In Araraquara, state of São Paulo, researchers detected the P.1 variant in 93% of samples from patients diagnosed at a primary healthcare facility in the first two months of the year (Brazilian Army sanitizing public places; photo: City of Araraquara)

Published on 03/15/2021

By Karina Toledo  |  Agência FAPESP – Preliminary results of a study show that the P.1 variant of the novel coronavirus endemic to the Brazilian Amazon is already predominant in Araraquara, a medium-sized city in the interior of the state of São Paulo in Brazil’s Southeast region. 

Araraquara is more than 2,500 km from Manaus, the capital of Amazonas State. Every single one of its COVID-19 intensive care beds were occupied on February 15, and since then new cases, hospital admissions, and deaths from the disease have continued to rise to record levels.

Researchers at the University of São Paulo’s Institute of Tropical Medicine (IMT-USP) analyzed 57 nasopharyngeal swabs from patients diagnosed with the disease between January 25 and February 23, 2021, at a primary healthcare facility in Araraquara. The test detected the presence of the P.1 variant, considered twice as transmissible as its predecessor (lineage B.1.128), in 93% of the samples.

“Until February 17, we were still detecting cases of infection by other lineages. Since then, they have all been P.1,” Camila Romano, principal investigator for the study, told Agência FAPESP.

The study was part of a project, supported by FAPESP, to assess transmission of SARS-CoV-2 within households in Araraquara. As soon as P.1 was detected in the city and local health services showed signs of being overwhelmed, the IMT-USP group contacted collaborators to obtain samples from residents and try to measure the prevalence of the novel variant.

The analysis began with material from 22 hospitalized patients. “They had been in hospital for many days, and viral load was expected to be so low that it would be hard to measure, but to our surprise, we were able to sequence the viral RNA in 14 samples, and 12 of them tested positive for P.1,” Romano said.

The next step was to analyze samples from the community, especially patients processed by primary healthcare facilities who did not necessarily have co-morbidities or progress to severe forms of the disease, so as to obtain a more representative portrait of the city’s population.

To make the analysis faster and cheaper, the IMT-USP group developed a methodology that dispenses viral genome sequencing. “We designed a novel RT-PCR trial similar to the test used for diagnosis but capable of distinguishing positive cases of COVID-19 that correlate with the presence of P.1,” Romano said.

The strategy was possible, she explained, because a key mutation in the gene NSP6 of the Brazilian variant resembles a mutation found in the more transmissible variants first detected in the United Kingdom (B.1.1.7) and South Africa (B.1.351). This gene encodes a protein that takes part in the viral replication process, and the mutation is characterized by deletion of three amino acids (equivalent to the loss of nine nucleotides).

“We added a primer to the RT-PCR protocol that ‘lights up’ only when this mutation in NSP6 is present,” she said. Primers in PCR are short sequences of nucleic acids that complement the genetic material being studied. They are used to hybridize with the sample DNA and define the region to be amplified. 

Adaptive advantage

Research on the evolution of SARS-CoV-2 suggests the mutation in NSP6 (which encodes non-structural protein 6) emerged concurrently in P.1, B.1.1.7, and B.1.351, and was not present in their common ancestor.

“This and other mutations in the spike protein [whereby the virus binds to human cells in order to enter and infect them] appear to give the virus an important adaptive advantage, enabling it to spread faster and wider,” Romano said. “In cities where P.1 has been detected, we’re seeing a significant change in the behavior of the epidemic.”

A study recently published by researchers affiliated with the Brazil-UK Center for Arbovirus Discovery, Diagnosis, Genomics and Epidemiology (CADDE) and also involved in the study led by Romano shows that the Brazilian variant emerged in November 2020 and in only seven weeks became the most prevalent strain of the virus in Manaus (read more at: agencia.fapesp.br/35414). 

“We don’t yet know exactly when P.1 arrived in Araraquara. We’re analyzing older samples to try to find out. The data we have so far suggests P.1 also became the most prevalent variant in the city in under two months,” Romano said. “It’s unusual for an emerging lineage to replace another completely in such a short time, especially while an outbreak is underway and a lot of people are infected. It’s alarming and evidences the power of this virus.”

 

Source: https://agencia.fapesp.br/35416