Aedes aegypti infected with chikungunya is identified in Northeast Brazil

Discovery published in PLOS Neglected Tropical Diseases reinforces evidence that A. aegypti was main vector involved in outbreaks of chikungunya seen in 2015 and 2016.

By Karina Toledo  |  Agência FAPESP – For the first time in Brazil a group of scientists have identified a mosquito of the species Aedes aegypti naturally infected by the virus that causes chikungunya fever. The discovery was made in Aracaju, the capital of Sergipe State. Their report has just been published in the journal PLOS Neglected Tropical Diseases.

According to the authors, the discovery reinforces the evidence that A. aegypti was the main vector involved in the outbreaks of chikungunya seen in 2015 and 2016, especially in Brazil’s Northeast region.

More than 265,000 probable cases of chikungunya fever had been reported in Brazil by December 2016, according to Health Ministry data. Almost 40,000 suspected cases were notified in 2015. Despite the larger number of cases, no infected mosquitoes have ever been found in Brazil before.

“In fact, for the Americas as a whole, there’s only been one description of a chikungunya-infected mosquito. It was a specimen of A. aegypti found in Mexico to be infected by the Asian genotype. The mosquito we found in Aracaju had been infected by the East/Central/South African or ECSA genotype. Both strains have been found in patients in Brazil,” said André Luis Costa-da-Silva, a postdoctoral fellow at the University of São Paulo’s Biomedical Science Institute (ICB-USP) and first author of the article.

The research was carried out under the aegis of the Zika Virus Research Network in São Paulo (Rede Zika), supported by FAPESP. The principal investigator was Margareth Capurro, a professor at ICB-USP.

Mosquitos were collected in February 2016 in six neighborhoods of Aracaju where there were large numbers of people with symptoms of infection by arthropod-borne viruses (arboviruses), such as high fever and muscle or joint ache. Strategic sites were located with the help of researchers at the Sergipe Central Public Health Laboratory (LACEN), assisted by a team from the Health Ministry. Other researchers from the University of São Paulo (USP) and Butantan Institute also took part in this multidisciplinary investigation in Aracaju.

The analysis focused on the three main arboviruses circulating at the time – dengue, Zika and chikungunya.

“The mosquitoes were captured alive using vacuum aspirators both inside homes and in their vicinity,” Costa-da-Silva said. “Each home had its own collection vial. The mosquitoes were sorted by species and sex in the lab. The females were subdivided into those with an engorged abdomen [showing they had recently fed on blood] and those with a normal abdomen. They were then frozen and sent for analysis in São Paulo.”

They collected 194 mosquitoes of the species Culex quinquefasciatus, the southern house mosquito. Next came A. aegypti, with 50 specimens. Two other species of the genus Aedes were also captured: A. scapularis (two specimens), and A. taeniorhynchus (also two specimens).

In the lab Capurro heads at USP, the genetic material in the mosquitoes was analyzed using a molecular biology technique called real-time polymerase chain reaction (PCR). Viral genotype sequencing and analysis were performed in partnership with US collaborators and researchers at USP’s Molecular Evolution & Bioinformatics Laboratory (LEMB) headed by Professor Paolo Zanotto.

In the case of male mosquitoes, all the RNA present in the insect’s body was analyzed together, but in the case of females, a more detailed analysis was performed using only RNA extracted from the thorax, where the salivary glands are located, in order to avoid a false-positive result.

“We took this precaution because females can feed on contaminated blood without becoming infected, in which case if we analyzed all the RNA present in their bodies we could detect the virus,” Costa-da-Silva said.

None of the 248 mosquitoes analyzed was infected with dengue or Zika virus, and only one female specimen of A. aegypti contained genetic material from chikungunya virus.

“It may sound negligible, but one infected female in a group of 50 is considered significant,” Costa-da-Silva said. “It suggests high viral circulation and possibly an ongoing epidemic.”

Zika

For Capurro, no Zika-infected mosquitoes were found in this study because the epidemic in the Northeast had already peaked when the specimens were collected. The number of cases began rising explosively in October 2015.

“The team were ready to go at once. FAPESP quickly approved the funding. It took longer to negotiate permission to collect specimens with the state health authorities. Despite the delay we succeeded in showing what no one had been able to confirm in Brazil hitherto,” Capurro said.

When the numbers of Zika and chikungunya cases began to surge, she recalled, there was a suspicion that the house mosquito might also be a vector of these diseases, but so far this has not been scientifically confirmed.

“Strong proof of the vector or vectors involved in an epidemic has a direct impact on control measures,” she said. “Different species require completely different control protocols. While this kind of knowledge is very important, almost all the investment in research on Zika has gone into the development of vaccines and diagnostic methods. There hasn’t been any investment in pilot programs for mosquito control.”

According to Costa-da-Silva, the group now plan to conduct similar collections and analyses to those described in this study in the Southeast, focusing on Zika virus detection. “This hasn’t been done so far for lack of promising areas for collection with large numbers of people who have the right symptoms,” he said.

The article “First report of naturally infected Aedes aegypti with chikungunya virus genotype ECSA in the Americas” by André Luis Costa-da-Silva, Rafaella Sayuri Ioshino, Vivian Petersen, Antonio Fernando Lima, Marielton dos Passos Cunha, Michael R. Wiley, Jason T. Ladner, Karla Prieto, Gustavo Palacios, Danuza Duarte Costa, Lincoln Suesdek, Paolo Marinho de Andrade Zanotto and Margareth Lara Capurro can be read at: journals.plos.org/plosntds/article?id=10.1371/journal.pntd.0005630.