## 4 Discussion
In addition, only phylogenetic trees based on both ORF1 N-proximal and C-proximal clustered the isolates based on their host species (Figure 2). This evidence raises the question of whether RdRP (ORF1) is a better candidate for genetic diversity and phylogenetic studies of the emaraviruses compared to NP. Similar findings have been reported for the potato leafroll virus, which has a very conserved genome, and P0 was a better candidate in comparison to the coat protein for clustering the isolates [36]. More sequence isolates from different species of emaraviruses are needed to evaluate this idea. ASaV RNA3 and ASaV ORF3, which encode the nucleoproteins in all sequenced isolates, showed at least 91.97% and 90.93% identity at the nt level, despite differences in host plant origins and geographic distribution. As with other emaraviruses, the RNA3 identity varies across species. Blackberry leaf motile-associated virus (BLMaV) (based on ORF3), rose rosette virus (RRV) (based on ORF3), redbud yellow ringspot-associated virus (RYRSaV) (based on ORF3), and jujube yellow mottle-associated virus (JYMaV) (based on partial RNA3: 885 nt) exhibit the highest NP identity, reported to be 95% to 100% [18; 28; 30; 32]. The RNA3 identity in pear chloride leaf spot-associated virus (PCLSaV) (based on partial RNA3: 725 bp) isolates has a range from 91% to 99.9% [32]. In comparison, ASMaV (based on near full-length), and pigeonpea sterility mosaic virus 1 (PPSMV-1) and 2 (PPSMV-2) (based on partial RNA3: 947 bp and 995 bp) exhibit a sequence identity among isolates of 86.8% to 100% [21; 28]. All ASaV isolates could be categorized based on RNA3 and ORF3, making this genomic region a good candidate for phylogenetic analysis beside the ORF1 N-proximal. A phylogenetic analysis of RNA3 revealed three distinct clades, indicating significant genetic divergence despite the absence of host or geographic structuring. This divergence was supported by high \(F_{ST}\) values (\(>\) 0.6) and elevated synonymous substitution rates (\(K_{\mathrm{s}}\)\({}^{*}\) = 2.17-2.99). Both the Snn and \(Z^{*}\) statistics were significant (\(p\) < 0.01), highlighting strong genetic differentiation among clades, likely shaped by historical population bottlenecks, founder events, or selective pressures. While the lack of host specificity suggests free viral movement among _Faximus_ spp. trees, occasional long-distance dispersal may contribute to the observed geographic distribution of RNA3 clades. Notably, ASaV has only been detected in _Faximus_ spp., with an unconfirmed report from _Pisum sativum_[37], leaving its potential host range uncertain. Our findings also revealed that ASaV RNA3 and RNA5 were more divergent than other genomic RNA molecules (Table 2). These differences likely reflect distinct functional and selective constraints across the genome. The high identity in RNA2 and RNA4 suggests that these regions were under stronger evolutionary pressure to maintain essential functions. In contrast, the variability in RNA3 and RNA5 suggests their roles in viral adaptation to diverse host and environmental conditions. RNA3, which encodes the nucleocapsid protein, underscores its significance in diversity studies, with its variability potentially facilitating adaptation to different hosts. RNA5, showing higher genetic variability in comparison to the other genomic regions, may be involved in host interactions or regulatory functions, further highlighting its importance in viral evolution. RNA2 showed at least 92.75% identity, and it was under purifying selection (Table 2), which can explain the lower genetic diversity of RNA2. The RNA2 of emaraviruses is known to encode glycoprotein precursors (GPPs), which are predicted to contain N-terminal signal peptides. In all ASaV isolates, the putative cleavage site is located at residues 22-23, as also observed in ASMaV and PPSMV-1. Four N-glycosylation sites and four transmembrane helix (TMH) regions were predicted in all ASaV isolates, though the number of such sites and regions varies among different emaraviruses [19; 21].