On the lookout for influenza viruses in Italy during the 2021-2022 season: Along came A(H3N2) viruses with a new phylogenetic makeup of their hemagglutinin.

AIMS: To assess influenza viruses (IVs) circulation and to evaluate A(H3N2) molecular evolution during the 2021-2022 season in Italy. MATERIALS AND METHODS: 12,393 respiratory specimens (nasopharyngeal swabs or broncho-alveolar lavages) collected from in/outpatients with influenza illness in the period spanning from January 1, 2022 (week 2022-01) to May 31, 2022 (week 2022-22) were analysed to identify IV genome and were molecularly characterized by 12 laboratories throughout Italy. A(H3N2) evolution was studied by conducting an in-depth phylogenetic analysis of the hemagglutinin (HA) gene sequences. The predicted vaccine efficacy (pVE) of vaccine strain against circulating A(H3N2) viruses was estimated using the sequence-based Pepitope model. RESULTS: The overall IV-positive rate was 7.2% (894/12,393), all were type A IVs. Almost all influenza A viruses (846/894; 94.6%) were H3N2 that circulated in Italy with a clear epidemic trend, with 10% positivity rate threshold crossed for six consecutive weeks from week 2022-11 to week 2022-16. According to the phylogenetic analysis of a subset of A(H3N2) strains (n=161), the study HA sequences were distributed into five different genetic clusters, all of them belonging to the clade 3C.2a, sub-clade 3C.2a1 and the genetic subgroup 3C.2a1b.2a.2. The selective pressure analysis of A(H3N2) sequences showed evidence of diversifying selection particularly in the amino acid position 156. The comparison between the predicted amino acid sequence of the 2021-2022 vaccine strain (A/Cambodia/e0826360/2020) and the study strains revealed 65 mutations in 59 HA amino acid positions, including the substitution H156S and Y159N in antigenic site B, within major antigenic sites adjacent to the receptor-binding site, suggesting the presence of drifted strains. According to the sequence-based Pepitope model, antigenic site B was the dominant antigenic site and the p(VE) against circulating A(H3N2) viruses was estimated to be -28.9%. DISCUSSION AND CONCLUSION: After a long period of very low IV activity since public health control measures have been introduced to face COVID-19 pandemic, along came A(H3N2) with a new phylogenetic makeup. Although the delayed 2021-2022 influenza season in Italy was characterized by a significant reduction of the width of the epidemic curve and in the intensity of the influenza activity compared to historical data, a marked genetic diversity of the HA of circulating A(H3N2) strains was observed. The identification of the H156S and Y159N substitutions within the main antigenic sites of most HA sequences also suggested the circulation of drifted variants with respect to the 2021-2022 vaccine strain. Molecular surveillance plays a critical role in the influenza surveillance architecture and it has to be strengthened also at local level to timely assess vaccine effectiveness and detect novel strains with potential impact on public health.

Multicenter epidemiological investigation and genetic characterization of respiratory syncytial virus and metapneumovirus infections in the pre-pandemic 2018-2019 season in northern and central Italy.

Respiratory syncytial virus (RSV) and human metapneumovirus (HMPV) cause a high burden of disease, particularly in children and the elderly. With the aim to add knowledge on RSV and HMPV infections in Italy, a prospective, multicenter study was conducted by eight centers of the Working Group on Respiratory Virus Infections (GLIViRe), from December 2018-April 2019. Weekly distribution and patients' demographic and clinical data were compared in 1300 RSV and 222 HMPV-positive cases. Phylogenetic analysis of the G-glycoprotein coding region was performed to characterize circulating strains. RSV positivity ranged from 6.4% in outpatients of all ages to 31.7% in hospitalized children; HMPV positivity was 4-1.2% with no age-association. RSV season peaked in February and ended in mid-April: HMPV circulation was higher when RSV decreased in early spring. RSV was more frequent in infants, whereas HMPV infected comparatively more elderly adults; despite, their clinical course was similar. RSV-B cases were two-thirds of the total and had similar clinical severity compared to RSV-A. Phylogenetic analysis showed the circulation of RSV-A ON1 variants and the predominance of RSV-B genotype BA10. HMPV genotype A2c was the prevalent one and presented insertions of different lengths in G. This first multicenter Italian report on seasonality, age-specific distribution, and clinical presentation of RSV and HMPV demonstrated their substantial disease burden in young patients but also in the elderly. These data may provide the basis for a national respiratory virus surveillance network.

Comparison between Dexamethasone and Methylprednisolone Therapy in Patients with COVID-19 Pneumonia Admitted to Non-Intensive Medical Units.

(1) Background: Data on different steroid compounds for the treatment of hospitalized COVID-19 (coronavirus disease 2019) patients are still limited. The aim of this study was to compare COVID-19 patients admitted to non-intensive units and treated with methylprednisolone or dexamethasone. (2) Methods: This was a single-center retrospective study that included consecutive patients with COVID-19 hospitalized in medical wards during the second wave of the pandemic. Thirty-day mortality and the need for intensive or semi-intensive care were the main clinical outcomes analyzed in patients receiving methylprednisolone (60 mg/day) compared with dexamethasone (6 mg/day). Secondary outcomes included complication rates, length of hospital stay, and time to viral clearance. (3) Results: Two-hundred-forty-six patients were included in the analysis, 110 treated with dexamethasone and 136 with methylprednisolone. No statistically significant differences were found between the two groups of patients regarding 30-day mortality (OR 1.35, CI95% 0.71-2.56, p = 0.351) and the need for intensive or semi-intensive care (OR 1.94, CI95% 0.81-4.66, p = 0.136). The complication rates, length of hospital stay, and time to viral clearance did not significantly differ between the two groups. (4) Conclusions: In patients hospitalized for COVID-19 in non-intensive units, the choice of different steroid compounds, such as dexamethasone or methylprednisolone, did not affect the main clinical outcomes.

COVID-19 in B Cell-Depleted Patients After Rituximab: A Diagnostic and Therapeutic Challenge.

B cell-targeting strategies such as rituximab are widely used in B cell hematologic malignancies, rheumatologic and musculoskeletal diseases and a variety of autoimmune disorders. The purpose of this paper is to illustrate how exposure to anti-CD20 treatment profoundly affects B cell functions involved in anti-SARS-CoV-2 immunity and significantly impacts on the clinical and serological course of SARS-CoV-2 infection, long term immunity and vaccine responses. The data presented here suggest that the effects of B cell-depleting agents on adaptive immunity should be taken into account for the proper selection and interpretation of SARS-CoV-2 diagnostics and to guide appropriate therapeutic approaches and protective measures. Combination therapeutic strategies including immunotherapy in association with prolonged antiviral treatment may play a decisive role in the setting of B cell immune deficiencies.

A Severe Acute Respiratory Syndrome Coronavirus 2 detection method based on nasal and nasopharyngeal lavage fluid: A pilot feasibility study.

OBJECTIVE: Nose and nasopharyngeal swab is the preferred and worldwide-accepted method to detect the Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) within the nose and nasopharynx. This method may be linked with possible difficulties, such as patient's discomfort or complications. This article shows a pilot study of SARS-CoV-2 detection with nasal and nasopharyngeal lavage fluids (level of evidence: 3). METHODS: Nasal lavage fluid was collected from patients who were submitted to SARS-CoV-2 screening test, due to a preceding positive rapid antigen test. A control group was enrolled among health care professionals whose nasopharyngeal swab tested negative. Nasal lavages were performed using isotonic saline solution injected through a nasal fossa. Both lavage fluid and traditional nasopharyngeal swab were analyzed by real-time (RT) PCR and antigenic test. RESULTS: A total of 49 positive subjects were enrolled in the study. Results of the analysis on lavages and nasopharyngeal swabs were concordant for 48 cases, regardless of the antigenic and molecular test performed. RT-PCR resulted weakly positive at swab in one case and negative at lavage fluid. Among the control group (44 subjects), nasopharyngeal swab and lavage fluid analyses returned a negative result. Sensitivity of the molecular test based on nasal lavage fluid, compared to traditional nasal swab, was 97.7%, specificity was 100%, and accuracy was 98.9%, with high agreement (Cohen's κ, 0.978). CONCLUSION: Nasal and nasopharyngeal lavages resulted to be highly reliable and well tolerated. A larger series is needed to confirm these results. This approach may potentially represent a valid alternative to the traditional swab method in selected cases. LEVEL OF EVIDENCE: 3.

Association between hepatitis C and B viruses and head and neck squamous cell carcinoma.

BACKGROUND: Hepatitis B and C viruses are known to be carcinogenic and have been associated with the development of non-Hodgkin's lymphoma as well as hepatocellular carcinoma. The incidence of head and neck cancer is increasing worldwide, and early diagnosis is vital in order to achieve good oncological outcomes. OBJECTIVES: To investigate the association between chronic hepatitis B and C infection, and head and neck squamous cell carcinoma (HNSCC). STUDY DESIGN: We performed a retrospective case control study with 774 head and neck squamous cell carcinoma (HNSCC) patients undergoing treatment, and 1518 cancer-free controls undergoing hernia surgery. Hepatitis B and C serologies were tested prior to treatment, and cases and controls were age- and sex-matched before analysing rates of infection. RESULTS: HNSCC patients were more likely than controls to have evidence of chronic hepatitis B (OR = 2.76; CI 95 %, 1.64-4.64) and hepatitis C (OR = 2.59; 95 % CI, 1.46-4.60) infection. No substantial association was found between hepatitis B and C infection and other known risk factors for head and neck cancer. CONCLUSIONS: These findings suggest a positive association between both hepatitis B and hepatitis C chronic infection, and HNSCC. More work is needed to establish a causal role, however an awareness of the possibility of increased risk of HNSCC may lead to earlier diagnosis and better outcomes in patients with hepatitis B and C.

HPV testing for cervical cancer screening: technical improvement of laboratory logistics and good clinical performance of the cobas 6800 in comparison to the 4800 system.

BACKGROUND: European guidelines for cervical cancer screening now recommend the use of clinically validated assays for high-risk HPV-DNA sequences as primary test in women older than 30 years, performed in centralized laboratories, and run on systems providing automated solutions for all steps. METHODS: We conducted a comparison study, according to the international guidelines, nested within the organized population-based cervical screening program, between the cobas 4800 and 6800 systems (Roche Diagnostics), to evaluate accuracy and reproducibility of HPV test results and laboratory workflow. In Italy implementation of HPV cervical screening is under way on a regional basis; in Veneto it started in June 2015, following a piloting phase; the assay in use in the three centralized laboratories is the cobas 4800 HPV test, run on the cobas 4800 system. Comparison of HPV results with a new version of the assay (cobas 6800/8800 HPV) run on the cobas 6800 system, and intra- and inter-reproducibility analyses have been conducted in samples collected in PreservCyt medium (Hologic) from women without and with a subsequent diagnosis of high-grade lesion. RESULTS: Samples from women older than 30 years attending organized cervical cancer screening were used. Clinical sensitivity and specificity were evaluated on 60 cases and 925 controls, respectively; intra-laboratory reproducibility and inter-laboratory agreement by the 6800 system were evaluated on 593 and 460 specimens, respectively. Our results showed a very high agreement (> 98%) for overall qualitative results between the two systems; clinical sensitivity and specificity of the HPV assay run on 6800 were non-inferior to those of the HPV assay run on 4800 (p = 0,0157 and p = 0,0056, respectively, at the recommended thresholds of 90 and 98%); kappa values of 0.967 and 0.969 were obtained for intra- and inter-laboratory reproducibility analyses in the 6800 system. The 6800 platform displayed several technological improvements over the 4800 system, with higher throughput and laboratory productivity, and lower operator's hands-on time. CONCLUSIONS: The new cobas 6800/8800 HPV assay run on the 6800 instrument is suitable for use in large centralized laboratories included within population-based cervical cancer screening programs.