Discussion
We observed a continent-wide allele-frequency cline with decreasing
frequencies in a northerly direction for both the DE and SAE loci; two
groups of microsatellites that are largely independent of one another
(interlocus distance <29 Mb at only two out of 72 DE-SAE locus
pairs). We could attribute these clines to selection (DE loci: P= 0.083, SAE loci: P = 0.043, all loci: P = 0.013).
Further, we observed an LD cline at the two most closely linked locus
pairs (P = 0.014, other three physically linked locus pairs:P = 0.99), with decreasing interlocus associations between
male-deleterious-trait-associated alleles in a northerly direction. Most
of the interlocus associations at the five physically linked locus pairs
were positive (P = 0.030). Continent-wide positive LD in
combination with an LD cline (combined P = 0.020) is indicative
of genome-wide selection resulting in increased frequencies of
deleterious-allele haplotypes (relative to linkage equilibrium).
Relatively high DE allele frequencies and near-significant positive LD
in East Africa indicate that the male-deleterious alleles are active
across the whole latitudinal range. Apparently, there is continent-wide
and genome-wide selection for male-deleterious traits with selection
strength decreasing from south to north. The only exceptions are the
populations in HiP and Nairobi NP, where male-deleterious alleles appear
to be under negative selection. According to Figure 2, these two
populations seem to have lost many (if not most in Nairobi NP) of their
male-deleterious alleles. The selection pressures in East and southern
Africa must be high to prevent destruction of the allele-frequency
clines and haplotypes by LD decay. These results provide strong support
for earlier reported observations of genome-wide selection in KNP and
HiP (van Hooft et al., 2018; van Hooft et al., 2019; van Hooft et al.,
2014).
Common explanations for allele-frequency clines due to selection assume
the advance of a favourable mutation (in our case male-deleterious
alleles), an environmental gradient, or a combination of the two
(Charlesworth & Charlesworth, 2010; de Jong, Collins, Beldade,
Brakefield, & Zwaan, 2013). The advance of a favourable mutation
assumes South Africa as the place of origin of the male-deleterious
alleles. However, we consider it unlikely that all male-deleterious
alleles originated in a relatively small area. The presence of a linear
environmental gradient seems an unlikely explanation considering the
wide range of habitats and climates in the sampled region that overall
are non-linearly distributed. To our knowledge, there is no linear
disease gradient in this region either. Further, the advance of a
favourable mutation, an environmental gradient or a disease gradient
would not explain the high frequencies of the male-deleterious alleles,
particularly those also deleterious to females (linked to the DE
alleles), which normally are under negative selection (Henn, Botigue,
Bustamante, Clark, & Gravel, 2015). It is also implausible that the
allele-frequency clines are a result of the selective agent being
present only in the most southern part of the range (e.g. South Africa)
considering the occurrence of positive LD in East Africa. LD decay is
expected to be very fast in the absence of selection (6-31% per
generation at the locus pairs analysed in East Africa, considering the
chromosomal distances involved).
Instead of the aforementioned explanations, we argue that the
continent-wide allele-frequency clines are caused by the same sex-ratio
meiotic gene-drive system that earlier has been hypothesized to be the
ultimate selective agent in KNP and HiP, although the male-deleterious
alleles and gene-drive system have been shown to interact with diseases,
such as BTB, and other environmental factors, such as droughts (van
Hooft et al., 2018; van Hooft et al., 2019; van Hooft et al., 2014). We
hypothesize that there is a similar continent-wide frequency cline of
gene-drive alleles, also with decreasing frequencies in a northerly
direction, considering that in KNP DE and SAE allele-frequency clines
co-occurred with an allele-frequency cline of a Y-chromosomal haplotype
linked to a Y-suppressor gene (Y haplotype 557) (van Hooft et al.,
2014). According to this hypothesis, the gene-drive system originated in
southern Africa and subsequently spread north, thereby forming a
selection gradient. Relatively low frequencies of Y haplotype 557 in
other southern African populations (except HiP) support this hypothesis
(frequency in KNP: 0.24, 95% CI: 0.19, 0.30; average frequency in four
other populations: 0.07, 95% CI: 0.02, 0.18 (Smitz et al., 2014; van
Hooft et al., 2010)). The male-deleterious alleles (which occur on the
autosomes) probably originated at various localities throughout Africa,
considering that mutations leading to such alleles can in principle
occur in any population, after which they formed an allele-frequency
cline in response to the aforementioned selection gradient (formed by
the gene-drive system, which occurs on the sex chromosomes).
When the gene-drive system is incomplete, selection may become negative,
resulting in relatively low DE and SAE allele frequencies. This has
earlier has been argued for HiP and may also apply for Nairobi NP (van
Hooft et al., 2019). In both cases, incompleteness of the gene drive
system may be due to strong genetic drift in these small and isolated
populations (HiP: ≤ 75 individuals between 1895 and 1930, Nairobi NP:
current census size ~150; these small sizes probably
contributed to the relatively large pairwise F STvalues with other populations, Figure S1) (Heller, Okello, &
Siegismund, 2010; van Hooft et al., 2019).
Our explanation for the allele-frequency clines implies that frequencies
of male-deleterious alleles and haplotypes on the autosomes are
maintained at specific equilibrium values in each population depending
on the local frequencies of the gene-drive alleles on the sex
chromosomes. This would require some form of balancing selection (which
does not preclude periods of positive or negative selection such as
observed in KNP and HiP, respectively (van Hooft et al., 2018; van Hooft
et al., 2019; van Hooft et al., 2014)). We previously hypothesized that
epigenetic suppression of male-deleterious alleles results in balancing
selection of Y-chromosomal genes in the gene-drive system (van Hooft et
al., 2018). Taking cognizance of an association between epigenetic
suppression and low pre-birth rainfall, we argued that males with many
active male-deleterious alleles should tend to produce offspring with
suppressed male-deleterious alleles and vice versa (males with
few active alleles producing offspring with many active alleles). We
further hypothesize here that the same process also results in balancing
selection of male-deleterious alleles and haplotypes.
Positive LD at linked loci indicates increased frequencies of
deleterious-allele haplotypes relative to linkage equilibrium. We
detected long-distance LD at two pairs of loci, both ~28
Mb apart; these pairs occur on chromosomes 1 and 3 in cattle.
Long-distance LD appears to occur genome-wide considering that
significant LD has also been observed in KNP for theINFNG -GLYCAM1 locus pair, which has an interlocus distance
of 18 Mb and occurs on chromosome 5 in cattle (D ’ = 0.28, i.e. LD
is at 28% of its maximum possible value) (Ihara et al., 2004;
Lane-deGraaf et al., 2015). This is a relatively large distance,
especially considering that high haploid diversity indicates a large
effective population size for the KNP buffalo (34 mitochondrial D-loop
haplotypes with H (gene diversity) = 0.94, 15 Y-chromosomal
haplotypes with H = 0.74; census size ~37,000 in
2010) (Greyling, 2007; van Hooft et al., 2018; van Hooft et al., 2007).
According to population genetic theory, a large effective population
size limits long-distance LD because of increased LD decay with
chromosomal distance (Slatkin, 2008). LD in buffalo populations extends
across much larger chromosomal distances than in other natural mammal
populations that we are aware of for which LD decay has been estimated.
In chimpanzees (Pan troglodytes ) and bonobos (Pan
paniscus ) LD extends to a distance of ~0.15 Mb, in
Arizona wild mice (Mus musculus domesticus ) to a distance of
~0.2 Mb, in Iberian wild boar (Sus scrofa ) to a
distance of ~0.5 Mb, and in gray wolf (Canus
lupus ) and coyote (Canus latrans ) to a distance of
~5 Mb, even in small or bottlenecked populations (except
for the wolf population of Isle Royale, which consisted of just 10-30
individuals) (De Manuel et al., 2016; Gray et al., 2009; Herrero-Medrano
et al., 2013; Laurie et al., 2007; Munoz et al., 2019). Further, the
half-length of LD (the distance at which LD is 50% of its maximal
value) in two isolated Canadian populations of bighorn sheep (Ovis
canadensis ) is only 17%-28% of that in KNP buffalo, despite their
small size of less than 200 individuals each (4.6-7.5 Mb vs. 26.4 Mb,
Text S4) (Miller, Poissant, Kijas, Coltman, & Int Sheep Genomics, 2011;
Miller, Poissant, Malenfant, Hogg, & Coltman, 2015).
LD between distant loci in large outbreeding buffalo populations despite
fast LD decay due to recombination (~20-31% per
generation), indicates strong selection pressures. However, simulation
studies indicate that multilocus selection may lower the minimum
selection pressure necessary for a given level of LD and may slow down
LD decay in a multilocus cline (although even then selection is probably
still relatively strong) (Baird, 1995; Hastings, 1984). Further, allele
frequency differences between male and female gametes due to
sex-specific selection, as hypothesized in KNP based on genetic data
from male and female calves (van Hooft et al., 2018), may have resulted
in admixture LD between distant loci, especially when selection is
strong (Úbeda et al., 2011).
A high number of high-frequency deleterious alleles of seemingly large
effect (van Hooft et al., 2018; van Hooft et al., 2014), with many
co-occurring in haplotypes (relative to what one would expect if no
linkage existed among these alleles), indicates a high genetic load.
However, most populations of African buffalo seem to be stable after
their recovery from the rinderpest pandemic of 1889-1895, which
decimated buffalo populations across the whole of Africa (Bengis, Kock,
& Fischer, 2002; van Hooft et al., 2000). At face value, this seems to
support the view, advocated by some population geneticists, that genetic
load plays a smaller role than one might expect in ecology (Agrawal &
Whitlock, 2012).
We suggest the following five reasons, among others that surely exist,
as non-mutually exclusive explanations why buffalo populations are still
stable.
- Male-deleterious alleles are epigenetically suppressed in a large
fraction of animals (van Hooft et al., 2018; van Hooft et al., 2019).
- Net-deleterious effects on female health are diminished or even
prevented by male-deleterious alleles or haplotypes with negative
phenotypic effects in males but positive phenotypic effects in females
(i.e., associated with SAE microsatellite alleles) (van Hooft et al.,
2014).
- Negative phenotypic effects only become evident in stressful periods,
such as those caused by droughts and disease outbreaks. For example,
in 2001 average body condition was lower in southern KNP, a region
characterized by relatively high frequencies of DE and SAE alleles,
than in northern KNP, but only significantly so at the end of the dry
season (Caron, Cross, & Du Toit, 2003). It may even be that
male-deleterious alleles are mainly expressed during times of high
environmental stress considering that most genetic samples used in the
studies on KNP and HiP were collected during dry seasons (van Hooft et
al., 2018; van Hooft et al., 2019; van Hooft et al., 2014), and then
mainly in animals born during stressful periods because these cohorts
do not experience epigenetic suppression (van Hooft et al., 2018; van
Hooft et al., 2019).
- Selection is mostly soft, where selective death due to
male-deleterious alleles would otherwise be replaced by nonselective
death due to environmental and ecological conditions, e.g. droughts,
diseases and intraspecific resource competition (Agrawal & Whitlock,
2012).
- Interspecific competition is limited, thereby minimizing the
ecological effects of genetic load (Agrawal & Whitlock, 2012). A
recent review has failed to find conclusive evidence for interspecific
competition between African buffalo and other large mammals (Prins,
2016), which may be due to ecological separation (Traill, 2004).
Our analyses and results reveal a continent-wide and genome-wide
distribution of high-frequency male-deleterious alleles in the African
buffalo, with many co-occurring in haplotypes (relative to what one
would expect if no linkage existed among these alleles). Since most
populations appear to be stable, this indicates that, under specific
circumstances, natural populations of mammals can withstand a high
genetic load. Nevertheless, we expect that a high genetic load makes
many buffalo populations vulnerable to environmental stresses such as
droughts and disease outbreaks. Since buffalo play an important role in
the maintenance and transmission of a variety of economically important
livestock diseases (Michel & Bengis, 2012), which may well have been
augmented by a high genetic load, particularly in southern Africa, our
results have relevance to livestock husbandry in areas were cattle graze
in close proximity to buffalo herds.