Distinct and dynamic activation profiles of circulating dendritic cells and monocytes in mild COVID-19 and after yellow fever vaccination.

Dysregulation of the myeloid cell compartment is a feature of severe disease in hospitalized COVID-19 patients. Here, we investigated the response of circulating dendritic cell (DC) and monocyte subpopulations in SARS-CoV-2 infected outpatients with mild disease and compared it to the response of healthy individuals to yellow fever vaccine virus YF17D as a model of a well-coordinated response to viral infection. In SARS-CoV-2-infected outpatients circulating DCs were persistently reduced for several weeks whereas after YF17D vaccination DC numbers were decreased temporarily and rapidly replenished by increased proliferation until 14 days after vaccination. The majority of COVID-19 outpatients showed high expression of CD86 and PD-L1 in monocytes and DCs early on, resembling the dynamic after YF17D vaccination. In a subgroup of patients, low CD86 and high PD-L1 expression were detected in monocytes and DCs coinciding with symptoms, higher age, and lower lymphocyte counts. This phenotype was similar to that observed in severely ill COVID-19 patients, but less pronounced. Thus, prolonged reduction and dysregulated activation of blood DCs and monocytes were seen in a subgroup of symptomatic non-hospitalized COVID-19 patients while a transient coordinated activation was characteristic for the majority of patients with mild COVID-19 and the response to YF17D vaccination.

Hybrid immunity by two COVID-19 mRNA vaccinations and one breakthrough infection provides a robust and balanced cellular immune response as basic immunity against severe acute respiratory syndrome coronavirus 2.

This longitudinal prospective controlled multicenter study aimed to monitor immunity generated by three exposures caused by breakthrough infections (BTI) after COVID-19-vaccination considering pre-existing cell-mediated immunity to common-corona-viruses (CoV) which may impact cellular reactivity against SARS-CoV-2. Anti-SARS-CoV-2-spike-IgG antibodies (anti-S-IgG) and cellular reactivity against Spike-(S)- and nucleocapsid-(N)-proteins were determined in fully-vaccinated (F) individuals who either experienced BTI (F+BTI) or had booster vaccination (F+Booster) compared to partially vaccinated (P+BTI) and unvaccinated (U) from 1 to 24 weeks post PCR-confirmed infection. High avidity anti-S-IgG were found in F+BTI compared to U, the latter exhibiting increased long-lasting pro-inflammatory cytokines to S-stimulation. CoV was associated with higher cellular reactivity in U, whereas no association was seen in F. The study illustrates the induction of significant S-specific cellular responses in F+BTI building-up basic immunity by three exposures. Only U seem to benefit from pre-existing CoV immunity but demonstrated inflammatory immune responses compared to F+BTI who immunologically benefit from enhanced humoral and cellular immunity after BTI. This study demonstrates that individuals with hybrid immunity from COVID-19-vaccination and BTI acquire a stable humoral and cellular immune response that is maintained for at least 6 months. Our findings corroborate recommendations by health authorities to build on basic immunity by three S-protein exposures.

Studying temporal titre evolution of commercial SARS-CoV-2 assays reveals significant shortcomings of using BAU standardization for comparison.

BACKGROUND: Measuring specific anti-SARS-CoV-2 antibodies has become one of the main epidemiological tools to survey the ongoing SARS-CoV-2 pandemic, but also vaccination response. The WHO made available a set of well-characterized samples derived from recovered individuals to allow normalization between different quantitative anti-Spike assays to defined Binding Antibody Units (BAU). METHODS: To assess sero-responses longitudinally, a cohort of ninety-nine SARS-CoV-2 RT-PCR positive subjects was followed up together with forty-five vaccinees without previous infection but with two vaccinations. Sero-responses were evaluated using a total of six different assays: four measuring anti-Spike proteins (converted to BAU), one measuring anti-Nucleocapsid proteins and one SARS-CoV-2 surrogate virus neutralization. Both cohorts were evaluated using the Euroimmun Anti-SARS-CoV-2-ELISA anti-S1 IgG and the Roche Elecsys Anti-SARS-CoV-2 anti-S1 assay. RESULTS: In SARS-CoV-2-convalesce subjects, the BAU-sero-responses of Euroimmun Anti-SARS-CoV-2-ELISA anti-S1 IgG and Roche Elecsys Anti-SARS-CoV-2 anti-S1 peaked both at 47 (43-51) days, the first assay followed by a slow decay thereafter (> 208 days), while the second assay not presenting any decay within one year. Both assay values in BAUs are only equivalent a few months after infection, elsewhere correction factors up to 10 are necessary. In contrast, in infection-naive vaccinees the assays perform similarly. CONCLUSION: The results of our study suggest that the establishment of a protective correlate or vaccination booster recommendation based on different assays, although BAU-standardised, is still challenging. At the moment the characteristics of the available assays used are not related, and the BAU-standardisation is unable to correct for that.

Neutrophils Contribute to Severity of Tuberculosis Pathology and Recovery From Lung Damage Pre- and Posttreatment.

BACKGROUND: Despite microbiological cure, about 50% of tuberculosis (TB) patients have poor lung recovery. Neutrophils are associated with lung pathology; however, CD16/CD62L-defined subsets have not been studied in TB. Using flow cytometry, we monitored frequencies, phenotype, and function of neutrophils following stimulation with Mycobacterium tuberculosis (Mtb) whole cell lysate (WCL) and ESAT-6/CFP-10 fusion protein (EC) in relation to lung pathology. METHODS: Fresh blood from 42 adult, human immunodeficiency virus (HIV)-negative TB patients were analyzed pre- and post-therapy, with disease severity determined using chest radiography and bacterial load. Flow cytometry was used to monitor frequencies, phenotype, and function (generation of reactive oxygen species [ROS], together with CD11b, tumor necrosis factor, and interleukin 10 [IL-10] expression) of neutrophils following 2-hour stimulation with Mtb-specific antigens. RESULTS: Total neutrophils decreased by post-treatment compared to baseline (P = .0059); however, CD16brCD62Lbr (segmented) neutrophils increased (P = .0031) and CD16dimCD62Lbr (banded) neutrophils decreased (P = .038). Banded neutrophils were lower in patients with severe lung damage at baseline (P = .035). Following WCL stimulation, ROS from segmented neutrophils was higher in patients with low Mtb loads even after adjusting for sex (P = .038), whereas IL-10-expressing CD16dimCD62Llo cells were higher in patients with mild damage (P = .0397) at baseline. CONCLUSIONS: High ROS generation, low levels of banded neutrophils, and high levels of IL-10-expressing CD16dimCD62Llo neutrophils are associated with reduced lung pathology at diagnosis. Hence, neutrophils are potential early indicators of TB severity and promising targets for TB host-directed therapy.

Long-term follow-up on HIV infected and non-infected women with cervical cancer from Tanzania: staging, access to cancer-directed therapies and associated survival in a real-life remote setting.

BACKGROUND: Worldwide 85% of cervical cancer (CC) related deaths occur in low- and middle-income countries. Sub-Saharan Africa is burdend by an overlapping high incidence of CC as well as HIV infection, a risk factor for HPV associated disease progression. Recent upscaling of CC screening activities increased the number of CC diagnoses in a previous unscreened population. The aim of the 2H study was to follow up on women with CC in the context of available health care services in Tanzania in relation to their HIV infection status. METHODS: This longitudinal observational cohort study included women with histological confirmed CC from Mbeya, Tanzania, between 2013-2019. All women were referred for CC staging and cancer-directed therapies (CDT), including surgery and/or radio-chemotherapy, or palliative care. Annual follow-up focused on successful linkage to CDT, interventions and survival. We assessed factors on compliance, used Kaplan-Meier-Survivor functions to evaluate survival time and poisson regression models to calculate incidence rate ratios on mortality (IRR) two years after diagnosis. RESULTS: Overall, 270 women with CC (123 HIV infected) were included. Staging information, available in 185 cases, showed 84.9% presented with advanced stage disease (FIGO ≥ IIB), no difference was seen in respect to HIV status. HIV-infected women were 12 years younger at the time of cancer diagnosis (median age 44.8 versus 56.4 years, p < 0.001). Median follow up period was 11.9 months (range 0.2-67.2). Survival information, available in 231 cases, demonstrated for women diagnosed in early-stage disease a median survival time of 38.3 months, in advanced-stage 16.0 months and late-stage disease 6.5 months after diagnosis. Of all women, 42% received CDT or palliative support. HIV co-infection and education were associated with higher health care compliance. CDT was significantly associated with lower 2-year mortality rates (IRR 0.62, p = 0.004). HIV coinfection did not impact mortality rates after diagnosis. CONCLUSION: High numbers of advanced and late staged CC were diagnosed, compliance to CDT was low. A beneficial impact of CDT on CC mortality could be demonstrated for local health care services. This study indicates challenges for successful linkage and supports an effective scale up of cancer care and treatment facilities.

TB sequel: incidence, pathogenesis and risk factors of long-term medical and social sequelae of pulmonary TB - a study protocol.

BACKGROUND: Up to fifty percent of microbiologically cured tuberculosis (TB) patients may be left with permanent, moderate or severe pulmonary function impairment. Very few studies have systematically examined pulmonary outcomes in patients to understand the pathophysiologic basis and long-term socio-economic consequences of this injury. The planned multi-country, multi-centre observational TB cohort study, aims to advance the understanding of the clinical, microbiological, immunological and socio-economic risk factors affecting long-term outcome of pulmonary TB. It will also determine the occurrence of reversible and irreversible socio-economic consequences to patients, their households and the health sector related to pulmonary TB disease and its treatment. METHODS: We will enrol up to 1.600 patients with drug sensitive and multidrug-resistant pulmonary TB who are treated according to the local standard of care by the respective National TB Program. Recruitment is taking place at the time of TB diagnosis at four African study clinics located in The Gambia, Mozambique, South Africa and Tanzania. The primary outcome is the proportion of TB patients with severe lung impairment measured by spirometry at 24 months after TB treatment initiation. Biological samples, including sputum, urine and blood, for studying host- and pathogenic risk factors will be collected longitudinally and examined in a nested case-control fashion. A standardized quality of life questionnaire will be used together with a novel version of WHO's generic patient cost instrument which has been adapted for the longitudinal study design. DISCUSSION: This study is an integral part of an overall strategy to fill a knowledge gap needed to improve TB treatment outcomes globally. The main scientific goal is to identify the major pathogenic mechanisms associated with poor TB treatment outcomes, so that such pathways can be interrupted to avert long term TB sequelae. National as well as supra-national stakeholders and decision makers have been integrated early in the study planning process to inform future treatment guidelines and national health policies. TRIAL REGISTRATION: ClinicalTrials.gov: NCT03251196 , August 16, 2017.

Preferential Targeting of Conserved Gag Regions after Vaccination with a Heterologous DNA Prime-Modified Vaccinia Virus Ankara Boost HIV-1 Vaccine Regimen.

Prime-boost vaccination strategies against HIV-1 often include multiple variants for a given immunogen for better coverage of the extensive viral diversity. To study the immunologic effects of this approach, we characterized breadth, phenotype, function, and specificity of Gag-specific T cells induced by a DNA-prime modified vaccinia virus Ankara (MVA)-boost vaccination strategy, which uses mismatched Gag immunogens in the TamoVac 01 phase IIa trial. Healthy Tanzanian volunteers received three injections of the DNA-SMI vaccine encoding a subtype B and AB-recombinant Gagp37 and two vaccinations with MVA-CMDR encoding subtype A Gagp55 Gag-specific T-cell responses were studied in 42 vaccinees using fresh peripheral blood mononuclear cells. After the first MVA-CMDR boost, vaccine-induced gamma interferon-positive (IFN-γ+) Gag-specific T-cell responses were dominated by CD4+ T cells (P < 0.001 compared to CD8+ T cells) that coexpressed interleukin-2 (IL-2) (66.4%) and/or tumor necrosis factor alpha (TNF-α) (63.7%). A median of 3 antigenic regions were targeted with a higher-magnitude median response to Gagp24 regions, more conserved between prime and boost, compared to those of regions within Gagp15 (not primed) and Gagp17 (less conserved; P < 0.0001 for both). Four regions within Gagp24 each were targeted by 45% to 74% of vaccinees upon restimulation with DNA-SMI-Gag matched peptides. The response rate to individual antigenic regions correlated with the sequence homology between the MVA- and DNA Gag-encoded immunogens (P = 0.04, r2 = 0.47). In summary, after the first MVA-CMDR boost, the sequence-mismatched DNA-prime MVA-boost vaccine strategy induced a Gag-specific T-cell response that was dominated by polyfunctional CD4+ T cells and that targeted multiple antigenic regions within the conserved Gagp24 protein.IMPORTANCE Genetic diversity is a major challenge for the design of vaccines against variable viruses. While including multiple variants for a given immunogen in prime-boost vaccination strategies is one approach that aims to improve coverage for global virus variants, the immunologic consequences of this strategy have been poorly defined so far. It is unclear whether inclusion of multiple variants in prime-boost vaccination strategies improves recognition of variant viruses by T cells and by which mechanisms this would be achieved, either by improved cross-recognition of multiple variants for a given antigenic region or through preferential targeting of antigenic regions more conserved between prime and boost. Engineering vaccines to induce adaptive immune responses that preferentially target conserved antigenic regions of viral vulnerability might facilitate better immune control after preventive and therapeutic vaccination for HIV and for other variable viruses.

Effect of Wuchereria bancrofti infection on HIV incidence in southwest Tanzania: a prospective cohort study.

BACKGROUND: The past decades have seen an ongoing controversial debate about whether the immune activation induced by helminths has an effect on the susceptibility of individuals to HIV. In view of this, we assessed the effect of lymphatic filariasis, a chronic helminth disease elicited by Wuchereria bancrofti, on HIV incidence in southwest Tanzania. METHODS: In this population-based cohort study, we enrolled a geographically stratified randomly chosen sample of about 10% of the households in nine distinct sites in southwest Tanzania. All household members present were followed up and tested for HIV and circulating filarial antigen, an indicator of W bancrofti adult worm burden. Our main outcome of interest was HIV incidence in participants with or without lymphatic filariasis. FINDINGS: Between May 29, 2006, and June 16, 2011, we enrolled 4283 households with roughly 18 000 participants. Of these, 2699 individuals from Kyela district participated in at least one round of the EMINI study. In the 1055 initially HIV-negative adolescents and adults with clearly defined lymphatic filariasis status, 32 new HIV infections were observed in 2626 person-years. HIV incidence in lymphatic filariasis-positive participants (1·91 cases per 100 person-years) was significantly higher than the incidence in lymphatic filariasis-negative participants (0·80 cases per 100 person-years). The age-adjusted and sex-adjusted incidence rate ratio was 2·17 (95% CI 1·08-4·37, p=0·0300). Lymphatic filariasis status remained an independent and significantly relevant risk factor for HIV infection when controlled for other known risk factors such as sexual behaviour and socioeconomic factors. INTERPRETATION: To our knowledge, this is the first prospective study demonstrating a significantly increased risk of acquiring HIV for lymphatic filariasis-infected individuals. Immunological studies and interventional treatment studies that eliminate the adult worms and not only the microfilariae are needed to follow up on the results presented. FUNDING: European Union as part of EuropAid; German Federal Ministry of Education and Research; German Center for Infection Research.

CD25+ FoxP3+ Memory CD4 T Cells Are Frequent Targets of HIV Infection In Vivo.

UNLABELLED: Interleukin 2 (IL-2) signaling through the IL-2 receptor alpha chain (CD25) facilitates HIV replication in vitro and facilitates homeostatic proliferation of CD25(+) FoxP3(+) CD4(+) T cells. CD25(+) FoxP3(+) CD4(+) T cells may therefore constitute a suitable subset for HIV infection and plasma virion production. CD25(+) FoxP3(+) CD4(+) T cell frequencies, absolute numbers, and the expression of CCR5 and cell cycle marker Ki67 were studied in peripheral blood from HIV(+) and HIV(-) study volunteers. Different memory CD4(+) T cell subsets were then sorted for quantification of cell-associated HIV DNA and phylogenetic analyses of the highly variable EnvV1V3 region in comparison to plasma-derived virus sequences. In HIV(+) subjects, 51% (median) of CD25(+) FoxP3(+) CD4(+) T cells expressed the HIV coreceptor CCR5. Very high frequencies of Ki67(+) cells were detected in CD25(+) FoxP3(+) memory CD4(+) T cells (median, 27.6%) in comparison to CD25(-) FoxP3(-) memory CD4(+) T cells (median, 4.1%; P < 0.0001). HIV DNA content was 15-fold higher in CD25(+) FoxP3(+) memory CD4(+) T cells than in CD25(-) FoxP3(-) T cells (P = 0.003). EnvV1V3 sequences derived from CD25(+) FoxP3(+) memory CD4(+) T cells did not preferentially cluster with plasma-derived sequences. Quasi-identical cell-plasma sequence pairs were rare, and their proportion decreased with the estimated HIV infection duration. These data suggest that specific cellular characteristics of CD25(+) FoxP3(+) memory CD4(+) T cells might facilitate efficient HIV infection in vivo and passage of HIV DNA to cell progeny in the absence of active viral replication. The contribution of this cell population to plasma virion production remains unclear. IMPORTANCE: Despite recent advances in the understanding of AIDS virus pathogenesis, which cell subsets support HIV infection and replication in vivo is incompletely understood. In vitro, the IL-2 signaling pathway and IL-2-dependent cell cycle induction are essential for HIV infection of stimulated T cells. CD25(+) FoxP3(+) memory CD4 T cells, often referred to as regulatory CD4 T cells, depend on IL-2 signaling for homeostatic proliferation in vivo Our results show that CD25(+) FoxP3(+) memory CD4(+) T cells often express the HIV coreceptor CCR5, are significantly more proliferative, and contain more HIV DNA than CD25(-) FoxP3(-) memory CD4 T cell subsets. The specific cellular characteristics of CD25(+) FoxP3(+) memory CD4(+) T cells probably facilitate efficient HIV infection in vivo and passage of HIV DNA to cell progeny in the absence of active viral replication. However, the contribution of this cell subset to plasma viremia remains unclear.

Association between different anti-Tat antibody isotypes and HIV disease progression: data from an African cohort.

BACKGROUND: The presence of IgG and IgM against Tat, an HIV protein important for viral replication and immune dysfunction, is associated with slow disease progression in clade B HIV-infected individuals. However, although Tat activities strictly depend on the viral clade, our knowledge about the importance of anti-Tat antibodies in non-clade B HIV infection is poor. The objective of this study was to investigate the association of different anti-Tat antibody isotypes with disease progression in non-clade B HIV-infected subjects and to study the relationship between anti-Tat humoral responses and immunological abnormalities. METHODS: Anti-clade B and -clade C Tat IgG, IgM and IgA titers were assessed in serum samples from 96 cART-naïve subjects with chronic HIV infection from Mbeya, Tanzania, and associated with CD4(+) T cell count, plasma viremia and CD4(+) and CD8(+) T cell phenotypes. RESULTS: Anti-Tat IgM were preferentially detected in chronic HIV-infected subjects with low T cell activation (p-value = 0.03) and correlated with higher CD4(+) T cell counts and lower viral loads irrespective of the duration of infection (p-value = 0.019 and p-value = 0.037 respectively). Conversely, anti-Tat IgA were preferentially detected in individuals with low CD4(+) T cell counts and high viral load (p-value = 0.02 and p-value < 0.001 respectively). The simultaneous presence of anti-Tat IgG and IgM protected from fast CD4(+) T cell decline (p-value < 0.01) and accumulation of CD38(+)HLADR(+)CD8(+) T cells (p- value = 0.029). CONCLUSIONS: Anti-Tat IgG alone are not protective in non-clade B infected subjects, unless concomitant with IgM, suggesting a protective role of persistent anti-Tat IgM irrespective of the infecting clade.

Helicobacter pylori Coinfection Is Associated With Decreased Markers of Immune Activation in ART-Naive HIV-Positive and in HIV-Negative Individuals in Ghana.

BACKGROUND: Helicobacter pylori coinfection in human immunodeficiency virus (HIV) patients has been associated with higher CD4+ cell counts and lower HIV-1 viral loads, with the underlying mechanisms being unknown. The objective of this study was to investigate the impact of H. pylori infection on markers of T-cell activation in HIV-positive and HIV-negative individuals. METHODS: In a cross-sectional, observational study, HIV patients (n = 457) and HIV-negative blood donors (n = 79) presenting to an HIV clinic in Ghana were enrolled. Data on clinical and sociodemographic parameters, CD4+/CD8+ T-cell counts, and HIV-1 viral load were recorded. Helicobacter pylori status was tested using a stool antigen test. Cell surface and intracellular markers related to T-cell immune activation and turnover were quantified by flow cytometry and compared according to HIV and H. pylori status. RESULTS: Helicobacter pylori infection was associated with decreased markers of CD4+ T-cell activation (HLA-DR+CD38+CD4+; 22.55% vs 32.70%; P = .002), cell proliferation (Ki67; 15.10% vs 26.80%; P = .016), and immune exhaustion (PD-1; 32.45% vs 40.00%; P = .005) in 243 antiretroviral therapy (ART)-naive patients, but not in 214 patients on ART. In HIV-negative individuals, H. pylori infection was associated with decreased frequencies of activated CD4+ and CD8+ T cells (6.31% vs 10.40%; P = .014 and 18.70% vs 34.85%, P = .006, respectively). CONCLUSIONS: Our findings suggest that H. pylori coinfection effectuates a systemic immune modulatory effect with decreased T-cell activation in HIV-positive, ART-naive patients but also in HIV-negative individuals. This finding might, in part, explain the observed association of H. pylori infection with favorable parameters of HIV disease progression. CLINICAL TRIALS REGISTRATION: Clinicaltrials.gov NCT01897909.

Pathogen-specific T cell depletion and reactivation of opportunistic pathogens in HIV infection.

During HIV infection, it is unclear why different opportunistic pathogens cause disease at different CD4 T cell count thresholds. Early work has shown that CD4 T cell depletion is influenced both by cellular activation status and expression of viral entry receptors. More recently, functional characteristics of the CD4 T cells, such as cytokine and chemokine production, have also been shown to influence cellular susceptibility to HIV. Here, we examine how functional differences in pathogen-specific CD4 T cells could lead to their differential loss during HIV infection. This may have implications for when different opportunistic infections occur, and a better understanding of the mechanisms for functional imprinting of antigen-specific T cells may lead to improvements in design of vaccines against HIV and opportunistic pathogens.

Cytokine response to selected MTB antigens in Ghanaian TB patients, before and at 2 weeks of anti-TB therapy is characterized by high expression of IFN-γ and Granzyme B and inter- individual variation.

BACKGROUND: There has been a long held belief that patients with drug-susceptible TB are non-infectious after two weeks of therapy. Recent microbiological and epidemiological evidence has challenged this dogma, however, the nature of the Mtb-specific cellular immune response during this period has not been adequately investigated. This knowledge could be exploited in the development of immunological biomarkers of early treatment response. METHODS: Cellular response to four Mtb infection phase-dependent antigens, ESAT-6/CFP-10 fusion protein and three DosR encoded proteins (Rv1733c, Rv2029c, Rv2628) were evaluated in a Ghanaian TB cohort (n=20) before and after 2 weeks of anti TB therapy. After 6-days in vitro stimulation, Peripheral blood mononuclear cell (PBMC) culture supernatant was harvested and the concentration of IFN-γ, Granzyme B, IL-10, IL-17, sIL2Rα and TNF-α were determined in a 6-plex Luminex assay. Frequencies of IFN-γ + CD4 and CD8 T cells were also determined in an intracellular cytokine assay. RESULTS: All antigens induced higher levels of IFN-γ, followed by Granzyme B, TNF-α and IL-17 and low levels of IL-10 and sIL-2R-α in PBMC before treatment and after 2 weeks of treatment. Median cytokine levels of IFN-γ, Granzyme B, IL-17 and sIL-2R-α increased during week two, but it was significant for only Rv1733-specific production of Granzyme B (P = 0. 013). The median frequency of antigen specific IFN-γ + CD4 T cells increased at week two; however, only the increase in the ESAT-6/CFP-10-specific response was significant (P = 0. 0008). In contrast, the median frequency of ESAT-6/CFP-10- specific IFN-γ + CD8 T cell responses declined during week two (P = 0. 0024). Additionally, wide inter-individual variation with three distinct patterns were observed; increase in all cytokine levels, decrease in all cytokine levels and fluctuating cytokine levels after 2 weeks of treatment. CONCLUSION: The second week of effective chemotherapy was characterized by a general increase in cytokine response to Mtb-specific antigens suggestive of an improvement in cellular response with therapy. However, the wide inter-individual variation observed would limit the utility of cytokine biomarkers during this period.

Evolution of protective SARS-CoV-2-specific B and T cell responses upon vaccination and Omicron breakthrough infection.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) Omicron breakthrough infection (BTI) induced better protection than triple vaccination. To address the underlying immunological mechanisms, we studied antibody and T cell response dynamics during vaccination and after BTI. Each vaccination significantly increased peak neutralization titers with simultaneous increases in circulating spike-specific T cell frequencies. Neutralization titers significantly associated with a reduced hazard rate for SARS-CoV-2 infection. Yet, 97% of triple vaccinees became SARS-CoV-2 infected. BTI further boosted neutralization magnitude and breadth, broadened virus-specific T cell responses to non-vaccine-encoded antigens, and protected with an efficiency of 88% from further infections by December 2022. This effect was then assessed by utilizing mathematical modeling, which accounted for time-dependent infection risk, the antibody, and T cell concentration at any time point after BTI. Our findings suggest that cross-variant protective hybrid immunity induced by vaccination and BTI was an important contributor to the reduced virus transmission observed in Bavaria in late 2022 and thereafter.

Diagnostic accuracy of a three-gene Mycobacterium tuberculosis host response cartridge using fingerstick blood for childhood tuberculosis: a multicentre prospective study in low-income and middle-income countries.

BACKGROUND: Childhood tuberculosis remains a major cause of morbidity and mortality in part due to missed diagnosis. Diagnostic methods with enhanced sensitivity using easy-to-obtain specimens are needed. We aimed to assess the diagnostic accuracy of the Cepheid Mycobacterium tuberculosis Host Response prototype cartridge (MTB-HR), a candidate test measuring a three-gene transcriptomic signature from fingerstick blood, in children with presumptive tuberculosis disease. METHODS: RaPaed-TB was a prospective diagnostic accuracy study conducted at four sites in African countries (Malawi, Mozambique, South Africa, and Tanzania) and one site in India. Children younger than 15 years with presumptive pulmonary or extrapulmonary tuberculosis were enrolled between Jan 21, 2019, and June 30, 2021. MTB-HR was performed at baseline and at 1 month in all children and was repeated at 3 months and 6 months in children on tuberculosis treatment. Accuracy was compared with tuberculosis status based on standardised microbiological, radiological, and clinical data. FINDINGS: 5313 potentially eligible children were screened, of whom 975 were eligible. 784 children had MTB-HR test results, of whom 639 had a diagnostic classification and were included in the analysis. MTB-HR differentiated children with culture-confirmed tuberculosis from those with unlikely tuberculosis with a sensitivity of 59·8% (95% CI 50·8-68·4). Using any microbiological confirmation (culture, Xpert MTB/RIF Ultra, or both), sensitivity was 41·6% (34·7-48·7), and using a composite clinical reference standard, sensitivity was 29·6% (25·4-34·2). Specificity for all three reference standards was 90·3% (95% CI 85·5-94·0). Performance was similar in different age groups and by malnutrition status. Among children living with HIV, accuracy against the strict reference standard tended to be lower (sensitivity 50·0%, 15·7-84·3) compared with those without HIV (61·0%, 51·6-69·9), although the difference did not reach statistical significance. Combining baseline MTB-HR result with one Ultra result identified 71·2% of children with microbiologically confirmed tuberculosis. INTERPRETATION: MTB-HR showed promising diagnostic accuracy for culture-confirmed tuberculosis in this large, geographically diverse, paediatric cohort and hard-to-diagnose subgroups. FUNDING: European and Developing Countries Clinical Trials Partnership, UK Medical Research Council, Swedish International Development Cooperation Agency, Bundesministerium für Bildung und Forschung; German Center for Infection Research (DZIF).

Sequential and parallel testing for microbiological confirmation of tuberculosis disease in children in five low-income and middle-income countries: a secondary analysis of the RaPaed-TB study.

BACKGROUND: Despite causing high mortality worldwide, paediatric tuberculosis is often undiagnosed. We aimed to investigate optimal testing strategies for microbiological confirmation of tuberculosis in children younger than 15 years, including the yield in high-risk subgroups (eg, children younger than 5 years, with HIV, or with severe acute malnutrition [SAM]). METHODS: For this secondary analysis, we used data from RaPaed-TB, a multicentre diagnostic accuracy study evaluating novel diagnostic assays and testing approaches for tuberculosis in children recruited from five health-care centres in Malawi, Mozambique, South Africa, Tanzania, and India conducted between Jan 21, 2019, and June 30, 2021. Children were included if they were younger than 15 years and had signs or symptoms of pulmonary or extrapulmonary tuberculosis; they were excluded if they weighed less than 2 kg, had received three or more doses of anti-tuberculosis medication at time of enrolment, were in a condition deemed critical by the local investigator, or if they did not have at least one valid microbiological result. We collected tuberculosis-reference specimens via spontaneous sputum, induced sputum, gastric aspirate, and nasopharyngeal aspirates. Microbiological tests were Xpert MTB/RIF Ultra (hereafter referred to as Ultra), liquid culture, and Löwenstein-Jensen solid culture, which were followed by confirmatory testing for positive cultures. The main outcome of this secondary analysis was categorising children as having confirmed tuberculosis if culture or Ultra positive on any sample, unconfirmed tuberculosis if clinically diagnosed, and unlikely tuberculosis if neither of these applied. FINDINGS: Of 5313 children screened, 975 were enrolled, of whom 965 (99%) had at least one valid microbiological result. 444 (46%) of 965 had unlikely tuberculosis, 282 (29%) had unconfirmed tuberculosis, and 239 (25%) had confirmed tuberculosis. Median age was 5·0 years (IQR 1·8-9·0); 467 (48%) of 965 children were female and 498 (52%) were male. 155 (16%) of 965 children had HIV and 110 (11%) children had SAM. 196 (82%) of 239 children with microbiological detection tested positive on Ultra. 110 (46%) of 239 were confirmed by both Ultra and culture, 86 (36%) by Ultra alone, and 43 (18%) by culture alone. 'Trace' was the most common semiquantitative result (93 [40%] of 234). 481 (50%) of 965 children had only one specimen type collected, 99 (21%) of whom had M tuberculosis detected. 484 (50%) of 965 children had multiple specimens collected, 141 (29%) of whom were positive on at least one specimen type. Of the 102 children younger than 5 years with M tuberculosis detected, 80 (78%) tested positive on sputum. 64 (80%) of 80 children who tested positive on sputum were positive on sputum alone; 61 (95%) of 64 were positive on induced sputum, two (3%) of 64 were positive on spontaneous sputum, and one (2%) was positive on both. INTERPRETATION: High rates of microbiological confirmation of tuberculosis in children can be achieved via parallel sampling and concurrent testing procedures. Sample types and choice of test to be used sequentially should be considered when applying to groups such as children younger than 5 years, living with HIV, or with SAM. FUNDING: European and Developing Countries Clinical Trials Partnership programme, supported by the EU, the UK Medical Research Council, Swedish International Development Cooperation Agency, Bundesministerium für Bildung und Forschung, the German Center for Infection Research, and Beckman Coulter.

Clinical and immunological benefits of full primary COVID-19 vaccination in individuals with SARS-CoV-2 breakthrough infections: A prospective cohort study in non-hospitalized adults.

BACKGROUND: SARS-CoV-2 variants of concern (VOC) may result in breakthrough infections (BTIs) in vaccinated individuals. The aim of this study was to investigate the effects of full primary (two-dose) COVID-19 vaccination with wild-type-based SARS-CoV-2 vaccines on symptoms and immunogenicity of SARS-CoV-2 VOC BTIs. METHODS: In a longitudinal multicenter controlled cohort study in Bavaria, Germany, COVID-19 vaccinated and unvaccinated non-hospitalized individuals were prospectively enrolled within 14 days of a PCR-confirmed SARS-CoV-2 infection. Individuals were visited weekly up to 4 times, performing a structured record of medical data and viral load assessment. SARS-CoV-2-specific antibody response was characterized by anti-spike-(S)- and anti-nucleocapsid-(N)-antibody concentrations, anti-S-IgG avidity and neutralization capacity. RESULTS: A total of 300 individuals (212 BTIs, 88 non-BTIs) were included with VOC Alpha or Delta SARS-CoV-2 infections. Full primary COVID-19 vaccination provided a significant effectiveness against five symptoms (relative risk reduction): fever (33 %), cough (21 %), dysgeusia (22 %), dizziness (52 %) and nausea/vomiting (48 %). Full primary vaccinated individuals showed significantly higher 50 % inhibitory concentration (IC50) values against the infecting VOC compared to unvaccinated individuals at week 1 (269 vs. 56, respectively), and weeks 5-7 (1,917 vs. 932, respectively) with significantly higher relative anti-S-IgG avidity (78% vs. 27 % at week 4, respectively). CONCLUSIONS: Full primary COVID-19 vaccination reduced symptom frequencies in non-hospitalized individuals with BTIs and elicited a more rapid and longer lasting neutralization capacity against the infecting VOC compared to unvaccinated individuals. These results support the recommendation to offer at least full primary vaccination to all adults to reduce disease severity caused by immune escape-variants.

Therapeutic Efficacy of a VSV-GP-based Human Papilloma Virus Vaccine in a Murine Cancer Model.

Human papilloma virus (HPV) infections are associated with almost all cervical cancers and to a lower extend also with anogenital or oropharyngeal cancers. HPV proteins expressed in HPV-associated tumors are attractive antigens for cancer vaccination strategies as self-tolerance, which is associated with most endogenous tumor-associated antigens, does not need to be overcome. In this study, we generated a live attenuated cancer vaccine based on the chimeric vesicular stomatitis virus VSV-GP, which has previously proven to be a potent vaccine vector and oncolytic virus. Genes at an earlier position in the genome more to the 3' end are expressed stronger compared to genes located further downstream. By inserting an HPV16-derived antigen cassette consisting of E2, E6 and E7 into VSV-GP either at first (HPVp1) or fifth (HPVp5) position in VSV-GP's genome we aimed to analyze the effect of vaccine antigen position and consequently expression level on viral fitness, immunogenicity, and anti-tumoral efficacy in a syngeneic mouse tumor model. HPVp1 expressed higher amounts of HPV antigens compared to HPVp5 in vitro but had a slightly delayed replication kinetic which overall translated into increased HPV-specific T cell responses upon vaccination of mice. Immunization with both vectors protected mice in prophylactic and in therapeutic TC-1 tumor models with HPVp1 being more effective in the prophylactic setting. Taken together, VSV-GP is a promising candidate as therapeutic HPV vaccine and first position of the vaccine antigen in a VSV-derived vector seems to be superior to fifth position.

Impact of Omicron Variant Infection on Assessment of Spike-Specific Immune Responses Using the EUROIMMUN Quan-T-Cell SARS-CoV-2 Assay and Roche Elecsys Anti-SARS-CoV-2-S.

The currently prevailing variants of SARS-CoV-2 are subvariants of the Omicron variant. The aim of this study was to analyze the effect of mutations in the Spike protein of Omicron on the results Quan-T-Cell SARS-CoV-2 assays and Roche Elecsys anti-SARS-CoV-2 anti-S1. Omicron infected subjects ((n = 37), vaccinated (n = 20) and unvaccinated (n = 17)) were recruited approximately 3 weeks after a positive PCR test. The Quan-T-Cell SARS-CoV-2 assays (EUROIMMUN) using Wuhan and the Omicron adapted antigen assay and a serological test (Roche Elecsys anti-SARS-CoV-2 anti-S1) were performed. Using the original Wuhan SARS-CoV-2 IGRA TUBE, in 19 of 21 tested Omicron infected subjects, a positive IFNy response was detected, while 2 non-vaccinated but infected subjects did not respond. The Omicron adapted antigen tube resulted in comparable results. In contrast, the serological assay detected a factor 100-fold lower median Spike-specific RBD antibody concentration in non-vaccinated Omicron infected patients (n = 12) compared to patients from the pre Omicron era (n = 12) at matched time points, and eight individuals remained below the detection threshold for positivity. For vaccinated subjects, the Roche assay detected antibodies in all subjects and showed a 400 times higher median specific antibody concentration compared to non-vaccinated infected subjects in the pre-Omicron era. Our results suggest that Omicron antigen adapted IGRA stimulator tubes did not improve detection of SARS-CoV-2-specific T-cell responses in the Quant-T-Cell-SARS-CoV-2 assay. In non-vaccinated Omicron infected individuals, the Wuhan based Elecsys anti-SARS-CoV-2 anti-S1 serological assay results in many negative results at 3 weeks after diagnosis.

Increased HIV Incidence in Wuchereria bancrofti Microfilaria Positive Individuals in Tanzania.

BACKGROUND: Infections with Wuchereria bancrofti are associated with reduced immunity against concomitant infections. Indeed, our previous study described a 2.3-fold increased HIV incidence among individuals with W. bancrofti infection, as measured by the circulating filarial antigen of the adult worm. This new study aimed to retrospectively determine microfilariae status of the participants to assess if the previously described increased HIV susceptibility was associated with the presence of MF in the same cohort. METHODS: CFA positive but HIV negative biobanked human blood samples (n = 350) were analyzed for W. bancrofti MF chitinase using real time PCR. RESULTS: The PCR provided a positive signal in 12/350 (3.4%) samples. During four years of follow-up (1109 person years (PY)), 22 study participants acquired an HIV infection. In 39 PY of W. bancrofti MF chitinase positive individuals, three new HIV infections occurred (7.8 cases per 100 PY), in contrast to 19 seroconversions in 1070 PY of W. bancrofti MF chitinase negative individuals (1.8 cases per 100 PY, p = 0.014). CONCLUSIONS: In the subgroup of MF-producing Wb-infected individuals, the HIV incidence exceeded the previously described moderate increased risk for HIV seen in all Wb-infected individuals (regardless of MF status) compared with uninfected persons from the same area.