Paola Stefanelli

Systemic innate immune responses during infant pertussis remain incompletely characterized, particularly in relation to disease severity, vaccination and respiratory viral coinfection. We conducted a prospective observational study of 61 infants younger than 12 months hospitalized with PCR-confirmed pertussis. Seven serum cytokines and chemokines were measured during early hospitalization, and their associations with clinical manifestations, vaccination and viral coinfection were investigated. Correlation analysis and principal component analysis identified a coordinated pro-inflammatory cluster comprising IL-1β, TNF-α, IL-6, IL-8 and CCL3 (PC1), and a distinct component mainly reflecting CXCL10 and IL-10 (PC2). No individual cytokine or composite profile was significantly associated with clinical manifestations after correction for multiple comparisons, although exploratory associations of CXCL10 and PC2 with respiratory distress were observed. Respiratory viral coinfection was independently associated with higher CXCL10 concentrations (2.21-fold increase; 95% CI 1.28–3.82; p = 0.0045; FDR q = 0.032). Prior vaccination and respiratory viral coinfection showed opposite trends in their associations with inflammatory mediators, with vaccination tending to be associated with stronger pro-inflammatory responses. Previously vaccinated infants also had higher anti-pertussis toxin IgG concentrations (p = 0.0006). These findings highlight distinct patterns of systemic immune activation during infant pertussis, influenced by respiratory viral detection and prior immune history, without clear associations with clinical severity.

Avian influenza viruses (AIVs) of H10 subtype are able to circulate in domestic and wild bird populations but can also spill over and adapt to mammals, posing a continuous risk to biodiversity conservation, veterinary health, and public health. In the present study, we assessed the zoonotic potential of nine H10 AIVs isolated from waterbirds during surveillance and research studies carried out in Italy between 1994 and 2007. Overall, six H10NX strains from wild mallards (n. 1 H10N2, n. 5 H10N7), one H10N7 strain from domestic mallards, and two H10N8 strains from Eurasian coots were sequenced by next-generation sequencing (NGS). HA phylogenetic analysis indicated a marked divergence between viruses from these two sympatric waterbird species and showed a close relationship between three H10N7 strains from wild mallard and one H10N7 isolate of domestic origin. Sequence analysis revealed the presence of several molecular markers, associated with increased zoonotic potential, including the PB2-A588V mutation found in the Eurasian coot H10N8 viruses and previously linked to mammalian adaptation in H10 strains. Molecular analysis also showed that all H10 viruses were susceptible to the major approved classes of influenza antivirals (inhibitors of neuraminidase, matrix-2, and polymerase acid protein). Moreover, phenotypic assay confirmed their susceptibility to oseltamivir and zanamivir drugs. From an ecological perspective, we found that different H10 gene pools seem to be harboured in different waterbird species sharing the same environment; additionally, a bidirectional transmission of H10 mallard isolates occurred between natural and anthropic ecosystems. Overall, our findings account for the need of continuous monitoring of AIVs belonging to the H10 subtype.

The continuous evolution of SARS-CoV-2 has led to the emergence of several variants representing significant challenges for public health. Many studies highlight the relevance of phylogenetic inference or mutational pattern analysis to understand the evolutionary relatedness of viral variants and to estimate the potential effect of new mutations on viral transmission, virulence and antigenicity. Here we describe an evolutionary investigation approach combined with mutational analyses of SARS-CoV-2 Spike gene to annotate and potentially track important amino acid site variation of specific functional domain relevant for viral survival. This approach was applied on XBB*, EG* and BA* and their sub-lineages (see materials and methods) available from GISAID. In addition, we considered the major variants of concern (Alpha, Delta, Omicron) and Wuhan-Hu-1 strain as references. Maximum likelihood phylogenetic tree was constructed from the complete dataset while selection pressure and mutational analyses were conducted on single variants separately. The obtained phylogenetic tree of Spike amino acid gene sequence showed a clear separation of viral variants as well as their expected appearance order. This result supported the significance of selection pressure analyses outcomes combined with amino acid mutational frequencies where in many cases they showed a linear and parallel trend. This allowed also to hypothesize the potential importance of low-frequency mutations in new potential virus variants. This study constitutes an asset of important insights to be considered in regular monitoring programs. In addition, the analysis framework described here introduces a starting point for further standardization, optimization and application on different data types and in large-scale studies.

The continuous emergence of SARS-CoV-2 variants and subvariants poses significant public health challenges. The latest designated subvariant JN.1, with all its descendants, shows more than 30 mutations in the spike gene. JN.1 has raised concerns due to its genomic diversity and its potential to enhance transmissibility and immune evasion. This study aims to analyse the molecular characteristics of JN.1-related lineages (JN.1*) identified in Italy from October 2023 to April 2024 and to evaluate the neutralization activity against JN.1 of a subsample of sera from individuals vaccinated with XBB.1.5 mRNA. The genomic diversity of the spike gene of 794 JN.1* strain was evaluated and phylogenetic analysis was conducted to compare the distance to XBB.1.5. Moreover, serum neutralization assays were performed on a subsample of 19 healthcare workers (HCWs) vaccinated with the monovalent XBB.1.5 mRNA booster to assess neutralizing capacity against JN.1. Sequence analysis displayed high spike variability between JN.1* and phylogenetic investigation confirmed a substantial differentiation between JN.1* and XBB.1.5 spike regions with 29 shared mutations, of which 17 were located within the RBD region. Pre-booster neutralization activity against JN.1 was observed in 42% of HCWs sera, increasing significantly post-booster, with all HCWs showing neutralization capacity three months after vaccination. A significant correlation was found between anti-trimeric Spike IgG levels and neutralizing titers against JN.1. The study highlights the variability of JN.1* in Italy. Results on a subsample of sera from HCWs vaccinated with XBB.1.5 mRNA booster vaccine suggested enhanced neutralization activity against JN.1.

In this pilot study, a multi-parametric analysis comparing immune responses in sera of adult healthy subjects (HS) or people with type 2 diabetes mellitus (T2D) undergoing the single or simultaneous administration of mRNA-based COVID-19 and cellular quadrivalent inactivated influenza vaccines was conducted. While SARS-CoV-2 antibodies remains comparable, influenza antibody titers and seroconversion were significantly higher upon simultaneous vaccination. Magnitude of anti-influenza humoral response closely correlated with an early innate immune signature, previously described for the COVID-19 vaccine, composed of IL-15, IL-6, TNF-α, IFN-γ, CXCL-10 and here extended also to acute-phase protein Pentraxin 3. People with T2D receiving simultaneous vaccination showed a protective response comparable to HS correlating with the early induction of IFN-γ/CXCL10 and a significant reduction of the circulating glucose level due to increased oxidation of glucose digestion and consumption. These data, although preliminary and in-need of validation in larger cohorts, might be exploited to optimize future vaccination in people with chronic disorders, including diabetes.

The Central Mediterranean migration route to Europe is one of the most significant and dangerous routes in the world, and it continues to be a major humanitarian concern. It is characterised by a mixed migration flow, with diverse groups of people from Africa, the Middle East, and Asia attempting to reach European shores in search […]

In this work, we studied the selective pressure and evolutionary analysis on the SARS-CoV-2 BF.7 and BQ.1.1 lineages circulating in Italy from July to December 2022. Two different datasets were constructed: the first comprised 694 SARS-CoV-2 BF.7 lineage sequences and the second comprised 734 BQ.1.1 sequences, available in the Italian COVID-19 Genomic (I-Co-Gen) platform and GISAID (last access date 15 December 2022). Alignments were performed with MAFFT v.7 under the Galaxy platform. The HYPHY software was used to study the selective pressure. Four positively selected sites (two in nsp3 and two in the spike) were identified in the BF.7 dataset, and two (one in ORF8 and one in the spike gene) were identified in the BQ.1.1 dataset. Mutation analysis revealed that R408S and N440K are very common in the spike of the BF.7 genomes, as well as L452R among BQ.1.1. N1329D and Q180H in nsp3 were found, respectively, at low and rare frequencies in BF.7, while I121L and I121T were found to be rare in ORF8 for BQ.1.1. The positively selected sites may have been driven by the selection for increased viral fitness, under circumstances of defined selective pressure, as well by host genetic factors.

Background: In Italy the introduction of meningococcal C conjugate vaccine in 2005 has led to a significant reduction of invasive meningococcal disease (IMD) caused by Neisseria meningitidis of serogroup C (MenC). However, this serogroup is still responsible of sporadic cases, clusters and local outbreaks. The study aims to investigate the genotype and antimicrobial susceptibility profile of MenC isolates collected in Italy from 2000 to 2020.Methods: Bacterial isolates and biological samples (blood or cerebrospinal fluid) from invasive meningococcal cases are collected and characterized at the National Reference Laboratory for IMD of Istituto Superiore di Sanità. Antimicrobial susceptibility was determined by MIC Test Strip Method and interpreted according to the EUCAST breakpoints guideline. Genotypic characteristics, including multi locus sequence typing (MLST), finetype, and antimicrobial resistance target genes were performed and analyzed using the PubMLST database. Genomic comparison of core genome MLST (cgMLST) of MenC genomes was also carried out.Results: From 2000 to 2020, a total of 665 MenC isolates were investigated for antimicrobial susceptibility and 301 for genotyping. Over two decades, almost all MenC isolates resulted susceptible to antimicrobials with few isolates resulting resistant to ciprofloxacin (N = 2), penicillin G (N = 13), and rifampicin (N = 9), respectively. Molecular typing of MenC obtained from isolates or clinical specimens identified mostly the genotype C:P1.5-1,10-8:F3-6:ST-11(cc11). However, phylogenetic analysis, performed on genomes from MenC isolates, identified two sub lineages, 11.1 and 11.2, among cc11, of which the sub lineage 11.2 was the predominant.Conclusion: Wider application of the genomic analysis and monitoring of antimicrobial susceptibility represent key aspects of IMD surveillance and to monitor the continued evolution of these hyperinvasive strains.

The "Investigating and translating genomic evidence for public health response to SARSCoV-2 (INSIDE SARS-CoV-2)" project is part of the initiative "Joint science and technology cooperation call for joint project proposals for the years 2021-2023" promoted by the Italian Ministry of Foreign Affairs and International Cooperation (MAECI) and the Republic of India. To start the project activities, the pandemic response and the epidemiological situation in Italy and in India, together with the genomic surveillance strategies for SARS-CoV-2 virus in the two countries, are here described.

Innate immune responses to coronavirus infections are highly cell specific. Tissue-resident macrophages, which are infected by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) in patients but are inconsistently infected in vitro, exert critical but conflicting effects by secreting both antiviral type I interferons (IFNs) and tissue-damaging inflammatory cytokines. Steroids, the only class of host-targeting drugs approved for the treatment of coronavirus disease 2019 (COVID-19), indiscriminately suppress both responses, possibly impairing viral clearance. Here, we established in vitro cell culture systems that enabled us to separately investigate the cell-intrinsic and cell-extrinsic proinflammatory and antiviral activities of mouse macrophages infected with the prototypical murine coronavirus MHV-A59. We showed that the nuclear factor κB–dependent inflammatory response to viral infection was selectively inhibited by loss of the lysine demethylase LSD1, which was previously implicated in innate immune responses to cancer, with negligible effects on the antiviral IFN response. LSD1 ablation also enhanced an IFN-independent antiviral response, blocking viral egress through the lysosomal pathway. The macrophage-intrinsic antiviral and anti-inflammatory activity of Lsd1 inhibition was confirmed in vitro and in a humanized mouse model of SARS-CoV-2 infection. These results suggest that LSD1 controls innate immune responses against coronaviruses at multiple levels and provide a mechanistic rationale for potentially repurposing LSD1 inhibitors for COVID-19 treatment.