scholarly journals Comparative genomics of global optrA-carrying Enterococcus faecalis uncovers a common chromosomal hotspot for optrA acquisition within a diversity of core and accessory genomes

2020 ◽  
Vol 6 (6) ◽  
Author(s):  
Ana R. Freitas ◽  
Ana P. Tedim ◽  
Carla Novais ◽  
Val F. Lanza ◽  
Luísa Peixe
Keyword(s):  
Enterococcus Faecalis ◽  
Type Species ◽  
De Novo ◽  
Integration Site ◽  
Medium Size ◽  
Phylogenetic Structure ◽  
Content Type ◽  
Link Type ◽  
Healthy Humans ◽  
Wide Range

Linezolid-resistant Enterococcus faecalis (LREfs) carrying optrA are increasingly reported globally from multiple sources, but we lack a comprehensive analysis of human and animal optrA-LREfs strains. To assess if optrA is dispersed in isolates with varied genetic backgrounds or with common genetic features, we investigated the phylogenetic structure, genetic content [antimicrobial resistance (AMR), virulence, prophages, plasmidome] and optrA-containing platforms of 27 publicly available optrA-positive E. faecalis genomes from different hosts in seven countries. At the genome-level analysis, an in-house database with 64 virulence genes was tested for the first time. Our analysis showed a diversity of clones and adaptive gene sequences related to a wide range of genera from Firmicutes . Phylogenies of core and accessory genomes were not congruent, and at least PAI-associated and prophage genes contribute to such differences. Epidemiologically unrelated clones (ST21, ST476-like and ST489) obtained from human clinical and animal hosts in different continents over eight years (2010–2017) could be phylogenetically related (3–126 SNPs difference). optrA was located on the chromosome within a Tn6674-like element (n=10) or on medium-size plasmids (30–60 kb; n=14) belonging to main plasmid families (RepA_N/Inc18/Rep_3). In most cases, the immediate gene vicinity of optrA was generally identical in chromosomal (Tn6674) or plasmid (impB-fexA-optrA) backbones. Tn6674 was always inserted into the same ∆radC integration site and embedded in a 32 kb chromosomal platform common to strains from different origins (patients, healthy humans, and animals) in Europe, Africa, and Asia during 2012–2017. This platform is conserved among hundreds of E. faecalis genomes and proposed as a chromosomal hotspot for optrA integration. The finding of optrA in strains sharing common adaptive features and genetic backgrounds across different hosts and countries suggests the occurrence of common and independent genetic events occurring in distant regions and might explain the easy de novo generation of optrA-positive strains. It also anticipates a dramatic increase of optrA carriage and spread with a serious impact on the efficacy of linezolid for the treatment of Gram-positive infections.

scholarly journals Comparative genomics of global optrA-carrying Enterococcus faecalis uncovers common genetic features and a chromosomal hotspot for optrA acquisition

2019 ◽  
Author(s):  
Ana R. Freitas ◽  
Ana P. Tedim ◽  
Carla Novais ◽  
Luísa Peixe
Keyword(s):  
Enterococcus Faecalis ◽  
De Novo ◽  
Integration Site ◽  
Medium Size ◽  
Multiple Sources ◽  
Animal Reservoir ◽  
Phylogenetic Structure ◽  
Healthy Humans ◽  
Wide Range ◽  
Genetic Features

AbstractLinezolid-resistant Enterococcus faecalis (LREfs) carrying optrA are increasingly reported globally from multiple sources, but we still lack a comprehensive analysis of human and animal optrA-LREfs strains. We investigated the phylogenetic structure, genetic content [antimicrobial resistance (AMR), virulence, prophages, plasmidome] and optrA-containing platforms of 28 publicly available optrA-positive E. faecalis genomes from different hosts in 7 countries. In the genome-level analysis, in house databases with 57 virulence and 391 plasmid replication genes were tested for the first time. Our analysis showed a diversity of clones and adaptive gene sequences related to a wide range of genera, mainly but not exclusive from Firmicutes. The content in AMR and virulence genes was highly identical in contrast to the diversity of phages and plasmids observed. Epidemiologically unrelated clones (ST476-like and ST21-like) obtained from human clinical and animal hosts in different continents over 5 years (2012-2017) were phylogenetically related (3-122 SNPs difference). They also exhibited identical AMR and virulence profiles, highlighting a global spread of optrA-positive strains with relevant adaptive traits in livestock and that they might originate from an animal reservoir. optrA was located on the chromosome within a Tn6674-like element (n=9) or on medium-size plasmids (30-60 kb; n=14) belonging to main plasmid families (RepA_N/Inc18/Rep_3). In most cases, the immediate gene vicinity of optrA was identical in chromosomal (Tn6674) and plasmid (impB-fexA-optrA) backbones. Tn6674 was always inserted in the same ΔradC integration site and embedded in a 32 kb chromosomal platform common to diverse strains from different origins (patients, healthy humans, and animals) in Europe, Africa, and Asia during 2012-2018. This platform is conserved among hundreds of E. faecalis genomes and we here propose a conserved chromosomal hotspot for optrA integration. The finding of optrA in strains sharing identical adaptive features and genetic backgrounds across different hosts and countries suggest the occurrence of common and independent genetic events occurring in distant regions, and might explain the easy de novo generation of optrA-positive strains. It also anticipates a dramatic increase of optrA carriage and spread with a serious impact in the efficacy of linezolid for the treatment of Gram-positive infections.

scholarly journals Characterization of φEf-vB1 prophage infecting oral Enterococcus faecalis and enhancing bacterial biofilm formation

2020 ◽  
Vol 69 (9) ◽  
pp. 1151-1168
Author(s):  
Ahmed Askora ◽  
Mohamed El-Telbany ◽  
Gamal El-Didamony ◽  
Eman Ariny ◽  
Momen Askoura
Keyword(s):  
Enterococcus Faecalis ◽  
Biofilm Formation ◽  
Type Species ◽  
Sequence Similarity ◽  
Bacterial Biofilm ◽  
Phenotypic Traits ◽  
High Sequence Similarity ◽  
Content Type ◽  
Link Type ◽  
Wide Range

Introduction. Enterococcus faecalis is a facultative, anaerobic, opportunistic pathogen associated with medical and dental diseases. Bacterial phenotypic traits and pathogenesis are often influenced by lysogeny. Aim. The aim of this study was to characterize both the morphology and complete genome sequences of induced prophages purified from E. faecalis clinical isolates. Methodology. E. faecalis isolates were recovered from the roots of teeth of patients attending an endodontic clinic. The morphological features of isolated phage were characterized using transmission electron microscopy (TEM). DNA sequencing was performed using the Illumina MiSeq platform. Results. TEM indicated that the isolated φEf-vB1 prophage belongs to the family Siphoviridae. The φEf-vB1 prophage was stable over a wide range of temperatures and pH. Sequencing of φEf-vB1 DNA revealed that the phage genome is 37 561 bp in length with a G+C content of 37.6mol% and contained 53 ORFs. Comparison with previously predicted prophage genomes using blast revealed that φEf-vB1 has a high sequence similarity to previously characterized phage genomes. The lysogenic E. faecalis strain exhibited a higher biofilm formation capacity relative to the non-lysogenic strain. Conclusion. The current findings highlight the role of lysogeny in modification of E. faecalis properties and reveal the potential importance of prophages in E. faecalis biology and pathogenesis.

Usitatibacter rugosus gen. nov., sp. nov. and Usitatibacter palustris sp. nov., novel members of Usitatibacteraceae fam. nov. within the order Nitrosomonadales isolated from soil

2021 ◽  
Author(s):  
Selma Vieira ◽  
Katharina J. Huber ◽  
Meina Neumann-Schaal ◽  
Alicia Geppert ◽  
Manja Luckner ◽  
...  
Keyword(s):  
16S Rrna ◽  
16S Rrna Gene ◽  
Type Strain ◽  
Type Species ◽  
Sequence Similarity ◽  
Rrna Gene ◽  
The Novel ◽  
Content Type ◽  
Link Type ◽  
Wide Range

Members of the metabolically diverse order Nitrosomonadales inhabit a wide range of environments. Two strains affiliated with this order were isolated from soils in Germany and characterized by a polyphasic approach. Cells of strains 0125_3T and Swamp67T are Gram-negative rods, non-motile, non-spore-forming, non-capsulated and divide by binary fission. They tested catalase-negative, but positive for cytochrome c-oxidase. Both strains form small white colonies on agar plates and grow aerobically and chemoorganotrophically on SSE/HD 1 : 10 medium, preferably utilizing organic acids and proteinaceous substrates. Strains 0125_3T and Swamp67T are mesophilic and grow optimally without NaCl addition at slightly alkaline conditions. Major fatty acids are C16 : 1  ω7c, C16 : 0 and C14 : 0. The major polar lipids are diphosphatidylglycerol, phosphatidylethanolamine and phosphatidyglycerol. The predominant respiratory quinone is Q-8. The G+C content for 0125_3T and Swamp67T was 67 and 66.1 %, respectively. The 16S rRNA gene analysis indicated that the closest relatives (<91 % sequence similarity) of strain 0125_3T were Nitrosospira multiformis ATCC 25196T, Methyloversatilis universalis FAM5T and Denitratisoma oestradiolicum AcBE2-1T, while Nitrosospira multiformis ATCC 25196T, Nitrosospira tenuis Nv1T and Nitrosospira lacus APG3T were closest to strain Swamp67T. The two novel strains shared 97.4 % 16S rRNA gene sequence similarity with one another and show low average nucleotide identity of their genomes (83.8 %). Based on the phenotypic, chemotaxonomic, genomic and phylogenetic analysis, we propose the two novel species Usitatibacter rugosus sp. nov (type strain 0125_3T=DSM 104443T=LMG 29998T=CECT 9241T) and Usitatibacter palustris sp. nov. (type strain Swamp67T=DSM 104440T=LMG 29997T=CECT 9242T) of the novel genus Usitatibacter gen. nov., within the novel family Usitatibacteraceae fam. nov.

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.

Idification and structural characterisation of a catecholate-type siderophore produced by Stenotrophomonas maltophilia K279a

Microbiology ◽  
2021 ◽  
Vol 167 (7) ◽  
Author(s):  
Atsushi Hisatomi ◽  
Yuh Shiwa ◽  
Nobuyuki Fujita ◽  
Hiroyuki Koshino ◽  
Naoto Tanaka
Keyword(s):  
Stenotrophomonas Maltophilia ◽  
Type Species ◽  
Monomer Unit ◽  
Reversed Phase ◽  
Negative Bacterium ◽  
Content Type ◽  
Structural Characterisation ◽  
Link Type ◽  
Wide Range ◽  
C Nmr

Siderophores are produced by several bacteria that utilise iron in various environments. Elucidating the structure of a specific siderophore may have valuable applications in drug development. Stenotrophomonas maltophilia , a Gram-negative bacterium that inhabits a wide range of environments and can cause pneumonia, produces siderophores. However, the structure was unknown, and therefore, in this study, we aimed to elucidate it. We purified siderophores from cultures of S. maltophilia K279a using preparative reversed-phase HPLC. The structure was analysed through LC-MS and 1H and 13C NMR. The results demonstrated that S. maltophilia K279a produces 2,3-dihydroxybenzoylserine (DHBS), a monomer unit of enterobactin. We suggested the uptake of Iron(III) by the DHBS complex. DHBS production by S. maltophilia K279a could be attributed to an incomplete enterobactin pathway. Drugs targeting DHBS synthesis could prevent S. maltophilia infection.

scholarly journals Harmonization of whole-genome sequencing for outbreak surveillance of Enterobacteriaceae and Enterococci

2021 ◽  
Vol 7 (7) ◽  
Author(s):  
Casper Jamin ◽  
Sien De Koster ◽  
Stefanie van Koeveringe ◽  
Dieter De Coninck ◽  
Klaas Mensaert ◽  
...  
Keyword(s):  
Whole Genome Sequencing ◽  
Genome Sequencing ◽  
Type Species ◽  
De Novo ◽  
Whole Genome ◽  
Data Generation ◽  
Sequencing Data ◽  
Content Type ◽  
Link Type ◽  
Antimicrobial Resistance Genes

Whole-genome sequencing (WGS) is becoming the de facto standard for bacterial typing and outbreak surveillance of resistant bacterial pathogens. However, interoperability for WGS of bacterial outbreaks is poorly understood. We hypothesized that harmonization of WGS for outbreak surveillance is achievable through the use of identical protocols for both data generation and data analysis. A set of 30 bacterial isolates, comprising of various species belonging to the Enterobacteriaceae family and Enterococcus genera, were selected and sequenced using the same protocol on the Illumina MiSeq platform in each individual centre. All generated sequencing data were analysed by one centre using BioNumerics (6.7.3) for (i) genotyping origin of replications and antimicrobial resistance genes, (ii) core-genome multi-locus sequence typing (cgMLST) for Escherichia coli and Klebsiella pneumoniae and whole-genome multi-locus sequencing typing (wgMLST) for all species. Additionally, a split k-mer analysis was performed to determine the number of SNPs between samples. A precision of 99.0% and an accuracy of 99.2% was achieved for genotyping. Based on cgMLST, a discrepant allele was called only in 2/27 and 3/15 comparisons between two genomes, for E. coli and K. pneumoniae, respectively. Based on wgMLST, the number of discrepant alleles ranged from 0 to 7 (average 1.6). For SNPs, this ranged from 0 to 11 SNPs (average 3.4). Furthermore, we demonstrate that using different de novo assemblers to analyse the same dataset introduces up to 150 SNPs, which surpasses most thresholds for bacterial outbreaks. This shows the importance of harmonization of data-processing surveillance of bacterial outbreaks. In summary, multi-centre WGS for bacterial surveillance is achievable, but only if protocols are harmonized.

scholarly journals Lysinibacillus acetophenoni sp. nov., a solvent-tolerant bacterium isolated from acetophenone

2015 ◽  
Vol 65 (Pt_6) ◽  
pp. 1741-1748 ◽  
Author(s):  
M. Azmatunnisa ◽  
K. Rahul ◽  
K. V. N. S. Lakshmi ◽  
Ch. Sasikala ◽  
Ch. V. Ramana
Keyword(s):  
Cell Wall ◽  
Type Species ◽  
Optimal Growth ◽  
Log P ◽  
Rrna Gene ◽  
Content Type ◽  
Link Type ◽  
Wide Range ◽  
Solvent Tolerant ◽  
Micro Organisms

A Gram-stain-positive, solvent-tolerating, aerobic, rod-shaped bacterium that formed terminal endospores was isolated from the organic solvent acetophenone. The strain, designated JC23T, was oxidase- and catalase-positive. The strain grew in the presence of a wide range of organic solvents with partition coefficients (log p values) between 1 and 4, which are exceptionally toxic to micro-organisms. Based on 16S rRNA gene sequence analysis, strain JC23T was identified as belonging to the genus Lysinibacillus and was most closely related to Lysinibacillus manganicus Mn1-7T (98.5 % similarity), L. massiliensis 440831T (97.2 %) and L. chungkukjangi 2RL3-2T (96.8 %). DNA–DNA relatedness of strain JC23T with the type strains of the closest species was <39 %. Strain JC23T grew chemo-organoheterotrophically with optimal growth at pH 7 (range pH 6–9) and at 35 °C (range 25–40 °C). The DNA G+C content was 41 mol%. Major cellular fatty acids of strain JC23T were iso-C15 : 0, iso-C16 : 0, anteiso-C15 : 0 and anteiso-C17 : 0. The cell-wall peptidoglycan type was determined to be A4α (l-Lys–d-Asp), which is in agreement with the cell-wall characteristics of the genus Lysinibacillus . The predominant quinone system was MK-7. Polar lipids of strain JC23T included diphosphatidylglycerol, phosphatidylglycerol, two unidentified glycolipids, β-gentiobiosyldiacylglycerol, two unidentified phospholipids and two unidentified lipids. On the basis of our morphological, physiological, genetic, phylogenetic and chemotaxonomic analyses, we conclude that strain JC23T should be assigned to a novel species of the genus Lysinibacillus , for which the name Lysinibacillus acetophenoni sp. nov. is proposed. The type strain is strain JC23T ( = CCUG 57911T = KCTC 13605T = NBRC 105754T = DSM 23394T).

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 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.

scholarly journals Bifidobacterium aesculapii sp. nov., from the faeces of the baby common marmoset (Callithrix jacchus)

2014 ◽  
Vol 64 (Pt_8) ◽  
pp. 2819-2827 ◽  
Author(s):  
M. Modesto ◽  
S. Michelini ◽  
I. Stefanini ◽  
A. Ferrara ◽  
S. Tacconi ◽  
...  
Keyword(s):  
16S Rrna ◽  
Type Species ◽  
Callithrix Jacchus ◽  
Rrna Gene ◽  
Positive Staining ◽  
Gene Sequences ◽  
Common Marmosets ◽  
Content Type ◽  
Link Type ◽  
Wide Range

Six Gram-positive-staining, microaerophilic, non-spore-forming, fructose-6-phosphate phosphoketolase-positive bacterial strains with a peculiar morphology were isolated from faecal samples of baby common marmosets (Callithrix jacchus). Cells of these strains showed a morphology not reported previously for a bifidobacterial species, which resembled a coiled snake, always coiled or ring shaped or forming a ‘Y’ shape. Strains MRM 3/1T and MRM 4/2 were chosen as representative strains and characterized further. The bacteria utilized a wide range of carbohydrates and produced urease. Glucose was fermented to acetate and lactate. Strain MRM 3/1T showed a peptidoglycan type unique among members of the genus Bifidobacterium . The DNA base composition was 64.7 mol% G+C. Almost-complete 16S rRNA, hsp60, clpC and rpoB gene sequences were obtained and phylogenetic relationships were determined. Comparative analysis of 16S rRNA gene sequences showed that strains MRM 3/1T and MRM 4/2 had the highest similarities to Bifidobacterium scardovii DSM 13734T (94.6 %) and Bifidobacterium stellenboschense DSM 23968T (94.5 %). Analysis of hsp60 showed that both strains were closely related to B. stellenboschense DSM 23968T (97.5 % similarity); however, despite this high degree of similarity, our isolates could be distinguished from B. stellenboschense DSM 23968T by low levels of DNA–DNA relatedness (30.4 % with MRM 3/1T). Strains MRM 3/1T and MRM 4/2 were located in an actinobacterial cluster and were more closely related to the genus Bifidobacterium than to other genera in the family Bifidobacteriaceae . On the basis of these results, strains MRM 3/1T and MRM 4/2 represent a novel species within the genus Bifidobacterium , for which the name Bifidobacterium aesculapii sp. nov. is proposed; the type strain is MRM 3/1T ( = DSM 26737T = JCM 18761T).

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