Phylogeography of the common vampire bat (Desmodus rotundus): Marked population structure, Neotropical Pleistocene vicariance and incongruence between nuclear and mtDNA markers

Background  

The common vampire bat Desmodus rotundus is an excellent model organism for studying ecological vicariance in the Neotropics due to its broad geographic range and its preference for forested areas as roosting sites. With the objective of testing for Pleistocene ecological vicariance, we sequenced a mitocondrial DNA (mtDNA) marker and two nuclear markers (RAG2 and DRB) to try to understand how Pleistocene glaciations affected the distribution of intraspecific lineages in this bat.     

Incidence and Risk Factors for Severe Bacterial Infections in People Living with HIV. ANRS CO3 Aquitaine Cohort, 2000–2012

Severe non-AIDS bacterial infections (SBI) are the leading cause of hospital admissions among people living with HIV (PLHIV) in industrialized countries. We aimed to estimate the incidence of SBI and their risk factors in a large prospective cohort of PLHIV patients over a 13-year period in France. Patients followed up in the ANRS CO3 Aquitaine cohort between 2000 and 2012 were eligible; SBI was defined as a clinical diagnosis associated with hospitalization of ≥48 hours or death. Survival analysis was conducted to identify risk factors for SBI.Total follow-up duration was 39,256 person-years [PY] (31,370 PY on antiretroviral treatment [ART]). The incidence of SBI decreased from 26.7/1000 PY [95% CI: 22.9–30.5] over the period 2000–2002 to 11.9/1000 PY [10.1–13.8] in 2009–2012 (p <0.0001). Factors independently associated to increased risk of SBI were: plasma HIVRNA>50 copies/mL (Hazard Ratio [HR] = 5.1, 95% Confidence Interval: 4.2–6.2), CD4 count <500 cells/mm³ and CD4/CD8 ratio <0.8 (with a dose-response relationship for both markers), history of cancer (HR = 1.4 [1.0–1.9]), AIDS stage (HR = 1.7 [1.3–2.1]) and HCV coinfection (HR = 1.4, [1.1–1.6]). HIV-positive patients with diabetes were more prone to SBI (HR = 1.6 [0.9–2.6]). Incidence of SBI decreased over a 13-year period due to the improvement in the virological and immune status of PLHIV on ART. Risk factors for SBI include low CD4 count and detectable HIV RNA, but also CD4/CD8 ratio, HCV coinfection, history of cancer and diabetes, comorbid conditions that have been frequent among PLHIV in recent years.     

Activation of G551D CFTR channel with MPB-91: regulation by ATPase activity and phosphorylation

We have designed and synthesizedbenzo[c]quinolizinium derivatives and evaluated their effects on theactivity of G551D cystic fibrosis transmembrane conductance regulator(CFTR) expressed in Chinese hamster ovary and Fisher ratthyroid cells. We demonstrated, using iodide efflux, whole cell patchclamp, and short-circuit recordings, that5-butyl-6-hydroxy-10-chlorobenzo[c]quinolizinium chloride (MPB-91)restored the activity of G551D CFTR (EC₅₀ = 85 µM)and activated CFTR in Calu-3 cells (EC₅₀ = 47 µM).MPB-91 has no effect on the ATPase activity of wild-type and G551DNBD1/R/GST fusion proteins or on the ATPase, GTPase, and adenylatekinase activities of purified NBD2. The activation of CFTR by MPB-91 isindependent of phosphorylation because 1) kinase inhibitors have no effect and 2) the compound still activated CFTRhaving 10 mutated protein kinase A sites (10SA-CFTR). The newpharmacological agent MPB-91 may be an important candidate drug toameliorate the ion transport defect associated with CF and to point outa new pathway to modulate CFTR activity.

     

Rapid generation of long tandem DNA repeat arrays by homologous recombination in yeast to study their function in mammalian genomes

We describe here a method to rapidly convert any desirable DNA fragment, as small as 100 bp, into long tandem DNA arrays up to 140 kb in size that are inserted into a microbe vector. This method includes rolling-circle phi29 amplification (RCA) of the sequence in vitro and assembly of the RCA products in vivo by homologous recombination in the yeast Saccharomyces cerevisiae. The method was successfully used for a functional analysis of centromeric and pericentromeric repeats and construction of new vehicles for gene delivery to mammalian cells. The method may have general application in elucidating the role of tandem repeats in chromosome organization and dynamics. Each cycle of the protocol takes ~ two weeks to complete.     

Evaluating the toxicity of environmental concentrations of waterborne chromium (VI) to a model teleost, Oncorhynchus mykiss: a comparative study of in vivo and in vitro

Toxic effects of environmental concentrations (50, 100, and 200μg/L) of waterborne chromium (VI) were evaluated in rainbow trout by comparison of in vitro and in vivo assays. Multiple biomarkers were measured including oxidative stress indices and antioxidant response parameters in liver and brain, as well as Na(+)-K(+)-ATPase in gill. Superoxide dismutase (SOD) and glutathione reductase (GR) activities were significantly induced (1.54-fold and 1.37-fold, respectively) in fish brain in vivo, but no significant differences were observed in any other biomarker or in vivo test group. Oxidative stress was apparent in vitro as significantly higher levels of oxidative indices, with the highest induction of TBARS and CP found in brain at 200μg/L Cr(VI) (2.41-fold and 1.95-fold, respectively), and SOD and GR activities and reduced glutathione in brain were significantly inhibited (65%, 44%, and 36%, respectively). In vitro Na(+)-K(+)-ATPase activity in gill was also significantly inhibited at concentrations of 100 and 200μg/L (69% and 45%, respectively). Short-term exposure to environmental concentrations of Cr(VI) does not therefore evoke marked effects in fish in vivo. Based on the present results, a set of in vitro tests with tissue homogenate can be evoked more remarkable effects by the lower concentrations of Cr(VI) than in vivo, which could provide some useful information and might be a potential alternative approach for monitoring heavy metal pollution in aquatic environments. However, it needs more detailed studies in other area, such as hormonal response or genotoxicity, before these findings could be applied in the field investigation.     

Hepatic proteome sensitivity in rainbow trout after chronically exposed to a human pharmaceutical verapamil

Verapamil (VRP), a cardiovascular pharmaceutical widely distributed and persistent in the aquatic environment, has potential toxicity to fish and other aquatic organisms. However, the molecular mechanisms that lead to these toxic effects are not well known. In the present study, proteomic analysis has been performed to investigate the protein patterns that are differentially expressed in liver of rainbow trout exposed to sublethal concentrations of VRP (0.5, 27.0, and 270 μg/liter) for 42 days. Two-dimensional electrophoresis coupled with MALDI-TOF/TOF mass spectrometry was employed to detect and identify the protein profiles. The analysis revealed that the expression of six hepatic acidic proteins were markedly altered in the treatment groups compared with the control group; three proteins especially were significantly down-regulated in fish exposed to VRP at environmental related concentration (0.5 μg/liter). These results suggested that the VRP induce mechanisms against oxidative stress (glucose-regulated protein 78 and 94 and protein disulfide-isomerase A3) and adaptive changes in ion transference regulation (calreticulin, hyperosmotic glycine-rich protein). Furthermore, for the first time, protein Canopy-1 was found to be significantly down-regulated in fish by chronic exposure to VRP at environmental related levels. Overall, our work supports that fish hepatic proteomics analysis serves as an in vivo model for monitoring the residual pharmaceuticals in aquatic environment and can provide valuable insight into the molecular events in VRP-induced toxicity in fish and other organisms.     

Poly(ethylene oxide), a new slowing agent for protozoa.

The water-soluble, viscoelastic resin Polyox WSR 301), a poly(ethylene oxide) of high molecular weight (approximately 4 million) is introduces as a new slowing agent for protozoa. Generally, as the kinetic viscosity of the resin increased from 0.25% to 1% (w/v), the swimming velocity of Euglena gracilis, Didnium nasutum, Paramecium aurelia, Blepharisma undulans, and Prorodon platyodon decreased. The 1.0% solution had the highest viscosity and decreased velocity more effectively than 1.0% methyl cellulose and Protoslo solutions. The Polyox solutions differed from those of methyl cellulose and Protoslo by having, in addition to viscous drag, an elastic recoil that pulled the protozoa backwards when their swimming efforts stopped. The toxicity of these slowing agents was determined using 10 P. aurelia/test slide preparation. Paramecium numbers decreased in 1.0% methyl cellulose and Protoslo to nearly zero by 24 hr; in Polyox, not only were most these ciliates alive after 24 hr, but many survived for 96 hr and divisions occurred in 0.25% and 0.50% solutions.     

Modulation of antioxidant defence system in brain of rainbow trout (Oncorhynchus mykiss) after chronic carbamazepine treatment

We investigated the effect of long-term exposure to CBZ on the antioxidant system in brain tissue of rainbow trout. Fish were exposed to sublethal concentrations of CBZ (1.0 μg/L, 0.2 mg/L or 2.0 mg/L) for 7, 21, and 42 days. Oxidative stress indices (LPO and CP) and activities of antioxidant enzymes (SOD, CAT, GPx and GR) in fish brain were measured. In addition, non-enzymatic antioxidant (GSH) was determined after 42 days exposure. Carbamazepine exposure at 0.2 mg/L led to significant increases (p < 0.05) of LPO and CP after 42 days and, at 2.0 mg/L, after 21 days. Activities of the antioxidant enzymes SOD, CAT, and GPx in CBZ-treated groups slightly increased during the first period (7 days). However, activities of all measured antioxidant enzymes were significantly inhibited (p < 0.05) at 0.2 mg/L exposure after 42 days and after 21 days at 2.0 mg/L. After 42 days, the content of GSH in fish brain was significantly lower (p < 0.05) in groups exposed to CBZ at 0.2 mg/L and 2.0 mg/L than in other groups. Prolonged exposure to CBZ resulted in excess reactive oxygen species formation, finally resulting in oxidative damage to lipids and proteins and inhibited antioxidant capacities in fish brain. In short, a low level of oxidative stress could induce the adaptive responses of antioxidant enzymes, but long-term exposure to CBZ could lead to serious oxidative damage in fish brain.