A colorimetric reverse transcription-loop mediated isothermal amplification (RT-LAMP) method for the rapid detection of SARS-CoV-2 in Thailand.

COVID-19, caused by the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), remains a global health threat. Timely identification of infected cases is important for appropriate patient management and the control of viral spread. Simple and cost-effective tests are required to increase access to testing and early case detection. Here, we describe a colorimetric reverse transcription-loop-mediated isothermal amplification (RT-LAMP) method to detect SARS-CoV-2. The RT-LAMP could amplify the orf1ab sequence detectable by visual color change within 45 min at 63 °C. The limit of detection (LoD) for SARS-CoV-2 RNA was less than 100 copies (13.36) per reaction with no cross-amplification with other related viruses. Clinical evaluation using leftover RNA samples extracted from 163 nasopharyngeal swab specimens showed perfect agreement in negative (n = 124) and positive samples with cycle thresholds (Ct) < 34 cycles (n = 33) detected by real-time reverse transcription-polymerase chain reaction (RT-PCR), targeting RdRp and N genes as a reference. Overall, the diagnostic accuracy, sensitivity, specificity, positive and negative predictive values of RT-LAMP in testing were 96.32% (95% CI: 92.16-98.64%), 84.62% (95% CI: 68.47-94.14%), 100% (95% CI: 97.07-100.0%), 100% (95% CI: 89.42-100.0%), and 95.38% (95% CI: 90.22-98.29), respectively. This RT-LAMP assay is simple and reliable, with the potential to be an alternative for the rapid detection of SAR-CoV-2 with minimal time and fewer resources compared to real-time RT-PCR.

One step for Legionella pneumophila detection in environmental samples by DNA-gold nanoparticle probe.

AIMS: To develop and evaluate a DNA-gold nanoparticle (DNA-AuNP) probe assay to detect Legionella pneumophila, which causes Legionnaires' disease, compared with the gold standard culture method. METHODS AND RESULTS: Gold nanoparticles (AuNPs) were conjugated with DNA probes to detect the mip gene of L. pneumophila. The DNA-AuNP probe assay was evaluated for its specificity, sensitivity and stability. The results showed that only L. pneumophila mixed with this probe resulted in a red solution that was easily detected by the naked eye, and the colour was stable when 10 mmol l-1 MgSO4 was added. The 100 Legionella isolates and 10 other bacteria led to 100% specificity. Compared with the culture method, our method showed a 100% negative predictive value, 100% sensitivity (kappa = 0·87), and a detection limit of 4·5 ng DNA µl-1 with a 6-min response time for the 124 colonies suspected of being Legionella. The DNA-AuNP probe reagents were stable for more than 6 months. CONCLUSIONS: The developed DNA-AuNP probe assay has good negative predictive value, sensitivity, rapidity and ease of use, which is helpful for ruling out negative samples. SIGNIFICANCE AND IMPACT OF THE STUDY: The DNA-AuNP probe assay can detect the mip gene of L. pneumophila. Therefore, it may be an alternative method for screening colonies suspected of being L. pneumophila.