Lack of phylogeographic structure of Nigerian chicken revealed by mitochondrial DNA

D-loop sequence analysis

A. O. Adebambo£, V. Mobegi*, J. M. Mwacharo*, G. Bjørnstad*§, H. Jianlin* and O.
Hanotte*&
£

Dept. Animal Breeding and Genetics, University of Agriculture, Abeokuta. P.M.B. 2240,

Abeokuta, Nigeria. *International Livestock Research Institute (ILRI), P.O. Box 30709,
Nairobi 00100, Kenya
§

Norwegian School of Veterinary Science, Department of Basic Sciences & Aquatic

Medicine, P.O. Box 8146 Dep., N-0033 Oslo, Norway
&

University of Nottingham, School of Biology, Nottingham NG7 2RD, United Kingdom

Correspondence: Department of Animal Breeding and Genetics, University of Agriculture,
P.M.B. 2240, Abeokuta, Nigeria. E-mail: tumininuadebambo@yahoo.com, Tel: (+234) 80
38239503

Keywords: Commercial chicken; D-loop, genetic relationship; indigenous village chicken

Running title: Genetic structure of Nigerian local chickens

1

2% of the total maternal variation occurs within populations. Thirty-one polymorphic sites which generate 30 haplotypes are identified. Phylogenetic analyses group Nigeria’s local chicken to a single clade and 96. Reference sequences representing the major chicken mtDNA lineages from Asia indicate the Indian sub-continent to be the main source of Nigeria local chickens. 2 .SUMMARY Genetic diversity studies that utilize phenotypic and genetic information are informative when formulating breeding and conservation plans. Lack of phylogeographic structure among Nigerian village chicken indicates extensive genetic intermixing. Our results call for separate breeding management of local and commercial chickens in Nigeria to maintain the genetic uniqueness of the former. Three haplotypes are specific to commercial stocks and 27 to Nigerian local chicken. The present study utilizes sequences of mitochondrial DNA (mtDNA) D-loop region of 172 chicken from Southern and Northern Nigeria to determine the origin and diversity of Nigerian local chickens.

2 Anak Titan (meat type) and 1 Yaffa (egg-type) – the latter two from Israel) found in the country were also included. Four commercial chickens (1 Giriraja sample (Indian dual purpose chicken). Liu et al. Five hundred and ninety two base pairs of the mtDNA D-loop region was amplified (Mobegi et al. 1999). Adebambo et al. In the present study. So far however no such data has been reported on Nigerian local chicken. diversity and origin of Nigerian local chicken. 2007). 1985. the hypervariable segment 1 (HV1) region of the D-loop was sequenced to investigate the phylogeographic structure. Such information if complemented with findings obtained using molecular markers could be useful in formulating long term inference/plans for genetic improvement programs. 33 from the Northeast region (NE). Chickens are the most widely distributed of all livestock species in Nigeria with a population of 166 million birds (FAOSTAT. Quantifying the structure of genetic diversity in different African chicken populations is of significance in optimizing conservation and utilization strategies. The description of Nigerian local chickens is based on phenotypic traits (Nwosu et al. 2005. MATERIALS AND METHODS Genomic DNA was extracted from 168 local unrelated chickens from three geographic regions in Nigeria. The D-loop region of mitochondrial DNA (mtDNA) has been used severally to assess the diversity and phylogeographic structure of various chicken populations (Niu et al. 38 samples from the Northwest region (NW). Desjardins and Morais 3 . Mobegi et al. Chicken play very significant sociocultural and economic roles in most African societies. 2006).INTRODUCTION Evidence confirms that chickens are derived from multiple maternal origins in Asia (Liu et al. 2006). 2002. 2005) using L16750 (5'AGGACTACGGCTTGAAAAGC-3'. 97 from the Southwest region (SW). accession number NC_001323.

All haplotypes have been deposited with the Genbank under accession numbers FJ851656-FJ851686. Two D-loop sequences of wild jungle fowl.1 (Applied Biosystems. 2003) as reverse primer. USA) using two internal primers CR-for (5'-TCTATATTCCACATTTCTC-3') and CR-rev (5'-GCGAGCATAACCAAATGG-3'). accession number AB098668. 1999) was also generated. bankiva (GenBank accession numbers AB007720 and AB007718). were included as outgroups. Tajima's D (Tajima 1989) and Fu's Fs (Fu 1997). Phylogenetic analysis (Figure 2) grouped Nigerian local chickens into a single clade (Clade IV) out of the 4 . including the HV1 region was subsequently used of analysis. Gallus gallus gallus and Gallus g. were calculated using Arlequin (Excoffier et al. Direct sequencing of PCR products was done with the BigDye® Terminator Cycle Sequencing Kit version 3. RESULTS The 168 sequences from local Nigerian and 4 commercial chicken populations generated 30 haplotypes from 31 polymorphic sites. The sequences used were those of the most common haplotype observed in the respective clades as reported by ILRI (2006). A Median Joining (MJ) network constructed using the program NETWORK 4.8 (Bandelt et al. A 397 bp long fragment. Reference sequences that correspond to different clades found in Asian domestic samples spanning the geographic range of wild red junglefowls (ILRI 2006) were also included in the analysis. USA).0 software (Kumar et al. Following 1000 bootstrap replicates.1990) as the forward primer and H547 (5'-ATGTGCCTGACCGAGGAACCAG-3'. Komiyama et al. and are shown in Figure 1. a Neighbour-Joining (NJ) tree was generated using MEGA 3.1. Genetic differentiation based on the analysis of molecular variance and population expansion inferred from mismatch distributions.0. PCR products were sequenced on ABI 3730 XL capillary sequencer (Applied Biosystems. 2004). 2005).

015 ± 0. Analysis of molecular variance show that 96. The mismatch distribution pattern (supplementary material 3) show a bell shaped curve indicative of signatures of population expansion (Kanginakudru et al. IIIc. DISCUSSION Results from the current study indicate that mtDNA (HV1) D-loop region is variable in Nigerian local chickens revealing a total of 30 haplotypes (27 in Nigerian chickens and 3 in commercial breeds). IIIb. One haplotype (Giriraja 57) occurred in Clade IIIC.735) were both significant (P < 0. Estimates of Tajima’s D (-1.188) were not significant. IIId and IV) identified in Asian domestic chicken (ILRI 2006).005) and Harpending’s Raggedness index r (0.05) and negative indicating departure from neutrality and therefore population expansion. Low haplotype (0. Supplementary material 2 shows the geographic distribution of haplotypes in Nigeria. 2008). the 27 haplotypes clustered together in Clade IV. All the mtDNA D-loop sequences observed in Nigerian village chickens 5 . the remaining is found between populations. Haplotype N84 is the most widely distributed and shows a Southwest to Northeast cline in its frequency of occurrence. As in the phylogenetic tree. IIIa.seven clades (Clade I. Positive values shown by SSD and Harpendings’s Raggedness also indicate increasing contribution of new and lowfrequency mutations to population variation typical in expanding populations. and therefore.015) were observed in the populations studied. The values of SSD (0. gave further support to the theory of population expansion among Nigerian local chickens.959) and Fu’s Fs (-11.2% of the maternal genetic variation occurs within populations. Southwest and Northwest populations share haplotypes NW24 and NW4 while Northwest and Northeast populations share haplotype NE36.3925 ± 0.049) and nucleotide diversities (0. II. Supplementary material 1 shows the MJ networks of the sampled Nigerian local chicken populations. The haplotypes observed in commercial chickens were absent in local chickens.

it appears that the Asian maternal ancestor of Nigerian local chicken would have originated initially from the Indian subcontinent as all haplotypes identified in local chicken belong to clade IV which most likely originates from the Indian subcontinent (ILRI 2006). which also predominates the Indian sub-continent (ILRI 2006). suggests a single geographic origin. 6 . most likely as a result of genetic intermixing between populations following human migrations and exchange through trade. (2006) also observed that majority of haplotypes found in West African village chicken populations cluster in clade IV. The D-loop region in Nigerian chicken shows departure from equilibrium (significant negative values for Tajima's D and Fu's Fs) suggesting population expansion. Mobegi et al. These results are consistent with the observed mismatch distribution patterns and non-significant value for Harpending's raggedness index. a population expansion of Nigerian local chicken likely followed their arrival in the country. Therefore.radiate from a single sequence (N84) belonging to Clade IV. Haplotype NE84 shows the widest geographic distribution. The ancestors of the today Nigerian chicken could have reached the country from the North of the African continent and/or could have originated from East Africa through migrations along the Sahelian belt. but with an increasing frequency in the Southwest direction. Low bootstrap values of the NJ tree indicate no clear genetic sub-structuring in Nigerian chicken populations. characterized by an expansion from a single ancestral haplotype (Supplementary material 1). Whatever the case. Our data do not allow us to distinguish between the two but the relative simple network expansion observed. The lack of phylogeographic structure observed among the different populations sampled from different geographic regions suggests population intermixing across large distances within the country. This possibly infers a North-South dispersal of this haplotype.

Bandelt. . Nigeria. Nigerian local chicken populations would have likely been introduced into the Northern parts of the country. 1999. Color variations and performance characteristics of the indigenous chicken of South Western Nigeria. Nairobi. 7 .A... C. Rohl. H. A. J. O. M. Acknowledgment The authors would like to acknowledge the assistance provided by the members of staff in the Biotechnology theme (International Livestock Research Institute.Conclusion Overall. A novel gene order in higher vertebrates. University of Agriculture..J.O. Morais. Ikeobi. Nigerian Journal of Animal Production 26:15-22. The lack of substructure in the chicken populations is evidence of a single maternal origin and extensive genetic intermixing in the past and present times. P. Biol 212:599-634.A. R.. Abeokuta. J Mo. Sequence and gene organization of the chicken mitochondrial genome. the Biotechnology Center and Department of Animal Breeding and Genetics. Mol Biol Evol 16:37-38. Desjardins. REFERENCES Adebambo. 1990.N. Adenowo. Median-joining networks for inferring intraspecific phylogenies. P. Ozoje.O... 1999. Kenya). Forster. Thereafter the chicken populations dispersed towards the southernmost region following human movements and commercial activities.

L. CDROM. Wu. Beja-Pereira. ARLEQUIN ver. The time is now ILRI Annual report 2006. K. R. 1997. FAOSTAT 2007.G. A. Zhang. M. T.. Pp: 108 Kanginakudru. Statistical tests of neutrality of mutations against population growth. L. Brief Bioinform 5:150-163. Metta. Y. Where is the origin of the Japanese gamecocks? Gene 317: 195-202. S. BMC Evol Biol 8:174. MEGA3: Integrated software for Molecular Evolutionary Genetics Analysis and sequence alignment. Schneider. 2003. Baig. Komiyama. Y.S.Excoffier. Safe guarding livestock diversity.. Laval. Multiple maternal origins of chickens: out of the Asian jungle.P..G.. Miao. Yao. G. Y. 2008. J.X. 2006...P.. Nei. Mol Phylogenet Evol 38:12-19.. Y.D. 2006. G.. Palanichamy. Kumar. hitchhiking and background selection.. M. S. 2004.. Tamura. K. Y.. Genetic evidence from Indian red jungle fowl corroborates multiple domestication of modern day chicken. 8 .. S. Ding.. M.. Nagaraju. Ikeo. Gojobori. Food and Agricultural Organization statistical databases.0: an integrated software package for population genetics data analysis. Fu. G. T.W. Liu. Luikart. Evol Bioinform Online 1:47-50. ILRI.. Jakati. 2005. Z. 3. M.. Genetics 147:915-925.

A. 2006. Tajima.Mobegi.. Statistical method for testing the neutral mutation hypothesis by DNA polymorphism. D. Mitochondrial DNA D-loop sequences reveal the genetic diversity of African chicken.. Tanzania Society for Animal Production. Nairobi (Kenya). Yu.. Nigerian Journal Animal Production 12:141-146...A. J. 9 . Proceedings of the 4th all Africa conference on animal agriculture and the 31st annual meeting of the Tanzania Society for Animal Production. All Africa Society for Animal Production. D. Sendalo..O. F.A. Fu. Nwosu. Chicken Diversity Consortium. X..M. Biochem Genet 40:163-174. Akpan. C.V. Y. Dar es Salaam. Nyamu. A. Niu. Dar es Salaam (Tanzania): TSAP. H. J. Onuora. p.. G..P. G.I. 1985. Zhang.). (eds. 293-298. Luo.C. 2002.E. The role of biotechnology in animal agriculture to address poverty in Africa: opportunities and challenges. Genetics 123:585-595. I. In Rege. Gowen. F.P. Y.. F. Obioha. Ruan. Chen.C. Biometrical study of the conformation of the native chicken. The origin and genetic diversity of Chinese native chicken breeds.. 1989...