Xylella fastidiosa releases outer membrane vesicles (OMVs) known to play a role in the systemic dissemination of this pathogen. OMVs inhibit bacterial attachment to xylem wall and traffic lipases/esterases that act on the degradation of plant cell wall. Here, we extended the characterization of X. fastidiosa OMVs by identifying proteins and metabolites potentially associated with OMVs produced by Temecula1, a Pierce’s disease strain, and by 9a5c and Fb7, two citrus variegated chlorosis strains. These results strengthen that one of the OMVs multiple functions is to carry determinants of virulence, such as lipases/esterases, adhesins, proteases, porins, and a pectin lyase-like protein. For the first time, we show that the two citrus variegated chlorosis strains produce X. fastidiosa diffusible signaling factor 2 (DSF2) and citrus variegated chlorosis-DSF (likewise, Temecula1) and most importantly, that these compounds of the DSF (X. fastidiosa DSF) family are associated with OMV-enriched fractions. Altogether, our findings widen the potential functions of X. fastidiosa OMVs in intercellular signaling and host–pathogen interactions.

Plasmids are a major source of horizontal gene transfer among bacteria contributing to their evolution and ecology. The known collection of plasmids carried by the plant pathogen Xylella fastidiosa increases as new strains from different origins and hosts are sampled and sequenced. Here we performed an extensive comparison of 61 publicly available sequences annotated as plasmids belonging to 38 X. fastidiosa strains isolated from different plant species and distinct geographical locations. Twenty-six strains exhibited at least one plasmid and up to four plasmids were found in a single strain. Plasmids sizes varied widely from 1.3 kbp to 64.3 kbp, ranging from 1 to 70 protein-coding sequences (CDS) encompassing 324 orthologs. Based on the presence of specific mobility proteins such as relaxases and type 4 secretion system-related genes, respectively 40 and 8 of the X. fastidiosa plasmids were classified as conjugative and mobilizable, while 13 were classified as non-mobilizable. X. fastidiosa plasmids did not carry known antibiotic resistance or virulence genes, and their stability seems to take advantage of toxin/antitoxin systems. The comparative analyses described here revealed similarity among plasmids of X. fastidiosa from different subspecies, geographical regions, and hosts, as well as with sequences found in plasmids from other bacterial species. Altogether, our results provide an in silico analysis of X. fastidiosa plasmid content and their main features, with applications in future studies of epidemiology, ecology, and evolution of this phytopathogen.

Xylella fastidiosa causes diseases in many plant species. Originally confined to the Americas, infecting mainly grapevine, citrus, and coffee, X. fastidiosa has spread to several plant species in Europe causing devastating diseases. Many pathogenicity and virulence factors have been identified, which enable the various X. fastidiosa strains to successfully colonize the xylem tissue and cause disease in specific plant hosts, but the mechanisms by which this happens have not been fully elucidated. Here we present thorough comparative analyses of 94 whole-genome sequences of X. fastidiosa strains from diverse plant hosts and geographic regions. Core-genome phylogeny revealed clades with members sharing mostly a geographic region rather than a host plant of origin. Phylogenetic trees for 1605 orthologous CDSs were explored for potential candidates related to host specificity using a score of mapping metrics. However, no candidate host-specificity determinants were strongly supported using this approach. We also show that X. fastidiosa accessory genome is represented by an abundant and heterogeneous mobilome, including a diversity of prophage regions. Our findings provide a better understanding of the diversity of phylogenetically close genomes and expand the knowledge of X. fastidiosa mobile genetic elements and immunity systems.

À Profa. Dra. Aline Maria da Silva por aceitar me orientar e por todo o suporte e orientação que me deu durante todo o período de meu mestrado.

Xylella fastidiosa subsp. pauca, once confined to South America and infecting mainly citrus and coffee plants, has been found to be associated with other hosts and in other geographic regions. We present high-quality draft genome sequences of X. fastidiosa subsp. pauca strains J1a12, B111, U24D, and XRB isolated from citrus plants in Brazil, strain Fb7 isolated from a citrus plant in Argentina and strains 3124, Pr8x, and Hib4 isolated, respectively, from coffee, plum, and hibiscus plants in Brazil. Sequencing was performed using Roche 454-GS FLX, MiSeq-Illumina or Pacific Biosciences platforms. These high-quality genome assemblies will be useful for further studies about the genomic diversity, evolution, and biology of X. fastidiosa.