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Xylella fastidiosa , the causal agent of Pierce's disease of grapevine, has been found in all major grape-growing regions in California, U.S.A. Large collections of X. fastidiosa isolates are available from these areas, which enable comparative studies of pathogen genetic traits and virulence. Owing to the significant resource requirements for experiments with X. fastidiosa in grapevine, however, most studies use only a single isolate to evaluate disease, and it is not clear how much variability between isolates impacts disease development in experimental or natural settings. In this study, a comprehensive panel of X. fastidiosa isolates from all California grape-growing regions was tested for virulence in susceptible grapevine and in the model host plant, tobacco. Seventy-one isolates were tested, 29 in both grapevine and tobacco. The results of this study highlight the inherent variability of inoculation experiments with X. fastidiosa , including variation in disease severity in plants inoculated with a single isolate, and variability between experimental replicates. There were limited differences in virulence between isolates that were consistent across experimental replicates, or across different host plants. This suggests that choice of isolate within the X. fastidiosa subsp. fastidiosa Pierce's disease group may not make any practical difference when testing in susceptible grape varieties, and that pathogen evolution has not significantly changed virulence of Pierce's disease isolates within California. The location of isolation also did not dictate relative disease severity. This information will inform experimental design for future studies of X. fastidiosa in grapevine and provide important context for genomic research., Competing Interests: The author(s) declare no conflict of interest.

Management of widespread plant pathogens is challenging as climatic differences among crop-growing regions may alter key aspects of pathogen spread and disease severity. Xylella fastidiosa is a xylem-limited bacterial pathogen that is transmitted by xylem sap-feeding insects. Geographic distribution of X. fastidiosa is limited by winter climate, and vines infected with X. fastidiosa can recover from infection when held at cold temperatures. California has a long history of research on Pierce's disease and significant geographic and climatic diversity among grape-growing regions. This background in combination with experimental disease studies under controlled temperature conditions can inform risk assessment for X. fastidiosa spread and epidemic severity across different regions and under changing climate conditions. California's grape-growing regions have considerable differences in summer and winter climate. In northern and coastal regions, summers are mild and winters are cool, conditions which favor winter recovery of infected vines. In contrast, in inland and southern areas, summers are hot and winters mild, reducing likelihood of winter recovery. Here, winter recovery of three table grape cultivars (Flame, Scarlet Royal, and Thompson Seedless) and three wine grape cultivars (Sauvignon Blanc, Cabernet Sauvignon, and Zinfandel) were evaluated under temperature conditions representative of the San Joaquin Valley, an area with hot summers and mild winters that has been severely impacted by Pierce's disease and contains a large portion of California grape production. Mechanically inoculated vines were held in the greenhouse under one of three warming treatments to represent different seasonal inoculation dates prior to being moved into a cold chamber. Winter recovery under all treatments was generally limited but with some cultivar variation. Given hot summer temperatures of many grape-growing regions worldwide, as well as increasing global temperatures overall, winter recovery of grapevines should not be considered a key factor limiting X. fastidiosa spread and epidemic severity in the majority of cases., Competing Interests: The author(s) declare no conflict of interest.

Xylella fastidiosa is a xylem-limited bacterial pathogen that causes Pierce's disease (PD) of grapevine. In host plants, this bacterium exclusively colonizes the xylem, which is primarily non-living at maturity. Understanding how X. fastidiosa interfaces with this specialized conductive tissue is at the forefront of investigation for this pathosystem. Unlike many bacterial plant pathogens, X. fastidiosa lacks a type III secretion system and cognate effectors that aid in host colonization. Instead, X. fastidiosa utilizes plant cell-wall hydrolytic enzymes and lipases as part of its xylem colonization strategy. Several of these virulence factors are predicted to be secreted via the type II secretion system (T2SS), the main terminal branch of the Sec-dependent general secretory pathway. In this study, we constructed null mutants in xpsE and xpsG , which encode for the ATPase that drives the T2SS and the major structural pseudopilin of the T2SS, respectively. Both mutants were non-pathogenic and unable to effectively colonize Vitis vinifera grapevines, demonstrating that the T2SS is required for X. fastidiosa infection processes. Furthermore, we utilized mass spectrometry to identify type II-dependent proteins in the X. fastidiosa secretome. In vitro, we identified six type II-dependent proteins in the secretome that included three lipases, a β-1,4-cellobiohydrolase, a protease, and a conserved hypothetical protein. [Formula: see text] Copyright © 2023 The Author(s). This is an open access article distributed under the CC BY-NC-ND 4.0 International license., Competing Interests: The author(s) declare no conflict of interest.