ABSTRACT
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 clustered regularly interspaced short palindromic repeats (CRISPR)-based detection using LwaCas13a and PsmCas13b to simultaneously diagnose severe acute respiratory syndrome coronavirus 2 (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 the 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 that can be locally manufactured-may enable sustainable use of CRISPR diagnostics technologies for COVID-19 and other diseases in POC settings.
Subject(s)
COVID-19 , SARS-CoV-2 , Humans , SARS-CoV-2/genetics , COVID-19/diagnosis , COVID-19 Testing , Pandemics , Point-of-Care Systems , CRISPR-Cas Systems/genetics , Gene EditingABSTRACT
We performed a genome-wide association study on 377 cases of neovascular age-related macular degeneration (AMD) and 1074 controls to determine the association of previously reported genetic variants associated with neovascular AMD in the Thai population. All patients were of Thai ancestry. We confirmed the association of age-related maculopathy susceptibility 2 (ARMS2) rs10490924 (P=7.38 × 10-17), HTRA1 rs11200638 (P=5.47 × 10-17) and complement factor H gene (CFH) rs800292 (P=2.53 × 10-8) with neovascular AMD, all loci passing the genome-wide significance level (P<5.22 × 10-8). We also found association of the previously reported CFH rs10737680 (P=1.76 × 10-6) locus in the discovery sample. Two loci not previously reported to be associated with neovascular AMD were selected for replication in 222 cases and 623 controls. The loci included LINCO1317 rs6733379 and rs2384550 on chromosome 12. LINCO1317 rs6733379 (P=3.85 × 10-2) remained significantly associated with neovascular AMD after replication. In conclusion, we confirm that ARMS2, HTRA1 and CFH variants are associated with neovascular AMD in the Thai population. Findings from this study also suggest that variants contributing to the susceptibility of neovascular AMD in the Thai population are mostly similar to other Asians with additional local genetic risks that may specifically be identified in Thai population.
