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International audience; An endolichenic strain of the Ascomycetaceous Xylaria hypoxylon, cultivated alone or in coculture with another endolichenic fungus Dendrothyrium variisporum, produced seven new bioactive eremophilane sesquiterpenes eremoxylarins D−J (1−7). The isolated compounds disclosed a high similarity with the eremophilane core of the bioactive integric acid, and structures were elucidated by 1D and 2D NMR spectra and electronic circular dichroism (ECD) analyses. Eremoxylarins D, F, G, and I showed a selective activity against Gram-positive bacteria such as methicillin-resistant Staphylococcus aureus with minimum inhibitory concentration (MIC) values between 0.39 and 12.5 μg/mL. Eremoxylarin I, the most antibacterial active sesquiterpene, was also active against HCoV-229E at a concentration nontoxic to the hepatoma Huh-7 cell line with an 50% inhibitory concentration (IC 50) of 18.1 μM and a 50% cytotoxic concentration (CC 50) of 46.6 μM.

During this global pandemic of the COVID-19 disease, a lot of information has arisen in the media and online without scientific validation, and among these is the possibility that this disease could be aggravated by a secondary bacterial infection such as

International audience; Salmonella Heidelberg is one of the most common serovar causing foodborne illnesses. To limit the development of digestive bacterial infection, food supplements containing probiotic bacteria can be proposed. Commensal non-toxigenic Bacteroides fragilis has recently been suggested as a next-generation probiotic candidate. By using an original triple co-culture model including Caco-2 cells (representing human enterocytes), HT29-MTX (representing mucus-secreting goblet cells), and M cells differentiated from Caco-2 by addition of Raji B lymphocytes, bacterial translocation was evaluated. The data showed that S. Heidelberg could translocate in the triple co-culture model with high efficiency, whereas for B. fragilis a weak translocation was obtained. When cells were exposed to both bacteria, S. Heidelberg translocation was inhibited. The cell-free supernatant of B. fragilis also inhibited S. Heidelberg translocation without impacting epithelial barrier integrity. This supernatant did not affect the growth of S. Heidelberg. The non-toxigenic B. fragilis confers health benefits to the host by reducting bacterial translocation. These results suggested that the multicellular model provides an efficient in vitro model to evaluate the translocation of pathogens and to screen for probiotics that have a potential inhibitory effect on this translocation.

International audience; The genus Roseburia consists of obligate Gram-positive anaerobic bacteria that are slightly curved, rod-shaped and motile by means of multiple subterminal flagella. It includes five species: Roseburia intestinalis, R. hominis, R. inulinivorans, R. faecis and R. cecicola. Gut Roseburia spp. metabolize dietary components that stimulate their proliferation and metabolic activities. They are part of commensal bacteria producing short-chain fatty acids, especially butyrate, affecting colonic motility, immunity maintenance and anti-inflammatory properties. Modification in Roseburia spp. representation may affect various metabolic pathways and is associated with several diseases (including irritable bowel syndrome, obesity, Type 2 diabetes, nervous system conditions and allergies). Roseburia spp. could also serve as biomarkers for symptomatic pathologies (e.g., gallstone formation) or as probiotics for restoration of beneficial flora.

International audience; We have previously demonstrated that out of the butyrolactones series synthesized based on the natural lichen metabolite lichesterinic acid, compound (B-13) was the most effective against oral bacteria. However, its antibacterial mechanism is still unknown. In this study, we have investigated its bacterial localization by synthesizing a fluorescently labeled B-13 with NBD while maintaining its antibacterial activity. We showed that this compound binds to Streptococcus gordonii cell surface, as demonstrated by HPLC analysis. By adhering to cell surface, B-13 induced cell wall disruption leading to the release of bacterial constituents and consequently, the death of S. gordonii, a Gram-positive bacterium. A Gram-negative counterpart, Porphyromanas gingivalis, showed also cracked and ruptured cells in the presence of B-13. Besides, we also demonstrated that the analog of B-13, B-12, has also induced disruption of P. gingivalis and S. gordonii. This study revealed that butyrolactones can be considered as potent antibacterial compounds against oral pathogens causing medical complications.

International audience; The oral bacteria not only infect the mouth and reside there, but also travel through the blood and reach distant body organs. If left untreated, the dental biofilm that can cause destructive inflammation in the oral cavity may result in serious medical complications. In dental biofilm, Streptococcus gordonii, a primary oral colonizer, constitutes the platform on which late pathogenic colonizers like Porphyromonas gingivalis, the causative agent of periodontal diseases, will bind. The aim of this study was to determine the antibacterial activity of eleven natural lichen compounds belonging to different chemical families and spanning from linear into cyclic and aromatic structures to uncover new antibiotics which can fight against the oral bacteria. The compounds were screened by broth microdilution assay. Three compounds were shown to have promising antibacterial activities where the depsidone core with certain functional groups constituted the best compound, psoromic acid, with the lowest MICs=11.72 and 5.86μg/mL against S. gordonii and P. gingivalis, respectively. The compounds screened had promising antibacterial activity which might be attributed to some important functional groups as discussed in our study. The best compounds did not induce the death of gingival epithelial carcinoma cells (Ca9-22). These results introduce new compounds having potent antibacterial activities against oral pathogens causing serious medical complications.

International audience; Novel butyrolactone analogues were designed and synthesized based on the known lichen antibacterial compounds, lichesterinic acids (B-10 and B-11), by substituting different functional groups on the butyrolactone ring trying to enhance its activity. All synthesized butyrolactone analogues were evaluated for their in vitro antibacterial activity against Streptococcus gordonii. Among the derivatives, B-12 and B-13 had the lowest MIC of 9.38 μg/mL where they have shown to be stronger bactericidals, by 2–3 times, than the reference antibiotic, doxycycline. These two compounds were then checked for their cytotoxicity against human gingival epithelial cell lines, Ca9–22, and macrophages, THP-1, by MTT and LDH assays which confirmed their safety against the tested cell lines. A preliminary study of the structure–activity relationships unveiled that the functional groups at the C4 position had an important influence on the antibacterial activity. An optimum length of the alkyl chain at the C5 position registered the best antibacterial inhibitory activity however as its length increased the bactericidal effect increased as well. This efficiency was attained by a carboxyl group substitution at the C4 position indicating the important dual role contributed by these two substituents which might be involved in their mechanism of action. © 2016 Elsevier Ltd

International audience; In this study, we investigated adherence and motility of the hypermutator Salmonella enterica Heidelberg B182 bovine strain related to a 12bp deletion in mutS. This mutator phenotype was associated with increased adherence to epithelial cells and with high expression of fimA as shown by real-time RT-PCR. Motility studies showed that fliC were up-regulated in the B182 strain, while fljA and fljB were down-regulated. In order to determine if mutated mutS is implicated in this genes expression, isogenic strains, derived from a WT strain, containing the 12bp deletion in mutS (Δ12bpmutS) or an inactivated mutS (ΔmutS) were generated. Δ12bpmutS and ΔmutS strains showed a spontaneous mutation rate similar to the environmental strain B182, but exhibited lower adherence capacity and fimA expression. In contrast to the fimbriae genes, in Δ12bpmutS, fliC expression was up-regulated, but fljA and fljB expression were decreased, as in the B182 strain. Only fljB expression was increased in ΔmutS mutants. Taken together, our data suggest that mutS alteration does not influence fimbriae expression but can impact flagella genes.

Human papillomavirus (HPV) infection, particularly type 16, is causally associated with cancer of the uterine cervix. The progression of cervical lesions suggests that viral antigens are not adequately presented to the immune system. The aim of this study was to determine whether HPV16 viral particles can influence the trafficking of human DC/Langerhans cells (LC), either by direct interactions with DC or following incubation with human normal keratinocytes that are in close contact with LC in the squamous epithelium. We first demonstrated that HPV16 L1 major capsid protein, when self-assembled into virus-like particles (VLP), is able to induce in DC an over-expression of CXC receptor 4 (CXCR4) via the activation of the NF-κB signaling pathway and to enhance DC motility in the presence of CXCL12, suggesting an ability to migrate towards lymph nodes. We also showed that conditioned media of HPV16 VLP-treated keratinocytes induce a lower LC migration than those from untreated keratinocytes and that prostaglandin E2 (PGE(2)), detected in HPV16 VLP-treated keratinocyte supernatants, may reduce LC recruitment into the squamous epithelium. Taken together, our data demonstrate that HPV16 VLP may differentially regulate the immune protective response according to their target cells.

A hepatitis E virus (HEV) vaccine would be valuable to reduce the morbidity and mortality associated with the infection in endemic areas. HEV pseudocapsids and epidermal delivery of HEV ORF2 DNA vaccine by gene-gun have been shown to confer protection against virus challenge in monkeys. Vectorization of a DNA vaccine by virus-like particles is a new immunization approach. We report here the successful immunization of mice with two ORF2 genes encapsidated into human papillomavirus type 31 virus-like particles. The HEV genes ORF2(112–660) and ORF2(112–608) were optimized for expression in mammalian cells and inserted in a baculovirus-derived vector for expression in insect cells. When expressed in Sf21 insect cells, ORF2(112–660) led to the production of irregular 15 nm particles that accumulated in the cytoplasm of the cells, whereas ORF2(112–608) induced the production of 18 nm particles that were present in both the cell culture medium and the cell cytoplasm. Anti-HEV immune responses were higher for the 15 nm particles (HEV112–660) than that for to the 18 nm particles (HEV112–608). Delivery into mice of two HEV ORF2 genes via a papillomavirus VLP was very effective in the induction of anti-HEV antibodies. In addition, an effective immune response to human papillomavirus capsids occurred. These engineered pseudoviruses were thus demonstrated to induce immune responses to both hepatitis E virus and human papillomavirus when they were administered to mice intramuscularly.

International audience; We have previously described that a strain of Salmonella Heidelberg with a hypermutator phenotype, B182, adhered strongly to HeLa cells. In this work, we showed that this hypermutator Salmonella strain invaded HeLa epithelial cells and induced cytoskeleton alteration. Those changes lead to HeLa cell death which was characteristic of apoptosis. For the first time, we showed that this hypermutator strain induced apoptosis associated with the activation of caspases 2, 9 and 3. Complementation of B182 strain showed a decrease in cells death induction. In the presence of other Salmonella Heidelberg with a normomutator phenotype, such as WT and SL486, cell death and caspase 3 were undetectable. These results suggested that early apoptosis and caspase 3 activation were specific to B182. Besides, B182 induced LDH release and caspase 3 activation in CaCo-2 and HCT116 cells. Heat-treated B182 and diffusible products failed to induce this phenotype. Epithelial cells treatment with cytochalasin D caused the inhibition of B182 internalisation and caspase 3 activation. These results showed that this cell death required active S. Heidelberg B182 protein synthesis and bacterial internalisation. However sipB and sopB, usually involved in apoptosis induced by Salmonella were not overexpressed in B182, contrary to fimA and fliC. Comparative genome analysis showed numerous mutations as in rpoS which would be more investigated. The role of the hypermutator phenotype might be suspected to be implicated in these specific features. This result expands our knowledge about strong mutators frequently found in bacterial organisms isolated from clinical specimens

International audience; Many β-lactamases have been described in various Gram-negative bacilli (Capnocytophaga, Prevotella, Fusobacterium, etc.) of the oral cavity, belonging to class A of the Ambler classification (CepA, CblA, CfxA, CSP-1 and TEM), class B (CfiA) or class D in Fusobacterium nucleatum (FUS-1). The minimum inhibitory concentrations of β-lactams are variable and this variation is often related to the presence of plasmids or other mobile genetic elements (MGEs) that modulate the expression of resistance genes. DNA persistence and bacterial promiscuity in oral biofilms also contribute to genetic transformation and conjugation in this particular microcosm. Overexpression of efflux pumps is facilitated because the encoding genes are located on MGEs, in some multidrug-resistant clinical isolates, similar to conjugative transposons harbouring genes encoding β-lactamases. All these facts lead us to consider the oral cavity as an important reservoir of β-lactam resistance genes and a privileged place for genetic exchange, especially in commensal strictly anaerobic Gram-negative bacilli.

Human papillomavirus (HPV) virus-like particles (VLP) and synthetic peptides corresponding to positively-charged sequences of the major and minor capsid proteins were tested for their efficacy in inhibiting the infectivity of HPV 31 pseudovirions by blocking virus entry into cells. A greater than 80% reduction of transfection was observed with one HPV-31 peptide at a concentration of 10 μg/ml. Moreover, the blocking was not type-specific since similar reduction in transfection was observed with peptides from other HPV types at a concentration of 60 μg/ml. This concentration was non-toxic for the cells. These findings indicate that some of the positively-charged sequences of the L1 and L2 HPV capsid proteins of papillomavirus are compounds that might be locally active against sexually transmitted papillomavirus. The findings provide further evidence that cellular glycosamino-glycans (GAGs) are functional receptors for HPVs. J. Med. Virol. 73:474–480, 2004. © 2004 Wiley-Liss, Inc.

In a case-control study in Uganda, we examined associations between different cancer sites or types in relation to antibodies against human papillomaviruses (HPV)-16, -18 and -45. For each cancer site or type, the controlgroup comprised all other cancers excluding those known, or thought to be associated with HPV infection (cancers of the uterine cervix, penis and eye). Among controls the seroprevalence of antibodies was 11% (68/616) against H PV-16, 5% (29/605) against HPV-18 and 6% (35/605) against HPV-45. Antibodies against HPV-16 were significantly associated with only two cancers: uterine cervix [prevalence of antibodies 27% (51/191); odds ratio (OR) 2.0, 95% confidence interval (Cl) 1.2-3.1, P=0.011 and penis [prevalence of antibodies 27% (4/15); OR 6.4, 95% Cl 1.7-24.3, P=0.01]. For both cancers, the risk Increased with increasing anti-HPV-16 antibody titre (P t r e n d =0.01 for each). No cancer site or type was significantly associated with antibodies against HPV-18 and -45.

Using synthetic peptides we have shown that positively charged sequences present at the C terminus of the L1 protein and the N and C termini of the L2 protein of human papillomavirus type 16 (HPV-16) bind to both DNA and heparan sulfate receptors. Moreover, these short amino acid sequences are sufficient to mediate gene transfer in COS-7 cells. The L1 proteins of other HPVs were shown to contain one or two DNA- and heparin-binding sequences that have the capacity to transfer genes. These DNA-binding sequences also recognized the enhancing packaging sequence of bovine papillomavirus type 1. The results suggest that the L2 protein could participate in DNA packaging during maturation of virions.

As part of a larger investigation of cancer in Uganda, we conducted a case–control study of conjunctival squamous cell carcinoma in adults presenting at hospitals in Kampala. Participants were interviewed about social and lifestyle factors and had blood tested for antibodies to HIV, KSHV and HPV-16, -18 and -45. The odds of each factor among 60 people with conjunctival cancer was compared to that among 1214 controls with other cancer sites or types, using odds ratios, estimated with unconditional logistic regression. Conjunctival cancer was associated with HIV infection (OR 10.1, 95% confidence intervals [CI] 5.2–19.4; P

The metabolic adaptation of strong mutator strains was studied to better understand the link between the strong mutator phenotype and virulence. Analysis of the growth curves of isogenic strains of Salmonella, which were previously grown in M63 glucose media, revealed that the exponential phase of growth was reached earlier in an M63 acetate medium with strong mutator strains (mutated in mutS or in mutL) than with normomutator strains (PSalmonella and that acetate is the principal short chain fatty acid of the human terminal ileum and colon where Salmonella infection is localized.

The major structural protein (VP1) of the BK polyomavirus (BKV) was expressed in the recombinant baculovirus expression system. Recombinant BKV VP1 was shown to self-assemble into virus-like particles (VLPs) with a diameter of 45–50 nm. As for other polyomaviruses, BKV VP1 has the capacity to bind to exogenous DNA. Furthermore, the potential of BKV VP1 VLPs was investigated for gene transfer into COS-7 cells using three methods for the formation of pseudo-virions: disassembly/reassembly, osmotic shock and direct interaction between VLPs and reporter plasmid DNA. The latter method was shown to be the most efficient when using linearized plasmid. Gene transfer efficiency with BKV pseudo-virions was of the same order as that observed with human papillomavirus type 16 L1 protein VLPs. In addition, it is demonstrated that cellular entry of BKV pseudo-virions is dependent on cell surface sialic acid.

International audience; In this prospective study, a strong mutator strain of Salmonella Typhimurium was isolated from a collection of 130 human clinical strains of Salmonella. Sequence analysis of the mutS, mutL, and mutH genes, which encode three proteins that are essential for initiation of methyl-directed DNA mismatch repair, revealed insertion of a short tandem repeat (STR) of leucine/alanine in the histidine kinase-like ATPase domain of MutL. The role of this STR in the acquisition of the strong mutator phenotype was confirmed by the construction of an isogenic mutant (6bpinsmutL) from a normomutator strain of Salmonella Heidelberg. This result adds to the sparse body of knowledge about strong mutators and highlights the role of this STR as a hotspot for the acquisition of a strong mutator phenotype in Salmonella.

In bacteria, normal mutation frequencies are mostly around 10 −10 per base pair. However, there exists natural isolates, called “mutators,” that exhibit permanent mutation occurrences up to 1,000-fold greater than usual. As mutations play essential roles, particularly in the evolution of antibiotic resistance, bacteria showing elevated mutation rates could have an important responsibility in the emergence of antibiotic resistance, especially in the clinical background. In this announcement, we report the first complete genome sequence of the Salmonella enterica subsp. enterica serotype Heidelberg B182 mutator strain, isolated from bovine feces (France), which consists of a 4,750,465-bp circular chromosome (cB182_4750; GC, 52.2%) and one circular plasmid of 37,581 bp (pB182_37; GC, 42.8%).

International audience; Heritable hypermutation in bacteria is mainly due to alterations in the methyl-directed mismatch repair (MMR) system. MMR-deficient strains have been described from several bacterial species, and all of the strains exhibit increased mutation frequency and recombination, which are important mechanisms for acquired drug resistance in bacteria. Antibiotics select for drug-resistant strains and refine resistance determinants on plasmids, thus stimulating DNA recombination via the MMR system. Antibiotics can also act as indirect promoters of antibiotic resistance by inducing the SOS system and certain error-prone DNA polymerases. These alterations have clinical consequences in that efficacious treatment of bacterial infections requires high doses of antibiotics and/or a combination of different classes of antimicrobial agents. There are currently few new drugs with low endogenous resistance potential, and the development of such drugs merits further research.

International audience; Citation Hubert P, Doyen J, Capelle X, Arafa M, Renoux V, Bisig B, Seidel L, Evrard B, Bousarghin L, Gerday C, Boniver J, Foidart J-M, Delvenne P, Jacobs N. Local applications of GM-CSF induce the recruitment of immune cells in cervical low-grade squamous intraepithelial lesions. Am J Reprod Immunol 2010; 64: 126–136 Problem Quantitative alterations of antigen-presenting cells (APC) in (pre)neoplastic lesions of the uterine cervix associated with human papillomavirus (HPV) infection suggest a diminished capacity to capture viral antigens and to induce a protective immune response. Method of study To test whether a cervical application of GM-CSF could restore an immune response against HPV in women with cervical low-grade squamous intraepithelial lesions (LSIL), we performed two clinical trials with 11 healthy women and 15 patients with LSIL. Results GM-CSF applications were well tolerated in all enrolled women, and no difference in toxicity between the treated and placebo groups was observed during the follow-up (until 30 months). Interestingly, in the GM-CSF treated group, a significant increase of APC and cytotoxic T-lymphocyte infiltration was observed in the cervical biopsies with no change in regulatory T cell numbers. All the HPV16+ patients exhibited an immune response against HPV16 after GM-CSF applications, as shown by NK and/or T cells producing IFN-γ whereas no cellular immune response was observed before the treatment. Moreover, the anti-virus-like particles antibody titers also increased after the treatment. Conclusion These encouraging results obtained from a limited number of subjects justify further study on the therapeutic effect of APC in cervical (pre)neoplastic lesions

Hypermutation is an important mechanism used by different Salmonella enterica subspecies enterica to regulate genetic stability in adaptation to changing environments, including antimicrobial treatments and industrial processes. Strong hypermutator strains generally contain a mutation in genes of the methyl mismatch repair (MMR) system and have mutation frequencies up to 1000-fold higher than wild type strains. The objectives of this study were to determine the distribution of mutation frequencies from a collection of 209 Salmonella strains, to genetically characterize a strong mutator, and to study MMR mutated protein-DNA binding interactions. Only one strain of S. Heidelberg was determined to have a hypermutator phenotype by virtue of its high mutation rate. Sequencing of genes of the MMR system showed a 12bp deletion in the mutS gene was present. The MMR mutated protein-DNA binding interactions were studied by bioanalysis, using the available crystal structure of a similar MutS protein from Escherichia coli. This analysis showed the small deletion in the Salmonella MutS was localized within the core domain. A retardation assay with MutS from hypermutable and wild type strains showed this mutation has no effect on MutS DNA binding. A better understanding of the genetic mechanisms of hypermutation will help to anticipate the behavior of hypermutator strains in various conditions.

Overexpression of human papillomavirus (HPV E6 and HPV E7) oncogenes in human cervical cells results in the development of cancer, and E6 and E7 proteins are therefore targets for preventing cervical cancer progression. Here, we describe the silencing of E6 and E7 expression in cervical carcinoma cells by RNA interference. In order to increase the efficacy of the RNA interference, HPV pseudovirions coding for a short hairpin RNA (shRNA) sequence were produced. The results indicated the degradation of E6 and E7 mRNAs when shRNA against E6 or E7 were delivered by pseudovirions in HPV-positive cells (CaSki and TC1 cells). E6 silencing resulted in the accumulation of cellular p53 and reduced cell viability. More significant cell death was observed when E7 expression was suppressed. Silencing E6 and E7 and the consequences for cancer cell growth were also investigated in vivo in mice using the capacity of murine TC1 cells expressing HPV-16 E6 and E7 oncogenes to induce fast-growing tumors. Treatment with lentiviruses and HPV virus-like particle vectors coding for an E7 shRNA sequence both resulted in dramatic inhibition of tumor growth. These results show the ability of pseudovirion-delivered shRNA to produce specific gene suppression and provide an effective means of reducing HPV-positive tumor growth. [Mol Cancer Ther 2009;8(2):357–65]

Les resultats de tolerance et d’immunogenicite obtenus avec les vaccins anti-papillomavirus composes de pseudo-particules virales sont tres encourageants. Les anticorps persistent pendant 4 a 5 ans a un titre au moins superieur a celui observe lors d’une infection naturelle. Cependant, des questions sont encore en suspens, comme la duree de la protection et la necessite d’injections de rappel au-dela de 5 ans, ou le titre en anticorps anti-HPV protectif. Cette vaccination devrait conduire en quelques decennies a une reduction significative des cancers du col de l’uterus et a plus court terme a une reduction des traitements du fait d’une reduction importante des lesions precancereuses.

Langerhans' cells (LCs) are a subset of immature dendritic cells (DCs) and play a key role in the initiation and regulation of immune responses. Functional studies of these cells have been hampered by difficulties in generating a large number of LCs in vitro. We describe a new method to efficiently generate immature DCs exhibiting morphological, immunohistochemical, and ultrastructural features of LCs (CD1a + , Birbeck Granules + , CD207 + , E-cadherin + , cutaneous lymphocyte-associated antigen + , and CCR6 + ) from a limited number of CD34 + cord blood progenitors. This method is based on a two-step procedure consisting of an amplification phase followed by a terminal differentiation induction. The amplification step is initiated with a combination of hematopoietic growth factors (thrombopoietin/stem cell factor/fetal liver tyrosine kinase-3 ligand), cytokines (granulocyte-macrophage colony-stimulating factor, tumor necrosis factor-α, and interleukin-4), and 5 ng/ ml of transforming growth factor (TGF)-β1. The differentiation is induced by increasing the concentration of TGF-β1 to 12.5 ng/ml. These culture conditions were efficient for generating a large number of immature LCs (8.74 x 10 6 ± 3.2) from 15 x 10 4 CD34 + progenitor cells. In addition, these LCs were shown to be able to infiltrate an in vitro reconstructed epithelium. Because LCs play an important role in the mucosal immunity, this technique could be useful to study their interactions with epithelial pathogenic agents and to perform pharmacological, toxicological, and clinical research.

Although human papillomavirus (HPV) DNA is detected in the majority of squamous intraepithelial lesions (SILs) and squamous cell carcinomas (SCCs) of the uterine cervix, the persistence and progression of cervical lesions suggest that viral antigens are not adequately presented to the immune system. This hypothesis is reinforced by the observation that most SILs show quantitative and functional alterations of Langerhans cells (LCs). The aim of this study was to determine whether modulation of E-cadherin-mediated homophilic and heterotypic interactions between keratinocytes and LCs is involved in these abnormalities of LCs in (pre)neoplastic cervical epithelium. Cell membrane expression of E-cadherin and the density of CD1a+ LCs were low in the epithelium of SILs and SCC biopsy specimens, compared with normal exocervical epithelium. Dendritic cells (DCs) and LCs generated in vitro were randomly distributed throughout the full thickness of organotypic cultures of E-cadherin− HPV-transformed cells. In contrast, these cells rapidly adhered to the keratinocyte cell layers when HPV-transformed cells transfected with E-cadherin were used. These data suggest that the E-cadherin-mediated contact between keratinocytes and LCs is potentially important for initiating or maintaining the immune response during chronic HPV infection. Copyright © 2005 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.

Langerhans cells (LC), the immature dendritic cells (DC) that reside in epithelial tissues are among the first immune cells to encounter human papillomavirus (HPV) and are not activated by HPV virus-like particles (VLPs) in contrast to DC. The notion that the differences in response to HPV VLPs between LC and DC are associated with different types of cell binding and intracellular trafficking has been addressed. Inhibition experiments with heparin and sodium chlorate showed that heparan sulfates are necessary for HPV 16 VLPs to bind to DC but not to LC. Electron microscopy analysis demonstrated a colocalization of HPV 16 VLPs and langerin, which is expressed only by LC. This colocalization was observed on the cell surface but also in cytoplasmic vesicles. As anti-langerin antibodies, HPV 16 VLPs were associated with a faster entry kinetics in LC, as reflected by the fact that VLPs were observed near the nuclear membrane of LC within 10 min whereas more than 60 min were needed in DC. However, no difference between LC and DC was observed for the endocytosis pathway. HPV 16 VLPs entered in both DC and LC by a clathrin-dependent-pathway and were then localized in large cytoplasmic vesicles resembling endosomes.

Human papillomavirus (HPV) virus-like particles (VLP) and synthetic peptides corresponding to positively-charged sequences of the major and minor capsid proteins were tested for their efficacy in inhibiting the infectivity of HPV 31 pseudovirions by blocking virus entry into cells. A greater than 80% reduction of transfection was observed with one HPV-31 peptide at a concentration of 10 microg/ml. Moreover, the blocking was not type-specific since similar reduction in transfection was observed with peptides from other HPV types at a concentration of 60 microg/ml. This concentration was non-toxic for the cells. These findings indicate that some of the positively-charged sequences of the L1 and L2 HPV capsid proteins of papillomavirus are compounds that might be locally active against sexually transmitted papillomavirus. The findings provide further evidence that cellular glycosamino-glycans (GAGs) are functional receptors for HPVs.

The early steps of the intracellular trafficking of human papillomavirus type 16 (HPV-16), -31, and -58 pseudovirions were studied by investigating the effects of drugs acting at defined points of endocytosis pathways on virus-like particle-mediated pseudoinfection by overexpression of a dominant-negative form of the Eps15 protein to inhibit clathrin-mediated endocytosis and by electron microscopy. The results obtained suggested the involvement of clathrin-mediated endocytosis in HPV-16 and HPV-58 entry and caveola-mediated endocytosis in HPV-31 entry.

As part of a larger investigation of cancer in Uganda, we conducted a case-control study of conjunctival squamous cell carcinoma in adults presenting at hospitals in Kampala. Participants were interviewed about social and lifestyle factors and had blood tested for antibodies to HIV, KSHV and HPV-16, -18 and -45. The odds of each factor among 60 people with conjunctival cancer was compared to that among 1214 controls with other cancer sites or types, using odds ratios, estimated with unconditional logistic regression. Conjunctival cancer was associated with HIV infection (OR 10.1, 95% confidence intervals [CI] 5.2-19.4; P0.001), and was less common in those with a higher personal income (OR=0.4, 95% CI 0.3-0.7; P0.001)[corrected]. The risk of conjunctival cancer increased with increasing time spent in cultivation and therefore in direct sunlight (chi2 trend=3.9, P=0.05), but decreased with decreasing age at leaving home (chi2 trend=3.9, P=0.05), perhaps reflecting less exposure to sunlight consequent to working in towns, although both results were of borderline statistical significance. To reduce confounding, sexual and reproductive variables were examined among HIV seropositive individuals only. Cases were more likely than controls to report that they had given or received gifts for sex (OR 3.5, 95% CI 1.2-10.4; P=0.03), but this may have been a chance finding as no other sexual or reproductive variable was associated with conjunctival cancer, including the number of self-reported lifetime sexual partners (P=0.4). The seroprevalence of antibodies against HPV-18 and -45 was too low to make reliable conclusions. The presence of anti-HPV-16 antibodies was not significantly associated with squamous cell carcinoma of the conjunctiva (OR 1.5, 95% CI 0.5-4.3; P=0.5) and nor were anti-KSHV antibodies (OR 0.9, 95% CI 0.4-2.1; P=0.8). The 10-fold increased risk of conjunctival cancer in HIV infected individuals is similar to results from other studies. The role of other oncogenic viral infections is unclear.

The neutralizing activities of polyclonal antibodies and monoclonal antibodies (MAbs) obtained by immunization of mice with L1 virus-like particles (VLPs) were investigated by using pseudovirion infectivity assays for human papillomavirus type 16 (HPV-16), HPV-31, HPV-33, HPV-45, HPV-58, and HPV-59 to obtain a better definition of cross-neutralization between high-risk HPVs. In this study, we confirmed and extended previous studies indicating that most genital HPV genotypes represent separate serotypes, and the results suggest that the classification of serotypes is similar to that of genotypes. In addition, three cross-neutralizing MAbs were identified (HPV-16.J4, HPV-16.I23, and HPV-33.E12). MAb HPV-16.J4 recognized a conserved linear epitope located within the FG loop of the L1 protein, and HPV-16.I23 recognized another located within the DE loop. The results suggested that reactivity of MAb HPV-16.I23 to L1 protein is lost when leucine 152 of the HPV-16 L1 protein is replaced by phenylalanine. This confirmed the existence of linear epitopes within the L1 protein that induce neutralizing antibodies, and this is the first evidence that such linear epitopes induce cross-neutralization. However, the cross-neutralization induced by L1 VLPs represents less than 1% of the neutralizing activity induced by the dominant conformational epitopes, and it is questionable whether this is sufficient to offer cross-protection in vivo.

The goal of this study was to develop a human papillomavirus (HPV) neutralization assay using HPV pseudovirions generated in vitro. For this purpose, gene transfer efficiency of HPV virus-like particles (VLPs) was improved by using direct interaction between a reporter plasmid and the VLPs. Electron microscopic observation of the interaction between DNA molecules and VLPs revealed that VLPs always interact with a single DNA molecule and that VLPs bind to the end of linearized DNA molecules. An 100-fold improvement in the gene transfer was obtained by simple interaction between a linearized DNA molecule and VLPs. Moreover, direct interaction methods offer the possibility of transferring plasmids a size higher than that of the papillomavirus genome. The approach that we developed to generate HPV-16 and HPV-31 pseudovirions proved to be suitable for testing neutralizing antibodies in human sera both after immunization and after natural infection.

Artificial viruses consisting of DNA plasmid packaged in vitro into virus-like particles (VLPs) are new vehicles for gene transfer. We therefore investigated the ability of nine human papillomavirus (HPV) VLPs to interact with heterologous DNA and transfer genes. HPV 16, 18, 31, 33, 39, 45, 58, 59, and 68 VLPs were able to bind heterologous DNA and to transfer genes into Cos-7 cells. Inhibition of gene transfer by preincubation of the pseudovirions with heparin confirmed that heparan sulfate on the cell surface plays a role as cell receptor for HPVs. As HPV neutralizing antibodies are mainly type-specific, gene transfer with different HPV pseudovirions offers the possibility of their sequential use in vivo for a greater efficacy.

Four C-terminal deletion mutants of the human papillomavirus 16 L1 protein were expressed in the baculovirus expression system. They consist of the deletion of amino acids 497-505, 477-505, 403-505 and 302-505 (delta C9, delta C31, delta C103 and delta C204 respectively). Only two of the C-terminally deleted proteins, delta C9 and delta C31, retained the ability to form virus-like particles (VLPs) resembling those obtained with the full length L1 protein. Analysis of deleted L1 proteins and corresponding VLPs indicated that the C-terminus was necessary both for DNA binding and DNA packaging. These results were corroborated by the loss of the gene transfer capacities of C-terminal deleted VLPs.

Correction to: British Journal of Cancer (2002) 87, 301–308. doi:10.1038/sj.bjc.6600451 In the sixth line of the abstract, the confidence interval in relation to personal income is incorrect. It should read: OR=0.4, 95% CI 0.3–0.7; P

La sclérose en plaques est une maladie neurodégénérative du système nerveux central. Comme son nom l’indique, elle est caractérisée par des plaques qui correspondent aux zones démyélinisées, responsables d’une altération de la conduction de l’influx nerveux. Son étiologie, encore mal connue, résulterait d’un ensemble de facteurs génétiques et environnementaux. Récemment,le rôle du microbiote intestinal dans la sclérose en plaques a été mis en évidence. Le lien entre le microbiote intestinal et le système nerveux est possible grâce à l’axe intestin-cerveau, qui permet une communication bidirectionnelle. Cette communication va être perturbée par la présence d’une dysbiose, avec une augmentation de l’abondance d’Akkermansia, Methanobrevibacter, Streptococcus et Bifidobacterium, et une diminution de Prevotella. Dans ce travail, nous avons regroupé les études les plus récentes, qui montrent un lien entre la dysbiose et l’impact dans la sclérose en plaques. Ces études ont montré le rôle du microbiote intestinal dans l’augmentation de l’inflammation, avec une augmentation des lymphocytes Th1 et Th17, et une diminution des lymphocytes Treg. Ces dernières années, des études font espérer de nouvelles perceptives thérapeutiques, par l’utilisation de probiotiques.

Pseudomonas aeruginosa est une bactérie ubiquitaire dans l'environnement. Grace à de nombreux facteurs de virulence, elle est capable d'infecter un large spectre d'hôtes, dont l'homme. Ces facteurs de virulence jouent un rôle déterminant dans la pathogenèse des infections dues à P. aeruginosa et dans la résistance de la bactérie aux traitements. P. aeruginosa est une bactérie opportuniste, elle est peu virulente chez les individus sains, mais elle est à l'origine de nombreuses infections chez les sujets les plus sensibles. La bactérie est notamment responsable d'infections pulmonaires, d'infections cutanées, d'infections urinaires et de bactériémies. P. aeruginosa est responsable de 8,4 % des infections nosocomiales en France. La prise en charge des infections dues à P. aeruginosa représente un enjeu de santé publique majeur de par la sévérité des infections et l'augmentation de la résistance aux antibiotiques de la bactérie. Le faible nombre de nouveaux traitements ainsi que l'absence de prophylaxie augmentent d'autant plus le risque d'impasse thérapeutique. Une attention accrue du respect du bon usage des antibiotiques demeure essentielle afin d'éviter la sélection des germes résistants dans l'attente de nouvelles thérapies.

The oral bacteria do not only infect the mouth and reside there, but also travel via the blood and reach distant body organs. If left untreated, the dental biofilm that can cause destructive inflammation in the oral cavity may result in serious systemic medical complications. In dental biofilm, Streptococcus gordonii, a primary oral colonizer, constitutes the platform on which late pathogenic colonizers like Porphyromonas gingivalis, the causative agent of periodontal diseases, will bind. The aim of the first study was to determine the antibacterial activity of eleven natural lichen compounds belonging to different chemical families to uncover new antibiotics which can fight against the oral bacteria. Three compounds were shown to have promising antibacterial activities where psoromic acid had the lowest MICs of 11.72 and 5.86 µg/mL against S. gordonii and P. gingivalis, respectively. Novel butyrolactone analogues were then designed and synthesized based on the known lichen antibacterial compounds, lichesterinic acids (B-10 and B-11), by substituting different functional groups on the butyrolactone ring trying to enhance its activity on S. gordonii and P. gingivalis.. Among the derivatives, B-12 and B-13 had the lowest MIC of 9.38 µg/mL where they have shown to be stronger bactericidals, by 2-3 times, than the reference antibiotic, doxycycline. B-12 and B-13 were also the most efficient on P. gingivalis exhibiting MIC of 0.037 and 0.293 µg/mL and MBC of 1.17 and 0.586 µg/mL, respectively. These 2 compounds were then checked for their cytotoxicity against human gingival epithelial cells and macrophages by MTT and LDH assays which confirmed their safety against the tested cell lines. A preliminary study of the structure-activity relationships unveiled the important dual role contributed by two substituents, alkyl chain at C4 and carboxyl group at C5 positions, in their mechanism of action. This was followed by the investigation of B-12 and B-13 for their antibiofilm activity against both oral strains using crystal violet assay and confocal microscopy. Both derivatives displayed a lowest concentration with maximal biofilm inhibition, LCMI, of 9.38 µg/mL against S. gordonii and 1.17 µg/mL against P. gingivalis. However, when sub-inhibitory concentrations of B-12 and B-13 were used, we demonstrated that the two investigated strains were able to form biofilms in vitro. Indeed, this antibiofilm activity decreased as indicated by the expression of the genes implicated in adhesion and biofilm formation. To better understand the mechanism of action of butyrolactones, we have investigated B-13 bacterial localization by synthesizing a fluorescently labeled B-13 with NBD (4-nitro-benzo[1,2,5]oxadiazole) conserving its antibacterial activity. By confocal microscope, we showed that this compound binds to S. gordonii cell surface and this was also demonstrated by HPLC analysis. By adhering to cell surface, B-13 induced cell wall disruption leading to the release of bacterial constituents and consequently, the death of S. gordonii, a Gram-positive bacterium. The expression of two genes, murA and alr, implicated in cell wall synthesis, was modified in the presence of this butyrolactone. Gram-negative bacteria such as P. gingivalis showed also cracked and ruptured cells in the presence of B-13, suggesting that this butyrolactone acts on Gram-positive and Gram-negative strains, but with greater efficacy against the Gram-negatives. Besides, we also demonstrated that the analogue of B-13, B-12, has also induced disruption of P. gingivalis and S. gordonii. All these studies demonstrated that butyrolactones derived from a lichen metabolite can be proposed as potent antibacterial agents against oral pathogens causing serious medical complications.; Les bactéries buccales n'infectent pas seulement la bouche mais y resident. Elles peuvent également passer dans la voie sanguine et atteindre des organes secondaires. S’il n'est pas traité, le biofilm dentaire peut provoquer une inflammation destructrice dans la cavité buccale, entrainant de graves complications médicales. Dans ce biofilm, Streptococcus gordonii, colonisateur oral primaire, constitue la plate-forme sur laquelle des colonisateurs pathogènes tardifs comme Porphyromonas gingivalis, l'agent causal des maladies parodontales, se lieront. L'objectif de la première partie de la thèse était de déterminer l'activité antibactérienne de onze composés de lichens appartenant à différentes familles chimiques, pour découvrir de nouveaux antibiotiques pouvant combattre ces bactéries buccales. Nous avons montré que trois composés avaient des activités antibactériennes prometteuses. L'acide psoromique enregistrait les CMIs le plus faibles. De nouveaux analogues de butyrolactone ont ensuite été conçus et synthétisés sur la base des composés antibactériens licheniques connus, les acides lichesteriniques, en substituant différents groupes fonctionnels sur le cycle butyrolactone pour améliorer son activité sur S. gordonii et P. gingivalis. Parmi les dérivés, B-12 et B-13 avaient la plus faible CMI où ils se sont révélés être des bactéricides plus forts, 2 à 3 fois plus, que l'antibiotique, doxycycline. B-12 et B-13 étaient également les plus efficaces vis-à-vis de P. gingivalis. La cytotoxicité de ces 2 composés a ensuite été vérifiée contre les cellulaires épithéliales gingivales humaines et les macrophages. Ils ne présentaient pas de toxicité contre les cellules testées. Une étude préliminaire de relation structure-activité a révélé le double rôle important apporté par deux substituants, chaîne alkyle en C5 et groupe carboxyle en C4 positions, dans leur mécanisme d'action. Ceci a été suivi par l'étude de l’activité antibiofilmique de B-12 et B-13 contre les deux souches orales en utilisant un test de cristal violet et microscopie confocale. Les deux dérivés ont montré, à une concentration plus faible, une inhibition maximale de la formation du biofilm, LCMI, de 9.38 μg/mL contre S. gordonii et 1.17 μg/mL contre P. gingivalis. Cependant, lorsque des concentrations sous-inhibitrices de B-12 et B-13 ont été utilisées, nous avons démontré que les deux souches étudiées pouvaient former des biofilms in vitro, accompagné d’une diminution de l'expression des gènes impliqués dans l'adhésion et la formation de biofilm. Pour mieux comprendre les mécanismes d'action des butyrolactones, nous avons étudié la localisation bactérienne du composé B-13 en synthétisant un B-13 marqué au NBD (4-nitro-benzo [1,2,5] oxadiazole) fluorescent conservant son activité antibactérienne. Par microscopie confocale et HPLC, nous avons montré que ce composé se lie à la surface cellulaire de S. gordonii. Ensuite, B-13 induit une rupture de la paroi cellulaire conduisant à la libération des constituants bactériens et par conséquent, à la mort de S. gordonii, une bactérie Gram-positive. L'expression de deux gènes, murA et alr, impliqués dans la synthèse de la paroi cellulaire, a été modifiée en présence de cette butyrolactone. Les bactéries Gram négatives telles que P. gingivalis ont également montré des cellules abimées présentant une rupture de la paroi en présence de B-13, ce qui suggère que cette butyrolactone agit sur des Gram-positives et Gram-négatives avec une plus grande efficacité contre les Gram-négatives. En outre, nous avons également démontré que l'analogue de B-13, B-12, induit une perturbation de la morphologie de P. gingivalis et S. gordonii. Toutes ces études ont démontré que les butyrolactones dérivées de lichen peuvent être proposés comme des composés antibactériens puissants contre les agents pathogènes oraux qui causent des complications médicales graves.

The oral bacteria do not only infect the mouth and reside there, but also travel via the blood and reach distant body organs. If left untreated, the dental biofilm that can cause destructive inflammation in the oral cavity may result in serious systemic medical complications. In dental biofilm, Streptococcus gordonii, a primary oral colonizer, constitutes the platform on which late pathogenic colonizers like Porphyromonas gingivalis, the causative agent of periodontal diseases, will bind. The aim of the first study was to determine the antibacterial activity of eleven natural lichen compounds belonging to different chemical families to uncover new antibiotics which can fight against the oral bacteria. Three compounds were shown to have promising antibacterial activities where psoromic acid had the lowest MICs of 11.72 and 5.86 µg/mL against S. gordonii and P. gingivalis, respectively. Novel butyrolactone analogues were then designed and synthesized based on the known lichen antibacterial compounds, lichesterinic acids (B-10 and B-11), by substituting different functional groups on the butyrolactone ring trying to enhance its activity on S. gordonii and P. gingivalis.. Among the derivatives, B-12 and B-13 had the lowest MIC of 9.38 µg/mL where they have shown to be stronger bactericidals, by 2-3 times, than the reference antibiotic, doxycycline. B-12 and B-13 were also the most efficient on P. gingivalis exhibiting MIC of 0.037 and 0.293 µg/mL and MBC of 1.17 and 0.586 µg/mL, respectively. These 2 compounds were then checked for their cytotoxicity against human gingival epithelial cells and macrophages by MTT and LDH assays which confirmed their safety against the tested cell lines. A preliminary study of the structure-activity relationships unveiled the important dual role contributed by two substituents, alkyl chain at C4 and carboxyl group at C5 positions, in their mechanism of action. This was followed by the investigation of B-12 and B-13 for their antibiofilm activity against both oral strains using crystal violet assay and confocal microscopy. Both derivatives displayed a lowest concentration with maximal biofilm inhibition, LCMI, of 9.38 µg/mL against S. gordonii and 1.17 µg/mL against P. gingivalis. However, when sub-inhibitory concentrations of B-12 and B-13 were used, we demonstrated that the two investigated strains were able to form biofilms in vitro. Indeed, this antibiofilm activity decreased as indicated by the expression of the genes implicated in adhesion and biofilm formation. To better understand the mechanism of action of butyrolactones, we have investigated B-13 bacterial localization by synthesizing a fluorescently labeled B-13 with NBD (4-nitro-benzo[1,2,5]oxadiazole) conserving its antibacterial activity. By confocal microscope, we showed that this compound binds to S. gordonii cell surface and this was also demonstrated by HPLC analysis. By adhering to cell surface, B-13 induced cell wall disruption leading to the release of bacterial constituents and consequently, the death of S. gordonii, a Gram-positive bacterium. The expression of two genes, murA and alr, implicated in cell wall synthesis, was modified in the presence of this butyrolactone. Gram-negative bacteria such as P. gingivalis showed also cracked and ruptured cells in the presence of B-13, suggesting that this butyrolactone acts on Gram-positive and Gram-negative strains, but with greater efficacy against the Gram-negatives. Besides, we also demonstrated that the analogue of B-13, B-12, has also induced disruption of P. gingivalis and S. gordonii. All these studies demonstrated that butyrolactones derived from a lichen metabolite can be proposed as potent antibacterial agents against oral pathogens causing serious medical complications.; Les bactéries buccales n'infectent pas seulement la bouche mais y resident. Elles peuvent également passer dans la voie sanguine et atteindre des organes secondaires. S’il n'est pas traité, le biofilm dentaire peut provoquer une inflammation destructrice dans la cavité buccale, entrainant de graves complications médicales. Dans ce biofilm, Streptococcus gordonii, colonisateur oral primaire, constitue la plate-forme sur laquelle des colonisateurs pathogènes tardifs comme Porphyromonas gingivalis, l'agent causal des maladies parodontales, se lieront. L'objectif de la première partie de la thèse était de déterminer l'activité antibactérienne de onze composés de lichens appartenant à différentes familles chimiques, pour découvrir de nouveaux antibiotiques pouvant combattre ces bactéries buccales. Nous avons montré que trois composés avaient des activités antibactériennes prometteuses. L'acide psoromique enregistrait les CMIs le plus faibles. De nouveaux analogues de butyrolactone ont ensuite été conçus et synthétisés sur la base des composés antibactériens licheniques connus, les acides lichesteriniques, en substituant différents groupes fonctionnels sur le cycle butyrolactone pour améliorer son activité sur S. gordonii et P. gingivalis. Parmi les dérivés, B-12 et B-13 avaient la plus faible CMI où ils se sont révélés être des bactéricides plus forts, 2 à 3 fois plus, que l'antibiotique, doxycycline. B-12 et B-13 étaient également les plus efficaces vis-à-vis de P. gingivalis. La cytotoxicité de ces 2 composés a ensuite été vérifiée contre les cellulaires épithéliales gingivales humaines et les macrophages. Ils ne présentaient pas de toxicité contre les cellules testées. Une étude préliminaire de relation structure-activité a révélé le double rôle important apporté par deux substituants, chaîne alkyle en C5 et groupe carboxyle en C4 positions, dans leur mécanisme d'action. Ceci a été suivi par l'étude de l’activité antibiofilmique de B-12 et B-13 contre les deux souches orales en utilisant un test de cristal violet et microscopie confocale. Les deux dérivés ont montré, à une concentration plus faible, une inhibition maximale de la formation du biofilm, LCMI, de 9.38 μg/mL contre S. gordonii et 1.17 μg/mL contre P. gingivalis. Cependant, lorsque des concentrations sous-inhibitrices de B-12 et B-13 ont été utilisées, nous avons démontré que les deux souches étudiées pouvaient former des biofilms in vitro, accompagné d’une diminution de l'expression des gènes impliqués dans l'adhésion et la formation de biofilm. Pour mieux comprendre les mécanismes d'action des butyrolactones, nous avons étudié la localisation bactérienne du composé B-13 en synthétisant un B-13 marqué au NBD (4-nitro-benzo [1,2,5] oxadiazole) fluorescent conservant son activité antibactérienne. Par microscopie confocale et HPLC, nous avons montré que ce composé se lie à la surface cellulaire de S. gordonii. Ensuite, B-13 induit une rupture de la paroi cellulaire conduisant à la libération des constituants bactériens et par conséquent, à la mort de S. gordonii, une bactérie Gram-positive. L'expression de deux gènes, murA et alr, impliqués dans la synthèse de la paroi cellulaire, a été modifiée en présence de cette butyrolactone. Les bactéries Gram négatives telles que P. gingivalis ont également montré des cellules abimées présentant une rupture de la paroi en présence de B-13, ce qui suggère que cette butyrolactone agit sur des Gram-positives et Gram-négatives avec une plus grande efficacité contre les Gram-négatives. En outre, nous avons également démontré que l'analogue de B-13, B-12, induit une perturbation de la morphologie de P. gingivalis et S. gordonii. Toutes ces études ont démontré que les butyrolactones dérivées de lichen peuvent être proposés comme des composés antibactériens puissants contre les agents pathogènes oraux qui causent des complications médicales graves.

La bronchiolite est un vrai problème de santé publique puisqu'elle atteint les tous petits en grand nombre et provoque une surcharge des urgences pédiatriques. Cette maladie est principalement due au virus respiratoire syncytial qui atteint les nourrissons chaque année lors d'épidémies hivernales. Elle se manifeste par une rhinopharyngite puis une toux et une dyspnée respiratoire. À l'heure actuelle, aucun vaccin anti-VRS n'est disponible ni même de traitement spécifique. La prise en charge de cette maladie repose essentiellement sur des mesures symptomatiques. La kinésithérapie respiratoire reste l'une des méthodes les plus efficaces pour soigner cette pathologie malgré ses critiques récentes, avec la désobstruction rhinopharyngée. Le pharmacien d'officine s'insère ici dans un rôle de proximité avec les parents. Il peut ainsi leur prodiguer divers conseils utiles afin de se préserver de l’épidémie et la conduite à tenir en cas de bronchiolite chez leur enfant.

The Hepatitis C Virus (HCV) affects approximately 170 million people worldwide. In France, there are about 220 000 people with chronic hepatitis C. Loss of liver function usually leads to liver transplantation in the most extreme cases. These hepatitis are currently treated with direct acting antivirals, the AAD, specifically targeting the virus multiplication mechanisms. I was led in my thesis to follow the cohort ANRS CO-­‐‑23 CUPILT conducted in Rennes University Hospital to study the kinetics of decrease in HCV viral load in a direct acting antivirals therapy in liver transplant recurrence of HCV infection. This in order to identify predictors of decrease in viral load and slower to estimate an impact. My study has shown that two parameters are associated with a slow decrease in viral load. This is a high viral load at baseline and genotype 1a. Furthermore, this study failed to show an impact on the kinetic decrease viral load on treatment efficacy.; Le Virus de l’hépatite C (VHC) touche 170 millions de personnes dans le monde. En France environ 220 000 personnes sont atteintes d'une hépatite C chronique. La perte de fonctionnalité hépatique conduit le plus souvent à une transplantation hépatique dans les cas les plus extrêmes. Ces hépatites sont actuellement traitées avec des antiviraux d'action directe, les AAD, ciblant plus précisément les mécanismes de multiplication du virus. J'ai été amené au cours de ma thèse à suivre la cohorte ANRS CO-23 CUPILT du CHU de Rennes, pour étudier les cinétiques de décroissance de la charge virale du VHC au cours d'un traitement par antiviraux d'action directe chez les transplantés hépatiques en récidive de l'infection par le VHC. Ceci dans le but d’identifier des facteurs prédictifs de décroissance de la charge virale lente et d'en estimer un impact. Mon étude a permis de montrer que deux paramètres sont associés à une décroissance de la charge virale lente. Il s'agit d’une charge virale élevée à l'inclusion et du génotype 1a. Par ailleurs cette étude n'a pas permis de mettre en évidence un impact de la cinétique de décroissance virale sur lefficacité des traitements.