In our previous screening of the transcriptome of the causal agent of the devastating pine wilt disease,pine wood nematode(PWN,Bursaphelenchus xylophilus),after treatment with the nematicide fomepizole,Surfeit locus g...In our previous screening of the transcriptome of the causal agent of the devastating pine wilt disease,pine wood nematode(PWN,Bursaphelenchus xylophilus),after treatment with the nematicide fomepizole,Surfeit locus gene sft-4,which encodes a regulatory factor,was found to be downregulated.In situ hybridization results showed that the sft-4 was continuously expressed from egg to adult and was especially high in the reproductive system.Here in a study of the effect of RNA interference(RNAi)of sft-4 and recombinant SFT-4 on PWN activity,treatment with sft-4 dsRNA inhibited feeding,reproduction,oviposition and egg hatching of PWN with the greatest inhibition on reproduction and oviposition,whereas recombinant SFT-4 had the opposite effect.In addition,RNAi of sft-4 changed the female–male ratio and lifespan of PWN.In bioassays of PWNs,with RNAi of sft-4 on seedlings and 2-year-old Pinus thunbergii trees,none of the treated plants developed symp-toms during the monitoring period,indicating that virulence of PWNs was either significantly weakened.These results indicate that the influence of sft-4 on PWN pathogenicity may be mainly through regulating reproductive function of PWN and its lifespan.展开更多
Bursaphelenchus xylophilus is the pathogen that causes pine wilt disease, which has greatly damaged forests and ecosystems in countries of East Asia and Europe.Bursaphelenchus mucronatus is closely related to B. xylop...Bursaphelenchus xylophilus is the pathogen that causes pine wilt disease, which has greatly damaged forests and ecosystems in countries of East Asia and Europe.Bursaphelenchus mucronatus is closely related to B. xylophilus in morphology and host plant specificity. A longrunning debate has existed regarding whether these two species can successfully produce hybrid offspring. In the present study, we performed in the laboratory, hybridization of two B. xylophilus nematode isolates from China and Japan and three B. mucronatus isolates from China, Japan and France. Nematode isolates of B. xylophilus were successfully crossed with B. mucronatus isolates, and the rate of hybridization was relatively high; however, some hybrid offspring died. Successful hybridization occurred between B. xylophilus and B. mucronatus isolates from China, and 22 generations of hybrids were produced. All F1 hybrids could be backcrossed with their parents and produce offspring. Variation in mucro length among the hybrid offspring and their parents was observed. The hybrid offspring and their parents were inoculated into 3-months-old black pine(Pinus tunbergii) seedlings. Weaker pathogenicity of hybrid offspring was observed compared with that of their parents, and significantly fewer offspring nematodes than parents were reisolated from pines. Therefore, the offspring of B. xylophilus and B. mucronatus may exist in the forest and could influence disease epidemics.展开更多
The method for pest risk analysis was used to make the potential risk analysis and assessment to the 8 Bursaphelenchus species found in the pines in Yuyao, Ningbao, and the corresponding risk management strategies wer...The method for pest risk analysis was used to make the potential risk analysis and assessment to the 8 Bursaphelenchus species found in the pines in Yuyao, Ningbao, and the corresponding risk management strategies were put forward.展开更多
Bursaphelenchus rainulfi isolated from dead pine trees in Zbejiang, China, is described and illustrated. It also provided some molecular characters of the Chinese population, including the PCR-RFLP and sequences of IT...Bursaphelenchus rainulfi isolated from dead pine trees in Zbejiang, China, is described and illustrated. It also provided some molecular characters of the Chinese population, including the PCR-RFLP and sequences of ITS region and D2-D3 expansion region of the large subunit （LSU） rRNA gene. Both the morphological characters and ITS-RFLP patterns match with the original description. The phylogenetic trees based on the 13 sequences of D2-D3 expansion region of the LSU rRNA gene and ITS region of Bursaphelenchus species were constructed, respectively, with the results showing the similar clades. The phylogenetic relationship based on the molecular data is similar to that with morphological characters. This is the first report of the species on pine wood in eastern China.展开更多
Pine wilt disease(PWD)is a devastating disease affecting the growth of Pinus massoniana,often leading to withering and death.To reveal the changes involved during disease progression,we investigated the mRNA expressio...Pine wilt disease(PWD)is a devastating disease affecting the growth of Pinus massoniana,often leading to withering and death.To reveal the changes involved during disease progression,we investigated the mRNA expression profile of P.massoniana infested by Bursaphelenchus xylophilus.The infestation resulted in the downregulation of genes involved in interactions with pathogenic pathways such as disease resistance gene,CC-NBS-LRR resistancelike protein,and the gene encoding a putative nematode resistance protein.Increased infestation pressure(number of nematodes inoculated)caused a continuous decline in the gene expression of stem samples.An infestation of P.massoniana also resulted in a pathway enrichment of genes involved in phenylpropanoid metabolism and flavonoid biosynthesis,which in turn reduced the levels of total phenols and total flavonoids.A downregulation of auxin responsive family protein was observed in infested samples,which resulted in a suppression of plant growth.Thus,upon B.xylophilus infestation,a downregulation of genes associated with the recognition of pathogens,PWD resistance,and growth regulation was observed in P.massoniana,together with a decrease in the levels of phytoalexinlike secondary substances,all of which resulted in withering and ultimately death of P.massoniana.展开更多
Genes homologous to members of the MRP gene family in Caenorhabditis elegans are important in drug resistance.To further explore the molecular mechanism of drug resistance in pine wood nematode(Bursaphelenchus xylophi...Genes homologous to members of the MRP gene family in Caenorhabditis elegans are important in drug resistance.To further explore the molecular mechanism of drug resistance in pine wood nematode(Bursaphelenchus xylophilus),we used bioinformatics approaches to analyze genomic data for B.xylophilus and identified Bx-MRP genes.We predicted the structure and function of the genes and encoded proteins.Using bioinformatics programs to predict and analyze various properties of the predicted proteins,including hydrophobicity,transmembrane regions,phosphorylation sites,and topologically isomeric structures,of these Bx-MRP genes,we determined that they function in transmembrane transport.From the results of RT-qPCR,the Bx-MRP family members confer significant differential resistance to different drug treatments.After treatment with different concentrations of emamectin benzoate,avermectin and matrine,the expression of each gene increased with increasing drug concentrations,indicating that the family members play a positive role in the regulation of multidrug resistance.展开更多
Bursaphelenchus xylophilus,causal agent of pine wilt disease,causes extensive damage worldwide.Strate-gies are needed to inhibit population growth or block the spread of the invasive nematode to control pine wilt dis-...Bursaphelenchus xylophilus,causal agent of pine wilt disease,causes extensive damage worldwide.Strate-gies are needed to inhibit population growth or block the spread of the invasive nematode to control pine wilt dis-ease.The gene daf-8 plays crucial roles in larval develop-ment and dauer formation in Caenorhabditis elegans,but little is known about its orthologue in B.xylophilus.In the present molecular characterization and functional analysis of daf-8 in B.xylophilus(Bx-daf-8),RT-qPCR showed that the expression of Bx-daf-8 gradually increased during the embryonic stage,peaked in the second-stage juvenile(J2),then dramatically dropped in the J3,and remained at that low level for the rest of its life.Bx-daf-8-transgenic C.elegans was employed to mimic the spatiotemporal expression of Bx-daf-8,which was expressed close to the pharynx during early embryogenesis and weakly throughout the whole body during late embryogenesis.It was observed in head neurons and tail ganglions throughout all postembryonic stages.B.xylophilus embryos were severely abnormal,and hatching rate decreased sharply after Bx-daf-8 knockdown.daf-16-1 and da-f16-2,downstream genes in the IIS pathway,also dropped sharply after Bx-daf-8 knockdown.We propose that TGFβmay crosstalk with the IIS pathway upstream of Bx-daf-16 and that daf-8 may act as a master regulator of daf-16 in B.xylophilus.However,knockdown of Bx-daf-8 did not lead to constitutive developmental arrest at the dauer larval stage,indicating that dauer entry in B.xylophilus might be controlled by several genes and is more complicated than in C.elegans.Bx-daf-8 alone did not control the dauer entry in B.xylophilus,but it was indispensable for embryogen-esis,providing a potential target for suppressing population growth of B.xylophilus.展开更多
This paper mainly summarizes recent advances in the classification of Bursaphelenchus species, introduces several common methods of molecular biologi- cal detection and identification methods, and analyzes the advanta...This paper mainly summarizes recent advances in the classification of Bursaphelenchus species, introduces several common methods of molecular biologi- cal detection and identification methods, and analyzes the advantages and disad- vantages of these methods.展开更多
Pine wilt disease caused by pine wood nema-todes is a deadly disease of the genus Pinus requiring strong quarantine measures.Since its discovery,it has been widely distributed throughout the world.China is one of the ...Pine wilt disease caused by pine wood nema-todes is a deadly disease of the genus Pinus requiring strong quarantine measures.Since its discovery,it has been widely distributed throughout the world.China is one of the coun-tries with a severe rate of infections due to its abundant pine resources.In this study,nematode-trapping fungi were collected from pine trees in Ninghai City,Zhejiang Prov-ince,which is the key area of pine wilt control in Febru-ary,May,September,October and November.The results showed that nematode-trapping fungi of pine are abundant,especially the number and species detected in each month and are quite different;species of fungi in July,Septem-ber and November were more numerous and had higher separation rates.The dominant species in November was Arthrobotrys oligospora followed by Arthrobotrys cladodes;July and September it was A.cladodes.In each part of the tree,A.cladodes was the most widely distributed with a high separation rate.Therefore,it is the dominant fungal species in the area.Arboreal Bursaphelenchus xylophilus-trapping fungi are dependent on pine wood nematodes,and none were isolated from healthy pine trees.More arboreal B.xylophilus-trapping fungi were found under the bark than in the xylem.There is a close relationship between arboreal B.xylophilus-trapping fungi and pine wood nematodes.How to make use of these resources to reduce the damage of pine wood nematode fusarium is the focus of future research.展开更多
Pine wood nematode （PWN）, Bursaphelenchus xylophilus （Steiner et Buhrer） Nickle is an important invasive alien species in forests of China and has become one of the most destructive forest diseases. In order to im...Pine wood nematode （PWN）, Bursaphelenchus xylophilus （Steiner et Buhrer） Nickle is an important invasive alien species in forests of China and has become one of the most destructive forest diseases. In order to improve the resistance and resilience of pine forest ecosystems against B. xylophilus invasion and make the pine forest ecosystem more timely responsive to PWN invasion, we made some recommendations based on five years of intensive observations. We advocate a set of management measures with the theme “Prevention is priority, but integrated with curative techniques and ecological resilience” on the pine forest ecosystem invaded by B. xylophilus; details of accurate measures are proposed. The aim is to discover the underlying problems of present pine forest ecosys-tems and to take, correspondingly, administrative measures and strategies, which will encourage the pine forest ecosystem to develop in a benign way.展开更多
基金supported by the Shandong Provincial Natural Science Foundation,China(ZR2020MC123)Qingdao Municipal People-benefitting Demonstration Project of Science and Technology,China(23-2-8-cspz-8-nsh).
文摘In our previous screening of the transcriptome of the causal agent of the devastating pine wilt disease,pine wood nematode(PWN,Bursaphelenchus xylophilus),after treatment with the nematicide fomepizole,Surfeit locus gene sft-4,which encodes a regulatory factor,was found to be downregulated.In situ hybridization results showed that the sft-4 was continuously expressed from egg to adult and was especially high in the reproductive system.Here in a study of the effect of RNA interference(RNAi)of sft-4 and recombinant SFT-4 on PWN activity,treatment with sft-4 dsRNA inhibited feeding,reproduction,oviposition and egg hatching of PWN with the greatest inhibition on reproduction and oviposition,whereas recombinant SFT-4 had the opposite effect.In addition,RNAi of sft-4 changed the female–male ratio and lifespan of PWN.In bioassays of PWNs,with RNAi of sft-4 on seedlings and 2-year-old Pinus thunbergii trees,none of the treated plants developed symp-toms during the monitoring period,indicating that virulence of PWNs was either significantly weakened.These results indicate that the influence of sft-4 on PWN pathogenicity may be mainly through regulating reproductive function of PWN and its lifespan.
基金supported by the special research program for non-profit forestry of the State Forestry Administration(201204501)National Nature Science Foundation of China(31170600)funded by the Priority Academic Program Development of Jiangsu Higher Education Institutions(PAPD)
文摘Bursaphelenchus xylophilus is the pathogen that causes pine wilt disease, which has greatly damaged forests and ecosystems in countries of East Asia and Europe.Bursaphelenchus mucronatus is closely related to B. xylophilus in morphology and host plant specificity. A longrunning debate has existed regarding whether these two species can successfully produce hybrid offspring. In the present study, we performed in the laboratory, hybridization of two B. xylophilus nematode isolates from China and Japan and three B. mucronatus isolates from China, Japan and France. Nematode isolates of B. xylophilus were successfully crossed with B. mucronatus isolates, and the rate of hybridization was relatively high; however, some hybrid offspring died. Successful hybridization occurred between B. xylophilus and B. mucronatus isolates from China, and 22 generations of hybrids were produced. All F1 hybrids could be backcrossed with their parents and produce offspring. Variation in mucro length among the hybrid offspring and their parents was observed. The hybrid offspring and their parents were inoculated into 3-months-old black pine(Pinus tunbergii) seedlings. Weaker pathogenicity of hybrid offspring was observed compared with that of their parents, and significantly fewer offspring nematodes than parents were reisolated from pines. Therefore, the offspring of B. xylophilus and B. mucronatus may exist in the forest and could influence disease epidemics.
文摘The method for pest risk analysis was used to make the potential risk analysis and assessment to the 8 Bursaphelenchus species found in the pines in Yuyao, Ningbao, and the corresponding risk management strategies were put forward.
基金Project (No.2002C34003) supported by the Science and Technology Department of Zhejiang Province,China
文摘Bursaphelenchus rainulfi isolated from dead pine trees in Zbejiang, China, is described and illustrated. It also provided some molecular characters of the Chinese population, including the PCR-RFLP and sequences of ITS region and D2-D3 expansion region of the large subunit （LSU） rRNA gene. Both the morphological characters and ITS-RFLP patterns match with the original description. The phylogenetic trees based on the 13 sequences of D2-D3 expansion region of the LSU rRNA gene and ITS region of Bursaphelenchus species were constructed, respectively, with the results showing the similar clades. The phylogenetic relationship based on the molecular data is similar to that with morphological characters. This is the first report of the species on pine wood in eastern China.
基金financially supported by the National Key Research and Development Program(2017YFD0600105)the National Natural Science Foundation of China(Grant No.31870641)+2 种基金the Research Foundation of Education Department of Fujian Province(No.JAT170882)Project of Financial Department of Fujian Province(Nos.K81139238 and K8911010)the Special Fund for Forestry Research in the Public Interest of China(No.201304401)
文摘Pine wilt disease(PWD)is a devastating disease affecting the growth of Pinus massoniana,often leading to withering and death.To reveal the changes involved during disease progression,we investigated the mRNA expression profile of P.massoniana infested by Bursaphelenchus xylophilus.The infestation resulted in the downregulation of genes involved in interactions with pathogenic pathways such as disease resistance gene,CC-NBS-LRR resistancelike protein,and the gene encoding a putative nematode resistance protein.Increased infestation pressure(number of nematodes inoculated)caused a continuous decline in the gene expression of stem samples.An infestation of P.massoniana also resulted in a pathway enrichment of genes involved in phenylpropanoid metabolism and flavonoid biosynthesis,which in turn reduced the levels of total phenols and total flavonoids.A downregulation of auxin responsive family protein was observed in infested samples,which resulted in a suppression of plant growth.Thus,upon B.xylophilus infestation,a downregulation of genes associated with the recognition of pathogens,PWD resistance,and growth regulation was observed in P.massoniana,together with a decrease in the levels of phytoalexinlike secondary substances,all of which resulted in withering and ultimately death of P.massoniana.
基金supported by the National Key Research and Development Program of China(2017YFD0600101).
文摘Genes homologous to members of the MRP gene family in Caenorhabditis elegans are important in drug resistance.To further explore the molecular mechanism of drug resistance in pine wood nematode(Bursaphelenchus xylophilus),we used bioinformatics approaches to analyze genomic data for B.xylophilus and identified Bx-MRP genes.We predicted the structure and function of the genes and encoded proteins.Using bioinformatics programs to predict and analyze various properties of the predicted proteins,including hydrophobicity,transmembrane regions,phosphorylation sites,and topologically isomeric structures,of these Bx-MRP genes,we determined that they function in transmembrane transport.From the results of RT-qPCR,the Bx-MRP family members confer significant differential resistance to different drug treatments.After treatment with different concentrations of emamectin benzoate,avermectin and matrine,the expression of each gene increased with increasing drug concentrations,indicating that the family members play a positive role in the regulation of multidrug resistance.
基金The work was supported by the National Natural Science Foundation of China(31870633,31700565,31670652,31570638 and 31270688)National Key Research and Development Plan of China(2017YFD0600102-03).
文摘Bursaphelenchus xylophilus,causal agent of pine wilt disease,causes extensive damage worldwide.Strate-gies are needed to inhibit population growth or block the spread of the invasive nematode to control pine wilt dis-ease.The gene daf-8 plays crucial roles in larval develop-ment and dauer formation in Caenorhabditis elegans,but little is known about its orthologue in B.xylophilus.In the present molecular characterization and functional analysis of daf-8 in B.xylophilus(Bx-daf-8),RT-qPCR showed that the expression of Bx-daf-8 gradually increased during the embryonic stage,peaked in the second-stage juvenile(J2),then dramatically dropped in the J3,and remained at that low level for the rest of its life.Bx-daf-8-transgenic C.elegans was employed to mimic the spatiotemporal expression of Bx-daf-8,which was expressed close to the pharynx during early embryogenesis and weakly throughout the whole body during late embryogenesis.It was observed in head neurons and tail ganglions throughout all postembryonic stages.B.xylophilus embryos were severely abnormal,and hatching rate decreased sharply after Bx-daf-8 knockdown.daf-16-1 and da-f16-2,downstream genes in the IIS pathway,also dropped sharply after Bx-daf-8 knockdown.We propose that TGFβmay crosstalk with the IIS pathway upstream of Bx-daf-16 and that daf-8 may act as a master regulator of daf-16 in B.xylophilus.However,knockdown of Bx-daf-8 did not lead to constitutive developmental arrest at the dauer larval stage,indicating that dauer entry in B.xylophilus might be controlled by several genes and is more complicated than in C.elegans.Bx-daf-8 alone did not control the dauer entry in B.xylophilus,but it was indispensable for embryogen-esis,providing a potential target for suppressing population growth of B.xylophilus.
基金Supported by Science Research Project of Ningbo Entry-Exit Inspection and Quarantine Bureau(YK12-2013)~~
文摘This paper mainly summarizes recent advances in the classification of Bursaphelenchus species, introduces several common methods of molecular biologi- cal detection and identification methods, and analyzes the advantages and disad- vantages of these methods.
基金The work was supported by the Fundamental Research Funds for the Central University(No.2572020DY11).
文摘Pine wilt disease caused by pine wood nema-todes is a deadly disease of the genus Pinus requiring strong quarantine measures.Since its discovery,it has been widely distributed throughout the world.China is one of the coun-tries with a severe rate of infections due to its abundant pine resources.In this study,nematode-trapping fungi were collected from pine trees in Ninghai City,Zhejiang Prov-ince,which is the key area of pine wilt control in Febru-ary,May,September,October and November.The results showed that nematode-trapping fungi of pine are abundant,especially the number and species detected in each month and are quite different;species of fungi in July,Septem-ber and November were more numerous and had higher separation rates.The dominant species in November was Arthrobotrys oligospora followed by Arthrobotrys cladodes;July and September it was A.cladodes.In each part of the tree,A.cladodes was the most widely distributed with a high separation rate.Therefore,it is the dominant fungal species in the area.Arboreal Bursaphelenchus xylophilus-trapping fungi are dependent on pine wood nematodes,and none were isolated from healthy pine trees.More arboreal B.xylophilus-trapping fungi were found under the bark than in the xylem.There is a close relationship between arboreal B.xylophilus-trapping fungi and pine wood nematodes.How to make use of these resources to reduce the damage of pine wood nematode fusarium is the focus of future research.
文摘Pine wood nematode （PWN）, Bursaphelenchus xylophilus （Steiner et Buhrer） Nickle is an important invasive alien species in forests of China and has become one of the most destructive forest diseases. In order to improve the resistance and resilience of pine forest ecosystems against B. xylophilus invasion and make the pine forest ecosystem more timely responsive to PWN invasion, we made some recommendations based on five years of intensive observations. We advocate a set of management measures with the theme “Prevention is priority, but integrated with curative techniques and ecological resilience” on the pine forest ecosystem invaded by B. xylophilus; details of accurate measures are proposed. The aim is to discover the underlying problems of present pine forest ecosys-tems and to take, correspondingly, administrative measures and strategies, which will encourage the pine forest ecosystem to develop in a benign way.
