Food safety is a global concern and foodborne illness outbreaks remain a significant challenge to public health and pose a huge economic burden worldwide. In the U.S., foodborne pathogens cause an estimated 9.4 million illnesses each year, including 56,000 hospitalizations and 1,400 deaths. Additionally, foodborne pathogens cause a 10% gastroenteritis in Europe annually. While 31 known pathogens cause foodborne illness, Salmonella, Campylobacter, Listeria monocytogenes, and E. coli O57:H7 have been implicated as the cause of many multi-state outbreaks of foodborne illness in recent years. Often the investigations of foodborne illness outbreaks fail to find the source, and the illness outbreak is referred to as caused by “unknown etiology” or by “unspecified agents.” Foodborne illnesses are preventable, yet they remain a significant challenge to the food industry and pose a huge public health and economic burden.
Pathogenic bacteria such as Salmonella, E. coli O157:H7, and L. monocytogenes are recognized as the most significant biological hazards, not only in ingredients, raw materials, and finished foods, but also in the food plant environment. Pathogens in the food plant environment can contaminate food, especially ready-to-eat (RTE) foods post processing and prior to packaging. Thus, the food industry and FDA are increasingly employing sampling food manufacturing facilities to isolate pathogenic organisms and characterize and subtype them to develop a microbiological profile of the processing facility that was sampled in addition to the products from that facility.
Similarly, epidemiological investigations by public health agencies (e.g., the Centers for Disease Control and Prevention (CDC) and state and local departments of public health) also involve pathogen isolation, characterization, and subtyping to identify which pathogen is causing an outbreak or recall and tracing the source involved in the outbreak. Clinical isolates obtained from patients affected by a foodborne illness can be compared with samples collected from foods and food plant environments to potentially identify a source of the pathogen that is causing the foodborne illness. When a match is made between clinical isolates and samples from a food or food plant, the scope and impact of the foodborne illness can be best understood. Product recalls may also be targeted based on the results of these efforts.
Food microbiologists have always been interested in methods of identification and characterization of microbial isolates in food and beverages. Early techniques included staining and microscopy; comparison of physiological, biochemical, and serological characteristics to discriminate species; and strains of microorganisms of interest. These techniques allowed for evaluation of the target organism; however, they did not have sufficient discriminatory power to allow precise identification of and differentiation between related strains of microorganisms. Also, these traditional methods are material and labor intensive, time consuming, and expensive for routine use in the identification, characterization, and subtyping of bacterial strains.
Advances in molecular biology during the last part of the 20th century have resulted in the development of efficient techniques that have made possible the rapid identification and characterization of microbial isolates. Next generation sequencing (NGS) methods have transformed from being solely research tools to being routinely applied in diagnostics, outbreak investigations, antimicrobial resistance, forensics, and food authenticity.
NGS is predominantly used in two ways:
- Determining the whole-genome sequence (WGS) of a single cultured isolate (e.g., a bacterial colony); and
- Application to a biological sample generating sequences of multiple (if not all) microorganisms in that sample (i.e., “metagenomics”).
WGS, which is a type of NGS that has a high discriminatory power when compared with traditional molecular typing tools, is increasingly replacing traditional microbial typing and characterization techniques.
WGS can differentiate microbial strains at a high enough resolution and is increasingly used by FDA, CDC, USDA Food Safety and Inspection Service (FSIS), and other public health agencies worldwide for epidemiological investigation of foodborne illnesses, identification of related cases, source attribution, and development of intervention strategies