Antagonistic Potential of Bacterial Species against Fungal Plant Pathogens(FPP)and Their Role in Plant Growth Promotion(PGP):A Review

Since the 19th century to date,the fungal pathogens have been involved in causing devastating diseases in plants.All types of fungal pathogens have been observed in important agricultural crops that lead to significant pre and postharvest losses.The application of synthetic fungicide against the fungal plant pathogens(FPP)is a traditional management practice but at the same time these fungicides kill other beneficial microbes,insects,animal,and humans and are harmful to environment.The antagonistic microorganism such as bacteria are being used as an alternate strategy to control the FPP.These antagonistic species are cost-effective and eco-friendly in nature.These biocontrol bacteria have a broad mechanism against fungal pathogens present in the phyllosphere and rhizosphere of the plant.The antagonistic bacteria have different strategies against the FPP,by producing siderophore,biofilm,volatile organic compounds(VOCs),through parasitism,antibiosis,competition for limited resources and induce systemic resistance(ISR)in the host plant by activating the immune systems.The commercial bio-products synthesized by the major bacterial species Pseudomonas syringae,Burkholderia cepacia,Streptomyces griseoviridis,Pseudomonas fluorescens and Bacillus subtilis are used to control Fusarium,Pythium,Rhizoctonia,Penicillium,Alternaria,and Geotrichum.The commercial bio-formulations of bacteria act as both antifungal and plant growth regulators.The Plant growth-promoting rhizobacteria(PGPR)played a significant role in improving plant health by nitrogen-fixing,phosphorus solubilization,phytohormones production,minimizing soil metal contamination,and by ACC deaminase antifungal activities.Different articles are available on the specific antifungal activity of bacteria in plant diseases.Therefore,this review article has summarized the information on biocontrol activity of bacteria against the FPP and the role of PGPR in plant growth promotion.This review also provided a complete picture of scattered information regarding antifungal activities of bacteria and the role of PGPR.

Viral pathogens and plant parasitic nematode

This special focus is dedicated to three parts：i）One of the most ubiquitous viral pathogens of stone fruit tree,Plum pox virus（PPV）;ii）a re-emerging pathogen,Wheat streak mosaic virus（WSMV）of cereal crops in Central Europe;and iii）a less studied plant parasitic,cyst-forming nematode in cereal crops Heterodera avenae.

A LAMP-assay-based specific microbiota analysis reveals community dynamics and potential interactions of 13 major soybean root pathogens

Soybean root diseases are associated with numerous fungal and oomycete pathogens;however,the community dynamics and interactions of these pathogens are largely unknown.We performed 13 loop-mediated isothermal amplification(LAMP)assays that targeted specific soybean root pathogens,and traditional isolation assays.A total of 159 samples were collected from three locations in the Huang-Huai-Hai region of China at three soybean growth stages(30,60,and 90 days after planting)in 2016.In LAMP results,we found that pathogen communities differed slightly among locations,but changed dramatically between soybean growth stages.Phytophthora sojae,Rhizoctonia solani,and Fusarium oxysporum were most frequently detected at the early stage,whereas Phomopsis longicolla,Fusarium equiseti,and Fusarium virguliforme were most common in the later stages.Most samples(86%)contained two to six pathogen species.Interestingly,the less detectable species tended to exist in the samples containing more detected species,and some pathogens preferentially co-occurred in diseased tissue,including P.sojae–R.solani–F.oxysporum and F.virguliforme–Calonectria ilicicola,implying potential interactions during infection.The LAMP detection results were confirmed by traditional isolation methods.The isolated strains exhibited different virulence to soybean,further implying a beneficial interaction among some pathogens.

Activity of Azoxystrobin and SHAM to Four Phytopathogens

The study was conducted to make clear the activity of azoxystrobin to 4 plant pathogens and the synergistic effects of salicylhydroxamic acid （SHAM）, which acted on the alternative oxidase. It was also conducted to be aware of the mechanism of azoxystrobin in inhibition on mycelial respiration and the influence of SHAM. The activity test of azoxystrobin and SHAM was carried out with a mycelial linear growth test and spore germination test. Other related biological properties were also observed. Inhibition of azoxystrobin and SHAM on 4 pathogens was determined by using SP-II oxygraph system. Azoxystrobin inhibited mycelial growth in Colletotrichum capsici, Botrytis cinerea, Rhizoctonia solani, and Magnaporthe grisea, respectively; it also inhibited conidia germination, and conidia production in C. capsici, B. cinerea M. grisea, and sclerotia formation in R. solani. Moreover, it created stayed pigment biosynthesis in C. capsici and M. grisea somehow. Salicylhydroxamic acid enhanced inhibition by azoxystrobin. An oxygen consuming test of the mycelia showed that azoxystrobin inhibited all the 4 fungi＇s respiration in the early stages. With the concentration rising up, the effectiveness increased. However, as time went on, the respiration of the mycelia treated with fungicides recovered and SHAM could not inhibit the oxygen consuming. This reaction between the mycelia and the fungicides appeared not to initiate alternative respiration but rather the other mechanism created a lack of efficacy.

Disruption of non-classically secreted protein(MoMtp)compromised conidiation,stress homeostasis,and pathogenesis of Magnaporthe oryzae

Blast disease,caused by the hemibiotrophic ascomycete fungus,Magnaporthe oryzae,is a significant threat to sustainable rice production worldwide.Studies have shown that the blast fungus secretes vast arrays of functionally diverse proteins into the host cell for a successful disease progression.However,the final destinations of these effector proteins inside the host cell and their role in advancing fungal pathogenesis remain a mystery.Here,we reported that a putative mitochondrial targeting non-classically secreted protein(MoMtp)positively regulates conidiogenesis and appressorium maturation in M.oryzae.Moreover,MoM TP gene deletion mutant strains triggered a hypersensitive response when inoculated on rice leaves displaying that MoMtp is essential for the virulence of M.oryzae.In addition,cell wall and oxidative stress results indicated that MoMtp is likely involved in the maintenance of the structural integrity of the fungus cell.Our study also demonstrates an upregulation in the expression pattern of the MoMTP gene at all stages of infection,indicating its possible regulatory role in host invasion and the infectious development of M.oryzae.Furthermore,Agrobacterium infiltration and sheath inoculation confirmed that MoMtpGFP protein is predominantly localized in the host mitochondria of tobacco leaf and rice cells.Taken together,we conclude that MoMtp protein likely promotes the normal conidiation and pathogenesis of M.oryzae and might have a role in disturbing the proper functioning of the host mitochondria during pathogen invasion.

In Vitro Antifungal Activity of the Extract and Compound from Acorus tatarinowii Against Seven Plant Pathogenic Fungi

Acorus tatarinowii Schott is a traditional Chinese medicine plant and has multiple bioactivities in medicine and pesticide field. In this study, the antifungal compound 1,2-dimethoxy-4（2-propenyl） benzene was isolated from A. tatarinowii Schott by activity-directed isolation method, and the inhibitory activity of the extract and 1,2-dimethoxy-4（2-propenyl） benzene against seven plant pathogenic fungi was evaluated. The results showed that the extract and 1,2-dimethoxy-4（2- propenyl） benzene had high inhibitory activity against hyphal growth of Thielaviopsis paradoxa （de Seynes） V. Hohnel, Pestalotia mangiferae P. Henn., Fusarium oxysporum f. sp. niveum （E. F. Smith） Syn. et Hans., Alternaria alternate Tanaka, Colletotrichum musae （Berk et Curt） V. Arx, Sphaceloma fawcettii Jenk., and Mycosphaerella sentina （Fr.） Schroter. The EC50 values of extract were 1.6162, 1.6811, 1.1253, 3.5771, 1.7024, 2.2284, and 2.2221 g L^-1, respectively, and the EC50 values of 1,2-dimethoxy-4（2-propenyl） benzene were 0.1021, 0.0997, 0.0805, 0.1742, 0.1503, 0.1853, and 0.1924 g L^-1, respectively. 1,2-Dimethoxy-4（2-propenyl） benzene also inhibited spores germination of T. paradoxa （de Seynes） V. Hohnel and F. oxysporum f. sp. niveum （E. F. Smith） Syn. et Hans., with the inhibitory rates of 98.81 and 100% at a concentration of 0.4 g L^-1 after 8 h, respectively. 1,2-Dimethoxy-4（2-propenyl） benzene is a potential botanical antifungal agent for controling of plant fungal diseases.

Functional identification of phenazine biosynthesis genes in plant pathogenic bacteria Pseudomonas syringae pv. tomato and Xanthomonas oryzae pv. oryzae

Phenazines are secondary metabolites with broad spectrum antibiotic activity and thus show high potential in biological control of pathogens. In this study, we identified phenazine biosynthesis （phz） genes in two genome-completed plant pathogenic bacteria Pseudomonas syringae pv. tomato （Pst） DC3000 and Xanthomonas oryzae pv. oryzae （Xoo） PXO99A. Unlike the phz genes in typical phenazine-producing pseudomonads, phz homologs in Pst DC3000 and Xoo PXO99A consisted of phzC/D/E/F/G and phzC/E1/E2/F/G, respectively, and the both were not organized into an operon. Detection experiments demonstrated that phenazine-l-carboxylic acid （PCA） of Pst DC3000 accumulated to 13.4 IJg L-1, while that of Xoo PXO99A was almost undetectable. Moreover, Pst DC3000 was resistant to 1 mg mL-1 PCA, while Xoo PXO99A was sensitive to 50 IJg mL ~ PCA. Furthermore, mutation of phzF blocked the PCA production and significantly reduced the pathogenicity of Pst DC3000 in tomato, while the complementary strains restored these phenotypes. These results revealed that Pst DC3000 produces low level of and is resistant to phenazines and thus is unable to be biologically controlled by phenazines. Additionally, phz-mediated PCA production is required for full pathogenicity of Pst DC3000. To our knowledge, this is the first report of PCA production and its function in pathogenicity of a plant pathogenic P. syringae strain.

Identification of candidate soil microbes responsible for small-scale heterogeneity in strawberry plant vigour

Studies were conducted to identify candidate soil microbes responsible for observed differences in strawberry vigour at a small spatial scale, which was not associated with visual disease symptoms. Samples were obtained from the soils close to the rhizosphere of ‘big＇ and ‘small＇ plants from small plots which exhibited large local heterogeneity in plant vigour. A metabarcoding approach was used to profile bacterial and fungal compositions, using two primer pairs for 16 S ribosomal RNA genes（16S r DNA） and one for the fungal internal transcribed spacer（ITS） region. Of the two 16 S r DNA primer sets, the 341F/805 R resulted in sequences of better quality. A total 28 operational taxonomic units（OTUs） had differential relative abundance between samples from ‘big＇ and ‘small＇ plants. However, plausible biological explanation was only possible for three fungal OTUs. Two were possible phytopathogens： Verticillium spp. and Alternaria alternata although the latter has never been considered as a main pathogen of strawberry in the UK. For samples from ‘small＇ plants, the abundance of these OTUs was much greater than from ‘big＇ plants. The opposite was true for a mycorrhizal OTU. These results suggest that soil microbes related to crop production can be identified using metabarcoding technique. Further research is needed to assess whether A. alternata and Verticillium spp. could affect strawberry growth in the field.

Formation of (2E)-4-Hydroxy-2-nonenal and (2E)-4-Hydroxy-2-hexenal by Plant Enzymes: A Review Suggests a Role in the Physiology of Plants

It is demonstrated that (3Z)-nonenal (NON) and (3Z)-hexenal (HEX) are oxidized in a cascade by lipoxygenase (LOX) and hydroperoxide peroxygenase (HP peroxygenase) into (2E)-4-hydroxy-2- nonenal (HNE) and (2E)-4-hydroxy-2-hexenal (HHE), respectively. In turn, HNE inactivates LOX terminating the cascade. The hydroxy-alkenals produced serve to inhibit plant pathogens, which initiated the cascade. In addition to LOX, other unknown oxygenases may be involved in the cascade.

Host niche, genotype, and field location shape the diversity and composition of the soybean microbiome

Plant-associated microbes represent a key determinant of plant fitness through acquiring nutrients,promoting growth,and resisting to abiotic and biotic stresses.However,an extensive characterization of the bacterial and fungal microbiomes present in different plant compartments of soybean in field conditions has remained elusive.In this study,we investigated the effects of four niches(roots,stems,leaves,and pods),four genotypes(Andou 203,Hedou 12,Sanning 16,and Zhonghuang 13),and three field locations(Jining,Suzhou,and Xuzhou)on the diversity and composition of bacterial and fungal communities in soybean using 16S and internal transcribed spacer rRNA amplicon sequencing,respectively.The soybean microbiome significantly differed across organs.Host genotypes explained more variation in stem bacterial community composition and leaf fungal community composition.Field location significantly affected the composition of bacterial communities in all compartments and the effects were stronger in the root and stem than in the leaf and pod,whereas field location explained more variation in stem and leaf fungal community composition than in the root and pod.The relative abundances of potential soybean fungal pathogens also differed among host organs and genotypes,reflecting the niches of these microbes in the host and probably their compatibility to the host genotypes.Systematic profiling of the microbiome composition and diversity will aid the development of plant protection technologies to benefit soybean health.

Antimicrobial,herbicidal and antifeedant activities of mansonone E from the heartwoods of Mansonia gagei Drumm

In vitro biological activities including anti-phytopathogenic fungi, antibacterial, antifeedant and herbicidal activities of the extracts from the heartwoods of Mansonia gagei Drumm. were evaluated. The dichloromethane （DCM） extract displayed antifungal activity against four plant pathogenic fungi （Altemaria porri, Colletotrichum gloeosporioides, Fusarium oxysporum and Phytophthora parasitica） higher than the methanolic （MeOH） extract. The separation of the DCM extract using bioassay guided antifungal activity against P. parasiUca led to the isolation of mansorins A, B, and C, mansonones C, E, G and H. Among isolated compounds, mansonone E displayed the highest antifungal activity against P. parasitica, followed by mansonone C, mansorin B and mansonone G. This potent compound revealed the same minimum inhibitory concentrations （MIC） of 31 μg mL-1 against C. gloeosporioides and P. parasitica, and minimum fungicidal concentration （MFC） of 31 and 125 μg mL-1, respectively. Moreover, mansonone E exhibited highly significant antibacterial activity against both Xanthomonas oryzae pv. oryzae （Xoo） and X. oryzae pv. oryzicola （Xoc） with MIC and minimum bactericidal concentration （MBC） as 7.8 and 〉500μg mL-1, respectively. This compound furthermore could inhibit the feed of Spodoptera litura with 45.9% antifeedant and significantly herbicidal activity reduced the shoot and root growth of Brassica chinensis, Oryza sativa, Mimosa pigra and Echinochlooa crus-galli. Mansonone E has potential as a new natural pesticide for agricultural plant pathogen management.

Minor antifungal aromatic glycosides from the roots of Gentiana rigescens(Gentianaceae)

Two new phenolic glycosides, 2,3-dihydroxybenzoic acid methyl ester 3-O-β-o-glucopyranosyl-（1-6）-β-D-glucopyranoside （1） and 2,5-dihydroxylbenzofuran 5-O-β-D-xylopyranosyl-（1-6）-O-β-D-glucopyranoside （2）, were isolated as the minor chemical constituents from the roots of Gentiana rigescens, along with 15 known compounds. Their structures were elucidated by detailed spectroscopic analysis, including 1D, 2D NMR and chemical method. All of these compounds were isolated for the first time from the title plant. Moreover, compounds 1 and 2 were tested for the antifungal activities on three plant pathogens Peronophythora litchi, Glomerella cingulata, and Glorosprium musarum.

Lasiodiplodia pseudotheobromae causes stem canker of Chinese hackberry in China

Chinese hackberry(Celtis sinensis Pers.)is an adaptable species widely growing in southern China.The symptoms of canker on stems of seedlings were discovered mid-July 2017 in Shuyang,Jiangsu Province.The diseased portions of the stems were dark brown due to discoloured xylem.Some seedlings showed symptoms of wilting,leaf fall,twig dieback,and tissue discolouration.The outbreak period was concentrated in July and August,suggesting that the disease spread during summer months.Possible fungal causal agents were isolated from naturally infected canker tissue and discoloured xylem.The isolate from xylem tissue with a high frequency(>50%)was named Ls7 type.Pathogenicity tests were carried out on 4-yearold seedlings.The symptoms of canker began to develop 20 days after inoculation with Ls7 isolate and by day 35,there were dark,enlarged longitudinal lesions.A phylogenetic tree of the isolate was developed using the internal transcribed spacer,elongation factor-1α(tef1-α),β-tubulin gene(TUB)and RNA polymerase II subunit primer genes(RPB2).Based on morphological features and phylogenetic information,the pathogen was identifi ed as Lasiodiplodia pseudotheobromae.This is the fi rst report of L.pseudotheobromae causing canker on Chinese hackberry stems in China.

Functional genomics in the rice blast fungus to unravel the fungal pathogenicity

A rapidly growing number of successful genome sequencing projects in plant pathogenic fungi greatly increase the demands for tools and methodologies to study fungal pathogenicity at genomic scale. Magnaporthe oryzae is an economically important plant pathogenic fungus whose genome is fully sequenced. Recently we have reported the development and application of functional genomics platform technologies in M. oryzae. This model approach would have many practical ramifications in design and implementation of upcoming functional genomics studies of filamentous fungi aimed at understanding fungal pathogenicity.

Lipopeptide Antibiotics Produced by the Engineered Strain Bacillus subtilis GEB3 and Detection of Its Bioactivity

MALDI-TOF-MS technology was used for identification of lipopeptide antibiotics producedby GEB3 strain, a derivative of Bacillus subtilis 168 which was transformed by lpaB3gene. The result showed GEB3 only produced lipopeptide antibiotic surfactin. The analysisby LC-MS demonstrated that GEB3 produced standard surfactin isoforms with side chainlengths of 13,14 and 15 carbon atoms. The bioactivity detection of surfactin indicatedthat the surfactin produced by GEB3 had inhibition effect on plant pathogens Rhizoctoniasolani and Pyricularia oryzae.

Use of Waste Vegetable Biomasses Treated by Steam Explosion for the Horticultural Crop Protection

The purpose of this study was to assess the suppressive effect of Waste Vegetable Biomasses （WVBs） treated by the Steam Explosion technique in a continuous plant, against soil-borne plant pathogens. In order to assess their disease suppression, five WVBs （Miscanthus biomass, durum wheat straw, rice straw, corn stalk and wood shaving） and commercial compost were tested in vivo at three different doses （10, 20 and 30% of potting mix） on seven horticultural pathosystems plant/fungus： tomato/Phytophthora nicotianae, cucumber/Pythium ultimum, lettuce/Fusariurn oxysporum f. sp. lactucae, melordFusariurn oxysporum f. sp. melonis, bearffRhizoctonia solani, eggplant/Verticillium dahlie and fennel/Sclerotinia sclerotiorum. The results showed that the corn stalk was more efficient respect to Miscanthus, compost, wheat straw, rice straw and wood shaving in all the patbosystems and at all the doses tested. The corn stalk suppression ranged from 97% in eggplant/F, dahliae to 35% in lettuce/F, oxysporum f. sp. lactucae, and it was significantly higher with respect to the other substrates. In general, the wheat straw, rice straw and wood shaving were statistically found less efficient as suppressive substrate with respect to corn stalk, Miscanthus and compost at the 30% dose in four pathosystems In particular, the wood shaving suppressiveness ranged from 48% in eggplant/V, dahliae to 12% in lettuce/F, oxysporum f. sp. lactucae. The different suppressiveness observed could be attributed to different concentration of the microbial inhibitory substances （furfurals, organic acids and lignosulfonates） produced during the processing of fresh biomass.

Rice(Oryza sativa L.)plant protection using dual biological control and plant growth-promoting agents:Current scenarios and future prospects

Various microorganisms live in association with different parts of plants and can be harmful,neutral,or beneficial to plant health.Some microbial inhabitants of plants can control plant diseases by contesting with,predating on,or antagonizing plant pathogens and by inducing systems for plant defense.A range of methods,including plant growth-promoting microorganisms(PGPMs)as biological control agents(BCAs)(BCA-PGPMs)are used for the biological management and control of plant pathogens.Some BCAs interact with plants by inducing resistance or priming plants without direct interaction with the pathogen.Other BCAs operate via nutrient competition or other mechanisms to modulate the growth conditions for the pathogen.Generally,PGPMs can be applied alone or together with other chemicals or carriers to control various crop diseases.This review highlights the effective types of BCA-PGPMs and their applications,roles,carrier based-formulations,and responses to rice(Oryza sativa L.)pathogens.Future plant disease management prospects are promising,and growers’increasing demand for BCA-PGPM products can be exploited as an effective approach to the management of plant diseases,as well as to improve yield,environmental protection,biological resources,and agricultural system sustainability.

First record of Eutypella vitis causing branch dieback on new host trees in Canada

Eutypella vitis is reported for the first time on several new host trees viz.Fagus grandifolia,Fraxinus pennsylvanica and Syringa reticulata with associated branch dieback symptoms in the Niagara Region of Southern Ontario,Canada.Morphological characteristics and phylogenetic analyses of ITS sequence data confirmed the species identification.Eutypella vitis is known as a woody plant pathogen causing dieback of grapevines and other economically important fruit trees widely cultivated in Ontario.The fungus ability to infect several alternate new hosts can promote its faster and wider spread across vineyards and consequently increase a dieback hazard to viticulture in Canada.

Endophytes versus biotrophic and necrotrophic pathogens—are fungal lifestyles evolutionarily stable traits?

Endophytes infect living plant tissues without causing symptoms of disease.Indeed,many of them contribute to the resistance phenotype of their host.However,fungal endophytes are generally closely related to plant pathogens,fungi that either develop within living host tissue(biotrophic fungi)or that kill the host cells and then live in the dead tissue(necrotrophic fungi).We adopted a phylogenetic approach to investigate whether these strategies represent evolutionarily stable lifestyles and to elucidate their general phylogenetic relationships.We analysed 163 fungal strains for which we found information on the sequence of the 5.8S rRNA gene and the flanking internal transcribed spacer regions,the identity of the host plant and the concrete phenotypic outcome of the infection.A Maximum-Likelihood analysis combined with ancestral character mapping by maximum parsimony revealed that some fungal lineages had switched multiple times between a necrotrophic and an endophytic lifestyle.Ancestral character mapping indicated a minimum of four changes from an endophytic to a necrotrophic lifestyle,four changes in the opposite direction and eight changes among these lifestyles for which the direction could not be determined unambiguously.By contrast,biotrophs formed five clusters that did not contain necrotrophs or endophytes.Once biotrophy evolves there is apparently no regression to one of the other two lifestyles.We conclude that biotrophy usually represents a derived and evolutionarily stable trait,whereas fungi easily can switch between an endophytic and necrotrophic lifestyle at the evolutionary and even the ecological timescale.Future experimental studies should focus on the environmental or genetic changes that cause the rapid switches between these two phenotypically different lifestyles.

Effective methods for isolation and purification of extracellular vesicles from plants

Plant extracellular vesicles(EVs)play critical roles in the cross-kingdom trafficking of molecules from hosts to interacting microbes,most notably in plant defense responses.However,the isolation of pure,intact EVs from plants remains challenging.A variety of methods have been utilized to isolate plant EVs from apoplastic washing fluid(AWF).Here,we compare published plant EV isolation methods,and provide our recommended method for the isolation and purification of plant EVs.This method includes a detailed protocol for clean AWF collection from Arabidopsis thaliana leaves,followed by EV isolation via differential centrifugation.To further separate and purify specific subclasses of EVs from heterogeneous vesicle populations,density gradient ultracentrifugation and immunoaffinity capture are then utilized.We found that immunoaffinity capture is the most precise method for specific EV subclass isolation when suitable specific EV biomarkers and their corresponding antibodies are available.Overall,this study provides a guide for the selection and optimization of EV isolation methods for desired downstream applications.