Partial sequencing of a putative Alstroemeria necrotic streak orthotospovirus isolate detected on lettuce in Colombia

ABSTRACT Lettuce is the most cultivated leafy salad vegetable in Colombia, being the municipality of Madrid, in the Department of Cundinamarca, the second largest producer. In this region, lettuce plants with foliar symptoms characterized by brown necrotic spots forming an extended necrotic area, chlorosis, leaf distortion and plant stunting have been detected, possibly caused by a viral infection associated with the Orthotospovirus genus. This study aimed to identify the orthotospovirus species associated with those symptoms, contributing to updating the lettuce phytosanitary status in this region. The presence of orthotospovirus was confirmed by the enzyme-linked immunosorbent assay (DAS-ELISA), although the sequence of the nucleocapsid (N) gene confirmed the presence of Alstroemeria necrotic streak orthotospovirus, disregarding the Tomato spotted wilt orthotospovirus and Impatiens necrotic spot orthotospovirus previously reported for this crop, being this its first report in lettuce crops in Colombia.

Genome Analysis of Melon Necrotic Spot Virus Incursions and Seed Interceptions in Australia

Melon necrotic spot virus (MNSV) was detected in field-grown Cucumis melo (rockmelon) and Citrullus lanatus (watermelon) plants in the Sunraysia district of New South Wales and Victoria, Australia, in 2012, 2013, and 2016, and in two watermelon seed lots tested at the Australian border in 2016. High-throughput sequencing was used to generate near full-length genomes of six isolates detected during the incursions and seed testing. Phylogenetic analysis of the genomes suggests that there have been at least two incursions of MNSV into Australia and none of the field isolates were the same as the isolates detected in seeds. The analysis indicated that one watermelon field sample (L10), the Victorian rockmelon field sample, and two seed interception samples may have European origins. The results showed that two isolates (L8 and L9) from watermelon were divergent from the type MNSV strain (MNSV-GA, D12536.2) and had 99% nucleotide identity to two MNSV isolates from human stool collected in the United States (KY124135.1, KY124136.1). These isolates also had high nucleotide pairwise identity (96%) to a partial sequence from a Spanish MNSV isolate (KT962848.1). The analysis supported the identification of three previously described MNSV genotype groups: EU-LA, Japan melon, and Japan watermelon. To account for the greater diversity of hosts and geographic regions of the MNSV isolates used in this study, it is suggested that the genotype groups EU-LA, Japan melon, and Japan watermelon be renamed to groups I, II, and III, respectively. The divergent isolates L8 and L9 from this study and the stool isolates from the United States formed a fourth genotype group, group IV. Soil collected from the site of the Victorian rockmelon MNSV outbreak was found to contain viable MNSV and the virus vector, a chytrid fungus, Olpidium bornovanus (Sahtiyanci) Karling, 18 months after the initial MNSV detection. This is a first report of O. bornovanus from soil sampled from an MNSV-contaminated site in Australia.

Thrips as the Transmission Bottleneck for Mixed Infection of Two Orthotospoviruses

Mixed infections provide opportunities for viruses to increase genetic diversity by facilitating genomic reassortment or recombination, and they may lead to the emergence of new virus species. Mixed infections of two economically important orthotospoviruses, Tomato spotted wilt orthotospovirus (TSWV) and Impatiens necrotic spot orthotospovirus (INSV), were found in recent years, but no natural reassortants between INSV and TSWV were ever reported. The goal of this study was to establish how vector preferences and the ability to transmit INSV and TSWV influence transmission and establishment of mixed infections. Our results demonstrate that thrips prefer to oviposit on TSWV and INSV mixed-infected plants over singly infected or healthy plants, providing young nymphs with the opportunity to acquire both viruses. Conversely, we observed that thrips served as a bottleneck during transmission and favored transmission of one of the two viruses over the second one, or over transmission of both viruses simultaneously. This constraint was relaxed in plants, when transmission of TSWV and INSV occurred sequentially, demonstrating that plants serve as orthotospovirus permissive hosts, while thrips serve as a bottleneck. Viral fitness, as measured by virus replication, transmission, and competition with other viral strains, is not well studied in mixed infection. Our study looks at the success of transmission during mixed infection of orthotopoviruses, enhancing the understanding of orthotospovirus epidemiology and evolution.