Exploring the Diversity of Plant DNA Viruses and Their Satellites Using Vector-Enabled Metagenomics on Whiteflies

Current knowledge of plant virus diversity is biased towards agents of visible and economically important diseases. Less is known about viruses that have not caused major diseases in crops, or viruses from native vegetation, which are a reservoir of biodiversity that can contribute to viral emergence. Discovery of these plant viruses is hindered by the traditional approach of sampling individual symptomatic plants. Since many damaging plant viruses are transmitted by insect vectors, we have developed “vector-enabled metagenomics” (VEM) to investigate the diversity of plant viruses. VEM involves sampling of insect vectors (in this case, whiteflies) from plants, followed by purification of viral particles and metagenomic sequencing. The VEM approach exploits the natural ability of highly mobile adult whiteflies to integrate viruses from many plants over time and space, and leverages the capability of metagenomics for discovering novel viruses. This study utilized VEM to describe the DNA viral community from whiteflies (Bemisia tabaci) collected from two important agricultural regions in Florida, USA. VEM successfully characterized the active and abundant viruses that produce disease symptoms in crops, as well as the less abundant viruses infecting adjacent native vegetation. PCR assays designed from the metagenomic sequences enabled the complete sequencing of four novel begomovirus genome components, as well as the first discovery of plant virus satellites in North America. One of the novel begomoviruses was subsequently identified in symptomatic Chenopodium ambrosiodes from the same field site, validating VEM as an effective method for proactive monitoring of plant viruses without a priori knowledge of the pathogens. This study demonstrates the power of VEM for describing the circulating viral community in a given region, which will enhance our understanding of plant viral diversity, and facilitate emerging plant virus surveillance and management of viral diseases

Generation and characterization of three monoclonal antibodies useful in detecting and distinguishing bean golden mosaic virus isolates

Two monoclonal antibodies prepared to purified virions of bean golden mosaic virus (BGMV) isolates from Guatemala (GA) and the Dominican Republic (DR) and one monoclonal antibody prepared to the coat protein of a BGMV isolate from Brazil (BZ) expressed in Escherichia coli were useful serological probes for detection of whitefly-transmitted geminiviruses. Abutilon mosaic, BGMV isolates from Puerto Rico (PR), and Homestead, Florida (H), BGMV-BZ, -DR, and -GA isolates, euphorbia mosaic, rhynchosia mosaic, squash leaf curl, soybean yellow mosaic, an isolate from Macroptilium lathyroides in Florida, and tomato mottle geminiviruses all reacted efficiently in Western blot and enzyme-linked immunosorbent assay with monoclonal antibody 3F7, indicating that this monoclonal antibody has a broad spectrum of reactivity. Another monoclonal antibody (2G5) reacted only with BGMV-DR, -GA, and -PR isolates. A third monoclonal antibody (5C5) reacted only with BGMV-BZ. A new geminivirus infecting cabbage (Brassicaceae) in Florida did not react with any of the three monoclonal antibodies