Pre-exposure prophylaxis with OspA-specific human monoclonal antibodies protects mice against tick transmission of Lyme disease spirochetes

Background. Tick transmission of Borrelia spirochetes to humans results in significant morbidity from Lyme disease worldwide. Serum concentrations of antibodies against outer surface protein A (OspA) were shown to correlate with protection from infection with Borrelia burgdorferi, the primary cause of Lyme disease in the United States. Methods. Mice transgenic for human immunoglobulin genes were immunized with OspA protein of B. burgdorferi to generate human monoclonal antibodies (HuMabs) against OspA. HuMabs were generated and tested in in vitro borreliacidal assays and animal protection assays. Results. Nearly 100 unique OspA specific HuMabs were generated and four HuMabs (221-7, 857-2, 319-44, and 212-55) were selected as lead candidates based on borreliacidal activity. HuMab 319-44, 857-2 and 212-55 were borreliacidal against one or two Borrelia genospecies, whereas 221-7 was borreliacidal (IC50 \u3c 1nM) against B. burgdorferi, B. afzelii and B. garinii, the three main genospecies endemic in the US, Europe and Asia. All four HuMabs completely protected mice from infection at 10 mg/kg in a murine model of tick-mediated transmission of B. burgdorferi. Conclusions. Our study indicates that OspA-specific HuMabs can prevent the transmission of Borrelia and administration of these antibodies could be employed as pre-exposure prophylaxis for Lyme disease

PLoS ONE

Lyme disease is a multisystemic disorder caused by B. burgdorferi sl. The molecular basis for specific organ involvement is poorly understood. The skin plays a central role in the development of Lyme disease as the entry site of B. burgdorferi in which specific clones are selected before dissemination. We compared the skin inflammatory response (antimicrobial peptides, cytokines and chemokines) elicited by spirochete populations recovered from patients presenting different clinical manifestations. Remarkably, these spirochete populations induced different inflammatory profiles in the skin of C3H/HeN mice. As spirochete population transmitted into the host skin is heterogeneous, we isolated one bacterial clone from a population recovered from a patient with neuroborreliosis and compared its virulence to the parental population. This clone elicited a strong cutaneous inflammatory response characterized by MCP-1, IL-6 and antimicrobial peptides induction. Mass spectrometry of this clone revealed 110 overexpressed proteins when compared with the parental population. We further focused on the expression of nine bacterial surface proteins. bb0347 coding for a protein that interacts with host fibronectin, allowing bacterial adhesion to vascular endothelium and extracellular matrix, was found to be induced in host skin with another gene bb0213 coding for a hypothetical protein. These findings demonstrate the heterogeneity of the B. burgdorferi ss population and the complexity of the interaction involved early in the skin

Microarray Analyses of Inflammation Response of Human Dermal Fibroblasts to Different Strains of Borrelia burgdorferi Sensu Stricto

In Lyme borreliosis, the skin is the key site of bacterial inoculation by the infected tick, and of cutaneous manifestations, erythema migrans and acrodermatitis chronica atrophicans. We explored the role of fibroblasts, the resident cells of the dermis, in the development of the disease. Using microarray experiments, we compared the inflammation of fibroblasts induced by three strains of Borrelia burgdorferi sensu stricto isolated from different environments and stages of Lyme disease: N40 (tick), Pbre (erythema migrans) and 1408 (acrodermatitis chronica atrophicans). The three strains exhibited a similar profile of inflammation with strong induction of chemokines (CXCL1 and IL-8) and IL-6 cytokine mainly involved in the chemoattraction of immune cells. Molecules such as TNF-alpha and NF-κB factors, metalloproteinases (MMP-1, -3 and -12) and superoxide dismutase (SOD2), also described in inflammatory and cellular events, were up-regulated. In addition, we showed that tick salivary gland extracts induce a cytotoxic effect on fibroblasts and that OspC, essential in the transmission of Borrelia to the vertebrate host, was not responsible for the secretion of inflammatory molecules by fibroblasts. Tick saliva components could facilitate the early transmission of the disease to the site of injury creating a feeding pit. Later in the development of the disease, Borrelia would intensively multiply in the skin and further disseminate to distant organs

Cutaneous inflammation during Lyme disease transmission, study on a murine model

La borréliose de Lyme est une infection bactérienne due à Borrelia burgdorferi sensu lato et transmise à l'Homme par piqûre de tique du genre Ixodes. Cette affection est caractérisée par des manifestations cliniques variées. La transmission de l'agent pathogène induit dans un premier temps une inflammation cutanée : l'érythème migrant. Après dissémination, les manifestations cliniques peuvent être de nature neurologique, articulaire ou cutanée. La peau est une interface essentielle dans la transmission du pathogène par le vecteur. Le rôle central de l'interface cutanée est abordé au cours de ce travail de thèse. D une part, nous nous sommes intéressés à la modulation de l'immunité cutanée par la salive de la tique, plus précisément à l'effet de la salive sur l'expression de peptides anti-microbiens (PAM), de cytokines pro-inflammatoires et de chimiokines. D autre part, les mécanismes responsables du tropisme préférentiel des bactéries vers les organes cibles ne sont pas connus à ce jour. Afin de répondre à ces questions, nous avons établi différents modèles expérimentaux. D une part, un modèle in vitro permettant d'analyser spécifiquement la réponse des cellules résidantes de la peau. D autre part, une approche in vivo d'infection chez des souris C3H/HeN. Au cours de ce travail, nous avons mis en évidence une immuno-modulation cutanée par la salive de tique durant la transmission de la borréliose de Lyme. Nous avons proposé un effet anti-alarmine de la salive sur la peau. Nous montrons également que la peau est un site de multiplication massive de la bactérie avant sa dissémination vers les organes cibles. De plus, la mise en évidence d'une réponse immunitaire différente selon le pathotype souligne le rôle potentiel de l'interface cutanée pour l'évolution de B. burgdorferi dans l'hôte vertébré.Lyme disease, is an infectious disorder caused by a tick-transmitted bacteria : Borrelia burgdorferi. The skin constitutes an essential interface in this arthropod borne disease. Indeed, the primary manifestation is a cutaneous inflammation, the erythema migrans. Dissemination of spirochetes from the site of inoculation can lead to other manifestations typically involving the skin, heart, joints or central nervous system. Mechanisms responsible of this specific dissemination are not known. In this project we focused on the cutaneous innate immune response during Lyme disease transmission. Part of skin innate immunity is constituted by the secretion of antimicrobial peptides (AMPs), cytokines and chemokines. We developed two experimental strategies. In vitro to measure the specific response from skin resident cells: keratinocytes. In vivo we challenged C3H/HeN mice with spirochetes from B. burgdoferi sensu stricto strains initially isolated from human clinical manifestation. In conclusion, we propose that tick saliva has a property not previously described : an anti-alarmin effect. Tick saliva is an essential actor in the pathogenesis of skin inflammation. Furthermore, we showed a clear difference in the skin innate immunity according to the strain tested. The skin by its immunity and the specificity of its different resident cells likely plays a major role in the development of Borrelia infection in the vertebrate host. There, an intense bacterial multiplication occurs. Some specific factors of both, the bacteria (like OspC and BBK32) and the host (like AMPs and MCP-1), display a sophisticated interaction that likely further orientate the bacterium in the rest of the body

Cutaneous inflammation during Lyme disease transmission, study on a murine model

La borréliose de Lyme est une infection bactérienne due à Borrelia burgdorferi sensu lato et transmise à l'Homme par piqûre de tique du genre Ixodes. Cette affection est caractérisée par des manifestations cliniques variées. La transmission de l'agent patLyme disease, is an infectious disorder caused by a tick-transmitted bacteria : Borrelia burgdorferi. The skin constitutes an essential interface in this arthropod borne disease. Indeed, the primary manifestation is a cutaneous inflammation, the erythem

Exploration de la diversité des bactéries marines dégradant les macroalgues par marquage isotopique

International audienc

Role of OspC, <i>I. ricinus</i> salivary gland extracts (SGE) and Salp15 in <i>Borrelia</i>-induced fibroblast inflammation

<p>(A) IL-8 synthesis induced by wild-type strain 297 (wt), OspC-deficient (OspC −/−), and OspC-deficient strain 297 complemented with a plasmid carrying the ospC gene (OspC cp) in fibroblasts. (B) IL-8 synthesis in fibroblasts induced by <i>B. burgdorferi</i> ss N40 (Bb) in absence or in presence of human anti-TLR2 antibody (Ab aTLR2) or isotype control antibody (Ab isotype control). (C) IL-8 synthesis in fibroblasts coincubated with 20 µg/ml SGE alone, 30 µg/ml Salp15 alone, <i>B. burgdorferi</i> ss N40 (Bb) alone, with the combination of <i>Borrelia</i> and SGE at 20 µg/ml (Bb + SGE), 5 µg/ml (Bb + SGE 1∶4), 1 µg/ml (Bb + SGE 1∶20), and 0.2 µg/ml (Bb + SGE 1∶100), with the combination of <i>Borrelia</i> and Salp15 (Bb + Salp15), or with the combination of <i>Borrelia</i> and 20 µg/ml SGE heat-denaturated at 56°C for 1 hour (Bb + SGE 56°C), or at 98°C for 3 minutes (Bb + SGE 98°C). For (A), (B) and (C) fibroblasts were incubated with <i>Borrelia</i> at MOI of 100∶1 for 24 hours. The negative control was unstimulated cells (NEG). Each bar shows the mean ± SDs of triplicate values (expressed as % stimulation of IL-8 synthesis induced by <i>Borrelia</i> alone) and is representative of three independent experiments. ***P<0.001; and *P<0.05 compared with the corresponding stimulation induced by <i>Borrelia</i> alone. (D) Images of fibroblast cell cultures stimulated with SGE, showing SGE-induced cytotoxic effect at 6 and 24 hours (h). Images were taken at 100x (I, II and III) or at 200x magnification (IV, V and VI).</p

Isotopic tracing reveals single-cell assimilation of a macroalgal polysaccharide by a few marine Flavobacteria and Gammaproteobacteria

International audienceAlgal polysaccharides constitute a diverse and abundant reservoir of organic matter for marine heterotrophic bacteria, central to the oceanic carbon cycle. We investigated the uptake of alginate, a major brown macroalgal polysaccharide, by microbial communities from kelp-dominated coastal habitats. Congruent with cell growth and rapid substrate utilization, alginate amendments induced a decrease in bacterial diversity and a marked compositional shift towards copiotrophic bacteria. We traced 13 C derived from alginate into specific bacterial incorporators and quantified the uptake activity at the single-cell level, using halogen in situ hybridization coupled to nanoscale secondary ion mass spectrometry (HISH-SIMS) and DNA stable isotope probing (DNA-SIP). Cell-specific alginate uptake was observed for Gammaproteobacteria and Flavobacteriales, with carbon assimilation rates ranging from 0.14 to 27.50 fg C µm −3 h −1. DNA-SIP revealed that only a few initially rare Flavobacteriaceae and Alteromonadales taxa incorporated 13 C from alginate into their biomass, accounting for most of the carbon assimilation based on bulk isotopic measurements. Functional screening of metagenomic libraries gave insights into the genes of alginolytic Alteromonadales active in situ. These results highlight the high degree of niche specialization in heterotrophic communities and help constraining the quantitative role of polysaccharide-degrading bacteria in coastal ecosystems

Gene expression profiles obtained from dermal fibroblasts stimulated with different strains of <i>B. burgdorferi</i> ss.

<p>(A) Venn diagram of genes significantly up-regulated (↑) or down-regulated (↓) after fibroblast stimulation with <i>Borrelia</i>, and compared with unstimulated fibroblasts. (B) Number of genes differentially expressed during fibroblast stimulation with <i>Borrelia</i>. The bars reflect the number of up-regulated genes (+) and down-regulated genes (-) for each strain. The light dotted areas correspond to gene expression changes of 1.7–5.0-fold, the grey hatched areas correspond to changes of 5.0–20.0-fold and black areas to changes ≥20.0-fold.</p

Measure of IL-8 secretion by fibroblasts co-incubated with different strains of <i>B. burgdorferi</i> ss.

<p>(A-C) IL-8 secretion of fibroblasts stimulated by different concentrations of <i>Borrelia</i> N40, Pbre, 1408 at MOI of 100∶1 (100B), 10∶1 (10B), and 1∶1 (1B) at 24 hours. (D-F) Kinetic studies of IL-8 secretions in the three strains. NEG: unstimulated fibroblasts. (A-F) Each bar shows the mean ± SDs of triplicate values and is representative of three independent experiments. ***P<0.001; **P<0.01; and *P<0.05 compared between stimulated and unstimulated cells.</p