Detection of Ampelovirus and Nepovirus by Lab-on-a-Chip: A Promising Alternative to ELISA Test for Large Scale Health Screening of Grapevine.

Ilaria Buja,Vito Elicio,Andrea Luvisi,Giuseppe Maruccio,Erika Sabella,Silvia Rizzato,Anna Grazia Monteduro,Lilia Formica,Luigi De Bellis

doi:10.3390/bios12030147

Ilaria Buja, Vito Elicio + Show 7 more

Open Access

https://doi.org/10.3390/bios12030147

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Journal: Biosensors	Publication Date: Feb 27, 2022
Citations: 7	License type: CC BY 4.0

Affiliation: University of Salento, INFN Sezione di Lecce

Abstract

The Ampelovirus Grapevine leafroll-associated virus 3 (GLRaV-3) and the Nepovirus Grapevine fanleaf virus (GFLV) are pathogens reported in many grapevine-growing areas all over the world, main causal agents of grapevine leafroll disease and grapevine fanleaf disease, respectively. Prevention of virus spread thanks to rapid diagnosis of infected plants is a key factor for control of both diseases. Although serological (e.g., enzyme-linked immunosorbent assay-ELISA test) and molecular methods are available to reveal the presence of the viruses, they turn out to be quite expensive, time-consuming and laborious, especially for large-scale health screening. Here we report the optimization of a lab-on-a-chip (LOC) for GLRaV-3 and GFLV detection, based on an electrochemical transduction and a microfluidic multichamber design for measurements in quadruplicate and simultaneous detection of both targets. The LOC detect GLRaV-3 and GFLV at dilution factors more than 15 times higher than ELISA, providing a higher sensitivity in the detection of both viruses. Furthermore, the platform offers several advantages as easy-to-use, rapid-test, portability and low costs, favoring its potential application for large-scale monitoring programs. Compared to other grapevine virus biosensors, our sensing platform is the first one to provide a dose-dependent calibration curve combined with a microfluidic module for sample analysis and a portable electronics providing an operator-independent read-out scheme.

Highlights

Introduction iationsAmpelovirus and Nepovirus genera include some of the most important viral pathogens of the grapevine, such as Grapevine leafroll-associated virus 3 (GLRaV-3) and Grapevine fanleaf virus (GFLV), respectively [1,2] (Figure 1)
In this paper we report, for the first time, the detection of an Ampelovirus (GLRaV3) and a Nepovirus (GFLV) through a LOC device connected to a portable potentiostat, with simultaneous electrochemical impedance measurements
Assays were performed with a dilution in Phosphate Buffered Saline (PBS) considering dilutions resulting negative in ELISA tests

Summary

Introduction

Introduction iationsAmpelovirus and Nepovirus genera include some of the most important viral pathogens of the grapevine, such as Grapevine leafroll-associated virus 3 (GLRaV-3) and Grapevine fanleaf virus (GFLV), respectively [1,2] (Figure 1). GLRaV-3 belongs to the family Closteroviridae, considered one of the most widespread pathogens associated with grapevine leafroll disease (GLD) [3], caused by various GLRaV-named viruses GLRaV-3 symptoms are evident in red cultivars (which exhibit leaf reddening maintaining green venation) during late summer-fall, but not so apparent in white ones (with mild yellowing or chlorosis interveinal area) and become apparent on mature leaves during the post-véraison period. In both cases phloem disruption and downward rolling of leaf margins are present [3].

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