MOLECULAR CHARACTERIZATION OF ANTIMICROBIAL RESISTANCE AND VIRULENCE GENES IN MILK WITH BOVINE Thesis Ref. No.____________ ADDIS ABABA UNIVERSITY STAPHYLOCOCCUS AUREUS ISOLATED FROM SUBCLINICAL MASTITIS IN CENTRAL ETHIOPIA
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Date
2019-06
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Abstract
Staphylococcus aureus (S.aureus) is one of the predominant causative agents of mastitis
in dairy herds. Mastitis has a negative impact through economic losses in the dairy sector
across the globe. A cross sectional study design was conducted from November 2018 to
May 2019 with the objectives to determine prevalence of bovine subclinical mastitis
,S.aureus and antimicrobial resistance profile and detect antimicrobial resistance and
virulence genes in S.aureus isolated from milk with bovine subclinical mastitis in Central
Ethiopia. A total of 265 lactating exotic zebu cross breed dairy cows were screened for
bovine sub clinical mastitis using California Mastitis Test (CMT). Different biochemical tests
and polymerase chain reaction (PCR) were used for the isolation and identification of
S.aureus isolates. PCR was performed for detection of antimicrobial resistance genes;
methicillin resistance (mecA) and erythromycin resistance (ermA, ermC and mrsA) genes
and virulence genes (clumping factor A (clfA), enterotoxin A (sea), exfoliative toxin A (eta),
beta-hemolysin (hlb), toxic shock syndrome toxin-1(tst-1) and intracellular adhesion D
(icaD)). Out of 265 lactating dairy cows screened, 49.06% (n=130) of them were positive
for subclinical mastitis. Out of 130 CMT positive samples,76.92% (n=100) of milk samples
were positive for S.aureus and subjected to nine antimicrobials agents for antimicrobial
resistance test. A large percent of the isolates were resistant to penicillin (93%) followed
by ampicillin (87%) and tetracycline (75%). Sixty eight PCR confirmed S.aureus isolates
were obtained from 100 phenotypic positive S.aureus isolates and from this, 17.64% (n=12)
samples were contained the mecA gene. No amplification was observed for erythromycin
resistance (ermA, ermC, and msrA) and virulence (tsst-1, sea, icaD and hlb) genes. The
frequency of clfA and eta genes in S.aureus isolates were 25% (n=17) and 22.05% (n=15),
respectively. The result obtained in this study suggests that poor management practices are
responsible for the increase in S.aureus isolation. The high resistance of S.aureus to
antimicrobials contribute in bovine mastitis in these farms may cause health problems in the
community consuming raw milk purchased from these farms.
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Keywords
Antimicrobial resistance gene, Bovine Subclinical Mastitis, Staphylococcus aureus