The American Chestnut Foundation’s Breeding Program for Blight Resistance

Frederick V. Hebard, William Y. C. White & Shawn YarnesMeadowview Research Farms14005 Glenbrook Ave.Meadowview, VA 24361 USAhttp://www.acffarms.org

TACF was created in 1983 as a private, non-profit organization.

Mission: to restore the American chestnut to American forests through a scientific program of breeding and cooperative research.

C x A F1 x A B1 x A B2 x A B3 x B3

B3-F2 x B3-F2

B3-F3

Expected ProportionChinese

1/2

1/4

1/8

1/16

1/16

1/16

Year Cross Made

1930s

1940s

1989-1995

1994-2002

1999-2009(estimate

2005-

Backcrossing• First, American and Chinese chestnut trees are crossed with each other.• The progeny from this first cross are then backcrossed to American chestnut.• Each cycle of backcrossing reduces the fraction of Chinese genes by a factor of one

half.• Finally, backcrosses are intercrossed to yield progeny that will breed true.• Blight resistance is retained by inoculating progeny and selecting resistant ones.

More than one American parent is used at each backcross to

avoid inbreeding. One set of American parents constitutes a

line, the different Americans are denoted A1 to A4 below.

C x A1F1 x A2B1 x A3B2 x A4B3

BackcrossingMore than one American chestnut line also is needed, also to avoid inbreeding, when B3s are intercrossed with each other.

C x A13F14 x A14B14 x A15B24 x A16B34

C x A9F13 x A10B13 x A11B23 x A12B33 x

C x A5F12 x A6B12 x A7B22 x A8B32

C x A1F11 x A2B11 x A3B21 x A4B31 x

B3-F212

B3-F234

Twenty B3 lines (families) eliminates most inbreeding

after intercrossing, enhancing the chance that a population

will not collapse.Effect of Partial Diallel Size on Inbreeding

Chapters add genetic diversity

The effect of adding sets of 20 B3-F2 progeny from our chapters on inbreeding effective population size, for one source of blight resistance. Restoration ecologists estimate that effective population size needs to be 50 to avoid immediate collapse from inbreeding depression and 500 for mutation to offset long-term erosion of genetic diversity by drift. Chapters also add adaptation to their local environment in addition to general genetic diversity.

Number of

Chapters

Inbreeding

Coefficient

Inbreeding Effective

Population Size

1 0.0207 72

2 0.0115 130

3 0.0085 176

4 0.0070 214

5 0.0060 248

Source Number Currently Used

C. mollissima 23

C. crenata 2

C. sativa 1

C. dentata 19

C. henryi & sequinii ?????

Our primary source of blight resistance is Chinese chestnut, because that was identified as the most resistant species by the USDA in the 1930s. No resistance was known in American chestnut until the 1980s. We also have a few crosses with Japanese and European chestnut. C. henryi and C. sequinii also may have individuals with high resistance, based on recent observations in China.

Sources of Blight Resistance

Castaneahenryi

Castaneaseguinii

Number of Trees at Meadowview in 2008

Type of Tree Number of

Nuts orTrees

Sources of Resistance

American Lines

American 1896 69

Chinese 1813 105

F1529 23 69

B1531 15 29

B22052 13 24

B32392 10 73

B4516 4 4

B1-F2471 4 4

B2-F2225 5 5

B3-F232422 3 49

B3-F3464 1 5

F2355 5 5

F36 2 2

Other 5931 73 116

Total 49603

TACF Meadowview Nut Harvest 2008Cross Type Bags Burs Nuts Cbags Cburs CNutsAxA 39 86 12 . . .B1

115 278 197 21 23 0B2

392 731 611 49 44 2B3

340 658 279 39 70 2B4

82 156 69 9 14 0Chapter 2664 7595 2446 290 741 8B1-F2

452 1073 346 112 70 0B1xC 225 422 87 18 32 0B2-F2

223 547 25 18 42 0B2xC 13 20 0 1 1 0CxC 233 566 131 25 58 0F1

298 100 210 22 34 5LSA 1820 3805 2488 193 313 25B1-F3

6128 17271

B2-F35644 12231

B3-F214999 24975

B3-F32421 4092

American 2273 2308

Chinese 3159

Chinkapin 42 50

Total 70987

B3-F2 family size needs to be 9 or 10 selected trees to capture most alleles from each B3. Obtaining fewer selections per parent would result in loss of genetic diversity.

0.00

0.25

0.50

0.75

1.00

0 5 10 15

Family Size

Pro

ba

bil

ity

of

Ca

ptu

rin

g A

ll A

lle

les

Effect of Family Size on the Probability of Capturing Both Alleles from 20 Independently Assorting Loci

B3-F2 Selection

B3-F2 Seed Orchard

Selection for Recurrent Type

• Trichome density and type

• Stipule size and shape

• Leaf shape

• Tooth shape

• Lenticel density, size and color

• Stem color

• Form

• Time of leaf emergence

• Vigor

Each backcross generation will vary in the remaining proportion of Chinese heritage. By picking trees with the fewest Chinese traits (except blight resistance), we accelerate the recovery of American type. At the present time, this selection is done using the following morphological traits:

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are needed to see this picture.

Price Farm viewed from Bryan Farm

Cutting disks of inoculum Inserting disk in treeTaping inocu-lation point

Elder Hostel Inoculation Crew Crew in Action

Selection for resistance We determine the blight resistance of chestnut trees by giving them the disease, inoculating them with the blight fungus, and observing the resultant cankers.

Highly blight-resistant and blight-susceptibleClapper x Graves B1-F2 progeny, characterizedby small and large cankers, respectively.

Table 1. Number of trees in canker size classes in 1993.

Cross Type Size Class (cm)                                   _________             

1.0-2.6 2.6-4.2 4.2-5.8 5.8-7.4 7.4-9.0 9.0-10.6 10.6-

______________________________________________________________________________

American 3 5 2

F1 Nanking 2 4 3

Seedling Chinese 2 7 3

Meiling Chinese 1 2 2

Nanking Chinese 3 2

F2 Mahogany 0 5 23 48 48 29 15

B1-F2 ClapperxGraves 4 25 84 116 112 54 4

______________________________________________________________________________

Table 2. Number of trees in canker size classes in 2003.

Cross Type Size Class (cm)                                      __________           

1.0-2.0 2.0-3.0 3.0-4.0 4.0-5.0 5.0-6.0 6.0-7.0 7.0-8.0

________________________________________________________________________________

American 4 2 2 2 1

F1 Nanking 1 2 3 1

Chinese 3 3 3 6

B2-F2 Clapper 3 11 15 37 16 12 3

B2-F2 Mahogany 3 11 21 31 14 14 1

________________________________________________________________________________

Chestnut Blight Resistance Rating Scheme

Ep155

SG1 2-3

small medium large larger largest

small medium largesmallsmall

StrainofCryphonectriaparasitica

Cankersize

Cankersize

Rating1 2 3 4 5

Number of Clapper B3-F2 seedlings in resistance classes after inoculation

with the blight fungus in 2004 and 2005.

Blight Resistance Class Susceptible Great-

Grandparent

LS Mean Resistance

Rating

Standard Deviation of Resistance

Rating

Number of

Progeny Tested

1 2 3 4 5

LFR4T3 4.0 A 0.8 91 0 3 17 40 31

LFR4T1 4.0 A 0.8 84 0 7 11 27 39

LFR4T14 3.9 A 0.9 145 1 7 30 42 65

LFR4T10 3.8 A 0.8 121 2 7 29 50 33

LFR4T9 3.7 A 1.0 245 3 28 82 69 63

LFR4T12 3.6 A 0.8 35 0 6 14 10 5

Blight-resistant Chinese to American B1-F2, ‘Clapper’ x ‘Graves.’

Obtaining trees like this suggests it should be possible to backcross the blight resistance of Chinese into American chestnut.

The goal.