COVID-19 typically manifests as a respiratory illness, but several clinical reports have described gastrointestinal symptoms. This is particularly true in children in whom gastrointestinal symptoms are frequent and viral shedding outlasts viral clearance from the respiratory system. These observations raise the question of whether the virus can replicate within the stomach. Here we generate gastric organoids from fetal, pediatric, and adult biopsies as in vitro models of SARS-CoV-2 infection. To facilitate infection, we induce reverse polarity in the gastric organoids. We find that the pediatric and late fetal gastric organoids are susceptible to infection with SARS-CoV-2, while viral replication is significantly lower in undifferentiated organoids of early fetal and adult origin. We demonstrate that adult gastric organoids are more susceptible to infection following differentiation. We perform transcriptomic analysis to reveal a moderate innate antiviral response and a lack of differentially expressed genes belonging to the interferon family. Collectively, we show that the virus can efficiently infect the gastric epithelium, suggesting that the stomach might have an active role in fecal-oral SARS-CoV-2 transmission.

Since 1997, G1-lineage H9N2 avian influenza viruses have been circulating in Asia and later on in the Middle East, and they have been associated to mild respiratory disease, drops in egg production and moderate mortality in chickens, in particular in the presence of concurrent infections. In this study, we investigated the importance of the G1-lineage H9N2 A/chicken/Israel/1163/2011 virus as a primary pathogen in layers, analyzing its tropism and binding affinity for the oviduct tissues, and investigating the long-term impact on egg production. Besides causing a mild respiratory infection, the virus replicated in the oviduct of 60% of the hens causing different degrees of salpingitis throughout the organ, in particular at the level of the infundibulum, where the detection of the virus was associated with severe heterophilic infiltrate, and necrosis of the epithelium. Binding affinity assays confirmed that the infundibulum was the most receptive region of the oviduct. The drop in egg production was at its peek at 2 weeks post-infection (pi) (60% decrease) and continued up to 80 days pi (35% decrease). On day 80 pi, non-laying birds showed egg yolk peritonitis, and histopathological analyses described profound alteration of the infundibulum architecture, duct ectasia and thinning of the epithelium, while the rest of the oviduct and ovary appeared normal. Our results show that this H9N2 virus is a primary pathogen in layer hens, and that its replication in the infundibulum is responsible for acute and chronic lesions that limits the effective functionality of the oviduct, compromising the commercial life of birds.

Bovine is considered the main reservoir of influenza D virus (IDV), however, low levels of seropositivity in other farmed species suggest a wide range of potential hosts. Nevertheless, it is not clear whether this scenario is the result of rare spillover events upon contact with bovines, or a lack of adaptation of IDV to these hosts. Among these species, sheep represents a crucial component of the rural economy in many developing countries, but little is known about its role in the ecology of the disease. To evaluate the susceptibility of sheep to IDV viruses of different origin, we used ovine respiratory tissues as an ex vivo model and investigated the infective phenotype of two IDV strains isolated from either bovine (IDV-BOV) or swine (IDV-SW). For translatability purposes, we included a parainfluenza type 3 virus, as positive control, given its known respiratory tropism in sheep. We performed a timed evaluation of the viral infectivity, cell tropism and the associated histopathology, by means of tissue culture infectious dose assays on supernatants and histological/immunohistochemical analyses on explanted tissues, respectively. To further investigate differences in the phenotype of these two strains and to identify the potential targets of replication in the most commonly land-based farmed mammalian species, we carried out virus binding assays on histological sections of the respiratory tract of bovine, caprine, ovine, horse and swine. Our results demonstrated that IDV successfully replicates in nasal, tracheal and lung ovine tissues, suggesting a moderate susceptibility of this species to IDV infection. Interestingly, despite the high genetic identity of these strains, IDV- BOV consistently replicated to higher titers than IDV-SW in all respiratory tracts, suggesting IDV viruses might display considerable levels of variability in their phenotype when crossing the species barrier. Virus binding assays confirmed a superior affinity of the IDV viruses for the bovine upper respiratory tract, and a preference for the pharyngeal epithelium of small ruminants, indicating possible targets to improve the sensitivity of virological sampling for diagnostic and post-mortem purposes. Further pathogenesis and cross-species transmission studies will be necessary to elucidate the ecology of IDV and eventually allow the design of cost-effective surveillance strategies.

Once low-pathogenic avian influenza viruses (LPAIVs) of the H5 and H7 subtypes from wild birds enter into poultry species, there is the possibility of them mutating into highly pathogenic avian influenza viruses (HPAIVs), resulting in severe epizootics with up to 100% mortality. This mutation from a LPAIV to HPAIV strain is the main cause of an AIV’s major economic impact on poultry production. Although AIVs are inextricably linked to their hosts in their evolutionary history, the contribution of host-related factors in the emergence of HPAI viruses has only been marginally explored so far. In this study, transcriptomic sequencing of tracheal tissue from chickens infected with four distinct LP H7 viruses, characterized by a different history of pathogenicity evolution in the field, was implemented. Despite the inoculation of a normalized infectious dose of viruses belonging to the same subtype (H7) and pathotype (LPAI), the use of animals of the same age, sex and species as well as the identification of a comparable viral load in the target samples, the analyses revealed a heterogeneity in the gene expression profile in response to infection with each of the H7 viruses administered.

Among flaviviruses, Zika virus (ZIKV) is the only arbovirus officially recognized as a teratogenic agent, as a consequence of its ability to infect and cross the placental barrier causing congenital malformation in the fetus. While many studies have focused on understanding ZIKV pathogenesis during pregnancy, the viral mechanisms affecting fetal development remain largely unclear. In this study, we investigated ZIKV virulence in placental trophoblasts, using viruses with distinct lipid profiles. Firstly, we propagated a ZIKV strain belonging to the Asian lineage in either mammalian or mosquito cells, obtaining two viral stocks, which were purified and analyzed to determine their genetic and lipid composition. Successively, we assessed the infectivity of the two stocks in placental cells using both immortalized cell lines and explants. We found that the two viral stocks displayed identical consensus sequences with homogeneous quasispecies composition. However, the lipid composition of their envelope significantly varied depending on the cell of origin, with the mammalian-derived viral stock characterized by a higher content of phosphatidylcholines compared to the virions originating from mosquito cells. Notably, ZIKV stock derived from mammalian cells showed a higher infectivity in immortalized villous trophoblasts and full-term placental explants of human origin. This increased infectivity was linked to enhanced fusion efficiency during the viral uncoating phase in trophoblast cells, as demonstrated using a lipophilic probe. Collectively, our data suggest a potential role of viral lipids as determinants of ZIKV infectivity in full-term placenta, underscoring the importance of lipidomic research in virology.

Since its emergence in 1996, highly pathogenic avian influenza (HPAI) viruses of the A/Goose/Guangdong/1/96 lineage have diversified into multiple clades, culminating in the 2020-2021 global panzootic caused by H5N1 viruses of the clade 2.3.4.4b. Further reassortment events have significantly diversified the phenotypes of these viruses, underscoring the need for continuous monitoring and strain characterization to better adjust control measures and mitigate the impact of the disease in wild birds and poultry. Standardized, ready-to-use ex vivo tissue platforms for rapid phenotyping of avian influenza viruses (AIVs) offer a valid alternative to in vivo models that are financially, ethically and logistically demanding. We optimized explant production and cryopreservation protocols for chicken and duck tracheal organ cultures (cTOCs and dTOCs) and precision-cut lung slices (cPCLS and dPCLS), assessing post-thaw viability, histological integrity, and susceptibility to AIV infection. Trehalose supplementation of cryopreservation solutions based on dimethyl sulfoxide (DMSO) and fetal bovine serum (FBS) significantly improved tissue viability. Although cryopreserved tissues were less viable than the fresh explants, viral replication was similar and only a modest reduction in susceptibility to infection was observed. Finally, we used duck and chicken TOCs to assess the ability of cryopreserved explants to discriminate viruses based on their divergent fitness and host preference. These findings underscore the potential of cryopreserved TOCs and PCLS as additional tools for the phenotypic characterisation of emerging AIVs.