Researchers at the Institute of Biomedical Sciences at the University of São Paulo (ICB-USP) managed to isolate the strain of the omicron variant of Sars-CoV-2.
Samples of the variant are being grown in cells and, within two weeks, will begin to be distributed to laboratories with biosafety level three (NB-3) and researchers from all regions of Brazil in order to help detect the spread of the omicron throughout parents. In addition, the isolation of the new strain will allow evaluating the effectiveness of vaccines against the new strain currently applied to the Brazilian population.
“It is the first time that the omicron strain has been isolated in Brazil”, says Edison Luiz Durigon, professor at ICB-USP and coordinator of the project, supported by FAPESP.
The strain of the omicron variant was detected in a couple of Brazilians who live in South Africa and who came to Brazil on a trip. The couple underwent tests at the Albert Einstein Hospital, in São Paulo, which detected the coronavirus infection and sent samples to the ICB-USP last Wednesday (08/12).
“This sample was quickly sequenced by the hospital, which confirmed that it was the omicron strain. We took this sample and put it in cell culture”, explains Durigon.
The group of researchers from ICB-USP was also the first to manage to isolate and cultivate in the laboratory the original strain of Sars-CoV-2 that arrived in Brazil at the end of February 2020.
At the time, aliquots of the virus cultivated in the laboratory were distributed to research groups and public and private clinical laboratories throughout the country and allowed for the development of the first diagnostic tests for Covid-19 in Brazil and the carrying out of studies on the disease.
“Now, we are preparing aliquots of the omicron strain to be able to distribute to laboratories and groups of researchers that want to standardize new tests to quickly identify this variant in other cities and states”, says Durigon.
The forecast is that within two weeks there will be a sufficient stock of cultivated virus to start distribution across the country. “For the laboratories that are most urgently in need, we were able to send some aliquots more quickly”, says Durigon.
Neutralization by cytopathic effect
The isolation of the original strain of Sars-CoV-2 and the omicron variant was carried out using a cytopathic neutralization reaction technique, known as VNT, which the ICB researchers improved during the Zika virus epidemic in Brazil.
“With the arrival of Sars-CoV-2 in Brazil, we were able to isolate the virus and use it as a standard neutralization control”, said Durigon in a lecture at the Symposium on Coronavac, held by the Butantan Institute and the pharmaceutical industry Sinovac Biotech, producer of the immunizing agent, between the 7th and 9th of December.
To isolate the virus, clinical samples from patients are incubated in Vero cell culture and taken to an incubator, where they remain for between 48 and 72 hours. After 48 hours, the researchers observed that Sars-CoV-2 causes very characteristic changes (cytopathic effects) in the host cells.
“The cells are quite rounded. Some variants of Sars-CoV-2 even cause syncytia [formação de células multinucleadas por fusão de células uninucleadas ou por muitas divisões celulares incompletas de células]”, says Durigon. “After 72 hours, we were able to isolate 100% of the samples with relative ease”, he says.
The neutralization technique was also used to assess the production of neutralizing antibodies in the serum or plasma of Covid-19 convalescents.
In this assay, the patient’s blood serum or plasma is serially diluted and incubated with known amounts of virus. The mixture of serum and virus is then transferred to Vero cell cultures and the cytopathic effect of the virus is assessed after 72 hours. The result is reported as the highest serum dilution capable of neutralizing the cytopathic effect of the virus.
“Plasma from convalescent patients, with high titers of neutralizing antibodies, is still used today in the treatment of patients with Covid-19 in serious condition and for early treatment, in the first seven days of infection”, explains Durigon.
The researchers also used the VNT technique to assess the amount of neutralizing antibodies induced by Coronavac.
The results of two studies, with the participation of 580 and 866 individuals vaccinated with the immunizing agent, respectively, showed that, after the second dose, Coronavac induces the production of neutralizing antibodies at high titers, reaching up to 640 for the Wuhan, P1 strains. (gamma), P2 (zeta) and delta. Titers peak between 30 and 60 days after the second dose. Then there is a decay, but they are still detectable up to five months after vaccination, and the immunological memory lasts.
“Unlike vaccines that are being used to induce the production of antibodies against the spike protein – used by Sars-CoV-2 to connect with the human cell receptor and enable the infection – Coronavac also induces antibodies against the nucleoprotein of the This greatly increases overall immunity,” says Durigon.
The follow-up of some patients vaccinated with Coronavac and who later contracted the disease also indicated that these individuals, even six months after receiving the second dose of the vaccine, present a very rapid response to the infection, with an abrupt increase in neutralizing antibody titers.
“This rapid response to infection allowed these patients to present mild cases of the disease. None of them had to be hospitalized,” said Durigon.
The idea now is to assess whether the omicron variant is able to escape or not the antibodies of patients who received the different vaccines applied in the country.