Analysis of volatile compounds production kinetics: A study of the impact of nitrogen addition and temperature during alcoholic fermentation.

Among the different compounds present in the must, nitrogen is an essential nutrient for the management of fermentation kinetics, also playing a major role in the synthesis of fermentative aromas. Fermentation temperature is yet another variable that affects fermentation duration and the production of fermentative aromas in wine. The main objective of this study was thus to evaluate the combined effects of nitrogen addition-at the start of the fermentation process or during the stationary phase-at different fermentation temperatures on both fermentation kinetics and aroma synthesis kinetics. To study the impact of these three parameters simultaneously, we used an innovative transdisciplinary approach associating an online GC-MS system with an original modeling approach: a Box-Behnken experimental design combined with response surface modeling and GAM modeling. Our results indicated that all three factors studied had significant effects on fermentation and aroma production kinetics. These parameters did not impact in the same way the different families of volatile compounds. At first, obtained data showed that reduction of ester accumulation in the liquid phase at high temperature was mainly due to important losses by evaporation but also to modifications of yeast metabolic capabilities to synthetize these compounds. In a noticeable way, optimal temperature changed for liquid accumulation of the two classes of esters-23°C for acetate ester and 18°C for ethyl esters-because biological impact of temperature was different for the two chemical families. Moreover, the study of these three factors simultaneously allowed us to show that propanol is not only a marker of the presence of assimilable nitrogen in the medium but above all a marker of cellular activity. Finally, this work enabled us to gain a deeper understanding of yeast metabolism regulation. It also underlines the possibility to refine the organoleptic profile of a wine by targeting the ideal combination of fermentation temperature with initial and added nitrogen concentrations. Such observation was particularly true for isoamyl acetate for which interactions between the three factors were very strong.

Influence of annual climatic variations, climate changes, and sociological factors on the production of the Périgord black truffle (Tuber melanosporum Vittad.) from 1903-1904 to 1988-1989 in the Vaucluse (France).

From 1903-1904 to 1988-1989, the two World Wars and sociological factors as rural desertification and changes in land uses mainly explained the decline of black truffle production in the Vaucluse department, which well reflects that of the whole of France. These can be correlated with the annual climatic variations as well as, from 1924-1925 to 1948-1949, the raw production rates of the managed truffle orchard of Pernes-les-Fontaines located in Vaucluse. The two methods used (correlation coefficients and Bayesian functional linear regression with Sparse Step functions) gave consistent results: the main factor explaining the annual variations of truffle production was the summer climatic water deficit of the year n. A general model including the rural exodus and the cumulated climatic water deficit of summer months both allowed to well explain the evolution of truffle production from 1903-1904 to 1988-1989 in the Vaucluse and its huge decrease. During that period, global warming had little effect. However, in the twenty-first century, all the scenarios predict increased summer water stress for the Mediterranean region, which could greatly affect black truffle production.

Molecular Markers and In Vitro Susceptibility to Doxycycline in Plasmodium falciparum Isolates from Thailand.

Determinations of doxycycline 50% inhibitory concentrations (IC50) for 620 isolates from northwest Thailand were performed via the isotopic method, and the data were analyzed by the Bayesian method and distributed into two populations (mean IC50s of 13.15 µM and 31.60 µM). There was no significant difference between the group with low IC50s versus the group with high IC50s with regard to copy numbers of the Plasmodium falciparum tetQ (pftetQ) gene (P = 0.11) or pfmdt gene (P = 0.87) or the number of PfTetQ KYNNNN repeats (P = 0.72).

PftetQ and pfmdt copy numbers as predictive molecular markers of decreased ex vivo doxycycline susceptibility in imported Plasmodium falciparum malaria.

BACKGROUND: The objective of this study was to evaluate the distribution of a series of independent doxycycline inhibitory concentration 50% (IC50) values to validate the trimodal distribution previously described and to validate the use of the pftetQ and pfmdt genes as molecular markers of decreased in vitro doxycycline susceptibility in Plasmodium falciparum malaria. METHODS: Doxycycline IC50 values, from 484 isolates obtained at the French National Reference Centre for Imported Malaria (Paris) between January 2006 and December 2010, were analysed for the first time by a Bayesian mixture modelling approach to distinguish the different in vitro phenotypic groups by their IC50 values. Quantitative real-time polymerase chain reaction was used to evaluate the pftetQ and pfmdt copy numbers of 89 African P. falciparum isolates that were randomly chosen from the phenotypic groups. RESULTS: The existence of at least three doxycycline phenotypes was demonstrated. The mean doxycycline IC50 was significantly higher in the group with a pftetQ copy number >1 compared to the group with a pftetQ copy number = 1 (33.17 µM versus 17.23 µM) and the group with a pfmdt copy number >1 (28.28 µM versus 16.11 µM). There was a significant difference between the combined low and medium doxycycline IC50 group and the high IC50 group in terms of the per cent of isolates with one or more copy numbers of the pftetQ gene (0% versus 20.69%) or pfmdt gene (8.33% versus 37.93%). In the logistic regression model, the pfmdt and pftetQ copy numbers >1 (odds ratio = 4.65 and 11.47) were independently associated with the high IC50 group. CONCLUSIONS: Copy numbers of pftetQ and pfmdt are potential predictive molecular markers of decreased susceptibility to doxycycline.

Development and validation of real-time one-step reverse transcription-PCR for the detection and typing of dengue viruses.

BACKGROUND: Dengue virus, transmitted by mosquitoes, causes every year 50 million cases of dengue fever. A standardize method for early diagnosis is still needed for clinical diagnosis and epidemiological studies. OBJECTIVE: To develop and validate for sensitivity, specificity, linearity and precision real-time one-step RT-PCR for the detection of dengue viruses. STUDY DESIGN: Multiple alignments of dengue virus sequence for each serotype were done and used to develop five systems of real-time RT-PCR to detect all dengue virus strains and then identify the serotype. These systems were validated on synthetic RNA transcripts for specificity, sensitivity, precision and linearity and then applied on series of human samples. RESULTS: The specificity of each system was determined by sequence alignments and experimentally tested on different flaviviruses. Methods precision and linearity were statistically validated. Each of these systems allowed the detection of less than one infectious particle and was able to detect and serotype quickly dengue virus in human samples where infectious virus cannot be isolated anymore. CONCLUSIONS: These systems are valuable tools for dengue virus diagnosis and epidemiological studies. Standardization and validation of these methods allow an easy transfer to diagnostic laboratories.

Plasmodium falciparum Na+/H+ exchanger 1 transporter is involved in reduced susceptibility to quinine.

Polymorphisms in the Plasmodium falciparum crt (Pfcrt), Pfmdr1, and Pfmrp genes were not significantly associated with quinine (QN) 50% inhibitory concentrations (IC(50)s) in 23 strains of Plasmodium falciparum. An increased number of DNNND repeats in Pfnhe-1 microsatellite ms4760 was associated with an increased IC(50) of QN (P = 0.0007). Strains with only one DNNND repeat were more susceptible to QN (mean IC(50) of 154 nM). Strains with two DNNND repeats had intermediate susceptibility to QN (mean IC(50) of 548 nM). Strains with three DNNND repeats had reduced susceptibility to QN (mean IC(50) of 764 nM). Increased numbers of NHNDNHNNDDD repeats were associated with a decreased IC(50) of QN (P = 0.0020). Strains with profile 7 for Pfnhe-1 ms4760 (ms4760-7) were significantly associated with reduced QN susceptibility (mean IC(50) of 764 nM). The determination of DNNND and NHNDNHNNDDD repeats in Pfnhe-1 ms4760 could be a good marker of QN resistance and provide an attractive surveillance method to monitor temporal trends in P. falciparum susceptibility to QN. The validity of the markers should be further supported by analyzing more isolates.

Social and environmental malaria risk factors in urban areas of Ouagadougou, Burkina Faso.

BACKGROUND: Despite low endemicity, malaria remains a major health problem in urban areas where a high proportion of fevers are presumptively treated using anti-malarial drugs. Low acquired malaria immunity, behaviour of city-dwellers, access to health care and preventive interventions, and heterogenic suitability of urban ecosystems for malaria transmission contribute to the complexity of the malaria epidemiology in urban areas. METHODS: The study was designed to identify the determinants of malaria transmission estimated by the prevalence of anti-circumsporozoite (CSP) antibodies, the prevalence and density of Plasmodium falciparum infection, and the prevalence of malarial disease in areas of Ouagadougou, Burkina-Faso. Thick blood smears, dried blood spots and clinical status have been collected from 3,354 randomly chosen children aged 6 months to 12 years using two cross-sectional surveys (during the dry and rainy seasons) in eight areas from four ecological strata defined according to building density and land tenure (regular versus irregular). Demographic characteristics, socio-economic information, and sanitary and environmental data concerning the children or their households were simultaneously collected. Dependent variables were analysed using mixed multivariable models with random effects, taking into account the clustering of participants within compounds and areas. RESULTS: Overall prevalences of CSP-antibodies and P. falciparum infections were 7.7% and 16.6% during the dry season, and 12.4% and 26.1% during the rainy season, respectively, with significant differences according to ecological strata. Malaria risk was significantly higher among children who i) lived in households with lower economic or education levels, iii) near the hydrographic network, iv) in sparsely built-up areas, v) in irregularly built areas, vi) who did not use a bed net, vii) were sampled during the rainy season or ii) had traveled outside of Ouagadougou. CONCLUSION: Malaria control should be focused in areas which are irregularly or sparsely built-up or near the hydrographic network. Furthermore, urban children would benefit from preventive interventions (e.g. anti-vectorial devices or chemoprophylaxis) aimed at reducing malaria risk during and after travel in rural areas.

The multiplicity of Plasmodium falciparum infections is associated with acquired immunity to asexual blood stage antigens.

We evaluated the relationship between immune response markers and the multiplicity of Plasmodium falciparum infections in order to assess the validity of the latter as an indicator of the acquisition of anti-malarial immunity. Parasite populations present during malaria episodes of 64 Gabonese children who presented with at least 4 such attacks during active follow-up over a 7-year period were characterized using MSP-1 and MSP-2 PCR-based methods. Plasma samples taken at healthy and parasite-free phase were used to measure P. falciparum antigen-specific antibody and cytokine activity. We found evidence of intra- and inter-individual variation in the number of parasite genotypes present in different malaria episodes, although in 72% of isolates no more than 2 parasite genotypes were detectable. Samples with the highest multiplicity were from children with significantly lower (p < 0.03) antibody responses to specific asexual stage antigens. Additionally, the whole blood interferon-gamma production capacity was significantly higher (p < 0.02) in those with lower infection multiplicity. Malaria episodes with multiple clones indeed reflect a low level of acquired immunity and a consequently poor capacity to control the infection. These findings suggest that the multiplicity of falciparum infection may be a potentially useful parameter in the evaluation of malaria control interventions.

Multinormal in vitro distribution model suitable for the distribution of Plasmodium falciparum chemosusceptibility to doxycycline.

The distribution and range of 50% inhibitory concentrations (IC(50)s) of doxycycline were determined for 747 isolates obtained between 1997 and 2006 from patients living in Senegal, Republic of the Congo, and Gabon and patients hospitalized in France for imported malaria. The statistical analysis was designed to answer the specific question of whether Plasmodium falciparum has different phenotypes of susceptibility to doxycycline. A triple normal distribution was fitted to the data using a Bayesian mixture modeling approach. The IC(50) geometric mean ranged from 6.2 microM to 11.1 microM according to the geographical origin, with a mean of 9.3 microM for all 747 parasites. The values for all 747 isolates were classified into three components: component A, with an IC(50) mean of 4.9 microM (+/-2.1 microM [standard deviation]); component B, with an IC(50) mean of 7.7 microM (+/-1.2 microM); and component C, with an IC(50) mean of 17.9 microM (+/-1.4 microM). According to the origin of the P. falciparum isolates, the triple normal distribution was found in each subgroup. However, the proportion of isolates predicted to belong to component B was most important in isolates from Gabon and Congo and in isolates imported from Africa (from 46 to 56%). In Senegal, 55% of the P. falciparum isolates were predicted to be classified as component C. The cutoff of reduced susceptibility to doxycycline in vitro was estimated to be 35 microM.

Dihydroethanoanthracene derivatives reverse in vitro quinoline resistance in Plasmodium falciparum malaria.

The capacity of ten molecules for reversing resistance in Plasmodium falciparum in vitro to quinoline antimalarial drugs, such as chloroquine (CQ), quinine (QN), mefloquine (MQ) and monodesethylamodiaquine (MDAQ), was assessed against 27 Plasmodium falciparum isolates. Four of these compounds were 9,10-dihydroethanoanthracene derivatives (DEAs). These DEAs reversed 75 to 92% of the CQ resistant strains. These synthetic compounds were more effective in combination with CQ than verapamil, ketotifen, chlorpromazine, reserpine or nicardipine, which reversed less than 50% of the CQ resistant strains. DEAs significantly reversed 67 to 100% of MDAQ resistant parasites. These compounds were more effective in combination with MDAQ than ketotifen (60% of reversal), chlorpromazine (45%), verapamil (33%), reserpine (30%) or nicardipine (9%). The reversal activity of MQ resistance was less pronounced, regardless of the molecule tested, and was homogeneous with a rate ranging from 42% for ketotifen to 58% for reserpine, nicardipine, verapamil and cyproheptadine. The four DEAs significantly reversed 50 to 55% of the parasites resistant to MQ. Fifty-six to 78 % of the QN resistant parasites were reversed by the synthetic DEAs. There were few differences in the rate of reversal activity on QN resistant strains between the ten compounds, with rates ranging between 56 to 78% for the ten chemosensitizers. The use of DEAs in combination with quinoline seems to be thus a promising strategy for limiting the development of drug resistant strains and for treating patients in drug resistant areas.

Antimalarial drug use in general populations of tropical Africa.

BACKGROUND: The burden of Plasmodium falciparum malaria has worsened because of the emergence of chloroquine resistance. Antimalarial drug use and drug pressure are critical factors contributing to the selection and spread of resistance. The present study explores the geographical, socio-economic and behavioural factors associated with the use of antimalarial drugs in Africa. METHODS: The presence of chloroquine (CQ), pyrimethamine (PYR) and other antimalarial drugs has been evaluated by immuno-capture and high-performance liquid chromatography in the urine samples of 3,052 children (2-9 y), randomly drawn in 2003 from the general populations at 30 sites in Senegal (10), Burkina-Faso (10) and Cameroon (10). Questionnaires have been administered to the parents of sampled children and to a random sample of households in each site. The presence of CQ in urine was analysed as dependent variable according to individual and site characteristics using a random - effect logistic regression model to take into account the interdependency of observations made within the same site. RESULTS: According to the sites, the prevalence rates of CQ and PYR ranged from 9% to 91% and from 0% to 21%, respectively. In multivariate analysis, the presence of CQ in urine was significantly associated with a history of fever during the three days preceding urine sampling (OR = 1.22, p = 0.043), socio-economic level of the population of the sites (OR = 2.74, p = 0.029), age (2-5 y = reference level; 6-9 y OR = 0.76, p = 0.002), prevalence of anti-circumsporozoite protein (CSP) antibodies (low prevalence: reference level; intermediate level OR = 2.47, p = 0.023), proportion of inhabitants who lived in another site one year before (OR = 2.53, p = 0.003), and duration to reach the nearest tarmacked road (duration less than one hour = reference level, duration equal to or more than one hour OR = 0.49, p = 0.019). CONCLUSION: Antimalarial drug pressure varied considerably from one site to another. It was significantly higher in areas with intermediate malaria transmission level and in the most accessible sites. Thus, P. falciparum strains arriving in cross-road sites or in areas with intermediate malaria transmission are exposed to higher drug pressure, which could favour the selection and the spread of drug resistance.

Inherited predisposition of lung cancer: a hierarchical modeling approach to DNA repair and cell cycle control pathways.

The DNA repair systems maintain the integrity of the human genome and cell cycle checkpoints are a critical component of the cellular response to DNA damage. We hypothesized that genetic variants in DNA repair and cell cycle control pathways will influence the predisposition to lung cancer, and studied 27 variants in 17 DNA repair enzymes and 10 variants in eight cell cycle control genes in 1,604 lung cancer patients and 2,053 controls. To improve the estimation of risks for specific variants, we applied a Bayesian approach in which we allowed the prior knowledge regarding the evolutionary biology and physicochemical properties of the variant to be incorporated into the hierarchical model. Based on the estimation from the hierarchical modeling, subjects who carried OGG1 326C/326C homozygotes, MGMT 143V or 178R, and CHEK2 157I had an odds ratio of lung cancer equal to 1.45 [95% confidence interval (95% CI), 1.05-2.00], 1.18 (95% CI, 1.01-1.40), and 1.58 (95% CI, 1.14-2.17). The association of CHEK2 157I seems to be overestimated in the conventional analysis. Nevertheless, this association seems to be robust in the hierarchical modeling. None of the pathways seem to have a prominent effect. In general, our study supports the notion that sequence variation may explain at least some of the variation of inherited susceptibility. In particular, further investigation of OGG1, MGMT, and CHEK2 focusing on the genetic regions where the present markers are located or the haplotype blocks tightly linked with these markers might be warranted.