rice yellow mottle virus in tanzania 1 distribution and genetic diversity n.
Download
Skip this Video
Loading SlideShow in 5 Seconds..
Rice Yellow Mottle Virus in Tanzania: 1. Distribution and Genetic Diversity PowerPoint Presentation
Download Presentation
Rice Yellow Mottle Virus in Tanzania: 1. Distribution and Genetic Diversity

Loading in 2 Seconds...

play fullscreen
1 / 25

Rice Yellow Mottle Virus in Tanzania: 1. Distribution and Genetic Diversity - PowerPoint PPT Presentation


  • 121 Views
  • Uploaded on

Rice Yellow Mottle Virus in Tanzania: 1. Distribution and Genetic Diversity. 1 Kanyeka, Z.L., E.Sangu 1 , D. Fargette 2 , A. Pinel-Galzi 2 & H. Hebrard 2 1 Department of Botany , UDSM, P. O. Box 35060 Dar es Salaam, Tanzania. 2 IRD, BP 34394 Montpellier Cedex 5, France . INTRODUCTION.

loader
I am the owner, or an agent authorized to act on behalf of the owner, of the copyrighted work described.
capcha
Download Presentation

PowerPoint Slideshow about 'Rice Yellow Mottle Virus in Tanzania: 1. Distribution and Genetic Diversity' - creola


An Image/Link below is provided (as is) to download presentation

Download Policy: Content on the Website is provided to you AS IS for your information and personal use and may not be sold / licensed / shared on other websites without getting consent from its author.While downloading, if for some reason you are not able to download a presentation, the publisher may have deleted the file from their server.


- - - - - - - - - - - - - - - - - - - - - - - - - - E N D - - - - - - - - - - - - - - - - - - - - - - - - - -
Presentation Transcript
rice yellow mottle virus in tanzania 1 distribution and genetic diversity

Rice Yellow Mottle Virus in Tanzania: 1.Distribution and Genetic Diversity

1Kanyeka, Z.L., E.Sangu1, D. Fargette2,

A. Pinel-Galzi2 & H. Hebrard2

1Department of Botany, UDSM, P. O. Box 35060 Dar es Salaam, Tanzania. 2IRD, BP 34394 Montpellier Cedex 5, France.

introduction
INTRODUCTION
  • Part 1:
    • Distribution and Genetic Diversity
  • Part 2: ( In Progress)
    • Resistance breaking isolates
    • More Surveys in TZ & MAL, MOZ. & ZAM
introd cont d
INTROD. cont’d
  • RYMV is a major constraint of rice production in Africa
  • Tanzania is severely affected
  • Devastating disease epidemics, reported in all rice growing areas
  • Hot spots areas: Mbeya, Morogoro, Arusha, Lake Victoria Zone, ZNZ & Pemba
why this rymv study
WHY THIS RYMV STUDY?
  • DITRIBUTION & DIVERSITY UNDER-ESTIMATED IN TZ
    • Few isolates characterized
    • Number of strains identified
    • Strain Localization
    • Habitat Fragmentation
    • Diversity btn & w/n strains
    • New rice growing areas
    • Intensification of the rice cultivation systems
    • Eastern Arc Mountains proposed Center of Origin
  • EMERGENCE OF RESISTANCE-BREAKING ISOLATES
  • SPREADING PATHWAYS FROM ITS CENTER OF ORIGIN
objectives
OBJECTIVES
  • Characterize the molecular variability of isolates
  • Identify and Determine the distribution of strains
  • Identify their phylogenic relationship with the known strains and isolates
  • Investigate resistance-breaking isolates (Pathogeny)
  • Propose possible spreading pathways from its Center of Origin (EAM)
materials and methods
MATERIALS AND METHODS
  • Field Collection of isolates in TZ
  • Recovery of isolates on S-CV
  • Transport of samples to IRD-Laboratory, Montpellier , France
  • Molecular Analysis of isolates
    • Immunological typing-Elisa tests
    • Molecular Typing
    • Coat Protein(CP) Gene Sequencing
field trip in tz
Field trip in TZ

Map 1. Field collection of RYMV isolates -May 2005.

molecular characterization of isolates
MOLECULAR CHARACTERIZATION OF ISOLATES
  • Immunological typing (ELISA Tests)
    • DAS-ELISA using polyclonal antibodies
      • (IgGs ant RYMV Mg diluted at 1/1000 with Carbonate buffer pH 9.6)
    • TAS-ELISA using monoclonal antibodies
      • MAb A, MAb M, MAb E, MAb D & MAb G
  • Molecular typing
    • Extraction of total RNA
      • Plant RNeasy Mini Kit
    • RT-PCR
      • Coat protein (CP) gene amplification
      • Primers Amerce M & III
rt pcr temperature conditions
RT-PCR Temperature Conditions
  • Denaturation (5min at 94°C),
    • 30 cycles
      • Denaturation (3min at 94°C),
      • Annealing (1min at 58°C),
      • Elongation (1min at 72°C
      • Final extension (10min at 72°C)
  • Visualisation on 1% Ethidium bromide stained agarose gel
    • 720bp band expected
sequencing of cp gene
SEQUENCING OF CP GENE
  • Sent to another istitute for sequencing
  • SEQUENCE ANALYSIS
    • Assembling (EditSeq) DNASTAR
    • Alignment CLASTAL V DNASTAR
    • Estimation of genetic distances (Distance matrix) (MegAlign) DANSTAR
  • PHYLOGENETIC ANALYSIS (PAUP) VERSION 4 (Swafford, 2000)
    • Phylogenetic relationship RYMV strain
      • DNA Polymorphism
results and discusion
RESULTS AND DISCUSION
  • ELISA Tests
  • RT-PCR Product
  • CP Sequences
  • Phylogeny
  • DNA-DIVERGENCE
  • POLYMORPHISM W/N & BTN CLADES
serological elisa tests results
Serological (ELISA Tests) Results
  • Two Serological Groups: Ser4 & Ser5 (Monoclonal Tests )
  • Several Variants of Ser4 & Ser5
    • All Ser4 variants had negative rxn with MAb M
    • Three Ser5 variants had negative rxn with Mab A
    • Two Ser5 variants had no rxn with Mabs A & G
    • Serotype variants in the same site showed similar rxn with Mabs.
rt pcr product
RT-PCR PRODUCT

M

1

2

3

4

5

6

7

8

9

10

11

  • Amplified fragments of the expected size (720bp) were obtained
  • No fragment was amplified for Health control plant (6) blank (7) & water (9)

2

Lane

M=maker 1=Ifakara 2=Lumemo 3=Hembenti

4=Dihombo 5=Malinyi

6=H/Plant 7 Blank

8=Ndungu 9=water

10=RNA 11=cDNA

cp sequence results
CP SEQUENCE RESULTS
  • Three strains identified; 9=S4, 1=S5 & 13= S6
  • No new strains were found
  • First record of strain S4 occurrence & distribution in EAM- confirming the highdiversity and proposal of RYMV Center of Origin
  • Most rare strain was - S5 observed twice in same area also Ali 1999
  • S4 is a highly diverse strain than S6
  • High molecular diversity btn & w/n strains
phylogeography of rymv in tz
Phylogeography of RYMV in TZ

C & WA

LV

S4

EAM

LM

S5

EAM

EA

S6

EAM

conclusions
Conclusions
  • No new RYM strain were found however, the study reconfirmed the high diversity of RYMV in TZ
  • Strain S4 isolates were recorded for the first time in the EAM Region and share the common ancestor with Lake Malawi group
  • Strain S5 isolates are rare and are confined only to the Kilombero Valley.
  • Strain S6 isolates are widely spread but restricted only to the EAM region
  • Reinforcement of the proposition that EAM is the center of origin of RYMV
acknowledgement
ACKNOWLEDGEMENT
  • UDSM-Sida/SAREC Project
    • Financial support
  • IRD-Montpellier, France
    • Facilitation of Laboratory analysis
slide25
ASANTE SANA

THANK YOU

MERCI BEAUCOUP