A Mismatch-Tolerant LAMP a visual determination method by adding colorimetric dyes. Because of its simplicity, all LAMP-based diagnostic assays may be easily updated to the newly improved version. The novel mismatch-tolerant LAMP method represents a simple, sensitive and promising approach for molecular diagnosis of highly variable viruses, and it is especially suited for application in resource-limited settings.
Human coronaviruses HCoV-OC43, HCoV-229E, HCoV-NL63 and HCoV-HKU1 are common respiratory viruses associated with acute respiratory infection. They have a global distribution. Rapid and accurate diagnosis of HCoV infection is important for the management and treatment of hospitalized patients with HCoV infection. Here, we developed a melting curve-based multiplex RT-qPCR assay for simultaneous detection of the four HCoVs. In the assay, SYTO 9 was used to replace SYBR Green I as the fluorescent dye, and GC-modified primers were designed to improve the melting temperature (Tm) of the specific amplicon. The four HCoVs were clearly distinguished by characteristic melting peaks in melting curve analysis. The detection sensitivity of the assay was 3 × 102 copies for HCoV-OC43, and 3 × 101 copies for HCoV-NL63, HCoV-229E and HCoV-HKU1 per 30 μL reaction. Clinical evaluation and sequencing confirmation demonstrated that the assay was specific and reliable. The assay represents a sensitive and reliable method for diagnosis of HCoV infection in clinical samples.
Background: The emergence and spread of HIV-1 drug resistance may compromise HIV control globally. In response to HIV/AIDS epidemic, China launched national HIV/AIDS treatment program in 2003, and started to accumulate drug resistance data since 2001. In this study we aimed to assess the level, trend and distribution of HIV-1 drug resistance during a period of 17 years from 2001 to 2017, and to characterize crucial drug resistance mutations. Methods: We systematically reviewed 4737 studies published between January 1, 2001 and March 31, 2019 in PubMed, Embase, China National Knowledge Infrastructure (CNKI), WanFang Database, Web of Science, conference abstracts from the Chinese Medical Association and the Chinese AIDS Academic Conferences, and selected 170 studies that met our study criteria. To assess the prevalence of drug resistance in whole country or a local region, we performed pooled analyses of raw data. The transformed proportions were pooled using the inverse variance fixed effects methods or the DerSimonian-Laired random effects methods. The temporal trend of transmitted drug resistance (TDR) was determined using generalized additive model implemented in the Mgcv version 1.8 package. HIV-1 genotypic resistance was analyzed using the Stanford HIVdb algorithm. Findings: We assembled 218 datasets from 170 selected studies (129 in Chinese and 41 in English), covering 21,451 ART-naïve and 30,475 ART-treated individuals with HIV-1 infection. The pooled prevalence of TDR was 3.0% (95%CI: 2.8À3.2), including 0.7% (95%CI: 0.4À1.0), 1.4% (95%CI: 1.3À1.6) and 0.5% (95%CI: 0.4À0.6) for nucleoside reverse transcriptase inhibitor (NRTI), non-NRTI (NNRTI) and protease inhibitor (PI) resistance, respectively. The acquired drug resistance (ADR) prevalence was 44.7% (95%CI: 39.3À50.2), including 31.4% (95%CI: 28.2À34.6), 39.5% (95%CI: 35.6À43.5) and 1.0% (95%CI: 0.8À1.2) for NRTI, NNRTI and PI resistance, respectively. TDR and ADR prevalence had characteristic regional patterns. The worst prevalence of drug resistance occurred in Central China, and higher ADR prevalence occurred in South China than North China. TDR in whole country has risen since 2012, and this rise was driven mainly by NNRTI resistance. One NRTI-associated (M184V/I) and three NNRTI-associated (K103N/S, Y181C/I and G190A/S) mutations had high percentages in ART-naïve and ARTtreated individuals, and these mutations conferred high-level resistance to 3TC, EFV and/or NVP. Interpretation: These findings suggest that the current available first-line ART regimens containing 3TC and/ or EFV or NVP need to be revised. In addition, scale-up of multiple viral load measurements per year and drug resistance testing prior to ART initiation are recommended. Furthermore, implementation of pre-treatment education and counseling to improve patient adherence to ART is encouraged.
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