Intermediate monocytes expansion and homing markers expression in COVID-19 patients associate with kidney dysfunction (preprint)

Patients with severe SARS-CoV-2 infection have an overwhelming inflammatory response characterized by remarkable organs monocyte infiltration. We performed an immunophenotypic analysis on circulating monocytes in 19 COVID-19 patients in comparison with 11 naïve HIV-1 patients and 10 healthy subjects. Reduced frequency of classical monocytes and increased frequency of intermediate monocytes characterized COVID-19 patients with respect to both HIV naïve patients and healthy subjects. Intensity of C-C motif chemokine receptor 2 (CCR2) monocyte expression highly correlated with parameters of kidney dysfunction. Our data indicate that highly activated monocytes of COVID-19 patients may be pathogenically associated to the development of renal disease.

High temporal resolution transcriptomic profiling delineates distinct patterns of interferon response following Covid-19 mRNA vaccination and SARS-CoV2 infection (preprint)

Knowledge of the factors contributing to the development of protective immunity after vaccination with COVID-19 mRNA vaccines is fragmentary. Thus we employed high-temporal-resolution transcriptome profiling and in-depth characterization of antibody production approaches to investigate responses to COVID-19 mRNA vaccination. There were marked differences in the timing and amplitude of the responses to the priming and booster doses. Notably, two distinct interferon signatures were identified, that differed based on their temporal patterns of induction. The first signature (S1), which was preferentially induced by type I interferon, peaked at day 2 post-prime and at day 1 post-boost, and in both instances was associated with subsequent development of the antibody response. In contrast, the second interferon signature (S2) peaked at day 1 both post-prime and post-boost but was found to be potently induced only post-boost, where it coincided with a robust inflammation peak. Notably, we also observed post-prime-like (S1++,S20/+) and post-boost-like (S1++,S2++) patterns of interferon response among COVID-19 patients. A post-boost-like signature was observed in most severely ill patients at admission to the intensive care unit and was associated with a shorter hospital stay. Interestingly, severely ill patients who stayed hospitalized the longest showed a peculiar pattern of interferon induction (S1-/0,S2+), that we did not observe following the administration of mRNA vaccines. In summary, high temporal resolution profiling revealed an elaborate array of immune responses elicited by priming and booster doses of COVID-19 mRNA vaccines. Furthermore, it contributed to the identification of distinct interferon-response phenotypes underpinning vaccine immunogenicity and the course of COVID-19 disease.

The longest persistence of viable SARS-CoV-2 with recurrence of viremia and relapsing symptomatic COVID-19 in an immunocompromised patient - a case study (preprint)

BackgroundImmunocompromised patients show prolonged shedding of SARS-CoV-2 in nasopharyngeal swabs. We report a case of a prolonged persistence of viable SARS-CoV-2 associated with clinical relapses of COVID-19 in a lymphoma patient. MethodsNasopharyngeal swabs and blood samples were tested for SARS-CoV-2 by Real time-PCR (RT-PCR). On five positive nasopharyngeal swabs, we performed viral culture and next generation sequencing. We analysed the patients adaptive and innate immunity to characterize T and NK cell subsets. FindingsSARS-CoV-2 RT-PCR on nasopharyngeal swabs samples remained positive with cycle threshold mean values of 22 {+/-} 1{middle dot}3 for over 8 months. All five performed viral cultures were positive and genomic analysis confirmed a persistent infection with the same strain. Viremia resulted positive in three out of four COVID-19 clinical relapses and cleared each time after remdesivir treatment. T and NK cells dynamic was different in aviremic and viremic samples and no SARS-CoV-2 specific antibodies were detected throughout the disease course. InterpretationIn our patient, SARS-CoV-2 persisted with proven infectivity for over eight months. Viremia was associated with COVID-19 relapses and remdesivir treatment was effective in viremia clearance and symptoms remission, although it was unable to clear the virus from the upper respiratory airways. During the viremic phase, we observed a low frequency of terminal effector CD8+ T lymphocytes in peripheral blood that are probably recruited in inflammatory tissue for viral eradication. In addition we found a high level of NK cells repertoire perturbation with a relevant involvement during SARS-CoV-2 viremia. FundingNone.

Bronchoalveolar lavage fluid characteristics and outcomes of invasively mechanically ventilated patients with COVID-19 pneumonia in Genoa, Italy. (preprint)

Background To describe the cellular characteristics of bronchoalveolar lavage fluid (BALF) of critically ill COVID-19 patients requiring invasive mechanical ventilation; the secondary outcome is to describe BALF findings between survivors vs non-survivors.Materials and Methods Patients positive for SARS-CoV-2 RT PCR, admitted to ICU between March and April 2020 were enrolled. At ICU admission, BALF were analyzed by flow cytometry. Univariate, multivariate and Spearman correlation analyses were performed.Results Sixty-four patients were enrolled, median age of 64 years (IQR 58–69). The majority cells in the BALF were neutrophils (70%, IQR 37.5–90.5) and macrophages (27%, IQR 7–49) while a minority were lymphocytes, 1%, TCD3 + 92% (IQR 82–95). The ICU mortality was 32.8%. Non-survivors had a significantly older age (p = 0.033) and peripheral lymphocytes (p = 0.012) were lower compared to the survivors. At multivariate analysis the percentage of macrophages in the BALF correlated with poor outcome (OR 1.336, CI95% 1.014–1.759, p = 0.039).Conclusions In critically ill patients, BALF cellularity is mainly composed of neutrophils and macrophages. The macrophages percentage in the BALF at ICU admittance correlated with higher ICU mortality. The lack of lymphocytes in BALF could partly explain a reduced anti-viral response.

Clinical Efficacy of Eucaloric Ketogenic Nutrition in the COVID-19 Cytokine Storm: a Retrospective Analysis of Mortality and ICU Admission (preprint)

Background: Some patients affected by COVID-19 present a life-threatening hyperinflammatory state known as cytokine storm syndrome (CSS) associated with a high mortality rate. Our hypothesis is that a eucaloric ketogenic diet (EKD) may be a safe and efficacious treatment option to reduce CSS and consequently to reduce the need for CPAP, ICU admission and COVID-19 mortality.Aim of the study: The primary objective is to explore the effect of an EKD on mortality, admission to the ICU and the need for NIV in hospitalized patients with COVID-19 in comparison to a eucaloric standard diet (ESD). The secondary objectives are to collect data about the safety and feasibility of an EKD during hospitalization and to evaluate the effect of the diet on biological and inflammatory parameters, particularly interleukin-6 (IL-6).Patients and methods: The study is a retrospective explorative analysis of 34 patients fed with an EKD during hospitalization for COVID-19 in comparison to 68 patients fed an ESD selected and matched using propensity score one-to-two to avoid the confounding effect of interfering variables.Results: A trend of reduced 30-day mortality (HR 0.416, 95% CI 0.122 – 1.413, P = 0.160) and a trend regarding the need for ICU admission (HR 0.357, 95% CI 0.045 – 2.847, P = 0.331) were observed in subjects treated with the EKD compared to patients fed with the standard diet. No significantly different risks in the need for CPAP (HR 0.968, CI 0.289 – 3.242, P = 0.958 for EKD) or the composite endpoint (HR 0.674, CI 0.233 – 1.949, P = 0.446 for EKD) were detectable between the two groups of dietary patterns.Furthermore, IL-6 concentrations between t 0 and t 7 (seven days after the beginning of the diet) in the ketogenic nutrition group showed a median difference of -26.0 ȵg/mL and a mean difference of -164 ȵg/mL (data from 23 of the 34 pairs) compared to controls, with a trend toward significance (P = 0.062). EKD was safe and no adverse events were observed in patients fed an EKD.Discussion and conclusions: These preliminary data on the clinical results for mortality, need for ICU admission and the effect on the IL-6 concentration during EKD feeding, collected in a retrospective way during the most aggressive period of the COVID-19 pandemic, suggest a favorable role of this dietary treatment in COVID-19 clinical management. The EKD was safe and well accepted by patients during hospitalization and seems to be an interesting tool in controlling COVID-19 CSS. The results of the prospective controlled randomized trial, currently underway with a large number of subjects, are necessary to confirm these preliminary data.

Tocilizumab and steroid treatment in patients with severe Covid-19 pneumonia. (preprint)

IntroductionCoronavirus disease 2019 (COVID-19) can lead to respiratory failure due to severe immune response. Treatment targeting this immune response might be beneficial but there is limited evidence on its efficacy. The aim of this study was to determine if early treatment of patients with COVID-19 pneumonia with tocilizumab and/or steroids was associated with better outcome. MethodsThis observational single-center study included patients with COVID-19 pneumonia who were not intubated and received either standard of care (SOC, controls) or SOC plus early (within 3 days from hospital admission) anti-inflammatory treatment. SOC consisted of hydroxychloroquine 400mg bid plus, in those admitted before March 24th, also darunavir/ritonavir. Anti-inflammatory treatment consisted of either tocilizumab (8mg/kg intravenously or 162mg subcutaneously) or methylprednisolone 1 mg/kg for 5 days or both. Failure was defined as intubation or death, and the endpoints were failure-free survival (primary endpoint) and overall survival (secondary) at day 30. Difference between the groups was estimated as Hazard Ratio by a propensity score weighted Cox regression analysis (HROW). ResultsOverall, 196 adults were included in the analyses. They were mainly male (67.4%), with comorbidities (78.1%) and severe COVID-19 pneumonia (83.7%). Median age was 67.9 years (range, 30-100) and median PaO2/FiO2 200 mmHg (IQR 133-289). Among them, 130 received early anti-inflammatory treatment with: tocilizumab (n=29, 22.3%), methylprednisolone (n=45, 34.6%), or both (n=56, 43.1%). The adjusted failure-free survival among tocilizumab/methylprednisolone/SOC treated patients vs. SOC was 80.8% (95%CI, 72.8-86.7) vs. 64.1% (95%CI, 51.3-74.0), HROW 0.48, 95%CI, 0.23-0.99; p=0.049. The overall survival among tocilizumab/methylprednisolone/SOC patients vs. SOC was 85.9% (95%CI, 80.7-92.6) vs. 71.9% (95%CI, 46-73), HROW 0.41, 95%CI: 0.19-0.89, p=0.025. ConclusionEarly adjunctive treatment with tocilizumab, methylprednisolone or both may improve outcomes in non-intubated patients with COVID-19 pneumonia.