| Endometritis,a multi-etiological ailment,is a challenge in the exploitation of optimum reproductive potential of animals compromising farm economy and health because of its occurrence rate sustaining as 35-45% in dairy sectors of developing countries.Among the etiological agents,Staphylococcus aureus(S.aureus)and Escherichia coli(E.coli)are known as the most significant causes of uterine ailments in bovines.The study was aimed to highlight the importance of the regular molecular epidemiological pattern of bacterial etiologies of endometritis especially overlooked bacteria,their drug resistance,and to find effective drug combinations to treat endometritis.The first experiment was designed to assess the prevalence of S.aureus and E.coli in bovine uterine samples along with contributing risk factors.These bacteria are usually overlooked for their prevalence and pathogenesis.Uterine flushing was obtained aseptically after ultrasonographic screening from n=304 postpartum cattle from different commercial dairy setups in and around District Khanewal,Punjab,Pakistan.A pre-designed questionnaire was filled on the spot to collect data about contributing risk factors.Samples were processed microbiologically and biochemically for isolation of S.aureus and E.coli.The study results showed a43.75% prevalence of endometritis.Microbiological assays were performed to identify S.aureus and E.coli.The experiment found 78.19% 72.18% of S.aureus and E.coli from endometritis samples.Most of the assumed risk factors were significantly associated endometritis.The second experiment was done for molecular conformation and genetic analysis of previously harvested isolates.This experiment was intended to find a possible route of spread of this pathogen and differences with the reported isolates.Antibiotic susceptibility profile was also determined to find resistance profile against multiple antibiotics.Local isolate of S.aureus Nuc gene(S.aureus-1,S.aureus-2,and S.aureus-3)sequence was closely related to the S.aureus Nuc gene isolated from Bos taurus and pork.The order of antibiotic resistance of S.aureus against antibiotics was trimethoprimsulphamethoxazole > ciprofloxacin ≥ enrofloxacin ≥ chloramphenicol >gentamicin > levofloxacin > linezolid > vancomycin > fusidic acid.Nucleotide analysis revealed that 23 S r RNA nucleotides sequence of local E.coli isolate showed 98% similarity to the Nucleotide sequence of the reference E.coli strain.The Nucleotide sequence of the 23 S r RNA gene of the local E.coli isolate was found to be closely related to that of the E.coli isolate from human rectal swab(Laos)and was clustered together with the E.coli isolated from human rectal swab(Laos),human urine(USA),swine liver(China),dairy manure(Canada),human feces(Singapore),chicken meat inner strip(Romania),human urine(China),human(Spain)and pig feces(Canada).The order of resistance of E.coli against different antibiotics was in the following order “fusidic acid >vancomycin>levofloxacin>linezolid>cefoxitin>gentamicin,≥enrofloxacin>cipro floxacin≥trimethoprim-sulfamethoxazole≥chloramphenicol”.In the third experiment,various antibiotics/drugs combination were tested to ensure an effective combination to treat the malaise and to give an idea of the use of antibiotics in rotation.For this purpose,well-diffusion test was used as primary in-vitro,and afterward,the broth micro-dilution method was used to determine synergistic combinations against S.aureus and E.coli.Based on the in-vitro trial,a field trial was conducted on E.coli endometritis and S.aureus endometritis.Drug combination analysis of well diffusion zones showed that there was a maximum increase in the inhibitory zone(188.89 %)for the combination of oxytetracycline with streptomycin when compared to oxytetracycline.Synergy testing of all the tested drug combinations against drug-resistant S.aureus isolates showed synergistic behavior of combinations;amoxicillin with gentamicin and metronidazole with enrofloxacin while antagonism was observed for combinations of oxytetracycline with amoxicillin,metronidazole,and enrofloxacin.The study(field trials against S.aureus based endometritis)noted the highest percentage recovery in the case of Genta+Enro,gentamicin alone,Co-amoxiclav+Gentamicin and oxytetracycline alone,and Metro+Enro combinations presenting 100%,100%,69.23%,57.14% and33.33% success rate in terms of recovery/healing of uterine wall.Synergy testing of all tested drugs combinations against E.coli isolates showed no synergistic effect while antagonism was noted for combinations of enrofloxacin and oxytetracycline.The field trial(E.coli based endometritis)showed the highest recovery rates in case of gentamicin + enrofloxacin(100%),gentamicin alone(100%),co-amoxiclav + gentamicin(84.61%),oxytetracycline alone(78.57%),and metronidazole + enrofloxacin(33.33%).The study in overall found higher prevalence of S.aureus and E.coli based bovine endometritis;genetic shifts in the targeted bacteria;higher resistance to commonly used antibiotics;and availability of wider range of effective antibiotic combinations to treat endometritis. |
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