The recent introductions of the bacterium Xylella fastidiosa (Xf) into Europe are linked to the international plant trade. However, both how and when these entries occurred remains poorly understood. Here, we show how almond scorch leaf disease, which affects ~79% of almond trees in Majorca (Spain) and was previously attributed to fungal pathogens, was in fact triggered by the introduction of Xf around 1993 and subsequently spread to grapevines (Pierceʼs disease). We reconstructed the progression of almond leaf scorch disease by using broad phylogenetic evidence supported by epidemiological data. Bayesian phylogenetic inference predicted that both Xf subspecies found in Majorca, fastidiosa ST1 (95% highest posterior density, HPD: 1990–1997) and multiplex ST81 (95% HPD: 1991–1998), shared their most recent common ancestors with Californian Xf populations associated with almonds and grapevines. Consistent with this chronology, Xf-DNA infections were identified in tree rings dating to 1998. Our findings uncover a previously unknown scenario in Europe and reveal how Pierce’s disease reached the continent.

Pathogen introductions have led to numerous disease outbreaks in naive regions of the globe. The plant pathogen Xylella fastidiosa has been associated with various recent epidemics in Europe affecting agricultural crops, such as almond, grapevine, and olive, but also endemic species occurring in natural forest landscapes and ornamental plants. We compared whole-genome sequences of X. fastidiosa subspecies multiplex from America and strains associated with recent outbreaks in southern Europe to infer their likely origins and paths of introduction within and between the two continents. Phylogenetic analyses indicated multiple introductions of X. fastidiosa subspecies multiplex into Italy, Spain, and France, most of which emerged from a clade with limited genetic diversity with a likely origin in California, USA. The limited genetic diversity observed in X. fastidiosa subspecies multiplex strains originating from California is likely due to the clade itself being an introduction from X. fastidiosa subspecies multiplex populations in the southeastern United States, where this subspecies is most likely endemic. Despite the genetic diversity found in some areas in Europe, there was no clear evidence of recombination occurring among introduced X. fastidiosa strains in Europe. Sequence type taxonomy, based on multilocus sequence typing (MLST), was shown, at least in one case, to not lead to monophyletic clades of this pathogen; whole-genome sequence data were more informative in resolving the history of introductions than MLST data. Although additional data are necessary to carefully tease out the paths of these recent dispersal events, our results indicate that whole-genome sequence data should be considered when developing management strategies for X. fastidiosa outbreaks. IMPORTANCE Xylella fastidiosa is an economically important plant-pathogenic bacterium that has emerged as a pathogen of global importance associated with a devastating epidemic in olive trees in Italy associated with X. fastidiosa subspecies pauca and other outbreaks in Europe, such as X. fastidiosa subspecies fastidiosa and X. fastidiosa subspecies multiplex in Spain and X. fastidiosa subspecies multiplex in France. We present evidence of multiple introductions of X. fastidiosa subspecies multiplex, likely from the United States, into Spain, Italy, and France. These introductions illustrate the risks associated with the commercial trade of plant material at global scales and the need to develop effective policy to limit the likelihood of pathogen pollution into naive regions. Our study demonstrates the need to utilize whole-genome sequence data to study X. fastidiosa introductions at outbreak stages, since a limited number of genetic markers does not provide sufficient phylogenetic resolution to determine dispersal paths or relationships among strains that are of biological and quarantine relevance.

Xylella fastidiosa (Xf) is a vascular plant pathogen native to the Americas. In 2013, it was first reported in Europe, implicated in a massive die-off of olive trees in Apulia, Italy. This finding prompted mandatory surveys across Europe, successively revealing that the bacterium was already established in some distant areas of the western Mediterranean. To date, the Balearic Islands (Spain) hold the major known genetic diversity of Xf in Europe. Since October 2016, four sequence types (ST) belonging to the subspecies fastidiosa (ST1), multiplex (ST7, ST81), and pauca (ST80) have been identified infecting 28 host species, including grapevines, almond, olive, and fig trees. ST1 causes Pierce’s disease (PD) and together with ST81 are responsible for almond leaf scorch disease (ALSD) in California, from where they were introduced into Mallorca in around 1993, very likely via infected almond scions brought for grafting. To date, almond leaf scorch disease affects over 81% of almond trees and Pierce’s disease is widespread in vineyards across Mallorca, although producing on average little economic impact. In this perspective, we present and analyze a large Xf-hosts database accumulated over four years of field surveys, laboratory sample analyses, and research to understand the underlying causes of Xf emergence and spread among crops and wild plants in the Balearic Islands. The impact of Xf on the landscape is discussed.

Plant pathogens pose increasing threats to global food security, causing yield losses that exceed 30% in food-deficit regions. Xylella fastidiosa (Xf) represents the major transboundary plant pest and one of the world’s most damaging pathogens in terms of socioeconomic impact. Spectral screening methods are critical to detect non-visual symptoms of early infection and prevent spread. However, the subtle pathogen-induced physiological alterations that are spectrally detectable are entangled with the dynamics of abiotic stresses. Here, using airborne spectroscopy and thermal scanning of areas covering more than one million trees of different species, infections and water stress levels, we reveal the existence of divergent pathogen- and host-specific spectral pathways that can disentangle biotic-induced symptoms. We demonstrate that uncoupling this biotic–abiotic spectral dynamics diminishes the uncertainty in the Xf detection to below 6% across different hosts. Assessing these deviating pathways against another harmful vascular pathogen that produces analogous symptoms, Verticillium dahliae, the divergent routes remained pathogen- and host-specific, revealing detection accuracies exceeding 92% across pathosystems. These urgently needed hyperspectral methods advance early detection of devastating pathogens to reduce the billions in crop losses worldwide.

An outbreak of Xylella fastidiosa subsp. multiplex sequence type ST6 was discovered in 2017 in mainland Spain affecting almond trees. Two cultured almond strains, “ESVL” and “IVIA5901,” were subjected to high throughput sequencing and the draft genomes assembled. Phylogenetic analysis conclusively indicated they belong to the subspecies multiplex, and pairwise comparisons of the chromosomal genomes showed an average nucleotide identity higher than 99%. Interestingly, the two strains differ for the presence of the plasmids pXF64-Hb_ESVL and pUCLA-ESVL detected only in the ESVL strain. The availability of these draft genomes contribute to extend the European genomic sequence dataset, a first step toward setting new research to elucidate the pathway of introduction and spread of the numerous strains of this subspecies so far detected in Europe.

We report the complete annotated genome sequence of the plant-pathogenic bacterium Xylella fastidiosa subsp. fastidiosa strain IVIA5235.

Identification of sequence types (ST) of Xylella fastidiosa based on direct MultiLocus Sequence Typing (MLST) of plant DNA samples is partly efficient. In order to improve the sensitivity of X. fastidiosa identification, we developed a direct nested-MLST assay on plant extracted DNA. This method was performed based on a largely used scheme targeting seven housekeeping gene (HKG) loci (cysG, gltT, holC, leuA, malF, nuoL, petC). Samples analyzed included 49 plant species and two insect species (Philaenus spumarius, Neophilaenus campestris) that were collected in 2017 (106 plant samples in France), in 2018 (162 plant samples in France, 40 plant samples and 26 insect samples in Spain), and in 2019 (30 plant samples in Spain). With the nested approach, a significant higher number of samples were amplified. The threshold was improved by 100 to 1000 times compared to conventional PCR. Using nested-MLST assay, plants that were not yet considered hosts tested positive and revealed novel alleles in France, whereas for Spanish samples it was possible to assign the subspecies or ST to samples considered as new hosts in Europe. Direct typing by nested-MLST from plant material has an increased sensitivity and may be useful for epidemiological purposes.

Xylella fastidiosa is an economically important plant pathogenic bacterium of global importance associated, since 2013, with a devastating epidemic in olive trees in Italy. Since then, several outbreaks of this pathogen have been reported in other European member countries including Spain, France, and Portugal. In Spain, the three major subspecies (subsp. fastidiosa, multiplex, and pauca) of the bacterium have been detected in the Balearic Islands, but only subspecies multiplex in the mainland (Alicante). We present the first complete genome sequences of two Spanish strains: X. fastidiosa subsp. fastidiosa IVIA5235 from Mallorca and X. fastidiosa subsp. multiplex IVIA5901 from Alicante, using Oxford Nanopore and Illumina sequence reads, and two hybrid approaches for genome assembly. These completed genomes will provide a resource to better understand the biology of these X. fastidiosa strains.