SARS-CoV-2 infection and replication in human fetal and pediatric gastric organoids

Abstract

Coronavirus disease 2019 (COVID-19) pandemic caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection is a global public health emergency. COVID-19 typically manifests as a respiratory illness but an increasing number of clinical reports describe gastrointestinal (GI) symptoms. This is particularly true in children in whom GI symptoms are frequent and viral shedding outlasts viral clearance from the respiratory system. By contrast, fetuses seem to be rarely affected by COVID-19, although the virus has been detected in placentas of affected women. These observations raise the question of whether the virus can infect and replicate within the stomach once ingested. Moreover, it is not yet clear whether active replication of SARS-CoV-2 is possible in the stomach of children or in fetuses at different developmental stages. Here we show the novel derivation of fetal gastric organoids from 8-21 post-conception week (PCW) fetuses, and from pediatric biopsies, to be used as an in vitro model for SARS-CoV-2 gastric infection. Gastric organoids recapitulate human stomach with linear increase of gastric mucin 5AC along developmental stages, and expression of gastric markers pepsinogen, somatostatin, gastrin and chromogranin A. In order to investigate SARS-CoV-2 infection with minimal perturbation and under steady-state conditions, we induced a reversed polarity in the gastric organoids (RP-GOs) in suspension. In this condition of exposed apical polarity, the virus can easily access viral receptor angiotensin-converting enzyme 2 (ACE2). The pediatric RP-GOs are fully susceptible to infection with SARS-CoV-2, where viral nucleoprotein is expressed in cells undergoing programmed cell death, while the efficiency of infection is significantly lower in fetal organoids. The RP-GOs derived from pediatric patients show sustained robust viral replication of SARS-CoV-2, compared with organoids derived from fetal stomachs. Transcriptomic analysis shows a moderate innate antiviral response and the lack of differentially expressed genes belonging to the interferon family. Collectively, we established the first expandable human gastric organoid culture across fetal developmental stages, and we support the hypothesis that fetal tissue seems to be less susceptible to SARS-CoV-2 infection, especially in early stages of development. However, the virus can efficiently infect gastric epithelium in pediatric patients, suggesting that the stomach might have an active role in fecal-oral transmission of SARS-CoV-2.

Reverse-transcribed SARS-CoV-2 RNA can integrate into the genome of cultured human cells and can be expressed in patient-derived tissues

“…To investigate the possibility that SARS-CoV-2 sequences integrated into the genome can be expressed, we analyzed published RNA-seq data from SARS-CoV-2–infected cells for evidence of chimeric transcripts ( 49 ). Examination of these datasets ( 50 – 55 ) ( SI Appendix , Fig. S5 ) revealed a number of human–viral chimeric reads ( SI Appendix , Fig.…”

“…To identify human–SARS-CoV-2 chimeric reads, published RNA-seq data were downloaded from Gene Expression Omnibus (GEO) with the accession numbers GSE147507 ( 50 ), GSE153277 ( 51 ), GSE156754 ( 52 ), GSE157852 ( 53 ), GSE153684 ( 54 ), and GSE154998 ( 55 ) (summarized in SI Appendix , Fig. S5 C ).…”

Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2): a Systemic Infection

“…Importantly, the intestines are not the only affected part of the digestive system; viral nucleocapsid protein was visualized in gastric tissue derived from COVID-19 patients ( 117 ). Unsurprisingly, human gastric organoids (HGOs) derived from pediatric patients supported SARS-CoV-2 replication ( 137 ). Of note, human organoids are not the only organoids permissive to novel coronavirus; bat intestinal organoids also support SARS-CoV-2 infection, which is in agreement with the virus origin predictions ( 126 , 138 ).…”

SARS-CoV-2 RNA reverse-transcribed and integrated into the human genome

“…To investigate the possibility of viral integration into virus infected cells we analyzed published RNA-Seq data from SARS-CoV-2 -infected cells for evidence of chimeric transcripts, which would be indicative of viral integration into the genome and expression. Examination of these data sets [24][25][26][27][28][29][30] (Fig. S1a-b) revealed a substantial number of host-viral chimeric reads (Fig. 1a-c, S1c).…”