Rodger A. Gwiazdowski

A phylogenetic analysis of armored scale insects (Hemiptera: Diaspididae), based upon nuclear, mitochondrial, and endosymbiont gene sequences

Armored scale insects (Hemiptera: Diaspididae) are among the most invasive insects in the world. They have unusual genetic systems, including diverse types of paternal genome elimination (PGE) and parthenogenesis. Intimate relationships with their host plants and bacterial endosymbionts make them potentially important subjects for the study of co-evolution. Here, we expand upon recent phylogenetic work (Morse and Normark, 2006) by analyzing armored scale and endosymbiont DNA sequences from 125 species of armored scale insect, represented by 253 samples and eight outgroup species. We used fragments of four different gene regions: the nuclear protein-coding gene Elongation Factor 1α (EF1α), the large ribosomal subunit (28S) rDNA, a mitochondrial region spanning parts of cytochrome oxidase I (COI) and cytochrome oxidase II (COII), and the small ribosomal subunit (16S) rDNA from the primary bacterial endosymbiont Uzinura diaspidicola. Maximum likelihood, and Bayesian analyses were performed producing highly congruent topological results. A comparison of two datasets, one with and one without missing data, found that missing data had little effect on topology. Our results broadly corroborate several major features of the existing classification, although we do not find any of the subfamilies, tribes or subtribes to be monophyletic as currently constituted. Using ancestral state reconstruction we estimate that the ancestral armored scale had the late PGE sex system, and it may as well have been pupillarial, though results differed between reconstruction methods. These results highlight the need for a complete revision of this family, and provide the groundwork for future taxonomic work in armored scale insects.

Survey for Winter Moth (Lepidoptera: Geometridae) in Northeastern North America with Pheromone-Baited Traps and Hybridization with the Native Bruce Spanworm (Lepidoptera: Geometridae)

ABSTRACT We used pheromone-baited traps to survey the distribution of winter moth, Operophtera brumata (L.) (Lepidoptera: Geometridae), a new invasive defoliator from Europe in eastern New England. The traps also attracted Bruce spanworm, Operophtera bruceata (Hulst) (Lepidoptera: Geometridae), native to North America. We distinguished between the two species by examining male genitalia and sequencing the mitochondrial cytochrome oxidase subunit 1 (COI) gene, the DNA barcoding region. In 2005, we recovered winter moths at sites stretching from eastern Long Island, southeastern Connecticut, all of Rhode Island, eastern Massachusetts, coastal New Hampshire, and southern coastal Maine. At sites further west and north we captured only Bruce spanworm. In 2006, we confirmed that both winter moth and Bruce spanworm are present in Nova Scotia and in coastal Maine, but only Bruce spanworm was recovered in coastal New Brunswick, Canada; Pennsylvania; Vermont; or Quebec City, Canada. In 2007, we collected Bruce spanworm, but no winter moths, in New Brunswick and the interior areas of Maine, New Hampshire, and New York. Winter moth and Brace spanworm differed in the COI sequence by 7.45% of their nucleotides. The prevalence of intermediate genitalia in the zone of overlap suggested that hybridization between the two species may be occurring. To confirm the presence of hybrids, we sequenced the nuclear gene, glucose-6phosphate dehydrogenase (G6PD). We identified six nucleotides that routinely distinguished winter moth and Bruce spanworm, of which three were always diagnostic. We showed that eggs produced by hybridizing the two species in the laboratory contained copies of both species at these six sites. We found that most of the moths collected in the field with intermediate genitalia had winter moth CO1 and G6PD sequences and thus were not hybrids (or at least F1 hybrids). We found three hybrids out of 158 moths with intermediate genitalia in the region where both species were caught. We conclude that hybrids occur in nature, but are not as common as previously reported. Introgression of genes between the two species may still be significant.

Phylogeographic Diversity of the Winter Moths Operophtera brumata and O. bruceata (Lepidoptera: Geometridae) in Europe and North America

ABSTRACT The European winter moth, Operophtera brumata (L.), an invasive forest defoliator, is undergoing a rapid range expansion in northeastern North America. The source of this invasion, and phylogeographic diversity throughout its native range, has not been explored. To do this, we used samples from a pheromone-baited trap survey of O. brumata collected across its native range in Europe, and invasive range in North America. Traps in North America also attract a congeneric species, the Bruce spanworm O. bruceata (Hulst), and the western Bruce spanworm O. b. occidentalis (Hulst). From this sampling, we sequenced two regions of the cytochrome c oxidase subunit I mitochondrial gene; one region corresponds to the DNA ‘barcode’ region, the other is a nonoverlapping section. We used these sequences, in combination with sequence data from a recent survey of the Geometridae in western North America, for phylogenetic and phylogeographic analyses to characterize genetic divergence and variation for O. brumata in North America and Europe, and O. bruceata and O. b. occidentalis in North America. We found O. brumata mtDNA diversity to be dominated by a single widespread, and common haplotype. In contrast, O. bruceata shows high haplotype diversity that is evenly distributed throughout North America. Phylogeographic patterns indicate an introduction of O. brumata in British Columbia likely originated from Germany, and suggest the invasive population in northeastern North America may have its origins in the United Kingdom, and/or Germany. We found uncorrected pairwise sequence divergence between Operophtera species to be ≈7%. O. b. occidentalis is ≈ 5% divergent from O. bruceata, has a restricted range in the Pacific Northwest, and has unique morphological characters. Together these lines of evidence suggest O. b. occidentalis may be deserving of species status. Additionally, a single morphologically unique Operophtera specimen, similar to O. bruceata, was collected in southern Arizona, far outside the known range of O. bruceata. This suggests that North America may contain further, unsampled, Operophtera diversity.

Laboratory Rearing of Common and Endangered Species of North American Tiger Beetles (Coleoptera: Carabidae: Cicindelinae)

ABSTRACT North American tiger beetles (Cicindela spp. L.) have been reared in the laboratory for more than a century, and here we summarize the relevant literature to develop a general rearing protocol. We used this protocol to experimentally overwinter adults in the laboratory and observe variation in oviposition and fecundity among several species. Overwintering experiments, involving five North East North American Cicindela species with spring-fall life histories—Cicindela repanda (Dejean), Cicindela hirticollis (Say), Cicindela purpurea (Olivier), Cicindela scutellaris (Say), and Cicindela tranquebarica (Herbst) —demonstrated that both a long cooldown (20 to 4°C by a degree a day) and a short photoperiod (8:16 [L:D] h) maximized survival and minimized overwintering weight loss, which varied between species and sex. Observations of oviposition, larval abundance and larval development involving five Cicindela species with summer life histories revealed that Cicindela punctulata (Olivier) produced more first-instar larvae than Cicindela abdominalis (F.), Cicindela dorsalis dorsalis (Say), Cicindela puritana (Horn), or Cicindela unipunctata (F.) and that high mortality due to accidental desiccation may be overcome by rearing larvae individually in tubes rather than in bins. We also present a first account of larval rearing of the federally threatened species C. puritana and the northern Marthas Vineyard population of the federally threatened species C. d. dorsalis. Notably, C. d. dorsalis produced fewer larvae than more common species reared in this study. We conclude that rearing large numbers of larvae is feasible with endangered as well as common species and we propose future improvements for rearing as part of conservation efforts.

Corroborating molecular species discovery: Four new pine-feeding species of Chionaspis (Hemiptera, Diaspididae)

Abstract The genus Chionaspis (Hemiptera, Diaspididae) includes two North American species of armored scale insects feeding on Pinaceae: Chionaspis heterophyllae Cooley, and Chionaspis pinifoliae (Fitch). Despite the economic impact of conifer-feeding Chionaspis on horticulture, the species diversity in this group has only recently been systematically investigated using samples from across the group’s geographic and host range. This paper provides morphological recognition characters for four new species that were recently hypothesized to exist on the basis of molecular evidence. The new species, here described, are Chionaspis brachycephalon Vea sp. n., Chionaspis caudata Vea sp. n., Chionaspis sonorae Vea sp. n. and Chionaspis torreyanae Vea sp. n. One of the new species, Chionaspis caudata Vea, has a gland spine at the apex of the pygidium, between the median lobes, unlike any other species of Chionaspis. An identification key to the species of Chionaspis feeding on pine in North America is provided.

An Unidentified Parasitoid Community (Chalcidoidea) Is Associated with Pine-Feeding Chionaspis Scale Insects (Hemiptera: Diaspididae)

ABSTRACT Many of the >22,000 described species of parasitoids in the Chalcidoidea attack armored scale insects, a group that may contain many cryptic species. Because the diversity of both these groups may be high, associations between them are likely to be underreported, and accurate reporting requires direct association between parasitoids and their scale insect hosts. One group of well-known armored scale insects are the pine-feeding Chionaspis of North America, which have been considered to be two species of economic pests, and have over a century of parasitoid records. These Chionaspis have been revealed to be a complex of at least 10, mostly cryptic, species. We have sequenced chalcidoid 28S rDNA from 50 parasitized Chionaspis hosts. We explore these host-parasitoid associations by placing the parasitoid sequences in a phylogenetic context. We aligned the sequences manually, based on secondary structure models, and added them to a previous 28S secondary structure alignment for the Chalcidoidea that includes 19 families, 72 subfamilies, 369 genera, and 685 species. The results of maximum likelihood analyses were interpreted using an updated, comprehensive list of parasitoids recorded from pine-feeding Chionaspis. The sequences are placed in nine clades, only two of which appear consistent with known Chionaspis parasitoids. Although pine-feeding Chionaspis are among the best-known armored scales, our results indicate that their parasitoid diversity has been underreported.