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Chemosphere Dec 2023Roxarsone (3-nitro-4-hydroxyphenylarsonic acid, Rox), a widely used organoarsenical feed additive, can enter soils and be further biotransformed into various arsenic...
Roxarsone (3-nitro-4-hydroxyphenylarsonic acid, Rox), a widely used organoarsenical feed additive, can enter soils and be further biotransformed into various arsenic species that pose human health and ecological risks. However, the pathway and molecular mechanism of Rox biotransformation by soil microbes are not well studied. Therefore, in this study, we isolated a Rox-transforming bacterium from manure-fertilized soil and identified it as Pseudomonas chlororaphis through morphological analysis and 16S rRNA gene sequencing. Pseudomonas chlororaphis was able to biotransform Rox to 3-amino-4-hydroxyphenylarsonic acid (3-AHPAA), N-acetyl-4-hydroxy-m-arsanilic acid (N-AHPAA), arsenate [As(V)], arsenite [As(III)], and dimethylarsenate [DMAs(V)]. The complete genome of Pseudomonas chlororaphis was sequenced. PcmdaB, encoding a nitroreductase, and PcnhoA, encoding an acetyltransferase, were identified in the genome of Pseudomonas chlororaphis. Expression of PcmdaB and PcnhoA in E. coli Rosetta was shown to confer Rox(III) and 3-AHPAA(III) resistance through Rox nitroreduction and 3-AHPAA acetylation, respectively. The PcMdaB and PcNhoA enzymes were further purified and functionally characterized in vitro. The kinetic data of both PcMdaB and PcNhoA were well fit to the Michaelis-Menten equation, and nitroreduction catalyzed by PcMdaB is the rate-limiting step for Rox transformation. Our results provide new insights into the environmental risk assessment and bioremediation of Rox(V)-contaminated soils.
Topics: Humans; Roxarsone; Pseudomonas chlororaphis; Soil; Acetyltransferases; RNA, Ribosomal, 16S; Escherichia coli; Arsenic; Biotransformation; Nitroreductases
PubMed: 37898462
DOI: 10.1016/j.chemosphere.2023.140558 -
Microbial Biotechnology Nov 2023Virulence factor modulating (VFM) is a quorum sensing (QS) signal shared by and specific to Dickeya bacteria, regulating the production of plant cell wall degrading...
Virulence factor modulating (VFM) is a quorum sensing (QS) signal shared by and specific to Dickeya bacteria, regulating the production of plant cell wall degrading enzymes (PCWDEs) and virulence of Dickeya. High polarity and trace of VFM signal increase the difficulty of signal separation and structure identification, and thus limit the development of quorum quenching strategy to biocontrol bacterial soft rot diseases caused by Dickeya. In order to high-throughput screen VFM quenching bacteria, a vfmE-gfp biosensor VR2 (VFM Reporter) sensitive to VFM signal was first constructed. Subsequently, two bacterial strains with high quenching efficiency were screened out by fluorescence intensity measurement and identified as Pseudomonas chlororaphis L5 and Enterobacter asburiae L95 using multilocus sequence analysis (MLSA). L5 and L95 supernatants reduced the expression of vfm genes, and both strains also decreased the production of PCWDEs of D. zeae MS2 and significantly reduced the virulence of D. oryzae EC1 on rice seedlings, D. zeae MS2 on banana seedlings, D. dadantii 3937 on potato and D. fangzhongdai CL3 on taro. Findings in this study provide a method to high-throughput screen VFM quenching bacteria and characterize novel functions of P. chlororaphis and E. asburiae in biocontrolling plant diseases through quenching VFM QS signal.
Topics: Virulence Factors; Dickeya; Quorum Sensing; Pseudomonas chlororaphis; Enterobacteriaceae; Plant Diseases
PubMed: 37815509
DOI: 10.1111/1751-7915.14351 -
Plants (Basel, Switzerland) Apr 2024Soil-borne spp. have been extensively studied for their biocontrol activities against pathogens and growth promotion ability in plants. However, the beneficial effect...
Soil-borne spp. have been extensively studied for their biocontrol activities against pathogens and growth promotion ability in plants. However, the beneficial effect of on inducing resistance against insect herbivores has been underexplored. Among diverse species, consistent with previous reports, we showed that root colonization by triggered induced systemic resistance (ISR) to the leaf-infecting hemibiotrophic fungal pathogens . Whether induces ISR to insect pests has not been tested before. In this study, we investigated whether affects jasmonic acid (JA) biosynthesis and defense against fall armyworm (FAW) and western corn rootworm (WCR). Unexpectedly, the results showed that colonization of maize seedlings grown in autoclaved soil suppressed wound-induced production of JA, resulting in reduced resistance to FAW. Similarly, the bacterial endophyte 30-84 was found to suppress systemic resistance to FAW due to reduced JA. Further comparative analyses of the systemic effects of these endophytes when applied in sterile or non-sterile field soil showed that both and 30-84 triggered ISR against C. in both soil conditions, but only suppressed JA production and resistance to FAW in sterile soil, while no significant impact was observed when applied in non-sterile soil. In contrast to the effect on FAW defense, colonization of maize roots suppressed WCR larvae survival and weight gain. This is the first report suggesting the potential role of as a biocontrol agent against WCR.
PubMed: 38732455
DOI: 10.3390/plants13091240 -
Journal of Agricultural and Food... Apr 2024Polyhydroxyalkanoates (PHAs) are promising alternatives to petroleum-based plastics, owing to their biodegradability and superior material properties. Here, the...
Polyhydroxyalkanoates (PHAs) are promising alternatives to petroleum-based plastics, owing to their biodegradability and superior material properties. Here, the controllable biosynthesis of scl--mcl PHA containing 3-hydroxybutyrate (3HB) and mcl 3-hydroxyalkanoates was achieved in HT66. First, key genes involved in fatty acid β-oxidation, the fatty acid biosynthesis pathway, and the -- operon were deleted to develop a chassis strain. Subsequently, an acetoacetyl-CoA reductase gene and a PHA synthase gene with broad substrate specificity were heterologously expressed for producing and polymerizing the 3HB monomer with mcl 3-hydroxyalkanoates under the assistance of native β-ketothiolase gene . Furthermore, the monomer composition of scl--mcl PHA was regulated by adjusting the amount of glucose and dodecanoic acid supplemented. Notably, the cell dry weight and scl--mcl PHA content reached 14.2 g/L and 60.1 wt %, respectively, when the engineered strain HT11Δ:: was cultured in King's B medium containing 5 g/L glucose and 5 g/L dodecanoic acid. These results demonstrated that can be a platform for producing scl--mcl PHA and has the potential for industrial application.
Topics: 3-Hydroxybutyric Acid; Pseudomonas chlororaphis; Acyltransferases; Polyhydroxyalkanoates; Glucose
PubMed: 38564621
DOI: 10.1021/acs.jafc.4c00777 -
Frontiers in Microbiology 2024The rhizosphere microbiome is critical to plant health and resistance. PGPR are well known as plant-beneficial bacteria and generally regulate nutrient utilization as...
INTRODUCTION
The rhizosphere microbiome is critical to plant health and resistance. PGPR are well known as plant-beneficial bacteria and generally regulate nutrient utilization as well as plant responses to environmental stimuli. In our previous work, one typical PGPR strain, IRHB3, isolated from the soybean rhizosphere, had positive impacts on soil-borne disease suppression and growth promotion in the greenhouse, but its biocontrol mechanism and application in the field are not unclear.
METHODS
In the current study, IRHB3 was introduced into field soil, and its effects on the local rhizosphere microbiome, disease resistance, and soybean growth were comprehensively analyzed through high-throughput sequencing and physiological and molecular methods.
RESULTS AND DISCUSSION
We found that IRHB3 significantly increased the richness of the bacterial community but not the structure of the soybean rhizosphere. Functional bacteria related to phosphorus solubilization and nitrogen fixation, such as , , , , and , were recruited in rich abundance by IRHB3 to the soybean rhizosphere as compared to those without IRHB3. In addition, the IRHB3 supplement obviously maintained the homeostasis of the rhizosphere microbiome that was disturbed by , resulting in a lower disease index of root rot when compared with . Furthermore, JA-mediated induced resistance was rapidly activated by IRHB3 following and expression, and meanwhile, a set of nodulation genes, , , and , were also considerably induced by IRHB3 to improve nitrogen fixation ability and promote soybean yield, even when plants were infected by . Thus, IRHB3 tends to synergistically interact with local rhizosphere microbes to promote host growth and induce host resistance in the field.
PubMed: 38380096
DOI: 10.3389/fmicb.2024.1328863 -
Microbial Biotechnology Jan 2024Rhamnolipids (RL) are biosurfactants naturally produced by the opportunistic pathogen Pseudomonas aeruginosa. Currently, RL are commercialized for various applications...
Rhamnolipids (RL) are biosurfactants naturally produced by the opportunistic pathogen Pseudomonas aeruginosa. Currently, RL are commercialized for various applications and produced by Pseudomonas putida due to the health risks associated with their large-scale production by P. aeruginosa. In this work, we show that RL containing one or two rhamnose moieties (mono-RL or di-RL, respectively) can be produced by the innocuous soil-bacterium Pseudomonas chlororaphis subsp chlororaphis ATCC 9446 at titres up to 66 mg/L (about 86% of the production of P. aeruginosa PAO1 in the same culture conditions). The production of RL depends on the expression of P. aeruginosa PAO1 genes encoding the enzymes RhlA, RhlB and RhlC. These genes were introduced in a plasmid, together with a transcriptional regulator (rhlR) forming part of the same operon, with and without RhlC. We show that the activation of rhlAB by RhlR depends on its interaction with P. chlororaphis endogenous acyl-homoserine lactones, which are synthetized by either PhzI or CsaI autoinducer synthases (producing 3-hydroxy-hexanoyl homoserine lactone, 3OH-C6-HSL, or 3-oxo-hexanoyl homoserine lactone, 3O-C6-HSL, respectively). P. chlororaphis transcriptional regulator couple with 3OH-C6-HSL is the primary activator of gene expression for phenazine-1-carboxylic acid (PCA) and phenazine-1-carboxamide (PCN) production in this soil bacterium. We show that RhlR coupled with 3OH-C6-HSL or 3O-C6-HSL promotes RL production and increases the production of PCA in P. chlororaphis. However, PhzR/3OH-C6-HSL or CsaR/3O-C6-HSL cannot activate the expression of the rhlAB operon to produce mono-RL. These results reveal a complex regulatory interaction between RhlR and P. chlororaphis quorum-sensing signals and highlight the biotechnology potential of P. chlororaphis ATCC 9446 expressing P. aeruginosa rhlAB-R or rhlAB-R-C for the industrial production of RL.
Topics: Pseudomonas chlororaphis; Acyl-Butyrolactones; Pseudomonas aeruginosa; Soil; Bacterial Proteins; Glycolipids; Pseudomonas; 4-Butyrolactone
PubMed: 38041625
DOI: 10.1111/1751-7915.14377 -
ACS Synthetic Biology Mar 2024The advancement of metabolic engineering and synthetic biology has promoted in-depth research on the nonmodel microbial metabolism, and the potential of nonmodel...
The advancement of metabolic engineering and synthetic biology has promoted in-depth research on the nonmodel microbial metabolism, and the potential of nonmodel organisms in industrial biotechnology is becoming increasingly evident. The nonmodel organism is a safe plant growth promoting bacterium for the production of phenazine compounds; however, its application is seriously hindered due to the lack of an effective gene expression precise regulation toolkit. In this study, we constructed a library of 108 promoter-5'-UTR (PUTR) and characterized them through fluorescent protein detection. Then, 6 PUTRs with stable low, intermediate, and high intensities were further characterized by report genes encoding β-galactosidase from K12 and encoding PCA monooxygenase from GP72 and thus developed as a static gene expression regulation system. Furthermore, the stable and high-intensity expressed PUTR was fused with the LacO operator to construct an IPTG-induced plasmid, and a self-induced plasmid was constructed employing the high-intensity PUTR regulated by cell density, resulting in a dynamic gene expression regulation system. In summary, this study established two sets of static and dynamic regulatory systems for , providing an effective toolkit for fine-tuning gene expression and reprograming the metabolism flux.
Topics: Pseudomonas chlororaphis; Metabolic Engineering; Gene Expression Regulation, Bacterial; Promoter Regions, Genetic; Bacterial Proteins
PubMed: 38377538
DOI: 10.1021/acssynbio.3c00714 -
JACS Au Nov 2023Chemical fertilizers have been crucial for sustaining the current global population by supplementing overused farmland to support consistent food production, but their...
Chemical fertilizers have been crucial for sustaining the current global population by supplementing overused farmland to support consistent food production, but their use is unsustainable. is a nitrogen-fixing bacterium that could be used as a fertilizer replacement, but this microbe is delicate. It is sensitive to stressors, such as freeze-drying and high temperatures. Here, we demonstrate protection of from freeze-drying, high temperatures (50 C), and high humidity using self-assembling metal-phenolic network (MPN) coatings. The composition of the MPN is found to significantly impact its protective efficacy, and with optimized compositions, no viability loss is observed for MPN-coated microbes under conditions where uncoated cells do not survive. Further, we demonstrate that MPN-coated microbes improve germination of seeds by 150% as compared to those treated with fresh . Taken together, these results demonstrate the protective capabilities of MPNs against environmental stressors and represent a critical step towards enabling the production and storage of delicate microbes under nonideal conditions.
PubMed: 38034965
DOI: 10.1021/jacsau.3c00426 -
Plants (Basel, Switzerland) Sep 2023The role of Calcium ions (Ca) is extensively documented and comprehensively understood in eukaryotic organisms. Nevertheless, emerging insights, primarily derived from... (Review)
Review
The role of Calcium ions (Ca) is extensively documented and comprehensively understood in eukaryotic organisms. Nevertheless, emerging insights, primarily derived from studies on human pathogenic bacteria, suggest that this ion also plays a pivotal role in prokaryotes. In this review, our primary focus will be on unraveling the intricate Ca toolkit within prokaryotic organisms, with particular emphasis on its implications for plant growth-promoting rhizobacteria (PGPR). We undertook an in silico exploration to pinpoint and identify some of the proteins described in the existing literature, including prokaryotic Ca channels, pumps, and exchangers that are responsible for regulating intracellular Calcium concentration ([Ca]), along with the Calcium-binding proteins (CaBPs) that play a pivotal role in sensing and transducing this essential cation. These investigations were conducted in four distinct PGPR strains: subsp. SMMP3, SVBP6, sp. BP01, and sp. 2A, which have been isolated and characterized within our research laboratories. We also present preliminary experimental data to evaluate the influence of exogenous Ca concentrations ([Ca]) on the growth dynamics of these strains.
PubMed: 37836138
DOI: 10.3390/plants12193398 -
Phytopathology Jun 2024Mixtures of fungicides with different modes of action are commonly used as disease and resistance management tools, but little is known of mixtures of natural and...
Mixtures of fungicides with different modes of action are commonly used as disease and resistance management tools, but little is known of mixtures of natural and synthetic products. In this study, mixtures of metabolites from the rhizobacterium Pseudomonas chlororaphis strain ASF009 formulated as Howler EVO with below label rates (50 µg/ml) of conventional sterol demethylation inhibitor (DMI) fungicides were investigated for control of anthracnose of cherry (Prunus avium) caused by Colletotrichum siamense. Howler mixed with metconazole or propiconazole synergistically reduced disease severity through lesion growth. Realtime PCR showed that difenoconazole, flutriafol, metconazole, and propiconazole induced the expression of DMI target genes CsCYP51A and CsCYP51B in C. siamense. The addition of Howler completely suppressed the DMI fungicide-induced expression of both CYP51 genes. We hypothesize that the downregulation of DMI fungicide-induced expression of the DMI target genes may, at least in part, explain the synergism observed in detached fruit assays.
PubMed: 38857059
DOI: 10.1094/PHYTO-03-24-0090-R