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Acidovorax citrulli is one of the most important pathogens of cucurbit crops, mainly melon and watermelon. Although A. citrulli is able to infect all aerial parts of the plant, fruits are highly sensitive to the bacterium. Therefore, the disease is known as bacterial fruit blotch (BFB). The unavailability of effective tools for managing BFB, including the lack of resistant varieties, exacerbates the threat this disease poses to the cucurbit industry. However, despite the economic importance of BFB, still little is known about basic aspects of A. citrulli-plant interactions. Here, we present diverse techniques that have recently been developed for investigation of basic aspects of BFB, including identification of virulence determinants of the pathogen., (© 2024. The Author(s), under exclusive license to Springer Science+Business Media, LLC, part of Springer Nature.)

Bacterial fruit blotch, caused by Acidovorax citrulli, is a serious disease of melon and watermelon. The strains of the pathogen belong to two major genetic groups: group I strains are strongly associated with melon, while group II strains are more aggressive on watermelon. A. citrulli secretes many protein effectors to the host cell via the type III secretion system. Here we characterized AopW1, an effector that shares similarity to the actin cytoskeleton-disrupting effector HopW1 of Pseudomonas syringae and with effectors from other plant-pathogenic bacterial species. AopW1 has a highly variable region (HVR) within amino acid positions 147 to 192, showing 14 amino acid differences between group I and II variants. We show that group I AopW1 is more toxic to yeast and Nicotiana benthamiana cells than group II AopW1, having stronger actin filament disruption activity, and increased ability to induce cell death and reduce callose deposition. We further demonstrated the importance of some amino acid positions within the HVR for AopW1 cytotoxicity. Cellular analyses revealed that AopW1 also localizes to the endoplasmic reticulum, chloroplasts, and plant endosomes. We also show that overexpression of the endosome-associated protein EHD1 attenuates AopW1-induced cell death and increases defense responses. Finally, we show that sequence variation in AopW1 plays a significant role in the adaptation of group I and II strains to their preferred hosts, melon and watermelon, respectively. This study provides new insights into the HopW1 family of bacterial effectors and provides first evidence on the involvement of EHD1 in response to biotic stress., (© 2023 The Authors. The Plant Journal published by Society for Experimental Biology and John Wiley & Sons Ltd.)

Acidovorax citrulli is the causal agent of bacterial fruit blotch (BFB) of cucurbit plants. In recent years, the disease has spread to many parts of the world, mainly via the inadvertent distribution of contaminated commercial seeds. Because of the costly lawsuits filed by growers against seed companies and the lack of efficient management methods, BFB represents a serious threat to the cucurbit industry, and primarily to watermelons and melons. Despite the economic importance of the disease, little is known about the basic aspects of A. citrulli pathogenesis. Nevertheless, the release of the genome of one A. citrulli strain, as well as the optimization of molecular manipulation and inoculation methods, has prompted basic studies and allowed advances towards an understanding of A. citrulli pathogenicity. In this article, we summarize the current knowledge about this important pathogen, with emphasis on its epidemiology and the factors involved in its pathogenicity and virulence., Taxonomy: Bacteria; Betaproteobacteria; order Burkholderiales; family C omamonadaceae; genus Acidovorax; species citrulli., Microbiological Properties: Gram-negative, strictly aerobic, rod-shaped; average dimensions of 0.5 μm × 1.7 μm; motile by means of an ~5.0-μm-long polar flagellum; colonies on King's medium B are round, smooth, transparent and nonpigmented; optimal temperatures for growth around 27-30 °C; induces a hypersensitive response on nonhost tobacco and tomato leaves., Host Range: Acidovorax citrulli strains are pathogenic to various species of the Cucurbitaceae family, including watermelon, melon, squash, pumpkin and cucumber. Significant economic losses have been reported in watermelon and melon., Disease Symptoms: Watermelon and melon seedlings and fruits are highly susceptible to A. citrulli. Typical seedling symptoms include water-soaked lesions on cotyledons that are often adjacent to the veins and later become necrotic, lesions on the hypocotyl, and seedling collapse and death. On watermelon fruits, symptoms begin as small, irregular, water-soaked lesions which later extend through the rind, turn brown and crack. On melon fruits, symptoms are characterized by small, often sunken rind lesions and internal fruit decay. Symptoms on the leaves of mature plants are difficult to diagnose because they are often inconspicuous or similar to those caused by other biotic or abiotic stresses. When they occur, leaf lesions can spread along the midrib and main veins. Lesions appear dark-brown to black on watermelon and light to reddish-brown on melon., Useful Websites: Bacterial fruit blotch of cucurbits at APSnet, http://www.apsnet.org/edcenter/intropp/lessons/prokaryotes/Pages/BacterialBlotch.aspx; bacterial fruit blotch guide from ASTA, http://www.amseed.com/pdfs/DiseaseGuide-BFB-English.pdf; Acidovorax citrulli AAC00-1 genome at JGI, http://genome.jgi-psf.org/aciav/aciav.info.html., (© 2012 THE AUTHORS. MOLECULAR PLANT PATHOLOGY © 2012 BSPP AND BLACKWELL PUBLISHING LTD.)

Bacterial fruit blotch caused by Acidovoraxcitrulli is a serious disease of cucurbit crops. Here we report characterization of a mutant strain of A. citrulli M6 defective in lip1, a gene encoding a lipolytic enzyme. The M6-lip1- mutant was detected in a mutant library screen aimed at identifying M6 mutants with altered levels of twitching motility. In this screen M6-lip1- was the only mutant that showed significantly larger twitching motility haloes around colonies than wild-type M6. Sequence analyses indicated that lip1 encodes a member of the GDSL family of secreted lipolytic enzymes. In line with this finding, lipolytic assays showed that the supernatants of M6-lip1- had lower lipolytic activity as compared with those of wild-type M6 and a lip1-complemented strain. The mutant was also affected in swimming motility and had compromised virulence on melon seedlings and on Nicotiana benthamiana leaves relative to wild-type and complemented strains. Lip1 contains a predicted N-terminal signal sequence for type II secretion. Evidence from our study confirms Lip1 is indeed secreted in a type II secretion-dependent manner, and this is required for full virulence of A. citrulli. To the best of our knowledge this is the first study reporting contribution of lipolytic activity to virulence of a plant-pathogenic Acidovorax species. [ABSTRACT FROM AUTHOR]

Acidovorax citrulli is the causal agent of bacterial fruit blotch disease of cucurbits. Strains of this pathogen are distributed into two major groups: Group I strains have been mainly isolated from melon and other non‐watermelon cucurbits, while Group II strains have been mainly recovered from watermelon. Here we report the characterization of strains T1 and EP isolated from diseased tomato and eggplant plants, respectively, and further confirmed to belong to A. citrulli species. Based on PCR, PFGE, and rep‐PCR, these strains showed high similarity to the Group II strain 7a1. Sequencing and comparative analyses revealed that the genomes of T1 and EP aligned with that of the Group II model strain AAC00‐1, over 97.88% and 99.22%, respectively. The virulence of T1, EP, and 7a1 determined on tomato, eggplant, and watermelon was similar and significantly higher than that of Group I strain M6. In contrast, M6 was more virulent on melon. Expression levels of seven virulence genes measured 24 hr after inoculation of tomato, eggplant, watermelon, and melon showed that the expression pattern was generally similar in strains 7a1, T1, and EP, whereas for M6 the expression was high only on melon. Overall, our results indicate that the solanaceous strains belong to Group II. To the best of our knowledge, this is the first study that reports characterization of A. citrulli strains isolated from solanaceous species. The fact that A. citrulli is able to naturally colonize and cause disease in non‐cucurbit crops poses additional challenges for management of this important pathogen. [ABSTRACT FROM AUTHOR]

Acidovorax citrulli is a seedborne bacterium that causes bacterial fruit blotch of cucurbit plants including watermelon and melon. A. citrulli strains can be divided into two major groups based on DNA fingerprint analyses and biochemical properties. Group I strains have been generally isolated from non-watermelon cucurbits, while group II strains are closely associated with watermelon. In the present study, we report the genome sequence of M6, a group I model A. citrulli strain, isolated from melon. We used comparative genome analysis to investigate differences between the genome of strain M6 and the genome of the group II model strain AAC00-1. The draft genome sequence of A. citrulli M6 harbors 139 contigs, with an overall approximate size of 4.85 Mb. The genome of M6 is ↑500 Kb shorter than that of strain AAC00-1. Comparative analysis revealed that this size difference is mainly explained by eight fragments, ranging from ↑35-120 Kb and distributed throughout the AAC00-1 genome, which are absent in the M6 genome. In agreement with this finding, while AAC00-1 was found to possess 532 open reading frames (ORFs) that are absent in strain M6, only 123 ORFs in M6 were absent in AAC00-1. Most of these M6 ORFs are hypothetical proteins and most of them were also detected in two group I strains that were recently sequenced, tw6 and pslb65. Further analyses by PCR assays and coverage analyses with other A. citrulli strains support the notion that some of these fragments or significant portions of them are discriminative between groups I and II strains of A. citrulli. Moreover, GC content, effective number of codon values and cluster of orthologs' analyses indicate that these fragments were introduced into group II strains by horizontal gene transfer events. Our study reports the genome sequence of a model group I strain of A. citrulli, one of the most important pathogens of cucurbits. It also provides the first comprehensive comparison at the genomic level between the two major groups of strains of this pathogen. [ABSTRACT FROM AUTHOR]

Acidovorax citrulli is the causal agent of bacterial fruit blotch of cucurbits. We have shown previously that type IV pili (TFP) are required for wild-type levels of virulence of A. citrulli on melon and that this pathogen can colonize and move thorough the xylem vessels of host seedlings. Here, comparative studies between wild-type and TFP mutant strains using microfluidic flow chambers demonstrated that TFP play a critical role in both the surface attachment and the biofilm formation of A. citrulli under a medium flow. Additionally, TFP null mutants were unable to perform twitching movement against the direction of medium flow. Assays using a flagellin mutant showed that, in contrast to TFP, polar flagella do not contribute to the adhesion and biofilm formation of A. citrulli under tested conditions. Also, flagellum-mediated swimming motility of wild-type strains was not observed under medium flow. These results imply that TFP may play an important role in colonization and spread in the xylem vessels under sap flow conditions, while polar flagella could be more important for spread during periods of time when xylem flow is minimal. [ABSTRACT FROM AUTHOR]

Bacterial fruit blotch (BFB) of cucurbits is caused by the Gram-negative bacterium Acidovorax avenae subsp. citrulli. The disease gained importance in the late 1980s, after devastating outbreaks in watermelon fields in several states in the US. Since then, BFB has spread worldwide, and has been reported in other cucurbits such as melon, pumpkin, squash, and cucumber. A. avenae subsp. citrulli is a seedborne pathogen of highly destructive potential. Under favorable conditions, the bacterium spreads rapidly throughout nurseries and in the field, leading to seedling blight or, at a later stage, fruit rot. Strategies for managing BFB are limited and there are no reliable sources of BFB resistance. The disease therefore represents a serious threat to the cucurbit industry. Despite its economic importance, there is little knowledge on basic aspects of the pathogen's biology or on the molecular basis of BFB pathogenesis. Recently, the genome sequence of one A. avenae subsp. citrulli strain was released, making a great contribution to the investigation of basic aspects of BFB. A better understanding of these aspects will contribute to improved screens for BFB resistance and to the development of more effective strategies to manage this threatening disease. [ABSTRACT FROM AUTHOR]

Bacterial fruit blotch (BFB), caused by Acidovorax avenae subsp. citrulli, is a serious disease of cucurbit plants. The first important occurrence of BFB in Israel was during 2000 to 2003 on watermelon and melon. Twelve bacterial isolates associated with these outbreaks were confirmed as A. avenae subsp, citrulli by pathogenicity assays, gas chromatography of fatty-acid methyl esters, and substrate-utilization profiles. The isolates were characterized in terms of their aggressiveness in different hosts by seed, seedling, and fruit inoculations, and according to their DNA fingerprinting profiles using pulse-field gel electrophoresis (PFGE) and repetitive-PCR approaches. Results from the present work agree with previous studies supporting the existence of two differentiated groups within A. avenae subsp, citrulli, one including strains that are more associated with watermelon (group II), the other consisting of strains that are usually associated with nonwatermelon cucurbits (group I). This study indicates that isolates from both groups have been introduced to Israel. PFGE analysis revealed that the 12 analyzed isolates can be divided into five different haplotypes, of which four were previously unreported. Additional differentiating features between group I and II strains are presented. [ABSTRACT FROM AUTHOR]

Strains of Acidovorax citrulli, the causal agent of bacterial fruit blotch (BFB) of cucurbits, can be assigned to two groups, I and II. The natural association of group I and II strains with different cucurbit species suggests host preference; however, there are no direct data to support this hypothesis under field conditions. Hence, the objective of this study was to assess differences in the prevalence of group I and II A. citrulli strains on cucurbit species in the field. From 2017 to 2019, we used group I and II strains to initiate BFB outbreaks in field plots planted with four cucurbit species. At different times, we collected symptomatic tissues and assayed them for group I and II strains using a group-specific PCR assay. Binary distribution data analysis revealed that the odds of melon, pumpkin, and squash foliage infection by group I strains were 21.7, 11.5, and 22.1 times greater, respectively, than the odds of watermelon foliage infection by the group I strain (P < 0.0001). More strikingly, the odds of melon fruit infection by the group I strain were 97.5 times greater than watermelon fruit infection by the same strain (P < 0.0001). Unexpectedly, some of the group II isolates recovered from the 2017 and 2019 studies were different from the group II strains used as inocula. Overall, data from these experiments confirm that A. citrulli strains exhibit a preference for watermelon and melon, which is more pronounced in fruit tissues. [ABSTRACT FROM AUTHOR]

Acidovorax citrulli , the causal agent of bacterial fruit blotch (BFB) disease, infects cucurbit crops including watermelon and melon. This bacterium can enter the viable but nonculturable (VBNC) state following exposure to copper sulfate. Moreover, copper-induced VBNC A. citrulli cells can be resuscitated by EDTA. In this study, isobaric tag for relative and absolute quantification (iTRAQ) was used to compare protein profiles of VBNC cells, resuscitated cells at different stages and log-phase cells of the A. citrulli model strain AAC00-1. A total of 2672 proteins were identified, with 60 being differentially abundant in VBNC cells compared with log-phase cells, and 469 being differentially abundant in resuscitated cells compared with VBNC cells. Proteins involved in the arginine and proline metabolism pathway and degradation of aromatic compounds could be important for the VBNC cells. In the early resuscitation process, proteins associated with carbon metabolism, and degradation of naphthalene and aromatic compounds were significantly enriched, while proteins involved in oxidative phosphorylation, bacterial chemotaxis, ABC transporters and quorum sensing were significantly enriched at the late resuscitation stages. This is the first study reporting thorough protein profile analyses of VBNC and resuscitating cells of a plant-pathogenic bacterium. The VBNC state is a dormant-like condition that was reported to occur in many bacterial species, upon facing a variety of environmental stresses. Acidovorax citrulli is a seed borne pathogenic bacterium that threatens cucurbit production worldwide. Moreover, A. citrulli can enter into the VBNC state after treatment of copper sulfate, thus increasing its survival and dissemination probabilities. This study enriches our understanding of the mechanisms of entrance into and resuscitation from the VBNC state of this important plant-pathogenic bacterium. This knowledge could be exploited in the future to develop novel approaches to interfere with these processes, thus contributing to a more efficient management of this pathogen. In a broader perspective, the knowledge emerging from this study has implications to the general understanding of the VBNC state in bacteria. Unlabelled Image • Differentially abundant proteins in VBNC state and resuscitation process of Acidovorax citrulli were identified using iTRAQ. • Most of the differentially abundant proteins were associated to general metabolism. • Energy production and oxidative stress response proteins were remarkably altered in their abundant level. • Some proteins involved in virulence were enriched during the resuscitation process. • This is the first study that handles mechanisms of VBNC state of a plant-pathogenic bacterium by a proteomic approach. [ABSTRACT FROM AUTHOR]