A single-dose live-attenuated YF17D-vectored SARS-CoV-2 vaccine candidate.

The expanding pandemic of coronavirus disease 2019 (COVID-19) requires the development of safe, efficacious and fast-acting vaccines. Several vaccine platforms are being leveraged for a rapid emergency response1. Here we describe the development of a candidate vaccine (YF-S0) for severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) that uses live-attenuated yellow fever 17D (YF17D) vaccine as a vector to express a noncleavable prefusion form of the SARS-CoV-2 spike antigen. We assess vaccine safety, immunogenicity and efficacy in several animal models. YF-S0 has an excellent safety profile and induces high levels of SARS-CoV-2 neutralizing antibodies in hamsters (Mesocricetus auratus), mice (Mus musculus) and cynomolgus macaques (Macaca fascicularis), and-concomitantly-protective immunity against yellow fever virus. Humoral immunity is complemented by a cellular immune response with favourable T helper 1 polarization, as profiled in mice. In a hamster model2 and in macaques, YF-S0 prevents infection with SARS-CoV-2. Moreover, a single dose conferred protection from lung disease in most of the vaccinated hamsters within as little as 10 days. Taken together, the quality of the immune responses triggered and the rapid kinetics by which protective immunity can be attained after a single dose warrant further development of this potent SARS-CoV-2 vaccine candidate.

Favipiravir at high doses has potent antiviral activity in SARS-CoV-2-infected hamsters, whereas hydroxychloroquine lacks activity.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) rapidly spread around the globe after its emergence in Wuhan in December 2019. With no specific therapeutic and prophylactic options available, the virus has infected millions of people of which more than half a million succumbed to the viral disease, COVID-19. The urgent need for an effective treatment together with a lack of small animal infection models has led to clinical trials using repurposed drugs without preclinical evidence of their in vivo efficacy. We established an infection model in Syrian hamsters to evaluate the efficacy of small molecules on both infection and transmission. Treatment of SARS-CoV-2-infected hamsters with a low dose of favipiravir or hydroxychloroquine with(out) azithromycin resulted in, respectively, a mild or no reduction in virus levels. However, high doses of favipiravir significantly reduced infectious virus titers in the lungs and markedly improved lung histopathology. Moreover, a high dose of favipiravir decreased virus transmission by direct contact, whereas hydroxychloroquine failed as prophylaxis. Pharmacokinetic modeling of hydroxychloroquine suggested that the total lung exposure to the drug did not cause the failure. Our data on hydroxychloroquine (together with previous reports in macaques and ferrets) thus provide no scientific basis for the use of this drug in COVID-19 patients. In contrast, the results with favipiravir demonstrate that an antiviral drug at nontoxic doses exhibits a marked protective effect against SARS-CoV-2 in a small animal model. Clinical studies are required to assess whether a similar antiviral effect is achievable in humans without toxic effects.

Molecular characterization of group A rotavirus strains detected in alpacas (Vicugna pacos) from Peru.

The alpaca is a very important social and economic resource for the production of fibre and meat for Andean communities. Peru is the main producer of alpacas. Group A rotavirus (RVA) has been sporadically detected in alpacas. In this study, a total of 1423 faecal samples from alpacas from different locations of the Puno department in Peru were collected and analysed by an antigen-capture ELISA in order to detect RVA. Four per cent of the samples were RVA-positive (57/1423). The genotype constellation of three selected alpaca RVA strains were G3/8 P[1/14]-I2-R2/5-C2/3-M2/3-A17-N2/3-T6-E3-H3. Two of the analysed strains presented a bovine-like genotype constellation, whereas the third strain presented six segments belonging to the AU-1-like genogroup (G3, M3, C3, N3, T3 and E3), suggesting reassorting events. Monitoring of the sanitary health of juvenile alpacas is essential to reduce the rates of neonatal mortality and for the development of preventive health strategies.

Phylogenetic analysis of open reading frame of 11 gene segments of novel human-bovine reassortant RVA G6P[1] strain in Pakistan.

Multiple Rotavirus A (RVA) strains are linked with gastrointestinal infections in children that fall in age bracket of 0 to 60 months. However, the problem is augmented with emergence of unique strains that reassort with RVA strains of animal origin. The study describes the sequence analysis of a rare G6P[1] rotavirus strain isolated from a less than 1 year old child, during rotavirus surveillance in Rawalpindi district, Pakistan in 2010. Extracted RNA from fecal specimen was subjected to high throughput RT-PCR for structural and nonstructural gene segments. The complete rotavirus genome of one isolate RVA/Human-wt/PAK/PAK99/2010/G6P[1] was sequenced for phylogenetic analysis to elucidate the evolutionary linkages and origin. Full genome examination of novel strain RVA/Human-wt/PAK/PAK99/2010/G6P[1] revealed the unique genotype assemblage: G6-P[1]-I2-R2-C2-M2-A3-N2-T6-E2-H1. The evolutionary analyses of VP7, VP4, NSP1 and NSP3 gene segments revealed that PAK99 clustered with bovine, or cattle-like rotavirus strains from other closely related species, in the genotypes G6, P[1], A3 and T6 respectively. Gene segments VP6, VP1, VP2, VP3, NSP2 and NSP4 all possessed the DS-1-like bovine genotype 2 and bovine (-like) RVA strains instead of RVA strains having human origin. However, the NSP5 gene was found to cluster closely with contemporary human Wa-like rotavirus strains of H1 genotype. This is the first report on bovine-human (Wa-like reassortant) genotype constellation of G6P[1] strain from a human case in Pakistan (and the second description worldwide). Our results emphasize the significance of incessant monitoring of circulating RVA strains in humans and animals for better understanding of RV evolution.

Evidence of zoonotic transmission of VP6 and NSP4 genes into human species A rotaviruses isolated in Pakistan in 2010.

Introduction of animal group A rotavirus (RVA) gene segments into the human RVA population is a major factor shaping the genetic landscape of human RVA strains. The VP6 and NSP4 genes of 74 G/P-genotyped RVA isolates collected in Rawalpindi during 2010 were analyzed, revealing the presence of VP6 genotypes I1 (60.8%) and I2 (39.2%) and NSP4 genotypes E1 (60.8%), E2 (28.3%) and E-untypable (10.8%) among the circulating human RVA strains. The typical human RVA combinations I1E1 and I2E2 were found in 59.4% and 24.3% of the cases, respectively, whereas 5.4% of the RVA strains were reassortants, i.e., either I1E2 or I2E1. The phylogeny of the NSP4 gene showed that one G2P[4] and two G1P[6] RVA strains clustered with porcine E1 RVA strains or RVA strains that were considered to be (partially) of porcine origin. In addition, the NSP4 gene segment of the unusual human G6P[1] RVA strains clustered closely with bovine E2 RVA strains, further strengthening the hypothesis of an interspecies transmission event. The study further demonstrates the role of genomic re-assortment and the involvement of interspecies transmission in the evolution of human RVA strains. The VP6 and NSP4 nucleotide sequences analyzed in the study received the GenBank accession numbers KC846908- KC846971 and KC846972-KC847037, respectively.

A Single Nucleoside Viral Polymerase Inhibitor Against Norovirus, Rotavirus, and Sapovirus-Induced Diarrhea.

A safe and highly efficient antiviral is needed for the prophylaxis and/or treatment of viral diarrhea. We here demonstrate the in vitro antiviral activity of four 2'-C-methyl nucleoside analogues against noro-, rota-, and sapoviruses. The most potent nucleoside analogue, 7-deaza-2'-C-methyladenosine, inhibits replication of these viruses with a 50% effective concentration < 5 µM. Mechanistically, we demonstrate that the 2'-C-methyl nucleoside analogues act by inhibiting transcription of the rotavirus genome. This provides the first evidence that a single viral-diarrhea-targeted treatment can be developed through a viral-polymerase-targeting small molecule.

Highly diverse population of Picornaviridae and other members of the Picornavirales, in Cameroonian fruit bats.

BACKGROUND: The order Picornavirales represents a diverse group of positive-stranded RNA viruses with small non-enveloped icosahedral virions. Recently, bats have been identified as an important reservoir of several highly pathogenic human viruses. Since many members of the Picornaviridae family cause a wide range of diseases in humans and animals, this study aimed to characterize members of the order Picornavirales in fruit bat populations located in the Southwest region of Cameroon. These bat populations are frequently in close contact with humans due to hunting, selling and eating practices, which provides ample opportunity for interspecies transmissions. RESULTS: Fecal samples from 87 fruit bats (Eidolon helvum and Epomophorus gambianus), were combined into 25 pools and analyzed using viral metagenomics. In total, Picornavirales reads were found in 19 pools, and (near) complete genomes of 11 picorna-like viruses were obtained from 7 of these pools. The picorna-like viruses possessed varied genomic organizations (monocistronic or dicistronic), and arrangements of gene cassettes. Some of the viruses belonged to established families, including the Picornaviridae, whereas others clustered distantly from known viruses and most likely represent novel genera and families. Phylogenetic and nucleotide composition analyses suggested that mammals were the likely host species of bat sapelovirus, bat kunsagivirus and bat crohivirus, whereas the remaining viruses (named bat iflavirus, bat posalivirus, bat fisalivirus, bat cripavirus, bat felisavirus, bat dicibavirus and bat badiciviruses 1 and 2) were most likely diet-derived. CONCLUSION: The existence of a vast genetic variability of picorna-like viruses in fruit bats may increase the probability of spillover infections to humans especially when humans and bats have direct contact as the case in this study site. However, further screening for these viruses in humans will fully indicate their zoonotic potential.

Human P[6] Rotaviruses From Sub-Saharan Africa and Southeast Asia Are Closely Related to Those of Human P[4] and P[8] Rotaviruses Circulating Worldwide.

BACKGROUND: P[6] rotaviruses have been circulating with a high prevalence in African and, to a more limited extent, Asian countries, but they have not been highly prevalent in other parts of the world. METHODS: To investigate the genomic relationship between African and Asian human P[6] rotaviruses and P[4] and P[8] rotaviruses circulating worldwide, we sequenced 39 P[6] strains, collected in Ghana, Mali, Kenya and Bangladesh, providing the largest data set of P[6] rotavirus genomes isolated in low-income countries or anywhere else in the world that has been published thus far. RESULTS: Overall, the data indicate that the genetic backbone of human P[6] strains from the low-income countries are similar to those of P[4] or P[8] strains circulating worldwide. CONCLUSIONS: The observation that gene segment 4 is the main differentiator between human P[6] and non-P[6] strains suggests that the VP4 spike protein is most likely one of the main reasons preventing the rapid spread of P[6] strains to the rest of the world despite multiple introductions. These observations reinforce previous findings about the receptor specificity of P[6] rotavirus strains.

Emerging OP354-Like P[8] Rotaviruses Have Rapidly Dispersed from Asia to Other Continents.

The majority of human group A rotaviruses possess the P[8] VP4 genotype. Recently, a genetically distinct subtype of the P[8] genotype, also known as OP354-like P[8] or lineage P[8]-4, emerged in several countries. However, it is unclear for how long the OP354-like P[8] gene has been circulating in humans and how it has spread. In a global collaborative effort 98 (near-)complete OP354-like P[8] VP4 sequences were obtained and used for phylogeographic analysis to determine the viral migration patterns. During the sampling period, 1988-2012, we found that South and East Asia acted as a source from which strains with the OP354-like P[8] gene were seeded to Africa, Europe, and North America. The time to the most recent common ancestor (TMRCA) of all OP354-like P[8] genes was estimated at 1987. However, most OP354-like P[8] strains were found in three main clusters with TMRCAs estimated between 1996 and 2001. The VP7 gene segment of OP354-like P[8] strains showed evidence of frequent reassortment, even in localized epidemics, suggesting that OP354-like P[8] genes behave in a similar manner on the evolutionary level as other P[8] subtypes. The results of this study suggest that OP354-like P[8] strains have been able to disperse globally in a relatively short time period. This, in combination with a relatively large genetic distance to other P[8] subtypes, might result in a lower vaccine effectiveness, underscoring the need for a continued surveillance of OP354-like P[8] strains, especially in countries where rotavirus vaccination programs are in place.

Complete genome characterization of recent and ancient Belgian pig group A rotaviruses and assessment of their evolutionary relationship with human rotaviruses.

UNLABELLED: Group A rotaviruses (RVAs) are an important cause of diarrhea in young pigs and children. An evolutionary relationship has been suggested to exist between pig and human RVAs. This hypothesis was further investigated by phylogenetic analysis of the complete genomes of six recent (G2P[27], G3P[6], G4P[7], G5P[7], G9P[13], and G9P[23]) and one historic (G1P[7]) Belgian pig RVA strains and of all completely characterized pig RVAs from around the globe. In contrast to the large diversity of genotypes found for the outer capsid proteins VP4 and VP7, a relatively conserved genotype constellation (I5-R1-C1-M1-A8-N1-T7-E1-H1) was found for the other 9 genes in most pig RVA strains. VP1, VP2, VP3, NSP2, NSP4, and NSP5 genes of porcine RVAs belonged to genotype 1, which is shared with human Wa-like RVAs. However, for most of these gene segments, pig strains clustered distantly from human Wa-like RVAs, indicating that viruses from both species have entered different evolutionary paths. However, VP1, VP2, and NSP3 genes of some archival human strains were moderately related to pig strains. Phylogenetic analysis of the VP6, NSP1, and NSP3 genes, as well as amino acid analysis of the antigenic regions of VP7, further confirmed this evolutionary segregation. The present results also indicate that the species barrier is less strict for pig P[6] strains but that chances for successful spread of these strains in the human population are hampered by the better adaptation of pig RVAs to pig enterocytes. However, future surveillance of pig and human RVA strains is warranted. IMPORTANCE: Rotaviruses are an important cause of diarrhea in many species, including pigs and humans. Our understanding of the evolutionary relationship between rotaviruses from both species is limited by the lack of genomic data on pig strains. In this study, recent and ancient Belgian pig rotavirus isolates were sequenced, and their evolutionary relationship with human Wa-like strains was investigated. Our data show that Wa-like human and pig strains have entered different evolutionary paths. Our data indicate that pig P[6] strains form the most considerable risk for interspecies transmission to humans. However, efficient spread of pig strains in the human population is most likely hampered by the adaptation of some crucial viral proteins to the cellular machinery of pig enterocytes. These data allow a better understanding of the risk for direct interspecies transmission events and the emergence of pig rotaviruses or pig-human reassortants in the human population.

Reassortment among picobirnaviruses found in wolves.

We conducted a viral metagenomics study in diarrheic free-ranging wolves in Portugal, revealing for the first time the presence of reassortant picobirnaviruses. These viruses shared identical capsid segments together with diverse RNA-dependent RNA polymerase segments. Even though causality between these picobirnaviruses and diarrhea could not be established, the study nonetheless confirms for the first time that wolves are a potential reservoir for picobirnaviruses, which might play a role as enteric pathogens.

Genetic diversity in three bovine-like human G8P[14] and G10P[14] rotaviruses suggests independent interspecies transmission events.

The group A rotavirus (RVA) P[14] genotype has been detected sporadically in humans and is thought to be acquired through zoonotic transmission. The present study describes the full-length genome analysis of two G8P[14] and one G10P[14] human RVAs detected in Italy. The strains possessed the typical bovine-like I2-R2-C2-M2-A3/A11-N2-T6-E2-H3 genotype constellation. All the segments of the two G8P[14] RVAs were most closely related to bovine(-like) strains but were relatively distant to each other, suggesting two independent interspecies transmission events. Likewise, the G10P[14] RVA gene segments were most similar to bovine(-like) RVAs but distinct from the G8 strains. The history of these strains probably involved the interspecies transmission of these viruses to humans from an as-yet-unidentified animal host, without evidence of reassortment events involving human RVAs. These results reinforce the potential of animal viruses to cross the host-species barrier, causing disease and increased viral genetic diversity in humans.

Fecal virome analysis of three carnivores reveals a novel nodavirus and multiple gemycircularviruses.

BACKGROUND: More knowledge about viral populations in wild animals is needed in order to better understand and assess the risk of zoonotic diseases. In this study we performed viral metagenomic analysis of fecal samples from three healthy carnivores: a badger (Meles meles), a mongoose (Herpestes ichneumon) and an otter (Lutra lutra) from Portugal. RESULTS: We detected the presence of novel highly divergent viruses in the fecal material of the carnivores analyzed, such as five gemycircularviruses. Four of these gemycircularviruses were found in the mongoose and one in the badger. In addition we also identified an RNA-dependent RNA polymerase gene from a putative novel member of the Nodaviridae family in the fecal material of the otter. CONCLUSIONS: Together these results underline that many novel viruses are yet to be discovered and that fecal associated viruses are not always related to disease. Our study expands the knowledge of viral species present in the gut, although the interpretation of the true host species of such novel viruses needs to be reviewed with great caution.

Evolution of DS-1-like human G2P[4] rotaviruses assessed by complete genome analyses.

Group A rotaviruses (RVAs) are a leading cause of viral gastroenteritis in children, with G2P[4] RVA being one of the most common human strains worldwide. The complete genome sequences of nine G2P[4] RVA strains, selected from a 26-year archival collection (1985-2011) established in Palermo, Italy, were determined. A strain associated with a peak of G2P[4] RVA activity in 1996 resembled a reassortant strain identified in Kenya in 1982 and differed completely in genomic make up from more recent strains that circulated during 2004-2011. Conversely, the 2004-2011 G2P[4] RVAs were genetically more similar to contemporary RVA strains circulating globally. Recent G2P[4] strains possessed either single or multiple genome segments (VP1, VP3 and/or NSP4) likely derived from ruminant viruses through intra-genotype reassortment. Amino acid substitutions were selected and maintained over time in the VP7 and VP8* antigenic proteins, allowing the circulation of two contemporary G2P[4] variants to be distinguished. Altogether, these findings suggest that major changes in the genomic composition of recent G2P[4] RVAs occurred in the early 2000s, leading to the appearance of a novel variant of the DS-1-like genotype constellation. Whether the modifications observed in the neutralizing antigens and in the genome composition of modern G2P[4] RVAs may affect the long-term effectiveness of the vaccination programmes remains to be explored.

Feline origin of rotavirus strain, Tunisia, 2008.

In Tunisia in 2008, an unusual G6P[9] rotavirus, RVA/human-wt/TUN/17237/2008/G6P[9], rarely found in humans, was detected in a child. To determine the origin of this strain, we conducted phylogenetic analyses and found a unique genotype constellation resembling rotaviruses belonging to the feline BA222-like genotype constellation. The strain probably resulted from direct cat-to-human transmission.

Molecular Epidemiology of Rotavirus A in Calves: Evolutionary Analysis of a Bovine G8P[11] Strain and Spatio-Temporal Dynamics of G6 Lineages in the Americas.

Rotavirus A (RVA) causes diarrhea in calves and frequently possesses the G6 and P[5]/P[11] genotypes, whereas G8 is less common. We aimed to compare RVA infections and G/P genotypes in beef and dairy calves from major livestock regions of Argentina, elucidate the evolutionary origin of a G8 strain and analyze the G8 lineages, infer the phylogenetic relationship of RVA field strains, and investigate the evolution and spatio-temporal dynamics of the main G6 lineages in American countries. Fecal samples (n = 422) from diarrheic (beef, 104; dairy, 137) and non-diarrheic (beef, 78; dairy, 103) calves were analyzed by ELISA and semi-nested multiplex RT-PCR. Sequencing, phylogenetic, phylodynamic, and phylogeographic analyses were performed. RVA infections were more frequent in beef (22.0%) than in dairy (14.2%) calves. Prevalent genotypes and G6 lineages were G6(IV)P[5] in beef (90.9%) and G6(III)P[11] (41.2%) or mixed genotypes (23.5%) in dairy calves. The only G8 strain was phylogenetically related to bovine and artiodactyl bovine-like strains. Re-analyses inside the G8 genotype identified G8(I) to G8(VIII) lineages. Of all G6 strains characterized, the G6(IV)P[5](I) strains from "Cuenca del Salado" (Argentina) and Uruguay clustered together. According to farm location, a clustering pattern for G6(IV)P[5] strains of beef farms was observed. Both G6 lineage strains together revealed an evolutionary rate of 1.24 × 10-3 substitutions/site/year, and the time to the most recent common ancestor was dated in 1853. The most probable ancestral locations were Argentina in 1981 for G6(III) strains and the USA in 1940 for G6(IV) strains. The highest migration rates for both G6 lineages together were from Argentina to Brazil and Uruguay. Altogether, the epidemiology, genetic diversity, and phylogeny of RVA in calves can differ according to the production system and farm location. We provide novel knowledge about the evolutionary origin of a bovine G8P[11] strain. Finally, bovine G6 strains from American countries would have originated in the USA nearly a century before its first description.

Complete molecular genome analyses of equine rotavirus A strains from different continents reveal several novel genotypes and a largely conserved genotype constellation.

In this study, the complete genome sequences of seven equine group A rotavirus (RVA) strains (RVA/Horse-tc/GBR/L338/1991/G13P[18], RVA/Horse-wt/IRL/03V04954/2003/G3P[12] and RVA/Horse-wt/IRL/04V2024/2004/G14P[12] from Europe; RVA/Horse-wt/ARG/E30/1993/G3P[12], RVA/Horse-wt/ARG/E403/2006/G14P[12] and RVA/Horse-wt/ARG/E4040/2008/G14P[12] from Argentina; and RVA/Horse-wt/ZAF/EqRV-SA1/2006/G14P[12] from South Africa) were determined. Multiple novel genotypes were identified and genotype numbers were assigned by the Rotavirus Classification Working Group: R9 (VP1), C9 (VP2), N9 (NSP2), T12 (NSP3), E14 (NSP4), and H7 and H11 (NSP5). The genotype constellation of L338 was unique: G13-P[18]-I6-R9-C9-M6-A6-N9-T12-E14-H11. The six remaining equine RVA strains showed a largely conserved genotype constellation: G3/G14-P[12]-I2/I6-R2-C2-M3-A10-N2-T3-E2/E12-H7, which is highly divergent from other known non-equine RVA genotype constellations. Phylogenetic analyses revealed that the sequences of these equine RVA strains are related distantly to non-equine RVA strains, and that at least three lineages exist within equine RVA strains. A small number of reassortment events were observed. Interestingly, the three RVA strains from Argentina possessed the E12 genotype, whereas the three RVA strains from Ireland and South Africa possessed the E2 genotype. The unusual E12 genotype has until now only been described in Argentina among RVA strains collected from guanaco, cattle and horses, suggesting geographical isolation of this NSP4 genotype. This conserved genetic configuration of equine RVA strains could be useful for future vaccine development or improvement of currently used equine RVA vaccines.

Genetic analyses reveal differences in the VP7 and VP4 antigenic epitopes between human rotaviruses circulating in Belgium and rotaviruses in Rotarix and RotaTeq.

Two live-attenuated rotavirus group A (RVA) vaccines, Rotarix (G1P[8]) and RotaTeq (G1-G4, P[8]), have been successfully introduced in many countries worldwide, including Belgium. The parental RVA strains used to generate the vaccines were isolated more than 20 years ago in France (G4 parental strain in RotaTeq) and the United States (all other parental strains). At present, little is known about the relationship between currently circulating human RVAs and the vaccine strains. In this study, we determined sequences for the VP7 and VP4 outer capsid proteins of representative G1P[8], G2P[4], G3P[8], G4P[8], G9P[8], and G12P[8] RVAs circulating in Belgium during 2007 to 2009. The analyses showed that multiple amino acid differences existed between the VP7 and VP4 antigenic epitopes of the vaccine viruses and the Belgian isolates, regardless of their G and P genotypes. However, the highest variability was observed among the circulating G1P[8] RVA strains and the G1 and P[8] components of both RVA vaccines. In particular, RVA strains of the P[8] lineage 4 (OP354-like) showed a significant number of amino acid differences with the P[8] VP4 of both vaccines. In addition, the circulating Belgian G3 RVA strains were found to possibly possess an extra N-linked glycosylation site compared to the G3 RVA vaccine strain of RotaTeq. These results indicate that the antigenic epitopes of RVA strains contained in the vaccines differ substantially from those of the currently circulating RVA strains in Belgium. Over time, these differences might result in selection for strains that escape the RVA neutralizing-antibody pressure induced by vaccines.

Atypical response to bacterial coinfection and persistent neutrophilic bronchoalveolar inflammation distinguish critical COVID-19 from influenza.

Neutrophils are recognized as important circulating effector cells in the pathophysiology of severe coronavirus disease 2019 (COVID-19). However, their role within the inflamed lungs is incompletely understood. Here, we collected bronchoalveolar lavage (BAL) fluids and parallel blood samples of critically ill COVID-19 patients requiring invasive mechanical ventilation and compared BAL fluid parameters with those of mechanically ventilated patients with influenza, as a non-COVID-19 viral pneumonia cohort. Compared with those of patients with influenza, BAL fluids of patients with COVID-19 contained increased numbers of hyperactivated degranulating neutrophils and elevated concentrations of the cytokines IL-1ß, IL-1RA, IL-17A, TNF-α, and G-CSF; the chemokines CCL7, CXCL1, CXCL8, CXCL11, and CXCL12α; and the protease inhibitors elafin, secretory leukocyte protease inhibitor, and tissue inhibitor of metalloproteinases 1. In contrast, α-1 antitrypsin levels and net proteolytic activity were comparable in COVID-19 and influenza BAL fluids. During antibiotic treatment for bacterial coinfections, increased BAL fluid levels of several activating and chemotactic factors for monocytes, lymphocytes, and NK cells were detected in patients with COVID-19 whereas concentrations tended to decrease in patients with influenza, highlighting the persistent immunological response to coinfections in COVID-19. Finally, the high proteolytic activity in COVID-19 lungs suggests considering protease inhibitors as a treatment option.

HIV protease inhibitors Nelfinavir and Lopinavir/Ritonavir markedly improve lung pathology in SARS-CoV-2-infected Syrian hamsters despite lack of an antiviral effect.

Nelfinavir is an HIV protease inhibitor that has been widely prescribed as a component of highly active antiretroviral therapy, and has been reported to exert in vitro antiviral activity against SARS-CoV-2. We here assessed the effect of Nelfinavir in a SARS-CoV-2 infection model in hamsters. Despite the fact that Nelfinavir, [50 mg/kg twice daily (BID) for four consecutive days], did not reduce viral RNA load and infectious virus titres in the lung of infected animals, treatment resulted in a substantial improvement of SARS-CoV-2-induced lung pathology. This was accompanied by a dense infiltration of neutrophils in the lung interstitium which was similarly observed in non-infected hamsters. Nelfinavir resulted also in a marked increase in activated neutrophils in the blood, as observed in non-infected animals. Although Nelfinavir treatment did not alter the expression of chemoattractant receptors or adhesion molecules on human neutrophils, in vitro migration of human neutrophils to the major human neutrophil attractant CXCL8 was augmented by this protease inhibitor. Nelfinavir appears to induce an immunomodulatory effect associated with increasing neutrophil number and functionality, which may be linked to the marked improvement in SARS-CoV-2 lung pathology independent of its lack of antiviral activity. Since Nelfinavir is no longer used for the treatment of HIV, we studied the effect of two other HIV protease inhibitors, namely the combination Lopinavir/Ritonavir (Kaletra™) in this model. This combination resulted in a similar protective effect as Nelfinavir against SARS-CoV2 induced lung pathology in hamsters.