Mosquito-borne arboviruses are transmitted to humans through the bite of an infected mosquito, and infections typically occur during the summer months when insects are most active. In the United States, the most common mosquito-borne illness is West Nile virus (WNV), a neurotropic human pathogen capable of spreading to the brain via hematogenous dissemination. Other neurotropic mosquito-borne arboviruses, including St. Louis encephalitis (SLE) virus and Japanese encephalitis virus, are less common but may cause similar symptoms.
Emerging arboviruses, including dengue, Zika, and chikungunya, have similar epidemiologies, transmission cycles, and clinical symptoms at onset (although complications vary substantially). Laboratory testing options for mosquito-borne arboviruses include serology and nucleic acid amplification testing (NAAT). The appropriate methodology depends on the suspected infection and duration of symptoms.
Quick Answers for Clinicians
Given the decline in Zika virus outbreaks, the CDC has updated its testing guidance for persons who live in or have recently traveled to areas with active dengue transmission and a risk of Zika virus infection. Zika virus testing is not advised in nonpregnant individuals. Refer to the Summary of Laboratory Testing Recommendations for Pregnant Women and Infants table for a full list of indications.
An unusually high number of Eastern equine encephalitis (EEE) cases (38 cases, including 19 deaths) were reported to the CDC in 2019. Most of these cases occurred in the northeastern United States. By contrast, an average of seven human cases of EEE were reported in the years between 2009 and 2018. Despite the rise in reported cases, EEE is relatively uncommon; only 4-5% of people bitten by an infected mosquito will develop the disease. Of those, roughly one-third of individuals who develop severe EEE die from the disease, whereas others may experience ongoing neurologic complications.
As with other mosquito-borne arboviruses, testing may be indicated in symptomatic individuals who have either lived in or traveled to areas where EEE is circulating. A panel that tests for related and/or cocirculating arboviral illnesses may be considered; if the patient presents with encephalitis, cerebrospinal fluid (CSF) testing is warranted.
Many mosquito-borne viruses cocirculate in some areas of the world and can cause similar clinical presentations. Moreover, serology testing can be complicated by cross-reactivity with related arboviruses. Therefore, adequate history of travel and vaccinations, especially if an individual has been in areas where viruses are cocirculating or where immunizations (eg, for yellow fever) are routine, can help determine the best strategy for further testing to identify the infecting virus. Plaque-reduction neutralization testing (PRNT) is recommended for confirmation because it can resolve nonspecific reactivity and, in some cases, identify the infecting virus.
Refer to the following sites for national and global maps of mosquito-borne virus circulation trends:
- CDC ArboNET : up-to-date provisional reporting of arboviruses circulating in the United States, including West Nile virus (WNV), St. Louis encephalitis (SLE) virus, Eastern equine encephalitis (EEE) virus, and dengue, chikungunya, and Zika viruses
- Zika virus travel information : global map of countries and territories with Zika virus cases
- DengueMap : global map of dengue virus risk throughout the world
- Chikungunya geographic distribution : map of countries and territories where chikungunya virus cases have been reported
Refer to the following disease-specific CDC sites for current testing information:
Neurotropic Mosquito-Borne Viruses
West Nile Virus
In the U.S., the most common mosquito-borne illness is WNV. WNV can range in severity from asymptomatic disease to severe meningitis and encephalitis. The milder version of WNV, referred to as West Nile fever, can be characterized by an abrupt onset of fever, headache, and possibly rash, among other symptoms. The more severe neuroinvasive disease affects one in 150 infected individuals and presents with meningitis, encephalitis, or flaccid paralysis syndrome.
Indications for Testing
Testing for WNV is indicated for patients who either live in or have traveled to areas where WNV is circulating during mosquito season and who present with acute, unexplained febrile illness, rash, and/or meningitis/encephalitis.
Diagnostic Laboratory Testing
Serology
Acute WNV infection is diagnosed by testing serum or cerebrospinal fluid (CSF) for WNV-specific immunoglobulin M (IgM) antibodies. WNV IgM antibodies are detectable 3 to 8 days after illness onset, but in some cases can persist for 30 to 90 days. IgG is detectable approximately 3 weeks postinfection and persists for years; the presence of IgG only indicates previous exposure. False-positive results can occur due to cross-reactivity with other arboviruses (eg, Eastern equine encephalitis [EEE] virus, Western equine encephalitis [WEE] virus, SLE virus) and dengue virus. Plaque-reduction neutralization testing (PRNT) may be required to determine the specific infecting virus.
Nucleic Acid Amplification Testing
NAAT is not recommended for diagnosis and is typically reserved for immunocompromised patients or for clarifying equivocal serology results.
Other Neurotropic Arboviruses
Although less common than WNV, the following arboviruses are endemic in the U.S. and can cause meningitis and encephalitis:
Japanese encephalitis virus, endemic in Asia, can also cause meningitis and encephalitis.
These viruses are diagnosed in a similar fashion as WNV. ARUP Laboratories offers an arboviral serology panel that detects arboviruses endemic to the U.S., and specific serology tests are available for Japanese encephalitis virus.
Nonneurotropic Mosquito-Borne Viruses
Zika Virus
Zika virus is a member of the Flavivirus genus and is transmitted to humans by Aedes spp mosquitos. Transmission is also possible during sexual contact, blood product transfusions, and pregnancy (from mother to fetus). Notably, infection with Zika virus during pregnancy has been linked to congenital microcephaly and other serious brain defects in fetuses and infants. Human-to-human transmission apart from sexual transmission has been described, but the mechanism of transmission is uncertain. The majority of individuals infected with Zika virus are asymptomatic. In symptomatic individuals, infection is usually mild and self-limiting and may last for several days to a week. Zika viral infection symptoms include fever, headache, retro-orbital pain, conjunctivitis, maculopapular rash, myalgia, and arthralgia. These symptoms are not unique to Zika virus and may be seen in patients infected with other arboviruses, such as dengue and chikungunya.
Diagnostic tests for Zika virus include NAAT and IgM serology. The test of choice depends on when the patient presents in relation to symptom onset or last possible exposure to Zika virus. Due to cocirculation and cross-reactivity, additional testing for dengue and chikungunya viruses should be considered.
Indications for Testing
Zika virus outbreaks have declined significantly since 2016 and are now outnumbered by reported dengue virus cases. Given this arboviral epidemiologic shift, Zika testing is not recommended for nonpregnant individuals; symptomatic nonpregnant individuals are advised to consider dengue virus testing. In the event that a country reports an outbreak of Zika virus, the CDC’s dengue and Zika virus diagnostic testing for patients with a clinically compatible illness and risk for infection with both viruses should be followed. The CDC’s current Zika virus testing guidance by population is detailed in the table below.
Population | Patient Characteristic(s) | Recommendation(s) |
---|---|---|
Pregnant women | Symptomatic |
Perform dengue and Zika virus NAAT testing on a serum specimen and Zika NAAT on a urine specimen Perform dengue IgM testing Refer to the CDC’s Zika and Dengue Testing Guidance (updated November 2019) for more information |
Asymptomatic, living in or with recent travel to the U.S. and its territories | Routine Zika testing is NOT recommended | |
Asymptomatic, living in or with recent travel to an area with a risk of Zika virus outside of the U.S. and its territories | Routine Zika testing is NOT recommended, but NAAT testing may still be considered | |
Prenatal ultrasound findings consistent with congenital Zika virus infection; residence in or travel to areas with a risk of Zika virus during pregnancy |
Perform Zika virus NAAT and IgM testing on maternal serum and NAAT on maternal urine If Zika NAAT is negative and IgM is positive, confirmatory PRNT should be performed against Zika and dengue If amniocentesis is performed as part of clinical care, Zika virus NAAT testing of amniocentesis specimens should also be performed; refer to the CDC’s Zika and Dengue Testing Guidance (updated November 2019) for more information |
|
Infants | Birth defects consistent with congenital Zika syndrome and born to mothers with possible Zika virus exposure during pregnancy, regardless of the mother’s Zika virus testing result |
Testing recommended Perform serum and urine NAAT and serum IgM serology concurrently If CSF is obtained for other purposes, perform NAAT and IgM antibody testing on CSF Cord testing is not recommended; refer to the CDC testing algorithm for infants with possible congenital Zika virus infection for more information |
No birth defects consistent with congenital Zika syndrome but born to mothers with laboratory evidence of possible Zika virus infection during pregnancy |
Testing recommended Perform serum and urine NAAT and serum IgM serology concurrently If CSF is obtained for other purposes, perform NAAT and IgM antibody testing on CSF Cord testing is not recommended; refer to the CDC testing algorithm for infants with possible congenital Zika virus infection for more information |
|
No birth defects consistent with congenital Zika syndrome and born to mothers without laboratory evidence of possible Zika virus infection during pregnancy | No testing recommended | |
Source: CDC, 2019 ; CDC, 2020 |
Diagnostic Laboratory Testing
Nucleic Acid Amplification Testing
The CDC recommends molecular testing by NAAT on serum and urine in symptomatic pregnant women who meet the criteria in the table above during the acute phase of the disease (<14 days since symptom onset). Refer to testing guidance for dengue virus if patients are symptomatic and not pregnant. Zika testing is not currently recommended for this group based on the current epidemiology of the virus.
NAAT tests may not detect Zika virus RNA in an infant who had a Zika virus infection in utero or in a child if the period of viremia has passed.
Serology
Zika virus-specific IgM and neutralizing antibodies typically develop toward the end of the first week of illness. IgM levels are variable but are generally positive beginning about 4 days after symptom onset and typically persist for up to 12 weeks after symptom onset or exposure (but may persist longer). False-positive Zika virus IgM serology results may occur in patients with a recent closely related arboviral infection (eg, dengue virus) or with recent vaccinations to related arboviruses (eg, Japanese encephalitis or yellow fever). Therefore, concurrent evaluation for other possible cocirculating arboviruses, especially dengue and chikungunya, should be considered. The CDC currently does not advise the use of Zika virus-specific serology testing in nonpregnant individuals.
Clinical decisions regarding patient management cannot be based on a positive IgM result alone. Confirmatory PRNT for Zika virus neutralizing antibodies is performed by the CDC or a CDC-designated laboratory on all presumptive positive Zika IgM results. ARUP provides these test results in the final patient chart.
Dengue Virus
Dengue virus and Zika virus are closely related arboviruses with similar symptoms and distribution patterns throughout the tropics; however, dengue virus is currently causing large outbreaks in many areas of the world, whereas Zika virus cases have declined. Dengue virus has four distinct serotypes (1, 2, 3, and 4) and is endemic throughout the tropics and subtropics. Although transmission most commonly occurs from the bite of an infected Aedes spp mosquito, dengue virus can also be transmitted during childbirth and through blood and breast milk. Clinical manifestations range from the more common self-limited dengue fever to dengue hemorrhagic fever with shock syndrome, the most serious manifestation. The risk of progression to severe disease is much higher in secondary dengue infection than in primary infection.
Diagnostic tests for dengue virus include serology, NS1 antigen testing (not currently performed at ARUP Laboratories), and NAAT. The test of choice depends on when the patient presents in relation to symptom onset or last possible exposure to dengue virus. Additional testing for other arboviruses, particularly Zika virus, is recommended if an outbreak is occurring or has occurred.
Indications for Testing
Testing for dengue virus should be considered in patients with appropriate clinical manifestations (eg, fever, headache, hemorrhage, retro-orbital pain) and recent travel to or residence in an area with known dengue transmission.
Diagnostic Laboratory Testing
Nucleic Acid Amplification Testing
NAAT or NS1 antigen and IgM antibody testing are recommended during the first week of illness. NAAT is the preferred method for diagnosis because it can confirm infection and identify the specific virus, distinguishing between dengue and Zika viruses that often cocirculate, for example. A negative NAAT or NS1 result should be followed with an IgM antibody test. Refer to the CDC for more information about differentiating between dengue and Zika virus.
Detected | Not Detected |
---|---|
Indicative of current infection No further testing indicated |
Does not exclude dengue virus infection If test was performed during the first 5 days of illness, follow with dengue IgM serology |
PCR, polymerase chain reaction |
Serology
Although generally the preferred method of diagnosis, NAAT can only detect dengue virus infection for a short period of time. Therefore, serology is usually used to diagnose dengue virus infection. After 7 days of illness, only serology testing is recommended. In primary dengue infection, IgM antibodies are reliably detectable after 7 days from symptom onset and can remain detectable for 3 months or longer. Although IgM antibody testing is important for diagnosis, false-positive dengue enzyme-linked immunosorbent assay (ELISA) results can occur due to cross-reactivity in patients infected with other flaviviruses such as Zika virus, WNV, and SLE virus. Therefore, concurrent evaluation for other possible cocirculating arboviruses, such as Zika virus, should be considered, and PRNT is recommended. Detailed vaccine, travel, and possible exposure history is key for determining the need for additional laboratory testing to identify the infecting virus. A patient with a positive dengue IgM-only result (negative for NAAT and NS1) is considered to have presumptive dengue infection only if no other arboviruses are circulating in the areas where the patient was exposed.
Chikungunya Virus
Chikungunya virus is primarily transmitted to humans by Aedes spp mosquitos. Bloodborne transmission is possible, and rare in utero and intrapartum transmissions have been documented. Unlike individuals with dengue and other mosquito-borne arboviruses, most individuals infected with chikungunya virus become symptomatic. Acute onset of fever and polyarthralgia are the most common clinical symptoms associated with chikungunya virus. Joint pain can be severe and may be persistent or relapsing in the months following acute illness.
Indications for Testing
Testing for chikungunya should be considered in patients with acute onset of fever and polyarthralgia and recent travel to or residence in an area with known chikungunya virus transmission.
Diagnostic Laboratory Testing
Nucleic Acid Amplification Testing
NAAT testing is appropriate for all individuals suspected of having chikungunya virus who present in the early stages of disease. A positive result indicates current infection, and no further testing is indicated. A negative PCR result does not exclude infection and should be followed by virus-specific IgM serology.
Serology
IgM serology is appropriate for individuals with clinical suspicion for chikungunya virus who present later in the course of illness (beyond the first week of illness). Virus-specific chikungunya IgM antibodies are detectable toward the end of the first week of illness and remain detectable for 3-4 months after infection, whereas IgG antibodies to chikungunya virus remain detectable for years.
As with Zika virus, false-positive chikungunya virus IgM serology results may occur in patients with closely related infections or with recent vaccinations to related viruses. Therefore, concurrent evaluation for other possible cocirculating viruses, especially Zika and dengue, should be considered.
ARUP Laboratory Tests
Order when WNV is suspected and patient presents with encephalitis
Semi-Quantitative Enzyme-Linked Immunosorbent Assay
Order when WNV is suspected and patient does NOT present with encephalitis
Semi-Quantitative Enzyme-Linked Immunosorbent Assay
Typically not a first-line test
Can be used to confirm positive WNV antibodies test result or to clarify equivocal serologic test results
Qualitative Polymerase Chain Reaction
Not a preferred test
Semi-Quantitative Enzyme-Linked Immunosorbent Assay
Semi-Quantitative Enzyme-Linked Immunosorbent Assay
Semi-Quantitative Enzyme-Linked Immunosorbent Assay
Semi-Quantitative Enzyme-Linked Immunosorbent Assay
Order when arboviral illness is suspected and patient presents with encephalitis
Semi-Quantitative Indirect Fluorescent Antibody/Semi-Quantitative Enzyme-Linked Immunosorbent Assay
Order when arboviral illness is suspected and patient does NOT present with encephalitis
Semi-Quantitative Indirect Fluorescent Antibody/Semi-Quantitative Enzyme-Linked Immunosorbent Assay
Panel components: WNV, SLE, California encephalitis, EEE, and WEE viruses
Use to detect presence of Japanese encephalitis-related IgG and IgM antibodies in individuals with evidence of acute encephalitis syndrome who have recently traveled to or resided in an endemic country in Asia or the western Pacific
Patient’s travel history is necessary to aid in test interpretation
Cotesting for other flaviviruses (eg, dengue fever virus) is recommended due to coexistence and antibody formation interference in individuals previously infected with flavivirus
Semi-Quantitative Enzyme-Linked Immunosorbent Assay
Not recommended as a stand-alone test
Semi-Quantitative Enzyme-Linked Immunosorbent Assay
Semi-Quantitative Enzyme-Linked Immunosorbent Assay
Recommended test for patients presenting in the acute phase of infection (<14 days after symptom onset or exposure)
Paired serum and urine specimens are preferred
Qualitative Polymerase Chain Reaction
Qualitative Polymerase Chain Reaction
Recommended screening test for patients with symptoms for ≥14 days
Follow-up test for patients with negative serum and urine NAAT
Semi-Quantitative Enzyme-Linked Immunosorbent Assay
Use to detect and differentiate dengue subtypes 1-4
Qualitative Polymerase Chain Reaction
Can be used to diagnose dengue virus during acute phase of disease (>5 days after symptom onset)
In endemic regions, cotesting is recommended for other arthropod-borne viruses, including chikungunya and Zika, due to overlapping symptoms
Semi-Quantitative Enzyme-Linked Immunosorbent Assay
May aid in diagnosis of dengue virus when timing of infection is uncertain
In endemic regions, cotesting is recommended for other arthropod-borne viruses, including chikungunya and Zika, due to overlapping symptoms
Semi-Quantitative Enzyme-Linked Immunosorbent Assay
Aids in detecting past dengue infection
In endemic regions, cotesting is recommended for other arthropod-borne viruses, including chikungunya and Zika, due to overlapping symptoms
Semi-Quantitative Enzyme-Linked Immunosorbent Assay
Use to detect chikungunya virus
Qualitative Polymerase Chain Reaction
May aid in diagnosis of chikungunya viral infection during acute phase of disease (>5 days after symptom onset)
In endemic regions, cotesting is recommended for dengue and Zika viruses because clinical picture of these diseases is similar
Semi-Quantitative Enzyme-Linked Immunosorbent Assay
Semi-Quantitative Enzyme-Linked Immunosorbent Assay
Aids in detecting past chikungunya viral infection
In endemic regions, cotesting is recommended for dengue virus because clinical picture of these diseases is similar
Semi-Quantitative Enzyme-Linked Immunosorbent Assay
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Medical Experts
Slev

Panel components: WNV, SLE, California encephalitis, EEE, and WEE viruses