Of all foodborne pathogens, Salmonella is one of the most difficult to isolate because of its homogeneity. Strains like Salmonella enteritidis and Salmonella Montevideo are, genetically speaking, almost indistinguishable from one another using conventional tools of forensic microbiology.
In fact, if you were to test a contaminated food sample with pulsed-field gel electrophoresis (PFGE), the standardized DNA fingerprinting used by the Centers for Disease Control and Prevention’s (CDC) PulseNet program to detect foodborne disease case clusters and identify common sources of outbreaks, S. enteritidis and S. Montevideo would appear to be the same pathogen.
That hamstrings investigators trying to accurately define the scope of an outbreak involving Salmonella. But next-generation genomic sequencing now under investigation at the U.S. Food and Drug Administration (FDA) could break that bottleneck.
In early 2010, about 300 people in the U.S. became ill with S. Montevideo, with victims in almost every state. Early in the investigation, it was thought the outbreak could be traced to contaminated pistachios, which had been implicated in a 2008 Salmonella outbreak. PFGE analysis couldn’t detect a difference between the strains involved in the two outbreaks—ultimately, field investigation narrowed the source of contamination to a spiced meat rub used in salami.
But which company produced the tainted product? Only in retrospect, using next-generation sequencing, were investigators at the FDA able to pinpoint the precise source of contamination—a single processing plant.
“This is the first time this technology has been used in a foodborne outbreak investigation, with enough strains to truly define the scope of an outbreak,” said Steven Musser, PhD, director of the FDA’s Office of Regulatory Science (ORS). Previously, next-generation sequencing had been used to differentiate among a handful of strains of an organism: in a Canadian tuberculosis study and an outbreak of cholera in Haiti. But in this case, researchers were attempting to distinguish among 30-40 Salmonella strains.
Dr. Musser explained the limitations of PFGE this way: “It chops DNA into very large pieces, separated electrophoretically into a pattern of about eight to 10 bands—sometimes more, sometimes less. This is a very nice technology for saying yes, it looks like we have a cluster of cases, but you may have a pattern that consistently shows up that you cannot differentiate further. It’s a high-level snapshot.”
But if PFGE is Google Earth at satellite-level view, next-generation sequencing is Google Earth at street level.
“We sequence all the DNA of a clinical strain,” Dr. Musser said. “We have every single base, and we know exactly how they define that particular organism. Even though 99% of the genetic information is very similar, this technology allows us to find the very small differences that define whether something has a high or low probability of being in the cluster. So with whole genome sequencing, we could easily say that the pistachios weren’t involved in the new outbreak.”
Next-generation sequencing can also include or exclude a particular patient from involvement in a specific outbreak. “That can be of great benefit to the epidemiologist, to help them determine whether they’re looking at multiple sources of contamination or one large, single outbreak,” Dr. Musser said.
Despite its advantages, whole genome sequencing will be limited primarily to retrospective investigations for the foreseeable future. That’s because of two major limitations: time and money.