Polymerase chain reaction detection of bacterial ribosomal genes from fresh and stored animal feeds

Abstract

AbstractFeed quality can be influenced by both the microbial population levels and types of organisms that become associated with the feed matrices during processing and storage. Evaluation of microbial quality of animal feeds is difficult, because feeds generally contain a diverse bacterial population that can fluctuate widely depending on a variety of factors. Microbial diversity may be investigated in animal feed using the polymerase chain reaction (PCR) and 16S rDNA primers. In this study a bacterial isolation step involving centrifugation was combined with several DNA extraction techniques, and PCR amplicons were visualised on electrophoresis gels. Seven different animal feeds and two commonly used feed ingredients, either fresh or stored for approximately 14 months at −20 °C, were chosen (18 feed sources in total) to represent a variety of different matrices, concentrations of macronutrients such as protein and fat, and particle sizes. DNA extraction involving polyethylene glycol 8000 appeared to be the most reliable protocol for the extraction of community DNA for PCR analysis of feeds. The majority of the feeds (14 out of 18 animal feeds and feed ingredients) examined in this study yielded at least one PCR‐positive replicate (1.1 kb band on gel), whereas no amplified products could be obtained from either of the other two DNA extraction protocols. Although some protocol refinement may be necessary for individual feeds, this approach has the potential to be a reliable method for monitoring microbial diversity changes in feeds and for rapid, simultaneous detection of a wide variety of micro‐organisms in animal feeds during processing and storage.© 2002 Society of Chemical Industry