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We conducted purification of filamentous temperature-sensitive protein Z of Streptococcus suis serotype 2 (S.suis 2) and measured its GTPase activity.The ftsz gene in the genome of the Chinese 05ZYH33 strain of S.suis 2 was successfully amplified using PCR,and then the ftsz gene was cloned into prokaryotic expression plasmid pET28a,and the recombinant plasmid pET28a-ftsz was transformed into E.coli BL21.After induction by IPTG,the isolated FtsZ protein was analyzed with SDS-PAGE.Then the recombinant protein was purified by Ni2+-NTA affinity chromatography.The rabbit serum was harvested after immunization with recombinant FtsZ protein,and was analyzed by indirect ELISA and Western blotting.The GTPase activity of FtsZ was measured with the malachite green method.Results showed that successfully constructed recombinant plasmid pET28a-ftsz and the recombinant protein with high purity was obtained;Western blot result indicated that FtsZ could react with the His-tag antibody and the rabbit serum;the polyclonal antibody titer of the rabbit serum reached 1 ∶ 13 107 200;FtsZ have GTPase activity.We successfully prepared S.suis 2 recombinant protein FtsZ having GTPase activity and high titer antiserum would be useful for the further study of S.suis 2 cell division mechanism.
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Objective To explore the interaction of streptococcus suis serotype 2 recombinant suilysin ( SLY ) with platelets, and provide the theoretical basis for clinic treatment of patients infected with S.suis.Methods The nickel column affinity chromatography was used to purify the recombinant SLY.The hemolytic acivity was identified by optical density before the platelets aggregation induced by a SLY was detected by a platelet aggregometer or electron microscope and the effect of aspirin on platelets aggregation was analyzed.The impact of wild type 05ZYH33 and sly-deficient mutant strainΔSLY on platelets of mice was compared to predict the interaction of the SLY with platelets in vivo.Results and Conclusion Hemolytic activity of recombinant SLY was 2000 hemolytic units( HU) and platelets aggregation was induced at 1 μg/ml.The aggregation can be inhibited by aspirin in 5 mmol/L.SLY can also increase the volume and reduce the amount of platelets in mice.
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ABSTRACT:The virulence genes ,hemolytic and antibiotic resistance characteristics from 19 isolated Streptococcus suis se‐rotype 2 in Anhui Province were investigated in this study .The PCR method was developed for detecting cps2J ,mrp ,ep f and sly gene;the plate method and micro‐ELISA were developed for detecting the hemolytic type and hemolytic titer ;the K‐B method was developed for detecting antibiotic resistance of 25 kinds of antibiotics .Results showed that there were 11 strains with the dominant virulence genotype of cps2J+ /mrp+ /ep f+ /sly+ accounting for 57 .9% .All strains were α‐hemolysis orβ‐hemolysis ,with hemolytic titer of 1∶4 to 1∶128 .Strains sensitive to rifapentine ,ceftazidime ,florfenicol and cefazolin was existed in high frequency with the sensitive rates for 84 .2% ,and the resistance to doxycycline ,tetracycline ,bacitracin and sul‐famethoxazole was existed in high frequency with the resistance rates for 82 .9% .The multi‐antibiogram typing was doxycyc‐line ,tetracycline ,bacitracin and sulfamethoxazole account for 63 .2% .In conclusion ,the distribution of virulence genes of S .suis 2 Anhui isolates was similar with that of domestic reported strains ,and there was some differences with that of over‐seas reported strains .CPS ,MRP ,EPF and SLY are important virulence factors and the incompleteness of sly gene had no effect to its haemolyticus .The multi‐antibiotic resistance of S .suis 2 Anhui isolates was serious .There is difference for anti‐biogram typing between S .suis 2 Anhui isolates and those in other areas .
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Streptococcus suis is considered worldwide as one of the pathogens of biggest health and economic impact in the swine industry. Among the serotypes described as zoonotic, serotype 2 is the most frequently isolated from diseased animals and humans in most countries. The study of the epidemiology of S. suis infections in Brazil is important and may help in the development of effective control measures. The aim of this study was to conduct a critical review of Brazilian literature, with support of the world literature, addressing the diagnosis of the agent and its prevalence in clinically ill animals and healthy carriers, especially regarding to the prevalence of the serotype 2 in the country.
Streptococcus suis é mundialmente considerado um dos patógenos de maior impacto sanitário e econômico na indústria suinícola. Dentre os sorotipos descritos como zoonóticos, o sorotipo 2 é o mais frequentemente isolado de animais e humanos doentes na maioria dos países. O estudo da epidemiologia das infecções por S. suis no Brasil é importante para a implantação de medidas efetivas de controle. O objetivo do presente trabalho foi realizar uma revisão crítica da literatura brasileira, com suporte da literatura mundial, abordando o diagnóstico do agente e sua prevalência em animais clinicamente doentes e portadores sadios, com destaque para a prevalência do sorotipo 2 no país.
Subject(s)
Animals , Swine Diseases , Food Safety , Noxae , Streptococcus suis , Prevalence , Health SurveillanceABSTRACT
To explore the molecular characteristics of Streptococcus suis serotype 2(ss2) isolated in Zhejiang province by deciding the variation loci and its variation frequency of Cps2J gene.The total DNA of 9 strains of ss2 isolated in Zhejiang province were extracted and amplifed by PCR. Then,the Cps2J fragments were cloned into plasmid carrier and completely sequenced after purification.Finally,the sequence results of all 9 ss2 isolates were compared with those obtained by other studies around the world.It was found that the open reading fragments of Cps2J in 9 SS2 isolates encoding 333 amino acids were 999 bp in length.Comparisons of this region among ss2 isolates revealed a similarity of between 98.8% and 99.9%, while the homology to ss1 strains varied between 56.8% and 57.0%.Our study shows the sequences of complete Cps2J segment are fairly stable and all these 9 ss2 strains of different sources possibly have the same evolutionary origin.
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A novel genomic island (GI) in Streptococcus suis serotype 2(SS2) was identified, which resided in the highly virulent strains but not in the hypo-virulent strains or avirulent strains of SS2 of the Chinese isolates. This newly discovered GI strain was designated as SSGI4 and its whole length of genome was 11 269 bps, sharing the typical properties of pathogenicity islands, such as the distinct G+C content, a mosaic architecture characteristics and the specificity for virulent isolates. There were 11 genes within SSGI4, in which some genes were putative cell surface protein genes and others were amino acid-binding protein genes. Our finding sheds light on the investigation of horizontal gene transfer in SS2 and their influence on pathogenicity.
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Objective To identify the presence of candidate pathogenicity island 89K DNA sequence of Streptococcus suis serotype 2 (SS2) strains isolated from patient in Zhejiang province. Methods Genes and DNA fragments were amplified by PCR, using specific primers, and three amplified fragments of the89K sequence were directly sequenced. The results were analyzed using software related to bioinformaties and epidemiology. Results 8 strains of SS2 all contained 89K sequence, cps2J and mrp virulent genes, and species-specific 16S rDNA. 3 amplified fragments of 89K candidate pathogenicity island of SS2 ZJ0501 were above 99% similar to SS2 strain identified from outbreaks in Jiangsu in 1998, and the gene fragment of coding DNA recombinant protein in the 89K sequence was highly homological with that of S. dysgalactiae and S. agalactiae. Conclusion In recent years SS2 strains isolaed from patients with clinical symptoms in Zhejiang province had been detected to have contained candidate pathogenicy 89K DNA fragment.
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To understand the enolase(eno)gene and its product in Streptococcus suis serotype 2(S.suis 2), bioinformatics was adopted to analyze the whole genome sequence of the Chinese strain 05ZYH33 of S.suis 2.A highly homologous eno gene was unveiled by the genome-wide mining.A pair of specific primers was designed for the eno,and the target DNA fragment of 1.3 kb was successfully amplified using the genomic template of 05ZYH33.Subsequently,eno gene was inserted into pMD18-T vector,and then subcloned into prokaryotic expression vector pET32a,generating a recombinant expression plasmid pET32a::eno.The resulting plasmid was confirmed by direct DNA sequencing and transformed into E.coli BL21(DE3) competent cells.Protein expression analysis showed that a 75 kD protein can be observed in 12% SDS-PAGE,indicating that the recombinant 6His-fused ENO protein can be produced in E.coli under the induction of IPTG.Westem-blot experiment demonstrated clearly it shares strong specific antigenicity. Moreover,ELISA result suggested that ENO can occur on the surface of 05ZYH33 strain.Together,our data ENO can function as a novel antigen,and may play pivotal roles in the severe infection of S.suis 2.