Core-Genome Multilocus Sequence Typing for Epidemiological and Evolutionary Analyses of Phytopathogenic Xanthomonas citri.

Xanthomonas citri subsp. citri is the cause of bacterial citrus canker, responsible for major economic losses to the citrus industry. X. citri subspecies and pathovars are responsible for diseases in soybean, common bean, mango, pomegranate, and cashew. X. citri disease has been tracked using several typing methods, but recent studies using genomic sequencing have been key to understanding the evolutionary relationships within the species, including fundamental differences among X. citri subsp. citri pathotypes. Here, we describe a core-genome multilocus sequence typing (cgMLST) scheme for X. citri based on 250 genomes comprising multiple examples of X. citri subsp. citri pathotypes A, A*, and Aw; X. citri subsp. malvacearum; X. citri pv. aurantifolii, pv. fuscans, pv. glycines, pv. mangiferaeindicae, pv. viticola, and pv. vignicola; and single isolates of X. citri pv. dieffenbachiae and pv. punicae. This data set included genomic sequencing of 100 novel X. citri subsp. citri isolates. cgMLST, based on 1,618 core genes across 250 genomes, is implemented at PubMLST (https://pubmlst.org/organisms/xanthomonas-citri/). GrapeTree minimum-spanning tree and Interactive Tree of Life (iTOL) neighbor-joining phylogenies generated from the cgMLST data resolved almost identical groupings of isolates to a core-genome single nucleotide polymorphism (SNP)-based neighbor-joining phylogeny. These resolved identical groupings of X. citri subsp. citri pathotypes and X. citri subspecies and pathovars. X. citri cgMLST should prove to be an increasingly valuable resource for the study of this key species of plant-pathogenic bacteria. Users can submit genomic data and associated metadata for comparison with previously characterized isolates at PubMLST to allow the rapid characterization of the local, national, and global epidemiology of these pathogens and examine evolutionary relationships. IMPORTANCE Xanthomonas citri is a plant pathogen that causes major economic losses to the citrus industry and sweet orange production in particular. Several subspecies and pathogens are recognized, with host ranges including soybean, common bean, mango, pomegranate, and cashew, among others. Recent genomic studies have shown that host-adapted X. citri subspecies and pathovars and X. citri subsp. citri pathotypes form distinct clades. In this study, we describe a core-genome multilocus sequence typing (cgMLST) scheme for this species that can rapidly and robustly discriminate among these ecologically distinct, host-adapted clades. We have established this scheme and associated databases containing genomic sequences and metadata at PubMLST, which users can interrogate with their own genome sequences to determine X. citri subspecies, pathovars, and pathotypes. X. citri cgMLST should prove to be an invaluable tool for the study of the epidemiology and evolution of this major plant pathogen.

Hexanoic acid: a new potential substitute for copper-based agrochemicals against citrus canker.

AIMS: The aim of the study is to evaluate hexanoic acid (HA) as an alternative to manage citrus canker. METHODS AND RESULTS: The minimal growth inhibitory concentration of HA against Xanthomonas citri subsp. citri was determined at 2·15 mmol l-1 using a respiratory activity assay. Growth curves at different pH values showed that growth inhibition was not due to media acidification induced by HA. The germination rate and root elongation of Lactuca sativa seeds exposed to different concentrations of HA (varying from 0·86 to 5·16 mmol l-1 ) were assessed to screen for phytotoxicity. The acid exhibited low phytotoxicity for L. sativa at 1·29 and 2·58 mmol l-1 . To evaluate the ability of HA to protect citrus against X. citri infection, leaves of Citrus sinensis were sprayed with the acid and subsequently challenged with X. citri. HA at 3·44 mmol l-1 was able to protect citrus against infection, showing a reduction of three orders of magnitude in the number of citrus canker lesions per cm2 when compared to the untreated negative control. CONCLUSION: HA is a potential alternative to copper for citrus canker management. SIGNIFICANCE AND IMPACT OF THE STUDY: HA inhibits X. citri growth, exhibits low phytotoxicity and is an alternative to copper for the protection of citrus plants against bacterial infection.