Getting to the root of Ralstonia invasion.

Plant diseases caused by soilborne pathogens are a major limiting factor in crop production. Bacterial wilt disease, caused by soilborne bacteria in the Ralstonia solanacearum Species Complex (Ralstonia), results in significant crop loss throughout the world. Ralstonia invades root systems and colonizes plant xylem, changing plant physiology and ultimately causing plant wilting in susceptible varieties. Elucidating how Ralstonia invades and colonizes plants is central to developing strategies for crop protection. Here we review Ralstonia pathogenesis from root detection and attachment, early root colonization, xylem invasion and subsequent wilting. We focus primarily on studies in tomato from the last 5-10 years. Recent work has identified elegant mechanisms Ralstonia uses to adapt to the plant xylem, and has discovered new genes that function in Ralstonia fitness in planta. A picture is emerging of an amazingly versatile pathogen that uses multiple strategies to make its surrounding environment more hospitable and can adapt to new environments.

LC-MS/MS-based phospholipid profiling of plant-pathogenic bacteria with tailored separation of methyl-branched species.

Plant-pathogenic bacteria are one of the major constraints on agricultural yield. In order to selectively treat these bacteria, it is essential to understand the molecular structure of their cell membrane. Previous studies have focused on analyzing hydrolyzed fatty acids (FA) due to the complexity of bacterial membrane lipids. These studies have highlighted the occurrence of branched-chain fatty acids (BCFA) alongside normal-chain fatty acids (NCFA) in many bacteria. As several FA are bound in the intact phospholipids of the bacterial membrane, the presence of isomeric FA complicates lipid analysis. Furthermore, commercially available reference standards do not fully cover potential lipid isomers. To address this issue, we have developed a reversed-phase high-performance liquid chromatography (RP-HPLC) method with tandem mass spectrometry (MS/MS) to analyze the phospholipids of various plant-pathogenic bacteria with a focus on BCFA containing phospholipids. The study revealed the separation of three isomeric phosphatidylethanolamines (PE) depending on the number of bound BCFA to NCFA. The validation of the retention order was based on available reference standards in combination with the analysis of hydrolyzed fatty acids through gas chromatography with mass spectrometry (GC/MS) after fractionation. Additionally, the transferability of the retention order to other major lipid classes, such as phosphatidylglycerols (PG) and cardiolipins (CL), was thoroughly examined. Using the information regarding the retention behavior, the phospholipid profile of six plant-pathogenic bacteria was structurally elucidated. Furthermore, the developed LC-MS/MS method was used to classify the plant-pathogenic bacteria based on the number of bound BCFA in the phospholipidome.

Genomic analysis provides novel insights into diversification and taxonomy of Allorhizobium vitis (i.e. Agrobacterium vitis).

BACKGROUND: Allorhizobium vitis (formerly named Agrobacterium vitis or Agrobacterium biovar 3) is the primary causative agent of crown gall disease of grapevine worldwide. We obtained and analyzed whole-genome sequences of diverse All. vitis strains to get insights into their diversification and taxonomy. RESULTS: Pairwise genome comparisons and phylogenomic analysis of various All. vitis strains clearly indicated that All. vitis is not a single species, but represents a species complex composed of several genomic species. Thus, we emended the description of All. vitis, which now refers to a restricted group of strains within the All. vitis species complex (i.e. All. vitis sensu stricto) and proposed a description of a novel species, All. ampelinum sp. nov. The type strain of All. vitis sensu stricto remains the current type strain of All. vitis, K309T. The type strain of All. ampelinum sp. nov. is S4T. We also identified sets of gene clusters specific to the All. vitis species complex, All. vitis sensu stricto and All. ampelinum, respectively, for which we predicted the biological function and infer the role in ecological diversification of these clades, including some we could experimentally validate. All. vitis species complex-specific genes confer tolerance to different stresses, including exposure to aromatic compounds. Similarly, All. vitis sensu stricto-specific genes confer the ability to degrade 4-hydroxyphenylacetate and a putative compound related to gentisic acid. All. ampelinum-specific genes have putative functions related to polyamine metabolism and nickel assimilation. Congruently with the genome-based classification, All. vitis sensu stricto and All. ampelinum were clearly delineated by MALDI-TOF MS analysis. Moreover, our genome-based analysis indicated that Allorhizobium is clearly separated from other genera of the family Rhizobiaceae. CONCLUSIONS: Comparative genomics and phylogenomic analysis provided novel insights into the diversification and taxonomy of Allorhizobium vitis species complex, supporting our redefinition of All. vitis sensu stricto and description of All. ampelinum. Our pan-genome analyses suggest that these species have differentiated ecologies, each relying on specialized nutrient consumption or toxic compound degradation to adapt to their respective niche.

Genomic and Biological Characterization of Ralstonia solanacearum Inovirus Brazil 1, an Inovirus that Alters the Pathogenicity of the Phytopathogen Ralstonia pseudosolanacearum.

Filamentous bacteriophages contain a single-stranded DNA genome and have a peculiar lifestyle, since they do not cause host cell lysis, but establish a persistent association with the host, often causing behavioral changes, with effects on bacterial ecology. Over the years, a gradual reduction in the incidence of bacterial wilt has been observed in some fields from Brazil. This event, which has been associated with the loss of pathogenicity of Rasltonia spp. isolates due to infection by filamentous viruses of the inovirus group, is widely reported for Ralstonia spp. Asian isolates infected by inoviruses. In an attempt to elucidate which factors are associated with the phenomenon reported in Brazil, we investigated one isolate of R. solanacearum (UB-2014), with unusual characteristics for R. solanacearum, obtained from eggplant with mild wilt symptoms. To verify if the presence of filamentous bacteriophage was related to this phenotype, we performed viral purification and nucleic acid extraction. The phage genome was sequenced, and phylogenetic analyses demonstrated that the virus belongs to the family Inoviridae and was named as Ralstonia solanacerarum inovirus Brazil 1 (RSIBR1). RSIBR1 was transmitted to R. pseudosolanacearum GMI1000, and the virus-infected GMI1000 (GMI1000 VI) isolate showed alterations in phenotypic characteristics, as well as loss of pathogenicity, similarly to that observed in R. solanacearum isolate UB-2014. The presence of virus-infected UB-2014 and GMI1000 VI plants without symptoms, after 3 months, confirms that the infected isolates can colonize the plant without causing disease, which demonstrates that the phage infection changed the behavior of these pathogens.

The γ-Core Motif Peptides of AMPs from Grasses Display Inhibitory Activity against Human and Plant Pathogens.

Antimicrobial peptides (AMPs) constitute an essential part of the plant immune system. They are regarded as alternatives to conventional antibiotics and pesticides. In this study, we have identified the γ-core motifs, which are associated with antimicrobial activity, in 18 AMPs from grasses and assayed their antimicrobial properties against nine pathogens, including yeasts affecting humans, as well as plant pathogenic bacteria and fungi. All the tested peptides displayed antimicrobial properties. We discovered a number of short AMP-derived peptides with high antimicrobial activity both against human and plant pathogens. For the first time, antimicrobial activity was revealed in the peptides designed from the 4-Cys-containing defensin-like peptides, whose role in plant immunity has remained unknown, as well as the knottin-like peptide and the C-terminal prodomain of the thionin, which points to the direct involvement of these peptides in defense mechanisms. Studies of the mode of action of the eight most active γ-core motif peptides on yeast cells using staining with propidium iodide showed that all of them induced membrane permeabilization leading to cell lysis. In addition to identification of the antimicrobial determinants in plant AMPs, this work provides short candidate peptide molecules for the development of novel drugs effective against opportunistic fungal infections and biopesticides to control plant pathogens.

Ralstonia solanacearum Type III Effector RipJ Triggers Bacterial Wilt Resistance in Solanum pimpinellifolium.

Ralstonia solanacearum causes bacterial wilt disease in solanaceous crops. Identification of avirulence type III-secreted effectors recognized by specific disease resistance proteins in host plant species is an important step toward developing durable resistance in crops. In the present study, we show that R. solanacearum effector RipJ functions as an avirulence determinant in Solanum pimpinellifolium LA2093. In all, 10 candidate avirulence effectors were shortlisted based on the effector repertoire comparison between avirulent Pe_9 and virulent Pe_1 strains. Infection assays with transgenic strain Pe_1 individually carrying a candidate avirulence effector from Pe_9 revealed that only RipJ elicits strong bacterial wilt resistance in S. pimpinellifolium LA2093. Furthermore, we identified that several RipJ natural variants do not induce bacterial wilt resistance in S. pimpinellifolium LA2093. RipJ belongs to the YopJ family of acetyltransferases. Our sequence analysis indicated the presence of partially conserved putative catalytic residues. Interestingly, the conserved amino acid residues in the acetyltransferase catalytic triad are not required for effector-triggered immunity. In addition, we show that RipJ does not autoacetylate its lysine residues. Our study reports the identification of the first R. solanacearum avirulence protein that triggers bacterial wilt resistance in tomato. We expect that our discovery of RipJ as an avirulence protein will accelerate the development of bacterial wilt-resistant tomato varieties in the future.[Formula: see text] Copyright © 2021 The Author(s). This is an open access article distributed under the CC BY 4.0 International license.

Antimicrobial activity of the volatile compound 3,5-dichloro-4-methoxybenzaldehyde, produced by the mushroom Porostereum spadiceum, against plant-pathogenic bacteria and fungi.

AIMS: In this study, volatile compounds released from mycelia of some aromatic mushrooms were investigated for their inhibitory activity against plant-pathogenic bacteria and fungi. METHODS AND RESULTS: A screening revealed that volatile compounds from mycelia of Porostereum spadiceum remarkably inhibited the colony formation of plant-pathogenic bacteria, including Clavibacter michiganensis subsp. michiganensis and Ralstonia solanacearum while also inhibiting the conidial germination of plant-pathogenic fungi including Alternaria brassicicola and Colletotrichum orbiculare. The volatile compounds were isolated from the culture filtrate of P. spadiceum, and 3,4-dichloro-4-methoxybenzaldehyde (DCMB) was identified as a major compound. DCMB significantly inhibited bacterial colonization at 10 µg ml-1 and fungal conidial germination at 0·1-1 µg ml-1 as a vapour. CONCLUSIONS: This is the first report on the production of the volatile compound DCMB by P. spadiceum and on the antimicrobial activity of DCMB against plant-pathogenic bacteria and fungi at low concentrations. It may be possible to use the compound as an agent for protecting crops from bacterial and fungal diseases during cultivation and storage. SIGNIFICANCE AND IMPACT OF THE STUDY: This study provides an understanding of antimicrobial activity of the mushroom volatile compound that may be useful as a novel biological control agent for protecting various plant diseases.

Development of a loop-mediated isothermal amplification assay for specific detection of all known subspecies of Clavibacter michiganensis.

AIMS: Clavibacter michiganensis is an important bacterial plant pathogen that causes vast destruction to agriculturally important crops worldwide. Early detection is critical to evaluate disease progression and to implement efficient control measures to avoid serious epidemics. In this study, we developed a sensitive, specific and robust loop-mediated isothermal amplification (LAMP) assay for detection of all known subspecies of C. michiganensis. METHODS AND RESULTS: Whole genome comparative genomics approach was taken to identify a unique and conserved region within all known subspecies of C. michiganensis. Primer specificity was evaluated in silico and with 64 bacterial strains included in inclusivity and exclusivity panels; no false positives or false negatives were detected. Both the sensitivity and spiked assay of the developed LAMP assay was 1 fg of the pathogen DNA per reaction. A 100% accuracy was observed when tested with infected plant samples. CONCLUSIONS: The developed LAMP assay is simple, sensitive, robust and easy to perform using different detection platforms and chemistries. SIGNIFICANCE AND IMPACT OF THE STUDY: The developed LAMP assay can detect all known subspecies of C. michiganensis. The LAMP process can be performed isothermally at 65°C and results can be visually assessed, which makes this technology a promising tool for monitoring the disease progression and for accurate pathogen detection at point-of-care.

Antimicrobial activities of an oxygenated cyclohexanone derivative isolated from Amphirosellinia nigrospora JS-1675 against various plant pathogenic bacteria and fungi.

AIMS: To evaluate the antimicrobial activities of an active compound isolated from the culture broth of Amphirosellinia nigrospora JS-1675 against various plant pathogenic bacteria and fungi. METHODS AND RESULTS: While screening for bioactive secondary metabolites from endophytic fungi, we found that A. nigrospora JS-1675 showed strong in vitro antibacterial activity against Ralstonia solanacearum. One compound (1) was isolated and identified as (4S, 5S, 6S)-5,6-epoxy-4-hydroxy-3-methoxy-5-methyl-cyclohex-2-en-1-one. Growth of most of the tested phytopathogenic bacteria was inhibited by compound 1 and the ethyl acetate (EtOAc) layer except Pseudomonas syringae pv. lachrymans. Compound 1 also inhibited the mycelial growth of several plant pathogenic fungi. Both compound 1 and the EtOAc layer reduced bacterial leaf spot disease in detached peach leaves. They also suppressed the development of bacterial wilt on tomato seedlings quite effectively. CONCLUSIONS: Amphirosellinia nigrospora JS-1675 showed antimicrobial activity against plant pathogenic bacteria and fungi by producing compound 1. SIGNIFICANCE AND IMPACT OF THE STUDY: This is the first report on the occurrence of compound 1 in A. nigrospora JS-1675 and its efficacy against plant pathogenic bacteria and fungi. Their strong disease control efficacy against tomato bacterial wilt suggests that this fungus can be used as a microbial bactericide.

The Role of Proteases in the Virulence of Plant Pathogenic Bacteria.

A pathogenic lifestyle is inextricably linked with the constant necessity of facing various challenges exerted by the external environment (both within and outside the host). To successfully colonize the host and establish infection, pathogens have evolved sophisticated systems to combat the host defense mechanisms and also to be able to withstand adverse environmental conditions. Proteases, as crucial components of these systems, are involved in a variety of processes associated with infection. In phytopathogenic bacteria, they play important regulatory roles and modulate the expression and functioning of various virulence factors. Secretory proteases directly help avoid recognition by the plant immune systems, and contribute to the deactivation of the defense response pathways. Finally, proteases are important components of protein quality control systems, and thus enable maintaining homeostasis in stressed bacterial cells. In this review, we discuss the known protease functions and protease-regulated signaling processes associated with virulence of plant pathogenic bacteria.

Insect-Borne Plant Pathogens and Their Vectors: Ecology, Evolution, and Complex Interactions.

The transmission of insect-borne plant pathogens, including viruses, bacteria, phytoplasmas, and fungi depends upon the abundance and behavior of their vectors. These pathogens should therefore be selected to influence their vectors to enhance their transmission, either indirectly, through the infected host plant, or directly, after acquisition of the pathogen by the vector. Accumulating evidence provides partial support for the occurrence of vector manipulation by plant pathogens, especially for plant viruses, for which a theoretical framework can explain patterns in the specific effects on vector behavior and performance depending on their modes of transmission. The variability in effects of pathogens on their vectors, however, suggests inconsistency in the occurrence of vector manipulation but also may reflect incomplete information about these systems. For example, manipulation can occur through combinations of specific effects, including direct and indirect effects on performance and behavior, and dynamics in those effects with disease progression or pathogen acquisition that together constitute syndromes that promote pathogen spread. Deciphering the prevalence and forms of vector manipulation by plant pathogens remains a compelling field of inquiry, but gaps and opportunities to advance it remain. A proposed research agenda includes examining vector manipulation syndromes comprehensively within pathosystems, expanding the taxonomic and genetic breadth of the systems studied, evaluating dynamic effects that occur during disease progression, incorporating the influence of biotic and abiotic environmental factors, evaluating the effectiveness of putative manipulation syndromes under field conditions, deciphering chemical and molecular mechanisms whereby pathogens can influence vectors, expanding the use of evolutionary and epidemiological models, and seeking opportunities to exploit these effects to improve management of insect-borne, economically important plant pathogens. We expect this field to remain vibrant and productive in its own right and as part of a wider inquiry concerning host and vector manipulation by plant and animal pathogens and parasites.

Die another day: Molecular mechanisms of effector-triggered immunity elicited by type III secreted effector proteins.

Bacterial pathogens inject type III secreted effector (T3SE) proteins into their hosts where they display dual roles depending on the host genotype. T3SEs promote bacterial virulence in susceptible hosts, and elicit immunity in resistant hosts. T3SEs are typically recognized when they modify a host target that is associated with a NOD-like receptor protein. We focus on the molecular mechanisms of T3SE recognition in plants. Plants guard multiple nodes of the immune signaling pathway, from recognition at the cell surface by receptor-like kinases to nuclear signaling. Some nodes are bacterial virulence targets, while other nodes are decoys that resemble true virulence targets.

Antibiotic Resistance in Plant Pathogenic Bacteria: Recent Data and Environmental Impact of Unchecked Use and the Potential of Biocontrol Agents as an Eco-Friendly Alternative.

The economic impact of phytopathogenic bacteria on agriculture is staggering, costing billions of US dollars globally. Pseudomonas syringae is the top most phytopathogenic bacteria, having more than 60 pathovars, which cause bacteria speck in tomatoes, halo blight in beans, and so on. Although antibiotics or a combination of antibiotics are used to manage infectious diseases in plants, they are employed far less in agriculture compared to human and animal populations. Moreover, the majority of antibiotics used in plants are immediately washed away, leading to environmental damage to ecosystems and food chains. Due to the serious risk of antibiotic resistance (AR) and the potential for environmental contamination with antibiotic residues and resistance genes, the use of unchecked antibiotics against phytopathogenic bacteria is not advisable. Despite the significant concern regarding AR in the world today, there are inadequate and outdated data on the AR of phytopathogenic bacteria. This review presents recent AR data on plant pathogenic bacteria (PPB), along with their environmental impact. In light of these findings, we suggest the use of biocontrol agents as a sustainable, eco-friendly, and effective alternative to controlling phytopathogenic bacteria.

Bacterial diversity, community structure and function in association of potato scabby tubers during storage in northern Thailand.

Potato scab is a common potato tuber disease that affects quality and cost in the marketplace, shortening storage, and increasing the chance for secondary infection. The tubers with disease severity of 1 to 4 are accepted and stored in potato storage for cheap selling in Thailand. However, there are few studies of the bacterial community of the scabby tuber during storage. Thus, we aim to elucidate the diversity, structure, and function of the bacterial community of 30-day storage potato scabby tubers stored in different temperatures using 16S amplicon metagenomic sequencing. Bacterial communities of storage potato scabby tubers (Spunta cultivar) collected from different storage temperatures, 4 °C (MEP1) and 6 °C (MEP2), were characterized using 16S rRNA amplicon metagenomic sequencing. The alpha-diversity abundance in the bacteriome of the scabby tubers stored at 6 °C was higher than in those stored at 4 °C. Actinobacteria (34.7%) was a dominant phylum in MEP1, while Proteobacteria (39.9%) was predominant in MEP2. The top 10 genera of both communities were Rhizobium group, Streptomyces, Pectobacterium, Ruminococcus, Cellulomonas, Promicromonospora, Prevotella, Enterobacter, Pedobacter, and Paenarthrobacter. Moreover, functional profile prediction of both communities reveals essential genes in the pathosystem: nos, bglA, and cebEFG-msiK for potato scab disease and phc and peh operons for rot disease. Our findings are the first study to explore details of the bacteriome of the accepted potato scabby tubers for selling during storage in Thailand and strongly indicate that although potatoes were stored at low temperatures, diseases still occur by secondary pathogens.

Comparison of the antimicrobial effects of semipurified cyclotides from Iranian Viola odorata against some of plant and human pathogenic bacteria.

AIMS: Cyclotides are mini-proteins that are synthesized via the ribosomal pathway. They have a variety of biological activities such as antimicrobial, antitumour, anti-HIV activities. Because of their various bioactivities and unique stability, they are suitable candidate in drug design applications. The main aim of this study was to determine new antimicrobial agents, which can be used instead of chemical antibiotics. For this reason, we compared the antimicrobial effects of semipurified cyclotides against human and plant pathogenic bacteria. METHODS AND RESULTS: The cyclotides were isolated from the Iranian plant Viola odorata by fractionation methods and semipurified on a SPE-C18 column chromatography. Antimicrobial activities of extracted cyclotides were studied by radial diffusion assays (RDAs), minimal inhibitory concentration (MIC) and minimal bactericidal concentration (MBC). Data analysis showed that MIC of semipurified cyclotides was 1.6 mg ml(-1) against Staphylococcus aureus, gram-positive bacteria. It was also revealed they are the most susceptible among human pathogenic bacteria used in this research. On the other hand, plant pathogenic bacteria are more susceptible than human pathogenic bacteria. CONCLUSION: The results of the study show that cyclotides from Iranian V. odorata have potent antimicrobial activity against gram-negative, plant pathogenic bacteria. SIGNIFICANCE AND IMPACT OF THE STUDY: This study is a part of our extended researches on finding new pharmaceutical potentials of plants and on developing new peptides for special purposes in a way that does not have harmful side effects or have the least side effects.

Differential Antimicrobial Effect of Three-Sized Biogenic Silver Nanoparticles as Broad-Spectrum Antibacterial Agents against Plant Pathogens.

BACKGROUND: Massive fruit losses are caused by microbial pathogens of unknown identities. Therefore, ecofriendly biocontrol measures are well sought after, and biogenic silver nanoparticles are plausible candidates. Here we investigate the antimicrobial effect of three different sized AgNPs samples on those pathogens. METHODOLOGY: Identities of three local pathogenic bacteria were investigated using molecular methods. Three different-sized samples of silver nanoparticles were bio-synthesized in the external solution of a cyanobacterial culture, characterized, and used in antimicrobial bioassay. RESULTS: The pathogens were identified as Erwinia pyrifoliae, Staphylococcus warneri, and Xanthomonas citri. UV-vis. and FTIR spectroscopy confirmed the biosynthesis of AgNPs. and their three different sizes were confirmed using Scanning electron microscopy. Growth of bacterial pathogens was inhibited by all three samples of AgNPs, but the largest inhibition zone was for the smallest sized AgNPs against Staphylococcus warneri (1.7 cm). DISCUSSION: The identity of the pathogens infecting different local fruits is reported for the first time. They belong to different bacterial lineages. The fact that biogenic AAgNPs were effective against all of them shows their broad-spectrum of antibacterial effect. Customized biosynthesis was successful in yielding different-sized AgNPs. The smaller the AgNPs, the stronger the antimicrobial impact. CONCLUSION: Local bacterial species infecting fruits are diverse. Customized biogenic AgNPs are effective broad-spectrum biocontrol agents against bacterial pathogens of local fruits and thereby help maintain food security and environmental sustainability.

Pest categorisation of Pantoea ananatis.

The EFSA Plant Health Panel performed a pest categorisation of Pantoea ananatis, a Gram-negative bacterium belonging to the Erwiniaceae family. P. ananatis is a well-defined taxonomic unit; nonetheless, its pathogenic nature is not well defined and non-pathogenic populations are known to occupy several, very different environmental niches as saprophytes, or as plant growth promoting bacteria or biocontrol agents. It is also described as a clinical pathogen causing bacteraemia and sepsis or as a member of the gut microbiota of several insects. P. ananatis is the causal agent of different diseases affecting numerous crops: in particular, centre rot of onion, bacterial leaf blight and grain discoloration of rice, leaf spot disease of maize and eucalyptus blight/dieback. A few insect species have been described as vectors of P. ananatis, among them, Frankliniella fusca and Diabrotica virgifera virgifera. This bacterium is present in several countries in Europe, Africa, Asia, North and South America, and Oceania from tropical and subtropical regions to temperate areas worldwide. P. ananatis has been reported from the EU territory, both as pathogen on rice and maize and as an environmental, non-pathogenic bacterium in rice marshes and poplar rhizosoil. It is not included in EU Commission Implementing Regulation 2019/2072. The pathogen can be detected on its host plants using direct isolation, or PCR-based methods. The main pathway for the entry of the pathogen into the EU territory is host plants for planting, including seeds. In the EU, there is a large availability of host plants, with onion, maize, rice and strawberry being the most important ones. Therefore, disease outbreaks are possible almost at any latitude, except in the most northern regions. P. ananatis is not expected to have frequent or consistent impact on crop production and is not expected to have any environmental impact. Phytosanitary measures are available to mitigate the further introduction and spread of the pathogen into the EU on some hosts. The pest does not satisfy the criteria, which are within the remit for EFSA to evaluate whether the pest meets the definition of a Union quarantine pest. P. ananatis is probably widely distributed in different ecosystems in the EU. It may impact some specific hosts such as onions while on other hosts such as rice it has been reported as a seed microbiota without causing any impact and can even be beneficial to plant growth. Hence, the pathogenic nature of P. ananatis is not fully established.

Investigation of the antibacterial mechanism of the novel bactericide dioctyldiethylenetriamine (Xinjunan).

BACKGROUND: Chemical control is an important method for tackling crop diseases. Clarifying the antibacterial mechanisms of bactericides is useful for developing new bactericides and for continuous plant disease control. In this study, the antibacterial mechanism of a novel bactericide, dioctyldiethylenetriamine (Xinjunan), which affects adenosine triphosphate (ATP) synthesis, was investigated. RESULTS: The results of an in vitro inhibition activity assay showed that dioctyldiethylenetriamine inhibited the growth of a variety of plant pathogenic bacteria, especially that of Xanthomonas spp. Scanning electron microscopy demonstrated that dioctyldiethylenetriamine caused cell distortion and rupture. To investigate the molecular mechanism underlying the antibacterial effect of dioctyldiethylenetriamine, transcriptome sequencing (RNA-seq) was performed for Xanthomonas oryzae pv. oryzae (Xoo, PXO99A) treated with dioctyldiethylenetriamine, which has strong antibacterial effects against xanthomonads. The results showed that differentially expressed genes were enriched mainly in the oxidative phosphorylation and tricarboxylic acid (TCA) cycle pathways after treatment. Moreover, the dioctyldiethylenetriamine treatment exhibited reduction in enzyme activities in the TCA cycle, decreased intracellular nicotinamide adenine dinucleotide and ATP contents, and increased accumulation of reactive oxygen species. In addition, dioctyldiethylenetriamine exhibited an inhibitory effect on the growth of other bacterial pathogens by reducing ATP synthesis. CONCLUSION: This is the first report of the mechanism by which dioctyldiethylenetriamine inhibits ATP synthesis by affecting oxidative phosphorylation and TCA cycle pathways in bacteria. © 2023 Society of Chemical Industry.

On the use of antibiotics to control plant pathogenic bacteria: a genetic and genomic perspective.

Despite growing attention, antibiotics (such as streptomycin, oxytetracycline or kasugamycin) are still used worldwide for the control of major bacterial plant diseases. This raises concerns on their potential, yet unknown impact on antibiotic and multidrug resistances and the spread of their genetic determinants among bacterial pathogens. Antibiotic resistance genes (ARGs) have been identified in plant pathogenic bacteria (PPB), with streptomycin resistance genes being the most commonly reported. Therefore, the contribution of mobile genetic elements (MGEs) to their spread among PPB, as well as their ability to transfer to other bacteria, need to be further explored. The only well-documented example of ARGs vector in PPB, Tn5393 and its highly similar variants (carrying streptomycin resistance genes), is concerning because of its presence outside PPB, in Salmonella enterica and Klebsiella pneumoniae, two major human pathogens. Although its structure among PPB is still relatively simple, in human- and animal-associated bacteria, Tn5393 has evolved into complex associations with other MGEs and ARGs. This review sheds light on ARGs and MGEs associated with PPB, but also investigates the potential role of antibiotic use in resistance selection in plant-associated bacteria.

Comparative Analysis of Novel Lytic Phages for Biological Control of Phytopathogenic Xanthomonas spp.

Xanthomonas is an important genus of plant-pathogenic bacteria that affects agronomic and economically important crops, causing serious economic losses. In fact, several Xanthomonas species are considered regulated quarantine pests. Due to the lack of effective control measures to treat plant-pathogenic bacteria, innovative control tools are needed to carry out integrated disease management. In this regard, bacteriophages (phages), viruses of bacteria, constitute a promising biocontrol tool. In this work, we report the isolation and characterization of 11 novel Xanthomonas arboricola pv. juglandis phages belonging to different families and genera of the class Caudoviricetes. Infectivity matrix in more than 60 isolates of different xanthomonads and other phytopathogenic bacteria suggests that these phages are specific to the Xanthomonas genus, with different host ranges depending on the isolates tested. Interestingly, some of these phages showed relevant features to be used as biocontrol tools to combat pathogenic Xanthomonas spp. as important as X. oryzae or X. citri. IMPORTANCE Phytopathogenic bacteria represent serious losses worldwide. The lack of current treatments has focused the spotlight on phages, viruses of bacteria, as very promising biocontrol tools. Phages are very specific and can help to control bacterial infections in crops, as is the case of xanthomonads-associated diseases. The discovery of new environmental phages with lytic capacity that can help to combat these pathogens is of special relevance, and it is necessary to implement phage isolation and characterization techniques to determine their host range and their genomic properties. The establishment of phage collections worldwide will allow their use as preventive, diagnostic, or therapeutic tools. Although there is still a long way to go, this work is a step forward in the implementation of new ecofriendly techniques to combat key pathogens in the field.