scholarly journals AsnB is responsible for peptidoglycan precursor amidation in Clostridium difficile in the presence of vancomycin

Microbiology ◽  
2020 ◽  
Vol 166 (6) ◽  
pp. 567-578 ◽  
Author(s):  
Fariza Ammam ◽  
Delphine Patin ◽  
Héloise Coullon ◽  
Didier Blanot ◽  
Thierry Lambert ◽  
...  
Keyword(s):  
Clostridium Difficile ◽  
Gene Cluster ◽  
Enterococcus Faecalis ◽  
Type Species ◽  
Asparagine Synthetase ◽  
Structure Modification ◽  
Content Type ◽  
E Coli ◽  
Link Type ◽  
Peptidoglycan Structure

Clostridium difficile 630 possesses a cryptic but functional gene cluster vanG Cd homologous to the vanG operon of Enterococcus faecalis . Expression of vanG Cd in the presence of subinhibitory concentrations of vancomycin is accompanied by peptidoglycan amidation on the meso-DAP residue. In this paper, we report the presence of two potential asparagine synthetase genes named asnB and asnB2 in the C. difficile genome whose products were potentially involved in this peptidoglycan structure modification. We found that asnB expression was only induced when C. difficile was grown in the presence of vancomycin, yet independently from the vanG Cd resistance and regulation operons. In addition, peptidoglycan precursors were not amidated when asnB was inactivated. No change in vancomycin MIC was observed in the asnB mutant strain. In contrast, overexpression of asnB resulted in the amidation of most of the C. difficile peptidoglycan precursors and in a weak increase of vancomycin susceptibility. AsnB activity was confirmed in E. coli . In contrast, the expression of the second asparagine synthetase, AsnB2, was not induced in the presence of vancomycin. In summary, our results demonstrate that AsnB is responsible for peptidoglycan amidation of C. difficile in the presence of vancomycin.

PBP4 and PBP5 are involved in regulating exopolysaccharide synthesis during Escherichia coli biofilm formation

Microbiology ◽  
2021 ◽  
Vol 167 (3) ◽  
Author(s):  
Sathi Mallick ◽  
Shanti Kiran ◽  
Tapas Kumar Maiti ◽  
Anindya S. Ghosh
Keyword(s):  
Escherichia Coli ◽  
Biofilm Formation ◽  
Type Species ◽  
Pentose Phosphate ◽  
Bacterial Cells ◽  
Surface Adhesion ◽  
Content Type ◽  
Penicillin Binding Proteins ◽  
E Coli ◽  
Link Type

Escherichia coli low-molecular-mass (LMM) Penicillin-binding proteins (PBPs) help in hydrolysing the peptidoglycan fragments from their cell wall and recycling them back into the growing peptidoglycan matrix, in addition to their reported involvement in biofilm formation. Biofilms are external slime layers of extra-polymeric substances that sessile bacterial cells secrete to form a habitable niche for themselves. Here, we hypothesize the involvement of Escherichia coli LMM PBPs in regulating the nature of exopolysaccharides (EPS) prevailing in its extra-polymeric substances during biofilm formation. Therefore, this study includes the assessment of physiological characteristics of E. coli CS109 LMM PBP deletion mutants to address biofilm formation abilities, viability and surface adhesion. Finally, EPS from parent CS109 and its ΔPBP4 and ΔPBP5 mutants were purified and analysed for sugars present. Deletions of LMM PBP reduced biofilm formation, bacterial adhesion and their viability in biofilms. Deletions also diminished EPS production by ΔPBP4 and ΔPBP5 mutants, purification of which suggested an increased overall negative charge compared with their parent. Also, EPS analyses from both mutants revealed the appearance of an unusual sugar, xylose, that was absent in CS109. Accordingly, the reason for reduced biofilm formation in LMM PBP mutants may be speculated as the subsequent production of xylitol and a hindrance in the standard flow of the pentose phosphate pathway.

scholarly journals d-Serine induces distinct transcriptomes in diverse Escherichia coli pathotypes

Microbiology ◽  
2021 ◽  
Vol 167 (10) ◽  
Author(s):  
James P. R. Connolly ◽  
Natasha C. A. Turner ◽  
Jennifer C. Hallam ◽  
Patricia T. Rimbi ◽  
Tom Flett ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Type Species ◽  
Pathogenic Bacteria ◽  
Neonatal Meningitis ◽  
Published Data ◽  
Toxic Metabolite ◽  
Transcriptional Responses ◽  
Content Type ◽  
E Coli ◽  
Link Type

Appropriate interpretation of environmental signals facilitates niche specificity in pathogenic bacteria. However, the responses of niche-specific pathogens to common host signals are poorly understood. d-Serine (d-ser) is a toxic metabolite present in highly variable concentrations at different colonization sites within the human host that we previously found is capable of inducing changes in gene expression. In this study, we made the striking observation that the global transcriptional response of three Escherichia coli pathotypes – enterohaemorrhagic E. coli (EHEC), uropathogenic E. coli (UPEC) and neonatal meningitis-associated E. coli (NMEC) – to d-ser was highly distinct. In fact, we identified no single differentially expressed gene common to all three strains. We observed the induction of ribosome-associated genes in extraintestinal pathogens UPEC and NMEC only, and the induction of purine metabolism genes in gut-restricted EHEC, and UPEC indicating distinct transcriptional responses to a common signal. UPEC and NMEC encode dsdCXA – a genetic locus required for detoxification and hence normal growth in the presence of d-ser. Specific transcriptional responses were induced in strains accumulating d-ser (WT EHEC and UPEC/NMEC mutants lacking the d-ser-responsive transcriptional activator DsdC), corroborating the notion that d-ser is an unfavourable metabolite if not metabolized. Importantly, many of the UPEC-associated transcriptome alterations correlate with published data on the urinary transcriptome, supporting the hypothesis that d-ser sensing forms a key part of urinary niche adaptation in this pathotype. Collectively, our results demonstrate distinct pleiotropic responses to a common metabolite in diverse E. coli pathotypes, with important implications for niche selectivity.

Emergence of a plasmid-borne tigecycline resistance in Klebsiella pneumoniae in Vietnam

2021 ◽  
Author(s):  
Aki Hirabayashi ◽  
Van Thi Thu Ha ◽  
An Van Nguyen ◽  
Son Thai Nguyen ◽  
Keigo Shibayama ◽  
...  
Keyword(s):  
Klebsiella Pneumoniae ◽  
Gene Cluster ◽  
Type Species ◽  
Bacterial Infections ◽  
Efflux Pump ◽  
Whole Genome Sequence ◽  
High Sequence Identity ◽  
Pump System ◽  
Content Type ◽  
Link Type

Tigecycline is a last-resort antimicrobial used to treat multidrug-resistant Gram-negative bacterial infections. One of the common antimicrobial resistance mechanisms is the efflux pump system composed of membrane protein complexes to excrete xenobiotic substrates. Recently, a novel gene cluster, tmexCD1-toprJ1, encoding the resistance–nodulation–cell division (RND) efflux pump was identified on plasmids in Klebsiella pneumoniae isolates in China. TMexCD1-TOprJ1 was found to be capable of excreting multiple antimicrobials, including tigecycline, which contributed to the strain's resistance. In this study, we identified K. pneumoniae isolates harbouring the tmexCD1-toprJ1 genes outside of China for the first time. Two tigecycline-resistant K. pneumoniae isolates belonging to ST273 by multilocus sequence typing were collected from different patients in a medical institution in Hanoi, Vietnam, in 2015. Whole-genome sequence analysis revealed that these isolates harboured a 288.0 kb tmexCD1-toprJ1–carrying plasmid with IncFIB and IncHI1B replicons. The tmexCD1-toprJ1 gene cluster was surrounded by several mobile gene elements, including IS26, and the plasmids had high sequence identity with that of K. pneumoniae isolated in China. Our finding suggests that the horizontal spread of tigecycline resistance mediated by tmexCD1-toprJ1–carrying plasmids has occurred in Vietnam and other countries, and raises concern about the further global dissemination.

Biochemical and functional characterization of Mycobacterium tuberculosis ketol-acid reductoisomerase

Microbiology ◽  
2021 ◽  
Vol 167 (9) ◽  
Author(s):  
Nirbhay Singh ◽  
Anu Chauhan ◽  
Ram Kumar ◽  
Sudheer Kumar Singh
Keyword(s):  
Amino Acids ◽  
Mycobacterium Tuberculosis ◽  
Type Species ◽  
Low Ph ◽  
Stress Conditions ◽  
Starvation Stress ◽  
Content Type ◽  
E Coli ◽  
Link Type

Branched-chain amino acids (BCAAs) are essential amino acids, but their biosynthetic pathway is absent in mammals. Ketol-acid reductoisomerase (IlvC) is a BCAA biosynthetic enzyme that is coded by Rv3001c in Mycobacterium tuberculosis H37Rv (Mtb-Rv) and MRA_3031 in M. tuberculosis H37Ra (Mtb-Ra). IlvCs are essential in Mtb-Rv as well as in Escherichia coli . Compared to wild-type and IlvC-complemented Mtb-Ra strains, IlvC knockdown strain showed reduced survival at low pH and under low pH+starvation stress conditions. Further, increased expression of IlvC was observed under low pH and starvation stress conditions. Confirmation of a role for IlvC in pH and starvation stress was achieved by developing E. coli BL21(DE3) IlvC knockout, which was defective for growth in M9 minimal medium, but growth could be rescued by isoleucine and valine supplementation. Growth was also restored by complementing with over-expressing constructs of Mtb-Ra and E. coli IlvCs. The E. coli knockout also had a survival deficit at pH=5.5 and 4.5 and was more susceptible to killing at pH=3.0. The biochemical characterization of Mtb-Ra and E. coli IlvCs confirmed that both have NADPH-dependent activity. In conclusion, this study demonstrates the functional complementation of E. coli IlvC by Mtb-Ra IlvC and also suggests that IlvC has a role in tolerance to low pH and starvation stress.

Functional characterization of a gene cluster responsible for inositol catabolism associated with hospital-adapted isolates of Enterococcus faecium

Microbiology ◽  
2021 ◽  
Vol 167 (9) ◽  
Author(s):  
Janetta Top ◽  
Jery Baan ◽  
Adinda Bisschop ◽  
Sergio Arredondo-Alonso ◽  
Willem van Schaik ◽  
...  
Keyword(s):  
Mouse Model ◽  
Gene Cluster ◽  
Enterococcus Faecium ◽  
Type Species ◽  
Integration Site ◽  
Functional Characterization ◽  
Donor Strain ◽  
Integrase Gene ◽  
Content Type ◽  
Link Type

Enterococcus faecium is a nosocomial, multidrug-resistant pathogen. Whole genome sequence studies revealed that hospital-associated E. faecium isolates are clustered in a separate clade A1. Here, we investigated the distribution, integration site and function of a putative iol gene cluster that encodes for myo-inositol (MI) catabolism. This iol gene cluster was found as part of an ~20 kbp genetic element (iol element), integrated in ICEEfm1 close to its integrase gene in E. faecium isolate E1679. Among 1644 E. faecium isolates, ICEEfm1 was found in 789/1227 (64.3 %) clade A1 and 3/417 (0.7 %) non-clade A1 isolates. The iol element was present at a similar integration site in 180/792 (22.7 %) ICEEfm1-containing isolates. Examination of the phylogenetic tree revealed genetically closely related isolates that differed in presence/absence of ICEEfm1 and/or iol element, suggesting either independent acquisition or loss of both elements. E. faecium iol gene cluster containing isolates E1679 and E1504 were able to grow in minimal medium with only myo-inositol as carbon source, while the iolD-deficient mutant in E1504 (E1504∆iolD) lost this ability and an iol gene cluster negative recipient strain gained this ability after acquisition of ICEEfm1 by conjugation from donor strain E1679. Gene expression profiling revealed that the iol gene cluster is only expressed in the absence of other carbon sources. In an intestinal colonization mouse model the colonization ability of E1504∆iolD mutant was not affected relative to the wild-type E1504 strain. In conclusion, we describe and functionally characterise a gene cluster involved in MI catabolism that is associated with the ICEEfm1 island in hospital-associated E. faecium isolates. We were unable to show that this gene cluster provides a competitive advantage during gut colonisation in a mouse model. Therefore, to what extent this gene cluster contributes to the spread and ecological specialisation of ICEEfm1-carrying hospital-associated isolates remains to be investigated.

Reclassification of Koreibacter algae as a later heterotypic synonym of Paraoerskovia marina and emended descriptions of the genus Paraoerskovia Khan et al. 2009 and of Paraoerskovia marina Khan et al. 2009

2013 ◽  
Vol 63 (Pt_1) ◽  
pp. 219-223 ◽  
Author(s):  
P. Schumann ◽  
R. Pukall ◽  
C. Spröer ◽  
E. Stackebrandt
Keyword(s):  
Type Species ◽  
Carbon Sources ◽  
Rrna Gene ◽  
Content Type ◽  
Link Type ◽  
Maldi Tof Mass Spectra ◽  
Peptidoglycan Structure ◽  
Lipid Pattern ◽  
Type Strains ◽  
Produce Acid

16S rRNA gene sequences deposited for the type strains of Paraoerskovia marina (CTT-37T; GenBank accession no. AB445007) and Koreibacter algae (DSW-2T; FM995611) show a similarity of 100 %. Consequently, the type strains were subjected to a polyphasic recharacterization under direct comparison in order to clarify their taxonomic position. PvuII RiboPrint patterns and quantitative ratios of cellular fatty acids revealed strain-specific differences between P. marina DSM 21750T ( = CTT-37T) and K. algae DSM 22126T ( = DSW-2T). The percentage of DNA–DNA binding of 94 % indicated that the two type strains belong to the same genomospecies. Agreement in the peptidoglycan structure and polar lipid pattern, highly similar fatty acid profiles and MALDI-TOF mass spectra, the ability to produce acid from the same carbon sources, corresponding enzymic activities and DNA G+C contents of 70.8±0.3 mol%, in addition to the consistent characteristics reported in the original descriptions, support the view that the two strains should be affiliated to the same species. According to Rules 38 and 42 of the Bacteriological Code, Koreibacter algae should be reclassified as later heterotypic synonym of Paraoerskovia marina , and the descriptions of the genus Paraoerskovia Khan et al. 2009 and of Paraoerskovia marina Khan et al. 2009 are emended accordingly.

scholarly journals bla OXA-48-like genome architecture among carbapenemase-producing Escherichia coli and Klebsiella pneumoniae in the Netherlands

2021 ◽  
Vol 7 (5) ◽  
Author(s):  
Antoni P. A. Hendrickx ◽  
Fabian Landman ◽  
Angela de Haan ◽  
Sandra Witteveen ◽  
Marga G. van Santen-Verheuvel ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Antibiotic Resistance ◽  
The Netherlands ◽  
Resistance Genes ◽  
Type Species ◽  
Antibiotic Resistance Genes ◽  
Gene Content ◽  
Content Type ◽  
E Coli ◽  
Link Type

Carbapenem-hydrolysing enzymes belonging to the OXA-48-like group are encoded by bla OXA-48-like alleles and are abundant among Enterobacterales in the Netherlands. Therefore, the objective here was to investigate the characteristics, gene content and diversity of the bla OXA-48-like carrying plasmids and chromosomes of Escherichia coli and Klebsiella pneumoniae collected in the Dutch national surveillance from 2014 to 2019 in comparison with genome sequences from 29 countries. A combination of short-read genome sequencing with long-read sequencing enabled the reconstruction of 47 and 132 complete bla OXA-48-like plasmids for E. coli and K. pneumoniae , respectively. Seven distinct plasmid groups designated as pOXA-48-1 to pOXA-48-5, pOXA-181 and pOXA-232 were identified in the Netherlands which were similar to internationally reported plasmids obtained from countries from North and South America, Europe, Asia and Oceania. The seven plasmid groups varied in size, G+C content, presence of antibiotic resistance genes, replicon family and gene content. The pOXA-48-1 to pOXA-48-5 plasmids were variable, and the pOXA-181 and pOXA-232 plasmids were conserved. The pOXA-48-1, pOXA-48-2, pOXA-48-3 and pOXA-48-5 groups contained a putative conjugation system, but this was absent in the pOXA-48-4, pOXA-181 and pOXA-232 plasmid groups. pOXA-48 plasmids contained the PemI antitoxin, while the pOXA-181 and pOXA-232 plasmids did not. Furthermore, the pOXA-181 plasmids carried a virB2-virB3-virB9-virB10-virB11 type IV secretion system, while the pOXA-48 plasmids and pOXA-232 lacked this system. A group of non-related pOXA-48 plasmids from the Netherlands contained different resistance genes, non-IncL-type replicons or no replicons. Whole genome multilocus sequence typing revealed that the bla OXA-48-like plasmids were found in a wide variety of genetic backgrounds in contrast to chromosomally encoded bla OXA-48-like alleles. Chromosomally localized bla OXA-48 and bla OXA-244 alleles were located on genetic elements of variable sizes and comprised regions of pOXA-48 plasmids. The bla OXA-48-like genetic element was flanked by a direct repeat upstream of IS1R, and was found at multiple locations in the chromosomes of E. coli . Lastly, K. pneumoniae isolates carrying bla OXA-48 or bla OXA-232 were mostly resistant for meropenem, whereas E. coli bla OXA-48, bla OXA-181 and chromosomal bla OXA-48 or bla OXA-244 isolates were mostly sensitive. In conclusion, the overall bla OXA-48-like plasmid population in the Netherlands is conserved and similar to that reported for other countries, confirming global dissemination of bla OXA-48-like plasmids. Variations in size, presence of antibiotic resistance genes and gene content impacted pOXA-48, pOXA-181 and pOXA-232 plasmid architecture.

scholarly journals Lipopolysaccharide core type diversity in the Escherichia coli species in association with phylogeny, virulence gene repertoire and distribution of type VI secretion systems

2021 ◽  
Vol 7 (9) ◽  
Author(s):  
Sébastien O. Leclercq ◽  
Maxime Branger ◽  
David G. E. Smith ◽  
Pierre Germon
Keyword(s):  
Escherichia Coli ◽  
Type Species ◽  
Virulence Gene ◽  
Uneven Distribution ◽  
Gene Repertoire ◽  
Content Type ◽  
E Coli ◽  
Link Type ◽  
Wide Range ◽  
K 12

Escherichia coli is a very versatile species for which diversity has been explored from various perspectives highlighting, for example, phylogenetic groupings and pathovars, as well as a wide range of O serotypes. The highly variable O-antigen, the most external part of the lipopolysaccharide (LPS) component of the outer membrane of E. coli , is linked to the innermost lipid A through the core region of LPS of which five different structures, denominated K-12, R1, R2, R3 and R4, have been characterized so far. The aim of the present study was to analyse the prevalence of these LPS core types in the E. coli species and explore their distribution in the different E. coli phylogenetic groups and in relationship with the virulence gene repertoire. Results indicated an uneven distribution of core types between the different phylogroups, with phylogroup A strains being the most diverse in terms of LPS core types, while phylogroups B1, D and E strains were dominated by the R3 type, and phylogroups B2 and C strains were dominated by the R1 type. Strains carrying the LEE virulence operon were mostly of the R3 type whatever the phylogroup while, within phylogroup B2, strains carrying a K-12 core all belonged to the complex STc131, one of the major clones of extraintestinal pathogenic E. coli (ExPEC) strains. The origin of this uneven distribution is discussed but remains to be fully explained, as well as the consequences of carrying a specific core type on the wider aspects of bacterial phenotype.

scholarly journals Identification of a repressor for the two iol operons required for inositol catabolism in Geobacillus kaustophilus

Microbiology ◽  
2020 ◽  
Author(s):  
Ken-ichi Yoshida ◽  
Yusuke Shirae ◽  
Ryo Nishimura ◽  
Kaho Fukui ◽  
Shu Ishikawa
Keyword(s):  
Dna Binding ◽  
Gene Cluster ◽  
Binding Sites ◽  
Type Species ◽  
Binding Property ◽  
Promoter Regions ◽  
Content Type ◽  
Link Type ◽  
Geobacillus Kaustophilus ◽  
Inositol Dehydrogenase

Geobacillus kaustophilus HTA426, a thermophilic Gram-positive bacterium, feeds on inositol as its sole carbon source, and an iol gene cluster required for inositol catabolism has been postulated with reference to the iol genes in Bacillus subtilis . The iol gene cluster of G. kaustophilus comprises two tandem operons induced in the presence of inositol; however, the mechanism underlying this induction remains unclear. B. subtilis iolQ is known to be involved in the regulation of iolX encoding scyllo-inositol dehydrogenase, and its homologue in HTA426 was found two genes upstream of the first gene (gk1899) of the iol gene cluster and was termed iolQ in G. kaustophilus . When iolQ was inactivated in G. kaustophilus , not only cellular myo-inositol dehydrogenase activity due to gk1899 expression but also the transcription of the two iol operons became constitutive. IolQ was produced and purified as a C-terminal histidine (His)-tagged fusion protein in Escherichia coli and subjected to an in vitro gel electrophoresis mobility shift assay to examine its DNA-binding property. It was observed that IolQ bound to the DNA fragments containing each of the two iol promoter regions and that DNA binding was antagonized by myo-inositol. Moreover, DNase I footprinting analyses identified two tandem binding sites of IolQ within each of the iol promoter regions. By comparing the sequences of the binding sites, a consensus sequence for IolQ binding was deduced to form a palindrome of 5′-RGWAAGCGCTTSCY-3′ (where R=A or G, W=A or T, S=G or C, and Y=C or T). IolQ functions as a transcriptional repressor regulating the induction of the two iol operons responding to myo-inositol.

scholarly journals Phylum barrier and Escherichia coli intra-species phylogeny drive the acquisition of antibiotic-resistance genes

2021 ◽  
Vol 7 (8) ◽  
Author(s):  
Marie Petitjean ◽  
Bénédicte Condamine ◽  
Charles Burdet ◽  
Erick Denamur ◽  
Etienne Ruppé
Keyword(s):  
Escherichia Coli ◽  
Antibiotic Resistance ◽  
Resistance Genes ◽  
Type Species ◽  
Antibiotic Resistance Genes ◽  
Resistance Pattern ◽  
Content Type ◽  
E Coli ◽  
Link Type ◽  
Specific Distribution

Escherichia coli is a ubiquitous bacterium that has been widely exposed to antibiotics over the last 70 years. It has adapted by acquiring different antibiotic-resistance genes (ARGs), the census of which we aim to characterize here. To do so, we analysed 70 301 E. coli genomes obtained from the EnteroBase database and detected 1 027 651 ARGs using the AMRFinder, Mustard and ResfinderFG ARG databases. We observed a strong phylogroup and clonal lineage specific distribution of some ARGs, supporting the argument for epistasis between ARGs and the strain genetic background. However, each phylogroup had ARGs conferring a similar antibiotic class resistance pattern, indicating phenotypic adaptive convergence. The G+C content or the type of ARG was not associated with the frequency of the ARG in the database. In addition, we identified ARGs from anaerobic, non- Proteobacteria bacteria in four genomes of E. coli , supporting the hypothesis that the transfer between anaerobic bacteria and E. coli can spontaneously occur but remains exceptional. In conclusion, we showed that phylum barrier and intra-species phylogenetic history are major drivers of the acquisition of a resistome in E. coli .

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