Prevalence and Molecular Analysis of Encephalomyocarditis Virus-2 in the Hazel Dormouse

Abstract

The hazel dormouse (Muscardinus avellanarius) population in the UK continues to decline due to habitat loss, despite reintroductions of captive-bred individuals being conducted nationally for over 30 years. Disease surveillance of captive-bred and wild dormice is performed to identify novel and existing disease threats which could impact populations. In this study, we firstly investigated cause of death in seven hazel dormice found dead in England, through next-generation sequencing identifying a virus closely related to a wood mouse encephalomyocarditis virus-2 (EMCV-2). Subsequently, lung tissue samples from 35 out of 44 hazel dormice tested positive for EMCV-2 RNA using a reverse transcriptase quantitative polymerase chain reaction (RT-qPCR) and Sanger sequencing methods developed in this study. Formalin-fixed tissues available for nine hazel dormice which tested positive for EMCV-2 RNA were examined microscopically. Three cases showed moderate interstitial pneumonia with minimal to mild lymphoplasmacytic myocarditis, but no evidence of encephalitis. However, the presence of possible alternative causes of death in these cases means that the lesions cannot be definitively attributed to EMCV-2. Here, we report the first detection of EMCV-2 in hazel dormice and conclude that EMCV-2 is likely to be endemic in the hazel dormouse population in England and may be associated with clinical disease.

Keywords: Cardiovirus; Muscardinus avellanarius; EMCV-2; Encephalomyocarditis virus; Hazel dormouse.

Figure 1

Phylogenetic analysis of a 379-bp DNA fragment of the encephalomyocarditis virus 3D polymerase gene. The DNA sequences were aligned with MUSCLE and the alignment was subjected to phylogenetic analysis using MEGA X software. The evolutionary history was inferred by using the maximum likelihood method, 500 bootstrap replicates and Tamura–Nei model (Tamura et al., 2021). The tree with the highest log likelihood (-2519.94) is shown. The percentage of trees in which the associated taxa clustered together is shown next to the branches. Initial tree(s) for the heuristic search were obtained automatically by applying Neighbor-Join and BioNJ algorithms to a matrix of pairwise distances estimated using the maximum composite likelihood (MCL) approach and then selecting the topology with superior log likelihood value. The tree is drawn to scale, with branch lengths measured in the number of substitutions per site. Cardiovirus B, GenBank accession number MK343442 was used as an outgroup. Bootstrap values above 50% are only shown in the tree.

Figure 2

a Distribution of 43 hazel dormice found dead in English counties and submitted for post-mortem examination. One hazel dormouse was submitted without location data. Counties with positive and negative EMCV-2 cases, detected by RT-qPCR on lung samples, are in blue and yellow, respectively. The number of positive cases and total number of submissions from the county are also reported in the map. b Map of English counties with and without reintroduction in the county.