| dc.description.abstract | The tick-borne encephalitis virus (TBEV), classified as an arbovirus within the Flavivirus genus, is responsible for tick-borne encephalitis. Ticks serve as main vector for TBEV. TBE is a serious disease condition in humans and several animal species. TBE has been a rising health concern in recent decades, not just in Europe but increasingly all over the world. The number of TBE cases seems to be increasing over time in Europe including Norway. Thus, the main purpose of the present study was to estimate the TBEV prevalence at Kilen, Mandal in southern Norway from questing nymphs collected in 2021.This study also addresses an amplification method and troubleshooting techniques to develop a method for WGS of low viral loads of TBEV in nymph samples.
In the present study a total of 518 nymph pools were screened and analyzed for TBEV. An overall TBEV prevalence of 0.7 % was estimated which is different from prevalence in 2018 and 2019 for the same site, when no TBEV was detected. This increase and change in prevalence indicated the monthly and yearly variation of TBEV prevalence in endemic regions of Norway.
The present study was also aimed to develop a WGS method of low virus load TBEV samples but, the study faced issues in obtaining desired 400bp PCR product for WGS method. Several amplification and troubleshooting experiments i.e., Inhibition tests and dilution tests were conducted out to explain the lack of required 400bp PCR product from nymph cDNA samples for WGS.
The result showed that there was no inhibition in the samples tested and the viral load in the samples may be too low to produce PCR product by the current PCR conditions. Spike-in methods were found helpful in improving PCR products. The study also found that cDNA generated by both the reverse transcription technique, High-Capacity cDNA, and Superscript IV cDNA, performs well with small fragment length overlapping primer pairs of the TBEV-Eu strain.
In conclusion, this study shows that there is a monthly and yearly variation of TBEV in Kilen Mandal. The developed amplification and troubleshooting procedures from this study may be further investigated to obtain an improved and desired PCR products for sequencing of whole viral genome of TBEV samples with low viral load. | |
