1. Breeding tomatoes for resistance to monopartite and bipartite leaf curl viruses of India
Dr. H. C. Prasanna
Indian Institute of Vegetable Research, Varanasi

2. Presentation Outline Pyramiding Ty-2 and Ty-3 genes
Response of Ty and pyramided lines to tomato leaf curl viruses of India
Implications of Ty-2 and Ty-3 in development of ToLCV resistant tomato hybrid
Toward diversification of Ty genes

3. ToLCV disease complex
A complex of monopartite and bipartite begomoviruses cause leaf curl disease on tomato
Both mono and bipartite viruses are distributed throughout India
As many as 15 different species are reported from India
Mixed infection are common in the field

4. Begomovirus species diversity
Rehman et al. 2009

5. Whitefly (Bemesia tabaci) diversity
34 distinct genetic groups of whitefly have been identified at the global level
17 genetic groups have so far been identified in Asia
India is home for at least 6 whitefly groups
Asia I
Asia II 1
Asia II 5
Asia II 7
Asia II 8
Middle East-Asia Minor 1
Whiteflies on eggplant

6. Non-GMO options for ToLCVD management
JASHS 2000
HortSci 2009
TAG 2009
Mol Breeding 2007
JASHS 2009
Non-GMO options for ToLCVD management
Ty-2
Ty-1/Ty-3
Ty-4
Ty-5
Chr. 6
Chr. 11
Chr. 3
Chr. 4
Ty-??
Chr. 10
Euphytica 2013

7. Molecular breeding platform for Ty genes
MAS based on the markers located on opposite ends of the introgression
Focused on reducing the possibility of selecting recombinants
Ty-2
Ty-3
JASHS 2009
HortSci 2009
P6-25

8. Fine map location of Ty-3 gene
C2_At5g61510
23.7
PLoS Genetics 2013

9. Fine map location of Ty-2
Mol Breeding 2014

10. Pyramiding strategy Backcross scheme Cross Scheme
Follows backcross method of breeding
MAS for both foreground and background selection
Expensive and product resembles recipient
Followed pedigree method of breeding
MAS for only the genes of interest
Economical and products can be diverse lines
VRT
VRT
VRT-6-1-4
VRT
VRT

11. Agroinoculation based screening
Tomato leaf curl Bangalore virus (ToLCBV-[IN:Hess:08] and ToLCBB-[IN:Hess:08])
Tomato leaf curl New Delhi virus (ToLNDV-IN[IN:Pune:JID:2010] and CLCuMB-[IN:Sri:02]
Tomato leaf curl Palampur virus (ToLCPalV-[IN:Punjab:Tom:10])
Overnight cultures (on LB media) used for inoculation using needle puncture method
Cultures of different components and satellites mixed equally
Two shots at weekly interval for a month old plants

12. Disease severity scale
Symptom less
Slight yellowing
Yellowing & Curling
Yellowing , Curling,
Cupping & stunted
growth
Yellowing , Curling & Cupping
Ann Appl Biol, 2002

13. Response to ToLCBaV-A + ToLCBaV-β (75dpi)
Punjab Chhuhara
Ty-1Ty-1 (DSI <1)
Ty-2Ty-2 (DSI -1.53)
Ty-3Ty-3 (DSI <1)
Pyramided line
Ty-1+Ty-2+Ty-3 (DSI <1)

14. Response to ToLCNDV-A + ToLCNDV-B + CLCMV-β (75dpi)
Kashi Vishesh (DSI ~4)
Ty-1Ty-1 (DSI ~1)
Ty-2Ty-2 (DSI ~3)
Ty-3Ty-3 (DSI <1)
Pyramided lines
Ty-1+Ty-2+Ty-3 (DSI <1)

15. Response to ToLCPV-A + ToLCPV-B (75dpi)
Kashi Vishesh (DSI ~4)
Ty-1Ty-1 (DSI ~2)
Ty-2Ty-2 (DSI -2.2)
Ty-3Ty-3 (DSI <1)
Pyramided lines
Ty-1+Ty-2+Ty-3 (DSI <1)

16. Disease severity scores on pyramided lines
Genotype
Tomato Leaf Curl New Delhi Virus
Tomato Leaf Curl Bangalore Virus
Tomato Leaf Curl Palampur Virus
30DPI
60DPI
Kashi Vishesh
3.67a±0.07
3.93a±0.07
3.67a±0.13
3.86a±0.7
3.80a±0.00
Punjab Chhuhara
3.4a±0.00
4.00a±0.00
2.87b±0.17
3.66a±0.07
3.73a±0.07
3.86a±0.07
LA3473 (Ty-1)
0.40c±0.20
0.66c±0.07
0.13d±0.07
0.33c±0.07
1.13b±0.07
2.06b±0.13
C-8-6-1(Ty-2)
1.80b±0.00
2.87b±0.07
1.13c±0.07
1.53b±0.13
1.53b±0.18
2.20b±0.11
D14-1-4(Ty-3)
0.13c±0.07
0.44c±0.08
0.00d±0.00
0.00c±0.00
Pyramided lines
0.47c±0.07
0.20c±0.00
0.27c±0.17
0.63c±0.00
0.27c±0.07
0.40c±0.00
Note: Means with the same superscript letter are non-significant based on Tukey’s HSD test

17. Field tests in disease hotspot
Genotype
45DAP
90DAP
Kashi Vishesh
3.16b±0.03
3.43b±0.03
Punjab Chhuhara
3.67a±0.03
3.93a±0.03
LA3473 (Ty-1)
1.20c±0.06
1.83d±0.07
C-8-6-1(Ty-2)
1.47c±0.07
2.13c±0.09
D14-1-4(Ty-3)
0.23f±0.07
0.70fg±0.06
Pyramided lines
0.33f±0.03
0.47g±0.03
0.87d±0.03
0.97e±0.03
Note: Means with the same superscript letter are non-significant based on Tukey’s HSD test. DAP: Days After Planting

18. Ty-3 dosage effect Ty-3 /Ty-3 Ty-3 /ty-3 ty-3 /ty-3
Response to ToLCNDV-A + ToLCNDV-B + CLCMV-β

19. Implication of pyramiding in hybrid development
ty/ty
Ty-2/ty-2
Ty-3 /ty-3
Ty-2/ty-2+Ty-3 /ty-3
Response to ToLCNDV-A + ToLCNDV-B + CLCMV-β

20. Implication of pyramiding in hybrid development
Response to ToLCNDV-A + ToLCNDV-B + CLCMV-β
ty/ty
F5 progenies
Ty-2/ty-2+Ty-3 /ty-3
Ty-3 /ty-3
Ty-2/ty-2
The Ty-2 gene in either homozygous or heterozygous state enhanced the level of resistance of plants with heterozygous Ty-3

21. Open pollinated variety- Kashi Aman (VRT0801)
Determinate variety
Recommended for Zone IV (Punjab, UP, Bihar & Jharkahand)
Average fruit weight ranges from g
ToLCVD resistance conferred byTy-3

22. Targeted wild relatives
Diversification of resistance genes/alleles
Targeted wild relatives
S. lycopersicum
LA1972 X F1
BC1
BC2
Marker analysis
Mapping of target traits
Target region capture (BC2F2)
NIL as potential pre-bred line
in BC2F2 or BC2F3
Embryo rescue
Embryo rescue
S. lycopersicum
Marker analysis
Solanum chilense

23. Variabilty in BC1F1
Kashi Amrit
Hybrid
S. chilense
BC1F1 generation

24. Diversification of resistance genes/alleles
Un-inoculated
Response to ToLCNDV-A + ToLCNDV-B + CLCMV-β
Susceptible
VRT
VRT

25. Our Team
D. P. Sinha G. K. Rai Sarvesh P. Kashyap Ramkrishna Nikhil Singh Dr. Major Singh Dr. V. G. Malathi
Acknowledgement
Dr. Peter Hanson Dr. Roger Chetelat, TGRC, UC Davis ICAR for funding in-house project on tomato breeding at IIVR

26. Thank you