Molecular typing of Escherichia coli isolates sourced from avian and aquatic species using Repetitive DNA sequences based PCR

Suryakant Mishra 1, S.S. Mishra 2, R.P. Sharma 1,
A. Dharani Kumar 3, B.J.R. Sharma 3 and S.R. Sharma 1

1. Project Directorate on Poultry, Rajendranagar, Hyderabad- 500 030,
2. Central Inland Fisheries Research Institute, Barrakpore, Kolkata- 700 120 &
3. Department of Microbiology, ANGRAU, Hyderabad-500 030

Repetitive Extragenic Palindromic-PCR (REP-PCR) based DNA fingerprinting techniques were applied to groups of E.coli isolates originating from various chicken samples and aquatic species for typing of and phylogeny analyses. A total of 9 chicken origins E.coli strains pre-typed and differentiated from each other (via surface antigen characteristics) were compared to 15 other E.coli strains which were sourced from both fresh-water fishes and Monodon (shrimp) samples using REP-PCR and Box-PCR as per standard protocols. The respective fingerprints were generated for all these isolates by electrophoresing the PCR products on a 2.5% agarose-gel matrix by UV illuminated-gel documentation. The results revealed that phylogenetic grouping of E.coli samples based on REP or Box based fingerprints individually, was not as efficient as it could be, based on combined profiles of REP and Box together. Though, both the REP and BOX fingerprints were individually able to discern one sample from the other, impeccably, the inter-sample genetic relationship on a similarity scale differed from one to the other. The REP fingerprints were more prolific (polymorphic information content) than the BOX profiles across the samples (12 versus 9 polymorphic bands), but the latter yielded more distinct source-specific fingerprint alleles. Phylogeny analysis of the combined (REP & Box together) profiles by standard softwares (NT-Sys and Phylip) demonstrated that all the E.coli samples could be grouped into 3 clusters (2 major and 1 minor). Bulks of the isolates from aquatic species origin were faithfully clustered into a major cluster with a higher genetic-homogeneity pattern (mean Band-sharing index: 73.8%). All the chicken origin-E.coli samples were accommodated in 2 clusters with an average inter-sample homogeneity of 58.2 percent (BS index). The study concluded that repetitive DNA fingerprints based on combined REP and BOX profiles could successfully establish sources of E.coli contamination as differentiated from chickens to aquatic species and would be useful in elucidating their clonal relationships. It was also possible to accurately identify the E.coli infections and trace them back to their sources using REP and Box-PCR, which could facilitate differential-diagnosis and appropriate health-care measures to support chicken farming.

Source : IPSACON-2005

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