الفهرس 
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Abstract
The current study was carried out on different dairy cattle farms in Kafr El Sheikh Governorate during the period extended from December 2017 till the end of June 2018 for isolation of mastitis causing bacteria from milk and environmental samples then determine the sensitivity of some recovered bacterial isolates to certain antibiotics commonly used in field treatment of mastitis and finally, evaluation of the efficiency of some chemical disinfectants commonly used in the veterinary field against the isolated bacteria as well as assessment the disinfecting effect Rosmarinus officinalis plant. 7.1. Isolation and identification of mastitis causing bacteria in different dairy farms: A total of 700 different samples were collected from dairy cows and their surrounding environment from different dairy farms located in Kafr El-Sheikh including; milk (150 samples from clinically suspected cases and another 150 samples from apparently healthy cows) and swabs from teats, milking machines and hands of workers as well as bedding materials (100 samples/each) for isolation and identification of significant mastitis causing bacteria including; Staphylococcus spp., Streptococcus spp. and Coliforms bacteria. At the beginning, California mastitis test (CMT) was carried out to detect sub-clinical mastitis in milk samples gathered from apparently healthy cows for selection of samples for bacterial culture and the recorded results revealed that the prevalence of subclinical mastitis was 54.67% (82 out of 150 tested milk samples). The frequency of isolation of Staphylococcus spp., Streptococcus spp. and Coliforms bacteria from milk samples of clinical mastitis was 65.33, 44 and 81.33 %, respectively while it was 62.19, 37.8 and 53.66 %, respectively in milk samples of sub clinical mastitis cases. Moreover, statistical analysis showed a significant association between the rates of isolation of identified mastitis causing bacteria where the highest frequency of isolation was noticed in in Staphylococcus spp. followed by Coliforms and lastly Streptococcus spp. On the other side, statistical analysis clarified a significant association between the rates of isolation of these bacteria from different environmental samples where Staphylococcus spp., scored the highest rate of isolation followed by coliforms and lastly Streptococcus spp. from the examined samples of teat swabs, milking machines swabs and hand swabs of workers. On contrary in case of the examined samples of bedding material, coliforms scored the highest rate of isolation (89%) followed by Streptococcus spp. (57%) and lastly, Staphylococcus spp. (15%). Concerning Staphylococcus spp., the results of coagulase test showed that the overall frequency of isolation of Staph. aureus was 63.81% and the highest frequency was observed in subclinical mastitis milk (90.2%) followed by clinical mastitis milk (78.57%) then teat swabs (67.03%), milking machines swabs (49.32%), bedding material (33.33%) and finally hand swabs of workers (28.89%) with significant association between the different rates of isolation. On the other side, the overall rate of isolation of Coagulase negative Staphylococci was 36.19% and the highest frequency was observed in hand swabs of workers (71.11%) followed by bedding material (66.67%), milking machines swabs (50.68%), teat swabs (32.97%) then samples of clinical mastitis milk (21.43%) and finally, samples of subclinical mastitis milk (9.80%) with significant association between the different rates of isolation. The biochemical identification of Str. agalactiae (an example of contagious mastitis causing streptococci) and Str. uberis (an example of environmental causing streptococci) was carries out. The recorded results clarified that the total rate of isolation of Str. agalactiae was 38.17% and the highest frequency was observed in the examined samples of subclinical mastitis milk and teat swabs (52.38% of each) followed by samples of milking machines swabs (41.86%), clinical mastitis milk (34.85%), hand swabs of workers (34.78%) and finally bedding material (21.05%) with significant association between the different rates of isolation. In addition, the overall frequency of isolation of Str. uberis was 35.11 % and the highest rate was observed in the examined samples of bedding (50.88%) followed by milking machine swabs (44.19%) then the hand swabs of workers (39.13%), teat swabs and finally milk samples of both clinical and subclinical mastitis cases (22% for each) with significant association between the different rates of isolation. Escherichia coli were identified from the isolated coliforms on basis of biochemical testing. It was found that the total rate of isolation of E. coli was 67.43% and the highest frequency of isolation was observed in the examined hand swabs of workers (84.37%) followed by clinical mastitis milk (78.69%) then subclinical mastitis milk (72.735%), teat swabs (71.26%) followed by samples of milking machines swabs (56.45%), and finally bedding material (47.19%) with significant association between the different rates of isolation. 7.2. In vitro antimicrobial susceptibility of the recovered bacterial isolates: In vitro antimicrobial susceptibility of the recovered bacterial isolates to some commonly antibiotics usually used for the treatment of mastitis in dairy farms was studied. Concerning coagulase positive Staph. aureus, statistical analysis showed a significant association between different tested antibiotic where it was noticed that all of the tested isolates (n=50) were found resistant to Amoxyclav (30 mcg), Penicillin-G (10 units) and Amoxicillin (10 mcg) while they were found to be sensitive for Norfloxacin (10 mcg) (64%) followed by Gentamicin (10 mcg) (62%) and Spiramycin (30 mcg) (52%) while in case of coagulase negative Staph. aureus, it was recorded that the highest antibiotic resistance was observed in Amoxicillin (10 mcg) (82%) followed by Penicillin-G (10 units) (78%) then Co-Trimoxazole (25 mcg) (76%) while the highest antibiotic sensitivity was observed in case of Gentamicin (10 mcg) (60%) followed by Spiramycin (30 mcg) (40%). Concerning coagulase Str. agalactiae, statistical analysis showed a significant association between different tested antibiotic where it was noticed that the highest antibiotic resistance was observed in Chloramphenicol (30 mcg) (90%) followed by Co-Trimoxazole (25 mcg) (78%) then Streptomycin (10 mcg) (64%) while the highest antibiotic sensitivity was observed in case of Spiramycin (30 mcg) (72%) followed by Gentamicin (10 mcg) (70%) while in case of Str. uberis, the highest antibiotic resistance was observed in Amoxicillin (10 mcg) (78%) followed by Amoxyclav (30 mcg) (72%) then Streptomycin (10 mcg) (64%) while the highest antibiotic sensitivity was observed in case of Spiramycin (30 mcg) (78%) followed by Streptomycin (10 mcg) (70%) then Cefotaxime (30 mcg) and Gentamicin (10 mcg) (64 and 60%, respectively). Results of the in vitro antimicrobial susceptibility test of E. coli clarified that all of the tested isolates (n=50) were found resistant to Amoxicillin (10 mcg) and Spiramycin (30 mcg) while Norfloxacin (10 mcg) scored the highest antibiotic sensitivity (64%) followed by Chloramphenicol (30 mcg) (42%) while in case of coliforms other than E. coli, it was recorded that the highest antibiotic resistance was observed in Cefotaxime (30 mcg) (86%) and Penicillin-G (10 units) (84%) followed by Spiramycin (30 mcg) (78%) then Amoxyclav (30 mcg) (66%) while the highest antibiotic sensitivity was observed in case of Co-Trimoxazole (25 mcg) (50%) followed by Chloramphenicol (30 mcg) and Spiramycin (30 mcg) (46%). 7.3. Molecular detection of resistance genes in different bacterial isolates: A total of 10 isolates of E. coli were screened for presence of blaTEM gene by PCR and they gave the characteristic band at 247 bp indicating the presence of target gene. On contrary, they gave no bands when they were screened for presence of genes blaSHV and group1blaCTX-M indicating the absence of these genes. Also, 10 isolates of Staphylococcus aureus were screened for presence of bla Z gene and they gave the characteristic band at 861 bp indicating the presence of target gene. 7.4. In vitro antiseptic sensitivity of some disinfectants against isolated bacteria: Four commercial types of disinfectant commonly used in dairy farms for teat dips and disinfection of dairies and milking machines including Iodine, Phenol, Potassium peroxy monosulfate (Virkon S®) and Quaternary Ammonium Compounds (TH4) were tested. Also, Rosemary (Rosmarinus officinalis) which is rich source of phenolic compounds with high antimicrobial activity against both Gram positive and Gram negative bacteria was tested for the antibacterial activity of rosemary extract against isolated organisms in vitro. The results in present study showed that the tested bacteria isolates exhibited resistance to all of the tested disinfectants including the extract of Rosemary on using the lowest tested concentration (0.25 %). On contrary, all of the tested bacterial isolates were found sensitive to all of the tested disinfectants including the extract of Rosemary on using the concentration of 2.0 % that is considered as double dose used in the veterinary practice.