Rifaximin Fails to Prevent Campylobacteriosis in the Human Challenge Model: A Randomized, Double-Blind, Placebo-Controlled Trial.

Background: Campylobacter species are a leading cause of diarrheal disease globally with significant morbidity. Primary prevention efforts have yielded limited results. Rifaximin chemoprophylaxis decreases rates of travelers' diarrhea and may be suitable for high-risk persons. We assessed the efficacy of rifaximin in the controlled human infection model for Campylobacter jejuni. Methods: Twenty-eight subjects were admitted to an inpatient facility and randomized to a twice-daily dose of 550 mg rifaximin or placebo. The following day, subjects ingested 1.7 × 105 colony-forming units of C. jejuni strain CG8421. Subjects continued prophylaxis for 3 additional days, were followed for campylobacteriosis for 144 hours, and were subsequently treated with azithromycin and ciprofloxacin. Samples were collected to assess immunologic responses to CG8421. Results: There was no difference (P = 1.0) in the frequency of campylobacteriosis in those receiving rifaximin (86.7%) or placebo (84.6%). Additionally, there were no differences in the clinical signs and symptoms of C. jejuni infection to include abdominal pain/cramps (P = 1.0), nausea (P = 1.0), vomiting (P = .2), or fever (P = 1.0) across study groups. Immune responses to the CG8421 strain were comparable across treatment groups. Conclusions: Rifaximin did not prevent campylobacteriosis in this controlled human infection model. Given the morbidity associated with Campylobacter infection, primary prevention efforts remain a significant need. Clinical Trials Registration: NCT02280044.

Plasmodium chabaudi infection induces AID expression in transitional and marginal zone B cells.

INTRODUCTION: Endemic Burkitt's lymphoma (eBL) is associated with Epstein-Barr virus and repeated malaria infections. A defining feature of eBL is the translocation of the c-myc oncogene to the control of the immunoglobulin promoter. Activation-induced cytidine deaminase (AID) has been shown to be critical for this translocation. Malaria infection induces AID in germinal center B cells, but whether malaria infection more broadly affects AID activation in extrafollicular B cells is unknown. METHODS: We either stimulated purified B cells from AID-green fluorescence protein (GFP) reporter mice or infected AID-GFP mice with Plasmodium chabaudi, AID fluorescence was monitored in B cell subsets by flow cytometry. RESULTS: In vitro analysis of B cells from these mice revealed that CpG (a Toll-like receptor 9 ligand) was a potent inducer of AID in both mature and immature B cell subsets. Infection of AID-GFP mice with Plasmodium chabaudi demonstrated that AID expression occurs in transitional and marginal zone B cells during acute malaria infection. Transitional B cells were also capable of differentiating into antibody secreting cells when stimulated in vitro with CpG when isolated from a P. chabaudi-infected mouse. CONCLUSIONS: These data suggest that P. chabaudi is capable of inducing AID expression in B cell subsets that do not participate in the germinal center reaction, suggesting an alternative role for malaria in the etiology of eBL.

Blk haploinsufficiency impairs the development, but enhances the functional responses, of MZ B cells.

Blk was identified two decades ago as a B-cell-specific member of the Src family of tyrosine kinases. Recent studies, however, have discovered that Blk is expressed in many cell types outside of the B lineage, including early thymic precursors, interleukin-17-producing γδ T cells and pancreatic ß-cells. In light of these recent discoveries, we performed a more comprehensive analysis of Blk expression patterns in hematopoietic cells and found that Blk is differentially expressed in mature B-cell subsets, with marginal zone (MZ) B cells expressing high levels, B1 B cells expressing intermediate-to-high levels and follicular (FO) B cells expressing low levels of Blk. To determine whether these differences in Blk expression levels reflected differential requirements for Blk in MZ, B1 and FO B-cell development, we analyzed the effects of reducing and eliminating Blk expression on B-cell development. We report that both Blk haploinsufficiency and Blk deficiency impaired the generation of MZ B cells. Moreover, although there were fewer MZ B cells in Blk(+/-) and Blk(-/-) mice as compared with Blk(+/+) mice, Blk-mutant MZ B cells were hyper-responsive to B-cell receptor stimulation, both in vitro and in vivo. Thus, this study has revealed a previously unappreciated role for Blk in the development and activation of MZ B cells.

Effects of aging on T cell function.

Immunosenescence influences many components of the immune system. Most importantly, profound changes in T cell function are evident in older individuals. The impact of aging on specific T cell subsets has been difficult to examine, but recent advances in murine model systems and new insights into T cell function have allowed for the more precise examination of how T cell responses change with aging. Importantly, recent studies have shown that age-related enhancement of both Th17 generation and regulatory T cell function may contribute to significant changes in immune function. In this review, we summarize the current views on how aging influences the factors that impact T cell function and how this can affect the immune response to infections, vaccinations, and tumors.

Bone marrow precursor cells from aged mice generate CD4 T cells that function well in primary and memory responses.

Understanding how aging impacts the function of memory CD4 T cells is critical for designing effective vaccines. Our studies show that immunological memory generated during youth functions well into old age, whereas that generated later in life functions poorly. This is the result of declines in the function of naive CD4 T cells from aged individuals and contributes to reduced efficacy of vaccines in the elderly. To begin to identify the cause of this defect, we examined the function of memory T cells generated from bone marrow precursor cells (BMPC) from young or aged mice in young hosts. In two different models, memory cells derived from young and aged BMPC exhibit good ex vivo and in vivo function. Importantly, memory CD4 T cells generated from aged BMPC exhibit potent cognate helper function for humoral responses, which are critical for effective immunization. These results indicate that there are no apparent age-related intrinsic defects in BMPC with regards to generation of functional memory T cells.

An ESAT-6:CFP10 DNA vaccine administered in conjunction with Mycobacterium bovis BCG confers protection to cattle challenged with virulent M. bovis.

The potency of genetic immunization observed in the mouse has demonstrated the utility of DNA vaccines to induce cell-mediated and humoral immune responses. However, it has been relatively difficult to generate comparable responses in non-rodent species. The use of molecular adjuvants may increase the magnitude of these suboptimal responses. In this study, we demonstrate that the co-administration of plasmid-encoded GM-CSF and CD80/CD86 with a novel ESAT-6:CFP10 DNA vaccine against bovine tuberculosis enhances antigen-specific cell-mediated immune responses. ESAT-6:CFP10+GM-CSF+CD80/CD86 DNA vaccinated animals exhibited significant (p<0.01) antigen-specific proliferative responses compared to other DNA vaccinates. Increased expression (p< or =0.05) of CD25 on PBMC from ESAT-6:CFP10+GM-CSF+CD80/CD86 DNA vaccinates was associated with increased proliferation, as compared to control DNA vaccinates. Significant (p<0.05) numbers of ESAT-6:CFP10-specific IFN-gamma producing cells were evident from all ESAT-6:CFP10 DNA vaccinated animals compared to control DNA vaccinates. However, the greatest increase in IFN-gamma producing cells was from animals vaccinated with ESAT-6:CFP10+GM-CSF+CD80/CD86 DNA. In a low-dose aerosol challenge trial, calves vaccinated as neonates with Mycobacterium bovis BCG and ESAT-6:CFP10+GM-CSF+CD80/CD86 DNA exhibited decreased lesion severity in the lung and lung-associated lymph nodes following viruluent M. bovis challenge compared to other vaccinated animals or non-vaccinated controls. These data suggest that a combined vaccine regimen of M. bovis BCG and a candidate ESAT-6:CFP10 DNA vaccine may offer greater protection against tuberculosis in cattle than vaccination with BCG alone.

Analysis of immune responses directed toward a recombinant early secretory antigenic target six-kilodalton protein-culture filtrate protein 10 fusion protein in Mycobacterium bovis-infected cattle.

Cell-mediated immune responses are critical for protective immunity to mycobacterial infections. Recent progress in defining mycobacterial antigens has determined that region of difference 1 (RD1) gene products induce strong T-cell responses, particularly the early secretory antigenic target 6-kDa (ESAT-6) protein and culture filtrate protein 10 (CFP10). However, comprehensive analysis of the immune response towards these antigens is incompletely characterized. To evaluate recall responses to ESAT-6 and CFP10, peripheral blood mononuclear cells from M. bovis-infected cattle were stimulated in vitro with a recombinant ESAT-6 (rESAT-6)-CFP10 fusion protein and compared to responses induced by M. bovis-derived purified protein derivative. Following antigenic stimulation, activation marker expression was evaluated. Significant proliferative responses (P < 0.05) were evident in CD4(+), CD8(+), immunoglobulin M-positive, and CD172a(+) cell fractions after 6 days of culture. Expression of CD25 and CD26 was increased (P < 0.05) on CD4(+), CD8(+), and gammadelta T-cell-receptor-positive cells. CD4(+) and CD8(+) cells also exhibited significant changes (P < 0.05) in expression of CD45 isoforms. Using a flow cytometry-based proliferation assay, it was determined that CD45R expression is downregulated (P < 0.05) and that CD45RO expression is upregulated (P < 0.05) on proliferating (i.e., activated) CD4(+) cells. Collectively, data indicate that recall immune responses directed toward the rESAT-6-CFP10 fusion protein or purified protein derivative are comparable and that recall to mycobacterial antigens correlates with a CD45RO(+) phenotype.

CD80 and CD86, but not CD154, augment DNA vaccine-induced protection in experimental bovine tuberculosis.

DNA vaccination is known to elicit robust cellular and humoral responses to encoded antigen. The co-administration of costimulatory molecules CD80 (B7-1), CD86 (B7-2) and CD154 (CD40L) has been shown to enhance immune responses in several murine models. The role of specific costimulatory molecules in non-rodent species remains incompletely characterized. In these studies, we demonstrate that the co-administration of CD80 and CD86, but not CD154, to an existing candidate subunit DNA vaccine (ESAT-6) against bovine tuberculosis, enhances protection after aerosol challenge with virulent Mycobacterium bovis. Additionally, we have shown that vaccination with M. bovis BCG is protective against tuberculosis following aerosol challenge in cattle. Two independent trials were conducted in cattle to determine the adjuvant effect of encoded antigen + CD80/CD86 and directly compare the adjuvant activities of CD80/CD86 to those of CD154. Co-administration of either CD80/CD86 or CD154 enhanced ESAT-6-specific IFN-gamma responses as compared to animals vaccinated with ESAT-6 DNA alone. However, following aerosol challenge, only animals vaccinated with CD80/CD86 possessed decreased pathology of the lungs and associated lymph nodes, as measured by gross examination, radiographic lesion morphometry and bacterial recovery. Collectively, these results demonstrate that the co-administration of costimulatory molecules with a protective antigen target enhances bovine immune responses to DNA vaccination, and that CD80/CD86 is superior to CD154 in augmenting DNA vaccine-induced protection in experimental bovine tuberculosis.