Rice Pathogen Research Articles

Phosphatidylcholine Transfer Protein OsPCTP Interacts with Ascorbate Peroxidase OsAPX8 to Regulate Bacterial Blight Resistance in Rice

Rice phosphatidylcholine transfer protein (PCTP), which contains a steroidogenic acute regulatory protein-related lipid transfer (START) domain, responds to bacterial blight disease. Overexpression of OsPCTP quantitatively enhances resistance to pathogen in rice, whereas depletion of it has the opposite effect. Further analysis showed that OsPCTP physically interacts with OsAPX8, a ROS scavenging enzyme, and influences ascorbate peroxidase enzymatic activity in vivo. In addition, the expression of pathogenesis-related genes PR1a, PR1b and PR10 were significantly induced in OsPCTP-OE plants compared with that in wild-type plants ZH11. Taken together, these results suggested that OsPCTP mediates bacterial blight resistance in rice through regulating the ROS defense pathway.

Evaluating the efficacy of endophytic bacteria in controlling rice sheath blight: In vitro and In vivo studies

Rice production is highly susceptible to various pathogens, including Rhizoctonia solani, Curvularia lunata, and Epicoccum rostratum, which are major threats in Asia. Exploring biological control methods using endophytic bacteria offers promising opportunities to enhance rice resilience against these lethal diseases. Using 16S RNA sequencing, we identified four endophytic isolates of Bacillus spp. from rice roots, stems, and leaves. We evaluated the antagonistic activity of these endophytic bacterial isolates against rice pathogens both in vitro and in vivo. These isolates inhibited the growth of C. lunata by 82 %, R. solani by 79 %, and E. rostratum by 88 % in vitro. The detached leaf assay for sheath blight (ShB) disease severity in strains ranged from 10.4 % to 73.3 %. In vivo results showed that B. amyloliquefaciens (R-19) exhibited the lowest disease intensity at 7.2 % and the highest disease suppression at 78.8 %. The fungicide propiconazole at 0.1 % treatment showed the lowest disease intensity of 7.7 % and the highest disease suppression of 73.4 %, compared to the infected control. Besides biocontrol efficacy, endophytic isolates enhance plant growth parameters, including shoot height, root length, fresh and dry weights, number of tillers, and grains per tiller. Plant hormones abscisic acid (ABA) and gibberellic acids (GA3) increased by 35 % and 53 %, respectively, due to B. subtilis (R-20) and B. amyloliquefaciens (R-19), while flavonoid and indole acetic acid (IAA) concentrations surged by 30%–80 %. Similarly, chlorophyll (a, b), carotenoids, antioxidant enzymatic activity, phenolic content, carbohydrates, and proline contents were higher compared to the control. This study provides a foundation for future studies on novel and eco-friendly biocontrol agents. In addition, our study recommends the integration of endophytic bacteria into sustainable agriculture for enhancing rice production and reducing disease impacts.

Natural Defense: The Inhibitory Effects of Endophytic Bacillus and Xylaria sp. against Rice Pathogens

Aim: Endophytes living inside the plant tissues are known to play a crucial role in the physiology and developmental functioning of host plants. Rice is the major food crop that feeds half of the world population. But pest and diseases lead to drastic yield reduction in rice. Sustainable and eco-friendly disease management strategies are urgently needed to reduce the reliance on chemical pesticides and mitigate the environmental impact of intensive agriculture.The aim of the study was the selection of natural suppressors of the brown spot and sheath blight diseases in rice. In this study, three bacterial endophytes viz., Bacillus subtilis isolate PAL 11, B. altitudins isolate TSL-4 and B. altitudinis isolate KAM-11 and one endophytic fungal isolate Xylaria sp. isolate KTD-2 were tested for their potential to inhibit Bipolaris oryzae and Rhizoctonia solani under in vitro conditions. Methodology: In vitro evaluation of endophytes against two major rice pathogens viz., Rhizoctonia solani and Bipolaris oryzae causing sheath blight and brown leaf spot respectively were tested by dual culture technique. Results: All the tested bacterial and fungal endophytes significantly inhibited both the pathogens under in vitro conditions. Among the bacterial endophytes, B. subtilis isolate PAL-11 exhibited highest inhibition of both the pathogens. The mycelial inhibition of B. oryzae was 56.04 and 74.48 per cent respectively at 5 and 8 days after dual culturing whereas the inhibition of R. solani was 80.31 and 80.22 per cent respectively at 3 and 5 days after dual culturing. The fungal endophyte Xylaria sp. isolate KTD-2 exhibited 63.2 and 67.95 per cent inhibition of mycelial growth of B. oryzae at 5 and 8 days respectively after the pathogen inoculation. The mycelial growth of R. solani was also significantly inhibited (54.95 %) by the fungal endophyte at 5 days after pathogen inoculation. Conclusion: This research highlights the potential role of endophytes in sustainable agriculture and suggests that integrating beneficial microbes into rice disease management strategies could help enhance plant health, increase crop yields, and contribute to global food security.

Limonene enhances rice plant resistance to a piercing-sucking herbivore and rice pathogens.

Terpene synthases (TPSs) are key enzymes in terpenoids synthesis of plants and play crucial roles in regulating plant defence against pests and diseases. Here, we report the functional characterization of OsTPS19 and OsTPS20, which were upregulated by the attack of brown planthopper (BPH). BPH female adults performed concentration-dependent behavioural responses to (S)-limonene showing preference behaviour at low concentrations and avoidance behaviour at high concentrations. Overexpression lines of OsTPS19 and OsTPS20, which emitted higher amounts of the monoterpene (S)-limonene, decreased the hatching rate of BPH eggs, reduced the lesion length of sheath blight caused by Rhizoctonia solani and bacterial blight caused by Xanthomonas oryzae. While knockout lines of OsTPS19 and OsTPS20, which emitted lower amounts of (S)-limonene, were more susceptible to these pathogens. Overexpression of OsTPS19 and OsTPS20 in rice plants had adverse effects on the incidence of BPH, rice blast, and sheath blight in the field and had no significant impacts on rice yield traits. OsTPS19 and OsTPS20 were found to be involved in fine-tuning the emission of (S)-limonene in rice plants and play an important role in defence against both BPH and rice pathogens.

OsHRZ1 negatively regulates rice resistant to Magnaporthe oryzae infection by targeting OsVOZ2.

Rice blast disease caused by Magnaporthe oryzae significantly reduces yield production. Blast resistance is closely associated with iron (Fe) status, but the mechanistic basis linking iron status to immune function in rice remains largely unknown. Here, iron-binding haemerythrin RING ubiquitin ligases OsHRZ1 was confirmed to play key roles in iron-mediated rice blast resistance. The expression of OsHRZ1 was suppressed by M. oryzae inoculation and high iron treatment. Both mutants of OsHRZ1 enhanced rice resistance to M. oryzae. OsPR1a was up-regulated in OsHRZ1 mutants. Yeast two-hybrid, bimolecular fluorescence complementation, and Co-IP assay results indicated that OsHRZ1 interacts with Vascular Plant One Zinc Finger 2 (OsVOZ2) in the nucleus. Additionally, the vitro ubiquitination assay indicated that OsHRZ1 can ubiquitinate OsVOZ2 and mediate the degradation of OsVOZ2. The mutants of OsVOZ2 showed reduced resistance to M. oryzae and down-regulated the expression of OsPR1a. Yeast one-hybrid, EMSA, and dual-luciferase reporter assay results indicated that OsVOZ2 directly binds to the promoter of OsPR1a, activating its expression. In summary, OsHRZ1 plays an important role in rice disease resistance by mediated degradation of OsVOZ2 thus shaping PR gene expression dynamics in rice cells. This highlights an important link between iron signaling and rice pathogen defenses.

Horizontal transmission of a rice bacterial pathogen Pantoea ananatis among plants by rice planthoppers.

Pantoea ananatis is a bacterium commonly found in various agronomic crops and agricultural pests. In this study, we present findings on a genome-reduced strain of P. ananatis, known as Lstr, which was initially isolated from Laodelphax striatellus (small brown rice planthopper, SBPH). We identified Lstr as a plant pathogen causing disease in rice using Koch's postulates. The pathogenicity of Lstr on rice is comparable to that of Xanthomonas oryzae pv. oryzae, the main causative agent of rice bacterial blight. Through a series of experiments involving live insects, molecular investigations, and microscopy, we find that Lstr can accumulate within SBPH. Subsequently, Lstr can be transmitted from SBPH to rice plants, resulting in leaf blight, and can also be transmitted to other SBPH individuals. Collectively, our results suggest that SBPH serves as a vector for P. ananatis Lstr in rice plants. P. ananatis may encounter susceptible insect populations and become endemic through horizontal transmission from these insects. This could also be valuable for predicting future occurrences of bacterial leaf blight in rice and other crops caused by P. ananatis.

Whole genome regulatory effect of MoISW2 and consequences for the evolution of the rice plant pathogenic fungus Magnaporthe oryzae.

Isw2 proteins, ubiquitous across eukaryotes, exhibit a propensity for DNA binding and exert dynamic influences on local chromosome condensation in an ATP-dependent fashion, thereby modulating the accessibility of neighboring genes to transcriptional machinery. Here, we report the deletion of a putative MoISW2 gene, yielding substantial ramifications on plant pathogenicity. Subsequent gene complementation and chromatin immunoprecipitation sequencing (ChIP-seq) analyses were conducted to delineate binding sites. RNA sequencing (RNA-seq) assays revealed discernible impacts on global gene regulation along chromosomes in both mutant and wild-type strains, with comparative analyses against 55 external RNA-seq data sets corroborating these findings. Notably, MoIsw2-mediated binding and activities delineate genomic loci characterized by pronounced gene expression variability proximal to MoIsw2 binding sites, juxtaposed with comparatively stable expression in surrounding regions. The contingent genes influenced by MoIsw2 activity predominantly encompass niche-determinant genes, including those encoding secreted proteins, secondary metabolites, and stress-responsive elements, alongside avirulence genes. Furthermore, our investigations unveil a spatial correlation between MoIsw2 binding motifs and known transposable elements (TEs), suggesting a potential interplay wherein TE transposition at these loci could modulate the transcriptional landscape of Magnaporthe oryzae in a strain-specific manner. Collectively, these findings position MoIsw2 as a plausible master regulator orchestrating the delicate equilibrium between genes vital for biomass proliferation, akin to housekeeping genes, and niche-specific determinants crucial for ecological adaptability. Stress-induced TE transposition, in conjunction with MoIsw2 activity, emerges as a putative mechanism fostering enhanced mutagenesis and accelerated evolution of niche-determinant genes relative to housekeeping counterparts.IMPORTANCEIsw2 proteins are conserved in plants, fungi, animals, and other eukaryotes. We show that a fungal Isw2 protein in the rice pathogen Magnaporthe oryzae binds to retrotransposon (RT) DNA motifs and affects the epigenetic gene expression landscape of the fungal genome. Mainly ecological niche determinant genes close to the binding motifs are affected. RT elements occur frequently in DNA between genes in most organisms. They move place and multiply in the genome, especially under physiological stress. We further discuss the Isw2 and RT combined activities as a possible sought-after mechanism that can cause biased mutation rates and faster evolution of genes necessary for reacting to abiotic and biotic challenges. The most important biotic challenges for plant pathogens are the ones from the host plants' innate immunity. The overall result of these combined activities will be an adaptation-directed evolution of niche-determinant genes.

Ustilaginoidea virens, an emerging pathogen of rice: the dynamic interplay between the pathogen virulence strategies and host defense.

The Ustilaginoidea virens -rice pathosystem has been used as a model for flower-infecting fungal pathogens. The molecular biology of the interactions between U. virens and rice, with an emphasis on the attempt to get a deeper comprehension of the false smut fungus's genomes, proteome, host range, and pathogen biology, has been investigated. Meta-QTL analysis was performed to identify potential QTL hotspots for use in marker-assisted breeding. The Rice False Smut (RFS) caused by the fungus Ustilaginoidea virens currently threatens rice cultivators across the globe. RFS infects rice panicles, causing a significant reduction in grain yield. U. virens can also parasitize other hosts though they play only a minor role in its life cycle. Furthermore, because it produces mycotoxins in edible rice grains, it puts both humans and animals at risk of health problems. Although fungicides are used to control the disease, some fungicides have enabled the pathogen to develop resistance, making its management challenging. Several QTLs have been reported but stable gene(s) that confer RFS resistance have not been discovered yet. This review offers a comprehensive overview of the pathogen, its virulence mechanisms, the genome and proteome of U. virens, and its molecular interactions with rice. In addition, information has been compiledon reported resistance QTLs, facilitatingthe development of a consensus genetic map using meta-QTL analysis for identifying potential QTL hotspots. Finally, this review highlights current developments and trends in U. virens-rice pathosystem research while identifying opportunities for future investigations.

Genome-Wide Transcriptome Analysis of a Virulent sRNA, Trans217, in Xanthomonas oryzae pv. oryzae (Xoo), the Causative Agent of Rice Bacterial Blight.

Small non-coding RNAs (sRNAs) act as post-transcriptional regulators to participate in many cellular processes. Among these, sRNA trans217 has been identified as a key virulent factor associated with pathogenicity in rice, triggering hypersensitive reactions in non-host tobacco and facilitating the secretion of the PthXo1 effector in Xanthomonas oryzae pv. oryzae (Xoo) strain PXO99A. Elucidating potential targets and downstream regulatory genes is crucial for understanding cellular networks governing pathogenicity and plant resistance. To explore the targets regulated by sRNA trans217, transcriptome sequencing was carried out to assess differential expression genes (DEGs) between the wild-type strain PXO99A and a mutant lacking the sRNA fragment under both virulence-inducing or normal growth conditions. DEG analysis revealed that sRNA trans217 was responsible for diverse functions, such as type III secretion system (T3SS), glutamate synthase activity, and oxidative stress response. Three genes were selected for further investigation due to their significant differential expression and biological relevance. Deletion of PXO_RS08490 attenuated the pathogenicity of Xoo in rice and reduced the tolerance level of PXO99A to hydrogen peroxide. These findings suggest a regulatory role of sRNA trans217 in modulating bacterial virulence through multiple gene targets, either directly or indirectly.

VmsR, a LuxR-Type Regulator, Contributes to Virulence, Cell Motility, Extracellular Polysaccharide Production and Biofilm Formation in Xanthomonas oryzae pv. oryzicola.

LuxR-type regulators play pivotal roles in regulating numerous bacterial processes, including bacterial motility and virulence, thereby exerting a significant influence on bacterial behavior and pathogenicity. Xanthomonas oryzae pv. oryzicola, a rice pathogen, causes bacterial leaf streak. Our research has identified VmsR, which is a response regulator of the two-component system (TCS) that belongs to the LuxR family. These findings of the experiment reveal that VmsR plays a crucial role in regulating pathogenicity, motility, biofilm formation, and the production of extracellular polysaccharides (EPSs) in Xoc GX01. Notably, our study shows that the vmsR mutant exhibits a reduced swimming motility but an enhanced swarming motility. Furthermore, this mutant displays decreased virulence while significantly increasing EPS production and biofilm formation. We have uncovered that VmsR directly interacts with the promoter regions of fliC and fliS, promoting their expression. In contrast, VmsR specifically binds to the promoter of gumB, resulting in its downregulation. These findings indicate that the knockout of vmsR has profound effects on virulence, motility, biofilm formation, and EPS production in Xoc GX01, providing insights into the intricate regulatory network of Xoc.

Morphogenetic characterization of Stenotrophomonas maltophilia infecting white stripe disease of rice (Oryza sativa L.)

Rice is a major cereal crop which ensure food security to more than half of the global population. Several biotic factors impact rice grain quality and its final production. White stripe disease, caused by pathogen Stenotrophomonas maltophilia is considered among the major limiting factor for reducing rice yields and quality. Present study was performed to understand the white stripe disease, which has been frequently misdiagnosed as bacterial leaf blight (BLB) due to similar symptoms. A survey was carried out based on accessibility and farmer participation to monitor incidence and sample collection. The survey was conducted in districts Faisalabad, Gujranwala, Sialkot, Sheikhupura, and Hafizabad, these districts were selected for their importance for rice cultivation in Pakistan. The total sample size was around 500 leaves distributed evenly throughout each study area. The results of study indicated presence of new pathogen of rice. These isolates were biochemically identified and confirmed by gram staining negative, 3% KOH positive, 5% salt tolerance positive, oxidase test negative, catalase test positive, starch hydrolysis test negative, nitrate reductase test positive, indole test negative, lactose test positive, maltose test positive, methyl red test negative, Voges-prokauer test negative and urea hydrolysis test negative. The pathogenicity test was confirmed that pathogen and the Sialkot isolate were the most aggressive isolate among the five isolates collected form the studied areas. The molecular characterization was accomplished by PCR and sequencing. The results of the phylogenetic study indicate that this pathogen belongs to a distinct group, as it is distantly related to Xanthomonas oryzae pv. oryzae (Xoo). This study provides important findings into a novel clade of pathogen causing white stripe disease in rice.

Multiple Chitin- or Avirulent Strain-Triggered Immunity Induces Microbiome Reassembly in Rice.

Magnaporthe oryzae is one of the most important fungal pathogens of rice. Chitin and avirulent strains can induce two layers of immunity response, pathogen-associated molecular pattern (PAMP)-triggered immunity (PTI) and effector-triggered immunity (ETI), in rice with cognate R genes. However, little is known about the assembly of the rice microbiome induced by PTI and ETI in rice. In this study, we investigate the impact of continuous treatment of the avirulent M. oryzae strain with AvrPi9 and chitin on the bacterial endophytic community of rice varieties harboring resistant gene Pi9 and their antagonistic activity against rice blast fungus. Analysis of the 16S rRNA showed a significant increase in the diversity and microbial co-occurrence network complexity and the number of beneficial taxa-Bacillus, Pseudomonas, Microbacterium, and Stenotrophomonas spp.-following the chitin and avirulent strain treatments. The antifungal assay with bacterial endophytes recovered from the leaves showed few bacteria with antagonistic potential in rice treated with avirulent strains, suggesting that the sequential treatment of the avirulent strain decreased the antagonistic bacteria against M. oryzae. Moreover, we identified Bacillus safensis Ch_66 and Bacillus altitudinis Nc_68 with overall antagonistic activities in vivo and in vitro. Our findings provide a novel insight into rice microbiome assembly in response to different innate immunity reactions.

Bacillus amyloliquefaciens LM-1 Affects Multiple Cell Biological Processes in Magnaporthe oryzae to Suppress Rice Blast.

Magnaporthe oryzae, one of the most destructive rice pathogens, causes significant losses during the rice harvest every year. Bacillus amyloliquefaciens has been explored in many crops as a potential biocontrol agent. However, the mechanisms of B. amyloliquefaciens controled rice blast are not fully understood. Here, a biocontrol strain LM-1, isolated from a contaminated medium, was identified as B. amyloliquefaciens using morphological observation, physiological and biochemical tests, and 16S rDNA sequencing. LM-1 inhibited the growth and pathogenicity of M. oryzae and Bipolaris oryzae (Breda de Haan) Shoem. The mycelia of M. oryzae co-cultured with LM-1 were enlarged and broken by fluorescence microscopy using calcofluor white. LM-1 inhibited the mycelia of M. oryzae from producing conidia. Genes itu, srf, and fenB were detected in LM-1. Furthermore, the supernatant of LM-1 interfered with the appressorium formation of M. oryzae, blocked conidial cell death, and reduced autophagy degradation but did not affect the normal germination of rice seeds and seeding growth. Additionally, we observed hypersensitivity reactions, reactive oxygen species, and iron accumulation reduction in rice cells inoculated with supernatant. Our study reveals that LM-1 has a control effect on rice blast and affects cell wall integrity, sporulation, appressorium formation, cell death, and autophagy.

The Identification and Gene Mapping of Spotted Leaf Mutant spl43 in Rice.

Our study investigates the genetic mechanisms underlying the spotted leaf phenotype in rice, focusing on the spl43 mutant. This mutant is characterized by persistent reddish-brown leaf spots from the seedling stage to maturity, leading to extensive leaf necrosis. Using map-based cloning, we localized the responsible locus to a 330 Kb region on chromosome 2. We identified LOC_Os02g56000, named OsRPT5A, as the causative gene. A point mutation in OsRPT5A, substituting valine for glutamic acid, was identified as the critical factor for the phenotype. Functional complementation and the generation of CRISPR/Cas9-mediated knockout lines in the IR64 background confirmed the central role of OsRPT5A in controlling this trait. The qPCR results from different parts of the rice plant revealed that OsRPT5A is constitutively expressed across various tissues, with its subcellular localization unaffected by the mutation. Notably, we observed an abnormal accumulation of reactive oxygen species (ROS) in spl43 mutants by examining the physiological indexes of leaves, suggesting a disruption in the ROS system. Complementation studies indicated OsRPT5A's involvement in ROS homeostasis and catalase activity regulation. Moreover, the spl43 mutant exhibited enhanced resistance to Xanthomonas oryzae pv. oryzae (Xoo), highlighting OsRPT5A's role in rice pathogen resistance mechanisms. Overall, our results suggest that OsRPT5A plays a critical role in regulating ROS homeostasis and enhancing pathogen resistance in rice.

Laser-disintegrated copper-activated salicylic acid (Cu-SA) nanoparticles suppressed the seed-borne pathogens of rice with enhanced seed invigoration parameters.

Leading phytopathological research is focused on managing seed-borne pathogens of rice through the utilization of engineered nanomaterials. Herein, blue laser-induced topo-morphologically nano-advanced copper salicylates (Cu-SA) (Cu/SA in 1:1 and 1:2 ratio) were prepared and evaluated for their augmented antifungal potential along seed invigoration effects in contrast to their prepared sonicated formulations. Laser disintegration on the Cu-SA (Cu/SA in 1:1 and 1:2 ratio) was achieved with high degree of success and precision using blue laser, which yielded uniformly distributed spherical nanoparticles with a narrow size distribution and better crystallinity than aqua-dispersed sonicated formulations. In vitro antifungal evaluation against seed-borne fungi of rice viz. Fusarium verticillioides and Fusarium fujikuroi revealed multiple times the augmented potential of laser-disintegrated nanoformulations (l-CuSA) than sonicated (s-CuSA) and bulk samples. Laser-induced nano-sodium bis(2-oxobenzoato)cuprate (II) (l-CuSA2) with Cu/SA in 1:2 ratio was the best to inhibit the in vitro fungal growth. Ultra-micrographs and fungal double-staining assay further rationalized the membrane disruption as the mode of action for the fungitoxicity. Nanopriming of fungal infested rice seeds with l-CuSA2 at 2500 μg/mL for 8 h showed the maximum reduction of seed rot (80.43%) and seedling blight (63.15%) with respect to control (untreated). The seed-invigorating factors of l-CuSA2 nanoprimed seeds were enhanced to maximum extent and showed the highest per cent germination (35.29%), shoot length (11.42%), root length (21.14%), dry weight (75.43%) and vigour index (81.04%) over the control. Inclusively, the seed nanopriming with l-CuSA2 proved as agro-compatible hypo-toxic semi natural nanoplatform for sustainable agriculture.

Plant growth-promoting rhizobacteria and biocontrol agents triggered plant defence responses against phytopathogenic fungi and improved rice growth

This study examined the antifungal activity and plant growth-promoting (PGP) traits of 30 beneficial bacterial strains isolated from Brassica campestris (L.) rhizosphere in rice. Among these, the PRB-8 strain exhibited potent antifungal activity against rice pathogens and produced a robust biofilm and a variety of PGP compounds. Based on our results, the PRB-8 showed distinct antifungal activity against fungal pathogens such as Rhizoctonia solani (75 %) > Alternaria alternata (74.5 %) > Sclerotium rolfsii (70 %) > Macrophomina phaselina (68 %) > Fusarium oxysporum (67 %). The isolate PRB-8 also increased the germination efficiency and length of rice seedlings under in-vitro conditions. The isolate PRB-8 was identified as Klebsiella pneumoniae (accession no. MT672588) and the absence of a hydrolysis zone on sheep blood agar plate suggests that PRB-8 is non-pathogenic. Inoculation of PRB-8 to R. solani and F. oxysporum-infected rice seedlings promotes length, dry biomass, and photosynthetic pigments and decreases the disease index in rice. Furthermore, PRB-8 potentially enhanced the defense responses in rice by increasing phenylalanine ammonia-lyase (PAL), β-1,3 glucanase, polyphenol oxidase (PPO), superoxide dismutase (SOD), catalase (CAT), and peroxidase (POD), which play a significant role in protecting plants from phytopathogens. Thus, it can be concluded that the use of the PRB-8 strain enhances rice growth and resistance to phytopathogens while simultaneously offering agroecosystems and environmentally friendly choices. Shortly, natural antagonistic PGPR may be used as biocontrol agents to sustainably enhance biotic stress tolerance and global food production.

Exploring the comparative genome of rice pathogen Burkholderia plantarii: unveiling virulence, fitness traits, and a potential type III secretion system effector

This study presents a comprehensive genomic analysis of Burkholderia plantarii, a rice pathogen that causes blight and grain rot in seedlings. The entire genome of B. plantarii KACC 18964 was sequenced, followed by a comparative genomic analysis with other available genomes to gain insights into its virulence, fitness, and interactions with rice. Multiple secondary metabolite gene clusters were identified. Among these, 12 demonstrated varying similarity levels to known clusters linked to bioactive compounds, whereas eight exhibited no similarity, indicating B. plantarii as a source of potentially novel secondary metabolites. Notably, the genes responsible for tropolone and quorum sensing were conserved across the examined genomes. Additionally, B. plantarii was observed to possess three complete CRISPR systems and a range of secretion systems, exhibiting minor variations among the analyzed genomes. Genomic islands were analyzed across the four genomes, and a detailed study of the B. plantarii KACC 18964 genome revealed 59 unique islands. These islands were thoroughly investigated for their gene contents and potential roles in virulence. Particular attention has been devoted to the Type III secretion system (T3SS), a crucial virulence factor. An in silico analysis of potential T3SS effectors identified a conserved gene, aroA. Further mutational studies, in planta and in vitro analyses validated the association between aroA and virulence in rice. Overall, this study enriches our understanding of the genomic basis of B. plantarii pathogenicity and emphasizes the potential role of aroA in virulence. This understanding may guide the development of effective disease management strategies.

Characterization and evaluation of native Trichoderma harzianum P39 for biocontrol potentials against five seed-borne fungal pathogens of rice

Plant seeds can harbour pathogens that limits crop growth and productivity. This study isolated, characterised and evaluated native Trichoderma harzianum for antagonism against five (Alternaria alternata, Aspergillus oryzae, Bipolaris oryzae, Curvularia lunata and Fusarium verticillioides) fungal pathogens isolated from rice seeds. Evaluation was through simultaneous inoculation, a pathogen against the biocontrol agent, using the dual culture method in a PDA growth medium. Results confirmed the T. harzianum isolate had 98.27% pairwise identity with T. harzianum P39 (KX381171.1. accession number). The isolate showed increasing percentage inhibition, against all the pathogens, with increasing days after inoculation. The highest values ranged from 53.02%, A. oryzae to 70.06%, F. verticillioides (the least and most susceptible respectively). The overall inhibition percentages of A. alterata and B. oryzae (12th day after inoculation) were not significantly different from that of F. verticillioides. Trichoderma harzianum P39 can be formulated as biological seed dressing agent.

Nitrogen fertilization modulates rice phyllosphere functional genes and pathogens through fungal communities

The phyllosphere is a vital yet often neglected habitat hosting diverse microorganisms with various functions. However, studies regarding how the composition and functions of the phyllosphere microbiome respond to agricultural practices, like nitrogen fertilization, are limited. This study investigated the effects of long-term nitrogen fertilization with different levels (CK, N90, N210, N330) on the functional genes and pathogens of the rice phyllosphere microbiome. Results showed that the relative abundance of many microbial functional genes in the rice phyllosphere was significantly affected by nitrogen fertilization, especially those involved in C fixation and denitrification genes. Different nitrogen fertilization levels have greater effects on fungal communities than bacteria communities in the rice phyllosphere, and network analysis and structural equation models further elucidate that fungal communities not only changed bacterial-fungal inter-kingdom interactions in the phyllosphere but also contributed to the variation of biogeochemical cycle potential. Besides, the moderate nitrogen fertilization level (N210) was associated with an enrichment of beneficial microbes in the phyllosphere, while also resulting in the lowest abundance of pathogenic fungi (1.14 %). In contrast, the highest abundance of pathogenic fungi (1.64 %) was observed in the highest nitrogen fertilization level (N330). This enrichment of pathogen due to high nitrogen level was also regulated by the fungal communities, as revealed through SEM analysis. Together, we demonstrated that the phyllosphere fungal communities were more sensitive to the nitrogen fertilization levels and played a crucial role in influencing phyllosphere functional profiles including element cycling potential and pathogen abundance. This study expands our knowledge regarding the role of phyllosphere fungal communities in modulating the element cycling and plant health in sustainable agriculture.

Screening Plant Growth-Promoting Bacteria with Antimicrobial Properties for Upland Rice.

This study explores beneficial bacteria isolated from the roots and rhizosphere soil of Khao Rai Leum Pua Phetchabun rice plants. A total of 315 bacterial isolates (KK001 to KK315) were obtained. Plant growth-promoting traits (phosphate solubilization and indole-3-acetic acid (IAA) production), and antimicrobial activity against three rice pathogens (Curvularia lunata NUF001, Bipolaris oryzae 2464, and Xanthomonas oryzae pv. oryzae) were assessed. KK074 was the most prolific in IAA production, generating 362.6 ± 28.0 μg/ml, and KK007 excelled in tricalcium phosphate solubilization, achieving 714.2 ± 12.1 μg/ml. In antimicrobial assays using the dual culture method, KK024 and KK281 exhibited strong inhibitory activity against C. lunata, and KK269 was particularly effective against B. oryzae. In the evaluation of antimicrobial metabolite production, KK281 and KK288 exhibited strong antifungal activities in cell-free supernatants. Given the superior performance of KK281, taxonomically identified as Bacillus sp. KK281, it was investigated further. Lipopeptide extracts from KK281 had significant antimicrobial activity against C. lunata and a minimum inhibitory concentration (MIC) of 3.1 mg/ml against X. oryzae pv. oryzae. LC-ESI-MS/MS analysis revealed the presence of surfactin in the lipopeptide extract. The crude extract was non-cytotoxic to the L-929 cell line at tested concentrations. In conclusion, the in vitro plant growth-promoting and disease-controlling attributes of Bacillus sp. KK281 make it a strong candidate for field evaluation to boost plant growth and manage disease in upland rice.