Livestock Research for Rural Development 21 (2) 2009 Guide for preparation of papers LRRD News

Citation of this paper

Genetic diversity and relationship of indigenous goats of Sub-saharan Africa using microsatellite DNA markers

E K Muema*,**, J W Wakhungu**, O Hanotte* and Han Jianlin*

*International Livestock Research Institute (ILRI), P.O. Box 30709, Nairobi 00100, Kenya
**Department of Animal Production, University of Nairobi, P.O. Box 29053, Nairobi, Kenya
emuema2003@yahoo.com

Abstract

Sub-Saharan African goats with an estimated population of 180 millions are major asset for farmer communities in a range of agro-ecological zones. This study was undertaken to determine the genetic diversity in and differentiation of relationships among 18 populations of goats from Uganda (4), Tanzania (5), Kenya (2), Mozambique (2), Nigeria (3), Mali (1) and Guinea Bissau (1). Heterozygosity, estimates of FST, genetic diversity and distances were performed using data from 11 microsatellite DNA loci.

 

Expected heterozygosity ranged from 0.450 in Guinea Bissau population to 0.541 in Mbeya population (Tanzania), while the observed heterozygosity ranged from 0.441 in Pafuri population (Mozambique) to 0.560 in Sebei population (Uganda). Mean number of alleles (MNA) per population ranged from 3.82 to 5.91. Gene differentiation (FST) among populations was low (5.3%), a result confirmed by genetic distances (DA).

 

Our results reveal that genetic relationships between populations reflect their geographical proximity rather than morphological classification.

Key Words: DNA markers, gene diversity, selection


Introduction

Africa comprises diverse agricultural environment determined by climate, natural resource and human population density. The economic importance of livestock in African farming systems increases with decreasing rainfall. Livestock production is vital to subsistence and economic development (Winrock International 1992). The ever increasing demand for livestock production to cater for the nutritional needs of rapidly growing human population has led to indiscriminate crossbreeding and replacement of indigenous goats with exotic breeds in an effort to improve productivity. These goats are endowed with unique qualities such as water economy, heat tolerance, disease resistance, mothering and walking abilities, and the ability to efficiently metabolize low quality feeds (Trail and Gregory 1984).

 

If genetic diversity is very low, none of the individuals in a population may have the characteristics needed to cope with the new environmental conditions or challenges. Such a population could be suddenly wiped out. Low amounts of genetic diversity increase the vulnerability of populations to catastrophic events such as disease outbreaks. Low genetic diversity may also indicate high levels of inbreeding with its associated problems of expression of deleterious alleles or loss of over-dominance. Change in the distribution of  the pattern of genetic diversity can destroy local adaptations and break up co-adapted gene complexes. These problems combine to lead to a poorer ‘match’ of the population to its habitat increasing and eventually leading to the probability of population or species extinction.

 

Additionally introgression from other populations, uncontrolled interbreeding among indigenous goat breeds, absence of breed improvement programme and political instability in some countries further endanger them. As a result some indigenous African goat breeds are at a risk of extinction, while others might be losing their unique genetic adaptation to local production systems. It is therefore urgent to understand the genetic diversity of indigenous goats of Africa to implement steps to ensure their conservation and rational utilization for improvement of productivity for the benefit of the farmers.

 

Materials and methods 

Sampling

 

A total of 749 blood samples were randomly collected in 19 populations (Table 1) including one population from Switzerland used as reference population. Genomic DNAs were extracted from the blood samples following the method of Sambrook et al (1989). Eleven microsatellite loci studied were: MAF209, INRA132, BM1818, ILSTS011, INRA063, SRCRSP03, BMS1494, ILSTS044, ILSTS005, ILSTS087 and MAF35.

 

PCR amplification

 

PCR amplification was done on 20 ng template DNA in a 10 μl reaction volume. The 10 μl reaction volume contains 1 μl template DNA, 1 μl 10 X PCR buffer, 0.5 μl dNTPs, 0.1 μl forward primer, 0.1 μl reverse primer, 0.1 μl Taq polymerase and 7.2 μl of double distilled water. Cycling profile included an initial denaturation step at 95oC for 5 minutes, followed by 35 cycles of 30 seconds at 95oC, 1 minute at 55-58oC depending on the primers used and 1 minute at 72oC, and a final extension step at 72oC for 10 minutes using a GeneAmp 9700 (Applied Biosystems) thermal cycler.


Table 1.  Sample information

 

Country

Population/breed

Sample size

1

Guinea Bissau

Gubu/Bissau

46

2

Kenya

Boran Galla

36

3

Kenya

Small East African

39

4

Mali

Maure

37

5

Mozambique

Pafuri

38

6

Mozambique

Landim

37

7

Nigeria

Red Sokoto

41

8

Nigeria

Born White

36

9

Nigeria

West African Dwarf

37

10

Switzerland

Grison Striped

31

11

Tanzania

Maasai

44

12

Tanzania

Small East African (Coat)

41

13

Tanzania

Small East Africa (Mbeya)

38

14

Tanzania

Ugogo

46

15

Tanzania

Ujiji

39

16

Uganda

Karamoja

36

17

Uganda

Teso

49

18

Uganda

Kigezi

38

19

Uganda

Sebei

40


Microsatellite genotyping

 

The resultant PCR products were electrophoresed for 2 hours through a 36 cm, 4.25% polyacrylamide denaturing gel on an ABI PrismTM 377 automated DNA sequencer to separate the alleles. The GenescanTM analysis software version 3.1.2 was used to size the resultant DNA fragments following electrophoresis. The Internal size standard used in this study was the GenescanTM 350-Tamra. The resultant data was imported into the GenotyperTM analysis Software version 2.0. The first step in Genotyper analysis was to check whether the internal size standard DNA fragments were accurately assigned (35, 50, 75, 100, 139, 150, 160, 200, 250, 300, 340 and 350 bp).

 

Results 

Genetic variation

 

A total of 101 alleles were detected at the 11 microsatellite loci for all populations. The highest number of alleles observed was 14 at BMS1494 while the lowest was five at MAF35.  Table 2 shows the observed and expected heterozygosities and the mean number of alleles (MNA) per population averaged across the 11 microsatellite loci. The expected heterozygosity ranged from 0.450 in Guinea Bissau population to 0.541 in Mbeya population. The observed heterozygosity ranged from 0.441 in Pafuri population to 0.559 in Sebei population. The MNA ranged from 3.82 ± 1.89 in Ujiji and Guinea Bissau populations to 5.91 ± 2.88 in Ugogo population.


Table 2.  Expected and observed heterozygosities and MNA with their standard errors within each population studied

Population

Sample size

He ± s.e.

Ho  ± s.e.

MNA

Karamoja

36

0.525 ± 0.060

0.495 ± 0.025

4.64 ± 1.63

Teso

49

0.515 ± 0.061

0.464 ± 0.022

5.09 ± 2.07

Kigezi

38

0.509 ± 0.060

0.506 ± 0.025

4.55 ± 2.11

Sebei

40

0.470 ± 0.064

0.559 ± 0.024

4.18 ± 1.72

Mbeya

38

0.541 ± 0.051

0.530 ± 0.025

5.00 ± 1.84

Coat

41

0.510 ± 0.074

0.465 ± 0.024

5.55 ± 2.07

Ugogo

46

0.527 ± 0.067

0.466 ± 0.022

5.91 ± 2.88

Maasai

44

0.482 ± 0.066

0.527 ± 0.023

4.27 ± 2.20

Ujiji

39

0.473 ± 0.066

0.459 ± 0.024

3.82 ± 1.89

Grison Striped

31

0.460 ± 0.089

0.468 ± 0.027

4.45 ± 2.77

Pafuri

38

0.458 ± 0.071

0.441 ± 0.024

5.00 ± 2.45

Landim

37

0.453 ± 0.072

0.435 ± 0.025

4.18 ± 1.40

Red sokoto

41

0.527 ± 0.062

0.516 ± 0.024

4.82 ± 2.44

Born white

36

0.531 ± 0.064

0.489 ± 0.025

5.09 ± 1.76

Maure

37

0.526 ± 0.061

0.492 ± 0.025

5.18 ± 1.99

West African Dwarf

37

0.506 ± 0.070

0.548 ± 0.025

4.64 ± 2.54

Small East African

39

0.504 ± 0.068

0.477 ± 0.024

4.18 ± 1.60

Boran Galla

36

0.492 ± 0.063

0.485 ± 0.025

4.18 ± 2.23

Guinea Bissau

46

0.450 ± 0.067

0.465 ± 0.022

3.82 ± 1.83


Genetic distances and relationships between populations

 

The highest DS value was observed between West African Dwarf (WAD) and Landim populations (0.111) and the lowest was between Karamoja and Teso populations (0.002) (Table 4). Large differences were observed between populations from different region: West Africa, East Africa and Southern Africa countries, but slight differences were observed between populations from the same country. The reference populations (Grison Striped and Guinea Bissau goats) had DS values significantly different from the rest of the populations. The largest DA distance was observed between WAD, Ujiji and Landim populations (0.095) and the lowest between Karamoja and Teso populations (0.028). The distances between all populations were generally low, but significantly different from the reference populations (Guinea Bissau and Grison Striped goats).

 

Population differentiation

 

Population differentiation was estimated using FST and GST values. The mean genetic differentiation within population (HS) was 0.498 and in the total populations (HT) was 0.524. Both values were lowest at MAF35 (0.146 and 0.149 respectively) and highest at BM1818 (0.727 and 0.748 respectively). The genetic differentiation (GST) ranged from 0.024 at MAF35 to 0.086 at BMS1494. The overall GST values for all markers across all populations was 0.05 indicating that 5% of the total genetic diversity was observed among populations, while 95% was within population.

 

In this study, a relatively low FST (5.3%) indicated that genetic differentiation among populations was limited. The FST estimator of Weir and Cockerham (1984) (5.3%) was close to the Nei’s GST estimator (1978) (5%). They both indicated that the genetic differentiation among the sub-Saharan goat populations is very low. 95% of the total allelic variations account for genetic variation among individuals within populations and only 5% of the total allelic variation account for genetic variation among populations.


Table 3.  Genetic diversity at the 11 microsatellite loci studied

Locus

    HO

    HS

    HT

   GST

FST

ILSTS087

0.276

0.322

0.334

0.036

0.035

ILSTS005

0.143

0.160

0.167

0.037

0.038

ILSTS044

0.502

0.626

0.660

0.051

0.055

ILSTS011

0.667

0.641

0.688

0.068

0.071

INRA132

0.538

0.551

0.565

0.025

0.025

INRA063

0.702

0.632

0.656

0.037

0.039

BM1818

0.648

0.727

0.748

0.028

0.029

BMS1494

0.511

0.613

0.671

0.086

0.095

MAF35

0.125

0.146

0.149

0.024

0.025

MAF209

0.516

0.516

0.561

0.080

0.080

SRCRSP03

0.749

0.543

0.570

0.049

0.051

Overall

0.489

0.498

0.524

0.050

0.053


Phylogenetic analysis

 

The DA NJ tree clustered the population into three major clusters mainly along the geographical locations. The first cluster consisted of 11 populations from East African countries. These populations included Coat, Ugogo, Maasai, Small East African, Boran Galla, Karamoja, Teso, Mbeya, Ujiji, Kigezi and Sebei populations. The second cluster comprised of Pafuri and Landim populations from Mozambique. The third cluster included four populations of WAD, Borno White, Red Sokoto and Maure from West Africa. The reference populations formed another group with the Grison Striped goats been closer to the West African populations compared to Guinea Bissau population.

 

Discussion 

The microsatellite DNA loci have shown a high genetic polymorphism. The sub-Saharan goats had a considerable amount of within population variation based on analysis of molecular variance, heterozygosities and number of alleles. The number of alleles ranged from 5 to 14 which are lower than Ethiopian goats (4 to 23) (Tesfaye 2004), West African Dwarf goats (4 to 21) (Mujibi 2005) and Swiss goat breeds (3 to19) (Saitbekova et al 1999). The relatively low number of heterozygotes indicates excess of homozygotes which could be due to locus under selection, null alleles, inbreeding or presence of population of substructure (Wahlund effect).

 

All populations studied showed significant differentiation and structuring within themselves. The FST and GST values overall populations was 0.053 and 0.05 respectively. This indicates that about 5% of the total genetic diversity was observed among populations and 95% was observed within populations. The total genetic diversity observed between populations was similar to other studies done on African goat populations namely West African Dwarf goats was 5.4% (Mujibi 2005). But populations outside Africa showed slightly higher between population variation [17% among Swiss goats (Saitbekova et al 1999), 11% among Italian goats (Ajamone-Marsan et al 2001) and 10.5% among Chinese goats (Li et al 2002)]. When a population is divided into isolated subpopulations, there is less heterozygosity than there would be if the population was undivided. Founder effects acting on different demes generally lead to subpopulation with allele frequencies that are different from the larger population.

 

The NJ tree classified the populations into three major clusters mainly along the geographical locations. The reference breeds stood out distinctly different from the rest thus suggesting a different ancestry. The information obtained in this study will aid their rational development, utilization and conservation.

 

References 

Ajmone-Marsan P, Negrini R, Crepaldi  P, Milanesi E, Gorni C, Valentini A and Cicogna M 2001 Assessing genetic diversity in Italian goat populations using AFLP® markers. Animal Genetics. Volume 32, Article 5. Retrieved October 2001 from http://www.blackwell-synergy.com/doi/abs/10.1046/j.1365-2052.2001.00789.x

 

Li M H, Zhao S H, Bian C, Wang H S, Wei H, Liu B, Yu M, Fan B, Chen S L, Zhu M. J, Li S J, Xiong T A and Li K 2002. Genetic relationships among twelve Chinese indigenous goat populations based on microsatellite analysis. Genetic Selection and Evolution. Volume 34 Article 6 Retrieved http://www.ncbi.nlm.nih.gov/pubmed/12473236

 

Mujibi N F 2005 Genetic characterization of West African Dwarf (WAD) goats using microsatellite markers. MSc thesis submitted to the Department of Biochemistry and Biotechnology, Kenyatta University, Nairobi, Kenya.

 

Nei M 1978 Estimation of average heterozygosity and genetic distance from a small number of individuals. Genetics. Volume 89, Article 3. http://www.pubmedcentral.nih.gov/picrender.fcgi?artid=1213855&blobtype=pdf

 

Saitbekova N, Gaillard C, Obexer-Ruff G and Dolf G 1999 Genetic diversity in Swiss goat breeds based on microsatellite analysis. Animal Genetics. Volume 30, Article 1 Retrieved February 1999, from http://www.blackwell-synergy.com/doi/abs/10.1046/j.1365-2052.1999.00429.x  

 

Sambrook J, Maniatis T and Fritsch E F 1989 Molecular cloning. A laboratory manual. Cold Spring Harbour Laboratory, New York. pp. 12.

 

Tesfaye A T 2004 Genetic characterization of indigenous goat populations of Ethiopia using microsatellite DNA markers. PhD thesis submitted to the National Dairy Research Institute (Deemed University), Karnal (Haryana), India.

 

Trail C M and Gregory K E 1984 Animal breeding in sub-Saharan Africa towards an integrated program for improving productivity in ‘livestock development in sub-Saharan Africa’. 1st edition. (Simpson J R and Evangolou P: editors). WesternPress, Boulder, Colorado. pp. 19-21.

 

Weir B S and Cockerham C C 1984 Estimating F-statistics for the analysis of population structure. Evolution. Volume 38, Article 6 http://www.jstor.org/sici?sici=0014-3820(198411)38%3A6%3C1358%3AEFFTAO%3E2.0.CO%3B2-0&cookieSet=1 

 

Winrock International 1992 Assessment of animal agriculture in sub-Saharan Africa. Morrilton, Arkansas, USA.



Received 25 June 2007; Accepted 5 April 2008; Published 1 February 2009

Go to top