A Preliminary Molecular Phylogeny of the Ant-Decapitating Flies, Genus Apocephalus
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A Preliminary Molecular Phylogeny of the Ant-decapitating Flies, Genus Apocephalus (Diptera: Phoridae) By Christine C. Hayes, B.S. A Thesis Submitted to the Department of Biology California State University Bakersfield In Partial Fulfillment for the Degree of Master of Science Spring 2013 Copyright By Christine C. Hayes 2013 A Preliminary Molecular Phylogeny ofthe Ant-decapitating Flies, GenusApocephalus (Diptera: Phoridae) By Christine C. Hayes This thesis or project has been accepted on behalf of the Department of Biology by their supervisory committee: Dr. Paul T. Smith Committee Chair Dr. Brian V. Brown Dr. Carl T. Kloock A Preliminary Molecular Phylogeny of the Ant-decapitating Flies, Genus Apocephalus (Diptera: Phoridae) Christine C. Hayes Department of Biology California State University, Bakersfield Abstract The phorid fly genus Apocephalus is the largest assemblage of ant-parasitizing Phoridae. Apocephalus is currently organized into two subgenera: A. (Apocephalus) and A. (Mesophora). The species of A. (Mesophora) attack a wide variety of non-ant hosts including stingless bees, spiders, wasps, bumble bees, and cantharoid beetles. The species of A. (Apocephalus) are the true “ant-decapitating flies” and are divided into six species groups: the A. attophilus group (parasitoids of attine leaf-cutting ants), “A. miricauda group” (parasitoids of ponerine ants), A. pergandei group (parasitoids of Camponotus carpenter ants), A. mucronatus group, A. feeneri group, and A. grandipalpus group. Here I report on a preliminary molecular phylogenetic study of Apocephalus, including representatives of both subgenera and exemplars of five currently recognized species groups. Maximum parsimony, maximum likelihood, and Bayesian phylogenies were inferred using four nuclear (AK, TPI, CAD, 28S) and four mitochondrial (12S, 16S, COI, ND1) gene fragments (4284 bp total). For all analyses Apocephalus was recovered as a monophyletic group relative to the outgroup taxa included in the study. In addition, subgenus A. (Mesophora) was recovered as a monophyletic group, but was not a sister group to the subgenus A. (Apocephalus). A phylogenetic hypothesis for exemplars of five Apocephalus species groups is presented and compared to hypotheses based on morphology. v Table of Contents List of Tables.………………………………………………………………..…… vii List of Figures……………………………………………………………………... viii A Preliminary Molecular Phylogeny of the Ant-decapitating Flies, Genus Apocephalus (Diptera: Phoridae) Introduction………………………………………………………………………... 1 Methods……………...………………………………………………………....…. 6 Results and Discussion……..……………………………………………………... 8 Acknowledgments ………………………………………………………………... 12 Literature Cited……………………………………………………………………. 23 vi List of Tables TABLE 1……………………………………………………………………………… 13 Taxa used in present study with current taxonomic distinctions. TABLE 2……………………………………………………………………………… 13 Oligonucleotide primers used in this study. TABLE 3……………………………………………………………………………… 14 Average nucleotide frequencies for all Apocephalus taxa by gene. TABLE 4……………………………………………………………………………… 15 Absolute pairwise distance matrix of all genes for 18 Apocephalus species. TABLE 5……………………………………………………………………………… 16 Within and between species group divergence values. Between group comparisons were calculated by averaging within group pairwise divergence values. TABLE 6……………………………………………………………………………… 16 Descriptive statistics for maximum parsimony trees inferred from three data partitions. PICs = parsimony informative characters, TL = tree length, EPTs = equally parsimonious trees, CI = consistency index, and RI = retention index. vii List of Figures FIGURE 1…………………………………………………………………….……….. 17 Strict consensus of three equally parsimonious trees with bootstrap support values. Tree length = 4472, CI = 0.499, RI = 0.520. FIGURE 2……………………………………………………………………………... 18 The single most parsimonious tree resulting from the nuclear gene only partition with bootstrap support values. TL = 1805, CI = 0.550, RI = 0.638. FIGURE 3……………………………………………………………………………... 19 Phylogenetic tree resulting from maximum likelihood analysis. Numbers at various nodes are bootstrap values in %. Final maximum likelihood optimization likelihood = log -25862.9. FIGURE 4……………………………………………………………………………... 20 Consensus tree resulting from 15,002 trees sampled from Bayesian analysis showing posterior probabilities. The GTR+I+G model was used for all genes except CAD, where GTR+G was determined to be the most appropriate model. FIGURE 5……………………………………………………………………………... 21 Composite tree depicting relationships of the various Apocephalus species groups. Branch labels indicate known host association for each clade. FIGURE 6……………………………………………………………………………... 22 The composite tree depicting relationships of the various Apocephalus species groups found in this study compared to the current phylogenetic hypothesis for the relationship of the ant host subfamilies viii A Preliminary Molecular Phylogeny of the Ant-decapitating Flies, Genus Apocephalus (Diptera: Phoridae) Introduction The family Phoridae is a highly diverse and speciose group of insects that are in need of much additional study. Researchers have suggested that only about 10% of the family has been described (Disney 1994, Brown & Smith 2010). Phorids are generally small insects and range in size from ~0.4-7 mm. Indeed the smallest known described fly is a member of the Phoridae (Brown 2012b). In addition to variation in size, there is also an incredible amount of morphological diversity and sexual dimorphism exhibited among the numerous phorid genera. Thus, because of their small size and extreme morphological variation, phorids are not as well studied as some other fly families. Perhaps one of the most spectacular features of phorid biology is the wide variety of larval feeding habits that exist. Members of this family are known to feed on a wide range of decaying organic material, while others are fungivores, herbivores, predators, parasites, and parasitoids (Binns 1980, Brown 1992, Disney 1994). In some cases, a single species may exhibit more than one type of larval feeding. For example, Megaselia scalaris, the most studied phorid species, has been known to feed on a wide variety of decaying organic material, as well as viable frog eggs (Villa & Townsend 1983, Disney 2008). Among the diverse larval feeding habits exhibited among phorids, the parasitoids of social insects are arguably the most fascinating because in some cases there has been selection for some atypical and extravagant body forms, which is an indication of some remarkable coevolutionary scenarios (Brown 1992, Feener & Brown 1992, Folgarait et al. 2002, Disney 1994). 1 The term parasitoid is functionally defined as an organism that develops on, or in, and extracts nourishment from another organism (usually during a specific host developmental stage); but in contrast to a basic parasite, this results in the death of the host organism (Eggleton & Gaston 1990). In general, the hymenopteran parasitoids have been more widely studied than the dipteran parasitoids. Hymenopteran parasitoids are thought to have diverged from a single evolutionary event; however, for the lesser known dipteran parasitoids, the lifestyle is thought to have evolved independently several times, including within Phoridae (Eggleton & Belshaw 1992, Feener & Brown 1997). Host organisms used by phorid parasitoids include both insects (e.g., cockroaches, termites, true bugs, beetles, moths/butterflies, flies, ants/bees/wasps), and non-insect terrestrial invertebrates (e.g., worms, snails, spiders, millipedes) (Disney 1994, Feener & Brown 1997). Some genera within Phoridae include a majority of species that specialize on a particular host group. For example, most Pseudacteon and Apocephalus species parasitize ants by laying their eggs in the body of the ant host; the larva develops by feeding on muscles and/or viscera, and then emerge from the host either prior to or following completion of pupation (Brown & Feener 1998, Mathis & Philpott 2012). Some unrelated phorid parasitoids may actually exhibit overlap in the species and/or individual host that is targeted. In the case of the latter, phorid parasitoids may oviposit in different body parts to reduce competition and/or hyperparasitism (Brown 1999). One of the largest and most diverse parasitoid groups in the Phoridae is the genus Apocephalus. In addition to morphology, the taxonomic division of species into species groups appears to relate somewhat with host associations. Although species often vary in their host specificity, and the general life history of many Apocephalus species is not well 2 understood, previous studies on Apocephalus (e.g., Brown & Feener 1991) have shed some light on the biology and natural history of some. For example, specific pheromonal olfactory cues are thought to be very important for Apocephalus paraponerae in locating its ant host, Paraponera clavata, indicating a high level of host specificity (Brown & Feener 1991). Moreover, research conducted on this same species-pair by Morehead & Feener (2000) found that, in addition to P. clavata, many other species could serve as suitable hosts, but that host range may be limited by the ability of the individual to locate the host. Additional studies such as the ones described above are sorely needed for most species of Apocephalus; however, before progress can be made toward a better understanding of the biology and natural history of Apocephalus in general, it is crucial to have a better understanding of the phylogenetic