Big, bad, and beautiful: phylogenetic relationships of the horned frogs (Anura: Ceratophryidae)

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

The Mycobacterium leprae hsp65 displays proteolytic activity. Mutagenesis studies indicate that the M. leprae hsp65 Proteolytic activity is catalytically related to the HslVU protease?

The present study reports, for the first time, that the recombinant hsp65 from Mycobacterium leprae (chaperonin 2) displays a proteolytic activity toward oligopeptides. The M. leprae hsp65 proteolytic activity revealed a trypsin-like specificity toward quenched fluorescence peptides derived from dynorphins. When other peptide substrates were used (β-endorphin, neurotensin, and angiotensin I), the predominant peptide bond cleavages also involved basic amino acids in P 1, although, to a minor extent, the hydrolysis involving hydrophobic and neutral amino acids (G and F) was also observed. The amino acid sequence alignment of the M. leprae hsp65 with Escherichia coli Hs1VU protease suggested two putative threonine catalytic groups, one in the N-domain (T 136, K 168, and Y 264) and the other in the C-domain (T 375, K 409, and S 502). Mutagenesis studies showed that the replacement of K 409 by A caused a complete loss of the proteolytic activity, whereas the mutation of K 168 to A resulted in a 25% loss. These results strongly suggest that the amino acid residues T 375, K 409, and S 502 at the C-domain form the catalytic group that carries out the main proteolytic activity of the M. leprae hsp65. The possible pathophysiological implications of the proteolytic activity of the M. leprae hsp65 are now under investigation in our laboratory.

Is Drosera meristocaulis a pygmy sundew? evidence of a long-distance dispersal between Western Australia and northern South America

Background and aims South America and Oceania possess numerous floristic similarities, often confirmed by morphological and molecular data. The carnivorous Drosera meristocaulis (Droseraceae), endemic to the Neblina highlands of northern South America, was known to share morphological characters with the pygmy sundews of Drosera sect. Bryastrum, which are endemic to Australia and New Zealand. The inclusion of D. meristocaulis in a molecular phylogenetic analysis may clarify its systematic position and offer an opportunity to investigate character evolution in Droseraceae and phylogeographic patterns between South America and Oceania. Methods Drosera meristocaulis was included in a molecular phylogenetic analysis of Droseraceae, using nuclear internal transcribed spacer (ITS) and plastid rbcL and rps16 sequence data. Pollen of D. meristocaulis was studied using light microscopy and scanning electron microscopy techniques, and the karyotype was inferred from root tip meristem. Key Results The phylogenetic inferences (maximum parsimony, maximum likelihood and Bayesian approaches) substantiate with high statistical support the inclusion of sect. Meristocaulis and its single species, D. meristocaulis, within the Australian Drosera clade, sister to a group comprising species of sect. Bryastrum. A chromosome number of 2n = approx. 32–36 supports the phylogenetic position within the Australian clade. The undivided styles, conspicuous large setuous stipules, a cryptocotylar (hypogaeous) germination pattern and pollen tetrads with aperture of intermediate type 7–8 are key morphological traits shared between D. meristocaulis and pygmy sundews of sect. Bryastrum from Australia and New Zealand. Conclusions The multidisciplinary approach adopted in this study (using morphological, palynological, cytotaxonomic and molecular phylogenetic data) enabled us to elucidate the relationships of the thus far unplaced taxon D. meristocaulis. Long-distance dispersal between southwestern Oceania and northern South America is the most likely scenario to explain the phylogeographic pattern revealed.

Filogeografia do complexo Ischnocnema lactea e Ischnocnema holti (Anura, Brachycephalidae), sudeste do Brasil

Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)

Molecular support for the establishment of the new genus Laurenciella within the Laurencia complex (Ceramiales, Rhodophyta)

Currently, five genera are assigned to red seaweeds of the Laurencia complex worldwide: Chondrophycus, Laurencia s.s., Osmundea, Palisada and Yuzurua. The genera are segregated on the basis of morphological characters, especially the reproductive traits, and molecular sequences of the plastid-encoded gene rbcL. Four of the genera have been resolved as monophyletic, but not Laurencia s.s. In this study based on an rbcL gene phylogeny we show the presence of a sixth lineage within the Laurencia complex, viz., Laurencia marilzae plus two unidentified species of Laurencia from Brazil. The phylogenetic position of this group, combined with the high genetic divergence from Laurencia s.s. (8.2-11%), strongly support the establishment of a sixth genus for the complex, proposed here as Laurenciella gen. nov. This new taxon differs from Laurencia s.s. and from the other genera of the complex by molecular sequence data, but is indistinguishable from Laurencia s.s. by the usual morphological features.

3-Hydroxypropionic Acid as an Antibacterial Agent from Endophytic Fungi Diaporthe phaseolorum

Endophytic fungi are considered a rich source of active compounds resulting from their secondary metabolism. Fungi from marine environment grow in a habitat with unique conditions that can contribute to the activation of metabolic pathways of synthesis of different unknown molecules. The production of these compounds may support the adaptation and survival of the fungi in the marine ecosystem. Mangroves are ecosystems situated between land and sea. They are frequently found in tropical and subtropical areas and enclose approximately 18.1 million hectares of the planet. The great biodiversity found in these ecosystems shows the importance of researching them, including studies regarding new compounds derived from the endophytic fungi that inhabit these ecosystems. 3-hydroxypropionic acid (3-HPA) has been isolated from the mangrove endophytic fungus Diaporthe phaseolorum, which was obtained from branches of Laguncularia racemosa. The structure of this compound was elucidated by spectroscopic methods, mainly 1D and 2D NMR. In bioassays, 3-HPA showed antimicrobial activities against both Staphylococcus aureus and Salmonella typhi. The structure of this antibiotic was modified by the chemical reaction of Fischer-Speier esterification to evaluate the biologic activity of its chemical analog. The esterified product, 3-hydroxypropanoic ethyl ester, did not exhibit antibiotic activity, suggesting that the free carboxylic acid group is important to the pharmacological activity. The antibiotic-producing strain was identified with internal transcribed spacer sequence data. To the best of our knowledge, this is the first report of antibacterial activity by 3-HPA against the growth of medically important pathogens.

Structural and Functional Insights from the Metagenome of an Acidic Hot Spring Microbial Planktonic Community in the Colombian Andes

A taxonomic and annotated functional description of microbial life was deduced from 53 Mb of metagenomic sequence retrieved from a planktonic fraction of the Neotropical high Andean (3,973 meters above sea level) acidic hot spring El Coquito (EC). A classification of unassembled metagenomic reads using different databases showed a high proportion of Gammaproteobacteria and Alphaproteobacteria (in total read affiliation), and through taxonomic affiliation of 16S rRNA gene fragments we observed the presence of Proteobacteria, micro-algae chloroplast and Firmicutes. Reads mapped against the genomes Acidiphilium cryptum JF-5, Legionella pneumophila str. Corby and Acidithiobacillus caldus revealed the presence of transposase-like sequences, potentially involved in horizontal gene transfer. Functional annotation and hierarchical comparison with different datasets obtained by pyrosequencing in different ecosystems showed that the microbial community also contained extensive DNA repair systems, possibly to cope with ultraviolet radiation at such high altitudes. Analysis of genes involved in the nitrogen cycle indicated the presence of dissimilatory nitrate reduction to N2 (narGHI, nirS, norBCDQ and nosZ), associated with Proteobacteria-like sequences. Genes involved in the sulfur cycle (cysDN, cysNC and aprA) indicated adenylsulfate and sulfite production that were affiliated to several bacterial species. In summary, metagenomic sequence data provided insight regarding the structure and possible functions of this hot spring microbial community, describing some groups potentially involved in the nitrogen and sulfur cycling in this environment. Citation: Jimenez DJ, Andreote FD, Chaves D, Montana JS, Osorio-Forero C, et al. (2012) Structural and Functional Insights from the Metagenome of an Acidic Hot Spring Microbial Planktonic Community in the Colombian Andes. PLoS ONE 7(12): e52069. doi:10.1371/journal.pone.0052069

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.

First Identification of Culex flavivirus (Flaviviridae) in Brazil

Introduction: Culex flavivirus (CxFV) was first isolated in 2007 from Culex pipiens in Japan and then identified in several other countries. Characterization of the CxFV showed that all strains are related to the cell fusing agent virus. In this manuscript we report the first identification of CxFV in South America. Material and Methods: We have collected Culex sp. mosquitoes using BG-Sentinel traps and manual aspirators. They were pooled according to genus, species, sex and location. Viral RNA was extracted and multiplex nested PCR was performed to test the presence of Flavivirus. The positive samples were isolated in C6/36 cells and sequenced for phylogenetic analyses. Results: 265 female Culex mosquitoes pooled in 83 pools were tested with specific CxFV, Saint Louis encephalitis virus (SLEV) and West Nile virus (WNV) primers. Our sequence data indicated maximum sequence similarity of 97% with CxFV. Discussion: In this study we report the circulation of CxFV in an urban setting where SLEV had previously caused an outbreak. In terms of public health, this is an important finding due to the assumption that the previous exposition of mosquitoes to CxFV might lessen the susceptibility of these mosquitoes to other flaviviruses. Copyright (C) 2012 S. Karger AG, Basel

MRKAd5 HIV-1 Gag/Pol/Nef Vaccine-Induced T-Cell Responses Inadequately Predict Distance of Breakthrough HIV-1 Sequences to the Vaccine or Viral Load

Background: The sieve analysis for the Step trial found evidence that breakthrough HIV-1 sequences for MRKAd5/HIV-1 Gag/Pol/Nef vaccine recipients were more divergent from the vaccine insert than placebo sequences in regions with predicted epitopes. We linked the viral sequence data with immune response and acute viral load data to explore mechanisms for and consequences of the observed sieve effect. Methods: Ninety-one male participants (37 placebo and 54 vaccine recipients) were included; viral sequences were obtained at the time of HIV-1 diagnosis. T-cell responses were measured 4 weeks post-second vaccination and at the first or second week post-diagnosis. Acute viral load was obtained at RNA-positive and antibody-negative visits. Findings: Vaccine recipients had a greater magnitude of post-infection CD8+ T cell response than placebo recipients (median 1.68% vs 1.18%; p = 0.04) and greater breadth of post-infection response (median 4.5 vs 2; p = 0.06). Viral sequences for vaccine recipients were marginally more divergent from the insert than placebo sequences in regions of Nef targeted by pre-infection immune responses (p = 0.04; Pol p = 0.13; Gag p = 0.89). Magnitude and breadth of pre-infection responses did not correlate with distance of the viral sequence to the insert (p. 0.50). Acute log viral load trended lower in vaccine versus placebo recipients (estimated mean 4.7 vs 5.1) but the difference was not significant (p = 0.27). Neither was acute viral load associated with distance of the viral sequence to the insert (p>0.30). Interpretation: Despite evidence of anamnestic responses, the sieve effect was not well explained by available measures of T-cell immunogenicity. Sequence divergence from the vaccine was not significantly associated with acute viral load. While point estimates suggested weak vaccine suppression of viral load, the result was not significant and more viral load data would be needed to detect suppression.

Climatic oscillations shape the phylogeographical structure of Atlantic Forest fire-eye antbirds (Aves: Thamnophilidae)

In the present study, mitochondrial (mt)DNA sequence data were used to examine the genetic structure of fire-eye antbirds (genus Pyriglena) along the Atlantic Forest and the predictions derived from the river hypothesis and from a Last Glacial Maximum Pleistocene refuge paleomodel were compared to explain the patterns of genetic variation observed in these populations. A total of 266 individuals from 45 populations were sampled over a latitudinal transect and a number of phylogeographical and population genetics analytical approaches were employed to address these questions. The pattern of mtDNA variation observed in fire-eye antbirds provides little support for the view that populations were isolated by the modern course of major Atlantic Forest rivers. Instead, the data provide stronger support for the predictions of the refuge model. These results add to the mounting evidence that climatic oscillations appear to have played a substantial role in shaping the phylogeographical structure and possibly the diversification of many taxa in this region. However, the results also illustrate the potential for more complex climatic history and historical changes in the geographical distribution of Atlantic Forest than envisioned by the refuge model. (c) 2012 The Linnean Society of London, Biological Journal of the Linnean Society, 2012, 105, 900824.

Pollination systems in Pogonieae (Orchidaceae: Vanilloideae): A hypothesis of evolution among reward and rewardless flowers

The tribe Pogonieae of Vanilloideae (Orchidaceae) consists of six genera, including Pogoniopsis, a mycoheterotrophic taxon with morphological characteristics distinct from the remaining of the tribe. A hypothesis about the phylogeny of the tribe was inferred, involving all currently recognized genera, based on isolated and combined sequence data of 5.8S, 18S and 26S (nrDNA) regions using parsimony and Bayesian analyses. Phylogenetic analyses show that inclusion of Pogoniopsis turns the tribe Pogonieae paraphyletic. All analyses reveal that Pogoniopsis is closely related to members of Epidendroideae. The pantropical Vanilla is monophyletic if Dictyophyllaria is assumed as synonym of Vanilla. Members of Pogonieae are pollinated by several groups of solitary and social bees, two pollination systems being recognized: reward-producing and deceptive. The molecular phylogeny suggests that ancestrals related to Pogonieae gave rise to two evolutionary lines: a tropical one with reward production of flowers, and a predominantly temperate regions invading line with deceptive flowers. Reward-producing flowers characterize the South and Central American clade (=Cleistes), while deceptive pollination is prominent in the clade that includes North American-Asiatic taxa plus the Amazonian genus Duckeella. (C) 2012 Elsevier GmbH. All rights reserved.