Coxiella burnetii inhibits host immunity by a protein phosphatase adapted from glycolysis.

Coxiella burnetii is a bacterial pathogen that replicates within host cells by establishing a membrane-bound niche called the Coxiella-containing vacuole. Biogenesis of this compartment requires effectors of its Dot/Icm type IV secretion system. A large cohort of such effectors has been identified, but the function of most of them remain elusive. Here, by a cell-based functional screening, we identified the effector Cbu0513 (designated as CinF) as an inhibitor of NF-κB signaling. CinF is highly similar to a fructose-1,6-bisphosphate (FBP) aldolase/phosphatase present in diverse bacteria. Further study reveals that unlike its ortholog from Sulfolobus tokodaii, CinF does not exhibit FBP phosphatase activity. Instead, it functions as a protein phosphatase that specifically dephosphorylates and stabilizes IκBα. The IκBα phosphatase activity is essential for the role of CinF in C. burnetii virulence. Our results establish that C. burnetii utilizes a protein adapted from sugar metabolism to subvert host immunity.

Caspase-8 activity mediates TNFα production and restricts Coxiella burnetii replication during murine macrophage infection.

Coxiella burnetii is an obligate intracellular bacteria that causes the global zoonotic disease Q Fever. Treatment options for chronic infection are limited, and the development of novel therapeutic strategies requires a greater understanding of how C. burnetii interacts with immune signaling. Cell death responses are known to be manipulated by C. burnetii, but the role of caspase-8, a central regulator of multiple cell death pathways, has not been investigated. In this research, we studied bacterial manipulation of caspase-8 signaling and the significance of caspase-8 to C. burnetii infection, examining bacterial replication, cell death induction, and cytokine signaling. We measured caspase, RIPK, and MLKL activation in C. burnetii-infected tumor necrosis factor alpha (TNFα)/cycloheximide-treated THP-1 macrophage-like cells and TNFα/ZVAD-treated L929 cells to assess apoptosis and necroptosis signaling. Additionally, we measured C. burnetii replication, cell death, and TNFα induction over 12 days in RIPK1-kinase-dead, RIPK3-kinase-dead, or RIPK3-kinase-dead-caspase-8-/- bone marrow-derived macrophages (BMDMs) to understand the significance of caspase-8 and RIPK1/3 during infection. We found that caspase-8 is inhibited by C. burnetii, coinciding with inhibition of apoptosis and increased susceptibility to necroptosis. Furthermore, C. burnetii replication was increased in BMDMs lacking caspase-8, but not in those lacking RIPK1/3 kinase activity, corresponding with decreased TNFα production and reduced cell death. As TNFα is associated with the control of C. burnetii, this lack of a TNFα response may allow for the unchecked bacterial growth we saw in caspase-8-/- BMDMs. This research identifies and explores caspase-8 as a key regulator of C. burnetii infection, opening novel therapeutic doors.

Modulation of innate immune signaling by a Coxiella burnetii eukaryotic-like effector protein.

The Q fever agent Coxiella burnetii uses a defect in organelle trafficking/intracellular multiplication (Dot/Icm) type 4b secretion system (T4SS) to silence the host innate immune response during infection. By investigating C. burnetii effector proteins containing eukaryotic-like domains, here we identify NopA (nucleolar protein A), which displays four regulator of chromosome condensation (RCC) repeats, homologous to those found in the eukaryotic Ras-related nuclear protein (Ran) guanine nucleotide exchange factor (GEF) RCC1. Accordingly, NopA is found associated with the chromatin nuclear fraction of cells and uses the RCC-like domain to interact with Ran. Interestingly, NopA triggers an accumulation of Ran-GTP, which accumulates at nucleoli of transfected or infected cells, thus perturbing the nuclear import of transcription factors of the innate immune signaling pathway. Accordingly, qRT-PCR analysis on a panel of cytokines shows that cells exposed to the C. burnetii nopA::Tn or a Dot/Icm-defective dotA::Tn mutant strain present a functional innate immune response, as opposed to cells exposed to wild-type C. burnetii or the corresponding nopA complemented strain. Thus, NopA is an important regulator of the innate immune response allowing Coxiella to behave as a stealth pathogen.

Tuning Subunit Vaccines with Novel TLR Triagonist Adjuvants to Generate Protective Immune Responses against Coxiella burnetii.

Coxiella burnetii is an obligate intracellular bacterium and the causative agent of Q fever. C. burnetii is considered a potential bioterrorism agent because of its low infectious dose; resistance to heat, drying, and common disinfectants; and lack of prophylactic therapies. Q-Vax, a formalin-inactivated whole-bacteria vaccine, is currently the only prophylactic measure that is protective against C. burnetii infections but is not U.S. Food and Drug Administration approved. To overcome the safety concerns associated with the whole-bacteria vaccine, we sought to generate and evaluate recombinant protein subunit vaccines against C. burnetii To accomplish this, we formulated C. burnetii Ags with a novel TLR triagonist adjuvant platform, which used combinatorial chemistry to link three different TLR agonists together to form one adjuvanting complex. We evaluated the immunomodulatory activity of a panel of TLR triagonist adjuvants and found that they elicited unique Ag-specific immune responses both in vitro and in vivo. We evaluated our top candidates in a live C. burnetii aerosol challenge model in C56BL/6 mice and found that several of our novel vaccine formulations conferred varying levels of protection to the challenged animals compared with sham immunized mice, although none of our candidates were as protective as the commercial vaccine across all protection criteria that were analyzed. Our findings characterize a novel adjuvant platform and offer an alternative approach to generating protective and effective vaccines against C. burnetii.

Long-term serological follow-up after primary Coxiella burnetii infection in patients with vascular risk factors for chronic Q fever.

We evaluated the long-term serological follow-up of patients with vascular risk factors for chronic Q fever that were previously Coxiella burnetii seropositive. C. burnetii phase I IgG titers were reevaluated in patients that gave informed consent or retrospectively collected in patients already deceased or lost to follow-up. Of 107 patients, 25 (23.4%) became seronegative, 77 (72.0%) retained a profile of past resolved Q fever infection, and five (4.7%) developed chronic Q fever. We urge clinicians to stay vigilant for chronic Q fever beyond two years after primary infection and perform serological testing based on clinical presentation.

Avoidance of the NLRP3 Inflammasome by the Stealth Pathogen, Coxiella burnetii.

Coxiella burnetii, a highly adapted obligate intracellular bacterial pathogen and the cause of the zoonosis Q fever, is a reemerging public health threat. C. burnetii employs a Type IV secretion system (T4SS) to establish and maintain its intracellular niche and modulate host immune responses including the inhibition of apoptosis. Interactions between C. burnetii and caspase-1-mediated inflammasomes are not fully elucidated. This study confirms that C. burnetii does not activate caspase-1 during infection of mouse macrophages in vitro. C. burnetii-infected cells did not develop NLRP3 and ASC foci indicating its ability to avoid cytosolic detection. C. burnetii is unable to inhibit the pyroptosis and IL-1ß secretion that is induced by potent inflammasome stimuli but rather enhances these caspase-1-mediated effects. We found that C. burnetii upregulates pro-IL-1ß and robustly primes NLRP3 inflammasomes via TLR2 and MyD88 signaling. As for wildtype C. burnetii, T4SS-deficient mutants primed and potentiated NLRP3 inflammasomes. An in vivo model of pulmonary infection in C57BL/6 mice was developed. Mice deficient in NLRP3 or caspase-1 were like wildtype mice in the development and resolution of splenomegaly due to red pulp hyperplasia, and histologic lesions and macrophage kinetics, but had slightly higher pulmonary bacterial burdens at the greatest measured time point. Together these findings indicate that C. burnetii primes but avoids cytosolic detection by NLRP3 inflammasomes, which are not required for the clinical resistance of C57BL/6 mice. Determining mechanisms employed by C. burnetii to avoid cytosolic detection via NLRP3 inflammasomes will be beneficial to the development of preventative and interventional therapies for Q fever.

Coxiella burnetii-Infected NK Cells Release Infectious Bacteria by Degranulation.

Natural killer (NK) cells are critically involved in the early immune response against various intracellular pathogens, including Coxiella burnetii and Chlamydia psittaciChlamydia-infected NK cells functionally mature, induce cellular immunity, and protect themselves by killing the bacteria in secreted granules. Here, we report that infected NK cells do not allow intracellular multiday growth of Coxiella, as is usually observed in other host cell types. C. burnetii-infected NK cells display maturation and gamma interferon (IFN-γ) secretion, as well as the release of Coxiella-containing lytic granules. Thus, NK cells possess a potent program to restrain and expel different types of invading bacteria via degranulation. Strikingly, though, in contrast to Chlamydia, expulsed Coxiella organisms largely retain their infectivity and, hence, escape the cell-autonomous self-defense mechanism in NK cells.

A possible link between recurrent upper respiratory tract infections and lower cytokine production in patients with Q fever fatigue syndrome.

Besides fatigue, many Q fever fatigue syndrome (QFS) patients also complain of frequently recurring upper respiratory tract infections with severe symptoms. We investigated whether immunologic dysregulation contributes to these complaints. Cytokine and chemokine production was measured after stimulating monocytes of QFS patients and age- and sex-matched healthy controls with LPS and several viral ligands. The H3K4me3 mark of open chromatin was measured at the promoter regions of cytokines and chemokines that differed significantly from healthy controls. Monocytes of QFS patients produced significantly less TNF-α (p = 0.032), IL-1ß (0.004, 0.024, and 0.008), IL-6 (0.043), RANTES (0.033), IP-10 (0.049), MCP-1 (0.022), IL- 13 (0.029), and IL-10 (0.026) than healthy controls when stimulated with various ligands. H3K4me3 expression was significantly lower in QFS patients than in healthy controls on the promoter regions of IL-1ß (p = 0.004), MCP-1 (<0.001 and <0.001), IP-10 (<0.001), IL-10 (0.041), and IL-13 (<0.001, <0.001, and 0.001). QFS patients showed diminished cytokine responses to various stimuli compared to age- and sex-matched healthy controls, likely due to epigenetic remodeling and long-term memory as a result from the acute Q fever infection. This might explain the upper respiratory tract ailments in QFS.

Comparison of Different Commercially Available Enzyme-Linked Immunosorbent Assays with Immunofluorescence Test for Detection of Phase II IgG and IgM Antibodies to Coxiella burnetii.

Several commercially available enzyme-linked immunosorbent assays (ELISAs) for the detection of phase II IgG or IgM antibodies against Coxiella burnetii were compared. In addition, an indirect immunofluorescence test was used as a confirmation test. In all, 70 serum samples for IgG and 43 serum samples for IgM were tested. The ELISAs showed large differences in sensitivity and specificity, which led to a partially high ratio of false-negative determinations. The most convincing test was PanBio from Abbott, which unfortunately can only test IgG but not IgM.

Chemokine Receptor 7 Is Essential for Coxiella burnetii Whole-Cell Vaccine-Induced Cellular Immunity but Dispensable for Vaccine-Mediated Protective Immunity.

BACKGROUND: Protective immunity against Coxiella burnetii infection is conferred by vaccination with virulent (PI-WCV), but not avirulent (PII-WCV) whole-cell inactivated bacterium. The only well-characterized antigenic difference between virulent and avirulent C. burnetii is they have smooth and rough lipopolysaccharide (LPS), respectively. METHODS: Mice were vaccinated with PI-WCV and PII-WCV. Humoral and cellular responses were evaluated using protein chip microarrays and ELISpots, respectively. Dendritic cell (DC) maturation after stimulation with PI-WVC and PII-WVC was evaluated using flow cytometry. Vaccine-challenge studies were performed to validate the importance of the receptor CCR7. RESULTS: Other than specific antibody response to PI-LPS, similar antibody profiles were observed but IgG titers were significantly higher after vaccination with PI-WCV. Furthermore, higher frequency of antigen-specific CD4+ T cells was detected in mice immunized with PI-WCV. PI-WCV-stimulated DCs displayed significantly higher levels of CCR7 and migratory ability to secondary lymphoid organs. Challenge-protection studies in wild-type and CCR7-deficient mice confirmed that CCR7 is critical for PI-WCV-induced cellular immunity. CONCLUSIONS: PI-WVC stimulates protective immunity to C. burnetii in mice through stimulation of migratory behavior in DCs for protective cellular immunity. Additionally, the humoral immune response to LPS is an important component of protective immunity.

Characterization of Early Stages of Human Alveolar Infection by the Q Fever Agent Coxiella burnetii.

Human Q fever is caused by the intracellular bacterial pathogen Coxiella burnetii Q fever presents with acute flu-like and pulmonary symptoms or can progress to chronic, severe endocarditis. After human inhalation, C. burnetii is engulfed by alveolar macrophages and transits through the phagolysosomal maturation pathway, resisting the acidic pH of lysosomes to form a parasitophorous vacuole (PV) in which to replicate. Previous studies showed that C. burnetii replicates efficiently in primary human alveolar macrophages (hAMs) in ex vivo human lung tissue. Although C. burnetii replicates in most cell types in vitro, the pathogen does not grow in non-hAM cells of human lung tissue. In this study, we investigated the interaction between C. burnetii and other pulmonary cell types apart from the lung environment. C. burnetii formed a prototypical PV and replicated efficiently in human pulmonary fibroblasts and in airway, but not alveolar, epithelial cells. Atypical PV expansion in alveolar epithelial cells was attributed in part to defective recruitment of autophagy-related proteins. Further assessment of the C. burnetii growth niche showed that macrophages mounted a robust interleukin 8 (IL-8), neutrophil-attracting response to C. burnetii and ultimately shifted to an M2-polarized phenotype characteristic of anti-inflammatory macrophages. Considering our findings together, this study provides further clarity on the unique C. burnetii-lung dynamic during early stages of human acute Q fever.

Coxiella burnetii Intratracheal Aerosol Infection Model in Mice, Guinea Pigs, and Nonhuman Primates.

Coxiella burnetii, the etiological agent of Q fever, is a Gram-negative bacterium transmitted to humans by inhalation of contaminated aerosols. Acute Q fever is often self-limiting, presenting as a febrile illness that can result in atypical pneumonia. In some cases, Q fever becomes chronic, leading to endocarditis that can be life threatening. The formalin-inactivated whole-cell vaccine (WCV) confers long-term protection but has significant side effects when administered to presensitized individuals. Designing new vaccines against C. burnetii remains a challenge and requires the use of clinically relevant modes of transmission in appropriate animal models. We have developed a safe and reproducible C. burnetii aerosol challenge in three different animal models to evaluate the effects of pulmonary acquired infection. Using a MicroSprayer aerosolizer, BL/6 mice and Hartley guinea pigs were infected intratracheally with C. burnetii Nine Mile phase I (NMI) and demonstrated susceptibility as determined by measuring bacterial growth in the lungs and subsequent dissemination to the spleen. Histological analysis of lung tissue showed significant pathology associated with disease, which was more severe in guinea pigs. Infection using large-particle aerosol (LPA) delivery was further confirmed in nonhuman primates, which developed fever and pneumonia. We also demonstrate that vaccinating mice and guinea pigs with WCV prior to LPA challenge is capable of eliciting protective immunity that significantly reduces splenomegaly and the bacterial burden in spleen and lung tissues. These data suggest that these models can have appreciable value in using the LPA delivery system to study pulmonary Q fever pathogenesis as well as designing vaccine countermeasures to C. burnetii aerosol transmission.

Coxiella burnetii Epitope-Specific T-Cell Responses in Patients with Chronic Q Fever.

Infection with Coxiella burnetii, the causative agent of Q fever, can result in life-threatening persistent infection. Reactogenicity hinders worldwide implementation of the only licensed human Q fever vaccine. We previously demonstrated long-lived immunoreactivity in individuals with past symptomatic and asymptomatic Coxiella infection (convalescents) to promiscuous HLA class II C. burnetii epitopes, providing the basis for a novel T-cell targeted subunit vaccine. In this study, we investigated in a cohort of 22 individuals treated for persistent infection (chronic Q fever) whether they recognize the same set of epitopes or distinct epitopes that could be candidates for a therapeutic vaccine or aid in the diagnosis of persistent infection. In cultured enzyme-linked immunosorbent spot (ELISpot) assays, individuals with chronic Q fever showed strong class II epitope-specific responses that were largely overlapping with the peptide repertoire identified previously for convalescents. Five additional peptides were recognized more frequently by chronic subjects, but there was no combination of epitopes uniquely recognized by or nonreactive in subjects with chronic Q fever. Consistent with more recent/prolonged exposure, we found, however, stronger ex vivo responses by direct ELISpot to both whole-cell C. burnetii and individual peptides in chronic patients than in convalescents. In conclusion, we have validated and expanded a previously published set of candidate epitopes for a novel T-cell targeted subunit Q fever vaccine in treated patients with chronic Q fever and demonstrated that they successfully mounted a T-cell response comparable to that of convalescents. Finally, we demonstrated that individuals treated for chronic Q fever mount a broader ex vivo response to class II epitopes than convalescents, which could be explored for diagnostic purposes.

Introduction to Measurement of Avidity of Anti-Coxiella burnetii IgG in Diagnosis of Q Fever.

Due to the atypical serological profile of some patients with primary Q fever infection who do not develop IgM against Coxiella burnetii, we developed an avidity test to distinguish recent or past infections. We tested 39 serum samples by immunofluorescence with conventional assay and after urea treatment from 26 patients at different stages of the disease. We observed a strong avidity in the 15 serum samples from patients with infections of >6 months and a low avidity for sera from patients with recent infections. A complete denaturation of the antibody-antigen complex was observed for patients for whom the time since the beginning of infection was <1 month and a mean of 2.06 ± 0.54 lowered titers when the infection was less than 3 months old. That was statistically significant compared to sera from patients with infections of greater than 6 months (mean 0.20 ± 0.41) and with infections between 3 and 6 months (mean, 1.17 ± 0.41) (P = 0.0022 and P < 0.0001, respectively). These results were visualized by Western blotting. We concluded that high avidity (≤1 lowered titer) ruled out infection during the last 6 months and that complete denaturation was related to an infection which had occurred within the previous 3 months. Between these two situations, the avidity test is inconclusive. We suggest using an avidity test for atypical Q fever serology that could be misclassified as residual antibodies (IgG against C. burnetii detected without active or recent infection) and for pregnant women risking obstetrical complications. This new test will dramatically improve the diagnosis and management of patients with Q fever.

Determination of immunodominant scaffolds of Com1 and OmpH antigens of Coxiella burnetii.

Today, there is an increasing emphasis on recombinant vaccines to eliminate the side effects of conventional vaccines such as whole-cell bacteria. Query fever is an emerging disease that causes irreparable complications for both humans and domestic animals. The cause of this disease is Coxiella burnetii, a gram-negative intracellular bacteria. In order to determine the most immunodominant epitopes of Com1 and OmpH antigens of C. burnetii, the most reliable bioinformatics tools with high rates of citation in predicting B cell and T cell epitopes were used. Finally, by comparing the results of all servers, the best overlapped epitopes with the highest antigenicity among different servers were selected. In this regard, epitopes in 18-27and 67-82 amino acids residues were introduced for MHCI and MHCII of T cell, respectively, whereas epitope in 16-25 amino acids residues was introduced for B cell of OmpH antigen. The epitopes in the range of 193-202, 100-108 and 215-223 amino acid residues were preferred for MHCI class of T cell, MHCII class of T cell and B cell of Com1 antigen, respectively. For each antigen, some empirical common epitopic regions were introduced, which included both T and B cells epitopes, 53-65 and 102-111 amino acid residues of OmpH antigen as well as 38-54 range of the amino acid of Com1 antigen. All the predicted epitopes were selected based on their high antigenicity scores and number of non-digestive enzymes. To optimize the application of reported epitopes, various orders of epitopes were arranged in three categories of B cell, T cell and common T and B cells epitopes for each antigen. Then, the best immunodominant scaffolds for each antigen were proposed in these categories. The results demonstrated that the scaffold arranged based on B cell epitopes had the highest antigenicity in both antigens.

Viable Coxiella burnetii Induces Differential Cytokine Responses in Chronic Q Fever Patients Compared to Heat-Killed Coxiella burnetii.

Cytokine responses of chronic Q fever patients to the intracellular bacterium Coxiella burnetii have mostly been studied using ex vivo stimulation of immune cells with heat-killed C. burnetii due to the extensive measures needed to work with viable biosafety level 3 agents. Whether research with heat-killed C. burnetii can be translated to immune responses to viable C. burnetii is imperative for the interpretation of previous and future studies with heat-killed C. burnetii Peripheral blood mononuclear cells (PBMCs) of chronic Q fever patients (n = 10) and healthy controls (n = 10) were stimulated with heat-killed or viable C. burnetii of two strains, Nine Mile and the Dutch outbreak strain 3262, for 24 h, 48 h, and 7 days in the absence or presence of serum containing anti-C. burnetii antibodies. When stimulated with viable C. burnetii, PBMCs of chronic Q fever patients and controls produced fewer proinflammatory cytokines (interleukin-6 [IL-6], tumor necrosis factor alpha, and IL-1ß) after 24 h than after stimulation with heat-killed C. burnetii In the presence of Q fever seronegative serum, IL-10 production was higher after stimulation with viable rather than heat-killed C. burnetii; however, when incubating with anti-C. burnetii antibody serum, the effect on IL-10 production was reduced. Levels of adaptive, merely T-cell-derived cytokine (gamma interferon, IL-17, and IL-22) and CXCL9 production were not different between heat-killed and viable C. burnetii stimulatory conditions. Results from previous and future research with heat-killed C. burnetii should be interpreted with caution for innate cytokines, but heat-killed C. burnetii-induced adaptive cytokine production is representative of stimulation with viable bacteria.

Coxiella burnetii Blocks Intracellular Interleukin-17 Signaling in Macrophages.

Coxiella burnetii is an obligate intracellular bacterium and the etiological agent of Q fever. Successful host cell infection requires the Coxiella type IVB secretion system (T4BSS), which translocates bacterial effector proteins across the vacuole membrane into the host cytoplasm, where they manipulate a variety of cell processes. To identify host cell targets of Coxiella T4BSS effector proteins, we determined the transcriptome of murine alveolar macrophages infected with a Coxiella T4BSS effector mutant. We identified a set of inflammatory genes that are significantly upregulated in T4BSS mutant-infected cells compared to mock-infected cells or cells infected with wild-type (WT) bacteria, suggesting that Coxiella T4BSS effector proteins downregulate the expression of these genes. In addition, the interleukin-17 (IL-17) signaling pathway was identified as one of the top pathways affected by the bacteria. While previous studies demonstrated that IL-17 plays a protective role against several pathogens, the role of IL-17 during Coxiella infection is unknown. We found that IL-17 kills intracellular Coxiella in a dose-dependent manner, with the T4BSS mutant exhibiting significantly more sensitivity to IL-17 than WT bacteria. In addition, quantitative PCR confirmed the increased expression of IL-17 downstream signaling genes in T4BSS mutant-infected cells compared to WT- or mock-infected cells, including the proinflammatory cytokine genes Il1a, Il1b, and Tnfa, the chemokine genes Cxcl2 and Ccl5, and the antimicrobial protein gene Lcn2 We further confirmed that the Coxiella T4BSS downregulates macrophage CXCL2/macrophage inflammatory protein 2 and CCL5/RANTES protein levels following IL-17 stimulation. Together, these data suggest that Coxiella downregulates IL-17 signaling in a T4BSS-dependent manner in order to escape the macrophage immune response.

Coxiella burnetii Inhibits Neutrophil Apoptosis by Exploiting Survival Pathways and Antiapoptotic Protein Mcl-1.

Our previous study demonstrated that neutrophils play an important role in host defense against Coxiella burnetii infection in mice. In this study, avirulent strain C. burnetii Nine Mile phase II (NMII) was used to examine if C. burnetii can modulate mouse bone marrow-derived neutrophil apoptosis. The results indicated that NMII can inhibit neutrophil apoptosis. Western blotting demonstrated that caspase-3 cleavage was decreased in NMII-infected neutrophils, while phosphorylated mitogen-activated protein kinase (MAPK) p38 and extracellular signal-regulated kinase 1 (Erk1) were increased. Additionally, p38, Erk1/2, phosphoinositide 3-kinase (PI3K), or NF-κB inhibitors reduced the ability of NMII to inhibit neutrophil apoptosis. These results suggest that NMII-mediated inhibition of neutrophil apoptosis depends on its ability to activate neutrophil MAPK pathways. Antiapoptotic protein myeloid cell leukemia-1 (Mcl-1) was significantly increased in NMII-infected neutrophils, and an Mcl-1 inhibitor significantly reduced the ability of NMII to inhibit neutrophil apoptosis. Mcl-1 protein stability was enhanced by phosphorylation at Thr-163 by Erk, and the protein levels were regulated by p38, Erk, PI3K, and NF-κB. Furthermore, the observation that a type IV secretion system (T4SS)-deficient dotA mutant showed a significantly reduced ability to inhibit neutrophil apoptosis compared to wild-type (WT) NMII suggests that T4SS-secreted factors may be involved in NMII-induced inhibition of neutrophil apoptosis. Collectively, these results demonstrate that NMII inhibits neutrophil apoptosis through inhibition of caspase-3 cleavage and activation of MAPK survival pathways with subsequent expression and stabilization of antiapoptotic protein Mcl-1, a process that may partially require the T4SS.

Both Major Histocompatibility Complex Class I (MHC-I) and MHC-II Molecules Are Required, while MHC-I Appears To Play a Critical Role in Host Defense against Primary Coxiella burnetii Infection.

To understand the role of class I major histocompatibility complex (MHC-I) and class II MHC (MHC-II) antigen presentation pathways in host defense against Coxiella burnetii infection, we examined whether MHC-I or MHC-II deficiency in mice would significantly influence their susceptibility to virulent C. burnetii Nine Mile phase I (NMI) infection. The results indicate that NMI infection induced more severe disease in both MHC-I-deficient and MHC-II-deficient mice than in wild-type (WT) mice, while only MHC-I-deficient mice developed a severe persistent infection and were unable to control bacterial replication. These results suggest that both MHC-I-restricted CD8+ T cells and MHC-II-restricted CD4+ T cells contribute to host defense against primary C. burnetii infection, while MHC-I-restricted CD8+ T cells appear to play a more critical role in controlling bacterial replication. Additionally, although NMI infection induced more severe disease in TAP1-deficient mice than in their WT counterparts, TAP1 deficiency in mice did not significantly influence their ability to eliminate C. burnetii This suggests that C. burnetii antigen presentation to CD8+ T cells by the MHC-I classical pathway may depend only partially on TAP1. Furthermore, granzyme B deficiency in mice did not significantly alter their susceptibility to C. burnetii infection, but perforin-deficient mice were unable to control host inflammatory responses during primary C. burnetii infection. These results suggest that perforin, but not granzyme B, is required for C. burnetii antigen-specific cytotoxic CD8+ T cells to control primary C. burnetii infection.

Seroprevalence of Coxiella burnetii antibodies and chronic Q fever among post-mortal and living donors of tissues and cells from 2010 to 2015 in the Netherlands.

BackgroundAfter a large Q fever outbreak in the Netherlands in the period from 2007 to 2010, the risk of Q fever transmission through tissue and cell transplantation from undiagnosed chronic Q fever cases became a potential issue. Aim: We aimed to evaluate the risk of Q fever transmission through tissue and cell transplantation. Methods: We performed a retrospective observational cohort study among 15,133 Dutch donors of tissues and stem cells from 2010 to 2015 to assess seroprevalence of Coxiella burnetii antibodies, to identify factors associated with presence of C. burnetii antibodies, and to assess the proportion of undiagnosed chronic Q fever cases. Results: The study population consisted of 9,478 (63%) femoral head donors, 5,090 (34%) post-mortal tissue donors and 565 (4%) cord blood donors. Seroprevalence of C. burnetii antibodies gradually decreased after the outbreak, from 2.1% in 2010 to 1.4% in 2015, with a significant trend in time (p < 0.001). Of 301 seropositive donors, seven (2.3%) were newly detected with chronic Q fever (0.05% of all screened donors). Conclusion: This study shows that seroprevalence of C. burnetii antibodies among donors of tissues and cells in the Netherlands after 2014 was similar to pre-outbreak levels in the general population. The proportion of newly detected chronic Q fever patients among donors of tissues and cells was smaller than 0.1%. This study may prompt discussion on when to terminate the screening programme for chronic Q fever in donors of tissues and cells in the Netherlands.