Embed Size (px)
Citation preview
Current status and progree of the cereal cyst nematodes Heterodera avenae and H. filipjevi on wheat in China“
Peng, D,, Nicol, J.M., Huang, W, Chen, S Zheng, J. Li, H,
Institute of Plant Protection, Chinese Academy of Agricultural Sciences, Beijing, China E-mail:[email protected]
4th Asian Conference on Plant Pathology， Darwin ，April 26-29
CCN is a Sedentary nematode forming white adult females on the root system which later become hard brown CYSTS which survive in soil.
• Below ground: bushy-knotted’ appearance usually with several females visible at each root, which later become cysts.
• Above ground: pale green patches of poor growth with yellow lowered leaves and generally plants with few tillers.
• Generally one generation per cropping season• Only cereal host (H. avenae)• Yield losses worldwide between 5-50%.• The damage is greater when the crop is stressed.
Cereal Cyst Nematodes caused seriouseffect on what yield and
The ‘knotted’ root system and white females which turn into
cysts
Symptoms：Yellow, darf
2005年菏泽
April,1992, dingzhou， Hebei Province
December，2007，Xuchang Henan
Confusion withnot speciality, confused with lack water
fertilizer Lack
2010/11/12Hufeng county，Sha’an Province,
Wheat variety Xiaoyan 22
CCN Symptoms: 2010/4/22, Xingyan, and XuchangHena
Quick spread by havesting tractor
Frequency 43%
Hubei 1989
The Cereal cyst nematode - CCN (Heterodera spp.) wasfirst found and reported from Hubei province in the centreof China in 1989 ( Peng, Chen, Wang, 1989)
CCN Distribution
Hubei, Tianmen, 1989 Peng Chen & WangHenan: Anyang, 1989 Liu, et al.
Zhenzhou 2005 Li et al.Hebei, Dingzhou， 1990 Peng et al.Beijing, Chanping 1991 Peng et al.Shanxi, Taigu 1992 Peng et al.Inner Mongolia, 1992 Peng et al.Qinghai 1992 Peng et al. Hou, 2006Anhui, 1998 Zheng, et al.,
2005, Huiping，Ian et al.,Shangdong 2005 Julie Deliang et al.Gansu 2007 Ian，Huixia et al,Shan’xi 2007 Peng et al.,Jiangsu 2010 Li, HM, et al.,Ningxia 2011.1 Huang, Peng
13 provinces
More than 300 counties
Province Year Scientist
CCN has been found in 13 provinces
CCN caused seriuos problem on wheat production, estimated occurrence area 40 Mha
5 province of large wheat produce in China
Henan： yield loss general 17-42%，serious 80%，17 MhaHebei： yield loss general 15-25%，serious 40%，7MhaShangdong：17MhaJiangsu： 3MhaAnhui： 5Mha
Hubei Sha′anxi Gansu Qinghai Ningxia Shanxi
Frequency of isolation and population density of CCN
Survey results from 1998-2008Global reports indicate a damaging threshold for CCN is 5 eggs/g soil
IMPACTS on wheat production in China
Cereal cyst nematodes: complex avenae filipjevi H filipjevi exist in China wheat field Population is higher than avenae population wheat production get more challenge and threaten
Heterodera filipjeviXuchang, Linying, WeihYanjing、Tangying
Species: H avenae H filipjevi
菲利普孢囊线虫H filipjeviNingxia Qinghai
Heterodera filipjevi
HinfI TaqI
M:100bp;: 1 positive control, HN-34 Henan xuchang，HN-29 Henan Xuchang， H glycines；HN-25zhenzhou，HB-22 Hebei， HB-23 Hebei， Shijiazhuang； QM-1； QH-1 ； SD-7 ; SD-8 ；
H. filipjevi: Linying, Xuchang, Weihui and Yanjing, Also found in Ningxia, Qinghai
H avenaeH filipjevi
Heterodera.filipjevi ITS-RFLP
Plant Disease ，2010.10：1262First Report of the Cyst Nematode (Heterodera filipjevi) on Wheat in Henan Province, China
Banpoupu
Xuchang
Henan
Henan privinceLinying (Peng Dl et.al., 2009)Xuchang (Peng Dl et.al., 2009)Weihui (Peng Dl et.al., 2009)Yanjing (Peng Dl et.al., 2009)Tanying，Henan province (Peng Dl et.al., 2010)Huaiyang (Li HL, 2010)Jiaozuo Boai (Li HL, 2010)Shangshui (Li HL, 2010)Lulong, luoyang (Li HL, 2010)
NX31 Ningxia (Peng Dl et.al., 2010)QH19 Qinghai (Peng Dl et.al., 2010)
Heterodera filipjevi found in 9 county, Henan
Sequences registration in GenBank
ITS sequences
Linying：GU083595,Xuchang：GU083596,Weihui：HM147944,Yanjing：HM147945
D2D3 region
Linying：GU083592Xuchang：GU083593Weihui： GU083594Yanjing： GU83597
http://apsjournals.apsnet.org/doi/abs/10.1094/PDIS-04-10-0309
Molecular characteristics analysis of Heterodera avenae, H filipjevi
Stucture of ribosomal DNA clusters
TW 81 - 5’ GTTTCCGTAGGTGAACCTGC-3’AB 28 - 5’ ATATGCTTAAGTTCAGCGGGT-3’
18 S 5.8 S 28 SITS1 ITS2
AB28TW81rDNA1 rDNA2
rDNA1.58S
IGS 5’-ETS 3’-ETS
The amplification of the rDNA ITS regions of each population yielded one fragment of approximately 1045(Heterodera avenae), 1054 bp ( H filipjevi ). No PCR products were obtained in the control lacking the DNA template
M：DL2000 marker；1-17：HY127B， HY65A，ZHZ162A，HY5B，CE18，DT142A，YL4B，GH132A，YBT11B，HY126B，HZXZA，DT2A，XS3，HG24，SF2，HA11，HA10；18： negative control.
1000bp→
M 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18
Molecular characteristics of Heterodera avenae and H filipjevi
H filipjevi HN-32,HN-34 codes:M-100bp, N-unrestricted PCR product , 1/2-AluI, 3/4-CfoI, 5/6-HifI, 7/8-SatI, 9/10-PstI, 11/12-RsaI, 13/14-TaqI, 15/16-Tru9I.
H avenae HN-16,HN-30 codes:M-100bp, N-unrestricted product , 1/2-AluI, 3/4-CfoI, 5/6-HifI, 7/8-SatI, 9/10-PstI, 11/12-RsaI, 13/14-TaqI, 15/16-Tru9I.
AluI CfoI HifI SatI PstI RsaI TaqI Tru9I AluI CfoI HifI SatI PstI RsaI TaqI Tru9I
Henan、Hebei, Shandong，Jiangsu，Gansu， Qinghai，
76 populations ITS
Henan、Hebei, Shandong，Jiangsu，Gansu，Qinghai，
76 populations D2D3
Heterodera avenae complex species
Europe
H. avenaeH. mani H. aucklandica
Asia
France
H. “avenae”
H. pratensis
H. ustinovi
H. filipjevi
H. australis
H. arenaria
with underbridge
without underbridge
rDNA-ITS and D2/D3 OF Heterodera avenae from Gansu province
rDNA-ITS of Heterodera avenae
A: AluⅠ；B: CfoⅠ；C: Bsh1236Ⅰ；D: AvaⅠ；E: RsaⅠ；F: BsuRⅠ；G: HinfⅠ；H: MvaⅠ；I: TaqⅠ). M: DL 100 bp marker）; U: uedigestion; 1-7: GsWx, GsGg, GsWw, GsMq, GsTs1, GsTs2, GsYz
rDNA-ITS-RFLP analysis of Heterodera avenae from Gansu province
Species specific primer for diagnosis Heterodera filipjevi
Forward: TW81 Reversed HfR1
cDNA library of H avenae J2, 6000 EST7 genes cloned already
species genes Accessionnumber
Year
Heterodera avenae Ha-eng-1 FJ839965 2009.3.19
Heterodera avenae Ha-CBP1 GQ178086 2009.05.21
Heterodera avenae Ha-PEL1 GQ998895 2009.09.25
Heterodera avenae Ha-PEL2 GU138156 2009.10.29
Heterodera avenae Ha-dsl-1 HM147942 201-04-27
Heterodera avenae Ha-dsl-2 HM147943 201-04-27
Heterodera avenae Ha-expb1 HQ386233 2010-10-12
PARASITISM GENES
A new endoglucanase-like gene (Ha-eng-1a GenBank ID: FJ839965) was cloned from H. avenae J2 by RACE
5’race 3’race
The full-length cDNA designated asHa-eng-1a is 1565 bp long
Conserved domain search shows the deduced protein Ha-eng-1a contains a catalytic domain and a carbohydrate-binding module of family 2 (CBM II)
Ha-eng-1a
Gts-eng-1
Gr-eng-1
Hs-eng-1
Hg-eng-1
cyst nema
Rs-eng-1a
Da-eng-1
Rs-eng-1b
Dd-eng-1
migratory nema
Mi-eng-1
GHF5
GHF45 Bx-eng-1
100
100
47
100
80
69
51
93
0.00.20.40.60.8
Phylogenetic tree of the new endogulcanase gene Ha-eng-1a.
Southern blot analysis shows Hg-eng-1a is a member of a multigene family
Ha-eng-1a clustered in a group of cyst nematode endogulcanases and classified into the GHF5.
Exon / intron organization of Ha-eng-1a gDNA
Ha-eng-1a gDNA clone is 2129 bp long containg eight introns
Ha-eng-1a specifically expressed in the two subventral gland cells of H. avenae
Recombinant protein Ha-eng-1a showed a strong cellulase activity in vitro
Cellulase activity assay on carboxymethylcellulose (CMC) agar plate test
Tissue localizaiton of Ha-eng-1a mRNA
A new cellulose binding protein gene (Ha-cbp-1， GQ178086 ) was cloned from H. avenae J2 by RACE
3’RACE 5’RACE
The full-length cDNA designated as Ha-CBP-1 is 964 bp long composed of a 393-bp open reading frame, a 55-bp 5’-untranslated region (5’-UTR) before the ATG initiation codon and a 513-bp 3’-UTR containing a polyadenlylation signal (AATAAA)
The predicted HA-CBP-1 shared similarities of 76% and 75%with the HG-CBP-1 and HS-CBP-1, respectively.
Sequence alignment between deduced amino acid sequence of cellulose binding protein HA-CBP-1 from H.avenae with a cellulose binding protein from other nematodes .
The genomic sequence of Hg-cbp-1 which contained two introns
Phylogenetic trees of six CBPs based on the CBPs amino acid sequence using ME method.
Capacity building to work with CCN in China
2006 – 1st 5.13-18, Cereal Cyst Nematode Workshop held in Hebei province with ATSE Crawford Foundation Australia, and CAAS.
2007 – 2nd 4.12-14, Cereal Cyst Nematode Workshop held in Beijing, IPP-CAAS
2008-3rd 8.3-5, Cereal Cyst Nematode Workshop held in Qinghai, IPP-CAAS
2009-4th 5.9-11, Cereal Cyst Nematode Workshop held in Zhengzhou, IPPCAAS，HNAU
2010-5th 6.13-15,Cereal Cyst Nematode Workshop held in Beijing, IPPCAAS
First China-CIMMYT workshop on CCN
Capacity building to work with CCN in China
CHINA-CIMMYT workshop on CCN, 2010, 6.12-15
10Th Chinese Nematology Symposoms， 2010, 8.13-15
CCN network in China
CIMMYTInstitute of plant Protection, CAASInstitute of plant Protection, HbAAFHenan Agricultural UniversityChina Agricultural UniversityHuazhong Agricultural UniversityHuanan Agricultural UniversityAnhui Agricultural UniversityInner Mongolia Agricultural UniversityQinghai Academy of Agricultural and Forestry SciencesGansu Agricultural UniversityShandong Agricultural UniversityQingdao Agricultural UniversityWest North Agricultural and Forestry UniversitySeveral local plant protection station from Henan, Hebei, Inner Mongolia, Qinghai and Gansu
2010/4/22 henan,
Training
Ningxia, 2010/6/9
Cereal cyst nematodes Projects in China
THREE PROJECTS ：
MOA-China project NO. 200903040NSF-CIMMYT project no. 30921140411MOST-China project NO. 2009DFB30230
National CCN Project in China
MOA-China project（NO. 200903040）Crop Cyst Nematode Managements Research and
Development (2009-2013） Leading Scientist：Deliang PengPartner – CAAS-IPP， 28 University，Institution，
Extension Station, about 50 scientists invovledHAAFS---Dr.Shulong Chen, HAU---Dr.Honglian Li,NAU---Dr.Hongmei Li, IMAU--Dr.Hongyou Zhou
Donor-MOF, MOA 3 M Euros
National CCN Project in China
Cereal cyst nematodes Project in China
NSF-CIMMYT project no. 30921140411:Development of molecular tools for Cereal Cyst Nematode (Heterodera spp.) diagnostics and identification of wheat resistance and nematode parasitsm genes Project Leader: Deliang Peng2010-2012. Partner – CAAS and other Chinese
HAU—Honglian LiHAAFS—Shulong Chen
Donor-NSF China
MOST-China-CIMMYT Project
MOST-China project NO. 2009DFB30230 Project title:
Study on the Pathotypes and and virulence genes of cereal cyst nematode in China 2009-2013
Leading Scientist: Deliang PengPartner – CAAS and associated Chinese NARsDonor-MOST
Ministry of Science and Technology, China
ONGOING RESEARCH
Risk Assessment – yield loss, damage thresholds and population dynamics
Develop a molecular diagnostics Parasitism gene and effector Pathogenecity, pathotype, ecotype, and biology Control of CCN through IPM practices including;Identification of resistance against known pathotype,
followed by subsequent breeding to incorporate resistance.Use of rotation Identification of Biological control agents
Cooperation with other countries ,scientists
ACKNOWLEDGMENT
Nature Sciences Foundation of China
Ministry of Agricultural
Ministry of Sciences and Technology
CIMMYT
SKLBPI
Acknowledgment
Research Team
Deliang PengDongsheng ZhangWenkun Huang Wenting He
Huan PengHaibo LongGaofeng WangXiaochuan GuWenxing Ye
Qiong Wang Binyu WangLingling Dai Xiaoli QiXiaoqiu Zong
The tentative agenda including1. Global status of the distribution of CCN2. The economic importance and population dynamics of CCN on wheat3. Control strategies of CCN in wheat using host resistance4. Control strategies of CCN other than host resistance5. Use of molecular tools for research with CCN (such as pathogen diagnostics, phylogeny studies and host resistance).6. CCN genome and parasitism genes7. Biological control and IPM
ORGANISERS
CIMMYT: Dr Julie NicolIPP-CAAS: Dr Deliang PengChinese Society of Nematologists
Registration and Accommodation
Registration fees are 250 Euros, Meals: Included
Hotel: Hubei Hotel, four Star hotel,
Hotel costs: 40 Euros standard room, 50 Euros single room
Budget hotels nearby
Sightseeing: Many amazing options
Shopping: Absolutely everything your heart desires
Proceeding: Official proceedings will be published.
Contact
Prof. Deliang Peng
Institute of Plant Protection, Chinese Academy of
Agricultural Sciences, Beijing 100193, China
E-mail: [email protected]
Tel ++ 86 10 62815611Fax + 86 10 62896114
Mobile: 86 13381056763
