Detection of DENV-2 and ZIKV coinfection in southeastern Brazil by serum and urine testing.

PURPOSE: Aedes aegypti mosquito-borne diseases have a significant impact on public health in Brazil. In this study, we investigated the presence of the Zika virus (ZIKV) and dengue virus (DENV) in serum and urine samples from symptomatic participants who attended an Emergency Care Unit located in a city in the northwestern region of São Paulo between February 2018 and April 2019. METHODS: Serum and urine samples were collected from participants suspected of having arbovirus infection. After the extraction of viral RNA, viral detection was performed by real-time quantitative reverse transcription polymerase chain reaction (RT-qPCR) (One-Step RT-qPCR). RESULTS: A total of 305 participants participated in this study. A total of 283 blood and 270 urine samples were collected. Of 305 patients, 36.4% (111/305) were positive for ZIKV, 43.3% (132/305) for DENV2, and 0.3% (1/305) for DENV1. Coinfection with ZIKV/DENV2 was observed in 13.1% of participants. If only serum samples were used, ZIKV detection would have decreased to 23.3% (71/305). Of all the participants included in the study, only one was suspected of having ZIKV infection based on clinical diagnosis, and the remaining participants were suspected of having DENV. CONCLUSION: By testing serum and urine samples, we increased the detection of both viruses and detected considerable levels of ZIKV and DENV-2 coinfection when compared to other studies. Additionally, we detected an unnoticed ZIKV outbreak in the city. These findings highlight the importance of the molecular diagnosis of arboviruses to aid public health surveillance and management strategies.

Detection of Zika virus in urine from randomly tested individuals in Mirassol, Brazil.

PURPOSE: Studies show that around 80% of Zika virus (ZIKV) infections are asymptomatic. The present study tested urine samples from volunteers, unsuspected of arboviral infection, which attended an emergency care unit (ECU) in Mirassol, Brazil, from March 2018 to April 2019. METHODS: The volunteers were divided into two groups. The first group was composed of outpatients who were not suspected to have an arbovirus infection. This first group was subdivided into two subgroups: outpatients with and without arbovirus-like symptoms. The second group consisted of companions of outpatients treated at the ECU. The second group was also subdivided into two subgroups: totally asymptomatic individuals and those who had arbovirus-like symptoms. RNA was extracted from urine samples, followed by RT-qPCR for ZIKV. RESULTS: We found that 11% (79/697) of the samples tested positive for ZIKV-RNA. Among the ZIKV-RNA-positive individuals, 16.5% (13/79) were companions, of which 61.5% (8/13) were totally asymptomatic and 38.5% (5/13) reported symptoms that could be suggestive of arbovirus infection. In addition, 83.5% (66/79) of the ZIKV-RNA-positive individuals were outpatients without a clinical diagnosis of arbovirus. Of these undiagnosed ZIKV-RNA-positive outpatients, 47% (31/66) had no arbovirus-related symptoms. CONCLUSION: Our study shows the effectiveness of urine as a non-invasive sample to detect the incidence of ZIKV infection. We also highlight the importance of ZIKV molecular diagnosis to aid public health surveillance and prevention of congenital Zika syndrome and other ZIKV-associated diseases.

Enteric viruses circulating in undiagnosed central nervous system infections at tertiary hospital in São José do Rio Preto, São Paulo, Brazil.

Enterovirus (EV) is commonly associated with central nervous system (CNS) syndromes. Recently, gastroenteric viruses, including rotavirus (RVA), human astrovirus (HAstV), and norovirus (NoV), have also been associated with CNS neurological disorders. The aim of the present study was to investigate the presence of EV, RVA, HAst, and NoV associated to CNS infections with undiagnosed etiology in Northwest region of São Paulo State, Brazil, and to conduct the molecular characterization of the positive samples detected. A total of 288 cerebrospinal fluid samples collected from July to December 2017 were tested for EV and NoV by quantitative real-time polymerase chain reaction (RT-qPCR), HAstV by conventional RT-PCR, and RVA by enzyme-linked immunosorbent assay. Positive-EV samples were inoculated in cells lines, amplified by RT-PCR and sequenced. RVA, NoV, and HAstV were not detected. EV infection was detected in 5.5% (16/288), and five samples successful genotyped: echovirus 3 (E3) (1/5), coxsackie virus A6 (CVA6) (1/5), and coxsackie virus B4 (CVB4) (3/5). Meningitis was the main syndrome observed (12/16; 75%). CVA6, CVB4, and E3 were identified associated with aseptic meningitis. Reports of CVA6 associated with aseptic meningitis are rare, E3 had not been previously reported in Brazil, and epidemiological data on CVB4 in the country is virtually unknown. The present investigation illustrates the circulation of diverse EV types in a small regional sample set and in a short period of time, highlighting the importance of an active EV surveillance system in CNS infections. Enhanced understanding of undiagnosed CNS infections will assist in public health and health care planning.

Genomic detection of a virus lineage replacement event of dengue virus serotype 2 in Brazil, 2019.

BACKGROUND Despite efforts to mitigate the impact of dengue virus (DENV) epidemics, the virus remains a public health problem in tropical and subtropical regions around the world. Most DENV cases in the Americas between January and July 2019 were reported in Brazil. São Paulo State in the southeast of Brazil has reported nearly half of all DENV infections in the country. OBJECTIVES To understand the origin and dynamics of the 2019 DENV outbreak. METHODS Here using portable nanopore sequencing we generated20 new DENV genome sequences from viremic patients with suspected dengue infection residing in two of the most-affected municipalities of São Paulo State, Araraquara and São José do Rio Preto. We conducted a comprehensive phylogenetic analysis with 1,630 global DENV strains to better understand the evolutionary history of the DENV lineages that currently circulate in the region. FINDINGS The new outbreak strains were classified as DENV2 genotype III (American/Asian genotype). Our analysis shows that the 2019 outbreak is the result of a novel DENV lineage that was recently introduced to Brazil from the Caribbean region. Dating phylogeographic analysis suggests that DENV2-III BR-4 was introduced to Brazil in or around early 2014, possibly from the Caribbean region. MAIN CONCLUSIONS Our study describes the early detection of a newly introduced and rapidly-expanding DENV2 virus lineage in Brazil.

Seroprevalence for dengue virus in a hyperendemic area and associated socioeconomic and demographic factors using a cross-sectional design and a geostatistical approach, state of São Paulo, Brazil.

BACKGROUND: São José do Rio Preto is one of the cities of the state of São Paulo, Brazil, that is hyperendemic for dengue, with the presence of the four dengue serotypes. OBJECTIVES: to calculate dengue seroprevalence in a neighbourhood of São José do Rio Preto and identify if socioeconomic and demographic covariates are associated with dengue seropositivity. METHODS: A cohort study to evaluate dengue seroprevalence and incidence and associated factors on people aged 10 years or older, was assembled in Vila Toninho neighbourhood, São José do Rio Preto. The participant enrolment occurred from October 2015 to March 2016 (the first wave of the cohort study), when blood samples were collected for serological test (ELISA IgG anti-DENV) and questionnaires were administrated on socio-demographic variables. We evaluated the data collected in this first wave using a cross-sectional design. We considered seropositive the participants that were positive in the serological test (seronegative otherwise). We modelled the seroprevalence with a logistic regression in a geostatistical approach. The Bayesian inference was made using integrated nested Laplace approximations (INLA) coupled with the Stochastic Partial Differential Equation method (SPDE). RESULTS: We found 986 seropositive individuals for DENV in 1322 individuals surveyed in the study area in the first wave of the cohort study, corresponding to a seroprevalence of 74.6% (95%CI: 72.2-76.9). Between the population that said never had dengue fever, 68.4% (566/828) were dengue seropositive. Older people, non-white and living in a house (instead of in an apartment), were positively associated with dengue seropositivity. We adjusted for the other socioeconomic and demographic covariates, and accounted for residual spatial dependence between observations, which was found to present up to 800 m. CONCLUSIONS: Only one in four people aged 10 years or older did not have contact with any of the serotypes of dengue virus in Vila Toninho neighbourhood in São José do Rio Preto. Age, race and type of house were associated with the occurrence of the disease. The use of INLA in a geostatistical approach in a Bayesian context allowed us to take into account the spatial dependence between the observations and identify the associated covariates to dengue seroprevalence.

Viral Load and Cytokine Response Profile Does Not Support Antibody-Dependent Enhancement in Dengue-Primed Zika Virus-Infected Patients.

Background: The pathogenesis of severe dengue disease involves immune components as biomarkers. The mechanism by which some dengue virus (DENV)-infected individuals progress to severe disease is poorly understood. Most studies on the pathogenesis of severe dengue disease focus on the process of antibody-dependent enhancement (ADE) as a primary risk factor. With the circulation of Zika virus (ZIKV) in DENV-endemic areas, many people infected by ZIKV were likely exposed to DENV. The influence of such exposure on Zika disease outcomes remains unknown. Methods: We investigated whether patients previously exposed to DENV exhibited higher viremia when exposed to a subsequent, heterologous dengue or Zika infection than those patients not previously exposed to dengue. We measured viral loads and cytokine profile during patients' acute infections. Results: Neither dengue nor Zika viremia was higher in patients with prior DENV infection, although the power to detect such a difference was only adequate in the ZIKV analysis. Of the 10 cytokines measured, only 1 significant difference was detected: Levels of interleukin 1ß (IL-1ß) were lower in dengue-infected patients who had experienced a previous dengue infection than patients infected with dengue for the first time. However, power to detect differences between groups was low. In Zika-infected patients, levels of IL-1ß showed a significant, positive correlation with viral load. Conclusions: No signs of ADE were observed in vivo in patients with acute ZIKV infection who had prior exposure to DENV.

Long-Term Viruria in Zika Virus-Infected Pregnant Women, Brazil, 2016.

During the 2016 Zika virus outbreak in Brazil, we detected Zika virus RNA in urine samples collected from Zika virus-positive pregnant women during different stages of pregnancy. Women had positive and negative intervals of viruria; 3 newborns had adverse outcomes. Further research is needed to clarify the relationship between viruria and outcomes for newborns.

Fetal Infection by Zika Virus in the Third Trimester: Report of 2 Cases.

Zika virus (ZIKV) infection acquired during pregnancy is associated with congenital microcephaly. We describe 2 cases of ZIKV infection in women in their 36th week of pregnancy whose fetuses had preserved head circumference at birth and findings of subependymal cysts and lenticulostriate vasculopathy in postnatal imaging. These represent the first signs of congenital brain injury acquired due to ZIKV in the third trimester.

First genome sequence of St. Louis encephalitis virus (SLEV) isolated from a human in Brazil.

St. Louis encephalitis virus (SLEV), a member of the family Flaviviridae, genus Flavivirus, is a causative agent of encephalitis in the Americas. In Brazil, sporadic cases of SLEV infection have been reported since 1953, but the first outbreak of SLEV in Brazil was identified only in 2007, concomitant with an outbreak of dengue virus (DENV) serotype 3. This finding, along with other reports, indicates that SLEV circulation in Brazil is largely unknown, and there may be epidemiological implications of the co-circulation of SLEV, DENV and other flaviviruses in Brazil. Here, we describe the first complete genome sequence of an SLEV strain isolated from a human patient in Brazil, strain BeH 355964. Phylogenetic analysis was performed to determine the genotype of BeH 355964 using the full-length genome and envelope (E) gene sequences separately. Both analyses showed that BeH 355964 could be classified as genotype V. Although the number of single gene sequences available is greater (such as for the E gene), the phylogenetic tree based on the complete genome sequence was better supported and provided further information about the virus.

Cryptic circulation of chikungunya virus in São Jose do Rio Preto, Brazil, 2015-2019.

BACKGROUND: Chikungunya virus (CHIKV) has spread across Brazil with varying incidence rates depending on the affected areas. Due to cocirculation of arboviruses and overlapping disease symptoms, CHIKV infection may be underdiagnosed. To understand the lack of CHIKV epidemics in São José do Rio Preto (SJdRP), São Paulo (SP), Brazil, we evaluated viral circulation by investigating anti-CHIKV IgG seroconversion in a prospective study of asymptomatic individuals and detecting anti-CHIKV IgM in individuals suspected of dengue infection, as well as CHIKV presence in Aedes mosquitoes. The opportunity to assess two different groups (symptomatic and asymptomatic) exposed at the same geographic region aimed to broaden the possibility of identifying the viral circulation, which had been previously considered absent. METHODOLOGY/PRINCIPAL FINDINGS: Based on a prospective population study model and demographic characteristics (sex and age), we analyzed the anti-CHIKV IgG seroconversion rate in 341 subjects by ELISA over four years. The seroprevalence increased from 0.35% in the first year to 2.3% after 3 years of follow-up. Additionally, we investigated 497 samples from a blood panel collected from dengue-suspected individuals during the 2019 dengue outbreak in SJdRP. In total, 4.4% were positive for anti-CHIKV IgM, and 8.6% were positive for IgG. To exclude alphavirus cross-reactivity, we evaluated the presence of anti-Mayaro virus (MAYV) IgG by ELISA, and the positivity rate was 0.3% in the population study and 0.8% in the blood panel samples. In CHIKV and MAYV plaque reduction neutralization tests (PRNTs), the positivity rate for CHIKV-neutralizing antibodies in these ELISA-positive samples was 46.7%, while no MAYV-neutralizing antibodies were detected. Genomic sequencing and phylogenetic analysis revealed CHIKV genotype ECSA in São José do Rio Preto, SP. Finally, mosquitoes collected to complement human surveillance revealed CHIKV positivity of 2.76% of A. aegypti and 9.09% of A. albopictus (although it was far less abundant than A. aegypti) by RT-qPCR. CONCLUSIONS/SIGNIFICANCE: Our data suggest cryptic CHIKV circulation in SJdRP detected by continual active surveillance. These low levels, but increasing, of viral circulation highlight the possibility of CHIKV outbreaks, as there is a large naïve population. Improved knowledge of the epidemiological situation might aid in outbreaks prevention.

Why Did ZIKV Perinatal Outcomes Differ in Distinct Regions of Brazil? An Exploratory Study of Two Cohorts.

The Zika virus (ZIKV) epidemic in Brazil occurred in regions where dengue viruses (DENV) are historically endemic. We investigated the differences in adverse pregnancy/infant outcomes in two cohorts comprising 114 pregnant women with PCR-confirmed ZIKV infection in Rio de Janeiro, Southeastern Brazil (n = 50) and Manaus, in the north region of the country (n = 64). Prior exposure to DENV was evaluated through plaque reduction neutralizing antibody assays (PRNT 80) and DENV IgG serologies. Potential associations between pregnancy outcomes and Zika attack rates in the two cities were explored. Overall, 31 women (27%) had adverse pregnancy/infant outcomes, 27 in Rio (54%) and 4 in Manaus (6%), p < 0.001. This included 4 pregnancy losses (13%) and 27 infants with abnormalities at birth (24%). A total of 93 women (82%) had evidence of prior DENV exposure, 45 in Rio (90%) and 48 in Manaus (75%). Zika attack rates differed; the rate in Rio was 10.28 cases/10,000 and in Manaus, 0.6 cases/10,000, p < 0.001. Only Zika attack rates (Odds Ratio: 17.6, 95% Confidence Interval 5.6-55.9, p < 0.001) and infection in the first trimester of pregnancy (OR: 4.26, 95% CI 1.4-12.9, p = 0.011) were associated with adverse pregnancy and infant outcomes. Pre-existing immunity to DENV was not associated with outcomes (normal or abnormal) in patients with ZIKV infection during pregnancy.

Prevalence of Measles Antibodies in São José do Rio Preto, São Paulo, Brazil: A serological survey model.

Measles is an acute and highly contagious but vaccine-preventable infectious disease. Despite years of being considered eliminated, decreased vaccination rates have produced virus reemergence in several countries, including Brazil. Measles can be controlled through immunization programs, through which aim to achieve 95% coverage with two doses of the vaccine. Measles can also be controlled if suspected cases can be properly identified in order to contain outbreaks. This cross-sectional study determined the prevalence of measles antibodies and their correlation with rubella antibodies (resulting from the combination vaccine used in Brazil's public immunization program) in individuals aged higher 10 years old in São José do Rio Preto, São Paulo State, Brazil, participants of a prospective cohort of arbovirosis surveillance before virus reemergence in the country. Our findings presented that 32.9% of individuals aged 10-40 years old had not antibodies against measles; 39.3% of total individuals with documented evidence of measles vaccination did not have anti-measles IgG, though only 20.2% of individuals with documented evidence of rubella vaccination lacked anti-rubella IgG. Besides, the most of measles cases reported in the city, following the virus spreading in the country, occurred especially in groups defined by us as susceptible. Because the combination MMR vaccine is part of Brazil's national vaccine schedule, the possible reasons for this relatively high rate of seronegativity need to be investigated further, once that it reflects outbreak risk.

Development of a rapid antiviral screening assay based on eGFP reporter virus of Mayaro virus.

Mayaro virus (MAYV) is a neglected mosquito-borne alphavirus that causes illness similar to Chikungunya (CHIKV), Dengue (DENV) and Zika virus (ZIKV). Currently, there is no specific treatment or vaccine against MAYV infection. To develop an efficient antiviral screening assay for MAYV, we constructed the infectious clones of MAYV strain BeAr 20290 and its eGFP reporter virus. The reporter virus exhibited high replication capacity indistinguishable with the wild type MAYV, and was genetically stable within at least five rounds of passages in BHK-21 cell. The expression of eGFP correlated well with the viral replication. Using the known inhibitor ribavirin, we confirmed that the MAYV-eGFP reporter virus could be used for antiviral screening to identify the specific inhibitors against MAYV. Using the MAYV-eGFP based antiviral assay, we found that the compound 6-Azauridine which had antiviral activity against CHIKV and SFV, showed a significant inhibitory effect on MAYV replication.

Unusual clinical manifestations of dengue disease - Real or imagined?

The disease caused by each of the four serotypes of dengue virus (DENV) have plagued humans since last century. Symptoms of dengue virus (DENV) infection range from asymptomatic to dengue fever (DF) to severe dengue disease (SDD). One third of the world's population lives in regions with active urban DENV transmission, and thousands of serologically naïve travelers visit these areas annually, making a significant portion of the human population at risk of being infected. Even though lifelong immunity to the homotypic serotype is achieved after a primary DENV infection. Heterotypic DENV infections may be exacerbated by a pre-existing immune memory to the primary infection and can result in an increased probability of severe disease. Not only, age, comorbidities and presence of antibodies transferred passively from dengue-immune mother to infants are considered risk factors to dengue severe forms. Plasma leakage and multiple organ impairment are well documented in the literature, affecting liver, lung, brain, muscle, and kidney. However, unusual manifestation, severe or not, have been reported and may require medical attention. This review will summarize and discuss the increasing reports of unusual manifestations in the clinical course of dengue infection.

Zika detection: comparison of methodologies.

Many countries in the Americas have detected local transmission of multiple arboviruses that cause febrile illnesses. Therefore, laboratory testing has become an important tool for confirming the etiology of these diseases. The present study aimed to compare the sensitivity and specificity of three different Zika virus detection assays. One hundred serum samples from patients presenting with acute febrile symptoms were tested using a previously reported TaqMan® RT-qPCR assay. We used a SYBR® Green RT-qPCR and a conventional PCR methodologies to compare the results. Of the samples that were determined to be negative by the TaqMan® RT-qPCR assay, 100% (Kappa=0.670) were also found to be negative by SYBR® Green RT-qPCR based on Tm comparison; however, 14% (Kappa=0.035) were found to be positive by conventional PCR followed by agarose gel electrophoresis. The differences between the ZIKV strains circulating worldwide and the low viremia period can compromise diagnostic accuracy and thereby the accuracy of outbreak data. Therefore, improved assays are required to improve the diagnosis and surveillance of arbovirus.

Evidence of natural Zika virus infection in neotropical non-human primates in Brazil.

In Africa, Old World Primates are involved in the maintenance of sylvatic circulation of ZIKV. However, in Brazil, the hosts for the sylvatic cycle remain unknown. We hypothesized that free-living NHPs might play a role in urban/periurban ZIKV dynamics, thus we undertook an NHP ZIKV investigation in two cities in Brazil. We identified ZIKV-positive NHPs and sequences obtained were phylogenetically related to the American lineage of ZIKV. Additionally, we inoculated four C. penicillata with ZIKV and our results demonstrated that marmosets had a sustained viremia. The natural and experimental infection of NHPs with ZIKV, support the hypothesis that NHPs may be a vertebrate host in the maintainance of ZIKV transmission/circulation in urban tropical settings. Further studies are needed to understand the role they may play in maintaining the urban cycle of the ZIKV and how they may be a conduit in establishing an enzootic transmission cycle in tropical Latin America.

Transverse Myelitis as an Unusual Complication of Dengue Fever.

Dengue fever is the most common arbovirus disease, and presents with a large spectrum of clinical manifestations ranging from asymptomatic disease through to the development of dengue hemorrhagic fever. These extreme cases can lead to dengue shock syndrome, and sometimes death. Spinal cord involvement in dengue virus (DENV) infections is rare. Here, we report a case in which the patient developed acute transverse myelitis (TM) without paraparesis following a DENV infection. This case highlights the importance of physicians' awareness of the possible link between DENV and TM in endemic areas.

Clinical, laboratory and virological data from suspected ZIKV patients in an endemic arbovirus area.

BACKGROUND: The emergence of Zika virus (ZIKV) presents new challenges to both clinicians and public health authorities. Overlapping clinical features between the diseases caused by ZIKV, dengue (DENV) and chikungunya (CHIKV) and the lack of validated serological assays for ZIKV make accurate diagnosis difficult. Brazilian authorities largely rely on clinical and epidemiological data for the epidemiological and clinical classifications of most ZIKV cases. OBJECTIVE: To report the laboratory and clinical profiles of patients diagnosed with Zika fever based only on clinical and epidemiological data. STUDY DESIGN: We analyzed 433 suspected cases of ZIKV identified by the attending physician based on proposed clinical criteria. The samples were also screened for ZIKV, DENV and CHIKV using PCR. RESULTS: Of the 433 patients analyzed, 168 (38.8%) were laboratory-confirmed for arboviruses: 96 were positive for ZIKV, 67 were positive for DENV (56 for DENV-2, 9 for DENV-1, and 2 for DENV-4), four were positive for co-infection with ZIKV/DENV-2, and one was positive for CHIKV. The most common signs or symptoms in the patients with laboratory-confirmed ZIKV were rash (100%), arthralgia (77.1%), fever (74.0%), myalgia (74.0%) and non-purulent conjunctivitis (69.8%). In patients with laboratory-confirmed DENV infections, the most frequently observed symptoms were rash (100%), fever (79.1%), myalgia (74.6%), headache (73.1%) and arthralgia (70.1%). The measure of association between clinical manifestations and laboratory manifestations among patients with ZIKV and DENV detected a statistically significant difference only in abdominal pain (p=0.04), leukopenia (p=0.003), and thrombocytopenia (p=0.01). CONCLUSION: Our data suggests that clinical and epidemiological criteria alone are not a good tool for ZIKV and DENV differentiation, and that laboratory diagnosis should be mandatory.

Mosquito-transmitted viruses - the great Brazilian challenge.

Arboviruses pose a serious threat to public health worldwide, overloading the healthcare system and causing economic losses. These viruses form a very diverse group, and in Brazil, arboviruses belonging to the families Flaviviridae and Togaviridae are predominant. Unfortunately, the number of arboviruses increases in proportion with factors such as deforestation, poor sanitation, climate changes, and introduction of new viruses like Chikungunya virus and Zika virus. In Brazil, dengue is endemic, along with the presence of other arboviruses. The situation is complicated by the scarcity of diagnostic infrastructure and the absence of approved vaccines for these diseases. Disease control, thus, relies solely on vector control. Therefore, enhanced clinical knowledge and improved general awareness about these arboviruses are indispensable to tackle diagnostic inadequacies.

Clinical and laboratory profile of Zika virus infection in dengue suspected patients: A case series.

BACKGROUND: The Zika virus (ZIKV) is an emerging arthropod-borne virus related to the dengue virus (DENV), and shows a similar clinical profile as other arboviral diseases, such as dengue and chikungunya virus (CHIKV). Historically, ZIKV has been associated with sporadic cases of human infection, but is now responsible for outbreaks worldwide. In Brazil, cases have been reported since 2015, with some cases causing severe disease. OBJECTIVE: To identify clinical symptoms of Zika in patients in Dengue suspected patients. STUDY DESIGN: Description of a series of cases, wherein we analyzed 100 clinical samples collected from patients who exhibited acute febrile disease for ≤5days, from January to February 2016. RESULTS: In this study, we report 13 cases of ZIKV infection in adults presenting dengue-like symptoms in a DENV endemic area. All patients presented with fever, with myalgia being the second most frequently observed symptom. Two patients had rashes, but none of them had conjunctivitis. Other less frequent manifestations included headache, arthralgia, diarrhea, and nausea. CONCLUSION: The co-circulation of ZIKV and DENV is a serious public health concern, since it represents both a clinical and diagnostic challenge in endemic areas, as well as in the field of travel medicine.