Morphological, molecular and cross-breeding analysis of geographic populations of coconut-mite associated predatory mites identified as Neoseiulus baraki: evidence for cryptic species?

Surveys were conducted in Brazil, Benin and Tanzania to collect predatory mites as candidates for control of the coconut mite Aceria guerreronis Keifer, a serious pest of coconut fruits. At all locations surveyed, one of the most dominant predators on infested coconut fruits was identified as Neoseiulus baraki Athias-Henriot, based on morphological similarity with regard to taxonomically relevant characters. However, scrutiny of our own and published descriptions suggests that consistent morphological differences may exist between the Benin population and those from the other geographic origins. In this study, we combined three methods to assess whether these populations belong to one species or a few distinct, yet closely related species. First, multivariate analysis of 32 morphological characters showed that the Benin population differed from the other three populations. Second, DNA sequence analysis based on the mitochondrial cytochrome oxidase subunit I (COI) showed the same difference between these populations. Third, cross-breeding between populations was unsuccessful in all combinations. These data provide evidence for the existence of cryptic species. Subsequent morphological research showed that the Benin population can be distinguished from the others by a new character (not included in the multivariate analysis), viz. the number of teeth on the fixed digit of the female chelicera.

Lack of COI variation for Clavelina oblonga (Tunicata, Ascidiacea) in Brazil: Evidence for its human-mediated transportation?

Recent studies indicate that ascidians are efficiently dispersed by human transport. We have chosen the mitochondrial gene cytochrome c oxidase subunit I (COI) to address whether Clavelina oblonga is an introduced species in the Brazilian coast. Colonies of C. oblonga were sampled in different localities along Atlantic coasts of USA, Panama, and Brazil. The sequencing of 92 colonies resulted in three haplotypes for the species, two unique to Florida and the other shared by exemplars collected in Brazil and Panama; the latter haplotype is identical to the published sequence of Azores. Our evidence, including the absence of C. oblonga in the country's northern tropical waters, its association with artificial habitats and lack of COI variation suggest that the species has been introduced in the southeastern and southern Brazilian coasts. Previous records (85 years old) suggest that it could be a relatively long-term introduction.

NEW RECORDS OF ANOPHELES HOMUNCULUS IN CENTRAL AND SERRA DO MAR BIODIVERSITY CORRIDORS OF THE ATLANTIC FOREST, BRAZIL

Two new records of Anopheles homunculus in the eastern part of the Atlantic Forest are reported. This species was found for the first time in Barra do Ouro district, Maquine municipality, Rio Grande do Sul state, located in the southern limit of the Atlantic Forest. The 2nd new record was in the Serra Bonita Reserve, Camacan municipality, southeast Bahia state. These records extend the geographical distribution of An. homunculus, suggesting that the species may be widely distributed in coastal areas of the Atlantic Forest. It is hypothesized that the disjunct distribution of the species may be caused by inadequate sampling, and also difficulties in species identification based only on female external characteristics. Species identification was based on morphological characters of the male, larva, and pupa, and corroborated by DNA sequence analyses, employing data from both 2nd internal transcribed spacer of nuclear ribosomal DNA and of mitochondrial cytochrome c oxidase subunit I.

Phylogenetic relationships of rock-wallabies, Petrogale (Marsupialia: Macropodidae) and their biogeographic history within Australia

The rock-wallaby genus Petrogale comprises a group of habitat-specialist macropodids endemic to Australia. Their restriction to rocky outcrops, with infrequent interpopulation dispersal, has been suggested as the cause of their recent and rapid diversification. Molecular phylogenetic relationships within and among species of Petrogale were analysed using mitochondrial (cytochrome oxidase c subunit 1, cytochrome b. NADH dehydrogenase subunit 2) and nuclear (omega-globin intron, breast and ovarian cancer susceptibility gene) sequence data with representatives that encompassed the morphological and chromosomal variation within the genus, including for the first time both Petrogale concinna and Petrogale purpureicollis. Four distinct lineages were identified, (1) the brachyotis group, (2) Petrogale persephone, (3) Petrogale xanthopus and (4) the lateralis-penicillata group. Three of these lineages include taxa with the ancestral karyotype (2n = 22). Paraphyletic relationships within the brachyotis group indicate the need for a focused phylogeographic study. There was support for P. purpureicollis being reinstated as a full species and P. concinna being placed within Petrogale rather than in the monotypic genus Peradorcas. Bayesian analyses of divergence times suggest that episodes of diversification commenced in the late Miocene-Pliocene and continued throughout the Pleistocene. Ancestral state reconstructions suggest that Petrogale originated in a mesic environment and dispersed into more arid environments, events that correlate with the timing of radiations in other arid zone vertebrate taxa across Australia. Crown Copyright (C) 2011 Published by Elsevier Inc. All rights reserved.

Redescriptions of two species and descriptions of three new species of the louse genus Bizarrifrons Eichler, 1938 (Phthiraptera: Ischnocera: Philopteridae)

Redescriptions of Bizarrifrons magus (Nitzsch [in Giebel], 1866), the type species of Bizarrifrons, and B. picturatus Carriker & Diaz-Ungria, 1961 are given based on material from their type hosts. The nymphal instars of these two species are described and illustrated for the first time. Also, three new species are named and described: B. latifrons, from the russet-backed oropendola, Psarocolius angustifrons alfredi (Des Murs, 1856); B. wecksteini, from the Amazonian oropendola, Psarocolius b. bifasciatus (Spix, 1824); and B. quasisymmetricus, from the solitary cacique, Cacicus solitarius (Vieillot, 1816) (Passeriformes: Icteridae). Two species-groups are proposed, and a checklist and a key for the species of Bizarrifrons are also included. Sequences of a portion of the mitochondrial cytochrome oxidase I (COI) and the nuclear elongation factor 1 alpha (EF-1 alpha) genes for two species are given for the first time in this genus.

Morphological and genetic variability in Hippolyte obliquimanus dana, 1852 (Decapoda, Caridea, Hippolytidae) from Brazil and the caribbean sea

Hippolyte obliquimanus is a marine shrimp reported from the Caribbean Sea and Brazil. The literature provides indications for morphological variation between populations from those regions and the species has a troubled taxonomic history. The aims of this study were to analyse morphological and genetic variation in the populations of H. obliquimanus from Brazil and the Caribbean Sea and to verify if those might support separation of H. obliquimanus into two or more species. This hypothesis was tested with the analysis of morphological and genetic data (mitochondrial gene 16S and the barcode region Cytochrome Oxidase I). The material analysed was obtained from samples and from loans of zoological collections. The rostrum as well as pereiopods 3, 4, and 5 were the adult morphological characters that showed variation, but this occurred in samples from both regions, Brazil and the Caribbean Sea. The sequences of the 16S gene were identical among all specimens analysed. There was, however, variation among the sequences of the barcoding gene COI (<2.0%); this divergence separated the specimens into two groups (Brazil versus the Caribbean) and these groups did not share haplotypes. In conclusion, specimens from the regions analysed showed both morphological and genetic variation, but these did not support the separation of H. obliquimanus into two or more species.

Cryptic species in the cosmopolitan Bugulan eritina complex (Bryozoa,Cheilostomata)

Previous analyses of the mitochondrial gene cytochrome c oxidase subunit 1 (COI) and γ-proteobacterial endosymbiont diversity have suggested that the marine bryozoan Bugula neritina is a complex of three cryptic species, namely Types S, D and N. Types D and N were previously reported to have restricted distributions along California (western USA) and Delaware and Connecticut (eastern USA), respectively, whereas Type S is considered widespread in tropical, subtropical and temperate regions due to anthropogenic transport. Here, Bayesian species delimitation analysis of a data set composed of two mitochondrial (COI and large ribosomal RNA subunit [16S]) and two nuclear genes (dynein light chain roadblock type-2 protein [DYN] and voltage-dependent anion-selective channel protein [VDAC]) demonstrated that Types S, D and N correspond to three biological species. This finding was significantly supported, in spite of the combinations of priors applied for ancestral population size and root age. Furthermore, COI sequences were used to assess the introduction patterns of the cosmopolitan Type S species. Two COI haplotypes of Type S (S1a and S1d) were found occurring at a global scale. Mantel tests showed correlation between these haplotypes and local sea surface temperature tolerance. Accordingly, the distributions of Type S haplotypes may reflect intraspecific temperature tolerance variation, in addition to the role of introduction vectors. Finally, we show that the Type N may also have been introduced widely, as this species was found for the first time in Central California and north-eastern Australia.

Systematics of the Oswaldoi Complex (Anopheles, Nyssorhynchus) in South America

Abstract Background Effective malaria control relies on accurate identification of those Anopheles mosquitoes responsible for the transmission of Plasmodium parasites. Anopheles oswaldoi s.l. has been incriminated as a malaria vector in Colombia and some localities in Brazil, but not ubiquitously throughout its Neotropical range. This evidence together with variable morphological characters and genetic differences supports that An. oswaldoi s.l. compromises a species complex. The recent fully integrated redescription of An. oswaldoi s.s. provides a solid taxonomic foundation from which to molecularly determine other members of the complex. Methods DNA sequences of the Second Internal Transcribed Spacer (ITS2 - rDNA) (n = 192) and the barcoding region of the Cytochrome Oxidase I gene (COI - mtDNA) (n = 110) were generated from 255 specimens of An. oswaldoi s.l. from 33 localities: Brazil (8 localities, including the lectotype series of An. oswaldoi), Ecuador (4), Colombia (17), Trinidad and Tobago (1), and Peru (3). COI sequences were analyzed employing the Kimura-two-parameter model (K2P), Bayesian analysis (MrBayes), Mixed Yule-Coalescent model (MYC, for delimitation of clusters) and TCS genealogies. Results Separate and combined analysis of the COI and ITS2 data sets unequivocally supported four separate species: two previously determined (An. oswaldoi s.s. and An. oswaldoi B) and two newly designated species in the Oswaldoi Complex (An. oswaldoi A and An. sp. nr. konderi). The COI intra- and inter-specific genetic distances for the four taxa were non-overlapping, averaging 0.012 (0.007 to 0.020) and 0.052 (0.038 to 0.064), respectively. The concurring four clusters delineated by MrBayes and MYC, and four independent TCS networks, strongly confirmed their separate species status. In addition, An. konderi of Sallum should be regarded as unique with respect to the above. Despite initially being included as an outgroup taxon, this species falls well within the examined taxa, suggesting a combined analysis of these taxa would be most appropriate. Conclusions: Through novel data and retrospective comparison of available COI and ITS2 DNA sequences, evidence is shown to support the separate species status of An. oswaldoi s.s., An. oswaldoi A and An. oswaldoi B, and at least two species in the closely related An. konderi complex (An. sp. nr. konderi, An. konderi of Sallum). Although An. oswaldoi s.s. has never been implicated in malaria transmission, An. oswaldoi B is a confirmed vector and the new species An. oswaldoi A and An. sp. nr. konderi are circumstantially implicated, most likely acting as secondary vectors.

Identification of epsilon PKC Targets During Cardiac Ischemic Injury

Background: Epsilon-protein kinase C (epsilon PKC) protects the heart from ischemic injury. However, the mechanism(s) of epsilon PKC cardioprotection is still unclear. Identification of the epsilon PKC targets may aid in elucidating the epsilon PKC-mediated cardioprotective mechanisms. Previous studies, using epsilon PKC transgenic mice and difference in gel electrophoresis, identified proteins involved in glucose metabolism, the expression of which was modified by epsilon PKC. Those studies were accompanied by metabolomic analysis, suggesting that increased glucose oxidation may be responsible for the cardioprotective effect of epsilon PKC. Whether these epsilon PKC-mediated alterations were because of differences in protein expression or phosphorylation was not determined. Methods and Results: In the present study, we used an epsilon PKC -specific activator peptide, psi epsilon RACK, combined with phosphoproteomics, to find epsilon PKC targets, and identified that the proteins whose phosphorylation was altered by selective activation of epsilon PKC were mostly mitochondrial proteins. Analysis of the mitochondrial phosphoproteome led to the identification of 55 spots, corresponding to 37 individual proteins, exclusively phosphorylated, in the presence of psi epsilon RACK. The majority of the proteins identified were involved in glucose and lipid metabolism, components of the respiratory chain as well as mitochondrial heat shock proteins. Conclusions: The protective effect of epsilon PKC during ischemia involves phosphorylation of several mitochondrial proteins involved in glucose and lipid metabolism and oxidative phosphorylation. Regulation of these metabolic pathways by epsilon PKC phosphorylation may lead to epsilon PKC-mediated cardioprotection induced by psi epsilon RACK. (Circ J 2012; 76: 1476-1485)

Genetic variability and demographic history of Heliothis virescens (Lepidoptera: Noctuidae) populations from Brazil inferred by mtDNA sequences

Intra-and inter-population genetic variability and the demographic history of Heliothis virescens (F.) populations were evaluated by using mtDNA markers (coxI, coxII and nad6) with samples from the major cotton-and soybean-producing regions in Brazil in the growing seasons 2007/08, 2008/09 and 2009/10. AMOVA indicated low and non-significant genetic structure, regardless of geographical scale, growing season or crop, with most of genetic variation occurring within populations. Clustering analyzes also indicated low genetic differentiation. The haplotype network obtained with combined datasets resulted in 35 haplotypes, with 28 exclusive occurrences, four of them sampled only from soybean fields. The minimum spanning network showed star-shaped structures typical of populations that underwent a recent demographic expansion. The recent expansion was supported by other demographic analyzes, such as the Bayesian skyline plot, the unimodal distribution of paired differences among mitochondrial sequences, and negative and significant values of neutrality tests for the Tajima's D and Fu's F-S parameters. In addition, high values of haplotype diversity ((H) over cap) and low values of nucleotide diversity (pi), combined with a high number of low frequency haplotypes and values of theta(pi)<theta(W), suggested a recent demographic expansion of H. virescens populations in Brazil. This demographic event could be responsible for the low genetic structure currently found; however, haplotypes present uniquely at the same geographic regions and from one specific host plant suggest an initial differentiation among H. virescens populations within Brazil.

Cocaine effects on mouse incentive-learning and human addiction are linked to alpha 2 subunit-containing GABA(A) receptors

Because GABA(A) receptors containing alpha 2 subunits are highly represented in areas of the brain, such as nucleus accumbens (NAcc), frontal cortex, and amygdala, regions intimately involved in signaling motivation and reward, we hypothesized that manipulations of this receptor subtype would influence processing of rewards. Voltage-clamp recordings from NAcc medium spiny neurons of mice with alpha 2 gene deletion showed reduced synaptic GABA(A) receptor-mediated responses. Behaviorally, the deletion abolished cocaine`s ability to potentiate behaviors conditioned to rewards (conditioned reinforcement), and to support behavioral sensitization. In mice with a point mutation in the benzodiazepine binding pocket of alpha 2-GABA(A) receptors (alpha 2H101R), GABAergic neurotransmission in medium spiny neurons was identical to that of WT (i.e., the mutation was silent), but importantly, receptor function was now facilitated by the atypical benzodiazepine Ro 15-4513 (ethyl 8-amido-5,6-dihydro-5-methyl-6-oxo-4H-imidazo [1,5-a] [1,4] benzodiazepine-3-carboxylate). In alpha 2H101R, but not WT mice, Ro 15-4513 administered directly into the NAcc-stimulated locomotor activity, and when given systemically and repeatedly, induced behavioral sensitization. These data indicate that activation of alpha 2-GABA(A) receptors (most likely in NAcc) is both necessary and sufficient for behavioral sensitization. Consistent with a role of these receptors in addiction, we found specific markers and haplotypes of the GABRA2 gene to be associated with human cocaine addiction.

Calorie restriction increases cerebral mitochondrial respiratory capacity in a NO center dot-mediated mechanism: Impact on neuronal survival

Calorie restriction (CR) enhances animal life span and prevents age-related diseases, including neurological decline. Recent evidence suggests that a mechanism involved in CR-induced life-span extension is NO-stimulated mitochondrial biogenesis. We examine here the effects of CR on brain mitochondrial content. CR increased eNOS and nNOS and the content of mitochondria] proteins (cytochrome c oxidase, citrate synthase, and mitofusin) in the brain. Furthermore, we established an in vitro system to study the neurological effects of CR using serum extracted from animals on this diet. In cultured neurons, CR serum enhanced nNOS expression and increased levels of nitrite (a NO product). CR serum also enhanced the levels of cytochrome c oxidase and increased citrate synthase activity and respiratory rates in neurons. CR serum effects were inhibited by L-NAME and mimicked by the NO donor SNAP. Furthermore, both CR sera and SNAP were capable of improving neuronal survival. Overall, our results indicate that CR increases mitochondrial biogenesis in a NO-mediated manner, resulting in enhanced reserve respiratory capacity and improved survival in neurons. (C) 2012 Elsevier Inc. All rights reserved.

(+)alpha-Tocopheryl succinate inhibits the mitochondrial respiratory chain complex I and is as effective as arsenic trioxide or ATRA against acute promyelocytic leukemia in vivo

The vitamin E derivative (+)alpha-tocopheryl succinate (alpha-TOS) exerts pro-apoptotic effects in a wide range of tumors and is well tolerated by normal tissues. Previous studies point to a mitochondrial involvement in the action mechanism; however, the early steps have not been fully elucidated. In a model of acute promyelocytic leukemia (APL) derived from hCG-PML-RAR alpha transgenic mice, we demonstrated that alpha-TOS is as effective as arsenic trioxide or all-trans retinoic acid, the current gold standards of therapy. We also demonstrated that alpha-TOS induces an early dissipation of the mitochondrial membrane potential in APL cells and studies with isolated mitochondria revealed that this action may result from the inhibition of mitochondrial respiratory chain complex I. Moreover, alpha-TOS promoted accumulation of reactive oxygen species hours before mitochondrial cytochrome c release and caspases activation. Therefore, an in vivo antileukemic action and a novel mitochondrial target were revealed for alpha-TOS, as well as mitochondrial respiratory complex I was highlighted as potential target for anticancer therapy. Leukemia (2012) 26, 451-460; doi:10.1038/leu.2011.216; published online 26 August 2011

Tamoxifen Subcellular Localization; Observation of Cell-Specific Cytotoxicity Enhancement by Inhibition of Mitochondrial ETC Complexes I and III

Recently, a nongenomic cytotoxic component of the chemotherapeutic agent tamoxifen (TAM) has been identified that predominantly triggers mitochondrial events. The present study delineates the intracellular fate of TAM and studies its interaction with a spectrum of cell homeostasis modulators primarily relevant to mitochondria. The subcellular localization of TAM was assessed by confocal fluorescence microscopy. The effect of the modulators on TAM cytotoxicity was assessed by standard MTT assays. Our findings show that in estrogen receptor positive MCF7 breast adenocarcinoma cells and DU145 human prostate cancer cells, TAM largely accumulates in the mitochondria and endoplasmic reticulum, but not lysosomes. Our results further demonstrate that in MCF7, but not in DU145 cells, mitochondrial electron transport chain complex I and III inhibitors exacerbate TAM toxicity with an order of potency of myxothiazol = stigmatellin > rotenone > antimycin A, suggesting a cell-specific cytotoxic interplay between mitochondrial complex I and III function and TAM action.

Expression of NADPH Oxidase by Trophoblast Cells: Potential Implications for the Postimplanting Mouse Embryo

Cytochemical localization of hydrogen peroxide-generating sites suggests NADPH (nicotinamide adenine dinucleotide 3-phosphate [ reduced form]) oxidase expression at the maternal-fetal interface. To explore this possibility, we have characterized the expression and activity of the NADPH oxidase complex in trophoblast cells during the postimplantation period. Implantation sites and ectoplacental cones (EPCs) from 7.5-gestational day embryos from CD1 mice were used as a source for expression analyses of NADPH oxidase catalytic and regulatory subunits. EPCs grown in primary culture were used to investigate the production of superoxide anion through dihydroxyethidium oxidation in confocal microscopy and immunohistochemical assays. NADPH subunits Cybb (gp91phox), Cyba (p22phox), Ncf4 (p40phox), Ncf1 (p47phox), Ncf2 (p67phox), and Rac1 were expressed by trophoblast cells. The fundamental subunits of membrane CYBB and cytosolic NCF2 were markedly upregulated after phorbol-12-myristate-13-acetate (PMA) treatment, as detected by quantitative real-time PCR, Western blotting, and immunohistochemistry. Fluorescence microscopy imaging showed colocalization of cytosolic and plasma membrane NADPH oxidase subunits mainly after PMA treatment, suggesting assembly of the complex after enzyme activation. Cultured EPCs produced superoxide in a NADPH-dependent manner, associating the NADPH oxidase-mediated superoxide production with postimplantation trophoblast physiology. NADPH-oxidase cDNA subunit sequencing showed a high degree of homology between the trophoblast and neutrophil isoforms of the oxidase, emphasizing a putative role for reactive oxygen species production in phagocytic activity and innate immune responses.