Elevated resource availability sufficient to turn opportunistic into virulent fish pathogens.

There is mounting evidence that organic or inorganic enrichment of aquatic environments increases the risk of infectious diseases, with disease agents ranging from helminth parasites to fungal, bacterial, and viral pathogens. The causal link between microbial resource availability and disease risk is thought to be complex and, in the case of so-called "opportunistic pathogens," to involve additional stressors that weaken host resistance (e.g., temperature shifts or oxygen deficiencies). In contrast to this perception, our experiment shows that the link between resource levels and infection of fish embryos can be very direct: increased resource availability can transform benign microbial communities into virulent ones. We find that embryos can be harmed before further stresses (e.g., oxygen depletion) weaken them, and treatment with antibiotics and fungicides cancels the detrimental effects. The changed characteristics of symbiotic microbial communities could simply reflect density-dependent relationships or be due to a transition in life-history strategy. Our findings demonstrate that simple microhabitat changes can be sufficient to turn "opportunistic" into virulent pathogens.

Intestinal bacteria condition dendritic cells to promote IgA production.

Immunoglobulin (Ig) A represents the predominant antibody isotype produced at the intestinal mucosa, where it plays an important role in limiting the penetration of commensal intestinal bacteria and opportunistic pathogens. We show in mice that Peyer's Patch-derived dendritic cells (PP-DC) exhibit a specialized phenotype allowing the promotion of IgA production by B2 cells. This phenotype included increased expression of the retinaldehyde dehydrogenase 1 (RALDH1), inducible nitric oxide synthase (iNOS), B cell activating factor of the tumor necrosis family (BAFF), a proliferation-inducing ligand (APRIL), and receptors for the neuropeptide vasoactive intestinal peptide (VIP). The ability of PP-DC to promote anti-CD40 dependent IgA was partially dependent on retinoic acid (RA) and transforming growth factor (TGF)-beta, whilst BAFF and APRIL signaling were not required. Signals delivered by BAFF and APRIL were crucial for CD40 independent IgA production, although the contribution of B2 cells to this pathway was minimal. The unique ability of PP-DC to instruct naïve B cells to differentiate into IgA producing plasma cells was mainly imparted by the presence of intestinal commensal bacteria, and could be mimicked by the addition of LPS to the culture. These data indicate that exposure to pathogen-associated molecular patterns present on intestinal commensal bacteria condition DC to express a unique molecular footprint that in turn allows them to promote IgA production.

Multi-faceted functions of secretory IgA at mucosal surfaces.

Secretory IgA (SIgA) plays an important role in the protection and homeostatic regulation of intestinal, respiratory, and urogenital mucosal epithelia separating the outside environment from the inside of the body. This primary function of SIgA is referred to as immune exclusion, a process that limits the access of numerous microorganisms and mucosal antigens to these thin and vulnerable mucosal barriers. SIgA has been shown to be involved in avoiding opportunistic pathogens to enter and disseminate in the systemic compartment, as well as tightly controlling the necessary symbiotic relationship existing between commensals and the host. Clearance by peristalsis appears thus as one of the numerous mechanisms whereby SIgA fulfills its function at mucosal surfaces. Sampling of antigen-SIgA complexes by microfold (M) cells, intimate contact occurring with Peyer's patch dendritic cells (DC), down-regulation of inflammatory processes, modulation of epithelial, and DC responsiveness are some of the recently identified processes to which the contribution of SIgA has been underscored. This review aims at presenting, with emphasis at the biochemical level, how the molecular complexity of SIgA can serve these multiple and non-redundant modes of action.

Gut physiology mediates a trade-off between adaptation to malnutrition and susceptibility to food-borne pathogens.

The animal gut plays a central role in tackling two common ecological challenges, nutrient shortage and food-borne parasites, the former by efficient digestion and nutrient absorption, the latter by acting as an immune organ and a barrier. It remains unknown whether these functions can be independently optimised by evolution, or whether they interfere with each other. We report that Drosophila melanogaster populations adapted during 160 generations of experimental evolution to chronic larval malnutrition became more susceptible to intestinal infection with the opportunistic bacterial pathogen Pseudomonas entomophila. However, they do not show suppressed immune response or higher bacterial loads. Rather, their increased susceptibility to P. entomophila is largely mediated by an elevated predisposition to loss of intestinal barrier integrity upon infection. These results may reflect a trade-off between the efficiency of nutrient extraction from poor food and the protective function of the gut, in particular its tolerance to pathogen-induced damage.

Emerging bacterial pathogens: the past and beyond.

Since the 1950s, medical communities have been facing with emerging and reemerging infectious diseases, and emerging pathogens are now considered to be a major microbiologic public health threat. In this review, we focus on bacterial emerging diseases and explore factors involved in their emergence as well as future challenges. We identified 26 major emerging and reemerging infectious diseases of bacterial origin; most of them originated either from an animal and are considered to be zoonoses or from water sources. Major contributing factors in the emergence of these bacterial infections are: (1) development of new diagnostic tools, such as improvements in culture methods, development of molecular techniques and implementation of mass spectrometry in microbiology; (2) increase in human exposure to bacterial pathogens as a result of sociodemographic and environmental changes; and (3) emergence of more virulent bacterial strains and opportunistic infections, especially affecting immunocompromised populations. A precise definition of their implications in human disease is challenging and requires the comprehensive integration of microbiological, clinical and epidemiologic aspects as well as the use of experimental models. It is now urgent to allocate financial resources to gather international data to provide a better understanding of the clinical relevance of these waterborne and zoonotic emerging diseases.

Host innate immune responses to microbial pathogens.

Sepsis is among the leading causes of death worldwide and its incidence is increasing. Defined as the host response to infection, sepsis is a clinical syndrome considered to be the expression of a dysregulated immune reaction induced by danger signals that may lead to organ failure and death. Remarkable progresses have been made in our understanding of the molecular basis of host defenses in recent years. The host defense response is initiated by innate immune sensors of danger signals designated under the collective name of pattern-recognition receptors. Members of the family of microbial sensors include the complement system, the Toll-like receptors, the nucleotide-binding oligomerization domainlike receptors, the RIG-I-like helicases and the C-type lectin receptors. Ligand-activated pattern-recognition receptors kick off a cascade of intracellular events resulting in the expression of co-stimulatory molecules and release of effector molecules playing a fundamental role in the initiation of the innate and adaptive immune responses. Fine tuning of proinflammatory and anti-inflammatory reactions is critical for keeping the innate immune response in check. Overwhelming or dysregulated responses induced by infectious stimuli may have dramatic consequences for the host as shown by the profound derangements observed in sepsis. Unfortunately, translational research approaches aimed at the development of therapies targeting newly identified innate immune pathways have not held their promises. Indeed, all recent clinical investigations of adjunctive anti-sepsis treatments had little, if any, impact on morbidity and all-cause mortality of sepsis. Dissecting the mechanisms underlying the transition from infection to sepsis is essential for solving the sepsis enigma. Important components of the puzzle have already been identified, but the hunt must go on in the laboratory and at the bedside.

Dendritic cells: the host Achille's heel for mucosal pathogens?

Mucosal surfaces represent the main sites of interaction with environmental microorganisms and antigens. Sentinel cells, including epithelial cells and dendritic cells (DCs), continuously sense the environment and coordinate defenses for the protection of mucosal tissues. DCs play a central role in the control of adaptive immune responses owing to their capacity to internalize foreign materials, to migrate into lymph nodes and to present antigens to naive lymphocytes. Some pathogenic microorganisms trigger epithelial responses that result in the recruitment of DCs. These pathogens hijack the recruited DCs to enable them to infect the host, escape the host's defense mechanisms and establish niches at remote sites.

High number of CD56(bright) NK-cells and persistently low CD4+ T-cells in a hemophiliac HIV/HCV co-infected patient without opportunistic infections.

BACKGROUND: Both the human immunodeficiency virus (HIV) and hepatitis C virus (HCV), either alone or as coinfections, persist in their hosts by destroying and/or escaping immune defenses, with high morbidity as consequence. In some cases, however, a balance between infection and immunity is reached, leading to prolonged asymptomatic periods. We report a case of such an indolent co-infection, which could be explained by the development of a peculiar subset of Natural Killer (NK) cells. RESULTS: Persistently high peripheral levels of CD56+ NK cells were observed in a peculiar hemophiliac HIV/HCV co-infected patient with low CD4 counts, almost undetectable HIV viral load and no opportunistic infections. Thorough analysis of NK-subsets allowed to identify a marked increase in the CD56bright/dim cell ratio and low numbers of CD16+/CD56- cells. These cells have high levels of natural cytotoxicity receptors but low NCR2 and CD69, and lack both CD57 and CD25 expression. The degranulation potential of NK-cells which correlates with target cytolysis was atypically mainly performed by CD56bright NK-cells, whereas no production of interferon γ (IFN-γ) was observed following NK activation by K562 cells. CONCLUSIONS: These data suggest that the expansion and lytic capacity of the CD56bright NK subset may be involved in the protection of this « rare » HIV/HCV co-infected hemophiliac A patient from opportunistic infections and virus-related cancers despite very low CD4+ cell counts.

Insight into cross-talk between intra-amoebal pathogens.

Abstract: Background: Amoebae are phagocytic protists where genetic exchanges might take place between amoeba-resistant bacteria. These amoebal pathogens are able to escape the phagocytic behaviour of their host. They belong to different bacterial phyla and often show a larger genome size than human-infecting pathogens. This characteristic is proposed to be the result of frequent gene exchanges with other bacteria that share a sympatric lifestyle and contrasts with the genome reduction observed among strict human pathogens.Results: We sequenced the genome of a new amoebal pathogen, Legionella drancourtii, and compared its gene content to that of a Chlamydia-related bacterium, Parachlamydia acanthamoebae. Phylogenetic reconstructions identified seven potential horizontal gene transfers (HGTs) between the two amoeba-resistant bacteria, including a complete operon of four genes that encodes an ABC-type transporter. These comparisons pinpointed potential cases of gene exchange between P. acanthamoebae and Legionella pneumophila, as well as gene exchanges between other members of the Legionellales and Chlamydiales orders. Moreover, nine cases represent possible HGTs between representatives from the Legionellales or Chlamydiales and members of the Rickettsiales order.Conclusions: This study identifies numerous gene exchanges between intracellular Legionellales and Chlamydiales bacteria, which could preferentially occur within common inclusions in their amoebal hosts. Therefore it contributes to improve our knowledge on the intra-amoebal gene properties associated to their specific lifestyle.

Discovery of new intracellular pathogens by amoebal coculture and amoebal enrichment approaches.

Intracellular pathogens such as legionella, mycobacteria and Chlamydia-like organisms are difficult to isolate because they often grow poorly or not at all on selective media that are usually used to cultivate bacteria. For this reason, many of these pathogens were discovered only recently or following important outbreaks. These pathogens are often associated with amoebae, which serve as host-cell and allow the survival and growth of the bacteria. We intend here to provide a demonstration of two techniques that allow isolation and characterization of intracellular pathogens present in clinical or environmental samples: the amoebal coculture and the amoebal enrichment. Amoebal coculture allows recovery of intracellular bacteria by inoculating the investigated sample onto an amoebal lawn that can be infected and lysed by the intracellular bacteria present in the sample. Amoebal enrichment allows recovery of amoebae present in a clinical or environmental sample. This can lead to discovery of new amoebal species but also of new intracellular bacteria growing specifically in these amoebae. Together, these two techniques help to discover new intracellular bacteria able to grow in amoebae. Because of their ability to infect amoebae and resist phagocytosis, these intracellular bacteria might also escape phagocytosis by macrophages and thus, be pathogenic for higher eukaryotes.

Histone deacetylase inhibitors impair innate immune responses

SUMMARYThe innate immune system plays a central role in host defenses against invading pathogens. Innate immune cells sense the presence of pathogens through pattern recognition receptors that trigger intracellular signaling, leading to the production of pro-inflammatory mediators like cytokines, which shape innate and adaptive immune responses. Both by excess and by default inflammation may be detrimental to the host. Indeed, severe sepsis and septic shock are lethal complications of infections characterized by a dysregulated inflammatory response.In recent years, members of the superfamily of histone deacetylases have been the focus of great interest. In mammals, histone deacetylases are broadly classified into two main subfamilies comprising histone deacetylases 1-11 (HDAC1-11) and sirtuins 1-7 (SIRT1-7). These enzymes influence gene expression by deacetylating histones and numerous non-histone proteins. Histone deacetylases have been involved in the development of oncologic, metabolic, cardiovascular, neurodegenerative and autoimmune diseases. Pharmacological modulators of histone deacetylase activity, principally inhibitors, have been developed for the treatment of cancer and metabolic diseases. When we initiated this project, several studies suggested that inhibitors of HDAC 1-11 have anti-inflammatory activity. Yet, their influence on innate immune responses was largely uncharacterized. The present study was initiated to fill in this gap.In the first part of this work, we report the first comprehensive study of the effects of HDAC 1- 11 inhibitors on innate immune responses in vitro and in vivo. Strikingly, expression studies revealed that HDAC1-11 inhibitors act essentially as negative regulators of basal and microbial product- induced expression of critical immune receptors and antimicrobial products by mouse and human innate immune cells like macrophages and dendritic cells. Furthermore, we describe a new molecular mechanism whereby HDAC1-11 inhibitors repress pro-inflammatory cytokine expression through the induction of the expression and the activity of the transcriptional repressor Μί-2β. HDAC1-11 inhibitors also impair the potential of macrophages to engulf and kill bacteria. Finally, mice treated with an HDAC inhibitor are more susceptible to non-severe bacterial and fungal infection, but are protected against toxic and septic shock. Altogether these data support the concept that HDAC 1-11 inhibitors have potent anti-inflammatory and immunomodulatory activities in vitro and in vivo.Macrophage migration inhibitory factor (MIF) is a pro-inflammatory cytokine that plays a central role in innate immune responses, cell proliferation and oncogenesis. In the second part of this manuscript, we demonstrate that HDAC1-11 inhibitors inhibit MIF expression in vitro and in vivo and describe a novel molecular mechanism accounting for these effects. We propose that inhibition of MIF expression by HDAC 1-11 inhibitors may contribute to the antitumorigenic and anti-inflammatory effects of these drugs.NAD+ is an essential cofactor of sirtuins activity and one of the major sources of energy within the cells. Therefore, sirtuins link deacetylation to NAD+ metabolism and energy status. In the last part of this thesis, we report preliminary results indicating that a pharmacological inhibitor of SIRT1-2 drastically decreases pro-inflammatory cytokine production (RNA and protein) and interferes with MAP kinase intracellular signal transduction pathway in macrophages. Moreover, administration of the SIRT1-2 inhibitor protects mice from lethal endotoxic shock and septic shock.Overall, our studies demonstrate that inhibitors of HDAC1-11 and sirtuins are powerful anti-inflammatory molecules. Given their profound negative impact on the host antimicrobial defence response, these inhibitors might increase the susceptibility to opportunistic infections, especially in immunocompromised cancer patients. Yet, these inhibitors might be useful to control the inflammatory response in severely ill septic patients or in patients suffering from chronic inflammatory diseases.

Ant queens (Hymenoptera: Formicidae) are attracted to fungal pathogens during the initial stage of colony founding

Ant queens that attempt to disperse and found new colonies independently face high mortality risks. The exposure of queens to soil entomopathogens during claustral colony founding may be particularly harmful, as founding queens lack the protection conferred by mature colonies. Here, we tested the hypotheses that founding queens (I) detect and avoid nest sites that are contaminated by fungal pathogens, and (II) tend to associate with other queens to benefit from social immunity when nest sites are contaminated. Surprisingly, in nest choice assays, young Formica selysi BONDROIT, 1918 queens had an initial preference for nest sites contaminated by two common soil entomopathogenic fungi, Beauveria bassiana and Metarhizium brunneum. Founding queens showed a similar preference for the related but non-entomopathogenic fungus Fusarium graminearum. In contrast, founding queens had no significant preference for the more distantly related nonentomopathogenic fungus Petromyces alliaceus, nor for heat-killed spores of B. bassiana. Finally, founding queens did not increase the rate of queen association in presence of B. bassiana. The surprising preference of founding queens for nest sites contaminated by live entomopathogenic fungi suggests that parasites manipulate their hosts or that the presence of specific fungi is a cue associated with suitable nesting sites.

Social evolution and defences against pathogens in ants

Pathogens represent a threat to all organisms, which generates a coevolutionary arms race. Social insects provide an interesting system to study host-pathogen interactions, because their defences depend on both the individual and collective responses, and involve genetic, physiological, behavioral and organizational mechanisms. In this thesis, I studied the evolutionary ecology of the resistance of ant queens and workers to natural fungal pathogens. Mechanisms that increase within-colony genetic diversity, like polyandry and polygyny, decrease relatedness among colony mates, which reduces the strength of selection for the evolution and maintenance of altruistic behavior. A leading hypothesis posits that intracolonial genetic diversity is adaptive because it reduces the risk of pathogen transmission. In chapter 1, I examine individual resistance in ant workers of Formica selysi, a species that shows natural variation in colony queen number. I discuss how this variation might be beneficial to resist natural fungal pathogens in groups. Overall my results indicate that there is genetic variation for fungal resistance in workers, a requirement for the 'genetic diversity for pathogen resistance' hypothesis. However I was not able to detect direct evidence that group diversity improves the survival of focal ants or reduces pathogen transmission. Thus, although the coexistence of multiple queens increases the within-colony variance in worker resistance, it remains unclear whether it protects ant colonies from pathogens and whether it is comparable to polyandry in other social insects. Traditionally, it was thought that the immune system of invertebrates lacked memory and specificity. In chapter 2, I investigate individual immunity in ant queens and show that they may be able to adjust their pathogen defences in response to their current environment by means of immune priming, which bears similarities with the adaptive immunity of vertebrates. However, my results indicate that the expression of immune priming in ant queens may be influenced by factors like mating status, mating conditions or host species. In addition, I showed that mating increases pathogen resistance in çhe two ant species that I studied (F. selysi and Lasius niger). This raises the question of how ant queens invest heavily in both maintenance and reproduction, which I discuss in the context of the evolution of social organization. In chapter 3,1 investigate if transgenerational priming against a fungal pathogen protects the queen progeny. I failed to detect this effect, and discuss why the detection of transgenerational immune priming in ants is a difficult task. Overall, this thesis illustrates some of the individual and collective mechanisms that likely played a role in allowing ants to become one of the most diverse and ecologically successful groups of organisms. -- Les pathogènes représentent une menace pour tous les organismes, ce qui a engendré l'évolution d'une course aux armements. Les insectes sociaux sont un système intéressant permettant d'étudier les interactions hôtes-pathogènes, car leurs défenses dépendent de réponses aussi bien individuelles que collectives, et impliquent des mécanismes génétiques, physiologiques, comportementaux et organisationnels. Dans cette thèse, j'ai étudié l'écologie évolutive de la résistance des reines et des ouvrières de fourmis exposées à des champignons pathogènes. Les facteurs augmentant la diversité génétique à l'intérieur de la colonie, comme la polyandrie et la polygynie, diminuent la parenté, ce qui réduit la pression de sélection pour l'évolution et la maintenance des comportements altruistes. Une hypothèse dominante stipule que la diversité génétique à l'intérieur de la colonie est adaptative car elle réduit le risque de transmission des pathogènes. Dans le chapitre 1, nous examinons la résistance individuelle à des pathogènes fongiques chez les ouvrières de Formica selysi, une espèce présentant une variation naturelle dans le nombre de reines par colonie. Nous discutons aussi de la possibilité que ces variations individuelles augmentent la capacité du groupe à résister à des champignons pathogènes. Dans l'ensemble, nos résultats indiquent une variation génétique dans la résistance aux champignons chez les ouvrières, un prérequis à l'hypothèse que la diversité génétique du groupe augmente la résistance aux pathogènes. Cependant, nous n'avons pas pu détecter une preuve directe que la diversité du groupe augmente la survie de fourmis focales ou réduise la transmission des pathogènes. Ainsi, bien que la coexistence de plusieurs reines augmente la variance dans la résistance des ouvrières à l'intérieur de la colonie, la question de savoir si cela protège les colonies de fourmis contre les pathogènes et si cela est comparable à la polyandrie chez d'autres insectes sociaux reste ouverte. Traditionnellement, il était admis que le système immunitaire des invertébrés ne possédait pas de mémoire et était non-spécifique. Dans le chapitre 2, nous avons étudié l'immunité individuelle chez des reines de fourmis. Nous avons montré que les reines pourraient être capables d'ajuster leurs défenses contre les pathogènes en réponse à leur environnement, grâce à une pré-activation du système immunitaire (« immune priming ») ressemblant à l'immunité adaptative des vertébrés. Cependant, nos résultats indiquent que cette pré-activation du système immunitaire chez les reines dépend du fait d'être accouplée ou non, des conditions d'accouplement, ou de l'espèce. De plus, nous avons montré que l'accouplement augmente la résistance aux pathogènes chez les deux espèces que nous avons étudié (F. selysi et Lasius niger). Ceci pose la question de la capacité des reines à investir fortement aussi bien dans la maintenance que dans la reproduction, ce que nous discutons dans le contexte de l'évolution de l'organisation sociale. Dans le chapitre 3, nous étudions si la pré-activation trans-générationelle du système immunitaire [« trans-generational immune priming ») protège la progéniture de la reine contre un champignon pathogène. Nous n'avons par réussi à détecter cet effet, et discutons des raisons pour lesquelles la détection de la pré-activation trans-générationelle du système immunitaire chez les fourmis est une tâche difficile. Dans l'ensemble, cette thèse illustre quelques-uns des mécanismes individuels et collectifs qui ont probablement contribué à la diversité et à l'important succès écologique des fourmis.