Gut Microbiota-Induced Immunoglobulin G Controls Systemic Infection by Symbiotic Bacteria and Pathogens.

The gut microbiota is compartmentalized in the intestinal lumen and induces local immune responses, but it remains unknown whether the gut microbiota can induce systemic response and contribute to systemic immunity. We report that selective gut symbiotic gram-negative bacteria were able to disseminate systemically to induce immunoglobulin G (IgG) response, which primarily targeted gram-negative bacterial antigens and conferred protection against systemic infections by E. coli and Salmonella by directly coating bacteria to promote killing by phagocytes. T cells and Toll-like receptor 4 on B cells were important in the generation of microbiota-specific IgG. We identified murein lipoprotein (MLP), a highly conserved gram-negative outer membrane protein, as a major antigen that induced systemic IgG homeostatically in both mice and humans. Administration of anti-MLP IgG conferred crucial protection against systemic Salmonella infection. Thus, our findings reveal an important function for the gut microbiota in combating systemic infection through the induction of protective IgG.

Intraspecific variation in immune gene expression and heritable symbiont density.

Host genetic variation plays an important role in the structure and function of heritable microbial communities. Recent studies have shown that insects use immune mechanisms to regulate heritable symbionts. Here we test the hypothesis that variation in symbiont density among hosts is linked to intraspecific differences in the immune response to harboring symbionts. We show that pea aphids (Acyrthosiphon pisum) harboring the bacterial endosymbiont Regiella insecticola (but not all other species of symbionts) downregulate expression of key immune genes. We then functionally link immune expression with symbiont density using RNAi. The pea aphid species complex is comprised of multiple reproductively-isolated host plant-adapted populations. These 'biotypes' have distinct patterns of symbiont infections: for example, aphids from the Trifolium biotype are strongly associated with Regiella. Using RNAseq, we compare patterns of gene expression in response to Regiella in aphid genotypes from multiple biotypes, and we show that Trifolium aphids experience no downregulation of immune gene expression while hosting Regiella and harbor symbionts at lower densities. Using F1 hybrids between two biotypes, we find that symbiont density and immune gene expression are both intermediate in hybrids. We propose that in this system, Regiella symbionts are suppressing aphid immune mechanisms to increase their density, but that some hosts have adapted to prevent immune suppression in order to control symbiont numbers. This work therefore suggests that antagonistic coevolution can play a role in host-microbe interactions even when symbionts are transmitted vertically and provide a clear benefit to their hosts. The specific immune mechanisms that we find are downregulated in the presence of Regiella have been previously shown to combat pathogens in aphids, and thus this work also highlights the immune system's complex dual role in interacting with both beneficial and harmful microbes.

Macrophage-derived LTB4 promotes abscess formation and clearance of Staphylococcus aureus skin infection in mice.

The early events that shape the innate immune response to restrain pathogens during skin infections remain elusive. Methicillin-resistant Staphylococcus aureus (MRSA) infection engages phagocyte chemotaxis, abscess formation, and microbial clearance. Upon infection, neutrophils and monocytes find a gradient of chemoattractants that influence both phagocyte direction and microbial clearance. The bioactive lipid leukotriene B4 (LTB4) is quickly (seconds to minutes) produced by 5-lipoxygenase (5-LO) and signals through the G protein-coupled receptors LTB4R1 (BLT1) or BLT2 in phagocytes and structural cells. Although it is known that LTB4 enhances antimicrobial effector functions in vitro, whether prompt LTB4 production is required for bacterial clearance and development of an inflammatory milieu necessary for abscess formation to restrain pathogen dissemination is unknown. We found that LTB4 is produced in areas near the abscess and BLT1 deficient mice are unable to form an abscess, elicit neutrophil chemotaxis, generation of neutrophil and monocyte chemokines, as well as reactive oxygen species-dependent bacterial clearance. We also found that an ointment containing LTB4 synergizes with antibiotics to eliminate MRSA potently. Here, we uncovered a heretofore unknown role of macrophage-derived LTB4 in orchestrating the chemoattractant gradient required for abscess formation, while amplifying antimicrobial effector functions.

The clinical significance of IL-6 s and IL-27 s in Bronchoalveolar lavage fluids from children with mycoplasma pneumoniae pneumonia.

BACKGROUND: IL-6 was associated with the severity of mycoplasma pneumoniae pneumonia (MPP). But the relationship between IL-27 and MPP was unknown. METHODS: Ninety-eight patients with MPP < 14 years old were enrolled in this study and divided into groups by severity (mild cases and severe cases), infection types (MP single infection group and MP mixed infection group) and DNA loads (low MP DNA loads group and high MP DNA loads group), respectively. Fifteen children with foreign bodies in bronchus were also enrolled as control. IL-6 s and IL-27 s in bronchoalveolar lavage fluids (BALFs) from these children were measured by ELISA. RESULTS: There were significant differences in IL-6 s of BALFs from patients between mild cases and severe cases, MP single infection group and MP mixed infection group, and low MP DNA loads group and high MP DNA loads group, respectively (P < 0.05). Compared with IL-6 s of BALFs from control, IL-6 s in BALFs from the 6 patient groups were significantly higher (P < 0.05). IL-27 s in BALFs from MP mixed infection group were significantly lower than those from MP single infection group and control (P < 0.05) respectively. CONCLUSION: IL-6 was firmly associated with MPP and had potential application in clinical practice while IL-27 was not related to MP infection.

Sequence tag-based analysis of microbial population dynamics.

We describe sequence tag-based analysis of microbial populations (STAMP) for characterization of pathogen population dynamics during infection. STAMP analyzes the frequency changes of genetically 'barcoded' organisms to quantify population bottlenecks and infer the founding population size. Analyses of intraintestinal Vibrio cholerae revealed infection-stage and region-specific host barriers to infection and showed unexpected V. cholerae migration counter to intestinal flow. STAMP provides a robust, widely applicable analytical framework for high-confidence characterization of in vivo microbial dissemination.

Azithromycin 1.5g Over 5 Days Compared to 1g Single Dose in Urethral Mycoplasma genitalium: Impact on Treatment Outcome and Resistance.

BACKGROUND: We evaluated the impact of extended azithromycin (1.5g over 5 days) on selection of macrolide resistance and microbiological cure in men with Mycoplasma genitalium urethritis during 2013-2015 and compared this to cases treated with azithromycin 1g in 2012-2013. METHODS: Microbiological cure was determined for men with M. genitalium urethritis treated with azithromycin 1.5g using quantitative polymerase chain reaction specific for M. genitalium DNA on samples 14-100 days post-treatment. Pre- and post-treatment macrolide resistance mutations were detected by sequencing the 23 S gene. RESULTS: There was no difference in proportions with microbiological cure between azithromycin 1.5g and 1g: 62/106 (58%; 95% confidence interval [CI], 49%, 68%) and 56/107 (52%; 95%CI 42-62%), P = .34, respectively. Also, there was no difference in the proportion of wild-type 23 S rRNA (presumed macrolide sensitive) infections cured after 1.5g and azithromycin 1g: 28/34 (82%; 95%CI 65-92%) and 49/60 (82%; 95%CI 70-90%), P=1.0, respectively. There was no difference between 1.5g and 1g in the proportions of wild-type infections with post-treatment resistance mutations: 4/34 (12%; 95%CI 3-27%) and 11/60 (18%; 95%CI 10-30%), respectively, P = .40. Pre-treatment resistance was present in 51/98 (52%; 95%CI 42-62%) cases in 2013-2015 compared to 47/107 (44%; 95%CI 34-54%) in 2012-2013, P = .25. CONCLUSIONS: Extended azithromycin 1.5g was no more effective than a single 1g dose at achieving cure of M. genitalium urethritis and importantly did not reduce the selection of macrolide resistance. Nonmacrolide and new approaches for the treatment of M. genitalium urethritis are required.

Effect of cadexomer iodine on the microbial load and diversity of chronic non-healing diabetic foot ulcers complicated by biofilm in vivo.

Objectives: The performance of cadexomer iodine was determined against microbial populations from chronic non-healing diabetic foot ulcers (DFUs) complicated by biofilm in vivo , using molecular, microscopy and zymography methods. Methods: Chronic non-healing DFUs due to suspected biofilm involvement were eligible for enrolment. DNA sequencing and real-time quantitative PCR was used to determine the microbial load and diversity of tissue punch biopsies obtained pre- and post-treatment. Scanning electron microscopy and/or fluorescence in situ hybridization confirmed the presence or absence of biofilm. Zymography was used to determine levels of wound proteases. Results: Seventeen participants were recruited over a 6 month period. Scanning electron microscopy and or fluorescence in situ hybridization confirmed the presence of biofilm in all samples. Eleven participants exhibited log 10 reductions in microbial load after treatment (range 1-2 log 10 ) in comparison with six patients who experienced <1 log 10 reduction ( P = 0.04). Samples were tested for levels of wound proteases pre- and post-treatment. Reductions in the microbial load correlated to reductions in wound proteases pre- and post-treatment ( P = 0.03). Conclusions: To the best of our knowledge, this study represents the first in vivo evidence, employing a range of molecular and microscopy techniques, of the ability of cadexomer iodine to reduce the microbial load of chronic non-healing DFUs complicated by biofilm. Further analyses correlating log reductions to optimal duration of therapy and improvements in clinical parameters of wound healing in a larger cohort are required.

Silkworm model for Francisella novicida infection.

Understanding the virulence and pathogenesis of human pathogens using insect models is an increasingly popular method. Francisella novicida, which is virulent in mice but non-pathogenic to immunocompetent humans, is widely used as an ideal candidate for Francisella research. In this study, we developed a silkworm (Bombyx mori) infection model for F. novicida by inoculating the hemocoels of silkworms with F. novicida. We found that silkworms died within 3-7 days of F. novicida infection. However, the deletion mutant of DotU, the core part of type VI secretion systems, failed to kill silkworm. In whole silkworm bodies, the bacterial load of the DotU deletion mutant was significantly less than that of the wild-type strain. Approximately 10-fold increase in bacterial load was recorded in hemolymph and subcutaneous tissues compared with that in the silk gland, Malpighian tubule, and reproductive organs. The CFU count of the DotU deletion mutant in all organs was similar results to the whole body CFU count. Confocal microscopy further confirmed the arrested growth of the mutant strain within hemocytes. The intracellular growth of F. novicida strains was also analyzed using the silkworm ovary-derived cell line BmN4. In BmN4, both CFU count assay and confocal microscopy revealed extensive growth of the wild-type strain compared with that of the mutant strain. Francisella DotU has already been proven as a virulence factor in mammals, and it was also found to be an essential virulence factor in our silkworm infection model. Therefore, this silkworm infection model is suitable for identifying new virulence factors of Francisella.

Hepcidin as a Major Component of Renal Antibacterial Defenses against Uropathogenic Escherichia coli.

The iron-regulatory peptide hepcidin exhibits antimicrobial activity. Having previously shown hepcidin expression in the kidney, we addressed its role in urinary tract infection (UTI), which remains largely unknown. Experimental UTI was induced in wild-type (WT) and hepcidin-knockout (Hepc-/-) mice using the uropathogenic Escherichia coli CFT073 strain. Compared with infected WT mice, infected Hepc-/- mice showed a dramatic increase in renal bacterial load. Moreover, bacterial invasion was significantly dampened by the pretreatment of WT mice with hepcidin. Infected Hepc-/- mice exhibited decreased iron accumulation in the renal medulla and significant attenuation of the renal inflammatory response. Notably, we demonstrated in vitro bacteriostatic activity of hepcidin against CFT073. Furthermore, CFT073 repressed renal hepcidin, both in vivo and in cultured renal cells, and reduced phosphorylation of SMAD kinase in vivo, suggesting a bacterial strategy to escape the antimicrobial activities of hepcidin. In conclusion, we provide new mechanisms by which hepcidin contributes to renal host defense and suggest that targeting hepcidin offers a strategy to prevent bacterial invasion.

Test of Cure for Anogenital Gonorrhoea Using Modern RNA-Based and DNA-Based Nucleic Acid Amplification Tests: A Prospective Cohort Study.

BACKGROUND: The use of nucleic acid amplification tests (NAATs) to diagnose Neisseria gonorrhoeae infections complicates the performance of a test of cure (TOC) to monitor treatment failure, if this is indicated. As evidence for the timing of TOC using modern NAATs is limited, we performed a prospective cohort study to assess time to clearance when using modern RNA- and DNA-based NAATs. METHODS: We included patients with anogenital gonorrhoea visiting the Sexually Transmitted Infection Clinic Amsterdam from March through October 2014. After treatment with ceftriaxone mono- or dual therapy (with azithromycin or doxycycline), anal, vaginal, or urine samples were self-collected during 28 consecutive days, and analyzed using an RNA-based NAAT (Aptima Combo 2) and a DNA-based NAAT (Cobas 4800). Clearance was defined as 3 consecutive negative results, and blips as isolated positive results following clearance. RESULTS: We included 77 patients; 5 self-cleared gonorrhoea before treatment and 10 were lost to follow-up. Clearance rate of the remaining 62 patients was 100%. Median time to clearance was 2 days, with a range of 1-7 days for RNA-based NAAT and 1-15 days for DNA-based NAAT. The risk of finding a blip after clearance was 0.8% and 1.5%, respectively. One patient had a reinfection. CONCLUSIONS: If indicated, we recommend that TOC be performed for anogenital gonorrhoea at least 7 or 14 days after administering therapy, when using modern RNA- or DNA-based NAATs, respectively. When interpreting TOC results for possible treatment failure, both the occurrence of blips and a possible reinfection need to be taken into account.

Interleukin-10 plays a key role in the modulation of neutrophils recruitment and lung inflammation during infection by Streptococcus pneumoniae.

Streptococcus pneumoniae is a major aetiological agent of pneumonia worldwide, as well as otitis media, sinusitis, meningitis and sepsis. Recent reports have suggested that inflammation of lungs due to S. pneumoniae infection promotes bacterial dissemination and severe disease. However, the contribution of anti-inflammatory molecules to the pathogenesis of S. pneumoniae remains unknown. To elucidate whether the production of the anti-inflammatory cytokine interleukin-10 (IL-10) is beneficial or detrimental for the host during pneumococcal pneumonia, we performed S. pneumoniae infections in mice lacking IL-10 (IL-10(-/-) mice). The IL-10(-/-) mice showed increased mortality, higher expression of pro-inflammatory cytokines, and an exacerbated recruitment of neutrophils into the lungs after S. pneumoniae infection. However, IL-10(-/-) mice showed significantly lower bacterial loads in lungs, spleen, brain and blood, when compared with mice that produced this cytokine. Our results support the notion that production of IL-10 during S. pneumoniae infection modulates the expression of pro-inflammatory cytokines and the infiltration of neutrophils into the lungs. This feature of IL-10 is important to avoid excessive inflammation of tissues and to improve host survival, even though bacterial dissemination is less efficient in the absence of this cytokine.

Accumulation of clinically relevant antibiotic-resistance genes, bacterial load, and metals in freshwater lake sediments in Central Europe.

Wastewater treatment plants (WWTP) receive the effluents from various sources (communities, industrial, and hospital effluents) and are recognized as reservoir for antibiotic-resistance genes (ARGs) that are associated with clinical pathogens. The aquatic environment is considered a hot-spot for horizontal gene transfer, and lake sediments offer the opportunity for reconstructing the pollution history and evaluating the impacts. In this context, variation with depth and time of the total bacterial load, the abundance of faecal indicator bacteria (FIB; E. coli and Enterococcus spp. (ENT)), Pseudomonas spp., and ARGs (blaTEM, blaSHV, blaCTX-M, blaNDM, and aadA) were quantified in sediment profiles of different parts of Lake Geneva using quantitative PCR. The abundance of bacterial marker genes was identified in sediments contaminated by WWTP following eutrophication of the lake. Additionally, ARGs, including the extended-spectrum ß-lactam- and aminoglycoside-resistance genes, were identified in the surface sediments. The ARG and FIB abundance strongly correlated (r ≥ 0.403, p < 0.05, n = 34) with organic matter and metal concentrations in the sediments, indicating a common and contemporary source of contamination. The contamination of sediments by untreated or partially treated effluent water can affect the quality of ecosystem. Therefore, the reduction of contaminants from the source is recommended for further improvement of water quality.

Performance evaluation of three commercial molecular assays for the detection of Mycobacterium tuberculosis from clinical specimens in a high TB-HIV-burden setting.

BACKGROUND: A major challenge faced by countries with a high burden of tuberculosis (TB) is early detection especially in individuals with paucibacillary disease which is common in HIV endemic settings. Remarkable efforts have been made globally to accelerate the development and expansion of new diagnostic technologies that allow better and earlier diagnosis of active tuberculosis particularly directly from clinical specimens with a few commercial options available. These include GenoType MTBDRplus Version 2.0 (Hain Lifescience), Xpert® MTB/RIF (Cepheid) and Anyplex™ plus MTB/NTM/DR-TB Real-time detection (Seegene). We evaluated the diagnostic performance of these three commercial molecular assays for the detection of Mycobacterium tuberculosis complex from clinical specimens in a high TB-HIV-burden setting. METHODS: This was a retrospective laboratory-based study using stored remnant sediments from clinical specimens of presumptive pulmonary TB cases. A stratified sample of smear positive TB, smear negative TB and TB culture negatives was included. All the samples were tested on the three molecular assays following the manufacturers' instructions; except for Anyplex™plus, for which DNA extraction was performed using the NucliSENS® easyMAG® platform (bioMerieux). Samples were also processed for liquid TB culture and time-to-culture positivity was recorded. RESULTS: Of the 90 sediments processed, 81 were analyzable across all three systems. The overall sensitivity was highest for Xpert® MTB/RIF (89.1%) followed by GenoType MTBDRplus (70.9%) and Anyplex™ plus (65.5%). The specificity and sensitivity in smear positive cases was comparable across all systems. There was a significant difference in sensitivity between Xpert® MTB/RIF and the other two assays for smear-negative cases (P < 0.05). The performance in cases where the time-to-culture positivity was ≥ 20 days was also significantly poorer for both Anyplex™ plus and GenoType MTBDRplus compared to Xpert® MTB/RIF (P < 0.05). Xpert® MTB/RIF achieved 100% specificity, while Anyplex™ plus and GenoType MTBDRplus achieved 96.2 and 92.3% respectively. CONCLUSION: The Xpert® MTB/RIF was superior to the other two assays for the detection of TB in smear negative specimens notably when bacterial loads are very low in sputum. It is important that studies reporting on test performance stratify their results by time-to-culture positivity to accurately assess clinical performance especially in high HIV settings.

Treatment efficacy, treatment failures and selection of macrolide resistance in patients with high load of Mycoplasma genitalium during treatment of male urethritis with josamycin.

BACKGROUND: Azithromycin has been widely used for Mycoplasma genitalium treatment internationally. However, the eradication efficacy has substantially declined recent decade. In Russia, josamycin (another macrolide) is the recommended first-line treatment for M. genitalium infections, however, no data regarding treatment efficacy with josamycin and resistance in M. genitalium infections have been internationally published. We examined the M. genitalium prevalence in males attending an STI clinic in Moscow, Russia from December 2006 to January 2008, investigated treatment efficacy with josamycin in male urethritis, and monitored the M. genitalium DNA eradication dynamics and selection of macrolide resistance in M. genitalium during this treatment. METHODS: Microscopy and real-time PCRs were used to diagnose urethritis and non-viral STIs, respectively, in males (n = 320). M. genitalium positive patients were treated with recommended josamycin regimen and treatment efficacy was monitored using quantitative real-time PCR. Macrolide resistance mutations were identified using sequencing of the 23S rRNA gene. RESULTS: Forty-seven (14.7%) males were positive for M. genitalium only and most (85.1%) of these had symptoms and signs of urethritis. Forty-six (97.9%) males agreed to participate in the treatment efficacy monitoring. All the pre-treatment M. genitalium specimens had wild-type 23S rRNA. The elimination of M. genitalium DNA was substantially faster in patients with lower pre-treatment M. genitalium load, and the total eradication rate was 43/46 (93.5%). Of the six patients with high pre-treatment M. genitalium load, three (50%) remained positive post-treatment and these positive specimens contained macrolide resistance mutations in the 23S rRNA gene, i.e., A2059G (n = 2) and A2062G (n = 1). CONCLUSIONS: M. genitalium was a frequent cause of male urethritis in Moscow, Russia. The pre-treatment M. genitalium load might be an effective predictor of eradication efficacy with macrolides (and possibly additional antimicrobials) and selection of macrolide resistance. Additional in vivo and in vitro data are crucial to support the recommendation of using josamycin as first-line treatment for M. genitalium infections in Russia. It would be valuable to develop international M. genitalium management guidelines, and quantitative diagnostic PCRs determining also M. genitalium load and resistance mutations (for macrolides and ideally also moxifloxacin) should ideally be recommended.

Influenza A exacerbates Staphylococcus aureus pneumonia by attenuating IL-1ß production in mice.

Pneumonia is a leading cause of death worldwide. Staphylococcal aureus can be a cause of severe pneumonia alone or a common pathogen in secondary pneumonia following influenza. Recently, we reported that preceding influenza attenuated the Type 17 pathway, increasing the lung's susceptibility to secondary infection. IL-1ß is known to regulate host defense, including playing a role in Th17 polarization. We examined whether IL-1ß signaling is required for S. aureus host defense and whether influenza infection impacted S. aureus-induced IL-1ß production and subsequent Type 17 pathway activation. Mice were challenged with S. aureus (USA 300), with or without preceding Influenza A/PR/8/34 H1N1 infection. IL-1R1(-/-) mice had significantly higher S. aureus burden, increased mortality, and decreased Type 17 pathway activation following S. aureus challenge. Coinfected mice had significantly decreased IL-1ß production versus S. aureus infection alone at early time points following bacterial challenge. Preceding influenza did not attenuate S. aureus-induced inflammasome activation, but there was early suppression of NF-κB activation, suggesting an inhibition of NF-κB-dependent transcription of pro-IL-1ß. Furthermore, overexpression of IL-1ß in influenza and S. aureus-coinfected mice rescued the induction of IL-17 and IL-22 by S. aureus and improved bacterial clearance. Finally, exogenous IL-1ß did not significantly rescue S. aureus host defense during coinfection in IL-17RA(-/-) mice or in mice in which IL-17 and IL-22 activity were blocked. These data reveal a novel mechanism by which Influenza A inhibits S. aureus-induced IL-1ß production, resulting in attenuation of Type 17 immunity and increased susceptibility to bacterial infection.

TLR8 gene polymorphism and association in bacterial load in southern Punjab of Pakistan: an association study with pulmonary tuberculosis.

Only 5-10% of people infected with Mycobacterium tuberculosis develop active tuberculosis which suggests a role of genetic variation in host immunity. Genetic variants in TLRs are potential indicator for host susceptibility and outcome of several diseases. We explored the association of nonsynonymous genetic variants (Met1Val) with Toll-like receptor 8 in Pakistani population. Genotypic and allelic distribution of TLR8 polymorphism (rs3764880) in patients with TB and healthy donors from different areas of southern Punjab, Pakistan, was determined. Results provide that our population is highly influenced by TLR8 Met1Val SNP for TB, and G allele appeared to increase TB susceptibility. Mutant genotype GG or G/- and G allele was significantly higher among all the categories of cases than in controls. Among different levels of bacillary load and genotypes, GG or G/- and G allele significantly supports the incidence of 2 + class for bacterial load.

Toll-like receptor 4 is essential to preserving cardiac function and survival in low-grade polymicrobial sepsis.

BACKGROUND: Toll-like receptor 4 (TLR4), the receptor for endotoxin, mediates hyperinflammatory response and contributes to high mortality during both endotoxin shock and severe sepsis. However, little is known about the role of TLR4 in the pathogenesis of low-grade polymicrobial sepsis, which is often associated with immunosuppression. METHODS: Low-grade polymicrobial sepsis was generated by cecum ligation and puncture. Mortality was monitored in wild- type (C57BL/10ScSn) and TLR4def (C57BL/10ScCr) mice. Ex vivo heart and individual cardiomyocyte function were assessed in Langendorff (Hugo Sachs Elektronik; Harvard Apparatus, Holliston, MA) and IonOptix systems (IonOptix, Milton, MA), respectively. Serum chemistry was tested for liver and kidney injury. Cytokines were examined using a multiplex immunoassay. Neutrophil migratory and phagocytic functions were assessed using flow cytometry. Reactive oxygen species were measured using redox-sensitive dichlorodihydrofluorescein dye. RESULTS: Following cecum ligation and puncture, wild-type mice developed bacterial peritonitis with mild cardiac dysfunction (n=3 in sham and n=8 in cecum ligation and puncture) and a mortality of 23% within 14 days (n=22). In comparison, septic TLR4def mice had deleterious cardiac dysfunction (n=6 in sham and n=10 in cecum ligation and puncture), kidney and liver injury (n=7), and much higher mortality at 81% (n=21). The deleterious effects observed in septic TLR4def mice were associated with increased local and systemic cytokine response, reduced neutrophil migratory and phagocytic function, increased reactive oxygen species generation in leukocytes, and impaired bacterial clearance. CONCLUSION: TLR4 plays an essential role in host defense against low-grade polymicrobial sepsis by mediating neutrophil migratory/phagocytic functions, attenuating inflammation, reducing reactive oxygen species generation, and enhanced bacterial clearance.

Marginal zone B cell is a major source of Il-10 in Listeria monocytogenes susceptibility.

Rag-1-knockout (KO) mice are highly resistant to Listeria monocytogenes infection. The role played by the many Rag-1-dependent lymphocyte lineages was studied using a genetic approach in which each Rag-1-dependent lymphocyte lineage was eliminated one at a time. Only B cell-deficient Igh-KO mice displayed reduced bacterial load and improved survival upon Listeria infection. Listeria infection of Rag-1-KO and Il-10-KO hosts that had been adoptively transferred with wild-type marginal zone B (MZB) cells, but not follicular B cells, resulted in heightened bacterial load and increased Il-10 production in the spleen, but not the liver. This MZB cell-dependent increase in bacterial load was eliminated by anti-Il-10 mAb. In addition, Listeria infection of MZB cell-deficient Rbpj-cKO mice showed decreased bacterial load and increased survival. Whereas multiple cell types have been shown to be capable of Il-10 production, our results indicate that the MZB cell is the most dominant and relevant Il-10 source in the context of Listeria susceptibility. In marked contrast to the generally protective nature of MZB cells in defending against pathogenic infection, our results demonstrate that MZB cells play a detrimental role in Listeria infection and possibly other infections as well.

IL-23-dependent IL-17 drives Th1-cell responses following Mycobacterium bovis BCG vaccination.

The generation of effective type 1 T helper (Th1)-cell responses is required for immunity against intracellular bacteria. However, some intracellular bacteria require interleukin (IL)-17 to drive Th1-cell immunity and subsequent protective host immunity. Here, in a model of Mycobacterium bovis Bacille Calmette-Guerin (BCG) vaccination in mice, we demonstrate that the dependence on IL-17 to drive Th1-cell responses is a host mechanism to overcome bacteria-induced IL-10 inhibitory effects. We show that BCG-induced prostaglandin-E2 (PGE2) promotes the production of IL-10 which limits Th1-cell responses, while simultaneously inducing IL-23 and Th17-cell differentiation. The ability of IL-17 to downregulate IL-10 and induce IL-12 production allows the generation of subsequent Th1-cell responses. Accordingly, BCG-induced Th17-cell responses precede the generation of Th1-cell responses in vivo, whereas the absence of the IL-23 pathway decreases BCG vaccine-induced Th17 and Th1-cell immunity and subsequent vaccine-induced protection upon M. tuberculosis challenge. Importantly, in the absence of IL-10, BCG-induced Th1-cell responses occur in an IL-17-independent manner. These novel data demonstrate a role for the IL-23/IL-17 pathway in driving Th1-cell responses, specifically to overcome IL-10-mediated inhibition and, furthermore, show that in the absence of IL-10, the generation of BCG-induced Th1-cell immunity is IL-17 independent.

Differential effects of interleukin-17 receptor signaling on innate and adaptive immunity during central nervous system bacterial infection.

Although IL-17A (commonly referred to as IL-17) has been implicated in the pathogenesis of central nervous system (CNS) autoimmune disease, its role during CNS bacterial infections remains unclear. To evaluate the broader impact of IL-17 family members in the context of CNS infection, we utilized IL-17 receptor (IL-17R) knockout (KO) mice that lack the ability to respond to IL-17, IL-17F and IL-17E (IL-25). In this article, we demonstrate that IL-17R signaling regulates bacterial clearance as well as natural killer T (NKT) cell and gamma-delta (γδ) T cell infiltrates during Staphylococcus aureus-induced brain abscess formation. Specifically, when compared with wild-type (WT) animals, IL-17R KO mice exhibited elevated bacterial burdens at days 7 and 14 following S. aureus infection. Additionally, IL-17R KO animals displayed elevated neutrophil chemokine production, revealing the ability to compensate for the lack of IL-17R activity. Despite these differences, innate immune cell recruitment into brain abscesses was similar in IL-17R KO and WT mice, whereas IL-17R signaling exerted a greater influence on adaptive immune cell recruitment. In particular, γδ T cell influx was increased in IL-17R KO mice at day 7 post-infection. In addition, NK1.1high infiltrates were absent in brain abscesses of IL-17R KO animals and, surprisingly, were rarely detected in the livers of uninfected IL-17R KO mice. Although IL-17 is a key regulator of neutrophils in other infection models, our data implicate an important role for IL-17R signaling in regulating adaptive immunity during CNS bacterial infection.