HIV-1-infected persons are at higher risk of lower respiratory tract infections than HIV-1-uninfected individuals. This suggests strongly that HIV-infected persons have specific impairment of pulmonary immune responses, but current understanding of how HIV alters pulmonary immunity is incomplete. Alveolar macrophages (AMs), comprising small and large macrophages, are major effectors of innate immunity in the lung. We postulated that HIV-1 impairs pulmonary innate immunity through impairment of AM physiological functions. AMs were obtained by bronchoalveolar lavage from healthy, asymptomatic, antiretroviral therapy-naive HIV-1-infected and HIV-1-uninfected adults. We used novel assays to detect in vivo HIV-infected AMs and to assess AM functions based on the HIV infection status of individual cells. We show that HIV has differential effects on key AM physiological functions, whereby small AMs are infected preferentially by the virus, resulting in selective impairment of phagocytic function. In contrast, HIV has a more generalized effect on AM proteolysis, which does not require direct viral infection. These findings provide new insights into how HIV alters pulmonary innate immunity and the phenotype of AMs that harbors the virus. They underscore the need to clear this HIV reservoir to improve pulmonary immunity and reduce the high incidence of lower respiratory tract infections in HIV-1-infected individuals.

Background In low-income countries, like Malawi, important public health measures including social distancing or a lockdown, have been challenging to implement owing to socioeconomic constraints, leading to predictions that the COVID-19 pandemic would progress rapidly. However, due to limited capacity to test for severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection, there are no reliable estimates of the true burden of infection and death. We, therefore, conducted a SARS-CoV-2 serosurvey amongst health care workers (HCW) in Blantyre city to estimate the cumulative incidence of SARS-CoV-2 infection in urban Malawi. Methods Five hundred otherwise asymptomatic HCWs were recruited from Blantyre City (Malawi) from 22nd May 2020 to 19th June 2020 and serum samples were collected all participants. A commercial ELISA was used to measure SARS-CoV-2 IgG antibodies in serum. We run local negative samples (2018 - 2019) to verify the specificity of the assay. To estimate the seroprevalence of SARS CoV-2 antibodies, we adjusted the proportion of positive results based on local specificity of the assay. Results Eighty-four participants tested positive for SARS-CoV-2 antibodies. The HCW with a positive SARS-CoV-2 antibody result came from different parts of the city. The adjusted seroprevalence of SARS-CoV-2 antibodies was 12.3% [CI 9.0-15.7]. Using age-stratified infection fatality estimates reported from elsewhere, we found that at the observed adjusted seroprevalence, the number of predicted deaths was 8 times the number of reported deaths. Conclusion The high seroprevalence of SARS-CoV-2 antibodies among HCW and the discrepancy in the predicted versus reported deaths, suggests that there was early exposure but slow progression of COVID-19 epidemic in urban Malawi. This highlights the urgent need for development of locally parameterised mathematical models to more accurately predict the trajectory of the epidemic in sub-Saharan Africa for better evidence-based policy decisions and public health response planning.

Controlled human infection model (CHIM) studies have pivotal importance in vaccine development, being useful for proof of concept, pathogenesis, down-selection and immunogenicity studies.  To date, however, they have seldom been carried out in low and middle income countries (LMIC), which is where the greatest burden of vaccine preventable illness is found.  This workshop discussed the benefits and barriers to CHIM studies in Malawi.  Benefits include improved vaccine effectiveness and host country capacity development in clinical, laboratory and governance domains.  Barriers include acceptability, safety and regulatory issues. The report suggests a framework by which ethical, laboratory, scientific and governance issues may be addressed by investigators considering or planning CHIM in LMIC.

Pneumococcal carriage is both immunising and a pre-requisite for mucosal and systemic disease. Murine models of pneumococcal colonisation show that IL-17A-secreting CD4+ T-cells (Th-17 cells) are essential for clearance of pneumococci from the nasopharynx. Pneumococcal-responding IL-17A-secreting CD4+ T-cells have not been described in the adult human lung and it is unknown whether they can be elicited by carriage and protect the lung from pneumococcal infection. We investigated the direct effect of experimental human pneumococcal nasal carriage (EHPC) on the frequency and phenotype of cognate CD4+ T-cells in broncho-alveolar lavage and blood using multi-parameter flow cytometry. We then examined whether they could augment ex vivo alveolar macrophage killing of pneumococci using an in vitro assay. We showed that human pneumococcal carriage leads to a 17.4-fold (p = 0.007) and 8-fold (p = 0.003) increase in the frequency of cognate IL-17A+ CD4+ T-cells in BAL and blood, respectively. The phenotype with the largest proportion were TNF+/IL-17A+ co-producing CD4+ memory T-cells (p<0.01); IFNγ+ CD4+ memory T-cells were not significantly increased following carriage. Pneumococci could stimulate large amounts of IL-17A protein from BAL cells in the absence of carriage but in the presence of cognate CD4+ memory T-cells, IL-17A protein levels were increased by a further 50%. Further to this we then show that alveolar macrophages, which express IL-17A receptors A and C, showed enhanced killing of opsonised pneumococci when stimulated with rhIL-17A (p = 0.013). Killing negatively correlated with RC (r = −0.9, p = 0.017) but not RA expression. We conclude that human pneumococcal carriage can increase the proportion of lung IL-17A-secreting CD4+ memory T-cells that may enhance innate cellular immunity against pathogenic challenge. These pathways may be utilised to enhance vaccine efficacy to protect the lung against pneumonia.

RationaleHIV-infected adults are at an increased risk of lower respiratory tract infections. HIV infection impairs systemic acquired immunity, but there is limited information in humans on HIV-related cell-mediated immune defects in the lung.ObjectiveTo investigate antigen-specific CD4+ T cell responses to influenza virus, Streptococcus pneumoniae and Mycobacterium tuberculosis antigens in bronchoalveolar lavage (BAL) and peripheral blood between HIV-infected individuals and HIV-uninfected Malawian adults.MethodsWe obtained BAL fluid and blood from HIV-infected individuals (n=21) and HIV-uninfected adults (n=24). We determined the proportion of T cell subsets including naive, memory and regulatory T cells using flow cytometry, and used intracellular cytokine staining to identify CD4+ T cells recognising influenza virus-, S pneumoniae- and M tuberculosis-antigens.Main resultsCD4+ T cells in BAL were predominantly of effector memory phenotype compared to blood, irrespective of HIV status (p<0.001). There was immune compartmentalisation with a higher frequency of antigen-specific CD4+ T cells against influenza virus, S pneumoniae and M tuberculosis retained in BAL compared to blood in HIV-uninfected adults (p<0.001 in each case). Influenza virus- and M tuberculosis-specific CD4+ T cell responses in BAL were impaired in HIV-infected individuals: proportions of total antigen-specific CD4+ T cells and of polyfunctional IFN-γ and TNF-α-secreting cells were lower in HIV-infected individuals than in HIV-uninfected adults (p<0.05 in each case).ConclusionsBAL antigen-specific CD4+ T cell responses against important viral and bacterial respiratory pathogens are impaired in HIV-infected adults. This might contribute to the susceptibility of HIV-infected adults to lower respiratory tract infections such as pneumonia and tuberculosis.

Alveolar macrophages (AM) are critical to the homeostasis of the inflammatory environment in the lung. Differential expression of surface markers classifies macrophages to either classically (M1) or alternatively activated (M2). We investigated the phenotype of human alveolar macrophages (AM) in adults living in two different geographical locations: UK and Malawi. We show that the majority of AM express high levels of M1 and M2 markers simultaneously, with the M1/M2 phenotype being stable in individuals from different geographical locations. The combined M1/M2 features confer to AM a hybrid phenotype, which does not fit the classic macrophage classification. This hybrid phenotype may confer to alveolar macrophages an ability to quickly switch between M1 or M2 associated functions allowing for appropriate responses to stimuli and tissue environment.Electronic supplementary materialThe online version of this article (10.1186/s12931-018-0777-0) contains supplementary material, which is available to authorized users.

In this study, we detail a novel approach that combines bacterial fitness fluorescent reporter strains with scRNA-seq to simultaneously acquire the host transcriptome, surface marker expression, and bacterial phenotype for each infected cell. This approach facilitates the dissection of the functional heterogeneity of M. tuberculosis–infected alveolar (AMs) and interstitial macrophages (IMs) in vivo. We identify clusters of pro-inflammatory AMs associated with stressed bacteria, in addition to three different populations of IMs with heterogeneous bacterial phenotypes. Finally, we show that the main macrophage populations in the lung are epigenetically constrained in their response to infection, while inter-species comparison reveals that most AMs subsets are conserved between mice and humans. This conceptual approach is readily transferable to other infectious disease agents with the potential for an increased understanding of the roles that different host cell populations play during the course of an infection.

Infectious challenge of the human nasal mucosa elicits immune responses that determine the fate of the host-bacterial interaction; leading either to clearance, colonisation and/or disease. Persistent antigenic exposure from pneumococcal colonisation can induce both humoral and cellular defences that are protective against carriage and disease. We challenged healthy adults intra-nasally with live 23F or 6B Streptococcus pneumoniae in two sequential cohorts and collected nasal wash, bronchoalveolar lavage (BAL) and blood before and 6 weeks after challenge. We hypothesised that both cohorts would successfully become colonised but this did not occur except for one volunteer. The effect of bacterial challenge without colonisation in healthy adults has not been previously assessed. We measured the antigen-specific humoral and cellular immune responses in challenged but not colonised volunteers by ELISA and Flow Cytometry. Antigen-specific responses were seen in each compartment both before and after bacterial challenge for both cohorts. Antigen-specific IgG and IgA levels were significantly elevated in nasal wash 6 weeks after challenge compared to baseline. Immunoglobulin responses to pneumococci were directed towards various protein targets but not capsular polysaccharide. 23F but not 6B challenge elevated IgG anti-PspA in BAL. Serum immunoglobulins did not increase in response to challenge. In neither challenge cohort was there any alteration in the frequencies of TNF, IL-17 or IFNγ producing CD4 T cells before or after challenge in BAL or blood. We show that simple, low dose mucosal exposure with pneumococci may immunise mucosal surfaces by augmenting anti-protein immunoglobulin responses; but not capsular or cellular responses. We hypothesise that mucosal exposure alone may not replicate the systemic immunising effect of experimental or natural carriage in humans.