Performance Assessment of the LIAISON® SARS-CoV-2 Antigen Assay On Nasopharyngeal Swabs.

The SARS-CoV-2 pandemic is ongoing worldwide, causing prolonged pressure on molecular diagnostics. Viral antigen (Ag) assays have several advantages, ranging from lower cost to shorter turnaround time to detection. Given the rare occurrence of low-load viremia, antigen assays for SARSCoV-2 have focused on nasopharyngeal swab and saliva as biological matrices, but their effectiveness must be validated. We assayed here the performances of the novel quantitative Liaison® SARSCoV-2 Ag assay on 119 nasopharyngeal swabs and obtained results were compared with Hologic Panther and Abbott m2000 RT-qPCR. The Ag assay demonstrated a good correlation with viral load, shorter turnaround time, and favorable economics. The best performance was obtained in the acute phase of disease.

Treatment of severe post-traumatic bone defects with autologous stem cells loaded on allogeneic scaffolds.

Mesenchymal stem cells may differentiate into angiogenic and osteoprogenitor cells. The effectiveness of autologous pluripotent mesenchymal cells for treating bone defects has not been investigated in humans. We present a case series to evaluate the rationale of using nucleated cells from autologous bone marrow aspirates in the treatment of severe bone defects that failed to respond to traditional treatments. Ten adult patients (mean age, 49.6-years-old) with severe bone defects were included in this study. Lower limb bone defects were >or=5 cm3 in size, and upper limb defects .or=2 cm3. Before surgery, patients were tested for antibodies to common pathogens. Treatment consisted of bone allogeneic scaffold enriched with bone marrow nucleated cells harvested from the iliac crest and concentrated using an FDA-approved device. Postsurgery clinical and radiographic follow-up was performed at 1, 3, 6, and 12 months. To assess viability, morphology, and immunophenotype, bone marrow nucleated cells were cultured in vitro, tested for sterility, and assayed for the possible replication of adventitious (contaminating) viruses. In 9 of 10 patients, both clinical and radiographic healing of the bone defect along with bone graft integration were observed (mean time, 5.6 months); one patient failed to respond. No post-operative complications were observed. Bone marrow nucleated cells were enriched 4.49-fold by a single concentration step, and these enriched cells were free of microbial contamination. The immunophenotype of adherent cells was compatible with that of mesenchymal stem cells. We detected the replication of Epstein-Barr virus in 2/10 bone marrow cell cultures tested. Hepatitis B virus, cytomegalovirus, parvovirus B19, and endogenous retrovirus HERV-K replication were not detected. Overall, 470 to 1,150 million nucleated cells were grafted into each patient. This case series, with a mean follow-up of almost 2 years, demonstrates that an allogeneic bone scaffold enriched with concentrated autologous bone marrow cells obtained from the iliac crest provides orthopedic surgeons a novel option for treating important bone defects that are unresponsive to traditional therapies.

Detection of Torquetenovirus and Redondovirus DNA in Saliva Samples from SARS-CoV-2-Positive and -Negative Subjects.

OBJECTIVES: Torquetenovirus (TTV) and Redondovirus (ReDoV) are the most prevalent viruses found in the human respiratory virome in viral metagenomics studies. A large-scale epidemiological study was performed to investigate their prevalence and loads in saliva samples according to SARS-CoV-2 status. METHODS: Saliva samples from 448 individuals (73% SARS-CoV-2 negative and 27% SARS-CoV-2 positive) aged 23-88 years were tested. SARS-CoV-2 and TTV were determined in saliva by specific qualitative and quantitative real-time PCRs, respectively. A sub-cohort of 377 subjects was additionally tested for the presence and load of ReDoV in saliva, and a different sub-cohort of 120 subjects for which paired saliva and plasma samples were available was tested for TTV and ReDoV viremia at the same timepoints as saliva. RESULTS: TTV in saliva was 72% prevalent in the entire cohort, at a mean DNA load of 4.6 log copies/mL, with no difference regardless of SARS-CoV-2 status. ReDoV was found in saliva from 61% of the entire cohort and was more prevalent in the SARS-CoV-2-negative subgroup (65% vs. 52%, respectively). In saliva, the total mean load of ReDoV was very similar to the one of TTV, with a value of 4.4 log copies/mL. The mean viral loads in subjects infected with a single virus, namely, those infected with TTV or ReDoV alone, was lower than in dually infected samples, and Tukey's multiple-comparison test showed that ReDoV single-infected samples resulted in the only true outlier (p = 0.004). Differently from TTV, ReDoV was not detected in any blood samples. CONCLUSIONS: This study establishes the prevalence and mean value of TTV and ReDoV in saliva samples and demonstrates the existence of differences between these two components of the human virome.

Mucosal immune response in BNT162b2 COVID-19 vaccine recipients.

BACKGROUND: Although the BNT162b2 COVID-19 vaccine is known to induce IgG neutralizing antibodies in serum protecting against COVID-19, it has not been studied in detail whether it could generate specific immunity at mucosal sites, which represent the primary route of entry of SARS-CoV-2. METHODS: Samples of serum and saliva of 60 BNT162b2-vaccinated healthcare workers were collected at baseline, two weeks after the first dose and two weeks after the second dose. Anti-S1-protein IgG and IgA total antibodies titres and the presence of neutralizing antibodies against the Receptor Binding Domain in both serum and saliva were measured by quantitative and by competitive ELISA, respectively. FINDINGS: Complete vaccination cycle generates a high serum IgG antibody titre as a single dose in previously infected seropositive individuals. Serum IgA concentration reaches a plateau after a single dose in seropositive individuals and two vaccine doses in seronegative subjects. After the second dose IgA level was higher in seronegative than in seropositive subjects. In saliva, IgG level is almost two orders of magnitude lower than in serum, reaching the highest values after the second dose. IgA concentration remains low and increases significantly only in seropositive individuals after the second dose. Neutralizing antibody titres were much higher in serum than in saliva. INTERPRETATION: The mRNA BNT162b2 vaccination elicits a strong systemic immune response by drastically boosting neutralizing antibodies development in serum, but not in saliva, indicating that at least oral mucosal immunity is poorly activated by this vaccination protocol, thus failing in limiting virus acquisition upon its entry through this route. FUNDING: This work was funded by the Department of Medicine and Surgery, University of Insubria, and partially supported by Fondazione Umberto Veronesi (COVID-19 Insieme per la ricerca di tutti, 2020).

Saliva is a reliable tool to detect SARS-CoV-2.

OBJECTIVES: This study analyzed salivary samples of COVID-19 patients and compared the results with their clinical and laboratory data. METHODS: Salivary samples of 25 COVID-19 patients were analyzed by rRT-PCR. The following data were collected: age, sex, comorbidities, drugs. Lactate dehydrogenase (LDH) and ultrasensitive reactive C protein (usRCP) values were registered on the same day when a salivary swab was collected. Prevalence of positivity in saliva and association between clinical data and the cycle threshold as a semiquantitative indicator of viral load were considered. RESULTS: Twenty-five subjects were recruited into this study, 17 males and 8 females. The mean age was 61.5 +/- 11.2 years. Cardiovascular and/or dysmetabolic disorders were observed in 65.22% of cases. All the samples tested positive for the presence of SARS-CoV-2, while there was an inverse association between LDH and Ct values. Two patients showed positive salivary results on the same days when their pharyngeal or respiratory swabs showed conversion. CONCLUSIONS: Saliva is a reliable tool to detect SARS-CoV-2. The role of saliva in COVID-19 diagnosis could not be limited to a qualitative detection of the virus, but it may also provide information about the clinical evolution of the disease.