Combining Insect Resistance with Virus Resistance to Construct a Dual System of Virus Control

Transcription

1 Combining Insect Resistance with Virus Resistance to Construct a Dual System of Virus Control Martha A. Mutschler, Darlene DeJong Dept. of Plant Breeding and Genetics Jessica Houle, George Kennedy Dept of Entomology, NCSU

2 Potential Impact of Breeding for Pest and Virus Resistance Control of insects to manage direct damage can be difficult and is expensive Pest control products are becoming more expensive and narrower in spectrum Control of virus through vector control using pesticides is not sufficient Virus resistance can also be of limited effect Questions for the Cornell breeding program: What is the potential for using vector resistance as an additional tool for both insect and virus management? Is the potential greater through combining vector resistance with virus resistance?

3 Acylsugars Provide Effective Insect Control Some of Pests Controlled by Acylsugar of S. pennellii LA716 Potato aphid* Green peach aphid* Tomato Psyllids Fruitworm* + Tomato fruitborer Armyworm* + Silverleaf whitefly (Bemisia) * Acylsugars are non-volatile, on the surface, effective on contact. Acylsugar affect a wide range of insect pests, including important virus vectors Control is based on deterrence of feeding and/or oviposition (species with *). Other species also show some level of antibiosis (species with +). Greenhouse whitefly* Leafminer* Thrips Spider mites Silverleaf Whitefly S. Bauer Thrips tabaci Frankliniella occidentalis A. N. Sparks

4 Acylsugars & Trichomes of S. pennellii & Tomato Tomato S. pennellii Type V nonglandular trichome Type VI glandular trichome Type IV glandular trichome Acylsugar droplet Acylsugars exuded from type IV glandular trichomes. These trichomes are present on epidermis of all green portions of plants. Acylsugars constitute ca. 95% of type IV glandular trichome exudates. Tomato produces extremely low levels of acylsugars Highest level of production in S. pennellii accessions Acylsugar production is NOT controlled by a single gene

5 Acylsugars Deter Feeding/Oviposition On Tomato by Silverleaf Whitefly In No-Choice Tests. Acylsugars No. of Eggs/leaflet Nymphs/leaflet (ug /cm 2 ) cages Mean No. Mean No a a b b c 1.38 c c 0.00 c c 0.00 c Means followed by the same letter are not signficantly different (P > 0.05; Fisher's Protected LSD). Data analyzed using square root. Egg counts taken 48 hr after infestation Liedl et al, 1995 J. of Econ. Ent. 88:

6 Acylsugars of 97FL for Silverleaf Whitefly. Bradenton, FL, Spring 99 Plant data M82 97FL No. Plants Acylsugars (ug/cm2) ** Silverleaf Whitefly No. Hatched Eggs Silverleaf Whitefly No. First Instars Silverleaf Whitefly No. 2nd & 3rd Instars Silverleaf Whitefly No. 4th Instars ** * ** ** *, ** egg or infestation means that are significantly different from tomato mean at P = 0.05, 0.01 levels using Dunnett's test for planned comparisons. Mutschler and Schuster,

7 Breed 2 nd Generation Acylsugar Lines In further backcrosses, select both for acylsugar levels and with markers to find plants with shorter/fewer S. pennellii introgressions This accelerated breeding process Created a series of >30 2nd generation acylsugar lines 4 to 6 introgressions, 10 to 15% of genome S. pennellii DNA Were these lines also capable of controlling whitefly?

8 Test Whitefly Generation Line and Controls Spring 2006 Florida Field Trial (a) Tomato control Insect pressure very high, as demonstrated by tomato control vs. 97FL control. (b) 97FL

9 Acylsugar Levels And Silverleaf Whitefly Counts On Acylsugar Lines Wimauma, FL, 2007 Line Mean acylsugar level ug sucrose/mg dried leaf tissue Mean egg number Mean number 1 st instar per leaflet Mean No. 2nd/3rd instar per leaflet Mean No. 4th instar per leaflet FLA F 3.0 A 8.8 A 37.3 A 72.9 A DE 3.0 ABC 2.4 B 3.5 B 1.8 C DE 2.9 ABC 0.9 BC 2.0 BC 2.1 CD BC 3.5 ABC 1.0 BC 0.7 D 1.0 CD BC 2.4 BCD 0.9 BC 1.9 CD 0.6 CD CD 1.1 BCD 1.2 BC 2.6 BC 0.7 CD B 1.7 BCD 1.0 BC 0.3 D 0.4 CD BC 0.6 CD 0.5 BC 0.6 D 0.2 D 97FL 46.9 A 0.0 D 0.0 C 0.0 D 0.0 CD Levels not connected by same letter are significantly different at 0.05 level Cooperative trial performed by Dave Schuster, U of Fl

10 Benchmark Acylsugar Line: CU CU is a BC 6 derived line. Contains 4 S. pennellii introgressions S. pennellii DNA accounts for ca. 10% genome Accumulates 45% the acylsucrose of CU97FL 84 cm 84 cm 7 cm 7 cm 18 cm 9 cm 18 cm 9 cm Use 97FL and CU to see if acylsugar mediated control of vector control reduce infestation by virus 12 cm 12 cm

11 Affect Of Acylsugars On Infection Rates of Greenhouse Whitefly Transmitted TICV (San Diego) Tomato Variety C Percent Visually Symptomatic Percent Infected by Dot Blot Assay M82 41% 59% S. pennellii LA716 0% 27% M82 x 97FL 0% 31% IL3-2 x 97FL 7% 31% IL6-3 x 97FL 3% 33% IL11-2 x 97FL 0% 26% Symptoms determined by visual observation. Infection confirmed by dot-blot hybridization using virus-specific probes. (Mutschler and Wintermantel 2005)

12 Thrips (F. occidentalis) Adults & Larvae, &TSWV Incidence vs. Acylsugar Levels Levels not connected by same letter are significantly different at 0.05 level by Tukey test. Test performed by George Kennedy in a field trial in NC in 2009.

13 Control of Virus Is More Demanding Than Control of Pests Insect damage must be controlled to threshold of economic damage, complete control not needed Infection by as few as one viral particle can result in systemic disease and major loss of yield and quality Virus resistance genes can fail as solo means of control Acylsugar mediated insect resistance at current acylsugar levels and types impacts incidence of viral diseases, but does not provide complete control Possible approaches to control insects and insect-vectored virus Add QTL to raise acylsugar level or alter acylsugar chemotype to further enhance insect control Combine insect resistance and virus resistance to create dual layer control Combination of both these approaches

14 Process for Combining Acylsugar Production and Virus Resistance Cross tomato line with virus resistance gene x CU Backcross resulting F1 to CU Select by marker analysis for BC1F1 plants with the virus resistance gene and homozygous S. pennellii introgressions Select BC1F2 for plants homozygous for virus resistance gene and all S. pennellii introgressions Resulting lines equivalent to CU for introgressions but possesses the virus resistance gene This process used to create acylsugar lines also possessing TY1/TY3, or Ty3, or Sw7

15 Acylsugar Tomato Lines Also Possessing TYLCV Resistance Genes CU Ty1+Ty3/acylsugar line Aug Ty3/acylsugar line Aug. 2010

16 Acylsugar Tomato Lines Possessing TY1+Ty3 with Improved Chromosome 3 Introgression Ty1+Ty3/acylsugar line Red fruited Ty1+Ty3/ acylsugar plants after removal of inversion and r gene (2010) TY1+Ty3/acylsugar line with additional modification of Chr. 3 introgression (2011)

17 Sw7 Lines Chromosome 12 Sw7 introgression relatively large, location of Sw7 within introgression not precisely known Introgression used also carries a recessive fruit color abnormality that must be eliminated Selected for BC1F1 plants with recombinations in Chromosome 12 introgression in process of breeding Sw7/acylsugar lines Need to select for this recombination led to selection of plants not homozygous or possessing all S. pennellii introgressions (lengthen breeding process) Had to test for presence of Sw7 in lines with different subregions of this introgression

18 2011 Sw Field Trial, North Carolina Seed Source Pedigree Fruit mottle associated full SW7 introgresion? AS as % a 41.0 clet8k c2_at5g c2_at2g g T CT ssr20 June 22nd planting Total number plants assayed b Percentage infected b CK-12 (F5) from CT230-4 Sw-7/Sw-7 stock from BYU yes NA Grower planting no NA control no 100 A SW7 Acylsugar line no 72.1 B SW7 Acylsugar line no 63.0 BC SW7 Acylsugar line no 52.2 BC SW7 Acylsugar line no 40.1 CD SW7 Acylsugar line yes 45.6 CD SW7 Acylsugar line yes 40.3 CD SW7 Acylsugar line no 20.6 DE a Lines lacking some S. pennellii introgressions have lower levels of acylsugars b Cooperative Sw7 experiment performed by Jessica Houle and George Kennedy, 2011 trial, NC

19 Sw5 Acylsugar Lines Sw5 acylsugar lines produced by crossing Sw5 line with 97FL as acylsugar parent Had to select for fewer/smaller introgressions, significant differences in acylsugar level Resulted in a series of lines with different introgressions and some with higher acylsugar levels Field trial attempted in NC summer 2011, few insects and no virus even in susceptible controls at that location

20 Current and Future Work: Whitefly/TYLCV TY1/Ty3, and Ty3 acylsugar lines Producing seed of new Ty1/Ty3 and Ty3 lines and hybrids using these lines Arranging for field trials in Florida, where both insect and virus pressure are fairly reliable How will acylsugar hybrids with different levels of acylsugar impact insect presence on plants How will acylsugar hybrids with different levels of acylsugar, and without virus resistance or either homozygous or heterozygous for virus resistance affect virus infection?

21 Current and Future Work: Thrips/TSWV Continuing to refine Sw7 and Sw5 acylsugar lines Incorporating modified Chromosome 3 in Sw7, Sw5, Sw5+Sw7 acylsugar lines Combine Sw5 and Sw7 in acylsugar lines Field trials using Sw7 and Sw5 acylsugar hybrids Arranging for field trials in North Carolina, where is both insect and virus pressure How will acylsugar lines with and without virus resistance impact direct and indirect damage due to Thrips/TSWV?

22 Acylsugar Cooperators and Contributors Martha A. Mutschler Lab Post Doctoral Fellow: Brian Leckie Res. Support. Stephen Southwick, & Richard Gaisser Technician Darlene DeJong David Schuster (FSU, Whitefly) Peter Hanson (AVRDC, TY1, TY2, TY3 genes) George Kennedy (Entomology, Thrips/SWV tests) Mike Stevens (BYU, SW7 gene) Jay Scott (FSU, TY3 gene) This work supported in part with funds from Hatch funds, the Vegetable Breeding Institute, Gates Foundation.