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Polyandry is often difficult to explain because benefits of the behaviour have proved elusive. In social insects, polyandry increases the genetic diversity of workers within a colony and this has been suggested to improve the resistance of the colony to disease. Here we examine the possible impact of host genetic diversity on parasite evolution by carrying out serial passages of a virulent fungal pathogen through leaf-cutting ant workers of known genotypes. Parasite virulence increased over the nine-generation span of the experiment while spore production decreased. The effect of host relatedness upon virulence appeared limited. However, parasites cycled through more genetically diverse hosts were more likely to go extinct during the experiment and parasites cycled through more genetically similar hosts had greater spore production. These results indicate that host genetic diversity may indeed hinder the ability of parasites to adapt while cycling within social insect colonies.
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- Materials and methods
Understanding the evolution of multiple mating by females (polyandry) is problematic because the behaviour is probably costly and yet benefits have often been hard to establish (Arnqvist & Nilsson, 2000; Jennions & Petrie, 2000; Simmons, 2001). Most social insect species in the order Hymenoptera (ants, bees and waSPS) are more or less monoandrous, but a number of derived genera have evolved high and obligate levels of polyandry (Boomsma & Ratnieks, 1996; Strassmann, 2001). These groups represent interesting cases because some potential material benefits of polyandry are unlikely to apply and because the behaviour is probably particularly costly for the females of most of the species. One leading hypothesis to explain polyandry in social insects is that it results in a more genetically diverse worker population and thereby improves the colony's resistance to parasites (Hamilton, 1987; Sherman et al., 1988).
There are a number of ways by which having a genetically diverse worker population may improve a colony's resistance to disease. The underlying assumption is that there is genetic variation for resistance to disease, which has been demonstrated in several social insect species (Baer & Schmid-Hempel, 2003; Palmer & Oldroyd, 2003; Hughes & Boomsma, 2004), and is well established in many other animals (Ebert et al., 1998; Little & Ebert, 1999, 2000, 2001; Carius et al., 2001). Variation in resistance may relate either to preventing an infection and/or to reducing within-host parasite growth. Several studies have produced evidence that directly support the hypothesis that genetically diverse colonies may be more resistant to disease (Baer & Schmid-Hempel, 1999, 2001; Hughes & Boomsma, 2004), or may be less prone to extreme rates of infection (Tarpy, 2003). However, to date, none have considered how genetically diverse worker populations impact upon the evolution of a parasite. Social insect colonies can contain thousands or even millions of individuals and can survive for many years. Most parasites, particularly the viruses, bacteria and fungi, have extremely short generation times. Social insect colonies therefore provide the potential, both in terms of number of host individuals and time, for parasites to pass many generations within the worker population. This carries with it the potential for the parasite to evolve whilst cycling within the colony.
Parasites will normally encounter a range of host genotypes. Assuming these differ in their resistance then a particular parasite genotype will be more or less adapted to the different host genotypes. Consequently, parasite virulence (here defined as the negative effect upon host fitness and including both the probability of infection and the impact of a successful infection) will vary depending upon the specific interaction between host and parasite genotypes. Parasites will evolve towards optimal (for the parasite) levels of virulence for the host genotypes that they interact with. Obtaining optimal virulence will be constrained by having to interact with a range of different host genotypes with different resistance properties. If a parasite is presented with a host population of low genetic diversity, then it will be easier for it to adapt to the restricted range of host genotypes in the population. This generally results in the parasite evolving heightened virulence and transmission characters (Ebert & Hamilton, 1996). The classic example is the evolution of increased virulence in parasites that exploit the monocultures that characterize modern agriculture (Brown, 1994). Very low genetic diversity is something that also characterizes the colonies of most social insects and they therefore represent prime targets for parasites to exploit (Schmid-Hempel, 1998; Boomsma et al., 2005). If colony populations are of higher genetic diversity then this will hinder the evolution of the parasite and may thus reduce the harm that the parasite causes to the colony over successive parasite generations.
Serial passage experiments provide excellent ways of examining how parasites adapt over successive generations. They involve parasites being transferred from one host to another, with the characters of the derived strain then being compared with those of the ancestral strain (Ebert, 2000). This most commonly results in an increase in parasite virulence, sometimes within only a few generations (Ebert, 1998, 2000). The increase in virulence has been suggested to be due to one or more of three reasons (Ebert, 1998, 2000). (1) That the hosts used in serial passage experiments are often genetically similar and therefore easier for the parasite to adapt to than the genetically diverse hosts found in most natural populations. (2) That parasite virulence is normally balanced by the cost of prematurely ending parasite growth by killing the host too quickly, and that this cost of high virulence does not apply to parasites in serial passage experiments because transmission is artificially ensured as part of the experiment. (3) That the artificial transmission that characterizes some serial passage experiments reduces the need for transmission stages and this allows parasites to divert more resources to within-host growth. In a passage experiment using the protozoan parasite Crithidia bombi, an increase in virulence was found to occur in colonies of the monoandrous bumblebee Bombus terrestris (Schmid-Hempel, 2001). After just three transfers between full-sibling host individuals, the post-selection parasite reduced host condition to a greater degree than did the ancestral strain.
We examined the impact of host genetic diversity on parasite evolution using serial passage experiments with the leaf-cutting ant host Acromyrmex echinatior Forel (Hymenoptera: Formicidae: Attini) and the fungal parasite Metarhizium anisopliae var. anisopliae (Metschnikoff) (Deuteromycotina: Hyphomycetes). A. echinatior is highly polyandrous, with queens typically mating with around ten males (Sumner et al., 2004). M. anisopliae var. anisopliae is a virulent, generalist entomopathogen that infects leaf-cutting ants (Alves & Sosa-Gómez, 1983; Humber, 1992; Jaccoud et al., 1999; Hughes et al., 2002; Poulsen et al., 2002; Hughes et al., 2004a,b), as well as many other insects. It is a semelparous, ‘obligate killer’ (Ebert & Weisser, 1997), producing transmission propagules (spores) shortly after host death. Virulence is expressed during within-host vegetative growth by the direct invasion of host tissues by hyphae, the diversion of host resources to parasite growth and the production of compounds that inhibit the immune response but which are also toxic to the host (Boucias & Pendland, 1998). The within-host interaction of Metarhizium parasites is apparently impervious to relatedness, and, unlike certain other parasites (West & Buckling, 2003; Griffin et al., 2004), is characterized by scramble competition rather than cooperation even when within-host relatedness is high (Hughes et al., 2004b). Kermarrec et al. (1986) report that a strain of M. anisopliae had increased virulence after only 10 passages through leaf-cutting ants of unspecified relatedness.
We carried out two experiments. The first consisted of six treatments (Fig. 1). Four of these treatments (1a, b, c and d) involved serial passages of a single parasite strain through ants of different levels of genetic diversity. The two main predictions were that the parasite should evolve greater virulence over the course of the experiment, as seen in other serial passage experiments (Ebert, 1998, 2000), and that virulence and/or spore production at the end of the experiment should be negatively correlated with the genetic diversity of the host ants through which the parasite had been passaged. Parasites passaged through genetically similar host ants should thus be able to evolve to a higher virulence than those passaged through more genetically different host ants. The other two treatments in the first experiment (1e and f) were similar, but involved a mixture of two parasite strains (Fig. 1). The prediction here was that the greater diversity of parasites and the addition of within-host competition between parasite strains might stimulate a more rapid adaptation of the parasite than in the single strain treatments. The second experiment used a single parasite strain, but the serial passages involved groups of ants of either low or high diversity (Treatments 2a and b; Fig. 1). This experiment thus attempted to mimic more closely the natural situation in colonies. Competition will be more global in this second experiment and so the derived parasite strains may have lower virulence and greater spore production than the derived strains produced by the first experiment (Frank, 1998). The same logic will also apply within the second experiment. If the two treatments differ in the numbers of ants sporulating and thus being used for subsequent passages, then competition will be more global in the treatment with the greater number of ants sporulating.
Figure 1. Experimental design. Nine serial passages were carried out. This was followed by a post-selection assessment in which the derived strains resulting from the ninth passage were compared with the ancestral strain used to start the first passage. The study consisted of eight treatments differing either in the relatedness of hosts used in the different passages, the number of parasite strains used, or whether ants were maintained individually or in groups of three.
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