Evaluation of cultural methods and selective media for the isolation of <i>Campylobacter fetus</i> subsp <i>venerealis</i> from cattle

Hum, S.; Brunner, J.; McInnes, A. G.; Mendoza, Gloria; Stephens, June M.

doi:10.1111/j.1751-0813.1994.tb03385.x

Cited by 23 publications

(59 citation statements)

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“…venerealis à polimixina B foi semelhante aos resultados citados por Hum et al (1994). Estes autores sugerem que este fato possa ser responsável pelas limitações das técnicas de diagnóstico da CVB através de isolamento, uma vez que a maioria dos meios de transporte e cultura possui a polimixina B em sua composição.…”

Section: Methodsunclassified

Susceptibilidade antimicrobiana de Campylobacter fetus subsp. venerealis isolado de bovinos

et al. 2005

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Venereal campylobacteriosis is associated with infection of Campylobacter fetus subsp. fetus and Campylobacter fetus subsp. venerealis. The etiological agent is transmitted by natural bull breeding or artificial insemination using contaminated semen. The present study aimed to determine the in vitro susceptibility of C. fetus subsp. venerealis isolates to antimicrobial drugs generally used in clinical and semen treatment. Reference strains of C. fetus subsp. fetus and C. fetus subsp. venerealis and 21 C. fetus isolates were tested. The susceptibility test was performed by using the modified KirbyBauer diffusion disc method. C. fetus subsp. fetus reference strain was resistant to lincomycin and penicillin. C. fetus subsp. venerealis was susceptible to all antimicrobial tested, with exception to C.C. KrewerDnalidixic acid. C. fetus subsp. venerealis samples were sensible to amikacin, ampicillin, cefalotin, streptomycin, gentamycin, penicillin and tetracycline. Drug resistance was observed on 42.86% of lincomycin, 4,76 % of enrofloxacin, and 100% to nalidixic acid. In addition 4.76% of the isolates showed intermediate susceptibility to enrofloxacin, neomycin, polimixin B and 9.52% to lincomycin. The susceptibility of C. fetus isolates to antimicrobial drugs commonly used in clinical and semen treatment was demonstrated.INDEX TERMS: Campylobacteriosis, cattle, semen, antibiomicrobials.

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Section: Methodsunclassified

Susceptibilidade antimicrobiana de Campylobacter fetus subsp. venerealis isolado de bovinos

et al. 2005

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“…The ELISA is known to produce false-positive and false-negative results, and a high percentage of C. fetus subsp. venerealis strains are susceptible to polymyxin B, an antibiotic used in all Campylobacter selective media and transport enrichment media (TEM) (20,26). Campylobacter colonies from preputial scrapes and vaginal mucus are visible within 48 h in a microaerobic environment, and the slow-growing C. fetus subsp.…”

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confidence: 99%

“…The diagnosis of infection is by the direct isolation of the causative agent by selective culture from semen, preputial smegma, or vaginal mucus (20,31) or through the detection of an immune response in cervico-vaginal mucus by using an enzyme linked immunosorbent assay (ELISA) (23). Several methods for the collection of preputial smegma and vaginal mucus have been investigated in order to improve the reliability of selective culture-based diagnostic procedures or ELISA.…”

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confidence: 99%

“…Molecular methods such as PCR (22), amplified fragment length polymorphism (57), and pulsed-field gel electrophoresis (17,44) have been used to discriminate between the two Campylobacter fetus subspecies. However, PCR has not been routinely applied for the diagnosis of bovine venereal campylobacteriosis and field studies continue to rely upon either selective culture (20), ELISA (21), or DIFT (38).…”

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confidence: 99%

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Comparison of Culture and a Novel 5′ Taq Nuclease Assay for Direct Detection of Campylobacter fetus subsp. venerealis in Clinical Specimens from Cattle

McMillen¹,

Fordyce²,

Doogan³

et al. 2006

J Clin Microbiol

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A Campylobacter fetus subsp. venerealis-specific 5 Taq nuclease PCR assay using a 3 minor groove binder-DNA probe (TaqMan MGB) was developed based on a subspecies-specific fragment of unknown identity (S. Hum, K. Quinn, J. Brunner, and S. L. On, Aust. Vet. J. 75:827-831, 1997). The assay specifically detected four C. fetus subsp. venerealis strains with no observed cross-reaction with C. fetus subsp. fetus-related Campylobacter species or other bovine venereal microflora. The 5 Taq nuclease assay detected approximately one single cell compared to 100 and 10 cells in the conventional PCR assay and 2,500 and 25,000 cells from selective culture from inoculated smegma and mucus, respectively. The respective detection limits following the enrichments from smegma and mucus were 5,000 and 50 cells/inoculum for the conventional PCR compared to 500 and 50 cells/inoculum for the 5 Taq nuclease assay. Field sampling confirmed the sensitivity and the specificity of the 5 Taq nuclease assay by detecting an additional 40 bulls that were not detected by culture. Urine-inoculated samples demonstrated comparable detection of C. fetus subsp. venerealis by both culture and the 5 Taq nuclease assay; however, urine was found to be less effective than smegma for bull sampling. Three infected bulls were tested repetitively to compare sampling tools, and the bull rasper proved to be the most suitable, as evidenced by the improved ease of specimen collection and the consistent detection of higher levels of C. fetus subsp. venerealis. The 5 Taq nuclease assay demonstrates a statistically significant association with culture ( 2 ‫؍‬ 29.8; P < 0.001) and significant improvements for the detection of C. fetus subsp. venerealis-infected animals from crude clinical extracts following prolonged transport.

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“…venerealis as culture has long been recognised as difficult and time-consuming (Clark, 1971;Eaglesome and Garcia, 1992;Hum et al, 1994a;Lander, 1990a;Monke et al, 2002). This is due to the limited survival of C. fetus subsp.…”

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confidence: 99%

Molecular diagnostic protocols for bovine genital campylobacteriosis using comparative genomics and virulence studies

Indjein¹

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Bovine genital campylobacteriosis is difficult to identify due to problems with the isolation of its causative agent Campylobacter fetus subspecies venerealis. To optimise isolation methods, three selective culture and three transport and enrichment media (TEMs) were evaluated using a pure C. fetus subsp. venerealis culture, with and without simulated field contamination by Pseudomonas aeruginosa. For optimal C. fetus subsp. venerealis isolation and growth, TEMs should be returned from the field within 48h of collection and plated within 4h onto Campylobacter selective agar plates incubated for 48-72h at 37°C under microaerophilic conditions. Longer transport or TEM incubation resulted in overgrowth of P. aeruginosa limiting C. fetus subsp. venerealis recovery.In order to establish an isolate collection, the parA real-time PCR assay was used to screen bull preputial secretions prior to culturing positive samples. A more sensitive real-time PCR assay was designed from the ISCfe1 insertion element and used to assess additional preputial samples, compared to the parA real-time assay. A proportion of real-time negative samples was also cultured. Fourteen cultures were obtained from either ISCfe1 (8/43; 18.6%) or parA (6/51; 11.8%) negative samples. ISCfe1 and parA sequences were obtained from isolates not confirmed by conventional methods suggesting that both parA and ISCfe1 may be found in organisms other than C. fetus subsp. venerealis. Molecular testing of cultures (n=84) confirmed the presence of these sequences in isolates identified as other Campylobacter spp. or Arcobacter-like organisms, while they were not always detected in all biochemically confirmed C. fetus subsp. venerealis isolates. The lack of specificity of all methods highlights the need for more reliable C. fetus subsp. venerealis-specific gene targets.The draft genomes of two C. fetus subsp. venerealis biovars aligned to C. fetus subsp. fetus genome provided 26 unique C. fetus subsp. venerealis regions. In order to identify new diagnostic targets, 13 putative conventional and one real-time PCR assays designed from contigs common to both C. fetus subsp. venerealis biovars were developed to screen the Campylobacter-like culture collection (n=84). Isolates biochemically identified as organisms other than C. fetus subsp.venerealis were positive on the new real-time and six conventional PCR assays, indicating that they were not suitable subspecies-specific markers. Seven assays detected isolates phenotyped as C. fetus subsp. venerealis except for two aerotolerant isolates recognized as C. fetus subsp. venerealis by iii other published methods. These seven gene targets could be further investigated for real-time PCR assay development as in silico BLAST analysis confirmed their conservation in the genomes of five of our C. fetus subsp. venerealis isolates. However, further bioinformatics analysis revealed that they were found in several C. fetus subsp. fetus strains, underlining that a pan-genomics study including multiple C. fetus genomes is ne...

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Evaluation of cultural methods and selective media for the isolation of Campylobacter fetus subsp venerealis from cattle

Cited by 23 publications

References 10 publications

Susceptibilidade antimicrobiana de Campylobacter fetus subsp. venerealis isolado de bovinos

Susceptibilidade antimicrobiana de Campylobacter fetus subsp. venerealis isolado de bovinos

Comparison of Culture and a Novel 5′ Taq Nuclease Assay for Direct Detection of Campylobacter fetus subsp. venerealis in Clinical Specimens from Cattle

Molecular diagnostic protocols for bovine genital campylobacteriosis using comparative genomics and virulence studies

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