SPOTTED OUT: A new assay test for HIV drug resistance colors individual virus molecules as either resistant [red] or non-resistant [green] and can test the same sample for resistance to different drugs [left v. right; see arrows]. Image: COURTESY OF FENG GAO
A new type of test may markedly improve the ability to detect potentially drug-resistant HIV infections using a patient's blood sample. The technique works by scanning individual virus molecules for genetic mutations that are known to confer immunity to existing treatments.
Although the test still has to prove it can benefit large numbers of patients and work in a real-world setting, "this to me looks faster, cheaper and easier" than existing assays for HIV drug resistance, says HIV/AIDS researcher Douglas Richman of the University of California, San Diego, who was not involved in the research. "So it may be a significant improvement."
In general, drug resistance occurs when some fraction of the viruses or bacteria causing an infection are immune to a drug. (Researchers do not know the minimum amount for HIV.) As a result, instead of eradicating the infection, the drug neutralizes the sensitive pathogens and allows the resistant ones to multiply, rendering the treatment ineffective.
In the U.S. and Europe, 10 to 20 percent of new HIV infections carry at least one drug-resistant mutation. Researchers have therefore developed assays to identify populations of resistant viruses in a patient's blood. Existing commercial assays may fail to spot the potential for drug resistance, however, because they can only detect resistant viruses if they make up 20 percent or more of those in a sample.
"Sooner or later [HIV patients] are going to generate enough resistant virus to make the patient fail the treatment," says Feng Gao of Duke University Medical Center, leader of the group that developed the new assay. "If you choose the drugs wisely, you can delay the resistance."
The test that Gao and his colleagues designed can spot drug-resistant HIV at levels of 0.1 to 0.01 percent, they report in this month's Nature Methods. In principle, the assay should be able to scan a single sample for all 30 or so known resistance mutations, says Gao, whose group hopes to start a company to commercialize the technology by 2009.
The researchers smeared each of 13 test samples from HIV-positive individuals on to its own polymer gel, which held the sample's virus particles in place. Next, they introduced a short piece of DNA that attached to the virus's genetic material adjacent to where a particular drug-resistant mutation would occur. To distinguish resistant from sensitive strains, they added two additional units of DNA (called bases), each labeled with a fluorescent marker of a different color. A resistant strain would incorporate one of the bases, making its location on the gel glow one color; a drug-sensitive strain would take up the other base and glow another color.
The researchers were able to repeat the assay on the same virus particles but for different drug-resistance mutations, in essence allowing them to identify individual viruses resistant to multiple drugs, they report.
For the time being, Richman says, "it's not going to replace any of the commercial assays right now because they are high-throughput," meaning they can be performed rapidly and at low cost. "From a research tool that looks very promising to a commercially viable assay there's a lot steps."