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Subject: Veterinary Public Health × End date: 2023 × Start date: 2023 × Thesis Type: M.V.Sc. ×
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    ThesisItemOpen Access
    MOLECULAR CHARACTERIZATION OF Clostridium perfringens ISOLATED FROM ANIMALS AND HUMANS
    (SRI VENKATESWARA VETERINARY UNIVERSITY TIRUPATI - 517 502. (A.P.) INDIA, 2023-03) SHARMILA GELLANKI; SRINIVASA RAO .T (MAJOR); SUBHASHINI .N; SUDHAKAR .K
    The present study was undertaken for isolation and characterization of C. perfringens from animals, foods of animal origin and human samples. Out of 395 samples analyzed, 22.7% (90/395) were positive for C. perfringens by cultural isolation and confirmation by biochemical tests. Out of 90 phenotypically positive C. perfringens isolates from different sources, 34 (37.7%) isolates were further confirmed by species specific PCR. The highest rate of occurrence of C. perfringens was detected in mutton samples (18.1%) while lowest in human stool swab (5%) samples. All the C. perfringens isolates (100%) showed lecithinase activity on egg yolk agar, 82.3% isolates exhibited congo red binding activity, 73.5% isolates showed gelatinase activity and 88.2% isolates showed hemolytic activity on sheep blood agar plates. Eight (23.5%) C. perfringens isolates carried cpa gene indicating toxinotype A, four (11.7%) isolates harboured both cpa and cpe genes indicating toxinotype F and three (8.8%) isolates carried cpe gene alone. None of the isolates did not carry other three virulent genes like etx, cpb and cpi. Among 34 C. perfringens isolates higher resistance was observed towards erythromycin, linezolid, metronidazole, pencillin G, rifampicin (100%), followed by gentamicin (91.1%), tetracycline (88.2%), teicoplanin (85.2%), ciprofloxacin (73.5%) and chloramphenicol (32.3%). The sensitivity percentages were highest against imipenem (88.2%) followed by ceftriaxone (14.7%). Intermediate resistant patterns were observed against ceftriaxone (85.2%), chloramphenicol (67.6%) and ciprofloxacin (26.4%). All the 34 C. perfringens isolates were MDR and MAR indexing of all isolates yielded 6 MAR index groups. Out of 34 C. perfringens isolates 19 were identified as ESBL suspects by PST and ESBL production was confirmed in ten isolates by CDM and in four isolates by DDST. All the 19 phenotypic ESBL suspects did not carry any of the beta-lactamase genes (blaTEM, blaSHV and blaOXA) by multiplex PCR. A greater degree of heterogeneity was observed among 34 C. perfringens isolates from different sources by ERIC and REP-PCR. ERIC-PCR distinguished 33 genotypes and REP-PCR distinguished 34 genotypes of C. perfringens isolates. Genotyping of C. perfringens by ERIC-PCR and REP-PCR was highly significant since the discriminatory power is >0.9 (0.998 for ERIC–PCR and 1 for REP-PCR). Cluster analysis also revealed a greater degree of homogeneity and heterogeneity among different isolates recovered from different sources indicating the chances of cross contamination.
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    ThesisItemOpen Access
    MOLECULAR STUDIES ON Acinetobacter baumannii ISOLATES OF ANIMAL AND HUMAN ORIGIN
    (SRI VENKATESWARA VETERINARY UNIVERSITY TIRUPATI - 517 502. (A.P.) INDIA, 2023-03) RESHMA MARADANA; SRINIVASA RAO .T (MAJOR); BINDU KIRANMAYI .CH; ASWANI KUMAR .K
    The present study was undertaken for isolation and characterization of Acinetobacter baumannii from foods of animal origin, rectal/cloacal swabs of animals/ birds and human clinical samples. Out of 420 samples analyzed, 21 (5%) samples were positive for A. baumannii by m-PCR. About 12% of human blood samples, 10% of milk samples, 6.6% of chicken samples, 4% of human urine samples and 2.8% of sheep rectal swabs samples were positive for A. baumannii. Fifty two percent of A. baumannii isolates showed gelatinase activity on gelatin agar and none of the isolates exhibited biofilm production ability on congo red agar. About 28.57% of A. baumannii isolates carried the virulence genes csgA gene and 14% of the isolates harboured fimH gene whereas cnf1 and afa/draBC genes were not found in any of the A. baumannii isolates. Antibiogram of 21 A. baumannii isolates revealed 100% resistance towards penicillin-G and polymyxin B, followed by co-trimoxazole 90.4%, gentamicin 61.9%, amikacin 47.6%, cefepime and colistin (42.8%). Higher sensitivity was observed for ciprofloxacin and tigecycline (100%), followed by tetracycline (80.9%), imipenem (71.4%) and meropenem (66.6%). Colistin resistance genes mcr-1 and mcr-2 were detected in two isolates of human blood samples out of nine isolates which were phenotypically resistant to colistin. A total of three (14.3%) human isolates were phenotypically confirmed as ESBL producers and blaOXA was the only β- lactamase gene detected in two A. baumannii isolates of human blood samples. ERIC-PCR genotyping distinguished 19 genotypes among 21 isolates of A. baumannii. REP-PCR genotyping distinguished 20 genotypes among 21 A. baumannii isolates. ERIC-PCR and REP-PCR analysis revealed a greater degree of heterogeneity among 21 A. baumannii isolates from different sources. The discriminatory power of ERIC-PCR and REP-PCR was found to be 0.990 and 0.995, respectively indicating both the genotyping methods were highly significant. Cluster analysis also revealed a great degree of homogeneity and heterogeneity among different isolates recovered from different sources thereby indicating that possible chance of interactions among different environments.
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    ThesisItemOpen Access
    DETECTION OF ANTIBIOTIC RESISTANCE, GENETIC DIVERSITY IN Klebsiella pneumoniae ISOLATED FROM FOODS OF ANIMAL ORIGIN AND ITS PUBLIC HEALTH SIGNIFICANCE
    (SRI VENKATESWARA VETERINARY UNIVERSITY TIRUPATI - 517 502. (A.P.) INDIA, 2023-03) VENKATALAXMI ADIMULAM; SUBHASHINI .N (MAJOR); SRINIVASA RAO .T; ASWANI KUMAR .K
    The present study was undertaken to characterize K. pneumoniae in foods of animal origin and human clinical samples based on cultural and molecular techniques. A total of 402 samples comprising different foods like milk, meat, egg and fish, human clinical samples such as blood, urine and nasal swabs, rectal/cloacal swabs of pigs and chickens were analyzed for the presence of K. pneumoniae. The overall occurrence of K. pneumoniae was found to be 6.47% (26/402) by cultural and molecular methods in samples obtained from different sources. Out of 26 K. pneumoniae isolates, highest occurrence was found in urine samples (7/26, 15.21%) followed by pork (6/44, 13.64%), eggs (2/15, 13.33%), milk (5/58, 8.62%), chicken (2/42, 4.76%) and blood (4/102, 3.92%) samples. Phenotypic virulence factors such as capsule, hypermucoviscosity and biofilm formation were detected in 26 (100%), 11(42.31%) and eight (30.77%) isolates, respectively whereas virulence genes such as uge, rmpA and Kfu were observed in eight (30.77%), four (15.38%) and three (11.54%) isolates, respectively. None of the isolates carried the aerobactin gene. Antibiogram profiling of K. pneumoniae isolates indicated the highest resistance to erythromycin, vancomycin and ampicillin in 100%, 100% and 92.3%, respectively followed by cefpodoxime (80.77%), amoxicillin-clavulanic acid (76.92%), cefixime (57.69%), ceftriaxone (34.61%). All the isolates were sensitive to imipenem followed by piperacillin-tazobactam (84.61%). Intermediate resistance was observed to colistin (69.23%). All K. pneumoniae isolates were found to be MDR and MAR index value ranged between 0.18 to 0.65. ESBL production was confirmed in 14 K. pneumoniae isolates by both phenotypic and molecular methods and blaOXA, was the predominant gene which was found in eight among 14 isolates, blaCTX-M1 and blaCTX-M9 were found in four and two isolates, respectively and the β-lactamase gene blaAmpC was detected only in four isolates. A greater degree of heterogeneity was observed among 14 ESBL-positive K. pneumoniae isolated from different sources as revealed by presence of 14 genotypes each by ERIC and REP-PCR analysis. All the 14 different K. pneumoniae subtypes were differentiated by ERIC-PCR and REP-PCR. Genotyping of ESBL-producing K. pneumoniae by ERIC-PCR and REP-PCR was found to be highly significant since the discriminatory power >0.9 is considered highly significant (one for both ERIC-PCR and REP-PCR). Cluster analysis also revealed a great degree of heterogeneity and also the homogeneity among different isolates recovered from different sources thereby indicating that there is a chance of cross-contamination between various foods of animal origin.
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    ThesisItemOpen Access
    ISOLATION AND MOLECULAR CHARACTERIZATION OF Bacillus cereus ISOLATED FROM MILK AND MILK PRODUCTS
    (2023-04) KUPPILI, PRIYANKA; Ch, BINDU KIRANMAYI; T, SRINIVASA RAO; K, ASWANI KUMAR
    The present study was undertaken to characterize the incidence, enterotoxigenic gene profile, antibiogram profile and genetic diversity of Bacillus cereus isolated from milk and different milk products. A total of 320 samples comprising of 150 raw milk and 170 milk product samples were analyzed for the presence of B. cereus. Overall prevalence of B. cereus in milk and milk products samples was found to be 9.68% (31/320) by species specific PCR targeting gyrB gene with 8% (12/150), 15% (3/20), 12.1% (4/33), 11.8% (2/17), 10.8% (4/37) and 21.4% (6/28) from raw milk, basundi, burfi, colostrum milk pudding (junnu), kalakhand and khoa samples, respectively. Out of 31 genotypically positive B. cereus isolates, 23 (74.1%) isolates were carrying atleast one of the virulence genes with nheA in 10 isolates (32.3%), nheB in 12 isolates (38.7%), nheC in 16 (51.6%) isolates and entFM in 14 isolates (45.2%) and none of the isolates were positive for hblA, hblC, hblD and cytK genes. Antibiogram profiling of 31 B. cereus isolates revealed that all the isolates (100%) were resistant to pencillin and amoxicillin-clavulanic acid followed by cefepime (87.1%), clindamycin (32.31%), rifampicin (29.0%), vancomycin (25.8%), colistin (19.4%) ciprofloxacin, chloramphenicol and erythromycin (6.5%) each and none of the isolates were resistant to gentamicin and tetracycline. None of the B. cereus isolates were positive for any of the ESBL genes (bla TEM, bla SHV, blaOXA and bla CTX-M). A greater degree of heterogeneity among the genotypically positive B. cereus isolates isolated from milk and different milk products was revealed by the presence of 28 genotypes by ERIC-PCR and 29 genotypes by REP-PCR analysis. Genotyping of B. cereus by ERIC-PCR and REP-PCR were found to be highly suitable since the discriminatory power above 0.9 are considered as highly significant (0.997 for ERIC PCR and 0.995 for REP-PCR). As the discriminatory power of ERIC-PCR is more than the REP-PCR, ERIC-PCR was able to differentiate the isolates better than REP-PCR. Cluster analysis using dendrograms revealed molecular heterogeneity and varying homogeneity among different isolates recovered from varied sources