Identification of a Promising Novel Genetic Source for Rice Root-Knot Nematode Resistance through Markers Associated with Trait-Specific Quantitative Trait Loci.

Direct-seeded rice (DSR) is gaining popularity among farmers due to its environmentally safe and resource-efficient production system. However, managing the rice root-knot nematode (RRKN), Meloidogyne graminicola, remains a major challenge in DSR cultivation. Developing genetic resistance is a pragmatic and effective approach compared to using hazardous pesticides. Pusa Basmati 1121 (PB1121) is the most popular Basmati rice variety, but it is highly susceptible to RRKN. In contrast, Phule Radha (PR) has shown highly resistant reaction to RRKN, as reported in our earlier study. We generated an F2:3 population from the cross of PB1121/PR and evaluated it for RRKN resistance-related traits under artificial inoculation conditions. The distribution pattern of traits in the F2:3 population indicated that resistance may be governed by a few major-effect genes and many minor-effect genes. The molecular markers reported to be associated with QTLs governing RRKN resistance traits were used to test in the current population. Although the simple linear regression identified significant associations between the markers and RRKN resistance-associated traits, these associations were spurious as the LOD score was below the threshold limit. This indicates that PR possesses novel genomic regions for resistance to RRKN as it does not possess any of the earlier reported QTLs.

Polystyrene nanoplastics induce cell type-dependent secondary wall reinforcement in rice (Oryza sativa) roots and reduce root hydraulic conductivity.

Nanoplastics (NPs) have been demonstrated the ability to penetrate plant roots and cause stress. However, the extent of NPs penetration into various root tissues and the corresponding plant defense mechanisms remain unclear. This study examined the penetration and accumulation patterns of polystyrene nanoplastics (PS-NPs) in different cell types within rice roots, and explored how the roots quickly modify their cell wall structure in response. The findings showed that fully developed sclerenchyma cells in rice roots effectively prevented the invasion of PS-NPs. Meanwhile, PS-NPs triggered the accumulation of lignin and suberin in specific cells such as the exodermis, sclerenchyma, and xylem vessels. PS-NPs at a concentration of 50 mg L-1 increased cell wall thickness by 18.6 %, 21.1 %, and 22.4 % in epidermis, exodermis, and sclerenchyma cells, respectively, and decreased root hydraulic conductivity by 14.8 %. qPCR analysis revealed that PS-NPs influenced the cell wall synthesis pathway, promoting the deposition of lignin and suberin monomers on the secondary wall through the up-regulation of genes such as OsLAC and OsABCG. These results demonstrate that PS-NPs can induce cell type-specific strengthening of secondary walls and barrier formation in rice roots, suggesting the potential role of plant secondary wall development in mitigating NPs contamination risks in crops.

Endophytic diazotrophic communities from rice roots are diverse and weakly associated with soil diazotrophic community composition and soil properties.

AIM: Bacteria that promote plant growth, such as diazotrophs, are valuable tools for achieving a more sustainable production of important non-legume crops like rice. Different strategies have been used to discover new bacteria capable of promoting plant growth. This work evaluated the contribution of soil diazotrophs to the endophytic communities established in the roots of rice seedlings cultivated on seven representative soils from Uruguay. METHODS AND RESULTS: The soils were classified into two groups according to the C and clay content. qPCR, terminal restriction fragment length polymorphism (T-RFLP), and 454-pyrosequencing of the nifH gene were used for analyzing diazotrophs in soil and plantlets' roots grown from seeds of the same genotype for 25 days under controlled conditions. A similar nifH abundance was found among the seven soils, roots, or leaves. The distribution of diazotrophs was more uneven in roots than in soils, with dominance indices significantly higher than in soils (nifH T-RFLP). Dominant soils' diazotrophs were mainly affiliated to Alphaproteobacteria and Planctomycetota. Conversely, Alpha, Beta, Gammaproteobacteria, and Bacillota were predominant in different roots, though undetectable in soils. Almost no nifH sequences were shared between soils and roots. CONCLUSIONS: Root endophytic diazotrophs comprised a broader taxonomic range of microorganisms than diazotrophs found in soils from which the plantlets were grown and showed strong colonization patterns.

Plant growth regulators mitigate oxidative damage to rice seedling roots by NaCl stress.

The aim of this experiment was to investigate the effects of exogenous sprays of 5-aminolevulinic acid (5-ALA) and 2-Diethylaminoethyl hexanoate (DTA-6) on the growth and salt tolerance of rice (Oryza sativa L.) seedlings. This study was conducted in a solar greenhouse at Guangdong Ocean University, where 'Huanghuazhan' was selected as the test material, and 40 mg/L 5-ALA and 30 mg/L DTA-6 were applied as foliar sprays at the three-leaf-one-heart stage of rice, followed by treatment with 0.3% NaCl (W/W) 24 h later. A total of six treatments were set up as follows: (1) CK: control, (2) A: 40 mg⋅ L-1 5-ALA, (3) D: 30 mg⋅ L-1 DTA-6, (4) S: 0.3% NaCl, (5) AS: 40 mg⋅ L-1 5-ALA + 0.3% NaCl, and (6) DS: 30 mg⋅ L-1 DTA-6+0.3% NaCl. Samples were taken at 1, 4, 7, 10, and 13 d after NaCl treatment to determine the morphology and physiological and biochemical indices of rice roots. The results showed that NaCl stress significantly inhibited rice growth; disrupted the antioxidant system; increased the rates of malondialdehyde, hydrogen peroxide, and superoxide anion production; and affected the content of related hormones. Malondialdehyde content, hydrogen peroxide content, and superoxide anion production rate significantly increased from 12.57% to 21.82%, 18.12% to 63.10%, and 7.17% to 56.20%, respectively, in the S treatment group compared to the CK group. Under salt stress, foliar sprays of both 5-ALA and DTA-6 increased antioxidant enzyme activities and osmoregulatory substance content; expanded non-enzymatic antioxidant AsA and GSH content; reduced reactive oxygen species (ROS) accumulation; lowered malondialdehyde content; increased endogenous hormones GA3, JA, IAA, SA, and ZR content; and lowered ABA content in the rice root system. The MDA, H2O2, and O2- contents were reduced from 35.64% to 56.92%, 22.30% to 53.47%, and 7.06% to 20.01%, respectively, in the AS treatment group compared with the S treatment group. In the DS treatment group, the MDA, H2O2, and O2- contents were reduced from 24.60% to 51.09%, 12.14% to 59.05%, and 12.70% to 45.20%. In summary, NaCl stress exerted an inhibitory effect on the rice root system, both foliar sprays of 5-ALA and DTA-6 alleviated damage from NaCl stress on the rice root system, and the effect of 5-ALA was better than that of DTA-6.

Brown planthopper infestation on rice reduces plant susceptibility to Meloidogyne graminicola by reducing root sugar allocation.

Plants are simultaneously attacked by different pests that rely on sugars uptake from plants. An understanding of the role of plant sugar allocation in these multipartite interactions is limited. Here, we characterized the expression patterns of sucrose transporter genes and evaluated the impact of targeted transporter gene mutants and brown planthopper (BPH) phloem-feeding and oviposition on root sugar allocation and BPH-reduced rice susceptibility to Meloidogyne graminicola. We found that the sugar transporter genes OsSUT1 and OsSUT2 are induced at BPH oviposition sites. OsSUT2 mutants showed a higher resistance to gravid BPH than to nymph BPH, and this was correlated with callose deposition, as reflected in a different effect on M. graminicola infection. BPH phloem-feeding caused inhibition of callose deposition that was counteracted by BPH oviposition. Meanwhile, this pivotal role of sugar allocation in BPH-reduced rice susceptibility to M. graminicola was validated on rice cultivar RHT harbouring BPH resistance genes Bph3 and Bph17. In conclusion, we demonstrated that rice susceptibility to M. graminicola is regulated by BPH phloem-feeding and oviposition on rice through differences in plant sugar allocation.

NRTPredictor: identifying rice root cell state in single-cell RNA-seq via ensemble learning.

BACKGROUND: Single-cell RNA sequencing (scRNA-seq) measurements of gene expression show great promise for studying the cellular heterogeneity of rice roots. How precisely annotating cell identity is a major unresolved problem in plant scRNA-seq analysis due to the inherent high dimensionality and sparsity. RESULTS: To address this challenge, we present NRTPredictor, an ensemble-learning system, to predict rice root cell stage and mine biomarkers through complete model interpretability. The performance of NRTPredictor was evaluated using a test dataset, with 98.01% accuracy and 95.45% recall. With the power of interpretability provided by NRTPredictor, our model recognizes 110 marker genes partially involved in phenylpropanoid biosynthesis. Expression patterns of rice root could be mapped by the above-mentioned candidate genes, showing the superiority of NRTPredictor. Integrated analysis of scRNA and bulk RNA-seq data revealed aberrant expression of Epidermis cell subpopulations in flooding, Pi, and salt stresses. CONCLUSION: Taken together, our results demonstrate that NRTPredictor is a useful tool for automated prediction of rice root cell stage and provides a valuable resource for deciphering the rice root cellular heterogeneity and the molecular mechanisms of flooding, Pi, and salt stresses. Based on the proposed model, a free webserver has been established, which is available at https://www.cgris.net/nrtp .

An Insight into Occurrence, Biology, and Pathogenesis of Rice Root-Knot Nematode Meloidogyne graminicola.

Rice (Oryza sativa L.) is one of the most widely grown crops in the world, and is a staple food for more than half of the global total population. Root-knot nematodes (RKNs), Meloidogyne spp., and especially M. graminicola, seem to be significant rice pests, which makes them the most economically important plant-parasitic nematode in this crop. RKNs develop a feeding site in galls by causing host cells to differentiate into hypertrophied, multinucleate, metabolically active cells known as giant cells. This grazing framework gives the nematode a constant food source, permitting it to develop into a fecund female and complete its life cycle inside the host root. M. graminicola effector proteins involved in nematode parasitism, including pioneer genes, were functionally characterized in earlier studies. Molecular modelling and docking studies were performed on Meloidogyne graminicola protein targets, such as ß-1,4-endoglucanase, pectate lyase, phospholipase B-like protein, and G protein-coupled receptor kinase, to understand the binding affinity of Beta-D-Galacturonic Acid, 2,6,10,15,19,23-hexamethyltetracosane, (2S)-2-amino-3-phenylpropanoic acid, and 4-O-Beta-D-Galactopyranosyl-Alpha-D-Glucopyranose against ligand molecules of rice. This study discovered important molecular aspects of plant-nematode interaction and candidate effector proteins that were regulated by M. graminicola-infected rice plants. To the best of our knowledge, this is the first study to describe M. graminicola's molecular adaptation to host parasitism.

Improvement of growth, yield and associated bacteriome of rice by the application of probiotic Paraburkholderia and Delftia.

Plant probiotic bacteria enhance growth and yield of crop plants when applied at the appropriate time and dose. Two rice probiotic bacteria, Paraburkholderia fungorum strain BRRh-4 and Delftia sp. strain BTL-M2 promote growth and yield of plants. However, no information is available on application of these two bacteria on growth, yield, and diversity and population of bacteriome in roots and rhizosphere soils of the treated rice plants. This study aimed to assess the effect of BRRh-4 and BTL-M2 application on growth, yield and bacteriome in roots and rhizosphere soil of rice under varying doses of N, P and K fertilizers. Application of BRRh-4 and BTL-M2 strains significantly (p < 0.05) increased seed germination, growth and yield of rice compared to an untreated control. Interestingly, the grain yield of rice by these bacteria with 50% less of the recommended doses of N, P, and K fertilizers were statistically similar to or better than the rice plants treated with 100% doses of these fertilizers. Targeted amplicon (16S rRNA) sequence-based analysis revealed significant differences (PERMANOVA, p = 0.00035) in alpha-diversity between the root (R) and rhizosphere soil (S) samples, showing higher diversity in the microbial ecosystem of root samples. Additionally, the bacteriome diversity in the root of rice plants that received both probiotic bacteria and chemical fertilizers were significantly higher (PERMANOVA, p = 0.0312) compared to the rice plants treated with fertilizers only. Out of 185 bacterial genera detected, Prevotella, an anaerobic and Gram-negative bacterium, was found to be the predominant genus in both rhizosphere soil and root metagenomes. However, the relative abundance of Prevotella remained two-fold higher in the rhizosphere soil metagenome (52.02%) than in the root metagenome (25.04%). The other predominant bacterial genera detected in the rice root metagenome were Bacillus (11.07%), Planctomyces (4.06%), Faecalibacterium (3.91%), Deinococcus (2.97%), Bacteroides (2.61%), and Chryseobacterium (2.30%). On the other hand, rhizosphere soil metagenome had Bacteroides (12.38%), Faecalibacterium (9.50%), Vibrio (5.94%), Roseomonas (3.40%), and Delftia (3.02%). Interestingly, we found the presence and/or abundance of specific genera of bacteria in rice associated with the application of a specific probiotic bacterium. Taken together, our results indicate that improvement of growth and yield of rice by P. fungorum strain BRRh-4 and Delftia sp. strain BTL-M2 is likely linked with modulation of diversity, structures, and signature of bacteriome in roots and rhizosphere soils. This study for the first time demonstrated that application of plant growth promoting bacteria significantly improve growth, yield and increase the diversity of bacterial community in rice.

Shifts in the diversity of root endophytic microorganisms across the life cycle of the ratooning rice Jiafuzhan.

The diversity of root endophytic microorganisms, which is closely related to plant life activities, is known to vary with the plant growth stage. This study on the ratooning rice Jiafuzhan explored the diversity of the root endophytic bacteria and fungi and their dynamics during the plant life cycle. By sequencing the 16S ribosomal ribonucleic acid (16S rRNA) and internal transcribed spacer (ITS) genes, 12,154 operational taxonomic units (OTUs) and 497 amplicon sequence variants (ASVs) were obtained, respectively. The root endophytic microorganisms of rice in the seedling, tillering, jointing, heading, and mature stages of the first crop and at 13, 25, and 60 days after regeneration (at the heading, full heading, and mature stages of the second crop, respectively) were analyzed using diversity and correlation analyses. There were significant differences in the α-diversity and ß-diversity of root endophytic bacteria and fungi in the growth stage. Additionally, linear discriminant analysis (LDA) effect size (LEfSe) analysis revealed biomarker bacteria for each growth stage, but biomarker fungi did not exist in every stage. Moreover, the correlation analysis showed that the bacterial and fungal biomarkers interacted with each other. Furthermore, the nitrogen-fixing genus Bradyrhizobium existed in all growth stages. These findings indicate the pattern of root endophytic microorganisms of ratooning rice at different growth stages, and they provide new insights into the high yield of the second crop of ratooning rice (in light of the abundance of various bacteria and fungi).

Nuclear translocation of OsMADS25 facilitated by OsNAR2.1 in reponse to nitrate signals promotes rice root growth by targeting OsMADS27 and OsARF7.

Nitrate is an important nitrogen source and signaling molecule that regulates plant growth and development. Although several components of the nitrate signaling pathway have been identified, the detailed mechanisms are still unclear. Our previous results showed that OsMADS25 can regulate root development in response to nitrate signals, but the mechanism is still unknown. Here, we try to answer two key questions: how does OsMADS25 move from the cytoplasm to the nucleus, and what are the direct target genes activated by OsMADS25 to regulate root growth after it moves to the nucleus in response to nitrate? Our results demonstrated that OsMADS25 moves from the cytoplasm to the nucleus in the presence of nitrate in an OsNAR2.1-dependent manner. Chromatin immunoprecipitation sequencing, chromatin immunoprecipitation qPCR, yeast one-hybrid, and luciferase experiments showed that OsMADS25 directly activates the expression of OsMADS27 and OsARF7, which are reported to be associated with root growth. Finally, OsMADS25-RNAi lines, the Osnar2.1 mutant, and OsMADS25-RNAi Osnar2.1 lines exhibited significantly reduced root growth compared with the wild type in response to nitrate supply, and expression of OsMADS27 and OsARF7 was significantly suppressed in these lines. Collectively, these results reveal a new mechanism by which OsMADS25 interacts with OsNAR2.1. This interaction is required for nuclear accumulation of OsMADS25, which promotes OsMADS27 and OsARF7 expression and root growth in a nitrate-dependent manner.

Calcium enhances phosphorus reclamation during biochar formation: Mechanisms and potential application as a phosphorus fertilizer in a paddy soil.

Transformation of phosphorus (P) species during pyrolytic production of biochar from P-rich biowastes with a subsequent soil amendment is important to P reclamation. Aiming at increasing the content of plant-available P and restraining the formation of easily mobile P in pyrolysis product, this study used exogenous calcium ions (20 wt% CaCl2) addition prior to pyrolysis to regulate the pyrolytic transformation of P chemical fractions from sewage sludge and bone dreg. Results showed that active Ca catalyzed the decomposition of organic P to transform into inorganic orthophosphate. Based on Hedley's sequential extraction method, this study found that addition of Ca ions remarkably reduced the content of soluble P, exchange P, Fe/Al bound P, and occluded P in biochar, while increased Ca bound P from 78 to 85% to 85-96%. Liquid 31P NMR indicated that exogenous Ca induced the crack of the P-O-P bond in pyrophosphate to become orthophosphates. It also explained why new orthophosphates including chlorapatite (Ca5(PO4)3Cl) and calcium hydroxyapatite (Ca10(PO4)6(OH)2) appeared in the Ca-composite biochar compared to pristine biochar. Combined with rapid P-release test in paddy soil (pH 6.27) and 30-days rice seedling growth test under flooded condition (10 wt% biochar addition ratio), it was confirmed that compared to pristine biochar, Ca-composite biochar released more P in paddy soil, but also promoted more P to be taken in by rice root and stalk. These results suggested that pretreating biowaste with Ca ion was a friendly approach to enhance P reclamation during biochar formation, making it a promising P fertilizer.

Diproline-induced resistance to parasitic nematodes in the same and subsequent rice generations: Roles of iron, nitric oxide and ethylene.

Induced resistance (IR) is a plant phenotype characterized by lower susceptibility to biotic challenges upon elicitation by so-called IR stimuli. Earlier, we identified diproline (cyclo(l-Pro-l-Pro)) as IR stimulus that protects rice (Oryza sativa) against the root-knot nematode Meloidogyne graminicola (Mg). In the current study, detailed transcriptome analyses at different time points, and under uninfected and nematode-infected conditions revealed that this rice IR phenotype is correlated with induction of genes related to iron (Fe), ethylene (ET) and reactive oxygen species (ROS)/reactive nitrogen species (RNS) metabolism. An infection experiment under Fe limiting conditions confirmed that diproline-IR is only effective under optimal Fe supply. Although total root Fe levels were not affected in diproline-treated plants, phytosiderophore secretion was found to be induced by this treatment. Experiments on mutant and transgenic rice lines impaired in ET or ROS/RNS metabolism confirmed that these metabolites are involved in diproline-IR. Finally, we provide evidence for transgenerational inheritance of diproline-IR (diproline-TIR), as two successive generations of diproline-treated ancestors exhibited an IR phenotype while themselves never being exposed to diproline. Transcriptome analyses on the offspring plants revealed extensive overlap between the pathways underpinning diproline-IR and diproline-TIR. Although diproline induces significant systemic changes in global DNA methylation levels early after treatment, such changes in DNA methylation were not detected in the descendants of these plants. To our knowledge, this is the first report of TIR in rice and the first transcriptional assessment of TIR in monocots.

Response mechanism and rapid detection of phenotypic information in rice root under heavy metal stress.

The root is an important organ affecting cadmium accumulation in grains, but there is no comprehensive research involving rice root phenotype under cadmium stress yet. To assess the effect of cadmium on root phenotypes, this paper investigated the response mechanism of phenotypic information including cadmium accumulation, adversity physiology, morphological parameters, and microstructure characteristics, and explored rapid detection methods of cadmium accumulation and adversity physiology. We found that cadmium had the effect of "low-promotion and high-inhibition" on root phenotypes. In addition, the rapid detection of cadmium (Cd), soluble protein (SP), and malondialdehyde (MDA) were achieved based on spectroscopic technology and chemometrics, where the optimal prediction model was least squares support vector machine (LS-SVM) based on the full spectrum (Rp=0.9958) for Cd, competitive adaptive reweighted sampling-extreme learning machine (CARS-ELM) (Rp=0.9161) for SP and CARS-ELM (Rp=0.9021) for MDA, all with Rp higher than 0.9. Surprisingly, it took only about 3 min, which was more than 90% reduction in detection time compared with laboratory analysis, demonstrating the excellent ability of spectroscopy for root phenotype detection. These results reveal response mechanism to heavy metal and provide rapid detection method for phenotypic information, which can substantially contribute to crop heavy metal control and food safety supervision.

The effects of different iron and phosphorus treatments on the formation and morphology of iron plaque in rice roots (Oryza sativa L).

Introduction: Rice (Oryza sativa L.) is a pivotal cereal crop worldwide. It relies heavily on the presence of iron plaque on its root surfaces for optimal growth and enhanced stress resistance across diverse environmental conditions. Method: To study the crystallographic aspects of iron plaque formation on rice roots, the concentrations of Fe2+ and PO4 3- were controlled in this study. The effects of these treatments were assessed through comprehensive analyzes encompassing root growth status, root surface iron concentration, root vitality, enzyme activities, and microstructural characteristics using advanced techniques such as root analysis, scanning electron microscopy (SEM), and ultrathin section transmission electron microscopy (TEM). Results: The results demonstrated that an increase in the Fe2+ concentration or a decrease in the PO4 3- concentration in the nutrient solution led to improvements in various root growth indicators. There was an elevation in the DCB (dithionite-citrate-bicarbonate) iron content within the roots, enhanced root vitality, and a significant increase in the activities of the superoxide dismutase (SOD), peroxidase (POD) and catalase (CAT) enzymes. Moreover, as the Fe2+ concentration increased, amorphous iron oxide minerals on the root surface were gradually transformed into ferrihydrite particles with sizes of approximately 200 nm and goethite particles with sizes of approximately 5 µm. This study showed that an increase in the Fe2+ concentration and a decrease in the PO4 3- concentration led to the formation of substantial iron plaque on the root surfaces. It is noteworthy that there was a distinct gap ranging from 0.5 to 3 µm between the iron plaque formed through PO4 3- treatment and the cellular layer of the root surface. Discussion: This study elucidated the impacts of Fe2+ and PO4 3- treatments on the formation, structure, and morphology of the iron plaque while discerning variations in the spatial proximity between the iron plaque and root surface under different treatment conditions.

Engineering D-glucose utilization in Azospirillum brasilense Sp7 promotes rice root colonization.

Bacteria of the genus Azospirillum include several plant associated bacteria which often promote the growth of their host plants. Although the host range of Azospirillum brasilense Sp7 is much wider than its close relative Azospirillum lipoferum 4B, it lacks the ability to efficiently utilize D-glucose for its growth. By comparing the genomes of both the species, the genes of A. lipoferum 4B responsible for conferring D-glucose utilization ability in A. brasilese Sp7 were identified by cloning individual or a combination of genes in a broad host range expression vector, mobilizing them in A. brasilense Sp7 and examining the ability of exconjugants to use D-glucose as sole carbon source for growth. These genes also included the homologs of genes involved in N-acetyl glucosamine utilization in Pseudomonas aeruginosa PAO1. A transcriptional fusion of the 5 genes encoding glucose-6-phosphate dehydrogenase and 4 components of glucose phosphotransferase system were able to improve D-glucose utilization ability in A. brasilense Sp7. The A. brasilense Sp7 strain engineered with D-glucose utilization ability showed significantly improved root colonization of rice seedling. The improvement in the ability of A. brasilense Sp7 to colonize rice roots is expected to bring benefits to rice by promoting its growth. KEY POINTS: • Genes required for glucose utilization in Azospirillum lipoferum were identified. • A gene cassette encoding glucose utilization was constructed. • Transfer of gene cassette in A. brasilense improves glucose utilization and rice root colonization..

Year-Long Assessment of Soil Nematode Diversity and Root Inhibition-Indicator Nematode Genera in Rice Fields.

Soil nematodes contribute to nutrient cycling. This year-long study aimed to investigate the changes in the diversity of soil nematodes during the spring, summer, and winter seasons in rice fields at 24 sites and to determine the indicator nematode genera that inhibit the roots of rice plants. A total of 216 soil samples were collected during three seasons, and the collection of 72 root samples was carried out during rice cropping. Forty-four soil nematode genera were identified. They exhibited significant changers in their abundance, which were dependent on the seasons and on soil characteristics. In particular, the abundance of plant-parasitic nematodes (PPNs) and free-living soil nematodes was 49% and 15% higher during the summer than during the spring and winter seasons, respectively. Soil characteristics, such as soil nitrogen (N) contents, carbon (C) contents, and soil moisture were significantly higher during the summer than in the spring and winter seasons, but soil pH was significantly lower during the summer than in the spring and winter seasons. Moreover, Hirschmanniella,Meloidogyne, and Heterodera emerged as good indicators for rice root inhibition, corroborating the frequency, density, and prominence value of PPNs of the sampled soil and rice roots. This study also indicated that free-living nematode genera, such as Rhabdolaimus, Diplogaster, and Rhabditis, might function as ecological indicators for soil health.

Temporal dynamics of total and active root-associated diazotrophic communities in field-grown rice.

Plant-associated nitrogen-fixing microorganisms (diazotrophs) are essential to host nutrient acquisition, productivity and health, but how host growth affects the succession characteristics of crop diazotrophic communities is still poorly understood. Here, Illumina sequencing of DNA- and RNA-derived nifH genes was employed to investigate the dynamics of total and active diazotrophic communities across rhizosphere soil and rice roots under four fertilization regimes during three growth periods (tillering, heading and mature stages) of rice in 2015 and 2016. Our results indicated that 71.9-77.2% of the operational taxonomic units (OTUs) were both detected at the DNA and RNA levels. According to the nonmetric multidimensional scaling ordinations of Bray-Curtis distances, the variations in community composition of active rhizosphere diazotrophs were greater than those of total rhizosphere diazotrophs. The community composition (ß-diversity) of total and active root-associated diazotrophs was shaped predominantly by microhabitat (niche; R 2 ≥ 0.959, p < 0.001), followed by growth period (R 2 ≥ 0.15, p < 0.001). The growth period had a stronger effect on endophytic diazotrophs than on rhizosphere diazotrophs. From the tillering stage to the heading stage, the α-diversity indices (Chao1, Shannon and phylogenetic diversity) and network topological parameters (edge numbers, average clustering coefficient and average degree values) of total endophytic diazotrophic communities increased. The proportions of OTUs shared by the total rhizosphere and endophytic diazotrophs in rhizosphere diazotrophs gradually increased during rice growth. Moreover, total diazotrophic α-diversity and network complexity decreased from rhizosphere soil to roots. Collectively, compared with total diazotrophic communities, active diazotrophic communities were better indicators of biological response to environmental changes. The host microhabitat profoundly drove the temporal dynamics of total and active root-associated diazotrophic communities, followed by the plant growth period.

Increasing phosphorus inhibits the retention and prevention of cadmium by iron plaque and promotes cadmium accumulation in rice plants.

Iron (Fe) plaque is an important component of rice roots because it influences the uptake and transport of cadmium (Cd) in rice. In this study, a hydroponic experiment was developed to investigate the influence of phosphorus (P) on the formation of iron plaque on the root surface as well as the influence on Cd uptake in rice plants. Three important results were obtained, as follows. (1) During the formation of iron plaque induced by exogenous Fe, P supply was beneficial for the iron plaque formation, but it restrained the Cd retention capability, resulting in a decrease in Cd in iron plaque by 35.48-61.93%, and leading to an increase in Cd in rice roots from 72.13 mg kg-1 to 112.78 mg kg-1 (2) After the iron plaque induction, the formation of iron plaque was inhibited by P supply, resulting in the amount of iron plaque decreasing by 18.46-54.57%, and the Cd in iron plaque decreased by 3.93-31.78%. Then, the Cd retention capability and the prevention effect simultaneously decreased, and as a result, the Cd in rice roots increased from 100.83 mg kg-1 to 146.03 mg kg-1 (3) Without exogenous Fe induction, P sufficiency continued to increase the amount of iron plaque and decrease the Cd in iron plaque, and increase the Cd in rice plants. These results suggested that P supply increases the amount of iron plaque, which is ineffective for Cd retention, such as non-reddish-brown iron plaque (NIP), and then decreases the capacity of iron plaque to retain Cd. Additionally, the P supply decreased the amount of formed iron plaque, causing the decreasing prevention effect. Therefore, excessive application of P fertilizer should be avoided in Cd-contaminated paddy fields.

First report of Pythium aristosporum causing root rot on rice seedling in Taiwan.

Rice (Oryza sativa L.) is the principle staple crops in the World and its production can be severely damaged by Pythium species. Several Pythium species including P. afertile, P. arrhenomanes, P. dissotocum, P. elongatum, P. spinosum, have been recorded to cause rice seedling root rot in Taiwan (List of Plant Diseases in Taiwan edited by Tzean et al., 2019). During the survey of rice seedling diseases, we identified a new species of Pythium that causes seedling root rot on rice in commercial nursery trays in two nursery fields in 2019 in Taichung, Taiwan. Stunting and root rot symptom were found on the affected plants and up to 20% seedlings in a nursery tray showed similar symptoms. To isolate the pathogen, symptomatic roots were surface sterilized with 75% ethanol for 1 min and rinsed in sterile water. The margin of lesion was cut off, placed on 1.5% water agar and incubated at 28 ℃. After 24 h, the hyphal tips of a white colony growing from the diseased region were transferred to potato dextrose agar (PDA) medium. Koch's postulates were fulfilled by inoculating the germinated rice seeds with mycelia. Rice seeds of O. sativa var. Tainan11 (TN11) were treated with 75% ethanol and then 1.2% NaOCl for 15 min. The sterilized seeds were soaked in sterile water under dark condition for 3 days and the water was replaced every day. Five of the pre-germinated seeds with 2~5 mm embryonic shoot were placed in a sterile petri-dish and inoculated with 3-ml mycelial suspension (OD600 = 0.045) prepared by blending the mycelia of a 3-days PDA culture using an Oster 10 speed blender 6640 (Oster, USA). The seeds-mycelia were then covered with sterilized soil mixture of Akadama soil and rice husk (1:1, volume to volume) and incubated in a growth chamber at 28 ℃. Seven days post-inoculation, the inoculated seedlings showed stunting with short and necrotic roots (Fig. S1). The pathogen was reisolated from the diseased seedlings and identified with morphology and molecular methods. For morphological characterization, the pathogen was cultured on V8 agar to produce oogonia and zoospore (Chamswarng and Cook 1985). Globose oogonia with multiple antheridia (1-5 per oogonium), inflated filamentous sporangia, vesicle with abundant zoospores, main hypha with up to 6.57 µm wide and mature aplerotic oospores with diameter 24.35-30.81 µm (average= 27.22 µm; n=20) were observed (Fig. S1) that are similar to the descriptions for P. aristosporum (van der Plaats-Niterink 1981). Genomic DNA was extracted with CTAB method (Wang and White 1997) and the sequences of the internal transcribed spacer (ITS) region and gene region of ß-tubulin (tub) and cytochrome c oxidase subunit II (cox II) were amplified with published primers (Villa et al., 2006). The obtained sequences were submitted to GenBank (accession nos: OL701302 (ITS), OL763269 (tub), and OL763270 (cox II); Fig. S2). Phylogenetic relationships between this Pythium pathogen and other 55 Pythium isolates, including the type species of P. aristosporum (ATCC11101), were conducted with the concatenated sequences of tub and cox II and analyzed by Bayesian interference (Fig. S3). Based on the tree built with tub and cox II sequences, this pathogen was identified as P. aristosporum that has not been reported in rice and other plants in Taiwan. It was observed in laboratory assays that this pathogen caused significant root-rot symptoms on several major rice varieties grown in Taiwan, including TN11, Tainung67 and Kaoshiung139. It may potentially cause severe crop loss in rice production, especially in nurseries. This identification provides important information on rice disease management.

Ultrahigh-Performance Liquid Chromatography-Mass Spectrometry Analysis of Carotenoid-Derived Hormones and Apocarotenoids in Plants.

Carotenoid oxidative cleavage products, apocarotenoids (APOs), are a class of important plant secondary metabolites, which include phytohormones abscisic acid (ABA) and strigolactones (SLs), and growth regulators and signaling molecules such as ß-cyclocitral, zaxinone, anchorene, ß-apo-11-carotenoids, and retinal. Qualitative and quantitative analysis of these bioactive compounds is crucial for understanding their metabolism and may also enable discovering further regulatory APOs. The state-of-the-art mass spectrometry (MS) technology has advanced the detection of plant APOs; however, it is still challenging to perform an accurate analysis of the low-level phytohormones ABA and SL and the structurally diverse APOs from complex plant matrices. Here, we describe ultrahigh-performance liquid chromatography-MS (UHPLC-MS) methods to determine carotenoid-derived hormones and APOs from plants by integrating ultrasound-assisted extraction and solid-phase extraction. These assays enable an accurate quantification of carotenoid-derived hormones and APOs from plant tissues by using an UHPLC hybrid quadrupole-Orbitrap mass spectrometer. © 2022 Wiley Periodicals LLC. Basic Protocol 1: UHPLC-MS analysis of APOs from rice roots Support Protocol: Preparation of dried plant root powder Basic Protocol 2: UHPLC-MS analysis of SLs from rice roots Basic Protocol 3: UHPLC-MS analysis of ABA from rice roots.