ISOLATION AND IDENTIFICATION OF PASTEURELLA MULTOCIDA OF ANIMAL AND AVIAN ORIGIN BY CULTURAL, BIOCHEMICAL AND MOLECULAR TECHNIQUES

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dc.contributor.advisorROY, ASHISH
dc.contributor.authorJAVIA, BHAVESHKUMAR B .
dc.date.accessioned2018-05-21T06:47:23Z
dc.date.available2018-05-21T06:47:23Z
dc.date.issued2004
dc.description.abstractThe present study was undertaken with a view of isolation and identification of P. multocida from suspected cases of pasteurellosis in different animals and birds. A total of 89 samples were collected from suspected cases, of which 13 (14.6 %) P. multocida isolates (3 from buffalo, 2 from rabbit, 1 from sheep and 7 from poultry) were isolated using blood agar as primary culture medium. These isolates were studied for their biochemical behaviours, in vitro antibiotic sensitivity pattern, P. multocida species specific PCR (PM-PCR) and molecular characterization by PCR-SSCP and repetitive sequence based PCR. All the isolates produce non-haemolytic colonies on blood agar and failed to grow on MacConkey agar. On biochemical behaviours, all the 13 isolates (100%) were found positive for oxidase, catalase, indole production, nitrate reduction and fermentation of glucose, mannitol, sucrose and mannose. All the 13 isolates (100%) v'cre negetive for citrate utilization test and could not ferment maltube, arabinose, lactose, dulitol, salicin and trehalose. All the isolates revealed similar biochemical behaviour. All the isolates (100%) were sensitive to enrofloxacin, flumequine, chloramphenicol and cephalexin, while twelve isolates (92.31%) were sensitive to petloxacin, six isolates (46.15%) were sensitive to ciprofloxacin and four isolates (30.77%) were sensitive to amoxycillin. All the thirteen P. multocida isolates (100%) were found to be resistant to sulphadiazine. All the 13 field isolates along with one vaccine strain (P. multocida P52) and five other bacterial cultures were tested by PM-PCR, which revealed an amplified product of approximately 465 bp size for all the P. multocida isolates, while other bacterial culture did not. The PM-PCR product was subjected to SSCP analysis on 0.5X MDE gel with 10% glycerol. Total six different SSCP band patterns were obtained. These patterns contained a minimum one to maximum four ssDNA bands. Total numbers of bands across the patterns were found to be eight. In this study capability of rep-PCR [repetitive BOX elements, enterobacterial repetitive intergenic consensus (ERIC) and repetitive extragenic palindromic (REP) polymerase chain reaction (PCR)] was tested for molecular characterization of P. multocida isolates. Fingerprinting with BOX-, ERIC- and REP-PCR generated 14, 16, and 13 different bands with molecular weight ranging from 310-1160 bp, 55-900 bp and 44- 500 bp, respectively. The number of bands amplified in different samples by BOX-, ERIC-, and REP-PCR were varied from 2 to 7, 6 to 11, and 1 to 10 with the band frequency ranging from 0.1429 to 0.7857. 0.0714 to 1.0000 and 0.1429 to 0.9286 respectively. The bands of BOX-, ERIC-, and REP-PCR were 100 %. 75 % and 100 % polymorphic, respectively. Dendrogram were constructed by POPGENE software using bands information from BOX-, ERIC-, and REP-PCR. Dendrogram of BOX-PCR data showed 61 % similarity among all buffalo isolates including vaccine strain and 91 % similarity among both rabbit isolates. Poultry isolates were formed two groups with no similarity. Dendrogram of ERIC-PCR data showed 100 % similarity among all buffalo isolates including vaccine strain and between both the rabbit isolates. All the poultry isolates showed 92 % similarity while sheep isolate showed 77 % similarity with all poultry isolates. Dendrogram of REP-PCR showed 100 % similarity among all buffalo isolates including vaccine strain, between both the rabbit isolates and also among all the poultry isolates. Sheep isolate was branched out separately. REP-PCR found most efficient to group isolates from different host origin. Dendrogram based on overall pooled data showed 89 % similarity among all buffalo isolates including vaccine strain, 80 % similarity between both rabbit isolates and 65 % similarity among all poultry isolates. Sheep isolate showed 36 % similarity to poultry isolates. Similar groups were also formed by NETWORK analysis. It is concluded that PM-PCR are useful for rapid and specific detection of P. multocida isolates. SSCP and rep-PCR are useful to identify and characterize P. multocida isolates which can not be classified and characterized by conventional ways.en_US
dc.identifier.urihttp://krishikosh.egranth.ac.in/handle/1/5810046863
dc.keywordsISOLATION AND IDENTIFICATION OF PASTEURELLA MULTOCIDA, ANIMAL AND AVIAN ORIGIN, CULTURAL, BIOCHEMICAL AND MOLECULAR TECHNIQUESen_US
dc.language.isoenen_US
dc.publisherAAU, Ananden_US
dc.research.problemISOLATION AND IDENTIFICATION OF PASTEURELLA MULTOCIDA OF ANIMAL AND AVIAN ORIGIN BY CULTURAL, BIOCHEMICAL AND MOLECULAR TECHNIQUESen_US
dc.subVeterinary Microbiologyen_US
dc.subjectVERTINARY MICROBIOLOGYen_US
dc.subjectTECHNIQUEen_US
dc.themeISOLATION AND IDENTIFICATION OF PASTEURELLA MULTOCIDA OF ANIMAL AND AVIAN ORIGIN BY CULTURAL, BIOCHEMICAL AND MOLECULAR TECHNIQUESen_US
dc.these.typeM.V.Sc.en_US
dc.titleISOLATION AND IDENTIFICATION OF PASTEURELLA MULTOCIDA OF ANIMAL AND AVIAN ORIGIN BY CULTURAL, BIOCHEMICAL AND MOLECULAR TECHNIQUESen_US
dc.typeThesisen_US
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