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Phylogenetic diversity and molecular evolution of Hantaan virus harbored by Apodemus chejuensis on Jeju Island, Republic of Korea, 2022-2023.

BACKGROUND: Hantaan virus (HTNV), hosted by Apodemus spp., is a well-recognized causative agent of hemorrhagic fever with renal syndrome (HFRS) and poses a crucial global public health concern. Based on the current evidence, HTNV carried by A. chejuensis is proposed as the likely etiological agent of HFRS on Jeju Island, Republic of Korea (ROK). METHODOLOGY/PRINCIPAL FINDINGS: In this study, 50 small mammals were collected from five locations in Seogwipo-si and Jeju-si on Jeju Island, ROK, during 2022-2023. Serological and molecular analyses revealed HTNV prevalence rates of 34% (16/47) and 27.7% (13/47), respectively. Using a multiplex polymerase chain reaction-based nanopore sequencing approach, nine complete HTNV genomes were sequenced from the lung tissues of A. chejuensis, representing the first comprehensive genomic characterization of HTNV from Seogwipo-si (Hogeun-dong) and Jeju-si (Sangdae-ri). Phylodynamic analyses suggest evolutionary divergence and phylogeographic diversity, with four unique amino acid substitutions identified in HTNV genomes from Seogwipo-si. CONCLUSION/SIGNIFICANCE: This study provides important insights into the genomic surveillance, genetic diversity, and evolutionary dynamics of orthohantaviruses, which are essential for guiding effective public health strategies to control and prevent future HFRS outbreaks in the ROK.

Animals

Sensitive and Visualized Detection of Hantavirus Using CRISPR/Cas12a Based on AutoCORDSv2 Design.

In recent years, detection technologies based on the CRISPR/Cas12a method have been extensively utilized in the fields of nucleic acid, enzyme, and macromolecule detection, thereby reinforcing their significant role in the detection landscape. Enhancing the simplicity of design, efficiency, and automation of the CRISPR/Cas12a detection system is essential for advancing its application in diagnostics. Recently, we developed an automated CRISPR/Cas12a design system named AutoCORDSv2. This system can process published genomic sequences of pathogenic bacteria in a high-throughput manner and automatically generate conserved and highly specific crRNA sequences, along with primer sequences for target amplification. This capability facilitates the specific and precise design of the CRISPR/Cas12a detection system. In this study, crRNAs targeting the Hantaan virus (HTNV) and Seoul virus (SEOV), as well as RT-PCR primers and RT-RPA primers, were designed using AutoCORDSv2. The experimental results demonstrated that the CRISPR/Cas12a system, automatically designed by AutoCORDSv2, was specific for the detection of both the HTNV and SEOV, with no cross-reactivity observed with other pathogens. The detection sensitivity reached 6 copies/μL (equivalent to 111 copies per amplification reaction), whether measured by a microplate reader or directly observed with the naked eye. The detection results for 50 samples were consistent with those obtained from commercial RT-qPCR kits, indicating high precision. Furthermore, the CRISPR/Cas12a system designed by AutoCORDSv2 can also be utilized for the development of a single-tube detection system with a sensitivity of 42 copies per reaction. This system combined with a 5-min extraction step and RT-RPA, further underscoring its potential for application.

CRISPR-Cas Systems