Natural competence and intersubspecific recombination in Xylella fastidiosa; a causal agent of emerging plant diseases

Abstract

Xylella fastidiosa is a Gram-negative, vector-transmitted, plant-pathogenic bacterium causing incurable diseases to economically important crops such as grapevine, citrus, almond, and coffee. Recent epidemiological studies show that X. fastidiosa is expanding its plant-host range and is spreading to new geographical locations. Moreover, strains isolated from these new infections were genetically different and showed presence of homologous recombination (HR) in some cases. HR between subspecies of X. fastidiosa has been described as a major evolutionary force in creating genetic diversity and causing plant-host shift. As X. fastidiosa is naturally competent, this could be an explanation for intersubspecific recombination and disease emergence. However, limited information exists regarding natural competence of X. fastidiosa in natural habitats and the factors that influence it, the degree to which strains are naturally competent, whether intersubspecific recombination can be experimentally demonstrated, and if intersubspecific recombination can be detected in the genomes of X. fastidiosa. Therefore, experiments were performed by co-culturing combinations of recipient strains and donor DNA (plasmids, marker tagged live- and heat-killed cells) under various growth conditions; by selecting the recombinants based on the acquisition of marker, and testing their phenotypes in vitro and in planta; and by sequencing and comparing whole genomes of the parent and recombinant strains. Results demonstrated that natural competence readily occurs under in vitro conditions that mimic the natural habitats of the bacterium. Moreover, widespread natural competence ability was detected among strains isolated from diverse host plants, and intersubspecific recombination was validated experimentally. The rate of recombination was iii significantly correlated with growth rate and twitching motility. Whole genome sequence analysis of in vitro generated recombinants and virulent strains isolated from infected fields demonstrated the presence of intersubspecific recombination in up to 10kb genomic regions. Results of this study are useful for policy makers in formulating strict quarantine measures to limit the introduction of new X. fastidiosa genotypes and prevent both intra- and intersubspecific recombination between the introduced and native strains, which will minimize the risk of new disease emergence and reduce economic losses due to the incurable diseases caused by X. fastidiosa.