Virulence Of Bacterial Pathogens Research Articles

Molecular characterisation of virulence genes in bacterial pathogens from daycare centres in Ile-Ife, Nigeria: implications for infection control

BackgroundDaycare centres play a critical role in early childhood development but are high-risk environments for infectious disease transmission due to close physical contact, shared toys, inadequate hygiene, and poor ventilation. These risks are especially concerning in low- and middle-income countries (LMICs) like Nigeria, where resources for infection control may be limited. This study aimed to identify and characterise virulence genes in bacterial isolates from daycare centres in Ile-Ife, Nigeria, to assess infection risks.MethodsBetween November 2017 and July 2019, 233 samples were collected from 76 children, 33 daycare workers, and 124 fomites in 17 daycare centres. The bacterial isolates were analysed using conventional PCR and RAPD analysis to detect the presence of virulence genes. The frequency of crucial virulence genes and the prevalence of each bacterial species were recorded.ResultsKey virulence genes were detected, including fimH in Klebsiella species (22.73% of Gram-negative isolates), algD in Pseudomonas aeruginosa (50%), and icaA and cna in Staphylococcus aureus (16.67%). Staphylococcus aureus was the most prevalent species (35%), followed by Klebsiella (28%) and Pseudomonas aeruginosa (20%).ConclusionThis study highlights the presence of virulent bacterial pathogens in daycare environments, posing a severe infection risk to children. To mitigate these risks, it is essential to implement enhanced infection control measures, such as regular microbial screening, improved hand hygiene practices, and disinfection protocols for fomites. Training programs for daycare workers on hygiene practices and routine monitoring could also significantly reduce infection transmission. These interventions are vital for safeguarding the health of daycare children in Nigeria and similar settings globally.

In silico elucidation of protein-protein interaction network in fish pathogen Flavobacterium Columnare

Flavobacterium columnare is a virulent intracellular bacterial pathogen that causes an infection known as columnaris in many species of fish. Some economically important fish species are strongly affected by columnaris, leading to a high mortality rate and significant economic losses. Previous in silico studies have provided various biological insights into F. columnare, including its interaction with MHC class I alleles and the epitopic region within outer membrane proteins. However, the protein-protein interaction networks underlying the growth, defense, and pathogenesis of F. columnare remain largely unknown. This study was conducted to identify the protein-protein interaction (PPI) networks and hub proteins of F. columnare that can be used as drug or vaccine targets. A total of 500 protein sequences were retrieved from UniprotKB in FASTA format and analyzed using VaxiJen, PSORTb, STRING, Cytoscape, and BLASTp programs. The results demonstrated that 60% of F. columnare proteins were predicted as antigenic proteins, most of which were associated with catalytic activity and metabolic processes, identified as cytoplasmic proteins. Ten hub proteins with the highest number of functional interactions were identified, which were also antigenic and non-host homologous. In conclusion, F. columnare hub proteins represent potential therapeutic targets in drug and vaccine development against columnaris infection.

Cytochrome oxidase requirements in Bordetella reveal insights into evolution towards life in the mammalian respiratory tract.

Little is known about oxygen utilization during infection by bacterial respiratory pathogens. The classical Bordetella species, including B. pertussis, the causal agent of human whooping cough, and B. bronchiseptica, which infects nearly all mammals, are obligate aerobes that use only oxygen as the terminal electron acceptor for electron transport-coupled oxidative phosphorylation. B. bronchiseptica, which occupies many niches, has eight distinct cytochrome oxidase-encoding loci, while B. pertussis, which evolved from a B. bronchiseptica-like ancestor but now survives exclusively in and between human respiratory tracts, has only three functional cytochrome oxidase-encoding loci: cydAB1, ctaCDFGE1, and cyoABCD1. To test the hypothesis that the three cytochrome oxidases encoded within the B. pertussis genome represent the minimum number and class of cytochrome oxidase required for respiratory infection, we compared B. bronchiseptica strains lacking one or more of the eight possible cytochrome oxidases in vitro and in vivo. No individual cytochrome oxidase was required for growth in ambient air, and all three of the cytochrome oxidases conserved in B. pertussis were sufficient for growth in ambient air and low oxygen. Using a high-dose, large-volume persistence model and a low-dose, small-volume establishment of infection model, we found that B. bronchiseptica producing only the three B. pertussis-conserved cytochrome oxidases was indistinguishable from the wild-type strain for infection. We also determined that CyoABCD1 is sufficient to cause the same level of bacterial burden in mice as the wild-type strain and is thus the primary cytochrome oxidase required for murine infection, and that CydAB1 and CtaCDFGE1 fulfill auxiliary roles or are important for aspects of infection we have not assessed, such as transmission. Our results shed light on the environment at the surface of the ciliated epithelium, respiration requirements for bacteria that colonize the respiratory tract, and the evolution of virulence in bacterial pathogens.

Long-term balancing selection for pathogen resistance maintains trans-species polymorphisms in a planktonic crustacean

Balancing selection is an evolutionary process that maintains genetic polymorphisms at selected loci and strongly reduces the likelihood of allele fixation. When allelic polymorphisms that predate speciation events are maintained independently in the resulting lineages, a pattern of trans-species polymorphisms may occur. Trans-species polymorphisms have been identified for loci related to mating systems and the MHC, but they are generally rare. Trans-species polymorphisms in disease loci are believed to be a consequence of long-term host-parasite coevolution by balancing selection, the so-called Red Queen dynamics. Here we scan the genomes of three crustaceans with a divergence of over 15 million years and identify 11 genes containing identical-by-descent trans-species polymorphisms with the same polymorphisms in all three species. Four of these genes display molecular footprints of balancing selection and have a function related to immunity. Three of them are located in or close to loci involved in resistance to a virulent bacterial pathogen, Pasteuria, with which the Daphnia host is known to coevolve. This provides rare evidence of trans-species polymorphisms for loci known to be functionally relevant in interactions with a widespread and highly specific parasite. These findings support the theory that specific antagonistic coevolution is able to maintain genetic diversity over millions of years.

Characterization of Ssc, an N-acetylgalactosamine-containing Staphylococcus aureus surface polysaccharide.

Surface polysaccharides play crucial roles in the biology and virulence of bacterial pathogens. Staphylococcus aureus produces four major types of polysaccharides that have been well-characterized. In this study, we identified a new surface polysaccharide containing N-acetylgalactosamine (GalNAc). This marks the first report of GalNAc-containing polysaccharide in S. aureus. Our discovery lays the groundwork for further investigations into the chemical structure, surface location, and role in pathogenesis of this new polysaccharide.

Investigating antimicrobial resistance genes in Kenya, Uganda and Tanzania cattle using metagenomics.

Antimicrobial resistance (AMR) is a growing problem in African cattle production systems, posing a threat to human and animal health and the associated economic value chain. However, there is a poor understanding of the resistomes in small-holder cattle breeds in East African countries. This study aims to examine the distribution of antimicrobial resistance genes (ARGs) in Kenya, Tanzania, and Uganda cattle using a metagenomics approach. We used the SqueezeMeta-Abricate (assembly-based) pipeline to detect ARGs and benchmarked this approach using the Centifuge-AMRplusplus (read-based) pipeline to evaluate its efficiency. Our findings reveal a significant number of ARGs of critical medical and economic importance in all three countries, including resistance to drugs of last resort such as carbapenems, suggesting the presence of highly virulent and antibiotic-resistant bacterial pathogens (ESKAPE) circulating in East Africa. Shared ARGs such as aph(6)-id (aminoglycoside phosphotransferase), tet (tetracycline resistance gene), sul2 (sulfonamide resistance gene) and cfxA_gen (betalactamase gene) were detected. Assembly-based methods revealed fewer ARGs compared to read-based methods, indicating the sensitivity and specificity of read-based methods in resistome characterization. Our findings call for further surveillance to estimate the intensity of the antibiotic resistance problem and wider resistome classification. Effective management of livestock and antibiotic consumption is crucial in minimizing antimicrobial resistance and maximizing productivity, making these findings relevant to stakeholders, agriculturists, and veterinarians in East Africa and Africa at large.

Metal selectivity and translocation mechanism characterization in proteoliposomes of the transmembrane NiCoT transporter NixA from Helicobacter pylori.

Essential trace metals play key roles in the survival, replication, and virulence of bacterial pathogens. Helicobacter pylori (H. pylori), the main bacterial cause of gastric ulcers, requires Ni(ii) to colonize and persist in the acidic environment inside the stomach, exploiting the nickel-containing enzyme urease to catalyze the hydrolysis of urea to ammonia and bicarbonate and create a pH-buffered microenvironment. Urease utilizes Ni(ii) as a catalytic cofactor for its activity. In ureolytic bacteria, unique transmembrane (TM) transporters evolved to guarantee the selective uptake and efflux of Ni(ii) across cellular membranes to meet the cellular requirements. NixA is an essential Ni(ii) transporter expressed by H. pylori when the extracellular environment experiences a drop in pH. This Class I nickel-cobalt transporter of the NiCoT family catalyzes the uptake of Ni(ii) across the inner membrane from the periplasm. In this study, we characterized NixA using a platform whereby, for the first time on a NiCoT transporter, recombinantly expressed and purified NixA and key mutants in the translocation pathway have been reconstituted in artificial lipid bilayer vesicles (proteoliposomes). Fluorescent sensors responsive to Ni(ii) transport (Fluozin-3-Zn(ii)), luminal pH changes (pyranine), and membrane potential (oxonol VI) were encapsulated in the proteoliposomes lumen to monitor, in real-time, NixA transport properties and translocation mechanism. Kinetic transport analysis revealed that NixA is highly selective for Ni(ii) with no substrate promiscuity towards Co(ii), the other putative metal substrate of the NiCoT family, nor Zn(ii). NixA-mediated Ni(ii) transport exhibited a Michaelis-Menten-type saturable substrate concentration dependence, with an experimental KM, Ni(ii) = 31.0 ± 1.2 μM. Ni(ii) transport by NixA was demonstrated to be electrogenic, and metal translocation did not require a proton motive force, resulting in the generation of a positive-inside transmembrane potential in the proteoliposome lumen. Mutation analysis characterized key transmembrane residues for substrate recognition, binding, and/or transport, suggesting the presence of a three-step transmembrane translocation conduit. Taken together, these investigations reveal that NixA is a Ni(ii)-selective Class I NiCoT electrogenic uniporter. The work also provides an in vitro approach to characterize the transport properties of metal transporters responsible for Ni(ii) acquisition and extrusion in prokaryotes.

The effect of metal remediation on the virulence and antimicrobial resistance of the opportunistic pathogen Pseudomonas aeruginosa.

Anthropogenic metal pollution can result in co-selection for antibiotic resistance and potentially select for increased virulence in bacterial pathogens. Metal-polluted environments can select for the increased production of siderophore molecules to detoxify non-ferrous metals. However, these same molecules also aid the uptake of ferric iron, a limiting factor for within-host pathogen growth, and are consequently a virulence factor. Anthropogenic methods to remediate environmental metal contamination commonly involve amendment with lime-containing materials. However, whether this reduces in situ co-selection for antibiotic resistance and siderophore-mediated virulence remains unknown. Here, using microcosms containing non-sterile metal-contaminated river water and sediment, we test whether liming reduces co-selection for these pathogenicity traits in the opportunistic pathogen Pseudomonas aeruginosa. To account for the effect of environmental structure, which is known to impact siderophore production, microcosms were incubated under either static or shaking conditions. Evolved P. aeruginosa populations had greater fitness in the presence of toxic concentrations of copper than the ancestral strain and showed increased resistance to the clinically relevant antibiotics apramycin, cefotaxime and trimethoprim, regardless of lime addition or environmental structure. Although we found virulence to be significantly associated with siderophore production, neither virulence nor siderophore production significantly differed between the four treatments. Furthermore, liming did not mitigate metal-imposed selection for antibiotic resistance or virulence in P. aeruginosa. Consequently, metal-contaminated environments may select for antibiotic resistance and virulence traits even when treated with lime.

Dietary iron regulates intestinal goblet cell function and alleviates Salmonella typhimurium invasion in mice.

Iron is an important micronutrient that plays a vital role in host defenses and bacterial pathogenicity. As iron treatments increase the risk of infection by stimulating the growth and virulence of bacterial pathogens, their roles in anti-infection immunity have frequently been underestimated. To estimate whether adequate dietary iron intake would help defend against pathogenic bacterial infection, mice were fed iron-deficient (2 mg kg-1 feed), iron-sufficient (35 mg kg-1 feed), or iron-enriched diet (350 mg kg-1 feed) for 12 weeks, followed by oral infection with Salmonella typhimurium. Our results revealed that dietary iron intake improved mucus layer function and decelerated the invasion of the pathogenic bacteria, Salmonella typhimurium. Positive correlations between serum iron and the number of goblet cells and mucin2 were found in response to total iron intake in mice. Unabsorbed iron in the intestinal tract affected the gut microbiota composition, and the abundance of Bacteroidales, family Muribaculaceae, was positively correlated with their mucin2 expression. However, the results from antibiotic-treated mice showed that the dietary iron-regulated mucin layer function was not microbial-dependent. Furthermore, in vitro studies revealed that ferric citrate directly induced mucin2 expression and promoted the proliferation of goblet cells in both ileal and colonic organoids. Thus, dietary iron intake improves serum iron levels, regulates goblet cell regeneration and mucin layer function, and plays a positive role in the prevention of pathogenic bacteria.

The LiaFSR-LiaX System Mediates Resistance of Enterococcus faecium to Peptide Antibiotics and to Aureocin A53- and Enterocin L50-Like Bacteriocins.

Multidrug-resistant Enterococcus faecium strains are currently a leading cause of difficult-to-treat nosocomial infections. The emerging resistance of enterococci to last-resort antibiotics, such as daptomycin, prompts a search for alternative antimicrobials. Aureocin A53- and enterocin L50-like bacteriocins are potent antimicrobial agents that form daptomycin-like cationic complexes and have a similar cell envelope-targeting mechanism of action, suggesting their potential as next-generation antibiotics. However, to ensure their safe use, the mechanisms of resistance to these bacteriocins and cross-resistance to antibiotics need to be well understood. Here, we investigated the genetic basis of E. faecium's resistance to aureocin A53- and enterocin L50-like bacteriocins and compared it with that to antibiotics. First, we selected spontaneous mutants resistant to the bacteriocin BHT-B and identified adaptive mutations in the liaFSR-liaX genes encoding the LiaFSR stress response regulatory system and the daptomycin-sensing protein LiaX, respectively. We then demonstrated that a gain-of-function mutation in liaR increases the expression of liaFSR, liaXYZ, cell wall remodeling-associated genes, and hypothetical genes involved in protection against various antimicrobials. Finally, we showed that adaptive mutations or overexpression of liaSR or liaR alone results in cross-resistance to other aureocin A53- and enterocin L50-like bacteriocins, as well as antibiotics targeting specific components of the cell envelope (daptomycin, ramoplanin, gramicidin) or ribosomes (kanamycin and gentamicin). Based on the obtained results, we concluded that activation of the LiaFSR-mediated stress response confers resistance to peptide antibiotics and bacteriocins via a cascade of reactions, eventually leading to cell envelope remodeling. IMPORTANCE Pathogenic enterococci carry virulence factors and a considerable resistome, which makes them one of the most serious and steadily increasing causes of hospital epidemiological risks. Accordingly, Enterococcus faecium is classified into a top-priority ESKAPE (Enterococcus faecium, Staphylococcus aureus, Klebsiella pneumoniae, Acinetobacter baumannii, Pseudomonas aeruginosa, and Enterobacter species) group of six highly virulent and multidrug-resistant (MDR) bacterial pathogens for which novel antimicrobial agents need to be developed urgently. Alternative measures, such as the use of bacteriocins, separately or in combination with other antimicrobial agents (e.g., antibiotics), could be a potential solution, especially since several international health agencies recommend and support the development of such interventions. Nevertheless, in order to exploit their efficacy, more basic research on the mechanisms of cell killing and the development of resistance to bacteriocins is needed. The present study fills some of the knowledge gaps regarding the genetic basis of the development of resistance to potent antienterococcal bacteriocins, pointing out the common and divergent features regarding the cross-resistance to antibiotics.

Computational Modelling, Functional Characterization and Molecular Docking to Lead Compounds of Bordetella pertussis Diaminopimelate Epimerase.

Bordetella pertussis, the causative agent of whooping cough, is an opportunistic virulent bacterial pathogen that is resistant to a wide range of antibiotics due to a variety of resistance mechanisms. Looking at the increasing number of infections caused by B. pertussis and its resistance to diverse antibiotics, it is essential to develop alternative strategies to fight against B. pertussis. Diaminopimelate epimerase (DapF) is an important enzyme of the lysine biosynthesis pathway in B. pertussis that catalyzes the formation of meso-2, 6-diaminoheptanedioate (meso-DAP), which is an important step in lysine metabolism. Therefore, Bordetella pertussis diaminopimelate epimerase (DapF) becomes an ideal target for antimicrobial drug development. In the present study, computational modelling, functional characterization, binding studies, and docking studies of BpDapF with lead compounds were carried out using different in silico tools. In silico prediction results in the secondary structure, 3-D structure analysis, and protein-protein interaction analysis of BpDapF. Docking studies further showed the respective amino acid residues for ligands in the phosphate‑binding loop of BpDapF play a vital role in the formation of H‑bonds with these ligands. The site where the ligand was bound is a deep groove, which is regarded as the binding cavity of the protein. Biochemical studies indicated that Limonin (binding energy - 8.8kcal/mol), Ajmalicine (binding energy - 8.7kcal/mol), Clinafloxacin (binding energy - 8.3kcal/mol), Dexamethasone (binding energy - 8.2kcal/mol), and Tetracycline (binding energy - 8.1kcal/mol) exhibited promising binding towards the drug target DapF of B. pertussis in comparison with the binding between other drugs and act as the potential inhibitors of BpDapF that eventually can reduce the catalytic activity of BpDapF.

Pseudomonas aeruginosa Cytotoxins: Mechanisms of Cytotoxicity and Impact on Inflammatory Responses.

Pseudomonas aeruginosa is one of the most virulent opportunistic Gram-negative bacterial pathogens in humans. It causes many acute and chronic infections with morbidity and mortality rates as high as 40%. P. aeruginosa owes its pathogenic versatility to a large arsenal of cell-associated and secreted virulence factors which enable this pathogen to colonize various niches within hosts and protect it from host innate immune defenses. Induction of cytotoxicity in target host cells is a major virulence strategy for P. aeruginosa during the course of infection. P. aeruginosa has invested heavily in this strategy, as manifested by a plethora of cytotoxins that can induce various forms of cell death in target host cells. In this review, we provide an in-depth review of P. aeruginosa cytotoxins based on their mechanisms of cytotoxicity and the possible consequences of their cytotoxicity on host immune responses.

Characterization of Virulent Bacterial Isolates Associated with Multi-Drug Resistance among Patients with Surgical Site Infections in Selected Specialist Hospitals in Calabar, Nigeria

Aims: Increasing research findings have documented the continuous emergence and threats posed by drug resistant clinical isolates from post-operative wound infections to commonly used antibiotics globally. This hospital-based study investigated virulent bacterial pathogens implicated with post-operative wound infections among surgical site infection (SSI) patients in Calabar, Nigeria and determined their antibiotic resistance pattern.
 Methodology: A total of 127 bacterial isolates of different genus from 110 SSI patients, were isolated from pus and surgical wound exudates and fully characterized using standard bacteriological procedures. Antimicrobial susceptibility patterns of isolates were determined using Kirby- Bauer disk diffusion method, following the guidelines by Clinical Laboratory Standard Institute (CLSI).
 Results: Multi-drug resistant bacteria isolated and their percentage frequency were coagulase Negative Staphylococci (21.3%), Staphylococcus aureus (19.7%), Pseudomonas aeruginosa (14.2%), Escherichia coli (11.8%), Klebsiella pneumoniae (9.4%), Enterococcus faecium (6.3), Enterobacter cloacae (4.7%), Proteus mirabilis (4.7%), Acinetobacter baumannii (3.1%), Pseudomonas putida (3.1%) and Aerococcus viridans (1.6%). Among gram-positive bacteria isolated, S. aureus showed highest resistance to several antimicrobials (100% to oxacillin, 96% to ciprofloxacin, 92% to levofloxacin, and 76% resistance to vancomycin). All recovered S. aureus isolates were cefoxitin screen positive indicating possible MRSA isolates. Additionally, among Gram-negative isolates K. pneumoniae was found to possess higher resistance to several antibiotics (66.7% resistance to each of ciprofloxacin, levofloxacin, ceftazidime, trimethoprim /sulfamethoxazole, cefazolin, ampicillin, tobramycin and 58.3% resistance to each of ceftriaxone, gentamicin, and ampicillin/sulbactam). Statistical analysis of categorical variables of study subjects revealed that length of hospital stay, type of surgery, previous admission history, antibiotic use, and age were significant (p<0.05) in SSI outcome of patients, while patients’ gender was not significant (p>0.05) in SSI outcome.
 Conclusion: Adherence to measures of strict infection control, optimal preoperative, intraoperative, and postoperative patient care, including multifaceted approaches involving surveillance, and antimicrobial stewardship, are vital to SSI treatment outcomes.

Removal of clinically significant antibiotics from aqueous solutions by applying unique high-throughput continuous-flow plasma pencil and plasma brush systems

High demands of medical and husbandry sectors led to a massive annual production of 100,000 tonnes of various antibiotics on a global scale. Considering that an access to sublethal doses of these pharmaceuticals causes subsequent increase in resistance of virulent human bacterial pathogens, there is a substantial need for development of novel methods and techniques for degradation of antibiotics from aqueous solutions. Here, high-throughput continuous flow plasma pencil and plasma brush methodologies were developed and verified for their applicability for removal of antibiotics from solutions, as estimated by high-performance liquid chromatography coupled with diode-array detection (HPLC-DAD) or ultraperformance liquid chromatography coupled with tandem mass spectrometry (UPLC-MS/MS). The removal efficiency of drugs treated by the plasma pencil was in the range from 3 to 24%, and was corresponded to with 1.39% to 10.76% losses in the antibacterial properties of the plasma-treated solutions. However, application of a unique plasma brush increased the antibiotics degradation rates significantly, now falling in the range from 29 to 67% that led to 12.06–81.59% decrease in the antimicrobial properties of the plasma-exposed antibiotics solutions. The deviations in the effectiveness of the plasma pencil and the plasma brush methods were assigned with the types and amounts of generated species. Various intermediate degradation products were also detected in the post-plasma solutions, which derived from the oxygen attack to nucleophilic sides of the antibiotic molecules. We assume that such novel high-throughput, continuous flow methodological setups may find application in hospitals as an end-of-pipe technology for routine elimination of pollutants before reaching municipal sewage treatment plants.

Arginine Catabolism and Polyamine Biosynthesis Pathway Disparities Within Francisella tularensis Subpopulations.

Francisella tularensis is a highly infectious zoonotic pathogen with as few as 10 organisms causing tularemia, a disease that is fatal if untreated. Although F. tularensis subspecies tularensis (type A) and subspecies holarctica (type B) share over 99.5% average nucleotide identity, notable differences exist in genomic organization and pathogenicity. The type A clade has been further divided into subtypes A.I and A.II, with A.I strains being recognized as some of the most virulent bacterial pathogens known. In this study, we report on major disparities that exist between the F. tularensis subpopulations in arginine catabolism and subsequent polyamine biosynthesis. The genes involved in these pathways include the speHEA and aguAB operons, along with metK. In the hypervirulent F. tularensis A.I clade, such as the A.I prototype strain SCHU S4, these genes were found to be intact and highly transcribed. In contrast, both subtype A.II and type B strains have a truncated speA gene, while the type B clade also has a disrupted aguA and truncated aguB. Ablation of the chromosomal speE gene that encodes a spermidine synthase reduced subtype A.I SCHU S4 growth rate, whereas the growth rate of type B LVS was enhanced. These results demonstrate that spermine synthase SpeE promotes faster replication in the F. tularensis A.I clade, whereas type B strains do not rely on this enzyme for in vitro fitness. Our ongoing studies on amino acid and polyamine flux within hypervirulent A.I strains should provide a better understanding of the factors that contribute to F. tularensis pathogenicity.

Complete Genome Sequences of Four Strains ofErwiniatracheiphila: A Resource for Studying aBacterial Plant Pathogen with a Highly Complex Genome.

Complete Genome Sequences of Four Strains ofErwiniatracheiphila: A Resource for Studying aBacterial Plant Pathogen with a Highly Complex Genome.

MTO1-RESPONDING DOWN 1 (MRD1) is a transcriptional target of OZF1 for promoting salicylic acid-mediated defense in Arabidopsis.

OZF1 promotes the transcription of MRD1, which is essential for SA-mediated defense against virulent and avirulent bacterial pathogens in Arabidopsis. Salicylic acid (SA) is critical for defense against biotrophic pathogens. A trans-activator protein NPR1 plays significant roles in SA-signaling. However, evidences suggest the existence of NPR1-independent pathways for SA signaling in plants. Previously, we reported Arabidopsis OXIDATION-RELATED ZN-FINGER PROTEIN1 (OZF1) as a positive regulator of NPR1-independent SA-signaling. However, the mechanism or components of OZF1-mediated SA signaling was not known. Through the analysis of differentially expressing genes, we report the identification of MTO1-RESPONDING DOWN 1 (MRD1) as a transcriptional target of OZF1. Expressions of MRD1 and its overlapping gene in Arabidopsis genome, HEI10 increase upon pathogen inoculation in an OZF1-dependent manner. Their mutants are susceptible to both virulent and avirulent bacterial pathogens and show compromised SA-mediated immunity. Overexpression of MRD1 but not the HEI10 rescues the loss-of-resistance phenotype of the ozf1 mutant. OZF1 physically associates at the MRD1 promoter area upon pathogen inoculation. Results altogether support that MRD1 is a transcriptional target of OZF1 for promoting SA-mediated defense in Arabidopsis.

Xenogeneic nucleoid-associated EnrR thwarts H-NS silencing of bacterial virulence with unique DNA binding

Type III and type VI secretion systems (T3/T6SS) are encoded in horizontally acquired genomic islands (GIs) that play crucial roles in evolution and virulence in bacterial pathogens. T3/T6SS expression is subjected to tight control by the host xenogeneic silencer H-NS, but how this mechanism is counteracted remains to be illuminated. Here, we report that xenogeneic nucleoid-associated protein EnrR encoded in a GI is essential for virulence in pathogenic bacteria Edwardsiella and Salmonella. We showed that EnrR plays critical roles in T3/T6SS expression in these bacteria. Various biochemical and genetic analyses demonstrated that EnrR binds and derepresses the promoter of esrB, the critical regulator of T3/T6SS, to promote their expression by competing with H-NS. Additionally, EnrR targets AT-rich regions, globally modulates the expression of ∼363 genes and is involved in various cellular processes. Crystal structures of EnrR in complex with a specific AT-rich palindromic DNA revealed a new DNA-binding mode that involves conserved HTH-mediated interactions with the major groove and contacts of its N-terminal extension to the minor groove in the symmetry-related duplex. Collectively, these data demonstrate that EnrR is a virulence activator that can antagonize H-NS, highlighting a unique mechanism by which bacterial xenogeneic regulators recognize and regulate foreign DNA.

Response of tomatoes primed by mycorrhizal colonization to virulent and avirulent bacterial pathogens

Most plants interact with arbuscular mycorrhizal fungi, which enhance disease resistance in the host plant. Because the effects of resistance against bacterial pathogens are poorly understood, we investigated the effects of mycorrhizal colonization on virulent and avirulent pathogens using phytopathological and molecular biology techniques. Tomato plants colonized by Gigaspora margarita acquired resistance not only against the fungal pathogen, Botrytis cinerea, but also against a virulent bacterial pathogen, Pseudomonas syringae pv. tomato DC3000 (Pst). In G. margarita-colonized tomato, salicylic acid (SA)- and jasmonic acid (JA)-related defense genes were expressed more rapidly and strongly compared to those in the control plants when challenged by Pst, indicating that the plant immunity system was primed by mycorrhizal colonization. Gene expression analysis indicated that primed tomato plants responded to the avirulent pathogen, Pseudomonas syringae pv. oryzae, more rapidly and strongly compared to the control plant, where the effect on the JA-mediated signals was stronger than in the case with Pst. We found that the resistance induced by mycorrhizal colonization was effective against both fungal and bacterial pathogens including virulent and avirulent pathogens. Moreover, the activation of both SA- and JA-mediated signaling pathways can be enhanced in the primed plant by mycorrhizal colonization.

RGI‐GOLVEN signaling promotes cell surface immune receptor abundance to regulate plant immunity

Plant immune responses must be tightly controlled for proper allocation of resources for growth and development. In plants, endogenous signaling peptides regulate developmental and growth‐related processes. Recent research indicates that some of these peptides also have regulatory functions in the control of plant immune responses. This classifies these peptides as phytocytokines as they show analogies with metazoan cytokines. However, the mechanistic basis for phytocytokine‐mediated regulation of plant immunity remains largely elusive. Here, we identify GOLVEN2 (GLV2) peptides as phytocytokines in Arabidopsis thaliana. GLV2 signaling enhances sensitivity of plants to elicitation with immunogenic bacterial elicitors and contributes to resistance against virulent bacterial pathogens. GLV2 is perceived by ROOT MERISTEM GROWTH FACTOR 1 INSENSITIVE (RGI) receptors. RGI mutants show reduced elicitor sensitivity and enhanced susceptibility to bacterial infection. RGI3 forms ligand‐induced complexes with the pattern recognition receptor (PRR) FLAGELLIN SENSITIVE 2 (FLS2), suggesting that RGIs are part of PRR signaling platforms. GLV2‐RGI signaling promotes PRR abundance independent of transcriptional regulation and controls plant immunity via a previously undescribed mechanism of phytocytokine activity.