Phylogeography of the cactophilic species Drosophila gouveai: demographic events and divergence timing in dry vegetation enclaves in eastern Brazil

Aim  The aim of this study was to assess the causal mechanisms underlying populational subdivision in Drosophila gouveai , a cactophilic species associated with xeric vegetation enclaves in eastern Brazil. A secondary aim was to investigate the genetic effects of Pleistocene climatic fluctuations on these environments.
Location  Dry vegetation enclaves within the limits of the Cerrado domain in eastern Brazil.
Methods  We determined the mitochondrial DNA haplotypes of 55 individuals (representing 12 populations) based on sequence data of a 483-bp fragment from the cytochrome c oxidase subunit II (COII) gene. Phylogenetic and coalescent analyses were used to test for the occurrence of demographic events and to infer the time of divergence amongst genetically independent groups.
Results  Our analyses revealed the existence of two divergent subclades (G1 and G2) plus an introgressed clade restricted to the southernmost range of D. gouveai . Subclades G1 and G2 displayed genetic footprints of range expansion and segregated geographical distributions in south-eastern and some central highland regions, east and west of the Paraná River valley. Molecular dating indicated that the main demographic and diversification events occurred in the late to middle Pleistocene.
Main conclusions  The phylogeographical and genetic patterns observed for D. gouveai in this study are consistent with changes in the distribution of dry vegetation in eastern Brazil. All of the estimates obtained by molecular dating indicate that range expansion and isolation pre-dated the Last Glacial Maximum, occurring during the late to middle Pleistocene, and were probably triggered by climatic changes during the Pleistocene. The current patchy geographical distribution and population subdivision in D. gouveai is apparently closely linked to these past events.     

Predicting Carriers of Ongoing Selective Sweeps without Knowledge of the Favored Allele

Methods for detecting the genomic signatures of natural selection have been heavily studied, and they have been successful in identifying many selective sweeps. For most of these sweeps, the favored allele remains unknown, making it difficult to distinguish carriers of the sweep from non-carriers. In an ongoing selective sweep, carriers of the favored allele are likely to contain a future most recent common ancestor. Therefore, identifying them may prove useful in predicting the evolutionary trajectory—for example, in contexts involving drug-resistant pathogen strains or cancer subclones. The main contribution of this paper is the development and analysis of a new statistic, the Haplotype Allele Frequency (HAF) score. The HAF score, assigned to individual haplotypes in a sample, naturally captures many of the properties shared by haplotypes carrying a favored allele. We provide a theoretical framework for computing expected HAF scores under different evolutionary scenarios, and we validate the theoretical predictions with simulations. As an application of HAF score computations, we develop an algorithm (PreCIOSS: Predicting Carriers of Ongoing Selective Sweeps) to identify carriers of the favored allele in selective sweeps, and we demonstrate its power on simulations of both hard and soft sweeps, as well as on data from well-known sweeps in human populations.     

Culture of chondrocytes in medium supplemented with fetal calf serum or a serum substitute: Ultroser G

Fetal calf serum and a serum substitute, Ultroser G, were compared for their effects on the growth curves, clonal growth and cell cycle progression of rabbit chondrocytes in primary culture and during at least three cell passages and included a screen for the maintenance of cartilage-like differentiation i.e. the presence of type II collagen. Proliferation was also compared with another serum substitute, Nu-Serum. Ultroser G is shown to be equivalent to fetal calf serum as far as chondrocyte proliferation is concerned, clonal growth is improved and biosynthesis of type II collagen is maintained in primary culture.     

Liposome-mediated DNA transfer in eukaryotic cells: Dependence of the transfer efficiency upon the type of liposomes used and the host cell cycle stage

The pBR322 plasmid containing the sequence encoding β-lactamase, the enzyme conferring resistance to ampicillin, was encapsulated in liposomes of different phospholipid composition and incubated with synchronized cells. In mitotic cells as compared to cells synchronized in G₁, twice as many exogeneous DNA molecules were found associated with the cell nuclear DNA, when fluid, neutral liposomes were used. These liposomes are taken up by the cells mainly via endocytosis. When fluid, negatively charged liposomes were used as carriers about the same number of exogeneous DNA molecules were found associated with the nuclear DNA both in mitotic and in G₁-synchronized cells. The efficiency for gene transfer of liposomes entering the cells by different mechanisms was further studied and expressed both by the fraction of the radioactive plasmid associated with the nuclear DNA and by the level of the β-lactamase activity detected in the transfected cells. It appears that liposomes entering the cells mainly via an energy-dependent mechanism are more efficient for this type of DNA transfer.     

Properties of succinylated wheat-germ agglutinin.

The physicochemical and binding properties of succinylated wheat germ agglutinin are described in comparison with these of unmodified wheat germ agglutinin. Succinylated wheat germ agglutinin is an acidic protein with a pI of 4.0 +/- 0.2 while the native lectin is basic, pI of 8.5. The solubility of succinylated wheat germ agglutinin is about 100 times higher than that of the unmodified lectin at neutral pH. Both lectins are dimeric at pH down to 5, and the dissociation occurs at pH lower than 4.5. The binding of oligosaccharides of N-acetylglucosamine to both lectins is very similar on the basis of fluorescence and phosphorescence studies. The minimal concentration required to agglutinate rabbit red blood cells is about 2 microgram/ml with both lectins and the concentrations of N-acetylglucosamine and di-N-acetylchitobiose which inhibit agglutination are similar with both lectins. The number of succinylated wheat germ agglutinin molecules bound to the surface of mouse thymocytes was ten times lower than that of the unmodified lectin although the apparent binding constant was only slightly different between the two lectins. The dramatic decrease of the apparent number of cell surface receptors upon succinylation of the lectin is discussed on the basis of the decrease of the isoelectric point and of the acidic properties of the cell surface.     

Homology Modeling and Molecular Docking Studies of Glutamate Dehydrogenase (GDH) from Cyanobacterium Synechocystis sp. PCC 6803

Glutamate dehydrogenase (GDH), which is present in most bacteria and eukaryotes’ mitochondria, plays an important role in amino acid metabolism. In g     

In silico design and in vitro expression of novel multiepitope DNA constructs based on HIV-1 proteins and Hsp70 T-cell epitopes

Objectives

Epitope-driven vaccines carrying highly conserved and immunodominant epitopes have emerged as promising approaches to overcome human immunodeficiency virus-1 (HIV-1) infection.

Methods

Two multiepitope DNA constructs encoding T cell epitopes from HIV-1 Gag, Pol, Env, Nef and Rev proteins alone and/or linked to the immunogenic epitopes derived from heat shock protein 70 (Hsp70) as an immunostimulatory agent were designed. In silico analyses were applied including MHC-I and MHC-II binding, MHC-I immunogenicity and antigen processing, population coverage, conservancy, allergenicity, toxicity and hemotoxicity. The peptide-MHC-I/MHC-II molecular docking and cytokine production analyses were carried out for predicted epitopes. The selected highly immunogenic T-cell epitopes were then used to design two multiepitope fusion constructs. Next, prediction of the physicochemical and structural properties, B cell epitopes, and constructs-toll-like receptors (TLRs) molecular docking were performed for each construct. Finally, the eukaryotic expression plasmids harboring totally 12 cytotoxic T Lymphocyte (CTL) and 10 helper T lymphocytes (HTL) epitopes from HIV-1 proteins (i.e., pEGFP-N1-gag-pol-env-nef-rev), and linked to 2 CTL and 2 HTL epitopes from Hsp70 (i.e., pEGFP-N1-hsp70-gag-pol-env-nef-rev) were generated and transfected into HEK-293 T cells for evaluating the percentage of multiepitope peptides expression using flow cytometry and western blotting.

Results

The designed DNA constructs could be successfully expressed in mammalian cells. The expression rates of Gag-Pol-Env-Nef-Rev-GFP and Hsp70-Gag-Pol-Env-Nef-Rev-GFP were about 56–60% as the bands of?~?63 and?~?72 kDa confirmed in western blotting, respectively.

Conclusion

The combined in silico/in vitro methods indicated two multiepitope constructs can be produced and used as probable effective immunogens for HIV-1 vaccine development.

     

Non-native prey species supporting fish assemblage biomass in a Neotropical reservoir

The present study aimed to investigate the role of four non-native invertebrates in supporting fish biomass as well as their influence on the carbon flow into the Volta Grande reservoir food web. The fish samples were carried out quarterly between October 2015 and July 2016 using gillnets. At the sampled sites, four non-native invertebrates (golden mussel, Asian clam, trumpet snail and Amazonian prawn), which are potential prey for fish in the Volta Grande reservoir, were collected by targeted sampling using a Petersen-type bottom dredger and semi-circular sieves. The gut contents of the fish were collected and analyzed under stereoscope, and samples of muscle tissue of these fish, as well as the four non-native invertebrate species sampled, were submitted for isotopic analysis. Results obtained by the present study, by both gut content and stable isotopic analyses, pointed to a trophic structure where non-native species represent not only a strong component of the fish community, but also their main carbon source. Based on gut contents and isotopic mixing models, we found that together, non-native prey are essential carbon sources for the fish fauna, fuelling more than 40.0% of the biomass in four dominant fish species. The consumption rate of non-native bivalves by the native omnivorous fish Leporinus friderici suggested these filter-feeding organisms potentially constitute an important trophic connection between littoral consumers and pelagic energy sources. In addition, non-native prey were also prominent carbon sources for non-native fish, fuelling more than half of the biomass in peacock bass and silver croaker, suggesting these prey have a fundamental role in maintaining non-native fish populations in this system. Our results may help to understand fundamental ecological issues bringing to light the extent to which these new combinations of species influence the energy flow and ecosystem properties of the Volta Grande reservoir.

     

A finite element model of stress-mediated vascular adaptation: application to abdominal aortic aneurysms

Despite rapid expansion of our knowledge of vascular adaptation, developing patient-specific models of diseased arteries is still an open problem. In this study, we extend existing finite element models of stress-mediated growth and remodelling of arteries to incorporate a medical image-based geometry of a healthy aorta and, then, simulate abdominal aortic aneurysm. Degradation of elastin initiates a local dilatation of the aorta while stress-mediated turnover of collagen and smooth muscle compensates the loss of elastin. Stress distributions and expansion rates during the aneurysm growth are studied for multiple spatial distribution functions of elastin degradation and kinetic parameters. Temporal variations of the degradation function are also investigated with either direct time-dependent degradation or stretch-induced degradation as possible biochemical and biomechanical mechanisms for elastin degradation. The results show that this computational model has the capability to capture the complexities of aneurysm progression due to variations of geometry, extent of damage and stress-mediated turnover as a step towards patient-specific modelling.