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July 7, 2019

New Delhi metallo-ß-lactamase-1-producing Klebsiella pneumoniae, Florida, USA(1).

New Delhi metallo-ß-lactamase (NDM)–producing Enterobacteriaceae have swiftly spread worldwide since an initial report in 2008 from a patient who had been transferred from India back home to Sweden (1). Epidemiologically, the global diffusion of NDM-1 producers has been associated with the Indian subcontinent and the Balkan region, which are considered the primary and secondary reservoirs of these pathogens, respectively (1). However, recent reports suggest that countries in the Middle East may constitute another potential reservoir for NDM-1 producers (1). More than 100 NDM-producing isolates have been reported in the United States, most of which were associated with recent travel from the Indian subcontinent (2,3). We report an NDM-1–producing Klebsiella pneumoniae strain that was recovered from a patient who had been transferred from Iran to a hospital in Florida, United States.


July 7, 2019

Complete genome sequence of thermophilic Bacillus smithii type strain DSM 4216(T).

Bacillus smithii is a facultatively anaerobic, thermophilic bacterium able to use a variety of sugars that can be derived from lignocellulosic feedstocks. Being genetically accessible, it is a potential new host for biotechnological production of green chemicals from renewable resources. We determined the complete genomic sequence of the B. smithii type strain DSM 4216(T), which consists of a 3,368,778 bp chromosome (GenBank accession number CP012024.1) and a 12,514 bp plasmid (GenBank accession number CP012025.1), together encoding 3880 genes. Genome annotation via RAST was complemented by a protein domain analysis. Some unique features of B. smithii central metabolism in comparison to related organisms included the lack of a standard acetate production pathway with no apparent pyruvate formate lyase, phosphotransacetylase, and acetate kinase genes, while acetate was the second fermentation product.


July 7, 2019

Evidence for an opportunistic and endophytic lifestyle of the Bursaphelenchus xylophilus-associated bacteria Serratia marcescens PWN146 isolated from wilting Pinus pinaster.

Pine wilt disease (PWD) results from the interaction of three elements: the pathogenic nematode, Bursaphelenchus xylophilus; the insect-vector, Monochamus sp.; and the host tree, mostly Pinus species. Bacteria isolated from B. xylophilus may be a fourth element in this complex disease. However, the precise role of bacteria in this interaction is unclear as both plant-beneficial and as plant-pathogenic bacteria may be associated with PWD. Using whole genome sequencing and phenotypic characterization, we were able to investigate in more detail the genetic repertoire of Serratia marcescens PWN146, a bacterium associated with B. xylophilus. We show clear evidence that S. marcescens PWN146 is able to withstand and colonize the plant environment, without having any deleterious effects towards a susceptible host (Pinus thunbergii), B. xylophilus nor to the nematode model C. elegans. This bacterium is able to tolerate growth in presence of xenobiotic/organic compounds, and use phenylacetic acid as carbon source. Furthermore, we present a detailed list of S. marcescens PWN146 potentials to interfere with plant metabolism via hormonal pathways and/or nutritional acquisition, and to be competitive against other bacteria and/or fungi in terms of resource acquisition or production of antimicrobial compounds. Further investigation is required to understand the role of bacteria in PWD. We have now reinforced the theory that B. xylophilus-associated bacteria may have a plant origin.


July 7, 2019

Complete genome of the starch-degrading myxobacteria Sandaracinus amylolyticus DSM 53668T.

Myxobacteria are members of d-proteobacteria and are typified by large genomes, well-coordinated social behavior, gliding motility, and starvation-induced fruiting body formation. Here, we report the 10.33 Mb whole genome of a starch-degrading myxobacterium Sandaracinus amylolyticus DSM 53668(T) that encodes 8,962 proteins, 56 tRNA, and two rRNA operons. Phylogenetic analysis, in silico DNA-DNA hybridization and average nucleotide identity reveal its divergence from other myxobacterial species and support its taxonomic characterization into a separate family Sandaracinaceae, within the suborder Sorangiineae. Sequence similarity searches using the Carbohydrate-active enzymes (CAZyme) database help identify the enzyme repertoire of S. amylolyticus involved in starch, agar, chitin, and cellulose degradation. We identified 16 a-amylases and two ?-amylases in the S. amylolyticus genome that likely play a role in starch degradation. While many of the amylases are seen conserved in other d-proteobacteria, we notice several novel amylases acquired via horizontal transfer from members belonging to phylum Deinococcus-Thermus, Acidobacteria, and Cyanobacteria. No agar degrading enzyme(s) were identified in the S. amylolyticus genome. Interestingly, several putative ß-glucosidases and endoglucanases proteins involved in cellulose degradation were identified. However, the absence of cellobiohydrolases/exoglucanases corroborates with the lack of cellulose degradation by this bacteria. © The Author 2016. Published by Oxford University Press on behalf of the Society for Molecular Biology and Evolution.


July 7, 2019

Molecular evolution of a Klebsiella pneumoniae ST278 isolate harboring blaNDM-7 and involved in nosocomial transmission.

During 2013, ST278 Klebsiella pneumoniae with blaNDM-7 was isolated from the urine (KpN01) and rectum (KpN02) of a patient in Calgary, Canada. The same strain (KpN04) was subsequently isolated from another patient in the same unit. Interestingly, a carbapenem-susceptible K. pneumoniae ST278 (KpN06) was obtained 1 month later from the blood of the second patient. Next-generation sequencing (NGS) revealed that the loss of carbapenem-resistance in KpN06 was due to a 5-kb deletion on the blaNDM-7-harboring IncX3 plasmid. In addition, an IncFIB plasmid in KpN06 had a 27-kb deletion that removed genes encoding for heavy metal resistance. Phylogenetic analysis showed that the K. pneumoniae ST278 from patient 2 was likely a descendant of KpN02 and that KpN06 was a close progenitor of an environmental ST278. It is unclear whether KpN06 lost the blaNDM-7 gene in vivo. This study detailed the remarkable plasticity and speed of evolutionary changes in multidrug-resistant K. pneumoniae, demonstrating the highly recombinant nature of this species. It also highlights the ability of NGS to clarify molecular microevolutionary events within antibiotic-resistant organisms.© The Author 2016. Published by Oxford University Press for the Infectious Diseases Society of America. All rights reserved. For permissions, e-mail journals.permissions@oup.com.


July 7, 2019

Large-scale maps of variable infection efficiencies in aquatic Bacteroidetes phage-host model systems.

Microbes drive ecosystem functioning and their viruses modulate these impacts through mortality, gene transfer and metabolic reprogramming. Despite the importance of virus-host interactions and likely variable infection efficiencies of individual phages across hosts, such variability is seldom quantified. Here, we quantify infection efficiencies of 38 phages against 19 host strains in aquatic Cellulophaga (Bacteroidetes) phage-host model systems. Binary data revealed that some phages infected only one strain while others infected 17, whereas quantitative data revealed that efficiency of infection could vary 10 orders of magnitude, even among phages within one population. This provides a baseline for understanding and modeling intrapopulation host range variation. Genera specific host ranges were also informative. For example, the Cellulophaga Microviridae, showed a markedly broader intra-species host range than previously observed in Escherichia coli systems. Further, one phage genus, Cba41, was examined to investigate nonheritable changes in plating efficiency and burst size that depended on which host strain it most recently infected. While consistent with host modification of phage DNA, no differences in nucleotide sequence or DNA modifications were detected, leaving the observation repeatable, but the mechanism unresolved. Overall, this study highlights the importance of quantitatively considering replication variations in studies of phage-host interactions. © 2016 Society for Applied Microbiology and John Wiley & Sons Ltd.


July 7, 2019

Use of genomic approaches in understanding the role of Actinomycetes as PGP in grain legumes

The advancement in molecular technologies has given a breakthrough to explore the untapped and novel microbial isolates for characterization in every aspect as we can consider microbes as an important primary natural store house for key secondary metabolites and enzymes. Actinomycetes are the most fruitful source of microorganisms for all types of bioactive secondary metabolites, including agroactive-antibiotic molecules that are best recognized and most valuable for their role in agriculture and industries. In agriculture, actinomycetes are used as biocontrol agents against some pests and pathogenic organisms as well as plant growth-promoting (PGP) agents for crops. Use of different molecular methods, e.g., metagenomics, metatranscriptomics, genetic fingerprinting, proteogenomics, and metaproteomics, are more significant for classifying and discovering the immense diversity in microbial population and for understanding their interactions with other abiotic and biotic environmental elements. The opportunity of accessing inexpensive sequencing techniques has led to the assemblies of copious genomic data for actinomycetes, such as Streptomyces and related species, with the goal of discovering novel bioactive metabolic and their utility as PGP; however, the use of actinomycetes in agriculture using genomic approaches is in its initial stages.


July 7, 2019

Plasmid-mediated colistin resistance in a patient infected with Klebsiella pneumoniae.

It is alarming that the plasmid-mediated mcr-1-encoded colistin resistance discovered by Yi-Yun Liu and colleagues,1 probably selected in cows and pigs as discussed by Marisa Haenni and colleagues2 and Surbhi Malhorta-Kumar and colleagues,3 is now spreading globally in Gram-negative pathogens.4 Moreover, colistin-resistant Escherichia coli without the canonical mcr-1 gene suggest that other (transferable) colistin-resistant mechanisms exist.3


July 7, 2019

Comparative evaluation of the genomes of three common Drosophila-associated bacteria.

Drosophila melanogaster is an excellent model to explore the molecular exchanges that occur between an animal intestine and associated microbes. Previous studies in Drosophila uncovered a sophisticated web of host responses to intestinal bacteria. The outcomes of these responses define critical events in the host, such as the establishment of immune responses, access to nutrients, and the rate of larval development. Despite our steady march towards illuminating the host machinery that responds to bacterial presence in the gut, there are significant gaps in our understanding of the microbial products that influence bacterial association with a fly host. We sequenced and characterized the genomes of three common Drosophila-associated microbes: Lactobacillus plantarum, Lactobacillus brevis and Acetobacter pasteurianus For each species, we compared the genomes of Drosophila-associated strains to the genomes of strains isolated from alternative sources. We found that environmental Lactobacillus strains readily associated with adult Drosophila and were similar to fly isolates in terms of genome organization. In contrast, we identified a strain of A. pasteurianus that apparently fails to associate with adult Drosophila due to an inability to grow on fly nutrient food. Comparisons between association competent and incompetent A. pasteurianus strains identified a short list of candidate genes that may contribute to survival on fly medium. Many of the gene products unique to fly-associated strains have established roles in the stabilization of host-microbe interactions. These data add to a growing body of literature that examines the microbial perspective of host-microbe relationships. © 2016. Published by The Company of Biologists Ltd.


July 7, 2019

Survival and evolution of a large multidrug resistance plasmid in new clinical bacterial hosts.

Large conjugative plasmids are important drivers of bacterial evolution and contribute significantly to the dissemination of antibiotic resistance. Although plasmid borne multidrug resistance is recognized as one of the main challenges in modern medicine, the adaptive forces shaping the evolution of these plasmids within pathogenic hosts are poorly understood. Here we study plasmid-host adaptations following transfer of a 73?kb conjugative multidrug resistance plasmid to naïve clinical isolates of Klebsiella pneumoniae and Escherichia coli. We use experimental evolution, mathematical modelling and population sequencing to show that the long-term persistence and molecular integrity of the plasmid is highly influenced by multiple factors within a 25?kb plasmid region constituting a host-dependent burden. In the E. coli hosts investigated here, improved plasmid stability readily evolves via IS26 mediated deletions of costly regions from the plasmid backbone, effectively expanding the host-range of the plasmid. Although these adaptations were also beneficial to plasmid persistence in a naïve K. pneumoniae host, they were never observed in this species, indicating that differential evolvability can limit opportunities of plasmid adaptation. While insertion sequences are well known to supply plasmids with adaptive traits, our findings suggest that they also play an important role in plasmid evolution by maintaining the plasticity necessary to alleviate plasmid-host constrains. Further, the observed evolutionary strategy consistently followed by all evolved E. coli lineages exposes a trade-off between horizontal and vertical transmission that may ultimately limit the dissemination potential of clinical multidrug resistance plasmids in these hosts.© The Author 2016. Published by Oxford University Press on behalf of the Society for Molecular Biology and Evolution.


July 7, 2019

Genome-directed analysis of prophage excision, host defence systems, and central fermentative metabolism in Clostridium pasteurianum.

Clostridium pasteurianum is emerging as a prospective host for the production of biofuels and chemicals, and has recently been shown to directly consume electric current. Despite this growing biotechnological appeal, the organism’s genetics and central metabolism remain poorly understood. Here we present a concurrent genome sequence for the C. pasteurianum type strain and provide extensive genomic analysis of the organism’s defence mechanisms and central fermentative metabolism. Next generation genome sequencing produced reads corresponding to spontaneous excision of a novel phage, designated f6013, which could be induced using mitomycin C and detected using PCR and transmission electron microscopy. Methylome analysis of sequencing reads provided a near-complete glimpse into the organism’s restriction-modification systems. We also unveiled the chief C. pasteurianum Clustered Regularly Interspaced Short Palindromic Repeats (CRISPR) locus, which was found to exemplify a Type I-B system. Finally, we show that C. pasteurianum possesses a highly complex fermentative metabolism whereby the metabolic pathways enlisted by the cell is governed by the degree of reductance of the substrate. Four distinct fermentation profiles, ranging from exclusively acidogenic to predominantly alcohologenic, were observed through redox consideration of the substrate. A detailed discussion of the organism’s central metabolism within the context of metabolic engineering is provided.


July 7, 2019

Isolation and plasmid characterization of carbapenemase (IMP-4) producing Salmonella enterica Typhimurium from cats.

Carbapenem-resistant Enterobacteriaceae (CRE) are a pressing public health issue due to limited therapeutic options to treat such infections. CREs have been predominantly isolated from humans and environmental samples and they are rarely reported among companion animals. In this study we report on the isolation and plasmid characterization of carbapenemase (IMP-4) producing Salmonella enterica Typhimurium from a companion animal. Carbapenemase-producing S. enterica Typhimurium carrying blaIMP-4 was identified from a systemically unwell (index) cat and three additional cats at an animal shelter. All isolates were identical and belonged to ST19. Genome sequencing revealed the acquisition of a multidrug-resistant IncHI2 plasmid (pIMP4-SEM1) that encoded resistance to nine antimicrobial classes including carbapenems and carried the blaIMP-4-qacG-aacA4-catB3 cassette array. The plasmid also encoded resistance to arsenic (MIC-150?mM). Comparative analysis revealed that the plasmid pIMP4-SEM1 showed greatest similarity to two blaIMP-8 carrying IncHI2 plasmids from Enterobacter spp. isolated from humans in China. This is the first report of CRE carrying a blaIMP-4 gene causing a clinical infection in a companion animal, with presumed nosocomial spread. This study illustrates the broader community risk entailed in escalating CRE transmission within a zoonotic species such as Salmonella, and in a cycle that encompasses humans, animals and the environment.


July 7, 2019

Use of WGS data for investigation of a long-term NDM-1-producing Citrobacter freundii outbreak and secondary in vivo spread of blaNDM-1 to Escherichia coli, Klebsiella pneumoniae and Klebsiella oxytoca.

An outbreak of NDM-1-producing Citrobacter freundii and possible secondary in vivo spread of blaNDM-1 to other Enterobacteriaceae were investigated.From October 2012 to March 2015, meropenem-resistant Enterobacteriaceae were detected in 45 samples from seven patients at Aalborg University Hospital, Aalborg, Denmark. In silico resistance genes, Inc plasmid types and STs (MLST) were obtained from WGS data from 24 meropenem-resistant isolates (13 C. freundii, 6 Klebsiella pneumoniae, 4 Escherichia coli and 1 Klebsiella oxytoca) and 1 meropenem-susceptible K. oxytoca. The sequences of the meropenem-resistant C. freundii isolates were compared by phylogenetic analyses. In vitro susceptibility to 21 antimicrobial agents was tested. Furthermore, in vitro conjugation and plasmid characterization was performed.From the seven patients, 13 highly clonal ST18 NDM-1-producing C. freundii were isolated. The ST18 NDM-1-producing C. freundii isolates were only susceptible to tetracycline, tigecycline, colistin and fosfomycin (except for the C. freundii isolates from Patient 2 and Patient 7, which were additionally resistant to tetracycline). The E. coli and K. pneumoniae from different patients belonged to different STs, indicating in vivo transfer of blaNDM-1 in the individual patients. This was further supported by in vitro conjugation and detection of a 154 kb IncA/C2 plasmid with blaNDM-1. Patient screenings failed to reveal any additional cases. None of the patients had a history of recent travel abroad and the source of the blaNDM-1 plasmid was unknown.To our knowledge, this is the first report of an NDM-1-producing C. freundii outbreak and secondary in vivo spread of an IncA/C2 plasmid with blaNDM-1 to other Enterobacteriaceae.© The Author 2016. Published by Oxford University Press on behalf of the British Society for Antimicrobial Chemotherapy. All rights reserved. For Permissions, please e-mail: journals.permissions@oup.com.


July 7, 2019

Genome and plasmid analysis of blaIMP-4 -carrying Citrobacter freundii B38.

Sequencing of the blaIMP-4 -carrying C. freundii B38 using PacBio SMRT technique revealed that the genome contained a chromosome of 5,134,500 bp, and three plasmids, pOZ172 (127,005 bp), pOZ181 (277,592 bp), and pOZ182 (18,467 bp). Plasmid pOZ172 was identified as IncFIIY, like pP10164-NDM and pNDM-EcGN174. It carries a class 1 integron with four cassettes: blaIMP-4-qacG2-aacA4-aphA15, and a complete hybrid tni module (tniR-tniQ-tniB-tniA). The recombination of tniR from Tn402 (identical) with tniQBA (99%) from Tn5053 occurred within the res site of Tn402/5053. The Tn402/5053-like integron, named Tn6017, was inserted into Tn1722 at the res II site. The replication, partitioning and transfer systems of pOZ181 were similar to IncHI2 (e.g. R478) and contained a sul1-type class 1 integron with the cassette array: orf-dfrA1-orf-gcu37-aadA5 linked to an upstream Tn1696 tnpA-tnpR and to a downstream 3′ CS and ISCR1 A Tn2 transposon with a blaTEM-1b ß-lactamase was identified on pOZ182. Other interesting resistance determinants on the B38 chromosome included MDR efflux pumps, AmpC ß-lactamase, and resistances to Cu, Ag, As, and Zn. This is the first report of a complete tni module linked to a blaIMP- 4 carrying class 1 integron, and together with other recently reported non-sul1 integrons, represents the emergence of a distinct evolutionary lineage of class 1 integrons lacking a 3′ -CS (qacE?1-sul1). The unique cassette array, complete tni module of Tn6017, and incompatibility group of pOZ172 suggests a different blaIMP-4 evolutionary pathway in C. freundii B38 compared to other blaIMP-4 foundin Gram-negative bacteria in the Western Pacific Region. Copyright © 2016, American Society for Microbiology. All Rights Reserved.


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