How are ticks tested for encephalitis? - briefly
Laboratory analysis of ticks involves extracting RNA and performing reverse‑transcriptase PCR to detect viral genomes linked to encephalitis. Positive samples are confirmed by sequencing or virus isolation in cell culture.
How are ticks tested for encephalitis? - in detail
Ticks collected from endemic areas are first sorted by species and developmental stage, because vector competence varies among taxa. Each specimen is placed in a sterile tube, kept on dry ice or at –80 °C until processing to preserve viral RNA.
The laboratory workflow proceeds as follows:
- Homogenisation – individual or pooled ticks are ground in a buffered solution containing proteinase inhibitors. Mechanical disruption (bead mill or mortar) ensures release of intracellular particles.
- Nucleic‑acid extraction – silica‑column or magnetic‑bead kits isolate total RNA. Extraction controls (e.g., spiked RNA) verify efficiency.
- Reverse transcription quantitative PCR (RT‑qPCR) – primer‑probe sets targeting conserved regions of encephalitis‑causing flaviviruses (e.g., TBEV, Powassan virus) amplify viral genomes. Amplification curves are interpreted against positive, negative and no‑template controls.
- Sequencing confirmation – amplicons from positive reactions undergo Sanger or next‑generation sequencing to determine strain identity and detect mutations.
- Virus isolation (optional) – homogenates from RT‑qPCR‑positive ticks are inoculated onto susceptible cell lines such as Vero or BHK‑21. Cytopathic effect is monitored, and immunofluorescence staining with virus‑specific antibodies confirms replication.
- Serological assays (optional) – enzyme‑linked immunosorbent assay (ELISA) or immunoblotting on tick extracts detects viral antigens when nucleic‑acid methods are inconclusive.
Quality‑assurance measures include duplicate testing, inclusion of extraction blanks, and periodic proficiency testing with reference materials. Results are reported as presence or absence of viral RNA, with cycle‑threshold values indicating viral load, and, when available, genotype information from sequencing data. This systematic approach provides reliable detection of encephalitic viruses in tick vectors.