Search Results (41)

Citrus canker, caused by Xanthomonas citri subsp. citri, is a devastating disease that affects citrus production and trade worldwide. Traditional control methods, based on copper compounds, are effective but pose environmental and health risks due to their toxicity and potential for bioaccumulation. This study evaluates the synergistic potential of essential oils (EOs) and rhamnolipids as sustainable alternatives for disease management. Four EOS (citronella, palmarosa, geranium, and clove) were tested for their antibacterial activity. Citronella EO showed a 90% inhibitory concentration (IC 90) of 0.15% (v/v) and a minimum bactericidal concentration of 0.25% (v/v), while the other EOs showed IC 90 and bactericidal activity at 0.06% (v/v). Rhamnolipids (RHLs), biosurfactants produced by Pseudomonas aeruginosa, inhibited X. citri at a concentration of 0.3% (v/v). The combination of citronella EO and RHLs showed a synergistic effect, reducing the inhibitory concentration of citronella by 50% and that of RHLs by more than 90%. In addition, the combined formulation permeabilized more than 80% of bacterial membranes and reduced biofilm formation. In contrast, other oils tested in combination with rhamnolipid showed independent effects. These results indicate that EOs and rhamnolipids represent an environmentally safe strategy for the control of X. citri subsp. citri that overcomes the limitations of conventional methods while reducing environmental and health impacts. Full article

Citrus canker, caused by Xanthomonas citri subsp. citri, and bacterial blight, caused by Xanthomonas citri subsp. malvacearum, results in substantial economic losses worldwide, and searching for new antibacterial agents is a critical challenge. In this study, regional isolates AE28 and RQ3 were obtained from characteristic lesions on Citrus limon and Gossypium hirsutum, respectively. Essential oils extracted by steam distillation from the fresh aerial parts of Pelargonium graveolens and Schinus molle exhibited complete (100%) inhibition of bacterial growth in vitro at a concentration of 1000 ppm, as determined by diffusion tests. To evaluate the potential of these essential oils for controlling Xanthomonas-induced diseases, in vivo assays were conducted on lemon leaves and cotton cotyledons inoculated with the regional AE28 and RQ3 strains. Two treatment approaches were tested: preventive application (24 h before inoculation) and curative application (24 h after inoculation). Preventive and curative treatments with P. graveolens essential oil significantly reduced citrus canker severity, whereas S. molle essential oil did not show a significant reduction compared to the control. In contrast, regardless of the treatment’s timing, both essential oils effectively reduced bacterial blight severity in cotton cotyledons by approximately 1.5-fold. Gas chromatography–mass spectrometry (GC-MS) analysis identified geraniol and citronellol as the major components of P. graveolens essential oil, while limonene and t-cadinol were predominant in S. molle. These findings highlight the promising potential of botanical products as bactericidal agents, warranting further research to optimize their application and efficacy. Full article

Xanthomonas citri subsp. citri (Xcc) is a Gram-negative bacterium responsible for citrus canker, a significant threat to citrus crops. ClpV is a critical protein in the type VI secretion system (T6SS) as an ATPase involved in bacterial motility, adhesion, and pathogenesis to the host for some pathogenic bacteria. In order to investigate the function of clpV gene in Xcc, the clpV-deletion strain ΔclpV was constructed, its biological properties were evaluated, and the differences in gene expression levels between the wild-type strain and ΔclpV were analyzed by transcriptomics. The results exhibited significantly reduced biofilm formation, extracellular polysaccharide synthesis, and swarming motility in ΔclpV compared to the wild-type strain. Although the clpV-deletion did not significantly affect bacterial growth or pathogenicity in terms of disease symptoms on citrus leaves, the mutant showed increased sensitivity to environmental stresses (NaCl, SDS, and H₂O₂) and antibiotics (β-lactams and aminoglycosides). Transcriptome analysis revealed that clpV-deletion altered the expression of motility-related genes and the efflux pump gene mexH. Our findings underscore the importance of ClpV in maintaining biofilm integrity and suggest a multifaceted role in adaptive strategies of Xcc, positioning ClpV as a potential target for mitigating citrus canker disease. Full article

Citrus bacterial canker disease, caused by Xanthomonas citri subsp. citri (Xcc), poses a significant global threat to the citrus industry. Lateral organ boundaries 1 (Lob1) is confirmed as a citrus susceptibility gene that induces pathogenesis by interaction with the PthA4 effector of Xcc. Citron C-05 (Citrus medica) is a Citrus genotype resistant to Xcc. However, there is little information available on the regulation of Lob1 in resistant genotypes, which is important for the breeding of citrus cultivars resistant to canker disease. This study aimed to identify upstream regulatory factors of Lob1 in Citron C-05 and to investigate its function in disease resistance. ‘Bingtang’ sweet orange (C. sinensis), a susceptible genotype, was utilized as the control. cDNA yeast libraries of Xcc-induced Citron C-05 and ‘Bingtang’ sweet orange were constructed. The capacities of ‘Bingtang’ and Citron C-05 were 1.896 × 10⁷ and 2.154 × 10⁷ CFU, respectively. The inserted fragments ranged from 500 to 2000 bp with a 100% recombination rate. The promoter of Lob1 was segmented into two pieces and the P1 fragment from both genotypes was used to construct a bait yeast (PAbAi-CsLob1-P1; PAbAi-CmLob1-P1). Through library screening with the bait yeast, upstream regulators interacting with the Lob1-P1 promoter were identified and then validated using Y1H and dual-luciferase tests. The expression analysis of the three transcript factors indicated that RMA3 was upregulated by inoculation with Xcc in the resistant Citron C-05, but not in the susceptible sweet orange. The overexpression of CsRMA3 in ‘Bingtang’ sweet orange led to reduced canker symptoms, with a significantly lower pathogen density in the leaves following Xcc inoculation. When CmRMA3 was silenced by virus-induced gene silencing (VIGS) in Citron C-05, typical canker symptoms appeared on the CmRMA3-silenced leaves at 15 days post-inoculation with Xcc. Further expression analyses revealed that the CmRMA3 transcription factor suppressed the expression of Lob1. These results suggest that RMA3 participates in the resistant reaction of Citron C-05 to Xcc infection, and such a response might be in relation to its suppression of the expression of the pathogenic gene Lob1. Full article

Peptidoglycan hydrolases are enzymes responsible for breaking the peptidoglycan present in the bacterial cell wall, facilitating cell growth, cell division and peptidoglycan turnover. Xanthomonas citri subsp. citri (X. citri), the causal agent of citrus canker, encodes an Escherichia coli M23 peptidase EnvC homolog. EnvC is a LytM factor essential for cleaving the septal peptidoglycan, thereby facilitating the separation of daughter cells. In this study, the investigation focused on EnvC contribution to the virulence and cell separation of X. citri. It was observed that disruption of the X. citri envC gene (ΔenvC) led to a reduction in virulence. Upon inoculation into leaves of Rangpur lime (Citrus limonia Osbeck), the X. citri ΔenvC exhibited a delayed onset of citrus canker symptoms compared with the wild-type X. citri. Mutant complementation restored the wild-type phenotype. Sub-cellular localization confirmed that X. citri EnvC is a periplasmic protein. Moreover, the X. citri ΔenvC mutant exhibited elongated cells, indicating a defect in cell division. These findings support the role of EnvC in the regulation of cell wall organization, cell division, and they clarify the role of this peptidase in X. citri virulence. Full article

Citrus bacterial canker (CBC) is a severe bacterial infection caused by Xanthomonas citri subsp. citri (Xcc), which continues to adversely impact citrus production worldwide. Members of the GATA family are important regulators of plant development and regulate plant responses to particular stressors. This report aimed to systematically elucidate the Citrus sinensis genome to identify and annotate genes that encode GATAs and evaluate the functional importance of these CsGATAs as regulators of CBC resistance. In total, 24 CsGATAs were identified and classified into four subfamilies. Furthermore, the phylogenetic relationships, chromosomal locations, collinear relationships, gene structures, and conserved domains for each of these GATA family members were also evaluated. It was observed that Xcc infection induced some CsGATAs, among which CsGATA12 was chosen for further functional validation. CsGATA12 was found to be localized in the nucleus and was differentially upregulated in the CBC-resistant and CBC-sensitive Kumquat and Wanjincheng citrus varieties. When transiently overexpressed, CsGATA12 significantly reduced CBC resistance with a corresponding increase in abscisic acid, jasmonic acid, and antioxidant enzyme levels. These alterations were consistent with lower levels of salicylic acid, ethylene, and reactive oxygen species. Moreover, the bacteria-induced CsGATA12 gene silencing yielded the opposite phenotypic outcomes. This investigation highlights the important role of CsGATA12 in regulating CBC resistance, underscoring its potential utility as a target for breeding citrus varieties with superior phytopathogen resistance. Full article

Citrus canker caused by Xanthomonas citri subsp. citri is a devastating bacterial disease with severe implications for the citrus industry. Microorganisms possessing biocontrol capabilities against X. citri subsp. citri offer a highly promising strategy for healthy citrus management. In the present study, a broad-spectrum antagonist strain ZJLMBA1908 with potent antibacterial activity against X. citri subsp. citri was isolated from symptomatic lemon leaves, and identified as Bacillus amyloliquefaciens. Cell-free supernatant (CFS) of strain ZJLMBA1908 also exhibited remarkable antimicrobial activity, especially suppressing the growth of X. citri subsp. citri and Nigrospora oryzae, with inhibition rates of 27.71% and 63.75%, respectively. The antibacterial crude extract (CE) derived from the CFS displayed effective activity against X. citri subsp. citri. A preventive treatment using the CE significantly reduced the severity and incidence of citrus canker in a highly susceptible citrus host. Additionally, the CE maintained activity in the presence of protease and under a wide range of temperature and pH treatments. Applying high-performance liquid chromatography (HPLC) to separate and purify the CE resulted in the discovery of one highly potent anti-X. citri subsp. citri subfraction, namely CE3, which could completely inhibit the growth of X. citri subsp. citri. Liquid chromatography–electrospray ionization–mass spectrometry (LC–ESI–MS) analysis revealed that CE3 mainly consisted of palmitic acid, surfactin C15, phytosphingosine and dihydrosphingosine. Taken together, the results contribute to the possible biocontrol mechanisms of B. amyloliquefaciens ZJLMBA1908, as well as providing a promising new candidate strain as a biological control agent for controlling citrus canker. Full article

To explore more efficient and less toxic antibacterial and antifungal pesticides, we utilized 2,6-difluorobenzamide as a starting material and ultimately synthesized 23 novel benzoylurea derivatives containing a pyrimidine moiety. Their structures were characterized and confirmed by ¹H NMR, ¹³C NMR, ¹⁹F NMR, and HRMS. The bioassay results demonstrated that some of the title compounds exhibited moderate to good in vitro antifungal activities against Botrytis cinerea in cucumber, Botrytis cinerea in tobacco, Botrytis cinerea in blueberry, Phomopsis sp., and Rhizoctonia solani. Notably, compounds 4j and 4l displayed EC₅₀ values of 6.72 and 5.21 μg/mL against Rhizoctonia solani, respectively, which were comparable to that of hymexazol (6.11 μg/mL). Meanwhile, at 200 and 100 concentrations, the target compounds 4a–4w exhibited lower in vitro antibacterial activities against Xanthomonas oryzae pv. oryzicola and Xanthomonas citri subsp. citri, respectively, compared to those of thiodiazole copper. Furthermore, the molecular docking simulation demonstrated that compound 4l formed hydrogen bonds with SER-17 and SER-39 of succinate dehydrogenase (SDH), providing a possible explanation for the mechanism of action between the target compounds and SDH. This study represents the first report on the antifungal and antibacterial activities of novel benzoylurea derivatives containing a pyrimidine moiety. Full article

Citrus canker, caused by the bacterium Xanthomonas citri (Xcc), is one of the most devastating diseases for the citrus industry. Xylose is a constituent of the cell wall of plants, and the ability of Xcc to use this carbohydrate may play a role in virulence. Xcc has two genes codifying for xylose isomerase (XI), a bifunctional enzyme that interconverts D-xylose into D-xylulose and D-glucose into D-fructose. The aim of this work was to investigate the functional role of the two putative XI ORFs, XAC1776 (xylA1) and XAC4225 (xylA2), in Xcc pathogenicity. XI-coding genes of Xcc were deleted, and the single mutants (XccΔxylA1 or XccΔxylA2) or the double mutant (XccΔxylA1ΔxylA2) remained viable. The deletion of one or both XI genes (xylA1 and/or xylA2) increased the aggressiveness of the mutants, causing disease symptoms. RT-qPCR analysis of wild strain and xylA deletion mutants grown in vivo and in vitro revealed that the highest expression level of hrpX and xylR was observed in vivo for the double mutant. The results indicate that XI depletion increases the expression of the hrp regulatory genes in Xcc. We concluded that the intracellular accumulation of xylose enhances Xcc virulence. Full article

Biological control of plant diseases has gained attraction for controlling various bacterial diseases at a field trial stage. An isolated endophytic bacterium, Bacillus velezensis 25 (Bv-25), from Citrus species had strong antagonistic activity against Xanthomonas citri subsp. citri (Xcc), which causes citrus canker disease. When Bv-25 was incubated in Landy broth or yeast nutrient broth (YNB), the ethyl acetate extract of Landy broth exhibited higher levels of antagonistic activity against Xcc compared to that of YNB. Therefore, the antimicrobial compounds in the two ethyl acetate extracts were detected by high performance liquid chromatography–mass spectrometry. This comparison revealed an increase in production of several antimicrobial compounds, including difficidin, surfactin, fengycin, and Iturin-A or bacillomycin-D by incubation in Landy broth. RNA sequencing for the Bv-25 grown in Landy broth were performed, and the differential expressions were detected for the genes encoding the enzymes for the synthesis of antimicrobial compounds, such as bacilysin, plipastatin or fengycin, surfactin, and mycosubtilin. Combination of metabolomics analysis and RNA sequencing strongly suggests that several antagonistic compounds, especially bacilysin produced by B. velezensis, exhibit an antagonistic effect against Xcc. Full article

Isolation for antibacterial compounds from natural plants is a promising approach to develop new pesticides. In this study, two compounds were obtained from the Chinese endemic plant Piper austrosinense using bioassay-guided fractionation. Based on analyses of ¹H-NMR, ¹³C-NMR, and mass spectral data, the isolated compounds were identified as 4-allylbenzene-1,2-diol and (S)-4-allyl-5-(1-(3,4-dihydroxyphenyl)allyl)benzene-1,2-diol. 4-Allylbenzene-1,2-diol was shown to have strong antibacterial activity against four plant pathogens, including Xanthomonas oryzae pathovar oryzae (Xoo), X. axonopodis pv. citri (Xac), X. oryzae pv. oryzicola (Xoc) and X. campestris pv. mangiferaeindicae (Xcm). Further bioassay results exhibited that 4-allylbenzene-1,2-diol had a broad antibacterial spectrum, including Xoo, Xac, Xoc, Xcm, X. fragariae (Xf), X. campestris pv. campestris (Xcc), Pectobacterium carotovorum subspecies brasiliense (Pcb) and P. carotovorum subsp. carotovorum (Pcc), with minimum inhibitory concentration (MIC) values ranging from 333.75 to 1335 μmol/L. The pot experiment showed that 4-allylbenzene-1,2-diol exerted an excellent protective effect against Xoo, with a controlled efficacy reaching 72.73% at 4 MIC, which was superior to the positive control kasugamycin (53.03%) at 4 MIC. Further results demonstrated that the 4-allylbenzene-1,2-diol damaged the integrity of the cell membrane and increased cell membrane permeability. In addition, 4-allylbenzene-1,2-diol also prevented the pathogenicity-related biofilm formation in Xoo, thus limiting the movement of Xoo and reducing the production of extracellular polysaccharides (EPS) in Xoo. These findings suggest the value of 4-allylbenzene-1,2-diol and P. austrosinense could be as promising resources for developing novel antibacterial agents. Full article

Citrus canker (CC), caused by one of the most destructive subfamilies of the bacterial phytopathogen Xanthomonas citri subsp. Citri (Xcc), poses a serious threat to the significantly important citrus fruit crop grown worldwide. This has been the subject of ongoing epidemiological and disease management research. Currently, five different forms have been identified of CC, in which Canker A (Xanthomonas citri subsp. citri) being the most harmful and infecting the majority of citrus cultivars. Severe infection symptoms include leaf loss, premature fruit drop, dieback, severe fruit blemishing or discoloration, and a decrease in fruit quality. The infection spreads rapidly through wind, rain splash, and warm and humid climates. The study of the chromosomal and plasmid DNA of bacterium has revealed the evolutionary pattern among the pathovars, and research on the Xcc genome has advanced our understanding of how the bacteria specifically recognize and infect plants, spread within the host, and propagates itself. Quarantine or exclusion programs, which prohibit the introduction of infected citrus plant material into existing stock, are still in use. Other measures include eliminating sources of inoculum, using resistant hosts, applying copper spray for protection, and implementing windbreak systems. The main focus of this study is to highlight the most recent developments in the fields of Xcc pathogenesis, epidemiology, symptoms, detection and identification, host range, spread, susceptibility, and management. Additionally, it presents an analysis of the economic impact of this disease on the citrus industry and suggests strategies to reduce its spread, including the need for international collaboration and research to reduce the impact of this disease on the global citrus industry. Full article

Xanthomonas citri pv. citri (Xcc) (X. citri subsp. citri) type A is the causal agent of citrus bacterial canker (CBC) on most Citrus spp. and close relatives. Two narrow-host-range strains of Xcc, A^w and A*, from Florida and Southwest Asia, respectively, infect only Mexican lime (Citrus aurantifolia) and alemow (C. macrophylla). In the initial stage of infection, these xanthomonads enter via stomata to reach the apoplast. Herein, we investigated the differences in chemotactic responses for wide and narrow-host-range strains of Xcc A, X. euvesicatoria pv. citrumelonis (X. alfalfae subsp. citrumelonis), the causal agent of citrus bacterial spot, and X. campestris pv. campestris, the crucifer black rot pathogen. These strains of Xanthomonas were compared for carbon source use, the chemotactic responses toward carbon compounds, chemotaxis sensor content, and responses to apoplastic fluids from Citrus spp. and Chinese cabbage (Brassica pekinensis). Different chemotactic responses occurred for carbon sources and apoplastic fluids, depending on the Xanthomonas strain and the host plant from which the apoplastic fluid was derived. Differential chemotactic responses to carbon sources and citrus apoplasts suggest that these Xanthomonas strains sense host-specific signals that facilitate their location and entry of stomatal openings or wounds. Full article

Citrus canker, caused by the bacterium Xanthomonas citri subsp. citri (Xcc), is a disease that causes serious problems to the global citrus industry. Matrix-Assisted Laser Desorption/Ionization Time-of-Flight (MALDI-TOF) Mass Spectrometry (MS) has been used in human medicine to diagnose various diseases caused by both fungi and bacteria. In agriculture, this technique has potential for the diagnosis of diseases due to the low cost of large-scale analysis and quickness. This study showed that MALDI-TOF MS combined with chemometric analysis was effective for differentiating the macromolecule profile of orange leaves with canker lesions, healthy leaves, and leaves with phytotoxicity symptoms, proving that this technique may be used for the rapid diagnosis of citrus canker. Full article

Citrus canker is a quarantined disease caused by the bacterial plant pathogen Xanthomonas citri subsp. citri (Xcc), which causes persistent surface damage, leaf and fruit drop, and tree decline in citrus plants. The citrus cultivar Citron C-05 (Citrus medica L.) is a disease-resistant genotype identified after years of screening at the National Center for Citrus Improvement (Changsha), which displays allergic, necrotic, and disease-resistant responses to Xcc. In this study, the BAK1 gene was identified in this cultivar to be a disease resistance gene involved in plant-microbe interaction between citrus and Xcc. Functional investigations of this gene revealed that both CsBAK1 (C. sinensis BAK1) or CmBAK1(C. medica BAK1) could inhibit the growth of Xcc to some extent when transiently expressed in the susceptible ‘Bingtang’ genotype of sweet orange. Critical regions of the CmBAK1 promoter sequence were identified by creating downstream deletions and exposing mutants to Xcc to determine effects on the resistance phenotype; a 426 bp region (−2000~–1574) was identified as a key functional region responsible for eliciting the hypersensitive response in plants. Through screening arrayed Citron C-05 cDNA libraries by yeast one-hybrid assays, a basic APETALA2/ETHYLENE RESPONSE FACTOR (AP2/ERF) transcription factor of CmRAP2-13 that binds directly to the 426 bp key sequence and activates expression of CmBAK1 was identified. Moreover, transcriptional analysis revealed an obvious increase in transcript levels of CsRAP2-13 in Citron C-05, American citron, and Finger citron. In this study, we present the identification of transcriptional activators that are found to interact with BAK1 proteins in response to Xcc. These results reveal a coordinated regulatory mechanism of RAP2-13, which may be involved in defence responses through the regulation of BAK1. Full article