A Map of the Whole Genome Tracks Outbreaks
During April and May this year, 14 people in the United States were infected by an outbreak of food-borne E. coli. One of them died, and public health officials are still hunting for the source of the outbreak.
Such detective work normally involves using a decades-old DNA fingerprinting technique to identify and trace the strains of microbes responsible. But a technology for whole-genome analysis could lead to quicker identification and more comprehensive analysis. More broadly, the technique could help researchers better understand the structure of genomes—disease-related or not—and the order of genes within.
Food-borne infections are typically caused by strains of bacteria that produce a potentially deadly toxin. Investigators use enzymes to cut suspect DNA and create a ladderlike pattern of DNA chunks that is unique to that strain. By cross-referencing the pattern against a library of known microbes, they can finger the bacterial culprit. The technique does not, however, keep track of the cut pieces of DNA in order. That limits investigators' ability to understand the genetic content of the pathogenic microbes.
An enhanced version of a technique known as optical mapping could offer a more ordered look at the guilty genome. Optical mapping involves stretching single pieces of DNA across a glass plate and then cutting them with sequence-specific enzymes. The tension in the DNA means that when it is cut, it recoils a bit, leaving easy-to-see gaps; because the DNA is affixed to a glass plate, researchers can add the order of cut sites to their detective work. That way, when DNA-staining fluorescent dye lights up the cut genetic material, a machine can measure the length of each piece, and computational analysis can create a map of the genome.
"We can create ordered maps based on the actual order of the sequence that exists in that particular chromosome," says Doug White, CEO of OpGen, which sells the optical mapping technology.
The method is not just useful for public health officials. Although the field of genomics is strewn with the phrase "whole-genome sequencing," DNA sequencing machines do not give a complete picture of the whole genome. The sequences are produced in bits and pieces that usually cannot be assembled into a complete chromosome. By combining genome sequencing with optical mapping, researchers can achieve a fuller picture.
Several genome sequencing centers already use optical mapping to get closer to a truly whole genome. The position of genes in a microbe's DNA can affect their function, which is an important consideration for genes that lead to antibiotic resistance or toxin production.
Furthermore, optical mapping can reveal repeated sequences and other structural quirks in a genome that high-throughput sequencing technologies can miss. In the analysis of the sequence data generated by those technologies, the pieces of identical sequence can be mistakenly assumed to belong to the same spot in the chromosome, resulting in an incorrect number of copies of the same sequence being represented in the final genome sequence. But because optical mapping measures the true length of chromosomes, genome scientists can determine the true number of repeats found within a chromosome.
While high-throughput sequencing technologies can analyze pieces of DNA from about 75 to 1,000 base pairs in length, optical mapping gives information in the length of 250,000 base pairs, according to OpGen. The company's chief scientific officer, Rich Moore, says the technology "gives long-range information across the entire genome."
Peter Gerner-Smidt, chief of the CDC's Enteric Diseases Laboratory, says the cost of the technology has kept it out of reach in public health. But the CDC is currently using optical mapping to facilitate the annotation of whole genomic sequences, and the agency is "exploring if there is a place for the technology as a supplement in our surveillance," he says.
Last month, OpGen announced that a dozen public health agencies, including the CDC, had joined a consortium to evaluate what role the company's technology could play in enhanced genotyping of outbreak-causing bacteria.
The technique has been used by public health officials elsewhere. In 2011, officials in Germany took just 48 hours to determine that an E. coli outbreak blamed for infecting some 850 people and killing 32 came from a single source of a unique strain of the bacteria, indicated by a unique and consistent pattern in the optical map.
Since it was founded in 2002, OpGen has focused on the genomes of microbes. But now it says its data analysis has improved to the point where the large chromosomes of humans can be mapped. Cancer-related changes in the genome often include chromosomal rearrangements that can alter the function of genes responsible for regulating cell growth and division. Some neurological diseases, such as Huntington's disease and a form of Lou Gehrig's disease, are caused by DNA repeats that extend to the point of disrupting cellular function.
The number of repeats often correlates with the rate of disease progression, says Albert La Spada, a clinician-scientist at the University of California, San Diego, who studies Huntington's and other repeat-based diseases. Existing high-throughput DNA sequencing can miss abnormal repeat numbers, he says, but optical mapping is well suited to the task. And because the technique looks at a single chromosome at a time, researchers could be better informed about which parent passed on a repeat-based disease. They could also identify genetic variants near the repeat expansion that could also play a role in the disease. "This type of approach could fill a niche," La Spada says.