Native chromatin immunoprecipitation (N-ChIP) and ChIP-Seq of Schistosoma mansoni: Critical experimental parameters.

International audienceHistone modifications are important epigenetic marks that influence chromatin structure and consequently play a role in the control of eukaryotic transcription. Several histone modifying enzymes have been characterized in Schistosoma mansoni and it has been suggested that the regulation of gene transcription in schistosomes may require the action of these enzymes. However, the influence of chromatin structure on gene transcription in schistosomes has never been investigated. Chromatin immunoprecipitation (ChIP) is the technique of choice to study the relationship between histone modifications and gene expression. Although this technique has been widely used with cultured cells from model organisms and with many unicellular organisms, it remains challenging to apply this technique to non-conventional organisms that undergo complex life cycles. In this work, we describe a native ChIP procedure that is applicable to all the stages of the S. mansoni life cycle and does not require expensive equipment. Immunoprecipitated DNA was analysed on a whole-genome scale using massively parallel sequencing (ChIP-Sequencing or ChIP-Seq). We show that ChIP-Seq and conventional quantitative PCR deliver comparable results for a life-cycle regulated locus, smRHO, that encodes a guanine-protein coupled receptor. This is the first time that the ChIP-Seq procedure has been applied to a parasite. This technique opens new ways for analyzing epigenetic mechanisms in S. mansoni at a whole-genome scale and on the level of individual loci

Female biased sex-ratio in Schistosoma mansoni after exposure to an allopatric intermediate host strain of Biomphalaria glabrata.

International audience: For parasites that require multiple hosts to complete their development, the interaction with the intermediate host may have an impact on parasite transmission and development in the definitive host. The human parasite Schistosoma mansoni needs two different hosts to complete its life cycle: the freshwater snail Biomphalaria glabrata (in South America) as intermediate host and a human or rodents as final host. To investigate the influence of the host environment on life history traits in the absence of selection, we performed experimental infections of two B. glabrata strains of different geographic origin with the same clonal population of S. mansoni. One B. glabrata strain is the sympatric host and the other one the allopatric host. We measured prevalence in the snail, the cercarial infectivity, sex-ratio, immunopathology in the final host and microsatellite frequencies of individual larvae in three successive generations. We show that, even if the parasite population is clonal based on neutral markers, S. mansoni keeps the capacity of generating phenotypic plasticity and/or variability for different life history traits when confront to an unusual environment, in this study the intermediate host. The most dramatic change was observed in sex-ratio: in average 1.7 times more female cercariae were produced when the parasite developed in an allopatric intermediate host

Whole-genome in-silico subtractive hybridization (WISH) - using massive sequencing for the identification of unique and repetitive sex-specific sequences: the example of Schistosoma mansoni

<p>Abstract</p> <p>Background</p> <p>Emerging methods of massive sequencing that allow for rapid re-sequencing of entire genomes at comparably low cost are changing the way biological questions are addressed in many domains. Here we propose a novel method to compare two genomes (genome-to-genome comparison). We used this method to identify sex-specific sequences of the human blood fluke <it>Schistosoma mansoni</it>.</p> <p>Results</p> <p>Genomic DNA was extracted from male and female (heterogametic) <it>S. mansoni </it>adults and sequenced with a Genome Analyzer (Illumina). Sequences are available at the NCBI sequence read archive <url>http://www.ncbi.nlm.nih.gov/Traces/sra/</url> under study accession number SRA012151.6. Sequencing reads were aligned to the genome, and a pseudogenome composed of known repeats. Straightforward comparative bioinformatics analysis was performed to compare male and female schistosome genomes and identify female-specific sequences. We found that the <it>S. mansoni </it>female W chromosome contains only few specific unique sequences (950 Kb i.e. about 0.2% of the genome). The majority of W-specific sequences are repeats (10.5 Mb i.e. about 2.5% of the genome). Arbitrarily selected W-specific sequences were confirmed by PCR. Primers designed for unique and repetitive sequences allowed to reliably identify the sex of both larval and adult stages of the parasite.</p> <p>Conclusion</p> <p>Our genome-to-genome comparison method that we call "whole-genome <it>in-silico </it>subtractive hybridization" (WISH) allows for rapid identification of sequences that are specific for a certain genotype (e.g. the heterogametic sex). It can in principle be used for the detection of any sequence differences between isolates (<it>e.g</it>. strains, pathovars) or even closely related species.</p

(Epi)genetic Inheritance in Schistosoma mansoni: A Systems Approach

International audienceThe G×E concept, in which genotype × environment interactions bring about the phenotype, is widely used to describe biological phenomena. We propose to extend the initial notion of the concept, replacing G by ‘inheritance system’. This system, comprised of both genome and epigenome components, collectively interacts with the environment to shape the development of a phenotype. In the case of the human blood fluke Schistosoma mansoni, responsible for intestinal bilharzia, the phenotypic trait that is most relevant to global health is infection success. Taking a systems biology view we show how genetic and epigenetic interactions result in ephemeral, but also heritable, phenotypic variations that are important for infection success

Exposure to hycanthone alters chromatin structure around specific gene functions and specific repeats in Schistosoma mansoni.

International audienceSchistosoma mansoni is a parasitic plathyhelminth responsible for intestinal schistosomiasis (or bilharzia), a disease affecting 67 million people worldwide and causing an important economic burden. The schistosomicides hycanthone, and its later proxy oxamniquine, were widely used for treatments in endemic areas during the twentieth century. Recently, the mechanism of action, as well as the genetic origin of a stably and Mendelian inherited resistance for both drugs was elucidated in two strains. However, several observations suggested early on that alternative mechanisms might exist, by which resistance could be induced for these two drugs in sensitive lines of schistosomes. This induced resistance appeared rapidly, within the first generation, but was metastable (not stably inherited). Epigenetic inheritance could explain such a phenomenon and we therefore re-analyzed the historical data with our current knowledge of epigenetics. In addition, we performed new experiments such as ChIP-seq on hycanthone treated worms. We found distinct chromatin structure changes between sensitive worms and induced resistant worms from the same strain. No specific pathway was discovered, but genes in which chromatin structure modifications were observed are mostly associated with transport and catabolism, which makes sense in the context of the elimination of the drug. Specific differences were observed in the repetitive compartment of the genome. We finally describe what types of experiments are needed to understand the complexity of heritability that can be based on genetic and/or epigenetic mechanisms for drug resistance in schistosome

Epigenetic and phenotypic variability in populaitons of Schistosoma mansoni - a possible kick-off for adaptative host/parasite evolution

International audienceEpigenetics, the science of heritable but modifiable information, is now a well-accepted component of many research fields. Nevertheless, epigenetics has not yet found broad appreciation in one of the most exciting fields of biology: the comprehension of evolution. This is surprising, since the reason for the existence of this alternative information-transmitting system lies certainly in the evolutionary advantage it provides. Theoretical considerations support a model in which epigenetic mechanisms allow for increasing phenotypic variability and permit populations to explore the adaptive landscape without modifications of the genotype. The data presented here support the view that modulating the epigenotype of the human bloodfluke Schistosoma mansoni by treatment of larvae with histone deacetylase inhibitor leads indeed to an increase of phenotypic variability. It is therefore conceivable that environmentally induced changes in the epigenotype release new phenotypes on which selection can act and that this process is the first step in adaptive evolution

Chromatin structure changes around satellite repeats on the Schistosoma mansoni female sex chromosome suggest a possible mechanism for sex chromosome emergence.

International audienceBACKGROUND: In the leuphotrochozoan parasitic platyhelminth Schistosoma mansoni, male individuals are homogametic (ZZ) whereas females are heterogametic (ZW). To elucidate the mechanisms that led to the emergence of sex chromosomes we compared the genomic sequence and the chromatin structure of male and female individuals. As for many eukaryotes, the lower estimate for the repeat content is 40%, with an unknown proportion of domesticated repeats. We used massive sequencing to de novo assemble all repeats, and identify unambiguously Z-specific, W-specific and pseudoautosomal regions of the S. mansoni sex chromosomes.RESULTS: We show that 70-90 % of S. mansoni W and Z are pseudoautosomal. No female specific gene could be identified. Instead, the W-specific region is composed almost entirely of 36 satellite repeat families, of which 33 were previously unknown. Transcription and chromatin status of female specific repeats are stage specific: for those repeats that are transcribed, transcription is restricted to the larval stages lacking sexual dimorphism. In contrast, in the sexually dimorphic adult stage of the life cycle, no transcription occurs. In addition, the euchromatic character of histone modifications around the W-specific repeats decreases during the life cycle. Recombination repression occurs in this region even if homologous sequences are present on both the Z and W chromosomes.CONCLUSION: Our study provides for the first time evidence for the hypothesis that, at least in organisms with a ZW type of sex chromosomes, repeat-induced chromatin structure changes could indeed be the initial event in sex chromosome emergence

Proteobacteria from the human skin microbiota: Species-level diversity and hypotheses

The human skin microbiota is quantitatively dominated by Gram-positive bacteria, detected by both culture and metagenomics. However, metagenomics revealed a huge variety of Gram-negative taxa generally considered from environmental origin. For species affiliation of bacteria in skin microbiota, clones of 16S rRNA gene and colonies growing on diverse culture media were analyzed. Species-level identification was achieved for 81% of both clones and colonies. Fifty species distributed in 26 genera were identified by culture, mostly belonging to Actinobacteria and Firmicutes, while 45 species-level operational taxonomic units distributed in 30 genera were detected by sequencing, with a high diversity of Proteobacteria. This mixed approach allowed the detection of 100% of the genera forming the known core skin Gram-negative microbiota and 43% of the known diversity of Gram-negative genera in human skin. The orphan genera represented 50% of the current skin pan-microbiota. Improved culture conditions allowed the isolation of Roseomonas mucosa, Aurantimonas altamirensis and Agrobacterium tumefaciens strains from healthy skin. For proteobacterial species previously described in the environment, we proposed the existence of skin-specific ecotypes, which might play a role in the fine-tuning of skin homeostasis and opportunistic infections but also act as a shuttle between environmental and human microbial communities. Therefore, skin-associated proteobacteria deserve to be considered in the One-Health concept connecting human health to the health of animals and the environment

Genes Related to Ion-Transport and Energy Production Are Upregulated in Response to CO₂-Driven pH Decrease in Corals: New Insights from Transcriptome Analysis

Since the preindustrial era, the average surface ocean pH has declined by 0.1 pH units and is predicted to decline by an additional 0.3 units by the year 2100. Although subtle, this decreasing pH has profound effects on the seawater saturation state of carbonate minerals and is thus predicted to impact on calcifying organisms. Among these are the scleractinian corals, which are the main builders of tropical coral reefs. Several recent studies have evaluated the physiological impact of low pH, particularly in relation to coral growth and calcification. However, very few studies have focused on the impact of low pH at the global molecular level. In this context we investigated global transcriptomic modifications in a scleractinian coral (Pocillopora damicornis) exposed to pH 7.4 compared to pH 8.1 during a 3-week period. The RNAseq approach shows that 16% of our transcriptome was affected by the treatment with 6% of upregulations and 10% of downregulations. A more detailed analysis suggests that the downregulations are less coordinated than the upregulations and allowed the identification of several biological functions of interest. In order to better understand the links between these functions and the pH, transcript abundance of 48 candidate genes was quantified by q-RT-PCR (corals exposed at pH 7.2 and 7.8 for 3 weeks). The combined results of these two approaches suggest that pH ≥ 7.4 induces an upregulation of genes coding for proteins involved in calcium and carbonate transport, conversion of CO₂ into HCO₃⁻ and organic matrix that may sustain calcification. Concomitantly, genes coding for heterotrophic and autotrophic related proteins are upregulated. This can reflect that low pH may increase the coral energy requirements, leading to an increase of energetic metabolism with the mobilization of energy reserves. In addition, the uncoordinated downregulations measured can reflect a general trade-off mechanism that may enable energy reallocation