Optimization, gene cloning, expression, and molecular docking insights for enhanced cellulase enzyme production by Bacillus amyloliquefaciens strain elh1.

BACKGROUND: In this study, we isolated a cellulase-producing bacterium, Bacillus amyloliquefaciens strain elh, from rice peel. We employed two optimization methods to enhance the yield of cellulase. Firstly, we utilized a one-variable-at-a-time (OVAT) approach to evaluate the impact of individual physical and chemical parameters. Subsequently, we employed response surface methodology (RSM) to investigate the interactions among these factors. We heterologously expressed the cellulase encoding gene using a cloning vectorin E. coli DH5α. Moreover, we conducted in silico molecular docking analysis to analyze the interaction between cellulase and carboxymethyl cellulose as a substrate. RESULTS: The bacterial isolate eh1 exhibited an initial cellulase activity of 0.141 ± 0.077 U/ml when cultured in a specific medium, namely Basic Liquid Media (BLM), with rice peel as a substrate. This strain was identified as Bacillus amyloliquefaciens strain elh1 through 16S rRNA sequencing, assigned the accession number OR920278 in GenBank. The optimal incubation time was found to be 72 h of fermentation. Urea was identified as the most suitable nitrogen source, and dextrose as the optimal sugar, resulting in a production increase to 5.04 ± 0.120 U/ml. The peak activity of cellulase reached 14.04 ± 0.42 U/ml utilizing statistical optimization using Response Surface Methodology (RSM). This process comprised an initial screening utilizing the Plackett-Burman design and further refinement employing the BOX -Behnken Design. The gene responsible for cellulase production, egl, was effectively cloned and expressed in E. coli DH5α. The transformed cells exhibited a cellulase activity of 22.3 ± 0.24 U/ml. The egl gene sequence was deposited in GenBank with the accession number PP194445. In silico molecular docking revealed that the two hydroxyl groups of carboxymethyl cellulose bind to the residues of Glu169 inside the binding pocket of the CMCase. This interaction forms two hydrogen bonds, with an affinity score of -5.71. CONCLUSIONS: Optimization of cultural conditions significantly enhances the yield of cellulase enzyme when compared to unoptimized culturing conditions. Additionally, heterologous expression of egl gene showed that the recombinant form of the cellulase is active and that a valid expression system can contribute to a better yield of the enzyme.

Phosphatidylethanolamines link ferroptosis and autophagy during appressorium formation of rice blast fungus.

A cell death pathway, ferroptosis, occurs in conidial cells and is critical for formation and function of the infection structure, the appressorium, in the rice blast fungus Magnaporthe oryzae. In this study, we identified an orthologous lysophosphatidic acid acyltransferase (Lpaat) acting at upstream of phosphatidylethanolamines (PEs) biosynthesis and which is required for such fungal ferroptosis and pathogenicity. Two PE species, DOPE and SLPE, that depend on Lpaat function for production were sufficient for induction of lipid peroxidation and the consequent ferroptosis, thus positively regulating fungal pathogenicity. On the other hand, both DOPE and SLPE positively regulated autophagy. Loss of the LPAAT gene led to a decrease in the lipidated form of the autophagy protein Atg8, which is probably responsible for the autophagy defect of the lpaatΔ mutant. GFP-Lpaat was mostly localized on the membrane of lipid droplets (LDs) that were stained by the fluorescent dye monodansylpentane (MDH), suggesting that LDs serve as a source of lipids for membrane PE biosynthesis and probably as a membrane source of autophagosome. Overall, our results reveal novel intracellular membrane-bound organelle dynamics based on Lpaat-mediated lipid metabolism, providing a temporal and spatial link of ferroptosis and autophagy.

The ACE1 secondary metabolite gene cluster is a pathogenicity factor of wheat blast fungus.

Wheat blast caused by Pyricularia oryzae pathotype Triticum is now becoming a very serious threat to global food security. Here, we report an essential pathogenicity factor of the wheat blast fungus that is recognized and may be targeted by a rice resistance gene. Map-based cloning of Pwt2 showed that its functional allele is the ACE1 secondary metabolite gene cluster of the wheat blast fungus required for its efficient penetration of wheat cell walls. ACE1 is required for the strong aggressiveness of Triticum, Eleusine, and Lolium pathotypes on their respective hosts, but not for that of Oryza and Setaria pathotypes on rice and foxtail millet, respectively. All ACE1 alleles found in wheat blast population are recognized by a rice resistance gene, Pi33, when introduced into rice blast isolates. ACE1 mutations for evading the recognition by Pi33 do not affect the aggressiveness of the rice blast fungus on rice but inevitably impair the aggressiveness of the wheat blast fungus on wheat. These results suggest that a blast resistance gene already defeated in rice may be revived as a durable resistance gene in wheat by targeting an Achilles heel of the wheat blast fungus.

Enhancing Cordycepin in Functional Porridge through Rice Varietal Fermentation with Cordyceps militaris (Ascomycetes) and Utilizing Non-Targeted Metabolomics for Process Enhancement.

Cordyceps militaris, a medicinal fungus rich in cordycepin, shows promise in treating diseases such as cancer, respiratory issues, and COVID-19. This study examines the impact of different Taiwanese rice varieties on its solid-state fermentation, focusing on optimizing cordycepin production. The results indicated that the cordycepin yield was indeed affected by the type of rice used. In terms of the fruiting bodies, germ rice resulted in the highest yield (13.1 ± 0.36 mg/g), followed by brown rice (11.9 ± 0.26 mg/g). In the rice culture medium (RCM), brown rice led to the highest yield (4.77 ± 0.06 mg/g). Using gas chromatography-mass spectrometry and untargeted metabolomics, the study identifies four key volatile components linked to cordycepin, providing insights into developing functional rice porridge products. These findings are significant for advancing cordycepin mass production and offering dietary options for older individuals.

Hijacking of the methylglyoxal detoxification pathway: a new tactic of Xoo pathogenesis in rice.

Xanthomonas oryzae pv. oryzae (Xoo), the causative agent of bacterial blight (BB), has developed a unique strategy to infect rice by hijacking the host's methylglyoxal (MG) detoxification pathway. This results in an over-accumulation of MG, which facilitates tissue colonization and evasion of host's immune responses. While MG role in abiotic stresses is well-documented, its involvement in biotic stresses has not been extensively explored. Recently, Fu et al. (2024) provided the first evidence of MG role in promoting Xoo pathogenesis in rice. This new virulence strategy contributes to the pathogen's remarkable adaptability and survival. In this mechanism of hijacking of MG detoxification pathway, Xoo induces OsWRKY62.1 to inhibit OsGLY II expression, leading to MG overaccumulation in infected rice cells. This excess MG hinders plant cell organelle function, creating a favorable environment for Xoo by compromising the rice defense system. In this article, we have presented our perspectives on how the BB pathogen adapts its virulence mechanisms to infect and cause disease in rice.

Defining genomic landscape for identification of potential candidate resistance genes associated with major rice diseases through MetaQTL analysis.

Rice production is severely affected by various diseases such as bacterial leaf blight (BLB), brown spot (BS), false smut (FS), foot rot (FR), rice blast (RB), and sheath blight (SB). In recent years, several quantitative trait loci (QTLs) studies involving different populations have been carried out, resulting in the identification of hundreds of resistance QTLs for each disease. These QTLs can be integrated and analyzed using meta-QTL (MQTL) analysis for better understanding of the genetic architecture underlying multiple disease resistance (MDR). This study involved an MQTL analysis on 661 QTLs (378, 161, 21, 41, 44, and 16 QTLs for SB, RB, BLB, BS, FS, and FR, respectively) retrieved from 50 individual studies published from 1995 to 2021. Of these, 503 QTLs were projected finally onto the consensus map saturated with 6,275 markers, resulting in 73 MQTLs, including 27 MDR-MQTLs conferring resistance to three or more diseases. Forty-seven MQTLs were validated using marker-trait associations identified in published genome-wide association studies. A total of 3,310 genes, including both R and defense genes, were also identified within some selected high-confidence MQTL regions that were investigated further for the syntenic relationship with barley, wheat, and maize genomes. Thirty-nine high-confidence candidate genes were selected based on their expression patterns and recommended for future studies involving functional validation, genetic engineering, and gene editing. Nineteen MQTLs were co-localized with 39 known R genes for BLB and RB diseases. These results could pave the way to utilize candidate genes in a marker-assisted breeding program for MDR in rice.

A novel MAP kinase-interacting protein MoSmi1 regulates development and pathogenicity in Magnaporthe oryzae.

The cell wall is the first barrier against external adversity and plays roles in maintaining normal physiological functions of fungi. Previously, we reported a nucleosome assembly protein, MoNap1, in Magnaporthe oryzae that plays a role in cell wall integrity (CWI), stress response, and pathogenicity. Moreover, MoNap1 negatively regulates the expression of MoSMI1 encoded by MGG_03970. Here, we demonstrated that deletion of MoSMI1 resulted in a significant defect in appressorium function, CWI, cell morphology, and pathogenicity. Further investigation revealed that MoSmi1 interacted with MoOsm1 and MoMps1 and affected the phosphorylation levels of MoOsm1, MoMps1, and MoPmk1, suggesting that MoSmi1 regulates biological functions by mediating mitogen-activated protein kinase (MAPK) signalling pathway in M. oryzae. In addition, transcriptome data revealed that MoSmi1 regulates many infection-related processes in M. oryzae, such as membrane-related pathway and oxidation reduction process. In conclusion, our study demonstrated that MoSmi1 regulates CWI by mediating the MAPK pathway to affect development and pathogenicity of M. oryzae.

Identification of plant based potential antifungal compounds against BMK-1 protein of Bipolaris oryzae using molecular docking approach.

Rice brown spot is an important disease of rice worldwide that inflicts substantial yield losses. The antimicrobial potential of methanol, acetone and dimethyl sulfoxide (DMSO) extracts of different medicinal plants, viz., Syzygium aromaticum, Saussurea costus, Acorus calamus, Bergenia ciliate, Geranium pratense, Mentha longifolia, Inula racemosa, Podophyllum hexandrum, Heracleum candicans and Picrorhiza kurroa, against the brown spot pathogen Bipolaris oryzae in vitro was evaluated via mycelial growth inhibition and spore germination inhibition assays. Among the plant extracts tested, 100% mycelial inhibition was observed for the methanol extract of Syzygium aromaticum at all three concentrations (2000 ppm, 3000 ppm and 4000 ppm), followed by the methanol extract of Inula racemosa (90.33%) at 4000 ppm. A maximum conidial germination inhibition of 83.54% was exhibited by the Heracleum candicans leaf extract. Phytochemical profiling of Syzygium aromaticum and Inula racemosa through liquid chromatography and mass spectrometry (HR-LCMS) revealed the presence of several compounds, such as eugenol, ursolic acid, quercetin, chlorogenic acid, and noscapine. A molecular docking approach was used to identify key inhibitory molecules against B. oryzae. Among the compounds detected in S. aromaticum and Inula racemosa, ursolic acid and noscapine were found to have the greatest binding affinity for the Big Mitogen Activated Protein Kinase (BMK-1) enzyme present in B. oryzae. In conclusion, S. aromaticum and Inula racemosa are potent compounds that could serve as lead compounds for drug discovery in the future.

OsPUB9 Gene Edited by CRISPR/Cas9 Enhanced Resistance to Bacterial Leaf Blight in Rice (Oryza sativa L.).

Ubiquitination plays a crucial role in regulating signal pathways during the post-translation stage of protein synthesis in response to various environmental stresses. E3 ubiquitin ligase has been discovered to ultimately control various intracellular activities by imparting specificity to proteins to be degraded. This study was conducted to confirm biological and genetic functions of the U-box type E3 ubiquitin ligase (PUB) gene against biotic stress in rice (Oryza sativa L.). OsPUB9 gene-specific sgRNA were designed and transformants were developed through Agrobacterium-mediated transformation. Deep sequencing using callus was performed to confirm the mutation type of T0 plants, and a total of three steps were performed to select null individuals without T-DNA insertion. In the case of the OsPUB9 gene-edited line, a one bp insertion was generated by gene editing, and it was confirmed that early stop codon and multiple open reading frame (ORF) sites were created by inserting thymine. It is presumed that ubiquitination function also changed according to the change in protein structure of U-box E3 ubiquitin ligase. The OsPUB9 gene-edited null lines were inoculated with bacterial leaf blight, and finally confirmed to have a resistance phenotype similar to Jinbaek, a bacterial blight-resistant cultivar. Therefore, it is assumed that the amino acid sequence derived from the OsPUB9 gene is greatly changed, resulting in a loss of the original protein functions related to biological mechanisms. Comprehensively, it was confirmed that resistance to bacterial leaf blight stress was enhanced when a mutation occurred at a specific site of the OsPUB9 gene.

Dihydroorotase MoPyr4 is required for development, pathogenicity, and autophagy in rice blast fungus.

Dihydroorotase (DHOase) is the third enzyme in the six enzymatic reaction steps of the endogenous pyrimidine nucleotide de novo biosynthesis pathway, which is a metabolic pathway conserved in both bacteria and eukaryotes. However, research on the biological function of DHOase in plant pathogenic fungi is very limited. In this study, we identified and named MoPyr4, a homologous protein of Saccharomyces cerevisiae DHOase Ura4, in the rice blast fungus Magnaporthe oryzae and investigated its ability to regulate fungal growth, pathogenicity, and autophagy. Deletion of MoPYR4 led to defects in growth, conidiation, appressorium formation, the transfer and degradation of glycogen and lipid droplets, appressorium turgor accumulation, and invasive hypha expansion in M. oryzae, which eventually resulted in weakened fungal pathogenicity. Long-term replenishment of exogenous uridine-5'-phosphate (UMP) can effectively restore the phenotype and virulence of the ΔMopyr4 mutant. Further study revealed that MoPyr4 also participated in the regulation of the Pmk1-MAPK signaling pathway, co-localized with peroxisomes for the oxidative stress response, and was involved in the regulation of the Osm1-MAPK signaling pathway in response to hyperosmotic stress. In addition, MoPyr4 interacted with MoAtg5, the core protein involved in autophagy, and positively regulated autophagic degradation. Taken together, our results suggested that MoPyr4 for UMP biosynthesis was crucial for the development and pathogenicity of M. oryzae. We also revealed that MoPyr4 played an essential role in the external stress response and pathogenic mechanism through participation in the Pmk1-MAPK signaling pathway, peroxisome-related oxidative stress response mechanism, the Osm1-MAPK signaling pathway and the autophagy pathway.

Bioengineering a plant NLR immune receptor with a robust binding interface toward a conserved fungal pathogen effector.

Bioengineering of plant immune receptors has emerged as a key strategy for generating novel disease resistance traits to counteract the expanding threat of plant pathogens to global food security. However, current approaches are limited by rapid evolution of plant pathogens in the field and may lack durability when deployed. Here, we show that the rice nucleotide-binding, leucine-rich repeat (NLR) immune receptor Pik-1 can be engineered to respond to a conserved family of effectors from the multihost blast fungus pathogen Magnaporthe oryzae. We switched the effector binding and response profile of the Pik NLR from its cognate rice blast effector AVR-Pik to the host-determining factor pathogenicity toward weeping lovegrass 2 (Pwl2) by installing a putative host target, OsHIPP43, in place of the native integrated heavy metal-associated domain (generating Pikm-1OsHIPP43). This chimeric receptor also responded to other PWL alleles from diverse blast isolates. The crystal structure of the Pwl2/OsHIPP43 complex revealed a multifaceted, robust interface that cannot be easily disrupted by mutagenesis, and may therefore provide durable, broad resistance to blast isolates carrying PWL effectors in the field. Our findings highlight how the host targets of pathogen effectors can be used to bioengineer recognition specificities that have more robust properties compared to naturally evolved disease resistance genes.

A fatty acid elongase complex regulates cell membrane integrity and septin-dependent host infection by the rice blast fungus.

Very-long-chain fatty acids (VLCFAs) regulate biophysical properties of cell membranes to determine growth and development of eukaryotes, such as the pathogenesis of the rice blast fungus Magnaporthe oryzae. The fatty acid elongase Elo1 regulates pathogenesis of M. oryzae by modulating VLCFA biosynthesis. However, it remains unknown whether and how Elo1 associates with other factors to regulate VLCFA biosynthesis in fungal pathogens. Here, we identified Ifa38, Phs1 and Tsc13 as interacting proteins of Elo1 by proximity labelling in M. oryzae. Elo1 associated with Ifa38, Phs1 and Tsc13 on the endoplasmic reticulum (ER) membrane to control VLCFA biosynthesis. Targeted gene deletion mutants Δifa38, Δphs1 and Δtsc13 were all similarly impaired as Δelo1 in vegetative growth, conidial morphology, stress responses in ER, cell wall and membrane. These deletion mutants also displayed severe damage in cell membrane integrity and failed to organize the septin ring that is essential for penetration peg formation and pathogenicity. Our study demonstrates that M. oryzae employs a fatty acid elongase complex to regulate VLCFAs for maintaining or remodelling cell membrane structure, which is important for septin-mediated host penetration.

Fishwaste Derived Hydroxyapatite Nanostructure Combined with Black Rice Wine for Potential Antioxidant and Antimicrobial Response.

Hospital-acquired infection remains a serious threat globally, due to development of resistance to conventional antibiotics, which necessitates the urge for alternative therapy. Green nanotechnology has emerged as a holistic approach to address antibiotic resistance by combining environmental sustainability with improved therapeutic outcome. Nanostructure hydroxyapatite (HAP) has received significant attention in therapeutic and regenerative purposes due to its porous scaffold structure and biocompatible nature. In the present study, hydroxyapatite (HAP) nanoparticle was fabricated from the fish scale waste of red snapper fish. Black rice wine (BRW) was extracted from black rice commonly termed as Karupu kavuni/forbidden rice known for its nutritious effects. The present study focused on encapsulation of BRW within HAP nanoparticles (HAP@BRW) and evaluated its potential against nosocomial infections. Spectral and microscopic characterization of HAP@BRW revealed uniform encapsulation of BRW in HAP nanoparticles, aggregated irregular-shaped morphology of size 117.6 nm. Maximum release of BRW (72%) within 24 h indicates HAP as suitable drug delivery system suitable for biomedical applications. Antimicrobial studies revealed that HAP@BRW exhibited potent bactericidal effect against MRSA, MSSA, and Pseudomonas aeruginosa. Furthermore, HAP@BRW significantly inhibited the biofilm forming ability of MSSA and P. aeruginosa. Rich antioxidant property of HAP@BRW might be due to the presence of rich source of total polyphenolic, flavonoid, and anthocyanin content in BRW. In vitro and in vivo toxicity studies revealed biocompatible nature of HAP@BRW. Antibiofilm, antimicrobial, antioxidant, and biocompatible nature of HAP@BRW makes it a promising candidate for coating medical implants to avoid implant-associated infections and nosocomial infections.

The Similarities in Microbial and Chemical Patterns of Fermentation in Two Open Environments were Promoted by Using 150-Year-Old Nukadoko as Starters.

Nukadoko, a fermented rice bran employed in traditional Japanese pickling, uses lactic acid bacteria to ferment vegetables. Here, we report the microbial and chemical data of a mixture of matured 150-year-old nukadoko and commercially available rice bran placed in two open environments over 29 days. Across the two environments, Loigolactobacillus was identified as the dominant microbial genera in the later stages of fermentation in nukadoko. The period of increase in the relative abundance of Loigolactobacillus correlated with a decrease in pH and Oxidation-Reduction Potential (ORP) values. While the two environments showed a difference in the rate of change in microbial diversity, they shared the common process through which Loigolactobacillus outcompeted adventitious bacteria in nukadoko, as indicated by the alpha and beta diversity index. Thus, the similarities in microbial and chemical data across two open environments during fermentation using starters indicate that starters contribute to the stability of fermentation in open environments.

Crenobacter oryzisoli sp. nov., a novel phosphate-solubilizing bacterium isolated from the paddy soil.

Two Gram-staining-negative, facultative anaerobic, rod-shaped and phosphate-solubilizing strains designated SG2303T and SG2305, were isolated from paddy soil in China. Phylogenetic analysis based on 16 S rRNA gene sequences indicated that SG2303T and SG2305 represented a member of the genus Crenobacter within the family Neisseriaceae of the phylum Pseudomonadota. Strain SG2303T displayed higher 16 S rRNA gene sequence similarities with members of the genus Crenobacter ranging from 93.5 to 94.0%. Strains C. luteus YIM 78141T and C. cavernae K1W11S-77T were closest related to the isolated strains and were considered as type strains. Growth of strain SG2303T occurred at 10-55 °C (optimum 37 °C), pH 5.0-9.0 (optimum pH 6.0-7.0) and 0-1% (w/v) NaCl (optimum 0%). The average nucleotide identity (ANI) and digital DNA-DNA hybridization (dDDH) values between strain SG2303T and its closely related taxa were 76.1-78.2% and 20.5-22.1%, respectively. The genomic DNA G + C content was 62.2%. The quinone of strain SG2303T was Q-8. The major fatty acids (> 10%) of strain SG2303T were C16:0 (30.6%), summed feature 3 (C16:1ω7c and/or C16:1ω6c) (26.0%) and C12:0 3OH (12.1%). The polar lipids were phosphatidylglycerol (PG), diphosphatidylglycerol (DPG), phosphatidylethanolamine (PE), phospholipids (PL), glycolipid (GL) and unidentified lipids (UL). Based on the results of the phylogenetic, physiological, biochemical, and morphological analysis, strain SG2303T is recognized as a novel species of the genus Crenobacter, for which the name Crenobacter oryzisoli sp. nov. is proposed. The type strain is SG2303T (= GDMCC 1.3970T = JCM 36468T). In addition, SG2303T was also able of phosphorus solubilization and promoting the growth of rice seeds. Strain SG2303T exhibited a relatively high dissolvable phosphorus content of 2.52 µg·mL- 1.

Screening, identification, and characterization of molds for brewing rice wine: Scale-up production in a bioreactor.

Rice wine, well known for its unique flavor, rich nutritional value, and health benefits, has potential for extensive market development. Rhizopus and Aspergillus are among several microorganisms used in rice wine brewing and are crucial for determining rice wine quality. The strains were isolated via Rose Bengal and starch as a combined separation medium, followed by oenological property and sensory evaluation screening. The strain exhibiting the best performance can be screened using the traditional rice wine Qu. The strains YM-8, YM-10, and YM-16, which exhibited strong saccharification and fermentation performance along with good flavor and taste, were obtained from traditional rice wine Qu. Based on ITS genetic sequence analysis, the YM-8, YM-10, and YM-16 strains were identified as Rhizopus microsporus, Rhizopus arrhizus, and Aspergillus oryzae. The optimum growth temperature of each of the three strains was 30°C, 32°C, and 30°C, and the optimum initial pH was 6.0, 6.5, and 6.5, respectively. The activities of α-amylase, glucoamylase, and protease of YM-16 were highest at 220.23±1.88, 1,269.04±30.32, and 175.16±1.81 U/g, respectively. The amino acid content of rice wine fermented in a 20-L bioreactor with the three mold strains was higher than that of the control group, except for arginine, which was significantly lower than that of the control group. The total amino acid content and the total content of each type of amino acid were ranked as YM-16 > YM-8 > YM-10 > control group, and the amino acid content varied greatly among the strains. The control group had a higher content, whereas YM-8 and YM-16 had lower contents of volatile aroma components than the control group and had the basic flavor substances needed for rice wine, which is conducive to the formation of rice wine aroma. This selected strain, YM-16, has strong saccharification and fermentation ability, is a rich enzyme system, and improves the flavor of rice wine, thereby demonstrating its suitability as a production strain for brewing.

Inhibition and Mechanism of Citral on Bacillus cereus Vegetative Cells, Spores, and Biofilms.

Bacillus cereus causes food poisoning by producing toxins that cause diarrhea and vomiting and, in severe cases, endocarditis, meningitis, and other diseases. It also tends to form biofilms and spores that lead to contamination of the food production environment. Citral is a potent natural antibacterial agent, but its antibacterial activity against B. cereus has not been extensively studied. In this study, we first determined the minimum inhibitory concentrations and minimum bactericidal concentrations, growth curves, killing effect in different media, membrane potential, intracellular adenosine triphosphate (ATP), reactive oxygen species levels, and morphology of vegetative cells, followed by germination rate, morphology, germination state of spores, and finally biofilm clearance effect. The results showed that the minimum inhibitory concentrations and minimum bactericidal concentrations of citral against bacteria ranged from 100 to 800 µg/mL. The lag phase of bacteria was effectively prolonged by citral, and the growth rate of bacteria was slowed down. Bacteria in Luria-Bertani broth were reduced to below the detection limit by citral at 800 µg/mL within 0.5 h. Bacteria in rice were reduced to 3 log CFU/g by citral at 4000 µg/mL within 0.5 h. After treatment with citral, intracellular ATP concentration was reduced, membrane potential was altered, intracellular reactive oxygen species concentration was increased, and normal cell morphology was altered. After treatment with citral at 400 µg/mL, spore germination rate was reduced to 16.71%, spore morphology was affected, and spore germination state was altered. It also had a good effect on biofilm removal. The present study showed that citral had good bacteriostatic activity against B. cereus vegetative cells and its spores and also had a good clearance effect on its biofilm. Citral has the potential to be used as a bacteriostatic substance for the control of B. cereus in food industry production.

Novel trans-Resveratrol Derivatives: Design, Synthesis, Antibacterial Activity, and Mechanisms.

Rice bacterial leaf blight and rice bacterial leaf streak have induced tremendous damage to production of rice worldwide. To discover an effective novel antibacterial agent, a series of novel trans-resveratrol (RSV) derivatives containing 1,3,4-oxadiazole and amide moieties were designed and synthesized for the first time. Most of them showed excellent antibacterial activities against Xanthomonas oryzae pv oryzicola and Xanthomonas oryzae pv oryzae. Especially, compound J12 had the best inhibitory with the half-maximal effective concentration values of 4.2 and 5.0 mg/L, respectively, which were better than that of RSV (63.7 and 75.4 mg/L), bismerthiazol (79.5 and 89.6 mg/L), and thiodiazole copper (105.4 and 112.8 mg/L). Furthermore, compound J12 had an excellent control effect against rice bacterial leaf streak and rice bacterial leaf blight, with protective activities of 46.2 and 42.1% and curative activities of 44.5 and 41.7%, respectively. Preliminary mechanisms indicated that compound J12 could not only remarkably decrease biofilm formation, extracellular polysaccharide production, and the synthesis of extracellular enzymes but also destroy bacterial cell surface morphology, thereby reducing the pathogenicity of bacteria. In addition, compound J12 could increase the activity of defense-related enzymes and affect the expression of multiple pathogenic-related genes including plant-pathogen interaction, the MAPK signaling pathway, and phenylpropanoid biosynthesis, and this could improve the defense of rice against rice bacterial leaf streak infection. The present work indicates that the RSV derivatives can be used as promising candidates for the development of antibacterial agents.

As(III)-oxidizing and plant growth-promoting bacteria increase the starch biosynthesis-related enzyme activity, 2-AP levels, and grain quality of arsenic-stressed rice plants.

BACKGROUND: Grain quality is an important index of rice production, particularly when plants are grown under stress. Arsenic (As) contamination in paddy fields severely affects rice grain yield and quality. Here, the effects of As and combinations of As(III)-oxidizing bacteria (Pseudomonas stutzeri 4.25, 4.27, and 4.44) and plant growth-promoting bacteria (Delftia acidovorans KKU2500-12 and Cupriavidus taiwanensis KKU2500-3) on enzymes related to starch accumulation in grains and the grain quality of Khao Dawk Mali 105 rice cultivated in As-contaminated soil under greenhouse conditions were investigated. RESULTS: Arsenic affected the activities of starch biosynthesis-related enzymes, and decreases of up to 76.27%, 71.53%, 49.74%, 73.39%, and 47.46% in AGPase, SSS, GBSS, SBE, and SDBE activities, respectively, and 9.42-61.07% in starch accumulation in grains were detected after growth in As-contaminated soil. However, the KKU2500-3/4.25 and KKU2500-3/4.44 combinations yielded the greatest enzyme activities in grains, and compared with the results observed in uninoculated seedlings, increases in starch accumulation of up to 51.16% and 23.81% were found in the inoculated seedlings after growth in medium- and high-As-contaminated soils, at 10-17 and 10-24 days after anthesis, respectively. The bacteria increased the 2-AP content in rice under As stress, possibly via the induction of proline, a 2-AP substrate. Bacterium-inoculated rice had significantly greater 2-AP levels than uninoculated rice, and 2.16-9.93% and 26.57-42.04% increases were detected in rice plants grown in medium- and high-As-contaminated soils, respectively. CONCLUSIONS: Arsenic toxicity can be mitigated in rice growing under greenhouse conditions by maintaining starch biosynthesis, accumulating amylose, and increasing 2-AP content. The effectiveness of these bacteria should be validated in paddy fields; hence, safe rice grains with a good starch content and aroma could be produced.

Green synthesis and characterization of silver nanoparticles and its efficacy against Rhizoctonia solani, a fungus causing sheath blight disease in rice.

Rice (Oryza sativa) stands as a crucial staple food worldwide, especially in Bangladesh, where it ranks as the third-largest producer. However, intensified cultivation has made high-yielding rice varieties susceptible to various biotic stresses, notably sheath blight caused by Rhizoctonia solani, which inflicts significant yield losses annually. Traditional fungicides, though effective, pose environmental and health risks. To address this, nanotechnology emerges as a promising avenue, leveraging the antimicrobial properties of nanoparticles like silver nanoparticles (AgNPs). This study explored the green synthesis of AgNPs using Ipomoea carnea leaf extract and silver nitrate (AgNO3), and also examined their efficacy against sheath blight disease in rice. The biosynthesized AgNPs were characterized through various analytical techniques such as UV-vis spectrophotometer, X-ray Diffraction (XRD), Fourier Transform Infrared Spectroscopy (FTIR), Particle size analyzer, Zeta potential, Scanning Electron Microscope (SEM), Field Emission Scanning Electron Microscope (FESEM), Transmission Electron Microscope (TEM) for confirming their successful production and crystalline nature of nanoparticles. The results of UV-visible spectrophotometers revealed an absorption peak ranging from 421 to 434 nm, validated the synthesis of AgNPs in the solution. XRD, DLS, and TEM estimated AgNPs sizes were ~45 nm, 66.2nm, and 46.38 to 73.81 nm, respectively. SEM and FESEM demonstrated that the synthesized AgNPs were spherical in shape. In vitro assays demonstrated the significant inhibitory effects of AgNPs on mycelial growth of Rhizoctonia solani, particularly at higher concentrations and pH levels. Further greenhouse and field experiments validated the antifungal efficacy of AgNPs against sheath blight disease in rice, exhibiting comparable effectiveness to commercial fungicides. The findings highlight the potential of AgNPs as a sustainable and effective alternative for managing rice sheath blight disease, offering a safer solution amidst environmental concerns associated with conventional fungicides.