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Development of a new quantitative RT-PCR to detect lymphocytic choriomeningitis virus
Laura Herrero1,2, Nuria Labiod1,2, Francisca Molero1
1Centro Nacional de Microbiología, Instituto de Salud Carlos III, Madrid, Spain.
Abstract:
Lymphocytic choriomeningitis virus (LCMV) is a neglected rodent-borne virus, with a worldwide distribution. The common mouse Mus musculus acts as reservoir and vector in the biological cycle of the virus. Surveillance of LCMV infection in mice is of importance as they are a widely used animal model in research and, through contact with them or their fluids, humans can be infected. Although most human cases are asymptomatic, LCMV infection can cause mild to severe, even fatal, and new diagnostic tools need to be developed to improve its detection. In the present work we report the development of a new method for the detection of LCMV RNA by quantitative reverse transcription polymerase chain reaction (RT-qPCR), able to detect all LCMV strains described to date. RT-qPCR targeting the S segment was developed and evaluated. Specificity and sensitivity were determined, and its limit of detection (LOD) was defined. The method designed is able to detect all 5 LCMV lineages described to date, with a LOD of 5.6 genome copies/μL. Its design with a built-in internal amplification control allows the detection of false negative results. Other arenaviruses were found not to cross-react with the method designed. In conclusion, a new diagnostic RT-qPCR for the detection of LCMV have successfully designed and validated. Improved detection techniques allow to reduce the turnaround time in the diagnosis of infections and to improve epidemiological surveillance in humans and animals.
Insights
A new quantitative reverse transcription polymerase chain reaction (RT-qPCR) method effectively detects all Lymphocytic choriomeningitis virus (LCMV) strains. This advancement improves diagnostic accuracy and epidemiological surveillance for this neglected zoonotic virus.
Area of Science:
- Virology
- Molecular Biology
- Public Health
Background:
- Lymphocytic choriomeningitis virus (LCMV) is a globally distributed, neglected virus transmitted by rodents.
- Mice (Mus musculus) serve as reservoirs and vectors, posing risks to research and human populations.
- Existing LCMV diagnostic tools require improvement for enhanced detection and surveillance.
Purpose of the Study:
- To develop and validate a novel quantitative reverse transcription polymerase chain reaction (RT-qPCR) assay for LCMV detection.
- To ensure the assay can identify all known LCMV strains and lineages.
- To enhance diagnostic capabilities for LCMV surveillance in both animal models and humans.
Main Methods:
- Development of an RT-qPCR assay targeting the S segment of the LCMV genome.
- Evaluation of the assay's specificity, sensitivity, and limit of detection (LOD).
- Inclusion of an internal amplification control to identify false negatives.
Main Results:
- The developed RT-qPCR method successfully detected all five known LCMV lineages.
- The assay demonstrated a low limit of detection (LOD) of 5.6 genome copies/μL.
- The method showed no cross-reactivity with other arenaviruses and included an internal control for result validation.
Conclusions:
- A highly sensitive and specific RT-qPCR assay for LCMV detection has been successfully designed and validated.
- This new diagnostic tool can improve the speed and accuracy of LCMV diagnosis.
- The validated assay will aid in enhanced epidemiological surveillance of LCMV in humans and animals.
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