Following photosynthesis, sucrose is translocated to sink organs, where it provides the primary source of carbon and energy to sustain plant growth and development. Sugar transporters from the SWEET (sugar will eventually be exported transporter) family are rate-limiting factors that mediate sucrose transport across concentration gradients, sustain yields, and participate in reproductive development, plant senescence, stress responses, as well as support plant–pathogen interaction, the focus of this study. We identified 25 SWEET genes in the walnut genome and distinguished each by its individual gene structure and pattern of expression in different walnut tissues. Their chromosomal locations, cis-acting motifs within their 5′ regulatory elements, and phylogenetic relationship patterns provided the first comprehensive analysis of the SWEET gene family of sugar transporters in walnut. This family is divided into four clades, the analysis of which suggests duplication and expansion of the SWEET gene family in Juglans regia. In addition, tissue-specific gene expression signatures suggest diverse possible functions for JrSWEET genes. Although these are commonly used by pathogens to harness sugar products from their plant hosts, little was known about their role during Xanthomonas arboricola pv. juglandis (Xaj) infection. We monitored the expression profiles of the JrSWEET genes in different tissues of “Chandler” walnuts when challenged with pathogen Xaj417 and concluded that SWEET-mediated sugar translocation from the host is not a trigger for walnut blight disease development. This may be directly related to the absence of type III secretion system-dependent transcription activator-like effectors (TALEs) in Xaj417, which suggests different strategies are employed by this pathogen to promote susceptibility to this major aboveground disease of walnuts.

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.

Xylella fastidiosa colonizes the xylem of various cultivated and native plants worldwide. Citrus production in Brazil has been seriously affected, and major commercial varieties remain susceptible to Citrus Variegated Chlorosis (CVC). Collective cellular behaviors such as biofilm formation influence virulence and insect transmission of X. fastidiosa. The reference strain 9a5c produces a robust biofilm compared to Fb7 that remains mostly planktonic, and both were isolated from symptomatic citrus trees. This work deepens our understanding of these distinct behaviors at the molecular level, by comparing the cellular and secreted proteomes of these two CVC strains. Out of 1017 identified proteins, 128 showed differential abundance between the two strains. Different protein families were represented such as proteases, hemolysin-like proteins, and lipase/esterases, among others. Here we show that the lipase/esterase LesA is among the most abundant secreted proteins of CVC strains as well, and demonstrate its functionality by complementary activity assays. More severe symptoms were observed in Nicotiana tabacum inoculated with strain Fb7 compared to 9a5c. Our results support that systemic symptom development can be accelerated by strains that invest less in biofilm formation and more in plant colonization. This has potential application in modulating the bacterial-plant interaction and reducing disease severity.

The interaction between the plant host, walnut (Juglans regia; Jr), and a deadly pathogen (Xanthomonas arboricola pv. juglandis 417; Xaj) can lead to walnut bacterial blight (WB), which depletes walnut productivity by degrading the nut quality. Here, we dissect this pathosystem using tandem mass tag quantitative proteomics. Walnut hull tissues inoculated with Xaj were compared to mock-inoculated tissues, and 3972 proteins were identified, of which 3296 are from Jr and 676 from Xaj. Proteins with differential abundance include oxidoreductases, proteases, and enzymes involved in energy metabolism and amino acid interconversion pathways. Defense responses and plant hormone biosynthesis were also increased. Xaj proteins detected in infected tissues demonstrate its ability to adapt to the host microenvironment, limiting iron availability, coping with copper toxicity, and maintaining energy and intermediary metabolism. Secreted proteases and extracellular secretion apparatus such as type IV pilus for twitching motility and type III secretion effectors indicate putative factors recognized by the host. Taken together, these results suggest intense degradation processes, oxidative stress, and general arrest of the biosynthetic metabolism in infected nuts. Our results provide insights into molecular mechanisms and highlight potential molecular tools for early detection and disease control strategies.

Walnuts (Juglans regia L.) are a valuable dietary source of polyphenols and lipids, with increasing worldwide consumption. California is a major producer, with ’Chandler’ and ’Tulare’ among the cultivars more widely grown. ’Chandler’ produces kernels with extra light color at a higher frequency than other cultivars, gaining preference by growers and consumers. Here we performed a deep comparative proteome analysis of kernel pellicle tissue from these two valued genotypes at three harvest maturities, detecting a total of 4937 J. regia proteins. Late and early maturity stages were compared for each cultivar, revealing many developmental responses common or specific for each cultivar. Top protein biomarkers for each developmental stage were also selected based on larger fold-change differences and lower variance among replicates, including proteins for biosynthesis of lipids and phenols, defense-related proteins and desiccation stress-related proteins. Comparison between the genotypes also revealed the common and specific protein repertoires, totaling 321 pellicle proteins with differential abundance at harvest stage. The proteomics data provides clues on antioxidant, secondary, and hormonal metabolism that could be involved in the loss of quality in the pellicles during processing for commercialization.

Xylella fastidiosa is the causal agent of several plant diseases affecting fruit and nut crops. Methylobacterium mesophilicum strain SR1.6/6 was isolated from Citrus sinensis and shown to promote plant growth by producing phytohormones, providing nutrients, inhibiting X. fastidiosa, and preventing Citrus Variegated Chlorosis. However, the molecular mechanisms involved in the interaction among these microbes are still unclear. The present work aimed to analyze physiological and molecular aspects of M. mesophilicum SR1.6/6 and X. fastidiosa 9a5c in co-culture. The transcriptome and secretome analyses indicated that X. fastidiosa down-regulates cell division and transport genes and up-regulates stress via induction of chaperones and pathogenicity-related genes including, the lipase-esterase LesA, a protease, as well as an oligopeptidase in response to M. mesophilicum competition. On the other hand, M. mesophilicum also down-regulated transport genes, except for iron uptake, which was up-regulated. Secretome analysis identified four proteins in M. mesophilicum exclusively produced in co-culture with X. fastidiosa, among these, three are related to phosphorous uptake. These results suggest that M. mesophilicum inhibits X. fastidiosa growth mainly due to nutrient competition for iron and phosphorous, thus promoting X. fastidiosa starvation, besides producing enzymes that degrade X. fastidiosa cell wall, mainly hydrolases. The understanding of these interactions provides a direction for control and management of the phytopathogen X. fastidiosa, and consequently, helps to improve citrus growth and productivity.