Genomic Evolution of Sars-Cov-2 in Molnupiravir-Treated Patients Compared to Paxlovid-Treated and Drug-Naïve Patients: A Proof-of-Concept Study (preprint)

Molnupiravir and Paxlovid are the only antivirals approved for COVID-19 treatment. Previous studies have evaluated their efficacy, tolerability, and viral clearance, but little is known about SARS-CoV-2 evolution under their pressure. Here the dynamics of genomic evolution of SARS-CoV-2 in 8 Molnupiravir-treated, 7 Paxlovid-treated and 5 drug-naïve individuals at 4 time-points (Day 0, Day 2, Day 5 of treatment and Day 7) were in-depth investigated. SARS-CoV-2 strains under Molnupiravir pressure were characterized by a higher genetic diversity compared to Paxlovid and no-drug pressure (mean ± SE: 18.66x10− 4±2.06x10− 4 vs. 3.34x10− 4±0.84x10− 4 vs. 3.10x10− 4±0.84x10− 4, P = 0.0003), with a peak between Day 2 and Day 5. Molnupiravir drove the emergence of more G-A and C-T transitions than other mutations (P = 0.031), regardless of SARS-CoV-2 genes. SARS-CoV-2 under Molnupiravir pressure did not show selective evolution different than that under Paxlovid or no-drug pressure, with the only exception of orf8 (dN > dS, P = 0.001); few amino acid mutations were enriched consistently at specific sites. No evidence of RdRp or Mpro mutations conferring resistance to Molnupiravir or Paxlovid was found. This proof-of-concept study defines the SARS-CoV-2 within-host evolution during antiviral treatment, confirming the higher in vivo variability induced by Molnupiravir respect to Paxlovid and controls, albeit not resulting in selection of resistance mutations.

Epidemiological Characterization of SARS-CoV-2 Variants in Children Over the Four COVID-19 Waves and Correlation with Clinical Presentation (preprint)

Since the start of SARS-CoV-2 pandemic, children aged ≤12 years have always been defined as underrepresented in terms of SARS-CoV-2 infections’ frequency and severity. By correlating SARS-CoV-2 transmission dynamics with clinical and virological features in 612 SARS-CoV-2 positive patients aged ≤12 years, we demonstrated a sizeable circulation of different SARS-CoV-2 lineages over the four pandemic waves in paediatric population, sustained by local transmission chains. Age <5 years, highest viral load, gamma and delta clades positively influence this local transmission. No correlations between COVID-19 manifestations and lineages or transmission chains are seen, except for a negative correlation between B.1.1.7 and hospitalization.

SARS-COV2 Seroprevalence in a Cohort of Perinatally HIV Infected Children and Young Adults From May to December 2020 in Italy (preprint)

SARS-CoV2 is a new coronavirus which started spreading in December 2019 from Wuhan, China. The seroprevalence of SARS-CoV2 antibodies allows to define a better picture of the spread of SARS-CoV2 infection in the population. The duration of SARS-CoV2 antibodies in the healthy population as well as in immunocompromised patients is still a topic of debate. HIV-infected people are at increased risk of developing complications from contracting a viral illness. Furthermore,their ability to develop and maintain an optimal immunological response to any kind of pathogen appears to be reduced.We analyzed the overall seroprevalence of SARS-CoV2 antibodies in 85 HIV infected-people on ART aged between 5 and 34 years old from May to January 2021. 88,2%of patients were in a good state of viroimmunological control: 23 showed a VL<40cp/ml and 52 had an undetectable VL. When positive for SARS-CoV2 serology, a confirmatory nasopharyngeal swab for PCR assessment and a second serological assay would be performed.Out of the 85 patients, 5 proved to be positive for SARS-CoV2 antibodies (rate of prevalence 5.8%). In all 5 cases the nasopharyngeal swabs were negative and the second assay for SARS-CoV2 antibodies performed in 4 out of 5 patients a week later was negative as well. The anamnestic recall brought no elements of suspicion for a past infection.The duration of SARS-CoV2 antibodies after COVID19 disease is still poorly understood in healthy population and additional studies will be needed to define the durability of humoral responses in immunocompromised children and in particular in HIV infected children under effective ART. It is still unknown whether ART or their immunological impairment may in part mitigate the pathogenesis of SARS-CoV-2 infection. Also, it will be interesting to analyze the impact of vaccination against SARS-CoV2 in HIV infected patients with a satisfactory virological control.

HUMORAL AND CELLULAR IMMUNOGENICITY and SAFETY UP TO 4 MONTHS AFTER VACCINATION WITH BNT162B2 mRNA COVID-19 VACCINE IN HEART AND LUNG TRANSPLANTED YOUNG ADULTS (preprint)

BackgroundImmunizations among vulnerable population, including solid organ transplant recipients (SOT), present suboptimal responses at vaccination and over time. We investigated safety and immunogenicity of the BNT162B2 mRNA COVID-19 vaccine in 34 SOT young adults as compared to 36 healthy controls (HC). Methodsimmunogenicity was measured through the analysis of anti SARS-CoV2 IgG Antibodies and antigen specific CD4 T cells (CD40L+), detected by flow cytometry before vaccination, 21 days after priming (T21), 7 days after booster dose (T28) and 2-4 months after priming (T120). Baseline T and B cell immune phenotype was deeply investigated. The safety profile was investigated by close monitoring and self-reported diary. ResultsAnti-S and anti-Trimeric Ab responses were significantly lower in SOT vs HC at T21 (p<0.0001) and at T28 (p<0.0001). Ten out of 34 SOT (29%) at T28 and 3 out of 33 (9%) at T120 had undetectable SARS-CoV-2 IgG. The analysis of SARS-CoV-2 specific CD4 T cells showed lower CD40L expression after in vitro stimulation in SOT compared to HC. Lower frequencies of memory B cells were found in patients not responding to vaccination. Lack of seroconversion was higher in patients treated with mycophenolate (p=0.0005). The vaccination was safe and well tolerated. Only short-term adverse events, were reported and no hospitalization or graft rejection were observed after vaccinations. ConclusionsThese data show that SOT have a suboptimal immune response following mRNA vaccinations as compared to HC. Alternative strategies should be investigated to improve the immunization against SARS-CoV-2 in these patients.

VIROLOGICAL AND IMMUNOLOGICAL FEATURES OF SARS-COV-2 INFECTED CHILDREN WITH DISTINCT SYMPTOMATOLOGY (preprint)

BACKGROUND: Despite SARS-CoV-2 immunizations have started in most countries, children are not currently included in the vaccination programs, thus it remains crucial to define their anti-SARS-CoV-2 immune response in order to minimize the risk for other epidemic waves. This study seeks to provide a description of the virology ad anti-SARS-CoV-2 immunity in children with distinct symptomatology. METHODS: Between March and July 2020, we recruited 15 SARS-CoV-2 asymptomatic (AS) and 51 symptomatic children (SY), stratified according to WHO clinical classification. We measured SARS-CoV-2 viral load using ddPCR and qPCR in longitudinally collected nasopharyngeal swabs samples. To define anti-SARS-CoV-2 antibodies we measured neutralization activity and total IgG load (Diasorin). We also evaluated antigen-specific B and CD8+T-cells, using a labelled S1+S2 protein and ICAM expression, respectively. Plasma protein profiling was performed with Olink. RESULTS: Virological profiling showed that AS had lower viral load at diagnosis (p=0.004) and faster virus clearance (p=0.0002) compared to SY. Anti-SARS CoV-2 humoral and cellular response did not appear to be associated with the presence of symptoms. AS and SY showed similar titers of SARS-CoV-2 IgG, levels of neutralizing activity, and frequency of Ag-specific B and CD8+T-cells. Whereas pro-inflammatory plasma protein profile was associated to symptomatology. CONCLUSION: We demonstrated the development of anti-SARS-CoV-2 humoral and cellular response with any regards to symptomatology, suggesting the ability of both SY and AS to contribute towards herd immunity. The virological profiling of AS suggested that they have lower virus load associated with faster virus clearance.

Prioritizing the first doses of SARS-CoV-2 vaccine to save the elderly: the case study of Italy (preprint)

Many countries are currently facing high mortality caused by the circulation of SARS-CoV-2 among the elderly not yet vaccinated. Vaccine shortage poses relevant challenges to health authorities, called to act in a timely manner, and with scarcity of vaccine, and data. We have developed a model for estimating of the impact of vaccination on the mortality of the elderly following a schedule of mRNA SARS-CoV-2 vaccine that prioritize first dose administration, as alternative to the standard schedule of two doses administered 3 to 4 weeks apart. We studied the Italian scenario, considering it representative of other Countries facing similar conditions in terms of virus circulation, mortality, and vaccine shortage, in the period from February 10 to April 14 2021. Under different conditions of quantity of vaccine administration, the schedule prioritizing first doses showed always significant increase of protected individuals, and a decrease of deaths, up to 19.8% less than the standard schedule. These findings support the vaccination option of prioritizing first dose in the elderly until vaccine supplies are adequate.

Post-Discharge Telephonic Follow-Up of Pediatric Patients Affected by SARS-CoV2 Infection: A Safe and Feasible Way to Monitor Children After Hospitalization. The Experience of a Single Italian Pediatric COVID Center. (preprint)

BackgroundSARS-CoV-2 infection in children is often non severe and in the majority of cases does not require long term hospitalization, nevertheless it is burdened with social issues and managing difficulties. To our knowledge there is no literature on telephonic follow up in pediatric patients with positive PCR for SARS-CoV-2 on rhino-pharyngeal swab after discharge. The aim of the study is to describe our experience in a telephonic follow up which can allow early and safe discharge from hospital while keeping the patients under close clinical monitoring.Materials and Methods65 children were admitted for SARS-CoV-2 infection at Bambino Gesù Pediatric Hospital COVID Center from 16th March to 3rd July. We monitored through a telephonic follow-up, using a specific survey, the patients discharged still presenting a positive PCR for SARS-CoV-2. We checked if any symptoms occurred at home until recovery, defined as two consecutive negative PCR for SARS-CoV-2 on rhino-pharyngeal swabs.ResultsDuring the follow up 7 patients had mild and self-limited symptoms related to SARS-CoV-2 infection, while 2 patients were re-hospitalized. 1 patient had Multisystem Inflammatory Syndrome in Children (MIS-C), the other patient had an increase in troponin and D-dimers. We also monitored the average time of viral shedding, resulting in a median duration of 28 days.ConclusionOur experience describes the daily telephonic follow up as safe in pediatric patients discharged with positive PCR. As a matter of fact it could avoid long term hospitalization and allow to promptly re-hospitalize children with major complications such as MIS-C.

Post-discharge telephonic follow-up of pediatric patients affected by SARS-CoV2 infection: a safe and feasible way to monitor children after hospitalization.The experience of a single Italian Pediatric COVID Center. (preprint)

BackgroundIn early January 2020, a novel type of Coronavirus was identified in a patient affected by pneumonia of unknown origin. The virus will be named SARS-CoV-2 and the disease COVID-19 a month later by the International Committee on Virus Taxonomy.Italy is one of the first countries in the world affected by the COVID-19 pandemic, with 1.2% of all patients represented by children. Although the infection in children is often non severe and in the majority of cases does not require long term hospitalization, it is burdened with social issues and managing difficulties.To our knowledge there is no literature on telephonic follow up in pediatric patients with positive rhino-pharyngeal swab for SARS-CoV-2 after discharge.Materials and MethodsWe monitored through a telephonic follow-up, using a specific survey, 19 children aged between 8 months and 15 years, hospitalized in the “Ospedale Pediatrico Bambino Gesù” COVID Center with positive rhino-pharyngeal swab at discharge. We checked if any symptoms occurred at home until recovery, defined as two consecutive negative rhino-pharyngeal swabs.ResultsDuring the follow up 7 patients had mild and self-limited symptoms related to SARS-CoV-2 infection, while 2 patients were re-hospitalized, 1 patient had Multisystem Inflammatory Syndrome in Children (MIS-C), the other patient had an increase in troponin a D-dimers.We didn’t miss any patient during the follow up.ConclusionWe demonstrated that daily telephonic follow up is safe in pediatric patients discharged with positive swab, it allows to avoid long term hospitalization and to promptly re-hospitalize children with major complication such as MIS-C.