Effect of Holding Conditions on the Detection of Chikungunya and Venezuelan Equine Encephalitis Viruses in Mosquito Pools.

This research investigates how varying storage temperatures affect the detectability of viruses in mosquito samples. The study finds that while heat can lower the presence of active viruses, the viral RNA (genetic material) in the samples remains detectable in all conditions. This finding could ease viral surveillance efforts, as it suggests that maintaining a cold chain may not be crucial.

Experimental Design

• The study involved two specific species of mosquitoes: Aedes albopictus and Aedes taeniorhynchus.
• Each type of mosquito was infected with a virus: the chikungunya virus was introduced to the Aedes albopictus, while the Venezuelan equine encephalitis virus was used for the Aedes taeniorhynchus.
• The mosquitoes were stored at 26°C for 8 days post-infection.
• Single infected mosquitoes were then added to vials of 24 non-infected mosquitoes.
• The resulting groups, or “pools”, were stored at various temperatures (-80°C, -20°C, 4°C, 22°C, or 35°C) for up to 14 days.

Virus and RNA Detection Method

• The pools were analyzed in two ways: for the presence of infectious viruses via plaque assay, and for viral RNA using a method known as reverse transcriptase-quantitative polymerase chain reaction (RT-qPCR).
• Infectious viruses are those that are still capable of causing infection, while the RNA is the genetic material of the virus that provides instructions for replication.
• The cycle threshold (Ct) values, an indicator of the detected viral load, were also noted.

Findings

• The results showed that at higher storage temperatures, the amount of infectious virus diminished rapidly.
• However, samples stored at 4°C or lower demonstrated a more stable virus count.
• Contrarily, the viral RNA detected via RT-qPCR was present in pools held at all tested temperatures and holding times.
• The Ct values increased as temperatures and holding times increased, indicating a decrease in detected viral load.

Conclusion and Implications

• This study’s findings indicate that although higher temperatures may reduce the number of infectious viruses, viral RNA detection is still possible.
• Consequently, if the goal is to detect viral RNA rather than infectious viruses, arbovirus surveillance can be achieved without the necessity of maintaining a strict cold chain.
• This suggests that practical field-based virus monitoring programmes could be made more feasible, especially in areas where cold storage might not be available.