Meridian Bioscience's Illumigene nucleic acid amplification test for Streptococcus pyogenes, or Group A Streptococcus, has a clinical sensitivity of 99 percent and specificity of 99.6 percent when compared to the diagnostic gold standard of culture and discrepant analysis using another nucleic acid amplification test, according to a study published last week in the Journal of Clinical Microbiology.
The results of the study validate Meridian's test as an "answer to the challenge" of developing improved laboratory tests to supplant current methods of diagnosing GAS pharyngitis using patient symptoms and clinical algorithms followed by culture-based confirmation, according to its authors.
In addition, Illumigene's high sensitivity, specificity, reproducibility, and fast turnaround time make it an "attractive alternative" to other rapid diagnostic tests for GAS, such as antigen-detection tests and laboratory-developed PCR-based assays, the authors said.
Meridian's Illumigene GAS test uses loop-mediated isothermal amplification, or LAMP, to target a highly conserved 206-bp sequence of the S. pyogenes pyrogenic exotoxin gene, SpeB, in throat swab samples. Nucleic acid amplification is detected indirectly by the presence of pyrophosphate that precipitates during elongation of the DNA template and is detectable using Meridian's Illumipro-10 device. In general, the assay can provide results in less than one hour.
The test received regulatory clearance from the US Food and Drug Administration in September, making it the only FDA-approved nucleic acid amplification test for the bacteria (PCR Insider, 9/20/2012).
Current methods for diagnosing GAS and distinguishing it from viral pharyngitis are based on physical examination by a physician usually followed by culture verification using nasophayngeal swab specimens.
However, the clinical manifestations of GAS pharyngitis and viral pharyngitis are similar, and although culture-based methods are highly sensitive, they can take up to two days to produce a result.
Meridian is betting that its Illumigene test can quickly produce at least a comparably accurate result, allowing physicians to determine more or less on the spot whether to begin antibiotic treatment.
"Current clinical practice for GAS detection is: if a patient has GAS symptoms, a rapid diagnostic test will be performed at a doctor’s office or sample will be submitted to a laboratory for culture," Slava Elagin, executive vice president of research and product development at Meridian Bioscience, told PCR Insider in an email.
"In case of [a] negative rapid test, a sample will be submitted to a laboratory for culture test," Elagin said. "The whole idea for the Illumigene GAS test is to eliminate culture confirmation … and provide rapid [turnaround time]. Current culture will take one to three days for a final result. Meanwhile, a physician might do empiric treatment or wait for a result — meaning a kid will go to school and spread a disease."
Other rapid diagnosis methods exist, including point-of-care antibody-based detection of the Group A carbohydrate antigen. This assay has a turnaround time of minutes, and is highly specific, but can fall short in terms of sensitivity compared to culture-based approaches.
Rapid molecular testing methods include the GAS Direct Test, a hybridization-based assay from Hologic's Gen-Probe business; and the LightCycler Strep-A assay from Roche, which uses PCR-based DNA amplification to detect GAS in patient specimens. However, the former has only modest sensitivity and specificity when compared with culture methods, and requires multiple instruments and steps; while the latter is in use only as a laboratory-developed test in the US, hindering its widespread adoption. And neither one is as rapid as the Illumigene assay.
In the JCM study, which received funding from Meridian, a group led by clinical researchers from the Medical College of Wisconsin evaluated the performance of the Illumigene assay in 796 pharyngeal swabs collected at three separate clinical centers.
The researchers compared results of the test with direct and extracted culture methods using a routine "direct" culture method and an enriched "extracted" culture method, which provides greater sensitivity than routine culture. In addition, the researchers resolved any discrepant results using the laboratory-developed LightCycler Strep-A assay.
The Illumigene assay detected GAS in 74 of 74 direct culture-positive specimens and 100 of 102 extracted culture-positive specimens. Illumigene also detected an additional 42 specimens that were direct culture-negative and 16 specimens that were extracted culture-negative.
Discrepant analysis using the PCR-based assay detected GAS nucleic acid in 13 of 16 "false positive" specimens and one of two "false negative" specimens, resulting in an overall sensitivity of 99 percent and specificity of 99.6 percent for detecting GAS in pharyngeal swabs using Illumigene.
Two specimens reported as negative by Illumigene were found to be positive by enriched culture, and both were negative by routine culture, suggesting, the researchers wrote, that the failure of Illumigene to detect GAS in these samples was the result of a low concentration of organisms.
Of the 16 apparent false positive results reported by Illumigene, 13 were also positive using the PCR assay. The researchers noted that these 13 samples may represent the presence of GAS nucleic acid in the absence of viable organisms "from patients with resolving infections or already undergoing antimicrobial therapy." They also noted that current rapid antigen POC tests also do not differentiate between viable and non-viable organisms.
The authors underscored several limitations of the Illumigene assay as a sole means for diagnosing pharyngitis.
"While the assay is very sensitive, it will only identify pharyngitis caused by GAS," they wrote. "Culture is still needed to detect other causes of pharyngitis (i.e. Group C Streptococcus or Group G Streptococcus, and viral etiologies)." Though not as prevalent as GAS, these non-GAS streptococci are responsible for up to 5 percent of the total cases of pediatric pharyngitis, they noted.
In addition, Illumigene is a qualitative assay, and is thus "not able to distinguish between active infection and low-level or asymptomatic carriage of GAS as part of the normal pharyngeal flora," the researchers wrote.
This, they noted, is also a limitation of antigen-based tests, but cautioned that pharyngeal carriage rates have been reported to be as high as 33 percent in some regions, and that positive results in these cases may not warrant antibiotic administration.
Despite these limitations, the researchers said that the performance of the Illumigene test is a boon, especially in light of guidelines recently issued by the Infectious Diseases Society of America that recommended the diagnosis of GAS pharyngitis be made by laboratory results rather than by clinical algorithms.
"This is a challenge given the poor sensitivity of currently available rapid tests," the researchers wrote. "In fact, improvement of rapid tests for the detection of GAS pharyngitis was specifically identified as an area in need of further research."
Illumigene GAS, however, is rapid and accurate; has high sensitivity and specificity; and is easy to perform and provides reproducible results among different users in different settings, making it applicable to a variety of clinical environments, they noted.
"These aspects make it a useful diagnostic tool, an attractive alternative to other rapid diagnostic tests for the GAS pharyngitis, and an answer to the challenge of rapid test improvement put forth by recent IDSA guidelines," the researchers concluded.