Shifts in species richness, herbivore specialization, and plant resistance along elevation gradients.

Environmental gradients have been postulated to generate patterns of diversity and diet specialization, in which more stable environments, such as tropical regions, should promote higher diversity and specialization. Using field sampling and phylogenetic analyses of butterfly fauna over an entire alpine region, we show that butterfly specialization (measured as the mean phylogenetic distance between utilized host plants) decreases at higher elevations, alongside a decreasing gradient of plant diversity. Consistent with current hypotheses on the relationship between biodiversity and the strength of species interactions, we experimentally show that a higher level of generalization at high elevations is associated with lower levels of plant resistance: across 16 pairs of plant species, low-elevation plants were more resistant vis-à-vis their congeneric alpine relatives. Thus, the links between diversity, herbivore diet specialization, and plant resistance along an elevation gradient suggest a causal relationship analogous to that hypothesized along latitudinal gradients.

The resistance of seeds of cowpea (Vigna unguiculata) to the cowpea weevil (Callosobruchus maculatus)

Cowpea (Vigna unguiculata) seeds are heavily damaged during storage by the bruchid Callosobruchus maculatus. Seeds of some Nigerian varieties showed a strong resistance to this bruchid. By utilizing biochemical and entomological techniques we were able to rule out the paticipation of proteolytic enzyme (trypsin, chimotrypsin, subtilisin and papain) inhibitors, lectins, and tannins in the resistance mechanisms. Fractionation of the seed meal of a resistant variety suggests that the factor(s) responsible for the effect is (are) concentrate in the globulin fraction.

CYP3A activity measured by the midazolam test is not related to 3435 C >T polymorphism in the multiple drug resistance transporter gene.

A recent study with 69 Japanese liver transplants treated with tacrolimus found that the MDR13435 C >T polymorphism, but not the MDR12677 G >T polymorphism, was associated with differences in the intestinal expression level of CYP3A4 mRNA. In the present study, over 6 h, we measured the kinetics of a 75 microg oral dose of midazolam, a CYP3A substrate, in 21 healthy subjects genotyped for the MDR13435 C >T and 2677 G >T polymorphism. No statistically significant differences were found in the calculated pharmacokinetic parameters between the three 3435 C >T genotypes (TT, CT and CC group, respectively: Cmax (mean +/- SD: 0.30 +/- 0.08 ng/ml, 0.31 +/- 0.09 ng/ml and 0.31 +/- 0.11 ng/ml; Apparent clearance: 122 +/- 29 l/h, 156 +/- 92 l/h and 111 +/- 35 l/h; t1/2: 1.9 +/- 1.1 h, 1.6 +/- 0.90 h and 1.7 +/- 0.7 h). In addition, the 30-min 1'OH midazolam to midazolam ratio, a marker of CYP3A activity, determined in 74 HIV-positive patients before the introduction of antiretroviral treatment, was not significantly different between the three 3435 C >T genotypes (mean ratio +/- SD: 3.65 +/- 2.24, 4.22 +/- 3.49 and 4.24 +/- 2.03, in the TT, CT and CC groups, respectively). Similarly, no association was found between the MDR12677 G >T polymorphism and CYP3A activity in the healthy subjects or in the HIV-positive patients. The existence of a strong association between the activity of CYP3A and MDR13435 C >T and 2677 G >T polymorphisms appears unlikely, at least in Caucasian populations and/or in the absence of specific environmental factors.

Molecular mechanisms of drug resistance in clinical Candida species isolated from tunisian hospitals.

Antifungal resistance of Candida species is a clinical problem in the management of diseases caused by these pathogens. In this study we identified from a collection of 423 clinical samples taken from Tunisian hospitals two clinical Candida species (Candida albicans JEY355 and Candida tropicalis JEY162) with decreased susceptibility to azoles and polyenes. For JEY355, the fluconazole (FLC) MIC was 8 μg/ml. Azole resistance in C. albicans JEY355 was mainly caused by overexpression of a multidrug efflux pump of the major facilitator superfamily, Mdr1. The regulator of Mdr1, MRR1, contained a yet-unknown gain-of-function mutation (V877F) causing MDR1 overexpression. The C. tropicalis JEY162 isolate demonstrated cross-resistance between FLC (MIC > 128 μg/ml), voriconazole (MIC > 16 μg/ml), and amphotericin B (MIC > 32 μg/ml). Sterol analysis using gas chromatography-mass spectrometry revealed that ergosterol was undetectable in JEY162 and that it accumulated 14α-methyl fecosterol, thus indicating a perturbation in the function of at least two main ergosterol biosynthesis proteins (Erg11 and Erg3). Sequence analyses of C. tropicalis ERG11 (CtERG11) and CtERG3 from JEY162 revealed a deletion of 132 nucleotides and a single amino acid substitution (S258F), respectively. These two alleles were demonstrated to be nonfunctional and thus are consistent with previous studies showing that ERG11 mutants can only survive in combination with other ERG3 mutations. CtERG3 and CtERG11 wild-type alleles were replaced by the defective genes in a wild-type C. tropicalis strain, resulting in a drug resistance phenotype identical to that of JEY162. This genetic evidence demonstrated that CtERG3 and CtERG11 mutations participated in drug resistance. During reconstitution of the drug resistance in C. tropicalis, a strain was obtained harboring only defective Cterg11 allele and containing as a major sterol the toxic metabolite 14α-methyl-ergosta-8,24(28)-dien-3α,6β-diol, suggesting that ERG3 was still functional. This strain therefore challenged the current belief that ERG11 mutations cannot be viable unless accompanied by compensatory mutations. In conclusion, this study, in addition to identifying a novel MRR1 mutation in C. albicans, constitutes the first report on a clinical C. tropicalis with defective activity of sterol 14α-demethylase and sterol Δ(5,6)-desaturase leading to azole-polyene cross-resistance.

Revisiting the diacylglycerol-induced insulin resistance hypothesis.

Obesity is associated with skeletal muscle insulin resistance, which is a crucial step in the development of type 2 diabetes. Among the mechanisms by which obesity may lead to insulin resistance, lipotoxicity is one of the hypotheses being explored; others include inflammation or the oxidative stress hypotheses. This review focuses on the role of diacylglycerols (DAG), a family of lipid metabolites implicated in the pathogenesis of lipotoxicity and insulin resistance. While recent studies report contradictory results in humans with regard to the importance of DAG-induced insulin resistance in skeletal muscle, other current literature highlight a potential role for DAG as signalling molecules. This review will discuss possible hypotheses explaining these contradictory results and the need to explore further the role of DAG in human metabolism.

Decreasing population selection rates of resistance mutation K65R over time in HIV-1 patients receiving combination therapy including tenofovir.

OBJECTIVES: The use of tenofovir is highly associated with the emergence of mutation K65R, which confers broad resistance to nucleoside/nucleotide analogue reverse transcriptase inhibitors (NRTIs), especially when tenofovir is combined with other NRTIs also selecting for K65R. Although recent HIV-1 treatment guidelines discouraging these combinations resulted in reduced K65R selection with tenofovir, updated information on the impact of currently recommended regimens on the population selection rate of K65R is presently lacking. METHODS: In this study, we evaluated changes over time in the selection rate of resistance mutation K65R in a large population of 2736 HIV-1-infected patients failing combination antiretroviral treatment between 2002 and 2010. RESULTS: The K65R resistance mutation was detected in 144 patients, a prevalence of 5.3%. A large majority of observed K65R cases were explained by the use of tenofovir, reflecting its wide use in clinical practice. However, changing patterns over time in NRTIs accompanying tenofovir resulted in a persistent decreasing probability of K65R selection by tenofovir-based therapy. The currently recommended NRTI combination tenofovir/emtricitabine was associated with a low probability of K65R emergence. For any given dual NRTI combination including tenofovir, higher selection rates of K65R were consistently observed with a non-nucleoside reverse transcriptase inhibitor than with a protease inhibitor as the third agent. DISCUSSION: Our finding of a stable time trend of K65R despite elevated use of tenofovir illustrates increased potency of current HIV-1 therapy including tenofovir.

Mefloquine resistant malaria in cameroon and correlation with resistance quinine

Based on the results of in vitro sensitivity of Plasmodium falciparum to chloroquine, quinine and mefloquine, and evaluation of drug consumption conducted in 1987-1988 in four areas in the noth and south-west of Cameron, two opposite situations were encountered in this country. In northern Cameron where mefloquine resistance is prevalent a close correlation was found between the responses of P. falciparum to mefloquine and to quinine, but not between mefloquine and chloroquine. In the south, where chloroquine resistance is highly prevalent, no correlation was found neither between mefloquine and chloroquine nor mefloquine and quinine, but the responses to quinine and chloroquine appear partly correlated. These lead to formulate the hypothesis of a "southern" type of P. falciparum submitted to a high chloroquine drug pressure inducing a secondary cross resistance, whilst a "northern"type submitted to a relatively high and abortive quinine drug pressure inducing a primary quinine resistance and a secondary cross resistance with mefloquine.

Epidemiological distribution of Plasmodium falciparum drug resistance in Brazil and its relevance to the treatment and control of malaria

With the use of a simple formulary, field by health agents was ewstablished a monitoring programme for responses of P. falciparum to the antimalarial drugs. This monitoring programme is emphasized for knowledge of the epidemiology of the drug resistance and the control of malaria falciparum in Amazonan Basin where occurs more than 95% of Brazilian malaria cases every year. It was demonstrated that still now 4-aminoquinolines have a great importance for the mortality control in areas where just SUCAM (National Health Foundation - Health Ministry) agenst are present without any medical assistance. The results obtained permitted the simplification of malaria treatment in Brazil Important conclusions were established in the field of malaria drug resistance.

Human IgE responses to Schistosoma mansoni and resistance to reinfection

Schistosoma mansoni infected Kenyan patients were treated and the intensities of their reinfections were followed over the next two years. in addition, their pre- and six month post-treatment serum levels of IgG1-4, IgM, and IgE, specific for schistosoma, egg and adult worm, were measured in ELISA. No reinfection took place before six months post-treatment. Reinfection intensities varied with age; the younger children becoming reinfected at significantly higher intensities than older individuals. When antibody and reinfection levels were compared, only the six month post-treatment IgE response against adult worm correlated negatively with intensities of reinfection and, therefore, was predictive of resistance or immunity to reinfection. IgE and IgG specific Western Blots were carried out. The adult worm antigens recognized by IgE were restricted compared with the IgG responses of the same patients, although no individual antigen was uniquely recognized by the IgE isotype. A dominant 22 kDa antigen was recognized by most but not all high IgE responders. Patients with IgE responses against this antigen suffered significantly lower subsequent levels of reinfection, compared with non-responders. A monospecific rabbit antiserum against the 22KDa adult worm antigen showed that this antigen is specifically located in the tegument of the adult worm and of 'lung' and 'liver' stage schistosomula, but is absent from the early 'skin' schistosomula. It is possible that this antigen is a target for human IgE mediated immune effector mechanisms active against the post skin stage schistosomula and that this is boosted by the death of adult worms.

Role of the stress sigma factor RpoS in GacA/RsmA-controlled secondary metabolism and resistance to oxidative stress in Pseudomonas fluorescens CHA0.

In Pseudomonas fluorescens biocontrol strain CHA0, the two-component system GacS/GacA positively controls the synthesis of extracellular products such as hydrogen cyanide, protease, and 2,4-diacetylphloroglucinol, by upregulating the transcription of small regulatory RNAs which relieve RsmA-mediated translational repression of target genes. The expression of the stress sigma factor sigmaS (RpoS) was controlled positively by GacA and negatively by RsmA. By comparison with the wild-type CHA0, both a gacS and an rpoS null mutant were more sensitive to H2O2 in stationary phase. Overexpression of rpoS or of rsmZ, encoding a small RNA antagonistic to RsmA, restored peroxide resistance to a gacS mutant. By contrast, the rpoS mutant showed a slight increase in the expression of the hcnA (HCN synthase subunit) gene and of the aprA (major exoprotease) gene, whereas overexpression of sigmaS strongly reduced the expression of these genes. These results suggest that in strain CHA0, regulation of exoproduct synthesis does not involve sigmaS as an intermediate in the Gac/Rsm signal transduction pathway whereas sigmaS participates in Gac/Rsm-mediated resistance to oxidative stress.

Resistance to oxidative stress is associated with metastasis in mucocutaneous leishmaniasis.

Mucocutaneous leishmaniasis (MCL) in South and Central America is characterized by the dissemination (metastasis) of Leishmania Viannia subgenus parasites from a cutaneous lesion to nasopharyngeal tissues. Little is known about the pathogenesis of MCL, especially with regard to the virulence of the parasites and the process of metastatic dissemination. We previously examined the functional relationship between cytoplasmic peroxiredoxin and metastatic phenotype using highly, infrequently, and nonmetastatic clones isolated from an L. (V.) guyanensis strain previously shown to be highly metastatic in golden hamsters. Distinct forms of cytoplasmic peroxiredoxin were identified and found to be associated with the metastatic phenotype. We report here that peroxidase activity in the presence of hydrogen peroxide and infectivity differs between metastatic and nonmetastatic L. (V.) guyanensis clones. After hydrogen peroxide treatment or heat shock, peroxiredoxin was detected preferentially as dimers in metastatic L. (V.) guyanensis clones and in L. (V.) panamensis strains from patients with MCL, compared with nonmetastatic parasites. These data provide evidence that resistance to the first microbicidal response of the host cell by Leishmania promastigotes is linked to peroxiredoxin conformation and may be relevant to intracellular survival and persistence, which are prerequisites for the development of metastatic disease.

Rapid detection of bacterial resistance to antibiotics using AFM cantilevers as nanomechanical sensors.

The widespread misuse of drugs has increased the number of multiresistant bacteria, and this means that tools that can rapidly detect and characterize bacterial response to antibiotics are much needed in the management of infections. Various techniques, such as the resazurin-reduction assays, the mycobacterial growth indicator tube or polymerase chain reaction-based methods, have been used to investigate bacterial metabolism and its response to drugs. However, many are relatively expensive or unable to distinguish between living and dead bacteria. Here we show that the fluctuations of highly sensitive atomic force microscope cantilevers can be used to detect low concentrations of bacteria, characterize their metabolism and quantitatively screen (within minutes) their response to antibiotics. We applied this methodology to Escherichia coli and Staphylococcus aureus, showing that live bacteria produced larger cantilever fluctuations than bacteria exposed to antibiotics. Our preliminary experiments suggest that the fluctuation is associated with bacterial metabolism.

Potassium restores vasorelaxation of resistance arterioles in non-hypertensive DOCA/salt fed mice.

BACKGROUND: Potassium-enriched diets exert renal and cardiovascular protective effects, but the underlying mechanisms are largely unknown. METHODS: Using the dorsal skinfold chamber model for intravital microscopy, we examined endothelium-dependent vasorelaxation of precapillary resistance arterioles in response to acetylcholine or the NO donor SNAP in awake mice. Experiments were performed in uni-nephrectomized one renin gene (Ren-1c) C57BL/6 mice (control group) and in mice having received a continuous administration of deoxycorticosterone acetate and a dietary supplementation of 1% sodium chloride for 8weeks (DOCA/salt group). An additional group of DOCA/salt treated animals received a dietary supplement of 0.4% KCl for 3weeks prior to the experiments (DOCA/salt + potassium group). RESULTS: DOCA/salt treatment for 8weeks resulted in hypokalemia, but blood pressure remained unchanged. In DOCA/salt mice, relaxation of resistance arterioles was blunted in response to acetylcholine, and to a lesser extent to SNAP, suggesting endothelial dysfunction. Endothelium-dependent vasorelaxation was restored by the potassium-enriched diet. CONCLUSION: This study is the first to demonstrate a protective effect of potassium on endothelium-dependent vasorelaxation in the absence of confounding anti-hypertensive effects, as observed in most animal models and the clinical situation. We propose that the known cardio- and nephro-protective effects of potassium might - at least in part - be mediated by the salutary effects on endothelium-dependent arteriolar relaxation.

Infected erythrocyte choline carrier inhibitors: exploring some potentialities inside Plasmodium phospholipid metabolism for eventual resistance acquisition

We have developed a model for designing antimalarial drugs based on interference with an essential metabolism developed by Plasmodium during its intraerythrocytic cycle, phospholipid (PL) metabolism. The most promising drug interference is choline transporter blockage, which provides Plasmodium with a supply of precursor for synthesis of phosphatidylcholine (PC), the major PL of infected erythrocytes. Choline entry is a limiting step in this metabolic pathway and occurs by a facilitated-diffusion system involving an asymmetric carrier operating according to a cyclic model. Choline transport in the erythrocytes is not sodium dependent nor stereospecific as demonstrated using stereoisomers of alpha and beta methylcholine. These last two characteristics along with distinct effects of nitrogen substitution on transport rate demonstrate that choline transport in the infected erythrocyte possesses characteristics quite distinct from that of the nervous system. This indicates a possible discrimination between the antimalarial activity (inhibition of choline transport in the infected erythrocyte) and a possible toxic effect through inhibition of choline entry in synaptosomes. Apart from the de novo pathway of choline, PC can be synthesized by N-methylation from phosphatidylethanolamine (PE). There is a de novo pathway for PE biosynthesis from ethanolamine in infected cells but phosphatidylserine (PS) decarboxylation also occurs. In addition, PE can be directly and abundantly synthesized from serine decarboxylation into ethanolamine, a pathway which is absent from the host. The variety of the pathways that exist for the biosynthesis of one given PL led us to investigate whether an equilibrium can occur between all PL metabolic pathways. Indeed, if alternative (compensative) pathway(s) can operate after blockage of the de novo PC biosynthesis pathway this would indicate a potential mechanism for resistance acquisition. Up until now, there is no evidence of such a compensative process occurring in Plasmodium-infected erythrocytes under physiological conditions. Besides, the discovery of a highly parasite-specific pathway (serine decarboxylation and the presence of PS synthase) constitutes a very attractive and promising target, which could be attacked if resistances are built up against choline analogs. Indeed, potential inhibitions of the serine decarboxylase pathway could be very useful in acting instead of, or in surgery with, choline analogs.