After completing the first genome sequencing of the monkeypox virus (MPXV) in Brazil, scientists at the University of São Paulo (USP) are culturing the virus in cell lines with the aim of distributing samples to public and private laboratories throughout the country for use in diagnostic testing, research on the evolution of the virus, and development of novel treatments and vaccines.

The work is ongoing at the Virology Laboratory (LIM52) of the Institute of Tropical Medicine (IMT-USP), and led by virologist Lucy dos Santos Vilas Boas.

“We received a clinical sample from the first patient diagnosed in Brazil and inoculated it into a culture of Vero cells. After 24 hours, we were able to observe morphological changes in the cells that are typical of monkeypox. This was confirmed by RT-PCR,” Vilas Boas told Agência FAPESP. Vero cells are derived from kidney epithelial cells extracted from an African green monkey, and widely used as a model for studying viruses.

The specific RT-PCR test for MPXV was developed at the Albert Einstein Jewish Hospital (HIAE) in São Paulo by a team led by Dr. João Renato Rebello Pinho, who is also a researcher at IMT-USP. The method is the same as for diagnosing COVID-19 and many other viral diseases, but each pathogen requires specific reagents so that the genetic material present in the clinical sample can be amplified and detected in the laboratory.

“Until that day [June 10], no lab anywhere in Brazil had a RT-PCR kit set up for this virus,” Pinho said. “We’d already studied the sequence [of nucleotides complementing the viral DNA, required to amplify the genetic material during the test] and asked a Brazilian firm to synthesize it. All that was missing was a positive control, which we obtained from the first confirmed local case. The positive control guarantees the test is working. Just like in the case of COVID-19, we’re seeing large-scale collaboration between public and private institutions, which is really important.”

After confirmation that the virus that multiplied in the Vero cells was a monkeypox virus, the next step was to extract it from the culture medium and inactivate it so that it could be shipped safely to other centers, Vilas Boas explained. “Infected cells die and release viral particles into the supernatant – the nutrient-rich liquid in which the cells are cultured. We collect this liquid and add a reagent that inactivates the virus,” she said. “We’re now sending samples of this material to public and private labs that have asked for them. At these other centers, the viral DNA can be extracted and used as a positive control in RT-PCR tests, expanding the nation’s testing capacity.”