RESUMO
Point-of-care (POC) nucleic acid detection technologies are poised to aid gold-standard technologies in controlling the COVID-19 pandemic, yet shortcomings in the capability to perform critically needed complex detection--such as multiplexed detection for viral variant surveillance--may limit their widespread adoption. Herein, we developed a robust multiplexed CRISPR-based detection using LwaCas13a and PsmCas13b to simultaneously diagnose SARS-CoV-2 infection and pinpoint the causative SARS-CoV-2 variant of concern (VOC)-- including globally dominant VOCs Delta (B.1.617.2) and Omicron (B.1.1.529)--all while maintaining high levels of accuracy upon the detection of multiple SARS-CoV-2 gene targets. The platform has several attributes suitable for POC use: premixed, freeze-dried reagents for easy use and storage; convenient direct-to-eye or smartphone-based readouts; and a one-pot variant of the multiplexed detection. To reduce reliance on proprietary reagents and enable sustainable use of such a technology in low- and middle-income countries, we locally produced and formulated our own recombinase polymerase amplification reaction and demonstrated its equivalent efficiency to commercial counterparts. Our tool--CRISPR-based detection for simultaneous COVID-19 diagnosis and variant surveillance which can be locally manufactured--may enable sustainable use of CRISPR diagnostics technologies for COVID- 19 and other diseases in POC settings.
RESUMO
@#Introduction: Rapid diagnosis for influenza virus infection is essential for proper patient management, delivering prompt treatment and reducing unnecessary antiviral therapy. Early diagnosis helps in disease prevention and control. Real-time reverse transcription-polymerase chain reaction (RT-PCR) assay yields high sensitivity and specificity in detecting influenza virus infection. However, it is relatively expensive and requires trained personnel and special equipment. In this study, we compared two rapid influenza diagnostic tests (RIDTs): digital readout systems (STANDARD™ F Influenza A/B FIA, fluorescence immunoassay) and conventional visual confirmation (QuickNavi™-Flu2, chromatography immunoassay) with the real-time RT-PCR assay. Methods: Two hundred ninety-eight respiratory samples were obtained from patients suspected of influenza infection at Siriraj Hospital from December 2018 to December 2019. Results: Real-time RT-PCR results showed the detection of influenza A virus in 99 samples (60%), influenza B virus in 61 samples (37%) and co-infection by both viruses in 5 samples (3%) by the real-time RT-PCR assay. The QuickNavi™-Flu2 sensitivity for detecting influenza A and B viruses were 81.73% and 84.85%, and the specificity was 100%. The STANDARD™ F Influenza A/B FIA sensitivity for detecting influenza A and B viruses were 84.62% and 83.33%, respectively. The specificity for influenza A virus detection was 99.25% and 94.74% for influenza B virus. Conclusion: The STANDARD™ F Influenza A/B FIA and the QuickNavi™-Flu2 showed acceptable and comparable sensitivity and specificity. Both RIDTs are potential alternative methods of real-time RT-PCR for rapid screening of influenza virus infection.
RESUMO
Objective@#To investigate the distribution of human papillomavirus (HPV) genotypes in low-grade squamous intraepithelial lesion (LSIL) cytology and the immediate risk of cervical intraepithelial neoplasia grade 2 or higher (CIN2+) lesions. @*Methods@#This prospective cross-sectional study enrolled women aged ≥21 years that were diagnosed with LSIL cytology at Siriraj Hospital (Bangkok, Thailand) during 2017-2019. Anyplex II HPV testing was performed to detect 14 high-risk HPV cases prior to colposcopy-directed biopsy. @*Results@#In total, 318 patients were included in the final analysis. Of those, 24 (7.5%), 241 (75.8%), 53 (16.7%) were aged 21- 25 years, 25-50 years, and ≥50 years, respectively. Eighty-two patients (25.8%) had abnormal screening results within the previous 5 years. High-risk HPV infection was found in 188 patients (59.1%) with 127 (39.9%) having single and 61 (19.2%) having multiple infections. The five most common HPV genotypes were HPV 66 (18.6%), HPV51 (9.7%), HPV58 (9.4%), HPV16 (9.1%), and HPV56 (8.2%). The immediate risk of CIN2+ was 6% in LSIL, regardless of the HPV status, 8% in high-risk HPV-positive LSIL, and 3.1% in high-risk HPV-negative LSIL. When using 6% as the threshold risk for colposcopy, performing reflex HPV testing in LSIL cytology can decrease the number of colposcopies by 40.9%, with an area under the receiver operating characteristic curve of 0.6 (95% confidence interval, 0.5-0.7). @*Conclusion@#The study findings support the idea that geographic variations affect the HPV genotype. Reflex HPV testing may decrease the number of colposcopies in cytology-based screening regions with a high prevalence of low-carcinogenic HPV.
