Whole Genome Sequencing: A Magic Bullet for Outbreaks?

Whole genome sequencing (WGS) technology has been explored extensively at food safety conferences and mentioned in news articles on foodborne illness outbreaks. Why? One reason is that the new technology is used as a new tool to identify isolates for foodborne investigations by regulatory agencies such as FDA and CDC.

Developed in the 1970s, DNA sequencing has since advanced drastically, especially during the human WGS project completed in 2003. This development brought in the Next Generation Sequence (NGS) technology era around 2004. During that era, high throughput desktop type-sequencers and software for complex data assembly and analysis became available for scientists to acquire outcomes faster with lower cost, making it possible for research institutes to adapt the technology for further applications—including food safety.

One advantage of WGS is that it can reveal a complete DNA sequence of a living organism. Genetic information of target organisms, such as bacteria, can be analyzed for mutations, virulence characteristics, and antimicrobial resistance. Using single nucleotide polymorphisms (SNPs)—a simple type of genetic variation (mutation) with changes of a DNA nucleotide at certain DNA locations—in comprehensive analysis based on WGS is helpful for understanding the target organism’s evolutionary history.

SNPs can act as biomarkers and be used for subtyping of organisms with high resolution and for comparative analysis with other related organisms. Other molecular subtyping techniques such as pulsed-field gel electrophoresis (PFGE) have some limitations in distinguishing highly related organisms (e.g., Salmonella) because those techniques tend to look at fragmented DNA patterns or particular parts of gene sets. Another significance of WGS is that organism sequence data are uploaded and stored in the public domain (e.g., FDA GenomeTrakr) and can be shared by researchers for real-time analysis and for faster identification and action on disease outbreak causative organisms. WGS also has been used to solve environmental pathogen contamination incidents. Better understanding of virulence factors of pathogenic organisms using WGS may help in developing strategies for preventing foodborne outbreaks. With technology still emerging, the main challenges may be establishing national or global standards and quality thresholds for sequencing methods, data analysis, and data handling/storage.

To ensure the presence of bacterial pathogens is absolutely minimized, many food companies use control programs that include incoming ingredient sampling and testing as verification and environmental sampling for sanitation and hygiene program verification. Conducting WGS on bacteria isolated from these samples generates detailed information about detected pathogen(s) which can be compared to publicly available genomic information. Regulators also may compare WGS results to foodborne illness case databases, which may reveal information about the route and/or source of the potential contamination and/or identify which, if any, preventive or sanitary controls may have been compromised.

Information available through WGS also could reveal that the initially identified pathogen is not from a manufacturing facility or its ingredients. Unlike historical fingerprinting techniques that show if there is a potential match or not, WGS shows actual relatedness of different bacteria. As we continue to sequence organisms, the public database may become robust enough to show where a pathogen, or its relative, has been found before.

In April 2008, CDC and state officials began investigating a Salmonella Saintpaul outbreak. Initial reports from case patients led investigators to believe tomatoes to be the source. But it was later determined that the source was jalapeno and Serrano peppers from Mexico. According to then FDA Associate Commissioner for Foods David Acheson, case-control studies early in the investigation focused on individual patients and strongly suggested tomatoes as the culprit. However, studies of later-occurring case clusters, which public health officials say are a more powerful epidemiologic tool, pointed toward the peppers. This misidentification prevented public health resources from being employed against the proper target. Who knows how many of the 1,442 illnesses and at least 286 hospitalizations could have been avoided by more accurate information generated by WGS?

It also caused the tomato industry to suffer economic harm. Many consumers avoided all tomatoes, even though FDA said some varieties were safe. Estimates from the Florida Tomato Growers Exchange put the loss at $500 million, the value of a full year’s tomato crop. What if WGS of the human isolates could have shown that a similar sequence had been found in Mexico but not Florida, or in peppers but not tomatoes? Of course, these clues need to be supported by epidemiological information, but, optimistically speaking, WGS will provide information that can aid investigators and further protect public health.

The molecular tools employed in the tomato-pepper investigation included what has been termed the “gold standard” PFGE method. However, WGS is already transforming public health microbiology, and may be used to subtype pathogens with greater precision than PFGE. In the September 2015 outbreak involving a California cheese manufacturer, WGS analysis of two environmental samples showed the presence of Listeria monocytogenes that was highly related to the outbreak strains. WGS analysis also determined that five environmental samples from the same facility in 2010 were highly related to the outbreak strains. These outcomes wouldn’t have been readily available in the past.

On the horizon, WGS will allow regulatory agencies and the industry greater precision in the identification of microbes responsible for foodborne illnesses. This added intricacy and detail is a welcome addition to the food scientist’s toolbox. Not only can successful implementation prevent economic hardship, it can target limited public health resources toward the correct offender—an admirable accomplishment for all.

References

Allard, M. W., Y. Luo, E. Strain, C. Li, C. E. Keys, I. Son, R. Stones, S. M. Musser, and E. W. Brown. 2012. High resolution clustering of Salmonella enterica serovar Montevideo strains using a Next-Generation Sequencing approach. BMC Genomics 13:32. http://bit.ly/1OqO45S.

CDC 2008, Outbreak of Salmonella Serotype Saintpaul Infections Associated with Multiple Raw Produce Items—United States, 2008, http://1.usa.gov/1PpEDSA.

CDC 2013. Next Generation PulseNet, http://www.cdc.gov/amd/. Accessed 30 Oct 2015.

Chen, J, A. Van Stelten, C. Cummings, C. Lee, E. Levandowsky, H. Maguire, H. den Bakker, and K. Nightingale. 2013. Whole Genome Sequencing and Phenotypic Characterization of Listeria monocytogenes Isolates from the 2011 Cantaloupe Outbreak Reveals Three Distinct Genetic Clades with Different Phenotypic Traits. Poster Presentations. IAFP, July 28-31, 2013, Charlotte, North Carolina, http://bit.ly/1WCsTgf. Accessed 30 Oct 2015.

FDA. 2015a. Whole Genome Sequencing (WGS) Program, http://1.usa.gov/1kw9Xnd. Accessed 30 Oct 2015.

FDA. 2015b. Genome Trakr Network, http://1.usa.gov/1FD0IXy. Accessed 30 Oct 2015

Food Tracks: Update: September 23, 2015. Karoun Dairy

IFSH 2015. Workshop (agenda): Whole Genome Sequencing for Food Safety—Opportunities and Applications to the Food Industry, http://bit.ly/1l9kYeK. Accessed 30 Oct 2015.

Marler, W., 2008, http://bit.ly/1XSuReT.

Myerson, A., An Analysis of the First-Order Economic Costs of the 2008 FDA Tomato Warning. The Leonard N. Stern School of Business, Glucksman Institute for Research in Securities Markets, April 2009