Background
Begomoviruses are dicot-infecting, whitefly-transmitted viruses with a genome comprised of one or two molecules of circular, single-stranded DNA. In Brazil, tomato-infecting begomoviruses have emerged as serious pathogens since the introduction of a new biotype of the insect vector in the mid-1990's. Tomato rugose mosaic virus (ToRMV) and Tomato severe rugose virus (ToSRV) are often found in tomato fields. The complete sequence of the DNA-B components of ToSRV and ToRMV show an identity of 98.2%. Additionally, the high nucleotide identity (96.2%) between their common regions indicates that these two viruses may share the same DNA-B.
Conclusions
ToRMV and ToSRV form viable pseudorecombinants in their natural host (Solanum lycopersicum) and share the same DNA-B. ToRMV DNA components are preferentially replicated over ToSRV components. These results indicate that the emergence of ToRMV involved both recombination and pseudorecombination, further highlighting the importance of these mechanisms in the emergence and adaptation of begomoviruses.
Methods
Tomato seedlings were biolistically inoculated with ToSRV (DNA-A and DNA-B) and ToRMV (DNA-A and DNA-B) infectious clones in every possible combination of single or mixed infection. Symptom expression was evaluated for up to 35 days post-inoculation (dpi). DNA was extracted at 28 dpi and the presence of each viral genomic component was examined by rolling circle amplification (RCA) followed by digestion, as well as by quantitative, real-time PCR. Sequence comparisons, recombination and phylogenetic analyzes were performed using EMBOSS needle, RDP program and maximum likelihood inference, respectively.
Results
Symptoms in tomato plants inoculated with the different combinations of ToRMV and ToSRV DNA-A and DNA-B components consisted of a typical mosaic in all combinations. Pseudorecombinants were formed in all possible combinations. When two DNA-A or two DNA-B components were inoculated simultaneously, the ToRMV components were detected preferentially in relation to the ToSRV components. The combination of minor changes in both the Rep protein and the CR may be involved in the preferential replication of ToRMV components. Recombination and phylogenetic analyzes support the exchange of genetic material between ToRMV and ToSRV. Conclusions: ToRMV and ToSRV form viable pseudorecombinants in their natural host (Solanum lycopersicum) and share the same DNA-B. ToRMV DNA components are preferentially replicated over ToSRV components. These results indicate that the emergence of ToRMV involved both recombination and pseudorecombination, further highlighting the importance of these mechanisms in the emergence and adaptation of begomoviruses.