Um newly published article by researchers from the Oswaldo Cruz Institute (IOC/Fiocruz) in the international scientific journal The Lancet Infectious Diseases presents the first complete genetic sequencing of a Zika virus linked to a case of microcephaly in Brazil.
The sequenced virus was isolated in November 2015 from the amniotic fluid of a pregnant woman in Paraíba whose baby was diagnosed with the malformation through ultrasound examination. At the time, evidence that the virus could cross the placental barrier and reach the fetus was reported to the Ministry of Health, helping to direct investigations into the growing number of cases of microcephaly in the country and its potential relationship with the Zika virus.
The finding was also cited by the Pan American Health Organization (PAHO) in an alert about the possible association between the virus and the malformation issued on December 1, 2015.
Through metagenomics, the researchers obtained the complete sequencing of the genome of the Zika virus present in the sample. Full sequencing demonstrated 97% to 100% homology to the genome of the strain associated with the outbreak in French Polynesia in 2013.
Zika virus was found in amniotic fluid more than two months after pregnant women showed symptoms of the disease. Photo: Gutemberg Brito
"We decided to use metagenomics to investigate not only the Zika virus but also whether other possible infectious agents, such as viruses and bacteria, could be present in the amniotic fluid. The results of the analyzes revealed that the only sequences obtained in highly relevant numbers were 'aligned' with the genome of the Zika virus. The phylogenetic analyzes of the sequences obtained showed that the Zika virus present in the amniotic fluid was genetically very close to the strain that circulated during the epidemic that occurred in French Polynesia in 2013, when an increase in the occurrence of Zika virus was also identified. congenital malformations," says virologist Ana Bispo, head of the Flavivirus Laboratory at IOC and one of the study coordinators, alongside researcher Amílcar Tanuri, from the Federal University of Rio de Janeiro (UFRJ).
The research also had the participation of scientists from the Evandro Chagas National Institute of Infectology (INI/Fiocruz) and the National Institute of Control and Quality (INCQS/Fiocruz), in Rio de Janeiro, and the Professor Joaquim Amorim Neto Research Institute (Ipesq), in Paraíba.
About the study
The article describes the analyzes that were performed, in November 2015, on amniotic fluid samples from two pregnant women from Paraíba whose fetuses were diagnosed with microcephaly by ultrasound examinations.
In both cases, molecular tests detected the presence of genetic material from the Zika virus in the samples. The patients presented Zika symptoms between the 10th and 18th week of pregnancy, more than two months before the collection of amniotic fluid, which occurred in the 28th week. For the scientists, the presence of the genetic material of the virus in the intrauterine environment so long after the acute phase of the disease suggests persistence of viral replication, which could in part be explained by the immature immunity of the fetus.
“Although the Zika virus was the only agent found in the amniotic fluid, further studies are still needed to confirm that this virus is the only cause of microcephaly in these cases, even though it is the strongest evidence so far. urgently needed to understand the biological mechanisms involved in the infection and its possible connection with fetal malformations", emphasizes Ana Bispo.
One of the research leaders, virologist Ana Bispo says that the Zika virus challenges the scientific community. Photo: Gutemberg Brito
Complete sequencing of the Zika virus genome was possible from samples from one of the cases. The virus genome sequence has been deposited in the GenBank international genome bank, which can be accessed online. While the similarity with the French Polynesian lineage that circulated in 2013 was 97% to 100%, the comparison with two viruses from Africa, identified in Uganda and Senegal, showed between 87% and 90% similarity.
The scientists also built phylogenetic trees (which indicate the genetic proximity between microorganisms in a similar way to genealogical trees, which indicate degrees of kinship). The study also revealed that the Zika virus detected in Brazil in a sample of amniotic fluid is part of the Asian lineage – the same phylogenetic group of viruses detected in the Americas, Southeast Asia and Pacific islands. On the other hand, there are the viruses detected in Africa, which are part of the African lineage. According to the researchers, phylogenetic analysis indicates that these two lineages have been evolving in isolation for over 50 years.
A comparison with genomes of other viruses belonging to the genus Flavivirus showed that, from a genetic point of view, Zika viruses, including the newly sequenced strain, are closer to Japanese encephalitis and yellow fever viruses than to dengue viruses. . The analysis found no evidence of mutation in the virus isolated from amniotic fluid. “The Zika virus challenges the scientific community with a series of questions. Our goal is to contribute to the global research effort and generate relevant knowledge on the subject”, summarizes the virologist.
Previous contributions by Fiocruzi
In 2015, the Fiocruz created the Cabinet to Combat the Sanitary Emergency of Public Health of National Importance (ESPIN/Fiocruz), which aims to make the most of the capabilities and resources available at the institution to meet the need to provide objective answers to the Ministry of Health and the population regarding the emergency situation regarding dengue, chikungunya and Zika in the country.
In November 2015, the IOC, through the Flavivirus Laboratory, concluded laboratory diagnoses that confirmed the presence of the Zika virus genome in samples of amniotic fluid from two pregnant women in the state of Paraíba, whose fetuses had microcephaly detected through ultrasound examinations. From these samples, a study was carried out on the genome of the virus recently published in The Lancet Infectious Diseases.
In January 2016, F.iocruz announced the creation of the Discriminatory NAT Kit for Dengue, Zika and Chikungunya. The innovation will ensure greater agility for the tests carried out in the Ministry of Health's laboratory network, in addition to reducing costs and allowing the replacement of foreign inputs by a national product. Conceived by IOC and developed in partnership with the Institute of Molecular Biology of Paraná (IBMP), the novelty has the support of Fiocruz-Paraná, from Fiocruz-Pernambuco and the Institute of Technology in Immunobiologicals (Bio-Manguinhos/Fiocruz).
Also in January, the Fiocruz-Paraná developed a study that confirmed the interplacental transmission of the Zika virus after analyzing the sample from the placenta of a pregnant woman in the Northeast region, who presented symptoms compatible with infection by the virus and who suffered a miscarriage – when the fetus stops developing inside the womb – in the first trimester of pregnancy. The research was carried out in partnership with the Pontifical Catholic University of Paraná (PUC-PR).
On February 5, the identification of active Zika virus particles in saliva and urine samples from patients collected during the presentation of symptoms compatible with the infection was announced. The pioneering study was led by the Flavivirus Molecular Biology Laboratory of the IOC, in collaboration with the Evandro Chagas National Institute of Infectology (INI/Fiocruz) and the Laboratory of Mosquitoes Transmitting Hematozoa of the IOC.
It was the first time that viruses with the potential to cause infection were found in these fluids, suggesting the need to investigate the epidemiological relevance of potential alternative routes of viral transmission.
Um newly published article by researchers from the Oswaldo Cruz Institute (IOC/Fiocruz) in the international scientific journal The Lancet Infectious Diseases presents the first complete genetic sequencing of a Zika virus linked to a case of microcephaly in Brazil.
The sequenced virus was isolated in November 2015 from the amniotic fluid of a pregnant woman in Paraíba whose baby was diagnosed with the malformation through ultrasound examination. At the time, evidence that the virus could cross the placental barrier and reach the fetus was reported to the Ministry of Health, helping to direct investigations into the growing number of cases of microcephaly in the country and its potential relationship with the Zika virus.
The finding was also cited by the Pan American Health Organization (PAHO) in an alert about the possible association between the virus and the malformation issued on December 1, 2015.
Through metagenomics, the researchers obtained the complete sequencing of the genome of the Zika virus present in the sample. Full sequencing demonstrated 97% to 100% homology to the genome of the strain associated with the outbreak in French Polynesia in 2013.
Zika virus was found in amniotic fluid more than two months after pregnant women showed symptoms of the disease. Photo: Gutemberg Brito
"We decided to use metagenomics to investigate not only the Zika virus but also whether other possible infectious agents, such as viruses and bacteria, could be present in the amniotic fluid. The results of the analyzes revealed that the only sequences obtained in highly relevant numbers were 'aligned' with the genome of the Zika virus. The phylogenetic analyzes of the sequences obtained showed that the Zika virus present in the amniotic fluid was genetically very close to the strain that circulated during the epidemic that occurred in French Polynesia in 2013, when an increase in the occurrence of Zika virus was also identified. congenital malformations," says virologist Ana Bispo, head of the Flavivirus Laboratory at IOC and one of the study coordinators, alongside researcher Amílcar Tanuri, from the Federal University of Rio de Janeiro (UFRJ).
The research also had the participation of scientists from the Evandro Chagas National Institute of Infectology (INI/Fiocruz) and the National Institute of Control and Quality (INCQS/Fiocruz), in Rio de Janeiro, and the Professor Joaquim Amorim Neto Research Institute (Ipesq), in Paraíba.
About the study
The article describes the analyzes that were performed, in November 2015, on amniotic fluid samples from two pregnant women from Paraíba whose fetuses were diagnosed with microcephaly by ultrasound examinations.
In both cases, molecular tests detected the presence of genetic material from the Zika virus in the samples. The patients presented Zika symptoms between the 10th and 18th week of pregnancy, more than two months before the collection of amniotic fluid, which occurred in the 28th week. For the scientists, the presence of the genetic material of the virus in the intrauterine environment so long after the acute phase of the disease suggests persistence of viral replication, which could in part be explained by the immature immunity of the fetus.
“Although the Zika virus was the only agent found in the amniotic fluid, further studies are still needed to confirm that this virus is the only cause of microcephaly in these cases, even though it is the strongest evidence so far. urgently needed to understand the biological mechanisms involved in the infection and its possible connection with fetal malformations", emphasizes Ana Bispo.
One of the research leaders, virologist Ana Bispo says that the Zika virus challenges the scientific community. Photo: Gutemberg Brito
Complete sequencing of the Zika virus genome was possible from samples from one of the cases. The virus genome sequence has been deposited in the GenBank international genome bank, which can be accessed online. While the similarity with the French Polynesian lineage that circulated in 2013 was 97% to 100%, the comparison with two viruses from Africa, identified in Uganda and Senegal, showed between 87% and 90% similarity.
The scientists also built phylogenetic trees (which indicate the genetic proximity between microorganisms in a similar way to genealogical trees, which indicate degrees of kinship). The study also revealed that the Zika virus detected in Brazil in a sample of amniotic fluid is part of the Asian lineage – the same phylogenetic group of viruses detected in the Americas, Southeast Asia and Pacific islands. On the other hand, there are the viruses detected in Africa, which are part of the African lineage. According to the researchers, phylogenetic analysis indicates that these two lineages have been evolving in isolation for over 50 years.
A comparison with genomes of other viruses belonging to the genus Flavivirus showed that, from a genetic point of view, Zika viruses, including the newly sequenced strain, are closer to Japanese encephalitis and yellow fever viruses than to dengue viruses. . The analysis found no evidence of mutation in the virus isolated from amniotic fluid. “The Zika virus challenges the scientific community with a series of questions. Our goal is to contribute to the global research effort and generate relevant knowledge on the subject”, summarizes the virologist.
Previous contributions by Fiocruzi
In 2015, the Fiocruz created the Cabinet to Combat the Sanitary Emergency of Public Health of National Importance (ESPIN/Fiocruz), which aims to make the most of the capabilities and resources available at the institution to meet the need to provide objective answers to the Ministry of Health and the population regarding the emergency situation regarding dengue, chikungunya and Zika in the country.
In November 2015, the IOC, through the Flavivirus Laboratory, concluded laboratory diagnoses that confirmed the presence of the Zika virus genome in samples of amniotic fluid from two pregnant women in the state of Paraíba, whose fetuses had microcephaly detected through ultrasound examinations. From these samples, a study was carried out on the genome of the virus recently published in The Lancet Infectious Diseases.
In January 2016, F.iocruz announced the creation of the Discriminatory NAT Kit for Dengue, Zika and Chikungunya. The innovation will ensure greater agility for the tests carried out in the Ministry of Health's laboratory network, in addition to reducing costs and allowing the replacement of foreign inputs by a national product. Conceived by IOC and developed in partnership with the Institute of Molecular Biology of Paraná (IBMP), the novelty has the support of Fiocruz-Paraná, from Fiocruz-Pernambuco and the Institute of Technology in Immunobiologicals (Bio-Manguinhos/Fiocruz).
Also in January, the Fiocruz-Paraná developed a study that confirmed the interplacental transmission of the Zika virus after analyzing the sample from the placenta of a pregnant woman in the Northeast region, who presented symptoms compatible with infection by the virus and who suffered a miscarriage – when the fetus stops developing inside the womb – in the first trimester of pregnancy. The research was carried out in partnership with the Pontifical Catholic University of Paraná (PUC-PR).
On February 5, the identification of active Zika virus particles in saliva and urine samples from patients collected during the presentation of symptoms compatible with the infection was announced. The pioneering study was led by the Flavivirus Molecular Biology Laboratory of the IOC, in collaboration with the Evandro Chagas National Institute of Infectology (INI/Fiocruz) and the Laboratory of Mosquitoes Transmitting Hematozoa of the IOC.
It was the first time that viruses with the potential to cause infection were found in these fluids, suggesting the need to investigate the epidemiological relevance of potential alternative routes of viral transmission.
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