Root Tissue Samples Research Articles

MYB transcription factors in Peucedanum Praeruptorum Dunn: the diverse roles of the R2R3-MYB subfamily in mediating coumarin biosynthesis

BackgroundThe MYB superfamily (v-myb avian myeloblastosis viral oncogene homolog) plays a role in plant growth and development, environmental stress defense, and synthesis of secondary metabolites. Little is known about the regulatory function of MYB genes in Peucedanum praeruptorum Dunn, although many MYB family members, especially R2R3-MYB genes, have been extensively studied in model plants.ResultsA total of 157 R2R3-MYB transcription factors from P. praeruptorum were identified using bioinformatics analysis. Comprehensive analyses including chromosome location, microsynteny, gene structure, conserved motif, phylogenetic tree, and conserved domain were further performed. The length of the 157 transcription factors ranged from 120 to 1,688 amino acids (molecular weight between 14.21 and 182.69 kDa). All proteins were hydrophilic. Subcellular localization predictions showed that 155 PpMYB proteins were localized in the nucleus, with PpMYB12 and PpMYB157 localized in the chloroplasts and mitochondria, respectively. Ten conserved motifs were identified in the PpMYBs, all of which contained typical MYB domains. Transcriptome analysis identified 47,902 unigenes. Kyoto Encyclopedia of Genes and Genomes analysis revealed 136 pathways, of which 524 genes were associated with the phenylpropanoid pathway. Differential expressed genes (DEGs) before and after bolting showed that 11 genes were enriched in the phenylpropanoid pathway. Moreover, the expression patterns of transcription genes were further verified by qRT-PCR. With high-performance liquid chromatography (HPLC), 8 coumarins were quantified from the root, stem, and leaf tissue samples of P. praeruptorum at different stages. Praeruptorin A was found in both roots and leaves before bolting, whereas praeruptorin B was mainly concentrated in the roots, and the content of both decreased in the roots and stems after bolting. Praeruptorin E content was highest in the leaves and increased with plant growth. The correlation analysis between transcription factors and coumarin content showed that the expression patterns of PpMYB3 and PpMYB103 in roots align with the accumulation trends of praeruptorin A, praeruptorin B, praeruptorin E, scopoletin, and isoscopoletin, which declined in content after bolting, suggesting that these genes may positively regulate the biosynthesis of coumarins. Eleven distinct metabolites and 48 DEGs were identified. Correlation analysis revealed that the expression of all DEGs were significantly related to the accumulation of coumarin metabolites, indicating that these genes are involved in the regulation of coumarin biosynthesis.ConclusionsR2R3-MYB transcription factors may be involved in the synthesis of coumarin. Our findings provide basic data and a rationale for future an in-depth studies on the role of R2R3-MYB transcription factors in the growth and regulation of coumarin synthesis.

Evaluation and Validation of Reliable Reference Genes for Quantitative Real-Time PCR Analysis of the Gene Expression in Macadamia integrifolia

Macadamia is an economically significant crop, with its kernel oil being abundant in monounsaturated fatty acids (MUFA). Analyzing the expression of genes related to MUFA biosynthesis is essential for understanding the complex regulatory networks in Macadamia. However, there are few reports on the identification of suitable reference genes for use as internal controls in this species. Consequently, selecting a reliable reference gene for gene expression studies under various conditions is critical. In this study, we evaluated the expression stability of 11 traditional housekeeping genes: α-tubulin (TUBa), β-tubulin (TUBb), malate dehydrogenase (MDH), 18S ribosomal RNA (18S), glyceraldehyde-3-phosphate dehydrogenase (GAPDH), α-elongation factor 1 (EF1a), β-elongation factor 1 (EF1b), ubiquitin (UBQ), ubiquitin-conjugating enzyme (UBC), cyclophilin (CYP), and actin (ACT) under abiotic stresses, hormonal treatments and in variety of plant tissues using the online tool RefFinder, which integrates four commonly used software programs: ΔCt, geNorm (version 3.4), NormFinder (version 0953), and BestKeeper (version 1.0). A comprehensive expression stability ranking was established by integrating results from these four methods based on the geometric mean. The findings indicated that ACT was the most stable gene across all samples, including those subjected to cold stress, NaCl stress, PEG stress, ABA treatment, MeJA treatment, and both stem and leaf tissues. EF1b was identified as the most stable gene in GA treatment and heat stress samples, while UBC and CYP were ranked highest in ethrel treatment and root tissue samples, respectively. Finally, the reliability of these findings was further validated using the target gene SAD through qRT-PCR. In summary, this study evaluated and validated appropriate reference genes for qRT-PCR, which will facilitate future investigations into the molecular mechanisms in Macadamia.

First Peek into the Transcriptomic Response in Heat-Stressed Tomato Inoculated with Septoglomus constrictum.

In this study, we report the interaction between an arbuscular mycorrhizal fungus, Septoglomus constrictum, and tomato plants under heat stress. For the first time, this interaction was studied by Illumina RNA-seq, followed by a comprehensive bioinformatic analysis that investigated root and leaf tissue samples. The genome-wide transcriptional profiling displayed fewer transcriptomic changes in the root under heat-stress conditions caused by S. constrictum. The top 50 DEGs suggested significant changes in the expression of genes encoding heat-shock proteins, transporter proteins, and genes of phytohormone metabolism involving jasmonic acid signalling. S. constrictum induced the upregulation of genes associated with pathways such as 'drought-responsive' and the 'development of root hair' in the root, as well as 'glycolipid desaturation', 'intracellular auxin transport', and 'ethylene biosynthesis' in the leaf. The pathways 'biotin biosynthesis' and 'threonine degradation' were found in both investigated tissue types. Expression analysis of transcription factors showed 2 and 11 upregulated transcription factors in heat-stressed root and leaf tissues, respectively. However, we did not find shared transcription factors. Heat-stressed arbuscular mycorrhizal plants suffered less oxidative stress when exposed to high temperatures. Colorimetric tests demonstrated less accumulation of H2O2 and MDA in heat-stressed mycorrhizal plants. This phenomenon was accompanied by the higher expression of six stress genes that encode peroxidases, glutathione S-transferase and ubiquitin carboxyl-terminal hydrolase in roots and leaves. Our findings provide a new perspective on elucidating the functional metabolic processes of tomato plants under mycorrhizal-heat stressed conditions.

First report of Stagonosporopsis pogostemonis causing root rot on Strawberry in China.

In Henan, strawberry cultivation occurs on approximately 10,000 hectares, with annual production approaching 230,000 tons. In April 2022, a root rot disease with a 10% incidence rate was observed on the strawberry cultivars 'Ningyu' and 'Sweet Charlie' grown in plastic greenhouses (0.7 ha) located in Xingyang (113.39°E, 34.79°N), Henan, China. Disease symptoms included yellowing of the outer mature leaves, stunted growth, and subsequent wilting of the entire plant. The roots developed dark brown spots, which gradually turned necrotic (Figures 1a, 1b). To determine the causal agent, four symptomatic plants (two plants per cultivar) were collected. Twelve symptomatic root tissues (three root tissue samples per plant) were surface sterilized with 75% ethanol and 0.1% mercuric chloride, washed thrice in sterile water, air dried, and then placed on PDA at 25°C for 3 days. Eight pure isolates were obtained by hyphal-tip isolation (Fang 2007). Each colony had a dark olivaceous green to brown, cottony appearance with a round margin, and the reverse side was grey-black near the center (Figure 1c). Conidia were ellipsoidal, aseptate, with rounded ends, and 3.1 to 4.8 μm × 1.0 to 2.5 μm in size (Figure 1d). Chlamydospores were ellipsoidal, pale brown, and 7.9 to 11.9 μm × 7.6 to 10.7 μm in size (Figure 1e). A representative fungus isolate, designated as Z5, was selected for further molecular identification. Genomic DNA was extracted from the mycelia of isolate Z5, and four gene partial regions (ITS, TUB2, RPB2, and LSU) were amplified using the primer pairs ITS1/ITS4, Bt-2a/Bt-2b, RPB2-5F/RPB2-7CR and LROR/LR5, respectively (White et al.1990, O'Donnell et al.1997, Reeb et al. 2004, Rehner and Samuels 1994). PCR products were sequenced and submitted to GenBank with the following accession numbers OQ130480 (ITS), OQ190093 (TUB2), OQ190092 (RPB2), and OQ255570 (LSU). BLASTn search revealed that the ITS, TUB2, RPB2, and LSU gene sequences of isolate Z5 showed 99.42% (513/516 bp), 99.69% (320/321 bp), 100% (1071/1071 bp), and 100% (857/857 bp) identity with those of ex-type S. pogostemonis stain ZHKUCC 21-0001 (Dong et al. 2021), respectively. A phylogenetic tree was constructed showing that Z5 clustered with S. pogostemonis (Figure 2). The isolates in this study were identified as S. pogostemonis based on morphological and molecular evidence. To confirm pathogenicity, five one-month-old 'Ningyu' cultivar strawberry seedlings were planted in sterilized nursery soil mixed with wheat grains (0.5% w/w) coated with Z5 mycelia (Fang 2007). An equal number of strawberry seedlings were placed in pots filled with non-infected potting mix to serve as controls. The seedlings were kept in a greenhouse under a 12 h light/dark photoperiod at 25°C. After two weeks, the inoculated seedlings displayed symptoms such as leaf wilting and root necrosis, similar to those observed in the greenhouses, while the control seedlings showed no symptoms (Figures 1f, 1g). The experiment was performed thrice. The identical fungus was re-isolated from the symptomatic roots and identified as S. pogostemonis based on morphological characteristics and molecular analysis, thus fulfilling Koch's postulates. This is the first report of S. pogostemonis causing root rot on strawberries worldwide. Our findings will contribute to a more comprehensive study on investigating and managing this disease.

Inducible defense phytohormones in annual bluegrass (Poa annua) and creeping bentgrass (Agrostis stolonifera) in response to annual bluegrass weevil (Listronotus maculicollis) infestation.

The annual bluegrass weevil (Listronotus maculicollis) is the most damaging insect pest of short-mown turfgrass on golf courses in eastern North America. Listronotus maculicollis larvae cause limited visible damage as stem-borers (L1-3), compared to the crown-feeding (L4-5) developmental instars. Prolonged larval feeding results in discoloration and formation of irregular patches of dead turf, exposing soil on high-value playing surfaces (fairways, collars, tee boxes, and putting greens). Annual bluegrass (Poa annua) is highly susceptible to L. maculicollis compared to a tolerant alternate host plant, creeping bentgrass (Agrostis stolonifera). This study explored whether defense signaling phytohormones contribute to A. stolonifera tolerance in response to L. maculicollis. Concentrations (ng/g) of salicylic acid (SA), jasmonic acid (JA), jasmonic-isoleucine (JA-Ile), 12-oxophytodienoic acid (OPDA), and abscisic acid (ABA) were extracted from turfgrass (leaf, stem, and root) tissue samples as mean larval age reached 2nd (L2), 3rd (L3), and 4th (L4) instar. Poa annua infested with L. maculicollis larvae (L2-4) possessed significantly greater SA in above-ground tissues than A. stolonifera. Levels of constitutive JA, JA-Ile, OPDA, and ABA were significantly higher within non-infested A. stolonifera aboveground tissues compared to P. annua. Inducible defense phytohormones may play a role in P. annua susceptibility to L. maculicollis but are unlikely to provide tolerance in A. stolonifera. Additional studies in turfgrass breeding, particularly focusing on cultivar selection for increased constitutive JA content, could provide a non-chemical alternative management strategy for L. maculicollis for turfgrass managers.

Exploring endophytic bacteria communities of Vanilla planifolia

BackgroundRhizosphere bacterial community and endophytes are now known to influence plant health and response to environmental stress. Very few studies have reported the diversity of endophytic bacterial communities of Vanilla planifolia and their potential roles in promoting plant growth or contributing to aromatic quality.ResultsIn this study, the composition and diversity of the Vanilla rhizosphere bacterial community were explored by analyzing rhizosphere soil and root tissue samples as well as green pods of three accessions of Vanilla planifolia grown on different types of substrates (compost and leaf litter). In addition, the endophytic bacterial diversity of roots and green pods as well as the evolution of endophytic bacteria after the curing process of vanilla green pods were analyzed based on a metabarcoding approach. The results showed that bacterial species richness and diversity were higher in the compost. The analysis of the soil bacterial composition displayed that Halomonas, Pseudoalteromonas, Enterobacter and Bradyrhizobium were the most abundant genera. Moreover, the results indicated that the soil bacterial community structure was linked to the host plant genotype. Regarding the roots endophytic bacteria composition, the genera Halomonas, Pseudoalteromonas, Bacillus and Carboxydocella genera were present in all samples, independently from the substrate nature. Several genera including Bacillus, Bradyrhizobium, Burkholderia and Halomonas were transmitted internally from the roots to the green pods. The curing process reduced the bacterial richness and bacterial diversity associated with the green pods. Halomonas, Pseudoalteromonas, Bacillus, and Carboxydocella are the dominant genera in the pods after the curing process.ConclusionsThis study provides an overview of changes of the bacterial communities dynamics especially endophytic in the roots and the green pods. It highlighted bacterial genera (Halomonas, Pseudoalteromonas, Bacillus, and Carboxydocella) potentially implicated in the formation of aroma compounds of vanilla beans.

Insights into chickpea (Cicer arietinum L.) genotype adaptations to terminal drought stress: Evaluating water-use patterns, root growth, and stress-responsive proteins

Chickpea (Cicer arietinum L.), a rainfed crop in semi-arid regions, experiences frequent intermittent drought stress, hindering growth. This study explored morpho-physiological and molecular responses of six chickpea genotypes (ICC 4958, BG 4005, Pusa 362, BGM 10218, RSG 888, ICC 1882) under terminal drought to enhance yield and adaptability. Drought stress was imposed during early podding in a controlled glasshouse experiment. Various physiological, root growth, fertility, and yield parameters were measured. Proteomics analysis was conducted on leaf, pod, and root tissue samples, collected at ∼0.3 fraction of transpirable soil water (FTSW). Significant differences occurred for various traits, including phenologies, water relations, pollen viability and germination, flower and pod numbers, abortion rates, photosynthetic apparatus and pigments, osmotic adjustments, canopy temperatures, root length, and densities, and overall yield and biomass. FTSW dynamics can aid genotype screening during early drought. Characterizing stress-responsive proteins can provide insights into cellular processes and adaptations. Terminal drought affected leaf central shoot metabolism, reducing photosynthetic capacity, while roots and pods reconfigured enzyme abundances to promote root growth and pod development. The extensive genotypic variation in proteome data, morpho-physiological traits, and water-use patterns presents opportunities for developing drought-adapted cultivars.

Composition, function and driving factors of microbial communities in rhizosphere soil and root endophyte of Codonopsis pilosula in different years

Root microbial communities influence plant health and response to environmental stress. While the succession of rhizosphere soil and endophytic microbial communities in the growth stage of perennial medicinal plant Codonopsis pilosula remains unclear. In this study, rhizosphere soil and root tissue samples of one-year, two-year and three-year C. pilosula were selected to determine the community composition and diversity of rhizosphere soil and root endophyte microorganisms based on high-throughput sequencing technology. The results showed that the composition and structure of microbial community in rhizosphere soil and root endophyte of C. pilosula were different in different planting years. The abundance of Proteobacteria has decreased significantly, while that of Acidobacteria and Ascomycota have increased significantly. The neutral community model showed that the bacteria were mainly affected by random assembly and fungi by deterministic assembly. The complexity of microbial network in rhizosphere soil was higher than that in the root endophyte, and the complexity of bacterial network was higher than that of fungal community. Environmental factors significantly affected the microbial diversity of C. pilosula root system, while the growth years had little effect on it. Soil protease (PRO), soil total nitrogen (TN) and microbial biomass carbon (MBC) were the key factors causing the changes of C. pilosula microbiome. This study provides insights into the dynamic changes and driving factors of root microbial community of C. pilosula, and provides a basis for regulating root microbial community in C. pilosula cultivation to improve the sustainability of agricultural production.

The Zoige pioneer plant Leymus secalinus has different endophytic bacterial community structures to adapt to environmental conditions.

Leymus secalinus is a pioneer plant grown in the Zoige desertified alpine grassland and it is also one of the dominant plant species used for environmental remediation. L. secalinus plays a large role in vegetation reconstruction in sandy land, but the abundance and diversity of its endophytes have not yet been investigated. This study was performed to investigate the changes in the endophytic bacterial community structure of L. secalinus under different ecological environments and to analyze the effects of environmental changes and different plant tissues on the L. secalinus endophytic bacteria. Leaf, stem, and root tissue samples of L. secalinus were collected from Zoige Glassland (Alpine sandy land) and an open field nursery (Control). DNA was extracted and the 16S ribosomal DNA was amplified. The sequence library was sequenced on an Illumina MiSeq platform and clustered by operational taxonomic units (OTUs). α-diversity and β-diversity analyses, species diversity analyses, functional prediction, and redundancy (RDA) analyses for the soil physicochemical properties were conducted. α-diversity and β-diversity analyses showed that the endophytic bacteria in L. secalinus varied in different areas and tissues. The abundance of Allorhizobium-Neorhizobium-Pararhizobium-Rhizobium, which is related to nitrogen fixation, increased significantly in the L. secalinus found in the Zoige Grassland.Moreover, the abundance of nutrition metabolism and anti-stress abilities increased in functional prediction in the desert samples. The soil physicochemical properties had an insignificant influence on bacterial diversity. The changes in the endophytic bacterial community structure in L. secalinus were significant and were caused by environmental alterations and plant choice. The endophytic bacteria in L. secalinus grown in alpine sandy land may have greater anti-stress properties and the ability to fix nitrogen, which has potential value in environmental remediation and agricultural production.

Fusarium spp. associated with Chenopodium quinoa crops in Colombia

Quinoa is a plant commonly-resistance to adverse biotic and abiotic factors. However, this crop can be affected by phytopathogenic fungi. There is a lack of knowledge about the fungi associated with quinoa plants in Colombia. Through morphological and molecular identification in this study were identified four Fusarium species associated with quinoa crops: Fusarium oxysporum, Fusarium graminearum, Fusarium equiseti, and Fusarium culmorum. For this, we collected samples of panicles, leaf tissue, root tissue, and soil for isolation of different isolates of Fusarium. We performed a pathogenicity test of the fungi strains, under greenhouse conditions to evaluate the pathogenicity in seedlings of the Piartal cultivar with two inoculation methods. First inoculating the stem through a nodal wound or second inoculating the abaxial face with a brush. The results indicate the presence of four species with both molecular markers, phylogenetically distributed in these groups. The four species turned out to be pathogenic but with different degrees of virulence with significant differences between F. graminearum and F. oxysporum depending on the inoculation method. This is the first report on the presence of Fusarium species isolated from Quinoa in Colombia.

TMT-Based Comparative Peptidomics Analysis of Rice Seedlings under Salt Stress: An Accessible Method to Explore Plant Stress-Tolerance Processing.

Rice (Oryza sativa L.) is an important staple crop, particularly in Asia, and abiotic stress conditions easily reduce its yields. Salt stress is one of the critical factors affecting rice growth and yield. In this study, a tandem mass tag (TMT)-based comparative peptidomics analysis of rice seedlings under salt stress was conducted. Rice seedlings were exposed to 50 and 150 mM NaCl for 24 and 72 h, respectively, and the root and shoot tissues of different treatment groups were collected separately for peptidomics analysis. A total of 911 and 1263 nonredundant peptides were identified in two pooled shoot tissue samples, while there were 770 and 672 nonredundant peptides in two pooled root tissue samples, respectively. Compared with the control groups, dozens to hundreds of differentially expressed peptides (DEPs) were characterized in all treatment groups. To explore the potential functions of these DEPs, we analyzed the basic characteristics of DEPs and further analyzed the annotated Gene Ontology terms according to their precursor proteins. Several DEP precursor proteins were closely related to the response to salt stress, and some were derived from the functional domains of their corresponding precursors. The germination rate and cotyledon greening rate of transgenic Arabidopsis expressing two DEPs, OsSTPE2 and OsSTPE3, were significantly enhanced under salt stress. The described workflow enables the discovery of a functional pipeline for the characterization of the plant peptidome and reveals two new plant peptides that confer salinity tolerance to plants. Data are available via ProteomeXchange with identifier PXD037574.

Genome-wide association, RNA-seq and iTRAQ analyses identify candidate genes controlling radicle length of wheat.

The radicle, present in the embryo of a seed, is the first root to emerge at germination, and its rapid growth is essential for establishment and survival of the seedling. However, there are few studies on the critical mechanisms underlying radicle and then radicle length in wheat seedlings, despite its importance as a food crop throughout the world. In the present study, 196 wheat accessions from the Huanghuai Wheat Region were screened to measure radicle length under 4 hydroponic culture environments over 3 years. Different expression genes and proteins (DEGs/DEPs) between accessions with extremely long [Yunong 949 (WRL1), Zhongyu 9,302 (WRL2)] and short roots [Yunong 201 (WRS1), Beijing 841 (WRS2)] were identified in 12 sets of root tissue samples by RNA-seq and iTRAQ (Isobaric tags for relative and absolute quantification). Phenotypic results showed that the elongation zone was significantly longer in root accessions with long roots compared to the short-rooted accessions. A genome-wide association study (GWAS) identified four stable chromosomal regions significantly associated with radicle length, among which 1A, 4A, and 7A chromosomes regions explained 7.17% to12.93% of the phenotypic variation. The omics studies identified the expression patterns of 24 DEGs/DEPs changed at both the transcriptional and protein levels. These DEGs/DEPs were mainly involved in carbon fixation in photosynthetic organisms, photosynthesis and phenylpropanoid biosynthesis pathways. TraesCS1A02G104100 and TraesCS2B02G519100 were involved in the biosynthesis of tricin-lignins in cell walls and may affect the extension of cell walls in the radicle elongation zone. A combination of GWAS and RNA-seq analyses revealed 19 DEGs with expression changes in the four accessions, among which, TraesCS1A02G422700 (a cysteine-rich receptor-like protein kinase 6, CRK6) also showed upregulation in the comparison group by RNA-seq, iTRAQ, and qRT-PCR. BSMV-mediated gene silencing also showed that TaCRK6 improves root development in wheat. Our data suggest that TaCRK6 is a candidate gene regulating radicle length in wheat.

First report of Fusarium oxysporum causing root rot of Gentiana scabra bunge (Adenophora capillaris) in China.

Gentiana scabra bunge (Adenophora capillaris) is a traditional Chinese medicine crop in the northeast China. In July 2019, the root rot symptoms of A. capillaris were observed in its production field (approximately 1.3 hectares) iningyuan Manchu Autonomous County (41º47'28" N, 124º21'35" E), Fushun City in Liaoning province, China. Typical symptom included wilting, darkening, and rotting of the root collar and vascular bundle, leading to plant defoliation and death. Approximately 25% of the plants in the field showed the symptoms, with 2-year-old plants having more severe symptoms. Root tissue samples were collected from the diseased plants, surface-sterilized with 75% ethanol for 30 s, followed by 2% NaClO for 5 min, rinsed three times in sterile distilled water, and plated onto potato dextrose agar (PDA). After 3 days of incubation at 25 °C, white Fusarium-like colonies grew out from the symptomatic root tissue pieces. Five single-spore isolates were obtained from 10-day-old cultures using single-spore isolating technique. The fungus produced many macroconidia with the typical macroconidia of Fusarium spp. on carnation leaf agar (CLA) after 18 days of incubation at 26°C. They were falculate, slender and slightly curved, and their cells at both ends were sharp. Macroconidia had 3 to 5 septa, measuring 24.8 to 48.6 × 3.5 to 6.4 μm (n=50). Microconidia had 1 to 2 septa, elliptical, rounded tip, measuring 6.7 to 22.5 × 2.4 to 5.5 μm (n=5). Morphologically, the isolates were identified as Fusarium oxysporum (Leslie and Summerell 2006). For molecular identification, the internal transcribed spacer (ITS) region of rDNA and the translation elongation factor-1a (TEF-1α) of Isolate LD528-1 were amplified with the general primer ITS1/ITS4 and TEF-1α primer EF-1/EF-2 (O'Donnell et al. 1998). The resulting sequences were deposited in GenBank (acc. nos. MW418098 and MW423622). BLASTn analysis of the ITS sequence (KU939043) and TEF sequence (MW423622) revealed 99.06% sequence identity with F. oxysporum (KU939043) and 100% with F. oxysporum (MN892354), respectively. For pathogenicity test, a pot experiment was conducted in a greenhouse with 22 to 28°C and 65 to 90% relative humidity. Roots of A. capillaris were dipped in a spore suspension (1×107 conidia/ml) of Isolate LD528-1 for approximately 5 min, and then planted into the pots filled with sterilized field soil. Root dipped in sterilized water served as the controls. There were five pots for the inoculation treatment and three pots for the control treatment. All treated pots were placed and maintained in the greenhouse. After 15 days, 80% of inoculated plants were infected, with the symptoms similar to those observed in the field. The plants in the control treatment did not develop any symptoms. The same fungus was re-isolated from the diseased root tissue and confirmed by morphological and molecular assays as described above. This is the first report of F. oxysporum causing root rot of gentiana scabra bunge in China. This disease can become one of most important diseases in gentiana scabra bunge in China.

Comparison of mangrove (Avicennia marina) metal tissue concentrations to ambient sediment with an extensive range of contaminant levels in a highly-modified estuary (Sydney estuary, Australia)

Statistically distinct ‘high’, ‘moderate’ and ‘low’ metal contamination positions were distinguished across five Sydney estuary embayments for total sedimentary metal concentrations of Ni, Pb and Zn. While statistically distinct total sedimentary As, Cd, Co and Cu concentrations were indicated for the ‘low’ position samples. Mangrove (Avicennia marina) pneumatophore tissue metal concentrations from ‘high’ position trees were statistically distinct for Cd, Ni, Pb and Zn, while root tissue samples from ‘low’ position trees were statistically distinct for As, Cd and Zn. Outcomes from CAP, simple linear regression and DISTLM modelling indicated sedimentary metals were predominantly assimilated into mangrove root tissue, with smaller uptake into pneumatophore tissue. A comparison of floristic tissue metal concentrations indicated mangrove (A. marina) root tissue assimilated higher sedimentary metal concentrations than leaf and root tissue of seagrasses (Zostera capricorni and Halophila ovalis), which had relatively higher metal concentrations than those from pneumatophore and leaf mangrove tissue.

Assessment of environmental microbial effects of insect-resistant transgenic Populus × euramericana cv. ‘74/76’ based on high-throughput sequencing

Pests and diseases present major obstacles to poplar growth, which have been overcome via transgenic engineering applications. However, given the long growth cycle of transgenic poplars, long-term environmental safety assessments are needed for transgenic poplars planted in the field. Therefore, we investigated the effects of exogenous genes in transgenic poplar on the diversity of endophytic and rhizosphere soil bacterial communities in poplars. The composition and diversity of bacterial communities in trunk tissue, root tissue, and rhizosphere soil samples of transgenic Populus × euramericana cv. ‘74/76’ containing the high-resistance gene HPb1 (hereafter, HPb1) and non-transgenic Populus × euramericana cv. ‘74/76’ (hereafter, 107) in a 4-year-old experimental forest were analyzed using high-throughput sequencing. In terms of bacterial abundance at the phylum level, in trunk tissue, Bacteroidetes was the dominant phylum in both poplar lines (HPb1: 32.17%; 107: 47.42%). In root tissue, Proteobacteria was the dominant phylum in both lines (HPb1: 67.52%; 107: 65.96%). In rhizosphere soil, Proteobacteria was the dominant phylum in both lines (HPb1: 39.75%; 107: 37.98%). There were no significant differences in the richness and diversity of bacterial communities between HPb1 and 107. Finally, the bacterial community compositions in root tissue and rhizosphere soil were highly similar, both of which differed somewhat from that in trunk samples. This study provides a reference for ecological safety assessments of transgenic poplars and allows for a more systematic understanding of the potential ecosystem risk posed by environmental release of transgenic poplars.

An improved strategy to analyse strigolactones in complex sample matrices using UHPLC\u2013MS/MS

BackgroundStrigolactones represent the most recently described group of plant hormones involved in many aspects of plant growth regulation. Simultaneously, root exuded strigolactones mediate rhizosphere signaling towards beneficial arbuscular mycorrhizal fungi, but also attract parasitic plants. The seed germination of parasitic plants induced by host strigolactones leads to serious agricultural problems worldwide. More insight in these signaling molecules is hampered by their extremely low concentrations in complex soil and plant tissue matrices, as well as their instability. So far, the combination of tailored isolation—that would replace current unspecific, time-consuming and labour-intensive processing of large samples—and a highly sensitive method for the simultaneous profiling of a broad spectrum of strigolactones has not been reported.ResultsDepending on the sample matrix, two different strategies for the rapid extraction of the seven structurally similar strigolactones and highly efficient single-step pre-concentration on polymeric RP SPE sorbent were developed and validated. Compared to conventional methods, controlled temperature during the extraction and the addition of an organic modifier (acetonitrile, acetone) to the extraction solvent helped to tailor strigolactone isolation from low initial amounts of root tissue (150 mg fresh weight, FW) and root exudate (20 ml), which improved both strigolactone stability and sample purity. We have designed an efficient UHPLC separation with sensitive MS/MS detection for simultaneous analysis of seven natural strigolactones including their biosynthetic precursors—carlactone and carlactonoic acid. In combination with the optimized UHPLC–MS/MS method, attomolar detection limits were achieved. The new method allowed successful profiling of seven strigolactones in small exudate and root tissue samples of four different agriculturally important plant species—sorghum, rice, pea and tomato.ConclusionThe established method provides efficient strigolactone extraction with aqueous mixtures of less nucleophilic organic solvents from small root tissue and root exudate samples, in combination with rapid single-step pre-concentration. This method improves strigolactone stability and eliminates the co-extraction and signal of matrix-associated contaminants during the final UHPLC–MS/MS analysis with an electrospray interface, which dramatically increases the overall sensitivity of the analysis. We show that the method can be applied to a variety of plant species.

The use of infrared spectroscopy and machine learning tools for detection ofMeloidogyneinfestations

AbstractPlant parasitic nematodes are generally soilborne pathogens that attack plants and cause economic losses in many crops. The infested plants show nonspecific symptoms or, often, are symptomless; therefore, diagnosis is performed by taking soil and root tissue samples. Here, we show that a combination of different infrared spectra analysis and machine learning algorithms can be used to detect plant parasitic nematode infestations before symptoms become visible, using leaves instead of roots and soil as samples. We found that tomato and guava plants infested withMeloidogyne enterorlobiiproduced different spectral patterns compared to uninfested plants. Using partial spectra from 1,450 to 900/cm as the "fingerprint region", principal component analyses indicated that after 5 (tomatoes) or 8 weeks (guava), plants with no visible symptoms of infestations were positively diagnosed. To improve the early detection response, we used machine learning modelling. A support vector machine (SVM) was used to obtain more robust, accurate models. The SVM model contained 34 support vectors, 17 for each level. The overall performance of the model was >97% and the total accuracy was significantly higher, demonstrating the absence of chance prediction. The best prediction of infestation was obtained at the second and fourth weeks for tomatoes and guavas, respectively, reducing the diagnostic time by half. The combined application of these techniques reduces the processing time from field to laboratory and shows enormous advantages by avoiding root and soil sampling.

Shedding light on response of Triticum aestivum cv. Kharchia Local roots to long-term salinity stress through transcriptome profiling

Among the various abiotic stresses, salinity is one of the major limitations for production and productivity of wheat. Kharchia Local is the most salt-tolerant wheat cultivar developed from farmers’ selection on salt affected areas of India. Here, to investigate the molecular response of Kharchia Local under salinity stress, transcriptome sequencing of root tissue samples at the anthesis stage was performed. Illumina sequencing generated a total of 82.84 million clean reads and were assembled into 1,18,200 unigenes. A set of 10,805 unigenes were differentially expressed in response to salinity stress. Around 8232 unigene-derived SSRs were mined from these DEGs that can be used as functional molecular markers. Expression pattern of salinity stress-responsive unigenes was validated using real time PCR and results were found to be consistent with that of transcriptome profiling. Functional annotation of DEGs against GO, KEGG, COG and BLASTX using nr protein database was performed. This revealed the upregulation of genes involved in various biological processes including ROS homeostasis, ion transport, signal transduction, ABA biosynthesis and osmoregulation. Genes encoding expansin, xyloglucan endotransglucosylase/hydrolase, dehydrins and peroxidases that take part in enhancement of root growth were found to be upregulated under salinity. This could be the reason for better root growth of Kharchia Local under long-term salinity stress as compared to its susceptible counterpart. The present investigation provides primary information on transcriptome profiling of Kharchia Local roots under long-term salinity stress at the anthesis stage. In conclusion, the data generated in this study provide useful insights in understanding the molecular mechanism of salinity stress tolerance and will also serve as a valuable genomic reservoir for functional characterization of salinity responsive genes to develop tolerant genotypes.

Trichoderma erinaceum Bio-Priming Modulates the WRKYs Defense Programming in Tomato Against the Fusarium oxysporum f. sp. lycopersici (Fol) Challenged Condition.

The beneficial association and interaction of rhizocompetent microorganisms are widely used for plant biofertilization and amelioration of stress-induced damage in plants. To explore the regulatory mechanism involved in plant defense while associating with beneficial microbial species, and their interplay when co-inoculated with pathogens, we evaluated the response of tomato defense-related WRKY gene transcripts. The present study was carried out to examine the qRT–PCR-based relative quantification of differentially expressed defense-related genes in tomato (Solanum lycopersicum L.; variety S-22) primed with Trichoderma erinaceum against the vascular wilt pathogen (Fusarium oxysporum f. sp. lycopersici). The tissue-specific and time-bound expression profile changes under the four different treatments “(unprimed, Fol challenged, T. erinaceum primed and Fol+ T. erinaceum)” revealed that the highest upregulation was observed in the transcript profile of SlWRKY31 (root) and SlWRKY37 (leaf) in T. erinaceum bioprimed treated plants at 24 h with 16.51- and 14.07-fold increase, respectively. In contrast, SlWRKY4 showed downregulation with the highest repression in T. erinaceum bioprimed root (24 h) and leaf (48 h) tissue samples with 0.03 and 0.08 fold decrease, respectively. Qualitative expression of PR proteins (chitinases and glucanases) was found elicited in T. erinaceum primed plants. However, the antioxidative activity of tomato superoxide dismutase and catalase increased with the highest upregulation of SOD and SlGPX1 in Fol + T. erinaceum treatments. We observed that these expression changes were accompanied by 32.06% lesser H2O2 production in T. erinaceum bioprimed samples. The aggravated defense response in all the treated conditions was also reflected by an increased lignified stem tissues. Overall, we conclude that T. erinaceum bio-priming modulated the defense transcriptome of tomato after the Fol challenged conditions, and were accompanied by enhanced accumulation of defense-related WRKY transcripts, increased antioxidative enzyme activities, and the reinforcements through a higher number of lignified cell layers.

Understanding the Role of the WRKY Gene Family under Stress Conditions in Pigeonpea (Cajanus Cajan L.).

Pigeonpea (Cajanus cajan L.), a protein-rich legume, is a major food component of the daily diet for residents in semi-arid tropical regions of the word. Pigeonpea is also known for its high level of tolerance against biotic and abiotic stresses. In this regard, understanding the genes involved in stress tolerance has great importance. In the present study, identification, and characterization of WRKY, a large transcription factor gene family involved in numerous biological processes like seed germination, metabolism, plant growth, biotic and abiotic stress responses was performed in pigeonpea. A total of 94 WRKY genes identified in the pigeonpea genome were extensively characterized for gene-structures, localizations, phylogenetic distribution, conserved motif organizations, and functional annotation. Phylogenetic analysis revealed three major groups (I, II, and III) of pigeonpea WRKY genes. Subsequently, expression profiling of 94 CcWRKY genes across different tissues like root, nodule, stem, petiole, petal, sepal, shoot apical meristem (SAM), mature pod, and mature seed retrieved from the available RNAseq data identified tissue-specific WRKY genes with preferential expression in the vegetative and reproductive stages. Gene co-expression networks identified four WRKY genes at the center of maximum interaction which may play a key role in the entire WRKY regulations. Furthermore, quantitative real-time polymerase chain reaction (qRT-PCR) expression analysis of WRKY genes in root and leaf tissue samples from plants under drought and salinity stress identified differentially expressed WRKY genes. The study will be helpful to understand the evolution, regulation, and distribution of the WRKY gene family, and additional exploration for the development of stress tolerance cultivars in pigeonpea and other legumes crops.