conjunctivitis. She had no fever until day 4, when fever (38°C), nausea, and vomiting began. Infection with SARS-CoV-2 was confirmed by performing real-time reverse transcription polymerase chain reaction (RT-PCR) assay on sputum samples (cycle threshold value [Ct], 16.1) on the admission day, followed by viral M gene sequencing (GenBank accession number MT008022), and virus isolation on Vero E6 cell line (2019-nCoV/Italy-INMI1). The full genome sequence was obtained from either clinical sample and or culture isolate (GISAID accession numbers EPI_ISL_410545 and EPI_ISL_410546). At admission, no other respiratory infections were detected (QIAstat-Dx® Respiratory Panel; Qiagen).
On day 3 after hospital admission, owing to the persistence of conjunctivitis, an ocular swab was collected and viral RNA was detected (Ct, 21.6). Subsequent ocular samples collected with almost daily frequency resulted positive up to day 21, with declining virus concentration (increased Ct values). Conjunctivitis greatly improved at day 15 and apparently resolved at day 20.
Five days after it became undetectable, SARS-CoV-2 RNA was detected again (Ct, 36.25) in the ocular swab sample collected at day 27 (Table) . SARS-CoV-2 RNA was detected in ocular swabs days after it was undetectable in nasal swabs (Table) . In addition, the Ct values detected in the late ocular samples were lower than those observed in the nasal swabs (Table 1) , suggesting sustained replication in conjunctiva. With the aim of demonstrating that viral genomes detected in ocular swabs represented infectious virus, the first RNApositive ocular sample was inoculated in Vero E6 cells, and cytopathic effect was observed 5 days postinoculum. Viral replication was confirmed by real-time RT-PCR on RNA purified from spent cell growth medium.
Discussion: We found that ocular fluids from SARS-CoV-2-infected patients may contain infectious virus, and hence may be a potential source of infection. These findings highlight the importance of control measures, such as avoiding touching the nose, mouth, and eyes and frequent hand washing. A related implication is the importance of appropriate use of personal protective equipment for ophthalmologists during clinical examination, because ocular mucosa may be not only a site of virus entry but also a source of contagion. Furthermore, we observed that ocular involvement of SARS-CoV-2 may occur early in the COVID-19 course, suggesting This article was published at Annals.org on 17 April 2020. 