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Enhanced rabies surveillance in roadkill specimens by real-time RT-PCR
Crystal M Gigante1, Claire Hartloge1, Rene Edgar Condori1
1Poxvirus and Rabies Branch, Division of High-Consequence Pathogens and Pathology, United States of America Centers for Disease Control and Prevention, Atlanta, Georgia, United States of America.
Plos Neglected Tropical Diseases
|July 18, 2025
Summary
Real-time reverse transcriptase PCR (RT-PCR) effectively detects rabies virus RNA in degraded roadkill samples, improving wildlife rabies surveillance. This method identifies rabid animals that antigen detection tests would miss.
Area of Science:
- Veterinary Virology
- Wildlife Disease Ecology
- Molecular Diagnostics
Background:
- Roadkill specimens are crucial for enhanced rabies surveillance (ERS) where other sample collection is difficult.
- The physical state of roadkill often prevents accurate rabies diagnosis using traditional antigen detection methods.
- Real-time reverse transcriptase PCR (RT-PCR) offers high sensitivity for detecting viral RNA in compromised samples.
Purpose of the Study:
- To evaluate the efficacy of real-time RT-PCR for detecting rabies virus RNA in roadkill specimens.
- To assess the suitability of RT-PCR for rabies surveillance using samples unsuitable for antigen-based tests.
Main Methods:
- Collected 299 roadkill specimens (raccoons, skunks, foxes, coyotes, bobcat, domestic cat) from 2018-2021 across ten US states.
- Utilized the LN34 pan-lyssavirus real-time RT-PCR assay to test samples for rabies virus RNA.
- Compared RT-PCR results with limitations of antigen detection methods like direct fluorescent antibody test (DFA) and direct rapid immunohistochemistry test (DRIT).
Main Results:
- Eight samples (2.7%) tested positive for rabies virus RNA using the real-time RT-PCR assay.
- The positive cases involved animals from areas critical for wildlife rabies management.
- These positive findings would likely have been missed by conventional diagnostic methods due to sample quality.
Conclusions:
- Real-time RT-PCR is a valuable tool for rabies virus detection in roadkill samples, even those in poor condition.
- This molecular method significantly enhances the capacity for rabies surveillance in wildlife populations.
- RT-PCR provides a reliable alternative to antigen-based tests when dealing with challenging sample matrices in rabies control programs.
