TY - JOUR
T1 - Integrated approach for detection of SARS-CoV-2 and its variant by utilizing LAMP and ARMS-PCR
AU - Nawab, Maryam
AU - Riaz, Syeda Kiran
AU - Ismail, Eiman
AU - Ahamed, Alfar
AU - Tariq, Aaysha
AU - Malik, Muhammad Faraz Arshad
AU - Qusty, Naeem F.
AU - Bantun, Farkad
AU - Slama, Petr
AU - Umair, Massab
AU - Haque, Shafiul
AU - Bonilla-Aldana, D. Katterine
AU - Rodriguez-Morales, Alfonso J.
N1 - Publisher Copyright:
© 2024, The Author(s).
PY - 2024/12
Y1 - 2024/12
N2 - Global impact of COVID-19 pandemic has heightened the urgency for efficient virus detection and identification of variants such as the Q57H mutation. Early and efficient detection of SARS-CoV-2 among densely populated developing countries is paramount objective. Although RT-PCR assays offer accuracy, however, dependence on expansive kits and availability of allied health resources pose an immense challenge for developing countries. In the current study, RT-LAMP based detection of SARS-Cov-2 with subsequent confirmation of Q57H variant through ARMS-PCR was performed. Among the 212 collected samples, 134 yielded positive results, while 78 tested negative using RT-LAMP. Oropharyngeal swabs of suspected individuals were collected and processed for viral RNA isolation. Isolated viral RNA was processed further by using either commercially available WarmStart Master Mix or our in house developed LAMP master mix separately. Subsequently, the end results of each specimen were evaluated by colorimetry. For LAMP assays, primers targeting three genes (ORF1ab, N and S) were designed using PrimerExplorer software. Interestingly, pooling of these three genes in single reaction tube increased sensitivity (95.5%) and specificity (93.5%) of LAMP assay. SARS-CoV-2 positive specimens were screened further for Q57H mutation using ARMS-PCR. Based on amplicon size variation, later confirmed by sequencing, our data showed 18.5% samples positive for Q57H mutation. Hence, these findings strongly advocate use of RT-LAMP-based assay for SARS-CoV-2 screening within suspected general population. Furthermore, ARMS-PCR also provides an efficient mean to detect prevalent mutations against SARS-Cov-2.
AB - Global impact of COVID-19 pandemic has heightened the urgency for efficient virus detection and identification of variants such as the Q57H mutation. Early and efficient detection of SARS-CoV-2 among densely populated developing countries is paramount objective. Although RT-PCR assays offer accuracy, however, dependence on expansive kits and availability of allied health resources pose an immense challenge for developing countries. In the current study, RT-LAMP based detection of SARS-Cov-2 with subsequent confirmation of Q57H variant through ARMS-PCR was performed. Among the 212 collected samples, 134 yielded positive results, while 78 tested negative using RT-LAMP. Oropharyngeal swabs of suspected individuals were collected and processed for viral RNA isolation. Isolated viral RNA was processed further by using either commercially available WarmStart Master Mix or our in house developed LAMP master mix separately. Subsequently, the end results of each specimen were evaluated by colorimetry. For LAMP assays, primers targeting three genes (ORF1ab, N and S) were designed using PrimerExplorer software. Interestingly, pooling of these three genes in single reaction tube increased sensitivity (95.5%) and specificity (93.5%) of LAMP assay. SARS-CoV-2 positive specimens were screened further for Q57H mutation using ARMS-PCR. Based on amplicon size variation, later confirmed by sequencing, our data showed 18.5% samples positive for Q57H mutation. Hence, these findings strongly advocate use of RT-LAMP-based assay for SARS-CoV-2 screening within suspected general population. Furthermore, ARMS-PCR also provides an efficient mean to detect prevalent mutations against SARS-Cov-2.
KW - ARMS-PCR
KW - ORF1ab
KW - Q57H
KW - RT-LAMP
KW - SARS-CoV-2
UR - http://www.scopus.com/inward/record.url?scp=85183712229&partnerID=8YFLogxK
U2 - 10.1186/s12941-023-00665-0
DO - 10.1186/s12941-023-00665-0
M3 - Original Article
C2 - 38303011
AN - SCOPUS:85183712229
SN - 1476-0711
VL - 23
JO - Annals of Clinical Microbiology and Antimicrobials
JF - Annals of Clinical Microbiology and Antimicrobials
IS - 1
M1 - 11
ER -