Divergent peanut stunt virus isolates from black locust (Robinia pseudoacacia L.) in Slovakia retain infectivity across multiple leguminous hosts

Abstract Peanut stunt virus (PSV; genus Cucumovirus , family Bromoviridae ) was identified infecting black locust ( Robinia pseudoacacia L.) in Slovakia, representing an unusual host for this legume-infecting virus. Using high-throughput sequencing, three nearly complete tripartite genomes of PSV (designated AG6, AG19, and AG25) were obtained from naturally infected black locust trees. These isolates showed high mutual nucleotide identity (96.8–99.0%) but approximately 20% divergence from typical PSV isolates originating from legume hosts. Phylogenetic analyses placed the Slovak isolates within a distinct clade together with previously reported black locust PSV isolates from Hungary, supporting host-associated divergence. Additional RT-PCR screening revealed that 7 of 30 tested black locust trees were PSV-positive. All infected trees exhibited symptoms, while no asymptomatic PSV infections were detected. Notably, the black locust isolates were not detected using commercially available polyclonal anti-PSV antibodies, likely due to substantial divergence in capsid protein amino acid sequences. Biological assays demonstrated that black locust isolate AG6 remained infectious to several typical Fabaceae hosts following mechanical inoculation, including peanut, common bean, pea, and soybean, as well as Nicotiana benthamiana , with cultivar-dependent symptom expression observed in common bean and pea. Furthermore, PSV-associated satellite RNA was detected in two of seven PSV-positive samples, and its complete genomic sequence was determined, representing the first report of this satellite RNA in black locust. Altogether, these findings demonstrate the presence of genetically and serologically distinct PSV isolates in R. pseudoacacia and provide new insights into PSV diversification and host adaptation.