Infectious SARS-CoV-2 Particles from Rectal Swab Samples from COVID-19 Patients in Brazil.

The main objective of this study was to investigate the dynamic of SARS-CoV-2 viral excretion in rectal swab (RS), saliva, and nasopharyngeal swab (NS) samples from symptomatic patients and asymptomatic contacts. In addition, in order to evaluate the replication potential of SARS-CoV-2 in the gastrointestinal (GI) tract and the excretion of infectious SARS-CoV-2 from feces, we investigated the presence of subgenomic nucleoprotein gene (N) mRNA (sgN) in RS samples and cytopathic effects in Vero cell culture. A prospective cohort study was performed to collect samples from symptomatic patients and contacts in Rio de Janeiro, Brazil, from May to October 2020. One hundred and seventy-six patients had samples collected at home visits and/or during the follow up, resulting in a total of 1633 RS, saliva, or NS samples. SARS-CoV-2 RNA was detected in 130 (73.9%) patients who had at least one sample that tested positive for SARS-CoV-2. The presence of replicating SARS-CoV-2 in RS samples, measured by the detection of sgN mRNA, was successfully achieved in 19.4% (6/31) of samples, whilst infectious SARS-CoV-2, measured by the generation of cytopathic effects in cell culture, was identified in only one RS sample. Although rare, our results demonstrated the replication capacity of SARS-CoV-2 in the GI tract, and infectious viruses in one RS sample. There is still a gap in the knowledge regarding SARS-CoV-2 fecal-oral transmission. Additional studies are warranted to investigate fecal or wastewater exposure as a risk factor for transmission in human populations.

Recovery of Synthetic Zika Virus Based on Rio-U1 Isolate Using a Genetically Stable Two Plasmid System and cDNA Amplification.

In 2016, the world experienced the unprecedented Zika epidemic. The ZIKV emerged as a major human pathogen due to its association with the impairment of perinatal development and Guillain-Barré syndrome. The occurrence of these severe cases of Zika points to the significance of studies for understanding the molecular determinants of flavivirus pathogenesis. Reverse genetics is a powerful method for studying the replication and determinants of pathogenesis, virulence, and viral attenuation of flaviviruses, facilitating the design of vaccines and therapeutics. However, the main hurdle in the development of infectious clones is the instability of full-length cDNA in Escherichia coli. Here, we described the development of a genetically stable and efficient infectious clone based on the ZIKV Rio-U1 isolated in the 2016 epidemic in Brazil. The employed strategy consisted of cloning the viral cDNA genome into two stable plasmid subclones and obtaining a high-quality cDNA template with increment in DNA mass for in vitro transcription by PCR amplification. The strategy for developing a ZIKV infectious cDNA clone designed in this study was successful, yielding a replicative and efficient clone-derived virus with high similarities with its parental virus, Rio-U1, by comparison of the proliferation capacity in mammal and insect cells. The infection of AG129 immunocompromised mice caused identical mortality rates, with similar disease progression and morbidity in the animals infected with the parental and the cDNA-derived virus. Histopathological analyses of mouse brains infected with the parental and the cDNA-derived viruses revealed a similar pathogenesis degree. We observed meningoencephalitis, cellular pyknosis, and neutrophilic invasion adjacent to the choroid plexus and perivascular cuffs with the presence of neutrophils. The developed infectious clone will be a tool for genetic and functional studies in vitro and in vivo to understand viral infection and pathogenesis better.