ANDRZEJ KRAUZEIn June 2009, two male patients were independently admitted to the Heartland Regional Medical Center in northwestern Missouri with fever, headache, muscle pain, nausea, and diarrhea—all classic signs of ehrlichiosis, a common tick-borne disease in the region. Although both men reported having recently been bitten by ticks, blood and serum samples sent to microbiologist William Nicholson, chief of Pathogen Biology and Disease Ecology at the Centers for Disease Control and Prevention (CDC), came back negative for Ehrlichia chaffeensis, the disease-causing bacterium.
Nevertheless, Nicholson says, when the researchers plated the samples over a culture of canine tumor cells, they started to see signs of a pathogen. First, they noticed increased vacuole formation in the cells. “When we see that, within a day or two we usually see Ehrlichia,” Nicholson explains. But in this case, no Ehrlichia appeared, and the cells eventually began to fall apart. Then, the single layer of cells that lined the bottom of the flask started to detach earlier than normal—within 6–7 days, instead of 2 weeks. Nicholson and his colleagues continued to transfer the cells to fresh media, “and then it’d do it again,” he says. “That was an indication that we have something in there, we just can’t see it.”
After finding none of the various pathogen culprits familiar to the researchers, Nicholson’s group turned to their colleagues in the CDC’s electron microscopy (EM) department for help. When they got the transmission EM images back, “you could clearly see the cell just loaded with virus,” he says. “That was a nice bit of detective work,” says Sam Telford, an epidemiologist at Tufts University.
Based on the virus’s shape and size, the researchers suspected it belonged to the family Bunyaviridae. To get a more precise identification, Nicholson turned to Laura McMullan in the CDC’s viral special pathogens group, which had recently purchased a 454 sequencer. Its sequence revealed the virus to be a novel Bunyaviridae species belonging to the genus Phlebovirus, and the researchers named it the Heartland virus (HRTV) following the convention of naming viruses after their region of origin, which was coincidentally the name of the hospital where it was discovered (N Engl J Med, 367:834-41, 2012).
The next step was to determine the virus’s vector. Interestingly, the closest known relative of HRTV was the severe fever thrombocytopenia syndrome virus (SFTSV), a tick-borne Phlebovirus identified in 2011 after causing several cases of severe fever in China. Indeed, with both Missouri patients having reported tick bites, the researchers suspected that HRTV might also be carried by the arachnids.
In April, June, and August 2012, Nicholson and his colleagues collected more than 56,000 ticks of various species and life stages from several sites in northwestern Missouri, including the farms of both HRTV patients. They froze them in vials and sent them off to the CDC center at Fort Collins, Colorado, for molecular analysis. Sure enough, some of the ticks—specifically nymphs of the lone star tick Amblyomma americanum—carried HRTV, including those found at the farm of one of the patients (Am J Trop Med Hyg, doi:10.4269/ajtmh.13-0209, 2013). All told, however, the virus was relatively rare, Nicholson says, estimated to be present in about 1 in every 500 ticks. For comparison, Ehrlichia is found in some 10 percent of ticks. This rarity could explain why no virus was found in the ticks at the second farm, where the researchers were not able to collect nearly as many animals.
The researchers suspect that the ticks are becoming infected from the blood meal they ingest as larvae, after which they fall to the ground and burrow into the soil, where they will develop and molt into the nymphal stage. Then, when the nymphs emerge in the spring looking for their next meal, they can pass the infection on to people. Of course, “this is speculation based on the fact that we’re getting these hot ticks in the spring,” Nicholson says.
“It just goes to show that the diversity of potential pathogens carried by ticks is fairly large.—Sam Telford, Tufts University
To get more answers, the team has been out in the field again this year, and is expanding its search for the virus from just ticks to the vertebrates that A. americanum generally feeds on, such as wild turkey, deer, raccoons, and gray squirrels.
As for the virus’s origin, “none of us believe that this is a new introduction,” says Telford. More likely, “it’s been under our noses all along. It just goes to show that the diversity of potential pathogens carried by ticks is fairly large.”
One possible explanation for the virus’s recent emergence as a disease-causing pathogen, then, is the country’s changing demographic. “The American population as a whole is aging,” Telford notes. “Previously, maybe something like this was infecting perfectly healthy younger farmers in Missouri, and they just sort of shrugged it off.” Indeed, the two case patients were 57 and 67 years old. “It’s a pattern that we’ve seen in infectious biology all along—that as people age they become immune-compromised and far more susceptible to severe disease,” says Telford, who in 1997 discovered a flavivirus carried by deer ticks—which also transmit Lyme disease—that has shown up on the radars of epidemiologists only in the last 5 years.
“It’s much more than just a story of pathogen discovery and a new threat from ticks,” he adds. “I think the more interesting stuff is how these [pathogen] communities evolved, where they come from, and what are the things that lead us to recognize them as potential causes of disease.”