Molecular and New-Generation Techniques for Rapid Detection of Foodborne Pathogens and Characterization of Microbial Communities in Poultry Meat

Abstract

Poultry meat processing is a complex operation, and there are numerous opportunities for poultry meat to come in contact with microorganisms during the various steps of processing. In addition, the microbial profile certainly changes as antimicrobial interventions are introduced at different steps in the processing plant. Consequently, the microbial communities associated with poultry meat can be highly diverse and complex. Characterizing these microbial populations has traditionally been conducted with culture-based media, but recent developments in molecular techniques have generated opportunities for more in-depth analyses. This review provides an overview of molecular techniques such as polymerase chain reaction (PCR), denaturing gradient gel electrophoresis (DGGE), and next-generation sequencing (NGS) and their applications for rapid detection and typing analysis in poultry microbiology and food safety. Multiplex PCR is a modified version of conventional PCR that can amplify multiple DNA fragments simultaneously, thus having considerable utility for pathogen identification, single-nucleotide polymorphism genotyping, and mutation analysis. Quantitative PCR, one of the more recent methods to quantify genes and organisms, allows absolute or relative quantification of target sequence in a high-throughput format. Quantitative PCR can be used for diagnostic applications in clinical microbiology to detect infectious disease agents, to detect genetically modified organisms, and to quantify genotype strains. Characterizing the microbial community of meat samples is important because it can not only help with detection of pathogens but also enable profiling of potential spoilage microbiota. One of the many types of approaches to investigate the poultry meat microbiome is band pattern-based analysis using DGGE- or DNA-based characterization by next-generation sequencing (NGS). Analysis by DGGE was first introduced to analyze the microbial diversity in mixed, complex microbial populations. However, modifications of DGGE and TGGE greatly expanded the usage of this technique for applications in genetic fingerprinting, monitoring microbial community status and subsequent enrichment of selected bacteria, gene detection, clone library screening, and even determining PCR and cloning biases. Next-generation sequencing (NGS), also referred to as parallel sequencing, is a more recent sequencing technology developed after Sanger sequencing had served as the standard. Millions of DNA fragments from a mixed culture sample can be sequenced by NGS; thus application of this technology can be unlimited. Examples of applicable fields include therapeutic and clinical use to determine genetic disease.