الفهرس 
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Abstract
A total of 237 samples were collected from 79 Animals (66 cows and 13 buffaloes)
.from each animal, we collected 3 samples including milk sample, teat swab and nasal swab.in addition to 15 samples were collected from dairy utensils and 52 human samples including nasal swabs and hand swabs from four different dairy farms in Qalyobia Governorate then transport to the laboratory to make Isolation and biochemical identification of the microbe from the collected samples, characterization of virulence factors using molecular biology methods, Detection of antibiotic resistance of the isolates., study of the risk factors associated with milk contamination in dairy farms, and Study the genetic sequence and phylogenetic analysis of these isolates.
The results showed the following:
The occurrence of S.aureus in the examined 66 cows was 4.55% from nasal swabs (33.33% with nasal catarrh and 3.17% apparently healthy animals), 6.06 % from teat swabs (25% with skin lesion and 3.45% apparently healthy animals) and 7.58% from milk. Concerning the occurrence of S.aureus in the examined 13 buffaloes, it was 15.38% from nasal swabs (100% with nasal catarrh and 8.33% apparently healthy animals), 15.38 % from teat swabs (100% with skin lesion and 0% apparently healthy animals).
Regarding to the occurrence of S.aureus in examined dairy utensils (26.67%), it was higher in hand milking (50%) than machine milking (11.11%).
Examined dairy workers reveled that out of 26 nasal swabs, 4 isolates (15.38%) (100% with nasal catarrh and 12% apparently healthy humans) were obtained and 6 isolates (23.08%) (57.14% with skin lesion and 10.53% apparently healthy humans) were obtained from hand swabs.
High resistance to ampicillin (90%) and amoxicillin/clavulanic acid (96.7%) were found in the current study’s findings.The current study also revealed resistance to cefotaxime (53.3%). The current study revealed that 100% of all S.aureus isolates were sensitive to
levofloxacin, (56.7%) were sensitive to erythromycin and azithromycin, higher sensitivity to ciprofloxacin (70%) and (26.7%) for ceftriaxone.
The isolation of S.aureus in milk that collected either automated or hand was 1.92% and 14.8% respectively and the isolation rate is higher in mastitic animals than apparently healthy animals.
Molecular identification of S. aureus isolates were confirmed as S.aureus by using PCR for nuc gene , all the twelve Staphylococcus aureus isolates were positive for nuc gene in PCR (12/12)100% as nuc gene is frequently employed as a particular target for the detection of S. aureus by PCR.
The ability of Staphylococcus aureus to form biofilm is considered to be a major virulence factor influencing its survival and persistence in both the environment and the host. So, in this study using PCR to target bap gene responsible for biofilm formation. Only 10 out of the 12 S.aureus isolates were positive in PCR (10/12) (83.33%) targeting bap gene. In this study, two Samples were negative for bap (nasal and teat swab of buffalo respectively . By targeting smr gene responsible for resistance to antiseptics, all the 12 S.aureus isolates were positive in PCR (12/12)100% for smr gene.
Three of the positive PCR products from each of S. aureus bap and smr genes were used for sequencing and phylogenetic analysis by using the programs Bio Edit 7.0.4.1 and MUSCLE were used to examine the nucleotide sequences acquired in this work. Using a neighbour-joining technique of the aligned sequences then Six nucleotide sequences for S.aureus isolates were produced and deposited in GenBank under the accession numbers OR344353 to OR344355 for the bap gene, OR344356 to OR344358 for the smr gene, and showed typical amino acid sequences (100%) of S. aureus isolates from cattle milk and dairy utensils while the human isolate showed major mutations through change and insertion (addition), almost along the DNA sequence especially from nucleotide 53 to nucleotide 68 for bap gene.
Targeting the bap gene, the phylogenetic tree formed four clades where all the S. aureus isolates recovered from cattle milk (OR344353) and dairy utensils (OR344354) showed a high homology (74%) with that of S.epidermidis isolated from human nosocomial infections (EU011247) in Brazil and low homology (47%) with S. aureus isolate from bovine milk (JX403946) in India .
S. aureus isolate (OR344355) from a human nasal swab in Egypt showed high homology with
S. aureus isolates (MF278359 (88%) & MF278360 (78%)) isolated from bovine milk in India. This showed the possibility of transfer of bap gene between human and animal Staphylococcus Spp. and shows the zoonotic importance of not only S. aureus but also coagulase negative staphylococci especially S. epidermidis.
Concerning the smr gene, all the S. aureus isolates from study (OR344356- OR344358) showed a high homology (43%) with S. epidermidis isolated from ovine milk (MK933771) in Italy.
The Comparative alignment was recorded for the three translated smr gene sequences, showed almost typical amino acid sequences of S. aureus isolates from cattle milk and dairy utensils while mutation by change and addition in human isolates (OR344358) especially from nucleotide 1 to nucleotide 90, almost along the DNA sequence.