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Background Whitefly, Bemisia tabaci (Genn.) (Hemiptera: Aleyrodidae), is one of the most harmful pests in greenhouses and in open fields. Main body The present study aimed to assess in vitro virulence of 3 entomopathogenic fungal strains (EPFs) of Lecanicillium lecanii 3 (V-3), 4 (V-4), and 5 (V-5) against the whitefly, Bemisia tabaci (Genn.) (Hemiptera: Aleyrodidae), by spray method. The 3 disparate bioassays were performed encompassed of conidial concentrations and fungal filtrate of the strains, V-3, V-4, and V-5, and also their binary combinations. In the filtrate bioassay, 2 ml of fungal filtrate of each strain was used. In the conidial bioassay, 3 disparate concentrations (1 × 105, 1 × 106, and 1 × 107 conidia ml−1) were used for each fungal strain, while in the binary combinations (1 ml filtrate + 1 ml conidia) of V-3 × V-3, V-4 × V-4, and V-5 × V-5 were used. Mortality rates against the whitefly were recorded on the 7th day. In the conidial bioassay, maximum mortality rates were found at V-3 strain (90.6%), V-4 strain (78.4%), and V-5 strain (83.6%) at the highest concentration (1 × 107 conidia ml−1) on the 7th day. In the filtrate bioassay, V-3 strain revealed a maximum mortality (93%), V-4 strain (85%), and then V-5 strain (87%) on the 7th day. Moreover, in the bioassay of binary combinations, the highest mortality rate of the whitefly was counted in V-3 × V-3 strain (84.6%), V-4 × V-4 strain (70.6%), and V-5 × V-5 strain (79.8%) on the 7th day. Conclusion All treatments had the potential to control B. tabaci significantly. In all bioassays, the V-3 strain was the extreme virulent, and the filtrate application of V-3 strain was the utmost impressive against B. tabaci.

Plants rely on various physiological and molecular defense mechanisms against biotic stresses such as herbivore insects. Many entomopathogenic fungi synthesize protein molecules that can trigger these plant defenses. This laboratory study characterized the bioactivity of a partially purified protein derived from Beauveria bassiana (ARSEF 2860) against whitefly, Bemisia tabaci (Hemiptera: Aleyrodidae), which is an economically important pest of agricultural and horticultural crops worldwide. Different concentrations (i.e., 0.021, 0.042 and 0.063 &mu, M) of fungal protein were bioassayed to determine their sub-lethal effect on the survival percentage and fecundity rate of B. tabaci on tomato (Lycopersicon esculentum) plants. In addition, the putative role of this partially purified B. bassiana protein in the defense mechanisms of plant was assessed through the expression analyses of important genes related to salicylic acid (SA)&mdash, and jasmonic acid (JA)&mdash, associated pathways using RT-qPCR. Results revealed a significant suppression of the survival percentage and fecundity rate of B. tabaci by the fungal protein. Lowest survival (41%) was recorded for the highest concentration of protein (0.063 &mu, M), whereas mean survival for the other two protein concentrations (0.042 and 0.021 &mu, M) were 62 and 71%, respectively. Likewise, the highest and lowest mean fecundity rates were observed for the control and the highest protein concentration (i.e., 3.3 and 1.8 eggs day&minus, 1 female&minus, 1, respectively). Furthermore, the exogenous application of B. bassiana-derived protein on tomato plants strongly up-regulated the SA-related genes (PAL, PR1, BGL2 and EDS1) and slightly up-regulated the JA-related genes (AOC, AOS, OPR3 and LOX) as compared to the control plants. These findings demonstrate the putative role of this partially purified B. bassiana protein fraction in inducing systemic resistance in the tomato plants against B. tabaci, suggesting its further purification and characterization to be used as novel biological pest control tool against B. tabaci and other sap-sucking insect pests.

Bacterial fruit blotch (BFB), caused by Acidovorax citrulli, seriously affects watermelon and other cucurbit crops, resulting in significant economic losses. However, the pathogenicity mechanism of A. citrulli is not well understood. Plant pathogenic bacteria often suppress the plant immune response by secreting effector proteins. Thus, identifying A. citrulli effector proteins and determining their functions may improve our understanding of the underlying pathogenetic mechanisms. In this study, a novel effector, AopN, which is localized on the cell membrane of Nicotiana benthamiana, was identified. The functional analysis revealed that AopN significantly inhibited the flg22-induced reactive oxygen species burst. AopN induced a programmed cell death (PCD) response. Unlike its homologous protein, the ability of AopN to induce PCD was dependent on two motifs of unknown functions (including DUP4129 and Cpta_toxin), but was not dependent on LXXLL domain. More importantly, the virulence of the aopN mutant of A. citrulli in N. benthamiana significantly decreased, indicating that it was a core effector. Further analysis revealed that AopN interacted with watermelon ClHIPP and ClLTP, which responds to A. citrulli strain Aac5 infection at the transcription level. Collectively, these findings indicate that AopN suppresses plant immunity and activates the effector-triggered immunity pathway.

Huanglongbing (HLB) is a devastating citrus disease that has caused massive economic losses to the citrus industry worldwide. The disease is endemic in most citrus-producing areas of southern China, especially in the sweet orange orchards where soil acidification has intensified. In this work, we used lime as soil pH amendment to optimize soil pH and enhance the endurance capacity of citrus against Candidatus Liberibacter asiaticus (CLas). The results showed that regulation of soil acidity is effective to reduce the occurrence of new infections and mitigate disease severity in the presence of HLB disease. We also studied the associated molecular mechanism and found that acid soil improvement can (i) increase the root metabolic activity and up-regulate the expression of ion transporter-related genes in HLB-infected roots, (ii) alleviate the physiological disorders of sieve tube blockage of HLB-infected leaves, (iii) strengthen the citrus immune response by increasing the expression of genes involved in SAR and activating the salicylic acid signal pathway, (iv) up-regulate 55 proteins related to stress/defence response and secondary metabolism. This study contributes to a better understanding of the correlation between environment factors and HLB disease outbreaks and also suggests that acid soil improvement is of potential value for the management of HLB disease in southern China.

Huanglongbing (HLB) is a devastating citrus disease that has caused massive economic losses to the citrus industry worldwide. The disease is endemic in most citrus-producing areas of southern China, especially in the sweet orange orchards where soil acidification has intensified. In this work, we used lime as soil pH amendment to optimize soil pH and enhance the endurance capacity of citrus against Candidatus Liberibacter asiaticus (CLas). The results showed that regulation of soil acidity is effective to reduce the occurrence of new infections and mitigate disease severity in the presence of HLB disease. We also studied the associated molecular mechanism and found that acid soil improvement can (i) increase the root metabolic activity and up-regulate the expression of ion transporter-related genes in HLB-infected roots, (ii) alleviate the physiological disorders of sieve tube blockage of HLB-infected leaves, (iii) strengthen the citrus immune response by increasing the expression of genes involved in SAR and activating the salicylic acid signal pathway, (iv) up-regulate 55 proteins related to stress/defence response and secondary metabolism. This study contributes to a better understanding of the correlation between environment factors and HLB disease outbreaks and also suggests that acid soil improvement is of potential value for the management of HLB disease in southern China.

A multipartite DNA virus was isolated from a plant pathogenic fungus, which is infectious as cloned DNA and purified virions., Here, we describe a tripartite circular single-stranded (ss) DNA mycovirus, named Fusarium graminearum gemytripvirus 1 (FgGMTV1). The genome of FgGMTV1 comprises three circular ssDNA segments (DNA-A, DNA-B, and DNA-C). Sequence alignments and phylogenetic analyses showed that FgGMTV1 is nested within the family Genomoviridae. We also constructed the first infectious DNA clones of a DNA mycovirus. Our results show that DNA-A and DNA-B are mutually interdependent for their replication and are associated with severely reduced colony growth and hypovirulence. DNA-C relies on DNA-A and DNA-B for replication and is necessary for the recovery of abnormal fungal phenotypes. DNA-C also enhances the accumulation of viral DNA in infected fungi and permits stable colonization and easy transmission via conidia. This is the first multipartite DNA virus isolated from a fungus. Our phylogenetic analyses also suggest that the multipartite genome of FgGMTV1 may have evolved from a monopartite genome of an ancient genomovirus.

PevD1, an Aa1-like protein secreted by Verticillium dahlia, promotes infection by inhibiting the antifungal activity of Gossypium hirsutum PR5. ., Alt a 1 family proteins (AA1s) have only been observed in the Dothideomycetes and Sordariomycetes classes of fungi, and their biological functions have remained poorly understood. Verticillium dahliae, a soil-borne pathogen that causes plant wilt disease, secretes hundreds of proteins during the process of pathogenic infection, including the AA1 member PevD1. In this study, we found that the pevd1 transcript was present in all of the hosts studied (cotton, Arabidopsis, tomato, and tobacco) and showed elevated expression throughout the infection process. Furthermore, pevd1 knockout mutants displayed attenuated pathogenicity compared with the wild-type (WT) strain and complemented strains in hosts. A partner protein of PevD1, pathogenesis-related protein 5 (PR5)-like protein GhPR5, was isolated from cotton (Gossypium hirsutum) plants by co-purification assays, and the PevD1–GhPR5 interaction was determined to be localized in the C-terminus (PevD1b, amino acids residues 113–155) by pull-down and yeast two-hybrid techniques. Re-introduction of the pevd1b gene into a pevd1 knockout mutant resulted in restoration of the virulence phenotype to WT levels. In addition, PevD1b, which is similar to PevD1, decreased the antifungal activity of GhPR5 in vitro. Our findings reveal an infection strategy in which V. dahliae secretes PevD1 to inhibit GhPR5 antifungal activity in order to overcome the host defence system.

Here we describe a novel (-)ssRNA mycovirus, Fusarium graminearum negative-stranded RNA virus 1 (FgNSRV-1), isolated from Fusarium graminearum strain HN1. The genome of FgNSRV-1 is 9072 nucleotides in length, with five discontinuous but linear ORFs (ORF I-V). Phylogenetic analysis based on entire L polymerase sequences indicated that FgNSRV-1 is related to the (-)ssRNA mycovirus Sclerotinia sclerotiorum negative-stranded RNA virus 1 (SsNSRV-1), and other mycoviruses. Our data suggest that FgNSRV-1 can be classified into the family Mymonaviridae, order Mononegavirales. Putative enveloped virion-like structures with filamentous morphology similar to SsNSRV-1 were observed in virion preparation samples. The L proteins of FgNSRV-1, and other fungal mononegaviruses, were found to be related to L protein-like sequences in some fungal genome, supporting the hypothesis that there is coevolution occurring between mycoviruses and fungi. Besides, clearing the virus from the infected host fungus resulted in no discernable phenotypic change.

Southern corn leaf blight (SCLB) caused by Bipolaris maydis is an important foliar disease of maize. In this study, a nematode-symbiotic bacterium Xenorhabdus budapestensis strain C72 was identified with remarkable inhibiting effect on mycelial growth and spore germination of B. maydis. The in vitro assay revealed that C72 cell-free culture media (CFCM) with thermostability exhibited broad-spectrum antifungal activities against other several important plant pathogenic fungi. The early colonization of B. maydis were significantly impaired by CFCM treatment under phytotron condition. This antagonism is likely to be the main contributor to the highly efficient plant protection of 40% (v/v) CFCM treatment against B. maydis, and the relative control effect reached to 59.15% and 77.96% in greenhouse and field experiments, which was comparable to the effect of fungicides. Moreover, we found that extracellular enzymes secreted by symbiotic bacterium may be one of the reasons for the antifungal potential of C72. Beside direct antagonistic effects provided by the bacterium, defense related genes were induced in maize after CFCM treatment. In summary, this study reported the first systematic evaluation of the effect of X. budapestensis C72 in controlling SCLB and exploration of its mode of action, then indicating that entomopathogenic bacteria have the potential to become a new and efficient biological control resource for plant fungal disease management.

The Botrytis cinerea BcSpl1 protein is a member of the cerato-platanin family, and consists of 137 amino acids and two disulfide bridges. This protein induces the onset of necrosis in infiltrated plant hosts. Recombinant BcSpl1 proteins produced in Pichia pastoris (pBcSpl1) and Escherichia coli (eBcSpl1) were initially compared regarding their abilities to induce necrosis and systemic acquired resistance (SAR). The pBcSpl1 and eBcSpl1 treatments led to the development of necrotic lesions on tomato leaves, and provided tomato plants with SAR to B. cinerea. The lesion area of leaves infiltrated with the BcSpl1 proteins decreased by 22.7% (pBcSpl1) and 21.8% (eBcSpl1). Additionally, eBcSpl1 up-regulated the expression levels of some defense-related genes, including PR-1a, prosystemin, PI I, and PI II, as well as SIPK and TPK1b, which encode two protein kinases. Furthermore, eBcSpl1 exhibited chitin-binding properties. Our data revealed that the E. coli expression system produces higher BcSpl1 yields than the P. pastoris system. This high-yield expression of BcSpl1 may be relevant for future large-scale applications of this elicitor to improve crop production.

Backgroud Huanglongbing (HLB) is currently one of the most destructive citrus disease worldwide. It is caused by Candidatus Liberibacter asiaticus (CLas), a nonculturable alpha-proteobacterium, which it resides exclusively in the phloem tissues. Therefore, understanding the early CLas-responsive proteins in citrus petiole where pathogenic bacteria colonized will help to investigate plant resistance to the pathogen.Results In this study, a comparative proteomic approach was applied to identify the petiole proteins associated with the response to CLas infection. A total of 777 proteins were differentially expressed in response to CLas. Among them, 499 proteins were up-regulated and 278 were down-regulated. Among the most highly up-regulated differentially expressed proteins (DEPs) were salicylate carboxymethyltransferase, ubiquitin carboxyl-terminal hydrolase 13, trans-resveratrol di-O-methyltransferase, linoleate 13S-lipoxygenase 2-1, granule-bound starch synthase 1 and thaumatin-like proteins. While the most highly down-regulated DEPs were oxygen-evolving enhancer proteins, ribulose bisphosphate carboxylase/oxygenase activase, peroxidases and photosystem reaction center subunits. The results of qPCR analysis of a number of indicated DEPs and western blotting further validated four representative DEPs, including salicylate carboxymethyltransferase, linoleate 13S-lipoxygenase 2-1, granule-bound starch synthase 1 and photosystem I reaction center subunit showed that most of detected DEPs were positively correlated with their mRNA and protein levels.Conclusions Our comparative proteomic analysis first profiling reveals early and primary proteome alterations in CLas-infected citrus petiole, where pathogens reside in. The DEPs results demonstrate that CLas infection could promote the carbohydrate metabolism, depress the photosystem and activate/inhibit defense responses.

In recent years, it is a hotspot research field on interaction mechanism between elicitor and plant. In this study, a novel hypersensitive response (HR)-inducing protein elicitor was isolated from the culture filtrate of Fusarium oxysporum f. sp. cubense and named PeFOC1, which consisted of 321 amino acids with a molecular weight of approximately 35 kDa. After the inducible expression in Escherichia coli and purification by AKTA explore system, the recombinant PeFOC1 also triggered a typical HR in tobacco. In addition, PeFOC1 induced a cascade of defense response in tobacco including production of hydrogen peroxide, deposition of callose, and accumulation of phenolic compounds. Moreover, PeFOC1 significantly improved systemic resistance of tobacco seedlings to tobacco mosaic virus and Pseudomonas syringae pv. tabaci. Real-time quantitative-PCR analysis indicated that several defense-related genes in tobacco, such as NtPR1a, NtNPR1, NtPAL, NtEDS1, NtPDF, and NtLOX, were all up-regulated by the treatment of PeFOC1. All these results collectively demonstrated that PeFOC1 triggered defense response and systemic acquired resistance (SAR) in tobacco. This research not only provides further research on immune mechanism between plant and elicitor, but also sheds new light on strategy for biocontrol in the future.

Panama disease, or Fusarium wilt, the most serious disease in banana cultivation, is caused by Fusarium oxysporum f. sp. cubense (FOC) and has led to great economic losses worldwide. One effective way to combat this disease is by enhancing host plant resistance. The cerato-platanin protein (CPP) family is a group of small secreted cysteine-rich proteins in filamentous fungi. CPPs as elicitors can trigger the immune system resulting in defense responses in plants. In this study, we characterized a novel cerato-platanin-like protein in the secretome of Fusarium oxysporum f. sp. cubense race 4 (FOC4), named FocCP1. In tobacco, the purified recombinant FocCP1 protein caused accumulation of reactive oxygen species (ROS), formation of necrotic reaction, deposition of callose, expression of defense-related genes, and accumulation of salicylic acid (SA) and jasmonic acid (JA) in tobacco. These results indicated that FocCP1 triggered a hypersensitive response (HR) and systemic acquired resistance (SAR) in tobacco. Furthermore, FocCP1 enhanced resistance tobacco mosaic virus (TMV) disease and Pseudomonas syringae pv. tabaci 6605 (Pst. 6605) infection in tobacco and improved banana seedling resistance to FOC4. All results provide the possibility of further research on immune mechanisms of plant and pathogen interactions, and lay a foundation for a new biological strategy of banana wilt control in the future.

Secreted effectors from Magnaporthe oryzae play critical roles in the interaction with rice to facilitate fungal infection and disease development. M. oryzae-secreted protein MoHrip1 can improve plant defense as an elicitor in vitro, however, its biological function in fungal infection is not clear. In this study, we found that the expression of mohrip1 was significantly induced in the stages of fungal penetration and colonization. Although dispensable for the growth and conidiation, MoHrip1 was necessary for the full virulence of M. oryzae. Deletion of mohrip1 remarkably compromised fungal virulence on rice seedlings and even on rice leaves with wounds. Rice sheath inoculation assay further demonstrated the defects of mohrip1-deleted mutants on penetration and proliferation in rice cells. Additionally, compared with WT and complementation strain, the inoculation of mohrip1-deleted mutants induced a higher expression of specific defense related genes and a higher production of specific defensive compounds in rice leaves. These data collectively indicated that MoHrip1 is necessary for fungal penetration and invasive expansion, and further full virulence of rice blast fungus.

PevD1, a fungal effector secreted by Verticillium dahliae, could induce hypersensitive responses-like necrosis and systemic acquired resistance (SAR) in cotton and tobacco plants. PevD1 could drastically induce the expression of Nbnrp1, which is an asparagine-rich protein (NRP) of Nicotiana benthamiana. Our previous research indicated that Nbnrp1 positively regulated PevD1-induced cell necrosis and disease resistance. In this study, we further investigated PevD1-induced immune responses in both wild-type (WT) and Nbnrp1-RNAi lines through RNA-seq, in order to reveal the underlying mechanism of Nbnrp1-modulated PevD1-induced disease resistance in N. benthamiana. Results showed that Nbnrp1-RNAi lines exhibited reduced PevD1-induced immune responses, like inhibiting H2O2 accumulation and MAPK phosphorylation. To silence Nbnrp1 inhibited the expression of PevD1-induced differential expression genes (DEGs) involved in pathways associated with sesquiterpenoid and triterpenoid biosynthesis, flavone and flavonol biosynthesis, plant-pathogen interaction and phenylpropanoid biosynthesis, etc. It is worth noting that sesquiterpene phytoalexin capsidiol accumulation were obviously decreased in Nbnrp1-RNAi plants after PevD1 treatment, accompanied with the down-expression of EAS and EAH, which were two key genes related to capsidiol biosynthesis. These results suggested that Nbnrp1 mediates PevD1-induced defense responses by regulating sesquiterpenoid phytoalexins biosynthesis pathway.

Phytophthora species cause enormous economic loss every year worldwide. Xenocoumacin 1 (Xcn1), isolated from the bacterium Xenorhabdus nematophilus, is a broad-spectrum antibiotic against agricultural pathogens, especially Phytophthora. To understand the inhibitory mode of Xcn1 toward Phytophthora pathogens, we determined the inhibitory effects of Xcn1 on Phytophthora capsici both in vitro and in vivo. In vitro, Xcn1 inhibited different stages in the life cycle of P. capsici, including sporangium formation, zoospore germination, and mycelial growth, with 50% effective concentration (EC50) values of 0.037, 0.81, and 2.44 μg ml−1, respectively. Xcn1 also reduced zoospore motility. In vivo, Xcn1 efficiently controlled the Phytophthora blight of pepper with a disease reduction of 99% at a concentration of 5 μg ml−1 assessed on the third day after incubation of wound stem plants. In addition, Xcn1-treated P. capsici mycelia exhibited increased mycelial branch spacing, evident plasmolysis, and leakage of intracellular components. In conclusion, in the presence of Xcn1, several stages in the life cycle of P. capsici were inhibited, and the hyphae exhibited obvious morphological changes.

Here we reported a novel protein elicitor from Bacillus amyloliquefaciens NC6 induced systemic resistance (ISR) in tobacco. The purification was executed by ion-exchange chromatography, native-page extraction and HPLC, and the amino acid sequence was identified by mass spectrometry. This recombinant elicitor protein, expressed in Escherichia coli by an E1 expression vector, had good thermal stability, and the elicitor caused a clearly defined hypersensitive response (HR) necrosis in tobacco leaves. It could also trigger early defence events, including generation of reactive oxygen species (H2O2 and O2 (-)) and phenolic-compound accumulation. Quantitative real-time PCR (Q-RT-PCR) results indicated that several plant defence genes, including the salicylic acid (SA)-responsive PR1a, PR1b, PR5, and phenylalanine ammonia lyase (PAL), as well as the jasmonic acid (JA)-responsive PDF1.2 and CORONATINE INSENSITIVE 1 (COI1), were all up-regulated. Moreover, infiltration conferred systemic resistance against a broad spectrum of pathogens, including Tobacco mosaic virus (TMV) and the fungal pathogen Botrytis cinerea.

The protein elicitor PevD1, isolated from Verticillium dahlia, could enhance resistance to TMV in tobacco and Verticillium wilt in cotton. Here, the pevd1 gene was over-expressed in wild type (WT) Arabidopsis, and its biological functions were investigated. Our results showed that the transgenic lines were more resistant to Botrytis cinerea and Pseudomonas syringae pv. tomato DC3000 than the WT line was. In transgenic plants, both the germination time and bolting time required were significantly shorter and fresh weights and plant heights were significantly higher than those in the WT line. A transcriptomics study using digital gene expression profiling (DGE) was performed in transgenic and WT Arabidopsis. One hundred and thirty-six differentially expressed genes were identified. In transgenic Arabidopsis, three critical regulators of JA biosynthesis were up-regulated and JA levels were slightly increased. Three important repressors of the ABA-responsive pathway were up-regulated, indicating that ABA signal transduction may be suppressed. One CML and two WRKY TFs involved in Ca(2+)-responsive pathways were up-regulated, indicating that this pathway may have been triggered. In conclusion, we show that PevD1 is involved in regulating several plant endogenous signal transduction pathways and regulatory networks to enhance resistance and promote growth and development in Arabidopsis.

The Cucumber aphid (Myzus persicae), a destructive cucumber aphid usually managed by chemical pesticides, is responsible for enormous annual agricultural losses. A protein elicitor, PeBL1, was investigated in the present work for its ability to induce a defense response against M. persicae in cucumber. The rates of population growth (Intrinsic rate of increase) of M. persicae (second and third generations) decreased with PeBL1-treated cucumber seedlings as compared to positive (water) and negative 70.58 &mu, g mL&minus, 1 controls (50 mM Tris-HCl, pH 8.0). In an assay on host selection, M. persicae had a preference for colonizing control plants as compared to the PeBL1-treated cucumber seedlings. The nymphal development time of the aphid was extended with the PeBL1-treated cucumber seedlings. Likewise, fecundity was reduced, with less offspring produced in the PeBL1-treated cucumber seedlings as compared to the positive (water) and negative 70.58 &mu, 1 controls (50 mM Tris-HCl, pH 8.0). The cucumber leaves treated with PeBL1 had a hazardous surface environment for M. persicae, caused by trichomes and wax formation. Jasmonic acid (JA), salicylic acid (SA), and ethylene (ET) levels were significantly higher, exhibiting significant accumulation in the PeBL1-treated cucumber seedlings. The following results showed that PeBL1 considerably altered the height of the cucumber plant and the surface structure of the leaves to minimize M. persicae reproduction, and it prevented colonization. Defensive processes also included the activation of pathways (JA, SA, and ET). This study provides evidence of biocontrol for the use of PeBL1 in cucumber defense against M. persicae.

This research reported the in vitro pathogenicity of Verticillium lecanii strains, L2 and L5, and Beauveria bassiana strains, B76 and B252, against Megoura japonica using leaf-dip method. The virulence potential of these four entomopathogenic fungal strains of V. lecanii and of B. bassiana were compared between fungi conidia (concentrations 1 &times, 106, 1 &times, 107, and 1 &times, 108 conidia mL&minus, 1) and culture filtrate. Moreover, binary combination of four different fungal strains (L2 + B76, B76 + L5, L2 + B252, and L2 + B76 + B252 + L5) were evaluated against M. japonica under control condition. Aphid mortality was recorded after two, four, six, and eight days of post-treatment. In the conidial concentration bioassay, strain B76 showed maximal mortality (85.3%) against bean aphid, and strain L5 showed the lowest effect (60.0%) at the highest concentration (1 &times, 1) at eight days post-treatment. Whereas, binary combinations of B76 and L2 strains showed the highest effect against M. japonica (90.5%) than other combinations. Moreover, in comparison with the effect of filtrate and conidia bioassay, 91.4% and 84.1% were achieved in strain B76, and the lowest effect (63.8% and 55.1%) was recorded in strain L5.

This study reports the characterization of protein elicitor PeBb1 derived from entomopathogenic fungus Beauveria bassiana ARSEF-2860 strain and its putative role in induced systemic resistance in Brassica rapa ssp. pekinensis against green peach aphid Myzus persicae. The sequence of purified elicitor protein was matched with the genomic sequence of a hypothetical protein BBA_10269 from B. bassiana ARSEF-2860 (GenBank Accession No. XP_008603588.1). The protein-encoding gene PeBb1 contained 534 bp cDNA encoding a polypeptide of 177 amino acids with a molecular mass of 19 kDa. The recombinant elicitor protein was expressed in Escherichia coli using pET-28a (+) expression vector and induced necrosis in the leaves of tobacco. The effects of elicitor protein on aphid M. persicae was determined by applying three different concentrations of PeBb1 (i.e., 26, 35, 53 &mu, M) on B. rapa plants at 4-leaf stage and the treated plants were exposed to newly emerged (0&ndash, 6 h old) apterous adult aphids. Bioassay results showed significant (p &lt, 0.05) sub-lethal effects of the exogenous application of PeBb1 elicitor on M. persicae. Moreover, the RT-qPCR gene expression analyses showed a significant up-regulation of most of the key genes linked to ethylene (ET)- and jasmonic acid (JA)-associated plant defense pathways in elicitor-treated plants. These results not only recommend the putative utilization of PeBb1 elicitor protein in future biological pest control strategies against phloem-feeding insect pests such as M. persicae, but also help in better comprehension of the mechanisms through which beneficial fungi trigger the induced plant resistance.

Many biotrophic and necrotrophic fungi synthesize proteins that may elicit induced plant resistance against different herbivore pests. This in-vitro study elucidates the sub-lethal effect of a partially-purified protein derived from the entomopathogenic fungus Lecanicillium lecanii (Zimmerman) (Hypocreales: Clavicipitaceae) against green peach aphid Myzus persicae (Sulzer) (Hemiptera: Aphididae), an economically important pest of many solanaceous crops including tomato. Bioassays were conducted to determine the impact of different concentrations of protein (i.e. 0.018, 0.036 and 0.054 µM) on the survival and fecundity of M. persicae on tomato (Lycopersicon esculentum) plants. Moreover, the potential role of this exogenous protein in the plant defense mechanism was assessed by expression analyses of key genes associated with salicylic acid (SA) and jasmonic acid (JA) pathways using RT-qPCR. The results indicated a significant negative effect of all protein concentrations on the survivorship and fecundity of M. persicae. The highest concentration (0.054 µM) resulted in lowest survival (46%) of aphids at 7th day post-treatment, while two other concentrations (0.036 and 0.018 µM) resulted in 61 and 71% survival rate, respectively. Similarly, lowest and highest mean fecundity rates were recorded for the highest protein concentration and the control (1.5 and 2.4 nymphs day−1 female−1), respectively. Moreover, L. lecanii-derived protein strongly upregulated the SA associated genes PR1, BGL2 and PAL, and moderately upregulated the JA associated genes LOX, AOS and AOC in protein-treated tomato plants compared to the control plants. These findings demonstrate the systemic resistance induced in tomato plants against M. persicae by the exogenous application of partially-purified protein extracted from L. lecanii, suggesting its further purification and characterization as a novel biological pest management tool against aphids and other phloem-feeding insect pests.

Elicitors are biofactors that induce resistance in plants against different insect pests. This in vitro study evaluated the impact of a novel elicitor protein PeBC1, extracted from a necrotrophic fungus Botrytis cinerea, on the development and fecundity parameters of green peach aphid (Myzus persicae) on common beans (Phaseolus vulgaris L.). Three different concentrations of PeBC1 elicitor (i.e., 33.56, 25.43, 19.33 µg mL−1) were applied at three different temperature regimes (i.e., 18, 21, and 25 °C). Elicitor treatments were applied topically on the bean plants at 3-leaf stage and newly emerged (0–6 h old) apterous adult aphids were exposed to these treated leaves. In addition to the biological parameters of aphids, the relative expression levels of key genes associated with jasmonic acid (JA) and salicylic acid (SA) plant defense pathways were also determined through RT-qPCR. Results of bioassays revealed that the application of PeBC1 elicitor protein exhibited pronounced and significant (p < 0.05) sub-lethal effects on green peach aphids. The fecundity was reduced and the nymphal development time was prolonged by different concentrations of PeBC1 elicitor and temperature regimes. Gene expression studies showed that the exogenous application of PeBC1 induced a significant upregulation of the expression levels of JA and SA pathway-associated genes in bean plants. As compared to control, elicitor-treated plants exhibited an induced resistance against aphids. Our findings suggest the potential use of PeBC1 elicitor protein in future bio-intensive management strategies against sap-sucking insect pests such as green peach aphids.

Green peach aphid, Myzus persicae (Hemiptera: Aphididae), is an economically important pest of crops within more than 40 plant families all over the world. This study encompasses in-vitro pathogenicity of two strains of Beauveria bassiana (BB-72 and BB-252) and one strain of Lecanicillium lecanii (V-4) against green peach aphid (M. persicae). Using a leaf-dip method, three different bioassays were conducted comprised of filtrates and conidial concentrations of BB-72, BB-252 and V-4 fungal strains and their binary combinations. Infiltrate bioassays, 2 mL fungal filtrate of each strain was used. In conidial bioassays, three different concentrations (i.e., 1 &times, 106, 1 &times, 107 and 1 &times, 108 conidia mL&minus, 1) of each fungal strain were used, while in binary combination bioassays, LC50 and LC33 of these fungal strains were evaluated. According to the results, maximum pathogenicity against test aphids was observed at 10th day of all treatments. Both strains of B. bassiana (BB-72 and BB-252) exhibited higher mortality of aphids (up to 95 and 91%, respectively) than L. lecanii (V-4) strain (87%) in all three types of bioassays. Moreover, binary combinations of BB-72 and BB-252 strains showed significantly higher aphid mortality (94%) than other combinations. Reduced mortality in case of fungal combinations of V-4 strain might be due to the antagonistic effect of L. lecanii strain along with both strains of B. bassiana. Nevertheless, evaluation of combined pathogenicity of the fungal strains constitutes the novelty of this in-vitro study which revealed that both strains of B. bassiana (i.e., BB-72 and BB-252) are mutually compatible and can be utilized together as new biocontrol tools against destructive insect pests such as M. persicae.

The protein elicitor PeaT1 was found in Alternaria tenuissima and exerted broad spectrum resistance in wheat, cotton, and rice. Recently, we found that overexpressing PeaT1 rice (OE) could enhance plant drought tolerance. Elucidating some elevated drought stress-related proteins and associated mechanisms is inevitable for improving drought tolerance in rice. In this study, combining a label-free quantitative proteomic method, multiple proteins were differentially accumulated in OE plants. Among these, a total of 57 significant changed proteins (including 32 up-regulated and 25 down-regulated) were mainly involved in metabolic, cellular, biological progress, and stress response. Using the RT-qPCR assay, 18 proteins’ relative abundance was detected mostly consistent with the proteins abundance in proteomic data. Specially, proteins involved in abiotic stress, such as OsSKIPa and OsPP2C, which were significantly induced in early after dehydration treatment in transgenic rice, and the other stress response genes (prohibitin protein, PsbP protein, msrB Protein) also changed in PeaT1 OE lines. Taken together, these results suggested that these differential proteins would be helpful for understanding the functional molecular mechanism of PeaT1 in rice.

Botrytis cinerea is one of the most notorious pathogenic species that causes serious plant diseases and substantial losses in agriculture throughout the world. We identified BcXyl1 from B. cinerea that exhibited xylanase activity. Expression of the BcXyl1 gene was strongly induced in B. cinerea infecting Nicotiana benthamiana and tomato plants, and BcXyl1 deletion strains severely compromised the virulence of B. cinerea. BcXyl1 induced strong cell death in several plants, and cell death activity of BcXyl1 was independent of its xylanase activity. Purified BcXyl1 triggered typically PAMP-triggered immunity (PTI) responses and conferred resistance to B. cinerea and TMV in tobacco and tomato plants. A 26-amino acid peptide of BcXyl1 was sufficient for elicitor function. Furthermore, the BcXyl1 death-inducing signal was mediated by the plant LRR receptor-like kinases (RLKs) BAK1 and SOBIR1. Our data suggested that BcXyl1 contributed to B. cinerea virulence and induced plant defense responses.

Verticillium dahliae is a wide-host-range fungal pathogen that causes soil-borne disease in hundreds of dicotyledonous hosts. In search of V. dahliae Vd991 cell death-inducing proteins, we identified a pectate lyase (VdPEL1) that exhibited pectin hydrolytic activity, which could induce strong cell death in several plants. Purified VdPEL1 triggered defense responses and conferred resistance to Botrytis cinerea and V. dahliae in tobacco and cotton plants. Our results demonstrated that the mutant VdPEL1rec lacking the enzymatic activity lacked functions to induce both cell death and plant resistance, implying that the enzymatic activity was necessary. In addition, VdPEL1 was strongly induced in V. dahliae infected Nicotiana benthamiana and cotton roots, and VdPEL1 deletion strains severely compromised the virulence of V. dahliae. Our data suggested that VdPEL1 contributed to V. dahliae virulence and induced plant defense responses. These findings provide a new insight for the function of pectate lyase in the host–pathogen interaction.

Botrytis cinerea is one of the most notorious pathogenic species that causes serious plant diseases and substantial losses in agriculture throughout the world. We identified BcXyl1 from B. cinerea that exhibited xylanase activity. Expression of the BcXyl1 gene was strongly induced in B. cinerea infecting Nicotiana benthamiana and tomato plants, and BcXyl1 deletion strains severely compromised the virulence of B. cinerea. BcXyl1 induced strong cell death in several plants, and cell death activity of BcXyl1 was independent of its xylanase activity. Purified BcXyl1 triggered typically PAMP-triggered immunity (PTI) responses and conferred resistance to B. cinerea and TMV in tobacco and tomato plants. A 26-amino acid peptide of BcXyl1 was sufficient for elicitor function. Furthermore, the BcXyl1 death-inducing signal was mediated by the plant LRR receptor-like kinases (RLKs) BAK1 and SOBIR1. Our data suggested that BcXyl1 contributed to B. cinerea virulence and induced plant defense responses.

Background BcGs1, a cell wall-degrading enzyme (CWDE), was originally derived from Botrytis cinerea. Our previous study revealed that BcGs1 could trigger defense responses and protect plants against various pathogens. We researched the defense response mechanism underlying this BcGs1 elicitation in tomato. Results We revealed that the two domains were required for BcGs1’s full necrosis activity. According to analysis and quantitative real-time PCR of the up-regulated proteins and genes filtered by iTRAQ-based quantitative proteome approach, oxidative metabolism and phenylpropanoid metabolism were speculated to be involved in BcGs1-triggered defense response in tomato. Furthermore, experimental evidence showed that BcGs1 triggered reactive oxygen species (ROS) burst and increased the level of phenylalanine-ammonia lyase (PAL) and peroxidase (POD) enzyme activity, as well as lignin accumulation. Moreover, histochemical analysis revealed that infiltration of BcGs1 in tomato leaves exhibited cell wall thickening compared with untreated plants. Conclusions The results suggested that BcGs1 activated the basal defense response included lignin metabolism contributed to BcGs1-induced resistance to Botrytis. cinerea infection in tomato. Electronic supplementary material The online version of this article (10.1186/s12870-018-1319-0) contains supplementary material, which is available to authorized users.

PevD1 is a fungal protein secreted by Verticillium dahliae. Our previous researches showed that this protein could induce hypersensitive responses-like necrosis and systemic acquired resistance (SAR) in cotton and tobacco. To understand immune activation mechanisms whereby PevD1 elicits defense response, the yeast two-hybrid (Y2H) assay was performed to explore interacting protein of PevD1 in Arabidopsis thaliana, and a partner AtNRP (At5g42050) was identified. Here, AtNRP homolog in Nicotiana benthamiana was identified and designated as Nbnrp1. The Nbnrp1 could interact with PevD1 via Y2H and bimolecular fluorescence complementation (BiFC) analyses. Moreover, truncated protein binding assays demonstrated that the C-terminal 132 amino acid (development and cell death, DCD domain) of Nbnrp1 is required for PevD1-Nbnrp1 interaction. To further investigate the roles of Nbnrp1 in PevD1-induced defense response, Nbnrp1-overexpressing and Nbnrp1-silence transgenic plants were generated. The overexpression of Nbnrp1 conferred enhancement of PevD1-induced necrosis activity and disease resistance against tobacco mosaic virus (TMV), bacterial pathogen Pseudomonas syringae pv. tabaci and fungal pathogen V. dahliae. By contrast, Nbnrp1-silence lines displayed attenuated defense response compared with the wild-type. It is the first report that an asparagine-rich protein Nbnrp1 positively regulated V. dahliae secretory protein PevD1-induced cell death response and disease resistance in N. benthamiana.

We describe here a double-stranded RNA mycovirus, termed Fusarium graminearum alternavirus 1 (FgAV1/AH11), from the isolate AH11 of the phytopathogenic fungus F. graminearum . Phylogenetic analysis showed that FgAV1/AH11 belongs to a newly proposed family, Alternaviridae . This is the first report of a mycovirus in the family Alternaviridae that infects F. graminearum .

Figure S1. Purification and necrosis activity of the protein BcGs1. A: SDS-PAGE analysis of purified BcGs1. M, Protein marker. 1, Purified BcGs1. B: Necrosis activity of BcGs1 in tomato, tobacco, cucumber and pea leaves. (DOCX 673Â kb)

We report the identification, characterization, and gene cloning of a novel protein elicitor (PeBL1) secreted from Brevibacillus laterosporus strain A60. Through a purification process consisting of ion-exchange chromatography and high-performance liquid chromatography (HPLC), we isolated a protein that was identified by electrospray ionization quadrupole time of flight tandem mass spectrometry (ESI–Q-TOF–MS-MS). The 351-bp PeBL1 gene produces a 12,833-Da protein with 116 amino acids that contains a 30-residue signal peptide. The PeBL1 protein was expressed in Escherichia coli . The recombinant protein can induce a typical hypersensitive response (HR) and systemic resistance in Nicotiana benthamiana , like the endogenous protein. PeBL1-treated N. benthamiana exhibited strong resistance to the infection of tobacco mosaic virus-green fluorescent protein (TMV-GFP) and Pseudomonas syringae pv. tabaci compared to control N. benthamiana . In addition, PeBL1 triggered a cascade of events that resulted in defense responses in plants, including reactive oxygen species (ROS) production, extracellular-medium alkalization, phenolic-compound deposition, and expression of several defense-related genes. Real-time quantitative-PCR analysis indicated that the known defense-related genes PR-1 , PR-5 , PDF1.2 , NPR1 , and PAL were upregulated to varying degrees by PeBL1. This research not only provides insights into the mechanism by which beneficial bacteria activate plant systemic resistance, but also sheds new light on a novel strategy for biocontrol using strain A60.

Plant bHLH transcription factors have important roles in regulation of developmental processes and responses to environmental stresses, but little is known about their functions in wheat in response to infection by pathogens. A new bHLH-type gene TabHLH060 was cloned from bread wheat ( Triticum aestivum ) and characterized. TabHLH060 was strongly expressed in wheat leaves and involved in response to infection by the obligate pathogen Puccinia striiformis f. sp. tritici . Treatments with defense-related hormones salicylic acid, methyl jasmonate and particularly ethylene suppressed the transcription level of TabHLH060 to different extents. Susceptibility of TabHLH060 -overexpressing Arabidopsis plants to Pseudomonas syringae DC3000 was significantly enhanced. TabHLH060 overexpression resulted in suppression of transcription levels of resistance-related genes PR1 , PR2 and PR5 involved in SA signaling pathway, but enhanced genes PDF1.2 and ORA59 involved in the jasmonic acid and ethylene signaling pathways. TabHLH060 is a novel bHLH member of the Ⅳc subgroup, and negatively regulates plant disease resistance through jasmonic acid and ethylene signaling.

During pathogenic infection, hundreds of proteins that play vital roles in the Verticillium dahliae-host interaction are secreted. In this study, an integrated proteomic analysis of secreted V. dahliae proteins was performed, and a conserved secretory protein, designated VdCP1, was identified as a member of the SnodProt1 phytotoxin family. An expression analysis of the vdcp1 gene revealed that the transcript is present in every condition studied and displays elevated expression throughout the infection process. To investigate the natural role of VdCP1 in V. dahliae, two vdcp1 knockout mutants and their complementation strains were generated. Bioassays of these mutants revealed no obvious phenotypic differences from the wild-type (WT) in terms of mycelial growth, conidial production or mycelial/spore morphology. However, compared with the WT, the vdcp1 knockout mutants displayed attenuated pathogenicity in cotton plants. Furthermore, treating plants with purified recombinant VdCP1 protein expressed in Pichia pastoris induced the accumulation of reactive oxygen species (ROS), expression of several defense-related genes, leakage of ion electrolytes, enhancement of defense-related enzyme activity and production of salicylic acid. Moreover, VdCP1 conferred resistance to Botrytis cinerea and Pseudomonas syringae pv. tabaci in tobacco and to V. dahliae in cotton. Further research revealed that VdCP1 possesses chitin-binding properties and that the growth of vdcp1 knockout mutants was more affected by treatments with chitinase, indicating that VdCP1 could protect V. dahliae cell wall from enzymatic degradation, which suggests an effector role of VdCP1 in infecting hosts.

MoHrip1, a protein isolated from Magnaporthe oryzae, belongs to the Alt A 1 (AA1) family. mohrip1 mRNA levels showed inducible expression throughout the infection process in rice. To determine the location of MoHrip1 in M. oryzae, a mohrip1-gfp mutant was generated. Fluorescence microscopy observations and western blotting analysis showed that MoHrip1 was both present in the secretome and abundant in the fungal cell wall. To obtain MoHrip1 protein, we carried out high-yield expression of MoHrip1 in Pichia pastoris. Treatment of tobacco plants with MoHrip1 induced the formation of necrosis, accumulation of reactive oxygen species and expression of several defense-related genes, as well as conferred disease resistance. By fusion to green fluorescent protein, we showed that MoHrip1 was able to bind to the tobacco and rice plant plasma membrane, causing rapid morphological changes at the cellular level, such as cell shrinkage and chloroplast disorganization. These findings indicate that MoHrip1 is a microbe-associated molecular pattern that is perceived by the plant immune system. This is the first study on an AA1 family protein that can bind to the plant plasma membrane.

PeaT1 is a proteinaceous elicitor from fungal pathogen Alternaria tenuissima. Our previous research revealed that this elicitor could induce defense response and enhance disease resistance in various plants including Nicotiana plants. However, immune activation mechanisms whereby PeaT1 elicits defense response remain unclear. In this study, the association between elicitor protein PeaT1 and the plasma membrane was assessed using the FITC (Fluorescein isothiocyanate) labeling method. A PeaT1-interacting protein was isolated via 125I-PeaT1 cross-linking and Far Western blot analyses, and designated PtBP1 (PeaT1 Binding Protein 1). From the data of Mass spectrometry (MS) and bioinformatics analysis, the 22 kDa plasma membrane protein PtBP1 was inferred to be a member of DREPP (developmentally regulated plasma membrane polypeptide) family that is induced in plants under stress conditions and might get involved in downstream signaling. For further verification of this association, Far Western blot, co-immunoprecipitation and bimolecular fluorescence complementation (BiFC) analyses were performed, showing PtBP1 could bind with PeaT1 in vitro and in vivo. Virus-induced gene silencing (VIGS) analysis exhibited that PtBP1 silencing in Nicotiana benthamiana attenuated tobacco mosaic virus (TMV) resistance compared to the tobacco rattle virus (TRV) control after PeaT1 treatment.

Abiotic stresses, especially drought, seriously threaten cereal crops yields and quality. In this study, we observed that the rice plants of overexpression the Alternariatenuissima PeaT1 gene showed enhanced drought stress tolerance and increased the survival rate following a drought treatment. In PeaT1-overexpressing (PeaT1OE) plants, abscisic acid and chlorophyll content significantly increased, while the malondialdehyde (MDA) content decreased compared with the wild-type plants. Additionally, we confirmed that the transcript levels of drought-responsive genes, including OsAM1, OsLP2, and OsDST, were prominently lower in the PeaT1OE plants. In contrast, expression levels of genes encoding positive drought stress regulators including OsSKIPa, OsCPK9, OsNAC9, OSEREBP1, and OsTPKb were upregulated in PeaT1OE plants. Furthermore, combing the yeast two-hybrid assay, we found that PeaT1 could interact with amyo-inositol oxygenase (OsMIOX), which was verified by pull-down assay. Interestingly, OsMIOX was highly expressed in PeaT1OE plants during the drought treatment. Additionally, the OsMIOX-GFP fusion protein co-localized with the endoplasmic reticulum (ER) marker in tobacco protoplasts, suggesting OsMIOX performs its function in ER. Therefore, our results are useful for elucidating the molecular mechanism underlying the improvement of drought tolerance by PeaT1.

We describe a novel positive single-stranded RNA virus, termed Fusarium langsethiae hypovirus 1 (FlHV1), from the isolate AH32 of the phytopathogenic fungus Fusarium langsethiae . The properties of FlHV1 permit assignment to the genus Alphahypovirus in the family Hypoviridae . This is the first report of a mycovirus identified in F. langsethiae .

The soil-borne fungal pathogen Verticillium dahliae infects a wide range of dicotyledonous plants including cotton, tobacco, and Arabidopsis. Among the effector proteins secreted by V. dahliae, the 16 kDa PevD1 induces a hypersensitive response in tobacco. Here we report the high-resolution structure of PevD1 with folds resembling a C2 domain-like structure with a calcium ion bound to the C-terminal acidic pocket. A yeast two-hybrid screen, designed to probe for molecular functions of PevD1, identified Arabidopsis asparagine-rich protein (NRP) as the interacting partner of PevD1. Extending the pathway of V. dahliae effects, which include induction of early flowering in cotton and Arabidopsis, NRP was found to interact with cryptochrome 2 (CRY2), leading to increased cytoplasmic accumulation of CRY2 in a blue light-independent manner. Further physiological and genetic evidence suggests that PevD1 indirectly activates CRY2 by antagonizing NRP functions. The promotion of CRY2-mediated flowering by a fungal effector outlines a novel pathway by which an external stimulus is recognized and transferred in changing a developmental program.

Our studies indicate a potential important elicitor candidate which can aid in the fight against a worldwide disease, rice blast. In this study, we report the purification, identification, characterization, and gene cloning of a novel hypersensitive response-inducing protein elicitor (MoHrip2) secreted from an important pathogenic fungus, Magnaporthe oryzae. The protein fraction was isolated from the culture filtrate of M. oryzae and identified by de novo sequencing. The elicitor-encoding gene mohrip2 was cloned following sequence comparison and PCR amplification. This 459-bp gene encodes a 152-residue polypeptide that contains an 18-residue signal peptide and exhibits a pI of 4.72 and an apparent molecular mass of 16 kDa. The hypothetical protein, MoHrip2, was expressed in Escherichia coli, and both the recombinant and the endogenous protein caused necrotic lesions in tobacco leaves. In addition to phenolic compound deposition and alkalization of the extracellular medium, MoHrip2 also induced hydrogen peroxide production and nitric oxide accumulation in tobacco cells. Moreover, rice seedlings treated with MoHrip2 exhibited pronounced resistance to M. oryzae compared with control seedlings.

In our previous study, PevD1 was characterized as a novel protein elicitor produced by Verticillium dahliae inducing hypersensitive response (HR) and systemic acquired resistance (SAR) in tobacco plants; however, the detailed mechanisms of PevD1's elicitor activity remain unclear. In this study, five mutant fragments of PevD1 were generated by polymerase chain reaction-based mutagenesis and the truncated proteins expressed in Escherichia coli were used to test their elicitor activities. Biological activity analysis showed that the N-terminal and C-terminal of PevD1 had distinct influence on HR and SAR elicitation. Fragment PevD1ΔN98, which spans the C-terminal 57 amino acids of PevD1, was critical for the induction of HR in tobacco plants. In contrast, fragment PevD1ΔC57, the N-terminal of 98 amino acids of PevD1, retained the ability to induce SAR against tobacco mosaic virus (TMV) but not induction of HR, suggesting that the induction of HR is not essential for SAR mediated by PevD1. Our results indicated that fragment PevD1ΔC57 could be a candidate peptide for plant protection against pathogens without causing negative effects.

We previously identified a novel protein elicitor, PebC1, from Botrytis cinerea and described its enhancement of plant growth, drought tolerance and disease resistance in tomato. Here, we have investigated the defense-associated molecular responses in Arabidopsis thaliana after treatment with recombinant PebC1. PebC1 was expressed in Escherichia coli. Recombinant protein treatments improved plant resistance to Botrytis infection and maintained plant defenses for more than 21 days. The purified protein at 10 μg ml(-1) activated extracellular medium alkalization (pH) and reactive oxygen species and nitric oxide generation and also induced defense gene expression. Arabidopsis mutants that are insensitive to salicylic acid had increased resistance to Botrytis infection after PebC1 treatment but PebC1 did not affect the resistance of mutants with jasmonic acid and ethylene transduction pathways. The results suggest that PebC1 can function as an activator of plant disease resistance and can promote disease resistance to Botrytis in A. thaliana through the ethylene signal transduction pathway.

Panama disease, or Fusarium wilt, the most serious disease in banana cultivation, is caused by Fusarium oxysporum f. sp. cubense (FOC) and has led to great economic losses worldwide. One effective way to combat this disease is by enhancing host plant resistance. The cerato-platanin protein (CPP) family is a group of small secreted cysteine-rich proteins in filamentous fungi. CPPs as elicitors can trigger the immune system resulting in defense responses in plants. In this study, we characterized a novel cerato-platanin-like protein in the secretome of Fusarium oxysporum f. sp. cubense race 4 (FOC4), named FocCP1. In tobacco, the purified recombinant FocCP1 protein caused accumulation of reactive oxygen species (ROS), formation of necrotic reaction, deposition of callose, expression of defense-related genes, and accumulation of salicylic acid (SA) and jasmonic acid (JA) in tobacco. These results indicated that FocCP1 triggered a hypersensitive response (HR) and systemic acquired resistance (SAR) in tobacco. Furthermore, FocCP1 enhanced resistance tobacco mosaic virus (TMV) disease and Pseudomonas syringae pv. tabaci 6605 (Pst. 6605) infection in tobacco and improved banana seedling resistance to FOC4. All results provide the possibility of further research on immune mechanisms of plant and pathogen interactions, and lay a foundation for a new biological strategy of banana wilt control in the future.

Elicitors are biofactors that induce resistance in plants against different insect pests. This in vitro study evaluated the impact of a novel elicitor protein PeBC1, extracted from a necrotrophic fungus Botrytis cinerea, on the development and fecundity parameters of green peach aphid (Myzus persicae) on common beans (Phaseolus vulgaris L.). Three different concentrations of PeBC1 elicitor (i.e., 33.56, 25.43, 19.33 &micro, g mL&minus, 1) were applied at three different temperature regimes (i.e., 18, 21, and 25 &deg, C). Elicitor treatments were applied topically on the bean plants at 3-leaf stage and newly emerged (0&ndash, 6 h old) apterous adult aphids were exposed to these treated leaves. In addition to the biological parameters of aphids, the relative expression levels of key genes associated with jasmonic acid (JA) and salicylic acid (SA) plant defense pathways were also determined through RT-qPCR. Results of bioassays revealed that the application of PeBC1 elicitor protein exhibited pronounced and significant (p &lt, 0.05) sub-lethal effects on green peach aphids. The fecundity was reduced and the nymphal development time was prolonged by different concentrations of PeBC1 elicitor and temperature regimes. Gene expression studies showed that the exogenous application of PeBC1 induced a significant upregulation of the expression levels of JA and SA pathway-associated genes in bean plants. As compared to control, elicitor-treated plants exhibited an induced resistance against aphids. Our findings suggest the potential use of PeBC1 elicitor protein in future bio-intensive management strategies against sap-sucking insect pests such as green peach aphids.

A double-stranded (ds) RNA element, sized at approximately 13 kb pairs, was purified from a field isolate, HN10, of Fusarium graminearum. The coding strand of the dsRNA was 13,023 nucleotides (nt) long (excluding the 3' poly(A) tail) and was predicted to contain two discontiguous open reading frames (ORF A and ORF B). The 5' proximal ORF A of 531 nt encoded a protein of 176 amino acids (aa), and a BLAST search showed it to be similar to the putative papain-like protease domains encoded by Valsa ceratosperma hypovirus 1 (35% identity) and Cryphonectria hypovirus 4 (CHV4) (31% identity). The 3' proximal ORF B of 11,118nt encoded a large polyprotein with three conserved domains, including papain-like protease, RNA-dependent RNA polymerase and RNA helicase domains. The polyprotein shared significant aa identities with CHV1 (32%) and CHV2 (32%). Both the genome organization and phylogenetic analysis suggested that the characterized RNA represented a novel hypovirus, designated "Fusarium graminearum hypovirus 1 (FgHV1)", which was closely related to CHV1 and CHV2 in the Hypoviridae family. Elimination of the virus resulted in no dramatic phenotypic alteration of the fungus.