Development of Molecular Markers for Detection of Acidovorax citrulli Strains Causing Bacterial Fruit Blotch Disease in Melon.

Md Rafiqul Islam,Md Abuyusuf,Jong-In Park,Denison Michael Immanuel Jesse,Hee-Jeong Jung,Hoy-Taek Kim,Ill-Sup Nou,Mohammad Rashed Hossain

doi:10.3390/ijms20112715

Md Rafiqul Islam, Md Abuyusuf + Show 6 more

Open Access

https://doi.org/10.3390/ijms20112715

Copy DOI

Abstract

Acidovorax citrulli (A. citrulli) strains cause bacterial fruit blotch (BFB) in cucurbit crops and affect melon significantly. Numerous strains of the bacterium have been isolated from melon hosts globally. Strains that are aggressively virulent towards melon and diagnostic markers for detecting such strains are yet to be identified. Using a cross-inoculation assay, we demonstrated that two Korean strains of A. citrulli, NIHHS15-280 and KACC18782, are highly virulent towards melon but avirulent/mildly virulent to the other cucurbit crops. The whole genomes of three A. citrulli strains isolated from melon and three from watermelon were aligned, allowing the design of three primer sets (AcM13, AcM380, and AcM797) that are specific to melon host strains, from three pathogenesis-related genes. These primers successfully detected the target strain NIHHS15-280 in polymerase chain reaction (PCR) assays from a very low concentration of bacterial gDNA. They were also effective in detecting the target strains from artificially infected leaf, fruit, and seed washing suspensions, without requiring the extraction of bacterial DNA. This is the first report of PCR-based markers that offer reliable, sensitive, and rapid detection of strains of A. citrulli causing BFB in melon. These markers may also be useful in early disease detection in the field samples, in seed health tests, and for international quarantine purposes.

Highlights

Acidovorax citrulli, formerly Acidovorax avenae subspecies citrulli (A. citrulli), is an aerobic, mesophilic, gram-negative, and seed-borne bacterium belonging to the beta subdivision of the Proteobacteria [1]
Whole-genome sequences of three A. citrulli strains M6, Pslb65, and DSM 17060 that were isolated from melon hosts and three A. citrulli strains AAC00-1, KACC17005, and tw6 that were isolated from watermelon hosts were retrieved from the National Center for Biotechnology Information database
This study exploited the variation within the whole-genome sequences of A. citrulli strains to develop polymerase chain reaction (PCR)-based markers for detecting A. citrulli strains that are associated with melon hosts

Summary

Introduction

Acidovorax citrulli, formerly Acidovorax avenae subspecies citrulli (A. citrulli), is an aerobic, mesophilic, gram-negative, and seed-borne bacterium belonging to the beta subdivision of the Proteobacteria [1]. This bacterial pathogen causes bacterial fruit blotch (BFB) disease [1,2] and represents a serious threat to cucurbit crops, mainly melon (Cucumis melo L., Cucurbitaceae) and watermelon (Citrullus lanatus L., Cucurbitaceae), worldwide, including South Korea [3,4,5,6]. In South Korea, BFB in melon was first reported in 1990, and several outbreaks have been observed in recent years [21]

Objectives

Methods

Conclusion