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1. A faster and less costly alternative for RNA extraction of SARS-CoV-2 using proteinase k treatment followed by thermal shock.

Author

Adolfo Marcelo Ñique, Fiorella Coronado-Marquina, Jairo Andrés Mendez Rico, María Paquita García Mendoza, Nancy Rojas-Serrano, Paulo Vitor Marques Simas, Cesar Cabezas Sanchez, and Jan Felix Drexler

Subjects
Medicine, Science
Abstract

One of the biggest challenges during the pandemic has been obtaining and maintaining critical material to conduct the increasing demand for molecular tests. Sometimes, the lack of suppliers and the global shortage of these reagents, a consequence of the high demand, make it difficult to detect and diagnose patients with suspected SARS-CoV-2 infection, negatively impacting the control of virus spread. Many alternatives have enabled the continuous processing of samples and have presented a decrease in time and cost. These measures thus allow broad testing of the population and should be ideal for controlling the disease. In this sense, we compared the SARS-CoV-2 molecular detection effectiveness by Real time RT-PCR using two different protocols for RNA extraction. The experiments were conducted in the National Institute of Health (INS) from Peru. We compared Ct values average (experimental triplicate) results from two different targets, a viral and internal control. All samples were extracted in parallel using a commercial kit and our alternative protocol-samples submitted to proteinase K treatment (3 μg/μL, 56°C for 10 minutes) followed by thermal shock (98°C for 5 minutes followed by 4°C for 2 minutes); the agreement between results was 100% in the samples tested. In addition, we compared the COVID-19 positivity between six epidemiological weeks: the initial two in that the Real time RT-PCR reactions were conducted using RNA extracted by commercial kit, followed by two other using RNA obtained by our kit-free method, and the last two using kit once again; they did not differ significantly. We concluded that our in-house method is an easy, fast, and cost-effective alternative method for extracting RNA and conducing molecular diagnosis of COVID-19.

Published
2021
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2. Climate and land-use shape the spread of zoonotic yellow fever virus

Author

Sarah C. Hill, Simon Dellicour, Ingra M. Claro, Patricia C. Sequeira, Talita Adelino, Julien Thézé, Chieh-Hsi Wu, Filipe Romero Rebello Moreira, Marta Giovanetti, Sabrina L. Li, Jaqueline G. de Jesus, Felipe J. Colón-González, Heather R. Chamberlain, Oliver Pannell, Natalia Tejedor-Garavito, Fernanda de Bruycker-Nogueira, Allison A. Fabri, Maria Angélica Mares-Guia, Joilson Xavier, Alexander E. Zarebski, Arran Hamlet, Maria Anice Mureb Sallum, Antonio C. da Costa, Erika R. Manuli, Anna S. Levin, Luís Filipe Mucci, Rosa Maria Tubaki, Regiane Maria Tironi de Menezes, Juliana Telles de Deus, Roberta Spinola, Leila Saad, Esper G. Kallas, G.R. William Wint, Pedro S. Peixoto, Andreza Aruska de Souza Santos, Jane P. Messina, Oliver J. Brady, Andrew J. Tatem, Marc A. Suchard, Jairo A. Mendez-Rico, André Abreu, Renato Santana Aguiar, Oliver G. Pybus, Guy Baele, Philippe Lemey, Felipe Iani, Mariana S. Cunha, Ana M. Bispo de Filippis, Ester C. Sabino, Nuno R. Faria, Royal Veterinary College [London], University of London [London], Department of Zoology, University of Oxford, Spatial Epidemiology Lab (SpELL), Université libre de Bruxelles (ULB), Rega Institute for Medical Research [Leuven, België], Catholic University of Leuven - Katholieke Universiteit Leuven (KU Leuven), Faculdade de Medicina da Universidade de São Paulo [São Paulo, Brésil], The Abdul Latif Jameel Institute for Disease and Emergency Analytics, Fundação Oswaldo Cruz / Oswaldo Cruz Foundation (FIOCRUZ), Réseau International des Instituts Pasteur (RIIP), Fundação Ezequiel Dias (FUNED), Unité Mixte de Recherche d'Épidémiologie des maladies Animales et zoonotiques (UMR EPIA), VetAgro Sup - Institut national d'enseignement supérieur et de recherche en alimentation, santé animale, sciences agronomiques et de l'environnement (VAS)-Institut National de Recherche pour l’Agriculture, l’Alimentation et l’Environnement (INRAE), School of Mathematics [Southampton], University of Southampton, Institute for Life Sciences, Laboratório de Biologia Molecular de Flavivírus [Rio de Janeiro], Instituto Oswaldo Cruz / Oswaldo Cruz Institute [Rio de Janeiro] (IOC), Réseau International des Instituts Pasteur (RIIP)-Réseau International des Instituts Pasteur (RIIP)-Fundação Oswaldo Cruz / Oswaldo Cruz Foundation (FIOCRUZ), Réseau International des Instituts Pasteur (RIIP)-Réseau International des Instituts Pasteur (RIIP), Universidade Federal de Minas Gerais = Federal University of Minas Gerais [Belo Horizonte, Brazil] (UFMG), School of Geography and the Environment [Oxford] (SoGE), School of Geography [Nottingham], University of Nottingham, UK (UON), Department of Infectious Disease Epidemiology, Faculty of Epidemiology and Population, Instituto de Ciências Biológicas [Goiânia, Brésil] (ICB), Imperial College London, Unité de Catalyse et Chimie du Solide - UMR 8181 (UCCS), Université d'Artois (UA)-Centrale Lille-Institut de Chimie du CNRS (INC)-Université de Lille-Centre National de la Recherche Scientifique (CNRS), and Department of Zoology [Oxford]

Subjects
[SDV.SPEE]Life Sciences [q-bio]/Santé publique et épidémiologie
Abstract

Zoonotic viruses that originate in wildlife harm global human health and economic prosperity1. Understanding virus transmission at the human-animal-environment interface is a key component of pandemic risk-reduction2,3. Zoonotic disease emergence is highest in biodiverse, tropical forests undergoing intensive land-use change4,5. Phylodynamic analyses of virus genomes can powerfully test epidemiological hypotheses, but are rarely applied to viruses of animals inhabiting these habitats. Brazil’s densely-populated Atlantic Forest and Cerrado region experienced in 2016–2021 an explosive human outbreak of sylvatic yellow fever, caused by repeated virus spillover from wild neotropical primates6. Here we use yellow fever virus (YFV) genome sequences and epidemiological data from neotropical primates, humans, and mosquito vectors to identify the environmental, demographic, and climatic factors determining zoonotic virus spread. Using portable sequencing approaches we generated 498 YFV genomes, resulting in a well-sampled dataset of zoonotic virus genomes sampled from wild mammals. YFV dispersal velocity was slower at higher elevation, in colder regions, and further away from main roads. Virus lineage dispersal was more frequent through wetter areas, areas with high neotropical primate density and through landscapes covered by mosaic vegetation. Higher temperatures were associated with higher virus effective population sizes, and peaks of transmission in warmer, wetter seasons were associated with higher virus evolutionary rates. Our study demonstrates how zoonotic disease transmission is linked to land-use and climate, underscoring the need for One-Health approaches to reducing the rate of zoonotic spillover.

Published
2023

3. SARS-CoV-2 Variants of Interest and Concern naming scheme conducive for global discourse

Author

Leo L.M. Poon, Lorenzo Subissi, Bas B. Oude Munnink, Malik Peiris, Brett N Archer, Lisa L. Carter, Edward C. Holmes, Oliver G. Pybus, Amal Barakat, John Ziebuhr, Jairo Andres Mendez Rico, Suxiang Tong, Maria D. Van Kerkhove, Ihab El Masry, Jinal N. Bhiman, Paola Cristina Resende, Mark J. Pallen, Frank Konings, Richard A. Neher, Bette T. Korber, Alexander E. Gorbalenya, Anne Cullinane, Mark Perkins, Esther L Hamblion, Emma B. Hodcroft, Belinda Louise Herring, Tulio de Oliveira, Andrew Rambaut, Marion Koopmans, Vincent J. Munster, Erik Alm, George F. Gao, Marco Marklewitz, Shagun Khare, Jens H. Kuhn, Trevor Bedford, Sebastian Maurer-Stroh, Anne von Gottberg, Julian Druce, Sylvie van der Werf, Manish Kakkar, Leon Caly, Roger Evans, Juliana Leite, Duncan MacCannell, Boris I. Pavlin, Volker Thiel, University Hospital of Cologne [Cologne], European Centre for Disease Prevention and Control [Stockholm, Sweden] (ECDC), Institut National d'Hygiène [Maroc], University of Washington [Seattle], Fred Hutchinson Cancer Research Center [Seattle] (FHCRC), Department of Biochemistry and Molecular Biology, Mayo Clinic, Victorian Infectious Diseases Reference Laboratory, Food and Agriculture Organization of the United Nations [Rome, Italie] (FAO), University of Chinese Academy of Sciences [Beijing] (UCAS), WHO, Regional Office for Africa [Brazzaville, Republic of the Congo], Génétique Moléculaire des Virus à ARN - Molecular Genetics of RNA Viruses (GMV-ARN (UMR_3569 / U-Pasteur_2)), Institut Pasteur [Paris] (IP)-Centre National de la Recherche Scientifique (CNRS)-Université Paris Cité (UPCité), Centre National de Référence des virus des infections respiratoires (dont la grippe) - National Reference Center Virus Influenzae [Paris] (CNR - laboratoire coordonnateur), Institut Pasteur [Paris] (IP), and Virology

Subjects
Microbiology (medical), Scheme (programming language), 2019-20 coronavirus outbreak, Coronavirus disease 2019 (COVID-19), [SDV]Life Sciences [q-bio], viruses, Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), Immunology, Global Health, World Health Organization, Applied Microbiology and Biotechnology, Microbiology, World health, 03 medical and health sciences, Terminology as Topic, Genetics, Global health, Humans, 030304 developmental biology, computer.programming_language, 0303 health sciences, SARS-CoV-2, 030306 microbiology, business.industry, Communication, COVID-19, Cell Biology, Public relations, Coronavirus, business, computer
Abstract

A group convened and led by the Virus Evolution Working Group of the World Health Organization reports on its deliberations and announces a naming scheme that will enable clear communication about SARS-CoV-2 variants of interest and concern.

Published
2021

4. Epidemiologic History and Genetic Diversity Origins of Chikungunya and Dengue Viruses, Paraguay

Author

Julien Thézé, Shirley Villalba, Juan Torales, Sandra Irala, María Liz Gamarra, Marta Giovanetti, Jairo Andres Mendez Rico, Andrea Gómez, Cynthia Vazquez, Tulio de Oliveira, Ingra Morales Claro, Vasco Azevedo, Fernanda de Bruycker-Nogueira, Leticia Franco, Ana Maria Bispo de Filippis, Carlos F. Campelo de Albuquerque, Joilson Xavier, Vagner Fonseca, Marcos Cesar Lima de Mendonça, Luiz Carlos Junior Alcantara, Edward C. Holmes, Jaqueline Goes de Jesus, Tiago Gräf, Instituto Gonçalo Moniz, Partenaires INRAE, Department of Infectious Diseases, Istituto Superiore di Sanita [Rome], Universidade Federal de Minas Gerais, and University of Oxford [Oxford]

Subjects
Serotype, Epidemiology, viruses, vector-borne infections, Infectious and parasitic diseases, RC109-216, MESH: Chikungunya Fever, MESH: Dengue, Dengue virus, medicine.disease_cause, MESH: Dengue Virus, Dengue fever, Dengue, 0302 clinical medicine, MESH: Arboviruses, 030212 general & internal medicine, Chikungunya, MESH: Genetic Variation, MESH: Phylogeny, MESH: Paraguay, Phylogeny, Phylogenetic tree, Zika Virus Infection, virus diseases, 3. Good health, Infectious Diseases, arboviruses, Medicine, Brazil, Microbiology (medical), 030231 tropical medicine, MESH: Zika Virus, Biology, Arbovirus, Virus, 03 medical and health sciences, MESH: Zika Virus Infection, parasitic diseases, medicine, Humans, Genetic diversity, chikungunya virus, MESH: Humans, dengue virus, Research, Genetic Variation, Zika Virus, biochemical phenomena, metabolism, and nutrition, medicine.disease, Virology, Epidemiologic History and Genetic Diversity Origins of Chikungunya and Dengue Viruses, Paraguay, genomic surveillance, 13. Climate action, Paraguay, nanopore sequencing, Chikungunya Fever, [SDV.SPEE]Life Sciences [q-bio]/Santé publique et épidémiologie, MESH: Brazil
Abstract

International audience; C hikungunya virus (CHIKV), dengue virus (DENV), and Zika virus (ZIKV) are 3 of the most common arthropod-borne viruses (arboviruses) that infect humans. All are transmitted by the anthropophilic and urban-adapted Aedes aegypti and Ae. albopictus mosquito vectors (1). Driven by human movement and climate trends, the distribution of these mosquitoes is expanding along with the arboviruses they transmit (2). In Latin America, CHIKV and ZIKV have emerged since the mid-2000s, joining DENV, which is already endemic there (3). In this region, only Uruguay and Chile did not report autochthonous transmissions of one of these arboviruses during 2014-2019, highlighting the current state of endemicity (4). Paraguay is a landlocked country in the center of South America; it borders Bolivia, Brazil, and Argentina. DENV is endemic to Paraguay, and all 4 serotypes (DENV-1-4) have been detected there; in some seasons, multiple serotypes co-circulate (5,6). Phylogenetic analysis has shown that DENV genetic diversity in Paraguay is closely related to that in neighboring countries, particularly Brazil (7,8). However, more genomic surveillance studies in Paraguay are needed to learn more about this epidemiologic pattern. Cases

Published
2021

5. A faster and less costly alternative for RNA extraction of SARS-CoV-2 using proteinase k treatment followed by thermal shock

Author

Cesar Cabezas Sanchez, María Paquita García Mendoza, Jairo Andrés Mendez Rico, Nancy Rojas-Serrano, Jan Felix Drexler, Adolfo Marcelo Ñique, Paulo Vitor Marques Simas, and Fiorella Coronado-Marquina

Subjects
0301 basic medicine, RNA viruses, Viral Diseases, Coronaviruses, Artificial Gene Amplification and Extension, Polymerase Chain Reaction, Biochemistry, Geographical locations, 0302 clinical medicine, COVID-19 Testing, Medical Conditions, Sense (molecular biology), Peru, 030212 general & internal medicine, Pathology and laboratory medicine, Mathematics, Virus Testing, Alternative methods, education.field_of_study, Multidisciplinary, biology, Proteases, Medical microbiology, Enzymes, Nucleic acids, Infectious Diseases, Viruses, Medicine, RNA, Viral, RNA extraction, Endopeptidase K, SARS CoV 2, Pathogens, Research Article, Coronavirus disease 2019 (COVID-19), SARS coronavirus, Science, Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), Population, Real-Time Polymerase Chain Reaction, Microbiology, 03 medical and health sciences, Extraction techniques, Diagnostic Medicine, Genetics, Humans, education, Molecular Biology Techniques, Pandemics, Molecular Biology, Medicine and health sciences, Chromatography, Biology and life sciences, Clinical Laboratory Techniques, Diagnostic Tests, Routine, SARS-CoV-2, Organisms, Viral pathogens, RNA, COVID-19, Proteins, Covid 19, Reverse Transcriptase-Polymerase Chain Reaction, South America, Proteinase K, Microbial pathogens, Research and analysis methods, 030104 developmental biology, biology.protein, Enzymology, Gene expression, People and places, RNA transport
Abstract

One of the biggest challenges during the pandemic has been obtaining and maintaining critical material to conduct the increasing demand for molecular tests. Sometimes, the lack of suppliers and the global shortage of these reagents, a consequence of the high demand, make it difficult to detect and diagnose patients with suspected SARS-CoV-2 infection, negatively impacting the control of virus spread. Many alternatives have enabled the continuous processing of samples and have presented a decrease in time and cost. These measures thus allow broad testing of the population and should be ideal for controlling the disease. In this sense, we compared the SARS-CoV-2 molecular detection effectiveness by Real time RT-PCR using two different protocols for RNA extraction. The experiments were conducted in the National Institute of Health (INS) from Peru. We compared Ct values average (experimental triplicate) results from two different targets, a viral and internal control. All samples were extracted in parallel using a commercial kit and our alternative protocol–samples submitted to proteinase K treatment (3 μg/μL, 56°C for 10 minutes) followed by thermal shock (98°C for 5 minutes followed by 4°C for 2 minutes); the agreement between results was 100% in the samples tested. In addition, we compared the COVID-19 positivity between six epidemiological weeks: the initial two in that the Real time RT-PCR reactions were conducted using RNA extracted by commercial kit, followed by two other using RNA obtained by our kit-free method, and the last two using kit once again; they did not differ significantly. We concluded that our in-house method is an easy, fast, and cost-effective alternative method for extracting RNA and conducing molecular diagnosis of COVID-19.

Published
2020

6. Portable sequencing in the field and the classroom: a retrospective examination of the circulation of DENV1 and DENV2 in Brazil

Author

Luiz Henrique Ferraz Demarchi, Marina Castilhos Souza Umaki Zardin, José Lourenço, Aristóteles Góes Neto, Jurandy Júnior Ferraz de Magalhães, Ronaldo de Jesus, Elaine Cristina de Oliveira, Cynthia Vazquez, Dario Brock Ramalho, Jairo Andres Mendez Rico, Priscila Leite, Luiz Carlos Junior Alcantara, Felipe Campos de Melo Iani, Vagner Fonseca, Joscélio Aguiar Silva, Ana Flávia Mendonça, Zoraida del Carmen Fernandez Grillo, Fernanda Khouri Barreto, Talita Émile Ribeiro Adelino, Álvaro Salgado de Abreu, Lívia Carla Vinhal Frutuoso, Wanderson Kleber de Oliveira, Victor Pimentel, Gislene Garcia de Castro Lichs, Joilson Xavier, Luiz Augusto Pereira, Allison Fabri, Flávia Löwen Levy Chalhoub, Vasco Azevedo, Valdinete Alves do Nascimento, Fátima María Cardozo Segovia, Stephane Tosta, Rodrigo Fabiano do Carmo Said, Roselene Hans Santos, Noely Fabiana Oliveira de Moura, Agenor de Castro Moreira dos Santos, Jorge Antônio Chamon Júnior, Arabela Leal e Silva de Mello, Arnaldo Correia de Medeiros, Marluce Aparecida Assunção Oliveira, Rita Maria Ribeiro Nogueira, Rivaldo Venâncio da Cunha, Vinícius Lemes da Silva, Edward C. Holmes, Svetoslav Nanev Slavov, Felicidade Mota Pereira, Poliana da Silva Lemos, Maricélia Maia de Lima, Carlos F. Campelo de Albuquerque, Ana Maria Bispo de Filippis, Gabriel Muricy Cunha, Alexander A. Martínez, Fernanda de Bruycker-Nogueira, André Luiz de Abreu, Crhistinne Cavalheiro Maymone Gonçalves, Emerson Luiz Lima Araújo, Marta Giovanetti, Simone Kashima Haddad, and Tulio de Oliveira

Subjects
medicine.medical_specialty, biology, Transmission (medicine), Public health, Dengue virus, biology.organism_classification, medicine.disease_cause, Zika virus, Geography, Epidemiology, Case fatality rate, medicine, Nanopore sequencing, Clade, Demography
Abstract

BackgroundBrazil has experienced a large dengue virus (DENV) epidemic in 2019, highlighting a continual struggle with effective control and public health preparedness. Brazil is a world leader in real-time genomic surveillance of arboviruses, although such technology and expertise remains inaccessible for the vast majority of local researchers and public health workers. In 2019, we led field and classroom initiatives for the genomic surveillance of DENV in Brazil.MethodsOxford Nanopore MinION technology was used for sequencing, focusing on generating DENV1 and DENV2 complete genomes. Using phylogenetic and epidemiological approaches conducted in real-time during a training program and subsequently through online channels, we explored the recent spatio-temporal evolution and spread of these viruses in Brazil.FindingsIn the years following the Zika virus epidemic (2017-2018) reporting was at an all-time low, and significant increases in reported cases and deaths in 2019 did not reflect a higher case fatality ratio. Estimated transmission potential and reporting of other arboviruses suggests that neither arboviral reporting saturation nor climatic factors can easily explain the post-Zika period and resurgence in 2019 (respectively). Phylogenetic analysis revealed complex patterns of transmission, with lineage co-circulation and replacement, in which the North and the Southeast acted as sources of dispersion to other regions. We identified two lineages within the already reported DENV2 BR-4 clade, for which the effective reproduction number had seasonal signatures alike reported cases, with a temporal increase towards 2019 mirroring the large epidemic that year.InterpretationWe describe the recent evolution and diffusion of DENV1 and DENV2 in Brazil. Importantly, the surveillance outputs and training initiative here described serve as proof-of-concept of the potential of portable sequencing for both research and local capacity building in the area of genomic surveillance of arboviruses.FundingDecit, SCTIE, BrMoH, CNPq, CAPES, EU Horizon 2020 through ZIKAlliance and STARBIOS2.

Published
2020

7. Diagnostic Laboratory Testing and Clinical Preparedness for Dengue Outbreaks during the COVID-19 Pandemic

Author

Luis Gerardo Castellanos, Gabriela Paz-Bailey, Stephen H. Waterman, Jose Luis San Martin, Gamaliel Gutierrez, and Jairo A. Mendez-Rico

Subjects
Mosquito Control, Coronavirus disease 2019 (COVID-19), Pneumonia, Viral, Dengue fever, Dengue, Betacoronavirus, Virology, Pandemic, medicine, Animals, Humans, Diagnostic laboratory, Letter to the Editor, Pandemics, Tropical Climate, biology, SARS-CoV-2, business.industry, Community Participation, COVID-19, Outbreak, medicine.disease, biology.organism_classification, Pneumonia, Culicidae, Infectious Diseases, Preparedness, Parasitology, Public Health, Coronavirus Infections, business
Published
2020

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