key: cord-0748800-72z4wbzl
authors: de Siqueira, Isadora Cristina; Camelier, Aquiles Assunção; Maciel, Elves A.P.; Nonaka, Carolina Kymie Vasques; Neves, Margarida Celia L.C.; Macêdo, Yasmin Santos Freitas; de Sousa, Karoline Almeida Félix; Araujo, Victor Costa; Paste, Aurea Angelica; Souza, Bruno Solano de Freitas; Gräf, Tiago
title: Early detection of P.1 variant of SARS-CoV-2 in a cluster of cases in Salvador, Brazil
date: 2021-05-12
journal: Int J Infect Dis
DOI: 10.1016/j.ijid.2021.05.010
sha: addc955978c890d5470499fcf9d43a35922da1e9
doc_id: 748800
cord_uid: 72z4wbzl

We report three cases of severe COVID-19 due to the SARS-CoV-2 P.1 lineage in a familial cluster detected in Salvador, Bahia-Brazil. All cases were linked to travel by family members from the state of Amazonas to Bahia in late December 2020. This report indicates the cryptic transmission of the SARS-CoV-2 P.1 lineage across Brazil and highlights the importance of genomic surveillance to track the emergence of new SARS-CoV-2 variants of concern.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), which causes coronavirus disease 2019 , reportedly emerged in December 2019 in Wuhan, China (Zhu et al., 2020) . SARS-CoV-2 rapidly spread around the world, and on March 11, 2020, the WHO declared the outbreak as a pandemic (WHO/Europe, 2020).

More than one year since its emergence, with more than 153,954,491 reported cases and Africa (Tegally et al., 2021) .

In early January 2021, another SARS-CoV-2 VOC, termed P.1 (or 20J/501Y.V3), was first identified (Fujino et al., 2021) . This variant is believed to have emerged in early December in the city of Manaus, located in northern Brazil, and has been associated with increased transmissibility and high viral load (Faria et al., 2021; Naveca et al., 2021) .

Herein, we report and characterize three cases of severe COVID-19 linked to the SARS-CoV-2 P.1 lineage detected in Salvador, Bahia located in northeastern Brazil, part of a familial cluster of infections resulting from travel between the states of Amazonas and Bahia in late December 2020.

Three members of the same family, residents of the city of Manaus (Amazonas, Brazil), traveled to Salvador (Bahia, Brazil) on December 19, 2020, to visit two relatives for the holidays. Table 1 lists the clinical characteristics and laboratory results from all five cases of infection.

J o u r n a l P r e -p r o o f Case 1. On December 23, 2020, a 40 year-old female with no comorbidities began to experience myalgia and nasal obstruction lasting for three days, and then promptly recovered.

Case 2. A previously healthy 19 year-old male reported fever and myalgia on December 27, and then completely recovered after three days of mild symptoms.

Case 3. On December 25, a 69 year-old male with systemic arterial hypertension and dyslipidemia began to experience fever, chills, headache, and myalgia. After ten days of symptoms and the onset of dyspnea, he was admitted to the emergency service of a local hospital. His pulse oxygen saturation (SpO2) level was 92% when breathing room air and High-Resolution Computed Tomography (HRCT) of the thorax revealed 50-75 % of bilateral ground-glass pulmonary opacities. The day after admission, the patient was transferred to the hospital's Intensive Care Unit (ICU) for oxygen support by high-flow nasal cannula (HFNC). He remained in the ICU for 12 days and was discharged after 19 days. Methylprednisolone was administered during his hospital stay.

A 41 year-old male reported myalgia and fever on December 27. After nine days of persistent fever and the onset of dyspnea, he was admitted to the same hospital for oxygen support. His SpO2 was 90% when breathing room air, with an arterial oxygen pressure (pO2) of 60.5 mmHg. Thorax HRCT revealed 50% of bilateral ground-glass pulmonary opacities. After three days, he was transferred to the ICU due to worsening respiratory symptoms. During 11 days in the ICU, the patient required oxygen delivery by HFNC. Methylprednisolone was administered, and the patient was discharged after 21 days.

Case 5. On December 25, a 71 year-old female with Chronic Obstructive Pulmonary Disease (COPD) and dyslipidemia experienced fever, odynophagia and diarrhea. After seven days, worsening dyspnea prompted her to seek medical attention, and upon arrival J o u r n a l P r e -p r o o f at the same hospital she was admitted to the ICU due to severe respiratory distress. Her SpO2 was 80% when breathing room air, and thorax HRCT revealed 50-75% of bilateral ground-glass pulmonary opacities. The patient initially received non-invasive oxygen support with HFNC, but due to worsening respiratory distress, endotracheal intubation and invasive mechanical ventilation support for moderate acute respiratory distress syndrome (ARDS) were required. Methylprednisolone was administered upon admission.

Her course became complicated by nosocomial infection, persistent respiratory failure, and multiple organ failure, culminating in death after 65 days in the ICU.

All five cases reported herein tested positive for SARS-CoV-2 by RT-PCR.

Nasopharyngeal swab samples were tested by multiplex real-time PCR using the Allplex™ SARS-CoV-2 assay (Seegene Inc, Seoul, Korea) on an ABI7500 Fast real-time PCR instrument (Thermo Fisher Scientific). SARS-CoV-2 viral genome sequencing was run on a PGM Ion System; reads were analyzed using Torrent Suite™ Software v.5.12.1 (Thermo Fisher Scientific).

High quality SARS-CoV-2 genome sequences were generated for Cases 3, 4 and 5, which were then submitted to the online pangolin tool for lineage classification (Rambaut et al., 2020) . All three sequences were classified as the P.1 lineage and are available at the EpiCov database, maintained by the GISAID initiative, with the accession codes EPI_ISL_1443197, EPI_ISL_1443196 and EPI_ISL_1443198. A phylogenetic tree was constructed considering the available P.1 diversity. All high-quality Brazilian P.1 sequences (>29,000 bp and <1% Ns) were retrieved from GISAID on March 25, 2021.

Alignment was performed using the MAFFT sequence alignment program, and the phylogenetic tree was inferred using the IQ-TREE software package with Maximum Likelihood (ML) analysis, as previously described (Naveca et al, 2021) . This approach resulted in a tree trunk with very low branch support, reflecting the rapid spread of P.1 and low genetic genome diversity in this early phase of dissemination (Figure 1 ). The majority of the obtained sequences branching out from the tree trunk had been isolated in the Amazonas region or were cases of individuals with a history of travel to the region.

Cases 3, 4, and 5 clustered within the obtained sequences, indicating that these also originated from the early diversity of P.1, prior to the further diversification that can be observed in more recently sampled genomes.

The emergence of SARS-CoV-2 VOCs in different parts of the world presents new challenges to the control of the COVID-19 pandemic. Increased transmissibility (Tegally et al., 2021) and mortality (Davies et al., 2021) have been associated with VOCs, and cases of reinfection have also been reported (Naveca et al., 2021b) . Finally, the capacity to decrease the efficacy of currently available vaccines (Madhi et al., 2021) represents an additional threat.

With COVID-19 incidence of 7,104 cases/100,000 individuals, corresponding to a mortality rate of 197 cases /100,000 individuals, Brazil is currently facing an extremely critical phase in the pandemic, with alarming increases in cases and deaths notified across all regions of the country (https://covid.saude.gov.br/ , accessed on May 5, 2021). One of the underlying causes behind the surge in case numbers is the emergence of SARS-CoV-2 lineage P.1, which has already been detected in all regions of Brazil (http://www.genomahcov.fiocruz.br , accessed on April 21, 2021).

The three cases of P.1 lineage identified herein exemplify the cryptic transmission of this VOC throughout Brazil prior to its identification. It is possible that P.1 could have been introduced in other Brazilian states due to holiday-associated travel at the end of 2020 and early 2021. The cases reported here predates 11 other cases identified in the state of Bahia in mid-January, all of which were associated with travel from/to Manaus (Tosta et al, 2021) . For instance, in the Southern Brazilian region, community transmission of P.1 was already detected in January 2021 (Martins, et al, 2021) , and the first local P.1 case was identified in a popular Brazilian tourist destination (Steiner et al., 2021) .

In conclusion, the present report details a familial cluster of COVID-19 linked to the SARS-CoV-2 P.1 lineage. Notably, three of the five reported cases evolved as severe J o u r n a l P r e -p r o o f COVID-19, entailing long ICU stays with one associated death. One patient, a 41-yearold male, had no prior risk factors that would predispose him to severe disease. It has been assumed that the P.1 lineage could imply an increased risk of severe infection or higher mortality, based on the alarming surge in COVID-19 deaths recently reported in Brazil. While this association remains speculative, future studies are urgently needed to assess this assumption more comprehensively.

Moreover, the present report further supports the cryptic transmission of the SARS-CoV-2 P.1 lineage across Brazil and highlights the importance of genomic surveillance to track the emergence of new viral variants, which associated with a coordinated implementation of non-pharmaceutical interventions, including social distance, airport screening and quarantine for travelers, are pivotal tools to slow-down viral transmission and diminish the burden on the national public health. 

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The authors are grateful to the physicians and nurses involved in the patient's clinical care, to the GISAID team, and to all researchers who submitted sequences to the EpiCoV database. The GISAID table containing detailed data on the sequences used in this study, as well as the associated researchers, is include here as Appendix Table 1 . The authors also thank Andris K. Walter for English language revision and manuscript copy editing assistance.