The virulent strain B6 of Curvularia lunata was screened out from 65 fungus strains isolated from the diseased leaves of barnyardgrass （Echinochloa crus-galli）. Greenhouse and field studies were conducted to evaluat...The virulent strain B6 of Curvularia lunata was screened out from 65 fungus strains isolated from the diseased leaves of barnyardgrass （Echinochloa crus-galli）. Greenhouse and field studies were conducted to evaluate the feasibility of the strain being exploited as a mycoherbicide for barnyardgrass control in paddy fields. The results of pathogenicity experiments showed that this strain was highly pathogenic to barnyardgrass at the 1- to 2.5-leaf stages. The fresh weight reduction increased with the increase of inoculated conidial concentrations and the prolongation of dew duration. Strain B6 provided excellent barnyardgrass control when it was applied at the concentration of 1×104-1×106 conidia mL-1 in paddy fields with water layer. This strain was very safe to rice and the most plant species except wheat, barley and corn. Findings of this study indicated that this strain could be a potential mycoherbicide for barnyardgrass control in paddy fields in the future.展开更多
Curvularia leaf spot, caused mainly by Curvularia lunata, is a widespread plant disease in China. In the recent years, di- rectional host selection by the pathogen, which likely results in the virulence differentiatio...Curvularia leaf spot, caused mainly by Curvularia lunata, is a widespread plant disease in China. In the recent years, di- rectional host selection by the pathogen, which likely results in the virulence differentiation in pathogens, is widely reported. Among the hallmarks potentially associated to pathogen variation in virulence, superoxide dismutase gene Sod has been found to be closely related to the enhancement of virulence. In the present study, the full-length of Sod was obtained via Blastn alignment against GenBank and the whole genome of C. lunata. In order to understand the role of Sod in the vir- ulence variation in C. lunata, targeted gene disruption was performed to construct Sod mutants. The cell wall degrading enzyme （CWDE） activities and toxin production of ASod were not distinctly different from wild-type strain CX-3 and its complon. However, at an early stage of infection, 3Sod virulence appeared to be lower than CX-3 and the complon, while at a later stage, its virulence gradually returned to the level of CX-3 and the complon. Furthermore, the melanin production of ASod was significantly reduced compared to CX-3 and the complon, suggesting that Sod gene influences the virulence by regulating melanin production at an early stage of infection but is not essential for pathogenicity. However, the disruption of Sod did not significantly affect the transcriptional expression of the melanin biosynthesis-associated genes, bml and scd. Therefore, we infer that Sod in C. lunata are involved, to some extent, with the virulence in maize leaf, but still needs further studies to have a clear understanding of its mechanism.展开更多
Mitogen-activated protein kinase （MAPK） cascades play an important role in extracellular signal transduction and are involved in the pathogenicity of fungal pathogens to host plants. In Curvularia lunata, the roles ...Mitogen-activated protein kinase （MAPK） cascades play an important role in extracellular signal transduction and are involved in the pathogenicity of fungal pathogens to host plants. In Curvularia lunata, the roles of two MAPK genes, Clkl and CIm 1, have already been studied. Clkl is involved in conidia formation and pathogenicity, and Clmf is closely related to pathogen cell wall formation and pathogenicity to maize leaves. In this study, a third C. lunata MAPK gene, Clhl, which is homologous to hog1, was successfully cloned. We found that a Clhl deletion mutant had lower intracellular glycerol accumulation than the wild-type stain and was unable to grow normally under osmotic stress conditions. Furthermore, the deletion mutants of three C. lunata MAPK genes （Clkl, Clml and Clhl） had lower levels of acetyI-CoA, which is an important intermediate product in the synthesis of melanin and furan toxin, and down-regulated expression of pathogenicity-associated genes. Furthermore, pathogenicity and the ability to produce toxin were restored after adding acetyI-CoA to the culture medium, suggesting that acetyI-CoA is closely involved in the pathogen MAPK signaling pathway.展开更多
The purpose of this study was to identify the dominant pathogens of Curvularia leaf spot and their pathogenicity variation in Huanghuaihai Region of China in recent years.In 2013 and 2016–2017,the occurrences of Curv...The purpose of this study was to identify the dominant pathogens of Curvularia leaf spot and their pathogenicity variation in Huanghuaihai Region of China in recent years.In 2013 and 2016–2017,the occurrences of Curvularia leaf spots on maize were investigated in fields located in Henan,Hebei,Shandong,and Anhui provinces,and 292 fungi were isolated from diseased leaves.These fungal isolates were subjected to morphological identification,and 232 isolates were found to have about 70%uncurved conidia and were identified as Curvularia lunata var.Most of the conidia of 2 representative isolates,namely,HNWB-131 and HNWB-185,were oblong with parallel septations and were distinctly different from a reference isolate CX-3.For further determination,the internal transcribed spacer(ITS),glyceraldehyde 3-phosphate dehydrogenase(GPDH),the large subunit(LSU),and translation elongation factor 1-alpha(EF1-α)sequences of HNWB-131,HNWB-185,and CX-3 were amplified and sequenced.The results of sequence analysis showed that the 4 gene sequences from the 3 isolates had a similarity of more than 99%to C.lunata.Based on the sequences of ITS and the combined data of the 4 genes,neighbor-joining trees were constructed for phylogenetic analysis.The results indicated that these 3 isolates were clustered together with C.lunata.The expression of Clg2 p and ClUrase genes in mycelia and conidia was significantly(P<0.05)higher in CX-3 than in HNWB-131 and HNWB-185.This study found that the dominant pathogen of Curvularia leaf spot was a new variety of C.lunata with morphological variations in Huanghuaihai Region from 2013 to 2017.The pathogenicity of the C.lunata var.was not significantly enhanced,and the expression of Clg2 p and ClUrase genes of C.lunata var.was decreased.展开更多
Twelve isolates of Curvularia lunata and six related species from maize or other host plants are examined for the analysis of polyacrylamide gel electrophoresis(SDS PAGE) patterns of their soluble proteins. Difference...Twelve isolates of Curvularia lunata and six related species from maize or other host plants are examined for the analysis of polyacrylamide gel electrophoresis(SDS PAGE) patterns of their soluble proteins. Differences exist significantly among different species and the same species. There is a special protein band for Curvularia at relative mobility value 0.177 (Rf 0.177) and a special protein band for C. lunata at Rf 0.225. Two kinds of primary antibodies to C. lunata are used to indirect ELISA and Avidin Biotin peroxidase Complex (ABC) immunoassay to evaluate their detecting sensitivity and specificity. The sensitivity of ABC immuno assay is higher than indirect ELISA. APAbs to C. lunata is special to C. lunata.展开更多
One-dimensional electrophoresis （1-DE） of proteins, two-dimensional electrophoresis （2-DE） of proteins and cloning of cDNA sequence were used to study the virulence differentiation of Curvularia lunata （Wakker） ...One-dimensional electrophoresis （1-DE） of proteins, two-dimensional electrophoresis （2-DE） of proteins and cloning of cDNA sequence were used to study the virulence differentiation of Curvularia lunata （Wakker） Boed. isolated from maize （Zea maydis L.） in China. From 1-DE gel profiles of proteins, 110 reproducible bands were separated from six isolates of C. lunata CX-3, SD-6, C-152, C107-1, DD-60 and W-18. Sixty-eight bands （61.82%） were polymorphic, suggesting huge biodiversities among the isolates. All isolates for the experiment were clustered into three groups consisting of different virulent types by coefficient value of 0.605. Group 1, consisting of CX-3, SD-6 and C- 152 with high virulence displayed more protein bands than Groups 2 and 3, consisting of C107-1 and DD-60 with low virulence. Proteomics approaches based on 2-DE techniques were applied to identify specific proteins associated with the virulence differentiation in CX-3 and DD-60. A total of 423 protein spots were separated. Out of them 75 specific protein spots were displayed in 2-DE gels. Among them 28 protein spots were unique in CX-3 and eight in DD-60, and 39 protein spots were shown on both 2-DE gels but expressed differently in intensity. Twenty protein spots including three unique protein spots and 17 differentially expressed protein spots （more than two-fold DD- 60） in CX-3 were further identified with MALDI-TOF MS/MS. Results indicated that most of the identified proteins were found to be associated with virulence differentiation, metabolisms, stress response and signal transduction. One of them was identified as Brnl protein, which had been reported to be related to melanin biosynthesis and the virulence differentiation in fungi. Combined with our previous findings, we assumed that Brnl protein and its regulating products might be involved in the virulence differentiation of C. lunata. Consequently, we cloned a Brnl cDNA fragment and aligned it with the fragments in other fungi. Results indicated that the 633-bp sequence of Brnl cloned in C. lunata was highly homological with the compared fungi. Further work for the exact gene roles of Brnl in our case is underway.展开更多
基金supported by the the High Technology R&D Program of China (2006AA10A214)
文摘The virulent strain B6 of Curvularia lunata was screened out from 65 fungus strains isolated from the diseased leaves of barnyardgrass （Echinochloa crus-galli）. Greenhouse and field studies were conducted to evaluate the feasibility of the strain being exploited as a mycoherbicide for barnyardgrass control in paddy fields. The results of pathogenicity experiments showed that this strain was highly pathogenic to barnyardgrass at the 1- to 2.5-leaf stages. The fresh weight reduction increased with the increase of inoculated conidial concentrations and the prolongation of dew duration. Strain B6 provided excellent barnyardgrass control when it was applied at the concentration of 1×104-1×106 conidia mL-1 in paddy fields with water layer. This strain was very safe to rice and the most plant species except wheat, barley and corn. Findings of this study indicated that this strain could be a potential mycoherbicide for barnyardgrass control in paddy fields in the future.
基金The National Natural Science Foundation of China(31171798 and 31471734)the China Agriculture Research System,China(CARS-02)
文摘Curvularia leaf spot, caused mainly by Curvularia lunata, is a widespread plant disease in China. In the recent years, di- rectional host selection by the pathogen, which likely results in the virulence differentiation in pathogens, is widely reported. Among the hallmarks potentially associated to pathogen variation in virulence, superoxide dismutase gene Sod has been found to be closely related to the enhancement of virulence. In the present study, the full-length of Sod was obtained via Blastn alignment against GenBank and the whole genome of C. lunata. In order to understand the role of Sod in the vir- ulence variation in C. lunata, targeted gene disruption was performed to construct Sod mutants. The cell wall degrading enzyme （CWDE） activities and toxin production of ASod were not distinctly different from wild-type strain CX-3 and its complon. However, at an early stage of infection, 3Sod virulence appeared to be lower than CX-3 and the complon, while at a later stage, its virulence gradually returned to the level of CX-3 and the complon. Furthermore, the melanin production of ASod was significantly reduced compared to CX-3 and the complon, suggesting that Sod gene influences the virulence by regulating melanin production at an early stage of infection but is not essential for pathogenicity. However, the disruption of Sod did not significantly affect the transcriptional expression of the melanin biosynthesis-associated genes, bml and scd. Therefore, we infer that Sod in C. lunata are involved, to some extent, with the virulence in maize leaf, but still needs further studies to have a clear understanding of its mechanism.
基金supported by the National Natural Science Foundation of China (31471734 and 31672072)the earmarked fund for China Agriculture Research System (CARS-02)
文摘Mitogen-activated protein kinase （MAPK） cascades play an important role in extracellular signal transduction and are involved in the pathogenicity of fungal pathogens to host plants. In Curvularia lunata, the roles of two MAPK genes, Clkl and CIm 1, have already been studied. Clkl is involved in conidia formation and pathogenicity, and Clmf is closely related to pathogen cell wall formation and pathogenicity to maize leaves. In this study, a third C. lunata MAPK gene, Clhl, which is homologous to hog1, was successfully cloned. We found that a Clhl deletion mutant had lower intracellular glycerol accumulation than the wild-type stain and was unable to grow normally under osmotic stress conditions. Furthermore, the deletion mutants of three C. lunata MAPK genes （Clkl, Clml and Clhl） had lower levels of acetyI-CoA, which is an important intermediate product in the synthesis of melanin and furan toxin, and down-regulated expression of pathogenicity-associated genes. Furthermore, pathogenicity and the ability to produce toxin were restored after adding acetyI-CoA to the culture medium, suggesting that acetyI-CoA is closely involved in the pathogen MAPK signaling pathway.
基金supported by the earmarked fund for the China Agriculture Research System (CARS-02)the National Key Research and Development Program of China (2017YFD0200400)
文摘The purpose of this study was to identify the dominant pathogens of Curvularia leaf spot and their pathogenicity variation in Huanghuaihai Region of China in recent years.In 2013 and 2016–2017,the occurrences of Curvularia leaf spots on maize were investigated in fields located in Henan,Hebei,Shandong,and Anhui provinces,and 292 fungi were isolated from diseased leaves.These fungal isolates were subjected to morphological identification,and 232 isolates were found to have about 70%uncurved conidia and were identified as Curvularia lunata var.Most of the conidia of 2 representative isolates,namely,HNWB-131 and HNWB-185,were oblong with parallel septations and were distinctly different from a reference isolate CX-3.For further determination,the internal transcribed spacer(ITS),glyceraldehyde 3-phosphate dehydrogenase(GPDH),the large subunit(LSU),and translation elongation factor 1-alpha(EF1-α)sequences of HNWB-131,HNWB-185,and CX-3 were amplified and sequenced.The results of sequence analysis showed that the 4 gene sequences from the 3 isolates had a similarity of more than 99%to C.lunata.Based on the sequences of ITS and the combined data of the 4 genes,neighbor-joining trees were constructed for phylogenetic analysis.The results indicated that these 3 isolates were clustered together with C.lunata.The expression of Clg2 p and ClUrase genes in mycelia and conidia was significantly(P<0.05)higher in CX-3 than in HNWB-131 and HNWB-185.This study found that the dominant pathogen of Curvularia leaf spot was a new variety of C.lunata with morphological variations in Huanghuaihai Region from 2013 to 2017.The pathogenicity of the C.lunata var.was not significantly enhanced,and the expression of Clg2 p and ClUrase genes of C.lunata var.was decreased.
文摘Twelve isolates of Curvularia lunata and six related species from maize or other host plants are examined for the analysis of polyacrylamide gel electrophoresis(SDS PAGE) patterns of their soluble proteins. Differences exist significantly among different species and the same species. There is a special protein band for Curvularia at relative mobility value 0.177 (Rf 0.177) and a special protein band for C. lunata at Rf 0.225. Two kinds of primary antibodies to C. lunata are used to indirect ELISA and Avidin Biotin peroxidase Complex (ABC) immunoassay to evaluate their detecting sensitivity and specificity. The sensitivity of ABC immuno assay is higher than indirect ELISA. APAbs to C. lunata is special to C. lunata.
基金Supported by the National Natural Science Foundation of China （30471057 and 30370917）, the State Key Basic Research and Development Plan of China （2006CB101901）, the National Science ＆ Technology Supporting Programme （2006BAD08A06）, the Postdoctoral Foundation of China （20060390638） and the Postdoctoral Foundation of Shanghai （06R214135）.
文摘One-dimensional electrophoresis （1-DE） of proteins, two-dimensional electrophoresis （2-DE） of proteins and cloning of cDNA sequence were used to study the virulence differentiation of Curvularia lunata （Wakker） Boed. isolated from maize （Zea maydis L.） in China. From 1-DE gel profiles of proteins, 110 reproducible bands were separated from six isolates of C. lunata CX-3, SD-6, C-152, C107-1, DD-60 and W-18. Sixty-eight bands （61.82%） were polymorphic, suggesting huge biodiversities among the isolates. All isolates for the experiment were clustered into three groups consisting of different virulent types by coefficient value of 0.605. Group 1, consisting of CX-3, SD-6 and C- 152 with high virulence displayed more protein bands than Groups 2 and 3, consisting of C107-1 and DD-60 with low virulence. Proteomics approaches based on 2-DE techniques were applied to identify specific proteins associated with the virulence differentiation in CX-3 and DD-60. A total of 423 protein spots were separated. Out of them 75 specific protein spots were displayed in 2-DE gels. Among them 28 protein spots were unique in CX-3 and eight in DD-60, and 39 protein spots were shown on both 2-DE gels but expressed differently in intensity. Twenty protein spots including three unique protein spots and 17 differentially expressed protein spots （more than two-fold DD- 60） in CX-3 were further identified with MALDI-TOF MS/MS. Results indicated that most of the identified proteins were found to be associated with virulence differentiation, metabolisms, stress response and signal transduction. One of them was identified as Brnl protein, which had been reported to be related to melanin biosynthesis and the virulence differentiation in fungi. Combined with our previous findings, we assumed that Brnl protein and its regulating products might be involved in the virulence differentiation of C. lunata. Consequently, we cloned a Brnl cDNA fragment and aligned it with the fragments in other fungi. Results indicated that the 633-bp sequence of Brnl cloned in C. lunata was highly homological with the compared fungi. Further work for the exact gene roles of Brnl in our case is underway.
