Inception meeting presentation on cassava disease by Wilmer Cuellar. Plant Virologist CIAT.

1. RTB-Rapid response to
transboundary and emerging
diseases
Wilmer J. Cuellar, Ph.D.
Plant Virologist
CIAT-AgBio
Sept 11th, 2019. Vientiane, Laos

2. Flagship Project 3: RESILIENT CROPS

3. Building a sustainable futureciat.cigar.org
Mosaic diseases in Asia and the Americas
Cassava Common Mosaic Disease
The Americas
Cassava Frogskin Disease
The Americas
Cassava Mosaic DIsease
Africa, Asia
C
Similar symptoms different pathogens
BA

4. Sri Lankan cassava mosaic virus
(SLCMV) group
African cassava mosaic virus
(EACMV & ACMV) groups
Related viruses
isolated from related
Euphorbia, Jathropha
and Pedianthus hosts
Cassava mosaic geminiviruses tree
Indian cassava mosaic virus
(ICMV) group

5. First report in Cambodia
Key information missing
Prevalence, severity of symptoms,
size of the sampled field, etc.

6. The challenge of communication
?????????Cassava Mosaic Disease
?????????

7. 7
Reports of CMD caused by SLCMV: 1998 - 2019

8. Feb 2016-Comparison of available tests for SLCMV
Evidence for:
- Mixed infections
- Latent/symptomless infections
- PCR more sensitive than ELISA

9. Fig. 1. Representative circular genomic map of cassava mosaic geminiviruses from Africa adn Asia. (A) African cassava
mosaic virus (ACMV) DNA-A (AJ427910), B) East African cassava mosaic Cameroon virus (EACMCV) DNA-A (AF112354). C.
Sri Lankan cassava mosaic virus (SLCMV) DNA-A (KT861468).
Genome characterization (DNA-A)
***
*
https://pestdisplace.org/sequence/CMDAFRICA

10. Replication Associated Protein sequence alignment
Fig. 4. Alignment of nucleotide sequences from Asia and Africa. Nucleotide sequences were aligned using sequence of Replication Associated Protein. The restriction enzyme site StuI are
shown in gray and CMBRep/F primer are shown in red square .
CRTCAATGACGTTGTACCAStu I
CMBRep/F

11. Fig. 3. Results of PCR on samples infected with cassava mosaic geminiviruses. (A) multiplex PCR for the detection of ACMV and EACMCV using replicase gene-
specific primers, (B) PCR using EACMCV-specific primers, (C) restriction enzyme analysis using StuI from PCR bands amplified with the EACMV specific primers,
(D) PCR for the detection of SLCMV using coat protein gene-specific primers. Samples from Malawi (lanes 1-5), Vietnam (lanes 6-10) and Cambodia (lanes 11-15),
C+: Undigested positive control, C-: healthy cassava, W: water control, M: 1 Kb Plus DNA Ladder (Invitrogen) and black arrows head indicated 650 bp.
A quick test for ‘false positives’
Primer name
Product
size (bp)
# mismatches vs.
SLCVM
Region Reference
SLCMV-F
771 - CP Dutt et al. (2005)
SLCMV-R
CMBRep/F -
RepACMVRep/R 368 1 (in 3’) Alabi et al. (2008)
EACMVRep/R 650 3 (2 in 5’)
CMBCP/F 2 (5’ and 3’)
ACMVCP/R 655 5 (3 in 3’) CP Alabi et al. (2008)
EACMVCP/R 524 8 (4 in 5’)
ACMV-AL1/F
1063
7 (4 in 5’) Intergenic
region and
MP
Zhou et al. (1997)
ACMV-AR0/R 10
UV-AL3/F
1086
3 (2 in 3’)
Rep - Ren Zhou et al. (1997)
UV-AL1/R2 8
ICMV-F
460
3 (2 in 5’)
Rep
Duraisamy et al.
(2013)ICMV-R 6 (4 in 3’)
SLCMV-REP/F
1056
-
Rep
Duraisamy et al.
SLCMV-REP/R - (2013)
PA
560
-
MP
Wang et al.
PB 1 (in 5’) (2016)

12. DNA -> digital -> BigData: Phylogenetics
- Collect, organize, analyze

13. DNA -> digital -> BigData: Phylogenetics
- Collect, organize, analyze

14. Phylogeography
- Collect, organize, analyze
https://nextstrain.org/community/pestdisplace/CMDASIA
Using Open Source tools e.g. Nextstrain. We prepared an initial tree for SLCMV,
you can contribute by submitting your sequences to: CIAT-PestDisPlace@cgiar.org

15. Symptom characterization, insect-proof propagation, use
of grafting and mechanical inoculation protocols
Clean and well characterized planting material
available for in planta tests
Germplasm collection-indicator plants-grafting tests

16. RNA/DNA from 20 mg of dried leaves
0
500
1000
1500
2000
2500
3000
3500
4000
Casanare 2013 Meta 2014 Sincelejo 2014 Sucre 4to muestreo 2014
Sucre cosecha 2015 Argentina 2015 Hawaii 2015 Costa Norte 2018
[ng/µl]
Standardization of procedures in the lab
siRNA
sequencing
Pooling samples
Standard field sampling

17. Molecular diagnostics supported by AI mobile apps

18. Building a sustainable futureciat.cigar.org
https://pestdisplace.org
Mobile phone AI apps
Scientific reports,
social media, news
Molecular diagnostics
Sequence data

19. Building a sustainable futureciat.cigar.org
A platform for early communication: diagnostics
and surveillance
You will need:
-Define the field sampling
strategy (contact us for support).
-Obtain your ORCID number:
www.orcid.org and log-in in
https://pestdisplace.org
-Activate the GPS in your mobile
phone and you are ready to go.

20. Thank you!