Menu

Asset Tag: Microbiology

April 21, 2020

Agricultural intensification reduces microbial network complexity and the abundance of keystone taxa in roots.

Root-associated microbes play a key role in plant performance and productivity, making them important players in agroecosystems. So far, very few studies have assessed the impact of different farming systems on the root microbiota and it is still unclear whether agricultural intensification influences the structure and complexity of microbial communities. We investigated the impact of conventional, no-till, and organic farming on wheat root fungal communities using PacBio SMRT sequencing on samples collected from 60 farmlands in Switzerland. Organic farming harbored a much more complex fungal network with significantly higher connectivity than conventional and no-till farming systems. The abundance of keystone taxa was the highest under organic farming where agricultural intensification was the lowest. We also found a strong negative association (R2?=?0.366; P?

Read more
April 21, 2020

High-Resolution Evolutionary Analysis of Within-Host Hepatitis C Virus Infection.

Despite recent breakthroughs in treatment of hepatitis C virus (HCV) infection, we have limited understanding of how virus diversity generated within individuals impacts the evolution and spread of HCV variants at the population scale. Addressing this gap is important for identifying the main sources of disease transmission and evaluating the risk of drug-resistance mutations emerging and disseminating in a population.We have undertaken a high-resolution analysis of HCV within-host evolution from 4 individuals coinfected with human immunodeficiency virus 1 (HIV-1). We used long-read, deep-sequenced data of full-length HCV envelope glycoprotein, longitudinally sampled from acute to chronic HCV infection to investigate the underlying viral population and evolutionary dynamics.We found statistical support for population structure maintaining the within-host HCV genetic diversity in 3 out of 4 individuals. We also report the first population genetic estimate of the within-host recombination rate for HCV (0.28 × 10-7 recombination/site/year), which is considerably lower than that estimated for HIV-1 and the overall nucleotide substitution rate estimated during HCV infection.Our findings indicate that population structure and strong genetic linkage shapes within-host HCV evolutionary dynamics. These results will guide the future investigation of potential HCV drug resistance adaptation during infection, and at the population scale. © The Author(s) 2019. Published by Oxford University Press for the Infectious Diseases Society of America.

Read more
April 21, 2020

Whole-Genome Analysis of Halomonas sp. Soap Lake #7 Reveals It Possesses Putative Mrp Antiporter Operon Groups 1 and 2.

The genus Halomonas possesses bacteria that are halophilic or halotolerant and exhibit a wide range of pH tolerance. The genome of Halomonas sp. Soap Lake #7 was sequenced to provide a better understanding of the mechanisms for salt and pH tolerance in this genus. The bacterium’s genome was found to possess two complete multiple resistance and pH antiporter systems, Group 1 and Group 2. This is the first report of both multiple resistance and pH antiporter Groups 1 and 2 in the genome of a haloalkaliphilic bacterium. © The Author(s) 2019. Published by Oxford University Press on behalf of the Society for Molecular Biology and Evolution.

Read more
April 21, 2020

Microbial community and fermentation characteristic of Italian ryegrass silage prepared with corn stover and lactic acid bacteria.

The bacterial community determined via PacBio single molecule, real-time sequencing technology (SMRT) and the fermentation characteristics of Italian ryegrass (IR, 82% moisture) silage prepared with corn stover (CS) were investigated. A selected strain of Lactobacillus plantarum (L694) and a commercial inoculant stain of Lactobacillus plantarum (LP) were used as additives. Lactic acid bacteria (LAB) effectively improved silage quality. After fermentation, Lactobacillus plantarum was the dominant species in IR?+?LP and IR?+?L694 treatments, which led to higher (P?

Read more
April 21, 2020

Genome Comparisons of Wild Isolates of Caulobacter crescentus Reveal Rates of Inversion and Horizontal Gene Transfer.

Since previous interspecies comparisons of Caulobacter genomes have revealed extensive genome rearrangements, we decided to compare the nucleotide sequences of four C. crescentus genomes, NA1000, CB1, CB2, and CB13. To accomplish this goal, we used PacBio sequencing technology to determine the nucleotide sequence of the CB1, CB2, and CB13 genomes, and obtained each genome sequence as a single contig. To correct for possible sequencing errors, each genome was sequenced twice. The only differences we observed between the two sets of independently determined sequences were random omissions of a single base in a small percentage of the homopolymer regions where a single base is repeated multiple times. Comparisons of these four genomes indicated that horizontal gene transfer events that included small numbers of genes occurred at frequencies in the range of 10-3 to 10-4 insertions per generation. Large insertions were about 100 times less frequent. Also, in contrast to previous interspecies comparisons, we found no genome rearrangements when the closely related NA1000, CB1, and CB2 genomes were compared, and only eight inversions and one translocation when the more distantly related CB13 genome was compared to the other genomes. Thus, we estimate that inversions occur at a rate of one per 10 to 12 million generations in Caulobacter genomes. The inversions seem to be complex events that include the simultaneous creation of indels.

Read more
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.

Read more
April 21, 2020

Whole-genome analysis of the colonization-resistant bacterium Phytobacter sp. SCO41T isolated from Bacillus nematocida B16-fed adult Caenorhabditis elegans.

Colonization resistance is an important attribute for bacterial interactions with hosts, but the mechanism is still not completely clear. In this study, we found that Phytobacter sp. SCO41T can effectively inhibit the in vivo colonization of Bacillus nematocida B16 in Caenorhabditis elegans, and we revealed the colonization resistance mechanism. Three strains of colonization-resistant bacteria, SCO41T, BX15, and BC7, were isolated from the intestines of the free-living nematode C. elegans derived from rotten fruit and soil. The primary characteristics and genome map of one of the three isolates was investigated to explore the underlying mechanism of colonization resistance in C. elegans. In addition, we performed exogenous iron supplementation and gene cluster knockout experiments to validate the sequencing results. The results showed that relationship was close among the three strains, which was identified as belonging to the genus Phytobacter. The type strain is SCO41T (=?CICC 24103T?=?KCTC 52362T). Whole genome analysis showed that csgA, csgB, csgC, csgE, csgF, and csgG were involved in the curli adhesive process and that fepA, fepB, fepC, fepD, and fepG played important roles in SCO41T against the colonization of B. nematocida B16 in C. elegans by competing for iron. Exogenous iron supplementation showed that exogenous iron can increase the colonization of B. nematocida B16, which was additionally confirmed by a deletion mutant strain. The csg gene family contributes to the colonization of SCO41T in C. elegans. Curli potentially contribute to the colonization of SCO41T in C. elegans, and enterobactin has a key role in SCO41T to resist the colonization of B. nematocida B16 by competing for iron.

Read more
April 21, 2020

Genetic characterization of an MDR/virulence genomic element carrying two T6SS gene clusters in a clinical Klebsiella pneumoniae isolate of swine origin.

Multiresistant Klebsiella pneumoniae isolates rarely cause infections in pigs. The aim of this study was to investigate a multiresistant porcine K. pneumoniae isolate for plasmidic and chromosomal antimicrobial resistance and virulence genes and their genetic environment.K. pneumoniae strain ZYST1 originated from a pig with pneumonia. Antimicrobial susceptibility testing was performed using broth microdilution. Conjugation experiments were conducted using Escherichia coli J53 as the recipient. The complete sequences of the chromosomal DNA and the plasmids were generated by WGS and analysed for the presence of resistance and virulence genes.The MDR K. pneumoniae ST1 strain ZYST1 contained three plasmids belonging to incompatibility groups IncFIIk5-FIB, IncI1 and IncX4, respectively. The IncFIIk5-FIB plasmid carried the resistance genes aadA2, mph(A), sul1 and aph(3′)-Ia, and the IncI1 plasmid carried aadA22 and erm(B). No resistance genes were present on the IncX4 plasmid. Plasmids related to the aforementioned three plasmids were also present in other Enterobacteriaceae species from humans, animals and the environment. Bioinformatic analyses identified a chromosomal 904?kb MDR element flanked by two copies of ISKpn26. This element included virulence factors, such as a type VI secretion system (T6SS) and genes for type 1 fimbriae, the toxin-antitoxin system HipA/HipB, antimicrobial resistance genes, such as blaSHV-187, mdtk, catA and the multiple antibiotic resistance operon marRABC, and heavy metal resistance determinants, such as chrB/chrA and tehA/tehB.This study reports a novel 904?kb MDR/virulence genomic element and three important plasmids coexisting in a clinical K. pneumoniae isolate of animal origin. © 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.

Read more
April 21, 2020

Co-circulation of Multidrug-resistant Shigella Among Men Who Have Sex With Men in Australia.

In urban Australia, the burden of shigellosis is either in returning travelers from shigellosis-endemic regions or in men who have sex with men (MSM). Here, we combine genomic data with comprehensive epidemiological data on sexual exposure and travel to describe the spread of multidrug-resistant Shigella lineages.A population-level study of all cultured Shigella isolates in the state of Victoria, Australia, was undertaken from 1 January 2016 through 31 March 2018. Antimicrobial susceptibility testing, whole-genome sequencing, and bioinformatic analyses of 545 Shigella isolates were performed at the Microbiological Diagnostic Unit Public Health Laboratory. Risk factor data on travel and sexual exposure were collected through enhanced surveillance forms or by interviews.Rates of antimicrobial resistance were high, with 17.6% (95/541) and 50.6% (274/541) resistance to ciprofloxacin and azithromycin, respectively. There were strong associations between antimicrobial resistance, phylogeny, and epidemiology. Specifically, 2 major MSM-associated lineages were identified: a Shigellasonnei lineage (n = 159) and a Shigella flexneri 2a lineage (n = 105). Of concern, 147/159 (92.4%) of isolates within the S. sonnei MSM-associated lineage harbored mutations associated with reduced susceptibility to recommended oral antimicrobials: namely, azithromycin, trimethoprim-sulfamethoxazole, and ciprofloxacin. Long-read sequencing demonstrated global dissemination of multidrug-resistant plasmids across Shigella species and lineages, but predominantly associated with MSM isolates.Our contemporary data highlight the ongoing public health threat posed by resistant Shigella, both in Australia and globally. Urgent multidisciplinary public health measures are required to interrupt transmission and prevent infection. © The Author(s) 2019. Published by Oxford University Press for the Infectious Diseases Society of America. All rights reserved. For permissions, e-mail: journals.permissions@oup.com.

Read more
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.

Read more
April 21, 2020

Molecular Epidemiology of Candida auris in Colombia Reveals a Highly Related, Countrywide Colonization With Regional Patterns in Amphotericin B Resistance.

Candida auris is a multidrug-resistant yeast associated with hospital outbreaks worldwide. During 2015-2016, multiple outbreaks were reported in Colombia. We aimed to understand the extent of contamination in healthcare settings and to characterize the molecular epidemiology of C. auris in Colombia.We sampled patients, patient contacts, healthcare workers, and the environment in 4 hospitals with recent C. auris outbreaks. Using standardized protocols, people were swabbed at different body sites. Patient and procedure rooms were sectioned into 4 zones and surfaces were swabbed. We performed whole-genome sequencing (WGS) and antifungal susceptibility testing (AFST) on all isolates.Seven of the 17 (41%) people swabbed were found to be colonized. Candida auris was isolated from 37 of 322 (11%) environmental samples. These were collected from a variety of items in all 4 zones. WGS and AFST revealed that although isolates were similar throughout the country, isolates from the northern region were genetically distinct and more resistant to amphotericin B (AmB) than the isolates from central Colombia. Four novel nonsynonymous mutations were found to be significantly associated with AmB resistance.Our results show that extensive C. auris contamination can occur and highlight the importance of adherence to appropriate infection control practices and disinfection strategies. Observed genetic diversity supports healthcare transmission and a recent expansion of C. auris within Colombia with divergent AmB susceptibility.

Read more
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.

Read more
April 21, 2020

Development of Tigecycline Resistance in Carbapenemase-Producing Klebsiella pneumoniae Sequence Type 147 via AcrAB Overproduction Mediated by Replacement of the ramA Promoter.

Carbapenem-resistant K. pneumoniae 2297, isolated from a patient treated with tigecycline for pneumonia, developed tigecycline resistance, in contrast to carbapenem-resistant isolate 1215, which was collected four months prior to the 2297 isolate. Mechanisms underlying tigecycline resistance were elucidated for the clinical isolates.The tigecycline minimum inhibitory concentration (MIC) was determined using the broth microdilution method, with or without phenylalanine-arginine ß-naphthylamide (PABN), and whole-genome sequencing was carried out by single-molecule real-time sequencing. The expression levels of the genes acrA, oqxA, ramA, rarA, and rpoB were determined by reverse-transcription quantitative PCR.Both isolates presented identical antibiograms, except for tigecycline, which showed an MIC of 0.5 mg/L in 1215 and 2 mg/L in 2297. The addition of PABN to tigecycline-resistant 2297 caused a four-fold decrease in the tigecycline MIC to 0.5 mg/L, although acrA expression (encoding the AcrAB efflux pump) was upregulated by 2.5 fold and ramA expression (encoding the pump activator RamA) was upregulated by 1.4 fold. We identified a 6,096-bp fragment insertion flanking direct TATAT repeats that disrupted the romA gene located upstream of ramA in the chromosome of K. pneumoniae 2297; the insertion led the ramA gene promoter replacement resulting in stronger activation of the gene.The K. pneumoniae isolate developed tigecycline resistance during tigecycline treatment. It was related to the overexpression of the AcrAB resistance-nodulation-cell division efflux system due to promoter replacement. © The Korean Society for Laboratory Medicine.

Read more
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.

Read more
April 21, 2020

Identification of the novel class D ß-lactamase OXA-679 involved in carbapenem resistance in Acinetobacter calcoaceticus.

The aim of this study was to characterize the Acinetobacter calcoaceticus clinical isolate AC_2117 with the novel carbapenem-hydrolysing class D ß-lactamase (CHDL) OXA-679.Identification of the species and ß-lactamases was verified by genome sequencing (PacBio) and phylogenetic analyses. Antibiotic susceptibility of AC_2117 and transformants harbouring cloned blaOXA-679 was evaluated using antibiotic gradient strips and microbroth dilution. OXA-679 was purified heterologously and kinetic parameters were determined using spectrometry or isothermal titration calorimetry. The impact of OXA-679 production during imipenem therapy was evaluated in the Galleria mellonella infection model.Sequencing of the complete genome of the clinical A. calcoaceticus isolate AC_2117 identified a novel CHDL, termed OXA-679. This enzyme shared sequence similarity of 71% to each of the families OXA-143 and OXA-24/40. Phylogenetic analyses revealed that OXA-679 represents a member of a new OXA family. Cloning and expression of blaOXA-679 as well as measurement of kinetic parameters revealed the effective hydrolysis of carbapenems which resulted in reduced susceptibility to carbapenems in Escherichia coli and A. calcoaceticus, and high-level carbapenem resistance in Acinetobacter baumannii. Infection of larvae of G. mellonella with a sublethal dose of blaOXA-679-expressing A. baumannii could not be cured by high-dose imipenem therapy, indicating carbapenem resistance in vivo.We identified blaOXA-679 in a clinical A. calcoaceticus isolate that represents a member of the new OXA-679 family and that conferred high-level carbapenem resistance in vitro and in vivo. © 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.

Read more

Subscribe for blog updates:

Talk with an expert

If you have a question, need to check the status of an order, or are interested in purchasing an instrument, we're here to help.