Soil sickness is a critical problem for eggplant (Solanum melongena L.) under continuous cropping that affects sustainable eggplant production. Relay intercropping is a significant technique on promoting soil quality, improving eco-environment, and raising output. Field experiments were conducted from September 2010 to November 2012 in northwest China to determine the effects of relay intercropping eggplant with garlic (Allium sativum L.) on soil enzyme activities, available nutrient contents, and pH value under a plastic tunnel. Three treatments were in triplicate using randomized block design: eggplant monoculture (CK), eggplant relay intercropping with normal garlic (NG) and eggplant relay intercropping with green garlic (GG). The major results are as follows: (1) the activities of soil invertase, urease, and alkaline phosphatase were generally enhanced in NG and GG treatments; (2) relay intercropping significantly increased the soil available nutrient contents, and they were mostly higher in GG than NG. On April 11, 2011, the eggplant/garlic co-growth stage, the available nitrogen content in GG was 76.30 mg·kg(-1), significantly higher than 61.95 mg·kg(-1) in NG. For available potassium on April 17, 2012, they were 398.48 and 387.97 mg·kg(-1) in NG and GG, both were significantly higher than 314.84 mg·kg(-1) in CK; (3) the soil pH showed a significantly higher level in NG treatment, but lower in GG treatment compared with CK. For the last samples in 2012, soil pH in NG and GG were 7.70 and 7.46, the highest and lowest one among them; (4) the alkaline phosphatase activity and pH displayed a similar decreasing trend with continuous cropping. These findings indicate that relay intercropping eggplant with garlic could be an ideal farming system to effectively improve soil nutrient content, increase soil fertility, and alleviate soil sickness to some extent. These findings are important in helping to develop sustainable eggplant production.

A hydroponic co-culture system was adopted to determine the allelopathic potential of garlic on the growth of pepper plants. Different numbers of garlic plants (0, 2, 4, 8 and 12) were hydroponically co-cultured with two pepper plants to investigate allelopathic effects on the growth attributes and antioxidative defense system of the test pepper plants. The responses of the pepper plants depended on the number of garlic plants included in the co-culture system, indicating an association of pepper growth with the garlic root exudate concentration. When grown at a pepper/garlic ratio of 1:1 or 1:2, the pepper plant height, chlorophyll content, and peroxidase (POD), catalase (CAT) and phenylalanine ammonia-lyase (PAL) activities were significantly increased after 30 days of co-culture; in contrast, reduction in methane dicarboxylic aldehyde (MDA) content was observed. However, when the pepper/garlic ratio was 1:4 or higher, these morphological indices and protective enzyme activities were significantly inhibited, whereas MDA levels in the pepper leaves were significantly increased due to severe membrane lipid peroxidation. The results indicate that although low concentrations of garlic root exudates appear to induce protective enzyme systems and promote pepper growth, high concentrations have deleterious effects. These findings suggest that further investigations should optimize the co-culture pepper/garlic ratio to reduce continuous cropping obstacles in pepper production.

Pseudomonas aeruginosa causes acute and chronic infections in human. Its increasing resistance to antibiotics requires alternative treatments that are more effective than available strategies. Among the alternatives is the unconventional usage of conventional antibiotics, of which the macrolide antibiotic azithromycin (AZM) provides a paradigmatic example. AZM therapy is associated with a small but consistent improvement in respiratory function of cystic fibrosis patients suffering from chronic P. aeruginosa infection. Besides immunomodulating activities, AZM represses bacterial genes involved in virulence, quorum sensing, biofilm formation, and motility, all of which are due to stalling of ribosome and depletion of cellular tRNA pool. However, how P. aeruginosa responds to and counteracts the effects of AZM remain elusive. Here, we found that deficiency of PA3297, a gene encoding a DEAH-box helicase, intensified AZM-mediated bacterial killing, suppression of pyocyanin production and swarming motility, and hypersusceptibility to hydrogen peroxide. We demonstrated that expression of PA3297 is induced by the interaction between AZM and ribosome. Importantly, mutation of PA3297 resulted in elevated levels of unprocessed 23S-5S rRNA in the presence of AZM, which might lead to increased susceptibility to AZM-mediated effects. Our results revealed one of the bacterial responses in counteracting the detrimental effects of AZM.

The incorporation of plant residues into soil can be considered a keystone sustainability factor in improving soil structure function. However, the effects of plant residue addition on the soil microbial communities involved in biochemical cycles and abiotic stress phenomena are poorly understood. In this study, experiments were conducted to evaluate the role of raw garlic stalk (RGS) amendment in avoiding monoculture-related production constraints by studying the changes in soil chemical properties and microbial community structures. RGS was applied in four different doses, namely the control (RGS0), 1% (RGS1), 3% (RGS2), and 5% (RGS3) per 100 g of soil. The RGS amendment significantly increased soil electrical conductivity (EC), N, P, K, and enzyme activity. The soil pH significantly decreased with RGS application. High-throughput Illumina MiSeq sequencing revealed significant alterations in bacterial community structures in response to RGS application. Among the 23 major taxa detected, Anaerolineaceae, Acidobacteria, and Cyanobacteria exhibited an increased abundance level. RGS2 increased some bacteria reported to be beneficial including Acidobacteria, Bacillus, and Planctomyces (by 42%, 64%, and 1% respectively). Furthermore, internal transcribed spacer (ITS) fungal regions revealed significant diversity among the different treatments, with taxa such as Chaetomium (56.2%), Acremonium (4.3%), Fusarium (4%), Aspergillus (3.4%), Sordariomycetes (3%), and Plectosphaerellaceae (2%) showing much abundance. Interestingly, Coprinellus (14%) was observed only in RGS-amended soil. RGS treatments effectively altered soil fungal community structures and reduced certain known pathogenic fungal genera, i.e., Fusarium and Acremonium. The results of the present study suggest that RGS amendment potentially affects the microbial community structures that probably affect the physiological and morphological attributes of eggplant under a plastic greenhouse vegetable cultivation system (PGVC) in monoculture.

We have demonstrated that Na/K-ATPase acts as a receptor for reactive oxygen species (ROS), regulating renal Na+ handling and blood pressure. TALLYHO/JngJ (TH) mice are believed to mimic the state of obesity in humans with a polygenic background of type 2 diabetes. This present work is to investigate the role of Na/K-ATPase signaling in TH mice, focusing on susceptibility to hypertension due to chronic excess salt ingestion. Age-matched male TH and the control C57BL/6J (B6) mice were fed either normal diet or high salt diet (HS: 2, 4, and 8% NaCl) to construct the renal function curve. Na/K-ATPase signaling including c-Src and ERK1/2 phosphorylation, as well as protein carbonylation (a commonly used marker for enhanced ROS production), were assessed in the kidney cortex tissues by Western blot. Urinary and plasma Na+ levels were measured by flame photometry. When compared to B6 mice, TH mice developed salt-sensitive hypertension and responded to a high salt diet with a significant rise in systolic blood pressure indicative of a blunted pressure-natriuresis relationship. These findings were evidenced by a decrease in total and fractional Na+ excretion and a right-shifted renal function curve with a reduced slope. This salt-sensitive hypertension correlated with changes in the Na/K-ATPase signaling. Specifically, Na/K-ATPase signaling was not able to be stimulated by HS due to the activated baseline protein carbonylation, phosphorylation of c-Src and ERK1/2. These findings support the emerging view that Na/K-ATPase signaling contributes to metabolic disease and suggest that malfunction of the Na/K-ATPase signaling may promote the development of salt-sensitive hypertension in obesity. The increased basal level of renal Na/K-ATPase-dependent redox signaling may be responsible for the development of salt-sensitive hypertension in polygenic obese TH mice.

Cover crops are key determinants of the ecological stability and sustainability of continuous cropping soils. However, their agro-ecological role in differentially reshaping the microbiome structure and functioning under a degraded agroecosystem remains poorly investigated. Therefore, structural and metabolic changes in soil bacterial community composition in response to diverse plant species were assessed. Winter catch leafy vegetables crops were introduced as cover plants in a cucumber-fallow period. The results indicate that cover crop diversification promoted beneficial changes in soil chemical and biological attributes, which increased crop yields in a cucumber double-cropping system. Illumina high-throughput sequencing of 16S rRNA genes indicated that the bacterial community composition and diversity changed through changes in the soil properties. Principal component analysis (PCA) coupled with non-metric multidimensional scaling (NMDS) analysis reveals that the cover planting shaped the soil microbiome more than the fallow planting (FC). Among different cropping systems, spinach-cucumber (SC) and non-heading Chinese cabbage-cucumber (NCCC) planting systems greatly induced higher soil nutrient function, biological activity, and bacterial diversity, thus resulting in higher cucumber yield. Quantitative analysis of linear discriminant analysis effect size (LEfSe) indicated that Proteobacteria, Actinobacteria, Bacteroidetes, and Acidobacteria were the potentially functional and active soil microbial taxa. Rhizospheres of NCCC, leaf lettuce-cucumber (LLC), coriander-cucumber (CC), and SC planting systems created hotspots for metabolic capabilities of abundant functional genes, compared to FC. In addition, the predictive metabolic characteristics (metabolism and detoxification) associated with host-plant symbiosis could be an important ecological signal that provides direct evidence of mediation of soil structure stability. Interestingly, the plant density of non-heading Chinese cabbage and spinach species was capable of reducing the adverse effect of arsenic (As) accumulation by increasing the function of the arsenate reductase pathway. Redundancy analysis (RDA) indicated that the relative abundance of the core microbiome can be directly and indirectly influenced by certain environmental determinants. These short-term findings stress the importance of studying cover cropping systems as an efficient biological tool to protect the ecological environment. Therefore, we can speculate that leafy crop diversification is socially acceptable, economically justifiable, and ecologically adaptable to meet the urgent demand for intensive cropping systems to promote positive feedback between crop-soil sustainable intensification.

Garlic substrate could influence plant growth through affecting soil microbiome structure. The relationship mechanism between changes in soil microbial communities, disease suppression and plant development, however, remains unclear, particularly in the degraded soil micro-ecological environment. In this study, garlic substrates as a soil amendment were incorporated with different ratios (1:100, 3:100 and 5:100 g/100 g of soil) in a replanted disturbed soil of long-term cucumber monoculture (annual double cropping system in a greenhouse). The results indicated that higher amount of C-amended garlic substrate significantly induced soil suppressiveness (35.9% greater than control (CK) against the foliar disease incidence rate. This inhibitory effect consequently improved the cucumber growth performance and fruit yield to 20% higher than the non-amended soil. Short-term garlic substrate addition modified the soil quality through an increase in soil organic matter (SOM), nutrient availability and enzymatic activities. Illumina MiSeq sequencing analysis revealed that soil bacterial and fungal communities in the garlic amendment were significantly different from the control. Species richness and diversity indices significantly increased under treated soil. The correlation-based heat map analysis suggested that soil OM, nutrient contents and biological activators were the primary drivers reshaping the microbial community structure. Furthermore, garlic substrate inhibited soil-borne pathogen taxa (Fusarium and Nematoda), and their reduced abundances, significantly affecting the crop yield. In addition, the host plant recruited certain plant-beneficial microbes due to substrate addition that could directly contribute to plant-pathogen inhibition and crop biomass production. For example, abundant Acidobacteria, Ascomycota and Glomeromycota taxa were significantly associated with cucumber yield promotion. Firmicutes, Actinobacteria, Bacteroidetes, Basidiomycota and Glomeromycota were the associated microbial taxa that possibly performed as antagonists of Fusarium wilt, with plant pathogen suppression potential in monocropped cucumber-planted soil.

Amylases take part with vital role in industries such as food, fermentation; starch processing, textile and paper etc. Increasing amylases demand, high nutrient expenditure and environmental pollution have forced to utilize agro-industrial residues as a low-cost feedstock for enzyme production. In present study, three soil samples were collected from agro-industrial waste dumping areas in District Faisalabad. Ten thermophilic bacterial isolates were separated at 55 °C on the basis of colonial morphology, three isolates (F6, F11, F17) showed prominent zone of clearance applying iodine test on starch agar plates. Bacterial isolate F-11 showed highest amylase activity with DNS method and molecularly identified through 16S RNA sequencing as Bacillus sp. with Accession number MH917294. Four unconventional food wastes (banana, lemon, mango and potato) pretreated with 0.8% sulphuric acid concentrations taking 1000 g/L weight released the highest sugars contents and phenolic components. Maximum amylase activity i.e. 29.23 mg/ml was achieved in mango waste at, 40 °C, with pH 6.0 and 0.17% nitrogenous source adding 8% inoculum size (2 days old) using Response Surface Methodology (RSM) for optimization. Crude amylase confirmed its efficiency in starch hydrolysis that suggested it as potential candidate for application in starch industries.

Proteus mirabilis is commonly considered to be an opportunistic pathogen causing urinary tract infections (UTIs) in humans. However, some strains of P. mirabilis were found to be associated with food poisoning outbreaks, with the pathogenic mechanism still unclear. In our study, we described a novel strain of P. mirabilis C02011 isolated from patients' specimens in a food poisoning in China. In order to determine its gastrointestinal pathogenicity, experiments were performed to compare P. mirabilis B02005 strain (isolated from healthy people) and P. mirabilis American Type Culture Collection (ATCC) 29906 strain both in vitro [Caco-2 cells: bacterial adhesion and invasion assays, Giemsa staining, and transmission electron microscopy (TEM)] and in vivo [BALB/c mouse model: fecal character, colon injury, histological examination, immunochemistry, and western blotting (WB)]. According to the results, C02011 strain exhibited almost identical characteristics with B02005 strain in bacterial appearance and proliferation. In vitro, Caco-2 cells were infected with P. mirabilis C02011, B02005, and P. mirabilis ATCC 29906 strains. After that, Giemsa staining and TEM were used for observing the infection process of C02011 strain. Meanwhile, the adhesive abilities of different strains were rated as follows: P. mirabilis B02005 > P. mirabilis C02011 > P. mirabilis ATCC 29906 (P < 0.01). Invasive abilities of different strains were rated as follows: P. mirabilis C02011 > P. mirabilis B02005 > P. mirabilis ATCC 29906 (P < 0.01). In vivo, BALB/c mice were infected with P. mirabilis C02011 and B02005 strains. C02011 strain shows more virulence than B02005 strain in terms of the following indicators: (1) feces water content and fecal character; (2) colon length of mice; (3) histological examination on mouse intestine tissues; (4) ELISA for detecting TNF-α level in the colon; and (5) WB and immunohistochemistry (IHC) for detecting occludin protein expression in the colon. On the basis of these results, we firstly validated that the novel strain of P. mirabilis C02011 shows more gastrointestinal pathogenicity than the other strains isolated from a healthy individual. In addition, type IV secretion system (T4SS) was preliminarily confirmed to play an important role in the pathogenesis of diarrheal P. mirabilis isolated from the food poisoning incident.

Regulatory T (Treg) cells play a negative role in anti-tumor immunity against triple-negative breast cancer, so it is of great significance to find the potential therapeutic target of Treg cells.

Human muscarinic receptor M4 belongs to the class A subfamily of the G-protein-coupled receptors (GPCRs). M4 has emerged as an attractive drug target for the treatment of Alzheimer's disease and schizophrenia. Recent results showed that M4-mediated cholinergic transmission is related to motor symptoms in Parkinson's disease. Selective ligand design for the five muscarinic acetylcholine receptor (mAchR) subtypes currently remains challenging owing to the high sequence and structural similarity of their orthosteric binding pockets. In order to obtain M4-selective antagonists, a new approach was tried to lock M4 into an inactive form by rationally designing an N4497.49R mutation, which mimics the allosteric sodium binding in the conserved sodium site usually found in class A GPCRs. In addition, the crystal structure of the mutation-induced inactive M4 was determined. By comparative analysis with other mAchR structures, followed by functional assays, the N4497.49R mutation was shown to stabilize M4 into an inactive state. Virtual screening of a focused ligand library using the crystal structure showed that the inactive M4 prefers antagonists much more than agonists. This study provides a powerful mutation strategy to stabilize GPCRs in inactive states and facilitate their structure determination.

Collagen type X alpha 1 (COL10A1) is overexpressed in diverse tumors and displays vital roles in tumorigenesis. However, the prognostic value of COL10A1 in breast cancer remains unclear.

Endocrine disrupting chemicals (EDCs) are xenobiotics that mimic the interaction of natural hormones and alter synthesis, transport, or metabolic pathways. The prospect of EDCs causing adverse health effects in humans and wildlife has led to the development of scientific and regulatory approaches for evaluating bioactivity. This need is being addressed using high-throughput screening (HTS) in vitro approaches and computational modeling.

Pathogenic Vibrio spp., Aeromonas spp. and Plesiomonas shigelloides are associated with human gastroenteritis and wound infections, as well as fish diseases. The comprehensive and accurate identification of these pathogens is crucial for the current public health. The present study describes the development of a multiplex assay for the simultaneous identification of ten bacterial pathogens in a single reaction by using a multiplex ligation reaction based on probe melting curve analysis (MLMA). The specificity for target genes was 100%, as assessed with a panel of 67 bacterial pathogens, which indicated no cross-reactions. The detection limit of this assay ranged from 0.8 × 107 CFU/mL to 1.5 × 108 CFU/mL at the pure bacterial culture level and from 0.1 ng to 1.0 ng at the DNA level. The MLMA assay was used to detect ten species of pathogens in 269 clinical and seafood samples, and for further validation, the results were compared with the conventional culture method. The results indicated greater than 90% sensitivity and 100% specificity for each bacterial pathogen tested, and the kappa correlation for all the pathogens ranged from 0.95 to 1.00. Overall, this assay is well suited for public health laboratories for its high throughput, accuracy, and low cost.

Infections by Pseudomonas aeruginosa are difficult to cure due to its high intrinsic and acquired antibiotic resistance. Once colonized the human host, and thanks to antibiotic treatment pressure, P. aeruginosa usually acquires genetic mutations which provide bacteria with antibiotic resistance as well as ability to better adapt to the host environment. Deciphering the evolutionary traits may provide important insights into the development of effective combinatory antibiotic therapy to treat P. aeruginosa infections. In this study, we investigated the molecular mechanisms by which a clinical isolate (ISP50) yields a carbapenem-resistant derivative (IRP41). RNAseq and genomic DNA reference mapping were conducted to compare the transcriptional profiles and in vivo evolutionary trajectories between the two isolates. Our results demonstrated that oprD mutation together with ampC hyper-expression contributed to the increased resistance to carbapenem in the isolate IRP41. Furthermore, a ldcA (PA5198) gene, encoding murein tetrapeptide carboxypeptidase, has been demonstrated for the first time to negatively influence the ampC expression in P. aeruginosa.

Polymyxin B and E (colistin) are the last resorts to treat multidrug-resistant Gram-negative pathogens. Pseudomonas aeruginosa is intrinsically resistant to a variety of antibiotics. The PhoP-PhoQ two-component regulatory system contributes to the resistance to polymyxins by regulating an arnBCADTEF-pmrE operon that encodes lipopolysaccharide modification enzymes. To identify additional PhoP-regulated genes that contribute to the tolerance to polymyxin B, we performed a chromatin immunoprecipitation sequencing (ChIP-Seq) assay and found novel PhoP binding sites on the chromosome. We further verified that PhoP directly controls the expression of PA14_46900, PA14_50740 and PA14_52340, and the operons of PA14_11970-PA14_11960 and PA14_52350-PA14_52370. Our results demonstrated that mutation of PA14_46900 increased the bacterial binding and susceptibility to polymyxin B. Meanwhile, mutation of PA14_11960 (papP), PA14_11970 (mpl), PA14_50740 (slyB), PA14_52350 (ppgS), and PA14_52370 (ppgH) reduced the bacterial survival rates and increased ethidium bromide influx under polymyxin B or Sodium dodecyl sulfate (SDS) treatment, indicating roles of these genes in maintaining membrane integrity in response to the stresses. By 1-N-phenylnaphthylamine (NPN) and propidium iodide (PI) staining assay, we found that papP and slyB are involved in maintaining outer membrane integrity, and mpl and ppgS-ppgH are involved in maintaining inner membrane integrity. Overall, our results reveal novel PhoP-PhoQ regulated genes that contribute to polymyxin B tolerance.

Therapy for Pseudomonas aeruginosa infections is hard due to its high natural and acquirable antibiotic resistance. After colonization in the hosts, P. aeruginosa commonly accumulates genomic mutations which confer them antibiotic resistance and better adaptations to the host environment. Deciphering the mechanisms of antibiotic resistance development in the clinical setting may provide critical insights into the design of effective combinatory antibiotic therapies to treat P. aeruginosa infections. In this work, we demonstrate a resistance mechanism to aztreonam of a clinical isolate (ARP36) in comparison with a sensitive one (CSP18). RNAseq and genomic DNA resequencing were carried out to compare the global transcriptional profiles and in the clinical setting genomic profiles between these two isolates. The results demonstrated that hyperexpression of an efflux pump MexAB-OprM caused by a R70Q substitution in MexR, contributed to the increased resistance to aztreonam in the isolate ARP36. Simulation of mexR of ARP36 by gene editing in CSP18 conferred CSP18 an ARP36-like susceptibility to the aztreonam. The R70Q substitution prevented MexR from binding to the intergenic region between mexR and mexAB-oprM operon, with no impact on its dimerization. The presented experimental results explain for the first time why the clinically relevant R70Q substitution in the MexR derepresses the expression of mexAB-oprM in P. aeruginosa.

Chimeric antigen receptor (CAR) T cells are powerful in eradicating hematological malignancies, but their efficacy is limited in treating solid tumors. One of the barriers is the immunosuppressive response induced by immunomodulatory signaling pathways. Pharmacological targeting of these immunosuppressive pathways may be a simple way to improve the efficacy of CAR T cells. In this study, anti-CD133 and anti-HER2 CAR T cells were generated from healthy donors, and combination therapy using CAR T cells and small molecules targeting adenosine receptors was performed in vitro and in vivo with the goal of probing for potential synergistic antitumor activities. The adenosine A2b receptor agonist, BAY 60-6583, was found to significantly increase cytokine secretion of CD133-or HER2-specific CAR T cells when co-cultured with the respective target tumor cells. The in vitro cytotoxicity and proliferation of CAR T cells were also enhanced when supplied with BAY 60-6583. Furthermore, the combination with this small molecule facilitated the anti-HER2 CAR T cell-mediated elimination of tumor cells in a xenograft mouse model. However, the enhanced antitumor activities could not be suppressed by knockout of the adenosine A2b receptor in CAR T cells. Furthermore, mass spectrometry and computational methods were used to predict several potential alternative targets. Four potential targets (pyruvate kinase M (PKM), Talin-1, Plastin-2, and lamina-associated polypeptide 2) were captured by a photo-affinity probe, of which PKM and Talin-1 were predicted to interact with BAY 60-6583. Overall, our data suggest that BAY 60-6583 upregulates T cell functions through a mechanism independent of the adenosine A2b receptor.

The COVID-19, caused by SARS-CoV-2, is threatening public health, and there is no effective treatment. In this study, we have implemented a multi-targeted anti-viral drug design strategy to discover highly potent SARS-CoV-2 inhibitors, which simultaneously act on the host ribosome, viral RNA as well as RNA-dependent RNA polymerases, and nucleocapsid protein of the virus, to impair viral translation, frameshifting, replication, and assembly. Driven by this strategy, three alkaloids, including lycorine, emetine, and cephaeline, were discovered to inhibit SARS-CoV-2 with EC50 values of low nanomolar levels potently. The findings in this work demonstrate the feasibility of this multi-targeting drug design strategy and provide a rationale for designing more potent anti-virus drugs.

NrtR is a Nudix-related transcriptional regulator that is distributed among diverse bacteria and plays an important role in modulating bacterial intracellular NAD homeostasis. Previously, we showed that NrtR influences the T3SS expression and pathogenesis of Pseudomonas aeruginosa and demonstrated that NrtR mediates T3SS regulation through the cAMP/Vfr pathway. In the present study, we found that mutation of the nrtR gene leads to upregulation of the Hcp secretion island-I type VI secretion system (H1-T6SS). Further analysis revealed that mutation of the nrtR gene results in upregulation of regulatory RNAs (RsmY/RsmZ) that are known to control the H1-T6SS by sequestration of RsmA or RsmN. Simultaneous deletion of rsmY/rsmZ reduced the expression of H1-T6SS in the ΔnrtR mutant. In addition, overexpression of either rsmA or rsmN in ΔnrtR decreased H1-T6SS expression. Chromatin immunoprecipitation (ChIP)-Seq and electrophoretic mobility shift assay (EMSA) analyses revealed that NrtR directly binds to the promoters of rsmY, rsmZ and tssA1 (first gene of the H1-T6SS operon). Overall, the results from this study reveal the molecular details of NrtR-mediated regulation of H1-T6SS in P. aeruginosa. IMPORTANCE NrtR is a Nudix-related transcriptional regulator and controls the NAD cofactor biosynthesis in bacteria. P. aeruginosa NrtR binds to the intergenic region between nadD2 and pcnA to repress the expression of the two operons, therefore controlling the NAD biosynthesis. We have previously reported that NrtR controls T3SS expression via the cAMP/Vfr pathway in P. aeruginosa. However, the global regulatory function and direct binding targets of the NrtR remain elusive in P. aeruginosa. This study reveals novel direct regulatory targets of the NrtR in P. aeruginosa, elucidating the molecular mechanism of NrtR-mediated regulation of H1-T6SS.