Zika Virus Prevention and Treatment Recommendations
Key Points
- There has been a substantial reduction in the risk of Zika virus (ZIKV) infection among Canadian travellers. Accordingly, CATMAT no longer routinely recommends that pregnant travellers avoid travel to areas where Zika is known or suspected to occur, or that special precautions to prevent sexual transmission while abroad or upon return are necessary.
	+ Some travellers, for example based on their values and preferences, might choose to follow recommendations for prevention of sexual or vertical (mother to fetus) transmission of ZIKV because Zika continues to present a low risk in many tropical and subtropical areas.
- Given the low risk of ZIKV infection, CATMAT recommends against routine testing of asymptomatic pregnant women. The poor positive predictive value, especially for screening serology tests, means a positive test has a high likelihood of being a false positive, which may have significant adverse consequences. The low population prevalence of infection means a negative test result is of negligible clinical utility.
- In a situation where testing is completed and ZIKV is confirmed CATMAT recommends:
	+ women should wait at least 2 months after their return from a risk area or onset of symptoms (whichever occurs later) before trying to conceive/having unprotected sex.
	+ men should wait at least 3 months after their return from a risk area or onset of symptoms (whichever occurs later) before trying to conceive with their partner or engaging in unprotected sex.
- In the case of a confirmed or symptomatic case with compatible symptoms of ZIKV infection in the male partner, CATMAT recommends that pregnant couples abstain from unprotected sex for the duration of the pregnancy.
- Travellers and health care providers should remain vigilant for emergent information related to ZIKV outbreaks. Should a significantly increased risk of ZIKV transmission be identified in a specific travel destination, pre- and post-travel advice should be modified accordingly. This would include consideration of avoiding travel to the outbreak area during pregnancy.
- Travel during pregnancy typically poses multiple health risks. The risk of ZIKV infection should be included in a broader discussion about infectious diseases and other potential complications when considering travel while pregnant.
Preamble
The Committee to Advise on Tropical Medicine and Travel (CATMAT) provides the Public Health Agency of Canada with ongoing and timely medical, scientific, and public health advice relating to tropical infectious disease and health risks associated with international travel. The Agency acknowledges that the advice and recommendations set out in this statement are based upon the best current available scientific knowledge and medical practices, and is disseminating this document for information purposes to both travellers and the medical community caring for travellers.
Persons administering or using drugs, vaccines, or other products should also be aware of the contents of the product monograph(s) or other similarly approved standards or instructions for use. Recommendations for use and other information set out herein may differ from that set out in the product monograph(s) or other similarly approved standards or instructions for use by the licensed manufacturer(s). Manufacturers have sought approval and provided evidence as to the safety and efficacy of their products only when used in accordance with the product monographs or other similarly approved standards or instructions for use.
Summary
There has been a substantial decrease in reported rates of Zika virus (ZIKV) related disease in countries (primarily in the Americas) that suffered from the outbreak that started in 2015 and a corresponding decline (>90%) in reports of ZIKV infection among travellers visiting these countries. Accordingly, CATMAT no longer routinely recommends that: pregnant travellers avoid travel to areas where Zika is known or suspected to occur, or that special precautions to prevent sexual transmission while abroad or upon return are necessary. Rather, CATMAT suggests that precautions for preventing ZIKV transmission be considered within the general context of the infectious and non-infectious risks travel may pose to a pregnant woman. 
Introduction
Zika virus (ZIKV) infection is caused by a flavivirus transmitted through the bite of an infected Aedes mosquito, mainly *Aedes aegypti*. Although infections in humans have been known to occur for some time, ZIKV emerged on the global stage when it caused major outbreaks in many tropical and subtropical areas of the world, especially in the Americas ,. During these outbreaks, a previously unrecognized pattern of Zika-related adverse outcomes emerged. Pre-eminent among these were complications of pregnancy including major congenital anomalies. First identified through spatial and temporal clustering of ZIKV activity with an increased incidence of congenital microcephaly  -   is now known that these can be manifest as a constellation of fetal developmental problems that are collectively referred to as a congenital Zika syndrome (CZS) -  The risk of CZS is substantial if infected while pregnant. For example, of 972 completed pregnancies enrolled in the US Zika Pregnancy Registry, 5% showed evidence of CZS (95% CI = 4%-7%). The greatest impact was observed when infection occurred during the first trimester (15% with CZS) . Similar results have been described for: affected US Territories, with a CZS rate of 5% among completed affected pregnancies  and for French territories in the Americas where 7% (39/555) of fetuses and infants were identified as having suffered defects possibly associated with ZIKV infection. In this last study, effects were more common if infection occurred during the first trimester, but were also seen if infection occurred in the second (3.6%) or third (5.3%) trimesters . Recently, data have been described for Zika associated harms before and after birth in a population of children born to infected mothers in US territories and freely associated states . From a population of 1,450 children with follow-up care, 6% suffered from a Zika-associated birth defect(s), 9% had a neurodevelopmental abnormality(ies) possibly associated with congenital infection, and 1% had both.
Although disease is usually relatively benign in adults, ZIKV infection can cause neurologic sequelae such as Guillain-Barre syndrome (GBS) , , . A case control study done in French Polynesia estimated a 0.24 in 1,000 risk for developing GBS in persons infected with ZIKV. This is comparable to the GBS risk of 0.25 to 0.65/1,000 observed following Campylobacter jejuni infection. There are also reports of acute disseminated encephalomyelitis (ADEM) following ZIKV infection .
Complicating the picture for ZIKV epidemiology and prevention, the virus can be transmitted sexually, with most reported cases being from an infected male to a female, although female to male and male to male transmission have also been reported . The risk of such transmission remains difficult to quantify, though a significant proportion of symptomatic men shed viral RNA in their semen. For example, 22/36 (61%) of men in a US-based study had detectable viral RNA in their semen when tested within 30 days of illness onset . This dropped to < 10% at 3 - 4 months after illness, and approximately 1% at 5-6 months after illness. Importantly, recovery of infectious ZIKV was rarer, occurring in 3/78 (4%) samples with all positives being obtained within 30 days of illness onset , though the possibility of false negatives in this cohort cannot be ruled out. Complementing these observations, the median serial interval between onset of symptoms in couples where sexual transmission occurred was 12 days (interquartile range: 10-14.5 days) . Overall, these data suggest that the period during which sexual transmission (from a male) is likely to occur is much shorter than the period (often 6 months) for which sexual precautions had been previously recommended.
The purposes of this statement are to review current knowledge related to ZIKV infection and to provide guidelines for health care providers on prevention and management of ZIKV disease.
Methods
This statement was developed by a working group of the Committee to Advise on Tropical Medicine and Travel (CATMAT). Members of the working group were from CATMAT, the Public Health Agency of Canada (the Agency) and the Society of Obstetricians and Gynaecologists of Canada. Each member was a volunteer, and none declared a relevant conflict of interest. The working group was responsible for assessing available literature, synthesis and analysis of the evidence, drafting recommendations, and writing. Secretariat support was provided throughout the process by the Public Health Agency of Canada. The final statement was approved by the full CATMAT committee.
This guideline complements existing CATMAT statements including the   and  .
This document is an update to the CATMAT Zika statement published in February 2018. The most important changes are related to: the overall reduction in ZIKV transmission in many areas previously affected by the outbreak, the consequent significant reduction in risk for travellers (see Table 1); and, the availability of a systematic review related to sexual transmission .
Epidemiology
ZIKV was first isolated from monkeys in Uganda in 1947. Soon after, in 1952, human infections were detected in Uganda and Tanzania ,. However, human infections were rarely reported until 2007, when a large outbreak of ZIKV disease occurred on the island of Yap (Micronesia) . Between 2013 and 2015, additional outbreaks occurred on islands and archipelagos in the Pacific region  -  . In 2014, local transmission in the Americas was reported on Easter Island . Subsequently, ZIKV caused a large outbreak in the Americas  as well as in other countries/regions. Since the outbreak peaked in 2016, the number of locally acquired reported cases in the majority of affected countries has decreased substantially ,, as has the number of travel-associated cases reported in Canada and the United States ,. For example, in the first 11 months of 2018, only 58 cases of travel-associated ZIKV infection had been reported in the continental United States , compared to almost 5,000 cases during the entirety of 2016 and 437 in 2017 . Similar decreases in travel-associated infections have been observed in Canada, with the number of cases reported dropping from 468 in 2016 to 74 in 2017, and then to 21 in 2018 and without a concomitant decrease in testing.

# Transmission
The mosquitoes associated with ZIKV tend to be more active during the day but can bite at night, with peak activity often occurring in the morning and later in the afternoon. In vertebrate hosts, the incubation period is usually 3 - 14 days , with blood viremia (the period when ZIKV is present in the blood) usually lasting for about 2 weeks  -  , but with the possibility of longer periods of viral detection in blood and other body fluids . Vertical transmission between mother and developing fetus presumably occurs during the viremic period ,. Other described routes of transmission include blood product transfusion  and sexual transmission after symptomatic infection  -  . There is a report of sexual transmission from a symptomatic infected woman to a sexual partner , and another report of sexual transmission from an asymptomatically infected male to their partner ,.
Viral RNA has been detected in various biological fluids for prolonged periods, but isolation of infectious virus has only been documented at substantially lower rates and for shorter time intervals. Viral RNA has been detected in urine up to 6 weeks after illness onset; however, the median time until the loss of viral genome detection was determined to be 8 days . For individuals with detectable viral RNA in their serum the median time for loss of detection was 14 days, a small percentage of patients were reverse transcriptase polymerase chain reaction (RT-PCR) positive for more than a month . In particular, pregnant women carrying congenitally-infected fetuses may be symptomatically viremic for a prolonged period, thus, Zika should be considered in pregnant women with compatible exposure history and extended duration of fever, despite the typically short viremic phase of flaviviruses, in general ,.
Based on a cohort of symptomatic men followed in the US, a significant proportion (61%) had detectable viral RNA in their semen in the first month after illness onset . This decreases to 1% or less by 6 months after symptom onset. Duration of shedding increased modestly with increasing age, and with certain symptoms. Interestingly, persistence of RNA in semen was most strongly and inversely associated with frequency of ejaculation. For example, men who reported ejaculating 4 times per week were estimated to clear RNA 3 weeks earlier than men who reported ejaculating once per week. This raises the possibility of a modifiable risk factor for prolonged shedding. In this same study, infectious virus was isolated from 3/19 (16%) samples obtained within 30 days after illness onset but in none of the 59 samples that were obtained later. The 3 men in which infectious virus was isolated had relatively high (>7.0 log10) ZIKV RNA copies per milliliter of semen in their first sample which declined in subsequent samples to 5.8 (log10 copies/ml of semen), 3.1 (log10 copies/ml of semen), and undetectable, respectively. These follow-up samples were obtained at 38, 59 and 76 days following illness onset and infectious ZIKV was not detected in the two men who still had detectable ZIKV RNA . Nevertheless, it remains theoretically possible that some of the PCR positive but culture negative cases were contagious. To date the longest period after symptom onset at which replication-competent virus has been detected in semen was 69 days; this was reported in a single case report of a vasectomized man . ZIKV RNA has also been detected at lower concentrations in men who have undergone vasectomy, but at rates similar to men who have not had this procedure. Although not studied extensively, the persistent shedding of ZIKA RNA in semen is considered to be similar amongst infected symptomatic and asymptomatic men based on case reports of sexual transmission and on data from asymptomatic blood donors , , .
Neutralizing antibodies for ZIKV are detectable after infection, and by extrapolation from other flaviviruses that induce a humoral antibody response, post-infection immunity is presumed to be lifelong . Among couples where such data is available (n=15) the median time between onset of sexual partners' symptoms (serial interval) was 12 days (interquartile range: 1014.5 days) while the maximum was 44 days . Based on RT-PCR data, the median duration of ZIKV positivity was 13.9 days (95% CI: 7.2-19.6) for any fluid in the female genital tract, with a maximum of 37 days. There were too few data available for analysis of viral culture specimens in female genital tract fluids .ZIKV RNA has been detected in breast milk; however, there have not been any unequivocally documented reports of transmission to infants through breastfeeding . At this time, the World Health Organization (WHO) considers that "the benefits of breastfeeding for the infant and mother outweigh any potential risk of Zika virus transmission through breast milk" . Until further evidence is available regarding transmission through breast milk, CATMAT suggests that potential benefits and harms of breastfeeding during acute ZIKV infection should be discussed with each patient and decisions be made on an individual basis.

# Clinical Manifestations
Approximately 20-25% (possibly as high as 50%)  of persons infected with ZIKV will manifest symptoms, including fever, myalgia, pruritis, eye pain, and maculopapular rash , , .
Early clinical manifestations are similar to other arboviral infections including dengue and chikungunya ,. Thus, the differential diagnosis of a febrile returned traveller will likely include these arboviral infections, other viral illnesses ,, as well as  .
Post-infectious neurologic complications, such as GBS, have been reported from many countries that were affected by the outbreak , , , . They include French Polynesia where a case-control study estimated that the odds of positive ZIKV serology was substantially greater in GBS cases compared to matched controls (OR 59.7; 95% CI 10.4 to ) . In the same study, and based on a ZIKV population seroprevalence of 0.66, the risk of GBS following ZIKV infection was estimated at approximately 0.25/ 1000. Other neurological manifestations have also been reported in association with ZIKV infection, e.g., acute myelitis, meningoencephalitis, acute disseminated encephalomyelitis, and reverse sensory polyneuropathy , , ,  suggesting that the neurological spectrum of sequelae associated with ZIKV is relatively broad.
Clinically relevant thrombocytopenia and subcutaneous hematomas have been reported in a small number of cases ,. Deaths from other causes have also been reported ,.
Over 30 countries  have reported CZS . Common manifestations include microcephaly, cerebral atrophy, abnormal cortical development, callosal hypoplasia, and diffuse subcortical calcifications ,. Ocular abnormalities and other congenital malformations such as arthrogryposis and hydrops fetalis have also been described  -  .
Reviews of the epidemiology of ZIKV, as well as the causal association of ZIKV and microcephaly have been published .
Risk to travellers
There are multiple lines of evidence to indicate that the risk of ZIKV infection for travellers has decreased significantly. First, the number of cases reported in the epidemic regions of the Americas has decreased ,. For US territories specifically, more than 36,000 cases (presumed to be locally acquired) were reported in 2016, compared to 652 (98% decrease) cases reported in 2017  and 116 cases reported (as of December 4) in 2018 (> 99% decrease) ,. Second, the number of travel-related cases reported in Canada and the United States has decreased by approximately two orders of magnitude since 2016. In Canada, the overall decrease from 2016 to 2018 has been > 95% (468 cases in 2016 to 21 cases in 2018). A similar decrease has also occurred in the continental United States  where 4,897 cases (travel-related) were reported in 2016, 437 in 2017 (91% reduction) and 34 cases reported (as of August 1) in 2018 (> 99% decrease); and, in the European Union where 2,121 cases were reported in 2016 and 198 cases (93% reduction) were reported in 2017 .
Set against the total travel volumes to countries affected by the outbreak, which exceeds 7 million annually for Canadians , the estimated risk of a traveller being confirmed with a ZIKV infection is currently very low, e.g., < 1 case/200,000 trips (14 cases in 2018, adjusted for surveillance period). While this is an underestimate of the true risk of infection, (the majority of infections are likely not reported), it nevertheless supports that ZIKV-infection among Canadian travellers has become a rare event. Accordingly, adverse outcomes associated with ZIKV-infection must be rare events in absolute terms, even with significant under-reporting. Indeed, from October 2015 - December 2018, a total of 4 sexually transmitted cases have been reported in Canada . During the same time period, 47 cases have been reported among pregnant women in Canada . From March 2017 to December 2018, CZS had been reported in  5 infants in Canada .

# Areas of risk
Transmission of ZIKV can occur in most areas of the world where *Aedes aegypti*, the principal vector, occurs. This means that there is the potential for transmission through much of the tropical and subtropical world and beyond. As described above, the risk of transmission to travellers is considered low.
Previously, CATMAT recommendations were linked to the WHO country classification scheme for ZIKV to determine areas of risk or potential risk. In July 2019, the WHO implemented a new approach for categorizing countries according to the presence or absence of current and historical reported ZIKV transmission. It should be noted that monitoring and reporting in some areas is suboptimal, and that this approach provides only an approximation of the areas of risk. Due to a general absence of laboratory and surveillance data from many countries/territories, it provides a conservative risk assessment (i.e. includes countries/territories where cases have been reported but also includes countries where there is a known historical risk but cases are not currently being reported).
Notwithstanding the above, travellers and health care providers should remain vigilant for emergent information related to ZIKV that might influence decision-making, for example evidence that indicates a significant outbreak is occurring in a travel destination. If this is the case, pre- and post-travel advice should be modified accordingly, including potentially recommending avoiding travel to the outbreak area during pregnancy. 
Prevention - Decision to travel to areas of risk

# All travellers
Health care providers should discuss with travellers what is known and what is not known about ZIKV to help their patients make an informed choice about travel and precautions. Factors to consider include:
- The low absolute risk for ZIKV infection and consequent impact on travellers. Travel health providers should keep themselves informed of any evolving risks for transmission in destination regions, as with other endemic and potentially epidemic infectious risks.
- The potential for ZIKV infection during pregnancy to have a severe impact on the fetus.
- The possibility of serious sequelae such as post-infection neurologic complications (e.g., GBS, ADEM).
- The potential for sexual transmission, which is particularly relevant to couples who are actively trying to conceive.
- The potential for co-morbidities to predispose to more serious outcomes (there is little specific evidence in this regard, though it is reasonable to expect such impacts).
- Patients' values and preferences (including risk perception and risk tolerance).
- The potential impacts of following the recommendation on a couple's reproductive plans.

# Pregnant women and women who are planning a pregnancy
CATMAT no longer routinely recommends that pregnant women and those planning a pregnancy avoid travel to areas where Zika is known or suspected to occur.
Depending on individual values and preferences, including risk tolerance, some pregnant women or those planning a pregnancy might nevertheless choose to minimize risk by not travelling to these areas.
For pregnant travellers, there are many health considerations in addition to Zika. For more information in this regard, see the CATMAT  .
Prevention of mosquito-borne transmission
CATMAT recommends that all travellers to areas of risk should be advised to adhere to recommendations for the use of personal protective measures (PPM) against mosquito bites (see below). Because the mosquitoes that transmit ZIKV often bite during daylight hours, PPM should be used through all hours of the day and night. In addition to ZIKV, PPM provide protection against other vector-associated diseases such as malaria, dengue, and chikungunya. Recommendations for PPM can be found in CATMAT's  .
Prevention of sexual transmission
ZIKV RNA has been detected in semen 6 or more months after symptomatic infection. However, infectious (culturable) virus appears to be much less persistent , and mostly documented only for periods shorter than a month. At this time, all reported cases of sexual transmission have occurred within 41 days after illness onset in the source male partner, and the large majority have occurred within 20 days . Sexual transmission should be prevented by abstinence from exposure to semen, and minimized by proper condom use.

# Asymptomatic traveller
CATMAT no longer recommends routine use of measures to prevent sexual transmission from asymptomatic travellers while in or after returning from areas where Zika is known or suspected to occur. This applies to all travellers, including pregnant ones. The proportion of all travellers returning from tropical and subtropical areas who have been asymptomatically infected with ZIKV is considered to be negligible. Nevertheless, depending on individual values and preferences, including risk tolerance and the potential impact of an infection (e.g., during a pregnancy), travellers might still choose to minimize risk by using condoms and/or restricting sexual activity, and this should be discussed with each patient.

# Symptomatic traveller

## Compatible symptoms
CATMAT recommends that those with symptoms compatible with ZIKV infection, and no alternate diagnosis to explain their symptoms should discuss the likelihood of ZIKV infection (which will be low in most cases) with their health care provider when making the decision on whether to apply measures to prevent sexual transmission. Diagnostic testing can be considered after discussion of the risks of both false negative and false positive results.

## Confirmed case
CATMAT recommends that those who have confirmed ZIKV infection should follow recommendations for preventing sexual transmission. Based on current information on the incubation period and duration of viremia, and the unclear duration of viral persistence in tissues, travellers who choose to apply these recommendations should do so as follows:
- Women should wait at least 2 months after their return from an affected area or onset of symptoms (whichever occurs later) before trying to conceive/having unprotected sex.
- Men should wait at least 3 months after their return from an affected area or onset of symptoms (whichever occurs later) before trying to conceive with their partner or engaging in unprotected sex. In some circumstances, for example based on risk tolerance, men might wish to delay trying to conceive for up to 6 months, which represents the outer theoretical limit of the potentially contagious period.
In the case of a confirmed or symptomatic case with compatible symptoms of ZIKV infection in the male partner, CATMAT recommends that pregnant couples should abstain from unprotected sex for the duration of the pregnancy.

# Role of laboratory testing in transmission prevention or monitoring of pregnant women
Laboratory testing for ZIKV infection is fully described below. In theory, based on information from other similar viral infections, the absence of ZIKV-specific antibodies 2 weeks or more after the last possible exposure implies that the individual has never been infected, and is not contagious to sexual partners or to the fetus. Nevertheless, there are major limitations in the usefulness of ZIKV screening tests; neither serology nor molecular testing can ever be 100% sensitive. The absence of ZIKV RNA in a semen sample might indicate absence of contagiousness at that time, but intermittent shedding is possible and there are no data to support the use of this approach to defining a risk-free sexual contact. Of more concern, the current very low prevalence of ZIKV infection means that an increase in the false positive rate is to be expected, especially for serology when preliminary testing is carried out. As well, screening enzyme-linked immunosorbent assay (ELISA)s may detect cross reacting antibodies to related flaviviruses such as dengue viruses. Furthermore, in the context of previous travel, the presence of antibody (in the absence of detectable viral RNA) may indicate a remote / previous infection. These false positive results could result in significant anxiety, unnecessary delays in conception, and potentially inappropriate decisions regarding termination of pregnancy. Currently, testing in Canada is focused on symptomatic individuals coming from potentially endemic areas, and pregnant women. It has been observed that testing of asymptomatic individuals (men or non-pregnant women) has a very low yield of true positives. In addition to the poor specificity of a positive test (especially serologic screening tests), the very low population prevalence of infection means that a negative result is of negligible predictive utility. Therefore, routine testing of asymptomatic travellers and their partners is not recommended.
Laboratory Diagnosis
Molecular testing using RT-PCR and screening serology procedures are conducted by some provincial laboratories in Canada. The National Microbiology Laboratory (NML) provides RT-PCR, IgM and IgG ELISA serological testing support to provinces and territories, along with confirmatory plaque reduction neutralization test (PRNT) diagnostics. Sensitivity and specificity of molecular tests are presumed to be high during the initial few days of illness, since ZIKV appears to circulate in the blood for several days after onset of symptoms . ZIKV RNA may be present in urine for a week or more after symptom onset , . Information about NML's guidelines and testing recommendations are available on the .
Serologic testing for Zika has become increasingly problematic for several reasons. Specificity has always been relatively low due to cross reactivity with other related flaviviruses, and positive predictive value has fallen as prevalence has decreased in recent years. At the start of the epidemic, the presence of antibodies suggested recent exposure. At the present time, however, seropositivity may often represent remote exposure. In addition, data shows that IgM also may remain positive for over 2 years, thus limiting specificity for recent infection. Sensitivity and negative predictive value remain high when serology is performed at appropriate time intervals. 
At the NML, screening serology is currently performed using a US CDC based in-house IgM ELISA and a commercial IgG ELISA. Serum samples identified as being positive by one or both of the ELISAs are then tested by a confirmatory ZIKV PRNT to determine if viral specific antibodies are present . IgM antibodies appear approximately 5 to 6 days after onset of symptoms, may persist for long periods as mentioned above, but also may wane as early as 3 months after exposure, affecting sensitivity as well as specificity. IgG antibodies are usually detected a few days later, with maximal sensitivity about 2 months after infection . For the acutely unwell patient with less than 10 days of symptoms, RT-PCR should be ordered. Serology can be requested to maximize sensitivity, but poor specificity typically limits the usefulness of this test. For the convalescent patient with symptom onset over 10 days ago, only serology should be requested, although again specificity for recent infection is a problem. Serum samples collected from exposed individuals several months after returning from ZIKV endemic areas may no longer be positive for IgM antibodies, however, IgG antibodies probably persist for years. Appropriate diagnostic specimens for RT-PCR testing include plasma/serum, urine, cerebrospinal fluid (CSF), amniotic fluid and placental tissue. Serology is usually only performed on serum; however, viral antibodies may be detected in CSF in some cases of neurological disease. 
As stated previously ZIKV is a member of the Flaviviridae family, and serologic tests, including the IgM ELISA, may be cross-reactive with other flaviviruses such as dengue, West Nile, and Yellow Fever (including vaccine recipients) . Confirmation of ZIKV exposures therefore depends upon the amplification of viral RNA by RT-PCR, or by confirmatory PRNT serologic testing. Confirmatory testing generally requires neutralizing IgG production, which may appear later than IgM. The specificity of the IgM or IgG ELISAs is limited particularly during secondary flavivirus infections. Patients whose serum samples are IgM positive and have ZIKV-specific antibodies confirmed through PRNT are confirmed cases of viral infection. As previously noted individuals whose serum samples are IgG positive but negative for IgM and have PRNT documented ZIKV specific antibodies were most likely exposed several months before their serum sample(s) was collected, but estimating the timing of infection is usually difficult. Despite repeat testing, serological results may remain equivocal particularly in cases where the individual was previously infected with a related flavivirus. For secondary exposures an IgM positive result usually implies that a recent flavivirus infection is likely, but it may be unclear if this infection was due to Zika virus or a related virus such as dengue. This is because individuals previously infected with or vaccinated against non-Zika flaviviruses may also exhibit cross reactivity in PRNT tests making them difficult to interpret. It is also recommended for equivocal cases that acute and convalescent sera be collected 2 - 3 weeks apart to increase the likelihood of documenting a seroconversion or a diagnostic increase (four-fold or greater) in ZIKV specific neutralizing antibodies, which suggests recent infection when detected. As discussed above, a negative serology at least 2 weeks after the last possible exposure is presumed to indicate the absence of recent infection, although the precise sensitivity of currently available tests has not been determined. If a negative serological test result is being considered in order to support the discontinuation of transmission precautions, a careful risk assessment is required and collection of a second serum sample should also be considered at a later date to definitively rule out a ZIKV exposure.
In regions where the ZIKV outbreak has been extensive, positive serology (IgM, IgG and PRNT) may increasingly represent remote, rather than recent infection. As mentioned above, the declining incidence of transmission in most areas of the world will increase the false positive rates particularly for relatively non-specific tests such as IgM. Therefore, confirmation of recent infection, especially during pregnancy and for those with exposure in the past, now rests primarily on PCR.
PCR for ZIKV can be performed on amniotic fluid (when amniocentesis is technically feasible) to confirm infection of the fetus. At this time, the risk of adverse outcomes of pregnancy if the fetus is infected with ZIKV is at least 8% of adverse outcomes , so the risk of the procedure must be weighed against the clinical utility of this test result. A negative PCR result likely means that the fetus is not currently infected, but would not eliminate the possibility of previous infection. It is not known when ZIKV RNA would be expected to appear in amniotic fluid after infection, or how long it is likely to be detectable. There is some evidence that viral RNA may persist in amniotic fluid for months .
For postnatal diagnosis of congenital infection, PCR for ZIKV can be performed on placental tissue, umbilical cord blood or infant blood, and CSF for confirmation of congenital infection. It is possible, however, that infants or fetuses infected weeks prior to specimen sampling will no longer have detectable viral RNA.
Screening and Management

# Evaluation of non-pregnant travellers returning from countries with demonstrated or potential autochthonous Zika transmission
Testing for ZIKV infection using PCR should be considered in the diagnosis of any ill traveller with compatible epidemiologic and clinical history, when symptom onset is within 3 days after arrival in, to 14 days after departing from an area of risk as identified by the WHO. Testing for other similar viral infections and for malaria should also be done as appropriate.
Given the low incidence of infection in most regions, most testing should be limited to molecular techniques, performed within approximately 10 days of the onset of symptoms. It may often be appropriate to perform molecular tests for other similar arboviral infections on the same specimen, with the understanding that sensitivity wanes for all such assays as time elapses beyond the febrile period. For the convalescent patient later than 10 days after the onset of symptoms, there is currently no accurate diagnostic test available for Zika virus. Serology is no longer routinely recommended, due to its lack of specificity for diagnosing recent infection. Paired acute and convalescent serology may be informative, but PCR on the acute specimen is faster and easier to interpret..
Serologic testing could be considered in exceptional circumstances for male returned travellers from areas of risk whose clinically compatible illness has resolved, and are at least 2 weeks post exposure, and when it is impossible or dangerous to delay attempts at conception, in order to assess for potential contagiousness to sexual partners. It is always considered safer to delay conception until the period of potential viral shedding has passed, rather than depend on serologic testing. The high probability of false positive results must be considered and discussed with the patient, prior to testing. 
Serological testing of male individuals with a history of travel to an area of risk, but no history of related symptoms is not recommended, given the extremely low risk of infection and high risk of false positive serology. 
A negative test result for a symptomatic or asymptomatic patient, whether from an initial or follow-up serology (e.g., if the first test is equivocal) obtained over 2 weeks post potential exposure indicates that a recent ZIKV infection is very unlikely. However, the risk of false positive results, the time required for confirmatory serologic testing, and the relatively short period during which transmission prevention measures might be required all combine to limit the usefulness of such testing . The decision to test should be made in consultation with a health care provider, and should be set against the broader context of the likelihood of infection and patient values and preferences. Testing would only be appropriate when transmission prevention measures are planned during the testing period, which may take several weeks.
The potential for viremia or transmission following a second exposure is unknown. Given that neurologic disorders like GBS have occurred following ZIKV infection, returning travellers should be counselled to report any neurologic symptoms to their doctor. In the event of the diagnosis of GBS or other unusual neurologic syndrome, a travel history for the patient and any male sexual partners should be elicited. If ZIKV infection is thought to be potentially associated with the illness, a specialist should be consulted.
Evaluation in the context of pregnancy

# Evaluation of pregnant women with a travel history to an area of risk
Health care providers should take a travel history from their pregnant patients including relevant information related to the travel history of their partner(s). Any patient who indicates that they or their partner have recently travelled to areas of transmission should be further evaluated.
Screening of asymptomatic pregnant women with possible exposure during pregnancy or during the peri-conception period should be discussed on a case-by-case basis between the woman and her health care provider. The declining incidence of Zika transmission in most areas of the world means that infection rates in this population will be extremely low, and significantly lower than in a population with symptoms compatible with infection. The risk of false positive laboratory results, particularly for serology, would be correspondingly elevated in an asymptomatic population. False positive diagnoses would have important implications for adverse events related to unwarranted additional testing and anxiety, as well as resource utilization. The decision whether to screen should take into account intensity of the potential exposure, the use of prevention measures, the likelihood of remote infection, and the transmission trends at the location of potential exposure. In most cases, screening is NOT recommended. Exceptions could be considered when the risk of exposure is particularly high, and the psychological benefit of a negative result clearly outweighs the harms which could arise from a false positive result. Screening would be done as for the symptomatic pregnancy patient, see below. The decision to test should include consideration of how the results of the screening tests would be used to inform subsequent decisions. The likelihood of false negative, and especially false positive results in the absence of a recent infection always require careful discussion with the patient. Diagnosis and identification of poor fetal outcomes will allow for appropriate counselling. 
Pregnant women and their partners may be justifiably concerned about the risk of ZIKV infection to their fetus and may want to receive counselling to decide the best course of action, including the question of termination. The risk of vertical infection (with clinical sequelae) in the setting of symptomatic or asymptomatic maternal infection appears highest in the first trimester . However severe sequelae have been reported after infection at all stages of pregnancy . This uncertainty makes pregnancy counselling a difficult prospect. Regardless, discussion and informed decision making regarding options for management of ZIKV infection in pregnancy (much like any other congenital infection or congenital anomaly) requires thorough consultation with a Maternal Fetal Medicine Specialist or another specialist familiar with reproductive infectious diseases.

# Evaluation of pregnant women with symptoms compatible with ZIKV infection
Testing should be offered to pregnant women with acute signs and symptoms compatible with ZIKV. Given the reports of longer periods of viremia in some pregnant women, for the patient with symptoms during the preceding 12 weeks, RT-PCR (on blood and urine) is the preferred testing modality. Serology is not recommended for routine testing and should only be requested very judiciously as it is not appropriate in most cases. A negative result has a high negative predictive value and serology at least 2 weeks after the last potential exposure provides reassurance if negative. However, the poor specificity must be clearly discussed and appreciated by the patient, and a clear and logical approach to a positive result should be established before ordering the test. The potential reassurance of a negative test must be weighed against the potentially important harms and likelihood of a false positive. Given the current epidemiology, it is reasonable to assume that for most women, a positive serology will NOT indicate exposure during the pregnancy. Exceptions would include the investigation of suspected congenital Zika syndrome. For the convalescent patient with symptom onset over 12 weeks ago, RT\_PCR will be of minimal value. Prolonged fever may be associated with persistent viremia and PCR testing is indicated in the investigation of these cases. Repeated ultrasound monitoring is indicated, unless the woman is found to be negative on appropriate laboratory testing, including negative serology at least 2 weeks after the last possible exposure. A woman whose fetus is suspected of having a congenital anomaly should also be offered testing if she or her partner has travelled to any location where ZIKV transmission may be occurring   even at a low level.
Although measurements of head circumference and biparietal diameter may occur as early as 15 weeks, there is no defined gestational age by which microcephaly and other intracranial abnormalities can be ruled out. Serial monitoring by ultrasound with close attention to measurement trends over time is recommended. It is possible that changes in intracranial anatomy may not be elucidated until well into the third trimester, or later.
Counselling recommendations are as in the section on the asymptomatic woman, above.

# Evaluation of the fetus among pregnant women diagnosed with ZIKV infection
Serial ultrasounds (every 3 - 4 weeks) are recommended in pregnant women with confirmed or suspected (if testing results are pending or equivocal) ZIKV infection in pregnancy. Should CNS calcifications or fetal microcephaly be noted at ultrasonography of an asymptomatic pregnant returned traveller, then specific ZIKV testing of the fetus (e.g., amniocentesis), in addition to other investigations to elucidate alternate aetiologies, should be considered to help define the likely cause of the anomaly.

# Evaluation of the infant born to a woman diagnosed with ZIKV infection or with suspected congenital ZIKV infection or CZS
Infants born to women with confirmed or suspected ZIKV infection in pregnancy, or those with unexplained microcephaly, intracranial calcifications, ventriculomegaly or major structural central nervous system abnormalities or other symptoms of congenital ZIKV infection in whom the mother had potential exposure to the virus, should be tested. This testing should include serology, PCR of serum (umbilical cord or infant sample), and PCR of placenta; if CSF is sampled, this can also be sent for PCR and serology. Management is evolving and infants with suspected or confirmed CZS should also undergo further work-up including: routine lab tests (CBC and liver enzymes), head ultrasound, ophthalmologic examination, and hearing evaluation as outlined in current guidelines by the Canadian Paediatric Society (CPS) . Care should be taken to ensure a thorough work up for other important and treatable causes of congenital infections, such as cytomegalovirus and *Toxoplasma*. Infants with confirmed CZS should have neurodevelopmental monitoring throughout infancy to assess the potential for long-term sequelae.
Infants born to women with symptoms of active ZIKV infection around the time of delivery are at risk for perinatal transmission of the disease. In the limited number of reported cases to date, perinatally infected infants have exhibited either no or mild symptoms and laboratory findings (rash, thrombocytopenia) . Regardless, such infants should be monitored closely given the unclear spectrum of potential illness in this emerging infection. Testing with serology and serum PCR during acute illness is recommended. Long term follow-up with a pediatric infectious disease specialist is suggested.
Treatment
Currently there is no specific therapy for the treatment of ZIKV infection. Treatment is supportive with antipyretics (acetaminophen in pregnancy), hydration and rest. Aspirin and other non-steroidal anti-inflammatory drugs (NSAIDs) should be avoided until dengue can be ruled out to reduce the risk of hemorrhage . Symptomatic disease typically lasts for up to 7 days. Urgent medical care is recommended for any symptoms associated with GBS, and treating health care providers should be made aware of recent travel to area with ZIKV circulation and/or symptoms of ZIKV infection.
If ZIKV infection is confirmed in the setting of pregnancy, referral to a Maternal Fetal Medicine Specialist or specialist familiar with Reproductive Infectious Diseases should be made. If microcephaly, intracranial calcifications or other abnormalities are identified, appropriate counselling by a Neonatologist and Pediatric Infectious Diseases Specialist on potential neurodevelopmental outcome should be offered to parents.
Summary of Recommendations for ZIKV
Discussion should occur around decision to travel, based on individual values and preferences, including risk of ZIKV infection. Some pregnant women or those planning a pregnancy may choose to minimize risk by:- not travelling to these areas or
- postponing travel until after pregnancy or
- postponing pregnancy until after travel.
If ZIKV infection is confirmed, CATMAT recommends:- women wait at least 2 months after their return from these areas or onset of symptoms (whichever occurs later) before trying to conceive and/or having unprotected sex;
- male partners wait at least 3 months after their return from these areas or onset of symptoms (whichever occurs later) before trying to conceive and/or having unprotected sex.
In the case of a confirmed or clinically compatible ZIKV infection in the male partner, couples planning a pregnancy might wish to delay conception for up to 6 months, depending on personal risk tolerance. The 6 months represents the outer theoretical limit of the potentially contagious period for a male partner.
As designated by the WHO