Mycolactone toxin induces an inflammatory response by targeting the IL-1ß pathway: Mechanistic insight into Buruli ulcer pathophysiology.

Mycolactone, a lipid-like toxin, is the major virulence factor of Mycobacterium ulcerans, the etiological agent of Buruli ulcer. Its involvement in lesion development has been widely described in early stages of the disease, through its cytotoxic and immunosuppressive activities, but less is known about later stages. Here, we revisit the role of mycolactone in disease outcome and provide the first demonstration of the pro-inflammatory potential of this toxin. We found that the mycolactone-containing mycobacterial extracellular vesicles produced by M. ulcerans induced the production of IL-1ß, a potent pro-inflammatory cytokine, in a TLR2-dependent manner, targeting NLRP3/1 inflammasomes. We show our data to be relevant in a physiological context. The in vivo injection of these mycolactone-containing vesicles induced a strong local inflammatory response and tissue damage, which were prevented by corticosteroids. Finally, several soluble pro-inflammatory factors, including IL-1ß, were detected in infected tissues from mice and Buruli ulcer patients. Our results revisit Buruli ulcer pathophysiology by providing new insight, thus paving the way for the development of new therapeutic strategies taking the pro-inflammatory potential of mycolactone into account.

Dental caries and enamelin haplotype.

In the literature, the enamelin gene ENAM has been repeatedly designated as a possible candidate for caries susceptibility. Here, we checked whether ENAM variants could increase caries susceptibility. To this aim, we sequenced coding exons and exon-intron boundaries of ENAM in 250 children with a severe caries phenotype and in 149 caries-free patients from 9 French hospital groups. In total, 23 single-nucleotide polymorphisms (SNPs) were found, but none appeared to be responsible for a direct change of ENAM function. Six SNPs had a high minor allele frequency (MAF) and 6 others were identified for the first time. Statistical and evolutionary analyses showed that none of these SNPs was associated with caries susceptibility or caries protection when studied separately and challenged with environmental factors. However, haplotype interaction analysis showed that the presence, in a same variant, of 2 exonic SNPs (rs7671281 and rs3796704; MAF 0.12 and 0.10, respectively), both changing an amino acid in the protein region encoded by exon 10 (p.I648T and p.R763Q, respectively), increased caries susceptibility 2.66-fold independent of the environmental risk factors. These findings support ENAM as a gene candidate for caries susceptibility in the studied population.

[Leprosy, a pillar of human genetics of infectious diseases]. / La lèpre, pilier de la génétique des maladies infectieuses.

Despite a natural reservoir of Mycobacterium leprae limited to humans and free availability of an effective antibiotic treatment, more than 200,000 people develop leprosy each year. This disease remains a major cause of disability and social stigma worldwide. The cause of this constant incidence is currently unknown and indicates that important aspects of the complex relationship between the pathogen and its human host remain to be discovered. An important contribution of host genetics to susceptibility to leprosy has long been suggested to account for the considerable variability between individuals sustainably exposed to M. leprae. Given the inability to cultivate M. leprae in vitro and in the absence of relevant animal model, genetic epidemiology is the main strategy used to identify the genes and, consequently, the immunological pathways involved in protective immunity to M. leprae. Recent genome-wide studies have identified new pathophysiological pathways which importance is only beginning to be understood. In addition, the prism of human genetics placed leprosy at the crossroads of other common diseases such as Crohn's disease, asthma or myocardial infarction. Therefore, novel lights on the pathogenesis of many common diseases could eventually emerge from the detailed understanding of a disease of the shadows.

A major gene effect controls resistance to caries.

Despite recent advances revealing genetic factors influencing caries susceptibility, questions regarding the model of inheritance involved are yet to be addressed. We conducted a Complex Segregation Analysis on decayed teeth in a sample of homogenous, isolated families recruited from the Brazilian Amazon. A dominant, major gene effect controlling resistance to phenotype was detected. The frequency of the resistance allele "A" was 0.63; mean numbers of decayed teeth were 1.53 and 9.53 for genotypes AA/AB and BB, respectively. These results represent a step toward a description of the exact nature of the genetic risk factors controlling human susceptibility to caries.

Impact of age and sex on mycobacterial immunity in an area of high tuberculosis incidence.

SETTING: The extent of immune reactivity measured by the tuberculin skin test (TST) and interferon-gamma (IFN-gamma) T-cell assays is usually not analysed. OBJECTIVE: To determine the impact of age and sex on assay positivity and on the extent of reactivity of both TST and T-cell assays in young persons in an area of South Africa with high TB transmission. RESULTS: Age had a strong impact on assay positivity for all seven immune phenotypes tested (P < 0.0007). Among positive responders, the extent of purified protein derivative (PPD) triggered IFN-gamma release (P < 0.003) was sensitive to age. ESAT-6 triggered IFN-gamma release (day 7, P = 0.03) and the frequency of PPD-specific IFN-gamma(+)CD4(+) (P = 0.03) and IFN-gamma(+)CD8(+) cells (P = 0.04) were weakly dependent on age. By contrast, the extent of TST induration was insensitive to age (P > 0.05), and sex had no significant impact on any phenotype measured (P > 0.05). The high proportion of positive responders in the 1-10 year age-group observed with long-term whole blood assays, but not with 3-day assays and TST, suggests that long-term whole blood assays may be confounded by bacille Calmette-Guérin vaccination in this age group. CONCLUSION: There is a significant impact of age, but not sex, on different assays of immune reactivity in this high TB transmission setting.

An extensive comparison of quantitative trait Loci mapping methods.

BACKGROUND: The choices of study design and statistical approach for mapping a quantitative trait (QT) are of great importance. Larger sibships and a study design based upon phenotypically extreme siblings can be expected to have a greater statistical power. On the other hand, selected samples and/or deviation from normality can influence the robustness and power. Unfortunately, the effects of violation of multivariate normality assumptions and/or selected samples are only known for a limited number of methods. Some recommendations are available in the literature, but an extensive comparison of robustness and power under several different conditions is lacking. METHODS: We compared eight freely available and commonly applied QT mapping methods in a Monte-Carlo simulation study under 36 different models and study designs (three genetic models, three selection schemes, two family structures and the possible effect of deviation from normality). RESULTS: Empirical type I error fractions and empirical power are presented and explained as a whole and for each method separately, followed by a thorough discussion. CONCLUSIONS: The results from this extensive comparison could serve as a valuable source for the choice of the study design and the statistical approach for mapping a QT.

Mycobacterial infections: PARK2 and PACRG associations in leprosy.

An overview of investigations indicating an important role of host genetics, both major histocompatibility complex (MHC) and non-MHC, in leprosy.

Segregation of HLA/TNF region is linked to leprosy clinical spectrum in families displaying mixed leprosy subtypes.

Each year an estimated 600000 new leprosy cases are diagnosed worldwide. The spectrum of the disease varies widely from limited tuberculoid forms to extensive lepromatous forms. A measure of the risk to develop lepromatous forms of leprosy is provided by the extent of skin reactivity to lepromin (Mitsuda reaction). To address a postulated oligogenic control of leprosy pathogenesis, we investigated in the present study linkage of leprosy susceptibility, leprosy clinical subtypes, and extent of the Mitsuda reaction to six chromosomal regions carrying known or suspected leprosy susceptibility loci. The only significant result obtained was linkage of leprosy clinical subtype to the HLA/TNF region on human chromosome 6p21 (P(corrected)=0.00126). In addition, we established that within the same family different HLA/TNF haplotypes segregate into patients with different leprosy subtypes directly demonstrating the importance of this genome region for the control of clinical leprosy presentation.

[Human mycobacterial infections: impact of host genetic factors]. / Infections mycobactériennes humaines: impact des facteurs génétiques de l'hôte.

Humans are exposed worldwide to a variety of environmental mycobacteria (EM) and most children are inoculated with live Bacille Calmette-Guérin (BCG) vaccine. Although rarely pathogenic, poorly virulent mycobacteria, including BCG and most EM, may cause a variety of clinical diseases. M. tuberculosis and M. leprae are more virulent, causing tuberculosis, and leprosy, respectively. Remarkably, only a minority of individuals develop clinical disease, even if infected with virulent mycobacteria. There is now accumulating evidence that the large interindividual variability of clinical outcome results in part from variability in the human genes that control host defense. We review here in current knowledge about genetic predisposition to common (leprosy and tuberculosis) and rare (BCG and EM infections) mycobacterial infections.

[Efficacy of the combination of DEET (20%) and EHD (15%) against mosquito bites. Results of a study carried out in Senegal]. / Efficacité de l'association DEET (20%) et EHD (15%) contre les piqûres de moustiques. Résultats d'une étude de terrain effectuée au Sénégal.

The authors report the results of a survey on the efficacy against mosquito bites of a repellent, Mousticologne Spécial Zones Infestées (DEET 20%, EHD 15%). Two forms of the product, spray and gel, were tested in Senegal. Repellent efficacy was evaluated by exposing volunteers, both repellent-treated and untreated, to mosquito bites. The number of mosquito bites per person and per night was 0.63 in the spray treated group (group 1), 6.03 in the gel treated group (group 2) and 94.17 in the untreated group (group 3). The analysis of these results showed a significant difference between treated and untreated persons. Untreated persons were not protected against mosquito bites, persons treated with the spray were protected for 12 hours and those treated with the gel had over 8 hours' protection. We concluded that a single application of the repellent Mousticologne in the field is capable of ensuring all-night protection against mosquito bites.

Incorporation of covariates in multipoint model-free linkage analysis of binary traits: how important are unaffecteds?

When the mode of inheritance is unknown, genetic linkage analysis of binary trait is commonly performed using affected-sib-pair approaches. When there is evidence that some covariates influence the phenotype, incorporation of this information is expected to increase the power of the analysis since it allows (1) a better specification of the phenotype and (2) to take into account unaffected subjects. Here, we show how to account for covariates in the sibship-oriented Maximum-Likelihood-Binomial (MLB) linkage method by means of Pearson's logistic regression residuals which are computed using phenotypic and covariate information on both affected and unaffected subjects. These residuals are subsequently analysed as a quantitative phenotype with the corresponding extension of the MLB approach which can be used without any assumption on the distribution of these residuals. Then, a large simulation study is performed to study the relative power of incorporating or not unaffected sibs. To this aim, two different strategies in the multipoint analysis of family data are compared: (1) using residuals of the whole sibships (ie both covariate and genotypic information on unaffecteds is needed), and (2) using affecteds only (no information on unaffecteds is needed), under different generating models according to genetic and covariate effects. The results show that there is a clear increment in the power to detect the susceptibility locus when making use of the information carried by unaffecteds, in particular for dominant mode of inheritance and when values of the covariates influencing the disease are shared by all the members of the family.

An autosome-wide search for loci underlying wheezing age of onset in German asthmatic children identifies a new region of interest on 6q24-q25.

While numerous familial studies of asthma have identified several distinct chromosomal regions, no linkage studies have been performed taking into account the age of onset of disease. Here, we performed a genome-wide scan to search for loci linked either to asthma or wheezing age of onset in a population of German asthmatic children by incorporating survival analysis techniques in the maximum-likelihood-binomial approach. In addition to several regions already reported in asthma, wheezing age of onset was found to be strongly linked to chromosome 6q24-q25 (lod score = 3.56). Interestingly, this region contains some candidates genes such as the gene coding for the IFN-gamma receptor ligand-binding chain.

Longitudinal changes in the ductus venosus, cerebral transverse sinus and cardiotocogram in fetal growth restriction.

OBJECTIVE: To evaluate the changes in flow velocity waveforms in the transverse cerebral sinus in growth-restricted fetuses and to correlate these changes with (1) flow velocity waveforms in the ductus venosus and (2) changes in computerized analysis of the fetal cardiotocogram. DESIGN: Fetuses between 22 and 37 weeks' gestation with an estimated fetal weight below the fifth centile were included in this prospective longitudinal study. Doppler measurements of the umbilical artery, descending aorta, middle cerebral artery, transverse cerebral sinus and ductus venosus were recorded. Fetal heart rate was analyzed by a computer system according to the Dawes-Redman criteria. RESULTS: We measured a significant correlation between pulsatility index in the cerebral transverse sinus and in the ductus venosus over the study period and at delivery. There was a negative correlation between these indices and short- and long-term variability of the fetal heart rate. There was a parallel increase in pulsatility in the ductus venosus and the transverse cerebral sinus. These changes were inversely correlated with fetal heart rate variability and preceded fetal distress. CONCLUSION: Cerebral venous blood flow in IUGR fetuses may be a useful additional investigation to discriminate between fetal adaptation and fetal decompensation in chronic hypoxemia.

Linkage analysis of quantitative trait loci: sib pairs or sibships?

Sib pair linkage studies are now widely used to investigate the genetic factors implicated in complex quantitative traits. To increase the power of these approaches, it has been proposed to select extremely discordant (ED) sib pairs which are expected to contain the highest linkage information. However, it is known that sibships of larger size contain more linkage information than independent sib pairs. In this paper we compare, in terms of power and cost considerations, the ED strategy, which uses information on sib pairs only, to the recently developed 'Maximum Likelihood Binomial' sibship-oriented method performed on the whole sibships from which the ED sib pairs have been extracted. We show that the use of these whole sibships is an efficient alternative to approaches focusing on ED sib pairs only.

Granulomatous reaction to intradermal injection of lepromin (Mitsuda reaction) is linked to the human NRAMP1 gene in Vietnamese leprosy sibships.

The Mitsuda test, which measures the specific immune response against intradermally injected lepromin, has a high prognostic value for susceptibility or resistance to the lepromatous form of leprosy. A sib-pair linkage analysis between the Mitsuda response and the NRAMP1 gene was done among 20 nuclear families with leprosy (totaling 118 sibs) from Ho Chi Minh City, Vietnam. All family subjects were genotyped for several intragenic and flanking NRAMP1 markers, leading to the definition of a fully informative NRAMP1 haplotype. Significant linkage was observed between NRAMP1 and Mitsuda reaction when considered either as a quantitative (P<.002) or as a categorical (P=.001) trait. Separate analyses among healthy and affected sibs showed evidence for linkage in both subsamples, indicating that linkage between the Mitsuda reaction and NRAMP1 is independent of leprosy status. These results support the view that NRAMP1 plays a regulatory role for the development of acquired antimycobacterial immune responses as determined by in vivo Mitsuda test reaction.

Genetic model-free linkage analysis using the maximum-likelihood-binomial method for categorical traits.

Within the simulated data of the 11th Genetic Analysis Workshop, we searched for the genes controlling the disease. We analyzed 200 families from Studies 2 and 3 presenting both mild and severe forms of disease. Linkage analysis was performed using the recently developed genetic model-free maximum-likelihood-binomial (MLB) method which overcomes the problem of multiple sibs by considering the sibship as a whole. The MLB allowed us to consider the disease as either a binary (affected/unaffected) or an ordered categorical (differentiating the two forms of disease and including effects of environmental factors) phenotype. In both studies, two regions provided evidence for linkage at a significance level below 10(-4). One is located on chromosome 3 (from D3G041 to D3G047), and the other on chromosome 5 (from D5G034 to D5G041). In Study 2, the most significant results were obtained by combining both forms of disease, suggesting that they are under the same genetic control, while in Study 3, the stronger results were obtained when considering severe subjects alone, suggesting that only the severe form is under the control of both locus B and C. The subsequent knowledge of the true model allowed a posterior interpretation of our results, in particular the difference in optimal coding schemes observed between Studies 2 and 3, and the failure to locate locus A.

Maximum-Likelihood-Binomial method for genetic model-free linkage analysis of quantitative traits in sibships.

Sib-pair linkage studies are widely used to investigate the genetic factors implicated in complex quantitative traits. To analyze these data, we propose a Maximum-Likelihood-Binomial (MLB) approach, which considers the sibship as a whole and relies on the idea of binomial distributions of parental alleles among offsprings. The method is based on the introduction of a latent binary variable capturing the linkage information between the observed quantitative trait and the marker, and the final likelihood can be expressed assuming a parametric distribution for the studied trait but also without any assumption on this distribution. The test for linkage is a simple likelihood ratio test involving a single parameter. The performances of the MLB method are assessed by a simulation study in different kinds of family samples. In the case of families with various sibship sizes, both MLB approaches (assuming or not a parametric distribution for the quantitative trait) provide very consistent results in terms of type I errors and yield power levels generally higher than those of the classical Haseman-Elston method. In the case of extremely discordant sib pairs, we analytically show that, for a common asymptotic type I error, the distribution-free MLB statistic is expected to be more powerful than the test proposed by Risch and Zhang [(1995) Science 268:1584-1589]. In samples including both extremely concordant and discordant sib-pairs, simulation studies show that the MLB approach is at least as powerful as the EDAC method [Gu et al. (1996) Genet Epidemiol 13:513-533]. This MLB method, which can be easily extended to perform multipoint analysis and to account for genetic heterogeneity, appears to be quite an interesting alternative for mapping quantitative trait loci in humans.