CD4+ T cell count and HIV-1 viral load dynamics positively impacted by H. pylori infection in HIV-positive patients regardless of ART status in a high-burden setting.

BACKGROUND: There is a widespread co-infection of HIV and Helicobacter pylori (H. pylori) globally, particularly in developing countries, and it has been suggested that this co-infection may affect the course of HIV disease. However, the interplay between H. pylori infection and HIV disease progression is not fully elucidated. In this study, we investigated the effect of H. pylori co-infection on CD4+ T cell count and HIV viral load dynamics in HIV-positive individuals in a high co-endemic setting. METHODS: A comparative cross-sectional study was conducted among 288 HIV-positive and 175 HIV-negative individuals, both with and without H. pylori infection. Among HIV-positive participants, 195 were on antiretroviral therapy (ART) and 93 were ART-naïve. CD4+ T cell count and HIV-1 viral load were measured and compared between H. pylori-infected and -uninfected individuals, taking into account different HIV and ART status. RESULT: Our study demonstrated that individuals infected with H. pylori had a significantly higher CD4+ T cell count compared to uninfected controls among both HIV-negative and HIV-positive participants, regardless of ART therapy. Conversely, HIV/H. pylori co-infected participants had lower HIV-1 viral load than those without H. pylori infection. Linear regression analysis further confirmed a positive association between H. pylori infection, along with other clinical factors such as BMI, ART, and duration of therapy, with CD4+ T cell count while indicating an inverse relationship with HIV-1 viral load in HIV-positive patients. Additionally, factors such as khat chewing, age and WHO clinical stage of HIV were associated with reduced CD4+ T cell count and increased HIV-1 viral load. CONCLUSION: Our study demonstrates that H. pylori co-infection was associated with higher CD4+ T cell count and lower HIV-1 viral load in HIV-positive patients, regardless of ART status. These findings show a positive effect of H. pylori co-infection on the dynamics of HIV-related immunological and virological parameters. Further studies are needed to elucidate the underlying mechanisms of the observed effects.

Prevalence and Antimicrobial Resistance of Campylobacter Species and Associated Factors Among Under-Five Children with Diarrhea at Randomly Selected Public Health Facilities in Mekelle, Tigray, Ethiopia.

Background: Campylobacter species are the most predominant bacterial agents to cause diarrhea in under-five children. It poses a serious challenge to public health worldwide with ongoing acquisition of resistance to different antimicrobials with multiple patterns. Thus, this study aimed to determine the prevalence, and antimicrobial resistance of Campylobacter species, and associated factors among under-five children with diarrhea in selected public health facilities. Methods: A cross-sectional study was conducted among under-five children with diarrhea using convenient sampling. Health facilities were selected using a simple random sampling method. The stool samples collected from 214 study participants were transported and processed following standard microbiological protocols. Campylobacter isolates were identified using Gram staining, biochemical test, serological test, and aerobic growth at 25°C. Antimicrobial susceptibility profiles of isolates were performed using the Kirby-Bauer method. Data were analyzed using SPSS ver. 25.0. Association between variables was assessed using Chi-square test and Logistic regression, with P ≤ 0.05. Results: The subject's mean age was 31.3 (±3.9) months. Of the 214 samples cultured, 14 (6.5%) of them were positive for Campylobacter species with 95% CI (3.3-10.3). Out of the isolated species, 12 (85.7%) were Campylobacter jejuni /Campylobacter coli and 2 (14.3%) were other Campylobacter species. Bottle feeding and history of direct contact to domestic animals were associated with Campylobacter species (AOR=5.13, CI=1.21-21.6, p=0.026 and AOR=4.93, CI=1.33-18.17, P=0.016), respectively. Campylobacter isolates were highly resistant to ciprofloxacin 5 (35.7%), and tetracycline 3 (21.4%). Conclusion: A higher incidence of Campylobacter species was obtained in children who were bottle-fed and who had a history of direct contact with domestic animals. The isolates were highly resistant to ciprofloxacin and tetracycline. These findings indicate that special attention is needed for better management of Campylobacter drug resistance in under-five children. To enhance and support our current findings, further research using molecular techniques is needed to identify the resistant and virulent genes of the bacterial isolates.

Shigella virulence protein VirG is a broadly protective antigen and vaccine candidate.

Diarrhea caused by Shigella has been associated with high morbidity and mortality in young children worldwide. There are no licensed vaccines, and those clinically advanced have restricted coverage as they elicit serotype-specific immunity while disease is caused by multiple circulating serotypes. Our group had previously reported a close association between serum antibodies to the Shigella virulence factor VirG (or IcsA) and clinical protection in infected individuals. VirG is highly conserved among Shigella strains and appealing as a broad-spectrum vaccine candidate. In this study, we investigated the immunogenicity and protective capacity of VirG as a subunit vaccine in mice. The surface-exposed alpha (α) domain of VirG (VirGα) was produced as a recombinant protein. This region has almost identical immune reactivity to full-length VirG. Administered intramuscularly with alum, VirGα elicited robust immune responses and high protective efficacy against S. flexneri 2a and S. sonnei. Almost complete protection was afforded by VirGα given intranasally with the E. coli double mutant heat-labile toxin (dmLT). VirGα-specific antibodies recognized VirG expressed on live Shigella, and blocked Shigella adhesion and invasion to human colonic cells. These results show for the first time that VirGα is a promising cross-protective vaccine candidate to prevent Shigella infection.

A Novel Shigella O-Polysaccharide-IpaB Conjugate Vaccine Elicits Robust Antibody Responses and Confers Protection against Multiple Shigella Serotypes.

Shigella is responsible for high burdens of diarrhea and dysentery globally. Children living in areas of endemicity are the most affected, and currently, there are no licensed vaccines to prevent shigellosis. Vaccine approaches have traditionally targeted the bacterial lipopolysaccharide as a protective antigen. Shigella O-polysaccharide (OPS) conjugated to recombinant Pseudomonas aeruginosa exotoxin A (rEPA) or tetanus toxoid (TT) is advanced in clinical evaluation. Adequate efficacy of these vaccines, particularly in the infant target group, remains to be demonstrated. A major limitation of the OPS-glycoconjugate concept is its limited coverage, since immunity to the O antigen is serotype specific, and there are multiple disease-causing serotypes. Another concern is the use of protein carriers already included in multiple other childhood vaccines. This study reports a novel Shigella OPS conjugate vaccine that uses the Shigella invasion plasmid antigen B (IpaB) as the carrier protein. IpaB is a virulence factor component of the Shigella type III secretion system and highly conserved among Shigella serotypes. It is robustly immunogenic and a protective antigen. IpaB and IpaB containing nonnative amino acids (nnAA) were produced at large scale using cell-free protein synthesis. Incorporation of nnAA enabled site-specific conjugation of IpaB to Shigella flexneri 2a OPS using click chemistry, yielding OPS-IpaB glycoconjugate. Parenteral immunization of mice with the OPS-IpaB vaccine resulted in high levels of OPS- and IpaB-specific serum IgG and robust protection against lethal S. flexneri 2a or Shigella sonnei challenge. The OPS-IpaB vaccine is a promising new vaccine candidate with the capacity to confer broad protection against clinically relevant Shigella serotypes. IMPORTANCE Diarrhea caused by Shigella species results in long-term disability and mortality globally, disproportionally affecting younger children living in poor countries. Although it is treatable by antibiotics, the rapid and widespread emergence of resistant strains and the highly contagious nature of the disease compel the development of preventive tools. Currently, several Shigella OPS conjugate vaccines are being evaluated in clinical studies, but these rely exclusively on immunity against the bacterial O antigen, which limits their coverage to only the immunizing serotype; multivalent vaccines are needed to protect against the most prevalent serotypes. This is the first report of a novel Shigella OPS-conjugate vaccine that uses Shigella IpaB as a carrier and protective antigen. This vaccine, administered parenterally, elicited robust immunity and protected mice against lethal infection by S. flexneri 2a or S. sonnei. The OPS-IpaB vaccine is a promising candidate for evaluation in vulnerable populations.

Efficient production of immunologically active Shigella invasion plasmid antigens IpaB and IpaH using a cell-free expression system.

Shigella spp. invade the colonic epithelium and cause bacillary dysentery in humans. Individuals living in areas that lack access to clean water and sanitation are the most affected. Even though infection can be treated with antibiotics, Shigella antimicrobial drug resistance complicates clinical management. Despite decades of effort, there are no licensed vaccines to prevent shigellosis. The highly conserved invasion plasmid antigens (Ipa), which are components of the Shigella type III secretion system, participate in bacterial epithelial cell invasion and have been pursued as vaccine targets. However, expression and purification of these proteins in conventional cell-based systems have been challenging due to solubility issues and extremely low recovery yields. These difficulties have impeded manufacturing and clinical advancement. In this study, we describe a new method to express Ipa proteins using the Xpress+TM cell-free protein synthesis (CFPS) platform. Both IpaB and the C-terminal domain of IpaH1.4 (IpaH-CTD) were efficiently produced with this technology at yields > 200 mg/L. Furthermore, the expression was linearly scaled in a bioreactor under controlled conditions, and proteins were successfully purified using multimode column chromatography to > 95% purity as determined by SDS-PAGE. Biophysical characterization of the cell-free synthetized IpaB and IpaH-CTD using SEC-MALS analysis showed well-defined oligomeric states of the proteins in solution. Functional analysis revealed similar immunoreactivity as compared to antigens purified from E. coli. These results demonstrate the efficiency of CFPS for Shigella protein production; the practicality and scalability of this method will facilitate production of antigens for Shigella vaccine development and immunological analysis. KEY POINTS : • First report of Shigella IpaB and IpaH produced at high purity and yield using CFPS • CFPS-IpaB and IpaH perform similarly to E. coli-produced proteins in immunoassays • CFPS-IpaB and IpaH react with Shigella-specific human antibodies and are immunogenic in mice.

Effect of pregnancy and HIV infection on detection of latent TB infection by Tuberculin Skin Test and QuantiFERON-TB Gold In-Tube assay among women living in a high TB and HIV burden setting.

OBJECTIVE: To investigate the effect of pregnancy and Human immunodeficiency virus (HIV) infection on detection performances of tuberculin skin test (TST) and QuantiFERON-TB Gold In-Tube (QFTGIT) for the diagnosis of latent tuberculosis infection (LTBI) among women living in high TB and HIV endemic setting. METHOD: A cross-sectional study was conducted among women with and without pregnancy and HIV infection. Three-hundred twenty women were enrolled in this study and were diagnosed using TST and QFTGIT for the detection of LTBI. RESULTS: Overall prevalence of LTBI among the enrolled women was 55.6%, 46.3% and 51.1% as determined by TST, QFTGIT and concordant TST/QFTGIT results, respectively. Our study revealed that pregnancy or HIV infection reduced the rate of detection of LTBI by TST and QFTGIT tests, with the utmost effect observed in HIV-positive pregnant women. Additionally, we observed that the concordance between TST and QFTGIT among women increased with the presence of pregnancy and/or HIV infection. A history of contact with TB patients was significantly associated with positivity of TST and QFTGIT. CONCLUSION: This study demonstrated that both pregnancy and HIV infection profoundly affected the detection performance of TST and QFTGIT, which may be associated with immunosuppression of anti-mycobacterial immunity in women with pregnancy and/or HIV infection.

Pregnancy suppresses Mycobacterium tuberculosis-specific Th1, but not Th2, cell-mediated functional immune responses during HIV/latent TB co-infection.

Immunity to tuberculosis (TB) is suppressed due to HIV coinfection and this suppression could further be enhanced by pregnancy. However, the effect of pregnancy on Mycobacterium tuberculosis (M. tuberculosis)-specific immune response during HIV/latent TB co-infection is not well understood. Here we investigated the changes in M. tuberculosis-specific functional Th1, Th2 and antibody responses in pregnant women with HIV/latent TB co-Infection. Pregnancy, concurrent with HIV infection, triggers a substantial suppression of M. tuberculosis-specific IFN-γ responses in a CD4+ T cell count dependent manner with an insignificant change in IL-4 and IgG responses. Conversely, M. tuberculosis-specific IL-10 production was markedly augmented in latent TB infected pregnant women with a lesser extent during HIV co-infection. These findings reveal that pregnancy suppresses anti-mycobacterial protective immune response in a CD4+ T cell count dependent manner during HIV/latent TB co-infection, suggesting a higher risk of developing active TB during pregnancy as a result of failing to control TB infection.

Enhanced IFN-γ, but not IL-2, response to Mycobacterium tuberculosis antigens in HIV/latent TB co-infected patients on long-term HAART.

BACKGROUND: HIV-infected individuals with latent TB infection are at increased risk of developing active TB. HAART greatly reduces the incidence rate of TB in HIV-infected patients and reconstitutes Mycobacterium tuberculosis (M. tuberculosis)-specific immune response in the first 12 months of therapy. The durability of the anti-mycobacterial immune restoration after a year of HAART however remains less investigated. METHOD: A cross-sectional study was conducted to evaluate M. tuberculosis-specific functional immune responses in HIV/latent TB co-infected patients who were on HAART for at least 1.5 up to 9 years as compared to HAART-naïve patients. Three-hundred sixteen HIV-infected patients without active TB were screened by tuberculin skin testing for M. tuberculosis infection and peripheral blood mononuclear cells (PBMCs) were isolated from 61 HIV/latent TB co-infected patients (30 HAART-naïve and 31 HAART-treated). IFN-γ and IL-2 ELISPOT as well as CFSE cell proliferation assays were performed after stimulation with M. tuberculosis antigens PPD and ESAT-6. RESULT: The median frequency of PPD and ESAT-6 specific IFN-γ secreting cells was significantly higher in the HAART-treated patients as compared to HAART-naïve patients, p = 0.0021 and p = 0.0081 respectively. However, there was no significant difference in the median frequency of IL-2 secreting cells responding to PPD (p = 0.5981) and ESAT-6 (p = 0.3943) antigens between HAART-naïve and-treated groups. Both IFN-γ and IL-2 responses were independent of CD4+ T cell count regardless of the HAART status. Notably, the frequency of PPD and ESAT-6 specific IL-2 secreting cells was positively associated with CD4+ T cell proliferation while inversely correlated with duration of HAART, raising the possibility that M. tuberculosis-specific IL-2 response that promote the antigen-specific CD4+ T cell proliferation diminish with time on antiretroviral therapy in HIV/latent TB co-infected patients. CONCLUSION: This study shows an increased M. tuberculosis-specific IFN-γ, but not IL-2, response in HIV/latent TB co-infected patients with long-term HAART, consistent with only partial immune restoration. Future studies should, therefore, be done to prospectively define the rate and extent to which functional immune responses to M. tuberculosis are restored after long-term HAART.

Inflammation triggers immediate rather than progressive changes in monocyte differentiation in the small intestine.

Bone marrow-derived circulating monocytes contribute to the replenishment and maintenance of the intestinal macrophage population. Intestinal monocytes undergo context-dependent phenotypic and functional adaptations to either maintain local immune balance or support intestinal inflammation. Here we use monocyte adoptive transfer to dissect the dynamics of monocyte-to-macrophage differentiation in normal and inflamed small intestine. We find that during homeostasis CCR2 and ß7-integrin mediate constitutive homing of monocytes to the gut. By contrast, intestinal inflammation increases monocyte recruitment via CCR2, but not ß7-integrin. In the non-inflamed intestine, monocytes gradually differentiate to express genes typically associated with tolerogenic macrophage functions. Conversely, immediately upon entry into the inflamed intestine, monocytes adapt a different expression pattern in a partly Trem-1-dependent manner. Our observations suggest that inflammation fundamentally changes the kinetics and modalities of monocyte differentiation in tissues.

The effect of maternal helminth infection on maternal and neonatal immune function and immunity to tuberculosis.

BACKGROUND: M. tuberculosis and helminth infection each affects one third of the world population. Helminth infections down regulate cell mediated immune responses and this may contribute to lower efficacy of BCG vaccination and higher prevalence of tuberculosis. OBJECTIVE: To determine the effect of maternal helminth infection on maternal and neonatal immune function and immunity to TB. METHODS: In this cross sectional study, eighty five pregnant women were screened for parasitic and latent TB infections using Kato-Katz and QFT-GIT tests, respectively. IFN-γ and IL-4 ELISpot on Cord blood Mononuclear Cells, and total IgE and TB specific IgG ELISA on cord blood plasma was performed to investigate the possible effect of maternal helminth and/or latent TB co-infection on maternal and neonatal immune function and immunity to TB. RESULT: The prevalence of helminth infections in pregnant women was 27% (n = 23), with Schistosoma mansoni the most common helminth species observed (20% of women were infected). Among the total of 85 study participants 25.8% were QFT-GIT positive and 17% had an indeterminate result. The mean total IgE value of cord blood was significantly higher in helminth positive than negative women (0.76 vs 0.47, p = 0.042). Cross placental transfer of TB specific IgG was significantly higher in helminth positive (21.9 ± 7.9) than negative (12.3 ± 5.1), p = 0.002) Latent TB Infection positive participants. The IFN-γ response of CBMCs to ESAT-6/CFP-10 cocktail (50 vs 116, p = 0.018) and PPD (58 vs 123, p = 0.02) was significantly lower in helminth positive than negative participants. There was no significant difference in IL-4 response of CBMCs between helminth negative and positive participants. CONCLUSIONS: Maternal helminth infection had a significant association with the IFN-γ response of CBMCs, total IgE and cross placental transfer of TB specific IgG. Therefore, further studies should be conducted to determine the effect of these factors on neonatal immune response to BCG vaccination.