Uncover a microbiota signature of upper respiratory tract in patients with SARS-CoV-2 + .

The outbreak of Coronavirus disease 2019 (COVID-19), caused by SARS-CoV-2, forced us to face a pandemic with unprecedented social, economic, and public health consequences. Several nations have launched campaigns to immunize millions of people using various vaccines to prevent infections. Meanwhile, therapeutic approaches and discoveries continuously arise; however, identifying infected patients that are going to experience the more severe outcomes of COVID-19 is still a major need, to focus therapeutic efforts, reducing hospitalization and mitigating drug adverse effects. Microbial communities colonizing the respiratory tract exert significant effects on host immune responses, influencing the susceptibility to infectious agents. Through 16S rDNAseq we characterized the upper airways' microbiota of 192 subjects with nasopharyngeal swab positive for SARS-CoV-2. Patients were divided into groups based on the presence of symptoms, pneumonia severity, and need for oxygen therapy or intubation. Indeed, unlike most of the literature, our study focuses on identifying microbial signatures predictive of disease progression rather than on the probability of infection itself, for which a consensus is lacking. Diversity, differential abundance, and network analysis at different taxonomic levels were synergistically adopted, in a robust bioinformatic pipeline, highlighting novel possible taxa correlated with patients' disease progression to intubation.

Virus-Derived DNA Forms Mediate the Persistent Infection of Tick Cells by Hazara Virus and Crimean-Congo Hemorrhagic Fever Virus.

Crimean-Congo hemorrhagic fever (CCHF) is a severe disease of humans caused by CCHF virus (CCHFV), a biosafety level (BSL)-4 pathogen. Ticks of the genus Hyalomma are the viral reservoir, and they represent the main vector transmitting the virus to its hosts during blood feeding. We have previously shown that CCHFV can persistently infect Hyalomma-derived tick cell lines. However, the mechanism allowing the establishment of persistent viral infections in ticks is still unknown. Hazara virus (HAZV) can be used as a BSL-2 model virus instead of CCHFV to study virus/vector interactions. To investigate the mechanism behind the establishment of a persistent infection, we developed an in vitro model with Hyalomma-derived tick cell lines and HAZV. As expected, HAZV, like CCHFV, persistently infects tick cells without any sign of cytopathic effect, and the infected cells can be cultured for more than 3 years. Most interestingly, we demonstrated the presence of short viral-derived DNA forms (vDNAs) after HAZV infection. Furthermore, we demonstrated that the antiretroviral drug azidothymine triphosphate could inhibit the production of vDNAs, suggesting that vDNAs are produced by an endogenous retrotranscriptase activity in tick cells. Moreover, we collected evidence that vDNAs are continuously synthesized, thereby downregulating viral replication to promote cell survival. Finally, vDNAs were also detected in CCHFV-infected tick cells. In conclusion, vDNA synthesis might represent a strategy to control the replication of RNA viruses in ticks allowing their persistent infection. IMPORTANCE Crimean-Congo hemorrhagic fever (CCHF) is an emerging tick-borne viral disease caused by CCHF virus (CCHFV). Ticks of the genus Hyalomma can be persistently infected with CCHFV representing the viral reservoir, and the main vector for viral transmission. Here we showed that tick cells infected with Hazara virus, a nonpathogenic model virus closely related to CCHFV, contained short viral-derived DNA forms (vDNAs) produced by endogenous retrotranscriptase activity. vDNAs are transitory molecules requiring viral RNA replication for their continuous synthesis. Interestingly, vDNA synthesis seemed to be correlated with downregulation of viral replication and promotion of tick cell viability. We also detected vDNAs in CCHFV-infected tick cells suggesting that they could represent a key element in the cell response to nairovirus infection and might represent a more general mechanism of innate immunity against RNA viral infection.

Lacticaseibacillus paracasei DG enhances the lactoferrin anti-SARS-CoV-2 response in Caco-2 cells.

The novel severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is causing the ongoing global pandemic of coronavirus disease 2019 (COVID-19), which primarily manifests with respiratory distress and may also lead to symptoms associated with the gastrointestinal tract. Probiotics are living microorganisms that have been shown to confer immune benefits. In this study, we investigated the immunomodulatory effects and anti-SARS-CoV-2 activity of three different Lacticaseibacillus probiotic strains, either alone or in combination with lactoferrin, using the intestinal epithelial Caco-2 cell line. Our results revealed that the Lacticaseibacillus paracasei DG strain significantly induced the expression of genes involved in protective antiviral immunity and prevented the expression of proinflammatory genes triggered by SARS-CoV-2 infection. Moreover, L. paracasei DG significantly inhibited SARS-CoV-2 infection in vitro. L. paracasei DG also positively affected the antiviral immune activity of lactoferrin and significantly augmented its anti-SARS-CoV-2 activity in Caco-2 intestinal epithelial cells. Overall, our work shows that the probiotic strain L. paracasei DG is a promising candidate that exhibits prophylactic potential against SARS-CoV-2 infection.

Hypothiocyanite and Hypothiocyanite/Lactoferrin Mixture Exhibit Virucidal Activity In Vitro against SARS-CoV-2.

SARS-CoV-2 replicates efficiently in the upper airways during the prodromal stage, resulting in environmental viral shedding from patients with active COVID-19 as well as from asymptomatic individuals. There is a need to find pharmacological interventions to mitigate the spread of COVID-19. Hypothiocyanite and lactoferrin are molecules of the innate immune system with a large spectrum cidal activity. The Food and Drug Administration and the European Medicines Agency designated the hypothiocyanite and lactoferrin combination as an orphan drug. We report an in vitro study showing that micromolar concentrations of hypothiocyanite exhibit dose- and time-dependent virucidal activity against SARS-CoV-2 and that the latter is slightly enhanced by the simultaneous presence of lactoferrin.

Protective Effects of Lactoferrin against SARS-CoV-2 Infection In Vitro.

SARS-CoV-2 is a newly emerging virus that currently lacks curative treatments. Lactoferrin (LF) is a naturally occurring non-toxic glycoprotein with broad-spectrum antiviral, immunomodulatory and anti-inflammatory effects. In this study, we assessed the potential of LF in the prevention of SARS-CoV-2 infection in vitro. Antiviral immune response gene expression was analyzed by qRT-PCR in uninfected Caco-2 intestinal epithelial cells treated with LF. An infection assay for SARS-CoV-2 was performed in Caco-2 cells treated or not with LF. SARS-CoV-2 titer was determined by qRT-PCR, plaque assay and immunostaining. Inflammatory and anti-inflammatory cytokine production was determined by qRT-PCR. LF significantly induced the expression of IFNA1, IFNB1, TLR3, TLR7, IRF3, IRF7 and MAVS genes. Furthermore, LF partially inhibited SARS-CoV-2 infection and replication in Caco-2 intestinal epithelial cells. Our in vitro data support LF as an immune modulator of the antiviral immune response with moderate effects against SARS-CoV-2 infection.