A simplified polymerase chain reaction (PCR) assay was developed for fast and easy screening of mycoplasma mastitis in dairy cattle. Species of major mycoplasma strains [Mycoplasma (M.) bovis, M. arginini, M. bovigenitalium, M. californicum, M. bovirhinis, M. alkalescens and M. canadense] in cultured milk samples were detected by this simplified PCR-based method as well as a standard PCR technique. The minimum concentration limit for detecting mycoplasma by the simplified PCR was estimated to be about 2.5 x 10(3) cfu/mL and was similar to that of the standard PCR. We compared the specificity and sensitivity of the simplified PCR to those of a culture method. Out of 1,685 milk samples cultured in mycoplasma broth, the simplified PCR detected Mycoplasma DNA in 152 that were also positive according to the culture assay. The sensitivity and specificity of the simplified PCR were 98.7% and 99.7%, respectively, for detecting mycoplasma in those cultures. The results obtained by the simplified PCR were consistent with ones from standard PCR. This newly developed simplified PCR, which does not require DNA purification, can analyze about 300 cultured samples within 3 h. The results from our study suggest that the simplified PCR can be used for mycoplasma mastitis screening in large-scale dairy farms.
Animals ; Cattle ; Colony Count, Microbial/veterinary ; DNA, Bacterial/chemistry/genetics ; Disease Outbreaks/prevention & control/veterinary ; Female ; Mastitis, Bovine/diagnosis/*microbiology ; Milk/cytology/*microbiology ; Mycoplasma/genetics/*isolation & purification ; Mycoplasma Infections/diagnosis/microbiology/*veterinary ; Polymerase Chain Reaction/veterinary

This study evaluated how predictive the California Mastitis Test (CMT) is for sub-clinical mastitis under tropical smallholder dairy production conditions in Kenya. It intended to establish whether the CMT usage could be contributing to misdiagnosis and consequent mistreatment with animal drugs resulting in residue problems. Milk samples (n = 239) were aseptically collected from lactating cows in the Rift Valley of Kenya and tested using the CMT, somatic cell counts (SCC) and bacterial culture. The samples were also screened for violative drug residues using the commercial delvo test and compared to the milks mastitic status for possible association. There was a numerical but non-significant (p > 0.05) difference evident in the frequencies observed using the three different mastitis indicators. The prevalent bacterial species isolated from mammary glands with subclinical mastitis were Staphylococcus aureus (45.6%), coagulase-negative Staphylococci (13.0%), Streptococci (11.7%) and Escherichia coli 5.9%. There was an overall poor but significant (p < 0.05) correlation between the CMT and the violative antimicrobial residues in samples from all quarters, infected and non-infected respectively. The results suggest that the CMT use amongst the smallholder dairy sector as a mastitic indicator may not be a risk factor in violative antimicrobial residues problems in milk.
Animals ; Anti-Infective Agents/*chemistry ; Cattle ; Cell Count/veterinary ; Cross-Sectional Studies ; Dairying ; Drug Residues/*chemistry ; Escherichia coli/isolation&purification ; Female ; Kenya ; Mastitis, Bovine/*diagnosis/microbiology ; Milk/chemistry/cytology/*microbiology ; Reagent Kits, Diagnostic/standards/*veterinary ; Rural Population ; Staphylococcus aureus/isolation&purification ; Streptococcus/isolation&purification ; Tropical Climate