Simultaneous Detection of Both RNA and DNA Viruses Infecting Dry Bean and Occurrence of Mixed Infections by BGYMV, BCMV and BCMNV in the Central-West Region of Mexico.

Elizabeth Chiquito-Almanza,Nadia C García-Álvarez,Victor Montero-Tavera,Eduardo R Garrido-Ramírez,José L Anaya-López,Lorenzo Guevara-Olvera,Jorge A Acosta-Gallegos

doi:10.3390/v9040063

Abstract

A multiplex reverse transcription polymerase chain reaction (RT-PCR) assay was developed to simultaneously detect bean common mosaic virus (BCMV), bean common mosaic necrotic virus (BCMNV), and bean golden yellow mosaic virus (BGYMV) from common bean leaves dried with silica gel using a single total nucleic acid extraction cetyl trimethyl ammonium bromide (CTAB) method. A mixture of five specific primers was used to amplify three distinct fragments corresponding to 272 bp from the AC1 gene of BGYMV as well as 469 bp and 746 bp from the CP gene of BCMV and BCMNV, respectively. The three viruses were detected in a single plant or in a bulk of five plants. The multiplex RT-PCR was successfully applied to detect these three viruses from 187 field samples collected from 23 municipalities from the states of Guanajuato, Nayarit and Jalisco, Mexico. Rates of single infections were 14/187 (7.5%), 41/187 (21.9%), and 35/187 (18.7%), for BGYMV, BCMV, and BCMNV, respectively; 29/187 (15.5%) samples were co-infected with two of these viruses and 10/187 (5.3%) with the three viruses. This multiplex RT-PCR assay is a simple, rapid, sensitive, and cost-effective method for detecting these viruses in the common bean and can be used for routine molecular diagnosis and epidemiological studies.

Highlights

Virus diseases are among the major biotic constraints to legume production, in the tropics and subtropics [1,2]
We describe the development and evaluation of a multiplex reverse transcription polymerase chain reaction (RT-PCR) assay for the simultaneous detection and differentiation of bean common mosaic virus (BCMV), bean common mosaic necrotic virus (BCMNV) and bean golden yellow mosaic virus (BGYMV) in single and mixed infections of the common bean
Specificity of the Primers Designed for BGYMV Detection

Summary

Introduction

Virus diseases are among the major biotic constraints to legume production, in the tropics and subtropics [1,2]. In 2013, over 29 million hectares of tropical and temperate agricultural land in America, Europe, Africa and Asia were used for common bean production [4]. Among the viruses infecting beans, the potyviruses bean common mosaic virus (BCMV) and bean common mosaic necrosis virus (BCMNV) are most worldwide spread, and together with the geminiviruses bean golden mosaic virus. Viruses 2017, 9, 63; doi:10.3390/v9040063 www.mdpi.com/journal/viruses (BGMV) and bean golden yellow mosaic virus (BGYMV) are considered one of the major production constraints of common bean in Latin America, the Caribbean, and East, West and Southern Africa [5,6]. BCMV can remain viable in bean seeds for more than 30 years [9], and under field conditions the infected seedlings originated from

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