Search for alternate hosts of the coconut Cape St. Paul Wilt Disease
( CSPWD ) pathogen
Team Members
Egya Ndede Yankey CSIR-OPRI
Fabian Pilet CIRAD – France
Robert N. Quaicoe CSIR-OPRI Sylvester K. Dery CSIR- OPRI
Dollet Michel CIRAD – France
Victoria P. Dzogbefia KNUST-Kumasi
INTRODUCTION AND JUSTIFICATION
Lethal Yellowing ( LY ) locally called Cape Saint Paul Wilt Disease
(CSPWD )is caused by a phytoplasma
It was first observed in 1932 and has since
become the foremost threat to the coconut industry in Ghana
Ankobra river area
Asebu
Asebu
Ankobra area
Why alternate hosts studies
The lethal yellowing phytoplasma has other hosts other than coconut in other places:
In the Americas it is reported to affect at least 38 palm species (Harrison et. al. 1999 )
In Jamaica the weeds Emelia fosbergii and
synedrella nodiflora were reported to host the
pathogen ( Brown et al. 2007 )
In Ghana the history of the disease at Cape Three Points makes a strong case for the presence of
alternate hosts of the phytoplasma:
1964: Devastated palms in the area (Ofori and Nkansah, 1997 )
1977: wiped off varieties introduced for resistance screening by CSIR-CRI ( Dery et al., 1997 )
1981: With the exception of SGD, all other varieties planted in a screening plot under the Fr-GH-C.I coconut project were
decimated ( Dery et.al.,1997 )
1995-Date : Nearly cleared all varieties (62.3%) imported under
the EC-STD III project ( Anon, 2006 )
It is in the light of this history that the presence of alternate hosts was investigated. The hypothesis being that, the phytoplasma, an obligate parasite needed another host to survive when their primary hosts, the coconuts were unavailable due to death from the disease
The identification of alternate hosts will greatly increase our understanding of the spread of the disease and help evolve sound control measures against its spread
In this work 57 plant species belonging to 23
botanical families were screened for their host
status to the CSPWD causing pathogen.
METHODOLOGY
Sampling sites: Cape Three Points ( W/R ) Asebu ( C/R )
Sampling Period:
July-Dec ( Rainy season )
March- April ( Dry season )
DNA Extraction
DNA was extracted from the test plants
employing the protocol of Daire et al. 1997 2 individual plants were pooled into each DNA sample.
For each plant species 15 DNA samples
were prepared
PCR & Gel electrophoresis
• Screening was first carried out with phytoplasma universal primers P1/P7.
• CSPWD specific primers G813/GAKSR were applied to positives from the above test
• PCR products were analysed by Gel electrophoresis on a 0.8% agarose gel and viewed under UV light. Restriction Fragment Length Polymorphism analysis ( RFLP ) with RSA 1, EcoR1, Hind III and Alu 1 was carried out on positives from the G813/GAKSR test.
• Cloning and Sequencing was performed by
Cogenics- France
Results of assay of rainy season samples
Primers
P1/P7 G813/GAKSR
sampl(N) +ves %+ves sampl(N) +ves %+ves
PLANT SPECIES ( ASEBU )
Commelina beghalensis 15 7 46.7 7 0 0
Solanum torvum 15 0 0 0 0 0
Calapogon mucunoides 15 2 13.3 2 0 0
Desmodium adscendeus 15 13 86.7 13 13 100
Malacantha alnifolia 15 4 26.7 4 0 0
Panicum maximum 15 1 6.7 1 0 0
Citrus limon 15 3 20 3 0 0
Nephrolepis bisserata 15 0 0 0 0 0
Tridax procumbens 15 0 0 0 0 0
Asebu samples cont’d Primers
P1/P7 G813/GAKSR
sampl(N) +ves %+ves sampl(N) +ves %+ve
Pennisetum purpureum 15 0 0 0 0 0
Ficus exasperate 15 0 0 0 0 0
Oxythenanthera abyssinica 15 0 0 0 0 0
Chromolaena odorata 15 0 0 0 0 0
Rauvolfia vomitoria 15 0 0 0 0 0
Sida acuta 15 0 0 0 0 0
Cape 3 points samples
Primers
P1/P7 G813/GAKSR
sampl(N) +ves %+ves sampl(N) +ves %+ves PLANT SPECIES
Setaria megaphylla 15 0 0 0 0 0
Mimosa pudica 15 0 0 0 0 0
Stachytarpheta indica 15 0 0 0 0 0
Aspilia africana 15 0 0 0 0 0
Borreria scabra 15 0 0 0 0 0
Dissotis rotundifolia 15 0 0 0 0 0
Asystasia gangetia 15 0 0 0 0 0
Ananas sativa 15 0 0 0 0 0
Rauvolfia vomitoria 15 0 0 0 0 0
Flagellaria guineese 15 0 0 0 0 0
Pueraria phaseoloides 15 0 0 0 0 0
Primers
P1/P7 G813/GAKSR
sampl(N) +ves %+ves sampl(N) +ves %+ve
Pueraria phaseoloides 15 0 0 0 0 0
Manihot esculenta 15 0 0 0 0 0
Voacanga africana 15 0 0 0 0 0
Saccharum officinale 15 0 0 0 0 0
Results of assay of dry season samples
Primers
P1/P7 G813/GAKSR
sampl(N) +ves %+ves sampl(N) +ves %+ve PLANT SPECIES ( ASEBU )
Stachytarheta indica 15 0 0 0 0 0
Desmodium adscendeus 15 0 0 0 0 0 Synedrella nodiflora 10 0 0 0 0 0 Ipomoea involucrata 15 0 0 0 0 0
Lypersicon esculentum 4 0 0 0 0 0
Jussiae spp. 15 0 0 0 0 0
Sporobolus pyramidalis 15 0 0 0 0 0
Brachiara deflexa 15 0 0 0 0 0
Cymbopogon citratus 15 0 0 0 0 0
Capsicum annum 15 0 0 0 0 0
Asebu samples cont’d Primers
P1/P7 G813/GAKSR
sampl(N) +ves %+ves sampl(N) +ves %+ve
Eleusine indica 15 0 0 0 0 0
Digitaria adscendens 15 0 0 0 0 0
Euphorbia heterophylla 15 0 0 0 0 0
Solanum tuberosum 15 0 0 0 0 0
Primers
P1/P7 G813/GAKSR
sampl(N) +ves %+ves sampl(N) +ves %+ve PLANT SPECIES ( Cape 3 pts )
Sanseviera liberica 15 4 26.7 4 0 0
Panicum laxum 15 0 0 0 0 0
Cassytha filiformis 15 0 0 0 0 0
Lantana camara 15 0 0 0 0 0
Clerodendrom capitatum 15 0 0 0 0 0
Morinda lucida 15 0 0 0 0 0
Chassalia kolly 15 0 0 0 0 0
Rottboellia exaltata 15 0 0 0 0 0
Cyperus spp. 15 0 0 0 0 0
Alcherna cordifolia 15 0 0 0 0 0
Cont,d
Primers
P1/P7 G813/GAKSR
sampl(N) +ves %+ves sampl(N) +ves %+ve PLANT SPECIES(Cape 3 pts)
Pennisetum pedicellatum 15 0 0 0 0 0
Spigelia anthelmia 15 0 0 0 0 0 Phyllanthus amarus 15 0 0 0 0 0
Justica flava 15 0 0 0 0 0
Paspalum scrobiculatum 15 0 0 0 0 0
Amplification of Desmodium adscendeus samples with G813/GAKSR
M 1 2 3
M45 6 7 8 9 10 11 121314 15 3&15:positve controls
1&2,4-14: Desmodium samples
Restriction Fragment Length Polymorphism (RFLP) analysis of Desmodium adscendeus samples (G813/GAKSR products) using
3 restriction endonucleases
M 1 2 3 4 5 6 7 8 9 10 11 12 13 14
Controls: Lanes 1, 5 and 10
RSA 1 Hind III EcoR1
