Assessment of Bovine Mastitis in Lactating Cows, and Characterization of Staphylococcus Aureus from the Cows and Dairy Products in West Showa Zone, Ethiopia
| dc.contributor.advisor | Asnake Desalegn | |
| dc.contributor.author | Negassa Feyissa | |
| dc.date.accessioned | 2025-08-17T22:39:02Z | |
| dc.date.available | 2025-08-17T22:39:02Z | |
| dc.date.issued | 2024-06 | |
| dc.description.abstract | Mastitis, to which Staphylococcus aureus is one of the main causative agents, is the costly mammary disease which adversely affects dairy industry worldwide in general and in developing countries in particular. S. aureus is a Gram-positive coccus resides on the skin and mucus linings, and associated with various diseases in animal and human. The virulence factors of S. aureus associated with disease causing potential and/or survive in the harsh melena include different enzymes and toxin productions, antibiotic resistance, and survival in dry conditions and relatively high concentration of salt. A cross-sectional study with objectives of determining the prevalence of mastitis in lactating cows, and assessing the prevalence and virulence factors of S. aureus isolated from the lactating cows, and bulk milk, and milk products (yogurt and cheese) was, conducted from May 2020 to March 2021 in West Showa Zone of Oromia, central Ethiopia. Stratified random sampling method was applied to select the districts. The PAs, the farm (herds), and individual sampling units were selected randomly. Mastitis was diagnosed physically by observations and palpation for clinical mastitis, and by California Mastitis Test (CMT) method for subclinical mastitis. Teat milk from the lactating cows, bulk milk, yogurt, and cheese samples were collected for bacterial isolation. Farmers and milk product retailers were interviewed using structured questionnaire to assess the hygienic milk collection and storage practices in the study area. The biological samples were aseptically collected, labeled, and were transported under cold chain to the Zoonosis and Food Safety Research Laboratory of Ambo University for bacterial isolation. S. aureus was selectively isolated on a manitol salt agar at 37°C for 24 hours, and presumptively identified by growth and rapid manitol fermentation within 24 hours, Gram staining and observation under microscope, and catalase and coagulase tests. Finally, Matrix-Assisted Laser Desorption/Ionization Time-of-Flight (MALDI-ToF) test was used to confirm the species. The enzyme production ability of the isolates was determined using heated plasma agar for staphylokinase, butterfat-supplemented brain-heart infusion agar for lipase, and skimmed milk agar for protease. The enterotoxin genes, resistant genes (blaZ, and mecA), and thermo nuclease gene (nuc) were assessed using a specific primer-based conventional polymerase chain reaction (PCR). In addition, the antibiotic susceptibility pattern of the isolates was determined using the disc diffusion method on Muller Hinton agar. The data obtained from the field assessment, laboratory experiment, and questionnaire surveys were analyzed using SPSS version 20 and R-statistical software. . The data were summarized using frequencies‘ and percentage, and presented in graphs and tables. The risk-factors associated with the prevalence of mastitis and S. aureus , and virulence factors of S. aureus, were analysed and interpreted by using chi-square test, logistic regression, t-test, and ANOVA at 95% confidence interval (C.I.) with p<0.05 significance levels. Accordingly, 258 lactating cows were diagnosed for mastitis and 97 (37.6%) of them were mastitis-positive. The prevalence of mastaitis was found significantly increased with the advancement of lactation stage of the cows, in those milked with unwashed hands and unwashed udders before milking, and in cows infested by ticks when compared with early lactation stage, those milked with washed hands and udder before milking, and tick free cows respectively (p<0.05). Subclinical mastitis was significantly more prevalent than clinical mastitis (p<0.05). The result of the study also disclosed that 94 (15.36%) of 612 samples were positive for S. aureus. The isolation rate of S. aureus was significantly higher in mastitis-positive cows (35.05%) than in mastitis-negative cows (5.59%) (p<0.05). The prevalence of S. aureus was also significantly higher in cows reared in farms with large number of lactating cows and tick-infested cows than in cows of farms with small number of lactating cows and in tick free cows, respectively (p<0.05). In addition, 354 bulk milk and milk product samples were collected and the bacterium was significantly more frequent in bulk milk (20.3%) and cheese (14.9%) than in yogurt (4.8%). Moreover, 71 (77.2%), 51 (55.4%), and 62 (67.4%) of 92 S.aureus isolates were positive for protease, staphylokinase and lipase respectively. The PCR result revealed that the S.aureus isolates harbored sea, seb, and tsst-1 genes at rate of 12/24 (50%), 3/24 (12.5%), and 4/24 (16.6%) respectively. The S. aureus isolates were resistant to penicillin(100%), tetracycline (100%), oxy-tetracycline (100%), amoxicillin (100%), ampicillin (83.3%), and to norfloxacin (75%). Of the 92 isolates tested 12(13%) were found resistant to oxacillin (1 μg) and were considered MRSA. The PCR test of the resistant genes (mecA and blaZ) revealed that 10/44 (22.73%) were mecA positive; but none of them harbored the blaZ gene. In addition, all (100%) of the 34 isolates tested for the thermonuclease (nuc) gene were positive to the gene. The questionnaire survey also indicated that all of the 140 respondents milked the animals manually. More than 50% of them have not washed their hands before milking; have not used the udder drying clothes; and/or used it for more than one cows; used plastic utensils for collecting and storing milk and milk products; and have not smoked the utensils. Therefore, it is concluded that mastitis was prevalent in the study area, and S. aureus might be the major cause of the disease. In addition, the milk and fermented milk products might not be safe for human consumption. Moreover, the unhygienic milking practices and tick infestation were one of the factors playing role in the prevalence of mastitis in the study area. The milk collection and storage practices might exacerbate the cross contamination of the dairy products by pathogens from the miller‘s hand, utensils and/or the environment so that the source of the S. aureus could be the animals, the handlers or the utensils. The presence of the enterotoxin genes in the bacterium is indicative for the food intoxications caused by the toxins. Furthermore, different enzymes producing ability of the isolates, summed up with resistance to commonly used antibiotics revealed that the S. aureus isolates are highly virulent to cause serious and incurable infections. The availability of the resistant genes (mecA) particularly in the isolates from milk and milk products can be evident for the possible transfer of the gene to other bacterium if co-infection occurs or to the enterobactriacae found in the GIT. Therefore, other mastitis causing agents and their antibiogram characteristics and virulence factors, and the types of MRSA distributed in the area should be studied. In addition, in order to prevent and control the disease and cross contamination of the dairy product, the animal and dairy product handlers should be trained about the hygienic milking practices and food handling methods. | |
| dc.identifier.uri | https://etd.aau.edu.et/handle/123456789/6918 | |
| dc.language.iso | en_US | |
| dc.publisher | Addis Ababa University | |
| dc.subject | Lacattin Cows | |
| dc.subject | Mastitis | |
| dc.subject | Milk and Milk Products | |
| dc.subject | S. Aureus | |
| dc.subject | Virulence Factors | |
| dc.title | Assessment of Bovine Mastitis in Lactating Cows, and Characterization of Staphylococcus Aureus from the Cows and Dairy Products in West Showa Zone, Ethiopia | |
| dc.type | Thesis |