Genetic variation of Nigerian cattle inferred from maternal and paternal genetic markers

Introduction

The modern domestic cattle were initially domesticated about 10,000 years ago from two putative domestication centers, the Near East for Bos taurus and the Indian Sub-continent for B. indicus (Loftus et al., 1994a). In Africa, the modern domestic cattle were probably introduced at different times with B. taurus circa 7,000–4,000 years BP and B. indicus circa 4,000–2,000 years BP from their putative centers of domestication (Meghen, MacHugh & Bradley, 1994; Freeman et al., 2004). However, the zebu cattle were also reintroduced to the African continent by Arab traders around ~699–640 years AD following the death of the Prophet (Bradley et al., 1998). Penetration of the predominant African taurine to West Africa was amid 4,000 years BP (Marshall & Hildebrand, 2002). The post-introduction of the zebu cattle led to their spread sporadically to West Africa circa 1,400 years ago (Meghen, MacHugh & Bradley, 1994; Hanotte et al., 2002) from East Africa hypothesized to be the original entry point of zebus in Africa (Gifford-Gonzalez & Hanotte, 2011). To date, the origin and migration of ancient and modern African cattle is still under strong debate due to conflicting archaeological and genetic evidence (Hanotte et al., 2000, 2002; Magnavita, 2006), therefore, unravelling of any possible clues is still at large and with very crucial pressure sought-after by scientist and archaeologist.

In the past decade scientist have been using mitochondrial DNA (mtDNA) markers in the control region and the entire mtDNA genome to understand phylogenetic tree models for describing origin, divergence times and domestications of cattle and their population expansion to different parts of the globe (Achilli et al., 2008; Ho et al., 2008; Horsburgh et al., 2013; Olivieri et al., 2015). MtDNA studies on cattle have grouped them in two major lineages represented by haplogroup T for B. taurus (Troy et al., 2001) and haplogroups I1 and I2 (Chen et al., 2010) both defining the genetic lineage of B. indicus. Within the haplogroup T, sister sub-haplogroups T1, T2, T3, T4 and T5 were revealed (Mannen et al., 2004; Achilli et al., 2008, 2009). The haplotypes T2 and T3 are the dominant haplotypes in the Middle East and Europe and also in Africa at very low frequency (Beja-Pereira et al., 2006; Olivieri et al., 2015). Majority of the mtDNA haplotypes in African cattle are defined by T1 haplogroup (Troy et al., 2001; Horsburgh et al., 2013), with no evidence of zebu mtDNA haplotypes been reported so far. There has been some more discoveries of other haplotypes P, Q, R, E and C especially when analysis of whole mtDNA genome was applied (Achilli et al., 2008, 2009; Bonfiglio et al., 2010), with only haplotype Q (Q1) being reported recently in the African cattle (Olivieri et al., 2015).

Genetic studies on paternal Y-chromosomal DNA markers have previously reported the frequency of both haplogroups Y1 and Y2 for taurines and of Y3 haplogroup an exclusive for zebus (Götherström et al., 2005; Bonfiglio et al., 2012a; Ginja et al., 2010). Some authors like Perez-Pardal et al. (2010), Álvarez et al. (2017) and Chen et al. (2018) have reported both Y1 and Y2 haplotype frequencies in sub-Saharan and Mediterranean regions in modern African cattle at considerable varying frequencies. However, majority of the African cattle have been reported to be more of zebu background (Hanotte et al., 2000) solely defined by haplogroup Y3 and are distributed at much higher frequency than Y1 or Y2, enormously across Africa encompassing both the North, West, East, Central and southern African regions (Perez-Pardal et al., 2010, 2018; Álvarez et al., 2017; Ginja et al., 2019).

Nigeria, a country in West Africa harbors both the taurine and zebu cattle (Rege, 1999) and their crossbreds (Loftus et al., 1994a). Apart from their economic benefit such as meat, milk, and skin, the Nigerian cattle are observed as important idols in ceremonial rituals and also utilized in drafting and ploughing during farming. Acquisition of the current knowledge regarding the genetic status of Nigerian cattle is crucial for conservation and utilization of their genetic resources. Previous genetic diversity studies in West African cattle, particularly in Nigeria are limited by the few number of populations (Bradley et al., 1994; Loftus et al., 1994a, 1994b; Bradley et al., 1996; Perez-Pardal et al., 2018). This could imply that the actual extent of the genetic diversity of Nigerian cattle still remains an enigma.

To disclose the genetic diversity in Nigerian cattle, we employed the use of both mtDNA and Y-chromosomal markers, which have been widely used in assessing the diversity and phylogeographic structure of many domestic animals (Lindgren et al., 2004; Ramirez et al., 2009; Wang et al., 2014; Álvarez et al., 2017). In this study, we evaluated variation in the mtDNA D-loop and Y-chromosome using 139 Nigerian cattle samples. Due to the nature of husbandry management in most of African countries, the assessment of genetic variation in Nigerian cattle was conducted based on their sampled locations such as from North West (Zamfara, Kano, Katsina, Kaduna and Sokoto States), North East/central (Taraba and Plateau States) and western (Oyo State) regions of Nigeria (Fig. 1A).

Materials and Methods

Results

Genetic polymorphism of Y-chromosome and haplogroup distribution

In Fig. S3, we provide detailed information on haplogroup distribution based on Y-chromosomal markers in Africa (Perez-Pardal et al., 2018; Ginja, Telo da Gama & Penedo, 2009). The sequences generated in this study have been deposited in the GenBank with accession numbers MW284951–MW284970. Our results show that all Nigerian cattle belong to haplogroup Y3 solely a B. indicus haplogroup due to similar mutations (Table 2; Supplemental Data S1). Notably, in addition to previously reported mutations that further classify zebu cattle into haplotype Y3 families as pointed out by Chen et al. (2018) and Perez-Pardal et al. (2018), we also show new mutations in Nigerian cattle possibly not previously reported. These include those SNPs observed between g.784 and g.805 bp (Table 2; Supplemental Data S1). Multiple sequence alignment with haplotypes defining the three major haplogroups of cattle (Y1, Y2 and Y3) (Nijman et al., 2008; Ginja, Telo da Gama & Penedo, 2009 and Table S2) revealed the existence of seven mutations including mutations A > G, T > C and T > G that distinguish Y3a (or Y_C) and Y3b (or Y3_A, Y3_B) haplotype families (Table 2) grouping Nigerian cattle into zebu Y3b sub-haplotype family (Supplemental Data S1) due to having similar polymorphic information at sites A/A and T/T (Supplemental Data S1). Majority of the Nigerian cattle shared haplotype Hap_7Y3 together with cattle from Asia (Fig. 2) which may signify possible shared origin from the most recent common ancestor (TMRCA). The phylogenetic tree shows that Nigerian cattle cluster separately from European Y1 or Y2 haplogroups (Fig. S4). The phylogenetic tree showed some of the Nigerian cattle clustering together with B. grunniens (yak) and B. bison, these two latter populations are considered distantly related populations to the major lineages of domestic cattle (Fig. S4).

Table 2:

Mutations that describe the distinction of the three major haplogroups of cattle based on ZFY intron 10 gene. The two genomic sites that separate each distinct haplogroup are in bold. The seven mutations observed in Nigerian cattle are italicized.

ZFY intron 10
Haplogroup	Reference	Alternative	Publication
Y1	C	C
	GT	—
Y2	C	C	Götherström et al. (2005)
	GT	GT	Ginja, Telo da Gama & Penedo, 2009
Y3	C	T
	GT	GT
Y3	C	T
	GT	GT
g.797	A	G
g.786	A	T
g.800, 804	A	C	This study
g.784, 803	T	A
g.802, 805	T	G
g.802, 803, 804	T	C
g.801	C	T

DOI: 10.7717/peerj.10607/table-2

The overall genetic diversity for Nigerian cattle measured by the haplotype diversity and the mean number of pairwise difference is 1.000 ± 0.016 and 1.679 ± 1.027 respectively (Table S9). The genetic diversity detected in some African cattle populations/breeds as per Perez-Pardal et al. (2018) and Ginja et al. (2019) although different Y-chromosomal markers such as microsatellite markers were used, the estimates still provide reliable information needed for comparison with our samples (Table S9).

Discussion

In this study we examined the genetic variation across eight Nigerian cattle populations, which represent samples from one country in West Africa. The results based on mtDNA D-loop revealed 80 haplotypes from 119 Nigerian cattle sequences, which showed a haplotype diversity of 0.985 ± 0.005. The haplotype diversity in Nigerian cattle is lower compared to those of Egyptian cattle H = 1.0 ± 0.008 (Olivieri et al., 2015). These findings signify a high level of maternal genetic variation in Nigerian cattle. AMOVA on the other hand suggests a general absence of maternal genetic structuring in Nigerian cattle that may have been as a result of extensive genetic intermixing within the country. Studies show that genetic intermixing is caused by rigorous transportation of domestic animals like goats, cattle, and sheep from place to place as a result of their valuable resource for economic trade or cultural exchange (Tarekegn et al., 2018). Notably, recent study has revealed the impact of African cattle pastoralism in admixture of cattle across the continent, and this admixture or intermixing has been well observed in the African humped cattle (Kim et al., 2020). It is generally known that most of the genetic variation in cattle elucidated by matrilineal genetic information is attributable to geographical differences (Hanotte et al., 2002), which can be sourced at the regional or continental levels (Dadi et al., 2009) rather than by their morphological disparities or differences in their origins (Álvarez et al., 2017). We observed this scenario in our study samples where the variation was explained only within populations from different geographical regions in Africa such as between pairs of Nigerian cattle with other African cattle populations (Table S6). On the other hand, although majority of variation can be explained within a population at individual level, the variation has always been nonsignificant reflecting lack of genetic structuring. This trend of unstructured matrilineal populations within Africa has been reported consistently throughout the entire continent (Álvarez et al., 2017). One of the possible reasons could have been due to the influx of a continual zebu introgression especially following the great rinderpest disease that wiped out nearly 5.2 million head of cattle (Hanotte et al., 2000, 2002). Nonetheless, the hybridization process between zebu and taurine which is practiced throughout the continent, might somewhat explain this unstructured scenario. This is usually attributed by the need of farmers to have a breed of cattle that can have both of the genetic attribute including resistance to disease such as trypanosomiasis and at the same time to be capable to withstand adverse tropical environmental conditions such as drought or hot or humid climatic environments (Kim et al., 2017).

We also observed similar genetic diversity pattern based on Y-chromosome analysis where the haplotype diversity detected was higher compared to that observed in a previous study (Perez-Pardal et al., 2018) for Nigerian cattle possibly due to a single breed population sampled, albeit it was similar to haplotype diversity of cattle from India and Central Asia (Perez-Pardal et al., 2018). Generally, in comparison with other cattle populations in Africa, our study indicated the highest genetic diversity of 1.000 ± 0.016 with the lowest in Zebu_Peul breed population from Burkina Faso (Table S9). Nigerian male cattle samples have been observed with high genetic diversity extrapolated by Y-chromosomal single nucleotide polymorphic marker which is consistent with the scenario of the unique zebu alleles mostly found only in West Africa (Perez-Pardal et al., 2010, 2018). This was hypothesized by these authors that zebus in West Africa have got unique genetic background probably because of genetic contribution from local ancient humped cattle. However, even though recent zebu introgression could have occurred into this region, may have not contributed much to the genetic affinity with other zebus from elsewhere possibly because the introgressions were female mediated (Hanotte et al., 2000) which can also be described in the case of Kuri cattle samples of Lake Chad (Meghen et al., 2000) or perhaps the restocking of African zebu sires from East to West during the early 20th century was relatively low (Álvarez et al., 2017). The high genetic divergence between zebu of West Africa and those from Asia was recently unveiled by Perez-Pardal et al. (2018) which further strengthens this speculative hypothesis.

The present-day zebu-like cattle in West Africa were products of crossbreeding events between the West African taurine cattle such as N’Dama and imported zebu cattle directly from Asia (MacHugh et al., 1997; Kim et al., 2017) or rather indirectly with zebu from East Africa. We computed the genetic relationships among Nigerian cattle by using MJ network (Fig. 1B) and NJ tree (Fig. S1). We observed all Nigerian cattle to be exclusively of taurine T1, an African specific haplogroup, and morphological resemblance of B. indicus such as the presence of the humps. This scenario is in accordance with similar observations by Loftus et al. (1994a) but also a similar case was depicted in zebu breeds of the Americas where they were typically found with mitochondrial taurine haplogroup in their genomes (Ginja et al., 2019). These observations further explain that zebu introgression is typically male-mediated and that this scenario took place in many places around the globe. In the light of this argument with respect to Nigerian cattle in West Africa, most farmers in this region preferred zebu of male lineage due to their massive muscle size but also being resistant to rinderpest that taurines are not, even though them being less vigorous towards trypanosomiasis, a disease prevalent in tsetse regions of both West and Central Africa whereby taurine cattle depict resistance to the disease (Grigson, 1991; Ibeagha-Awemu et al., 2004) The complete absence of Asian zebu mtDNA in Nigerian cattle samples suggests that crossbreeding events were mainly through the imported Asian male zebus (Loftus et al., 1994a, 1994b; Bradley et al., 1996). Nonetheless, this scenario could have been attributed by the continued adoption of trypanotolerant breeds that probably led to total loss of the B. indicus mtDNA lineages or its vulnerability towards population bottleneck. This has also been observed elsewhere by several studies particularly cattle from North-East Asia (Mannen et al., 2004). Bradley et al. (1994) and Perez-Pardal et al. (2018) further stress this observation extrapolating the importation of probably only male zebus into West African region. Other factors such as droughts and the great rinderpest disease outbreak or unbiased selection of cattle breeds over zebu mitochondria may have contributed to the loss of any rare zebu mtDNA (Dadi et al., 2009).

We have also analyzed the Nigerian cattle using a single Y-chromosomal polymorphic marker in the X-degenerate region of the male specific Y-chromosome. Our findings based on mtDNA and Y-chromosome analyses conducted in this study have confirmed that Nigerian cattle are an influence of both taurine and zebu lineages from female and male genetic contributions respectively, with only zebu male specific haplotypes being detected. This illustrates a complex scenario of the genetic background in most of the present-day African cattle populations (Álvarez et al., 2017). Previously, mtDNA studies (Loftus et al., 1994a) found that African cattle are of taurine background. However, earlier studies conducted using Y-chromosomal markers (Bradley et al., 1994) found that African cattle had been genetically introgressed with zebu cattle from Asia. The introgressions were male mediated (Hanotte et al., 2000) and that majority of African cattle belong to the zebu specific haplogroup Y3. Patrilineal studies show that zebu cattle are distributed at varying frequency across the continent with more frequency observed in eastern and Central Africa but at a lower frequency in the western and southern parts of Africa (Fig. S3). This decrease in frequency of zebu haplogroup Y3 as one moves towards West and South of Africa is possibly due to the presence of B. taurus European haplogroups Y1 and Y2 in these regions (Fig. S3). A study by Decker et al. (2014) had previously demonstrated similar observations using autosomal single nucleotide polymorphisms (SNPs) markers. This unevenly distribution of zebu allele and introgression in the African continent is possibly attributed to selection of breeds by farmers based on disease tolerance such as tolerance to trypanosomiasis or ability to withstand adverse environmental conditions such as drought (Hanotte et al., 2000). Although, some of the previous studies conducted in West African cattle detected European Y1/Y2 haplogroups (Perez-Pardal et al., 2010; Ginja et al., 2019), it is surprisingly enough that our study samples from Nigeria did not yield any of these, but rather showed that all Nigerian cattle are of zebu Y3 origin. We believe that, the undetected Y1 or Y2 haplotypes in Nigerian cattle might possibly indicate low or no influence of European cattle in these studied samples, coinciding with what was reported previously by Hanotte et al. (2000) of 0% Y- chromosomal taurine alleles in Nigerian cattle. Nonetheless, it’s probable that our sampling coverage did not cover enough samples from western part of Nigeria where most cattle of haplotype Y1 and Y2 are likely to be found. Similarly, this was shown in phylogenetic tree (Fig. S4), where Nigerian cattle somehow clustered separately from European Y1/Y2 haplotypes. Some studies based on additional Y-chromosomal markers, including STRs, have further indicated that B. indicus haplogroup Y3 is composed of three sub-haplotype families which include Y3_A, Y3_B (also defined as Y3b nomenclature), and Y3_C (or Y3a) (Chen et al., 2018; Perez-Pardal et al., 2018), with Y3_A being a cosmopolitan haplotype, while Y3_B is exclusively found in West African cattle. These three haplotypic zebu families co-existed together with their counterpart taurine in approximately 200,000 years ago (Loftus et al., 1994a; Ho et al., 2008; Murray et al., 2010). However, they diverged at different times among themselves before the start of domestication, with the most recent divergence occurring between West and East African zebus (Perez-Pardal et al., 2018).

Moreover, our study observed that the Nigerian cattle of Y3 haplogroup can be further classified into sub-haplotype family Y3b based on the nomenclature adopted by Chen et al. (2018) (Supplemental Data S1) with several zebu strains (Fig. 2) which possibly reflect less intensive selection in African countries as observed in Nigeria compared to other regions for instance Europe (Xu et al., 2015). Our findings indicate a possibility of gene flow or backcrossing of cattle with wild stock from other related bovine species which may have had occurred probably during initial stages of early domestication in the Eurasia prior to population expansion or at later instances post domestication around 2,000 years ago (Chen et al., 2010; Perez-Pardal et al., 2018). These possible imprints of gene flow or introgression with wild stock or other related domestic Bovidae was inferred through the shared matrilineal mutation motifs in the wild bison (Bison bison), the domestic yak (B. grunniens) and Nigerian cattle. This was clearly confirmed by the patrilineal phylogeny where some of the Nigerian cattle clustered closely with yak, the B. grunniens and B. bison (Fig. S4). Our finding is coherent with a study by Chen et al. (2018), where they were able to detect evidence of gene flow and adaptive introgression between Chinese zebu cattle and the Banteng (B. javanicus). Nonetheless, previous studies have also reported possibility of secondary introgression events that might have occurred from the wild oxen stock into the gene pool of domestic cattle during early domestication (Achilli et al., 2008, 2009). The observed shared wild stock mutations in Nigerian cattle agrees with previous studies (Perez-Pardal et al., 2010, 2018) that the Y3_B, also termed Y3b (by Chen et al. (2018)) zebu lineage in West Africa were probably intermingled with ancient humped cattle which would currently be a significant representative reservoir of male zebu biodiversity. The legacy of the possibility of subsequent introgression from local ancient wild cattle partly explains the existing large divergence between West African zebus and zebus from Asia (Perez-Pardal et al., 2010, 2018). These observations maybe speculative, and could indicate a much more complex scenario of the origin of cattle that possibly involved multiple ancestral domestication populations (Perez-Pardal et al., 2018) especially when considering the discrete contribution of wild ox into the gene pools of the major descendant lineages of cattle (Murray et al., 2010) or perhaps the remnant traces left before the divergence between domestic Bos species and the ancient wild ox had taken place some million years ago (Buntjer et al., 2002).

Conclusions

This study reported the current genetic status and some possible new insights about the origin of cattle in West Africa using Nigerian samples from matrilineal and patrilineal perspectives. High level of maternal and paternal genetic diversity was observed in Nigerian cattle, with lack of phylogeographic structure possibly due to human mediated interventions that usually enhance severe intermixing as a result of improper husbandry management practices. The phylogenetic tree based on patrilineal analysis and the matrilineal haplogroup classification have both provided consensus evidence of a possible introgression and gene flow from wild ancient stock and other related bovine species into Nigerian cattle. We recommend carrying out in-depth population genetic studies using high-throughput technologies on complete mtDNA and autosomal genomes of Nigerian cattle in West Africa as well as the use of other Y-chromosomal markers in order to generate comprehensive genetic information on their adaptive traits, selection and demographic history.

Supplemental Information