Evidence that Xylella fastidiosa is the causal agent of almond leaf scorch disease in Alicante, mainland Spain (Iberian Peninsula)

In 2017, Xylella fastidiosa, a quarantine plant pathogenic bacterium in Europe, was detected in almond trees associated to leaf scorch symptoms in Alicante, a Mediterranean area in southeastern mainland Spain. The bacterium was detected by serological and molecular techniques, isolated in axenic culture from diseased almond trees, and identified as X. fastidiosa subsp. multiplex ST6. Inoculation experiments on almond plants in greenhouse trials with a characterized strain of X. fastidiosa subsp. multiplex ST6 isolated in the outbreak area have proved that it was able to multiply and systemically colonize inoculated plants. Disease symptoms characteristic of leaf scorch, as those observed in the field, were observed in the inoculated almond trees after one year. Furthermore, the pathogen was re-isolated and identified by molecular tests. With the fulfillment of the Koch’s postulates we have demonstrated that X. fastidiosa is the causal agent of the almond leaf scorch disease in the Alicante outbreak.

Landscape Epidemiology of Xylella fastidiosa in the Balearic Islands

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.

Phylogenetic inference enables reconstruction of a long-overlooked outbreak of almond leaf scorch disease (Xylella fastidiosa) in Europe

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.

Draft Genome Resources of Two Strains (“ESVL” and “IVIA5901”) of Xylella fastidiosa Associated with Almond Leaf Scorch Disease in Alicante, Spain

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.

Rootstock Effects on Almond Leaf Scorch Disease Incidence and Severity

A 5-year field study was conducted to evaluate effects of duration and exclusion of Xylella fastidiosa infections on young almond tree performance and their links to tree vigor. ‘Nemaguard,’ ‘Okinawa,’ ‘Nonpareil,’ and Y119 were used as rootstocks for almond scion ‘Sonora.’ Among X. fastidiosa-infected trees, there was significant etiological heterogeneity with i) absence of leaf scorching symptoms in the presence of reduced growth, ii) presence of leaf scorching symptoms in the absence of reduced growth, and iii) severe leaf scorching and reduced growth. Trunk cross sectional areas of X. fastidiosa-infected trees grafted on ‘Nemaguard’ and ‘Nonpareil’ rootstocks were significantly smaller than noninfected trees, whereas trunk size of trees grafted on ‘Okinawa’ and Y119 was not affected by infection status. Severity of leaf scorching symptoms was highest on trees grafted on ‘Nonpareil’ rootstock, intermediate on ‘Okinawa’ and Y119, and lowest on ‘Nemaguard.’ X. fastidiosa infections and seasonal leaf scorching symptoms persisted on most inoculated trees throughout the study, except on trees grafted on ‘Nemaguard’ that manifested complete leaf scorching symptom remission and apparent elimination of the pathogen after the second year. Results indicate that depending on rootstock type X. fastidiosa can affect trunk size in a relatively short period and/or persist for years as trees grow.

Population Structure of Xylella fastidiosa Associated with Almond Leaf Scorch Disease in the San Joaquin Valley of California

Xylella fastidiosa causes disease in many commercial crops, including almond leaf scorch (ALS) disease in susceptible almond (Prunus dulcis). In this study, genetic diversity and population structure of X. fastidiosa associated with ALS disease were evaluated. Isolates obtained from two almond orchards in Fresno and Kern County in the San Joaquin Valley of California were analyzed for two successive years. Multilocus simple-sequence repeat (SSR) analysis revealed two major genetic clusters that were associated with two host cultivars, ‘Sonora’ and ‘Nonpareil’, respectively, regardless of the year of study or location of the orchard. These relationships suggest that host cultivar selection and adaptation are major driving forces shaping ALS X. fastidiosa population structure in the San Joaquin Valley. This finding will provide insight into understanding pathogen adaptation and host selection in the context of ALS disease dynamics.

Susceptibility to Xylella fastidiosa in a First-generation Hybrid from a non-traditional Peach–Almond Cross

To facilitate development of Prunus L. rootstocks with desirable agronomic traits, domesticated peach (Prunus persica) and almond (P. dulcis) were crossed with wild almond relatives. This work reports that a hybrid from a P. webbii × P. persica cv. Harrow Blood cross is susceptible to almond leaf scorch disease (ALSD). ALSD is caused by the fastidious, xylem-limited bacterium Xylella fastidiosa. The P. webbii × ‘Harrow Blood’ hybrid, along with its parents, was inoculated with two ALSD-inducing strains (X. fastidiosa subsp. fastidiosa strain M23 and subsp. multiplex strain Dixon). Both X. fastidiosa strains grew to high titer in the susceptible P. webbii parent and in the interspecific hybrid; defoliation was also observed. As expected, ‘Harrow Blood’ did not exhibit defoliation symptoms or support growth of X. fastidiosa. This result contrasts with earlier work demonstrating that a P. persica × P. dulcis hybrid is not a suitable host for X. fastidiosa subsp. fastidiosa M23. It appears that the genetic basis of resistance/susceptibility differs between a P. persica × P. dulcis cross and the P. webbii × P. persica cross reported here. Understanding the degree of susceptibility to X. fastidiosa in complex hybrids of subgenus Amygdalus should be an important part of rootstock development.

Almond Leaf Scorch Disease Development on Almond Branches High-Grafted on Peach Rootstock

Development of almond leaf scorch (ALS) disease was monitored on young almond (Prunus dulcis ‘NePlus’ and ‘Peerless’) shoots produced from almond scion wood that was high-grafted on peach rootstocks (P. persica ‘Queencrest/Nemaguard’), after the almond shoots were mechanically inoculated with Xylella fastidiosa. The objective of this study was to evaluate the potential movement of X. fastidiosa through almond–peach graft unions. ALS symptoms developed on both X. fastidiosa-inoculated and uninoculated almond shoots that were high-grafted on different peach limbs of the same tree in September following inoculations with X. fastidiosa made in June and July 2002, respectively, when the average distance in peach rootstock between the two almond–peach graft unions was 30.5 cm or shorter. No ALS symptoms were observed on uninoculated almond shoots late in the growing season of the second year. The incidence of ALS-affected leaves on shoots inoculated with X. fastidiosa decreased in the second year on the inoculated shoots of Peerless as determined by the number of inoculated shoots showing ALS symptoms in 2002 versus 2003. No visible ALS symptoms were observed in NePlus late in the growing season of the second year, suggesting that survival of X. fastidiosa in NePlus was less than in Peerless. These data demonstrate that movement of X. fastidiosa through two almond–peach graft unions was possible during the year of inoculation when the peach rootstock stem distance between the two almond–peach graft unions was minimal. However, X. fastidiosa may not survive the winter season or survived so poorly as not to be able to produce ALS symptoms in the second growing season on the uninoculated shoots that previously showed ALS symptoms in the fall of the year when an adjacent shoot was inoculated. The results suggest that high-grafting almond scion wood on multiple peach rootstock limbs may provide a means to limit movement of X. fastidiosa from one almond limb to another on the same tree.

Phenology of Xylella fastidiosa and Its Vector Around California Almond Nurseries: An Assessment of Plant Vulnerability to Almond Leaf Scorch Disease

Management of almond leaf scorch disease requires knowledge of all possible infection pathways. The disease is caused by the xylem-limited bacterium Xylella fastidiosa, which is transmitted by several species of sharpshooters. The objectives of this research were to elucidate the fate of bacteria in planta after inoculations in almond nursery plants and to determine patterns of insect vector population dynamics and temporal distribution of X. fastidiosa–infected plants relative to host plant assemblages in habitats surrounding commercial nurseries. In an experimental nursery, disease incidence was markedly affected by rootstock type. Prior to bud grafting, ‘Nemaguard’ rootstock seedlings were not susceptible to bacterial infection. After bud grafting with a susceptible scion (‘Sonora’), scions were susceptible to infection regardless of rootstock genotype. Surveys near commercial nurseries revealed that only habitats with permanent grass cover sustained vector populations throughout the season. A total of 87 plant samples tested positive for X. fastidiosa (6.3%) using enzyme-linked immunosorbent assay (ELISA), with a higher number of X. fastidiosa–infected plants found in weedy alfalfa fields than in other habitat types. Among plant species infected by X. fastidiosa, 33% were winter annuals, 45% were biennials or perennials, and 22% were summer annuals. Collectively, these findings identified a potential pathway for X. fastidiosa infection of almonds in nursery situations.