Tree Genetics & Genomes (2011) 7:363–372
DOI 10.1007/s11295-010-0337-3

ORIGINAL PAPER

Genetic studies on resistance to Valsa canker
in apple: genetic variance and breeding values estimatedfrom intra- and inter-specific hybrid progeny populations
Kazuyuki Abe & Nobuhiro Kotoda & Hidenori Kato &Jun-ichi Soejima

Received: 8 February 2010 / Revised: 23 August 2010 / Accepted: 1 September 2010 / Published online: 2 October 2010
# Springer-Verlag 2010

Abstract Malus sieboldii Rehd. exhibits high levels of
resistance to Valsa canker caused by Valsa ceratosperma(Tode ex Fr.) Maire while cultivated apples (Malus domestica
Borkh.) are susceptible to the disease. In this study, progenies
from 23 full-sib families derived from both inter- and intraspecific hybridization among 16 Malus genotypes as parents
were assessed for resistance to V. ceratosperma (Vc) for two
seasons using an excised shoot assay to determine the patternof inheritance of the resistance and to also estimate thevariance components, narrow-sense heritability, and breedingvalues of parental genotypes. Generally, M. sieboldii×M.
domestica and its reciprocal crosses had more resistantprogenies to Vc than intra-specific crosses of M. domestica.
Resistance to Vc expressed as the relative lesion length amongprogenies showed continuous variation irrespective of cross,suggesting the quantitative nature of the resistance to the threevirulent isolates of Vc that were tested. Resistance to Vc using
the progeny population was analyzed using a mixed linear
model based on restricted maximum likelihood. The parental
effect (general combining ability (GCA)) was significant
while the interaction effect between parents (specific combining ability (SCA)) was relatively small and non-significant.
The ratio of SCA/GCA variance was about 32%, suggestingthat additive genetic variance had a major contribution to thetotal genetic variance for resistance to Vc. There was apositive correlation (r=0.49, p<0.01) between mid-parental
GCA and SCA predictions among 23 full-sib families for the
resistance. Narrow-sense
heritability
estimated by sib analysis
 2
was moderate b
h ¼ 0:29 . The predicted breeding values(BV) of the 16 parents indicated that M. sieboldii “Sanashi
63” and “Hayanarisanashi 1” would be useful for breeding forhigh levels of resistance to Vc.

Communicated by R. Burdon

Keywords Apple . Breeding value . Heritability . General
combining ability . Resistance . Valsa ceratosperma

K. Abe (*) : N. Kotoda : H. Kato : J.-i. SoejimaApple Breeding and Physiology Research Team,National Institute of Fruit Tree Science,
92-24 Shimo-kuriyagawa,Morioka, Iwate 020-0123, Japane-mail: kazuyuki@affrc.go.jp
Present Address:
N. Kotoda
Fruit Genome Research Team,National Institute of Fruit Tree Science,
485-6 Okitsu-nakacho,
Shimizu, Shizuoka 424-0292, Japan
Present Address:
H. KatoLocal Crop Breeding Research Team,National Agricultural Resarch Center for Hokkaido Region,
1 Hitsujigaoka, Toyohira,Sapporo, Hokkaido 062-8555, Japan

Introduction
Valsa canker, caused by Valsa ceratosperma (Tode ex Fr.)
Maire (syn. Valsa mali Miyabe et Yamada), is one of themost important diseases of apple (Malus) especially ineastern Asia (Sakuma 1990; Uhm and Sohn 1995). The
causal fungus infects trees through wounds at the pruning
ends with infected trunks and shoots appearing in the spring
while the canker develops rapidly in the late spring andearly summer. Infection produces localized cankers followed by death of twigs, limbs, or an entire tree. Due to theability of the mycelium of the causal fungus to thoroughly
invade healthy tissues of bark, phloem, and xylem, and theperennial nature of the canker, spray applications of364

Tree Genetics & Genomes (2011) 7:363–372

fungicides are not successful for the control of Valsa
canker.
Genetic resistance would seem to be a better means tocontrol the canker than preventative sprays. Several
genotypes belonging to Malus sieboldii Rehd. areresistant under field conditions, while all cultivatedapples are susceptible to the disease (Liu et al. 1990).
Since inter-specific hybridizations between cultivatedapples and various wild Malus species, including M.
sieboldii, are known to be successful (Korban 1986),
resistant genotypes of M. sieboldii would appear to be ofmuch value as resistant gene sources for Valsa canker.
However, only a few reports on the variability and genetic
control level of the disease have been published. This isdue to a lack of standardized inoculation and evaluation
methods that are reproducible, low cost and that require
minimal labor.
Knowledge of the genetic basis for resistance to Valsa
canker is important to inform breeding strategy. The
development of an artificial inoculation method to evaluate
the resistance in the laboratory (Bessho et al. 1994; Suzaki
et al. 1997) now allows labor-saving assessment that will
facilitate genetic studies using large datasets. We previously
attempted to obtain genetic information based on inheritance of Valsa canker with this approach and the results
suggested that the resistance in M. sieboldii to bequantitative (Abe et al. 2000). However, due to the small
size of the hybrid population, we were unable to estimate
genetic parameters.
The objective of the present study was to estimate thegenetic variance components necessary to determine themode of inheritance of Valsa canker resistance to inform
breeding strategy development. Furthermore, we intended
to rank parents, especially genotypes belonging to M.
sieboldii in our breeding population, their combining
ability.

Materials and methods
Plant materials
This study was carried out using F1 progenies from theapple breeding program at the Apple Research Center,
National Institute of Fruit Tree Science (NIFTS), Morioka,
Iwate. Sixteen apple cultivars/genotypes were used asparents (Tables 2 and 3). The population for the progeny
test consisted of 23 full-sib families with seven to 75
individuals per family (Table 1). Seventeen families(population 1) came from crosses between genotypes ofM. domestica made for the production of commercial
cultivars and, therefore, had no specific mating design. As
a result, the frequency of appearance of a genotype incrosses varied from 0.04 to 0.17 among parental genotypes.
Six families (population 2) came from inter-specific crosses
between M. domestica and M. sieboldii. Seedling progenies
were planted in 1994 to 1996 in the orchard; trees within afamily were planted side by side at a spacing of 0.5 m
within a row with 3 m between rows. Parental plants weregrown in neighboring fields.
Preparation of twigs for the excised shoot assay
Sixteen parental genotypes and a total of 492 offspring were
inoculated with a virulent isolate VC96-A of V. ceratosperma.
The parental genotypes and a total of 128 offspring derived
from three crosses, “Jonathan”×M. sieboldii “Mo 15,” M.
sieboldii “Hayanarisanashi 1”×“Megumi”, and M. sieboldii“Sanashi 63”×“Jonathan”, were also inoculated with twoother isolates of AVC-12 and AVC-55. “Fuji” was used as asusceptible control. Five dormant 1-year-old shoots fromeach genotype/progeny were collected from trees in thefields at the Apple Research Center (Morioka) in December
or January. After a mild rinse of the shoot surface with

Table 1 Summary of the inoculation test using intra- and inter-specific hybrid populations in this study
Population

1a
2b
2b
Total

Type of crossMalus domestica×M. domestica
Malus sieboldii×M. domestica
M. domestica×M. sieboldii

R resistantaIntra-specific hybrid populationb

Inter-specific hybrid populationc

RL<0.60

No. offamilies

17
3
3
23

No. of progeny used in
1998

1999

80
32
81
193

266
99
85
450

Progeny evaluated(pooled No.)

Percentage of
R progenyc

285
104
103
492

0
24.0
9.7
7.1

Tree Genetics & Genomes (2011) 7:363–372

running water, shoots were dried with a laboratory towel.
One 12-cm segment was then cut from the middle of eachshoot and used in the excised shoot assay. Five twigs per
genotype/progeny were subjected to the inoculation test.

365

estimated according to Durel et al. (1996) as follows:
IDð%Þ ¼ ðFM 0 FM i Þ  100=FM 0 ;ð2Þ

where FM0 is the overall family mean for F = 0 in
population 1, and FMi is the family mean for F=i.

Inoculum, inoculation, incubation, and disease assessment
Statistical analysis
The monoconidial isolates of VC96-A, originating from aninfected apple tree in Iwate Prefecture, Japan, and AVC-12
and AVC-55, originating from an infected apple tree in
Aomori Prefecture, Japan, were used. The isolates were
multiplied for 14 days on a PDA medium at 25°C in
darkness. The preparation of the inoculum and method of
inoculation followed the method developed by Suzaki et al.
(1997) with minor modifications based on Abe et al.
(2007). After a 14-day incubation, fungal mycelium wasadded to a 100 ml volume of sterilized water and thenmildly homogenized for about 30 s. The mixture after
homogenization was used as an inoculum for the excised
shoot assay. The distal cut end of twig segments was
subjected to a scorch treatment with a flat iron to kill thetissue there and to ensure easy invasion by the hyphae of
the inoculum into the twigs. After the scorch treatment, thefive twigs were tied up and placed vertically into a plastic
box so that the proximal cut end of each bundle of twigs
was about 1 cm deep in the water in a plastic box. A dropletof 10 μl of the inoculum was dripped onto the distal end ofeach twig at the scorch site and the plastic box was
immediately covered with a vinyl film to retain thehumidity around the inoculated twigs. The twigs were kept
in darkness at 25°C for 10 days. The lesion length of thenecrosis caused by the inoculum was measured at 10 days
post-inoculation for the excised shoot assay.
Data analysis
Since many families in population 1 have parents that are
more or less genetically related to one another, while someothers are not inbred, the families are different regarding
their inbreeding level. To evaluate the relationship between
the inbreeding coefficient (F) and inbreeding depression(ID) for resistance to Valsa canker, the inbreeding coefficient of each family (FX) in population 1 was calculated
according to Falconer and Mackay (1996) with slight
modification on the assumption that the earliest recorded
ancestors of each family were unrelated using the formula:X½ð1=2Þn ð1 þ FA Þ;
FX ¼
ð1Þ
A

where n is the number of genotypes (cultivars) in any pathof relationship counting the parents of X and the common
ancestor A. FA is the inbreeding coefficient of the common
ancestor A. For the families in population 1, ID was

Differences among genotypes and among isolates of V.
ceratosperma and the interaction between genotypes andisolates were analyzed by a two-way analysis of variance(ANOVA) for lesion length from 16 parental genotypes
inoculated with three virulent isolates of V. ceratosperma.
The following model was fitted to the data:
Yijk ¼ m þ Gi þ Isoj þ ðGIsoÞij þ eijk ;ð3Þ

where Yijk is the lesion length of the ith genotype inoculatedwith the jth isolate; μ is the overall mean; Gi is the effect of
the ith genotype; Isoj is the effect of the jth isolate; GIsoij isthe interaction between the ith genotype and the jth isolate;and eijk is the residual term indicating the within-genotype
deviation resulting from the effects of the kth shootcollected from the ith genotype.
Prior to estimating genetic parameters on the overallprogeny data, the following linear model was fitted to theprogeny data of population 1; this was done primarily totest the significance of the effect of inbreeding levels. A
result of non-significance means that it is possible toestimate genetic parameters without incorporating the Feffect into a model.
 
Yijkl ¼ m þ Yri þ IBj þ Fyk IBj þ IN Dl ðFyk Þ þ eijklm ;ð4Þ

where Yijkl is the lesion length of the lth offspring from an
intra-specific cross between the genotypes of M. domesticain the ith year; μ is the overall mean; Yri is the fixed effectof the ith year; inbreeding (IB)j is the random effect of the
IB level of each family; Fyk(IBj) is the random effects of thekth family nested in the jth IB; INDl(Fyk) is the random
effect of the lth individual tree (offspring) within kth
family; and eijkl is the residual term indicating the withintree deviation resulting from the effects of excised shoots of
the lth offspring.
For population 2, the following linear model was fitted
to the progeny data to test the significance of the effect ofcross-types between M. domestica×M. sieboldii and its
reciprocal:
Yijkl ¼ m þ Y ri þ CT j þ Fyk CT j þ IN Dl ðFyk Þ þ eijklm ;ð5Þ

where Yijkl is the lesion length of the lth offspring from aninter-specific cross between M. domestica and M. sieboldii
or its reciprocal in the ith year; μ is the overall mean; Yri is366Tree Genetics & Genomes (2011) 7:363–372

the fixed effect of the ith year; CTj is the random effect ofcross-type (M. domestica×M. sieboldii (MD×MS) or M.
sieboldii × M. domestica (MS × MD)) of each family;
Fyk(CTj) is the random effects of the kth family nested in
the jth cross-type; INDl(Fyk) is the random effect of the lth
individual tree (offspring) within the family; and eijkl is theresidual term indicating the within-tree deviation of the lth
offspring.
Since the effects of inbreeding level in population 1 and thecross-type in population 2 were non-significant (Table 5), thepooled progeny data was then fitted to the following linear
model to test the significance of the genetic group:
Yijkl ¼ m þ Y ri þ GGj þ Fyk GGj þ INDl ðFyk Þ þ eijklm ;ð6Þ

where Yijkl is the lesion length of the lth progeny from aninter-specific cross between M. domestica and M. sieboldii
or its reciprocal in the ith year; μ is the overall mean; Yri isthe fixed effect of the ith year; GGj is the random effect of
the genetic group (MD×MD or MD×MS and MS×MD) of
each family; Fyk(GGj) is the random effects of the kth family
nested in the jth genetic group; INDl(Fyk) is the random
effect of the lth individual tree (offspring) within the kth
family; and eijkl is the residual term. The statistical analyses
for each population described above were carried out using
SAS PROC GLM.
To estimate the breeding value of the parental genotypes
used in the study, the progeny test data was further
analyzed by fitting the following mixed linear model(Xiang and Li 2001; Iwanami et al. 2008):
Yijkl ¼ m þ Y ri þ Gjm þ Gkf þ Sjk þ eijkl ;ð7Þ

where Yijkl is the lesion length of the lth offspring from across between the jth male parent and the kth female parent
in the ith year; μ is the overall mean; Gj-m and Gk-f are therandom effects of the jth male parent and the kth female
parent, respectively, which are analogous to the parental
effect (i.e., the general combining ability (GCA)) of the jthand kth parents; Sjk is the random effect of the jth and kth
parent interactions, which is analogous to the interaction
effect between parents (i.e., the specific combining ability(SCA)); and eijkl is the residual term indicating the withinfamily deviation resulting from the effects of the lth
offspring of family jk. The estimation of the variancecomponents of the random effect in this model was basedon the restricted maximum likelihood (REML) algorithm in
SAS (SAS Institute, Cary, NC) using the MIXED procedure.
REML is well suited for unbalanced designs to estimate
variance components and can give the variance of male
GCA (σ 2GCA-m) and female GCA (σ 2GCA-f), the variance of
SCA (σSCA2), and the within-family variance (σ 2). Breedingvalue is generally expressed as twice GCA, which is definedwithout distinction between males and females in diallelmating designs. We therefore constructed dummy variables
for each parent using the IML procedures in SAS (Xiang and22
2
Li 2001) so that σ GCA-f =σ GCA-m =σGCA , and then thevariance component for GCA effect was estimated.
The narrow-sense heritability (h2) of lesion length wasestimated using the above results of sib analysis. Because
the covariance of half-sibs was COV(HS) =1/4 VA =σGCA2(where VA =additive genetic variance; Xiang and Li 2001),
narrow-sense heritability was estimated using the following
equation:


b
bA =V
bP ¼ 4sb 2 GCA = 2sb 2 GCA þ s
b2 SCA þ sb 2
h2 ¼ V(where VP =phenotypic variance). Additive genetic varianceestimated in an inbred population, however, is (1+F) VA,
where F is the inbreeding coefficient (Falconer and Mackay
1996; Kumar 2004). Since the F values were different
among the families used in the study, the mean of the Fvalues across families in populations 1 and 2 were
calculated and regarded as an approximate of both
populations. Using the F value, correct heritability on theslightly inbred population was calculated (Kumar 2004).
The standard error of the heritability was estimatedaccording to Falconer and Mackay (1996).

Results and discussion
Relationship between parental genotypes and pathogen
isolates
The mean lesion lengths of parental genotypes in excised
shoot assays caused by the inocula of V. ceratosperma
isolates VC96-A, AVC-12, and AVC-55 had various
values among genotypes across the isolates (Tables 2 and
3). Three genotypes belonging to M. sieboldii had low
values of lesion length, ranging from 5.2 to 7.2 mm in“Sanashi 61”, from 5.8 to 7.8 mm in “Mo 15”,and from
5.4 to 10.8 mm in “Hayanarisanashi 1”. Their values oflesion length (RL) relative to those of “Fuji” ranged from0.25 to 0.56, showing a high level of resistance to V.
ceratosperma. The mean lesion length of the other 13
genotypes belonging to M. domestica had higher values
than those of M. sieboldii, 13.4 to 20.8 mm to VC96-A,
15.4 to 21.4 mm to AVC-12, and 15.2 to 24.0 mm to AVC55. A two-way ANOVA indicated significant differencesamong genotypes (p < 0.001) and among isolates(p=0.038), while no statistically significant interaction
between genotypes and isolates was shown (Tables 2 and
3). The result of ANOVA suggests that screening tests
using parental genotypes and progeny populations for the
resistance to V. ceratosperma will be successful with asingle isolate as inoculum.

Tree Genetics & Genomes (2011) 7:363–372

367

Table 2 Lesion length on excised dormant shoots inoculated with three virulent isolates of Valsa ceratosperma in 16 apple genotypes
Genotype

Species

Resistance levelb

Lesion length (mm) caused by
VC96-A

AVC-12

AVC-55

Fuji
Tsugaru
JonathanGolden Delicious
Akane

Malus domestica Borkh.
M. domestica Borkh.
M. domestica Borkh.
M. domestica Borkh.
M. domestica Borkh.

16.2 (1)a16.4 (1.01)
15.6 (0.96)
15.4 (0.95)
13.4 (0.83)

19.2 (1)20.8 (1.08)
17.6 (0.92)
18.4 (0.96)
16.6 (0.86)

20.8 (1)18.0 (0.87)
20.0 (0.96)
16.6 (0.80)
15.2 (0.73)

S
S
S
S
S

Sansa
Himekami
IwakamiSenshuStarking Delicious
Splendor
Megumi
Alps Otome
Sanashi 63
Mo 15
Hayanarisanashi 1

M. domestica Borkh.
M. domestica Borkh.
M. domestica Borkh.
M. domestica Borkh.
M. domestica Borkh.
M. domestica Borkh.
M. domestica Borkh.
M. domestica Borkh.
Malus sieboldii Rehd.
M. sieboldii Rehd.
M. sieboldii Rehd.

13.8 (0.85)
15.8 (0.98)
16.0 (0.99)
15.6 (0.96)
20.8 (1.28)
16.0 (0.99)
14.6 (0.90)
18.6 (1.15)
7.2 (0.44)
5.8 (0.36)
7.0 (0.43)

16.4 (0.85)
18.6 (0.97)
15.4 (0.80)
16.8 (0.88)
21.4 (1.11)
19.4 (1.01)
16.6 (0.86)
20.8 (1.08)
8.2 (0.43)
6.0 (0.31)
10.8 (0.56)

19.0 (0.91)––
19.8 (0.95)
24.0 (1.15)
18.8 (0.90)
17.2 (0.83)
24.6 (1.18)
5.2 (0.25)
7.8 (0.38)
5.4 (0.26)

S
S
S
S
S
S
S
S
R
R
R

R resistant, MR moderately resistant; S susceptibleaRelative lesion length values (RL value, mean lesion length of the respective genotype/mean length of Fuji) were presented in the parenthesesb

RL<0.60 (R); 0.60≦RL≦0.70 (MR); RL>0.70 (S)Inheritance pattern of resistance
Data on the lesion length of offspring showed a continuous
variation across the families, with the family mean lesion
length around the mean length of parents. The distribution
pattern of the within-family offspring derived from interspecific hybridization indicated the quantitative nature of
the resistance in the genotypes of M. sieboldii tested in the
study (Fig. 1). In crosses in which genotypes of M. sieboldii
were used as parents, the distributions for the lesion lengthof progenies shifted toward lower values than those ofprogenies derived from crosses between genotypes of M.
domestica (Fig. 1, Table 1).
Table 3 Result of analysis of variance for the lesion length on excised
dormant shoots inoculated with three virulent isolates of Valsa
ceratosperma in 16 apple genotypes
Source of variation

df

MS

F

G
I
G×I
Residual

15
2
30
182

331.77140.81
3.38
42.19

7.86**
3.34*
0.08

df degrees of freedom, MS mean square, F F static, G genotype, I
isolate*p<0.05; **p<0.001

Generally, families in population 2 had more offspring
resistant to V. ceratosperma than those in population 1. On
the basis of the RL value of 0.6 as the cut off to separate theresistant (R) and remaining offspring (Abe et al. 2007), R
offspring were obtained with varying frequencies in thefamilies derived from M. sieboldii as a parent; the overallfrequency of R offspring in the families having M. sieboldii
as a parent was 0.17, whereas no R offspring was producedin the families made by the intra-specific crosses of M.
domestica (Table 1).
Relationship between the inbreeding level and resistance
The inbreeding level (IB) did not affect the family meanvalues of the lesion length (Table 4). There was no clear
trend of inbreeding depression for the resistant trait with anincrease of IB from F=0 to 0.25 in population 1. The resultof ANOVA performed for the four levels of the inbreeding
coefficient showed that the effect of IB was non-significant
with family variation (Table 5). The estimated amount ofcontribution to the total variance by the factor IB was 0%(Table 6). The result of ANOVA in population 2 showedthat the effect of cross-type (M. domestica×M. sieboldii or
M. sieboldii×M. domestica) was also statistically nonsignificant, although the estimated percentage of contribution to the total variance by the factor cross-type was 11.5%(Tables 5 and 6).

368

Tree Genetics & Genomes (2011) 7:363–372

Fig. 1 Frequency distribution of lesion length in the F1 progeny
derived from “Jonathan” and Malus sieboldii “Mo 15” (No. ofprogenies= 75) inoculated with a Valsa ceratosperma isolate VC96A; b isolate AVC-12; and c isolate AVC-55, in the F1 progeny
derived from M. sieboldii “Hayanarisanashi 1” and “Megumi” (No.
of progenies=32) inoculated with d isolate VC96-A; e isolate AVC-

12; and f isolate AVC-55, and in the F1 progeny derived from M.
sieboldii “Sanashi 63” and “Jonathan” (No. of progenies = 22)
inoculated with g isolate VC96-A; h isolate AVC-12; and i isolate
AVC-55. P1 and P2 are the lesion lengths of the female and maleparent, respectively

Due to insignificance of the factors by the IB and crosstype in each population, further ANOVA based on both
populations with a pooled data set was performed to test
only the effect of genetic group (intra- versus inter-specifichybrids); the effect was significant at the 5% probability

level (Table 5). The result suggested that the inter-specifichybridization between M. domestica and M. sieboldii or its
reciprocal was more effective to obtain resistant progenies
than the intra-specific hybridization among M. domestica
genotypes. The amount of the contribution to the total
variance by the factor genetic group was about 5.1%
(Table 6).

Table 4 Family mean values of the lesion length for the different F
levels of the families in populations 1 and 2
Population

Inbreeding levela

FM

1

0
0.06
0.13
0.25
0

18.3
18.5
16.9
18.7
13.5

2

FM family mean value of the lesion length (mm)a

Each value represent inbreeding coefficient (F value)

Variance component and heritability estimates by sib
analysis
Further analysis of the variance of two populations
combined for the resistance trait indicated that the parental
effect (i.e., the GCA) was significant (Table 7). As
contrasted with the GCA variance, the interaction effectbetween parents (i.e., the SCA) was relatively small andnon-significant. The ratio of SCA/GCA variance was about
32%, suggesting that additive genetic variance gave a major
contribution to the total genetic variance for the resistance.

Tree Genetics & Genomes (2011) 7:363–372

369

Table 5 Result of ANOVA performed for resistance to Valsa canker using intra- and inter-specific hybrid populations
Source of variation

Population 1

Population 2

Whole population

df

MS

F

df

MS

F

Genetic group
Inbreeding level
Cross-type
FamilyIndividual

–

–
0.83
–
6.57**
1.61**

–
–

13
268

–
489.71–591.19
89.98

1
4
208

–
–
7,375.26
1,299.35
101.54––5.68
12.80**
2.62**

Year
Residual

1
1102

56.47
55.75

1.01

1
1056

19.13
38.79

0.49

3
–

df

MS

F

21
476

6,944.04––
1,034.63
95.03

6.71*
–
–
10.89**
2.00**

1
2159

16.40
47.46

1
–
–

0.35

*p<0.05; **p<0.001

Usually, additive genetic variance is found to be abundantand more important than non-additive genetic variance forpreviously unselected material and in the first few generations of selected materials in tree species (Namkoong etal. 1988), which may also be the case in apple breeding
programs for resistance to V. ceratosperma. Since most fruit
tree breeding programs are based on the premise thatgenetic gains will be cumulative over generations, anadditive genetic effect is a major important source ofvariation to incorporate into breeding and production
programs, while non-additive variance can only originate
from the effects of gene pairs and would be considereddifficult to handle in a tree breeding program (Yanchuk
1996). The presence of additive genetic variance inpopulations of fruit trees for disease-resistance traits is
supported by several quantitative genetic studies, e.g., in
the pathosystems of Podosphaera leucotricha–M. domestica (Bus et al. 2005), Erwinia amylovora–Pyrus communis(Bagnara et al. 1996; Bell and Janick 1977; Quamme et al.
1990), Rubus spp. (Stewart et al. 2005), and Plasmopara
viticola–Vitis spp. (Brown et al. 1999).
2
The narrow-sense heritability ðbh Þ estimated by sib
analysis was moderate, 0.29 (Table 7). For a quantitative

Table 6 Estimates of variancecomponents for resistanceto Valsa canker using
intra- and inter-specifichybrid populations

Variance component

σGG2
σIB2
σCT2
σFy2
σIND2
σe2
Total

disease-resistance trait of fruit trees, it was consistent withthe estimates found for fire blight (Luby et al. 2002) andpowdery mildew (Oraguzie and Currie 2004) in apple, fire
blight in blackberry (Stewart et al. 2005) and Anthracnose
rot in blueberry (Polashock et al. 2005), but it was lower
than those reported for perennial canker in peach (Chang etal. 1991), downy mildew in table grape (Brown et al.
1999), Alternaria late blight in pistachio (Chao et al. 2001)
and scab in pecan (Thompson and Grauke 1994). While thenarrow-sense h2 estimate is not high, the value of 0.3–0.4 isstill favorable and, as Durel et al. (1998) pointed out,
should guarantee the efficiency of mass selection especiallyfor the trait of interest in apple.
Relationship between GCA and SCA values
There was a positive relationship between mid-parental
GCA and SCA predictions among 23 full-sib families forresistance to V. ceratosperma (Fig. 2). The GCA and SCA
correlation was 0.492, and it was significant at the 1%
probability level. This result of a significant positive
correlation between mid-parental GCA and SCA is, to ourknowledge, the first case in a fruit tree species. In forest tree

Population
1 (%)

2 (%)

Whole population (%)–––
7.47 (11.5)
8.21 (12.6)
10.57 (16.3)
38.79 (59.6)
65.04 (100)3.52 (5.1)––
9.01 (13.1)
8.93 (13.0)
47.46 (68.8)
68.93 (100)0 (0)–6.78 (9.8)
6.90 (9.9)
55.75 (80.3)
69.43 (100)370

Tree Genetics & Genomes (2011) 7:363–372

Table 7 Estimated variance components for apple Valsa canker
resistance using 23 families in apple
Variance components
Parental effect (GCA)
Parental interaction effect (SCA)
Within family
SCA/GCA (%)
Narrow-sense heritability

σ 2GCAσ 2SCAσ2h2

Estimates

p

5.26
1.69
56.46
32
0.29±0.097

0.04
0.14
<0.001

species, the same relationship has been reported by Wu andMatheson (2004) for the growth trait in radiata pine.
Determining the genetic cause of such a positive
correlation is a substantial challenge. One possible explanation of the observed correlation, genetic background, andsituation of parental genotypes could be a major contributing factor of the correlation. Linkage disequilibrium (LD)in a population produces a covariance between the additiveand dominance gene effect, as described by Falconer andMackay (1996); LD can generate a correlation between
additive and non-additive gene action. LD can be caused by
selection (Falconer and Mackay 1996), random genetic driftin populations of finite size (Barker 1979), or the mix ofpreviously isolated and genetically different populations (i.
e., from populations with different gene frequencies)(Barker 1979; Falconer and Mackay 1996). Most of theparental genotypes belonging to M. domestica used in our
experiment were selected from numerous apple cultivars/
genotypes for better fruit quality in recent apple breeding
programs. Use of the parental genotypes, given intensive
selection for fruit traits, is suspected to form a tentative LDin parental genotypes for genes associated with resistance toV. ceratosperma as well as for genes controlling fruit traits.

This could be a reason for the positive correlation between
GCA and SCA. The mixture of genetically different
genotypes in a population of parents in our experiment isanother possibility. Parental genotypes of M. sieboldii are
regarded as genetically different from those of M. domestica, a finding that is supported by molecular research on
genetic differentiation among Malus (apple) species (e.g.,
Hokanson et al. 2001) and by phylogenic studies on thegenus Malus Mill. (Forte et al. 2002; Harris et al. 2002;
Robinson et al. 2001). As a result, the use of geneticallyheterogeneous parents and the production of full-sib
families containing several families of inter-specifichybridization origin might be an alternative explanation
for the positive correlation.
Regardless of the genetic cause of such a relationshipbetween GCA and SCA, the positive correlation renders anadvantage in the practical use of SCA variance, i.e.,
selection of the best GCA will capture some portion of
SCA variance. It is of interest for practical breeding topredict how much extra genetic gain by SCA variance canbe captured from selections on the basis of the estimatedbreeding values.
Predicted breeding values of parental genotypes
Parental BVs were predicted using data from full-sib apple
families (Table 8). There was a marked difference among
parental genotypes and, on the basis of the predicted BVs,
Table 8 Estimated breeding values for 16 parental genotypes basedon best linear unbiased predictions (BLUPs)
Genotype

NPV

Mo15

−2.039

1.4

Hayanarisanashi 1
Sanashi 63
Akane
Sansa
MegumiIwakamiGolden Delicious
JonathanSenshuHimekami
Splendor
Fuji
Tsugaru
Alps Otome
Starking Delicious−1.750−1.701−0.208−0.111
0.081
0.081
0.274
0.322
0.322
0.370
0.419
0.467
0.515
1.045
1.575

−6.411
−7.77
2.11
−1.3
−2.4
2.170
−2.04
0.8
3.28
−1.49
3.88
0.417
−0.864−0.21
8.415

ra =0.622**
Fig. 2 The relationship between mid-parental general combining
ability (GCA) and specific combining ability (SCA) predictions among
23 full-sib families for resistance to Valsa ceratosperma

BV

NPV normalized phenotypic value, BV breeding valueaCorrelation coefficient between NPVs and BVs

Tree Genetics & Genomes (2011) 7:363–372

the best parents for transmitting resistance to V. ceratosperma were M. sieboldii “Sanashi 63” (−7.768) and“Hayanarisanashi 1” (−6.411). Among the parents belonging to M. domestica, “Megumi” (−2.402) and “Golden
Delicious” (−2.035) seemed to be good parents, while“Starking Delicious” (8.415), which was regarded as highly
susceptible to V. ceratosperma (Abe et al. 2007; Bessho etal. 1994), was the poorest. Together with the data on thephenotypic values of the parental genotypes, a significant
correlation (r=0.622) was observed between the normalized phenotypic values of parents and the BVs estimated bysib analysis.
Implications for breeding and cultivar development
The main purpose of our breeding program was the creationof new apple cultivars by intra-specific crossing among
typical cultivars, selections, and breeding materials and the
production of various breeding materials for increased
disease resistance by inter-specific crossing between M.
domestica and M. sieboldii for further breeding purposes in
the apple breeding program at NIFTS. The primaryobjective of the sib analysis was to analyze the genetic
variation available in a disease-resistance breeding programof apples at NIFTS, and the results are, for the most part,
specific to this program. However, they indicate the extent
and nature of the genetic variation available in a modern
apple breeding program in which resistance to V. ceratosperma has been introgressed from M. sieboldii, and it isnoteworthy that the methodology is of general relevance toapple breeding. Since many apple breeders worldwide areworking with similar plant materials, such as “Golden
Delicious”, “(Red) Delicious”, “Jonathan”, “Gala”, and“Fuji” (Laurens 1998; Kouassi et al. 2009), the results
maybe generalized to current breeding populations of apple.
The breeding values shown in Table 8 indicate that M.
sieboldii “Sanashi 63” and “Hayanarisanashi 1” were usefulwith respect to transmitting resistance to V. ceratosperma.
The result will justify the use of such genotypes of M.
sieboldii as a donor parent in a long-term apple breeding
program to improve the resistance level to V. ceratosperma.
To date, several accessions of M. sieboldii have been used
to improve some disease-resistance traits in practical apple
breeding programs, as they were found to possess thehighest level of resistance to fire blight (Gardner et al.
1980), powdery mildew (Schuster 2000), and crown gall(Moriya et al. 2010).
The goal of apple breeding for disease resistance is toincrease the resistance level while simultaneously improving or at least maintaining, original fruit quality. Our results
indicate that it would be feasible to develop cultivars withincreased resistance to Valsa canker. The experience withdisease resistance in apple, e.g., apple scab, implies that371

continuous breeding is required (Crosby et al. 1992;
Gessler et al. 2006). Further genetic information on Valsa
canker resistance will facilitate resistance breeding andcultivar development in apple.
Acknowledgements We are grateful to Dr. Koichi Suzaki forproviding the Valsa ceratosperma isolates used in this study. We also
thank Dr. Hiroshi Iwanami for his help with statistical analysis.

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