Phylogenetic reconstruction using secondary structures of Internal Transcribed Spacer 2 (ITS2, rDNA): finding the molecular and morphological gap in Caribbean gorgonian corals

Background  

Most phylogenetic studies using current methods have focused on primary DNA sequence information. However, RNA secondary structures are particularly useful in systematics because they include characteristics, not found in the primary sequence, that give "morphological" information. Despite the number of recent molecular studies on octocorals, there is no consensus opinion about a region that carries enough phylogenetic resolution to solve intrageneric or close species relationships. Moreover, intrageneric morphological information by itself does not always produce accurate phylogenies; intra-species comparisons can reveal greater differences than intra-generic ones. The search for new phylogenetic approaches, such as by RNA secondary structure analysis, is therefore a priority in octocoral research.     

A simple cryogenic holder for tensile testing of soft biological tissues

To overcome the difficulty of gripping soft biological materials for tensile test, a simple inexpensive cryogenic holder was developed which allows rapid (3 min) preparation of samples. It is made of 6 parts, built in a bakelite cloth, which is an excellent thermal isolant, and is used with rectangular (8x10(-2)x10(-2)x10(-2)m) samples. The holder with the sample inside is completely immersed in liquid nitrogen for 50 s. This duration allows the freezing of the sample ends on a 10(-2)m length and gives a very flat freezing surface throughout the sample cross section. The 6x10(-2)m central part of the sample remained at ambient temperature. Two parts of the holder help maintain the sample until its ends are vertically gripped in the tensile machine thus avoiding any sample deformation during this step. No pressure was applied on the frozen part of the sample by grips of the tensile machine and this avoids breaks in this region. The sample is fixed by adhesion forces (>1 kN) between its frozen parts and 2 pieces of the holder. The procedure has been successfully tested with bovine and salmon muscle samples and results show tensile breaks randomly distributed in the unfrozen region of the samples. Particular attention has been paid to obtain a very flat freezing surface so that the axial strain is equal throughout the sample and therefore any strain-related mechanical parameters can be accurately determined. The dimensions of the holder can be easily modified to fit other sample geometries and can be used with other biological materials.     

Direct and label‐free detection of the human growth hormone in urine by an ultrasensitive bimodal waveguide biosensor

A label‐free interferometric transducer showing a theoretical detection limit for homogeneous sensing of 5 × 10^–8 RIU, being equivalent to a protein mass coverage resolution of 2.8 fg mm^–2, is used to develop a high sensitive biosensor for protein detection. The extreme sensitivity of this transducer combined with a selective bioreceptor layer enables the direct evaluation of the human growth hormone (hGH) in undiluted urine matrix in the 10 pg mL^–1 range.

     

The role of surface charge and hydrophobic interaction in the activation of rat liver adenylate cyclase

Rat liver plasma membranes were incubated either with procaine, lidocaine or tetracaine to study the binding of glucagon to receptors and the responses of adenylate cyclase to glucagon or fluoride. Procaine treatment increased the glucagon and fluoride activation of the cyclase and the stimulation was concentration-dependent; this compound seemed to act at the G/F unit level since changes in the glucagon binding were not observed and the basal activity was not modified. Tetracaine inhibited the adenylate cyclase activity in the order glucagon greater than basal greater than fluoride; it seems that tetracaine acted at the receptor unit level since it reduced the binding affinity. Tetracaine at high concentration (10 mM) also inhibited the fluoride stimulation of the Lubrol PX-solubilized enzyme; apparently the anesthetic acts on the G/F unit and this would indicate the component is still bound to the catalytic unit. The solubilized enzyme is not longer stimulated by procaine. These data suggested that the F component site of the G/F units is in some aspects different to the G component and more resistant to the detergent. The results of this work allowed a clear distinction among the different components of the glucagon-stimulated adenylate cyclase system and showed the importance of surface charge and hydrophobic interactions as regulatory mechanisms.     

Discovery of Phytophthora infestans Genes Expressed in Planta through Mining of cDNA Libraries

Background

Phytophthora infestans (Mont.) de Bary causes late blight of potato and tomato, and has a broad host range within the Solanaceae family. Most studies of the Phytophthora – Solanum pathosystem have focused on gene expression in the host and have not analyzed pathogen gene expression in planta.

Methodology/Principal Findings

We describe in detail an in silico approach to mine ESTs from inoculated host plants deposited in a database in order to identify particular pathogen sequences associated with disease. We identified candidate effector genes through mining of 22,795 ESTs corresponding to P. infestans cDNA libraries in compatible and incompatible interactions with hosts from the Solanaceae family.

Conclusions/Significance

We annotated genes of P. infestans expressed in planta associated with late blight using different approaches and assigned putative functions to 373 out of the 501 sequences found in the P. infestans genome draft, including putative secreted proteins, domains associated with pathogenicity and poorly characterized proteins ideal for further experimental studies. Our study provides a methodology for analyzing cDNA libraries and provides an understanding of the plant – oomycete pathosystems that is independent of the host, condition, or type of sample by identifying genes of the pathogen expressed in planta.     

Hidden among Sea Anemones: The First Comprehensive Phylogenetic Reconstruction of the Order Actiniaria (Cnidaria,Anthozoa, Hexacorallia) Reveals a Novel Group of Hexacorals

Sea anemones (order Actiniaria) are among the most diverse and successful members of the anthozoan subclass Hexacorallia, occupying benthic marine habitats across all depths and latitudes. Actiniaria comprises approximately 1,200 species of solitary and skeleton-less polyps and lacks any anatomical synapomorphy. Although monophyly is anticipated based on higher-level molecular phylogenies of Cnidaria, to date, monophyly has not been explicitly tested and at least some hypotheses on the diversification of Hexacorallia have suggested that actiniarians are para- or poly-phyletic. Published phylogenies have demonstrated the inadequacy of existing morphological-based classifications within Actiniaria. Superfamilial groups and most families and genera that have been rigorously studied are not monophyletic, indicating conflict with the current hierarchical classification. We test the monophyly of Actiniaria using two nuclear and three mitochondrial genes with multiple analytical methods. These analyses are the first to include representatives of all three currently-recognized suborders within Actiniaria. We do not recover Actiniaria as a monophyletic clade: the deep-sea anemone Boloceroides daphneae, previously included within the infraorder Boloceroidaria, is resolved outside of Actiniaria in several of the analyses. We erect a new genus and family for B. daphneae, and rank this taxon incerti ordinis. Based on our comprehensive phylogeny, we propose a new formal higher-level classification for Actiniaria composed of only two suborders, Anenthemonae and Enthemonae. Suborder Anenthemonae includes actiniarians with a unique arrangement of mesenteries (members of Edwardsiidae and former suborder Endocoelantheae). Suborder Enthemonae includes actiniarians with the typical arrangement of mesenteries for actiniarians (members of former suborders Protantheae, Ptychodacteae, and Nynantheae and subgroups therein). We also erect subgroups within these two newly-erected suborders. Although some relationships among these newly-defined groups are still ambiguous, morphological and molecular results are consistent enough to proceed with a new higher-level classification and to discuss the putative functional and evolutionary significance of several morphological attributes within Actiniaria.     

Patterns of Symbiodinium spp. associations within the family Aiptasiidae,a monophyletic lineage of symbiotic of sea anemones (Cnidaria,Actiniaria)

Coral Reefs - Although the symbiotic relationships between dinoflagellates and cnidarians are well recognized, few studies have examined these associations from an evolutionary perspective. This is...     

Genetic diversity of the entomopathogenic fungus Cordyceps tenuipes in forests and butterfly gardens in Quindío,Colombia

Cordyceps tenuipes is an entomopathogenic fungus that infects mostly pupae of several lepidopteran families. In Colombia the species has been reported in non-disturbed tropical rain forests and more recently in butterfly gardens. The aim of this study was to assess the genetic diversity in populations of C. tenuipes present in natural (forests) and artificial (e.g. butterfly gardens) environments in the department of Quindío, Colombia, using three molecular nuclear markers ITS, TEF-1α and RPB1. All the samples evaluated corresponded morphologically and phylogenetically to C. tenuipes. The butterfly garden of Quindio Botanical Garden (QBG) showed the highest genetic diversity among all sampling localities and was very similar to that of its adjacent forest. The Amaranta Butterfly Garden (ABG), located north of QBG, showed lower genetic diversity as well as little genetic differentiation with QBG, consistent with the hypothesis of a pathogen transfer from QBG to ABG. Higher F_ST values were observed for TEF-1α and ITS, revealing genetic differentiation between all demes and the southern forest population. Our research constitutes the first study of the intraspecific diversity of C. tenuipes in Colombia and can serve as the first step in identifying diversity reservoirs and management of epizootic episodes caused by this fungal species.