Biovar 1A Research Articles

Characterisation of Yersinia Enterocolitica Strains Isolated from Wildlife in the Northwestern Italian Alps.

IntroductionYersiniosis is a zoonosis causing gastroenteritis, diarrhoea, and occasionally reactive arthritis and septicaemia. Cases are often linked to meat consumption and the most common aetiological agent is the Gram-negative bacilliform Yersinia enterocolitica bacterium. The occurrence of Yersinia spp. among wild animals has mostly been studied in wild boar, but it has seldom been in other species.Material and MethodsA total of 1,868 faecal samples from animals found dead or hunted were collected between 2015 and 2018 in the Valle d’Aosta region of the northwestern Italian Alps. Alpine ibex faecal samples were collected during a health monitoring program in 2018. Bacteria were isolated via PCR and confirmed as Y. enterocolitica biochemically. Strain antimicrobial susceptibility was tested by Kirby–Bauer disc diffusion, and the presence of virulence factors and antimicrobial resistance genes was investigated using whole-genome sequencing.ResultsYersinia enterocolitica strains of biotype 1A were detected in six faecal samples from red deer (0.93%), roe deer (0.49%) and red foxes (0.7%). Strains found in beech martens (3.57%) and Alpine ibex (2.77%) belonged to biotypes 1B and 5, respectively and harboured the pYPTS01 plasmid that had only been detected in Y. pseudotuberculosis PB1/+. All the isolates were resistant to ampicillin and erythromycin.ConclusionThe biovar 1A strains exhibited different virulence factors and behaved like non-pathogenic commensals. The strain from an Alpine ibex also harboured the self-transmissible pYE854 plasmid that can mobilise itself and the pYPTS01 plasmid to other strains. The beech marten could be considered a sentinel animal for Y. enterocolitica. Phenotypic resistance may account for the ability of all the strains to resist β-lactams.

Lon Protease- and Temperature-Dependent Activity of a Lysis Cassette Located in the Insecticidal Island of Yersinia enterocolitica.

The Yersinia genus comprises pathogens that can adapt to an environmental life cycle stage as well as to mammals. Yersinia enterocolitica strain W22703 exhibits both insecticidal and nematocidal activity conferred by the tripartite toxin complex (Tc) that is encoded on the 19-kb pathogenicity island Tc-PAI Ye All tc genes follow a strict temperature regulation in that they are silenced at 37°C but activated at lower temperatures. Four highly conserved phage-related genes, located within the Tc-PAI Ye , were recently demonstrated to encode a biologically functional holin-endolysin gene cassette that lyses its own host W22703 at 37°C. Conditions transcriptionally activating the cassette are not yet known. In contrast to Escherichia coli, the overproduction of holin and endolysin did not result in cell lysis of strain W22703 at 15°C. When the holin-endolysin genes were overexpressed at 15°C in four Y. enterocolitica biovars and in four other Yersinia spp., a heterogenous pattern of phenotypes was observed, ranging from lysis resistance of a biovar 1A strain to the complete growth arrest of a Y. kristensenii strain. To decipher the molecular mechanism underlying this temperature-dependent lysis, we constructed a Lon protease-negative mutant of W22703 in which the overexpression of the lysis cassette leads to cell death at 15°C. Overexpressed endolysin exhibited a high proteolytic susceptibility in strain W22703 but remained stable in the W22703 Δlon strain or in Y. pseudotuberculosis Although artificial overexpression was applied here, the data indicate that Lon protease plays a role in the control of the temperature-dependent lysis in Y. enterocolitica W22703.IMPORTANCE The investigation of the mechanisms that help pathogens survive in the environment is a prerequisite to understanding their evolution and their virulence capacities. In members of the genus Yersinia, many factors involved in virulence, metabolism, motility, or biofilm formation follow a strict temperature-dependent regulation. While the molecular mechanisms underlying the activation of determinants at body temperature have been analyzed in detail, the molecular basis of low-temperature-dependent phenotypes is largely unknown. Here, we demonstrate that a novel phage-related lysis cassette, which is part of the insecticidal and nematocidal pathogenicity island of Y. enterocolitica, does not lyse its own host following overexpression at 15°C and that the Lon protease is involved in this phenotype.

Green fluorescent protein labeling of food pathogens Yersinia enterocolitica and Yersinia pseudotuberculosis

Labeling of bacteria with marker genes, such as green fluorescent protein, is a useful and practicable tool for tracking and enumerating bacterial cells in a complex environment e.g. discrimination from the indigenous background population. In this study, novel TurboGFP prokaryotic expression vector was utilized for labeling of Yersinia species. Y. enterocolitica biovar 1A, biovar 2, biovar 4 and Y. pseudotuberculosis were successfully transformed with the vector and expressed bright green fluorescence that was even detectable visually by eye. No adverse effects were observed in growth behavior of the labeled strains compared to wild type (parental) strains and vector maintenance for longer time periods could be achieved for Y. enterocolitica biovar 1A, Y. enterocolitica biovar 2 and Y. pseudotuberculosis.

Resistance to amoxicillin-clavulanate and its relation to virulence-related factors in Yersinia enterocolitica biovar 1A

Recent studies have reported that the virulence factors (VFs) were detected more frequently in amoxicillin-clavulanate (AMC) susceptible clinical isolates of Escherichia coli. Here, we have evaluated the relationship between VFs and AMC-resistance phenotype in clinical isolates of Y. enterocolitica biovar 1A. The presence/absence of VFs was compared with their minimum inhibitory concentrations for AMC in strains of two serovars. We observed that the strains of the serovar O: 6, 30-6, 31 showed a similar relationship between the number of VFs and resistance to clavulanic acid as in E. coli but not of serovar O: 6, 30. Variations in the promoters/complete coding sequences (CCDSs) of β-lactamase gene (blaA) or the serological characteristics could not account for unusual susceptibility to AMC displayed by the strains of the serovar O: 6, 30. Therefore, we speculate that since the clinical strains of serovar O: 6, 30-6, 31 originated from the environment they were less exposed to antibiotics compared to clinical strains of serovar O: 6, 30. Thus, AMC susceptibility seems to be influenced by factors other than serotypes or promoters/CCDS of β-lactamase genes.

Analysis of iron acquisition and storage-related genes in clinical and non-clinical strains of Yersinia enterocolitica biovar 1A.

Possession of mechanisms for iron acquisition and its storage enhances the ability of the bacteria to survive in the iron-limiting environment of the host. In this study, 81 strains of Yersinia enterocolitica biovar 1A isolated from various clinical (n=51) and non-clinical (n=30) sources were investigated for the presence of the genes related to iron acquisition and storage. Important genes which were present in more than 85% of the strains included hasA, foxA, bfr, bfd, ftnA, and hmsT as well as the fhuCDB, fepBDGCfesfepA, feoAB, yfuABCD, hemPRSTUV, and hmsHFRS gene clusters. Majority of these genes is being reported for the first time in biovar 1A strains and showed significant homology with genes present in the known pathogenic biovars of Y.enterocolitica. However, no significant difference was observed in the distribution of iron acquisition and storage-related genes among clinical and non-clinical biovar 1A strains. Thus, it may be suggested that the presence of iron acquisition and storage-related genes per se might not be responsible for the supposedly better ability of clinical biovar 1A strains to cause infections in humans. However, in the backdrop of this data, the need to undertake functional studies are highly recommended.

Structural Variabilities in β-Lactamase (blaA) of Different Biovars of Yersinia enterocolitica: Implications for β-Lactam Antibiotic and β-Lactamase Inhibitor Susceptibilities.

Yersiniosis caused by Yersinia enterocolitica has been reported from all continents. The bacterial species is divided into more than fifty serovars and six biovars viz. 1A, 1B, 2, 3, 4 and 5 which differ in geographical distribution, ecological niches and pathogenicity. Most Y.enterocolitica strains harbor chromosomal genes for two β-lactamases, blaA an Ambler class A penicillinase and blaB an Ambler class C inducible cephalosporinase. In the present study, susceptibility to b-lactam antibiotics and β-lactamase inhibitor was studied for Y. enterocolitica strains of biovars 1A, 1B, 2 and 4. We observed that β-lactamases were expressed differentially among strains of different biovars. To understand the molecular mechanisms underlying such differential expression, the sequences of genes and promoters of blaA were compared. Also, the variants of blaA present in different biovars were modeled and docked with amoxicillin and clavulanic acid. The mRNA secondary structures of blaA variants were also predicted in-silico. Our findings indicated that neither variations in the promoter regions, nor the secondary structures of mRNA contributed to higher/lower expression of blaA in different biovars. Analysis of H-bonding residues of blaA variants with amoxicillin and clavulanic acid revealed that if amino acid residues of a β-lactamase interacting with amoxicillin and the clavulanic acid were similar, clavulanic acid was effective in engaging the enzyme, accounting for a significant reduction in MIC of amoxicillin-clavulanate. This finding might aid in designing better β-lactamase inhibitors with improved efficiencies in future.

Proteomic analysis of Yersinia enterocolitica biovar 1A under iron-rich and iron-poor conditions indicate existence of efficiently regulated mechanisms of iron homeostasis

The pathogenicity of Yersinia enterocolitica biovar 1A strains is controversial as these lack most of the known virulence factors. Acquisition of iron and presence of well-regulated iron homeostasis in bacteria represents an important virulence trait. Differential abundance of proteins was examined under iron-rich and iron-poor conditions in a clinical Y. enterocolitica biovar 1A strain IP27407. Whole cell protein profiles were analysed by 2D gel electrophoresis (2D-GE). Following statistical and MALDI-TOF MS analyses, 28 differentially abundant proteins were identified. Significant iron-responsive changes were observed in the proteins involved in iron acquisition or storage namely, hemin receptor (HemR), periplasmic Fe2+ transport protein (Tpd), periplasmic chelated iron-binding protein (YfeA) and bacterioferritin (Bfr). Quantitative real-time PCR (qRT-PCR) of eight mRNA transcripts revalidated the differential protein abundance. In silico analysis of iron homeostasis mediated by the bacterioferritin and bacterioferritin-associated ferredoxin (Bfr–Bfd) complex suggested two pathways for the release of reserve iron which might be operating under conditions of different iron availability. The study, for the first time, showed the existence of highly competent iron homeostasis mechanisms in Y. enterocolitica biovar 1A and identified the key proteins involved thereof. Such mechanisms might have implications for the pathogenicity of Y. enterocolitica biovar 1A strains. Biological significanceAlthough, a few studies have identified the differentially abundant bacterial proteins in response to iron starvation, little information is available in this regard for Y. enterocolitica (especially, the biovar 1A strains). In the present study, differential abundance of several proteins was identified under iron-rich and iron-poor conditions by 2D-GE and MALDI-TOF/MS analysis. These included proteins which may not only be directly implicated in iron acquisition or storage but also play crucial role in cellular metabolism. Given the absence of most known virulence factors in Y. enterocolitica biovar 1A strains, demonstration of well-regulated mechanisms for efficient iron homeostasis constitutes an important observation. The proteins, as identified in the present study, provide useful insights to further unravel the potential pathogenicity of the biovar 1A strains.

Yersinia enterocolitica: Revisitation of an Enduring Human Pathogen

Yersinia enterocolitica remains a charismatic microorganism. The research findings outlined in this article have uncovered new virulence factors, such as type III secretion systems, and have advanced our understanding of the pathogenesis of gastrointestinal infections caused by Y. enterocolitica. Furthermore, a clarification of the potential capability of biovar 1A strains, long regarded as avirulent, has now proven that they cause symptomatic infections in appropriate patient settings, such as immunosuppression or hemochromatosis. Researchers have also expanded the spectra of non-gastrointestinal clinical manifestations of disease to include cutaneous infections, endocarditis, pneumonia, pharyngitis, and meningitis.

Molecular modeling and docking of novel laccase from multiple serotype of Yersinia enterocolitica suggests differential and multiple substrate binding

Multi-copper oxidases (MCOs) are widely distributed in bacteria, where they are responsible for metal homeostasis, acquisition and oxidation. Using specific primers, yacK coding for MCO was amplified from different serotypes of Yersinia enterocolitica biovar 1A. Homology modeling of the protein followed by docking with five well-known substrates for different MCO’s (viz., 2,2′-azino-bis(3-ethylbenzthiazoline-6-sulphonic acid [ABTS], syringaldazine, l-tyrosine, ammonium ferrous sulfate and guaiacol), lignin monomers (Coniferyl alcohol, p-coumaryl alcohol and sinapyl alcohol) and two inhibitors i.e., kojic acid and N-hydroxyglycine was done. The docking gave maximum GoldScore i.e., 91.93 and 72.64 with ammonium ferrous sulfate and ABTS, respectively. Similarly, docking with ICM gave −82.10 and −83.61 docking score, confirming the protein to be true laccase with ferroxidase activity. Further, validation with ammonium ferrous sulfate as substrate gave laccase activity of 0.36Units/L/min. Guaiacol, l-tyrosine, and lignin monomers showed good binding affinity with protein models with GoldScores of 35.89, 41.82, 40.41, 41.12 and 43.10, respectively. The sequence study of all the cloned Yack genes showed serotype specific clade in dendrogram. There was distinct discrimination in the ligand binding affinity of Y. enterocolitica laccase, among strains of same clonal groups, suggesting it as a tool for phylogenetic studies.

Interaction of Yersinia enterocolitica biovar 1A with cultured cells in vitro does not reflect the two previously identified clonal groups

Yersinia enterocolitica biovar 1A strains have been delineated into two clonal groups (A and B) based on repetitive extragenic palindrome- and enterobacterial repetitive intergenic consensus-PCR genotyping. The present study investigated the interaction of Y. enterocolitica biovar 1A strains with cultured cells in vitro by their ability to adhere, invade and survive within these cells. The response of macrophages to these strains was also studied by quantifying the expression of inducible nitric oxide synthase, production of nitric oxide and cytokines, and activation of NFκB. The survival rate of clonal group B strains inside macrophages was significantly higher than that of clonal group A strains. In addition, strains harbouring the fepA gene showed better survival inside macrophages. However, the production of nitric oxide and cytokines and activation of NFκB did not show any significant differences between the two clonal groups. In this study, interaction of Y. enterocolitica biovar 1A with cultured cells in vitro did not reflect the previously identified clonal groups, but was more dependent on the characteristics of the individual strains. Therefore, a combination of genotype and phenotype data must be used to characterize this extremely heterogeneous organism.

THU0440 Yersinia Enterocolitica Biotype 1A Infection can Trigger Reactive Arthritis

BackgroundYersinia enterocolitica (YE) is a well-known trigger of reactive arthritis (ReA) [1]. Studies of YE-related ReA have concentrated on the traditionally pathogenic serotypes O:3 and O:9 (biotypes 4 and 2,...

Identification and distribution of putative virulence genes in clinical strains ofYersinia enterocoliticabiovar 1A by suppression subtractive hybridization

To detect putative virulence genes in clinical strains of Yersinia enterocolitica biovar 1A by suppression subtractive hybridization between two closely related strains of clinical and nonclinical origin having the same serotype (O:6,30-6,31). Suppression Subtractive Hybridization (SSH) was used to identify genomic differences between clinical (serotype O:6,30-6,31, from diarrhoeic human stools) and nonclinical (serotype O:6,30-6,31, from wastewater) strains of Y. enterocolitica biovar 1A. Following genomic subtraction and DNA sequencing, nine DNA sequences that were present only in clinical biovar 1A strains were identified. The sequences identified using SSH showed similarity to conserved hypothetical proteins, proteins related to iron acquisition and haemin storage, type 1 secretion proteins, flagellar hook proteins, exported protein and ABC transport system. All these sequences showed high similarity with Y. enterocolitica 8081 (biovar 1B). The distribution of these genes was further analysed using PCR in 26 clinical strains of Y. enterocolitica biovar 1A. The results revealed that the distribution of these genes was not uniform. Genes related to iron acquisition and storage, and flagellar proteins might be responsible for virulence of some of the clinical strains of Y. enterocolitica biovar 1A. Genes identified in this study might be useful in understanding the pathogenic potential of clinical strains of Y. enterocolitica biovar 1A.

Proteomic analysis of arsenite – mediated multiple antibiotic resistance in Yersinia enterocolitica biovar 1A

Arsenic is one of the most important global environmental pollutants. In the present study, fifty one clinical strains of Yersinia enterocolitica biovar 1A showed high resistance to arsenite and arsenate. The minimum inhibitory concentration (MIC) of arsenite (0.625-20 mM) was lower than arsenate (10-80 mM). Growth of Y. enterocolitica in 2 mM arsenite led to 2-8 fold increase in MICs of the five antibiotics (amikacin, ciprofloxacin, gentamycin, kanamycin and tetracycline), suggesting expression of arsenite-induced multiple antibiotic resistance among the strains. Proteomic analysis of Y. enterocolitica revealed differential expression of certain proteins following arsenite exposure, which included a putative outer membrane porin (OmpA) and a putative amino acid transporter protein. In conclusion, modulation of membrane permeability may be involved in the induction of arsenite-mediated expression of multiple antibiotic resistance in Y. enterocolitica.

Whole cell protein profiling reiterate phylogenetic relationships among strains of Yersinia enterocolitica biovar 1A as discerned earlier by different genotyping methods

To evaluate whole cell protein profiling vis-à-vis genotyping to discern phylogenetic relationships among strains of Y. enterocolitica biovar 1A. Whole cell protein profiling of Y. enterocolitica biovar 1A strains was performed using SDS-PAGE. Twenty-one distinct protein profile types were obtained among a collection of 81 strains isolated from clinical and nonclinical sources. Whole cell protein profiling exhibited discriminatory index (DI) of 0·80 and clustered the strains into two distinct clonal groups. The clinical and the aquatic serotype O:6,30-6,31 strains were clustered into two discrete subgroups. Whole cell protein profiling displayed sufficient diversity among strains of Y. enterocolitica biovar 1A, and the phylogenetic relationships obtained were in good agreement with those established earlier by genotyping techniques. Whole cell protein profiling was as discriminatory as some of the genotyping methods and has the potentiality to be used as an adjunct tool to study epidemiology of Y. enterocolitica.

YersiniaSpecies Isolated from Bats, Germany

<i>Yersinia</i>Species Isolated from Bats, Germany

Genetic relationships between clinical and non-clinical strains of Yersinia enterocolitica biovar 1A as revealed by multilocus enzyme electrophoresis and multilocus restriction typing

BackgroundGenetic relationships among 81 strains of Y. enterocolitica biovar 1A isolated from clinical and non-clinical sources were discerned by multilocus enzyme electrophoresis (MLEE) and multilocus restriction typing (MLRT) using six loci each. Such studies may reveal associations between the genotypes of the strains and their sources of isolation.ResultsAll loci were polymorphic and generated 62 electrophoretic types (ETs) and 12 restriction types (RTs). The mean genetic diversity (H) of the strains by MLEE and MLRT was 0.566 and 0.441 respectively. MLEE (DI = 0.98) was more discriminatory and clustered Y. enterocolitica biovar 1A strains into four groups, while MLRT (DI = 0.77) identified two distinct groups. BURST (Based Upon Related Sequence Types) analysis of the MLRT data suggested aquatic serotype O:6,30-6,31 isolates to be the ancestral strains from which, clinical O:6,30-6,31 strains might have originated by host adaptation and genetic change.ConclusionMLEE revealed greater genetic diversity among strains of Y. enterocolitica biovar 1A and clustered strains in four groups, while MLRT grouped the strains into two groups. BURST analysis of MLRT data nevertheless provided newer insights into the probable evolution of clinical strains from aquatic strains.

Multilocus variable number tandem repeat analysis as a tool to discern genetic relationships among strains ofYersinia enterocoliticabiovar 1A

To identify variable number tandem repeat (VNTR)-containing loci, and to use multilocus VNTR (MLVA) to discern genetic relationships among strains of Yersinia enterocolitica biovar 1A isolated from diverse sources. The whole genome sequence of Y. enterocolitica 8081 was analysed and eight VNTR loci with repeat sizes between 4 and 9 bp, and each containing more than four repeat copies were selected for MLVA typing of 88 strains of Y. enterocolitica. Of these, four loci were polymorphic and generated 26 MLVA genotypes among 81 strains of Y. enterocolitica biovar 1A. MLVA was found to be quite discriminatory (DI = 0.87). Cluster analysis and population modelling using minimum spanning tree (MST) clearly clustered Y. enterocolitica biovar 1A into two major groups. The MLVA is easy to perform and can be used to discern clonal relationship among strains of Y. enterocolitica. Also the phylogenetic relationships obtained with MLVA genotypes were in good agreement with those established by other typing methods. The MLVA method reported is relatively more discriminatory than the other genotyping methods and has the potential to be used as an epidemiological tool for the study of strains of Y. enterocolitica biovar 1A.

Molecular and biochemical characterization of urease and survival of Yersinia enterocolitica biovar 1A in acidic pH in vitro

BackgroundYersinia enterocolitica, an important food- and water-borne enteric pathogen is represented by six biovars viz. 1A, 1B, 2, 3, 4 and 5. Despite the lack of recognized virulence determinants, some biovar 1A strains have been reported to produce disease symptoms resembling that produced by known pathogenic biovars (1B, 2-5). It is therefore imperative to identify determinants that might contribute to the pathogenicity of Y. enterocolitica biovar 1A strains. Y. enterocolitica invariably produces urease and the role of this enzyme in the virulence of biovar 1B and biovar 4 strains has been reported recently. The objective of this work was to study genetic organization of the urease (ure) gene complex of Y. enterocolitica biovar 1A, biochemical characterization of the urease, and the survival of these strains under acidic conditions in vitro.ResultsThe ure gene complex (ureABCEFGD) of Y. enterocolitica biovar 1A included three structural and four accessory genes, which were contiguous and was flanked by a urea transport (yut) gene on the 3' side. Differences were identified in ure gene complex of biovar 1A strain compared to biovar 1B and 4 strains. This included a smaller ureB gene and larger intergenic regions between the structural genes. The crude urease preparation exhibited optimal pH and temperature of 5.5 and 65°C respectively, and Michaelis-Menten kinetics with a Km of 1.7 ± 0.4 mM urea and Vmax of 7.29 ± 0.42 μmol of ammonia released/min/mg protein. The urease activity was dependent on growth temperature and growth phase of Y. enterocolitica biovar 1A, and the presence of nickel in the medium. The molecular mass of the enzyme was > 545 kDa and an isoelectric point of 5.2. The number of viable Y. enterocolitica biovar 1A decreased significantly when incubated at pH 2.5 for 2 h. However, no such decrease was observed at this pH in the presence of urea.ConclusionsThe ure gene cluster of biovar 1A strains though similar to biovar 1B and 4 strains, exhibited important differences. The study also showed the ability of biovar 1A strains of Y. enterocolitica to survive at highly acidic pH in vitro in the presence of urea.

Characteristics of β-lactamases and their genes (blaA and blaB) in Yersinia intermedia and Y. frederiksenii

BackgroundThe presence of β-lactamases in Y. enterocolitica has been reported to vary with serovars, biovars and geographical origin of the isolates. An understanding of the β-lactamases in other related species is important for an overall perception of antibiotic resistance in yersiniae. The objective of this work was to study the characteristics of β-lactamases and their genes in strains of Y. intermedia and Y. frederiksenii, isolated from clinical and non-clinical sources in India.ResultsThe enzymes, Bla-A (a constitutive class A penicillinase) and Bla-B (an inducible class C cephalosporinase) were found to be present in all the clinical and non-clinical strains of Y. intermedia and Y. frederiksenii by double disc diffusion method. The results showed differential expression of Bla-A as indicated by presence/absence of synergy whereas expression of Bla-B was quite consistent. The presence of these enzymes was also reflected in the high minimum inhibitory concentrations, MIC50 (126–1024 mg/L) and MIC90 (256–1024 mg/L) of β-lactam antibiotics against these species. Restriction fragment length polymorphism (RFLP) revealed heterogeneity in both blaA and blaB genes of Y. intermedia and Y. frederiksenii. The blaA gene of Y. intermedia shared significant sequence identity (87–96%) with blaA of Y. enterocolitica biovars 1A, 1B and 4. The sequence identity of blaA of Y. frederiksenii with these biovars was 77–79%. The sequence identity of blaB gene of Y. intermedia and Y. frederiksenii was more (85%) with that of Y. enterocolitica biovars 1A, 1B and 2 compared to other species viz., Y. bercovieri, Y. aldovae and Y. ruckeri. Isoelectric focusing data further revealed that both Y. intermedia and Y. frederiksenii produced Bla-A (pI 8.7) and "Bla-B like" (pI 5.5–7.1) enzymes.ConclusionBoth Y. intermedia and Y. frederiksenii showed presence of blaA and blaB genes and unequivocal expression of the two β-lactamases. Limited heterogeneity was detected in blaA and blaB genes as judged by PCR-RFLP. Phylogenetic relationships showed that the two species shared a high degree of identity in their bla genes. This is the first study reporting characteristics of β-lactamases and their genes in strains of Y. intermedia and Y. frederiksenii isolated from Asian region.

The rrn locus and gyrB genotyping confirm the existence of two clonal groups in strains of Yersinia enterocolitica subspecies palearctica biovar 1A

Eighty-one strains of Yersinia enterocolitica biovar 1A representing several serotypes isolated from India, France, Germany and the USA were analyzed using ribotyping, 16S–23S rDNA intergenic spacer length polymorphism analysis (PCR-ribotyping) and gyrB restriction fragment length polymorphism. Ribotyping with BglI, NciI and EcoRV distinguished 81 strains into 4, 3 and 2 ribotypes respectively. BglI- NciI combination gave the highest Simpson's diversity index (DI = 0.43). Strains with identical ribotypes were further differentiated by PCR-ribotyping. The combination of BglI- NciI ribotyping with PCR ribotyping increased DI to 0.72. This suggested that the combination of the two may be used for molecular epidemiological studies of Y. enterocolitica biovar 1A. This approach clearly resolved the strains into two clonal groups, each comprising strains isolated from humans, swine, pork and wastewater. PCR-RFLP of the gyrB gene using three enzymes ( AluI, MspI and HinfI) distinguished strains into seven types and confirmed the existence of two clonal groups. Thus, assessment of heterogeneity based on chromosomal restriction analysis (ribotyping), rRNA spacer length polymorphism (PCR-ribotyping) and gyrB gene analysis were in concordance and provided unequivocal evidence for the presence of two groups amongst strains of Y. enterocolitica biovar 1A despite their diverse geographic origins. These data also grouped clinical and non-clinical strains of serotype O:6,30–6,31 into discrete subgroups.