Long-Term Monitoring of Aphid-Transmitted Viruses in Melon and Zucchini Crops: Genetic Diversity and Population Structure of Cucurbit Aphid-Borne Yellows Virus and Watermelon Mosaic Virus.

Understanding the emergence and prevalence of viral diseases in crops requires the systematic epidemiological monitoring of viruses, as well as the analysis of how ecological and evolutionary processes combine to shape viral population dynamics. Here, we extensively monitored the occurrence of six aphid-transmitted viruses in melon and zucchini crops in Spain for 10 consecutive cropping seasons between 2011 and 2020. The most prevalent viruses were cucurbit aphid-borne yellows virus (CABYV) and watermelon mosaic virus (WMV), found in 31 and 26% of samples with yellowing and mosaic symptoms. Other viruses, such as zucchini yellow mosaic virus, cucumber mosaic virus, Moroccan watermelon mosaic virus, and papaya ring spot virus, were detected less frequently (<3%) and mostly in mixed infections. Notably, our statistical analysis showed a significant association between CABYV and WMV in melon and zucchini hosts, suggesting that mixed infections might be influencing the evolutionary epidemiology of these viral diseases. We then carried out a comprehensive genetic characterization of the full-length genome sequences from CABYV and WMV isolates by using the Pacific Biosciences single-molecule real-time (PacBio) high-throughput technology to assess the genetic variation and structure of their populations. Our results showed that the CABYV population displayed seven codons under positive selection, and although most isolates clustered in the Mediterranean clade, a subsequent analysis of molecular variance revealed a significant, fine-scale temporal structure, which was in part explained by the level of the variance between isolates from single and mixed infections. In contrast, the WMV population genetic analysis showed that most of the isolates grouped into the Emergent clade, with no genetic differentiation and under purifying selection. These results underlie the epidemiological relevance of mixed infections for CABYV and provide a link between genetic diversity and CABYV dynamics at the whole-genome level.

Long-Term Cocirculation of Two Strains of Pepino Mosaic Virus in Tomato Crops and Its Effect on Population Genetic Variability.

Mixed viral infections are common in plants, and the evolutionary dynamics of viral populations may differ depending on whether the infection is caused by single or multiple viral strains. However, comparative studies of single and mixed infections using viral populations in comparable agricultural and geographical locations are lacking. Here, we monitored the occurrence of pepino mosaic virus (PepMV) in tomato crops in two major tomato-producing areas in Murcia (southeastern Spain), supporting evidence showing that PepMV disease-affected plants had single infections of the Chilean 2 (CH2) strain in one area and the other area exhibited long-term (13 years) coexistence of the CH2 and European (EU) strains. We hypothesized that circulating strains of PepMV might be modulating the differentiation between them and shaping the evolutionary dynamics of PepMV populations. Our phylogenetic analysis of 106 CH2 isolates randomly selected from both areas showed a remarkable divergence between the CH2 isolates, with increased nucleotide variability in the geographical area where both strains cocirculate. Furthermore, the potential virus-virus interaction was studied further by constructing six full-length infectious CH2 clones from both areas, and assessing their viral fitness in the presence and absence of an EU-type isolate. All CH2 clones showed decreased fitness in mixed infections and although complete genome sequencing indicated a nucleotide divergence of those CH2 clones by area, the magnitude of the fitness response was irrespective of the CH2 origin. Overall, these results suggest that although agroecological cropping practices may be particularly important for explaining the evolutionary dynamics of PepMV in tomato crops, the cocirculation of both strains may have implications on the genetic variability of PepMV populations.