Agdia (United States)

Tomato brown rugose fruit virus (ToBRFV) causes severe crop losses worldwide, infecting primarily tomato (Solanum lycopersicum) and pepper (Capsicum annuum). It was detected for the first time in 2014 in Israel (Luria et al., 2017), then in 2015 in Jordan (Salem et al., 2016), and has since spread rapidly to other countries. In the Americas, it was reported in Mexico (Cambrón-Crisantos et al., 2018) and USA (Ling et al., 2019). In December 2022, greenhouse-grown tomato plants from three different growers from Santa Lucía and Lavalle (Corrientes, Argentina) showed similar symptoms to those caused by tobamoviruses. The plants showed mosaic and leaf mottling, narrowing (needle-like) and chlorosis in young leaves. The fruits exhibited necrotic lesions and blotchy ripening (Figures 1-3). More than 50% of the plants in the affected greenhouses showed symptoms, the large incidence being consistent with mechanical transmission of the disease.

Effective use of diagnostic assays is essential for the early detection of plant pathogens and mitigation of potential disease impacts. Assay developers require a full understanding of the intended use of a test to address complicating factors that might be observed by an end user and limit the utility of the test and its scope of application in the field. The fitness of a test for a disease prevention application is determined by its performance characteristics, which are selected during assay design and defined during validation. This paper provides guidance to developers by standardizing the descriptions of key validation terms and concepts, including tiers that can be referenced in publications to better communicate the extent to which a test has been validated. These concepts are then applied in the broader context of a strategic approach to validation for various taxa and methods. The aim of this paper is to increase awareness of common pitfalls and gaps encountered during this process, with the goal of increased success in technology transfer. Recommendations are given for improving the efficiency and quality of test development through improved coordination among stakeholders. [Formula: see text] Copyright © 2023 The Author(s). This is an open access article distributed under the CC BY 4.0 International license .

Potato mop-top virus (PMTV) is considered an emerging threat to potato production in the United States. PMTV is transmitted by a soil-borne protist, Spongospora subterranean. Rapid, accurate, and sensitive detection of PMTV in leaves and tubers is an essential component in PMTV management program. A rapid test that can be adapted to in-field, on-site testing with minimal sample manipulation could help in ensuring the sanitary status of the produce in situations such as certification programs and shipping point inspections. Toward that goal, a rapid and highly sensitive recombinase polymerase amplification (RPA)-based test was developed for PMTV detection in potato tubers. The test combines the convenience of RPA assay with a simple sample extraction procedure, making it amenable to rapid on-site diagnosis of PMTV. Furthermore, the assay was duplexed with a plant internal control to monitor sample extraction and RPA reaction performance. The method described could detect as little as 10 fg of PMTV RNA transcript in various potato tissues, the diagnostic limit of detection (LOQ) similar to that of traditional molecular methods.

Diagnostic test results are used for many purposes and are heavily relied upon to prevent movement of pathogens from one country or region to another; to clear plants as free from disease for commerce; and to surveil against natural, accidental, or unintentional pathogen introductions and new and reemerging diseases. Diagnostic test results are also used by plant producers to make critical management decisions. The level of confidence in each assay is defined by a set of metrics that describe the performance of the assay under defined conditions. Collectively, these metrics are called “validation” of the assay. There are hundreds of diagnostic plant disease assays used every day in the United States that either are not validated or are validated in an ad hoc way. An endemic pathogen in one region can be an exotic pathogen in another. Therefore, this multitude of diagnostic assays for which performance criteria are not standardized impairs communication about confidence in the test outcome. We propose to create a framework for standardizing assay validation language and definitions within the United States across existing plant diagnostic networks. The long-term goal is to have operable standards, understanding the “trueness” of assay results, and sustained communications between diagnostic laboratories that use and those that develop plant disease diagnostic assays.

A rapid and reliable molecular diagnostic assay, AmplifyRP Acceler8, was developed for the direct detection of Fusarium oxysporum f. sp. vasinfectum (FOV) race 4, a virulent genotype of the Fusarium wilt pathogen of cotton (Gossypium spp.), in soil and cotton tissue. Unlike traditional polymerase chain reaction (PCR) assays, the recombinase polymerase amplification based-assay described here utilizes an advanced isothermal technology where the amplification is carried out at a single constant temperature, 39°C, without the need of a thermal cycler. The AmplifyRP Acceler8 diagnostic assay consistently detected FOV race 4 from all infected tissue samples. The test is rapid, simple, and more sensitive than conventional PCR. The AmplifyRP Acceler8 diagnostic assay detected DNA from FOV race 4 at concentrations of 1 ng/μL and above. In addition, it did not amplify DNA from other known FOV races (races 1, 2, 3, 6, and 8). The whole process from sample preparation to reading the results can be completed in as little as 30 min. The test can detect FOV race 4 from cotton taproots, petioles, and stems. Accepted for publication 6 January 2014. Published 15 March 2014.