Effects of Helicobacter pylori infection on T cell activation markers and regulatory T cells in people with and without HIV infection in Central Ethiopia

Effects of Helicobacter pylori infection on T cell activation markers and regulatory T cells in people with and without HIV infection in Central Ethiopia | Research Square window.SnipcartSettings = { analytics: { enabled: false } }; (function() { var accessVector = localStorage.getItem('access_vector') || ''; window.dataLayer = window.dataLayer || []; if (accessVector) { window.dataLayer.push({ user: { profile: { profileInfo: { snid: accessVector } } } }); } })(); (function(w,d,s,l,i){w[l]=w[l]||[];w[l].push({'gtm.start':new Date().getTime(),event:'gtm.js'});var f=d.getElementsByTagName(s)[0],j=d.createElement(s),dl=l!='dataLayer'?'&l='+l:'';j.async=true;j.src='https://www.googletagmanager.com/gtm.js?id='+i+dl;f.parentNode.insertBefore(j,f);})(window,document,'script','dataLayer','GTM-K279D39R'); Browse Preprints In Review Journals COVID-19 Preprints AJE Video Bytes Research Tools Research Promotion AJE Professional Editing AJE Rubriq About Preprint Platform In Review Editorial Policies Our Team Advisory Board Help Center Sign In Submit a Preprint Cite Share Download PDF Research Article Effects of Helicobacter pylori infection on T cell activation markers and regulatory T cells in people with and without HIV infection in Central Ethiopia Smaranda Gliga, Million Getechew Mesfun, Tafese Beyene Tufa, Andre Fuchs, and 5 more This is a preprint; it has not been peer reviewed by a journal. https://doi.org/ 10.21203/rs.3.rs-7058889/v1 This work is licensed under a CC BY 4.0 License Status: Published Journal Publication published 11 Feb, 2026 Read the published version in European Journal of Medical Research → Version 1 posted 12 You are reading this latest preprint version Abstract Background: Helicobacter pylori (H. pylori) is known to modulate host immunity and sustain chronic inflammation, yet most data come from HIV-negative populations. In people living with HIV, whose T cell compartments are already dysregulated, the way H. pylori shapes peripheral T cell phenotypes, and how those profiles change after eradication therapy, is still unclear. Because both infections are common in Central Ethiopia, we examined peripheral T cell phenotypes in adults with and without HIV according to H. pylori status and assessed the immunologic effects of antibiotic eradication. Materials and Methods We conducted a prospective study in people with and without HIV infection from Ethiopia. H. pylori status was determined by stool-antigen testing; a subset received standard triple therapy and was followed for 12 months. Multiparameter flow cytometry quantified T cell activation, proliferation, exhaustion, and regulatory T cells (T regs ) at baseline and after therapy. Results T cell analyses showed that participants with HIV had consistently higher proliferation (Ki67), exhaustion (PD-1, TIM3), and Th17 (CCR6⁺CD161⁺) markers than those without HIV. H. pylori -positive individuals exhibited higher T reg levels irrespective of HIV status (HIV-negative: median 2% vs 1.08%, p < 0.0001; HIV-positive: median 2.9% vs 1.62%, p = 0.009). Eradication therapy led to a significant reduction in T regs in both HIV-positive (SD 1.98%, p = 0.014) and HIV-negative (SD 1.5%, p = 0.023) groups. Conclusions H. pylori infection is associated with specific alterations in T cell profiles, in both HIV negative and positive populations. Eradication therapy, irrespective of success, reduces T regs and partially restores T cell function, offering insight into potential therapeutic strategies for managing immune dysregulation in co-infected populations. Trial registration Not applicable. This study was not registered in a clinical trial registry, as there was no focus on investigational treatment. The aspects of the study reported here were observational in character. Ethical approval was obtained from both Ethiopian and German ethic committees. The study adhered to the Declaration of Helsinki and ICHGCP guidelines. T cell exhaustion T cell proliferation Helicobacter pylori eradication therapy Immune modulation HIV Figures Figure 1 Figure 2 Figure 3 Figure 4 Background Helicobacter pylori (H. pylori) is a microaerophilic bacterium that establishes lifelong residence in the gastric epithelium after infection, which usually occurs in childhood [ 1 ]. Although most carriers remain symptom-free, a proportion progress to dyspepsia and other gastroduodenal disorders [ 2 ]. Early during colonization, innate sensing sparks the release of pro-inflammatory cytokines which in turn drive a strong T helper (Th) 1 and Th17 skewed T-cell response [ 3 , 4 ]. To prevent excessive tissue damage, the mucosa simultaneously produces counter-regulatory mediators such as transforming growth factor (TGF)-β and IL-10 [ 5 ]. TGF-β recruits T cells with anti-inflammatory activity, in particular regulatory T cells (T regs ). T regs derive from the same lineage as naïve CD4 + T cells and present the transcription factor forkhead box protein 3 (Foxp3) and the activating marker CD25 [ 6 ]. It was shown that a greater population of T regs is present in the gastric mucosa and in the peripheral blood of H. pylori positive individuals, while T regs are largely absent in H. pylori negative subjects [ 7 , 8 ]. Although T regs are activated to prevent hyperinflammatory conditions, they can also facilitate bacterial persistence by suppressing IFN-γ-producing effector T cells [ 9 ]. Consistent with this notion, experimental depletion of T regs with anti-CD25 antibodies in mice intensifies gastritis while reducing the gastric bacterial load, highlighting the delicate balance between protection and pathology [ 10 ]. Given that a significant proportion of the 39 million people living with HIV (PLWH) [ 11 ] worldwide reside in low- and middle-income countries, where H. pylori infection is highly prevalent, it is likely that many of these individuals also have H. pylori co-infection. HIV infection is characterized by a reduced CD4 + T cell count, which in time favors opportunistic infections, both viral and bacterial. While some publications show a high prevalence of H. pylori in PLWH versus those without HIV [ 12 ], most report a lower prevalence of H. pylori infection in PLWH, especially in patients with CD4 + T cells below 200/µL [ 13 – 16 ]. Furthermore, the introduction of combination antiretroviral therapy (cART) induced an increase in dyspeptic complications and their severity [ 17 ]. HIV infection is associated with a strong recruitment of Th2 cells and reduction of Th1/Th17cells, which in turn decrease the pro-inflammatory response against H. pylori and the intensity of gastritis [ 18 ]. A previous study our group performed in Ghana showed that H. pylori carriage coincided with reduced CD4⁺ T cell activation (HLA-DR⁺CD38⁺), proliferation (Ki67) and PD-1 expression in both ART-naïve PLWH and HIV-negative controls [ 15 ]. These results parallel other reports in which co-infected, treatment-naïve patients displayed comparatively higher CD4⁺ counts and diminished HIV-1 viral loads, supporting the notion that the bacterium can dampen systemic immune activation [ 19 , 20 ]. To date, however, no study has characterized these immunological interactions in Ethiopian cohorts, nor has any work prospectively examined how eliminating H. pylori affects T cell phenotypes in this setting. Accordingly, we designed the present prospective, randomized study to elucidate the mechanisms of systemic immune modulation linked to H. pylori infection by tracking activation, proliferation, and exhaustion signatures before and after standard eradication therapy. Materials and methods Study population Between March and June 2017, we enrolled adults attending the Asella Teaching and Referral Hospital (ATRH, Ethiopia). Two streams were used: (i) successive visitors to the HIV outpatient service and (ii) successive clients of the hospital’s voluntary counselling-and-testing unit. All recruits underwent H. pylori screening with a stool-antigen assay (see below). Eligibility criteria were 18–55 years of age, signed informed consent and for PLWH, a CD4⁺ cell count above 350 cells/µl. From the first 140 HIV-negative and 140 HIV-positive individuals who satisfied these criteria we applied additional exclusion filters: prior immunosuppression, malignancy, chronic viral hepatitis (B and C), tuberculosis, elevated C-reactive protein, parasitic infection, recent anti-helminthic/antibiotic therapy, upper-gastrointestinal symptoms, recent acute infection, hemoglobin < 10 g/dl or pregnancy, as described earlier ¹⁵. All PLWH were on cART. Ethics The protocol received approval in Ethiopia and Germany (refs A/U/H/S/C/87/6392; 3.10/271/2017; 5728). Eradication therapy Within the H. pylori –positive group, 26 / 123 HIV-negative individuals and 25 / 121 PLWH were randomized to receive a 14-day triple regimen: metronidazole 500 mg, clarithromycin 500 mg and pantoprazole 40 mg, each taken twice daily. This macrolide- and nitroimidazole-based schedule follows Ethiopian guidelines and deliberately avoids penicillin to prevent reactions in participants with undocumented β-lactam allergy, as previously mentioned [ 21 ]. Clinical review with paired blood and stool sampling was scheduled at baseline and at three, six and 12 months. Successful eradication was defined as a negative stool-antigen test at the three-month visit. H. pylori stool antigen test Antigen detection employed the GA Generic Assays kit (Blankenfelde, Germany) according to the manufacturer’s protocol. Stool microscopy with direct and formol-ether concentration techniques ruled out concomitant parasitic infection. Blood sample collection and laboratory analysis At every visit 15 ml of EDTA-anticoagulated blood and 5 ml of serum were collected. Serum at baseline was analyzed for hepatitis B surface antigen (HBsAg), anti-hepatitis C (HCV) antibodies (InTec Products Inc., China) and CRP (NADAL® CRP, nal von minden, Germany). Complete blood counts and, for PLWH, CD4⁺ T cell enumerations were performed locally on a FACSCalibur™ flow cytometer (BD Biosciences, NJ, USA). PBMC isolation and cryopreservation Peripheral blood mononuclear cells (PBMCs) were isolated by Ficoll-Hypaque density gradient, suspended in 20% dimethyl sulfoxide (DMSO)/ fetal calf serum (FCS) freezing medium, cooled at − 80°C for 24 h in a controlled-rate container and shipped on dry ice to Düsseldorf for batched analysis. Flow-cytometric assessment To evaluate how H. pylori clearance altered T cell characteristics, phenotypic read-outs recorded after therapy were compared with each participant’s own baseline. Cryopreserved PBMCs were rapidly thawed at 37°C, washed twice and resuspended in phosphate-buffered saline (PBS). Surface labelling was performed at 2–8°C in the dark, whereas intracellular markers were stained at room temperature. Antibody details are listed in Supplementary Table 1; four dedicated panels covered (1) activation, (2) exhaustion, (3) regulatory T cells and (4) Th17 signatures. All reagents were sourced from eBioscience (Frankfurt am Main, Germany). Viable cells were gated with a fixable viability dye. After surface staining, samples in panels 2 and 4 were held in flow-cytometry buffer for immediate acquisition, while those in panels 1 and 3 underwent fixation, permeabilization and 30-min intracellular staining before a final wash into buffer. Data were collected on a three-laser BD FACSCanto II. Spectral overlap was corrected with UltraComp beads (eBioscience, Germany), singularly stained for each fluorochrome. Files were processed in FlowJo v10.1 (Tree Star, OR, USA); the gating strategy appears in Supplementary Fig. 1. Phenotypic outputs were stratified by HIV status and by H. pylori infection, as well as by pre- versus post-eradication time points. Statistical Analysis Data from participants without and with HIV were processed independently with Prism version 9.0 (GraphPad Software, San Diego, California, USA) software. Normally distributed variables were compared by paired or unpaired t tests; results are given as mean ± standard deviation (SD) or median values. Skewed data were evaluated using the two-tailed Mann-Whitney test. Significance codes: n.s.=not significant; * p < 0.05; ** p < 0.01; *** p < 0.001; **** p < 0.0001, unless stated otherwise in the figure and text. Results Cohort A cohort of 306 people living with HIV and 201 people without HIV underwent screening to detect H. pylori colonization at enrolment. From these, we pragmatically selected 140 individuals in the HIV-positive cohort (78 colonized, 62 non-colonized) and an equal 140 individuals in the HIV-negative cohort (93 colonized, 47 non-colonized) for detailed evaluation of exclusion criteria, including co-infections, inflammation, medication, or pregnancy history. Subsequently, 27 H. pylori positive volunteers from each cohort were randomly allocated to a 14-day metronidazole–clarithromycin–pantoprazole regimen, constituting the study’s standard triple eradication intervention arm for bacterial clearance. After testing for confounding factors, two participants from the group with HIV and one participant from those without HIV infection were excluded from analysis as predefined. The rates of successful eradication in participants with HIV and those without HIV were previously reported [ 21 ]. Comparison of T cell profile in participants without vs. with HIV at baseline In participants with HIV, CD4 + T cells exhibited consistently higher levels of Ki67 (median 96% vs 88%, p < 0.0001) as proliferation markers, of PD-1 (median 19% vs 14%, p = 0.0003) and TIM3 (median 3.4% vs 1.6%, p < 0.0001) as exhaustion markers, and of Th17 Cells (CCR6 + CD161 + ; median 0.64% vs 0.3%, p < 0.0001) (Fig. 1 A). In addition, CD8 + T cells had higher levels of Ki67 (median 7.2% vs 5.6%, p = 0.002), PD-1 (median 0.84% vs 0.66%, p = 0.046) and TIM3 (median 7.2% vs 3.4%, p < 0.0001). (Fig. 1 B). T Cell Profile in Participants without HIV according to H. pylori status at baseline In the group of participants without HIV (n = 123 after exclusion of 17 participants with confounding factors), T cell markers were analyzed comparatively between subgroups with (n = 83) and without (n = 40) H. pylori infection. Results for CD4 + and CD8 + T cells are presented in Fig. 2 . The key findings of this analysis were that participants with H. pylori infection showed significantly lower percentages of activation markers (HLA-DR + CD38 + ) on CD4 + T cells compared to uninfected individuals (median 1.52% vs 2.24%, p = 0.003). Furthermore, they showed lower levels of T cell exhaustion markers (PD-1, TIM3) in CD4 + (PD-1: median 12.7% vs 16.05%, p = 0.03; TIM3: median 1.28% vs 2.39%, p = 0.002) and CD8 + T cells (PD-1: median 0.49% vs 1.12%, p = 0.001; TIM3: median 3.13% vs 5.61%, p = 0.007) and significantly higher levels of T regs (CD25 + Foxp3 + CD4 + ; median 2% vs 1.08%, p < 0.0001), highlighting their critical role in maintaining immune homeostasis by suppressing excessive immune activation. T Cell Profile in participants with HIV infection according to H. pylori status at baseline Among the 121 people living with HIV, 56 tested negative and 65 positive for H. pylori . Dual-infected individuals exhibited a higher proportion of regulatory T cells (median 2.9% vs 1.62%, p = 0.009), highlighting the contribution of T regs in moderating immune activation during coinfection (Fig. 3 ). In contrast, markers of T cell activation and exhaustion showed no significant variation between the two subgroups. Effect of H. pylori Eradication Therapy on T Cell Populations Of the H. pylori participants with HIV-infection (n = 25) and without HIV infection (n = 26) who were randomized to eradication therapy, 15/24 participants without HIV (62.5%) and 9/24 with HIV (37.5%) presented for the follow-up investigation three months after eradication therapy and were successfully eradicated (as confirmed by a negative H. pylori stool antigen test) and thus eligible for final analysis (for details see the previous publication by Mesfun et al ) [ 21 ]. In participants without HIV infection, successful H. pylori eradication resulted in increased proliferation marker expression (Ki67) in CD4 + T cells (n = 14; SD 27.4%, p = 0.049). CD8 + T cells showed an increase in exhaustion markers (CD57) (SD 17.05%, p = 0.0027; Fig. 4 A). We observed a decrease in T regs in HIV-negative participants after successful (SD 1.5%, p = 0.023), but also after unsuccessful (SD 0.72%, p = 0.0004; Fig. 4 C) H. pylori eradication therapy. Among PLWH, seven individuals who cleared H. pylori showed a rise in CD8⁺ T cell activation (HLA-DR⁺CD38⁺; SD 0.39%, p = 0.016) accompanied by a fall in Ki67 proliferation signals (SD 3.1%, p = 0.012; Fig. 4 B) and a parallel contraction of T regs (SD 1.98%, p = 0.014). Conversely, in those whose therapy failed we detected reduced Ki-67 on CD8⁺ T lymphocytes (SD 13.3%, p = 0.012), lower PD-1 on CD4⁺ (SD 8.43%, p = 0.0009) and CD8⁺ T cells (SD 1.05%, p = 0.01), together with elevated CD57 on CD4⁺ T cells (SD 10.14%, p = 0.04; Fig. 4 D). These shifts collectively suggest differential immune recalibration following attempted bacterial clearance in adults. Discussion Because few studies from sub-Saharan Africa have explored how H. pylori shapes immune responses, this study aimed to fill this critical gap by prospectively examining H. pylori –related effects on T cell populations, including markers of activation, exhaustion and T regs , in individuals with and without HIV infection. The rationale for such an investigation stems from the potential immune-modulating effects of H. pylori , which may be especially pertinent in the setting of HIV, where the immune system is already under strain. Our study revealed that T cell exhaustion markers, particularly PD-1, were consistently elevated in individuals with HIV, an observation aligning with well-established data on chronic immune dysregulation in HIV infection [ 22 , 23 ]. Moreover, despite limited regional data on H. pylori immune interactions in Sub-Saharan Africa, we confirmed that in this cohort, HIV infection correlated with characteristic shifts in T cell activation, proliferative activity, and the distribution of regulatory subsets. Building on these findings, we also evaluated Th17 cells, which tend to be depleted in untreated HIV but can be reconstituted under cART. Here, we noted persistently elevated Th17 markers in the HIV positive group, likely reflecting the relatively preserved CD4⁺ T cell counts afforded by ART. This backdrop provides important context for interpreting the role of H. pylori , a bacterium often linked to robust Th17 responses in gastric lesions [ 24 – 26 ]. In our cohort, however, H. pylori status did not appear to further alter Th17 frequency in people with HIV, indicating that cART-driven immune restoration may outweigh any additive influence from H. pylori . Studies examining H. pylori -driven changes in peripheral T cell activation, proliferation, and exhaustion remain limited. In our earlier work, colonization corresponded to diminished CD4⁺ T cell activation (HLA-DR⁺CD38⁺) and lower expression of exhaustion markers, PD-1 on CD8⁺ cells and TIM-3 on both CD4⁺ and CD8⁺ subsets [ 15 ]. A different group, however, documented increased HLA-DR expression in PBMCs, especially in individuals with peptic ulcers, implying that H. pylori could have variable immunomodulatory effects depending on clinical presentation [ 27 ]. Other contrasting results come from murine and human studies [ 28 , 29 ] describing elevated TIM3 expression in Th cells from H. pylori ‐infected hosts, and higher exhaustion levels on CD8⁺ T cells in gastric cancer patients [ 29 ]. Such inconsistencies may reflect differences in host genetics, regional factors, sample size or disease phenotypes (e.g., asymptomatic carriers vs. peptic ulcer disease vs. gastric cancer). Our exclusive enrollment of asymptomatic patients could partly explain why we observed lower levels of certain exhaustion or activation markers relative to other findings. A significant outcome of our investigation was the notably higher percentage of T regs among H. pylori -infected participants, regardless of HIV status, paralleling observations from histopathological studies of the gastric mucosa [ 7 – 9 , 24 , 30 ]. While most prior publications focused on local T regs in gastric tissue, only two reports have shown elevated circulating T regs in H. pylori ‐infected cohorts [ 8 , 24 ]. Our own earlier work hinted at a similar peripheral trend, suggesting that H. pylori –induced immune regulation may be systemic rather than confined solely to the gastric microenvironment [ 15 ]. Prior work by Eberhardt and colleagues [ 15 ] linked H. pylori infection to reduced T cell activation, proliferation and exhaustion markers, but could not definitively establish causality. Accordingly, we sought to determine whether eradicating H. pylori would lead to an immunological reversal of these trends. However, high levels of metronidazole resistance [ 31 ] in Ethiopia resulted in a low overall eradication rate, leaving only small subgroups available for a detailed pre- vs. post‐therapy comparison. Recognizing that H. pylori typically recruits T regs to facilitate its persistence [ 30 ], we hypothesized that T reg levels would drop if H. pylori were successfully eliminated. Indeed, we observed precisely that: a notable decline in T regs post‐eradication, regardless of HIV status. Yet, a parallel reduction occurred even in some participants without HIV who did not clear their infection, which we attribute to antibiotic‐induced dysbiosis [ 32 , 33 ]. This broader disruption of the microbiota may have secondary effects on T regs and Th17 subsets, mirroring data from other pathologies where antibiotic therapy modifies immune populations [ 34 ]. Interestingly, participants with HIV who were successfully eradicated showed increased activation of CD8⁺ T cells. We surmise that reduced T reg -mediated suppression could permit cytotoxic T lymphocytes to become more activated [ 35 ]. Unexpectedly, those who failed eradication still exhibited a decrease in T regs alongside reduced proliferation and PD‐1 expression, hinting at a complex interplay between T regs and PD‐1/PD‐L1 pathways [ 36 ]. These findings complicate a straightforward narrative of T reg modulation, pointing to additional factors, such as partial antibiotic effects, subclinical shifts in gut flora or baseline immune status, that may influence immune cell phenotypes regardless of eradication outcome. Although we document significant immunological shifts following H. pylori eradication, the limited sample size and suboptimal eradication success rate constrain our ability to draw broad conclusions. Larger-scale studies using alternative antibiotic regimens or susceptibility-guided therapy are needed to confirm these findings and clarify the complex interplay among H. pylori , HIV status, antibiotic resistance and host genetics. Moreover, while antibiotics may affect T regs and Th17 cells, we did not systematically evaluate broader gut microbiome shifts, which can transiently or persistently alter immune phenotypes and confound outcomes [ 37 , 38 ]. Future investigations should integrate microbiome analyses (e.g., 16S rRNA or metagenomic approaches) to distinguish the effects of H. pylori eradication from those of collateral disruptions in gut flora. Finally, despite observing immunological changes associated with H. pylori infection, the long-term consequences for HIV progression remain unclear. Some research suggests that chronic coinfections can modulate systemic immune activation in people with HIV, possibly influencing viral reservoirs or disease progression [ 39 , 40 ]. Whether H. pylori is ultimately protective, neutral or detrimental may depend on bacterial strain diversity, host genetics, cART status, and local factors. Further exploration of these variables is needed to unravel the multifaceted relationship between H. pylori and HIV. Conclusions This study underscores the importance of T regs in modulating immune responses during H. pylori infection, regardless of HIV status. T reg levels were consistently elevated in infected individuals, potentially aiding bacterial persistence, and successful H. pylori eradication led to a significant drop in T regs alongside improved immune activation and increased exhaustion. Although these findings suggest potential therapeutic benefits by modulating T regs , the low eradication rate and limited sample size limit broader conclusions. Larger studies are needed to confirm these results, clarify clinical applications, and investigate whether targeting T regs could benefit individuals with coinfections. Notably, although adoptive T reg therapy has reached Phase I/II trials in type 1 diabetes [ 41 ], no large trials have specifically addressed T regs in the context of coinfections. Abbreviations ATRH Asella Teaching and Referral Hospital cART Combination antiretroviral therapy CRP C-reactive protein DMSO Dimethyl sulfoxide FCS Fetal calf serum Foxp3 Forkhead box protein 3 H. pylori Helicobacter pylori n.s. Not significant PBMC Peripheral blood mononuclear cell PBS Phosphate-buffered saline PLWH People living with HIV SD Standard deviation TGF-β Transforming growth factor-β Th T helper cell Treg Regulatory T cell Declarations Conflicts of interest The authors have no financial or non-financial conflicts of interest to disclose in connection with this work. Funding Declaration This research received no external funding . Acknowledgments SG, TF, and MGM wrote and edited the manuscript. HMO, AF, TBT, EOK, and PAL designed the study protocol and recruited patients. MGM performed the flow cytometry experiments. TL provided critical scientific insight and oversight throughout the project. We also thank all staff and participants for their support. Data Availability The flow cytometry datasets generated and analysed during this study are not publicly archived to safeguard participant confidentiality, but they are available from the corresponding author on reasonable request. References Bücker R, Azevedo-Vethacke M, Groll C, Garten D, Josenhans C, Suerbaum S, et al. Helicobacter pylori colonization critically depends on postprandial gastric conditions. Sci Rep. 2012;2:994. Xie F-J. Helicobacter pylori infection and esophageal cancer risk: An updated meta-analysis. World J Gastroenterol. 2013;19:6098. Kivrak Salim D, Sahin M, Köksoy S, Adanir H, Süleymanlar I. Local Immune Response in Helicobacter pylori Infection. Medicine (Baltimore). 2016;95:e3713. Bamford KB, Fan X, Crowe SE, Leary JF, Gourley WK, Luthra GK, et al. Lymphocytes in the human gastric mucosa during Helicobacter pylori have a T helper cell 1 phenotype. Gastroenterology. 1998;114:482–92. Bornschein J, Kandulski A, Selgrad M, Malfertheiner P. From Gastric Inflammation to Gastric Cancer. Dig Dis. 2010;28:609–14. Curiel TJ. Tregs and rethinking cancer immunotherapy. J Clin Invest. 2007;117:1167–74. Lundgren A, Strömberg E, Sjöling Å, Lindholm C, Enarsson K, Edebo A, et al. Mucosal FOXP3 -Expressing CD4 + CD25 high Regulatory T Cells in Helicobacter pylori -Infected Patients. Infect Immun. 2005;73:523–31. Cook KW, Letley DP, Ingram RJM, Staples E, Skjoldmose H, Atherton JC, et al. CCL20/CCR6-mediated migration of regulatory T cells to the Helicobacter pylori -infected human gastric mucosa. Gut. 2014;63:1550–9. Kandulski A, Malfertheiner P, Wex T. Role of regulatory T-cells in H. pylori-induced gastritis and gastric cancer. Anticancer Res. 2010;30:1093–103. Rad R, Brenner L, Bauer S, Schwendy S, Layland L, Da Costa CP, et al. CD25+/Foxp3 + T Cells Regulate Gastric Inflammation and Helicobacter pylori Colonization In Vivo. Gastroenterology. 2006;131:525–37. UNAIDS/WHO estimates. Available from: https://www.who.int/images/default-source/departments/hiv/summary-of-the-global-hiv-epidemic-2022.png?sfvrsn=73ac5b6a_13 Abadi T, Teklu T, Wondmagegn T, Alem M, Desalegn G. Helicobacter pylori infection and associated risk factors among HIV-positive and HIV-negative individuals in Northern Ethiopia. J Infect Chemother. 2025;31:102517. Fialho AB, Braga-Neto MB, Guerra EJ, Fialho AM, Fernandes KC, Sun JL, et al. Low prevalence of H. pylori Infection in HIV-Positive Patients in the Northeast of Brazil. BMC Gastroenterol. 2011;11:13. Olmos M, Araya V, Pskorz E, Quesada EC, Concetti H, Perez H, et al. Coinfection: Helicobacter pylori/Human Immunodeficiency Virus. Dig Dis Sci. 2004;49:1836–9. Eberhardt KA, Sarfo FS, Dompreh A, Kuffour EO, Geldmacher C, Soltau M, et al. Helicobacter pylori Coinfection Is Associated With Decreased Markers of Immune Activation in ART-Naive HIV-Positive and in HIV-Negative Individuals in Ghana. Clin Infect Dis. 2015;61:1615–23. Cacciarelli AG, Marano BJ, Gualtieri NM, Zuretti AR, Torres RA, Starpoli AA, et al. Lower Helicobacter pylori infection and peptic ulcer disease prevalence in patients with AIDS and suppressed CD4 counts. Am J Gastroenterol. 1996;91:1783–4. Radovanović Spurnić A, Brmbolić B, Stojšić Z, Pekmezović T, Bukumirić Z, Korać M, et al. The increasing prevalence of HIV/ Helicobacter pylori co-infection over time, along with the evolution of antiretroviral therapy (ART). PeerJ. 2017;5:e3392. Krzyżek P, Gościniak G. Immunomodulatory influence of HIV and EBV on Helicobacter pylori infections – a review. Ann Parasitol. 2019;65:3–17. Abadi T, Teklu T, Wondmagegn T, Alem M, Desalegn G. 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. Eur J Med Res. 2024;29:178. Spurnic AR, Bukumiric Z, Jevtovic D, Brmbolic B, Pekmezovic T, Salemovic D, et al. Helicobacter pylori infection rates in dyspeptic Serbian HIV-infected patients compared to HIV-negative controls. PloS One. 2021;16:e0248041. Mesfun MG, Gliga S, Fuchs A, Orth HM, Schönfeld A, Luedde T, et al. Prevalence of H. pylori among asymptomatic HIV-positive and negative individuals in Central Ethiopia and efficacy of eradication therapy. IJID Reg. 2022;2:169–74. Trautmann L, Janbazian L, Chomont N, Said EA, Gimmig S, Bessette B, et al. Upregulation of PD-1 expression on HIV-specific CD8 + T cells leads to reversible immune dysfunction. Nat Med. 2006;12:1198–202. Breton G, Chomont N, Takata H, Fromentin R, Ahlers J, Filali-Mouhim A, et al. Programmed Death-1 Is a Marker for Abnormal Distribution of Naive/Memory T Cell Subsets in HIV-1 Infection. J Immunol. 2013;191:2194–204. Talayev V, Svetlova M, Zaichenko I, Voronina E, Babaykina O, Neumoina N, et al. CCR6 + T helper cells and regulatory T cells in the blood and gastric mucosa during Helicobacter pylori infection. Helicobacter. 2024;29:e13097. Pinchuk IV, Morris KT, Nofchissey RA, Earley RB, Wu J-Y, Ma TY, et al. Stromal cells induce Th17 during Helicobacter pylori infection and in the gastric tumor microenvironment. PloS One. 2013;8:e53798. Lal KG, Phuang-Ngern Y, Suhkumvittaya S, Leeansyah E, Alrubayyi A, Dias J, et al. Longitudinal Analysis of Peripheral and Colonic CD161 + CD4 + T Cell Dysfunction in Acute HIV-1 Infection and Effects of Early Treatment Initiation. Viruses. 2020;12:1426. Suleymanov Z. Expression of class I and II MHC receptors in Helicobacter pylori-positive patients with active gastritis and duodenal ulcer. Turk J Gastroenterol Off J Turk Soc Gastroenterol. 2003;14:168–72. Hu S, Xie Y, Zhou N, Jin L, Tan Y, Liu D, et al. Expression of T-cell immunoglobulin- and mucin-domain-containing molecules-1 and – 3 (Tim-1 and Tim-3) in Helicobacter pylori infection. Helicobacter. 2011;16:373–81. Shen P, Yue R, Tang J, Si H, Shen L, Guo C, et al. Preferential Tim-3 expression on Treg and CD8(+) T cells, supported by tumor-associated macrophages, is associated with worse prognosis in gastric cancer. Am J Transl Res. 2016;8:3419–28. Kandulski A, Wex T, Kuester D, Peitz U, Gebert I, Roessner A, et al. Naturally occurring regulatory T cells (CD4+, CD25high, FOXP3+) in the antrum and cardia are associated with higher H. pylori colonization and increased gene expression of TGF-beta1. Helicobacter. 2008;13:295–303. Asrat D, Kassa E, Mengistu Y, Nilsson I, Wadström T. Antimicrobial susceptibility pattern of Helicobacter pylori strains isolated from adult dyspeptic patients in Tikur Anbassa University Hospital, Addis Ababa, Ethiopia. Ethiop Med J. 2004;42:79–85. Pandiyan P, Bhaskaran N, Zou M, Schneider E, Jayaraman S, Huehn J. Microbiome Dependent Regulation of Tregs and Th17 Cells in Mucosa. Front Immunol. 2019;10:426. Yap TW-C, Gan H-M, Lee Y-P, Leow AH-R, Azmi AN, Francois F, et al. Helicobacter pylori Eradication Causes Perturbation of the Human Gut Microbiome in Young Adults. Badger JH, editor. PLOS ONE. 2016;11:e0151893. Rajendran M, Looney S, Singh N, Elashiry M, Meghil MM, El-Awady AR, et al. Systemic Antibiotic Therapy Reduces Circulating Inflammatory Dendritic Cells and Treg–Th17 Plasticity in Periodontitis. J Immunol. 2019;202:2690–9. Suvas S, Kumaraguru U, Pack CD, Lee S, Rouse BT. CD4 + CD25 + T cells regulate virus-specific primary and memory CD8 + T cell responses. J Exp Med. 2003;198:889–901. Nikolova M, Wiedemann A, Muhtarova M, Achkova D, Lacabaratz C, Lévy Y. Subset- and Antigen-Specific Effects of Treg on CD8 + T Cell Responses in Chronic HIV Infection. Silvestri G, editor. PLOS Pathog. 2016;12:e1005995. Francino MP. Antibiotics and the Human Gut Microbiome: Dysbioses and Accumulation of Resistances. Front Microbiol. 2015;6:1543. Langdon A, Crook N, Dantas G. The effects of antibiotics on the microbiome throughout development and alternative approaches for therapeutic modulation. Genome Med. 2016;8:39. Zilberman-Schapira G, Zmora N, Itav S, Bashiardes S, Elinav H, Elinav E. The gut microbiome in human immunodeficiency virus infection. BMC Med. 2016;14:83. Nwosu FC, Avershina E, Wilson R, Rudi K. Gut Microbiota in HIV Infection: Implication for Disease Progression and Management. Gastroenterol Res Pract. 2014;2014:803185. Bluestone JA, Buckner JH, Fitch M, Gitelman SE, Gupta S, Hellerstein MK, et al. Type 1 diabetes immunotherapy using polyclonal regulatory T cells. Sci Transl Med [Internet]. 2015 [cited 2025 Apr 11];7. Available from: https://www.science.org/doi/ 10.1126/scitranslmed.aad4134 Additional Declarations No competing interests reported. Supplementary Files SupplementaryTable1.docx SupplementaryFig1.docx Cite Share Download PDF Status: Published Journal Publication published 11 Feb, 2026 Read the published version in European Journal of Medical Research → Version 1 posted Editorial decision: Revision requested 22 Aug, 2025 Reviews received at journal 19 Aug, 2025 Reviews received at journal 18 Aug, 2025 Reviews received at journal 31 Jul, 2025 Reviewers agreed at journal 31 Jul, 2025 Reviewers agreed at journal 30 Jul, 2025 Reviewers agreed at journal 20 Jul, 2025 Reviewers agreed at journal 18 Jul, 2025 Reviewers invited by journal 18 Jul, 2025 Editor assigned by journal 08 Jul, 2025 Submission checks completed at journal 08 Jul, 2025 First submitted to journal 06 Jul, 2025 You are reading this latest preprint version Research Square lets you share your work early, gain feedback from the community, and start making changes to your manuscript prior to peer review in a journal. As a division of Research Square Company, we’re committed to making research communication faster, fairer, and more useful. We do this by developing innovative software and high quality services for the global research community. Our growing team is made up of researchers and industry professionals working together to solve the most critical problems facing scientific publishing. Also discoverable on Platform About Our Team In Review Editorial Policies Advisory Board Help Center Resources Author Services Accessibility API Access RSS feed Manage Cookie Preferences © Research Square 2026 | ISSN 2693-5015 (online) Privacy Policy Terms of Service Do Not Sell My Personal Information {"props":{"pageProps":{"initialData":{"identity":"rs-7058889","acceptedTermsAndConditions":true,"allowDirectSubmit":false,"archivedVersions":[],"articleType":"Research Article","associatedPublications":[],"authors":[{"id":488179921,"identity":"5b078a0a-b83a-4610-872b-fc18b7a1f092","order_by":0,"name":"Smaranda Gliga","email":"data:image/png;base64,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","orcid":"","institution":"University Hospital Düsseldorf, Heinrich Heine University","correspondingAuthor":true,"prefix":"","firstName":"Smaranda","middleName":"","lastName":"Gliga","suffix":""},{"id":488179922,"identity":"dfaad4e2-6585-438d-b238-58ae065a9b72","order_by":1,"name":"Million Getechew Mesfun","email":"","orcid":"","institution":"Hirsch Institute of Tropical Medicine","correspondingAuthor":false,"prefix":"","firstName":"Million","middleName":"Getechew","lastName":"Mesfun","suffix":""},{"id":488179925,"identity":"a5f503d0-99dd-4d45-b1a2-4df6d466c790","order_by":2,"name":"Tafese Beyene Tufa","email":"","orcid":"","institution":"University Hospital Düsseldorf, Heinrich Heine University","correspondingAuthor":false,"prefix":"","firstName":"Tafese","middleName":"Beyene","lastName":"Tufa","suffix":""},{"id":488179926,"identity":"51e48262-9cd1-4467-98e6-c7afca84e1f1","order_by":3,"name":"Andre Fuchs","email":"","orcid":"","institution":"University Hospital of Augsburg","correspondingAuthor":false,"prefix":"","firstName":"Andre","middleName":"","lastName":"Fuchs","suffix":""},{"id":488179927,"identity":"00a774b0-7c11-41b5-8889-504143cb04ad","order_by":4,"name":"Hans Martin Orth","email":"","orcid":"","institution":"University Hospital Düsseldorf, Heinrich Heine University","correspondingAuthor":false,"prefix":"","firstName":"Hans","middleName":"Martin","lastName":"Orth","suffix":""},{"id":488179928,"identity":"52091b9f-8eb4-4cbe-9da3-da09ebea0c5d","order_by":5,"name":"Edmund Osei Kuffour","email":"","orcid":"","institution":"University Hospital Düsseldorf, Heinrich Heine University","correspondingAuthor":false,"prefix":"","firstName":"Edmund","middleName":"Osei","lastName":"Kuffour","suffix":""},{"id":488179929,"identity":"9e298e99-f300-4770-ad63-acd679b07446","order_by":6,"name":"Philipp A Lang","email":"","orcid":"","institution":"University Hospital Düsseldorf, Heinrich Heine University","correspondingAuthor":false,"prefix":"","firstName":"Philipp","middleName":"A","lastName":"Lang","suffix":""},{"id":488179932,"identity":"0023b731-55ad-44ad-a087-f44ed394449c","order_by":7,"name":"Tom Luedde","email":"","orcid":"","institution":"University Hospital Düsseldorf, Heinrich Heine University","correspondingAuthor":false,"prefix":"","firstName":"Tom","middleName":"","lastName":"Luedde","suffix":""},{"id":488179933,"identity":"7f8e0d61-ba4e-4334-83dd-d6f9b4501fa2","order_by":8,"name":"Torsten Feldt","email":"","orcid":"","institution":"University Hospital Düsseldorf, Heinrich Heine University","correspondingAuthor":false,"prefix":"","firstName":"Torsten","middleName":"","lastName":"Feldt","suffix":""}],"badges":[],"createdAt":"2025-07-06 15:53:16","currentVersionCode":1,"declarations":"","doi":"10.21203/rs.3.rs-7058889/v1","doiUrl":"https://doi.org/10.21203/rs.3.rs-7058889/v1","draftVersion":[],"editorialEvents":[{"content":"https://doi.org/10.1186/s40001-026-04025-4","type":"published","date":"2026-02-11T15:57:20+00:00"}],"editorialNote":"","failedWorkflow":false,"files":[{"id":87346380,"identity":"2537bd84-018c-4578-9216-e478f0cf1609","added_by":"auto","created_at":"2025-07-23 02:25:19","extension":"jpeg","order_by":1,"title":"Figure 1","display":"","copyAsset":false,"role":"figure","size":526869,"visible":true,"origin":"","legend":"\u003cp\u003eComparison of CD4\u003csup\u003e+\u003c/sup\u003e (A) and CD8\u003csup\u003e+\u003c/sup\u003e (B) T cell markers (order: activation, proliferation, exhaustion and Th17) in participants without HIV vs those with HIV. The Mann-Whitney test was used for statistical analysis. For the \u003cem\u003ey\u003c/em\u003e axis a logarithmic scale was used. Bars indicate median and CI95 %; n.s.: not significant and *\u003cem\u003ep\u003c/em\u003e \u0026lt; 0.05, **\u003cem\u003ep\u003c/em\u003e\u0026nbsp;\u0026lt; 0.01, *** \u003cem\u003ep\u003c/em\u003e\u0026nbsp;\u0026lt; 0.001 and **** \u003cem\u003ep\u003c/em\u003e\u0026lt;0.0001.\u003c/p\u003e","description":"","filename":"Fig.1.jpeg","url":"https://assets-eu.researchsquare.com/files/rs-7058889/v1/e2b7d32393b31b6f99925c83.jpeg"},{"id":87346854,"identity":"a87e9089-964e-4d16-adca-4b921dd5319b","added_by":"auto","created_at":"2025-07-23 02:33:19","extension":"jpeg","order_by":2,"title":"Figure 2","display":"","copyAsset":false,"role":"figure","size":697561,"visible":true,"origin":"","legend":"\u003cp\u003ePeripheral T cell parameters compared between \u003cem\u003eH. pylori\u003c/em\u003e-uninfected and infected individuals without HIV. Statistical evaluation employed the Mann–Whitney U test. Graphs display medians with 95 % confidence intervals (CI): CD4⁺ data (upper row) use polygon symbols, whereas CD8⁺ data (lower row) use triangles throughout all scatter plots. n.s. denotes non-significant; * p \u0026lt; 0.05, ** p \u0026lt; 0.01, **** p \u0026lt; 0.0001.\u003c/p\u003e","description":"","filename":"Fig.2.jpeg","url":"https://assets-eu.researchsquare.com/files/rs-7058889/v1/e4d9f74d09933a9c1d40819d.jpeg"},{"id":87346387,"identity":"ad8186cd-4fd3-44ae-9c7d-bb92d2c0e187","added_by":"auto","created_at":"2025-07-23 02:25:19","extension":"jpeg","order_by":3,"title":"Figure 3","display":"","copyAsset":false,"role":"figure","size":669481,"visible":true,"origin":"","legend":"\u003cp\u003ePeripheral T cell parameters compared between \u003cem\u003eH. pylori\u003c/em\u003e-uninfected and infected individuals in people living with HIV (PLWH). Statistical evaluation employed the Mann–Whitney U test. Graphs display medians with 95 % confidence intervals (CI): CD4⁺ data (upper row) use polygon symbols, whereas CD8⁺ data (lower row) use triangles throughout all scatter plots. n.s. denotes non-significant; ** p \u0026lt; 0.01.\u003c/p\u003e","description":"","filename":"Fig.3.jpeg","url":"https://assets-eu.researchsquare.com/files/rs-7058889/v1/994940d995076af0b2cf8ea4.jpeg"},{"id":87346856,"identity":"c11b913d-9286-43e7-ba0c-7d767ed17726","added_by":"auto","created_at":"2025-07-23 02:33:19","extension":"jpeg","order_by":4,"title":"Figure 4","display":"","copyAsset":false,"role":"figure","size":1512674,"visible":true,"origin":"","legend":"\u003cp\u003eT cell profile before (BE) and after (AE) eradication. Successfully eradicated people without (Fig. 4A) and people with HIV (Fig. 4B). Unsuccessfully eradicated people without (Fig. 4C) and with HIV (Fig. 4D) paired \u003cem\u003et- test \u003c/em\u003ewas used for statistical analysis. Only statistically significant changes in phenotype and function are presented: *\u003cem\u003ep\u003c/em\u003e \u0026lt; 0.05, **\u003cem\u003ep\u003c/em\u003e \u0026lt; 0.01, *** \u003cem\u003ep\u003c/em\u003e \u0026lt; 0.001.\u003c/p\u003e","description":"","filename":"Fig.4.jpeg","url":"https://assets-eu.researchsquare.com/files/rs-7058889/v1/bd71578b0375d38ab55fecf3.jpeg"},{"id":102785710,"identity":"4a43c1fc-60a9-4350-8493-70874dd017e7","added_by":"auto","created_at":"2026-02-16 16:09:03","extension":"pdf","order_by":0,"title":"","display":"","copyAsset":false,"role":"manuscript-pdf","size":4210274,"visible":true,"origin":"","legend":"","description":"","filename":"manuscript.pdf","url":"https://assets-eu.researchsquare.com/files/rs-7058889/v1/da00d184-2458-4124-ae00-5ed728430d61.pdf"},{"id":87346375,"identity":"7d045988-8205-411e-9b84-bc56a01c5b73","added_by":"auto","created_at":"2025-07-23 02:25:18","extension":"docx","order_by":0,"title":"","display":"","copyAsset":false,"role":"supplement","size":14849,"visible":true,"origin":"","legend":"","description":"","filename":"SupplementaryTable1.docx","url":"https://assets-eu.researchsquare.com/files/rs-7058889/v1/69a3360277f8945fe80252ce.docx"},{"id":87346381,"identity":"7accec8b-f8a4-4f84-b4b2-88fe22bdf495","added_by":"auto","created_at":"2025-07-23 02:25:19","extension":"docx","order_by":1,"title":"","display":"","copyAsset":false,"role":"supplement","size":361775,"visible":true,"origin":"","legend":"","description":"","filename":"SupplementaryFig1.docx","url":"https://assets-eu.researchsquare.com/files/rs-7058889/v1/38399aed692bcb57ee3494ea.docx"}],"financialInterests":"No competing interests reported.","formattedTitle":"Effects of Helicobacter pylori infection on T cell activation markers and regulatory T cells in people with and without HIV infection in Central Ethiopia","fulltext":[{"header":"Background","content":"\u003cp\u003e\u003cem\u003eHelicobacter pylori (H. pylori)\u003c/em\u003e is a microaerophilic bacterium that establishes lifelong residence in the gastric epithelium after infection, which usually occurs in childhood [\u003cspan citationid=\"CR1\" class=\"CitationRef\"\u003e1\u003c/span\u003e]. Although most carriers remain symptom-free, a proportion progress to dyspepsia and other gastroduodenal disorders [\u003cspan citationid=\"CR2\" class=\"CitationRef\"\u003e2\u003c/span\u003e].\u003c/p\u003e\u003cp\u003eEarly during colonization, innate sensing sparks the release of pro-inflammatory cytokines which in turn drive a strong T helper (Th) 1 and Th17 skewed T-cell response [\u003cspan citationid=\"CR3\" class=\"CitationRef\"\u003e3\u003c/span\u003e, \u003cspan citationid=\"CR4\" class=\"CitationRef\"\u003e4\u003c/span\u003e]. To prevent excessive tissue damage, the mucosa simultaneously produces counter-regulatory mediators such as transforming growth factor (TGF)-β and IL-10 [\u003cspan citationid=\"CR5\" class=\"CitationRef\"\u003e5\u003c/span\u003e]. TGF-β recruits T cells with anti-inflammatory activity, in particular regulatory T cells (T\u003csub\u003eregs\u003c/sub\u003e). T\u003csub\u003eregs\u003c/sub\u003e derive from the same lineage as na\u0026iuml;ve CD4\u003csup\u003e+\u003c/sup\u003e T cells and present the transcription factor forkhead box protein 3 (Foxp3) and the activating marker CD25 [\u003cspan citationid=\"CR6\" class=\"CitationRef\"\u003e6\u003c/span\u003e]. It was shown that a greater population of T\u003csub\u003eregs\u003c/sub\u003e is present in the gastric mucosa and in the peripheral blood of \u003cem\u003eH. pylori\u003c/em\u003e positive individuals, while T\u003csub\u003eregs\u003c/sub\u003e are largely absent in \u003cem\u003eH. pylori\u003c/em\u003e negative subjects [\u003cspan citationid=\"CR7\" class=\"CitationRef\"\u003e7\u003c/span\u003e, \u003cspan citationid=\"CR8\" class=\"CitationRef\"\u003e8\u003c/span\u003e]. Although T\u003csub\u003eregs\u003c/sub\u003e are activated to prevent hyperinflammatory conditions, they can also facilitate bacterial persistence by suppressing IFN-γ-producing effector T cells [\u003cspan citationid=\"CR9\" class=\"CitationRef\"\u003e9\u003c/span\u003e]. Consistent with this notion, experimental depletion of T\u003csub\u003eregs\u003c/sub\u003e with anti-CD25 antibodies in mice intensifies gastritis while reducing the gastric bacterial load, highlighting the delicate balance between protection and pathology [\u003cspan citationid=\"CR10\" class=\"CitationRef\"\u003e10\u003c/span\u003e].\u003c/p\u003e\u003cp\u003eGiven that a significant proportion of the 39\u0026nbsp;million people living with HIV (PLWH) [\u003cspan citationid=\"CR11\" class=\"CitationRef\"\u003e11\u003c/span\u003e] worldwide reside in low- and middle-income countries, where \u003cem\u003eH. pylori\u003c/em\u003e infection is highly prevalent, it is likely that many of these individuals also have \u003cem\u003eH. pylori\u003c/em\u003e co-infection. HIV infection is characterized by a reduced CD4\u003csup\u003e+\u003c/sup\u003e T cell count, which in time favors opportunistic infections, both viral and bacterial. While some publications show a high prevalence of \u003cem\u003eH. pylori\u003c/em\u003e in PLWH versus those without HIV [\u003cspan citationid=\"CR12\" class=\"CitationRef\"\u003e12\u003c/span\u003e], most report a lower prevalence of \u003cem\u003eH. pylori\u003c/em\u003e infection in PLWH, especially in patients with CD4\u003csup\u003e+\u003c/sup\u003e T cells below 200/\u0026micro;L [\u003cspan additionalcitationids=\"CR14 CR15\" citationid=\"CR13\" class=\"CitationRef\"\u003e13\u003c/span\u003e\u0026ndash;\u003cspan citationid=\"CR16\" class=\"CitationRef\"\u003e16\u003c/span\u003e]. Furthermore, the introduction of combination antiretroviral therapy (cART) induced an increase in dyspeptic complications and their severity [\u003cspan citationid=\"CR17\" class=\"CitationRef\"\u003e17\u003c/span\u003e]. HIV infection is associated with a strong recruitment of Th2 cells and reduction of Th1/Th17cells, which in turn decrease the pro-inflammatory response against \u003cem\u003eH. pylori\u003c/em\u003e and the intensity of gastritis [\u003cspan citationid=\"CR18\" class=\"CitationRef\"\u003e18\u003c/span\u003e].\u003c/p\u003e\u003cp\u003eA previous study our group performed in Ghana showed that \u003cem\u003eH. pylori\u003c/em\u003e carriage coincided with reduced CD4⁺ T cell activation (HLA-DR⁺CD38⁺), proliferation (Ki67) and PD-1 expression in both ART-na\u0026iuml;ve PLWH and HIV-negative controls [\u003cspan citationid=\"CR15\" class=\"CitationRef\"\u003e15\u003c/span\u003e]. These results parallel other reports in which co-infected, treatment-na\u0026iuml;ve patients displayed comparatively higher CD4⁺ counts and diminished HIV-1 viral loads, supporting the notion that the bacterium can dampen systemic immune activation [\u003cspan citationid=\"CR19\" class=\"CitationRef\"\u003e19\u003c/span\u003e, \u003cspan citationid=\"CR20\" class=\"CitationRef\"\u003e20\u003c/span\u003e]. To date, however, no study has characterized these immunological interactions in Ethiopian cohorts, nor has any work prospectively examined how eliminating \u003cem\u003eH. pylori\u003c/em\u003e affects T cell phenotypes in this setting. Accordingly, we designed the present prospective, randomized study to elucidate the mechanisms of systemic immune modulation linked to \u003cem\u003eH. pylori\u003c/em\u003e infection by tracking activation, proliferation, and exhaustion signatures before and after standard eradication therapy.\u003c/p\u003e"},{"header":"Materials and methods","content":"\u003cp\u003e\u003cb\u003eStudy population\u003c/b\u003e\u003c/p\u003e\u003cp\u003eBetween March and June 2017, we enrolled adults attending the Asella Teaching and Referral Hospital (ATRH, Ethiopia). Two streams were used: (i) successive visitors to the HIV outpatient service and (ii) successive clients of the hospital\u0026rsquo;s voluntary counselling-and-testing unit. All recruits underwent \u003cem\u003eH. pylori\u003c/em\u003e screening with a stool-antigen assay (see below). Eligibility criteria were 18\u0026ndash;55 years of age, signed informed consent and for PLWH, a CD4⁺ cell count above 350 cells/\u0026micro;l. From the first 140 HIV-negative and 140 HIV-positive individuals who satisfied these criteria we applied additional exclusion filters: prior immunosuppression, malignancy, chronic viral hepatitis (B and C), tuberculosis, elevated C-reactive protein, parasitic infection, recent anti-helminthic/antibiotic therapy, upper-gastrointestinal symptoms, recent acute infection, hemoglobin\u0026thinsp;\u0026lt;\u0026thinsp;10 g/dl or pregnancy, as described earlier \u0026sup1;⁵. All PLWH were on cART.\u003c/p\u003e\u003cp\u003e\u003cb\u003eEthics\u003c/b\u003e\u003c/p\u003e\u003cp\u003eThe protocol received approval in Ethiopia and Germany (refs A/U/H/S/C/87/6392; 3.10/271/2017; 5728).\u003c/p\u003e\u003cp\u003e\u003cb\u003eEradication therapy\u003c/b\u003e\u003c/p\u003e\u003cp\u003eWithin the \u003cem\u003eH. pylori\u003c/em\u003e\u0026ndash;positive group, 26 / 123 HIV-negative individuals and 25 / 121 PLWH were randomized to receive a 14-day triple regimen: metronidazole 500 mg, clarithromycin 500 mg and pantoprazole 40 mg, each taken twice daily. This macrolide- and nitroimidazole-based schedule follows Ethiopian guidelines and deliberately avoids penicillin to prevent reactions in participants with undocumented β-lactam allergy, as previously mentioned [\u003cspan citationid=\"CR21\" class=\"CitationRef\"\u003e21\u003c/span\u003e]. Clinical review with paired blood and stool sampling was scheduled at baseline and at three, six and 12 months. Successful eradication was defined as a negative stool-antigen test at the three-month visit.\u003c/p\u003e\u003cp\u003e\u003cb\u003eH. pylori\u003c/b\u003e \u003cb\u003estool antigen test\u003c/b\u003e\u003c/p\u003e\u003cp\u003eAntigen detection employed the GA Generic Assays kit (Blankenfelde, Germany) according to the manufacturer\u0026rsquo;s protocol. Stool microscopy with direct and formol-ether concentration techniques ruled out concomitant parasitic infection.\u003c/p\u003e\u003cp\u003e\u003cb\u003eBlood sample collection and laboratory analysis\u003c/b\u003e\u003c/p\u003e\u003cp\u003eAt every visit 15 ml of EDTA-anticoagulated blood and 5 ml of serum were collected. Serum at baseline was analyzed for hepatitis B surface antigen (HBsAg), anti-hepatitis C (HCV) antibodies (InTec Products Inc., China) and CRP (NADAL\u0026reg; CRP, nal von minden, Germany). Complete blood counts and, for PLWH, CD4⁺ T cell enumerations were performed locally on a FACSCalibur\u0026trade; flow cytometer (BD Biosciences, NJ, USA).\u003c/p\u003e\u003cp\u003e\u003cb\u003ePBMC isolation and cryopreservation\u003c/b\u003e\u003c/p\u003e\u003cp\u003ePeripheral blood mononuclear cells (PBMCs) were isolated by Ficoll-Hypaque density gradient, suspended in 20% dimethyl sulfoxide (DMSO)/ fetal calf serum (FCS) freezing medium, cooled at \u0026minus;\u0026thinsp;80\u0026deg;C for 24 h in a controlled-rate container and shipped on dry ice to D\u0026uuml;sseldorf for batched analysis.\u003c/p\u003e\u003cp\u003e\u003cb\u003eFlow-cytometric assessment\u003c/b\u003e\u003c/p\u003e\u003cp\u003eTo evaluate how \u003cem\u003eH. pylori\u003c/em\u003e clearance altered T cell characteristics, phenotypic read-outs recorded after therapy were compared with each participant\u0026rsquo;s own baseline. Cryopreserved PBMCs were rapidly thawed at 37\u0026deg;C, washed twice and resuspended in phosphate-buffered saline (PBS). Surface labelling was performed at 2\u0026ndash;8\u0026deg;C in the dark, whereas intracellular markers were stained at room temperature. Antibody details are listed in Supplementary Table\u0026nbsp;1; four dedicated panels covered (1) activation, (2) exhaustion, (3) regulatory T cells and (4) Th17 signatures. All reagents were sourced from eBioscience (Frankfurt am Main, Germany). Viable cells were gated with a fixable viability dye. After surface staining, samples in panels 2 and 4 were held in flow-cytometry buffer for immediate acquisition, while those in panels 1 and 3 underwent fixation, permeabilization and 30-min intracellular staining before a final wash into buffer. Data were collected on a three-laser BD FACSCanto II. Spectral overlap was corrected with UltraComp beads (eBioscience, Germany), singularly stained for each fluorochrome. Files were processed in FlowJo v10.1 (Tree Star, OR, USA); the gating strategy appears in Supplementary Fig.\u0026nbsp;1. Phenotypic outputs were stratified by HIV status and by \u003cem\u003eH. pylori\u003c/em\u003e infection, as well as by pre- versus post-eradication time points.\u003c/p\u003e\u003cdiv id=\"Sec3\" class=\"Section2\"\u003e\u003ch2\u003eStatistical Analysis\u003c/h2\u003e\u003cp\u003eData from participants without and with HIV were processed independently with Prism version 9.0 (GraphPad Software, San Diego, California, USA) software. Normally distributed variables were compared by paired or unpaired \u003cem\u003et\u003c/em\u003e tests; results are given as mean\u0026thinsp;\u0026plusmn;\u0026thinsp;standard deviation (SD) or median values. Skewed data were evaluated using the two-tailed Mann-Whitney test. Significance codes: n.s.=not significant; * p\u0026thinsp;\u0026lt;\u0026thinsp;0.05; ** p\u0026thinsp;\u0026lt;\u0026thinsp;0.01; *** p\u0026thinsp;\u0026lt;\u0026thinsp;0.001; **** p\u0026thinsp;\u0026lt;\u0026thinsp;0.0001, unless stated otherwise in the figure and text.\u003c/p\u003e\u003c/div\u003e"},{"header":"Results","content":"\u003cp\u003e\u003cb\u003eCohort\u003c/b\u003e\u003c/p\u003e\u003cp\u003eA cohort of 306 people living with HIV and 201 people without HIV underwent screening to detect \u003cem\u003eH. pylori\u003c/em\u003e colonization at enrolment. From these, we pragmatically selected 140 individuals in the HIV-positive cohort (78 colonized, 62 non-colonized) and an equal 140 individuals in the HIV-negative cohort (93 colonized, 47 non-colonized) for detailed evaluation of exclusion criteria, including co-infections, inflammation, medication, or pregnancy history. Subsequently, 27 \u003cem\u003eH. pylori\u003c/em\u003e positive volunteers from each cohort were randomly allocated to a 14-day metronidazole\u0026ndash;clarithromycin\u0026ndash;pantoprazole regimen, constituting the study\u0026rsquo;s standard triple eradication intervention arm for bacterial clearance.\u003c/p\u003e\u003cp\u003eAfter testing for confounding factors, two participants from the group with HIV and one participant from those without HIV infection were excluded from analysis as predefined. The rates of successful eradication in participants with HIV and those without HIV were previously reported [\u003cspan citationid=\"CR21\" class=\"CitationRef\"\u003e21\u003c/span\u003e].\u003c/p\u003e\u003cp\u003e\u003cb\u003eComparison of T cell profile in participants without vs. with HIV at baseline\u003c/b\u003e\u003c/p\u003e\u003cp\u003eIn participants with HIV, CD4\u003csup\u003e+\u003c/sup\u003e T cells exhibited consistently higher levels of Ki67 (median 96% vs 88%, p\u0026thinsp;\u0026lt;\u0026thinsp;0.0001) as proliferation markers, of PD-1 (median 19% vs 14%, p\u0026thinsp;=\u0026thinsp;0.0003) and TIM3 (median 3.4% vs 1.6%, p\u0026thinsp;\u0026lt;\u0026thinsp;0.0001) as exhaustion markers, and of Th17 Cells (CCR6\u003csup\u003e+\u003c/sup\u003eCD161\u003csup\u003e+\u003c/sup\u003e; median 0.64% vs 0.3%, p\u0026thinsp;\u0026lt;\u0026thinsp;0.0001) (Fig.\u0026nbsp;\u003cspan refid=\"Fig1\" class=\"InternalRef\"\u003e1\u003c/span\u003eA).\u003c/p\u003e\u003cp\u003e\u003c/p\u003e\u003cp\u003eIn addition, CD8\u003csup\u003e+\u003c/sup\u003e T cells had higher levels of Ki67 (median 7.2% vs 5.6%, p\u0026thinsp;=\u0026thinsp;0.002), PD-1 (median 0.84% vs 0.66%, p\u0026thinsp;=\u0026thinsp;0.046) and TIM3 (median 7.2% vs 3.4%, p\u0026thinsp;\u0026lt;\u0026thinsp;0.0001). (Fig.\u0026nbsp;\u003cspan refid=\"Fig1\" class=\"InternalRef\"\u003e1\u003c/span\u003eB).\u003c/p\u003e\u003cp\u003e\u003cb\u003eT Cell Profile in Participants without HIV according to\u003c/b\u003e \u003cb\u003eH. pylori\u003c/b\u003e \u003cb\u003estatus at baseline\u003c/b\u003e\u003c/p\u003e\u003cp\u003eIn the group of participants without HIV (n\u0026thinsp;=\u0026thinsp;123 after exclusion of 17 participants with confounding factors), T cell markers were analyzed comparatively between subgroups with (n\u0026thinsp;=\u0026thinsp;83) and without (n\u0026thinsp;=\u0026thinsp;40) \u003cem\u003eH. pylori\u003c/em\u003e infection. Results for CD4\u003csup\u003e+\u003c/sup\u003e and CD8\u003csup\u003e+\u003c/sup\u003e T cells are presented in Fig.\u0026nbsp;\u003cspan refid=\"Fig2\" class=\"InternalRef\"\u003e2\u003c/span\u003e.\u003c/p\u003e\u003cp\u003e\u003c/p\u003e\u003cp\u003eThe key findings of this analysis were that participants with \u003cem\u003eH. pylori\u003c/em\u003e infection showed significantly lower percentages of activation markers (HLA-DR\u003csup\u003e+\u003c/sup\u003eCD38\u003csup\u003e+\u003c/sup\u003e) on CD4\u003csup\u003e+\u003c/sup\u003e T cells compared to uninfected individuals (median 1.52% vs 2.24%, p\u0026thinsp;=\u0026thinsp;0.003). Furthermore, they showed lower levels of T cell exhaustion markers (PD-1, TIM3) in CD4\u003csup\u003e+\u003c/sup\u003e (PD-1: median 12.7% vs 16.05%, p\u0026thinsp;=\u0026thinsp;0.03; TIM3: median 1.28% vs 2.39%, p\u0026thinsp;=\u0026thinsp;0.002) and CD8\u003csup\u003e+\u003c/sup\u003e T cells (PD-1: median 0.49% vs 1.12%, p\u0026thinsp;=\u0026thinsp;0.001; TIM3: median 3.13% vs 5.61%, p\u0026thinsp;=\u0026thinsp;0.007) and significantly higher levels of T\u003csub\u003eregs\u003c/sub\u003e (CD25\u003csup\u003e+\u003c/sup\u003eFoxp3\u003csup\u003e+\u003c/sup\u003eCD4\u003csup\u003e+\u003c/sup\u003e; median 2% vs 1.08%, p\u0026thinsp;\u0026lt;\u0026thinsp;0.0001), highlighting their critical role in maintaining immune homeostasis by suppressing excessive immune activation.\u003c/p\u003e\u003cp\u003e\u003cb\u003eT Cell Profile in participants with HIV infection according to\u003c/b\u003e \u003cb\u003eH. pylori\u003c/b\u003e \u003cb\u003estatus at baseline\u003c/b\u003e\u003c/p\u003e\u003cp\u003eAmong the 121 people living with HIV, 56 tested negative and 65 positive for \u003cem\u003eH. pylori\u003c/em\u003e. Dual-infected individuals exhibited a higher proportion of regulatory T cells (median 2.9% vs 1.62%, \u003cem\u003ep\u003c/em\u003e\u0026thinsp;=\u0026thinsp;0.009), highlighting the contribution of T\u003csub\u003eregs\u003c/sub\u003e in moderating immune activation during coinfection (Fig.\u0026nbsp;\u003cspan refid=\"Fig3\" class=\"InternalRef\"\u003e3\u003c/span\u003e). In contrast, markers of T cell activation and exhaustion showed no significant variation between the two subgroups.\u003c/p\u003e\u003cp\u003e\u003c/p\u003e\u003cp\u003e\u003cb\u003eEffect of\u003c/b\u003e \u003cb\u003eH. pylori\u003c/b\u003e \u003cb\u003eEradication Therapy on T Cell Populations\u003c/b\u003e\u003c/p\u003e\u003cp\u003eOf the \u003cem\u003eH. pylori\u003c/em\u003e participants with HIV-infection (n\u0026thinsp;=\u0026thinsp;25) and without HIV infection (n\u0026thinsp;=\u0026thinsp;26) who were randomized to eradication therapy, 15/24 participants without HIV (62.5%) and 9/24 with HIV (37.5%) presented for the follow-up investigation three months after eradication therapy and were successfully eradicated (as confirmed by a negative \u003cem\u003eH. pylori\u003c/em\u003e stool antigen test) and thus eligible for final analysis (for details see the previous publication by Mesfun \u003cem\u003eet al\u003c/em\u003e) [\u003cspan citationid=\"CR21\" class=\"CitationRef\"\u003e21\u003c/span\u003e].\u003c/p\u003e\u003cp\u003eIn participants without HIV infection, successful \u003cem\u003eH. pylori\u003c/em\u003e eradication resulted in increased proliferation marker expression (Ki67) in CD4\u003csup\u003e+\u003c/sup\u003e T cells (n\u0026thinsp;=\u0026thinsp;14; SD 27.4%, p\u0026thinsp;=\u0026thinsp;0.049). CD8\u003csup\u003e+\u003c/sup\u003e T cells showed an increase in exhaustion markers (CD57) (SD 17.05%, p\u0026thinsp;=\u0026thinsp;0.0027; Fig.\u0026nbsp;\u003cspan refid=\"Fig4\" class=\"InternalRef\"\u003e4\u003c/span\u003eA). We observed a decrease in T\u003csub\u003eregs\u003c/sub\u003e in HIV-negative participants after successful (SD 1.5%, p\u0026thinsp;=\u0026thinsp;0.023), but also after unsuccessful (SD 0.72%, p\u0026thinsp;=\u0026thinsp;0.0004; Fig.\u0026nbsp;\u003cspan refid=\"Fig4\" class=\"InternalRef\"\u003e4\u003c/span\u003eC) \u003cem\u003eH. pylori\u003c/em\u003e eradication therapy.\u003c/p\u003e\u003cp\u003e\u003c/p\u003e\u003cp\u003eAmong PLWH, seven individuals who cleared \u003cem\u003eH. pylori\u003c/em\u003e showed a rise in CD8⁺ T cell activation (HLA-DR⁺CD38⁺; SD 0.39%, p\u0026thinsp;=\u0026thinsp;0.016) accompanied by a fall in Ki67 proliferation signals (SD 3.1%, p\u0026thinsp;=\u0026thinsp;0.012; Fig.\u0026nbsp;\u003cspan refid=\"Fig4\" class=\"InternalRef\"\u003e4\u003c/span\u003eB) and a parallel contraction of T\u003csub\u003eregs\u003c/sub\u003e (SD 1.98%, p\u0026thinsp;=\u0026thinsp;0.014). Conversely, in those whose therapy failed we detected reduced Ki-67 on CD8⁺ T lymphocytes (SD 13.3%, p\u0026thinsp;=\u0026thinsp;0.012), lower PD-1 on CD4⁺ (SD 8.43%, p\u0026thinsp;=\u0026thinsp;0.0009) and CD8⁺ T cells (SD 1.05%, p\u0026thinsp;=\u0026thinsp;0.01), together with elevated CD57 on CD4⁺ T cells (SD 10.14%, p\u0026thinsp;=\u0026thinsp;0.04; Fig.\u0026nbsp;\u003cspan refid=\"Fig4\" class=\"InternalRef\"\u003e4\u003c/span\u003eD). These shifts collectively suggest differential immune recalibration following attempted bacterial clearance in adults.\u003c/p\u003e"},{"header":"Discussion","content":"\u003cp\u003eBecause few studies from sub-Saharan Africa have explored how \u003cem\u003eH. pylori\u003c/em\u003e shapes immune responses, this study aimed to fill this critical gap by prospectively examining \u003cem\u003eH. pylori\u003c/em\u003e\u0026ndash;related effects on T cell populations, including markers of activation, exhaustion and T\u003csub\u003eregs\u003c/sub\u003e, in individuals with and without HIV infection. The rationale for such an investigation stems from the potential immune-modulating effects of \u003cem\u003eH. pylori\u003c/em\u003e, which may be especially pertinent in the setting of HIV, where the immune system is already under strain.\u003c/p\u003e\u003cp\u003eOur study revealed that T cell exhaustion markers, particularly PD-1, were consistently elevated in individuals with HIV, an observation aligning with well-established data on chronic immune dysregulation in HIV infection [\u003cspan citationid=\"CR22\" class=\"CitationRef\"\u003e22\u003c/span\u003e, \u003cspan citationid=\"CR23\" class=\"CitationRef\"\u003e23\u003c/span\u003e]. Moreover, despite limited regional data on \u003cem\u003eH. pylori\u003c/em\u003e immune interactions in Sub-Saharan Africa, we confirmed that in this cohort, HIV infection correlated with characteristic shifts in T cell activation, proliferative activity, and the distribution of regulatory subsets.\u003c/p\u003e\u003cp\u003eBuilding on these findings, we also evaluated Th17 cells, which tend to be depleted in untreated HIV but can be reconstituted under cART. Here, we noted persistently elevated Th17 markers in the HIV positive group, likely reflecting the relatively preserved CD4⁺ T cell counts afforded by ART. This backdrop provides important context for interpreting the role of \u003cem\u003eH. pylori\u003c/em\u003e, a bacterium often linked to robust Th17 responses in gastric lesions [\u003cspan additionalcitationids=\"CR25\" citationid=\"CR24\" class=\"CitationRef\"\u003e24\u003c/span\u003e\u0026ndash;\u003cspan citationid=\"CR26\" class=\"CitationRef\"\u003e26\u003c/span\u003e]. In our cohort, however, \u003cem\u003eH. pylori\u003c/em\u003e status did not appear to further alter Th17 frequency in people with HIV, indicating that cART-driven immune restoration may outweigh any additive influence from \u003cem\u003eH. pylori\u003c/em\u003e.\u003c/p\u003e\u003cp\u003eStudies examining \u003cem\u003eH. pylori\u003c/em\u003e-driven changes in peripheral T cell activation, proliferation, and exhaustion remain limited. In our earlier work, colonization corresponded to diminished CD4⁺ T cell activation (HLA-DR⁺CD38⁺) and lower expression of exhaustion markers, PD-1 on CD8⁺ cells and TIM-3 on both CD4⁺ and CD8⁺ subsets [\u003cspan citationid=\"CR15\" class=\"CitationRef\"\u003e15\u003c/span\u003e]. A different group, however, documented increased HLA-DR expression in PBMCs, especially in individuals with peptic ulcers, implying that \u003cem\u003eH. pylori\u003c/em\u003e could have variable immunomodulatory effects depending on clinical presentation [\u003cspan citationid=\"CR27\" class=\"CitationRef\"\u003e27\u003c/span\u003e]. Other contrasting results come from murine and human studies [\u003cspan citationid=\"CR28\" class=\"CitationRef\"\u003e28\u003c/span\u003e, \u003cspan citationid=\"CR29\" class=\"CitationRef\"\u003e29\u003c/span\u003e] describing elevated TIM3 expression in Th cells from \u003cem\u003eH. pylori\u003c/em\u003e‐infected hosts, and higher exhaustion levels on CD8⁺ T cells in gastric cancer patients [\u003cspan citationid=\"CR29\" class=\"CitationRef\"\u003e29\u003c/span\u003e]. Such inconsistencies may reflect differences in host genetics, regional factors, sample size or disease phenotypes (e.g., asymptomatic carriers vs. peptic ulcer disease vs. gastric cancer). Our exclusive enrollment of asymptomatic patients could partly explain why we observed lower levels of certain exhaustion or activation markers relative to other findings.\u003c/p\u003e\u003cp\u003eA significant outcome of our investigation was the notably higher percentage of T\u003csub\u003eregs\u003c/sub\u003e among \u003cem\u003eH. pylori\u003c/em\u003e-infected participants, regardless of HIV status, paralleling observations from histopathological studies of the gastric mucosa [\u003cspan additionalcitationids=\"CR8\" citationid=\"CR7\" class=\"CitationRef\"\u003e7\u003c/span\u003e\u0026ndash;\u003cspan citationid=\"CR9\" class=\"CitationRef\"\u003e9\u003c/span\u003e, \u003cspan citationid=\"CR24\" class=\"CitationRef\"\u003e24\u003c/span\u003e, \u003cspan citationid=\"CR30\" class=\"CitationRef\"\u003e30\u003c/span\u003e]. While most prior publications focused on local T\u003csub\u003eregs\u003c/sub\u003e in gastric tissue, only two reports have shown elevated circulating T\u003csub\u003eregs\u003c/sub\u003e in \u003cem\u003eH. pylori\u003c/em\u003e‐infected cohorts [\u003cspan citationid=\"CR8\" class=\"CitationRef\"\u003e8\u003c/span\u003e, \u003cspan citationid=\"CR24\" class=\"CitationRef\"\u003e24\u003c/span\u003e]. Our own earlier work hinted at a similar peripheral trend, suggesting that \u003cem\u003eH. pylori\u003c/em\u003e\u0026ndash;induced immune regulation may be systemic rather than confined solely to the gastric microenvironment [\u003cspan citationid=\"CR15\" class=\"CitationRef\"\u003e15\u003c/span\u003e].\u003c/p\u003e\u003cp\u003ePrior work by Eberhardt and colleagues [\u003cspan citationid=\"CR15\" class=\"CitationRef\"\u003e15\u003c/span\u003e] linked \u003cem\u003eH. pylori\u003c/em\u003e infection to reduced T cell activation, proliferation and exhaustion markers, but could not definitively establish causality. Accordingly, we sought to determine whether eradicating \u003cem\u003eH. pylori\u003c/em\u003e would lead to an immunological reversal of these trends. However, high levels of metronidazole resistance [\u003cspan citationid=\"CR31\" class=\"CitationRef\"\u003e31\u003c/span\u003e] in Ethiopia resulted in a low overall eradication rate, leaving only small subgroups available for a detailed pre- vs. post‐therapy comparison. Recognizing that \u003cem\u003eH. pylori\u003c/em\u003e typically recruits T\u003csub\u003eregs\u003c/sub\u003e to facilitate its persistence [\u003cspan citationid=\"CR30\" class=\"CitationRef\"\u003e30\u003c/span\u003e], we hypothesized that T\u003csub\u003ereg\u003c/sub\u003e levels would drop if \u003cem\u003eH. pylori\u003c/em\u003e were successfully eliminated. Indeed, we observed precisely that: a notable decline in T\u003csub\u003eregs\u003c/sub\u003e post‐eradication, regardless of HIV status. Yet, a parallel reduction occurred even in some participants without HIV who did not clear their infection, which we attribute to antibiotic‐induced dysbiosis [\u003cspan citationid=\"CR32\" class=\"CitationRef\"\u003e32\u003c/span\u003e, \u003cspan citationid=\"CR33\" class=\"CitationRef\"\u003e33\u003c/span\u003e]. This broader disruption of the microbiota may have secondary effects on T\u003csub\u003eregs\u003c/sub\u003e and Th17 subsets, mirroring data from other pathologies where antibiotic therapy modifies immune populations [\u003cspan citationid=\"CR34\" class=\"CitationRef\"\u003e34\u003c/span\u003e].\u003c/p\u003e\u003cp\u003eInterestingly, participants with HIV who were successfully eradicated showed increased activation of CD8⁺ T cells. We surmise that reduced T\u003csub\u003ereg\u003c/sub\u003e-mediated suppression could permit cytotoxic T lymphocytes to become more activated [\u003cspan citationid=\"CR35\" class=\"CitationRef\"\u003e35\u003c/span\u003e]. Unexpectedly, those who failed eradication still exhibited a decrease in T\u003csub\u003eregs\u003c/sub\u003e alongside reduced proliferation and PD‐1 expression, hinting at a complex interplay between T\u003csub\u003eregs\u003c/sub\u003e and PD‐1/PD‐L1 pathways [\u003cspan citationid=\"CR36\" class=\"CitationRef\"\u003e36\u003c/span\u003e]. These findings complicate a straightforward narrative of T\u003csub\u003ereg\u003c/sub\u003e modulation, pointing to additional factors, such as partial antibiotic effects, subclinical shifts in gut flora or baseline immune status, that may influence immune cell phenotypes regardless of eradication outcome.\u003c/p\u003e\u003cp\u003eAlthough we document significant immunological shifts following \u003cem\u003eH. pylori\u003c/em\u003e eradication, the limited sample size and suboptimal eradication success rate constrain our ability to draw broad conclusions. Larger-scale studies using alternative antibiotic regimens or susceptibility-guided therapy are needed to confirm these findings and clarify the complex interplay among \u003cem\u003eH. pylori\u003c/em\u003e, HIV status, antibiotic resistance and host genetics.\u003c/p\u003e\u003cp\u003eMoreover, while antibiotics may affect T\u003csub\u003eregs\u003c/sub\u003e and Th17 cells, we did not systematically evaluate broader gut microbiome shifts, which can transiently or persistently alter immune phenotypes and confound outcomes [\u003cspan citationid=\"CR37\" class=\"CitationRef\"\u003e37\u003c/span\u003e, \u003cspan citationid=\"CR38\" class=\"CitationRef\"\u003e38\u003c/span\u003e]. Future investigations should integrate microbiome analyses (e.g., 16S rRNA or metagenomic approaches) to distinguish the effects of \u003cem\u003eH. pylori\u003c/em\u003e eradication from those of collateral disruptions in gut flora.\u003c/p\u003e\u003cp\u003eFinally, despite observing immunological changes associated with \u003cem\u003eH. pylori\u003c/em\u003e infection, the long-term consequences for HIV progression remain unclear. Some research suggests that chronic coinfections can modulate systemic immune activation in people with HIV, possibly influencing viral reservoirs or disease progression [\u003cspan citationid=\"CR39\" class=\"CitationRef\"\u003e39\u003c/span\u003e, \u003cspan citationid=\"CR40\" class=\"CitationRef\"\u003e40\u003c/span\u003e]. Whether \u003cem\u003eH. pylori\u003c/em\u003e is ultimately protective, neutral or detrimental may depend on bacterial strain diversity, host genetics, cART status, and local factors. Further exploration of these variables is needed to unravel the multifaceted relationship between \u003cem\u003eH. pylori\u003c/em\u003e and HIV.\u003c/p\u003e"},{"header":"Conclusions","content":"\u003cp\u003eThis study underscores the importance of T\u003csub\u003eregs\u003c/sub\u003e in modulating immune responses during \u003cem\u003eH. pylori\u003c/em\u003e infection, regardless of HIV status. T\u003csub\u003ereg\u003c/sub\u003e levels were consistently elevated in infected individuals, potentially aiding bacterial persistence, and successful \u003cem\u003eH. pylori\u003c/em\u003e eradication led to a significant drop in T\u003csub\u003eregs\u003c/sub\u003e alongside improved immune activation and increased exhaustion. Although these findings suggest potential therapeutic benefits by modulating T\u003csub\u003eregs\u003c/sub\u003e, the low eradication rate and limited sample size limit broader conclusions. Larger studies are needed to confirm these results, clarify clinical applications, and investigate whether targeting T\u003csub\u003eregs\u003c/sub\u003e could benefit individuals with coinfections. Notably, although adoptive T\u003csub\u003ereg\u003c/sub\u003e therapy has reached Phase I/II trials in type 1 diabetes [\u003cspan citationid=\"CR41\" class=\"CitationRef\"\u003e41\u003c/span\u003e], no large trials have specifically addressed T\u003csub\u003eregs\u003c/sub\u003e in the context of coinfections.\u003c/p\u003e"},{"header":"Abbreviations","content":"\u003cp\u003eATRH \u0026nbsp; \u0026nbsp;Asella Teaching and Referral Hospital\u003c/p\u003e\n\u003cp\u003ecART \u0026nbsp; \u0026nbsp;Combination antiretroviral therapy\u003c/p\u003e\n\u003cp\u003eCRP \u0026nbsp; \u0026nbsp;C-reactive protein\u003c/p\u003e\n\u003cp\u003eDMSO \u0026nbsp; \u0026nbsp;Dimethyl sulfoxide\u003c/p\u003e\n\u003cp\u003eFCS \u0026nbsp; \u0026nbsp; \u0026nbsp; Fetal calf serum\u003c/p\u003e\n\u003cp\u003eFoxp3 \u0026nbsp; \u0026nbsp;Forkhead box protein 3\u003c/p\u003e\n\u003cp\u003eH. pylori \u0026nbsp; \u0026nbsp;Helicobacter pylori\u003c/p\u003e\n\u003cp\u003en.s. \u0026nbsp;Not significant\u003c/p\u003e\n\u003cp\u003ePBMC \u0026nbsp; \u0026nbsp;Peripheral blood mononuclear cell\u003c/p\u003e\n\u003cp\u003ePBS \u0026nbsp; \u0026nbsp; Phosphate-buffered saline\u003c/p\u003e\n\u003cp\u003ePLWH \u0026nbsp;People living with HIV\u003c/p\u003e\n\u003cp\u003eSD \u0026nbsp; \u0026nbsp; Standard deviation\u003c/p\u003e\n\u003cp\u003eTGF-\u0026beta; Transforming growth factor-\u0026beta;\u003c/p\u003e\n\u003cp\u003eTh \u0026nbsp; \u0026nbsp; T helper cell\u003c/p\u003e\n\u003cp\u003eTreg \u0026nbsp; \u0026nbsp;Regulatory T cell\u003c/p\u003e"},{"header":"Declarations","content":"\u003ch2\u003eConflicts of interest\u003c/h2\u003e\u003cp\u003eThe authors have no financial or non-financial conflicts of interest to disclose in connection with this work.\u003c/p\u003e\u003ch2\u003eFunding Declaration\u003c/h2\u003e\n\u003cp\u003eThis research received \u003cstrong\u003eno external funding\u003c/strong\u003e\u003cstrong\u003e.\u003c/strong\u003e\u003c/p\u003e\u003ch2\u003eAcknowledgments\u003c/h2\u003e\u003cp\u003eSG, TF, and MGM wrote and edited the manuscript. HMO, AF, TBT, EOK, and PAL designed the study protocol and recruited patients. MGM performed the flow cytometry experiments. TL provided critical scientific insight and oversight throughout the project. We also thank all staff and participants for their support.\u003c/p\u003e\u003ch2\u003eData Availability\u003c/h2\u003e\u003cp\u003eThe flow cytometry datasets generated and analysed during this study are not publicly archived to safeguard participant confidentiality, but they are available from the corresponding author on reasonable request.\u003c/p\u003e"},{"header":"References","content":"\u003col\u003e\u003cli\u003e\u003cspan\u003eB\u0026uuml;cker R, Azevedo-Vethacke M, Groll C, Garten D, Josenhans C, Suerbaum S, et al. Helicobacter pylori colonization critically depends on postprandial gastric conditions. Sci Rep. 2012;2:994.\u003c/span\u003e\u003c/li\u003e\u003cli\u003e\u003cspan\u003eXie F-J. \u003cem\u003eHelicobacter pylori\u003c/em\u003e infection and esophageal cancer risk: An updated meta-analysis. World J Gastroenterol. 2013;19:6098.\u003c/span\u003e\u003c/li\u003e\u003cli\u003e\u003cspan\u003eKivrak Salim D, Sahin M, K\u0026ouml;ksoy S, Adanir H, S\u0026uuml;leymanlar I. Local Immune Response in Helicobacter pylori Infection. Medicine (Baltimore). 2016;95:e3713.\u003c/span\u003e\u003c/li\u003e\u003cli\u003e\u003cspan\u003eBamford KB, Fan X, Crowe SE, Leary JF, Gourley WK, Luthra GK, et al. Lymphocytes in the human gastric mucosa during Helicobacter pylori have a T helper cell 1 phenotype. Gastroenterology. 1998;114:482\u0026ndash;92.\u003c/span\u003e\u003c/li\u003e\u003cli\u003e\u003cspan\u003eBornschein J, Kandulski A, Selgrad M, Malfertheiner P. From Gastric Inflammation to Gastric Cancer. Dig Dis. 2010;28:609\u0026ndash;14.\u003c/span\u003e\u003c/li\u003e\u003cli\u003e\u003cspan\u003eCuriel TJ. Tregs and rethinking cancer immunotherapy. J Clin Invest. 2007;117:1167\u0026ndash;74.\u003c/span\u003e\u003c/li\u003e\u003cli\u003e\u003cspan\u003eLundgren A, Str\u0026ouml;mberg E, Sj\u0026ouml;ling \u0026Aring;, Lindholm C, Enarsson K, Edebo A, et al. Mucosal \u003cem\u003eFOXP3\u003c/em\u003e -Expressing CD4\u003csup\u003e+\u003c/sup\u003e CD25\u003csup\u003ehigh\u003c/sup\u003e Regulatory T Cells in \u003cem\u003eHelicobacter pylori\u003c/em\u003e -Infected Patients. Infect Immun. 2005;73:523\u0026ndash;31.\u003c/span\u003e\u003c/li\u003e\u003cli\u003e\u003cspan\u003eCook KW, Letley DP, Ingram RJM, Staples E, Skjoldmose H, Atherton JC, et al. CCL20/CCR6-mediated migration of regulatory T cells to the \u003cem\u003eHelicobacter pylori\u003c/em\u003e -infected human gastric mucosa. Gut. 2014;63:1550\u0026ndash;9.\u003c/span\u003e\u003c/li\u003e\u003cli\u003e\u003cspan\u003eKandulski A, Malfertheiner P, Wex T. Role of regulatory T-cells in H. pylori-induced gastritis and gastric cancer. Anticancer Res. 2010;30:1093\u0026ndash;103.\u003c/span\u003e\u003c/li\u003e\u003cli\u003e\u003cspan\u003eRad R, Brenner L, Bauer S, Schwendy S, Layland L, Da Costa CP, et al. CD25+/Foxp3\u0026thinsp;+\u0026thinsp;T Cells Regulate Gastric Inflammation and Helicobacter pylori Colonization In Vivo. Gastroenterology. 2006;131:525\u0026ndash;37.\u003c/span\u003e\u003c/li\u003e\u003cli\u003e\u003cspan\u003eUNAIDS/WHO estimates. Available from: \u003cspan class=\"ExternalRef\"\u003e\u003cspan class=\"RefSource\"\u003ehttps://www.who.int/images/default-source/departments/hiv/summary-of-the-global-hiv-epidemic-2022.png?sfvrsn=73ac5b6a_13\u003c/span\u003e\u003cspan address=\"https://www.who.int/images/default-source/departments/hiv/summary-of-the-global-hiv-epidemic-2022.png?sfvrsn=73ac5b6a_13\" targettype=\"URL\" class=\"RefTarget\"\u003e\u003c/span\u003e\u003c/span\u003e\u003c/span\u003e\u003c/li\u003e\u003cli\u003e\u003cspan\u003eAbadi T, Teklu T, Wondmagegn T, Alem M, Desalegn G. Helicobacter pylori infection and associated risk factors among HIV-positive and HIV-negative individuals in Northern Ethiopia. J Infect Chemother. 2025;31:102517.\u003c/span\u003e\u003c/li\u003e\u003cli\u003e\u003cspan\u003eFialho AB, Braga-Neto MB, Guerra EJ, Fialho AM, Fernandes KC, Sun JL, et al. Low prevalence of H. pylori Infection in HIV-Positive Patients in the Northeast of Brazil. BMC Gastroenterol. 2011;11:13.\u003c/span\u003e\u003c/li\u003e\u003cli\u003e\u003cspan\u003eOlmos M, Araya V, Pskorz E, Quesada EC, Concetti H, Perez H, et al. Coinfection: Helicobacter pylori/Human Immunodeficiency Virus. Dig Dis Sci. 2004;49:1836\u0026ndash;9.\u003c/span\u003e\u003c/li\u003e\u003cli\u003e\u003cspan\u003eEberhardt KA, Sarfo FS, Dompreh A, Kuffour EO, Geldmacher C, Soltau M, et al. \u003cem\u003eHelicobacter pylori\u003c/em\u003e Coinfection Is Associated With Decreased Markers of Immune Activation in ART-Naive HIV-Positive and in HIV-Negative Individuals in Ghana. Clin Infect Dis. 2015;61:1615\u0026ndash;23.\u003c/span\u003e\u003c/li\u003e\u003cli\u003e\u003cspan\u003eCacciarelli AG, Marano BJ, Gualtieri NM, Zuretti AR, Torres RA, Starpoli AA, et al. Lower Helicobacter pylori infection and peptic ulcer disease prevalence in patients with AIDS and suppressed CD4 counts. Am J Gastroenterol. 1996;91:1783\u0026ndash;4.\u003c/span\u003e\u003c/li\u003e\u003cli\u003e\u003cspan\u003eRadovanović Spurnić A, Brmbolić B, Stojšić Z, Pekmezović T, Bukumirić Z, Korać M, et al. The increasing prevalence of HIV/ \u003cem\u003eHelicobacter pylori\u003c/em\u003e co-infection over time, along with the evolution of antiretroviral therapy (ART). PeerJ. 2017;5:e3392.\u003c/span\u003e\u003c/li\u003e\u003cli\u003e\u003cspan\u003eKrzyżek P, Gościniak G. Immunomodulatory influence of HIV and EBV on Helicobacter pylori infections \u0026ndash; a review. Ann Parasitol. 2019;65:3\u0026ndash;17.\u003c/span\u003e\u003c/li\u003e\u003cli\u003e\u003cspan\u003eAbadi T, Teklu T, Wondmagegn T, Alem M, Desalegn G. CD4\u0026thinsp;+\u0026thinsp;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. Eur J Med Res. 2024;29:178.\u003c/span\u003e\u003c/li\u003e\u003cli\u003e\u003cspan\u003eSpurnic AR, Bukumiric Z, Jevtovic D, Brmbolic B, Pekmezovic T, Salemovic D, et al. Helicobacter pylori infection rates in dyspeptic Serbian HIV-infected patients compared to HIV-negative controls. PloS One. 2021;16:e0248041.\u003c/span\u003e\u003c/li\u003e\u003cli\u003e\u003cspan\u003eMesfun MG, Gliga S, Fuchs A, Orth HM, Sch\u0026ouml;nfeld A, Luedde T, et al. Prevalence of H. pylori among asymptomatic HIV-positive and negative individuals in Central Ethiopia and efficacy of eradication therapy. IJID Reg. 2022;2:169\u0026ndash;74.\u003c/span\u003e\u003c/li\u003e\u003cli\u003e\u003cspan\u003eTrautmann L, Janbazian L, Chomont N, Said EA, Gimmig S, Bessette B, et al. Upregulation of PD-1 expression on HIV-specific CD8\u0026thinsp;+\u0026thinsp;T cells leads to reversible immune dysfunction. Nat Med. 2006;12:1198\u0026ndash;202.\u003c/span\u003e\u003c/li\u003e\u003cli\u003e\u003cspan\u003eBreton G, Chomont N, Takata H, Fromentin R, Ahlers J, Filali-Mouhim A, et al. Programmed Death-1 Is a Marker for Abnormal Distribution of Naive/Memory T Cell Subsets in HIV-1 Infection. J Immunol. 2013;191:2194\u0026ndash;204.\u003c/span\u003e\u003c/li\u003e\u003cli\u003e\u003cspan\u003eTalayev V, Svetlova M, Zaichenko I, Voronina E, Babaykina O, Neumoina N, et al. CCR6\u003csup\u003e+\u003c/sup\u003e T helper cells and regulatory T cells in the blood and gastric mucosa during \u003cem\u003eHelicobacter pylori\u003c/em\u003e infection. Helicobacter. 2024;29:e13097.\u003c/span\u003e\u003c/li\u003e\u003cli\u003e\u003cspan\u003ePinchuk IV, Morris KT, Nofchissey RA, Earley RB, Wu J-Y, Ma TY, et al. Stromal cells induce Th17 during Helicobacter pylori infection and in the gastric tumor microenvironment. PloS One. 2013;8:e53798.\u003c/span\u003e\u003c/li\u003e\u003cli\u003e\u003cspan\u003eLal KG, Phuang-Ngern Y, Suhkumvittaya S, Leeansyah E, Alrubayyi A, Dias J, et al. Longitudinal Analysis of Peripheral and Colonic CD161\u0026thinsp;+\u0026thinsp;CD4\u0026thinsp;+\u0026thinsp;T Cell Dysfunction in Acute HIV-1 Infection and Effects of Early Treatment Initiation. Viruses. 2020;12:1426.\u003c/span\u003e\u003c/li\u003e\u003cli\u003e\u003cspan\u003eSuleymanov Z. Expression of class I and II MHC receptors in Helicobacter pylori-positive patients with active gastritis and duodenal ulcer. Turk J Gastroenterol Off J Turk Soc Gastroenterol. 2003;14:168\u0026ndash;72.\u003c/span\u003e\u003c/li\u003e\u003cli\u003e\u003cspan\u003eHu S, Xie Y, Zhou N, Jin L, Tan Y, Liu D, et al. Expression of T-cell immunoglobulin- and mucin-domain-containing molecules-1 and \u0026ndash;\u0026thinsp;3 (Tim-1 and Tim-3) in Helicobacter pylori infection. Helicobacter. 2011;16:373\u0026ndash;81.\u003c/span\u003e\u003c/li\u003e\u003cli\u003e\u003cspan\u003eShen P, Yue R, Tang J, Si H, Shen L, Guo C, et al. Preferential Tim-3 expression on Treg and CD8(+) T cells, supported by tumor-associated macrophages, is associated with worse prognosis in gastric cancer. Am J Transl Res. 2016;8:3419\u0026ndash;28.\u003c/span\u003e\u003c/li\u003e\u003cli\u003e\u003cspan\u003eKandulski A, Wex T, Kuester D, Peitz U, Gebert I, Roessner A, et al. Naturally occurring regulatory T cells (CD4+, CD25high, FOXP3+) in the antrum and cardia are associated with higher H. pylori colonization and increased gene expression of TGF-beta1. Helicobacter. 2008;13:295\u0026ndash;303.\u003c/span\u003e\u003c/li\u003e\u003cli\u003e\u003cspan\u003eAsrat D, Kassa E, Mengistu Y, Nilsson I, Wadstr\u0026ouml;m T. Antimicrobial susceptibility pattern of Helicobacter pylori strains isolated from adult dyspeptic patients in Tikur Anbassa University Hospital, Addis Ababa, Ethiopia. Ethiop Med J. 2004;42:79\u0026ndash;85.\u003c/span\u003e\u003c/li\u003e\u003cli\u003e\u003cspan\u003ePandiyan P, Bhaskaran N, Zou M, Schneider E, Jayaraman S, Huehn J. Microbiome Dependent Regulation of Tregs and Th17 Cells in Mucosa. Front Immunol. 2019;10:426.\u003c/span\u003e\u003c/li\u003e\u003cli\u003e\u003cspan\u003eYap TW-C, Gan H-M, Lee Y-P, Leow AH-R, Azmi AN, Francois F, et al. Helicobacter pylori Eradication Causes Perturbation of the Human Gut Microbiome in Young Adults. Badger JH, editor. PLOS ONE. 2016;11:e0151893.\u003c/span\u003e\u003c/li\u003e\u003cli\u003e\u003cspan\u003eRajendran M, Looney S, Singh N, Elashiry M, Meghil MM, El-Awady AR, et al. Systemic Antibiotic Therapy Reduces Circulating Inflammatory Dendritic Cells and Treg\u0026ndash;Th17 Plasticity in Periodontitis. J Immunol. 2019;202:2690\u0026ndash;9.\u003c/span\u003e\u003c/li\u003e\u003cli\u003e\u003cspan\u003eSuvas S, Kumaraguru U, Pack CD, Lee S, Rouse BT. CD4\u0026thinsp;+\u0026thinsp;CD25\u0026thinsp;+\u0026thinsp;T cells regulate virus-specific primary and memory CD8\u0026thinsp;+\u0026thinsp;T cell responses. J Exp Med. 2003;198:889\u0026ndash;901.\u003c/span\u003e\u003c/li\u003e\u003cli\u003e\u003cspan\u003eNikolova M, Wiedemann A, Muhtarova M, Achkova D, Lacabaratz C, L\u0026eacute;vy Y. Subset- and Antigen-Specific Effects of Treg on CD8\u0026thinsp;+\u0026thinsp;T Cell Responses in Chronic HIV Infection. Silvestri G, editor. PLOS Pathog. 2016;12:e1005995.\u003c/span\u003e\u003c/li\u003e\u003cli\u003e\u003cspan\u003eFrancino MP. Antibiotics and the Human Gut Microbiome: Dysbioses and Accumulation of Resistances. Front Microbiol. 2015;6:1543.\u003c/span\u003e\u003c/li\u003e\u003cli\u003e\u003cspan\u003eLangdon A, Crook N, Dantas G. The effects of antibiotics on the microbiome throughout development and alternative approaches for therapeutic modulation. Genome Med. 2016;8:39.\u003c/span\u003e\u003c/li\u003e\u003cli\u003e\u003cspan\u003eZilberman-Schapira G, Zmora N, Itav S, Bashiardes S, Elinav H, Elinav E. The gut microbiome in human immunodeficiency virus infection. BMC Med. 2016;14:83.\u003c/span\u003e\u003c/li\u003e\u003cli\u003e\u003cspan\u003eNwosu FC, Avershina E, Wilson R, Rudi K. Gut Microbiota in HIV Infection: Implication for Disease Progression and Management. Gastroenterol Res Pract. 2014;2014:803185.\u003c/span\u003e\u003c/li\u003e\u003cli\u003e\u003cspan\u003eBluestone JA, Buckner JH, Fitch M, Gitelman SE, Gupta S, Hellerstein MK, et al. Type 1 diabetes immunotherapy using polyclonal regulatory T cells. Sci Transl Med [Internet]. 2015 [cited 2025 Apr 11];7. Available from: \u003cspan class=\"ExternalRef\"\u003e\u003cspan class=\"RefSource\"\u003ehttps://www.science.org/doi/\u003c/span\u003e\u003cspan address=\"https://www.science.org/doi/\" targettype=\"URL\" class=\"RefTarget\"\u003e\u003c/span\u003e\u003c/span\u003e\u003cspan class=\"ExternalRef\"\u003e\u003cspan class=\"RefSource\"\u003e10.1126/scitranslmed.aad4134\u003c/span\u003e\u003cspan address=\"10.1126/scitranslmed.aad4134\" targettype=\"DOI\" class=\"RefTarget\"\u003e\u003c/span\u003e\u003c/span\u003e\u003c/span\u003e\u003c/li\u003e\u003c/ol\u003e"}],"fulltextSource":"","fullText":"","funders":[],"hasAdminPriorityOnWorkflow":false,"hasManuscriptDocX":true,"hasOptedInToPreprint":true,"hasPassedJournalQc":"","hasAnyPriority":false,"hideJournal":false,"highlight":"","institution":"","isAcceptedByJournal":true,"isAuthorSuppliedPdf":false,"isDeskRejected":"","isHiddenFromSearch":false,"isInQc":false,"isInWorkflow":false,"isPdf":false,"isPdfUpToDate":true,"isWithdrawnOrRetracted":false,"journal":{"display":true,"email":"[email protected]","identity":"european-journal-of-medical-research","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":false,"externalIdentity":"ejmr","sideBox":"Learn more about [European Journal of Medical Research](http://eurjmedres.biomedcentral.com)","snPcode":"40001","submissionUrl":"https://submission.nature.com/new-submission/40001/3","title":"European Journal of Medical Research","twitterHandle":"@BioMedCentral","acdcEnabled":true,"dfaEnabled":true,"editorialSystem":"em","reportingPortfolio":"BMC/SO AJ","inReviewEnabled":true,"inReviewRevisionsEnabled":true},"keywords":"T cell exhaustion, T cell proliferation, Helicobacter pylori eradication therapy, Immune modulation, HIV","lastPublishedDoi":"10.21203/rs.3.rs-7058889/v1","lastPublishedDoiUrl":"https://doi.org/10.21203/rs.3.rs-7058889/v1","license":{"name":"CC BY 4.0","url":"https://creativecommons.org/licenses/by/4.0/"},"manuscriptAbstract":"\u003ch2\u003eBackground:\u003c/h2\u003e\u003cp\u003e\u003cem\u003eHelicobacter pylori (H. pylori)\u003c/em\u003e is known to modulate host immunity and sustain chronic inflammation, yet most data come from HIV-negative populations. In people living with HIV, whose T cell compartments are already dysregulated, the way \u003cem\u003eH. pylori\u003c/em\u003e shapes peripheral T cell phenotypes, and how those profiles change after eradication therapy, is still unclear. Because both infections are common in Central Ethiopia, we examined peripheral T cell phenotypes in adults with and without HIV according to \u003cem\u003eH. pylori\u003c/em\u003e status and assessed the immunologic effects of antibiotic eradication.\u003c/p\u003e\u003ch2\u003eMaterials and Methods\u003c/h2\u003e\u003cp\u003eWe conducted a prospective study in people with and without HIV infection from Ethiopia. \u003cem\u003eH. pylori\u003c/em\u003e status was determined by stool-antigen testing; a subset received standard triple therapy and was followed for 12 months. Multiparameter flow cytometry quantified T cell activation, proliferation, exhaustion, and regulatory T cells (T\u003csub\u003eregs\u003c/sub\u003e) at baseline and after therapy.\u003c/p\u003e\u003ch2\u003eResults\u003c/h2\u003e\u003cp\u003eT cell analyses showed that participants with HIV had consistently higher proliferation (Ki67), exhaustion (PD-1, TIM3), and Th17 (CCR6⁺CD161⁺) markers than those without HIV. \u003cem\u003eH. pylori\u003c/em\u003e-positive individuals exhibited higher T\u003csub\u003ereg\u003c/sub\u003e levels irrespective of HIV status (HIV-negative: median 2% vs 1.08%, p\u0026thinsp;\u0026lt;\u0026thinsp;0.0001; HIV-positive: median 2.9% vs 1.62%, p\u0026thinsp;=\u0026thinsp;0.009). Eradication therapy led to a significant reduction in T\u003csub\u003eregs\u003c/sub\u003e in both HIV-positive (SD 1.98%, p\u0026thinsp;=\u0026thinsp;0.014) and HIV-negative (SD 1.5%, p\u0026thinsp;=\u0026thinsp;0.023) groups.\u003c/p\u003e\u003ch2\u003eConclusions\u003c/h2\u003e\u003cp\u003e\u003cem\u003eH. pylori\u003c/em\u003e infection is associated with specific alterations in T cell profiles, in both HIV negative and positive populations. Eradication therapy, irrespective of success, reduces T\u003csub\u003eregs\u003c/sub\u003e and partially restores T cell function, offering insight into potential therapeutic strategies for managing immune dysregulation in co-infected populations.\u003c/p\u003e\u003ch2\u003eTrial registration\u003c/h2\u003e\u003cp\u003eNot applicable. This study was not registered in a clinical trial registry, as there was no focus on investigational treatment. The aspects of the study reported here were observational in character. Ethical approval was obtained from both Ethiopian and German ethic committees. The study adhered to the Declaration of Helsinki and ICHGCP guidelines.\u003c/p\u003e","manuscriptTitle":"Effects of Helicobacter pylori infection on T cell activation markers and regulatory T cells in people with and without HIV infection in Central Ethiopia","msid":"","msnumber":"","nonDraftVersions":[{"code":1,"date":"2025-07-23 02:25:14","doi":"10.21203/rs.3.rs-7058889/v1","editorialEvents":[{"type":"communityComments","content":0},{"type":"decision","content":"Revision requested","date":"2025-08-22T18:04:10+00:00","index":"","fulltext":""},{"type":"editorInvitedReview","content":"","date":"2025-08-19T11:16:13+00:00","index":"hide","fulltext":""},{"type":"editorInvitedReview","content":"","date":"2025-08-18T18:07:21+00:00","index":"hide","fulltext":""},{"type":"editorInvitedReview","content":"","date":"2025-07-31T08:15:17+00:00","index":"hide","fulltext":""},{"type":"reviewerAgreed","content":"258120117577840984836770212716529281564","date":"2025-07-31T08:09:19+00:00","index":"hide","fulltext":""},{"type":"reviewerAgreed","content":"185177340155824235061412616331420409164","date":"2025-07-30T16:51:06+00:00","index":"hide","fulltext":""},{"type":"reviewerAgreed","content":"211353374188658641184091117272283321656","date":"2025-07-20T23:49:53+00:00","index":"hide","fulltext":""},{"type":"reviewerAgreed","content":"176408620302717806022454624533355090143","date":"2025-07-18T13:14:11+00:00","index":"hide","fulltext":""},{"type":"reviewersInvited","content":"","date":"2025-07-18T11:11:35+00:00","index":"","fulltext":""},{"type":"editorAssigned","content":"","date":"2025-07-08T15:03:39+00:00","index":"","fulltext":""},{"type":"checksComplete","content":"","date":"2025-07-08T11:34:50+00:00","index":"","fulltext":""},{"type":"submitted","content":"European Journal of Medical Research","date":"2025-07-06T15:50:38+00:00","index":"","fulltext":""}],"status":"published","journal":{"display":true,"email":"[email protected]","identity":"european-journal-of-medical-research","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":false,"externalIdentity":"ejmr","sideBox":"Learn more about [European Journal of Medical Research](http://eurjmedres.biomedcentral.com)","snPcode":"40001","submissionUrl":"https://submission.nature.com/new-submission/40001/3","title":"European Journal of Medical Research","twitterHandle":"@BioMedCentral","acdcEnabled":true,"dfaEnabled":true,"editorialSystem":"em","reportingPortfolio":"BMC/SO AJ","inReviewEnabled":true,"inReviewRevisionsEnabled":true}}],"origin":"","ownerIdentity":"afb81907-01ae-4bcf-8a4d-75372a593e3f","owner":[],"postedDate":"July 23rd, 2025","published":true,"recentEditorialEvents":[],"rejectedJournal":[],"revision":"","amendment":"","status":"published-in-journal","subjectAreas":[],"tags":[],"updatedAt":"2026-02-16T16:06:56+00:00","versionOfRecord":{"articleIdentity":"rs-7058889","link":"https://doi.org/10.1186/s40001-026-04025-4","journal":{"identity":"european-journal-of-medical-research","isVorOnly":false,"title":"European Journal of Medical Research"},"publishedOn":"2026-02-11 15:57:20","publishedOnDateReadable":"February 11th, 2026"},"versionCreatedAt":"2025-07-23 02:25:14","video":"","vorDoi":"10.1186/s40001-026-04025-4","vorDoiUrl":"https://doi.org/10.1186/s40001-026-04025-4","workflowStages":[]},"version":"v1","identity":"rs-7058889","journalConfig":"researchsquare"},"__N_SSP":true},"page":"/article/[identity]/[[...version]]","query":{"redirect":"/article/rs-7058889","identity":"rs-7058889","version":["v1"]},"buildId":"8U1c8b4HqxoKbykW_rLl7","isFallback":false,"isExperimentalCompile":false,"dynamicIds":[84888],"gssp":true,"scriptLoader":[]}