Detection of Enterotoxigenic and Methicillin-Resistant Staphylococcus aureus Isolated from Raw Milk of Cows Reared in Erbil-Kurdistan Region
The present study was conducted to evaluate raw cow milk in Erbil Governorate; total of 150 milk samples were randomly collected from farms and milk collection centers from the 1st April to the end of June 2018, this includes; 75 samples directly from animals (Direct Milk), and 75 samples from bulk tank milk (BTM). The samples were subjected for determination of prevalence rate, enumeration, identification of Staphylococcus aureus, and screening for the occurrence of enterotoxigenic and methicillin-resistant S. aureus strains (MRSA). The study revealed that 46 (30.6%) of the total raw milk samples were positive for S. aureus this comprised 14 (18.6%) and 32 (42.6%) from DM and BTM samples, respectively. Nearly half of samples 22 (47.8%) [3 (21.4%) form DM and 19 (60.3%) form BTM] contained S. aureus at the level 103 – 104 CFU/ml, which is an unsatisfactory range for dairy production. Statistical analyses indicated that the prevalence and number of S. aureus were significantly higher in BTM than DM samples.
Microbiological and biochemical characterization was carried out for the isolated bacteria, 46 (30.6%) isolates identified as S. aureus, and all isolates were confirmed by species-specific primers targeting 16S rRNA and nuc genes through PCR assay.
Staphylococcal Enterotoxins (SEs) production of S. aureus isolates were tested by Staphylococcal Enterotoxin-Reversed Passive Latex Agglutination (SET-RPLA) from SEA to SED, and related staphylococcal enterotoxin genes (sea-sed) were detected by multiplex PCR. Results displayed that 16 (34.7%) out of 46 isolates produced SEs. Nine isolates produced SEC (56.2%), 7 (43.7%) isolates produced SEA, while SEB and SED were not produced. Staphylococcal enterotoxin genes were determined in 30 (65.2%) isolates, the most frequently produced enterotoxin gene was gene type sea 18 (60%), followed by sec 9 (30%), sea+sec 3 (10%) but genes seb and sed were not detected in any of the isolates. A relatively high discrepancy was observed between the results of SET-RPLA and multiplex PCR, especially with SEA.
All the 46 isolates were examined for methicillin resistance gene (mecA) and were also tested phenotypically through disk diffusion method. Almost all (45) isolates were methicillin resistance Staphylococcus aureus (MRSA) phenotypically, while 32 isolates found possesses mecA gene. Low relation was observed among methods for determination of MRSA incidence, with statistical significance difference between them.
The antimicrobial sensitivity test exhibited obvious evidence of antimicrobial resistance among isolated bacteria. Among utilized antimicrobial agents, only amikacin and ciprofloxacin were effective against S. aureus isolates. The highest resistance was shown toward penicillin, cloxacillin, ceftazidime, methicillin, and metronidazole by 100% of isolates.
Our results highlight the potential risk of consuming raw cow milk, particularly in the absence of strict hygiene to keep away the existence of MRSA and enterotoxin producing strains of S. aureus in milk.