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Abstract Salicylic acid (SA) plays an essential role in plant defense against biotrophic and semi‐biotrophic pathogens. Following pathogen recognition, SA biosynthesis dramatically increases at the infection site of the host plant. The manner in which pathogens sense and tolerate the onslaught of SA stress to survive in the plant following infection remains to be understood. The objective of this work was to determine how the model phytopathogen Xanthomonas campestris pv. campestris (Xcc) senses and effluxes SA during infection inside host plants. First, RNA‐Seq analysis identified an SA‐responsive operon Xcc4167–Xcc4171, encoding a MarR family transcription factor HepR and an RND (resistance‐nodulation‐cell division) family efflux pump HepABCD in Xcc. Electrophoretic mobility shift assays and DNase I footprint analysis revealed that HepR negatively regulated hepABCD expression by specifically binding to an AT‐rich region of the promoter of the hepRABCD operon, Phep. Second, isothermal titration calorimetry and further genetic analysis suggest that HepR is a novel SA sensor. SA binding released HepR from its cognate promoter Phep and then induced the expression of hepABCD. Third, the RND family efflux pump HepABCD was responsible for SA efflux. The hepRABCD cluster was also involved in the regulation of culture pH and quorum sensing signal diffusible signaling factor turnover. Finally, the hepRABCD cluster was transcribed during the XC1 infection of Chinese radish and was required for the full virulence of Xcc in Chinese radish and cabbage. These findings suggest that the ability of Xcc to co‐opt the plant defense signal SA to activate the multidrug efflux pump may have evolved to ensure Xcc survival and virulence in susceptible host plants.

Abstract Xanthomonas campestris pv. campestris (Xcc) is the causal agent of black rot in Brassica vegetables, which can induce the host plant to produce salicylic acid and 4-hydroxybenzoic acid (4-HBA) during infection. Xcc was previously shown to sense and degrade host plant-derived 4-HBA via the sensor PobR and a PobA-dependent pathway. The degradation of 4-HBA is associated with Xcc virulence in cabbage. The present study generated a reporter strain XC1::PpobA-gusA to monitor pobA transcription. 4-HBA-like compounds were screened for their ability to interfere with pobA transcription. Benzoic acid (BA) was found to efficiently decrease pobA transcription in a dose-dependent manner. Xcc neither produced nor degraded BA; however, the exogenous addition of BA to the 4-HBA-containing Xcc culture significantly decreased the 4-HBA degradation rate. Furthermore, addition of BA into the Xcc culture did not significantly affect the transcription of pobA or pobR; however, addition of BA into the 4-HBA-containing culture significantly decreased the transcription of both genes. Isothermal titration calorimetry and an electrophoretic mobility shift assay revealed that BA binds to PobR with a moderate affinity, which interfered with the binding of 4-HBA/PobR complex to the pobA promoter and thereby inhibiting pobA transcription and 4-HBA degradation. The endogenous BA level of the infected cabbage leaves increased in response to Xcc infection. In the presence of BA, the virulence of Xcc on cabbage decreased significantly. Taken together, these results suggest that cabbage utilizes BA to interfere with 4-HBA degradation, thereby reducing Xcc virulence. Thus, BA has the potential to be developed as a bactericide against Xcc infection.

Abstract Pradimicin U is a new dihydrobenzo[a]naphthacenequinone compound found to be active on a screen designed to investigate compounds with antimicrobial activity, produced by the actinomycete designated strain FMUSA5-5T. The strain was isolated from a bio-fertilizer of Musa spp. collected from Suphanburi province, Thailand. The chemotaxonomic characteristics and 16S rRNA gene analysis revealed that strain FMUSA5-5T is a member of the genus Nonomuraea. Low genome-based taxonomic criteria, average nucleotide identity (ANI) (82.8–88.3%), average amino-acid identity (AAI) (79.4–87.3%), and digital DNA–DNA hybridization (dDDH) (29.5–38.5%) values and several phenotypic differences between strain FMUSA5-5T and its closest type strains of the genus Nonomuraea indicated that strain FMUSA5-5T represents a novel species of the genus Nonomuraea and the name Nonomuraea composti sp. nov. is proposed for the strain. The crude extract from the culture broth of strain FMUSA5-5T displayed promising antimicrobial activity against several pathogens and led to the isolation of a novel secondary metabolite, pradimicin U. Interestingly, this compound displayed a broad spectrum of biological activities such as antimalarial activity against Plasmodium falciparum K1 (IC50 value = 3.65 µg/mL), anti-Mycobacterium tuberculosis H37Ra (MIC value = 25.0 µg/mL), anti-Alternaria brassicicola BCC 42724 (MIC value = 25.0 µg/mL), anti-Bacillus cereus ATCC 11778 and anti-Staphylococcus aureus ATCC 29213 (MIC values = 6.25 and 1.56 µg/mL, respectively). Moreover, the compound possessed strong anti-human small cell lung cancer (NCI-H187) activity with IC50 value of 5.69 µg/mL, while cytotoxicity against human breast cancer (MCF-7) and Vero cells was very weak (IC50 values of 52.49 and 21.84 µg/mL, respectively).

Abstract Pseudomonas aeruginosa is a ubiquitous and metabolically versatile microorganism naturally found in soil and water. It is also an opportunistic pathogen in plants, insects, animals, and humans. In response to increasing cell density, P. aeruginosa uses two acyl‐homoserine lactone (AHL) quorum‐sensing (QS) signals (i.e., N‐3‐oxo‐dodecanoyl homoserine lactone [3‐oxo‐C12‐HSL] and N‐butanoyl‐homoserine lactone [C4‐HSL]), which regulate the expression of hundreds of genes. However, how the biosynthesis of these two QS signals is coordinated remains unknown. We studied the regulation of these two QS signals in the rhizosphere strain PA1201. PA1201 sequentially produced 3‐oxo‐C12‐HSL and C4‐HSL at the early and late growth stages, respectively. The highest 3‐oxo‐C12‐HSL‐dependent elastase activity was observed at the early stage, while the highest C4‐HSL‐dependent rhamnolipid production was observed at the late stage. The atypical regulator RsaL played a pivotal role in coordinating 3‐oxo‐C12‐HSL and C4‐HSL biosynthesis and QS‐associated virulence. RsaL repressed lasI transcription by binding the –10 and –35 boxes of the lasI promoter. In contrast, RsaL activated rhlI transcription by binding the region encoding the 5′‐untranslated region of the rhlI mRNA. Further, RsaL repressed its own expression by binding a nucleotide motif located in the –35 box of the rsaL promoter. Thus, RsaL acts as a molecular switch that coordinates the sequential biosynthesis of AHL QS signals and differential virulence in PA1201. Finally, C4‐HSL activation by RsaL was independent of the Las and Pseudomonas quinolone signal (PQS) QS signaling systems. Therefore, we propose a new model of the QS regulatory network in PA1201, in which RsaL represents a superior player acting at the top of the hierarchy.

Biocontrol strain Pseudomonas PA1201 produces pyoluteorin (Plt), which is an antimicrobial secondary metabolite. Plt represents a promising candidate pesticide due to its broad-spectrum antifungal and antibacterial activity. Although PA1201 contains a complete genetic cluster for Plt biosynthesis, it fails to produce detectable level of Plt when grown in media typically used for Pseudomonas strains. In this study, minimum medium (MM) was found to favor Plt biosynthesis. Using the medium M, which contains all the salts of MM medium except for mannitol, as a basal medium, we compared 10 carbon sources for their ability to promote Plt biosynthesis. Fructose, mannitol, and glycerol promoted Plt biosynthesis, with fructose being the most effective carbon source. Glucose or succinic acid had no significant effect on Plt biosynthesis, but effectively antagonized fructose-dependent synthesis of Plt. Promoter-lacZ fusion reporter strains demonstrated that fructose acted through activation of the pltLABCDEFG (pltL) operon but had no effect on other genes of plt gene cluster; glucose or succinic acid antagonized fructose-dependent pltL induction. Mechanistically, fructose-mediated Plt synthesis involved carbon catabolism repression. The two-component system CbrA/CbrB and small RNA catabolite repression control Z (crcZ) were essential for fructose-induced Plt synthesis. The small RNA binding protein Hfq and Crc negatively regulated fructose-induced Plt. Taken together, this study provides a new model of fructose-dependent Plt production in PA1201 that can help improve Plt yield by biosynthetic approaches.

Abstract Previous studies have demonstrated that bis-(3',5')-cyclic diguanosine monophosphate (bis-3',5'-c-di-GMP) is a ubiquitous second messenger employed by bacteria. Here, we report that 2',3'-cyclic guanosine monophosphate (2',3'-cGMP) controls the important biological functions, quorum sensing (QS) signaling systems and virulence in Ralstonia solanacearum through the transcriptional regulator RSp0980. This signal specifically binds to RSp0980 with high affinity and thus abolishes the interaction between RSp0980 and the promoters of target genes. In-frame deletion of RSp0334, which contains an evolved GGDEF domain with a LLARLGGDQF motif required to catalyze 2',3'-cGMP to (2',5')(3',5')-cyclic diguanosine monophosphate (2',3'-c-di-GMP), altered the abovementioned important phenotypes through increasing the intracellular 2',3'-cGMP levels. Furthermore, we found that 2',3'-cGMP, its receptor and the evolved GGDEF domain with a LLARLGGDEF motif also exist in the human pathogen Salmonella typhimurium. Together, our work provides insights into the unusual function of the GGDEF domain of RSp0334 and the special regulatory mechanism of 2',3'-cGMP signal in bacteria.

Abstract Tirandamycin (TAM B) is a tetramic acid antibiotic discovered to be active on a screen designed to find compounds with neuroprotective activity. The producing strain, SBST2-5T, is an actinobacterium that was isolated from wastewater treatment bio–sludge compost collected from Suphanburi province, Thailand. Taxonomic characterization based on a polyphasic approach indicates that strain SBST2-5T is a member of the genus Streptomyces and shows low average nucleotide identity (ANI) (81.7%), average amino-acid identity (AAI) (78.5%), and digital DNA-DNA hybridization (dDDH) (25.9%) values to its closest relative, Streptomyces thermoviolaceus NBRC 13905T, values that are significantly below the suggested cut-off values for the species delineation, indicating that strain SBST2-5T could be considered to represent a novel species of the genus Streptomyces. The analysis of secondary metabolites biosynthetic gene clusters (smBGCs) in its genome and chemical investigation led to the isolation of TAM B. Interestingly, TAM B at 20 µg/mL displayed a suppressive effect on beta-secretase 1 (BACE1) with 68.69 ± 8.84% inhibition. Molecular docking simulation reveals the interaction mechanism between TAM B and BACE1 that TAM B was buried in the pocket of BACE-1 by interacting with amino acids Thr231, Asp 228, Gln73, Lys 107 via hydrogen bond and Leu30, Tyr71, Phe108, Ile118 via hydrophobic interaction, indicating that TAM B represents a potential active BACE1 inhibitor. Moreover, TAM B can protect the neuron cells significantly (% neuron viability = 83.10 ± 9.83% and 112.72 ± 6.83%) from oxidative stress induced by serum deprivation and Aβ1–42 administration models at 1 ng/mL, respectively, without neurotoxicity on murine P19-derived neuron cells nor cytotoxicity against Vero cells. This study was reportedly the first study to show the neuroprotective and BACE1 inhibitory activities of TAM B.

Abstract Xanthomonas campestris pv. campestris (Xcc) is the causal agent of black rot of cruciferous plants, which harbor high levels of hydroxycinnamic acids (HCAs) in their above-ground parts. Thus, upon infection of the host plant, the pathogen experiences a complex cocktail of HCAs. The present study shows that Xcc can efficiently degrade the HCAs, 4-hydroxycinnamic acid (4-HCA), ferulic acid (FA) and sinapic acid (SiA), via an hca cluster which encodes putative genes for a 4-hydroxycinnamoyl-CoA synthetase/4-HCA ligase HcaL, a benzaldehyde dehydrogenase HcaD, a 4-hydroxycinnamoyl-CoA hydratase/lyase HcaH and a member of the MarR-family of transcriptional factors, HcaR. Xcc also degrades the HCA caffeic acid, but with an alternative mechanism. RT-PCR and subsequent GUS assays show that the hca cluster is transcribed within a single operon, and its transcription is specifically induced by 4-HCA, FA and SiA. Furthermore, we show that HcaR negatively regulates hca transcription when its ligand, the proposed degradation pathway intermediate HCA-CoA, is not present. HcaR specifically binds to a 25-bp site, which encompasses the -10 elements of the hca promoter. Finally, GUS histochemical staining and subsequent quantitative analysis shows that the hca cluster is transcribed in planta during pathogenesis of Chinese radish, and hca deletion mutant strains exhibit compromised virulence in cabbage. Together, these results suggest that the ability to degrade HCAs contributes to Xcc virulence by facilitating its growth and spread, and by protecting the pathogen from HCA toxicity. A working model to explain Xcc HCA sensing and subsequent induction of the HCA degradation process is proposed.

Histone-like nucleoid-structuring (H-NS) proteins are key regulators in gene expression silencing and in nucleoid compaction. The H-NS family member proteins MvaU in Pseudomonas aeruginosa are thought to bind the same AT-rich regions of chromosomes and function to coordinate the control of a common set of genes. Here, we explored the molecular mechanism by which MvaU controls PCA biosynthesis in P. aeruginosa PA1201. We present evidence suggesting that MvaU is self-regulated. Deletion of mvaU significantly increased PCA production, and PCA production sharply decreased when mvaU was over-expressed. MvaU transcriptionally repressed phz2 cluster expression and consequently reduced PCA biosynthesis. β-galactosidase assays confirmed that base pairing near the −35 box is required when MvaU regulates PCA production in PA1201. Electrophoretic mobility shift assays (EMSA) and additional point mutation analysis demonstrated that MvaU directly bound to an AT-rich motif within the promoter of the phz2 cluster. Chromatin immunoprecipitation (ChIP) analysis also indicated that MvaU directly bound to the P5 region of the phz2 cluster promoter. MvaU repression of PCA biosynthesis was independent of QscR and OxyR in PA1201 and neither PCA or H2O2 were the environmental signals that induced mvaU expression. These findings detail a new MvaU-dependent regulatory pathway of PCA biosynthesis in PA1201 and provide a foundation to increase PCA fermentation titer by genetic engineering.

ABSTRACT Plant colonization by phytopathogens is a very complex process in which numerous factors are involved. Upon infection by phytopathogens, plants produce salicylic acid (SA) that triggers gene expression within the plant to counter the invading pathogens. The present study demonstrated that SA signal also directly acts on the quorum-sensing (QS) system of the invading pathogen Xanthomonas campestris pv. campestris to affect its virulence by inducing turnover of the diffusible signaling factor (DSF) family QS signal. First, Xanthomonas campestris pv. campestris infection induces SA biosynthesis in the cabbage host plant. SA cannot be degraded by Xanthomonas campestris pv. campestris during culturing. Exogenous addition of SA or endogenous production of SA induces DSF signal turnover during late growth phase of Xanthomonas campestris pv. campestris in XYS medium that mimics plant vascular environments. Further, the DSF turnover gene rpfB is required for SA induction of DSF turnover. However, SA does not affect the expression of rpfB and DSF biosynthesis gene rpfF at the transcriptional level. SA induction of DSF turnover only occurs under acidic conditions in XYS medium. Furthermore, addition of SA to XYS medium significantly increased both culture and cytoplasmic pH. Increased cytoplasmic pH induced DSF turnover in a rpfB-dependent manner. In vitro RpfB-dependent DSF turnover activity increased when pH increased from 6 to 8. SA exposure did not affect the RpfB-dependent DSF turnover in vitro. Finally, SA-treated Xanthomonas campestris pv. campestris strain exhibited enhanced virulence when inoculated on cabbage. These results provide new insight into the roles of SA in host plants and the molecular interactions between Xanthomonas campestris pv. campestris and cruciferous plants. IMPORTANCE SA is a phenolic acid plant hormone that plays an essential role in plant defenses against biotrophic and semibiotrophic pathogens. Substantial progress has been made in understanding the pivotal role of SA in plant immunity. However, the roles of SA in inhibiting invading plant pathogens and the associated underlying molecular mechanisms are not yet fully understood. The present study demonstrated that the SA signal directly acts on the quorum-sensing (QS) system of the invading pathogen Xanthomonas campestris pv. campestris to affect its virulence by inducing turnover of the DSF family QS signal via a pH-dependent manner. These findings provide new insight into the roles of SA and expand our understanding of the molecular interactions between pathogens and plant hosts.

Evidence has shown that gut microbiome plays a role in modulating the development of diseases beyond the gastrointestinal tract, including skin disorders such as psoriasis. The gut–skin axis refers to the bidirectional relationship between the gut microbiome and skin health. This is regulated through several mechanisms such as inflammatory mediators and the immune system. Dysregulation of microbiota has been seen in numerous inflammatory skin conditions such as atopic dermatitis, rosacea, and psoriasis. Understanding how gut microbiome are involved in regulating skin health may lead to development of novel therapies for these skin disorders through microbiome modulation, in particularly psoriasis. In this review, we will compare the microbiota between psoriasis patients and healthy control, explain the concept of gut–skin axis and the effects of gut dysbiosis on skin physiology. We will also review the current evidence on modulating gut microbiome using probiotics in psoriasis.

Abstract Background Probiotics could prevent Pseudomonas aeruginosa colonization in lower respiratory tract (LRT) and reduced P. aeruginosa ventilator-associated pneumonia (VAP) rate. Recent studies also suggested that probiotics could improve lung inflammation in mice infected with P. aeruginosa. It seems that microbiota regulation may be a potential therapy for P. aeruginosa VAP patients. However, we know less about the LRT microbial composition and its correlation with prognosis in P. aeruginosa VAP patients. This study aimed to characterize LRT microbiota in P. aeruginosa VAP patients and explore the relationship between microbiota and patient prognosis. Methods Deep endotracheal secretions were sampled from subjects via intubation. Communities were identified by 16S ribosomal RNA gene sequencing. The relationship between microbiota and the prognosis of P. aeruginosa VAP patients were evaluated. Clinical pulmonary infection score and the survival of intensive care unit were both the indicators of patient prognosis. Results In this study, the LRT microbial composition of P. aeruginosa VAP patients was significantly different from non-infected intubation patients, and showed significant individual differences, forming two clusters. According to the predominant phylum of each cluster, these two clusters were named Pro cluster and Fir-Bac cluster respectively. Patients from Pro cluster were dominated by Proteobacteria (adj.P

Abstract The environmentally friendly antibiotic phenazine-1-carboxylic acid (PCA) protects plants, mammals and humans effectively against various fungal pathogens. However, the mechanism by which PCA inhibits or kills fungal pathogens is not fully understood. We analyzed the effects of PCA on the growth of two fungal model organisms, Saccharomyces cerevisiae and Candida albicans, and found that PCA inhibited yeast growth in a dose-dependent manner which was inversely dependent on pH. In contrast, the commonly used antibiotic hygromycin B acted in a dose-dependent manner as pH increased. We then screened a yeast mutant library to identify genes whose mutation or deletion conferred resistance or sensitivity to PCA. We isolated 193 PCA-resistant or PCA-sensitive mutants in clusters, including vesicle-trafficking- and autophagy-defective mutants. Further analysis showed that unlike hygromycin B, PCA significantly altered intracellular vesicular trafficking under growth conditions and blocked autophagy under starvation conditions. These results suggest that PCA inhibits or kills pathogenic fungi in a complex way, in part by disrupting vesicular trafficking and autophagy.

Epinecidin-1 is an antimicrobial peptide derived from the orange-spotted grouper (Epinephelus coioides). The mature epinecidin-1 peptide is predicted to have an amphipathic α-helical structure and a non-helical hydrophilic domain at the C-terminal RRRH. The majority of work studying the potential pharmacological activities of epinecidin-1, utilize synthesized epinecidin-1 (Epi-1), which is made up of 21 amino acids, from the amino acid sequence of 22–42 residues of Epi-1—GFIFHIIKGLFHAGKMIHGLV. The synthetized Epi-1 peptide has been demonstrated to possess diverse pharmacological activities, including antimicrobial, immunomodulatory, anticancer, and wound healing properties. It has also been utilized in different clinical and agricultural fields, including topical applications in wound healing therapy as well as the enhancement of fish immunity in aquaculture. Hence, the present work aims to consolidate the current knowledge and findings on the characteristics and pharmacological properties of epinecidin-1 and its potential applications.

Actinobacteria constitute prolific sources of novel and vital bioactive metabolites for pharmaceutical utilization. In recent years, research has focused on exploring actinobacteria that thrive in extreme conditions to unearth their beneficial bioactive compounds for natural product drug discovery. Natural products have a significant role in resolving public health issues such as antibiotic resistance and cancer. The breakthrough of new technologies has overcome the difficulties in sampling and culturing extremophiles, leading to the outpouring of more studies on actinobacteria from extreme environments. This review focuses on the diversity and bioactive potentials/medically relevant biomolecules of extremophilic actinobacteria found from various unique and extreme niches. Actinobacteria possess an excellent capability to produce various enzymes and secondary metabolites to combat harsh conditions. In particular, a few strains have displayed substantial antibacterial activity against methicillin-resistant Staphylococcus aureus (MRSA), shedding light on the development of MRSA-sensitive antibiotics. Several strains exhibited other prominent bioactivities such as antifungal, anti-HIV, anticancer, and anti-inflammation. By providing an overview of the recently found extremophilic actinobacteria and their important metabolites, we hope to enhance the understanding of their potential for the medical world.

Xanthomonas oryzae pv. oryzae, the bacterial blight pathogen of rice, produces diffusible signal factor (DSF) family quorum sensing signals to regulate virulence. The biosynthesis and perception of DSF family signals require components of the rpf (regulation of pathogenicity factors) cluster. In this study, we report that RpfB plays an essential role in DSF family signal turnover in X. oryzae pv. oryzae PXO99A. The production of DSF family signals was boosted by deletion of the rpfB gene and was abolished by its overexpression. The RpfC/RpfG-mediated DSF signaling system negatively regulates rpfB expression via the global transcription regulator Clp, whose activity is reversible in the presence of cyclic diguanylate monophosphate. These findings indicate that the DSF family signal turnover system in PXO99A is generally consistent with that in Xanthomonas campestris pv. campestris. Moreover, this study has revealed several specific roles of RpfB in PXO99A. First, the rpfB deletion mutant produced high levels of DSF family signals but reduced extracellular polysaccharide production, extracellular amylase activity, and attenuated pathogenicity. Second, the rpfB/rpfC double-deletion mutant was partially deficient in xanthomonadin production. Taken together, the RpfB-dependent DSF family signal turnover system is a conserved and naturally presenting signal turnover system in Xanthomonas spp., which plays unique roles in X. oryzae pv. oryzae adaptation and pathogenesis.

Two almost identical gene clusters (phz1 and phz2) are responsible for phenazine-1-carboxylic acid (PCA) production in Pseudomonas aeruginosa (P. aeruginosa) strain MSH (derived from strain PA1201). Here, we showed that the anti-activator QslA negatively regulated PCA biosynthesis and phz1 expression in strain PA1201 but had little effect on phz2 expression. This downregulation was mediated by a 56-bp region within the 5′-untranslated region (5′-UTR) of the phz1 promoter and was independent of LasR and RsaL signaling. QslA also negatively regulated Pseudomonas quinolone signal (PQS) production. Indeed, QslA controlled the PQS threshold concentration needed for PQS-dependent PCA biosynthesis. The quorum sensing regulator MvfR was required for the QslA-dependent inhibition of PCA production. We identified a direct protein–protein interaction between QslA and MvfR. The ligand-binding domain of MvfR (residues 123–306) was involved in this interaction. Our results suggested that MvfR bound directly to the promoter of the phz1 cluster. QslA interaction with MvfR prevented the binding of MvfR to the phz1 promoter regions. Thus, this study depicted a new mechanism by which QslA controls PCA and PQS biosynthesis in P. aeruginosa.

Bacteria use quorum-sensing (QS) systems to monitor and regulate their population density. Bacterial QS involves small molecules that act as signals for bacterial communication. Many Gram-negative bacterial pathogens use a class of widely conserved molecules, called diffusible signal factor (DSF) family QS signals. The measurement of DSF family signal molecules is essential for understanding DSF metabolic pathways, signaling networks, as well as regulatory roles. Here, we describe a method for the extraction of DSF family signal molecules from Xanthomonas oryzae pv. oryzae (Xoo) cell pellets and Xoo culture supernatant. We determined the levels of DSF family signals using ultra-performance liquid chromatographic system (UPLC) coupled with accurate mass time-of-flight mass spectrometer (TOF-MS). With the aid of UPLC/MS system, the detection limit of DSF was as low as 1 µM, which greatly improves the ability to detect DSF DSF family signal molecules in bacterial cultures and reaction mixtures.

Xanthomonas oryzae pv. oryzae, the causal agent of rice bacterial blight, produces membrane-bound yellow pigments, referred to as xanthomonadins. Xanthomonadins protect the pathogen from photodamage and host-induced perioxidation damage. They are also required for epiphytic survival and successful host plant infection. Here, we show that XanB2 encoded by PXO_3739 plays a key role in xanthomonadin and coenzyme Q8 biosynthesis in X. oryzae pv. oryzae PXO99A. A xanB2 deletion mutant exhibits a pleiotropic phenotype, including xanthomonadin deficiency, producing less exopolysaccharide (EPS), lower viability and H2O2 resistance, and lower virulence. We further demonstrate that X. oryzae pv. oryzae produces 3-hydroxybenzoic acid (3-HBA) and 4-hydroxybenzoic acid (4-HBA) via XanB2. 3-HBA is associated with xanthomonadin biosynthesis while 4-HBA is mainly used as a precursor for coenzyme Q (CoQ)8 biosynthesis. XanB2 is the alternative source of 4-HBA for CoQ8 biosynthesis in PXO99A. These findings suggest that the roles of XanB2 in PXO99A are generally consistent with those in X. campestris pv. campestris. The present study also demonstrated that X. oryzae pv. oryzae PXO99A has evolved several specific features in 3-HBA and 4-HBA signaling. First, our results showed that PXO99A produces less 3-HBA and 4-HBA than X. campestris pv. campestris and this is partially due to a degenerated 4-HBA efflux pump. Second, PXO99A has evolved unique xanthomonadin induction patterns via 3-HBA and 4-HBA. Third, our results showed that 3-HBA or 4-HBA positively regulates the expression of gum cluster to promote EPS production in PXO99A. Taken together, the results of this study indicate that XanB2 is a key metabolic enzyme linking xanthomonadin, CoQ, and EPS biosynthesis, which are collectively essential for X. oryzae pv. oryzae pathogenesis.

Xanthomonas campestris pv. campestris produces a membrane-bound yellow pigment called xanthomonadin. A diffusible factor (DF) has been reported to regulate xanthomonadin biosynthesis. In this study, DF was purified from bacterial culture supernatants using a combination of solvent extraction, flash chromatography, and high-performance liquid chromatography. Mass spectrometry and nuclear magnetic resonance analyses resolved the DF chemical structure as 3-hydroxybenzoic acid (3-HBA), which was further confirmed by synthetic 3-HBA. Significantly, bioassay and in silico analysis suggest that DF production is widely conserved in a range of bacterial species. Analysis of DF derivatives established the hydroxyl group and its position as the key structural features for the role of DF in xanthomonadin biosynthesis. In addition, we showed that DF is also associated with bacterial survival, H2O2 resistance, and systemic invasion. Furthermore, evidence was also presented that DF and diffusible signaling factor have overlapping functions in modulation of bacterial survival, H2O2 resistance, and virulence. Utilization of different mechanisms to modulate similar virulence traits may provide X. campestris pv. campestris with plasticity in response to various environmental cues.

BACKGROUND: Although the basic scorpion K(+) channel toxins (KTxs) are well-known pharmacological tools and potential drug candidates, characterization the acidic KTxs still has the great significance for their potential selectivity towards different K(+) channel subtypes. Unfortunately, research on the acidic KTxs has been ignored for several years and progressed slowly. PRINCIPAL FINDINGS: Here, we describe the identification of nine new acidic KTxs by cDNA cloning and bioinformatic analyses. Seven of these toxins belong to three new α-KTx subfamilies (α-KTx28, α-KTx29, and α-KTx30), and two are new members of the known κ-KTx2 subfamily. ImKTx104 containing three disulfide bridges, the first member of the α-KTx28 subfamily, has a low sequence homology with other known KTxs, and its NMR structure suggests ImKTx104 adopts a modified cystine-stabilized α-helix-loop-β-sheet (CS-α/β) fold motif that has no apparent α-helixs and β-sheets, but still stabilized by three disulfide bridges. These newly described acidic KTxs exhibit differential pharmacological effects on potassium channels. Acidic scorpion toxin ImKTx104 was the first peptide inhibitor found to affect KCNQ1 channel, which is insensitive to the basic KTxs and is strongly associated with human cardiac abnormalities. ImKTx104 selectively inhibited KCNQ1 channel with a K(d) of 11.69 µM, but was less effective against the basic KTxs-sensitive potassium channels. In addition to the ImKTx104 toxin, HeTx204 peptide, containing a cystine-stabilized α-helix-loop-helix (CS-α/α) fold scaffold motif, blocked both Kv1.3 and KCNQ1 channels. StKTx23 toxin, with a cystine-stabilized α-helix-loop-β-sheet (CS-α/β) fold motif, could inhibit Kv1.3 channel, but not the KCNQ1 channel. CONCLUSIONS/SIGNIFICANCE: These findings characterize the structural and functional diversity of acidic KTxs, and could accelerate the development and clinical use of acidic KTxs as pharmacological tools and potential drugs.

Streptomyces learnhanii sp. nov. MUM 203JT was isolated from Malaysia mangrove soil. This Gram-positive bacterium produces pale greenish-yellow aerial and greyish-yellow substrate mycelia on ISP 2 agar. The taxonomy status of strain MUM 203JT was determined via a polyphasic approach comprising phenotypic observations, genomic and phylogenetic analyses, and chemotaxonomic analyses. The strain demonstrated typical Streptomyces features based on a series of phenotypic and chemotaxonomic evaluations. In particular, the cell wall peptidoglycan contained LL-diaminopimelic acid, and the predominant menaquinones detected include MK9(H8). Analysis of whole-cell sugars revealed the presence of glucose, ribose and mannose. The polar lipid profile of the strain comprised lipid, glycolipid, phospholipid, phosphatidylglycerol, phosphatidylinositol, phosphatidylethanolamine, phosphoglycolipid, and diphosphatidylglycerol. The major cellular fatty acids (>10.0 %) were anteiso-C15:0 (24.7 %), anteiso-C17:0 (16.4 %), iso-C16:0 (15.7 %), iso-C15:0 (11.5 %), and C16:0 (11.1 %). The closely related type strains for strain MUM 203JT, as determined by phylogenetic analysis, include Streptomyces coeruleorubidus JCM 4359T (98.8 %), Streptomyces coeruleoprunus JCM 6919T (98.5 %), Streptomyces thermocarboxydovorans NBRC 16324T (98.1 %). The DNA–DNA relatedness values between strain MUM 203JT and closely related type strains ranged from 10.7 ± 0.6 % to 23.3 ± 4.7 %. Strain MUM 203JT has a genome size of 6,446,886 bp, with DNA G + C content of 72.26 mol%. Based on the polyphasic study of strain MUM 203JT, it can be concluded that this strain represents a novel species, for which the name Streptomyces learnhanii sp. nov. is proposed. The type strain is MUM 203JT (= NBRC 114250T = MCCC 1K04200T). [ABSTRACT FROM AUTHOR]