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African trypanosome parasites colonise the skin in a process important for parasite transmission. However, how the skin responses to trypanosome infection remain unresolved. Here, using a combination of spatial and single cell transcriptomics, coupled within vivogenetic models, we investigated the local immune response of the skin in both a murine model of infection. Our results provide several novel key findings previously unappreciated in the context of parasitic infections in the skin. First, we detected a significant expansion of a population of IL-17A-producing Vg6gdT cells in the infected murine skin compared to naïve controls that occur mainly in the subcutis, which we further validated at the protein level by flow cytometry. Second, interstitial preadipocytes located in the subcutis upregulate several genes involved in inflammatory signalling and antigen presentation, including T cell activation and survival.In silicocell-cell communication suggests that adipocytes triggergdT cell activation locallyvia Cd40, Il6, Il10,andTnfsf18signalling, amongst others. Third, mice deficient of IL-17A-producinggdT cells show extensive inflammation, increased frequency of dermal IFNg-producing CD8+T cells and limited subcutaneous adipose tissue wasting compared to wild-type infected controls, independent of TH1 CD4+T cells and parasite burden. Based on these observations, we proposed a model whereby adipocytes as well as Vg6gdT cells act concertedly in the subcutis to limit skin inflammation and tissue wasting. These studies shed light into the mechanisms ofgdT cell-mediated immunity in the skin in the context of African trypanosomes infection, as well as a potential role of immature and mature adipocytes as homeostatic regulators in the skin during chronic infection.&nbsp

The meningeal space is an important structure in the brain borders, which provides immunosurveillance for the central nervous system, but the impact of infections on the meningeal immune landscape is far from being fully understood.Trypanosoma brucei, the causative agents of Human African Trypanosomiasis or sleeping sickness, accumulates in the meningeal spaces, inducing severe meningitis and resulting in death if left untreated. Thus, sleeping sickness represents an attractive model to study immunological dynamics in the meninges during infection. Here, combining single cell transcriptomics and mass cytometry by time of flight (CyTOF), coupled within vivointerventions, we found that chronicT. bruceiinfection triggers the development of ectopic lymphoid aggregates (ELAs) in the murine meninges during chronic infection. These infection-induced ectopic structures are defined by the presence of ER-TR7+fibroblastic reticular cells (FRCs) and follicular dendritic cells (FDCs) that initiate a signalling cascade driving local T cell activation towards a T follicular helper (TFH)-like phenotype, as well as B cell class switching. Furthermore, the meningeal-resident plasma cells display activity typically associated with germinal centre reactions and are able to produce antibodies able to recognise mouse brain antigens, which correlates with cortical (and white matter) demyelination. Lastly, we found that systemic lymphotoxinb(LTb) signalling blockade led to a significant depletion of meningeal FDCs and autoreactive B cells, indicating that LTbsignalling is critical to induce and maintain local responses in the meninges. Taken together, we provide evidence that the meningeal immune response results in the acquisition of lymphoid tissue-like properties during chronicT. bruceiinfection. These findings have broader implications for understanding the mechanisms underlying the formation ELAs during chronic inflammation resulting in autoimmunity, as observed in other demyelinating neurodegenerative disorders such as multiple sclerosis.

The meningeal space is an important structure in the brain borders, which provides immunosurveillance for the central nervous system, but the impact of infections on the meningeal immune landscape is far from being fully understood. The extracellular protozoan parasite Trypanosoma brucei, which causes Human African Trypanosomiasis (HAT) or sleeping sickness, accumulate in the meningeal spaces, ultimately inducing severe meningitis and resulting in death if left untreated. Thus, sleeping sickness represents an attractive model to study immunological dynamics in the meninges during infection. Here, combining single cell transcriptomics and mass cytometry by time of flight (CyTOF), coupled with in vivo interventions, we found that chronic T. brucei infection triggers the development of ectopic lymphoid aggregates (ELAs) in the murine meninges during chronic infection. These infection-induced ectopic structures are defined by the presence of ER-TR7+ fibroblastic reticular cells (FRCs) and follicular dendritic cells (FDCs) that initiate a signalling cascade driving local T cell activation towards a T follicular helper (TFH)-like phenotype, as well as B cellclass switching. Furthermore, the GC-like B cells found in the infected meningesproduce high-affinity autoantibodies able to recognise mouse brain antigens. We found that systemic lymphotoxin β (LTβ) signalling blockade led to a significant depletion of meningeal FDC-like cells and autoreactive B cells, indicating that LTβ signalling is critical to induce and maintain local responses in the meninges. In humans, we identified the presence of autoreactive IgG antibodies able to recognise human brain lysates in the cerebrospinal fluid of second stage HAT patients compared to first stage HAT patients, consistent with our findings in experimental infections. Taken together, our data provide evidence that the meningeal immune response results in the acquisition of lymphoid tissue-like properties during chronic T. brucei infection, broadening our understanding of meningeal immunity in the context of chronic infections. These findings have wider implications for understanding the mechanisms underlying the formation ELAs during chronic inflammation resulting in autoimmunity in mice and humans, as observed in other autoimmune neurodegenerative disorders such as neuropsychiatric lupus and multiple sclerosis.

BackgroundHuman African trypanosomiasis (HAT) has caused social–economic burden in remote rural communities mostly in sub-Saharan Africa for over a century. The World Health Organization had targeted the year 2020 for the elimination of HAT caused by Trypanosoma brucei rhodesiense, which is mainly endemic in Malawi, Uganda, Tanzania, and Zambia. Significant progress has been made in reducing reported HAT cases in some countries. Area-specific updated epidemiological and clinical data may facilitate in understanding the progress of such efforts as well as the development of new intervention strategies.MethodsWe analyzed HAT prevalence and demographics from epidemiological surveys carried out from 2012 to 2020 obtained from the Ministry of Health, Malawi. In addition, we analyzed blood samples and clinical profiles of HAT patients surveyed between 2016 and 2020 from Rumphi and Nkhotakota districts. From the blood samples, parasite observations and speciation were carried out, whereas disease staging and severity were ascertained from the clinical profiles.ResultsMalawi reported 315 HAT cases from 2012 to 2020. The majority of HAT cases were men (70.2%), and the mean age was 29.9 ± 15.3 with all HAT fatalities resulting from stage 2 disease. Clinical symptoms were not significantly associated with disease outcome; however, swollen lymph nodes (p = 0.004), weight loss (p = 0.010), headache (p = 0.019), and sleep disturbance (p = 0.032) were significantly associated with the HAT stage of patients. About 50% of all HAT patients were reported within 2 years from 2019 to 2020, suggesting a HAT outbreak in Malawi.ConclusionThis study has highlighted the current epidemiological insights of the rHAT trend in Malawi. We have shown that rHAT clinical phenotypes in Malawi are focus-dependent and that there has been a steady increase in rHAT cases compared to all countries with incidences of rHAT. We have also highlighted an outbreak of rHAT that occurred in Malawi from 2019 to 2020 with almost 50% of the total rHAT cases that we have presented in this study reported within 2 years of the outbreak. These should call for a review of Malawi’s rHAT control and elimination strategies. A One-Health approach with the inclusion of key stakeholders such as the department of parks and wildlife may also be considered.

Background The diagnosis of gambiense human African trypanosomiasis (gHAT) typically involves 2 steps: a serological screen, followed by the detection of living trypanosome parasites in the blood or lymph node aspirate. Live parasites can, however, remain undetected in some seropositive individuals, who, we hypothesize, are infected with Trypanosoma brucei gambiense parasites in their extravascular dermis. Methods To test this hypothesis, we conducted a prospective observational cohort study in the gHAT focus of Forecariah, Republic of Guinea. Of the 5417 subjects serologically screened for gHAT, 66 were enrolled into our study and underwent a dermatological examination. At enrollment, 11 seronegative, 8 unconfirmed seropositive, and 18 confirmed seropositive individuals had blood samples and skin biopsies taken and examined for trypanosomes by molecular and immunohistological methods. Results In seropositive individuals, dermatological symptoms were significantly more frequent, relative to seronegative controls. T.b. gambiense parasites were present in the blood of all confirmed cases (n = 18) but not in unconfirmed seropositive individuals (n = 8). However, T. brucei parasites were detected in the extravascular dermis of all unconfirmed seropositive individuals and all confirmed cases. Skin biopsies of all treated cases and most seropositive untreated individuals progressively became negative for trypanosomes 6 and 20 months later. Conclusions Our results highlight the skin as a potential reservoir for African trypanosomes, with implications for our understanding of this disease’s epidemiology in the context of its planned elimination and underlining the skin as a novel target for gHAT diagnostics., Live trypanosomes can remain undetected in the blood of individuals seropositive for sleeping sickness. Here, we show that they could be infected with parasites in their extravascular dermis, highlighting the skin as a potential reservoir for trypanosomes.