First Report of <i>Fusarium striatum</i> Causing Root Rot Disease of <i>Panax notoginseng</i> in Yunnan,China

Panax notoginseng is a traditional Chinese medicinal plant. Root rot of P. notoginseng is one of the most serious diseases affecting P. notoginseng growth and causes wilted leaves, fewer lateral roots and rotten roots. Root rot is a soil-borne disease, and mainly occurs from June to August in Yunnan Province when the temperatures are high and the air is humid. In this study, the endophytic fungal genus Fusarium isolate E-2018.1.22-#3.2 was obtained from a P. notoginseng embryo. Fusarium isolate E-2018.1.22-#3.2 was identified as Fusarium striatum based on morphological characteristics and molecular analysis. The fungus was found to have conidiophores and macroconidia, and its ITS, LSU and TEF-1α genes shared 100%, 99.2% and 99% identities with the homologous genes of Fusarium striatum, respectively. Isolate F. striatum E-2018.1.22-#3.2 can cause root rot symptoms, including black, soft roots, fewer lateral roots and leaf wilt, in 93% of the experimental P. notoginseng plants, and could be re-isolated, fulfilling Koch’s postulates. When the P. notoginseng plants were treated with the fungicide pyraclostrobin, isolate F. striatum E-2018.1.22-#3.2 was unable to cause root rot. We have therefore demonstrated that F. striatum E-2018.1.22-#3.2 is able to cause root rot disease in P. notoginseng. This is the first report of root rot disease caused by F. striatum on P. notoginseng in China.     

Overexpression of <i>IbSINA5</i> Increases Cold Tolerance through a CBF SINA-COR Mediated Module in Sweet Potato

Seven in absentia (SINA) family proteins play a central role in plant growth, development and resistance to abiotic stress. However, their biological function in plant response to cold stress is still largely unknown. In this work, a seven in absentia gene IbSINA5 was isolated from sweet potato. Quantitative real-time polymerase chain reaction (qRT-PCR) analyses demonstrated that IbSINA5 was ubiquitously expressed in various tissues and organs of sweet potato, with a predominant expression in fibrous roots, and was remarkably induced by cold, drought and salt stresses. Subcellular localization assays revealed that IbSINA5-GFP fusion protein was mainly localized in cytoplasm and nucleus. Overexpression of IbSINA5 in sweet potato led to dramatically improved resistance to cold stress in transgenic plants, which was associated with the up-regulated expression of IbCOR (cold-regulated) genes, increased proline production, and decreased malondialdehyde (MDA) and H₂O₂ accumulation in the leaves of transgenic plants. Furthermore, transient expression of IbCBF3, a C-repeat binding factor (CBF) gene, in the leaf protoplasts of wild type sweet potato plants up-regulated the expression of both IbSINA5 and IbCOR genes. Our results suggest that IbSINA5 could function as a positive regulator in the cold signaling pathway through a CBF-SINA-COR mediated module in sweet potato, and have a great potential to be used as a candidate gene for the future breeding of new plant species with improved cold resistance.     

Differential Responses of <i>NHX1</i> and <i>SOS1</i> Gene Expressions to Salinity in two <i>Miscanthus sinensis</i> Anderss. Accessions with Different Salt Tolerance

The lignocellulosic crop Miscanthus spp. has been identified as a good candidate for biomass production. The responses of Miscanthus sinensis Anderss. to salinity were studied to satisfy the needs for high yields in marginal areas and to avoid competition with food production. The results indicated that the relative advantages of the tolerant accession over the sensitive one under saline conditions were associated with restricted Na⁺ accumulation in shoots. Seedlings of two accessions (salt-tolerant ‘JM0119’ and salt-sensitive ‘JM0099’) were subjected to 0 (control), 100, 200, and 300 mM NaCl stress to better understand the salt-induced biochemical responses of genes involved in Na⁺ accumulation in M. sinensis. The adaptation responses of genes encoding for Na⁺ /H⁺ antiporters, NHX1 and SOS1 to NaCl stress were examined in JM0119 and JM0099.The cDNA sequences of genes examined were highly conserved among the relatives of M. sinensis based on the sequencing on approximate 600 bp-long cDNA fragments obtained from degenerate PCR. These salt-induced variations of gene expression investigated by quantitative real-time PCR provided evidences for insights of the molecular mechanisms of salt tolerance in M. sinensis. The expression of NHX1 was up-regulated by salt stress in JM0119 shoot and root tissues. However, it was hardly affected in JM0099 shoot tissue except for a significant increase at the 100 mM salt treatment, and it was salt-suppressed in the JM0099 root tissue. In the root tissue, the expression of SOS1 was induced by the high salt treatment in JM0119 but repressed by all salt treatments in JM0099. Thus, the remarkably higher expression of NHX1 and SOS1 were associated with the resistance to Na⁺ toxicity by regulation of the Na⁺ influx, efflux, and sequestration under different salt conditions.     

Phenotype Analysis and Fine Mapping of the Male Sterile Mutant <i>ms10</i> in Rice (<i>Oryza sativa</i> L.)

There is a positive correlation between fertility and yield, and the decrease of fertility is bound to a greatly reduced crop yield. Male sterile mutants can be used in hybrid rice. Therefore, rice male sterility has an important value in research and application, and the study of related mutants is also very vital. The mutant ms10 (male sterile 10) reported in this study was induced by ethyl methane sulfonate (EMS) in the indica maintainer line Xinong 1B. There was no significant difference between the ms10 and wild type in the vegetative growth stage. However, in the reproductive growth stage, ms10 showed that the plant became shorter, the anther became smaller and the color became lighter, and finally showed the phenotype of male sterility in comparison to the wild type. I₂-KI staining showed that the pollen was malformed and only a little was active. Scanning electron microscopy observation showed that the exine waxy layer of the ms10 anther decreased, suggesting that the protective effect on pollen was decreased. This may be one of the reasons leading to the phenotype of male sterility. Finally, the pollen showed shrinkage and collapsed, and the structure of germinating pore cover disappeared. This may be the result of sterility. Genetic analysis showed that the male sterility phenotype of the mutant was controlled by a single recessive nuclear gene. MS10 was mapped between the molecular markers IND37 and IND51 on chromosome 4, with a physical distance of 178.6 kb. These results lay the foundation for further studies on MS10.     

AM Fungi and <i>Piriformospora indica</i> Improve Plant Growth of <i>Pinus elliottii</i> Seedlings

Pinus elliottii is an exotic afforestation pine extensively distributed in southern parts of China. In order to understand whether endophytic fungi can affect seedling growth of P. elliottii, Piriformospora indica (Pi), Funnelifcrmis mosseae (Fm), and Diversispora tortuosa (Dt) were inoculated respectively, and the non-inoculated group was set as control. The growth indexes, the contents of soluble sugar and soluble protein, and plant endogenous hormone levels in the leaves of P. elliottii, were analyzed. The results showed that Fm, Dt and Pi colonized the P. elliottii roots to form mycorrhizal structure and chlamydospores arranged in beads respectively. Three fungal inoculants exhibited the stimulated growth responses, whilst Dt illustrated the most positive effect on plant height, single fresh weight, trunk diameter and root system structure, compared with the control. On the other hand, the soluble sugar and soluble protein contents were increased distinctively in mycorrhizal plants. The endogenous IAA, GA3, ZR contents were increased, while the ABA contents were reduced in mycorrhizal plants versus non-mycorrhizal plants. The fungi-induced endogenous hormone changes triggered plant growth improvement of P. elliottii seedlings. This research unraveled the positive effect of AM fungi and P. indica on growth of pine seedlings, while, more application of endophytic fungi to fields needs to be explored.     

<i>Aspergillus tubingensis</i> Causes Leaf Spot of Cotton (<i>Gossypium hirsutum</i> L.) in Pakistan

Cotton (Gossypium hirsutum L.) is a key fiber crop of great commercial importance. Numerous phytopathogens decimate crop production by causing various diseases. During July-August 2018, leaf spot symptoms were recurrently observed on cotton leaves in Rahim Yar Khan, Pakistan and adjacent areas. Infected leaf samples were collected and plated on potato dextrose agar (PDA) media. Causal agent of cotton leaf spot was isolated, characterized and identified as Aspergillus tubingensis based on morphological and microscopic observations. Conclusive identification of pathogen was done on the comparative molecular analysis of CaM and β-tubulin gene sequences. BLAST analysis of both sequenced genes showed 99% similarity with A. tubingensis. Koch’s postulates were followed to confirm the pathogenicity of the isolated fungus. Healthy plants were inoculated with fungus and similar disease symptoms were observed. Fungus was re-isolated and identified to be identical to the inoculated fungus. To our knowledge, this is the first report describing the involvement of A. tubingensis in causing leaf spot disease of cotton in Pakistan and around the world.     

Development of a New Cold-Tolerant Maize (<i>Zea mays</i> L.) Germplasm Using the <i>ICE1</i> Gene from <i>Arabidopsis thaliana</i>

To develop cold-tolerant maize germplasms and identify the activation of INDUCER OF CRT/DRE-BINDING FACTOR EXPRESSION (ICE1) expression in response to cold stress, RT-PCR was used to amplify the complete open reading frame sequence of the ICE1 gene and construct the plant expression vector pCAMBIA3301-ICE1-Bar. Immature maize embryos and calli were transformed with the recombinant vector using Agrobacterium tumefaciens-mediated transformations. From the regenerated plantlets, three T1 lines were screened and identified by PCR. A Southern blot analysis showed that a single copy of the ICE1 gene was integrated into the maize (Zea mays L.) genomes of the three T1 generations. Under low temperature-stress conditions (4°C), the relative conductivity levels decreased by 27.51%–31.44%, the proline concentrations increased by 12.50%–17.50%, the malondialdehyde concentrations decreased by 16.78%–18.37%, and the peroxidase activities increased by 19.60%–22.89% in the T1 lines compared with those of the control. A real-time quantitative PCR analysis showed that the ICE1 gene was ectopically expressed in the roots, stems, and leaves of the T1 lines. ICE1 positively regulates the expression of the CBF genes in response to cold stress. Thus, this study showed the successful transformation of maize with the ICE1 gene, resulting in the generation of a new maize germplasm that had increased tolerance to cold stress.     

蔓割病不同抗性甘薯品种的茎部细胞结构观察

蔓割病是我国南方薯区甘薯主要病害之一,本研究采用直接观察、显微和超微结构观测等方法,对高抗、中感和高感蔓割病的3个甘薯品种(高抗品种:金山57,中感品种:热薯1号,高感品种:新种花)接种蔓割病菌28 d植株充分发病后其茎基部细胞的侵染结构进行了观察。结果表明,在用清水作对照处理时,3个品种茎部组织的细胞形态和结构正常且完整,细胞代谢强。高抗品种金山57无论是接种组还是对照组,均未发现病原菌丝的存在,其茎下部、中部和上部细胞结构相对完整。中感品种热薯1号,蔓割病菌从其茎基部侵入后导致茎基部细胞破损坏死,而中部寄主-病原互作较为活跃和典型,造成养分运输受阻,茎基部接种后病原菌丝会沿着寄主茎部的维管束和其他组织一直向寄主的茎部末端蔓延,遭受侵染后的部位其细胞反应与高感品种新种花类似。高感品种新种花遭受蔓割病原菌侵染后,病原菌菌丝从茎基部新鲜剪口侵入后进入表皮细胞、皮层、维管束并在甘薯茎基部蔓延至中部、上部,直至寄主整株枯死,甘薯茎的木质部导管出现侵填体,质壁分离;与此同时,细胞壁的沉积物及乳突在病原菌的入侵处形成,各种无定型物质或纤丝构成的织网迅速包围入侵菌丝。     

Sexual Morph of <i>Furcasterigmium furcatum</i> (Plectosphaerellaceae) from <i>Magnolia liliifera</i> Collected in Northern Thailand

We isolated an interesting fungus from dead leaves of Magnolia liliifera collected from Chiang Mai, Thailand. The novel strain is related to Plectosphaerellaceae based on the morphology of its asexual morph and the analysis of sequence data. Phylogenetic analyses using a combined gene analysis of LSU and ITS sequence data showed that this strain is clustered in the same clade with Furcasterigmium furcatum with high statistical support. The new strains produced the asexual morph in culture which is morphologically similar to F. furcatum. Thus, we identified this strain as the sexual morph of F. furcatum. This is the first record of sexual morph for the monotypic genus Furcasterigmium and the first record of this genus on Magnolia.     

Cloning and Characterization of <i>EuGID1</i> in <i>Eucommia ulmoides</i> Oliver

Gibberellic acid controlled the key developmental processes of the life cycle of landing plants, and regulated the growth and development of plants. In this study, a novel gibberellin receptor gene EuGID1 was obtained from Eucommia ulmoides Oliver. The cDNA of EuGID1 was 1556 bp, and the open reading frame was 1029 bp, which encoded 343 amino acids. EuGID1 had the homology sequence with the hormone-sensitive lipase family. Amino acid sequence alignment confirmed EuGID1 protein had the highest homology with the GID1 protein of Manihot esculenta. EuGID1 was located in the nucleus and cell membrane and had expression in four plant organs. Overexpression of EuGID1 in transgenic Arabidopsis plants promoted plant elongation and increased siliques yield.     

Effects of <i>Piriformospora indica</i> on the Respiration of <i>Taxus chinensis</i> var. <i>mairei</i> under Water Stress

Seedlings of Taxus chinensis var. mairei were used as experimental materials to study the adaptation of Piriformospora indica to this plant under water stress. The materials were divided into two groups, namely, with or without inoculation with P. indica. Each group was subjected to four different levels of water stress. Vitality and physiological and biochemical indexes of the roots of T. chinensis var. mairei were regularly measured. Under water stress, T. chinensis var. mairei had significantly decreased root vitality; root vitality was higher in inoculated roots than in uninoculated roots. Under intense water stress, the inoculated roots had a higher soluble sugar content than the uninoculated roots. Under water stress, T. chinensis var. mairei experienced decreased activity of aerobic respiratory metabolic enzymes. The activity of anaerobic respiratory metabolic enzymes and alcohol dehydrogenase initially increased and then decreased, whereas that of lactate dehydrogenase increased. The inoculated roots had a higher activity of respiratory metabolic enzymes than the uninoculated roots. As water stress was further intensified, the roots had significantly decreased activity of aerobic respiratory metabolic enzymes and significantly increased activity of anaerobic respiratory metabolic enzymes. The activity of respiratory metabolic enzymes decreased faster in the uninoculated roots than in the inoculated roots. This study demonstrated that Piriformospora indica plays a positive role in enhancing the antihypoxic ability of T. chinensis var. mairei, thereby alleviating plant damage due to water stress.     

Direct effect of biocontrol agents on wilt and root-rot diseases of sesame

In Egypt, sesame cultivation is subject to attack by wilt and root-rot diseases caused by Fusarium oxysporum f.sp. sesami (Zap) Cast. and Macrophomina phaseolina (Maubl) Ashby causing losses in quality and quantity of sesame seed yield. Bacillus subtilis and Trichoderma viride isolates which were isolated from sesame rhizosphere were the most effective to antagonise fungal pathogens, causing high reduction of hyphal fungal growth. Trichoderma viride was found to be mycoparasitic on Fusarium oxysporum f.sp. sesami and M. phaseolina causing morphological atternation of fungal cells and sclerotial formation. In general, Bacillus subtilis, T. viride, avirulent Fusarium oxysporum isolate and Glomus spp. (Amycorrhizae) significantly reduced wilt and root-rot incidence of sesame plants at artificially infested potted soil by each one or two pathogens. Data obtained indicate that Glomus spp significantly reduced wilt and disease severity development on sesame plants followed by T. viride. Meanwhile, avirulent Fusarium oxysporum isolate followed by Glomus spp. were effective against root-rot disease incidence caused by M. phaseolina. Glomus spp. followed by B. subtilis significantly reduced wilt and root-rot disease of sesame plants. All biotic agents significantly reduced F. oxysporum f.sp. sesami and M. phaseolina counts in sesame rhizosphere at the lowest level. Glomus spp. and the avirulent isolate of F. oxysporum eliminated M. phaseolina in sesame rhizosphere. Meanwhile T. viride was the best agent at reducing F. oxysporum at a lower level than other treatments. Application of VA mycorrhizae (Glomus spp.) in fields naturally infested by pathogens significantly reduced wilt and root-rot incidence and it significantly colonised sesame root systems and rhizospheres compared to untreated sesame transplantings.     

Antibacterial Activity and Mechanisms of Ethanol Extracts from <i>Scutellaria baicalensis</i> Georgi and <i>Magnolia officinalis</i> against <i>Phytophthora nicotianae</i>

Phytophthora nicotianae causes substantial economic losses in most countries where tobacco is produced. At present, the control of P. nicotianae mainly depends on chemical methods, with considerable environmental and health issues. We investigated the effects of ethanol extracts from Scutellaria baicalensis Georgi (SBG) and Magnolia officinalis (MO). On mycelial growth, sporangium formation, and zoospore release of P. nicotianae. Both extracts inhibited the growth of P. nicotianae, with mycelial growth inhibition rates of 88.92% and 93.92%, respectively, at 40 mg/mL, and EC50 values of 5.39 and 5.74 mg/mL, respectively. The underlying mechanisms were the inhibition of sporangium formation, the reduction of zoospore number, and the destruction of the mycelium structure. At an SBG extract concentration of 16.17 mg/mL, the inhibition rates for sporangia and zoospores were 98.66% and 99.39%, respectively. At an MO extract concentration of 2.87 mg/mL, the production of sporangia and zoospores was completely inhibited. The hyphae treated with the two plant extracts showed different degrees of deformation and damage. Hyphae treated with SBG extract showed adhesion and local swelling, whereas treatment with MO extract resulted in broken hyphae. Mixture of the extracts resulted in a good synergistic effect.     

Overexpression of Wheat <i>TaELF3-1BL</i> Delays Flowering in Arabidopsis

EARLY FLOWERING 3 (ELF3), a light zeitnehmer (time-taker) gene, regulates circadian rhythm and photoperiodic flowering in Arabidopsis, rice, and barley. The three orthologs of ELF3 (TaELF3-1AL, TaELF3-1BL, and TaELF3-1DL) have been identified in wheat too, and one gene, TaELF3-1DL, has been associated with heading date. However, the basic characteristics of these three genes and the roles of the other two genes, TaELF3-1BL and, TaELF3-1AL, remain unknown. Therefore, the present study obtained the coding sequences of the three orthologs (TaELF3-1AL, TaELF3-1BL, and TaELF3-1DL) of ELF3 from bread wheat and characterized them and investigated the role of TaELF3-1BL in Arabidopsis. Protein sequence comparison revealed similarities among the three TaELF3 genes of wheat; however, they were different from the Arabidopsis ELF3. Real-time quantitative PCR revealed TaELF3 expression in all wheat tissues tested, with the highest expression in young spikes; the three genes showed rhythmic expression patterns also. Furthermore, the overexpression of the TaELF3-1BL gene in Arabidopsis delayed flowering, indicating their importance in flowering. Subsequent overexpression of TaELF3-1BL in the Arabidopsis ELF3 nonfunctional mutant (elf3 mutant) eliminated its early flowering phenotype, and slightly delayed flowering. The wild-type Arabidopsis overexpressing TaELF3-1BL demonstrated reduced expression levels of flowering-related genes, such as CONSTANS (AtCO), FLOWERING LOCUS T (AtFT), and GIGANTEA (AtGI). Thus, the study characterized the three TaELF3 genes and associated TaELF3-1BL with flowering in Arabidopsis, suggesting a role in regulating flowering in wheat too. These findings provide a basis for further research on TaELF3 functions in wheat.     

Formulation and Delivery of the Bacterial Antagonist Bacillus subtilis for Management of Lentil Vascular Wilt Caused by Fusarium oxysporum f. sp. lentis

Different formulations of Bacillus subtilis were prepared using standard laboratory protocols. Bacillus subtilis survived in glucose and talc powders at 8.6 and 7.8 log₁₀ CFU/g, respectively, for 1 year of storage at room temperature compared with 3.5 log₁₀ CFU/g on a peat formulation. Glasshouse experiments using soil and seed treatments were conducted to test the efficacy of B. subtilis for protecting lentil against the wilt disease caused by Fusariumoxysporum f. sp. lentis. Seed treatments with formulations of B. subtilis on glucose, talc and peat significantly enhanced its biocontrol activity against Fusarium compared with a treatment in which spores were applied directly to seed. The formulations decreased disease severity by reducing colonization of plants by the pathogen, promoting their growth and increased the dry weight of lentil plants. Of these treatments the glucose and talc‐based powder formulations were more effective than the peat formulation and the spore application without a carrier. It was shown that the B. subtilis spores applied with glucose were viable for longer than those applied with other carriers. Seed treatment with these formulated spores is an effective delivery system that can provide a conducive environment for B. subtilis to suppress vascular wilt disease on lentil and has the potential for utilization in commercial field application.     

<i>DWARF</i> and <i>SMALL SEED1</i>, a Novel Allele of <i>OsDWARF</i>, Controls Rice Plant Architecture,Seed Size,and Chlorophyll Biosynthesis

Plant architecture is a vital agronomic trait to control yield in rice (Oryza sativa L.). A dwarf and small seed 1 (dss1) mutant were obtained from the ethyl methanesulfonate (EMS) mutagenized progeny of a Guizhou glutinous landrace cultivar, Lipingzabianhe. The dss1 mutant displayed phenotypes similar to those of brassinosteroid (BR) deficient mutants, such as dwarfing, dark green and rugose erect leaves, small seeds, and loner neck internode panicles with primary branching. In our previous study, the underlying DSS1 gene was isolated, a novel allele of OsDWARF (OsBR6ox) that encodes a cytochrome P450 protein involved in the BR biosynthetic pathway by MutMap technology. In this work, we confirmed that a Thr335Ile amino acid substitution residing in DSS1/OsDWARF was responsible for the dwarf, panicle architecture, and small seed phenotypes in the dss1 mutants by genetic transformation experiments. The overexpression of OsDWARF in the dss1 mutant background could not only recover dss1 to the normal plant height and panicle architecture but also rescued normal leaf angles, seed size, and leaf color. Thus, the specific mutation in DSS1/OsDWARF influenced plant architecture, seed size, and chlorophyll biosynthesis.