SARS-CoV-2 has emerged as a human pathogen, causing clinical signs, from fever to pneumonia-COVID-19-but may remain mild or asymptomatic. To understand the continuing spread of the virus, to detect those who are and were infected, and to follow the immune response longitudinally, reliable and robust assays for SARS-CoV-2 detection and immunological monitoring are needed. We quantified IgM, IgG, and IgA antibodies recognizing the SARS-CoV-2 receptor-binding domain (RBD) or the Spike (S) protein over a period of 6 months following COVID-19 onset. We report the detailed setup to monitor the humoral immune response from over 300 COVID-19 hospital patients and healthcare workers, 2500 University staff, and 198 post-COVID-19 volunteers. Anti-SARS-CoV-2 antibody responses follow a classic pattern with a rapid increase within the first three weeks after symptoms. Although titres reduce subsequently, the ability to detect anti-SARS-CoV-2 IgG antibodies remained robust with confirmed neutralization activity for up to 6 months in a large proportion of previously virus-positive screened subjects. Our work provides detailed information for the assays used, facilitating further and longitudinal analysis of protective immunity to SARS-CoV-2. Importantly, it highlights a continued level of circulating neutralising antibodies in most people with confirmed SARS-CoV-2.

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Background. SARS-CoV-2 infected cancer patients (CP) show worse outcomes compared with non-cancer patients (NCP). The humoral immune response (HIR) of CP against SARS-CoV-2 is not well characterized. To better understand it, we conducted a serological study of hospitalized SARS-CoV-2 CP. Materials and Methods. Unicentric, retrospective study enrolling adult SARS-CoV-2 patients admitted to a central hospital from March 15 to June 17, 2020, whose serum samples were quantified for anti-SARS-CoV-2 receptorbinding domain or spike protein immunoglobulin (Ig) M, G and A antibodies. The aims of the study were to assess the HIR to SARS-CoV-2, correlate it with different cancer types, stages and treatments, clarify the interplay between HIR of CP and clinical outcomes and to compare the HIR of SARS-CoV-2 CP and NCP. Results. We included 72 SARS-CoV-2 positive subjects (19 CP, 53 controls). About 90% of controls revealed a robust serological response. Among CP, a strong response was verified in 57.9% of them, with 42.1% showing a persistently weak response. Treatment with chemotherapy within 14 days before positivity was the only factor statistically shown to be associated with persistently weak serological responses among CP. No significant differences in outcomes were observed between CP with strong and weak responses. All IgG, IgM, IgA and total Ig antibody titers were significantly lower in CP when compared with NCP. Conclusions. A significant part of CP develops a proper HIR. Recent chemotherapy treatment may be associated with weak serological responses among CP. CP have a weaker SARS-CoV-2 antibody response compared with NCP. Implications for practice. Our results place the spotlight on cancer patients, particularly the ones actively treated with chemotherapy. These patients may potentially be more vulnerable to SARS-CoV-2 infection, being important to provide oncologists further theoretical support (with concrete examples and respective mechanistic correlations) for the decision of starting, maintaining or stopping antineoplastic treatments (particularly chemotherapy) not only on noninfected but also on infected cancer patients in accordance with cancer type, stage and prognosis, treatment agents, treatment setting and SARS-CoV-2 infection risks. The Oncologist ;9999:• •

SARS-CoV-2 has emerged as a novel human pathogen, causing clinical signs, from fever to pneumonia - COVID-19 - but may remain mild or even asymptomatic. To understand the continuing spread of the virus, to detect those who are and were infected, and to follow the immune response longitudinally, reliable and robust assays for SARS-CoV-2 detection and immunological monitoring are needed and have been setup around the world. We quantified immunoglobulin M (IgM), IgG and IgA antibodies recognizing the SARS-CoV-2 receptor-binding domain (RBD) or the Spike (S) protein over a period of five months following COVID-19 disease onset or in previously SARS-CoV-2 PCR-positive volunteers. We report the detailed setup to monitor the humoral immune response from over 300 COVID-19 hospital patients and healthcare workers, 2500 University staff and 187 post-COVID19 volunteers, and assessing titres for IgM, IgG and IgA. Anti-SARS-CoV-2 antibody responses followed a classic pattern with a rapid increase within the first three weeks after symptoms. Although titres reduce from approximately four weeks, the ability to detect SARS-CoV-2 antibodies remained robust for five months in a large proportion of previously virus-positive screened subjects. Our work provides detailed information for the assays used, facilitating further and longitudinal analysis of protective immunity to SARS-CoV-2. Moreover, it highlights a continued level of circulating neutralising antibodies in most people with confirmed SARS-CoV-2, at least up to five months after infection.

The metabolic capacity of many cells is tightly regulated and can adapt to changes in metabolic resources according to environmental changes. Tissue-resident memory (T RM ) CD8 + T cells are one of the most abundant T cell populations and offer rapid protection against invading pathogens, especially at the epithelia. T RM cells metabolically adapt to their tissue niche, such as the intestinal epithelial barrier. In the small intestine, the types of T RM cells are intraepithelial lymphocytes (IELs), which contain high levels of cytotoxic molecules and express activation markers, suggesting a heightened state of activation. We hypothesize that the tissue environment may determine IEL activity. We show that IEL activation, in line with its semiactive status, is metabolically faster than circulating CD8 + T cells. IEL glycolysis and oxidative phosphorylation (OXPHOS) are interdependently regulated and are dependent on rapid access to metabolites from the environment. IELs are restrained by local availability of metabolites, but, especially, glucose levels determine their activity. Importantly, this enables functional control of intestinal T RM cells by metabolic means within the fragile environment of the intestinal epithelial barrier.

Eimeria vermiformis is a tissue specific, intracellular protozoan that infects the murine small intestinal epithelia, which has been widely used as a coccidian model to study mucosal immunology. This mouse infection model is valuable to investigate the mechanisms of host protection against primary and secondary infection in the small intestine. Here, we describe the generation of an E. vermiformis stock solution, preparation of sporulated E. vermiformis to infect mice and determination of oocysts burden. This protocol should help to establish a highly reproducible natural infection challenge model to study immunity in the small intestine. The information obtained from using this mouse model can reveal fundamental mechanisms of interaction between the pathogen and the immune response, e.g., provided by intraepithelial lymphocytes (IEL) at the basolateral site of epithelial cells but also a variety of other immune cell populations present in the gut.

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Effective antibody responses are essential to generate protective humoral immunity. Different inflammatory signals polarize T cells towards an appropriate effector phenotype during an infection or immunization. Th1 and Th2 cells have been associated with the polarization of humoral responses for several decades. However, it is now established that T follicular helper cells (Tfh) have a unique ability to access the B cell follicle and support the Germinal Centre (GCs) responses by providing help to B cells. We investigated the specialization of Tfh cells induced under type-1 and type-2 conditions. We first studied homogenous Tfh cell populations generated by adoptively transferred TCR-transgenic T cells in mice immunized with type-1 and type-2 adjuvants. Using a machine learning approach, we established a gene expression signature that discriminates Tfh cells polarized towards type-1 and type-2 response, defined as Tfh1 and Tfh2 cells. The Tfh1 and Tfh2 distinct signature was validated against datasets of Tfh cells induced following LCMV or helminth infection. Using single-cell transcriptomics, we also dissected the heterogeneity of Tfh cells from the two immunizing conditions. Our results show that Tfh cells acquire a specialized function under distinct types of immune responses, but with the coexistence of a small population of Tfh cells of the alternative type. Furthermore, the specific molecular hallmarks of Tfh1 and Tfh2 cells identified herein offer putative new targets for tuning humoral responses.