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Seventy-seven cases of equine abortion from 49 Hungarian farms that occurred between 1998 and 2000 were investigated for the presence of chlamydiae by immunohistochemistry, PCR and/or MZN staining. Evidence of the presence of these bacteria was obtained in 64 cases (83.1%) from 41 (83.7%) different farms. Partial ompA gene sequencing of PCR products revealed that the agent was Chlamydophila psittaci. Based on the findings of microbial diagnosis, pathology and case history, chlamydial infection was considered to be the most likely cause of abortion in at least 11 (14.3%) cases. In the remaining 53 Chlamydophila-positive cases, either other bacterial and viral agents (n = 22 or 28.6%) as well as non-infectious factors (n = 14 or 18.2%) were identified as more probable primary causes of disease, or the role of chlamydiae remained unclear because lesions in fetuses and fetal membranes were absent (n = 17 or 22.1%). When chlamydial antigen was detected in aborted equine placental tissue using immunohistochemistry it was seen only in the chorionic epithelial cells, but not in other parts of the fetal membranes nor in any of the fetal tissues. In conclusion, chlamydial infection of the genital tract should be considered a possible factor in equine reproductive disorders. 相似文献
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Elschner M Schrader C Hotzel H Prudlo J Sachse K Eichhorn W Herbst W Otto P 《Veterinary microbiology》2005,105(2):123-129
A total of 26 rotavirus positive faecal samples of diarrhoeal foals, and 8 equine rotavirus isolates were examined. Viral RNA patterns were generated, G typing was performed by PCR, and a P[12]-specific DNA probe was developed for P typing. Furthermore, five equine rotavirus isolates were sequenced in the genomic regions coding for VP7 and part of VP4. Rotaviruses of genotype G3 P[12] were found in 22 faecal samples and G14 P[12] type could be found in 4 faecal samples. These findings confirm that in Germany G3 P[12] is the predominating type of equine rotaviruses. 相似文献
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Tissue samples from lungs, pulmonary lymph nodes, large intestine, and uteri of 14 wild boar bagged at a seasonal hunt were examined for the presence of chlamydiae, mycobacteria and mycoplasmas. Nested PCR detected chlamydial DNA in 57.1% of the animals, predominantly in the lung. DNA sequencing identified Chlamydophila psittaci as the predominant species, but Chlamydophila abortus and Chlamydia suis were also encountered. Immunohistochemical staining of tissue sections confirmed the presence of typical chlamydial inclusions in lungs and uteri. While the role of Chlamydiaceae as pathogens in wild boar has yet to be established, the present findings revealed a possible wildlife reservoir of these bacteria. 相似文献
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For molecular biological detection of Mycobacterium avium subsp. paratuberculosis (MAP), PCR methods with primers targeting different regions specific for MAP are used worldwide. However, some uncertainties exist concerning the specificity of certain target regions and the sensitivity. To identify the methods which are best suited for diagnostics, 8 single-round and 5 nested PCR systems including 12 different primer pairs based on IS900 (9 x), ISMav2 (1x), f57 (1x), and locus 255 (1x) sequences were compared regarding their analytical sensitivity and specificity under similar PCR conditions. Reference strains and field isolates of 17 Mycobacterium species and subspecies, 16 different non-mycobacterial bovine pathogens and commensals were included in this study. Single-round PCR resulted in a detection limit of 100 fg to 1 pg, and nested PCR in 10 fg or below. Depending on the specific primer sequences targeting IS900, false positive results occurred with one of the five single-round and two of the four nested PCR systems. This also applied to the single-round PCR based on ISMav2 and the nested PCR based on f57. A high number of non-specific products were primarily detected for the single-round PCR assay based on ISMav2, but also for a single-round PCR targeting the IS900 and the locus 255. In conclusion, stringent selection of IS900-specific primers ensures that IS900 remains a favourite target sequence for amplification of MAP specific loci. The studied PCR systems based on f57, and locus 255 can also be recommended. Revision of ISMav2 primers is necessary. Single-round PCR systems are very reliable. Nested PCR assays were occasionally disturbed by contaminations, thus bearing a risk for routine diagnostics. 相似文献
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Otto PH Ahmed MU Hotzel H Machnowska P Reetz J Roth B Trojnar E Johne R 《Veterinary microbiology》2012,156(1-2):8-15
Avian rotaviruses (AvRVs) represent a diverse group of intestinal viruses, which are suspected as the cause of several diseases in poultry with symptoms of diarrhoea, growth retardation or runting and stunting syndrome (RSS). To assess the distribution of AvRVs in chickens and turkeys, we have developed specific PCR protocols. These protocols were applied in two field studies investigating faecal samples or intestinal contents of diseased birds derived from several European countries and Bangladesh. In the first study, samples of 166 chickens and 33 turkeys collected between 2005 and 2008 were tested by PAGE and conventional RT-PCR and AvRVs were detected in 46.2%. In detail, 16.1% and 39.2% were positive for AvRVs of groups A or D, respectively. 11.1% of the samples contained both of them and only four samples (2.0%) contained rotaviruses showing a PAGE pattern typical for groups F and G. In the second study, samples from 375 chickens and 18 turkeys collected between 2009 and 2010 were analyzed using a more sensitive group A-specific and a new group D-specific real-time RT-PCR. In this survey, 85.0% were AvRV-positive, 58.8% for group A AvRVs, 65.9% for group D AvRVs and 38.9% for both of them. Although geographical differences exist, the results generally indicate a very high prevalence of group A and D rotaviruses in chicken and turkey flocks with cases of diarrhoea, growth retardation or RSS. The newly developed diagnostic tools will help to investigate the epidemiology and clinical significance of AvRV infections in poultry. 相似文献
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Dennis Rubbenstroth Helmut Hotzel Johannes Knobloch Lydia Teske Silke Rautenschlein Martin Ryll 《Veterinary microbiology》2011,147(1-2):103-112
Riemerella columbina (RC) and Riemerella anatipestifer (RA) belong to the genus Riemerella within the family Flavobacteriaceae. While RA is a well-described pathogen of waterfowl and other avian species, only little is known about RC. Previous work reporting the isolation of RC from internal organs of clinically diseased pigeons suggested a potential pathogenic role in this avian species. In this study we examined pharyngeal swabs collected from pigeons and found RC to be widely distributed also among healthy birds. Further characterization of 81 RC-isolates revealed several atypical strains, which differed from all previously described RC-isolates by the lack of aesculin-hydrolysis activity (17 isolates) or by expression of yellow or orange pigmentation (6 isolates). Sequence analysis of the 16S rRNA and outer membrane protein A (ompA) gene supported the affiliation of these strains to the species RC. Aesculin-hydrolysis negative isolates were found to be biochemically indistinguishable from RA. We demonstrated that bacterial fingerprinting using matrix assisted laser desorption/ionisation–time of flight mass spectrometry (MALDI–TOF MS) analysis is useful for the identification and differentiation of RC and RA. 相似文献
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Petra Mbius Isabel Fritsch Gabriele Luyven Helmut Hotzel Heike Khler 《Veterinary microbiology》2009,139(3-4):398-404
Mycobacterium avium subsp. paratuberculosis (MAP) strains with two new IS900 restriction fragment length polymorphism (RFLP) BstEII types intermediate suspected to belong to the MAP Type III group were isolated from migrating sheep in Germany. Such strains have only been sporadically identified in a few studies. For a better understanding of the genomic diversity of MAP with regard to specific host associations, geographic origin, and the discussed classification into Type I, Type II and Type III, these isolates were further characterized.Using IS900-RFLP, the isolates showed unique fingerprint patterns after BstEII-, PstI-, PvuII- and BamHI-digestion which had not been published before. Additionally, using gyrB-PCR-restriction endonuclease analysis (PCR/REA) and mycobacterial interspersed repetitive unit (MIRU)-PCR, the two strains showed differences to known patterns of the Type I as well as the Type II group. Unique genotypes were also obtained with multilocus short sequence repeat (MLSSR) sequencing and MIRU-variable-number tandem-repeat (VNTR) typing.As expected, genomic profiles identical to the Type I and different from the Type II group were detected by IS1311-PCR/REA, IS1311 sequencing as well as by Large Sequence Polymorphism analysis (LSPA 8, 17, 20, 4-II, and 18).In addition to distinct growth characteristics, the unique genotypes of the studied sheep strains support their affiliation to the assumed third group within the MAP subspecies and suggest the existence of different genotypes within this Type III group. The results could serve as further evidence that Type I and Type III groups are more closely related to each other than to the bovine Type II group. 相似文献
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Sachse K Laroucau K Vorimore F Magnino S Feige J Müller W Kube S Hotzel H Schubert E Slickers P Ehricht R 《Veterinary microbiology》2009,135(1-2):22-30
The avian and human pathogen Chlamydophila (C.) psittaci represents a genetically heterogeneous species. To facilitate epidemiological surveys, more rapid yet highly specific molecular tests are needed. Currently used typing methods, i.e. serotyping and PCR-RFLP, have only limited sensitivity and are incapable of covering the wide spectrum of naturally occurring types of C. psittaci strains. In the present study, a new DNA microarray assay based on the ArrayTube (AT) technology was used to genotype C. psittaci in 98 isolates and 23 clinical tissue samples. The present array carries 35 oligonucleotide probes derived from variable domains 2 and 4 of the ompA gene. The assay proved highly sensitive, allowing correct genotyping of DNA from 2 inclusion-forming units. The results of DNA microarray genotyping of cultured strains proved highly concordant with the data from PCR-RFLP typing and serotyping. Sequencing of the ompA gene served as the reference test to verify the accuracy of AT genotyping results. In 15 instances (15.3%), strains were successfully typed by the AT assay, while serotyping and/or PCR-RFLP genotyping failed to produce unambiguous results. Eleven of these samples were ompA sequenced to confirm the AT findings. In addition to the currently accepted nine ompA genotypes, the microarray test was shown to recognise new provisional genotypes, such as Mat116 and YP84. In conclusion, the new AT assay proved to be suitable for rapid, sensitive and reproducible genotyping of C. psittaci strains and can be recommended for routine diagnosis. 相似文献
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The species Campylobacter (C.) fetus is divided into the subspecies venerealis and fetus, which differ in epidemiology and clinical importance. The differences between these subspecies make an accurate distinction essential. Differentiation of C. fetus by traditional microbiological methods is only based on two reactions (tolerance to glycin, Na selenite reduction), in which C. fetus ssp. venerealis reacts negatively. However, the value of both reactions is limited. We used a specific PCR-based assay for identifying and differentiating the two C. fetus subspecies, which was recently developed by HUM et al. (1997). In this assay, a 764 bp amplicon is produced using primers MG3F and MG4R for both subspecies of C. fetus. In contrast to HUM et al. (1997), this amplicon was approximately 200 bp smaller. This discrepancy can't be explained. Afterwards, the primers VenSF and VenSR are used for differentiation. The identification of the sub-species venerealis is based on the presence of a 142 bp amplicon, which is not formed with subspecies fetus. The type strains of both C. fetus subspecies were used as positive controls. Non-specific reactions were not observed. In this PCR assay, 73 field strains were investigated (among them 24 C. fetus ssp. veneralis, 26 C. fetus ssp. fetus). In these investigations, the method has proved its diagnostic suitability. The results of the traditional microbiological differentiation of the C. fetus field strains could be confirmed by the PCR assay. In future, the traditional phenotypic characterization of C. fetus subspecies remains indispensable, but this PCR assay constitutes a valuable method for the confirmation of these results. 相似文献