Phylogenetic and population genetic analyses strongly support a model of recent sympatric speciation by host shift in the brood parasitic indigobirds (Sorenson et al
). Sympatric indigobirds associated with different hosts are in most cases morphologically distinct and show limited but often statistically significant differentiation at both nuclear and mitochondrial loci, suggesting limited gene flow between species (Sorenson et al. 2003
; Sefc et al. 2005
; ). Here, we use parentage analyses to directly assess the possibility of ongoing gene flow among sympatric indigobird species and host races, testing for imperfect fidelity of female indigobirds in mate choice and/or host choice, both of which appear to be dependent on the host species by which a particular female was raised (Payne et al. 2000
Consistent with strong host specificity by female indigobirds and assortative mating between males and females reared by the same host species (), pairs of close relatives identified in our analyses generally comprised two individuals associated with the same host species. In addition, all 184 males that were recorded and captured in breeding plumage were of the colour expected given their songs; thus, we detected no obvious cases of female indigobirds parasitizing the host of another indigobird species. Similarly, Payne et al. (1993)
found that just 4 out of 494 male indigobirds (0.8%) recorded in southern Africa mimicked the host of another indigobird species.
An alternative scenario yielding male relatives with different songs posits a female indigobird mating with a heterospecific male before laying an egg in a nest of her usual host (i.e. the species that raised her; b). In this case, the male offspring would be a hybrid genetically and might be expected to have intermediate plumage colour, unless differences in colour are controlled by a simple genetic polymorphism, with, for example, blue dominant to green or vice versa. Unfortunately, nothing is known of the genetics of plumage colour in indigobirds, though it does not appear to be a developmentally plastic trait (R. B. Payne 2005, personal communication). Although our analyses identified two PO pairs involving males of different species, the large number of pairwise comparisons completed in our search for close relatives resulted in an appreciable risk of false positives. Relatively low relatedness values for these pairs () and exclusion by additional sequence loci support the conclusion that both pairs were in fact false positives. In addition, if these were legitimate PO pairs, the results would require that each son inherited a dominant colour gene from his mother, as all four males involved had songs consistent with their morphology. Thus, our dataset does not include any strongly supported examples of interbreeding between indigobird species.
An inference of host specificity and assortative mating also applies to the two host races of V. camerunensis, in which first-degree male relatives generally mimicked songs of the same hosts, suggesting that two sympatric and morphologically indistinguishable populations are, for the most part, reproductively isolated due to their association with two different hosts. Our analyses, however, revealed four exceptions to this overall pattern. Two PO pairs and one significant HS pair comprised two blue-plumaged males mimicking the two different hosts of V. camerunensis. The two PO pairs were strongly supported in all of our analyses and were further supported by data from three additional, highly variable loci. Due to the absence of plumage colour differences between the V. camerunensis host races, there is no basis for distinguishing between the alternative scenarios described above. Either laying in an alternate host nest or mating with a male of the other host race could produce a father–son pair in which the two males mimic the songs of the two different hosts associated with this species. Likewise, either mechanism could produce a father–daughter pair in which the female prefers males of the other host race; the fourth significant PO pair involved a female whose host association was indirectly inferred based on capture near a male call site.
While the frequency of related individuals associated with different hosts was much lower than expected if female V. camerunensis
mated indiscriminately or were not generally faithful to their hosts, two to four such pairs in our relatively small sample of close relatives are potentially significant from a population genetic perspective. Imperfect fidelity in mating and/or egg-laying of the order of approximately 5 per cent could lead to a substantial level of ongoing gene flow, sufficient to counteract genetic drift and perhaps also impede adaptive differentiation of the two host races (Wright 1931
; Slatkin 1987
Song playback experiments suggest that V. camerunensis
host races at Tibati are behaviourally isolated (Balakrishnan & Sorenson 2006
); territorial males respond strongly to playbacks of homotypic song, whereas similarly weak responses are elicited by playback of the other V. camerunensis
host race or other indigobird species. Given that random mating by females should select for generalized territoriality by male indigobirds, this result is consistent with premating reproductive isolation among both species and host races. Whether the lack of plumage differences between host races makes mixed matings more likely than between indigobird species is not known, but the central importance of song in indigobird mate choice is clear (Payne et al. 2000
). Even if mating is perfectly assortative with respect to song, however, gene flow might continue via imperfect fidelity in host choice (a
Reproductive isolation arising as a consequence of imprinting on host song is the key element of the indigobird speciation model (Payne et al
). Our evidence of ongoing gene flow between V. camerunensis
host races, however, suggests that complete reproductive isolation may not be an automatic or immediate consequence of host colonization. Rather, following colonization, divergent natural and/or sexual selection on morphology, ecology and/or behaviour may be necessary to complete the speciation process. This is analogous to cases of allopatric speciation in which geographical separation alone is insufficient to produce reproductive isolation (e.g. Rice & Hostert 1993
). An important factor probably influencing the extent of reproductive isolation following a host shift by indigobirds is host nesting ecology. Among the four host species at Tibati, the two associated with V. camerunensis
are the most closely related phylogenetically (Sorenson et al
), show the greatest similarity in nestling morphology (Payne 2005
) and show the greatest overlap in nesting habitat at Tibati (C.N. Balakrishnan 2001–2003, personal observations). Together, these factors may facilitate both egg-laying errors by female indigobirds and subsequent survival of offspring in the nest of a closely related alternate host. Cophylogenetic analyses of brood parasitic finches and their hosts indicate a pattern of ‘clade-limited’ colonization wherein host shifts have most often involved closely related host species (Sorenson et al. 2004
). Thus, the initial colonization of hosts with similar nesting ecology and nestling morphology may be more frequent and/or successful, but the same factors may have the countervailing effect of facilitating ongoing gene flow following colonization.