Sensitive test for screening for <i>Staphylococcus aureus</i> in bovine mastitis by broth cultivation and PCR

Yamagishi, Norio; Jinkawa, Y; Omoe, Katsuhiko; Makino, Shinichiroh; Oboshi, Kenji

doi:10.1136/vr.161.11.381

Cited by 10 publications

(7 citation statements)

References 4 publications

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“…2). The enrichment of mastitic milk samples in the present study not only improved the sensitivity of PCR (Yamagishi et al 2007) but also helped to detect only live bacterial pathogens and eliminated the chances of detecting the dead bacteria contaminants, thereby avoiding the false positive results.…”

Section: Discussionmentioning

confidence: 91%

Evaluation of PCR test for detecting major pathogens of bubaline mastitis directly from mastitic milk samples of buffaloes

Kumar

2009

Trop Anim Health Prod

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Present study is aimed to evaluate the PCR test for detecting the major pathogens of bubaline mastitis (BM) directly from the mastitic milk samples of the buffaloes. Cell lysates of the enriched mastitic milk samples were directly used as template DNA in PCR and the reactivity of genus and/or species specific primers against the selected pathogens of BM was tested in individual simplex PCR assays. Out of the 60 mastitic milk samples tested 30 were positive for E. coli, 21 were positive for S. aureus, 4 were positive for S. agalactiae, 9 were positive for S. dysgalactiae and 7 were positive for S. uberis. Certain samples were positive for more than one pathogen thus indicating the mixed infection of the udder. Although certain mastitic milk samples reacted with Staphylococcus genus specific primers, they didn't react with S. aureus specific primers in PCR, which indicates the implication of other species of Staphylococcus in BM. The processing of mastitic milk samples was also simplified by eliminating the use of expensive reagents. The detection limits of PCR with the cell lysates are sensitive enough for PCR to be used as diagnostic tool for identifying the pathogens of BM.

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Section: Discussionmentioning

confidence: 91%

Evaluation of PCR test for detecting major pathogens of bubaline mastitis directly from mastitic milk samples of buffaloes

Kumar

2009

Trop Anim Health Prod

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“…Attempts to increase the sensitivity of detecting Staph. aureus have also been made by testing different enrichment procedures before bacteriological culturing with varying results (Thurmond et al, 1989;Sol et al, 2002;Yamagishi et al, 2007). In our study, neither enrichment of samples in nutrition broth nor processing by the Mastistrip method had any effect on the isolation of Staph.…”

Section: Discussionmentioning

confidence: 70%

An improved method to culture Staphylococcus aureus from bovine milk

Artursson

Nilsson-Öst

Waller

2010

Journal of Dairy Science

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Staphylococcus aureus is an important udder pathogen often associated with subclinical mastitis in dairy cows. Identification of Staph. aureus-positive udder quarters and cows is an important part of control programs to reduce spread of Staph. aureus within and between dairy herds. Therefore, accurate and easy-to-perform culturing methods of Staph. aureus in milk are needed. In the present study, 8 methods for isolation of Staph. aureus in bovine milk samples were investigated. The methods involved different culturing volumes, enrichment, incubation, and freezing processes as well as sedimentation and use of the Mastistrip cassette (SVA, Uppsala, Sweden). Three different sets of milk samples were collected, and 6, 5, and 4 methods were used in each subset of samples. Our results indicate an increased probability of detecting Staph. aureus in milk samples when a simple incubation step (37 degrees C for 18 h) without additives was included before culturing. Using this incubation method, the number of Staph. aureus-positive udder quarters and cows increased by 50 and 29%, respectively, compared with using the standard method of direct culturing of 10 microL of milk. The improved method may be especially useful for detection of low concentrations of Staph. aureus in milk; for example, when screening herds for Staph. aureus.

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“…aureus, might be present in very low numbers. 7,31 As an example, Staph. aureus occurred in the current study in 10 out of 40 samples in pure culture in numbers of 10 4 CFU/ml or fewer and was detected in only 2 of these samples.…”

Section: Discussionmentioning

confidence: 99%

Identification of pathogens in mastitis milk samples with fluorescent in situ hybridization

et al. 2013

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Abstract. Traditionally, the bacteriological examination of mastitis milk samples is performed by culture followed by biochemical tests on the cultured bacteria to allow identification of the causative pathogen. Depending on the species involved, this classic identification is time-consuming compared to other techniques such as fluorescent in situ hybridization (FISH), a culture-independent method that utilizes oligonucleotides (labeled with a fluorophore) that are specific to a string of target DNA/RNA. In the current study, the applicability of FISH was evaluated for the detection of mastitis pathogens directly in milk samples. To remove interfering lipids and proteins from mastitis milk samples prior to FISH, a previously published enzymatic treatment with savinase was evaluated. FISH was performed using oligonucleotides specific for Staphylococcus aureus, Streptococcus agalactiae, Streptococcus uberis, Enterococcus faecalis, Enterococcus faecium, Escherichia coli, and Trueperella (Arcanobacterium) pyogenes. The enzymatic pretreatment and the sensitivity of FISH were evaluated using spiked whole milk samples and mastitis milk samples with bacterial loads of less than 10 3 up to 10 8 colony-forming units (CFU)/ml. Bacteria were reliably detected in milk samples with bacterial numbers of 10 6 CFU/ml or higher. However, bacteria present in numbers below 10 6 CFU/ml were not detectable in all cases. The ability of FISH to identify mastitis-causing pathogens directly in milk samples, and therefore earlier than classical culture methods, can supplement the classic diagnostic procedures for mastitis milk samples.

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Sensitive test for screening for Staphylococcus aureus in bovine mastitis by broth cultivation and PCR

Cited by 10 publications

References 4 publications

Evaluation of PCR test for detecting major pathogens of bubaline mastitis directly from mastitic milk samples of buffaloes

Evaluation of PCR test for detecting major pathogens of bubaline mastitis directly from mastitic milk samples of buffaloes

An improved method to culture Staphylococcus aureus from bovine milk

Identification of pathogens in mastitis milk samples with fluorescent in situ hybridization

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