Whole Genome Sequencing (WGS) and Pulsed Field Gel Electrophoresis (PFGE) are important tools the Center for Disease Control and Prevention (CDC) have used in order to collect and compare data pertaining to different foodborne outbreaks across the nation. What is the difference between the two, and is it really important which method researchers use?
For starters, both techniques look at an organism’s DNA. Each individual organism (whether it be a whale, fish, or a toad), has a different genetic makeup, and therefore a unique genetic pattern. This pattern is composed of nucleotide bases (A, T, C, G), and if someone can identify the pattern in which these bases occur, then they have identified the organism’s “unique DNA fingerprint”. So if both techniques are used on an organism’s DNA and have the same end goal, what is the difference?
PFGE is an older technique used by the CDC, and what it does is separate large DNA molecules, in other words, the whole genome. This is the main process the CDC and the 82 state and local agencies used up until 2019, when WGS largely replaced it. While the PFGE only looks at approximately 15-30 bars in a certain DNA pattern at a time, WGS can look at millions, all the while comparing this newly-stored information to previous outbreaks.
The CDC compared the two techniques by saying, PFGE is like “looking at a few chapters of a book”, while WGS is like “looking at every word in the book”.
Furthermore, Whole Genome Sequencing is also known to be both more efficient, and much swifter. It allows epidemiologists, according to Zedic, “to more quickly identify genetically matched lab results from patients”, thus enabling health officials to identify the source of the outbreak more promptly than you would with PFGE, and to subsequently get the contaminated product off the shelves with more ease.
Both WGS and PFGE also use a system called Pulsenet, a national laboratory network that gathers the information from the 82 state and local agencies, and combines them, allowing for better communication. Beginning in 1996, Pulsenet, according to the CDC, has improved safety systems through identifying outbreaks early. This allows the investigators to find the source, alert the public sooner, and identify gaps in food safety systems that would not otherwise be recognized.
Since becoming the most widely used method in collecting DNA pertaining to foodborne illnesses, WGS has been used to solve a multitude of food poisoning outbreaks. To name a few, outbreaks associated with bacteria such as Campylobacter, Shiga toxin-producing E.coli (STEC), Salmonella, Vibrio, and Listeria have all been successfully investigated with WGS.
Since its claim to fame, WGS has not only improved surveillance, the CDC says, but it has also “enhanced our ability to detect trends in foodborne infections and antimicrobial resistance.” This is mainly due to the “detailed and precise” information this process yields.
How is it done, the people ask? According to the CDC, Scientists conduct whole genome sequencing by following these four main steps:
- DNA shearing: Scientists begin by using molecular scissors to cut the DNA, which is composed of millions of bases (A’s, C’s, T’s and G’s), into pieces that are small enough for the sequencing machine to read.
- DNA barcoding: Scientists add small pieces of DNA tags, or bar codes, to identify which piece of sheared DNA belongs to which bacteria. This is similar to how a bar code identifies a product at a grocery store.
- DNA sequencing: The bar-coded DNA from multiple bacteria is combined and put in a DNA sequencer. The sequencer identifies the A’s, C’s, T’s, and G’s, or bases, that make up each bacterial sequence. The sequencer uses the bar code to keep track of which bases belong to which bacteria.
- Data analysis: Scientists use computer analysis tools to compare sequences from multiple bacteria and identify differences. The number of differences can tell the scientists how closely related the bacteria are, and how likely it is that they are part of the same outbreak.
The PFGE technique, on the other hand, is much less complicated. While it does in fact “cut” the DNA like WGS does, the similarities end there. After the DNA has been “cut”, the DNA fragments for each isolate are embedded in a well at the top of a gel. The gel is then exposed to an electric field, which separates the fragments by size and charge, “creating a specific PFGE pattern”, says the Washington Department of Health.
Although this may seem easier, researchers have found that it is actually a much more lengthy task. On top of this, it is not nearly as affordable and is more technically challenging (therefore not as many laboratorians have the necessary training to complete the process).
Moving to WGS was a wise decision, one that undoubtedly saved lives. Yet, it is still important to recognize the part PFGE had to play in the investigation of foodborne outbreaks and the subsequent future development of WGS.