# Characterization of a Novel Emaravirus Affecting Ash Species (_Fraximus_ spp.) in Europe
## 3 Results
### Genetic Variability of ASaV Based on Partial Sequences  
A total of 118 partial ASaV PCR amplicons (RNA3 = 52, RNA4 = 17, RNA5 = 49) from infected _F. excelsior_ and _F. ornus_ were analyzed by pairwise sequence comparisons to assess genetic variability. All partial sequences lie within the coding region of the respective genome segment. Genetic distances calculated in MEGA-X are listed in Table 5.  
Results demonstrate a high level of conservation of partial sequences at both the nucleotide (nt) and amino acid (aa) level, with mean genetic dista  
\begin{table}
\begin{tabular}{c c c c} \hline \hline
**Location** & **Host Species** & **Samples** & **Symptomatic/ASaV** & **Asymptomatic/ASaV** \\  & & & **Infected** & **Infected** \\ \hline Germany 1 & _F. excelsior_ & 5 & 5/4 & - \\ Germany 2 & _F. ornus_ & 44 & 34/34 & 10/1 \\ Italy & _F. ornus_ & 11 & 9/9 & 2/2 \\ Sweden & _F. excelsior_ & 39 & 29/26 & 10/6 \\ Switzerland & _F. excelsior_ & 8 & 6/6 & 2/0 \\  & _F. excelsior_ & 52 & 40/36 (90\%) & 12/6 (50\%) \\ Total & _F. ornus_ & 57 & 45/44 (98\%) & 12/3 (25\%) \\  & Total & 109 & 85/80 (94\%) & 24/9 (38\%) \\ \hline \hline \end{tabular} 1  
\end{table}
Table 4: Detection of putative emaravirus ash shoestring-associated virus (ASaV) in symptomatic and asymptomatic ash samples, without suspected virus symptoms, collected in 2016–2019 at locations across Europe.  
\begin{table}
\begin{tabular}{c c c c c c c c c c c} \hline \hline
**Host Species and** & & & & & & & & & \\
**Genome** & **RNA** & **n 1** & **Nt 2** & **Min 3** & **Max 3** & **Mean** & **SEM 5** & **aa** & **Min 4** & **Max 4** & **Mean** & **SEM 5** \\  & & & & & & & & & & & \\ \hline _F. excelsior, F. ornus_ & & 52 & & 0 & 0.096 & 0.030 & 0.001 & & 0 & 0.063 & 0.012 & 0.001 \\ \hline _F. excelsior_ & 3 & 18 & 297 & 0 & 0.088 & 0.031 & 0.002 & 99 & 0 & 0.063 & 0.011 & 0.002 \\ \hline _F. omus_ & & 34 & & 0 & 0.089 & 0.025 & 0.001 & & 0 & 0.063 & 0.012 & 0.001 \\ \hline _F. excelsior, F. ornus_ & & 17 & & 0 & 0.062 & 0.025 & 0.001 & & 0 & 0.034 & 0.004 & 0.001 \\ \hline _F. excelsior_ & 4 & 10 & 453 & 0 & 0.055 & 0.026 & 0.003 & 151 & 0 & 0.034 & 0.007 & 0.002 \\ \hline _[PERSON]_ & & 7 & & 0.002 & 0.044 & 0.025 & 0.003 & & 0 & 0 & 0 \\ \hline _[PERSON], [PERSON]_ & & 49 & & 0 & 0.086 & 0.039 & 0.001 & & 0 & 0.107 & 0.040 & 0.001 \\ \hline _[PERSON]_ & 5 & 17 & 357 & 0 & 0.072 & 0.027 & 0.002 & 119 & 0 & 0.097 & 0.033 & 0.002 \\ \hline _[PERSON]_ & & 32 & & 0 & 0.086 & 0.036 & 0.001 & & 0 & 0.106 & 0.033 & 0.002 \\ \hline \hline \end{tabular} 1
[FOOTNOTE:1]Footnote 1: footnotetext: \({}^{\text{1}}\) Number of sequences analyzed; 2
[FOOTNOTE:2]Footnote 1: footnotetext: \({}^{\text{2}}\) Fragment length excluding primers; 3
[FOOTNOTE:3]Footnote 1: footnotetext: \({}^{\text{3}}\) Genetic distance calculated by maximum composite likelihood model implemented in Mega-X; 4
[FOOTNOTE:4]Footnote 4: footnotetext: \({}^{\text{4}}\) Genetic distance calculated by equal input model implemented in Mega-X; 5  
\end{table}
Table 5: Comparison of genetic distances of ASaV RNA3, RNA4 and RNA5 partial nucleotide and amino acid sequences. Pairwise comparisons conducted on all partial sequences for each genome segment (_F. excelsior, F. ornus_) and for sequences originating from each host species separately (_F. excelsior_ or _F. ornus_).  
0.039 (standard error 0.001-0.003) and 0-0.04 (standard error 0-0.002) at the nt and aa level, respectively. The results of this first variability analysis indicated there is no correlation between genetic variability and geographical location or ASaV host species.