Zika Incidence Timeline

In May of 2015, health officials at the Pan American Health Organization (PAHO) issued an alert regarding the first confirmed Zika virus infection in Brazil, where a sudden and sustained incidence of babies born with microcephaly was observed. These babies were born, in some cases, to mothers that had experienced a brief period of fever and rash during their pregnancy. 

On January 15, 2016 in response to a continued increase in the number of Zika cases and babies born with microcephaly, the Centers for Disease Control (CDC) issued a travel alert regarding Zika virus. In this alert it was noted that in May 2015 the World Health Organization (WHO) “reported the first local transmission of Zika virus in the Western Hemisphere, with autochthonous (locally acquired) cases identified in Brazil" . 

The reports from Brazil and other Central and South American countries regarding Zika have increased since this travel alert. As of March 31, “Microcephaly and other fetal malformations have been reported in Brazil (944 cases), Cabo Verde (2 cases), Colombia (32 cases), French Polynesia (8 cases), Martinique (1 case) and Panama (1 case). Two additional cases, linked to a stay in Brazil, were detected in the United States of America and Slovenia”, according to the Pan American Health Organization web site

History of Zika Virus

Although many of us had not heard of Zika virus before late 2015, it is not a new virus. Previous outbreaks have occurred in Asia, Africa and Pacific Islands. Zika virus was initially reported in Uganda in 1947 where it was isolated from a sentinel Rhesus monkey. In 1952 the first human cases of Zika infection were reported, presumed to have been transmitted from an infected monkey to a human by mosquitoes (Dick G.W. et al.). The name Zika comes from this first isolation in Uganda, which occurred near the Zika forest. These maps depict how the virus has spread globally since the 1940s.  

While Zika is not new, the virus is demonstrating frightening new casualties in humans, such as underdeveloped heads (microcephaly) in fetuses and newborns, as well as a marked increase in the incidence of Guillain-Barre syndrome in Zika-affected areas.

Virus Details

Zika virus is a mosquito-borne RNA virus of the family Flaviviridae; there are upwards of 70 viruses in this family. Some of the better-known flaviviruses include Dengue fever, Yellow fever and West Nile virus. RNA viruses have RNA as their genetic material instead of DNA. The tricky piece about RNA viruses, which include Ebola and HIV, is that they have a much higher mutation rate than DNA viruses. Viruses with higher mutation rates are more difficult vaccine targets. 

Zika, like other flaviviruses, is spread by mosquitoes of the Aedes family. In Brazil, one of the countries hardest hit by Zika virus recently, Aedes aegypti is the most common Aedes family member. This particular mosquito is also found in approximately 20 of the 50 United States, mostly in southern and eastern seaboard states

About the Mosquito

The Aedes sp. mosquitoes that are known to carry Zika virus, include Aedes africanus in Uganda, and Aedes aegypti and Aedes albopictus in the Americas. There are important differences in the behavior of these mosquitoes, in particular the time of day at which they feed and their choice of animal on which to feed. 

The CDC notes that “Aedes aegypti bites primarily during the day. This species is most active for approximately two hours after sunrise and several hours before sunset, but it can bite at night in well-lit areas. This mosquito can bite people without being noticed because it approaches from behind and bites on the ankles and elbows. Aedes aegypti prefers to bite people but it also bites dogs and other domestic animals, mostly mammals”. Only female Aedes sp. take blood meals. 

Aedes africanus, the mosquito that carries Zika virus in Uganda, on the other hand, is more reserved in its feeding behavior. This mosquito prefers monkeys to humans and feeds mostly in forested areas away from urban settings. Likewise, Aedes albopictus is known to prefer non-urban areas and animals to humans for feeding. 

In considering ways to avoid mosquitoes, it’s important to note that the lifespan of Aedes mosquitoes is fairly short, typically 4–6 weeks. The mosquitoes generally have a limited area of travel, living and feeding close to where eggs are laid and hatched.  

Zika Virus Research

Evidence of Zika virus has been found in tissues from pregnant women with symptoms such as fever and rash who later gave birth to microcephalic babies, and in babies born with microcephaly (Calvet G. et al.). In addition there is now evidence from laboratory studies demonstrating the ability of Zika virus to infect human skin (Hamel R. et al.) and neural progenitor cells (Tang H. et al.). These studies provide strong evidence of a link between Zika infection and microcephaly, in part by showing the ability of this virus to preferentially infect certain cell types. Here is a closer look at what these two reports revealed.  

Study Details

Hamel et al. examined skin cells for permissiveness to Zika virus entry. They showed that Zika can infect keratinocytes resulting in morphologic changes. In addition, they demonstrated that 50% of infected immature dendritic cells expressed viral envelope proteins, and saw that Zika virus induced an innate antiviral response in primary human skin fibroblasts.

Tang et al. looked for a target for Zika virus in human nervous system cells. They used human induced pluripotent stem cells that were differentiated into forebrain-specific human neural progenitor cells (hNPCs). They infected cells with Zika virus and quantitated infection by RT-PCR using primers specific to Zika strain MR766. They showed that 65–90% of cells were infected within 3 days. Moreover, these authors found that Zika virus infection of hNPCs attenuated cell growth in vitro, a step toward understanding the as-yet-unknown pathway of Zika virus to fetal brain tissue.

Detection is Key

In the Centers for Disease Control Jan. 15, 2016 health advisory it was noted that “there is no diagnostic test for Zika”. That was true on Jan. 15. However, on Jan. 29, 2016, a new detection test became available. This test kit was developed by GenArraytion, Inc. as a “For Research Use Only” means of detecting the presence of Zika virus in mosquito populations.

To develop the detection assay, GenArraytion scientists obtained Zika virus sequence information from GenBank and used in-house bioinformatics to evaluate the virus genome. From this information, they developed primers for 4 unique regions of the Zika virus genome, and then a new detection test for the virus. 

The test uses an RNA sample isolated from mosquitoes that is reverse transcribed to cDNA. The cDNA, GenArraytion MultiFLEX ™ kit primers and polymerase are added to a reaction tube or plate well, along with other essential PCR components, and PCR is run. (The GenArraytion MultiFLEX™ kit does not include a polymerase, but can be used and was validated with GoTaq® Probe 1-Step RT-qPCR System and GoTaq® Probe qPCR Master Mix from Promega.)  

The PCR product is then combined with a capture bead-based assay available from Luminex. Alternatively, real-time PCR can be performed on mosquito samples using fluorescent probes and a real-time PCR platform to detect results. It takes about 4 hours from the start of PCR to results.

Recent Research and Support

In an effort to support Zika virus researchers in their efforts, here are several research publications in which Promega products were used:

Mutebi, J–P. et al. (2004) Genetic Relationships and Evolution of Genotypes of Yellow Fever Virus and Other Members of the Yellow Fever Virus Groups within the Flavivirus Genus Based on the 3’ Noncoding Region. The researchers used pGEM®-T Easy Vectors (Cat.# A1360) to clone Zika virus PCR products.

Hamel, R. et al. (2015) Biology of Zika Virus Infection in Human Skin Cells. M-MLV Reverse Transcriptase (Cat.# M3681) was used with Zika virus RNA in real-time RT-PCR.

Tang, H. et al. (2016) Zika virus infects human cortical neural progenitors and attenuates their growth. GoTaq Green Master Mix (Cat.# M7122) was used in PCR.

Drs. Schaudies and Robinson, in developing the GenArraytion MultiFLEX™ kit, used GoTaq® Probe 1-Step RT-qPCR System (Cat.# A6120) and GoTaq® Probe qPCR Master Mix 
(Cat.# A6102).


Images were generated using information from the Centers for Disease Control.