Genes Involved in miRNA Biogenesis Are Not Downregulated in SARS-CoV-2 Infection.

miRNAs, small non-coding RNAs that regulate gene expression, are involved in various pathological processes, including viral infections. Virus infections may interfere with the miRNA pathway through the inhibition of genes involved in miRNA biogenesis. A reduction in the number and the levels of miRNAs expressed in nasopharyngeal swabs of patients with severe COVID-19 was lately observed by us, pointing towards the potential of miRNAs as possible diagnostic or prognostic biomarkers for predicting outcomes among patients with severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) infection. The objective of the present study was to investigate whether SARS-CoV-2 infection influences the expression levels of messenger RNAs (mRNAs) of key genes involved in miRNA biogenesis. mRNA levels of AGO2, DICER1, DGCR8, DROSHA, and Exportin-5 (XPO5) were measured by quantitative reverse-transcription polymerase chain reaction (RT-qPCR) in nasopharyngeal swab specimens from patients with COVID-19 and controls, as well as in cells infected with SARS-CoV-2 in vitro. Our data showed that the mRNA expression levels of AGO2, DICER1, DGCR8, DROSHA, and XPO5 were not significantly different in patients with severe COVID-19 when compared to patients with non-severe COVID-19 and controls. Similarly, the mRNA expression of these genes was not affected by SARS-CoV-2 infection in NHBE and Calu-3 cells. However, in Vero E6 cells, AGO2, DICER1, DGCR8, and XPO5 mRNA levels were slightly upregulated 24 h after infection with SARS-CoV-2. In conclusion, we did not find evidence for downregulation of mRNA levels of miRNA biogenesis genes during SARS-CoV-2 infection, neither ex vivo nor in vitro.

Viruses and Endocrine Diseases.

Viral infections have been frequently associated with physiological and pathological changes in the endocrine system for many years. The numerous early and late endocrine complications reported during the current pandemic of coronavirus disease 2019 (COVID-19) reinforce the relevance of improving our understanding of the impact of viral infections on the endocrine system. Several viruses have been shown to infect endocrine cells and induce endocrine system disturbances through the direct damage of these cells or through indirect mechanisms, especially the activation of the host antiviral immune response, which may lead to the development of local or systemic inflammation or organ-specific autoimmunity. In addition, endocrine disorders may also affect susceptibility to viral infections since endocrine hormones have immunoregulatory functions. This review provides a brief overview of the impact of viral infections on the human endocrine system in order to provide new avenues for the control of endocrine diseases.

Respiratory virus detection in returning travelers and pilgrims from the Middle East.

BACKGROUND: Pilgrims travelling to Saudi Arabia are commonly infected with respiratory viruses. Since the Middle East Respiratory Syndrome Coronavirus (MERS-CoV) emerged in 2012, patients with acute respiratory symptoms returning from an endemic area can be suspected to be infected by this virus. METHODS: 98 patients suspected to have MERS-CoV infection from 2014 to 2019 were included in this retrospective cohort study. Upper and lower respiratory tract samples were tested by real-time RT-PCR for the detection of MERS-CoV and other respiratory viruses. Routine microbiological analyses were also performed. Patient data were retrieved from laboratory and hospital databases retrospectively. RESULTS: All patients with suspected MERS-CoV infection travelled before their hospitalization. Most frequent symptoms were cough (94.4%) and fever (69.4%). 98 specimens were tested for MERS-CoV RNA and none of them was positive. Most frequently detected viruses were Enterovirus/Rhinovirus (40/83; 48.2%), Influenzavirus A (34/90; 37.8%) and B (11/90; 12.2%), H-CoV (229E and OC43 10/83; 12% and 7/83; 8.4%, respectively). CONCLUSION: From 2014 to 2019, none of 98 patients returning from endemic areas was MERS-CoV infected. However, infections with other respiratory viruses were frequent, especially with Enterovirus/Rhinoviruses and Influenzaviruses.

Serum neutralization of SARS coronavirus 2 Omicron sublineages BA.1 and BA.2 and cellular immune responses 3 months after booster vaccination.

OBJECTIVES: We investigated serum neutralizing activity against BA.1 and BA.2 Omicron sublineages and T cell response before and 3 months after administration of the booster vaccine in healthcare workers (HCWs). METHODS: HCWs aged 18-65 years who were vaccinated and received booster doses of the BNT162b2 vaccine were included. Anti-SARS coronavirus 2 IgG levels and cellular response (through interferon γ ELISpot assay) were evaluated in all participants, and neutralizing antibodies against Delta, BA.1, and BA.2 were evaluated in participants with at least one follow-up visit 1 or 3 months after the administration of the booster dose. RESULTS: Among 118 HCWs who received the booster dose, 102 and 84 participants attended the 1-month and 3-month visits, respectively. Before the booster vaccine dose, a low serum neutralizing activity against Delta, BA.1, and BA.2 was detectable in only 39/102 (38.2%), 8/102 (7.8%), and 12/102 (11.8%) participants, respectively. At 3 months, neutralizing antibodies against Delta, BA.1, and BA.2 were detected in 84/84 (100%), 79/84 (94%), and 77/84 (92%) participants, respectively. Geometric mean titres of neutralizing antibodies against BA.1 and BA.2 were 2.2-fold and 2.8-fold reduced compared with those for Delta. From 1 to 3 months after the administration of the booster dose, participants with a recent history of SARS coronavirus 2 infection (n = 21/84) had persistent levels of S1 reactive specific T cells and neutralizing antibodies against Delta and BA.2 and 2.2-fold increase in neutralizing antibodies against BA.1 (p 0.014). Conversely, neutralizing antibody titres against Delta (2.5-fold decrease, p < 0.0001), BA.1 (1.5-fold, p 0.02), and BA.2 (2-fold, p < 0.0001) declined from 1 to 3 months after the administration of the booster dose in individuals without any recent infection. DISCUSSION: The booster vaccine dose provided significant and similar response against BA.1 and BA.2 Omicron sublineages; however, the immune response declined in the absence of recent infection.

Immunogenicity of BNT162b2 vaccine booster against SARS-CoV-2 Delta and Omicron variants in nursing home residents: A prospective observational study in older adults aged from 68 to 98 years.

Background: The present study aimed to evaluate the persistent immunogenicity offered by a third dose of BNT162b2 against Delta and Omicron variants, in nursing home (NH) residents. Methods: In this monocenter prospective observational study, anti-spike IgG levels, S1 domain reactive T cell counts, serum neutralizing antibody titers against Delta and Omicron variants were compared before and up to three months after the BNT162b2 booster dose, in NH residents without COVID-19 (COVID-19 naive) or with COVID-19 prior to initial vaccination (COVID-19 recovered). Findings: 106 NH residents (median [interquartile range] age: 86·5 [81;91] years) were included. The booster dose induced a high increase of anti-spike antibody levels in all subjects (p < 0.0001) and a mild transient increase of specific T cells. Before the booster dose, Delta neutralization was detected in 19% (n = 8/43) and 88% (n = 37/42) of COVID-19 naive and COVID-19 recovered subjects, respectively. Three months after the booster dose, all NH residents developed and maintained a higher Delta neutralization (p < 0·0001). Before the booster dose, Omicron neutralization was detected in 5% (n = 2/43) and 55% (n = 23/42) of COVID-19 naive and COVID-19 recovered subjects, respectively, and three months after, in 84% and 95%, respectively. Neutralizing titers to Omicron were lower than to Delta in both groups with a 35-fold reduction compared to Delta. Interpretation: The booster dose restores high neutralization titers against Delta in all NH residents, and at a lower level against Omicron in a large majority of participants. Future studies are warranted to assess if repeated BNT162b2 booster doses or new specific vaccines might be considered for protecting such fragile patients against Omicron and/or future SARS-CoV-2 variants. Funding: French government through the Programme Investissement d'Avenir (I-SITE ULNE/ANR-16-IDEX-0004 ULNE) and the Label of COVID-19 National Research Priority (National Steering Committee on Therapeutic Trials and Other COVID-19 Research, CAPNET).

Evaluation of a rapid semiquantitative lateral flow assay for the prediction of serum neutralizing activity against SARS-CoV-2 variants.

BACKGROUND: Neutralizing antibodies (NAbs) against SARS-CoV-2 have been shown to correlate with protection against infection. Simple tools such as lateral flow assays (LFA) that can accurately measure NAbs may be useful for monitoring anti-SARS-CoV-2 immunity in the future. OBJECTIVES: We assessed the performance of the ichroma™ COVID-19 nAb test, a rapid semiquantitative LFA, for the prediction of serum neutralizing activity against SARS-CoV-2 variants. STUDY DESIGN: Serum samples were collected from COVID-19 recovered patients and vaccinated individuals. The result of the ichroma assay was provided as inhibition rate, and was compared to anti-SARS-CoV-2 IgG levels, and NAbs against Alpha, Delta and Omicron variants. RESULTS: A total of 90 sera from recovered unvaccinated patients and 209 sera from the vaccine cohort were included in this study. In post-infection samples, the ichroma inhbition rate was found to be correlated with IgG levels (ρ = 0.83), and with anti-Alpha NAbs levels (ρ = 0.78). In the vaccine cohort, a good correlation was also observed between the ichroma inhibition rate and IgG levels (ρ = 0.84), as well as NAbs against Alpha (ρ = 0.62), Delta (ρ = 0.88) and Omicron (ρ = 0.74). An ichroma inhbition rate of 77.2%, 90.8% and 99.6% accurately predicted neutralization against Alpha, Delta and Omicron variants respectively. CONCLUSIONS: The ichroma™ COVID-19 nAb assay, with appropriate variant cut-offs, can be useful for the monitoring of anti-SARS-CoV-2 immunization and may provide a rapid prediction of protection, especially in individuals with significant levels of NAbs.

Rapid syndromic testing for respiratory viral infections in children attending the emergency department during COVID-19 pandemic in Lille, France, 2021-2022.

OBJECTIVES: Viral respiratory infections are common in children, and usually associated with non-specific symptoms. Respiratory panel-based testing was implemented during the COVID-19 pandemic, for the rapid differentiation between SARS-CoV-2 and other viral infections, in children attending the emergency department (ED) of the teaching hospital of Lille, northern France, between February 2021 and January 2022. METHODS: Samples were collected using nasopharyngeal swabs. Syndromic respiratory testing was performed with two rapid multiplex molecular assays: the BioFire® Respiratory Panel 2.1 - plus (RP2.1 plus) or the QIAstat-Dx Respiratory SARS-CoV-2 Panel. SARS-CoV-2 variant was screened using mutation-specific PCR-based assays and genome sequencing. RESULTS: A total of 3517 children were included in the study. SARS-CoV-2 was detected in samples from 265 children (7.5%). SARS-CoV-2 infected patients were younger than those without SARS-CoV-2 infection (median age: 6 versus 12 months, p < 0.0001). The majority of infections (61.5%) were associated with the Omicron variant. The median weekly SARS-CoV-2 positivity rate ranged from 1.76% during the Alpha variant wave to 24.5% with the emergence of the Omicron variant. Most children (70.2%) were treated as outpatients, and seventeen patients were admitted to the intensive care unit. Other respiratory viruses were more frequently detected in SARS-CoV-2 negative children than in positive ones (82.1% versus 37.4%, p < 0.0001). Human rhinovirus/enterovirus and respiratory syncytial virus were the most prevalent in both groups. CONCLUSIONS: We observed a low prevalence of SARS-CoV-2 infection in children attending pediatric ED, despite the significant increase due to Delta and Omicron variants, and an important circulation of other respiratory viruses. Severe disease was overall rare in children.

Clofoctol inhibits SARS-CoV-2 replication and reduces lung pathology in mice.

Drug repurposing has the advantage of shortening regulatory preclinical development steps. Here, we screened a library of drug compounds, already registered in one or several geographical areas, to identify those exhibiting antiviral activity against SARS-CoV-2 with relevant potency. Of the 1,942 compounds tested, 21 exhibited a substantial antiviral activity in Vero-81 cells. Among them, clofoctol, an antibacterial drug used for the treatment of bacterial respiratory tract infections, was further investigated due to its favorable safety profile and pharmacokinetic properties. Notably, the peak concentration of clofoctol that can be achieved in human lungs is more than 20 times higher than its IC50 measured against SARS-CoV-2 in human pulmonary cells. This compound inhibits SARS-CoV-2 at a post-entry step. Lastly, therapeutic treatment of human ACE2 receptor transgenic mice decreased viral load, reduced inflammatory gene expression and lowered pulmonary pathology. Altogether, these data strongly support clofoctol as a therapeutic candidate for the treatment of COVID-19 patients.

Low Serum Levels of Interferon Alpha in COVID-19 Patients Are Associated with Older Age.

Innate immune response, especially type 1 interferon (IFN) response is considered to play a substantial role in the outcome of SARS-CoV-2 infection. A reduced and delayed IFN response has been associated with progression to severe COVID-19. In this study, we investigated levels of circulating IFNα and serum neutralizing activity in COVID-19 patients admitted to the intensive care unit. We found a significant association of levels of IFNα with age (p = 0.007). This association has also been observed in a cohort of COVID-19 outpatients with mild infection (p = 0.02). The impact of senescence on IFN response can explain the higher susceptibility of the elderly to severe COVID-19.

SARS-CoV-2 infection long time after full vaccination is related to a lack of neutralizing antibodies.

SARS-CoV-2 infections after COVID-19 vaccination are not unexpected, but those occurring more than 14 days after second vaccine dose need to be investigated. We describe a well-characterized infection which occurred almost 2 months after full vaccination, and provide the evidence of a link with a lack of anti-SARS-CoV-2 neutralizing antibodies.

Preanalytical Issues and Cycle Threshold Values in SARS-CoV-2 Real-Time RT-PCR Testing: Should Test Results Include These?

Since the emergence of SARS-CoV-2 pandemic, clinical laboratories worldwide are overwhelmed with SARS-CoV-2 testing using the current gold standard: real-time reverse-transcription polymerase chain reaction (RT-PCR) assays. The large numbers of suspected cases led to shortages in numerous reagents such as specimen transport and RNA extraction buffers. We try to provide some answers on how strongly preanalytical issues affect RT-PCR results by reviewing the utility of different transport buffer media and virus inactivation procedures and comparing the literature data with our own recent findings. We show that various viral inactivation procedures and transport buffers are available and are less of a bottleneck for PCR-based methods. However, efficient alternative lysis buffers remain more difficult to find, and several fast RT-PCR assays are not compatible with guanidine-containing media, making this aspect more of a challenge in the current crisis. Furthermore, the availability of different SARS-CoV-2-specific RT-PCR kits with different sensitivities makes the definition of a general cutoff level for the cycle threshold (Ct) value challenging. Only a few studies have considered how Ct values relate to viral infectivity and how preanalytical issues might affect viral infectivity and RNA detection. We review the current data on the correlation between Ct values and viral infectivity. The presence of the SARS-CoV-2 viral genome in its own is not sufficient proof of infectivity and caution is needed in evaluation of the infectivity of samples. The correlation between Ct values and viral infectivity revealed an RT-PCR cutoff value of 34 cycles for SARS-CoV-2 infectivity using a laboratory-developed RT-PCR assay targeting the RdRp gene. While ideally each clinical laboratory should perform its own correlation, we believe this perspective article could be a reference point for others, in particular medical doctors and researchers interested in COVID-19 diagnostics, and a first step toward harmonization.

Fluoxetine Can Inhibit SARS-CoV-2 In Vitro.

An outbreak of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) resulted in the coronavirus disease pandemic, drastically affecting global health and economy. Though the understanding of the disease has improved, fighting the virus remains challenging. One of the strategies is repurposing existing drugs as inhibitors of SARS-CoV-2. Fluoxetine (FLX), a selective serotonin reuptake inhibitor, reportedly inhibits the replication of RNA viruses, especially Coxsackieviruses B (CVB), such as CV-B4 in vitro and in vivo. Therefore, in this study, we investigated the in vitro antiviral activity of FLX against SARS-CoV-2 in a model of acute infection. When 10 µM of FLX was added to SARS-CoV-2-infected Vero E6 cells, the virus-induced cytopathic effect was not observed. In this model, the level of infectious particles in the supernatant was lower than that in controls. The level was below the limit of detection of the assay up to day 3 post-infection when FLX was administered before viral inoculation or simultaneously followed by daily inoculation. In conclusion, FLX can inhibit SARS-CoV-2 in vitro. Further studies are needed to investigate the potential value of FLX to combat SARS-CoV-2 infections, treat SARS-CoV-2-induced diseases, and explain the antiviral mechanism of this molecule to pave way for novel treatment strategies.

Spatial and Temporal Virus Load Dynamics of SARS-CoV-2: A Single-Center Cohort Study.

The severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has caused an ongoing pandemic. Reverse transcription polymerase chain reaction (RT-PCR) is the gold standard for the detection of SARS-CoV-2 and has been applied to different specimen types. Understanding the virus load and virus detection frequency in different specimen types is important to improve diagnosis and estimate the duration of potential infectivity. We conducted a retrospective single-center cohort study on hospitalized and outpatients with SARS-CoV-2 infection. We analyzed the frequency of virus detection, virus load, and duration of the virus excretion in upper and lower respiratory specimens as well as stool and plasma. We found that the frequency of SARS-CoV-2 detection, the virus load, and duration of virus excretion was higher in lower respiratory tract (LRT) than in upper respiratory tract (URT) specimens. The duration of virus excretion was longer in patients requiring intensive care unit (ICU) admission. In conclusion, LRT specimens are the most appropriate specimen type for the detection and follow-up of SARS-CoV-2 infection. Duration of virus excretion is longer in severe cases of SARS-CoV-2 infection.

Fluoxetine Can Inhibit SARS-CoV-2 In Vitro

An outbreak of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) resulted in the coronavirus disease pandemic, drastically affecting global health and economy. Though the understanding of the disease has improved, fighting the virus remains challenging. One of the strategies is repurposing existing drugs as inhibitors of SARS-CoV-2. Fluoxetine (FLX), a selective serotonin reuptake inhibitor, reportedly inhibits the replication of RNA viruses, especially Coxsackieviruses B (CVB), such as CV-B4 in vitro and in vivo. Therefore, in this study, we investigated the in vitro antiviral activity of FLX against SARS-CoV-2 in a model of acute infection. When 10 μM of FLX was added to SARS-CoV-2-infected Vero E6 cells, the virus-induced cytopathic effect was not observed. In this model, the level of infectious particles in the supernatant was lower than that in controls. The level was below the limit of detection of the assay up to day 3 post-infection when FLX was administered before viral inoculation or simultaneously followed by daily inoculation. In conclusion, FLX can inhibit SARS-CoV-2 in vitro. Further studies are needed to investigate the potential value of FLX to combat SARS-CoV-2 infections, treat SARS-CoV-2-induced diseases, and explain the antiviral mechanism of this molecule to pave way for novel treatment strategies.