Genetically Depauperate and Still Successful: Few Multilocus Genotypes of the Introduced Parthenogenetic Weevil Naupactus cervinus (Coleoptera: Curculionidae) Prevail in the Continental United States.

Naupactus cervinus is a parthenogenetic weevil native to South America that is currently distributed worldwide. This flightless species is polyphagous and capable of modifying gene expression regimes for responding to stressful situations. Naupactus cervinus was first reported in the continental United States in 1879 and has rapidly colonized most of the world since. Previous studies suggested that an invader genotype successfully established even in areas of unsuitable environmental conditions. In the present work, we analyze mitochondrial and nuclear sequences from 71 individuals collected in 13 localities across three states in the southern US, in order to describe the genetic diversity in this area of introduction that has not yet been previously studied. Our results suggest that 97% of the samples carry the most prevalent invader genotype already reported, while the rest shows a close mitochondrial derivative. This would support the hypothesis of a general purpose genotype, with parthenogenesis and its associated lack of recombination maintaining the linkage of genetic variants capable of coping with adverse conditions and enlarging its geographical range. However, demographic advantages related to parthenogenetic reproduction as the main driver of geographic expansion (such as the foundation of a population with a single virgin female) cannot be ruled out. Given the historical introduction records and the prevalence of the invader genotype, it is possible that the continental US may act as a secondary source of introductions to other areas. We propose that both the parthenogenesis and scarce genetic variation in places of introduction may, in fact, be an asset that allows N. cervinus to thrive across a range of environmental conditions.

Host-specific gene expression as a tool for introduction success in Naupactus parthenogenetic weevils.

Food resource access can mediate establishment success in invasive species, and generalist herbivorous insects are thought to rely on mechanisms of transcriptional plasticity to respond to dietary variation. While asexually reproducing invasives typically have low genetic variation, the twofold reproductive capacity of asexual organisms is a marked advantage for colonization. We studied host-related transcriptional acclimation in parthenogenetic, invasive, and polyphagous weevils: Naupactus cervinus and N. leucoloma. We analyzed patterns of gene expression in three gene categories that can mediate weevil-host plant interactions through identification of suitable host plants, short-term acclimation to host plant defenses, and long-term adaptation to host plant defenses and their pathogens. This approach employed comparative transcriptomic methods to investigate differentially expressed host detection, detoxification, immune defense genes, and pathway-level gene set enrichment. Our results show that weevil gene expression responses can be host plant-specific, and that elements of that response can be maintained in the offspring. Some host plant groups, such as legumes, appear to be more taxing as they elicit a complex gene expression response which is both strong in intensity and specific in identity. However, the weevil response to taxing host plants shares many differentially expressed genes with other stressful situations, such as host plant cultivation conditions and transition to novel host, suggesting that there is an evolutionarily favorable shared gene expression regime for responding to different types of stressful situations. Modulating gene expression in the absence of other avenues for phenotypic adaptation may be an important mechanism of successful colonization for these introduced insects.

Effect of ethylenethiourea on metamorphosis and ovary development: A comparative study of three larval frogs.

Amphibian endocrine systems interact with each other during normal development. Interference with one of the endocrine systems may influence others. We studied the effect of a thyroid inhibitor (ethylenethiourea [ETU]) on metamorphosis and ovary development of three species, Sphaerotheca pashchima, Indosylvirana caesari, and Euphlyctis cyanophlyctis with different larval durations. We treated the tadpoles of these species with 50, 100, and 200 mg/L concentrations of ETU and studied their larval duration, size at metamorphosis, and ovary development. The results revealed that ETU affects metamorphosis, depending on the species and concentration. ETU delayed metamorphosis of E. cyanophlyctis tadpoles and did not affect metamorphosis in S. pashchima tadpoles. Lower concentrations of ETU stimulated metamorphosis in I. caesari tadpoles while high concentration delayed metamorphosis. In the tadpoles (E. cyanophlyctis) treated with higher concentrations of ETU, ovary development was advanced with an increased size of the diplotene oocytes. Oocyte size was smaller in the tadpoles (of I. caesari) treated with lower concentrations of ETU. These results demonstrated that the tadpoles of these species show different responses to the thyroid inhibitor, possibly due to the differences in the larval duration and sensitivity. Inhibition or acceleration of metamorphosis did not interfere in the ovary development of E. cyanophlyctis and I. caesari. These results will be useful in understanding the impact of endocrine disruptors on the interaction between thyroid and sex steroid hormones.

Genetic status and timing of a weevil introduction to Santa Cruz Island, Galapagos.

Successful invasive species can overcome or circumvent the potential genetic loss caused by an introduction bottleneck through a rapid population expansion and admixture from multiple introductions. We explore the genetic makeup and the timing of a species introduction to Santa Cruz Island in the Galápagos archipelago. We investigate the presence of processes that can maintain genetic diversity in populations of the broad-nosed weevil Galapaganus howdenae howdenae. Analyses of combined genotypes for 8 microsatellite loci showed evidence of past population size reductions through moment and likelihood-based estimators. No evidence of admixture through multiple introductions was found, but substantial current population sizes (N0 298, 95% credible limits 50-2300), genetic diversity comparable with long-established endemics (Mean number of alleles = 3.875), and lack of genetic structure across the introduced range (F ST = 0.01359) could suggest that foundations are in place for populations to rapidly recover any loss of genetic variability. The time estimates for the introduction into Santa Cruz support an accidental transfer during the colonization period (1832-1959) predating the spurt in human population growth. Our evaluation of the genetic status of G. h. howdenae suggests potential for population growth in addition to our field observations of a concurrent expansion in range and feeding preferences towards protected areas and endemic host plants.

Comparative genetic structure and demographic history in endemic galapagos weevils.

The challenge of maintaining genetic diversity within populations can be exacerbated for island endemics if they display population dynamics and behavioral attributes that expose them to genetic drift without the benefits of gene flow. We assess patterns of the genetic structure and demographic history in 27 populations of 9 species of flightless endemic Galápagos weevils from 9 of the islands and 1 winged introduced close relative. Analysis of mitochondrial DNA reveals a significant population structure and moderately variable, though demographically stable, populations for lowland endemics (F(ST) = 0.094-0.541; π: 0.014-0.042; Mismatch P = 0.003-0.026; and D((Tajima)) = -0.601 to 1.203), in contrast to signals of past contractions and expansions in highland specialists on 2 islands (Mismatch P = 0.003-0.026 and D((Tajima)) = -0.601 to 1.203). We interpret this series of variable and highly structured population groups as a system of long-established, independently founded island units, where structuring could be a signal of microallopatric differentiation due to patchy host plant distribution and poor dispersal abilities. We suggest that the severe reduction and subsequent increase of a suitably moist habitat that accompanied past climatic variation could have contributed to the observed population fluctuations in highland specialists. We propose the future exploration of hybridization between the introduced and highland endemic species on Santa Cruz, especially given the expansion of the introduced species into the highlands, the sensitivity to past climatic variation detected in highland populations, and the potentially threatened state of single-island endemics.

The age and phylogeny of wood boring weevils and the origin of subsociality.

A large proportion of the hyperdiverse weevils are wood boring and many of these taxa have subsocial family structures. The origin and relationship between certain wood boring weevil taxa has been problematic to solve and hypotheses on their phylogenies change substantially between different studies. We aimed at testing the phylogenetic position and monophyly of the most prominent wood boring taxa Scolytinae, Platypodinae and Cossoninae, including a range of weevil outgroups with either the herbivorous or wood boring habit. Many putatively intergrading taxa were included in a broad phylogenetic analysis for the first time in this study, such as Schedlarius, Mecopelmus, Coptonotus, Dactylipalpus, Coptocorynus and allied Araucariini taxa, Dobionus, Psepholax, Amorphocerus-Porthetes, and some peculiar wood boring Conoderini with bark beetle behaviour. Data analyses were based on 128 morphological characters, rDNA nucleotides from the D2-D3 segment of 28S, and nucleotides and amino acids from the protein encoding gene fragments of CAD, ArgK, EF-1α and COI. Although the results varied for some of the groups between various data sets and analyses, one may conclude the following from this study: Scolytinae and Platypodinae are likely sister lineages most closely related to Coptonotus; Cossoninae is monophyletic (including Araucariini) and more distantly related to Scolytinae; Amorphocerini is not part of Cossoninae and Psepholax may belong to Cryptorhynchini. Likelihood estimation of ancestral state reconstruction of subsociality indicated five or six origins as a conservative estimate. Overall the phylogenetic results were quite dependent on morphological data and we conclude that more genetic loci must be sampled to improve phylogenetic resolution. However, some results such as the derived position of Scolytinae were consistent between morphological and molecular data. A revised time estimation of the origin of Curculionidae and various subfamily groups were made using the recently updated fossil age of Scolytinae (100 Ma), which had a significant influence on node age estimates.

Temporal lags and overlap in the diversification of weevils and flowering plants.

The extraordinary diversity of herbivorous beetles is usually attributed to coevolution with angiosperms. However, the degree and nature of contemporaneity in beetle and angiosperm diversification remain unclear. Here we present a large-scale molecular phylogeny for weevils (herbivorous beetles in the superfamily Curculionoidea), one of the most diverse lineages of insects, based on approximately 8 kilobases of DNA sequence data from a worldwide sample including all families and subfamilies. Estimated divergence times derived from the combined molecular and fossil data indicate diversification into most families occurred on gymnosperms in the Jurassic, beginning approximately 166 Ma. Subsequent colonization of early crown-group angiosperms occurred during the Early Cretaceous, but this alone evidently did not lead to an immediate and major diversification event in weevils. Comparative trends in weevil diversification and angiosperm dominance reveal that massive diversification began in the mid-Cretaceous (ca. 112.0 to 93.5 Ma), when angiosperms first rose to widespread floristic dominance. These and other evidence suggest a deep and complex history of coevolution between weevils and angiosperms, including codiversification, resource tracking, and sequential evolution.

Molecular cloning and characterization of patellin1, a novel sec14-related protein, from zucchini (Cucurbita pepo).

A full-length patellin1 (PATL1) cDNA was cloned and characterized from zucchini (Cucurbita pepo). PATL1, originally discovered in the higher plant Arabidopsis thaliana, is a plant Sec14-related protein that localizes to the cell plate during the late stages of cytokinesis. PATL1 is related in sequence to other eukaryotic proteins involved in membrane trafficking and is thought to participate in vesicle trafficking events associated with cell plate maturation. The zucchini PATL1 (CpPATL1) cDNA predicts a 605 amino acid protein which consists of an acidic N-terminal domain (pI=4.2) followed by a Sec14 lipid-binding domain and a C-terminal Golgi dynamics domain (GOLD). The predicted CpPATL1 protein sequence shows a high degree of similarity to Arabidopsis PATL1, especially in the Sec14 (84%) and GOLD domains (87%). A phylogenetic analysis of all available full-length PATL sequences revealed that the PATLs belong to four distinct clades; CpPATL1 is a member of the PATL1/2 clade. RT-PCR analysis showed that the CpPATL1 gene is highly expressed throughout the plant. The domain structure, as well as biochemical fractionation studies, which demonstrated that CpPATL1 is a peripheral membrane protein, support a role in membrane trafficking events.

Phylogenetic relationships in Dichroplus Stål (Orthoptera: Acrididae: Melanoplinae) inferred from molecular and morphological data: testing karyotype diversification.

The neotropical genus Dichroplus and related genera are characterized by a relatively uniform external morphology and a remarkably divergent male genitalia and hence its taxonomy is controversial. It also shows an extreme karyotypic diversification. In this study we used molecular and morphological characters to test the monophyly of the genus and to evaluate chromosome evolution. Twenty-seven species from Dichroplus and related genera were included in the analysis. Morphological characters refer to the general morphology, male genitalia and female structures. Molecular studies were performed, sequencing part of two mitochondrial genes, cytochrome oxidase I and II. Independent and combined phylogenetic analyses of the data were performed under maximum parsimony. The karyotypic characters (rearrangements) were either mapped onto the combined topology or combined with the other data sets. While the molecular analysis confirms some results attained with morphology, some others do not. All point towards the paraphyly of the genus. Our results show the relevance of morphological data in phylogenetic studies because morphology and molecules supply complementary evidence. The mapping of chromosome characters on the combined tree shows that the most extreme karyotype, in D. silveiraguidoi, is a derived condition, probably reached through several centric fusions, and that X-autosome centric fusions were recurrently fixed during the evolution of the group.

Evolutionary rates in the adaptive radiation of beetles on plants.

Herbivorous insects and other small consumers are often specialized both in use of particular host taxa and in use of particular host tissues. Such consumers also often seem to show consistent differences in the rates of evolution of these two dimensions of host use, implying common processes, but this has been little studied. Here we quantify these rates of change in host use evolution in a major radiation of herbivorous insects, the Chrysomeloidea, whose diversity has been attributed to their use of flowering plants. We find a significant difference in the rates of evolutionary change in these two dimensions of host use, with host taxon associations most labile. There are apparently similar differences in rates of host use evolution in other parasite groups, suggesting the generality of this pattern. Divergences in parasite form associated with use of different host tissues may facilitate resource partitioning among successive adaptive radiations on particular host taxa.

Molecular and morphological phylogenetics of weevils (coleoptera, curculionoidea): do niche shifts accompany diversification?

The main goals of this study were to provide a robust phylogeny for the families of the superfamily Curculionoidea, to discover relationships and major natural groups within the family Curculionidae, and to clarify the evolution of larval habits and host-plant associations in weevils to analyze their role in weevil diversification. Phylogenetic relationships among the weevils (Curculionoidea) were inferred from analysis of nucleotide sequences of 18S ribosomal DNA (rDNA; approximately 2,000 bases) and 115 morphological characters of larval and adult stages. A worldwide sample of 100 species was compiled to maximize representation of weevil morphological and ecological diversity. All families and the main subfamilies of Curculionoidea were represented. The family Curculionidae sensu lato was represented by about 80 species in 30 "subfamilies" of traditional classifications. Phylogenetic reconstruction was accomplished by parsimony analysis of separate and combined molecular and morphological data matrices and Bayesian analysis of the molecular data; tree topology support was evaluated. Results of the combined analysis of 18S rDNA and morphological data indicate that monophyly of and relationships among each of the weevil families are well supported with the topology ((Nemonychidae, Anthribidae) (Belidae (Attelabidae (Caridae (Brentidae, Curculionidae))))). Within the clade Curculionidae sensu lato, the basal positions are occupied by mostly monocot-associated taxa with the primitive type of male genitalia followed by the Curculionidae sensu stricto, which is made up of groups with the derived type of male genitalia. High support values were found for the monophyly of some distinct curculionid groups such as Dryophthorinae (several tribes represented) and Platypodinae (Tesserocerini plus Platypodini), among others. However, the subfamilial relationships in Curculionidae are unresolved or weakly supported. The phylogeny estimate based on combined 18S rDNA and morphological data suggests that diversification in weevils was accompanied by niche shifts in host-plant associations and larval habits. Pronounced conservatism is evident in larval feeding habits, particularly in the host tissue consumed. Multiple shifts to use of angiosperms in Curculionoidea were identified, each time associated with increases in weevil diversity and subsequent shifts back to gymnosperms, particularly in the Curculionidae.