Within a species there may be distinctive subspecies that have different allele frequencies. The differences are maintained because there is restricted gene (allele) flow between them. The two subspecies may look very different or they may be very similar in appearance.
Genetic exchange between the subspecies is often prevented because the subspecies are geographically separated. This is the first step on the path to allopatric speciation. But genetic exchange can also be restricted by other mechanisms, for example the timing of reproduction, that occurs even if the subspecies inhabit the same environment. This could lead to sympatric speciation.
In either case, the two subspecies will become distinct species—as defined by the biological species concept—when it becomes impossible to form hybrids due to genetic incompatibility. The study of actual speciation events is a hot topic in evolution these days. One of the goals is to identify the genes responsible for preventing the formation of fertile hybrids. The other goal is to identify the mechanism by which the alleles of these genes become fixed in the subspecies. Is it by natural selection or random genetic drift? (Shuker et al. 2005)
One of the best studied examples of speciation in action is due to the work of H.D. Bradshaw and Douglas Schemske at the University of Washington in Seattle, Washington (USA) (Schemske is now at Michigan State University). They studied two species of monkeyflowers that grow near streams and rivers in the mountains and valleys of western North America.
Mimulus lewisii (top) is found primarily at higher elevations (1600 m to 3000 m) while Mimulus cardinalis (right) grows at lower elevations (sea level to 2000 m). Their ranges overlap at moderate elevations in the mountains of California but hybrids are exceedingly rare.
The species differ in a number of characteristics including leaf shape and stem height but the most obvious differences are in the flowers. Mimulus lewsii has pink flowers that are quite open. They attract bumblebees and in the wild 100% of pollinations within this subspecies are by bees. Mimulus cardinalis has red flowers with a more narrow shape. These flowers attract hummingbirds who are responsible for 98% of pollination events in M. cardinalis.
When crossed in a greenhouse, the two species produce fertile hybrids so technically they are not really species but subspecies.
Ramsey et al. (2003) have studied the barriers to gene flow in the wild. Much of it is due to ecogeographic isolation, which is a fancy way of saying that the species don't often come in contact. They grow at different elevations and each species has become adapted to that elevation so that M. lewisii, for example, does not survive well at low elevations and M. cardinalis can't take the cold and the shorter growing season at high elevations.
The fact that the two species have different pollinators is a major factor in preventing gene flow between them. Hummingbirds hardly ever visit M. lewisii and in the overlapping zones there were very few recorded instances of bees visiting flowers from both species. Thus, the opportunities for cross-pollination were effectively zero. What this means is that, "even in sympatry these species are isolated to a large degree by pollinators" (Ramsey et al. 2003).
There are other factors contributing to genetic isolation. The hybrid plants are somewhat less fit and cross-pollination results in fewer seeds than pollination within a (sub)species. The sum of all these factors means that, in the wild, the total reproductive isolation between the two species is 0.9974 to 0.9998. In other words, they don't mix! (But recall that they can readily form fertile hybrids when crossed in the greenhouse.)
In this example, a major component of the restricted gene flow is due to physical separation of the species and that separation is the result of adaptation to different environments. In that sense, the path to speciation is driven, in part, by natural selection. The species are not genetically incompatible so we're not dealing with mutations that prevent hybridization as would be the case if they were true biological species.
Attention has focused on flower color and shape since that determines whether an individual is pollinated by bumblebees or hummingbirds. It's another step toward preventing gene flow between the species. Is it due primarily to selection or drift?
Schemske and Bradshaw (1999) identified a locus, called yellow upper (YUP), that plays a large role in determining flower color in the two species. The locus affects carotenoid distribution in the petals. In M. cardinalis carotenoids are found throughout the petals and the flowers are red. Bees are not attracted to red flowers. The YUP allele in M. lewisii results in less carotenoid and the flowers are pink. These flowers attract bees.
A subsequent study by Bradshaw and Schemske (2003) established that the YUP alleles are directly responsible for much of the pollinator discrimination observed in monkeyflowers. In the second study the authors created near-isogenic lines (NIL) that differed only at the YUP locus.
The normal M. lewisii flower is pink and the petals are in an open shape (a). The normal M. cardinalis flower is red and the shape of the flower is quite different (c). The dominant YUP allele from M. lewisii prevents carotenoid deposition and when it is bred into M. cardialis the flowers are pink (d). The recessive yup allele from M. cardinalis causes more carotenoid to be deposited making the flowers orange in an M. lewisii background (b).
The plants were tested in a natural environment where the ranges of the two species overlapped and both bees and humingbirds were common. Bees preferred the pink flowers whether they were in an M. lewisii background or an M. cardialis background. Conversely, hummingbirds preferred the orange and red flowers in both backgrounds. Thus, the two species have adapted to different pollinators and a large part of this adaptation is due to flower color.
Here's where it gets tricky. Is the switch from bee pollination to hummingbird pollination driven by natural selection? In other words, when the mutation causing red flowers first arose did it confer a fitness advantage on the individuals that came to be pollinated by hummingbirds?
Here's how Bradshaw and Schemske (2003) address this question,
As ‘mutations’ at the YUP locus decrease visitation by the current pollinator guild, and simultaneously increase visitation by a new pollinator guild, are there plausible ecological circumstances in which the mutant might be favoured by natural selection? The combined rate of bumblebee and hummingbird visitation to the yellow-orange-flowered ‘mutants’ of M. lewisii is just 26% of that to the wild-type pink flowers, and the combined rate for dark-pinkflowered ‘mutants’ of M. cardinalis is 95% of the wild type. This implies that a change in the relative abundance of bumblebees and hummingbirds, compared with the pollinator assemblage present during our field experiments, would be required for the mutant to be favoured by natural selection in the common ancestor of M. lewisii and M. cardinalis. The change in relative abundance of pollinators necessary to produce equal visitation to both flower colour phenotypes can be estimated from our data. A ninefold decrease in the relative abundance of bumblebees would produce equal combined visitation rates in the wild-type pink-flowered and ‘mutant’ yellow-orange-flowered M. lewisii NILs. At the equilibrium point, 99% of visitors to wild-type M. lewisii flowers would be bumblebees, whereas 87% of visitors to ‘mutants’ would be hummingbirds. In the M. cardinalis NILs, a twofold increase in the relative abundance of bumblebees would produce equal visitation rates to pink and red flowers. At the equilibrium point, hummingbirds would be virtually the only visitor to the wild-type red M. cardinalis flowers, and remain the major visitor (89% of visits) even to the dark-pink ‘mutants.’In order for the red flower allele to be fixed by natural selection there would have to be a significant decline in the bee population at the time the mutation arose. Presumably, this decline would have only occurred in a small part of the range leading to a subpopulation with red flowers while the main, wild-type, population (pink flowers) continued to be visited by bees.
The authors don't mention the other possibility; namely, that the red flower allele (yup) spread in a subpopulation by random genetic drift. In this scenario, there is no selective advantage to individual plants if they are pollinated by humingbirds. Clearly the evolution of pollinator discrimination by flower color will lead to restricted gene flow between the two species but it is not clear whether this epiphenomenon is due to selection for hummingbird pollination or random genetic drift.
[Photo Credits: Mimulus lewisii or Purple monkey-flower (top) is from flickr. Mimulus cardinalis or Cardinal monkeyflower (second from top) is from the Arizona-Sonore Desert Museum.
Bradshaw, H.D. Jr. and Schemske, D.W. (1999) Allele substitution at a flower colour locus produces a pollinator shift in monkeyflowers. Nature 426:176-178. [doi:10.1038/nature02106] [PDF]
Ramsey, J., Bradshaw, H.D. Jr., Schemske, D.W. (2003) Components of Reproductive Isolation between the Monkeyflowers Mimulus lewisii and M. cardinali (Phrymaceae). Evolution 57:1520-1534. [PDF]
Schemske, D.W. and Bradshaw, H.D. Jr. (2003) Pollinator preference and the evolution of floral traits in monkeyflowers (Mimulus). Proc. Natl. Acad. Sci. (USA) 96:11910-11915. PDF]
Shuker, D.M., Underwood, K., King, T.M., and Butlin, R.K. (2005) Patterns of male sterility in a grasshopper hybrid zone imply accumulation of hybrid incompatibilities without selection. Proc. Biol. Sci. 272:2491-2497. [DOI: 10.1098/rspb.2005.3242]