Repetitive elements sequence (REP/ERIC)-PCR based genotyping of clinical and environmental strains of Yersinia enterocolitica biotype 1A reveal existence of limited number of clonal groups

REP- and ERIC-PCR genotyping were used to assess genetic heterogeneity among 81 strains of Yersinia enterocolitica biotype 1A isolated from India, Germany, France and the USA. Although both gave comparable results, ERIC fingerprints discriminated the strains better. The rep- (REP and ERIC) PCR genotyping showed that strains having different serotypes produced identical rep-profiles indicating their limited genetic diversity. The concatenated dendrogram of REP- and ERIC-PCR fingerprints clustered the biotype 1A strains into two major groups. In each group, majority of the Indian, European and American strains exhibited similarities ranging from 85% to >95%. Similarity of rep-PCR fingerprints amongst strains isolated from widely separated geographical regions revealed existence of a limited number of clonal groups of Y. enterocolitica biotype 1A. The present study failed to reveal unequivocal relationships between rep-PCR genotypes and the source of isolation. However, the clinical serotype O:6,30-6,31 strains formed a tight cluster and the aquatic O:6,30-6,31 strains formed a yet another tight cluster.     

儿童牙齿表面的菌群分布与龋齿的关系

本实验从幼儿园和小学一年级的学生中选择５－８岁的无龋齿和有龋齿的儿童各４５例,进行牙齿表面的菌群分布与龋齿关系的研究,结果在有龋组中分离到变形链球菌８株,放线菌６株及拟杆菌２６株,而在无龋组中未分离到变形链球菌,分离到放线菌１株,拟杆菌１４株,两组比较,以上三种菌有显著差异（Ｘ２检验Ｐ＜０．０５）。说明除变形链球菌是主要的致龋菌外〔１,２〕,还应考虑放线菌和拟杆菌在龋齿发生上的作用。此外,我们还发现两组的兼性厌氧菌的分布也有所不同,无龋组中分离到棒状杆菌２９株,而有龋组中分离到棒状杆菌１１株,小肠结肠炎耶尔森氏菌７株,两组也有明显差异,故我们认为龋齿发生的原因可能是多方面的,就病原菌来说,也可能是多种细菌的混合作用,从两组兼性厌氧菌与无芽胞厌氧菌的分布不同来看,亦应考虑龋齿的发生与口腔内生态平衡有关     

粪便标本小肠结肠炎耶尔森菌筛检方法的比较

目的 比较分离培养法、反转录‒聚合酶链反应法对腹泻粪便中小肠结肠炎耶尔森菌的检测情况。方法 在2016年6月至2017年6月临床收治的腹泻患者中选取368例,均使用分离培养法、反转录‒聚合酶链反应法对其粪便中的小肠结肠炎耶尔森菌展开检验,对比分析两种方法的检验结果。结果 反转录‒聚合酶链反应法阳性率为77.17%,比分离培养法的51.09%高,差异有统计学意义（P＜0.05）;反转录‒聚合酶链反应法致病菌株检出率为83.10%,高于分离培养法的59.57%,差异有统计学意义（P＜0.05）。结论 对腹泻患者粪便标本中的小肠结肠炎耶尔森菌检测时,相较于分离培养法,反转录‒聚合酶链反应法阳性检出率更高,可使小肠结肠炎耶尔森菌性腹泻诊断准确率提升。     

Chromatographic Study of Spirosin by Use of Monoclonal Antibodies to Spirosin of Yersinia enterocolitica

Two monoclonal antibodies (MAbs) reacting with spirosins from Enterobacteriaceae were obtained in a course of screening MAbs to spirosin from Yersinia enterocolitica SYT-11-72 (YE72). The antibodies were designated MAbs-S44 and S50. They were IgG_2b and IgG_2a, respectively, both with k light chains. On Western blotting after limited proteolysis of YE72 spirosin with Staphylococcus aureus V8 protease, they reacted markedly with peptide fragments of 27 and 35 kDa, suggesting the presence of an antigenic determinant on the fragments. When supernatant cell lysate from Escherichia coli K12 was chromatographed on DEAE-cellulose and Sepharose CL-6B columns successively, a 96-kDa protein with alcohol dehydrogenase (ADH) activity was always associated with reactivity to MAb-S50. These findings combined with N-terminal amino acid sequences clearly indicate the identity of spirosin to ADH in E. coli.     

Characteristics and Pathogenicity of Non-Melibiose-Fermenting Strains of Yersinia pseudotuberculosis O3

The biological properties of non-melibiose-fermenting (NMF) strains of Yersinia pseudotuberculosis 03 were investigated. These strains were clearly distinguished from representative melibiose-fermenting (MF) strains of Y. pseudotuberculosis 03 by their pathogenicity in mice, sensitivity to some phages, production of catalase, restriction endonuclease analysis of virulence plasmid DNA with BamHI, detection of specific yersinia outer-membrane proteins with SDS-PAGE, antigenicity of the outer-membrane proteins and neutrophil resistance to phagocytosis. The pathogenicity of NMF strains was clearly less than that of MF strains. In addition, the resistance of NMF strains to phagocytosis and catalase activity was evidently weaker than that of MF strains. These results suggested that the difference of pathogenicity was due to the ability of catalase production. Although the relationship between the above characteristics and melibiose-fermentation was not analysed, the pathogenicity of Y. pseudotuberculosis 03 strains can probably be predicted by testing melibiose-fermentation and catalase production.     

Purification of lipopolysaccharide from strains of Yersinia enterocolitica belonging to serogroups 03 and 09

Lipopolysaccharide (LPS) was purified from strains of Yersinia enterocolitica belonging to serogroups 03 and 09, by three methods, and analysed by SDS-PAGE and silver staining for carbohydrate. SDS-PAGE of LPS prepared from whole-cells by digestion with proteinase-K, produced profiles containing high molecular mass LPS and a lower molecular mass region migrating as discrete bands. LPS prepared from strains belonging to serogroup 03, using a hot-phenol procedure alone was found to contain cellular proteins, and LPS prepared from strains of serogroup 09, by this method, did not contain high molecular mass carbohydrate. A novel method of preparing LPS by digesting bacterial outer membranes with proteinase-K prior to hot-phenol extraction produced protein-free LPS from strain of Y. enterocolitica 03 and high molecular mass LPS from strains belonging to serogroup 09.     

The polymerase chain reaction: an epidemiological tool to differentiate between two clusters of pathogenic Yersinia enterocolitica strains

A primer set designed to amplify the enterotoxin (yst) gene of pathogenic Yersinia enterocolitica strains generated two different electrophoretic profiles of the target sequence when a collection of strains of worldwide origin was screened. Serovars O:1,3; O:2a,3; O:3; O:5,27 and O:9, known as European strains, produced a 200-bp fragment that matched the size of the target sequence. However, serovars O:4,32; O:8; O:13a,13b; O:20 and O:21, known as American strains, generated two fragments of 1.4 and 1.6 kb. The amplified products of one American strain were sequenced and the presence of the yst gene was confirmed in both fragments. Thus, the potential of the polymerase chain reaction to be used as an epidemiological tool in differentiation between the two clusters of pathogenic strains of Y. enterocolitica could be demonstrated.     

Relevance of N-acyl-L-homoserine lactone production by Yersinia enterocolitica in fresh foods

AIMS: Determination of the food matrix impact on the potential for N-acyl-L-homoserine lactones (AHLs) production by Yersinia enterocolitica. METHODS AND RESULTS: Induction and inhibition of a sensor strain and a fluorescent assay were used to investigate Y. enterocolitica AHL production in artificial media, as well as in different food extracts. All Y. enterocolitica strains tested produced AHLs in artificial media. Thin Layer Chromatography analysis of Y. enterocolitica strains indicated the production of 3-oxo-hexanoyl homoserine lactone and hexanoyl homoserine lactone. Yersinia enterocolitica produced AHL principally in fish and meat extracts. CONCLUSIONS: AHL production by Y. enterocolitica was observed in products of animal origin, but were inhibited by some vegetables extracts. SIGNIFICANCE AND IMPACT OF THE STUDY: This study suggests that quorum sensing systems in Y. enterocolitica is significant in foods but depends upon the type of food. Determination of physiological functions in Y. enterocolitica which are regulated by quorum sensing and their relation to the production of AHLs in foods need to be further assessed.     

Molecular characterization of beta-lactamase genes blaA and blaB of Yersinia enterocolitica biovar 1A

The beta-lactamase genes blaA and blaB were detected by PCR amplification in strains of Yersinia enterocolitica biovar 1A isolated from India, Germany, France and the USA. Both genes were detected in all strains. Polymerase chain reaction-restriction fragment length polymorphism revealed genetic heterogeneity in blaA but not in blaB. Cluster analysis of blaA restriction profiles grouped the strains into three groups. The blaA gene of Y. enterocolitica biovar 1A showed a high degree of sequence homology to that of Y. enterocolitica 8081 (biovar 1B) and Y. enterocolitica Y-56 (biovar 4), whereas homology was low with class A beta-lactamase genes of other members of the family Enterobacteriaceae. The pI 8.7 of enzyme Bla-A of Y. enterocolitica biovar 1A was similar to that of biovars 2, 3 and 4. The enzyme Bla-B focused at 6.8 and 7.1, indicating that biovar 1A strains produced a 'B-like' enzyme. This is the first study to have investigated the genetic heterogeneity of the beta-lactamase genes of Y. enterocolitica.     

The serodiagnosis of human infections with Yersinia enterocolitica and Yersinia pseudotuberculosis

The techniques of sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) and immunoblotting were evaluated for the serodiagnosis of human infections with Yersinia enterocolitica and Yersinia pseudotuberculosis. Lipopolysaccharide (LPS) was prepared from strains comprising four serogroups of Y. enterocolitica and five serogroups of Y. pseudotuberculosis, tested against 200 sera submitted to the Laboratory of Enteric Pathogens for routine serodiagnosis, and shown to contain antibodies to Yersinia LPS by agglutination. Forty four sera were found to contain antibodies that bound to one of the LPS preparations used in the immunoassay. Thirty five of the sera contained antibodies to the LPS of Y. enterocolitica O3, whilst three contained antibodies to the LPS of Y. enterocolitica O5, 27 and Y. enterocolitica O9 LPS respectively. Two sera had antibodies to the LPS of Y. pseudotuberculosis II and a single serum contained antibodies to Y. pseudotuberculosis IV. The SDS-PAGE-immunoblotting procedure described proved to be a reliable procedure for the serodiagnosis of infections with Y. enterocolitica and Y. pseudotuberculosis.