Exposure to sublethal doses of fipronil and thiacloprid highly increases mortality of honeybees previously infected by Nosema ceranae

BACKGROUND: The honeybee, Apis mellifera, is undergoing a worldwide decline whose origin is still in debate. Studies performed for twenty years suggest that this decline may involve both infectious diseases and exposure to pesticides. Joint action of pathogens and chemicals are known to threaten several organisms but the combined effects of these stressors were poorly investigated in honeybees. Our study was designed to explore the effect of Nosema ceranae infection on honeybee sensitivity to sublethal doses of the insecticides fipronil and thiacloprid. METHODOLOGY/FINDING: Five days after their emergence, honeybees were divided in 6 experimental groups: (i) uninfected controls, (ii) infected with N. ceranae, (iii) uninfected and exposed to fipronil, (iv) uninfected and exposed to thiacloprid, (v) infected with N. ceranae and exposed 10 days post-infection (p.i.) to fipronil, and (vi) infected with N. ceranae and exposed 10 days p.i. to thiacloprid. Honeybee mortality and insecticide consumption were analyzed daily and the intestinal spore content was evaluated 20 days after infection. A significant increase in honeybee mortality was observed when N. ceranae-infected honeybees were exposed to sublethal doses of insecticides. Surprisingly, exposures to fipronil and thiacloprid had opposite effects on microsporidian spore production. Analysis of the honeybee detoxification system 10 days p.i. showed that N. ceranae infection induced an increase in glutathione-S-transferase activity in midgut and fat body but not in 7-ethoxycoumarin-O-deethylase activity. CONCLUSIONS/SIGNIFICANCE: After exposure to sublethal doses of fipronil or thiacloprid a higher mortality was observed in N. ceranae-infected honeybees than in uninfected ones. The synergistic effect of N. ceranae and insecticide on honeybee mortality, however, did not appear strongly linked to a decrease of the insect detoxification system. These data support the hypothesis that the combination of the increasing prevalence of N. ceranae with high pesticide content in beehives may contribute to colony depopulation.     

Why Do Sleeping Nematodes Adopt a Hockey-Stick-Like Posture?

A characteristic posture is considered one of the behavioral hallmarks of sleep, and typically includes functional features such as support for the limbs and shielding of sensory organs. The nematode C. elegans exhibits a sleep-like state during a stage termed lethargus, which precedes ecdysis at the transition between larval stages. A hockey-stick-like posture is commonly observed during lethargus. What might its function be? It was previously noted that during lethargus, C. elegans nematodes abruptly rotate about their longitudinal axis. Plausibly, these “flips” facilitate ecdysis by assisting the disassociation of the old cuticle from the new one. We found that body-posture during lethargus was established using a stereotypical motor program and that body bends during lethargus quiescence were actively maintained. Moreover, flips occurred almost exclusively when the animals exhibited a single body bend, preferentially in the anterior or mid section of the body. We describe a simple biomechanical model that imposes the observed lengths of the longitudinally directed body-wall muscles on an otherwise passive elastic rod. We show that this minimal model is sufficient for generating a rotation about the anterior-posterior body axis. Our analysis suggests that posture during lethargus quiescence may serve a developmental role in facilitating flips and that the control of body wall muscles in anterior and posterior body regions are distinct.     

The membrane proximal external region of the HIV-1 envelope glycoprotein gp41 contributes to the stabilization of the six-helix bundle formed with a matching N' peptide

The HIV-1 envelope glycoprotein gp41 undergoes a sequence of extensive conformational changes while participating in the fusion of the virus with the host cell. Since the discovery of its postfusion conformation, the structure and function of the protease-resistant six-helix bundle (6-HB) have been the subject of extensive investigation. In this work, we describe additional determinants (S528-Q540 and W666-N677) in the fusion peptide proximal region (FP-PR) and the membrane proximal external region (MPER) that stabilize the six-helix bundle and are involved in the interaction of T-20 (FUZEON, an anti-HIV-1 fusion inhibitor drug) with the gp41 FP-PR. Circular dichroism and sedimentation equilibrium measurements indicate that the 1:1 mixture of N' and C' peptides comprising residues A541-T569 and I635-K665 from the gp41 first and second helical repeats, HR1 and HR2, respectively, fail to form a stable six-helix bundle. Triglutamic acid and triarginine tags were added to these N' and C' peptides, respectively, at the termini distant from the FP-PR and the MPER to alter their pI and increase their solubility at pH 3.5. The tagged HR1 and HR2 peptides were elongated by addition of residues S528-Q540 from the FP-PR and residues W666-N677 from the MPER, respectively. A 1:1 complex of the elongated peptides formed a stable six-helix bundle which melted at 60 degrees C. These results underscore the importance of a detailed high-resolution characterization of MPER interactions, the results of which may improve our understanding of the structure-function relationship of gp41 and its role in HIV-1 fusion.     

CD47 ligation induces caspase-independent cell death in chronic lymphocytic leukemia.

Thrombospondin forms a 'molecular bridge' between phagocytic and apoptotic cells through interaction with alphavbeta3/CD36. We report here that engagement of CD47, a newly described thrombospondin receptor, by immobilized monoclonal antibody against CD47 or by thrombospondin induced in all B-cell chronic lymphocytic leukemia clones the cytoplasmic features of apoptosis (cell shrinkage, decrease in mitochondrial transmembrane potential and phosphatidylserine externalization) without the nuclear features (chromatin condensation, appearance of single-stranded DNA, DNA fragmentation and cleavage of poly ADP-ribose polymerase). These cytoplasmic events of apoptosis were not prevented by the addition of caspase inhibitor z-VAD-fmk, or by the presence of survival factors (such as interleukin-4 and gamma interferon) or cell activation. Morphological studies confirmed the integrity of the nucleus and showed swelling of the mitochondria. This caspase-independent death pathway may be relevant to the development of alternate therapeutic strategies in chronic lymphocytic leukemia, which remains an incurable disease.     

Studies designed to increase the stability and antiviral activity (HCMV) of the active benzimidazole nucleoside, TCRB.

The potent activity of 2,5,6-trichloro-1-(beta-D-ribofuranosyl)benzimidazole (TCRB) against Human Cytomegalovirus with the concomitant low cellular toxicity at concentrations that inhibit viral growth prompted considerable interest in this research area. This interest was moderated by the pharmacokinetic studies of TCRB in rats and monkeys that revealed the instability of TCRB in vivo. These studies suggested that the instability was due to a cleavage of the glycosidic bond in vivo which released the heterocycle (2,5,6-trichlorobenzimidazole) into the bloodstream. This prompted us to initiate synthetic studies designed to increase the stability of the glycosidic bond of TCRB and BDCRB. Several synthetic approaches to address this and other problems are presented.     

DPP4 Gene DNA Methylation in the Omentum is Associated With Its Gene Expression and Plasma Lipid Profile in Severe Obesity

Severely obese subjects with the metabolic syndrome (MS) have higher dipeptidyl peptidase‐4 (DPP4) expression in their visceral adipose tissue (VAT) compared to obese individuals without MS. We tested the hypothesis that methylation level of CpG sites in the DPP4 promoter CpG island in VAT was genotype‐dependent and associated with DPP4 mRNA abundance and MS‐related phenotypes. The VAT DNA was extracted in 92 severely obese premenopausal women undergoing biliopancreatic derivation for the treatment of obesity. Women were nondiabetic and none of them used medication to treat MS features. Cytosine methylation rates (%) of 102 CpG sites in the DPP4 CpG island were assessed by pyrosequencing of sodium bisulfite‐treated DNA. Methylation rates were >10% for CpG sites 94–102. Their mean methylation rate (%Meth_94–102) was different between genotypes for DPP4 polymorphisms rs13015258 (P = 0.001), rs17848915 (P = 0.0004), and c.1926 G>A (P = 0.001). The %Meth_94–102 correlated negatively with DPP4 mRNA abundance (r = ?0.25, P < 0.05) and positively with plasma high‐density lipoprotein (HDL) cholesterol concentrations (r = 0.22, P < 0.05), whereas DPP4 mRNA abundance correlated positively with plasma total‐/HDL‐cholesterol ratio (r = 0.25; P < 0.05). In the VAT of nondiabetic severely obese women, genotype‐dependent methylation levels of specific CpG sites in the DPP4 promoter CpG island were associated with DPP4 gene expression and variability in the plasma lipid profile. Higher DPP4 gene expression in VAT and its relationship with the plasma lipid profile may be explained by actually unknown DPP4 biological effect or, to another extent, may also be a marker of VAT inflammation known to be associated with metabolic disturbances.     

Proteome of Aedes aegypti larvae in response to infection by the intracellular parasite Vavraia culicis

We report on the modification of the Aedes aegypti larval proteome following infection by the microsporidian parasite Vavraia culicis. Mosquito larvae were sampled at 5 and 15 days of age to compare the effects of infection when the parasite was in two different developmental stages. Modifications of the host proteome due to the stress of infection were distinguished from those of a more general nature by treatments involving hypoxia. We found that the major reaction to stress was the suppression of particular protein spots. Older (15 days) larvae reacted more strongly to infection by V. culicis (46% of the total number of spots affected; 17% for 5 days larvae), while the strongest reaction of younger (5 days) larvae was to hypoxia for pH range 5-8 and to combined effects of infection and hypoxia for pH range 3-6. MALDI-TOF results indicate that proteins induced or suppressed by infection are involved directly or indirectly in defense against microorganisms. Finally, our MALDI-TOF results suggest that A. aegypti larvae try to control or clear V. culicis infection and also that V. culicis probably impairs the immune defense of this host via arginases-NOS competition.