Abstract
During the current pandemic of COVID-19, the authors observed that during screening test for SARS-CoV-2 targeting the E-gene by qRT-PCR, few nasopharyngeal/oropharyngeal samples showed amplification signals at late cycle threshold (C T -value) > 35 despite being negative for other confirmatory target genes. Thirty such samples (taken as cases) showing detectable C T of > 35 cycle in E-gene which were negative for other target genes of SARS-CoV-2 and 30 samples with undetectable fluorescence in E-gene were taken as controls for investigation. An in-vitro diagnostic approved commercial qRT-PCR multiplex kit detecting 33 respiratory pathogens which can also detect Haemophilus influenzae was used for screening the samples. It was observed that out of the 30 samples showing detectable C T > 35 in E-gene, 11 samples were positive for Haemophilus influenzae whereas in the controls only three samples were positive for H. influenzae (p-value: 0.03) which was statistically significant. Further, the probes and primers were screened against H. influenzae for matches in the genome. It was observed that all primers and probes for the E-gene of SARS-CoV-2 had over 13 bp long sequences matching 100% with multiple sites across the H. influenzae genome. This qRT-PCR primer & probes are being used extensively across India, and laboratories using them should be aware of the cross-reactivity of primers & probes with the H. influenzae genome. Further, the authors observed that 95.9% (5415/5642) of COVID-19 positive cases detected in their laboratory were asymptomatic at the time of collection of samples. This warrants further investigations.
Keywords: COVID-19; E-gene; Haemophilus influenzae; SARS-CoV-2; qRT-PCR.
【저자키워드】 COVID-19, SARS-CoV-2, qRT-PCR, E-gene, Haemophilus influenzae, 【초록키워드】 qRT-PCR, Genome, India, diagnostic, Laboratory, cross-reactivity, amplification, Asymptomatic, cycle threshold, Control, Pathogens, multiplex, screening test, Target genes, E-gene, Haemophilus influenzae, In-vitro, target gene, P-value, respiratory pathogen, sequence, probes, primer, positive, Primers, H. influenzae, pandemic of COVID-19, Haemophilus, probe, was used, detect, detectable, approved, screened, statistically significant, undetectable,