D. Jane Hata, Ph.D., is a Consultant for the Department of Laboratory Medicine and Pathology at Mayo Clinic in Jacksonville, Florida. She holds the academic rank of Assistant Professor of Laboratory Medicine and Pathology.
Contact us: MMLHotTopics@mayo.edu.
Transcript and References
Hi, I’m Matt Binnicker, the Director of Clinical Virology and Vice Chair of Practice in the Department of Laboratory Medicine and Pathology at Mayo Clinic. Zika virus has been in the news a lot lately, due to the large outbreak occurring in Central and South America and the virus being associated with significant complications, including Guillain-Barre syndrome and fetal microcephaly. In this month’s "Hot Topic," my colleague Dr. Jane Hata will provide you with an overview of the epidemiology of Zika virus, as well as the possible clinical features of disease and the recommended laboratory tests to assist in the diagnosis of Zika virus infection. I hope you enjoy this month’s "Hot Topic," and I want to personally thank you for allowing Mayo Clinic the opportunity to be a partner in your patients’ health care.
Thank you, Dr. Binnicker, for the introduction and also to the viewers for joining us today. This “Hot Topic” will provide an update on the clinical manifestations of Zika virus, in addition to information on treatment and prevention.
We’ll also review diagnostic testing options and algorithms for detection of Zika virus.
I am a current member of the College of American Pathologist Microbiology Resource Committee.
As you view this presentation, consider the following important points regarding testing:
- How is the test going to be used in your practice?
- When should these tests be used?
- And how will results impact patient management?
Since the emergence of Zika virus in the Western hemisphere in 2013/2014, a tremendous amount of information has been generated in many different aspects of this pathogen, from vector preference to public health implications.
In today’s presentation, I will discuss the characteristics of Zika virus and its pathogenic potential, with a discussion on Zika virus and pregnancy, its emerging epidemiology, and symptoms and treatment of the disease. Appropriate use of current diagnostic methods will be discussed, and I will conclude with potential strategies to prevent infection.
Zika virus is a single-strand RNA virus in the family Flaviviridae. Other members of this family include the yellow fever virus as well as West Nile virus. Zika is also phylogenetically closely related to St. Louis encephalitis virus.
Genomic data leads us to believe that there are two major lineages of Zika: African and Asian. Strains found in the Western Hemisphere demonstrate nucleotide identity with the Asian genotype.
Zika virus is most often transmitted via a mosquito vector: Aedes aegypti. Other Aedes species such as Aedes albopictus, Aedes hensilli, and others have also been implicated. Remember that Aedes aegypti and albopictus are also responsible for transmission of dengue fever and the chikungunya virus.
Zika virus breeds in the mosquito vector, which becomes infectious after a 15-day development period and remains so for the rest of its 50-day life. The virus is spread to the next susceptible host when the mosquito takes a blood meal.
Non-Vector Borne Transmission of Zika Virus
We now know that there are multiple methods of Zika transmission that are not dependent upon presence of a mosquito vector. A needle-stick injury was reported in a research laboratory worker, reminding us all of the need for good laboratory practice.
We will address maternofetal transmission in our discussion of Zika and pregnancy shortly.
Transfusion-transmitted Zika virus infections were always considered a theoretical possibility and were confirmed in 2016 in connection with the outbreak in Brazil. In addition, asymptomatic blood donors have been identified in surveys performed in French Polynesia, Puerto Rico, Texas, and Florida.
Both the U.S. Food and Drug Administration and the American Association of Blood Banks issued guidance in order to reduce the risk of Zika transmission in the U.S. blood supply. This is challenging when you consider that most donors may be asymptomatic. Due to this complicating factor, the recommendations chose to focus on methods for nucleic acid amplification-based screening of collected blood units or utilization of pathogen inactivation technologies such as ultraviolet light or detergent treatment of blood products prior to use.
Sexual Transmission of Zika Virus
Since 2008, there have been multiple confirmed reports of Zika virus by sexual transmission. Studies have demonstrated that Zika RNA can be detected in blood and semen for six months after the onset of symptoms or last possible viral exposure.
Women who have had possible Zika virus exposure through travel or sexual contact and do not have ongoing risks for exposure should wait at least eight weeks from symptom onset or last possible exposure (if they are asymptomatic) to attempt conception.
Men with possible Zika virus exposure, regardless of symptom status, should wait at least six months from symptom onset or last possible exposure (if they are asymptomatic) before attempting conception.
Zika virus testing is not recommended for non-pregnant persons with possible exposure who do not have symptoms of Zika virus disease, including persons who are planning to attempt conception or to assess the risk for sexual transmission. Zika virus testing for this purpose remains of uncertain value because current understanding of the duration and pattern of shedding of Zika virus in reproductive tissues is limited. Falsely positive tests can result in stress and expense, and falsely negative tests might provide false reassurance and lead to inadvertent fetal exposure to Zika virus.
History and Epidemiology
To review, Zika virus was initially isolated in 1947 in sentinel rhesus monkeys used for yellow fever research in Uganda. The first human infections were then reported in 1952 in Uganda and Tanzania.
In terms of confirmed human infections, sporadic cases were reported in Nigeria, Indonesia, Thailand, and Gabon from the early 1970s to 2010.
There was a significant epidemic on the Micronesian island of Yap in 2007. This affected nearly 2% of the population (although the infection rate may have been much higher), but there were no hospitalizations or deaths reported.
In late 2013 in French Polynesia, a significantly larger epidemic was noted. Nearly 11% of the population was affected, but again, no deaths were reported.
However, Zika virus was carried to Japan, France, Easter Island, and New Caledonia via travelers from French Polynesia. There were 114 cases of Zika with self-sustained transmission that occurred in New Caledonia secondary to exposure to a single infected traveler.
South America 2014–2015
In Brazil, late in 2014, a cluster of acute erythematous rash infections were noted from 18 states in Brazil. Zika virus was identified in April of 2015, and genetic studies indicated an Asian lineage of Zika was involved.
The epidemic peaked in May 2015 and affected all age groups. This pattern would be typical of spread of a new virus in a susceptible population as there was an equal distribution of disease between both males and females.
Infections were also confirmed in El Salvador, Guatemala, Colombia, Mexico, Panama, Paraguay, and Venezuela.
Based on past history of viral spread, there was a major concern for transmission of Zika virus in connection with the millions of spectators estimated to attend the 2016 Summer Olympic Games in Rio de Janeiro. Athletes were warned to take precautions, and some athletes and spectators did not attend due to the perceived risk.
In the end, the World Health Organization reported no confirmed cases of Zika among either athletes or visitors.
In 2015, a Zika outbreak was again reported in Cape Verde in Africa with 5,000 cases reported between September and December of that year.
Geographic Distribution of Zika Virus
Here is a map of the geographic distribution of Zika virus taken from the Centers for Disease Control (CDC) website, which contains an interactive map and is an excellent source of up-to-date information. The data are based on seroprevalence studies, epidemics, and tracking of individual cases, and it is continually changing.
We now see cases throughout most of South and Central America, including Mexico, the Caribbean and Puerto Rico, and also Africa, India, and Southeast Asia.
Since 2015, there have been more than 5,200 cases reported in the United States. The vast majority have been in travelers retuning from infected areas, but local mosquito-borne transmission has been confirmed in more than 200 cases. There have been 46 confirmed cases of sexual transmission.
Zika is now a nationally reportable disease. Laboratories and health care providers are required to report suspected cases to their local or state health departments in order to facilitate diagnosis and epidemiology.
Pathogenesis Due to Zika Virus
Zika virus is considered a “biosafety level 2” pathogen, so it may be handled routinely at the laboratory bench with appropriate personal protective equipment.
As with other flaviviruses, replication of Zika virus is thought to occur in the cytoplasm or nuclei of human host dendritic cells located near the site of the bite from the infected mosquito. The virus then spreads to the lymph nodes and bloodstream.
Studies in humans, mice, and primates have indicated that Zika virus demonstrates trophism for neural progenitor tissue, resulting in neuronal degeneration and cell death.
In primate studies, after either subcutaneous or intravenous inoculation, Zika virus could be recovered from blood, saliva, urine, CSF, genital fluids, lymph nodes, heart, spleen, and GI tissue, although specific pathogenesis at the non-neural sites has not been reported.
Zika Virus and Guillain-Barré Syndrome
There has been an association noted between Zika virus infection and the occurrence of Guillain-Barre syndrome (GBS). Remember that GBS is an autoimmune-mediated syndrome that can occur after viral infections, including dengue, West Nile Virus, and chikungunya virus. GBS is characterized by development of subacute or acute flaccid paralysis following a symptomatic disease presentation.
This association was first reported in the Zika outbreak in French Polynesia, with a total of 74 cases. There was a 20-fold increase in Guillain-Barre over the normal baseline prevalence in addition to a temporal relationship to the ongoing epidemic of Zika on that island. Increases in the number of cases of GBS have also been noted in Brazil, El Salvador, and Venezuela as related to ongoing Zika virus outbreaks.
Most GBS cases associated with Zika have recovered on their own, although the use of intravenous immunoglobulin, plasma exchange, and use of intensive care facilities have been necessary in some instances.
Zika Virus and Pregnancy
One of the most disturbing aspects of Zika virus infection is its potential effect on fetal development. As we have previously discussed, Zika virus exhibits tropism for neural progenitor cells.
A 20-fold increase in reports of microcephaly in Brazilian infants was reported in October 2015, resulting in the declaration of a national health emergency. An uptick in the prevalence of microcephaly was also reported retrospectively after the French Polynesia epidemic of 2014.
The greatest risk to the fetus occurs if infection takes place during the first trimester of pregnancy. The overall risk of microcephaly is estimated to be anywhere from 1% to 13%.
At birth, Zika virus can be detected in amniotic fluid of microcephalic infants, however, the maternal blood may be Zika RNA negative. Congenital infection is possible in asymptomatic women.
Microcephaly is associated with reduced head circumference and gestational weight, brain atrophy, and calcifications, resulting in significant developmental issues and death in some cases. Ocular cataracts and auditory impairment may lead to neurodevelopmental delays.
New recommendations for Zika virus testing were released by the CDC in July 2017 due to overall declines in prevalence of Zika disease in the Americas.
Current recommendations conclude that all pregnant women should be asked about Zika infection at each prenatal visit, regardless of symptoms. An understanding of geographic areas of concern and timeframe of potential exposure is essential in determination of risk of infection.
This includes individuals with recent travel or residence in a Zika-affected area within eight weeks before conception or individuals with sexual contact with a person who traveled to or lives in an area with risk of Zika.
For pregnant, symptomatic women with ongoing risk of Zika exposure, concurrent serologic and nucleic acid testing (also known as NAT testing) of serum and urine should be performed. A positive NAT result is confirmatory for maternal Zika infection. For asymptomatic women with ongoing risk of Zika exposure, NAT testing should be offered at the initiation of prenatal care and twice more during the duration of the pregnancy.
Due to IgM persistence, Zika infection before pregnancy may confound results of serologic testing performed during pregnancy, therefore, IgM testing is no longer recommended. This is a change from previous recommendations.
For asymptomatic pregnant women with recent possible, but not ongoing exposure (such as travel), routine testing for Zika is no longer recommended, due to the risk of increased false-positive test results in light of declining Zika prevalence. However, a conversation with your health care provider in order to assess the risk is strongly encouraged.
If there is presumptive or confirmed Zika virus infection in the mother, ultrasound studies should be performed every 3 to 4 weeks in order to monitor fetal anatomy and growth. In neonates at risk, both nucleic acid testing of serum and urine and IgM testing on serum should be performed. Testing of cord blood is not recommended.
A testing algorithm for the management of pregnant women is available on the CDC Zika virus website and should be consulted for the most up-to-date recommendations.
Symptoms of Zika Virus Infection
The incubation period of Zika virus in the human host is approximately 2 to 10 days after the bite from an infected mosquito. It is very important to note that in most individuals, the disease will be quite mild with only a few days of symptoms, and in many cases, people are completely asymptomatic. This will complicate epidemiologic follow-up of suspected cases.
In the 20% of individuals who are symptomatic, you may see mild fever, muscle or joint pain, a maculopapular rash, which is occasionally itchy, and nonsuppurative conjunctivitis. Occasionally, anorexia, vomiting, diarrhea, or constipation may occur. Headache, and in some cases, retro-orbital pain has been reported.
You may recognize many of these symptoms as also being similar to dengue fever and chikungunya virus, which are also found in the geographic areas affected by Zika. Again, this will complicate the diagnostic picture.
Severe disease requiring hospitalization and death is rare.
Here is a comparison of symptoms noted in Zika virus infection, dengue virus infection, and chikungunya virus infection. These diseases demonstrate significant overlap in terms of mosquito vector transmission, geographic prevalence, and symptomology.
Note that in Zika virus infection, when patients are symptomatic, characteristics such as edema of extremities, conjunctivitis, a maculopapular rash and the absence of hemorrhage, leukopenia, and thrombocytopenia may assist in determining a diagnosis. Among these symptoms, rash and conjunctivitis are significantly more common among Zika-positive patients as compared to dengue or chikungunya.
Diagnosis of Zika Virus Infection
The definitive diagnosis of Zika virus infection can only be made on the basis of laboratory testing.
Diagnostic testing is currently recommended for assessment of infection in individuals who are symptomatic, have a significant travel history to, or residence in, a Zika-affected area.
As discussed in a previous slide, all pregnant women should be asked about Zika exposure. Pregnant women should be tested based on presentation, and this includes pregnant women exposed via sexual partners who have lived in or traveled to a Zika-risk area.
Current testing methods for assessment of acute infection rely on serologic testing for presence of IgM to Zika virus and detection of Zika virus RNA in serum and urine.
Due to the many similarities in symptoms, providers should also consider also testing for dengue and chikungunya in symptomatic patients.
Serologic Testing for Zika Virus
Zika virus IgM may not be detectable until 4 days after onset of symptoms and remain detectable up to 12 weeks later.
The CDC Zika IgM Antibody Capture Enzyme-Linked Immunosorbent Assay, otherwise known as the “Zika MAC-ELISA test” is used in qualified public health and reference laboratories in the United States for the qualitative detection of Zika virus IgM antibodies in serum or cerebrospinal fluid collected from persons meeting the clinical and epidemiologic criteria for suspected Zika virus disease.
Confirmation of any non-negative IgM result will require the use of a plaque-reduction assay to detect specific neutralizing antibody to Zika virus. The PRNT assay is only performed at the CDC or certain designated state laboratories.
It should be noted that with current assays, “cross reactivity” with other flaviviruses leading to false-positive results is an issue. Not only does this apply to previous infections with viruses such as St. Louis encephalitis or West Nile virus, but Zika serologic false-positives could also occur in individuals who have received vaccinations for other flaviviruses such as yellow fever or Japanese encephalitis.
Therefore, the plaque-reduction neutralization test (or PRNT test) measures Zika virus-specific neutralizing antibody titers and should be performed to rule out false-positive ELISA results due to other flaviviruses.
RT-PCR (NAT) Testing for Zika Virus
Nucleic acid detection by reverse transcriptase-polymerase chain reaction targeting the non-structural protein 5 genomic region is the primary means of definitive diagnosis of Zika.
RT-PCR, or NAT testing for Zika virus, is extremely sensitive and specific; however, due to the short period of viremia (the presence of a virus in the blood), serum and urine samples must be collected within 14 days of the onset of symptoms. If possible, urine should always be collected with a patient-matched serum specimen.
If the specimen can be collected within 14 days of illness onset, the use of PCR will nullify the cross-reactivity issues noted with serologic testing for Zika.
If a PCR result is positive, that is considered confirmatory, and no additional testing is required. If molecular testing for Zika is negative, this does not rule out the possibility of Zika virus infection, and serological testing for Zika IgM is recommended.
There are now a number of molecular tests from different manufacturers that have obtained “emergency use authorization” (EUA) from the FDA for Zika diagnosis. Generally speaking, testing can be performed on serum, plasma, or urine.
Here are some factors to consider if your laboratory needs to pursue Zika virus testing. Both serological and molecular testing for Zika virus is available from reference laboratories and through certain state departments of health.
It is very important to have a conversation with your testing laboratory prior to specimen submission to determine if testing is available and what types of samples are acceptable, especially for molecular testing. The availability of testing and specimen requirements may vary, so ask first.
If you are submitting specimens for testing, you will need to supply the following information:
It is essential to know when samples were collected in relationship to symptoms and any pertinent travel, residence, and medical history (including pregnancy status) in order to determine the most appropriate testing.
Algorithms to guide when and how Zika testing should be performed are available at www.cdc.gov/zika.
Treatment of Zika Virus Infections
The treatment for Zika virus infections in symptomatic individuals consists of supportive care only and is mainly comprised of rest and fluids. This may also include aspirin, acetaminophen, or other non-steroidal anti-inflammatory agents for joint pain and headache.
Testing to rule out eengue fever should be performed before use of these agents due to the possibility of hemorrhage.
Antihistamines are helpful for an itchy rash.
No antiviral agents have demonstrated activity against Zika.
There are no specific recommendations for treatment of Zika virus in pregnant women or microcephalic infants.
Individuals infected with Zika should avoid mosquito exposure in order to reduce the risk of local transmission. The possibility of sexual transmission should also be considered, and condom use is encouraged.
Will We See a Zika Vaccine?
Will we see a Zika vaccine? It’s very possible. An amazing amount of research in this area in the last several years has given us the following possibilities for vaccine development:
Based on mouse and primate studies, a focus on development of Zika-neutralizing antibody in the host is believed to be effective in preventing infection. Toward that end, approaches based on development of a DNA vaccine expressing Zika pre-membrane and envelope protein sequences, an adenovirus vector using those same protein sequences or an inactivated purified Zika virus are being actively investigated.
All of these approaches have resulted in Phase 1 clinical trials to prove vaccine safety and immunogenicity. If efficacy is demonstrated, additional trials will continue. The major challenge with Zika-vaccine development will be a formulation that is sufficiently immunogenic, has acceptable side effects after administration, and is effective and safe for use in pregnant women.
Zika Transmission and Travel
Although we have started to see an overall decreased prevalence of infection, local transmission of Zika virus has occurred in Florida, Texas, and Hawaii. Most Zika cases have been directly associated with travel to an affected area, primarily South America or the Caribbean.
As the vast majority of cases have been travel-related, be an informed traveler before you go.
Recommendations will change, so consult the CDC’s “travel page” on its website for the most up-to-date information.
Most importantly, if you are pregnant, if at all possible, you should consider postponing travel to areas with reported Zika virus transmission. If you must travel, avoidance of mosquito bites and use of repellent will not only protect against Zika but also exposure to dengue, chikungunya, and malaria. Women who are trying to become pregnant should discuss risks of Zika with their medical provider before they travel.
If you do become ill after travel to an endemic area, see your medical provider without delay and provide a specific travel history.
Also, if you are ill, protect yourself against mosquito bites in order to prevent the possibility of viral transmission to others. Clean up the immediate environment by removing debris and sources of standing water where mosquitos breed.
Apply insecticides such as DEET or use permethrin-treated clothing. And use barriers such as long pants, long sleeves, and window screens.
To summarize today’s presentation:
Zika virus has rapidly emerged around the world, including the United States. Most cases are in association with a human-mosquito-human cycle. However, there is a risk of sexual-, laboratory-, and blood transfusion-transmitted infections. Of greatest concern is the possibility of Zika transmission in pregnant women due to the potentially devastating neurotrophic effects of Zika infection in the developing fetus.
Most infections in adults are asymptomatic or mild, consisting of fever, arthralgia, headache, and rash.
Symptoms can be confused with dengue or chikungunya. Conjunctivitis, extremity edema, and rash are more likely to occur in association with Zika virus infection.
Only supportive treatment is recommended at this time.
Rarely, more severe cases of Zika can result in manifestations such as Guillain-Barre syndrome.
Zika virus infection in expectant women has been positively liked to microcephaly and other fetal brain abnormalities. The overall risk of microcephaly if infection occurs during the first trimester is estimated to be between 1% and 13%.
If you are traveling to an area in which Zika has been reported, avoid mosquito bites. If you are ill, avoid mosquito bites in order to prevent transmission to others.
Finally, there are specific travel recommendations that have been issued by the CDC in response to this epidemic. Women who are pregnant are currently advised against travel to areas where Zika transmission is ongoing. These recommendations will change over time, so consult the CDC’s travel page on its website for the most up-to-date information.
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