RESUMO
Campylobacter jejuni is among the most prevalent causes of human bacterial gastroenteritis worldwide. Domesticated animals and, especially, chicken meat are considered to be the main sources of infections. However, the contribution of surface waters and wildlife in C. jejuni transmission to humans is not well understood. We have evaluated the source attribution potential of a six-gene multiplex PCR (mPCR) method coupled with STRUCTURE analysis on a set of 410 C. jejuni strains isolated from environment, livestock, food and humans in central Europe. Multiplex PCR fingerprints were analysed using Subclade prediction algorithm to classify them into six distinct mPCR clades. A subset of C. jejuni isolates (70%) was characterized by multilocus sequence typing (MLST) demonstrating 74% congruence between mPCR and MLST. The correspondence analysis of mPCR clades and sources of isolation indicated three distinct groups in the studied C. jejuni population-the first one associated with isolates from poultry, the second one with isolates from cattle, and the third one with isolates from the environment. The STRUCTURE analysis attributed 7.2% and 21.7% of human isolates to environmental sources based on MLST and mPCR fingerprints, respectively.
Assuntos
Infecções por Campylobacter/epidemiologia , Infecções por Campylobacter/microbiologia , Campylobacter jejuni/genética , Gado/microbiologia , Animais , Campylobacter jejuni/isolamento & purificação , Microbiologia Ambiental , Europa (Continente) , Humanos , Tipagem de Sequências Multilocus , Filogenia , Reação em Cadeia da Polimerase , ZoonosesRESUMO
REASONS FOR PERFORMING STUDY: The prevalence of Taylorella equigenitalis infection in Slovenia is unknown and methods used to refine identification in these stallions are required. HYPOTHESIS: In diagnosis of T. equigenitalis, polymerase chain reaction (PCR) would have advantages over culture methods, especially in cases where small numbers of causal agent or intensive contamination of genital swabs are involved. METHODS: Culture method and PCR were used to examine a total of 980 genital swabs from the urethra and fossa urethralis of 245 stallions for the presence of the contagious equine metritis organism. RESULTS: Among 245 examined stallions, 225 (91.8%) were negative to T. equigenitalis by both methods. From the swabs of 17 stallions (6.9%) T. equigenitalis was isolated at first and/or second sampling. Swabs of 3 (13%) stallions were PCR positive but the isolation of T. equigenitalis failed. The rate of T. equigenitalis detection was higher with PCR than with the classic bacteriological examination. CONCLUSIONS AND POTENTIAL RELEVANCE: PCR protocol used in this study provided a specific, sensitive, and simple tool for rapid detection of T. equigenitalis. PCR is especially valuable in cases of intensive bacterial and fungal contamination of swabs where the isolation of T. equigenitalis usually fails.
Assuntos
Infecções por Bactérias Gram-Negativas/veterinária , Doenças dos Cavalos/diagnóstico , Reação em Cadeia da Polimerase/veterinária , Taylorella equigenitalis/isolamento & purificação , Animais , Contagem de Colônia Microbiana/métodos , Contagem de Colônia Microbiana/veterinária , DNA Bacteriano/análise , Infecções por Bactérias Gram-Negativas/diagnóstico , Infecções por Bactérias Gram-Negativas/epidemiologia , Doenças dos Cavalos/epidemiologia , Cavalos , Masculino , Reação em Cadeia da Polimerase/métodos , Prevalência , Sensibilidade e Especificidade , Eslovênia/epidemiologia , Fatores de TempoRESUMO
Cystic forms (also called spheroplasts or starvation forms) and their ability to reconvert into normal motile spirochetes have already been demonstrated in the Borrelia burgdorferi sensu lato complex. The aim of this study was to determine whether motile B. garinii could develop from cystic forms, not only in vitro but also in vivo, in cyst-inoculated mice. The cysts prepared in distilled water were able to reconvert into normal motile spirochetes at any time during in vitro experiments, lasting one month, even after freeze-thawing of the cysts. Motile spirochetes were successfully isolated from 2 out of 15 mice inoculated intraperitoneally with cystic forms, showing the infectivity of the cysts. The demonstrated capacity of the cysts to reconvert into motile spirochetes in vivo and their surprising resistance to adverse environmental conditions should lead to further studies on the role and function of these forms in Lyme disease.
Assuntos
Borrelia/citologia , Borrelia/fisiologia , Animais , Anticorpos Antibacterianos/sangue , Borrelia/patogenicidade , Infecções por Borrelia/imunologia , Grupo Borrelia Burgdorferi/citologia , Grupo Borrelia Burgdorferi/fisiologia , Ensaio de Imunoadsorção Enzimática , Rim/microbiologia , Camundongos , Movimento , Técnicas de Cultura de Órgãos , Esferoplastos/fisiologia , Bexiga Urinária/microbiologiaRESUMO
Inbred C57BL/6 mice were challenged with Borrelia afzelii, Borrelia garinii and Borrelia burgdorferi sensu stricto and tested for antigen-specific T-cell response in vitro. The sonicated preparations of in vitro grown spirochetes were capable of stimulating polyclonal proliferation and specific cell-mediated response, depending on duration of the cell culture. Murine splenocytes previously sensitized to B. burgdorferi sensu lato (s.l. ), but not those from control mice, could be induced for antigen-specific proliferation in vitro. Moreover, detectable cell-mediated response could be induced only with antigen preparations derived from a corresponding strain but not with those obtained from other Borrelia genospecies as revealed by the [(3)H]thymidine incorporation assay. The current study considers that the strict B. burgdorferi s.l. antigen-specific response may also be expected in infections in humans and contributes to the explanation of the frequently poor antibody- and cell-mediated immune response observed in patients diagnosed with Lyme disease.
Assuntos
Infecções por Borrelia/imunologia , Borrelia , Epitopos de Linfócito T/imunologia , Animais , Especificidade de Anticorpos , Antígenos de Bactérias/imunologia , Células Cultivadas , Ensaio de Imunoadsorção Enzimática , Feminino , Soros Imunes/imunologia , Imunidade Celular , Ativação Linfocitária , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Baço/citologia , Fatores de TempoRESUMO
Inbred C57bl/6 mice were challenged with high-passage Borrelia afzelii, Borrelia garinii and Borrelia burgdorferi sensu stricto and tested for antigen specific T-cell response in vitro. Sonicated preparations of washed spirochetes were potent cell activators, capable of stimulating polyclonal proliferation after 72h of culture while increasing the incubation time up to 120h provoked specific cell-mediated response. Isolated murine spleocytes previously sensitized to B. burgdorferi sensu lato but not those from control mice could be induced for antigen-specific proliferation in vitro, as revealed by [3H]thymidine incorporation assay, Moreover, in mice presensitized to B. burgdorferi sensu lato, detectable cell-mediated response could be induced only with antigen preparations derived from a corresponding strain but not with those obtained from other Borrelia genospecies. The current study emphasises that the B. burgdorferi antigen-specific response may also be expected in different genospecies infections in men.
Assuntos
Antígenos de Bactérias/imunologia , Infecções por Borrelia/imunologia , Borrelia , Animais , Anticorpos Antibacterianos/análise , Infecções por Borrelia/sangue , Relação Dose-Resposta Imunológica , Imunidade Celular , Ativação Linfocitária , Camundongos , Camundongos Endogâmicos C57BL , Especificidade da Espécie , Baço/imunologia , Fatores de TempoRESUMO
A Gram-negative, microaerophilic slender rod, measuring approximately 10 mum long and approximately 1 microm wide, isolated from the gastric mucosa of a cat and designated strain M50(T), was subjected to a polyphasic taxonomic study. Despite its apparent lack of helical coils, the organism showed a corkscrew-like motion by means of multiple sheathed flagella located at both ends of the cell and by a periplasmic fibril coiled around the body. Strain M50(T) grew preferably on biphasic culture plates or on very moist agar. Coccoid forms predominated in cultures older than 4 days as well as in growth obtained on dry agar plates. The strain grew at 37 degrees C, but not at 25 or 42 degrees C and exhibited urease, oxidase and catalase activities. On the basis of 16S rRNA gene sequence analysis, the novel isolate was identified as a member of the genus Helicobacter and showed about 98 to 99 % sequence similarity to Helicobacter felis, Helicobacter bizzozeronii, Helicobacter salomonis, Helicobacter cynogastricus and 'Candidatus Helicobacter heilmannii', five highly related species previously detected in the feline or canine gastric mucosa. Protein profiling of strain M50(T) using SDS-PAGE revealed a pattern different from those of other Helicobacter species of mammalian gastric origin. Additionally, the urease and HSP60 gene sequences of strain M50(T) were different from those of H. felis, H. bizzozeronii, H. salomonis, H. cynogastricus and 'Ca. H. heilmannii'. It is thus proposed that strain M50(T) (=LMG 23839(T)=CCUG 53816(T)) represents a novel species within this genus, for which the name Helicobacter baculiformis sp. nov. is proposed.
Assuntos
Doenças do Gato/microbiologia , Mucosa Gástrica/microbiologia , Infecções por Helicobacter/veterinária , Helicobacter/classificação , Animais , Técnicas de Tipagem Bacteriana , Gatos , Chaperonina 60/genética , Chaperonina 60/metabolismo , DNA Bacteriano/análise , Genes de RNAr , Helicobacter/genética , Helicobacter/isolamento & purificação , Infecções por Helicobacter/microbiologia , Dados de Sequência Molecular , Fenótipo , Filogenia , RNA Ribossômico 16S/genética , Análise de Sequência de DNA , Especificidade da Espécie , Urease/genética , Urease/metabolismoRESUMO
The susceptibilities of Helicobacter felis (15 strains), H. bizzozeronii (7 strains), and H. salomonis (3 strains) to 10 antimicrobial agents were investigated by determination of the MIC using the agar dilution method. No consistent differences were noticed between the different Helicobacter species, which were all highly susceptible to ampicillin, clarithromycin, tetracycline, tylosin, enrofloxacin, gentamicin, and neomycin, as demonstrated by low MICs. Higher MICs were obtained for lincomycin (up to 8 microg/ml) and spectinomycin (up to 4 microg/ml). Two H. felis strains showed a MIC of 16 microg/ml for metronidazole, suggesting acquired resistance to this antimicrobial agent.