Species Delimitation: A Case Study in a Problematic Ant Taxon

Species delimitation has been invigorated as a discipline in systematics by an influx of new character sets, analytical methods, and conceptual advances. We use genetic data from 68 markers, combined with distributional, bioclimatic, and coloration information, to hypothesize boundaries of evolutionarily independent lineages (species) within the widespread and highly variable nominal fire ant species Solenopsis saevissima, a member of a species group containing invasive pests as well as species that are models for ecological and evolutionary research. Our integrated approach uses diverse methods of analysis to sequentially test whether populations meet specific operational criteria (contingent properties) for candidacy as morphologically cryptic species, including genetic clustering, monophyly, reproductive isolation, and occupation of distinctive niche space. We hypothesize that nominal S. saevissima comprises at least 4-6 previously unrecognized species, including several pairs whose parapatric distributions implicate the development of intrinsic premating or postmating barriers to gene flow. Our genetic data further suggest that regional genetic differentiation in S. saevissima has been influenced by hybridization with other nominal species occurring in sympatry or parapatry, including the quite distantly related Solenopsis geminata. The results of this study illustrate the importance of employing different classes of genetic data (coding and noncoding regions and nuclear and mitochondrial DNA [mtDNA] markers), different methods of genetic data analysis (tree-based and non-tree based methods), and different sources of data (genetic, morphological, and ecological data) to explicitly test various operational criteria for species boundaries in clades of recently diverged lineages, while warning against over reliance on any single data type (e.g., mtDNA sequence variation) when drawing inference

Gene discovery using massively parallel pyrosequencing to develop ESTs for the flesh fly Sarcophaga crassipalpis

<p>Abstract</p> <p>Background</p> <p>Flesh flies in the genus <it>Sarcophaga </it>are important models for investigating endocrinology, diapause, cold hardiness, reproduction, and immunity. Despite the prominence of <it>Sarcophaga </it>flesh flies as models for insect physiology and biochemistry, and in forensic studies, little genomic or transcriptomic data are available for members of this genus. We used massively parallel pyrosequencing on the Roche 454-FLX platform to produce a substantial EST dataset for the flesh fly <it>Sarcophaga crassipalpis</it>. To maximize sequence diversity, we pooled RNA extracted from whole bodies of all life stages and normalized the cDNA pool after reverse transcription.</p> <p>Results</p> <p>We obtained 207,110 ESTs with an average read length of 241 bp. These reads assembled into 20,995 contigs and 31,056 singletons. Using BLAST searches of the NR and NT databases we were able to identify 11,757 unique gene elements (E<0.0001) representing approximately 9,000 independent transcripts. Comparison of the distribution of <it>S. crassipalpis </it>unigenes among GO Biological Process functional groups with that of the <it>Drosophila melanogaster </it>transcriptome suggests that our ESTs are broadly representative of the flesh fly transcriptome. Insertion and deletion errors in 454 sequencing present a serious hurdle to comparative transcriptome analysis. Aided by a new approach to correcting for these errors, we performed a comparative analysis of genetic divergence across GO categories among <it>S. crassipalpis</it>, <it>D. melanogaster</it>, and <it>Anopheles gambiae</it>. The results suggest that non-synonymous substitutions occur at similar rates across categories, although genes related to response to stimuli may evolve slightly faster. In addition, we identified over 500 potential microsatellite loci and more than 12,000 SNPs among our ESTs.</p> <p>Conclusion</p> <p>Our data provides the first large-scale EST-project for flesh flies, a much-needed resource for exploring this model species. In addition, we identified a large number of potential microsatellite and SNP markers that could be used in population and systematic studies of <it>S. crassipalpis </it>and other flesh flies.</p

Distribution and Prevalence of Wolbachia Infections in Native Populations of the Fire Ant Solenopsis invicta (Hymenoptera: Formicidae)

Wolbachia are endosymbiotic bacteria that commonly infect arthropods. These bacteria induce a number of phenotypes in their hosts, including cytoplasmic incompatibility, thelytokous parthenogenesis, feminization, and male killing. We surveyed native South American populations of the fire ant Solenopsis invicta Buren for Wolbachia infections by using a diagnostic polymerase chain reaction assay. In addition, we determined the fidelity of vertical transmission of the bacteria from mother to offspring in this species by assaying daughters in 24 simple-family (monogyne) colonies. Infections were common in many parts of the extensive native range of S. invicta. However, the proportion of individuals infected varied greatly among samples, ranging from zero in several populations from the northerly parts of the range to >90% in more southerly populations. Possible explanations for this variation in the prevalence of Wolbachia infections are discussed. A survey of the two social forms of S. invicta from four geographic areas showed that the prevalence of Wolbachia infections consistently was higher in the monogyne form (single queen per colony) than the sympatric polygyne form (multiple queens per colony). One likely explanation for this trend is that the selective regimes acting on Wolbachia in the two forms differ because of the dissimilar reproductive strategies used by each form. Finally, overall transmission efficiency was found to be very high (>99%), making it unlikely that imperfect transmission prevents the spread of the microbe to near fixation in native population

Distribution of the Two Social Forms of the Fire Ant Solenopsis invicta (Hymenoptera: Formicidae) in the Native South American Range

Polygyne (multiple queen) colony social organization in the fire ant Solenopsis invicta Buren is always associated with the presence of a particular class of alleles at the gene Gp-9. We used diagnostic polymerase chain reaction assays capable of distinguishing these alleles to determine the location of polygyne populations in the native South American range of this species. We found that polygyny occurs in a mosaic pattern with respect to the more common monogyne (single queen) social form, a pattern superficially similar to that seen in the introduced range in the United States. However, polygyny appears to be relatively restricted in its geographical prevalence in the native range compared with the introduced range. This difference may stem from higher dispersal rates in the introduced range, which are associated with greater opportunities for human-mediated transport of mated queens or colony fragments. On the basis of our distributional data and results from other studies, the southern part of the native range of S. invicta, particularly northeastern Argentina, is emerging as the most likely geographic source of the founders of the U.S. populatio

The Molecular Clockwork of the Fire Ant Solenopsis invicta

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Molecular Variation at a Candidate Gene Implicated in the Regulation of Fire Ant Social Behavior

The fire ant Solenopsis invicta and its close relatives display an important social polymorphism involving differences in colony queen number. Colonies are headed by either a single reproductive queen (monogyne form) or multiple queens (polygyne form). This variation in social organization is associated with variation at the gene Gp-9, with monogyne colonies harboring only B-like allelic variants and polygyne colonies always containing b-like variants as well. We describe naturally occurring variation at Gp-9 in fire ants based on 185 full-length sequences, 136 of which were obtained from S. invicta collected over much of its native range. While there is little overall differentiation between most of the numerous alleles observed, a surprising amount is found in the coding regions of the gene, with such substitutions usually causing amino acid replacements. This elevated coding-region variation may result from a lack of negative selection acting to constrain amino acid replacements over much of the protein, different mutation rates or biases in coding and non-coding sequences, negative selection acting with greater strength on non-coding than coding regions, and/or positive selection acting on the protein. Formal selection analyses provide evidence that the latter force played an important role in the basal b-like lineages coincident with the emergence of polygyny. While our data set reveals considerable paraphyly and polyphyly of S. invicta sequences with respect to those of other fire ant species, the b-like alleles of the socially polymorphic species are monophyletic. An expanded analysis of colonies containing alleles of this clade confirmed the invariant link between their presence and expression of polygyny. Finally, our discovery of several unique alleles bearing various combinations of b-like and B-like codons allows us to conclude that no single b-like residue is completely predictive of polygyne behavior and, thus, potentially causally involved in its expression. Rather, all three typical b-like residues appear to be necessary

Data from: Genetic analyses reveal cryptic diversity in the native North American fire ants (Hymenoptera: Formicidae: Solenopsis)

The native North American fire ants (Solenopsis) comprise a difficult group taxonomically that has undergone multiple revisions in the past century yet remains in a state of taxonomic uncertainty. In this study, we utilized a large set of microsatellite markers to conduct the first robust genetic analysis of the nominal species. Our approach used a variety of methods to test operational criteria commonly employed in species delimitation, including genotypic clustering, reproductive isolation/cohesion, and monophyly. We conclude from our results that the recognized North American fire ant species represent evolutionarily independent entities and, moreover, we confirm the presumed sister status of the desert fire ants, S. aurea and S. amblychila. However, the presence of at least two genetically divergent populations within the nominal species boundaries, including a western form of S. xyloni and a distinct population of S. aurea endemic to the Coachella Valley of California, suggests that the current taxonomy does not fully capture the species-level diversity in this group. Our study provides the molecular foundation for future integrated studies of the taxonomy and evolution of this scientifically and economically important group of insects

Data from: Joint evolution of asexuality and queen number in an ant

Ants exhibit a striking diversity of reproductive systems, varying in traits such as the number of reproductives per colony [1], the mode of daughter production (sexual or asexual) [2], and the mode of caste determination (genetic or environmental) [3]. Species employing mixed reproductive systems present a unique opportunity to explore the causes and consequences of alternative breeding strategies. Mixed reproductive systems in ants include social polymorphism in colony queen number, whereby single-queen (monogyne) and multiple-queen (polygyne) colonies co-occur within species [4, 5, 6, 7], and facultative asexuality, in which female offspring may be produced sexually or asexually within colonies [8, 9, 10, 11, 12, 13]. Here, we document a remarkable confluence of multiple mixed reproductive systems in the tropical fire ant, Solenopsis geminata, in a population with three important features: (1) polygyne colonies produce workers sexually but queens asexually, whereas monogyne colonies produce both castes sexually; (2) polygyne queens mate with monogyne males to produce workers, but monogyne queens do not mate with polygyne males; and (3) different asexual/polygyne lineages evidently were founded separately by genetically distinct founder queens, which appear to have originated from the same neighboring monogyne population. Multiple asexual/polygyne genomes are transmitted undiluted in this system, but sterile workers produced with sperm from a sexually-reproducing/monogyne population are necessary for the persistence of these lineages. The intersection of social polymorphism, facultative asexuality, and genetic caste determination marks this population of S. geminata as an embodiment of the diversity of ant reproductive systems and suggests previously unknown connections between these phenomena

Data from: Disruption of gene expression in hybrids of the fire ants Solenopsis invicta and Solenopsis richteri

Transcriptome analysis is a powerful tool for unveiling the distribution and magnitude of genetic incompatibilities between hybridizing taxa. The nature of such incompatibilities is closely associated with the evolutionary histories of the parental species and may differ across tissues and between the sexes. In eusocial insects, the presence of castes that experience divergent selection regimes may result in additional distinct patterns of caste-specific hybrid incompatibilities. We analyzed levels of expression of >14,000 genes in two life stages of each caste and sex in the fire ants S. invicta and S. richteri and in their hybrids. We found strong contributions of both developmental stage and caste to gene expression patterns. Hybrid incompatibilities were surprisingly modest: only 32 genes were mis-expressed, indicating low levels of disruption in gene regulation in hybrids. The castes differed in numbers of mis-expressed genes, with males and workers each mis-expressing at least seven times as many genes as queens. Interestingly, homologues of many of the mis-expressed genes have been implicated in behavioral variation in Drosophila melanogaster. General expression profiles of hybrids were more similar to those of S. richteri than S. invicta, presumably because S. richteri trans-regulatory elements tend to be dominant to those of S. invicta and/or because there is an overall bias in the genetic composition of the hybrids towards S. richteri. Altogether, our results suggest that selection acting on each caste may contribute differently to interspecific divergence and speciation in this group of ants