DNA-Based Analysis Detects Group A Streptococcus

In clinical microbiology, standard cultivation methods are considered the ‘gold standard’ for microorganism detection because the phenotypic characterization of clinical isolates is important for determining the treatment directions, but such methods are laborious and time-consuming.

Medical microbiologist at the Tokyo Medical and Dental University (Japan) and their colleagues collected throat swab samples from October 2013 to June 2014 from students and staff from various departments of the university. Two throat swabs were collected from the upper pharyngeal region of each of the 148 volunteers. One swab was dipped into 500 μL of sterile Trypticase soy broth for the culture-dependent study. The other swab was dipped into 600 μL of the sterile bead solution from the PowerSoil DNA isolation kit (MO BIO Laboratories, Carlsbad, CA, USA) and stored.

Two methods, culture dependent and independent, were used to detect GAS in throat swabs from healthy adults. For the culture-dependent method, swabs were streaked onto two types of agar medium and in addition to streaking; each throat swab dipped tryptic soy broth (TSB) and was also added to 1.2 mL of fresh TSB for enrichment. For the culture-independent study DNA extracted from the other was amplified by polymerase chain reaction (PCR) with two GAS-specific primer pairs: one was a newly designed 16S rRNA-specific primer pair, the other a previously described V-Na+-ATPase primer pair. The cloned PCR products were sequenced using M13 primers and the 3130 Genetic Analyzer (Applied Biosystems, Foster City CA, USA).

The scientists found that although only five (3.4%) of the 148 samples were GAS-positive by the culture-dependent method, 146 (98.6%) were positive for the presence of GAS DNA by the culture-independent method. To obtain serotype information by M type protein gene (emm) typing, they performed nested PCR using newly designed emm primers. They detected the four different emm types in 25 (16.9%) samples, and these differed from the common emm types associated with GAS associated diseases in Japan. The different emm types detected in the healthy volunteers indicate that the presence of unique emm types might be associated with GAS carriage.

The authors concluded that culture-independent methods should be considered for profiling GAS in the healthy hosts, with a view to obtaining better understanding of these organisms. The GAS-specific primers (16S rRNA and V-Na+-ATPase) used in this study can be used to estimate the maximum potential GAS carriage in people. These results suggest that not only school-age children, but also healthy adults can serve as GAS carriers. The study was published on October 11, 2016, in the journal BMC Microbiology.

Related Links:
Tokyo Medical and Dental University
MO BIO Laboratories
Applied Biosystems