Investigation of the specificity and sensitivity of the speed mam colour ™ test in the diagnosis of important mastitis pathogens in cows

Mastitis, known as inflammation of the mammary gland, is a disease that is widely observed in dairy farming and causes the highest economic loss. Rapid diagnosis of the disease agents in field conditions, identification of the pathogens causing the disease and early initiation of treatment increases the chance of success and more effective use of antibiotics. The aim of this thesis is to investigate the reliability of Speed Mam ColourTM test in the diagnosis of Staphylococcus spp., Streptococcus spp., Streptococcus uberis and Escherichia coli, which are known as important mastitis pathogens in cows. Within the scope of the study, milk collected from healthy herds, which had given birth at least once, between the 5th and 200th days of lactation, healthy, without any clinical symptoms, without udder abnormalities, without antibiotic or nonsteroidal drug treatment until 15 days ago were used for the determination of the detection limit of the test, specificity and sensitivity studies. With milk samples contaminated with reference strains, 7 different dilutions were prepared for each target bacteria and each bacterial dilution was added to the microbiological culture in parallel with the test kit to control the dilution rates and left for incubation. The observation of 10-fold decreasing bacterial growth at the end of incubation showed that the dilution was performed correctly. The lowest concentration at which positive results were obtained with the test kit was determined as the detection limit of the kit. As a result of the tests performed accordingly, the detection limit results of the Speed Mam Colour™ kit at the end of 48 hours were 101 CFU/ml for Staphylococcus spp.; 103 CFU/ml for Streptococcus spp.; 103 CFU/ml for S. uberis; 101 CFU/ml for E. coli. In order to determine the sensitivity and specificity, the reference strain of each target bacterium was grown in nutrient broth liquid medium by culture method, the concentrations were adjusted according to McFarland 0.5 and different concentrations (>103 CFU/ml) above the detection limit specified in the Speed Mam Colour ™ package insert were diluted in 5 tubes containing healthy milk and the different concentrations obtained were randomly distributed to 25 tubes containing 4 ml of healthy milk by taking 1 ml each from these tubes. In this way, 25 contaminated 25 healthy milk samples obtained for each target were tested by microbiological culture method and Speed Mam Colour ™. The specificity of the Speed Mam Colour™ test was 100% for Staphylococcus spp., Streptococcus spp., S.uberis and E. coli , while the sensitivity was 100% for Staphylococcus spp., 92% for Streptococcus spp., 92% for S.uberis and 96% for E. coli . The positive predictive value (PPV) value was 100% for Staphylococcus spp., Streptococcus spp., S. uberis and E. coli, while the negative predictive value (NPV) value was 100%, 93%, 93% and 96%, and the accuracy (Ac) value was 100%, 96%, 96% and 98%, respectively. In the other stage of the study, which was the verification of the test, CMT was applied to the cows considering the above mentioned criteria and 318 milk samples showing positivity according to CMT results were collected. Microbiological culture method, which is accepted as the gold standard method, was used to compare 318 CMT positive milk samples with subclinical mastitis collected for the verification of Speed Mam Colour™ test. Milk samples tested by culture method were compared with the bacterial results analysed in the Speed Mam Colour™ test. Detection of more than 3 colonies in the milk sample was considered as contamination; when 2-3 colonies were detected, it was defined as mixed infection. While 5 different combinations of target bacteria were obtained in the study of milk samples with subclinical mastitis, when the growth status of some target bacteria in Speed Mam Colour ™ and microbiological culture was examined, the specificity of the Speed Mam Colour ™ test was determined as Staphylococcus spp, S.uberis and E. coli was 100%, for Streptococcus spp. 95%, sensitivity was 96% for Staphylococcus spp. 96%, for Streptococcus spp. 83%, for S.uberis and E. coli 100%. PPV value was calculated as 100% for Staphylococcus spp., S.uberis and E. coli and 94% for Streptococcus spp. NPV values were 95%, 87%, 100% and 100% for the related bacteria, and Ac values were 98%, 90%, 100% and 100%, respectively. As a result, it was thought that the Speed Mam Colour™ test could be a test that can be used to diagnose important mastitis pathogens causing mastitis under field conditions.