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July 7, 2019

Evolutionary dynamics and genomic features of the Elizabethkingia anophelis 2015 to 2016 Wisconsin outbreak strain.

An atypically large outbreak of Elizabethkingia anophelis infections occurred in Wisconsin. Here we show that it was caused by a single strain with thirteen characteristic genomic regions. Strikingly, the outbreak isolates show an accelerated evolutionary rate and an atypical mutational spectrum. Six phylogenetic sub-clusters with distinctive temporal and geographic dynamics are revealed, and their last common ancestor existed approximately one year before the first recognized human infection. Unlike other E. anophelis, the outbreak strain had a disrupted DNA repair mutY gene caused by insertion of an integrative and conjugative element. This genomic change probably contributed to the high evolutionary rate of the outbreak strain and may have increased its adaptability, as many mutations in protein-coding genes occurred during the outbreak. This unique discovery of an outbreak caused by a naturally occurring mutator bacterial pathogen provides a dramatic example of the potential impact of pathogen evolutionary dynamics on infectious disease epidemiology.

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July 7, 2019

Population genomic analysis of 1,777 extended-spectrum beta-lactamase-producing Klebsiella pneumoniae isolates, Houston, Texas: unexpected abundance of clonal group 307.

Klebsiella pneumoniae is a major human pathogen responsible for high morbidity and mortality rates. The emergence and spread of strains resistant to multiple antimicrobial agents and documented large nosocomial outbreaks are especially concerning. To develop new therapeutic strategies for K. pneumoniae, it is imperative to understand the population genomic structure of strains causing human infections. To address this knowledge gap, we sequenced the genomes of 1,777 extended-spectrum beta-lactamase-producing K. pneumoniae strains cultured from patients in the 2,000-bed Houston Methodist Hospital system between September 2011 and May 2015, representing a comprehensive, population-based strain sample. Strains of largely uncharacterized clonal group 307 (CG307) caused more infections than those of well-studied epidemic CG258. Strains varied markedly in gene content and had an extensive array of small and very large plasmids, often containing antimicrobial resistance genes. Some patients with multiple strains cultured over time were infected with genetically distinct clones. We identified 15 strains expressing the New Delhi metallo-beta-lactamase 1 (NDM-1) enzyme that confers broad resistance to nearly all beta-lactam antibiotics. Transcriptome sequencing analysis of 10 phylogenetically diverse strains showed that the global transcriptome of each strain was unique and highly variable. Experimental mouse infection provided new information about immunological parameters of host-pathogen interaction. We exploited the large data set to develop whole-genome sequence-based classifiers that accurately predict clinical antimicrobial resistance for 12 of the 16 antibiotics tested. We conclude that analysis of large, comprehensive, population-based strain samples can assist understanding of the molecular diversity of these organisms and contribute to enhanced translational research. IMPORTANCEKlebsiella pneumoniae causes human infections that are increasingly difficult to treat because many strains are resistant to multiple antibiotics. Clonal group 258 (CG258) organisms have caused outbreaks in health care settings worldwide. Using a comprehensive population-based sample of extended-spectrum beta-lactamase (ESBL)-producing K. pneumoniae strains, we show that a relatively uncommon clonal type, CG307, caused the plurality of ESBL-producing K. pneumoniae infections in our patients. We discovered that CG307 strains have been abundant in Houston for many years. As assessed by experimental mouse infection, CG307 strains were as virulent as pandemic CG258 strains. Our results may portend the emergence of an especially successful clonal group of antibiotic-resistant K. pneumoniae. Copyright © 2017 Long et al.

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July 7, 2019

The evolutionary life cycle of the polysaccharide biosynthetic gene cluster based on the Sphingomonadaceae.

Although clustering of genes from the same metabolic pathway is a widespread phenomenon, the evolution of the polysaccharide biosynthetic gene cluster remains poorly understood. To determine the evolution of this pathway, we identified a scattered production pathway of the polysaccharide sanxan by Sphingomonas sanxanigenens NX02, and compared the distribution of genes between sphingan-producing and other Sphingomonadaceae strains. This allowed us to determine how the scattered sanxan pathway developed, and how the polysaccharide gene cluster evolved. Our findings suggested that the evolution of microbial polysaccharide biosynthesis gene clusters is a lengthy cyclic process comprising cluster 1???scatter???cluster 2. The sanxan biosynthetic pathway proved the existence of a dispersive process. We also report the complete genome sequence of NX02, in which we identified many unstable genetic elements and powerful secretion systems. Furthermore, nine enzymes for the formation of activated precursors, four glycosyltransferases, four acyltransferases, and four polymerization and export proteins were identified. These genes were scattered in the NX02 genome, and the positive regulator SpnA of sphingans synthesis could not regulate sanxan production. Finally, we concluded that the evolution of the sanxan pathway was independent. NX02 evolved naturally as a polysaccharide producing strain over a long-time evolution involving gene acquisitions and adaptive mutations.

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July 7, 2019

Complete genome sequence of the aerobically denitrifying thermophilic bacterium Chelatococcus daeguensis TAD1

Chelatococcus daeguensis TAD1 is a themophilic bacterium isolated from a biotrickling filter used to treat NOx in Ruiming Power Plant, located in Guangzhou, China, which shows an excellent aerobic denitrification activity at high temperature. The complete genome sequence of this strain was reported in the present study. Genes related to the aerobic denitrification were identified through whole genome analysis. This work will facilitate the mechanism of aerobic denitrification and provide evidence for its potential application in the nitrogen removal.

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July 7, 2019

Biofilm formation potential of heat-resistant Escherichia coli dairy isolates and the complete genome of multidrug-resistant, heat-resistant strain FAM21845.

We tested the biofilm formation potential of 30 heat-resistant and 6 heat-sensitive Escherichia coli dairy isolates. Production of curli and cellulose, static biofilm formation on polystyrene (PS) and stainless steel surfaces, biofilm formation under dynamic conditions (Bioflux), and initial adhesion rates (IAR) were evaluated. Biofilm formation varied greatly between strains, media, and assays. Our results highlight the importance of the experimental setup in determining biofilm formation under conditions of interest, as correlation between different assays was often not a given. The heat-resistant, multidrug-resistant (MDR) strain FAM21845 showed the strongest biofilm formation on PS and the highest IAR and was the only strain that formed significant biofilms on stainless steel under conditions relevant to the dairy industry, and it was therefore fully sequenced. Its chromosome is 4.9 Mb long, and it harbors a total of five plasmids (147.2, 54.2, 5.8, 2.5, and 1.9 kb). The strain carries a broad range of genes relevant to antimicrobial resistance and biofilm formation, including some on its two large conjugative plasmids, as demonstrated in plate mating assays.IMPORTANCE In biofilms, cells are embedded in an extracellular matrix that protects them from stresses, such as UV radiation, osmotic shock, desiccation, antibiotics, and predation. Biofilm formation is a major bacterial persistence factor of great concern in the clinic and the food industry. Many tested strains formed strong biofilms, and especially strains such as the heat-resistant, MDR strain FAM21845 may pose a serious issue for food production. Strong biofilm formation combined with diverse resistances (some encoded on conjugative plasmids) may allow for increased persistence, coselection, and possible transfer of these resistance factors. Horizontal gene transfer may conceivably occur in the food production setting or the gastrointestinal tract after consumption. Copyright © 2017 Marti et al.

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July 7, 2019

Transcriptome Remodeling of Acinetobacter baumannii during Infection and Treatment.

Acinetobacter baumannii is an increasingly common multidrug-resistant pathogen in health care settings. Although the genetic basis of antibiotic resistance mechanisms has been extensively studied, much less is known about how genetic variation contributes to other aspects of successful infections. Genetic changes that occur during host infection and treatment have the potential to remodel gene expression patterns related to resistance and pathogenesis. Longitudinal sets of multidrug-resistant A. baumannii isolates from eight patients were analyzed by RNA sequencing (RNA-seq) to identify differentially expressed genes and link them to genetic changes contributing to transcriptional variation at both within-patient and population levels. The number of differentially expressed genes among isolates from the same patient ranged from 26 (patient 588) to 145 (patient 475). Multiple patients had isolates with differential gene expression patterns related to mutations in the pmrAB and adeRS two-component regulatory system genes, as well as significant differences in genes related to antibiotic resistance, iron acquisition, amino acid metabolism, and surface-associated proteins. Population level analysis revealed 39 genetic regions with clade-specific differentially expressed genes, for which 19, 8, and 3 of these could be explained by insertion sequence mobilization, recombination-driven sequence variation, and intergenic mutations, respectively. Multiple types of mutations that arise during infection can significantly remodel the expression of genes that are known to be important in pathogenesis. IMPORTANCE Health care-associated multidrug-resistant Acinetobacter baumannii can cause persistent infections in patients, but bacterial cells must overcome host defenses and antibiotic therapies to do so. Genetic variation arises during host infection, and new mutations are often enriched in genes encoding transcriptional regulators, iron acquisition systems, and surface-associated structures. In this study, genetic variation was shown to result in transcriptome remodeling at the level of individual patients and across phylogenetic groups. Differentially expressed genes include those related to capsule modification, iron acquisition, type I pili, and antibiotic resistance. Population level transcriptional variation reflects genome dynamics over longer evolutionary time periods, and convergent transcriptional changes support the adaptive significance of these regions. Transcriptional changes can be attributed to multiple types of genomic change, but insertion sequence mobilization had a predominant effect. The transcriptional effects of mutations that arise during infection highlight the rapid adaptation of A. baumannii during host exposure. Copyright © 2017 Wright et al.

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July 7, 2019

High-quality draft genome sequences of four lignocellulose-degrading bacteria isolated from Puerto Rican forest soil: Gordonia sp., Paenibacillus sp., Variovorax sp., and Vogesella sp.

Here, we report the high-quality draft genome sequences of four phylogenetically diverse lignocellulose-degrading bacteria isolated from tropical soil (Gordonia sp., Paenibacillus sp., Variovorax sp., and Vogesella sp.) to elucidate the genetic basis of their ability to degrade lignocellulose. These isolates may provide novel enzymes for biofuel production. Copyright © 2017 Woo et al.

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July 7, 2019

IncFII conjugative plasmid-mediated transmission of blaNDM-1 elements among animal-borne Escherichia coli strains.

This study aims to investigate the prevalence and transmission dynamics of the blaNDM-1 gene in animal Escherichia coli strains. Two IncFII blaNDM-1-encoding plasmids with only minor structural variation in the MDR region, pHNEC46-NDM and pHNEC55-NDM, were found to be responsible for the transmission of blaNDM-1 in these strains. The blaNDM-1 gene can be incorporated into plasmids and stably inherited in animal-borne E. coli strains that can be maintained in animal gut microflora even without carbapenem selection pressure. Copyright © 2016 American Society for Microbiology.

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July 7, 2019

Genetic and functional characterization of blaCTX-M-199, a novel tazobactam and sulbactam resistance-encoding gene located in a conjugative mcr-1-bearing IncI2 plasmid.

The study reported the genetic and functional characterization of a novel CTX-M-199 ß-lactamase, which was encoded by a blaCTX-M-64 variant gene found in a conjugative mcr-1-bearing IncI2 plasmid and exhibited resistance to ß-lactamase inhibitors, tazobactam and sulbactam. Copyright © 2017 American Society for Microbiology.

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July 7, 2019

Novel plasmid-mediated colistin resistance gene mcr-3 in Escherichia coli.

The mobile colistin resistance gene mcr-1 has attracted global attention, as it heralds the breach of polymyxins, one of the last-resort antibiotics for the treatment of severe clinical infections caused by multidrug-resistant Gram-negative bacteria. To date, six slightly different variants of mcr-1, and a second mobile colistin resistance gene, mcr-2, have been reported or annotated in the GenBank database. Here, we characterized a third mobile colistin resistance gene, mcr-3 The gene coexisted with 18 additional resistance determinants in the 261-kb IncHI2-type plasmid pWJ1 from porcine Escherichia colimcr-3 showed 45.0% and 47.0% nucleotide sequence identity to mcr-1 and mcr-2, respectively, while the deduced amino acid sequence of MCR-3 showed 99.8 to 100% and 75.6 to 94.8% identity to phosphoethanolamine transferases found in other Enterobacteriaceae species and in 10 Aeromonas species, respectively. pWJ1 was mobilized to an E. coli recipient by conjugation and contained a plasmid backbone similar to those of other mcr-1-carrying plasmids, such as pHNSHP45-2 from the original mcr-1-harboring E. coli strain. Moreover, a truncated transposon element, TnAs2, which was characterized only in Aeromonas salmonicida, was located upstream of mcr-3 in pWJ1. This ?TnAs2-mcr-3 element was also identified in a shotgun genome sequence of a porcine E. coli isolate from Malaysia, a human Klebsiella pneumoniae isolate from Thailand, and a human Salmonella enterica serovar Typhimurium isolate from the United States. These results suggest the likelihood of a wide dissemination of the novel mobile colistin resistance gene mcr-3 among Enterobacteriaceae and aeromonads; the latter may act as a potential reservoir for mcr-3IMPORTANCE The emergence of the plasmid-mediated colistin resistance gene mcr-1 has attracted substantial attention worldwide. Here, we examined a colistin-resistant Escherichia coli isolate that was negative for both mcr-1 and mcr-2 and discovered a novel mobile colistin resistance gene, mcr-3 The amino acid sequence of MCR-3 aligned closely with phosphoethanolamine transferases from Enterobacteriaceae and Aeromonas species originating from both clinical infections and environmental samples collected in 12 countries on four continents. Due to the ubiquitous profile of aeromonads in the environment and the potential transfer of mcr-3 between Enterobacteriaceae and Aeromonas species, the wide spread of mcr-3 may be largely underestimated. As colistin has been and still is widely used in veterinary medicine and used at increasing frequencies in human medicine, the continuous monitoring of mobile colistin resistance determinants in colistin-resistant Gram-negative bacteria is imperative for understanding and tackling the dissemination of mcr genes in both the agricultural and health care sectors. Copyright © 2017 Yin et al.

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July 7, 2019

Phenotypic and genomic survey on organic acid utilization profile of Pseudomonas mendocina strain S5.2, a vineyard soil isolate.

Root exudates are chemical compounds that are released from living plant roots and provide significant energy, carbon, nitrogen and phosphorus sources for microbes inhabiting the rhizosphere. The exudates shape the microflora associated with the plant, as well as influences the plant health and productivity. Therefore, a better understanding of the trophic link that is established between the plant and the associated bacteria is necessary. In this study, a comprehensive survey on the utilization of grapevine and rootstock related organic acids were conducted on a vineyard soil isolate which is Pseudomonas mendocina strain S5.2. Phenotype microarray analysis has demonstrated that this strain can utilize several organic acids including lactic acid, succinic acid, malic acid, citric acid and fumaric acid as sole growth substrates. Complete genome analysis using single molecule real-time technology revealed that the genome consists of a 5,120,146 bp circular chromosome and a 252,328 bp megaplasmid. A series of genetic determinants associated with the carbon utilization signature of the strain were subsequently identified in the chromosome. Of note, the coexistence of genes encoding several iron-sulfur cluster independent isoenzymes in the genome indicated the importance of these enzymes in the events of iron deficiency. Synteny and comparative analysis have also unraveled the unique features of D-lactate dehydrogenase of strain S5.2 in the study. Collective information of this work has provided insights on the metabolic role of this strain in vineyard soil rhizosphere.

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July 7, 2019

Genetic and biochemical characterization of HMB-1, a novel subclass B1 metallo-ß-lactamase found in a Pseudomonas aeruginosa clinical isolate.

To characterize a novel subclass B1 metallo-ß-lactamase (MBL) found in an MDR Pseudomonas aeruginosa clinical isolate.The isolate P. aeruginosa NRZ-03096 was recovered in 2012 from an anal swab from a patient hospitalized in Northern Germany and showed high MICs of carbapenems. MBL production was analysed by several phenotypic tests. Genetic characterization of the novel bla gene and MLST was performed by WGS. The novel bla gene was expressed in Escherichia coli TOP10 and the enzyme was subjected to biochemical characterization to determine the kinetic parameters K m and k cat .P. aeruginosa NRZ-03096 was resistant to all tested ß-lactams and showed an MBL phenotype. Shotgun cloning experiments yielded a clone producing a novel subclass B1 enzyme with only 74.3% identity to the next nearest relative, KHM-1. The novel MBL was named HMB-1 (for Hamburg MBL). Analysis of WGS data showed that the bla HMB-1 gene was chromosomally located as part of a Tn 3 family transposon that was named Tn 6345 . Expression of bla HMB-1 in E. coli TOP10 led to increased resistance to ß-lactams. Determination of K m and k cat revealed that HMB-1 had different hydrolytic characteristics compared with KHM-1, with lower hydrolytic rates for cephalosporins and a higher rate for imipenem.The identification of HMB-1 further underlines the ongoing spread and diversification of carbapenemases in Gram-negative human pathogens and especially in P. aeruginosa .

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July 7, 2019

2015 epidemic of severe Streptococcus agalactiae sequence type 283 infections in Singapore associated with the consumption of raw freshwater fish: a detailed analysis of clinical, epidemiological, and bacterial sequencing data.

Streptococcus agalactiae (group B Streptococcus [GBS]) has not been described as a foodborne pathogen. However, in 2015, a large outbreak of severe invasive sequence type (ST) 283 GBS infections in adults epidemiologically linked to the consumption of raw freshwater fish occurred in Singapore. We attempted to determine the scale of the outbreak, define the clinical spectrum of disease, and link the outbreak to contaminated fish.Time-series analysis was performed on microbiology laboratory data. Food handlers and fishmongers were screened for enteric carriage of GBS. A retrospective cohort study was conducted to assess differences in demographic and clinical characteristics of patients with invasive ST283 and non-ST283 infections. Whole-genome sequencing was performed on human and fish ST283 isolates from Singapore, Thailand, and Hong Kong.The outbreak was estimated to have started in late January 2015. Within the study cohort of 408 patients, ST283 accounted for 35.8% of cases. Patients with ST283 infection were younger and had fewer comorbidities but were more likely to develop meningoencephalitis, septic arthritis, and spinal infection. Of 82 food handlers and fishmongers screened, none carried ST283. Culture of 43 fish samples yielded 13 ST283-positive samples. Phylogenomic analysis of 161 ST283 isolates from humans and fish revealed they formed a tight clade distinguished by 93 single-nucleotide polymorphisms.ST283 is a zoonotic GBS clone associated with farmed freshwater fish, capable of causing severe disease in humans. It caused a large foodborne outbreak in Singapore and poses both a regional and potentially more widespread threat.

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July 7, 2019

Genomic epidemiology of NDM-1-encoding plasmids in Latin American clinical isolates reveals insights into the evolution of multidrug resistance

Bacteria that produce the broad-spectrum Carbapenem antibiotic New Delhi Metallo-ß-lactamase (NDM) place a burden on health care systems worldwide, due to the limited treatment options for infections caused by them and the rapid global spread of this antibiotic resistance mechanism. Although it is believed that the associated resistance gene blaNDM-1 originated in Acinetobacter spp., the role of Enterobacteriaceae in its dissemination remains unclear. In this study, we used whole genome sequencing to investigate the dissemination dynamics of blaNDM-1-positive plasmids in a set of 21 clinical NDM-1-positive isolates from Colombia and Mexico (Providencia rettgeri, Klebsiella pneumoniae, and Acinetobacter baumannii) as well as six representative NDM-1-positive Escherichia coli transconjugants. Additionally, the plasmids from three representative P. rettgeri isolates were sequenced by PacBio sequencing and finished. Our results demonstrate the presence of previously reported plasmids from K. pneumoniae and A. baumannii in different genetic backgrounds and geographically distant locations in Colombia. Three new previously unclassified plasmids were also identified in P. rettgeri from Colombia and Mexico, plus an interesting genetic link between NDM-1-positive P. rettgeri from distant geographic locations (Canada, Mexico, Colombia, and Israel) without any reported epidemiological links was discovered. Finally, we detected a relationship between plasmids present in P. rettgeri and plasmids from A. baumannii and K. pneumoniae. Overall, our findings suggest a Russian doll model for the dissemination of blaNDM-1 in Latin America, with P. rettgeri playing a central role in this process, and reveal new insights into the evolution and dissemination of plasmids carrying such antibiotic resistance genes.© The Author 2017. Published by Oxford University Press on behalf of the Society for Molecular Biology and Evolution.

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July 7, 2019

Genomic characterization of a large plasmid containing a bla NDM-1 gene carried on Salmonella enterica serovar Indiana C629 isolate from China.

The bla NDM-1 gene in Salmonella species is mostly reported in clinical cases, but is rarely isolated from red and white meat in China.A Salmonella Indiana (S. Indiana) isolate was cultured from a chicken carcass procured from a slaughterhouse in China. Antimicrobial susceptibility was tested against a panel of agents. Whole-genome sequencing of the isolate was carried out and data was analyzed.A large plasmid, denoted as plasmid pC629 (210,106 bp), containing a composite cassette, consisting of IS26-bla NDM-1-ble MBL -?trpF-tat-cutA-ISCR1-sul1-qacE?1-aadA2-dfrA12-intI1-IS26 was identified. The latter locus was physically linked with bla OXA-1, bla CTX-M-65, bla TEM-1-encoding genes. A mercury resistance operon merACDEPTR was also identified; it was flanked on the proximal side, among IS26 element and the distally located on the bla NDM-1 gene. Plasmid pC629 also contained 21 other antimicrobial resistance-encoding genes, such as aac(6′)-Ib-cr, aac(3)-VI, aadA5, aph(4)-Ia, arr-3, blmS, brp, catB3, dfrA17, floR, fosA, mph(A), mphR, mrx, nimC/nimA, oqxA, oqxB, oqxR, rmtB, sul1, sul2. Two virulence genes were also identified on plasmid pC629.To the best of our knowledge, this is the first report of bla NDM-1 gene being identified from a plasmid in a S. Indiana isolate cultured from chicken carcass in China.

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