Novel conopeptides of the I-superfamily occur in several clades of cone snails

The I-superfamily of conotoxins represents a new class of peptides in the venom of some Conus species. These toxins are characterized by four disulfide bridges and inhibit or modify ion channels of nerve cells. When testing venoms from 11 Conus species for a functional characterization, blocking activity on potassium channels (like Kv1.1 and Kv1.3 channels, but not Kv1.2 channels) was detected in the venom of Conus capitaneus, Conus miles, Conus vexillum and Conus virgo. Analysis at the cDNA level of these venoms using primers designed according to the amino acid sequence of a potassium channel blocking toxin (ViTx) from C. virgo confirmed the presence of structurally homologous peptides in these venoms. Moreover, peptides belonging to the I-superfamily, but with divergent amino acid sequences, were found in Conus striatus and Conus imperialis. In all cases, the sequences of the precursors' prepro-regions exhibited high conservation, whereas the sequences of the mature peptides ranged from almost identical to highly divergent between species. We then performed phylogenetic analyses of new and published mitochondrial 16S rDNA sequences representing 104 haplotypes from these and numerous other Conus species, using Bayesian, maximum-likelihood, maximum-parsimony and neighbor-joining methods of inference. Cone snails known to possess I-superfamily toxins were assigned to five different major clades in all of the resulting gene trees. Moreover, I-superfamily conopeptides were detected both in vermivorous and piscivorous species of Conus, thus demonstrating the widespread presence of such toxins in this speciose genus beyond evolutionary and ecological groups

Population structure and distribution patterns of the sibling mosquito apecies Culex pipiens and Culex torrentium (Diptera: Culicidae) reveal different evolutionary paths

Nowadays a number of endemic mosquito species are known to possess vector abilities for various diseases, as e.g. the sibling species Culex pipiens and Culex torrentium. Due to their morphological similarity, ecology, distribution and vector abilities, knowledge about these species' population structure is essential. Culicidae from 25 different sampling sites were collected from March till October 2012. All analyses were performed with aligned cox1 sequences with a total length of 658 bp. Population structure as well as distribution patterns of both species were analysed using molecular methods and different statistical tests like distance based redundancy analysis (dbDRA), analysis of molecular variances (AMOVA) or McDonald & Kreitman test and Tajima's D. Within both species, we could show a genetic variability among the cox1 fragment. The construction of haplotype networks revealed one dominating haplotype for Cx. pipiens, widely distributed within Germany and a more homogeneous pattern for Cx. torrentium. The low genetic differences within Cx. pipiens could be a result of an infection with Wolbachia which can induce a sweep through populations by passively taking the also maternally inherited mtDNA through the population, thereby reducing the mitochondrial diversity as an outcome of reproductive incompatibility. Pairwise population genetic differentiation (FST) ranged significantly from moderate to very great between populations of Cx. pipiens and Cx. torrentium. Analyses of molecular variances revealed for both species that the main genetic variability exists within the populations (Cx. pipiens [88.38%]; Cx. torrentium [66.54%]). Based on a distance based redundancy analysis geographical origin explained a small but significant part of the species' genetic variation. Overall, the results confirm that Cx. pipiens and Cx. torrentium underlie different factors regarding their mitochondrial differentiation, which could be a result of endosymbiosis, dispersal between nearly located populations or human introduction

RegIIA: An a4/7-conotoxin from the venom of Conus regius that potently blocks a3ß4 nAChRs

Neuronal nicotinic acetylcholine receptors (nAChRs) play pivotal roles in the central and peripheral nervous systems. They are implicated in disease states such as Parkinson's disease and schizophrenia, as well as addictive processes for nicotine and other drugs of abuse. Modulation of specific nAChRs is essential to understand their role in the CNS. a-Conotoxins, disulfide-constrained peptides isolated from the venom of cone snails, potently inhibit nAChRs. Their selectivity varies markedly depending upon the specific nAChR subtype/a-conotoxin pair under consideration. Thus, a-conotoxins are excellent probes to evaluate the functional roles of nAChRs subtypes. We isolated an a4/7-conotoxin (RegIIA) from the venom of Conus regius. Its sequence was determined by Edman degradation and confirmed by sequencing the cDNA of the protein precursor. RegIIA was synthesized using solid phase methods and native and synthetic RegIIA were functionally tested using two-electrode voltage clamp recording on nAChRs expressed in Xenopus laevis oocytes. RegIIA is among the most potent antagonist of the a3ß4 nAChRs found to date and is also active at a3ß2 and a7 nAChRs. The 3D structure of RegIIA reveals the typical folding of most a4/7-conotoxins. Thus, while structurally related to other a4/7 conotoxins, RegIIA has an exquisite balance of shape, charge, and polarity exposed in its structure to potently block the a3ß4 nAChRs

Population Structure and Distribution Patterns of the Sibling Mosquito Species <i>Culex pipiens</i> and <i>Culex torrentium</i> (Diptera: Culicidae) Reveal Different Evolutionary Paths

<div><p>Nowadays a number of endemic mosquito species are known to possess vector abilities for various diseases, as e.g. the sibling species <i>Culex pipiens</i> and <i>Culex torrentium</i>. Due to their morphological similarity, ecology, distribution and vector abilities, knowledge about these species' population structure is essential. Culicidae from 25 different sampling sites were collected from March till October 2012. All analyses were performed with aligned cox1 sequences with a total length of 658 bp. Population structure as well as distribution patterns of both species were analysed using molecular methods and different statistical tests like distance based redundancy analysis (dbDRA), analysis of molecular variances (AMOVA) or McDonald & Kreitman test and Tajima's D. Within both species, we could show a genetic variability among the cox1 fragment. The construction of haplotype networks revealed one dominating haplotype for <i>Cx. pipiens</i>, widely distributed within Germany and a more homogeneous pattern for <i>Cx. torrentium</i>. The low genetic differences within <i>Cx. pipiens</i> could be a result of an infection with <i>Wolbachia</i> which can induce a sweep through populations by passively taking the also maternally inherited mtDNA through the population, thereby reducing the mitochondrial diversity as an outcome of reproductive incompatibility. Pairwise population genetic differentiation (F_ST) ranged significantly from moderate to very great between populations of <i>Cx. pipiens</i> and <i>Cx. torrentium</i>. Analyses of molecular variances revealed for both species that the main genetic variability exists within the populations (<i>Cx. pipiens</i> [88.38%]; <i>Cx. torrentium</i> [66.54%]). Based on a distance based redundancy analysis geographical origin explained a small but significant part of the species' genetic variation. Overall, the results confirm that <i>Cx. pipiens</i> and <i>Cx. torrentium</i> underlie different factors regarding their mitochondrial differentiation, which could be a result of endosymbiosis, dispersal between nearly located populations or human introduction.</p></div

Haplotype networks of <i>Culex pipiens</i> and <i>Culex torrentium</i> for the cox1 gene segment calculated using statistical parsimony as implemented in TCS 1.21.

<p>The squares stand for the most probable ancestral haplotypes, the circle for all other haplotypes. The Numbers are equal to the haplotypes of each species. Each line represents a single mutation while small white dots symbolize hypothetical missing haplotypes. The size of the circles and the square is proportional to the number of the occurring haplotypes. The number of individuals can be derived from the scale which is given in the figure. Different colors represent the different geographical sampling localities. The colored area is proportional to the occurrence at the respective site.</p

Population pairwise F_ST from <i>Culex torrentium</i> calculated with Arlequin 3.5.1.2.

<p>Data based on the analyses of 193 sequences of the cox1 gene segment from 10 different sampling localities in Germany (FFM = Frankfurt/Main with KS, FB and FZ). Sample size are shown in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0102158#pone-0102158-t001" target="_blank">Table 1</a>, abbreviations of each locality are shown in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0102158#pone-0102158-t001" target="_blank">Table 1</a> and <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0102158#pone-0102158-t003" target="_blank">3</a>. Significant different F_ST values are shown in bold. For calculation only populations with 5 or more individuals were used.</p

Results of the Analysis of molecular variance (AMOVA) and level of genetic differentiation of <i>Culex pipiens</i> and <i>Culex torrentium</i> measured by F_CT, F_SC, and F_ST.

<p>All Data were calculated in Arlequin 3.5.1.2 <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0102158#pone.0102158-Excoffier2" target="_blank">[59]</a> based on the distance method of Tamura and Nei <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0102158#pone.0102158-Tamura3" target="_blank">[60]</a>.</p