Menu

Asset Tag: Microbiology

July 7, 2019

Characterization of VCC-1, a novel ambler class A carbapenemase from Vibrio cholerae isolated from imported retail shrimp sold in Canada.

One of the core goals of the Canadian Integrated Program for Antimicrobial Resistance Surveillance (CIPARS) is to monitor major meat commodities for antimicrobial resistance. Targeted studies with methodologies based on core surveillance protocols are used to examine other foods, e.g., seafood, for antimicrobial resistance to detect resistances of concern to public health. Here we report the discovery of a novel Ambler class A carbapenemase that was identified in a nontoxigenic strain of Vibrio cholerae (N14-02106) isolated from shrimp that was sold for human consumption in Canada. V. cholerae N14-02106 was resistant to penicillins, carbapenems, and monobactam antibiotics; however, PCR did not detect common ß-lactamases. Bioinformatic analysis of the whole-genome sequence of V. cholerae N14-02106 revealed on the large chromosome a novel carbapenemase (referred to here as VCC-1, for Vibrio cholerae carbapenemase 1) with sequence similarity to class A enzymes. Two copies of blaVCC-1 separated and flanked by ISVch9 (i.e., 3 copies of ISVch9) were found in an acquired 8.5-kb region inserted into a VrgG family protein gene. Cloned blaVCC-1 conferred a ß-lactam resistance profile similar to that in V. cholerae N14-02106 when it was transformed into a susceptible laboratory strain of Escherichia coli. Purified VCC-1 was found to hydrolyze penicillins, 1st-generation cephalosporins, aztreonam, and carbapenems, whereas 2nd- and 3rd-generation cephalosporins were poor substrates. Using nitrocefin as a reporter substrate, VCC-1 was moderately inhibited by clavulanic acid and tazobactam but not EDTA. In this report, we present the discovery of a novel class A carbapenemase from the food supply. Copyright © 2016, American Society for Microbiology. All Rights Reserved.

Read more
July 7, 2019

Horizontal gene acquisitions, mobile element proliferation, and genome decay in the host-restricted plant pathogen Erwinia tracheiphila.

Modern industrial agriculture depends on high-density cultivation of genetically similar crop plants, creating favorable conditions for the emergence of novel pathogens with increased fitness in managed compared with ecologically intact settings. Here, we present the genome sequence of six strains of the cucurbit bacterial wilt pathogen Erwinia tracheiphila (Enterobacteriaceae) isolated from infected squash plants in New York, Pennsylvania, Kentucky, and Michigan. These genomes exhibit a high proportion of recent horizontal gene acquisitions, invasion and remarkable amplification of mobile genetic elements, and pseudogenization of approximately 20% of the coding sequences. These genome attributes indicate that E. tracheiphila recently emerged as a host-restricted pathogen. Furthermore, chromosomal rearrangements associated with phage and transposable element proliferation contribute to substantial differences in gene content and genetic architecture between the six E. tracheiphila strains and other Erwinia species. Together, these data lead us to hypothesize that E. tracheiphila has undergone recent evolution through both genome decay (pseudogenization) and genome expansion (horizontal gene transfer and mobile element amplification). Despite evidence of dramatic genomic changes, the six strains are genetically monomorphic, suggesting a recent population bottleneck and emergence into E. tracheiphila’s current ecological niche. © The Author 2016. Published by Oxford University Press on behalf of the Society for Molecular Biology and Evolution.

Read more
July 7, 2019

Campylobacter fetus subspecies contain conserved type IV secretion systems on multiple genomic islands and plasmids.

The features contributing to differences in pathogenicity of the Campylobacter fetus subspecies are unknown. Putative factors involved in pathogenesis are located in genomic islands that encode a type IV secretion system (T4SS) and fic domain (filamentation induced by cyclic AMP) proteins, which may disrupt host cell processes. In the genomes of 27 C. fetus strains, three phylogenetically-different T4SS-encoding regions (T4SSs) were identified: one was located in both the chromosome and in extra-chromosomal plasmids; one was located exclusively in the chromosome; and one exclusively in extra-chromosomal plasmids. We observed that C. fetus strains can contain multiple T4SSs and that homologous T4SSs can be present both in chromosomal genomic islands (GI) and on plasmids in the C. fetus strains. The GIs of the chromosomally located T4SS differed mainly by the presence of fic genes, insertion sequence elements and phage-related or hypothetical proteins. Comparative analysis showed that T4SS sequences, inserted in the same locations, were conserved in the studied C. fetus genomes. Using phylogenetic analysis of the T4SSs, it was shown that C. fetus may have acquired the T4SS regions from other Campylobacter species by horizontal gene transfer. The identified T4SSs and fic genes were found in Cff and Cfv strains, although the presence of T4SSs and fic genes were significantly associated with Cfv strains. The T4SSs and fic genes could not be associated with S-layer serotypes or geographical origin of the strains.

Read more
July 7, 2019

Investigation of proposed ladderane biosynthetic genes from anammox bacteria by heterologous expression in E. coli.

Ladderanes are hydrocarbon chains with three or five linearly concatenated cyclobutane rings that are uniquely produced as membrane lipid components by anammox (anaerobic ammonia-oxidizing) bacteria. By virtue of their angle and torsional strain, ladderanes are unusually energetic compounds, and if produced biochemically by engineered microbes, could serve as renewable, high-energy-density jet fuel components. The biochemistry and genetics underlying the ladderane biosynthetic pathway are unknown, however, previous studies have identified a pool of 34 candidate genes from the anammox bacterium, Kuenenia stuttgartiensis, some or all of which may be involved with ladderane fatty acid biosynthesis. The goal of the present study was to establish a systematic means of testing the candidate genes from K. stuttgartiensis for involvement in ladderane biosynthesis through heterologous expression in E. coli under anaerobic conditions. This study describes an efficient means of assembly of synthesized, codon-optimized candidate ladderane biosynthesis genes in synthetic operons that allows for changes to regulatory element sequences, as well as modular assembly of multiple operons for simultaneous heterologous expression in E. coli (or potentially other microbial hosts). We also describe in vivo functional tests of putative anammox homologs of the phytoene desaturase CrtI, which plays an important role in the hypothesized ladderane pathway, and a method for soluble purification of one of these enzymes. This study is, to our knowledge, the first experimental effort focusing on the role of specific anammox genes in the production of ladderanes, and lays the foundation for future efforts toward determination of the ladderane biosynthetic pathway. Our substantial, but far from comprehensive, efforts at elucidating the ladderane biosynthetic pathway were not successful. We invite the scientific community to take advantage of the considerable synthetic biology resources and experimental results developed in this study to elucidate the biosynthetic pathway that produces unique and intriguing ladderane lipids.

Read more
July 7, 2019

Fully closed genome sequences of five type strains of the genus Cronobacter and one Cronobacter sakazakii strain.

Cronobacteris associated with infant infections and the consumption of reconstituted infant formula. Here we sequenced and closed six genomes ofC. condimenti(T),C. muytjensii(T),C. universalis(T),C. malonaticus(T),C. dublinensis(T), andC. sakazakiithat can be used as reference genomes in single nucleotide polymorphism (SNP)-based next-generation sequencing (NGS) analysis for source tracking investigations. Copyright © 2016 Moine et al.

Read more
July 7, 2019

A strange endocytobiont revealed as largest virus.

A lot of endocytobionts (or endosymbionts) have been discovered within free-living amoebae in recent years. In this article the results of a long lasting effort to derive valuable data about an extraordinary spore-like infectious microorganism (endocytobiont, endosymbiont) within host amoebae (Acanthamoeba sp.) recently isolated from the contact lens case of a patient with keratitis, are presented. It took some time until this endocytobiont could be attributed to the genus Pandoravirus following a publication of two other pandoraviruses isolated from aquatic environments. Consequently the molecular biological investigation led to the taxonomic affiliation of the endocytobiont with the genus Pandoravirus and to the description of a new Pandoravirus species, Pandoravirus inopinatum after whole-genome sequencing in 2015. The fact that it was isolated from a contact lens container of a keratitis patient gives another dimension to these findings showing paradigmatically, how readily these ‘new’ giant viruses get to humans. Copyright © 2016 Elsevier Ltd. All rights reserved.

Read more
July 7, 2019

Metabolomics-guided analysis of isocoumarin production by Streptomyces species MBT76 and biotransformation of flavonoids and phenylpropanoids.

Actinomycetes produce the majority of the antibiotics currently in clinical use. The efficiency of antibiotic production is affected by multiple factors such as nutrients, pH, temperature and growth phase. Finding the optimal harvesting time is crucial for successful isolation of the desired bioactive metabolites from actinomycetes, but for this conventional chemical analysis has limitations due to the metabolic complexity. This study explores the utility of NMR-based metabolomics for (1) optimizing fermentation time for the production of known and/or unknown bioactive compounds produced by actinomycetes; (2) elucidating the biosynthetic pathway for microbial natural products; and (3) facilitating the biotransformation of nature-abundant chemicals.

Read more
July 7, 2019

Genome sequence and analysis of Escherichia coli MRE600, a colicinogenic, nonmotile strain that lacks RNase I and the type I methyltransferase, EcoKI.

Escherichia coli strain MRE600 was originally identified for its low RNase I activity and has therefore been widely adopted by the biomedical research community as a preferred source for the expression and purification of transfer RNAs and ribosomes. Despite its widespread use, surprisingly little information about its genome or genetic content exists. Here, we present the first de novo assembly and description of the MRE600 genome and epigenome. To provide context to these studies of MRE600, we include comparative analyses with E. coli K-12 MG1655 (K12). Pacific Biosciences Single Molecule, Real-Time sequencing reads were assembled into one large chromosome (4.83 Mb) and three smaller plasmids (89.1, 56.9, and 7.1 kb). Interestingly, the 7.1-kb plasmid possesses genes encoding a colicin E1 protein and its associated immunity protein. The MRE600 genome has a G + C content of 50.8% and contains a total of 5,181 genes, including 4,913 protein-encoding genes and 268 RNA genes. We identified 41,469 modified DNA bases (0.83% of total) and found that MRE600 lacks the gene for type I methyltransferase, EcoKI. Phylogenetic, taxonomic, and genetic analyses demonstrate that MRE600 is a divergent E. coli strain that displays features of the closely related genus, Shigella. Nevertheless, comparative analyses between MRE600 and E. coli K12 show that these two strains exhibit nearly identical ribosomal proteins, ribosomal RNAs, and highly homologous tRNA species. Substantiating prior suggestions that MRE600 lacks RNase I activity, the RNase I-encoding gene, rna, contains a single premature stop codon early in its open-reading frame. © The Author 2016. Published by Oxford University Press on behalf of the Society for Molecular Biology and Evolution.

Read more
July 7, 2019

Comparative genomic analyses of the Moraxella catarrhalis serosensitive and seroresistant lineages demonstrate their independent evolution.

The bacterial species Moraxella catarrhalishas been hypothesized as being composed of two distinct lineages (referred to as the seroresistant [SR] and serosensitive [SS]) with separate evolutionary histories based on several molecular typing methods, whereas 16S ribotyping has suggested an additional split within the SS lineage. Previously, we characterized whole-genome sequences of 12 SR-lineage isolates, which revealed a relatively small supragenome when compared with other opportunistic nasopharyngeal pathogens, suggestive of a relatively short evolutionary history. Here, we performed whole-genome sequencing on 18 strains from both ribotypes of the SS lineage, an additional SR strain, as well as four previously identified highly divergent strains based on multilocus sequence typing analyses. All 35 strains were subjected to a battery of comparative genomic analyses which clearly show that there are three lineages-the SR, SS, and the divergent. The SR and SS lineages are closely related, but distinct from each other based on three different methods of comparison: Allelic differences observed among core genes; possession of lineage-specific sets of core and distributed genes; and by an alignment of concatenated core sequences irrespective of gene annotation. All these methods show that the SS lineage has much longer interstrain branches than the SR lineage indicating that this lineage has likely been evolving either longer or faster than the SR lineage. There is evidence of extensive horizontal gene transfer (HGT) within both of these lineages, and to a lesser degree between them. In particular, we identified very high rates of HGT between these two lineages for ß-lactamase genes. The four divergent strains aresui generis, being much more distantly related to both the SR and SS groups than these other two groups are to each other. Based on average nucleotide identities, gene content, GC content, and genome size, this group could be considered as a separate taxonomic group. The SR and SS lineages, although distinct, clearly form a single species based on multiple criteria including a large common core genome, average nucleotide identity values, GC content, and genome size. Although neither of these lineages arose from within the other based on phylogenetic analyses, the question of how and when these lineages split and then subsequently reunited in the human nasopharynx is explored. © The Author 2016. Published by Oxford University Press on behalf of the Society for Molecular Biology and Evolution.

Read more
July 7, 2019

Stability of the encoding plasmids and surface expression of CS6 differs in enterotoxigenic Escherichia coli (ETEC) encoding Different heat-stable (ST) enterotoxins (STh and STp).

Enterotoxigenic Escherichia coli (ETEC), one of the most common reasons of diarrhea among infants and children in developing countries, causes disease by expression of either or both of the enterotoxins heat-labile (LT) and heat-stable (ST; divided into human-type [STh] and porcine-type [STp] variants), and colonization factors (CFs) among which CS6 is one of the most prevalent ETEC CFs. In this study we show that ETEC isolates expressing CS6+STh have higher copy numbers of the cssABCD operon encoding CS6 than those expressing CS6+STp. Long term cultivation of up to ten over-night passages of ETEC isolates harboring CS6+STh (n = 10) or CS6+STp (n = 15) showed instability of phenotypic expression of CS6 in a majority of the CS6+STp isolates, whereas most of the CS6+STh isolates retained CS6 expression. The observed instability was a correlated with loss of genes cssA and cssD as examined by PCR. Mobilization of the CS6 plasmid from an unstable CS6+STp isolate into a laboratory E. coli strain resulted in loss of the plasmid after a single over-night passage whereas the plasmid from an CS6+STh strain was retained in the laboratory strain during 10 passages. A sequence comparison between the CS6 plasmids from a stable and an unstable ETEC isolate revealed that genes necessary for plasmid stabilization, for example pemI, pemK, stbA, stbB and parM, were not present in the unstable ETEC isolate. Our results indicate that stable retention of CS6 may in part be affected by the stability of the plasmid on which both CS6 and STp or STh are located.

Read more
July 7, 2019

Evolutionary history of the global emergence of the Escherichia coli epidemic clone ST131.

Escherichia colisequence type 131 (ST131) has emerged globally as the most predominant extraintestinal pathogenic lineage within this clinically important species, and its association with fluoroquinolone and extended-spectrum cephalosporin resistance impacts significantly on treatment. The evolutionary histories of this lineage, and of important antimicrobial resistance elements within it, remain unclearly defined. This study of the largest worldwide collection (n= 215) of sequenced ST131E. coliisolates to date demonstrates that the clonal expansion of two previously recognized antimicrobial-resistant clades, C1/H30R and C2/H30Rx, started around 25 years ago, consistent with the widespread introduction of fluoroquinolones and extended-spectrum cephalosporins in clinical medicine. These two clades appear to have emerged in the United States, with the expansion of the C2/H30Rx clade driven by the acquisition of ablaCTX-M-15-containing IncFII-like plasmid that has subsequently undergone extensive rearrangement. Several other evolutionary processes influencing the trajectory of this drug-resistant lineage are described, including sporadic acquisitions of CTX-M resistance plasmids and chromosomal integration ofblaCTX-Mwithin subclusters followed by vertical evolution. These processes are also occurring for another family of CTX-M gene variants more recently observed among ST131, theblaCTX-M-14/14-likegroup. The complexity of the evolutionary history of ST131 has important implications for antimicrobial resistance surveillance, epidemiological analysis, and control of emerging clinical lineages ofE. coli These data also highlight the global imperative to reduce specific antibiotic selection pressures and demonstrate the important and varied roles played by plasmids and other mobile genetic elements in the perpetuation of antimicrobial resistance within lineages.IMPORTANCEEscherichia coli, perennially a major bacterial pathogen, is becoming increasingly difficult to manage due to emerging resistance to all preferred antimicrobials. Resistance is concentrated within specificE. colilineages, such as sequence type 131 (ST131). Clarification of the genetic basis for clonally associated resistance is key to devising intervention strategies. We used high-resolution genomic analysis of a large global collection of ST131 isolates to define the evolutionary history of extended-spectrum beta-lactamase production in ST131. We documented diverse contributory genetic processes, including stable chromosomal integrations of resistance genes, persistence and evolution of mobile resistance elements within sublineages, and sporadic acquisition of different resistance elements. Both global distribution and regional segregation were evident. The diversity of resistance element acquisition and propagation within ST131 indicates a need for control and surveillance strategies that target both bacterial strains and mobile genetic elements. Copyright © 2016 Stoesser et al.

Read more
July 7, 2019

Complete genome sequence of Salmonella enterica serovar Typhimurium strain SO2 (sequence type 302) isolated from an asymptomatic child in Mexico.

The complete genome sequence of Salmonella enterica serovar Typhimurium strain SO2, isolated from an asymptomatic child in Mexico, was determined using PacBio single-molecule real-time technology. Strain SO2 has six complete chromosomal prophages, namely, ST104, Gifsy-2, ST64B, Gifsy-1, ELPhiS, and FSL SP-004, and carries a Salmonella virulence plasmid.

Read more
July 7, 2019

Complete genome sequence of Salmonella enterica serovar Typhimurium strain YU15 (sequence type 19) harboring the Salmonella genomic island 1 and virulence plasmid pSTV.

The complete genome of Salmonella enterica subsp. enterica serovar Typhimurium sequence type 19 (ST19) strain YU15, isolated in Yucatán, Mexico, from a human baby stool culture, was determined using PacBio technology. The chromosome contains five intact prophages and the Salmonella genomic island 1 (SGI1). This strain carries the Salmonella virulence plasmid pSTV.

Read more
July 7, 2019

Complete genome sequence of Enterococcus faecium commensal isolate E1002.

The emergence of vancomycin-resistant enterococci (VRE) has been associated with an increase in multidrug-resistant nosocomial infections. Here, we report the 2.614-Mb genome sequence of the Enterococcus faecium commensal isolate E1002, which will be instrumental in further understanding the determinants of the commensal and pathogenic lifestyle of E. faecium. Copyright © 2016 Tytgat et al.

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.