Brucella melitensis invades murine erythrocytes during infection.

Brucella spp. are facultative intracellular Gram-negative coccobacilli responsible for brucellosis, a worldwide zoonosis. We observed that Brucella melitensis is able to persist for several weeks in the blood of intraperitoneally infected mice and that transferred blood at any time point tested is able to induce infection in naive recipient mice. Bacterial persistence in the blood is dramatically impaired by specific antibodies induced following Brucella vaccination. In contrast to Bartonella, the type IV secretion system and flagellar expression are not critically required for the persistence of Brucella in blood. ImageStream analysis of blood cells showed that following a brief extracellular phase, Brucella is associated mainly with the erythrocytes. Examination by confocal microscopy and transmission electron microscopy formally demonstrated that B. melitensis is able to invade erythrocytes in vivo. The bacteria do not seem to multiply in erythrocytes and are found free in the cytoplasm. Our results open up new areas for investigation and should serve in the development of novel strategies for the treatment or prophylaxis of brucellosis. Invasion of erythrocytes could potentially protect the bacterial cells from the host's immune response and hamper antibiotic treatment and suggests possible Brucella transmission by bloodsucking insects in nature.

Critical role of regulatory T cells in Th17-mediated minor antigen-disparate rejection.

Th17-mediated immune responses have been recently identified as novel pathogenic mechanisms in a variety of conditions; however, their importance in allograft rejection processes is still debated. In this paper, we searched for MHC or minor Ag disparate models of skin graft rejection in which Th17 immune responses might be involved. We found that T cell-derived IL-17 is critical for spontaneous rejection of minor but not major Ag-mismatched skin grafts. IL-17 neutralization was associated with a lack of neutrophil infiltration and neutrophil depletion delayed rejection, suggesting neutrophils as an effector mechanism downstream of Th17 cells. Regulatory T cells (Tregs) appeared to be involved in Th17 reactivity. We found that in vivo Treg depletion prevented IL-17 production by recipient T cells. An adoptive cotransfer of Tregs with naive monospecific antidonor T cells in lymphopenic hosts biased the immune response toward Th17. Finally, we observed that IL-6 was central for balancing Tregs and Th17 cells as demonstrated by the prevention of Th17 differentiation, the enhanced Treg/Th17 ratio, and a net impact of rejection blockade in the absence of IL-6. In conclusion, the ability of Tregs to promote the Th17/neutrophil-mediated pathway of rejection that we have described should be considered as a potential drawback of Treg-based cell therapy.

IL-17 production elicited by allo-major histocompatibility complex class II recognition depends on CD25posCD4pos T cells.

BACKGROUND: Interleukin (IL)-17 is involved in autoimmune inflammatory disorders and naturally occurring CD25pos regulatory T cells were shown to promote IL-17 synthesis. Because IL-17 is overproduced in certain types of allograft rejection, it is important to characterize the cells responsible for IL-17 synthesis and to define how IL-17 is regulated during alloimmune responses. METHODS: Splenic CD4pos T cells were isolated from C57BL/6 mice and fractionated according to CD25 expression. T cells were stimulated by major histocompatibility complex class II-mismatched bone marrow-derived dendritic cells from bm12 mice, either immature or made mature by exposure to lipopolysaccharide. To track T cell populations, CD25negCD4pos and CD25posCD4pos were isolated from Thy1.1 and congenic Thy1.2 mice, respectively. Cell proliferation was quantified by CFSE dilution. IL-17-producing cells and FOXP3pos cells were enumerated by intracytoplasmic staining and cytokine levels in culture supernatants were measured by ELISA. RESULTS: Addition of CD25posCD4pos T cells to CD25negCD4pos T cells inhibited IL-2, interferon-[gamma], and IL-13 production but promoted IL-17 synthesis on stimulation by allogenic immature DC. In this setting, IL-17 originated from CD25intCD4posFOXP3neg memory T cells, which depend on IL-2 to produce IL-17. Alloreactive CD25negCD4pos T cells were also induced to produce IL-17 when stimulated by mature DC in the presence of CD25highCD4posFOXP3pos T cells. CONCLUSIONS: We conclude that (1) the cellular source of IL-17 during an antiallo major histocompatibility complex class II response depends on the maturation status of allogenic DC, (2) whereas suppressing Th1 and Th2 cytokine synthesis, naturally occurring regulatory T cells, allow IL-17 production by alloreactive CD4pos T cells.

The evolutionarily conserved protein Las1 is required for pre-rRNA processing at both ends of ITS2.

Ribosome synthesis entails the formation of mature rRNAs from long precursor molecules, following a complex pre-rRNA processing pathway. Why the generation of mature rRNA ends is so complicated is unclear. Nor is it understood how pre-rRNA processing is coordinated at distant sites on pre-rRNA molecules. Here we characterized, in budding yeast and human cells, the evolutionarily conserved protein Las1. We found that, in both species, Las1 is required to process ITS2, which separates the 5.8S and 25S/28S rRNAs. In yeast, Las1 is required for pre-rRNA processing at both ends of ITS2. It is required for Rrp6-dependent formation of the 5.8S rRNA 3' end and for Rat1-dependent formation of the 25S rRNA 5' end. We further show that the Rat1-Rai1 5'-3' exoribonuclease (exoRNase) complex functionally connects processing at both ends of the 5.8S rRNA. We suggest that pre-rRNA processing is coordinated at both ends of 5.8S rRNA and both ends of ITS2, which are brought together by pre-rRNA folding, by an RNA processing complex. Consistently, we note the conspicuous presence of ~7- or 8-nucleotide extensions on both ends of 5.8S rRNA precursors and at the 5' end of pre-25S RNAs suggestive of a protected spacer fragment of similar length.

Adenylate cyclases of Trypanosoma brucei inhibit the innate immune response of the host.

The parasite Trypanosoma brucei possesses a large family of transmembrane receptor-like adenylate cyclases. Activation of these enzymes requires the dimerization of the catalytic domain and typically occurs under stress. Using a dominant-negative strategy, we found that reducing adenylate cyclase activity by about 50% allowed trypanosome growth but reduced the parasite's ability to control the early innate immune defense of the host. Specifically, activation of trypanosome adenylate cyclase resulting from parasite phagocytosis by liver myeloid cells inhibited the synthesis of the trypanosome-controlling cytokine tumor necrosis factor-α through activation of protein kinase A in these cells. Thus, adenylate cyclase activity of lyzed trypanosomes favors early host colonization by live parasites. The role of adenylate cyclases at the host-parasite interface could explain the expansion and polymorphism of this gene family.

Synthesis of fiber Bragg grating parameters from experimental reflectivity: a simplex approach and its application to the determination of temperature-dependent properties.

A simple, accurate, and fast method to synthesize the physical parameters of a fiber Bragg grating numerically from its reflectivity is proposed and demonstrated. Our program uses the transfer matrix method and is based on a Nelder-Mead simplex optimization algorithm. It can be applied to both uniform and nonuniform (apodized and chirped) fiber Bragg gratings. The method is then used to synthesize a uniform Bragg grating from its reflectivity taken at different temperatures. It gives a good estimate of the thermal expansion coefficient and the thermo-optic coefficient of the fiber.