## 3 Results
### Analysis of the BSTV \(R\)-DNA Sequences  
Like the S-DNA, the R-DNA gene was also highly conserved. Except isolates BBTV_286_F2 and BBTV_273_F1, which presented identities between 93% and 96% when compared to each other, all the other isolates presented identities between 98% and 100%. When compared with the PIO isolates from GenBank, the identity of these two isolates ranged from 94% to 96%, while all other Mozambican isolates exhibited identities between 96% and 99%. On the other hand, when compared with the SEA isolates, the two isolates presented identities between 87 and 89%, while the others presented identities between 90 and 93%.  
When comparing the identities between the PIO isolates from GenBank, it was observed that they have 97% to 99% identity among themselves and between 90 and 93% identities with the SEA isolates. These results allowed us to infer that the isolates that occur in Mozambique are of the PIO type, i.e., they belong to the Indian-Pacific Oceans group.  
\begin{table}
\begin{tabular}{c c c} \hline \hline \multirow{2}{*}{**BSV Isolate Name**} & \multicolumn{3}{c}{**GenBank Accession Number**} \\ \cline{2-3}  & **R-DNA** & **S-DNA** \\ \hline BBTV 252 F1 & PP097007 & PP096947 \\ BBTV 260 F1 & PP097009 & PP096949 \\ BBTV 2  
The amino acid identities between the two isolates cited above (BBTV_273_F1 and BBTV_286_F2) and the other Mozambique isolates were lower than that of the nucleotides, between 91% and 93%, indicating that the nucleotide substitutions were nonsynonymous.  
The same was observed with isolate 3, whose nucleotide identity ranged from 94% to 99% and amino acid identity ranged from 91% to 97%. Most isolates presented identities of 100% when compared to each other and from 97% to 100% when compared to the isolates from the PIO group and 91% to 95% when compared to the isolates from the SEA group.  
Figure 3: Phylogenetic tree constructed based on the nucleotide sequence of the S-DNA of BBTV isolates collected in the district of Chókwé, Mozambique and isolates from GenBank. Bootstrap values were obtained using the MEGA 7.0 neighbor-joining program with 2000 repetitions.  
The phylogenetic tree based on the nucleotide sequences is illustrated in Figure 5. The isolates were grouped into two distinct clades, one with the isolates of the PIO group and the other with the isolates of the SEA group. All Mozambique isolates were grouped into a subclade with the Malawi isolate JQ820453. The other isolates of the PIO group were grouped in the same clade but in a distinct subclade.  
Figure 4: Phylogenetic tree constructed based on the amino acid sequence of the S-DNA of BBTV isolates collected in the district of Chökwé, Mozambique and isolates from GenBank. Bootstrap values were obtained using the MEGA 7.0 neighbor-joining program with 2000 repetitions.  
In the phylogenetic tree based on amino acid sequences (Figure 6), the distribution was slightly different. There was also a separation of the isolates into two clades, one with the isolates of the PIO group and the other with the isolates of the SEA group. However, in addition to isolate JQS20453 from Malawi, isolates KM607635 from DRC, JQS20465 from  
Figure 5: Phylogenetic tree constructed based on the nucleotide sequence of the R-DNA of BBTV isolates collected in the district of Chökwé, Mozambique and isolates from GenBank. Bootstrap values were obtained using the MEGA7.0 neighbor-joining program with 2000 repetitions.  
Rwanda and, a little further away, AF416467 from Tonga also grouped together with most of the Mozambican isolates. Another eight isolates from Mozambique were mixed with isolates from different parts of the world in different combinations. Both the identity and phylogenetic results support the classification of the studied isolates within the PIO group.