Genomic evidence for evolutionary history and local adaptation of two endemic apricots: Prunus hongpingensis and P. zhengheensis.

Apricot, belonging to the Armeniaca section of Rosaceae, is one of the economically important crop fruits that has been extensively cultivated. The natural wild apricots offer valuable genetic resources for crop improvement. However, some of them are endemic, with small populations, and are even at risk of extinction. In this study we unveil chromosome-level genome assemblies for two southern China endemic apricots, Prunus hongpingensis (PHP) and P. zhengheensis (PZH). We also characterize their evolutionary history and the genomic basis of their local adaptation using whole-genome resequencing data. Our findings reveal that PHP and PZH are closely related to Prunus armeniaca and form a distinct lineage. Both species experienced a decline in effective population size following the Last Glacial Maximum (LGM), which likely contributed to their current small population sizes. Despite the observed decrease in genetic diversity and heterozygosity, we do not observe an increased accumulation of deleterious mutations in these two endemic apricots. This is likely due to the combined effects of a low inbreeding coefficient and strong purifying selection. Furthermore, we identify a set of genes that have undergone positive selection and are associated with local environmental adaptation in PHP and PZH, respectively. These candidate genes can serve as valuable genetic resources for targeted breeding and improvement of cultivated apricots. Overall, our study not only enriches our comprehension of the evolutionary history of apricot species but also offers crucial insights for the conservation and future breeding of other endemic species amidst rapid climate changes.

Methylomonas defluvii sp. nov., a type I methane-oxidizing bacterium from a secondary sedimentation tank of a wastewater treatment plant.

An aerobic methanotroph was isolated from a secondary sedimentation tank of a wastewater treatment plant and designated strain OY6T. Cells of OY6T were Gram-stain-negative, pink-pigmented, motile rods and contained an intracytoplasmic membrane structure typical of type I methanotrophs. OY6T could grow at a pH range of 4.5-7.5 (optimum pH 6.5) and at temperatures ranging from 20 °C to 37 °C (optimum 30 °C). The major cellular fatty acids were C14 : 0, C16 : 1ω7c/C16 : 1ω6c and C16 : 1ω5c; the predominant respiratory quinone was MQ-8. The genome size was 5.41 Mbp with a DNA G+C content of 51.7 mol%. OY6T represents a member of the family Methylococcaceae of the class Gammaproteobacteria and displayed 95.74-99.64 % 16S rRNA gene sequence similarity to the type strains of species of the genus Methylomonas. Whole-genome comparisons based on average nucleotide identity (ANI) and digital DNA-DNA hybridisation (dDDH) confirmed that OY6T should be classified as representing a novel species. The most closely related type strain was Methylomonas fluvii EbBT, with 16S rRNA gene sequence similarity, ANI by blast (ANIb), ANI by MUMmer (ANIm) and dDDH values of 99.64, 90.46, 91.92 and 44.5 %, respectively. OY6T possessed genes encoding both the particulate methane monooxygenase enzyme and the soluble methane monooxygenase enzyme. It grew only on methane or methanol as carbon sources. On the basis of phenotypic, genetic and phylogenetic data, strain OY6T represents a novel species within the genus Methylomonas for which the name Methylomonas defluvii sp. nov. is proposed, with strain OY6T (=GDMCC 1.4114T=KCTC 8159T=LMG 33371T) as the type strain.

Spread of antibiotic resistance genes in drinking water reservoirs: Insights from a deep metagenomic study using a curated database.

The exploration of antibiotic resistance genes (ARGs) in drinking water reservoirs is an emerging field. Using a curated database, we enhanced the ARG detection and conducted a comprehensive analysis using 2.2 Tb of deep metagenomic sequencing data to determine the distribution of ARGs across 16 drinking water reservoirs and associated environments. Our findings reveal a greater diversity of ARGs in sediments than in water, underscoring the importance of extensive background surveys. Crucial ARG carriers-specifically Acinetobacter, Pseudomonas, and Mycobacterium were identified in drinking water reservoirs. Extensive analysis of the data uncovered a considerable concern for drinking water safety, particularly in regions reliant on river sources. Mobile genetic elements have been found to contribute markedly to the propagation of ARGs. The results of this research suggest that the establishment of drinking water reservoirs for supplying raw water may be an effective strategy for alleviating the spread of water-mediated ARGs.

Constructed Rice Tracers Identify the Major Virulent Transcription Activator-Like Effectors of the Bacterial Leaf Blight Pathogen.

Xanthomonas oryzae pv. oryzae (Xoo) injects major transcription activator-like effectors (TALEs) into plant cells to activate susceptibility (S) genes for promoting bacterial leaf blight in rice. Numerous resistance (R) genes have been used to construct differential cultivars of rice to identify races of Xoo, but the S genes were rarely considered. Different edited lines of rice cv. Kitaake were constructed using CRISPR/Cas9 gene-editing, including single, double and triple edits in the effector-binding elements (EBEs) located in the promoters of rice S genes OsSWEET11a, OsSWEET13 and OsSWEET14. The near-isogenic lines (NILs) were used as tracers to detect major TALEs (PthXo1, PthXo2, PthXo3 and their variants) in 50 Xoo strains. The pathotypes produced on the tracers determined six major TALE types in the 50 Xoo strains. The presence of the major TALEs in Xoo strains was consistent with the expression of S genes in the tracers, and it was also by known genome sequences. The EBE editing had little effect on agronomic traits, which was conducive to balancing yield and resistance. The rice-tracers generated here provide a valuable tool to track major TALEs of Xoo in Asia which then shows what rice cultivars are needed to combat Xoo in the field.

A New Biocontrol Agent Bacillus velezensis SF334 against Rubber Tree Fungal Leaf Anthracnose and Its Genome Analysis of Versatile Plant Probiotic Traits.

Natural rubber is an important national strategic and industrial raw material. The leaf anthracnose of rubber trees caused by the Colletotrichum species is one of the important factors restricting the yields of natural rubber. In this study, we isolated and identified strain Bacillus velezensis SF334, which exhibited significant antagonistic activity against both C. australisinense and C. siamense, the dominant species of Colletotrichum causing rubber tree leaf anthracnose in the Hainan province of China, from a pool of 223 bacterial strains. The cell suspensions of SF334 had a significant prevention effect for the leaf anthracnose of rubber trees, with an efficacy of 79.67% against C. siamense and 71.8% against C. australisinense. We demonstrated that SF334 can lead to the lysis of C. australisinense and C. siamense mycelia by causing mycelial expansion, resulting in mycelial rupture and subsequent death. B. velezensis SF334 also harbors some plant probiotic traits, such as secreting siderophore, protease, cellulase, pectinase, and the auxin of indole-3-acetic acid (IAA), and it has broad-spectrum antifungal activity against some important plant pathogenic fungi. The genome combined with comparative genomic analyses indicated that SF334 possesses most genes of the central metabolic and gene clusters of secondary metabolites in B. velezensis strains. To our knowledge, this is the first time a Bacillus velezensis strain has been reported as a promising biocontrol agent against the leaf anthracnose of rubber trees caused by C. siamense and C. australisinense. The results suggest that B. velezensis could be a potential candidate agent for the leaf anthracnose of rubber trees.

Suppression of host plant defense by bacterial small RNAs packaged in outer membrane vesicles.

Noncoding small RNAs (sRNAs) packaged in bacterial outer membrane vesicles (OMVs) function as novel mediators of interspecies communication. While the role of bacterial sRNAs in enhancing virulence is well established, the role of sRNAs in the interaction between OMVs from phytopathogenic bacteria and their host plants remains unclear. In this study, we employ RNA sequencing to characterize differentially packaged sRNAs in OMVs of the phytopathogen Xanthomonas oryzae pv. oryzicola (Xoc). Our candidate sRNA (Xosr001) was abundant in OMVs and involved in the regulation of OsJMT1 to impair host stomatal immunity. Xoc loads Xosr001 into OMVs, which are specifically ttransferred into the mechanical tissues of rice leaves. Xosr001 suppresses OsJMT1 transcript accumulation in vivo, leading to a reduction in MeJA accumulation in rice leaves. Furthermore, the application of synthesized Xosr001 sRNA to the leaves of OsJMT1-HA-OE transgenic line results in the suppression of OsJMT1 expression by Xosr001. Notably, the OsJMT1-HA-OE transgenic line exhibited attenuated stomatal immunity and disease susceptibility upon infection with ΔXosr001 compared to Xoc. These results suggest that Xosr001 packaged in Xoc OMVs functions to suppress stomatal immunity in rice.

Tal6b/AvrXa27A, a hidden TALE targeting the susceptibility gene OsSWEET11a and the resistance gene Xa27 in rice.

Xanthomonas oryzae pv. oryzae (Xoo) secretes transcription activator-like effectors (TALEs) to activate rice susceptibility (S) genes, causing bacterial blight (BB), as well as resistance (R) genes, leading to defense against BB. This activation follows a gene-for-gene paradigm that results in an arms race between the TALE of the pathogen and effector-binding elements (EBEs) in the promoters of host genes. In this study, we characterized a novel TALE, designated Tal6b/AvrXa27A, that activates the rice S gene OsSWEET11a and the rice R gene Xa27. Tal6b/AvrXa27A is a member of the AvrXa27/TalAO class and contains 16 repeat variable diresidues (RVDs); one RVD is altered and one is deleted in Tal6b/AvrXa27A compared with AvrXa27, a known avirulence (avr) effector of Xa27. Tal6b/AvrXa27A can transcriptionally activate the expression of Xa27 and OsSWEET11a via EBEs in their corresponding promoters, leading to effector-triggered immunity and susceptibility, respectively. The 16 RVDs in Tal6b/AvrXa27A have no obvious similarity to the 24 RVDs in the effector PthXo1, but EBETal6b and EBEPthXo1 are overlapped in the OsSWEET11a promoter. Tal6b/AvrXa27A is prevalent among Asian Xoo isolates, but PthXo1 has only been reported in the Philippine strain PXO99A. Genome editing of EBETal6b in the OsSWEET11a promoter further confirmed the requirement for OsSWEET11a expression in Tal6b/AvrXa27A-dependent susceptibility to Xoo. Moreover, Tal6b/AvrXa27A resulted in higher transcription of Xa27 than of OsSWEET11a, which led to a strong, rapid resistance response that blocked disease development. These findings suggest that Tal6b/AvrXa27A has a dual function: triggering resistance by activating Xa27 gene expression as an avirulence factor and inducing transcription of the S gene OsSWEET11a, resulting in virulence. Intriguingly, Tal6b/AvrXa27A, but not AvrXa27, can bind to the promoter of OsSWEET11a. The underlying recognition mechanism for this binding remains unclear but appears to deviate from the currently accepted TALE code.

Comparative Transcriptomic Analysis of Wheat Cultivars in Response to Xanthomonas translucens pv. cerealis and Its T2SS, T3SS, and TALEs Deficient Strains.

Xanthomonas translucens pv. cerealis causes bacterial leaf streak disease on small grain cereals. Type II and III secretion systems (T2SS and T3SS) play a pivotal role in the pathogenicity of the bacterium, while no data are available on the transcriptomic profile of wheat cultivars infected with either wild type (WT) or mutants of the pathogen. In this study, WT, TAL-effector mutants, and T2SS/T3SS mutants of X. translucens pv. cerealis strain NXtc01 were evaluated for their effect on the transcriptomic profile of two wheat cultivars, 'Chinese Spring' and 'Yangmai-158', using Illumina RNA-sequencing technology. RNA-Seq data showed that the number of differentially expressed genes (DEGs) was higher in Yangmai-158 than in Chinese Spring, suggesting higher susceptibility of Yangmai-158 to the pathogen. In T2SS, most suppressed DEGs were related to transferase, synthase, oxidase, WRKY, and bHLH transcription factors. The gspD mutants showed significantly decreased disease development in wheat, suggesting an active contribution of T2SS in virulence. Moreover, the gspD mutant restored full virulence and its multiplication in planta by addition of gspD in trans. In the T3SS-deficient strain, downregulated DEGs were associated with cytochrome, peroxidases, kinases, phosphatases, WRKY, and ethylene-responsive transcription factors. In contrast, upregulated DEGs were trypsin inhibitors, cell number regulators, and calcium transporter. Transcriptomic analyses coupled with quantitative real-time-PCR indicated that some genes are upregulated in Δtal1/Δtal2 compared with the tal-free strain, but no direct interaction was observed. These results provide novel insight into wheat transcriptomes in response to X. translucens infection and pave the way for understanding host-pathogen interactions.

Antagonistic transcriptome profile reveals potential mechanisms of action on Xanthomonas oryzae pv. oryzicola by the cell-free supernatants of Bacillus velezensis 504, a versatile plant probiotic bacterium.

Bacterial leaf streak (BLS) of rice is a severe disease caused by the bacterial pathogen Xanthomonas oryzae pv. oryzicola (Xoc) that has gradually become the fourth major disease on rice in some rice-growing regions in southern China. Previously, we isolated a Bacillus velezensis strain 504 that exhibited apparent antagonistic activity against the Xoc wild-type strain RS105, and found that B. velezensis 504 was a potential biocontrol agent for BLS. However, the underlying mechanisms of antagonism and biocontrol are not completely understood. Here we mine the genomic data of B. velezensis 504, and the comparative transcriptomic data of Xoc RS105 treated by the cell-free supernatants (CFSs) of B. velezensis 504 to define differentially expressed genes (DEGs). We show that B. velezensis 504 shares over 89% conserved genes with FZB42 and SQR9, two representative model strains of B. velezensis, but 504 is more closely related to FZB42 than SQR9, as well as B. velezensis 504 possesses the secondary metabolite gene clusters encoding the essential anti-Xoc agents difficidin and bacilysin. We conclude that approximately 77% of Xoc RS105 coding sequences are differentially expressed by the CFSs of B. velezensis 504, which significantly downregulates genes involved in signal transduction, oxidative phosphorylation, transmembrane transport, cell motility, cell division, DNA translation, and five physiological metabolisms, as well as depresses an additional set of virulence-associated genes encoding the type III secretion, type II secretion system, type VI secretion system, type IV pilus, lipopolysaccharides and exopolysaccharides. We also show that B. velezensis 504 is a potential biocontrol agent for bacterial blight of rice exhibiting relative control efficiencies over 70% on two susceptible cultivars, and can efficiently antagonize against some important plant pathogenic fungi including Colletotrichum siamense and C. australisinense that are thought to be the two dominant pathogenic species causing leaf anthracnose of rubber tree in Hainan province of China. B. velezensis 504 also harbors some characteristics of plant growth-promoting rhizobacterium such as secreting protease and siderophore, and stimulating plant growth. This study reveals the potential biocontrol mechanisms of B. velezensis against BLS, and also suggests that B. velezensis 504 is a versatile plant probiotic bacterium.

Detection and Diagnosis of Bacterial Leaf Streak on Small Grain Cereals: From Laboratory to Field.

Bacterial leaf streak of small-grain cereals is an economically important disease of wheat and barley crops. The disease occurs in many countries across the globe, with particular importance in regions characterized by high precipitation or areas in which sprinkler irrigation is used. Three genetically distinct lineages of the Gram-negative bacterium Xanthomonas translucens (X. translucens pv. undulosa, X. translucens pv. translucens, and X. translucens pv. cerealis) are responsible for most of the bacterial leaf streak infections on wheat and barley crops. Considering the seedborne nature of the pathogens, they are included in the A2 (high-risk) list of quarantine organisms for some European countries; hence, they are under strict quarantine control and zero tolerance. Due to the taxonomic complexities within X. translucens, the exact geographic distribution of each pathovar has not yet been determined. In this mini review, we provide an updated overview of the detection and diagnosis of the bacterial leaf streak pathogens. First, a short history of the leaf streak pathogens is provided, followed by the symptomology and host range of the causal agents. Then, the utility of conventional methods and high-throughput molecular approaches in the precise detection and identification of the pathogens is explained. Finally, we highlight the role of quarantine inspections and early detection of the pathogen in combating the risk of bacterial leaf streak in the 21st century's small-grains cereals' industry.

The occurrence, characteristics, and adaptation of A-to-I RNA editing in bacteria: A review.

A-to-I RNA editing is a very important post-transcriptional modification or co-transcriptional modification that creates isoforms and increases the diversity of proteins. In this process, adenosine (A) in RNA molecules is hydrolyzed and deaminated into inosine (I). It is well known that ADAR (adenosine deaminase acting on RNA)-dependent A-to-I mRNA editing is widespread in animals. Next, the discovery of A-to-I mRNA editing was mediated by TadA (tRNA-specific adenosine deaminase) in Escherichia coli which is ADAR-independent event. Previously, the editing event S128P on the flagellar structural protein FliC enhanced the bacterial tolerance to oxidative stress in Xoc. In addition, the editing events T408A on the enterobactin iron receptor protein XfeA act as switches by controlling the uptake of Fe3+ in response to the concentration of iron in the environment. Even though bacteria have fewer editing events, the great majority of those that are currently preserved have adaptive benefits. Interestingly, it was found that a TadA-independent A-to-I RNA editing event T408A occurred on xfeA, indicating that there may be other new enzymes that perform a function like TadA. Here, we review recent advances in the characteristics, functions, and adaptations of editing in bacteria.

The natural pyrazolotriazine pseudoiodinine from Pseudomonas mosselii 923 inhibits plant bacterial and fungal pathogens.

Natural products largely produced by Pseudomonads-like soil-dwelling microorganisms are a consistent source of antimicrobial metabolites and pesticides. Herein we report the isolation of Pseudomonas mosselii strain 923 from rice rhizosphere soils of paddy fields, which specifically inhibit the growth of plant bacterial pathogens Xanthomonas species and the fungal pathogen Magnaporthe oryzae. The antimicrobial compound is purified and identified as pseudoiodinine using high-resolution mass spectra, nuclear magnetic resonance and single-crystal X-ray diffraction. Genome-wide random mutagenesis, transcriptome analysis and biochemical assays define the pseudoiodinine biosynthetic cluster as psdABCDEFG. Pseudoiodinine biosynthesis is proposed to initiate from guanosine triphosphate and 1,6-didesmethyltoxoflavin is a biosynthetic intermediate. Transposon mutagenesis indicate that GacA is the global regulator. Furthermore, two noncoding small RNAs, rsmY and rsmZ, positively regulate pseudoiodinine transcription, and the carbon storage regulators CsrA2 and CsrA3, which negatively regulate the expression of psdA. A 22.4-fold increase in pseudoiodinine production is achieved by optimizing the media used for fermentation, overexpressing the biosynthetic operon, and removing the CsrA binding sites. Both of the strain 923 and purified pseudoiodinine in planta inhibit the pathogens without affecting the rice host, suggesting that pseudoiodinine can be used to control plant diseases.

CRISPR/FnCas12a-mediated efficient multiplex and iterative genome editing in bacterial plant pathogens without donor DNA templates.

CRISPR-based genome editing technology is revolutionizing prokaryotic research, but it has been rarely studied in bacterial plant pathogens. Here, we have developed a targeted genome editing method with no requirement of donor templates for convenient and efficient gene knockout in Xanthomonas oryzae pv. oryzae (Xoo), one of the most important bacterial pathogens on rice, by employing the heterologous CRISPR/Cas12a from Francisella novicida and NHEJ proteins from Mycobacterium tuberculosis. FnCas12a nuclease generated both small and large DNA deletions at the target sites as well as it enabled multiplex genome editing, gene cluster deletion, and plasmid curing in the Xoo PXO99A strain. Accordingly, a non-TAL effector-free polymutant strain PXO99AD25E, which lacks all 25 xop genes involved in Xoo pathogenesis, has been engineered through iterative genome editing. Whole-genome sequencing analysis indicated that FnCas12a did not have a noticeable off-target effect. In addition, we revealed that these strategies are also suitable for targeted genome editing in another bacterial plant pathogen Pseudomonas syringae pv. tomato (Pst). We believe that our bacterial genome editing method will greatly expand the CRISPR study on microorganisms and advance our understanding of the physiology and pathogenesis of Xoo.

A varied AvrXa23-like TALE enables the bacterial blight pathogen to avoid being trapped by Xa23 resistance gene in rice.

INTRODUCTION: Xa23 as an executor mediates broad-spectrum resistance to Xanthomonas oryzae pv. oryzae (Xoo), which contains a matching avirulence gene avrXa23, in rice for bacterial leaf blight (BLB). avrXa23 encodes a transcription activator-like effector (TALE) protein which binds to the EBE (effector-binding element) of the Xa23 promoter. It is unclear whether the considerable pressure of Xa23 leads to an emerging Xoo strain that overcomes Xa23 resistance. OBJECTIVES: This study aimed to uncover new Xoo isolate(s) that overcome Xa23-mediated resistance and to investigate how the pathogen evades the resistance. METHODS: Totally 185 Xoo isolates were used to screen possibly compatible strain(s) with Xa23-containing rice CBB23 by pathogenicity test. Genome Sequencing, Southern blot, tal gene cloning, Western blot, qRT-PCR and electrophoretic mobility shift assays (EMSA) were conducted to determine the mechanism of one Xoo isolate being compatible with Xa23-containing rice. RESULTS: One isolate AH28 from Anhui province is compatible with CBB23. AH28 strain contains an ortholog of avrXa23, tal7b and has 17 tal genes. The 4th RVD (repeat-variable diresidue) in Tal7b are missed and the 5th and 8th RVDs changed from NG and NS to NS and S*, respectively. These alternations made Tal7b unable to bind to the EBE of Xa23 promoter to activate the expression of Xa23 in rice. The ectopic expression of tal7b in a tal-free mutant PH of PXO99A did not alter the virulence of the strain PH, whereas avrXa23 made AH28 from compatibility to incompatibility with Xa23 rice. CONCLUSION: Best to our knowledge, this is the first insight of a naturally-emerging Xoo isolate that overcomes the broad-spectrum resistance of Xa23 by the variable AvrXa23-like TALE Tal7b. The RVD alteration in AvrXa23 may be a common strategy for the pathogen evolution to avoid being "trapped" by the executor R gene.

Next Generation Sequencing and Comparative Genomic Analysis Reveal Extreme Plasticity of Two Burkholderia glumae Strains HN1 and HN2.

Burkholderia glumae is an important rice pathogen, thus the genomic and evolutionary history may be helpful to control this notorious pathogen. Here, we present two complete genomes of the B. glumae strains HN1 and HN2, which were isolated from diseased rice seed in China. Average nucleotide identity (ANI) analysis shows greater than 99% similarity of the strains HN1 and HN2 with other published B. glumae genomes. Genomic annotation revealed that the genome of strain HN1 consists of five replicons (6,680,415 bp) with an overall G + C content of 68.06%, whereas the genome of strain HN2 comprises of three replicons (6,560,085 bp) with an overall G + C content of 68.34%. The genome of HN1 contains 5434 protein-coding genes, 351 pseudogenes, and 1 CRISPR, whereas the genome of HN2 encodes 5278 protein-coding genes, 357 pseudogenes, and 2 CRISPR. Both strains encode many pathogenic-associated genes (143 genes in HN1 vs. 141 genes in HN2). Moreover, comparative genomic analysis shows the extreme plasticity of B. glumae, which may contribute to its pathogenicity. In total, 259 single-copy genes were affected by positive selection. These genes may contribute to the adaption to different environments. Notably, six genes were characterized as virulence factors which may be an additional way to assist the pathogenicity of B. glumae.

The Xanthomonas type III effector XopAP prevents stomatal closure by interfering with vacuolar acidification.

Plant stomata close rapidly in response to a rise in the plant hormone abscisic acid (ABA) or salicylic acid (SA) and after recognition of pathogen-associated molecular patterns (PAMPs). Stomatal closure is the result of vacuolar convolution, ion efflux, and changes in turgor pressure in guard cells. Phytopathogenic bacteria secrete type III effectors (T3Es) that interfere with plant defense mechanisms, causing severe plant disease symptoms. Here, we show that the virulence and infection of Xanthomonas oryzae pv. oryzicola (Xoc), which is the causal agent of rice bacterial leaf streak disease, drastically increased in transgenic rice (Oryza sativa L.) plants overexpressing the Xoc T3E gene XopAP, which encodes a protein annotated as a lipase. We discovered that XopAP binds to phosphatidylinositol 3,5-bisphosphate (PtdIns(3,5)P2 ), a membrane phospholipid that functions in pH control in lysosomes, membrane dynamics, and protein trafficking. XopAP inhibited the acidification of vacuoles by competing with vacuolar H+ -pyrophosphatase (V-PPase) for binding to PtdIns(3,5)P2 , leading to stomatal opening. Transgenic rice overexpressing XopAP also showed inhibition of stomatal closure when challenged by Xoc infection and treatment with the PAMP flg22. Moreover, XopAP suppressed flg22-induced gene expression, reactive oxygen species burst and callose deposition in host plants, demonstrating that XopAP subverts PAMP-triggered immunity during Xoc infection. Taken together, these findings demonstrate that XopAP overcomes stomatal immunity in plants by binding to lipids.

The MinCDE Cell Division System Participates in the Regulation of Type III Secretion System (T3SS) Genes, Bacterial Virulence, and Motility in Xanthomonas oryzae pv. oryzae.

Xanthomonas oryzae pv. oryzae (Xoo) causes bacterial leaf blight (BLB) in rice, which is one of the most severe bacterial diseases in rice in some Asian countries. The type III secretion system (T3SS) of Xoo encoded by the hypersensitive response and pathogenicity (hrp) genes is essential for its pathogenicity in host rice. Here, we identified the Min system (MinC, MinD, and MinE), a negative regulatory system for bacterial cell division encoded by minC, minD, and minE genes, which is involved in negative regulation of hrp genes (hrpB1 and hrpF) in Xoo. We found that the deletion of minC, minD, and minCDE resulted in enhanced hrpB1 and hrpF expression, which is dependent on two key hrp regulators HrpG and HrpX. The minC, minD, and minCDE mutants exhibited elongated cell lengths, and the classic Min system-defective cell morphology including minicells and short filamentations. Mutation of minC in Xoo resulted in significantly impaired virulence in host rice, swimming motility, and enhanced biofilm formation. Our transcriptome profiling also indicated some virulence genes were differentially expressed in the minC mutants. To our knowledge, this is the first report about the Min system participating in the regulation of T3SS expression. It sheds light on the understanding of Xoo virulence mechanisms.

Xanthomonas oryzae Pv. oryzicola Response Regulator VemR Is Co-opted by the Sensor Kinase CheA for Phosphorylation of Multiple Pathogenicity-Related Targets.

Two-component systems (TCSs) (cognate sensor histidine kinase/response regulator pair, HK/RR) play a crucial role in bacterial adaptation, survival, and productive colonization. An atypical orphan single-domain RR VemR was characterized by the non-vascular pathogen Xanthomonas oryzae pv. oryzicola (Xoc) is known to cause bacterial leaf streak (BLS) disease in rice. Xoc growth and pathogenicity in rice, motility, biosynthesis of extracellular polysaccharide (EPS), and the ability to trigger HR in non-host tobacco were severely compromised in the deletion mutant strain RΔvemR as compared to the wild-type strain RS105. Site-directed mutagenesis and phosphotransfer experiments revealed that the conserved aspartate (D56) residue within the stand-alone phosphoacceptor receiver (REC) domain is essential for phosphorelay and the regulatory activity of Xoc VemR. Yeast two-hybrid (Y2H) and co-immunoprecipitation (co-IP) data identified CheA as the HK co-opting the RR VemR for phosphorylation. Affinity proteomics identified several downstream VemR-interacting proteins, such as 2-oxoglutarate dehydrogenase (OGDH), DNA-binding RR SirA, flagellar basal body P-ring formation protein FlgA, Type 4a pilus retraction ATPase PilT, stress-inducible sensor HK BaeS, septum site-determining protein MinD, cytoskeletal protein CcmA, and Type III and VI secretion system proteins HrpG and Hcp, respectively. Y2H and deletion mutant analyses corroborated that VemR interacted with OGDH, SirA, FlgA, and HrpG; thus, implicating multi-layered control of diverse cellular processes including carbon metabolism, motility, and pathogenicity in the rice. Physical interaction between VemR and HrpG suggested cross-talk interaction between CheA/VemR- and HpaS/HrpG-mediated signal transduction events orchestrating the hrp gene expression.