzi genetic heterogeneity, host immune response and genetic factors as plausible drivers of chronic chagasic cardiomyopathy.

Chagas disease is a complex tropical pathology caused by the kinetoplastid Trypanosoma cruzi. This parasite displays massive genetic diversity and has been classified by international consensus in at least six Discrete Typing Units (DTUs) that are broadly distributed in the American continent. The main clinical manifestation of the disease is the chronic chagasic cardiomyopathy (CCC) that is lethal in the infected individuals. However, one intriguing feature is that only 30-40% of the infected individuals will develop CCC. Some authors have suggested that the immune response, host genetic factors, virulence factors and even the massive genetic heterogeneity of T. cruzi are responsible of this clinical pattern. To date, no conclusive data support the reason why a few percentages of the infected individuals will develop CCC. Therefore, we decided to conduct a systematic review analysing the host genetic factors, immune response, cytokine production, virulence factors and the plausible association of the parasite DTUs and CCC. The epidemiological and clinical implications are herein discussed.


April 21, 2020  |  

Characterization of NDM-5- and CTX-M-55-coproducing Escherichia coli GSH8M-2 isolated from the effluent of a wastewater treatment plant in Tokyo Bay.

New Delhi metallo-ß-lactamase (NDM)-5-producing Enterobacteriaceae have been detected in rivers, sewage, and effluents from wastewater treatment plants (WWTPs). Environmental contamination due to discharged effluents is of particular concern as NDM variants may be released into waterways, thereby posing a risk to humans. In this study, we collected effluent samples from a WWTP discharged into a canal in Tokyo Bay, Japan.Testing included the complete genome sequencing of Escherichia coli GSH8M-2 isolated from the effluent as well as a gene network analysis.The complete genome sequencing of GSH8M-2 revealed that it was an NDM-5-producing E. coli strain sequence type ST542, which carries multiple antimicrobial resistance genes for ß-lactams, quinolone, tetracycline, trimethoprim-sulfamethoxazole, florfenicol/chloramphenicol, kanamycin, and fosfomycin. The blaNDM-5 gene was found in the IncX3 replicon plasmid pGSH8M-2-4. Gene network analysis using 142 IncX3 plasmid sequences suggested that pGSH8M-2-4 is related to both clinical isolates of  E. coli and Klebsiella species in Eastern Asia. GSH8M-2 also carries the blaCTX-M-55 gene in IncX1 plasmid pGSH8M-2-3.This is the first report of environmental NDM-5-producing E. coli isolated from a WWTP in Japan. NDM-5 detection is markedly increasing in veterinary and clinical settings, suggesting that dual ß-lactamases, such as NDM-5 and CTX-M-55, might be acquired through multiple steps in environment settings. Environmental contamination through WWTP effluents that contain producers of NDM variants could be an emerging potential health hazard. Thus, regular monitoring of WWTP effluents is important for the detection of antimicrobial-resistant bacteria that may be released into the waterways and nearby communities.


April 21, 2020  |  

Genome characterization of an extensively drug-resistant Streptococcus pneumoniae serotype 11A strain.

In this study, the whole genome sequences of two Streptococcus pneumoniae clinical isolates from South Korea were determined and compared. They were found to be the same serotype (11?A) and multilocus sequence typing analysis showed that they are single-locus variants (SLVs; ST8279 and ST166) of each other, differing at one allele (aroE). However, the ST8279 strain is extensively drug-resistant (XDR) whereas the ST166 strain is not. The genome of the XDR strain is very similar in structure to that of two previously reported genomes, AP200 (11?A:ST62) and 70585 (5:ST5803); however, some regions were inverted and there were some exogenous regions in the ST8279 strain. It was found that 6,502 single nucleotide polymorphisms are dispersed across the genome between the two serotype 11?A ST8279 and ST166 strains. Many of them are located in genes associated with antibiotic resistance. In addition, many amino acid differences were also identified in genes involved in DNA repair (mutL, uvrA and uvrC) and recombination (recU, recR and recA). On the basis of these results, it was inferred that the XDR strain did not evolve from its SLV via a single recombination event involving a large portion of the genome including the aroE gene. Rather, the strain likely evolved through many point mutations and recombination events involving small portions of the genome. © 2019 The Societies and John Wiley & Sons Australia, Ltd.


April 21, 2020  |  

WGS of 1058 Enterococcus faecium from Copenhagen, Denmark, reveals rapid clonal expansion of vancomycin-resistant clone ST80 combined with widespread dissemination of a vanA-containing plasmid and acquisition of a heterogeneous accessory genome.

From 2012 to 2015, a sudden significant increase in vancomycin-resistant (vanA) Enterococcus faecium (VREfm) was observed in the Capital Region of Denmark. Clonal relatedness of VREfm and vancomycin-susceptible E. faecium (VSEfm) was investigated, transmission events between hospitals were identified and the pan-genome and plasmids from the largest VREfm clonal group were characterized.WGS of 1058 E. faecium isolates was carried out on the Illumina platform to perform SNP analysis and to identify the pan-genome. One isolate was also sequenced on the PacBio platform to close the genome. Epidemiological data were collected from laboratory information systems.Phylogeny of 892 VREfm and 166 VSEfm revealed a polyclonal structure, with a single clonal group (ST80) accounting for 40% of the VREfm isolates. VREfm and VSEfm co-occurred within many clonal groups; however, no VSEfm were related to the dominant VREfm group. A similar vanA plasmid was identified in =99% of isolates belonging to the dominant group and 69% of the remaining VREfm. Ten plasmids were identified in the completed genome, and ~29% of this genome consisted of dispensable accessory genes. The size of the pan-genome among isolates in the dominant group was 5905 genes.Most probably, VREfm emerged owing to importation of a successful VREfm clone which rapidly transmitted to the majority of hospitals in the region whilst simultaneously disseminating a vanA plasmid to pre-existing VSEfm. Acquisition of a heterogeneous accessory genome may account for the success of this clone by facilitating adaptation to new environmental challenges. © 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  |  

The complete genome of the antifungal bacterium Pseudomonas sp. strain MS82

The genomic sequence of Pseudomonas sp. strain MS82 isolated from the rhizosphere of a soybean plant is reported and analyzed in relation to its extensive antifungal activity. Broth media used for production of the antifungal extract from strain MS82 against the mushroom pathogen Trichoderma viride were optimized using the routine plate bioassays. Culture extract of strain 82 in the peptone-K2HPO4-MgSO4 medium (PKM; peptone 20 g/L, K2HPO4 1.5 g/L, MgSO4 1.5 g/L and sterilized water) showed the best antifungal activity with an inhibition rate of 88.69thinspacetextpmthinspace3.87% to the fungal pathogen. Control efficacy of the T. viride contamination was investigated in mushroom production compost. The disease severity index of P. ostreatus hyphae infected by T. viride of treatment mixed with MS82 supernatant (38.33thinspacetextpmthinspace5.20%) was lower than that of the compost mixed with non-inoculated broth (97.50thinspacetextpmthinspace2.50%). The multilocus sequence analysis, containing four partial sequences from the gyrB, rpoB, recA and rpoD, suggests that strain MS82 is a Pseudomonas strain. The strain MS82 genome consists of a circular chromosome of 6,207,556 bp that was predicted to encode 5401 proteins and 131 RNA genes. Genome analysis revealed the presence of the gene clusters for biosynthesis of antifungal compounds, such as phenazine, pyocyanin, pyoverdine, volatile HCN and cyclic lipopeptides (arthrofactin). Genome analysis presented in the report will provide insights into development of biological control for fungal contamination in mushroom cultivation.


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