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Monitoring of genetic diversity in Taiwan conserved chickensassessed by pedigree and molecular data Michèle Tixier-Boicharde, Chih-Feng Chena,f, Yen-Pai Leea a Department of Animal Scien

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Monitoring of genetic diversity in Taiwan conserved chickens

assessed by pedigree and molecular data

Michèle Tixier-Boicharde, Chih-Feng Chena,f, Yen-Pai Leea

a

Department of Animal Science, National Chung-Hsing University, Taichung 40227, Taiwan

b

Faculty of Animal Science, Vietnam National University of Agriculture, Trau Quy Town, Gia Lam District, Ha Noi City, VietNam

c

Key Laboratory of Animal Cell Technology, National Institute of Animal Sciences, Tu Liem, Hanoi, VietNam

d

Institut de Recherche pour le Développement, UMR Diversité, Adaptation et Développement des Plantes (DIADE), Avenue Agropolis, BP 64501, 34394

Montpellier Cedex 5, France

e

INRA/AgroParisTech, UMR1313 Génétique Animale et Biologie Intégrative, Jouy-en-Josas, France

f

Research Center for Integrative and Evolutionary Galliformes Genomics (iEGG), National Chung-Hsing University, Taichung 40227, Taiwan

a r t i c l e i n f o

Article history:

Received 9 August 2015

Received in revised form

27 December 2015

Accepted 29 December 2015

Keywords:

Conservation priorities

Effective population sizes

Inbreeding

Molecular data

Pedigree information

a b s t r a c t

Local chicken breeds face high risks of extinction A conservation program has been set up for eight Taiwan conserved chicken populations (TCP) The research presented here aims at estimating effective population size (Ne) and conservation priorities of TCP populations using pedigree and molecular data Genome diversity was assessed by genotyping 22 microsatellite markers in 45–50 animals per breed Results from the pedigree-based analysis showed that most Nevalues ranged between 50 and 100 except the Shek-Ki breed which exhibited the smallest value (46) so that most breeds could be considered as safe from a conservation point of view The change in inbreeding per generation varied between 0.7% to 1.9% depending on breeds Nevalues estimated from molecular-based analysis were generally lower than those estimated from pedigree-based analysis, suggesting a loss of diversity between the onset of the conservation program (from 1983 to 1995) and the start of pedigree recording in 2002 According to Ne

values, the TCP populations do not appear to be at a high risk, but mating plans by a rotation mating system should be designed in order to limit the increase in inbreeding Regarding the conservation strategy within the TCP, the Shek-Ki and Hua-Tung breeds showed the highest priority for conservation

in terms of genetic risk status and contributions to total diversity across pedigree- and molecular-based approaches In conclusion, this study of TCP populations shows how different types of data can be combined to define conservation priorities considering risk, diversity, or utility of local chicken breeds

& 2016 Published by Elsevier B.V

1 Introduction

Local chicken breeds play an important role in Taiwan due to the

traditional cuisine and culture Local chicken breeds may carry

disease-resistant genes and show high abilities to adapt to

alter-native farming systems, such as organic, which will particularly

improve animal welfare and food safety (Fanatico et al., 2009;Pham

et al., 2012) as well as adaptation to harsh environmental conditions

(Tixier-Boichard et al., 2009) Phenotypic data and pedigree

information have proven to be useful for characterization and management of genetic diversity (Boichard et al., 1997; Tixier-Boi-chard et al., 2009;Lenstra et al., 2012) Unfortunately, phenotypic data and pedigree records of local chickens are rarely documented

in reality (Tixier-Boichard et al., 2009) Therefore, molecular mar-kers are used to monitor the loss of genetic diversity of populations and set priorities for conservation (Boettcher et al., 2010)

FAO (2014)reported that 21.3 percent of chicken breeds in the world were classified as being at risk of extinction, highlighting the importance to assess genetic diversity and the current popu-lation status This percentage might be higher than that because of

a large number of populations with an unknown status in devel-oping countries Basically, there are three strategies for setting priorities in conservation such as the maximum-risk strategy, the maximum-diversity strategy as well as the maximum-utility strategy (Bennewitz et al., 2007)

The maximum-risk strategy is based on the numbers of

Contents lists available atScienceDirect

Livestock Science

http://dx.doi.org/10.1016/j.livsci.2015.12.013

1871-1413/& 2016 Published by Elsevier B.V.

n Corresponding author at: Faculty of Animal Science, Vietnam National

Uni-versity of Agriculture, Trau Quy Town, Gia Lam District, Ha Noi City, VietNam.

E-mail addresses: phammanhhung@vnua.edu.vn (M.-H Pham),

hoantranvcn@yahoo.com (X.-H Tran),

c.berthouly@gmail.com (C Berthouly-Salazar),

michele.boichard@jouy.inra.fr (M Tixier-Boichard),

cfchen@dragon.nchu.edu.tw (C.-F Chen), yplee@mail.nchu.edu.tw (Y.-P Lee).

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breeding animals (FAO, 2000), inbreeding rate (EAAP, 1998;

Meu-wissen, 2009) and recommended effective population size (Ne)

(Meuwissen, 2009) The Nemeasures the number of breeding

in-dividuals in an idealized population in equilibrium that would

show a similar trend in inbreeding as the population under study,

it is one of the most pivotal parameters in both evolutionary

biology and conservation of genetic diversity (Waples and Do,

2010; Goyache et al., 2011; Leroy et al., 2013) This parameter is

considered as one of the major criteria for monitoring risk status

in livestock populations because it accounts for inbreeding and

loss of genetic diversity through random genetic drift (Falconer

and Mackay, 1996;Meuwissen, 2009).Leroy et al (2013)showed

that estimates of effective population size varied according to the

within-breed genetic structure, for different species (i.e cattle,

dog, horse and sheep)

The maximum-diversity strategy defines that a breed is

se-lected for conservation when it contributes significantly to the

overall genetic diversity weighted by both the between- and

within-breed diversity Breeds can be ranked according to their

contribution either to the actual or to the predicted future

di-versity (Bennewitz et al., 2007) For instance,Zanetti et al (2010)

set up conservation priorities forfive Italian local chicken breeds

undergoing in situ conservation using 20 microsatellites In the

case of Vietnamese domestic chickens,Pham et al (2013a)showed

with the highest priorities for conservation according toCaballero

and Toro (2002), andPetit et al (1998) approaches, taking into

account within- and between-breeds components of diversity

When possible, pedigree information and molecular data

should be combined for decision making of conservation priorities

(Zanetti et al., 2010) Pedigree-based and molecular-based

esti-mates of genetic diversity may be more or less correlated

de-pending on the pedigree completeness and the number of markers

(Toro et al., 2006) as illustrated in Iberian pigs with correlations

between pedigree inbreeding and marker homozygosity ranging

from 0.69 with 49 microsatellites (Toro et al., 2002) to 0.92 with

60 K SNPs (Silió et al., 2010) Furthermore, perfect correlations

between approaches cannot be reached because pedigree-based

estimates do not take into account Mendelian sampling, and it is

known that full-sibs would share between 45% and 55% of their

genes rather than exactly 50% in a traditional relationship matrix

(VanRaden and Tooker, 2007) An additional difference between

pedigree- and molecular-based analyses is the definition of the

founder population, which depends on the depth of pedigree for

pedigree-based analysis, the more complete the pedigree, the

more ancient the founder population (Falconer and Mackay, 1996)

Consequently, pedigree- and molecular-based analysis is using

different information, as pedigree-based analysis reflects only

di-versity due to relatively recent ancestry, depending on the

popu-lation history (Toro et al., 2006;Engelsma et al., 2012)

The purposes of this study were (i) to assess genetic diversity

with pedigree-based estimates and molecular-based estimates for

eight populations kept under a conservation program in Taiwan

since 1982, and (ii) to monitor trends in genetic diversity and to

make recommendations for conservation strategy

2 Material and methods

2.1 Data

Conservation of native chickens in Taiwan started from 1982,

when native chickens were collected around the islands and

conserved at National Chung-Hsing University (NCHU)

experi-mental farm Eight Taiwan conserved chicken populations (TCP: B

strain, L2 strain, Hsin-Yi, Hua-Tung, Ju-Chi, Nagoya, Quemoy and

Shek-Ki) have been conserved at NCHU experimental farm since then (Lee, 2006) The L2 and B strains were selected by NCHU from

strains were closed populations since their establishment in 1983, while B strain was a male line and a L2 female line for crossing to produce commercial meat-type chicken Since then, they have been selected for 24 and 26 generations, respectively, and have been extensively used in research as well as in production (Chao and Lee, 2001; Chen et al., 2007; Pham et al., 2013b) A small

man-agement of conserved chicken populations followed a routine procedure (Chao and Lee, 2001) Chicks were raised infloor pens until 16 week of age, when they were transferred to individual wirefloored cages Artificial insemination was individually used for the female chickens with sire known and dam known On the average, the generation interval of TCP populations was one gen-eration per year (Table 2) For the present study, pedigree in-formation recorded between 2002 (starting with ancestors in 2001) and 2008 were used to estimate the effective population size with different methods The pedigree information included a total of 4283 individuals and the numbers of founders at the onset

of pedigree recording are shown inTable 2 In addition, samples from 383 individuals (i.e 288 individuals from six TCP born in

2003 and 95 individuals from B and L2 strains born in 2008) were genotyped and part of the data was previously published ( Ber-thouly et al., 2008;Chang et al., 2012;Pham et al., 2013b) Briefly,

an average of 48 individuals per population was genotyped for 22

Table 1 Pedigree information in the first generation of the eight populations when con-servation program started.

Population First generation

N es , effective population size based on number of sires and dams; and ΔF, rate of hypothetical inbreeding (in percentage) for a population with such an effective population size.

Table 2 Number of founders at the onset of pedigree recording and average number of male and female in the 2002–2008 periods for the eight populations.

Population Founders in 2001 2002–2008 generations

L2 strain 16 328 61.0 0.82 20.8 194.2 75.2 0.67 5

N es , effective population size based on number of founder animals; ΔF, rate of hypothetical inbreeding (in percentage) expected for a population with N es ; N m , average number of male; N f , average number of female; N es , effective population size based on number of breeding animals; and ΔF, rate of hypothetical inbreeding (in percentage) expected for a population with N es across the 2002–2008 genera-M.-H Pham et al / Livestock Science 184 (2016) 85–91

86

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samples were used to estimate the contemporary effective

popu-lation size and contributions to diversity

2.2 Demographic and Pedigree-based analysis

2.2.1 Effective number of founders

The effective number of founders (fe) is the number of equally

contributing founders, which would give the same amount of

genetic diversity that is present in the current population This

was calculated as inLacy (1989), and it is usually much smaller

than the actual number of founders in pedigree (animals with both

parents unknown) because of unequal contributions of founders to

the current population

2.2.2 Effective number of ancestors

The effective number of ancestors (fa) was calculated as in

Boichard et al (1997), explaining the complete genetic diversity of

a population When compared with the effective number of

founders, it provides evidence of bottlenecks that occurred in

population in the past

2.2.3 Effective number of founder genomes

The loss of genetic diversity would occur due to genetic drift in

a small population, even if founders would contribute equally to

this population (Ballou and Lacy, 1995) Therefore, the effective

number of founder genomes or founder genome equivalent (fge) is

defined as the number of equally contributing founders with no

loss of founder alleles that would give the same amount of genetic

diversity as is present in the reference population The fge was

calculated as inCaballero and Toro (2000) It accounts for the loss

of genetic diversity that occurred in the population due to genetic

drift and bottlenecks

2.2.4 Effective number of nonfounders

The effective number of nonfounders (Nenf) was calculated as

Nenf f1 f1 ,

1

nonfounders and for loss of genetic diversity due to drift

accu-mulated over nonfounder generations (Caballero and Toro, 2000)

2.2.5 Measure of the loss of genetic diversity

Measures of the loss of genetic diversity can be derived from fe,

fge, and Nenf The amount of genetic diversity (GD) in the reference

f

1

2ge When expressed as 1GD, it measures the genetic

diversity loss in the population since the founder generation, as a

f

1

2e

1GD*measures the loss of genetic diversity that occurred in the

population due to the unequal contributions of founders before

their contributions converged (Caballero and Toro, 2000) The

difference between GD*and GD isGD* −GD= 2N1

enf, which mea-sures the loss of diversity by genetic drift accumulated over

non-founder generations (Caballero and Toro, 2000)

2.2.6 Effective population size based on the mating plan

The estimation of effective population size based on number of

sires and dams (Nes) followsWright’s (1931)model This method

makes possible to predict Nesunder several assumptions,

includ-ing random matinclud-ing, absence of selection and random variation

of progeny size across parents (Leroy et al., 2013) Computation

+

N N

es

4m f

(ΔF) is inversely proportional to the number of Nes: ΔF =

N

1

2es

2.2.7 Effective population size based on individual inbreeding rate The effective population size based on the individual increase

in inbreeding (Nei) was computed for each population (Gutiérrez

et al., 2008) The individual increase in inbreeding is defined as

ΔFi= –1 gi 1− 1–Fi, where Fi is the inbreeding coefficient for each individual i, and giis the equivalent complete generations ( Gu-tiérrez et al., 2009) The mean of theΔFivalues computed for the n

individuals belonging to a given population of individuals (ΔF ) can

be used to estimate Neiby the method ofCervantes et al (2008)

Δ

N

F

ei 21 2.2.8 Effective population size based on individual coancestry rate The realised effective population size (Nec) was calculated for each population based on the individual increase in coancestry rates (Cervantes et al., 2011) The increase in coancestry between

( + )

gj gk

where cjkis the inbreeding of an offspring from j and k, and gjand

in-dividual j and k, respectively By averaging the increase in coan-cestry for all pairs of individuals in a reference subpopulation, we can estimate an effective population size based on coancestries as

= Δ

1

2 The confidence interval of the estimated values of Nei

and Neccan be computed using the variance ofΔFiandΔcjkfrom the individuals in each population (Gutiérrez et al., 2008) The genealogical information was analyzed using the program Endog v4.8, a computer program for monitoring genetic variability of populations using pedigree information (Gutiérrez and Goyache,

2005)

2.3 Degree of nonrandom mating

correlation of genes within individuals relative to the correlation

of genes taken at random from the population as inCaballero and Toro (2000) This coefficient gives an indication of the deviation from Hardy–Weinberg equilibrium expectations and it is related to inbreeding and coancestry coefficient by (1F)¼(1f)  (1α), where F and f are the inbreeding and coancestry coefficients, re-spectively (Wright, 1969)

2.4 Molecular-based analysis 2.4.1 Microsatellites genotyping and polymorphism The presence of null alleles was tested using FreeNA software (Chapuis and Estoup, 2007) in which loci with estimated fre-quencies of null alleles above 0.2 are considered to be potentially problematic for calculations The null allele frequency estimated for the 22 loci was lower than 0.2 (data not shown) so we assumed that null alleles were absent and used for our analyses The matrix

of Nei's DAgenetic distances (Nei et al., 1983) was computed by Populations package 1.2.32 (Langella, 1999)

2.4.2 Contemporary effective population size Contemporary effective population size (NeLD) was computed from genotypic data by a point estimation method using linkage disequilibrium (Hill, 1981; Waples, 2006) This method was

cor-rects for biases resulting from the presence of a wide range of

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(2006) The NeLD could be calculated for unlinked loci

( × ( – ))

N

r

3

S

2 1 , where r is correlation among alleles and S is

sample size (Hill, 1981;Waples, 2006) All alleles with frequencies

less than the critical values (Pcrit) of 0.05 were excluded (Waples

and Do, 2008) A jackknife method was used to construct 95% CIs

of the estimates

2.5 Contributions to diversity

The contribution of each breed to total genetic diversity was

computed by three approaches: a method based on molecular

coancestry (Caballero and Toro, 2002), a method based on allelic

richness (Petit et al., 1998) and a Weitzman approach modified by

Ollivier and Foulley (2005)

The method described byCaballero and Toro (2002)uses as a

criterion for the maintenance of the maximum overallNei’s (1987)

gene diversity (GD) minimizing the average of molecular kinship

within subpopulations (fs), the average of molecular coancestry

genetic distance between subpopulations (Nei, 1987) Total gene

diversity (GDT) is GDT¼1fm Genetic diversity within

sub-populations is GDW¼1fs Genetic diversity between

subpopula-tions is GDB¼fsfm This approach estimates the genetic diversity

remaining when removing a breed Therefore, a positive value will

indicate that higher diversity is obtained when a breed is not

in-cluded in the dataset

The method described byPetit et al (1998), is using the rarefied

number of alleles per locus, and was applied to assess the

con-tribution of each subpopulation to total allelic richness (CT) in

meta-population The CTincluded the AR of within-subpopulation

diversity (CS) and its divergence from other subpopulations (CD)

and therefore taking into account private alleles In contrast to the

method of Caballero and Toro, this one estimates a contribution to

genetic diversity Therefore, positive value would indicate that

when population is included into the dataset it would increase

genetic diversity Contributions of the breeds to diversity were

computed using Molkin 3.0, a computer program for genetic

analysis of populations using molecular coancestry information

(Gutiérrez et al., 2005)

The method described by Ollivier and Foulley (2005),

con-tributions to between-breeds diversity (CB) were assessed by

using marginal loss of genetic diversity, based on DAgenetic

dis-tances (Nei et al., 1983), following Weitzman approach (Weitzman,

1993) implemented in WEITZPRO (Derban et al., 2002)

Within-breed contributions to diversity (CW) and aggregate diversity (D)

were calculated as suggested byOllivier and Foulley (2005) The D

index was obtained after weighting CB by FSTand CW by 1FSTas

the following formula: D¼FST CBþ(1FST) CW

3 Results and discussion 3.1 Pedigree-based analysis 3.1.1 Inbreeding and effective population size The estimates of hypothetical inbreeding increment per gen-eration are given in Table 2 The highest ΔF was approximately 1.3% found in Shek-Ki and B strain that means that in average 1.3%

of heterozygosity was lost per generation.Table 3showed that the highest increase in inbreeding rates was found for Ju-Chi, Hua-Tung and Shek-Ki populations at the values of 1.34%, 1.76% and 1.88%, respectively, which exceeded the maximum level of one

Additional management such as mating decisions (avoidance of mating between relatives by forming 20 mating pairs) and opti-mum contribution selection (Grundy et al., 1998) should be con-sidered Avoiding mating between close relatives can be easily

mated to a group of sisters/half-sisters, and the son of this male‘X’ will be mated to the daughters of another male‘Y’ which has no

coancestry coefficients and to design the mating plan in a way to minimize the average coancestry of parents by the software Fa-mily sizes should be kept homogenous, with always one son kept per sire family and at least one daughter kept per dam An increase

of generation interval for these breeds could also minimize in-breeding and thus a better conservation of their genetic diversity The estimation of Nevalues highly varied between breeds and

ranged from 38.3 in Shek-Ki to 75.2 in L2 strain, and were always lower than those estimated for 2001, except for B and L2 strains (Table 2) All but the Shek-Ki breed (Nei: 45.9) showed Neivalues within the recommended minimum levels between 50 and 100 to maintain genetic variation for a population in a long term ac-cording toMeuwissen (2009) The average coancestry rates, cor-responding to the expected inbreeding rates in the next genera-tion, varied from 0.77 in L2 strain to 3.14% in Shek-Ki (Table 3) The

Necvalues were approximately equal to one-third of the Neivalues,

as indicated by the Nec/Neiratio, and ranged from 21.0 in Shek-Ki to 35.9 in L2 strain (Table 3;Fig 1) The ratio between Nec and Nei

values was lower than one, which reflected that no substructure (Wahlund effect) is found within eight populations (Cervantes

et al., 2011)

Correlations between Nesand other Nemeasures (Nei, Nec) were positive but not significant (Spearman's rank correlation¼0.51, Table 3

Estimates of effective population sizes for each population obtained by the pedigree and molecular data.

Population Pedigree information a

Linkage disequilibrium b

N

ec ei

an, number of individuals; ΔF , the rates of inbreeding per generation (in percentage); Nei, effective population size based on individual increase in breeding; Δc, the rates

of coancestry per generation (in percentage); N ec , effective population size based on increase in coancestry; and ratioN

N

ec

ei

b

M.-H Pham et al / Livestock Science 184 (2016) 85–91 88

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0.57, P40.05).

3.1.2 Genetic contributions

For the period between 2002 and 2008, the estimated values of

the fe, fa, fge, Nenfand Na50in eight populations are given inTable 4

They were always higher than the real number of founders

re-corded in the early generations between 1983 and 1995 In

Shek-Ki, the estimated values were consistently lower than in the

re-maining seven populations, except for Nenf, which was lower in

Hua-Tung The fe, faand fgevalues for Shek-Ki were 21, 20 and 16,

respectively The numbers of ancestors needed to explain 50% of

the gene pool ranged from 7 in Shek-Ki to 16 in L2 strain These

results indicate that Shek-Ki has a narrow genetic base and

con-tributes less to the within-breed component of genetic diversity

than the remaining populations The effective number of

non-founders was greatly higher than effective number of founder

genomes, which indicates that loss of genetic diversity due to drift

was accumulated over nonfounder generations

The loss of overall genetic diversity observed in Hua-Tung and

Shek-Ki was 5.2% and 6.3%, respectively Thefirst loss ranged from

2.6% in Hua-Tung to 3.1% in Shek-Ki due to bottlenecks and genetic

drift (Fig 2) The second loss accounted for unequal founder

contribution and ranged from 1.7% in Hua-Tung to 2.4% in Shek-Ki

The remaining loss was due to genetic drift only

3.2 Degree of nonrandom mating

The average inbreeding coefficients were smaller than average

coancestries in all eight populations (Table 4) Therefore, the de-gree of nonrandom mating has been negative and mating of highly average-related individuals was successfully avoided The average inbreeding rates were moderate and ranged from 2.1% in Nagoya

to 7.2% in Shek-Ki for the last generation (Table 4) However, these values do not take into account the fact that the founders in 2001 could have been already inbred because each population was closed since several generations In practice, chickens showing a low individual inbreeding should be chosen for mating in order to maintain the recommended effective population sizes and to prevent the risk of extinction in small populations

3.3 Molecular-based analysis 3.3.1 Contemporary effective population size The estimates of NeLDwere lower than the ones of Neigiven by the pedigree-based analysis and that of Nesvalues (Tables 2and3; Fig 1) The correlations between NeLD and Nes was very low (Spearman’s rank correlation¼0.24); but it was significantly

correlation¼0.85, 0.83 and 0.83, respectively, Po0.05) The sam-ple size used for genotyping was included in the CIs of NeLD It is possible that this underestimation is due to the small number of molecular markers used (Waples, 2006; Engelsma et al., 2012) Engelsma et al (2010)showed that marker density was important

in order to assess genetic diversity with heterozygosity estimates but was not so much important when an IBD method was used to assess diversity

The difference between molecular-based estimate of Ne and pedigree-based estimates of Necould also occur if a loss of di-versity took place between the initial founder generation and the first generation included in the pedigree Pedigree-based esti-mates are as good as the pedigree data, which, in the present case, represents a quite recent part of the history of populations Ac-cording toToro et al (2006), some important loci might be highly differentiated because selective forces are strong enough at such loci to overcome the effect of low effective size In order to maintain genetic diversity in a gene bank of Holstein cattle, En-gelsma et al (2012) concluded that high density SNP-based di-versity provided a more detailed knowledge of didi-versity at the scale of chromosomal regions than pedigree-based estimate of diversity, which remains global

3.3.2 Contributions to diversity The conservation priorities were mostly consistent across three approaches we used Contribution to global diversity (GDG) was significantly negatively correlated with total allelic diversity (CT, Spearman’s rank correlation¼ 0.83, Po0.05) and with aggregate diversity (D, Spearman's rank correlation¼ 0.93, Po0.001) The

Fig 1 Effective population sizes for each population obtained by the pedigree and

molecular data N ei , effective population size based on individual increase in

breeding with its standard error bars; N ec , effective population size based on

in-crease in coancestry with its standard error bars; and N eLD , contemporary effective

population size for each population.

Table 4

Genetic contributions of eight Taiwan conserved chicken populations.

Population n f e f a f ge N enf N a50 F f F 2008 7SE

B strain 595 44 21 31.6 112.1 8 0.011 0.017 0.05170.006

L2 strain 1346 87 45 65.5 265.0 16 0.006 0.007 0.02570.003

Hsin-Yi 362 41 38 31.1 128.8 14 0.010 0.017 0.04870.015

Hua-Tung 386 29 27 19.3 57.7 10 0.019 0.026 0.06570.011

Ju-Chi 424 28 27 20.3 73.8 10 0.013 0.025 0.03870.004

Nagoya 428 38 31 30.1 144.8 11 0.008 0.018 0.02170.003

Quemoy 419 31 26 22.8 86.2 10 0.009 0.022 0.03570.004

Shek-Ki 323 21 20 16.0 67.2 7 0.020 0.031 0.07270.013

n, number of individuals; f e , estimates of effective number of founders; f a , effective

number of ancestors; f ge , effective number of founder genomes; N enf , effective

number of nonfounders; N a50 , number of ancestors explaining 50% of the gene

pool; F, inbreeding coefficient for each population in the 2002–2008 periods; f,

coancestry coefficient for each population in the 2002–2008 periods; and F 2008 ,

inbreeding coefficient of the last generation in 2008; SE, standard error.

Fig 2 Genetic diversity (GD) loss in eight populations in the period between 2002 and 2008 GD loss due to bottlenecks and genetic drift (1GD); due to unequal founder contribution (1 GD *

); and genetic drift only (GD * GD).

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D index was positively correlated with the CT (Spearman's rank

correlation¼0.91, Po0.01) Negative correlation is expected with

GDGsince it expresses a loss why other methods express a gain

The results showed that Shek-Ki had the highest contributions

(GDG¼ 2.47%, CT¼5.33% and D¼8.29%) to overall genetic

di-versity (Table 5) Nagoya ranked the second contributions to

di-versity (GDG¼ 1.76% and D¼7.02%), and the fourth of CT¼2.73%

GDWfor conservation, whereas the traditional Ju-Chi breed had

the smallest values of diversity (GDG¼1.21%, CT¼ 1.64% and

D¼ 0.47%) for conservation priorities Overall, the Shek-Ki,

Hua-Tung and Nagoya had high conservation priorities in terms of

genetic diversity However, Shek-Ki and Nagoya exhibited very low

within-breed diversity and it was highly differentiated from other

breeds and appeared as obvious conservation priorities When

analysed for six TCP (i.e not including B and L2 strains), the

Hua-Tung contributed the highest to the aggregate diversity (Berthouly

et al., 2008) Thus, using a different set of breeds seems to change

the relative contribution of each breed, as observed in 24

Vietna-mese domestic chicken populations (Pham et al., 2013a) In fact,

the main contribution of the Hua-Tung was based on its gene

di-versity and thus this breed had a high contribution to the gene

pool

3.4 Potential conservation

We have analyzed genetic diversity for eight Taiwan conserved

chicken breeds based on both pedigree and molecular data The

smallest values of effective population size based on pedigree

in-formation were observed in Shek-Ki and Hua-Tung breeds Thus,

the Shek-Ki and Hua-Tung breeds showed high priorities for

conservation in terms of genetic risk status and contributions to

diversity In spite of a very small number of founders in thefirst

generation, the Hua-Tung breed exhibited the highest contribution

to within-breed diversity It is likely that the initial founders were

very diverse because of the breed history: farmers kept this breed

together with others in the backyard, so that Hua-Tung might have

been crossed with imported larger game birds from Southeast Asia

(Lee, 2006;Berthouly et al., 2008), which have bright black feather

and large body size Quemoy exhibited also a relatively high NeLD

and Neiin spite of a very narrow base population Contrasting with

the history of the Hua-Tung breed, the Quemoy breed is expected

to be a true native since it was essentially isolated from outside the

world between 1949 and around 1990 Although chickens were

also kept in the backyard, the original owners intentionally kept

them as pure as possible (Chia-Juing Won, personal

communica-tion) However, the Quemoy conservation program is the most

recent of all populations, since it started in 1995 Thus, it is likely

that effects of genetic drift have been less important in this breed than in others Therefore, the effective numbers of founder gen-omes of Hua-Tung and Quemoy showed high values, for different reasons

Considering the maximum-utility approach is also needed be-fore making recommendations for conservation Conservation priorities should include the market demand, survivability and productivity related to specific genes under existing management conditions, scocio-economics and the needs for research and de-velopment (Toro et al., 2006;Pham et al., 2013a) In this respect, Shek-Ki has been selected as a sire line to be distributed to farmers producing the Three Yellow breed for meat consumption in Hong Kong (Berthouly et al., 2008) This breed showed the highest body weight at 16 weeks of age (Chang et al., 2012) Selection for a specific trait has resulted in a reduction of genetic variation (Toro

et al., 2011).Chang et al (2012)showed that Hsin-Yi, Shek-Ki and Hua-Tung males exhibited a better heat tolerance due to a lower panting rate than observed in others In addition, Hsin-Yi re-sponded to the highest antibody levels from Infectious Bursal Disease (IBD) vaccine The Quemoy exhibited high antibody re-sponse to low pathogenic avian influenza H6N1 virus, Newcastle Disease and IBD vaccines

4 Conclusions The Shek-Ki breed is of high priority for conservation con-sidering risk and utility, the Hua-Tung breed is of high priority considering risk and diversity, and the Quemoy breed is of high priority considering diversity and utility Then, the Nagoya breed may be of high priority considering between-breeds diversity, and the Hsin-Yi, B and L2 lines could be considered for conservation on the basis of utility only Thus, this study of TCP populations shows how different types of data can be combined to define conserva-tion priorities considering risk, diversity, or utility of local breeds

Conflict of interest statement

No actual or potential conflict of interest in relation to this ar-ticle exists

Acknowledgments

We sincerely appreciate the chicken caretaker at NCHU ex-perimental farm and graduate students for their assistance in pedigree recording We also would like to thank two anonymous

Table 5

Loss or gain of genetic diversity, contributions to allelic richness and aggregate diversity for each of the eight Taiwan conserved chicken breeds.

GD W , change in within-population genetic diversity after removing population i; GD B , change in between-population genetic diversity after removing population i; GD G , change in global diversity after removing population i; C S , a contribution to within-population genetic diversity; C D , a between-population genetic diversity; C T , Total diversity; CW, contribution to within-population diversity; CB, contribution to between-population diversity; and D, aggregate diversity (in percentage).

M.-H Pham et al / Livestock Science 184 (2016) 85–91 90

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reviewers to revise the manuscript with several improvements.

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