Identification of bacteria recovered from animals using the 16S ribosomal RNA gene with pyrosequencing and Sanger sequencing

Journal of Veterinary Diagnostic Investigation : Official Publication of the American Association of Veterinary Laboratory Diagnosticians, Inc
Deepanker TewariJulia Livengood

Abstract

Bacterial identification using genetic sequencing is fast becoming a confirmatory tool for microbiologists. Its application in veterinary diagnostic laboratories is still growing. In addition to availability of Sanger sequencing, pyrosequencing has recently emerged as a unique method for short-read DNA sequencing for bacterial identifications. Its ease of use makes it possible to diagnose infections rapidly at a low cost even in smaller laboratories. In the current study, pyrosequencing was compared with Sanger sequencing for identification of the bacterial organisms. Fifty-four bacterial isolates spanning 23 different bacterial families encountered in veterinary diagnostic microbiology laboratories were sequenced using 16S ribosomal RNA gene with pyrosequencing and Sanger sequencing. Pyrosequencing was able to identify 80% of isolates to the genus level, and 43% isolates to the species level. Sanger sequencing with approximately 500 bp performed better for both genus (100%) and species (87%) identification. Use of different sequence databases to identify bacteria isolated from animals showed relative importance of public databases compared to a validated commercial library. A time and limited cost comparison between pyrosequen...Continue Reading

References

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Methods Mentioned

BETA
PCR

Software Mentioned

ClustalW
MicroSEQ

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