Date: Mon 1 Jul 2019
Source: EurekAlert!, American Association for the Advancement of Science (AAAS) [edited]
The origin of the virus responsible for the ongoing yellow fever epidemic in Brazil, the worst for 40 years, has just been identified by scientists affiliated with 2 Brazilian institutions: Adolfo Lutz Institute (IAL) and the University of Sao Paulo (USP).
By means of a molecular study of yellow fever viruses found in dead monkeys and in mosquitoes, the group discovered that the strain behind the current epidemic originated in Para state in North Brazil in 1980.
The virus infected monkeys in Para and spread from there throughout the Amazon region until it reached Venezuela and Suriname. From 2000 on, always via infection of monkeys, the disease migrated to the Center-West and Southeast of Brazil, finally reaching Sao Paulo state in 2013. The 1st deaths of humans in Sao Paulo occurred in 2016.
Findings of the study, which was supported by Sao Paulo Research Foundation - FAPESP, are published in Scientific Reports [see reference below].
The investigation was led by Mariana Sequetin Cunha, a researcher in IAL's Vector-Borne Disease Group. Scientists at the University of Sao Paulo's Tropical Medicine Institute (IMT-USP), the Federal University of Para (UFPA) and the Federal University of Sao Paulo (UNIFESP) also took part. The project was also funded by Brazil's National Council for Scientific and Technological Development (CNPq).
Since mid-2016, when the ongoing yellow fever epidemic began, 2245 cases of the disease have been confirmed, with 764 deaths, according to the Health Ministry. The largest number of cases since 1980, when the government made notification mandatory, had previously been reported in 2000. In that year, 40 people died from yellow fever.
Another face of the problem is the infection of monkeys by the same mosquitoes that transmit the virus to humans. Since 2016, public health authorities responsible for epidemiological surveillance in the Center-West, Southeast, and South, where the epidemic is concentrated, have collected the carcasses of more than 10 000 monkeys found in forests and parks, mainly howler monkeys (_Alouatta_ spp.), marmosets (_Callithrix_ spp.) and capuchins (_Sapajus_ spp). Yellow fever virus was detected in 3403, according to the Health Ministry (Boletim Epidemiologico de Febre Amarela/Yellow Fever Epidemiological Bulletin). "More than 90% of the dead monkeys are believed to be _Alouatta guariba_ [brown howler monkey]. The species is extremely susceptible to yellow fever," said Ester Sabino, Director of IMT-USP. "Troops of more than 80 monkeys were entirely destroyed," Cunha said, referring to the deaths of howler monkeys from yellow fever in Horto Florestal, a nature reserve in the north of Sao Paulo city in late 2017.
Yellow fever is an acute disease caused by a virus transmitted to monkeys and humans through the bites of infected mosquitos. The symptoms include jaundice, a yellowish or greenish pigmentation of the skin and whites of the eyes due to high bilirubin levels, reflecting liver damage.
In the sylvatic (wild) transmission cycle, the yellow fever virus circulates between mosquitoes of the genera _Haemagogus_ and _Sabethes_ and monkeys. Humans are considered incidental hosts in the sense that people are infected only if they happen to live or work in tropical forests or travel on land through such areas. In the urban transmission cycle, the virus is transmitted to humans (the main host in this case) by the mosquito _Aedes aegypti_.
Yellow fever was endemic in the South and Southeast of Brazil in the early 20th century. Urban transmission has been eradicated thanks to vaccination and action against _Ae. aegypti_ breeding sites.
In the last 2 decades, transmission to humans has occurred outside the Amazon region, where yellow fever is still endemic. Cases have been reported in humans and monkeys in Bahia, Goias, Minas Gerais, Sao Paulo, Parana, and Rio Grande do Sul [states].
Since late 2016, the disease has spread faster and farther, reaching the Atlantic Rainforest biome, with all its extraordinary biodiversity, which includes many species of monkey. Yellow fever had not occurred in these areas for decades.
In search of the yellow fever virus, Cunha and her group investigated samples of brain, liver, and spleen tissue from dead monkeys found by state public health workers and compulsorily sent for analysis to IAL, the state reference laboratory. Samples from 430 dead monkeys were tested between July 2016 and March 2017. Most were _Alouatta_, _Callithrix_, and _Sapajus_, but there were some specimens of black-fronted titi (_Callicebus nigrifrons_) and golden lion tamarin (_Leontopithecus rosalia_), an endangered species.
Investigation in search of the yellow fever virus was carried out in each one of the species. The published study contributes to a better understanding of the biotic pathways involved in the virus's spread from the Amazon to the Southeast. "The study describes the evolution of the virus in different species. The disease is milder in capuchins than in howler monkeys and marmosets," Sabino said.
Not all the dead monkeys sent to IAL died from yellow fever. "Some had been run over and others electrocuted, for example," Cunha said. "The protocol requires analysis by the reference lab of tissue samples from all dead monkeys found, whatever the circumstances."
The presence of the virus was ruled out in most cases, and even in the minority in which it was confirmed, it was not always possible to be sure that death was due to yellow fever. The disease is practically a death sentence for howler monkeys. Marmosets are susceptible but do not always die. Capuchins are considered resistant.
By mid-2017, the epidemic that began in the north of Sao Paulo state in 2016 had spread to the Campinas region, not far from the state capital. "Yellow fever virus hadn't circulated in Campinas since the early 20th century," Cunha said.
The 1st infected monkey was confirmed by IAL in July 2016. It was a capuchin from the Ribeirao Preto region. The species is resistant, so the cause of death was not reported as having been caused by yellow fever, although the virus was found in its tissue. "The animal came into contact with the pathogen via a mosquito bite but died from other causes. We wanted to find out if capuchins were acting as natural reservoirs of the virus precisely because they're resistant," Cunha said.
Yellow fever virus was found in 67 out of the 430 samples analyzed by Cunha and colleagues at IMT-USP; 30 were from howler monkeys, 9 from marmosets, and 7 from capuchins. The rest were from monkeys of unidentified genera. "In these 21 cases, the material didn't indicate the genus, but we suspect they were _Alouatta_ owing to the high viral loads in the tissues analyzed," Cunha said.
The researchers isolated the virus from each of the 67 confirmed samples, sequenced the genomes, and compared the genomes with those (available online) of viruses from the outbreaks of yellow fever that occurred between 1980 and 2015 in Brazil and neighboring countries.
They discovered that the strain responsible for the current epidemic originated in Venezuela and in Roraima state and Para State in Brazil. This is in line with prior research suggesting that the 2016-17 epidemic began in the North region and spread to the Southeast by means of a long and continuous sylvatic cycle involving mosquitoes and monkeys.
The results of the study reveal an evolutionary journey of sizable proportions in both time and space. In 1980, yellow fever virus was endemic in Para. In 2000, it reached Mato Grosso do Sul, Goias, and Minas Gerais in the Center-West. By 2004, it had crossed into Venezuela, and by 2009, it had reached Trinidad and Tobago in the Caribbean. In 2010, it was present in Roraima in the [Brazilian] far North, while one strain was found in Rio Grande do Sul in the far South. The virus arrived in Sao Paulo state in 2013.
The molecular analyses performed by Cunha and colleagues showed that the virus was fully disseminated in most Brazilian states and in Suriname by 2017.
Other researchers at IAL and IMT-USP are now conducting similar studies involving the dead monkeys collected in Sao Paulo State during the 2nd wave of the epidemic, in July 2017-June 2018, and during the 3rd wave, which began in July 2018 and will fizzle out this year  with the end of the rainy season and the advent of winter, when the mosquitoes practically stop reproducing.
Depending on the results of these forthcoming studies, it may be possible to determine whether the current epidemic in Sao Paulo state is on the wane. Alternatively, despite mass vaccination the virus could still be spreading through the monkey population and fresh outbreaks may be in the offing.
Cunha MS, da Costa AC, de Azevedo Fernandes NCC, et al. Epizootics due to yellow fever virus in Sao Paulo state, Brazil: viral dissemination to new areas (2016-2017). Sci Rep. 2019; 9(1): 5474. doi: 10.1038/s41598-019-41950-3; <https://www.nature.com/articles/s41598-019-41950-3
The report above provides a fascinating account of the recent yellow fever (YF) outbreaks and traces the movement of the virus in Brazil over time. Fortunately, in none of the widely dispersed outbreaks were there any reports of urban (people-_Aedes aegypti_-people) transmission of YF virus.
The Brazil Ministry of Health has embarked on an ambitious effort to vaccinate everyone, starting with areas of highest risk. Although that should eliminate human cases, it will not affect the sylvan (forest) cycle involving non-human primates. Meanwhile, human YF cases continue to occur in several states in Brazil this year (2019) (See Yellow fever - Americas (09): Brazil, official summary http://promedmail.org/post/20190627.6541056)
. An official 8 Jun 2019 report stated that confirmed human yellow fever cases were registered in the states of Sao Paulo (68 cases), Parana (13), and Santa Catarina (1) this year (2019). Most of the cases were rural workers or those exposed in forested areas. The 1st case of yellow fever was confirmed in Santa Catarina state. Cases in non-human primates continue to be reported also. Confirmed epizootics in non-human primates were registered in Sao Paulo (13 cases), Rio de Janeiro (1), Parana (32), Para (1), and Santa Catarina (1). Most of the confirmed epizootics were registered in the southern region (69%; 33/48) with the majority of the epizootics occurring in Parana state.
Continued YF surveillance will be essential to track YF movement in nature over time. - ProMED Mod.TY]
[Maps of Brazil: