Design, development, and validation of multi-epitope proteins for serological diagnosis of Zika virus infections and discrimination from dengue virus seropositivity.

Zika virus (ZIKV), an arbovirus from the Flaviviridae family, is the causative agent of Zika fever, a mild and frequent oligosymptomatic disease in humans. Nonetheless, on rare occasions, ZIKV infection can be associated with Guillain-Barré Syndrome (GBS), and severe congenital complications, such as microcephaly. The oligosymptomatic disease, however, presents symptoms that are quite similar to those observed in infections caused by other frequent co-circulating arboviruses, including dengue virus (DENV). Moreover, the antigenic similarity between ZIKV and DENV, and even with other members of the Flaviviridae family, complicates serological testing due to the high cross-reactivity of antibodies. Here, we designed, produced in a prokaryotic expression system, and purified three multiepitope proteins (ZIKV-1, ZIKV-2, and ZIKV-3) for differential diagnosis of Zika. The proteins were evaluated as antigens in ELISA tests for the detection of anti-ZIKV IgG using ZIKV- and DENV-positive human sera. The recombinant proteins were able to bind and detect anti-ZIKV antibodies without cross-reactivity with DENV-positive sera and showed no reactivity with Chikungunya virus (CHIKV)- positive sera. ZIKV-1, ZIKV-2, and ZIKV-3 proteins presented 81.6%, 95%, and 66% sensitivity and 97%, 96%, and 84% specificity, respectively. Our results demonstrate the potential of the designed and expressed antigens in the development of specific diagnostic tests for the detection of IgG antibodies against ZIKV, especially in regions with the circulation of multiple arboviruses.

Ophthalmologic Findings in a Premature Infant leading to a Zika Diagnosis during the COVID-19 Pandemic: A Case Report.

We report on the first case of congenital Zika syndrome to be identified during the COVID-19 pandemic in Puerto Rico. The Zika virus (ZIKV) infection was first seen in Puerto Rico in December 2015. It is a flavivirus with vertical transmission, spreading from infected mothers to their fetuses and having a broad spectrum of clinical manifestations, of which microcephaly is the most worrisome. In Puerto Rico, routine ZIKV screening during pregnancy was implemented in October 2016. However, this practice has become less frequent over time. Nevertheless, the transmission of ZIKV continues, so it is important to ensure routine ZIKV screening in endemic regions, such as Puerto Rico.

Multiomics Approach Reveals Serum Biomarker Candidates for Congenital Zika Syndrome.

The Zika virus (ZIKV) can be vertically transmitted, causing congenital Zika syndrome (CZS) in fetuses. ZIKV infection in early gestational trimesters increases the chances of developing CZS. This syndrome involves several pathologies with a complex diagnosis. In this work, we aim to identify biological processes and molecular pathways related to CZS and propose a series of putative protein and metabolite biomarkers for CZS prognosis in early pregnancy trimesters. We analyzed serum samples of healthy pregnant women and ZIKV-infected pregnant women bearing nonmicrocephalic and microcephalic fetuses. A total of 1090 proteins and 512 metabolites were identified by bottom-up proteomics and untargeted metabolomics, respectively. Univariate and multivariate statistical approaches were applied to find CZS differentially abundant proteins (DAP) and metabolites (DAM). Enrichment analysis (i.e., biological processes and molecular pathways) of the DAP and the DAM allowed us to identify the ECM organization and proteoglycans, amino acid metabolism, and arachidonic acid metabolism as CZS signatures. Five proteins and four metabolites were selected as CZS biomarker candidates. Serum multiomics analysis led us to propose nine putative biomarkers for CZS prognosis with high sensitivity and specificity.

Case report: Zika surveillance complemented with wastewater and mosquito testing.

BACKGROUND: In June 2023, a local cluster of 15 Zika cases was reported in a neighbourhood in Northeastern Singapore. The last significant local transmission of Zika virus (ZIKV) with more than 450 cases was in 2016-2017. To monitor the situation and mitigate further transmission, case, entomological and wastewater-based surveillance were carried out. METHODS: Primary healthcare practitioners and the community were alerted to encourage timely case identification. Surveillance was enhanced through testing of Aedes mosquitoes collected from the National Gravitrap surveillance system, and wastewater samples were collected from a network of autosamplers deployed at manholes across the country. FINDINGS: ZIKV RNA was detected in mosquito pools (3/43; 7%) and individual mosquitoes (3/82; 3.7%) captured, and in wastewater samples (13/503) collected from the vicinity of the cluster of cases. Respective samples collected from other sites across the country were negative. The peak detection of ZIKV RNA in mosquitoes and wastewater coincided temporally with the peak in the number of cases in the area (15-25 May 2023). INTERPRETATION: The restriction of ZIKV signals from wastewater and mosquitoes within the neighbourhood suggested limited ZIKV transmission. The subsequent waning of signals suggested effectiveness of control measures. We demonstrate the utility of wastewater-based surveillance of ZIKV, which complements existing case- and entomological-based surveillance. The non-intrusive approach is particularly useful to monitor diseases such as Zika, which generally causes silent or mild infections, but may cause severe outcomes such as congenital Zika syndrome. FUNDING: This study was funded by Singapore's Ministry of Finance and the National Environment Agency, Singapore.

Clinical description of dengue and chikungunya virus infections amongst acute febrile patients in a malaria endemic area of Mfou, the Centre region of Cameroon.

This study aims to determine the frequency and clinical manifestations of dengue and chikungunya viral infections in the district hospital of Mfou, Centre region of Cameroon where malaria is endemic. Blood samples were collected from suspected cases and tested for Plasmodium parasites and for the molecular detection of viral RNAs (dengue, zika and chikungunya viruses) using TRIOPLEX qPCR. A total of 108 patients were clinically suspected among which 25 % were male and 50 % were less than 15.5 years old. Of these 14.8 % (16/108) and 2.8 % (3/108) had acute dengue and chikungunya fevers respectively. Co-infection with malaria was reported in 56.3 % (9/16) of Dengue cases and 33.3 % (1/3) of chikungunya cases. Clinical profiling further revealed that nausea and vomiting show a significant difference in dengue infected individuals to those of non-infected individuals (P = 0.027). The presence of dengue fever and chikungunya fever and the absence of specific clinical manifestations highlight the need to strengthen surveillance of acute febrile infections for a better estimation of the burden of arboviruses.

Clinical manifestations of dengue, Zika and chikungunya in the Pacific Islands: A systematic review and meta-analysis.

Dengue, Zika and chikungunya outbreaks pose a significant public health risk to Pacific Island communities. Differential diagnosis is challenging due to overlapping clinical features and limited availability of laboratory diagnostic facilities. There is also insufficient information regarding the complications of these arboviruses, particularly for Zika and chikungunya. We conducted a systematic review and meta-analysis to calculate pooled prevalence estimates with 95% confidence intervals (CI) for the clinical manifestations of dengue, Zika and chikungunya in the Pacific Islands. Based on pooled prevalence estimates, clinical features that may help to differentiate between the arboviruses include headache, haemorrhage and hepatomegaly in dengue; rash, conjunctivitis and peripheral oedema in Zika; and the combination of fever and arthralgia in chikungunya infections. We estimated that the hospitalisation and mortality rates in dengue were 9.90% (95% CI 7.67-12.37) and 0.23% (95% CI 0.16-0.31), respectively. Severe forms of dengue occurred in 1.92% (95% CI 0.72-3.63) of reported cases and 23.23% (95% CI 13.58-34.53) of hospitalised patients. Complications associated with Zika virus included Guillain-Barré syndrome (GBS), estimated to occur in 14.08 (95% CI 11.71-16.66) per 10,000 reported cases, and congenital brain malformations such as microcephaly, particularly with first trimester maternal infection. For chikungunya, the hospitalisation rate was 2.57% (95% CI 1.30-4.25) and the risk of GBS was estimated at 1.70 (95% CI 1.06-2.48) per 10,000 reported cases. Whilst ongoing research is required, this systematic review enhances existing knowledge on the clinical manifestations of dengue, Zika and chikungunya infections and will assist Pacific Island clinicians during future arbovirus outbreaks.

Detecting the impacts of humidity, rainfall, temperature, and season on chikungunya, dengue and Zika viruses in Aedes albopictus mosquitoes from selected sites in Cebu city, Philippines.

BACKGROUND: Aedes albopictus is the secondary vector for dengue virus (DENV) in the Philippines, and also harbors chikungunya (CHIKV) and Zika (ZIKV) viruses. This study aimed to determine the minimum infection rates (MIRs) of CHIKV, DENV serotypes, and ZIKV in Ae. albopictus collected from selected two-site categories by altitude (highland [H] and lowland [L] sites) in Cebu city, Philippines during the wet (WS) and dry seasons (DS) of 2021-2022, and to explore the relationships between these arboviral MIRs and the local weather. METHODS: The viral RNA extracts in pooled and reared adult Ae. albopictus collected during the DS and WS from two-site categories were subjected to RT-PCR to amplify and detect gene loci specific for CHIKV, DENV-1 to DENV-4, and ZIKV and analyzed with the weather data. RESULTS: The range of CHIKV MIRs was higher in the WS (13.61-107.38 infected individuals per 1,000 mosquitoes) than in the DS (13.22-44.12), but was similar between the two-site categories. Rainfall (RF) influenced the CHIKV MIR. The MIR ranges of both DENV-2 (WS: H = 0, L = 0; DS: H = 0-5.92; L = 0-2.6) and DENV-4 (WS: H = 0, L = 0-2.90; DS: H = 2.96-6.13, L = 0-15.63) differed by season but not between the two-site categories. Relative humidity (RH), RF, and temperature did not influence DENVs' MIRs. The MIR range of ZIKV was similar in both seasons (WS: 11.36-40.27; DS: 0-46.15) and two-site categories (H = 0-90.91, L = 0-55.56). RH and temperature influenced ZIKV MIR. CONCLUSIONS: RF influenced CHIKV MIR in Ae. albopictus, whereas RH and temperature influenced that of ZIKV. Season influenced the MIRs of CHIKV and DENVs but not in ZIKV. Ae. albopictus were co-infected with CHIKV, DENVs, and ZIKV in both highland and lowland sites in Cebu city. Recommendations include all-year-round implementation of the Philippine Department of Health's  4S enhanced strategy and installation of water pipelines in rural highlands for vector and disease control. Our findings are relevant to protect public health in the tropics in this climate change.

Proteomics and Metabolomics in Congenital Zika Syndrome: A Review of Molecular Insights and Biomarker Discovery.

Zika virus (ZIKV) infection can be transmitted vertically, leading to the development of congenital Zika syndrome (CZS) in infected fetuses. During the early stages of gestation, the fetuses face an elevated risk of developing CZS. However, it is important to note that late-stage infections can also result in adverse outcomes. The differences between CZS and non-CZS phenotypes remain poorly understood. In this review, we provide a summary of the molecular mechanisms underlying ZIKV infection and placental and blood-brain barriers trespassing. Also, we have included molecular alterations that elucidate the progression of CZS by proteomics and metabolomics studies. Lastly, this review comprises investigations into body fluid samples, which have aided to identify potential biomarkers associated with CZS.

Neuroinflammation, blood-brain barrier dysfunction, hippocampal atrophy and delayed neurodevelopment: Contributions for a rat model of congenital Zika syndrome.

The congenital Zika syndrome (CZS) has been characterized as a set of several brain changes, such as reduced brain volume and subcortical calcifications, in addition to cognitive deficits. Microcephaly is one of the possible complications found in newborns exposed to Zika virus (ZIKV) during pregnancy, although it is an impacting clinical sign. This study aimed to investigate the consequences of a model of congenital ZIKV infection by evaluating the histopathology, blood-brain barrier, and neuroinflammation in pup rats 24 h after birth, and neurodevelopment of the offspring. Pregnant rats were inoculated subcutaneously with ZIKV-BR at the dose 1 × 107 plaque-forming unit (PFU mL-1) of ZIKV isolated in Brazil (ZIKV-BR) on gestational day 18 (G18). A set of pups, 24 h after birth, was euthanized. The brain was collected and later evaluated for the histopathology of brain structures through histological analysis. Additionally, analyses of the blood-brain barrier were conducted using western blotting, and neuroinflammation was assessed using ELISA. Another set of animals was evaluated on postnatal days 3, 6, 9, and 12 for neurodevelopment by observing the developmental milestones. Our results revealed hippocampal atrophy in ZIKV animals, in addition to changes in the blood-brain barrier structure and pro-inflammatory cytokines expression increase. Regarding neurodevelopment, a delay in important reflexes during the neonatal period in ZIKV animals was observed. These findings advance the understanding of the pathophysiology of CZS and contribute to enhancing the rat model of CZS.

Detection of anti-premembrane antibody as a specific marker of four flavivirus serocomplexes and its application to serosurveillance in endemic regions.

In the past few decades, several emerging/re-emerging mosquito-borne flaviviruses have resulted in disease outbreaks of public health concern in the tropics and subtropics. Due to cross-reactivities of antibodies recognizing the envelope protein of different flaviviruses, serosurveillance remains a challenge. Previously we reported that anti-premembrane (prM) antibody can discriminate between three flavivirus infections by Western blot analysis. In this study, we aimed to develop a serological assay that can discriminate infection or exposure with flaviviruses from four serocomplexes, including dengue (DENV), Zika (ZIKV), West Nile (WNV) and yellow fever (YFV) viruses, and explore its application for serosurveillance in flavivirus-endemic countries. We employed Western blot analysis including antigens of six flaviviruses (DENV1, 2 and 4, WNV, ZIKV and YFV) from four serocomplexes. We tested serum samples from YF-17D vaccinees, and from DENV, ZIKV and WNV panels that had been confirmed by RT-PCR or by neutralization assays. The overall sensitivity/specificity of anti-prM antibodies for DENV, ZIKV, WNV, and YFV infections/exposure were 91.7%/96.4%, 91.7%/99.2%, 88.9%/98.3%, and 91.3%/92.5%, respectively. When testing 48 samples from Brazil, we identified multiple flavivirus infections/exposure including DENV and ZIKV, DENV and YFV, and DENV, ZIKV and YFV. When testing 50 samples from the Philippines, we detected DENV, ZIKV, and DENV and ZIKV infections with a ZIKV seroprevalence rate of 10%, which was consistent with reports of low-level circulation of ZIKV in Asia. Together, these findings suggest that anti-prM antibody is a flavivirus serocomplex-specific marker and can be employed to delineate four flavivirus infections/exposure in regions where multiple flaviviruses co-circulate.

Zika virus infection during pregnancy and vertical transmission: case reports and peptide-specific cell-mediated immune responses.

Zika virus (ZIKV) infection in pregnant women is associated with birth defects, which are more prevalent and severe the earlier in pregnancy the infection occurs. Pregnant women at risk of possible ZIKV exposure (n = 154) were screened using ELISA for ZIKV IgM and IgG. Nine of 154 (5.84%) pregnant women who underwent screening exhibited positive ZIKV serology. Of these, two maternal infections were confirmed by real-time RT-PCR and five were considered probable, but only three of those were retained for further analysis based on strict diagnostic criteria. Plaque reduction neutralization tests (PRNT) confirmed ZIKV infection in nine cases (5.84%). Two cases of vertical ZIKV transmission were confirmed by PCR. One infant showed no signs of congenital ZIKV syndrome and had a normal developmental profile despite first-trimester maternal infection. In the second case, pregnancy was terminated. Production of interferon γ (IFN-γ) by peripheral blood mononuclear cells obtained from pregnant women and umbilical cord blood was measured using enzyme-linked immunospot assay (ELISpot) after stimulation with panels of synthetic peptides derived from the sequence of ZIKV proteins. This analysis revealed that, among all peptide pools tested, those derived from the ZIKV envelope protein generated the strongest IFN-γ response.

Does a patient with acquired arbovirus infection have a hearing impairment? A scoping review of hearing changes in an adult with Dengue, Chikungunya, and Zika.

OBJECTIVES: To identify and understand the evidence regarding hearing changes related to acquired Dengue, Chikungunya, and Zika virus infection in adult individuals. METHODS: A scoping review was performed according to the recommendations of The Joanna Briggs Institute and guidelines of the Preferred Reporting Items for Systematic Reviews and Meta-Analyses extension for Scoping Reviews in the Embase, PubMed/Medline, ScienceDirect, Scopus, and Web of Science databases without restriction on language and year of publication. Case studies, observational studies, and clinical trials reporting hearing loss in adult subjects (>18-60 years of age) of both sexes with DENV, CHIKV, or ZIKV diagnosed by positive molecular/serological examination by RT-PCR or IgM/IgG by ELISA method were included. RESULTS: Thirteen studies met the inclusion criteria and were selected for review. The occurrence of auditory symptoms caused by arboviroses and the presence of permanent or transient sensorineural hearing loss was variable in adults. CONCLUSIONS: Dengue, Chikungunya, and Zika infections in adults are associated with a variety of auditory symptoms. The frequency of permanent or transient sensorineural hearing loss is low but not negligible.

Chikungunya virus antepartum transmission and abnormal infant outcomes in a cohort of pregnant women in Nigeria.

OBJECTIVES: Chikungunya virus (CHIKV), a reemerging global public health concern, which causes acute febrile illness, rash, and arthralgia and may affect both mothers and infants during pregnancy. Mother-to-child transmission (MTCT) of CHIKV in Africa remains understudied. METHODS: Our cohort study screened 1006 pregnant women with a Zika/dengue/CHIKV rapid test at two clinics in Nigeria between 2019 and 2022. Women who tested positive for the rapid test were followed through their pregnancy and their infants were observed for 6 months, with a subset tested by reverse transcription-polymerase chain reaction (RT-PCR) and neutralization, to investigate seropositivity rates and MTCT of CHIKV. RESULTS: Of the 1006, 119 tested positive for CHIKV immunoglobulin (Ig)M, of which 36 underwent detailed laboratory tests. While none of the IgM reactive samples were RT-PCR positive, 14 symptomatic pregnant women were confirmed by CHIKV neutralization test. Twelve babies were followed with eight normal and four abnormal outcomes, including stillbirth, cleft lip/palate with microcephaly, preterm delivery, polydactyly with sepsis, and jaundice. CHIKV IgM testing identified three possible antepartum transmissions. CONCLUSION: In Nigeria, we found significant CHIKV infection in pregnancy and possible CHIKV antepartum transmission associated with birth abnormalities.

Neurodevelopmental assessment of normocephalic children born to Zika virus exposed and unexposed pregnant people.

BACKGROUND: Studies examining the association between in utero Zika virus (ZIKV) exposure and child neurodevelopmental outcomes have produced varied results. METHODS: We aimed to assess neurodevelopmental outcomes among normocephalic children born from pregnant people enrolled in the Zika in Pregnancy in Honduras (ZIPH) cohort study, July-December 2016. Enrollment occurred during the first prenatal visit. Exposure was defined as prenatal ZIKV IgM and/or ZIKV RNA result at enrollment. Normocephalic children, >6 months old, were selected for longitudinal follow-up using the Bayley Scales of Infant and Toddler Development (BSID-III) and the Ages & Stages Questionnaires: Social-Emotional (ASQ:SE-2). RESULTS: One hundred fifty-two children were assessed; after exclusion, 60 were exposed and 72 were unexposed to ZIKV during pregnancy. Twenty children in the exposed group and 21 children in the unexposed group had a composite score <85 in any of the BSID-III domains. Although exposed children had lower cognitive and language scores, differences were not statistically significant. For ASQ:SE-2 assessment, there were not statistically significant differences between groups. CONCLUSIONS: This study found no statistically significant differences in the neurodevelopment of normocephalic children between in utero ZIKV exposed and unexposed. Nevertheless, long-term monitoring of children with in utero ZIKV exposure is warranted. IMPACT: This study found no statistically significant differences in the neurodevelopment in normocephalic children with in utero Zika virus exposure compared to unexposed children, although the exposed group showed lower cognitive and language scores that persisted after adjustment by maternal age and education and after excluding children born preterm and low birth weight from the analysis. Children with prenatal Zika virus exposure, including those normocephalic and have no evidence of abnormalities at birth, should be monitored for neurodevelopmental delays. Follow-up is important to be able to detect developmental abnormalities that might not be detected earlier in life.

ICP-MS and fluorescence dual-mode detection of ZIKV-RNA based on quantum dot labeling with hybridization chain reaction.

The detection of Zika virus (ZIKV) is of great significance to human life and health. Herein, we presented an ICP-MS and fluorescent dual-mode sensor for quantitative analysis of Zika virus RNA fragments (ZIKV-RNA), which employed quantum dots (QDs) as signal tags and combined with hybridization chain reaction (HCR). The dual-mode sensor realized cross-checking of the analysis results and improved the assay accuracy. Firstly, the single-stranded DNA (ssDNA) was anchored on the surface of magnetic beads (MBs). Afterward, HCR was conducted with probe DNA-CdSe quantum dots conjugates (pDNA-QDs) and link DNA (lDNA), producing the MBs-ssDNA-[pDNA-QDs-lDNA]n conjugates. In the presence of target ZIKV-RNA, a strand displacement reaction occurred, leading to the dissociation of the [pDNA-QDs-lDNA]n labels from the conjugates into the solution. Finally, the signal intensity was detected using ICP-MS and fluorescence analysis, with achieved limits of detection of 131 pM and 152 pM, respectively. The inter-assay RSD values of fluorescence and ICP-MS were 3.94 % and 4.26 % at 10 nM level, respectively, showing that the method had good precision. This method showed high selectivity and was applied to the analysis of biological fluids. There was no significant difference between the results of ICP-MS modes and fluorescence mode. This method offers a new strategy for sensitivity analysis of ZIKV-RNA and exhibits promise in clinical applications for diagnosis.

IgM-specific linear epitopes on the E2 protein for serodiagnosis of Chikungunya.

Chikungunya virus (CHIKV) and Dengue virus (DENV) are vector-borne diseases transmitted by Aedes aegypti and Aedes albopictus that pose a significant threat to global public health. Cases of acute Chikungunya fever often present similar clinical symptoms to other vector-borne diseases, such as Dengue fever. In regions where multiple vector-borne diseases coexist, CHIKV is often overlooked or misdiagnosed as Dengue virus, West Nile virus, Zika virus or other viral infections, which delays its prevention and control. However, IgM antibodies directed against the E2 protein of CHIKV have not yet been generalized to clinical settings due to the low sensitivity and high cost in commercial kits. Indirect ELISA with peptides provides an effective supplementary tool for detecting CHIKV IgM antibodies. Our study aims at examining the potential of linear epitopes on the E2 glycoprotein that specifically bind to IgM antibodies as serodiagnostic tool for CHIKV. The sensitivity of the established peptide indirect ELISA method for detecting clinical samples is significantly better than that of commercial kits, realizing a beneficial supplement to the existing IgM antibody assay. It also established the groundwork for comprehending the biological mechanisms of the CHIKV E2 protein and the advancement of innovative epitope peptide vaccines.

Detection and persistence of Zika virus in body fluids and associated factors: a prospective cohort study.

This study aimed to analyze the detection and duration of the Zika virus (ZIKV) in plasma, urine, saliva, sweat, rectal swabs, vaginal secretions, breast milk, and semen and to explore risk factors associated with prolonged viral persistence. A prospective cohort study of symptomatic patients and their household contacts was conducted in Brazil from July 2017 to June 2019. A total of 260 individuals (184 women and 76 men) with confirmed ZIKV infection were enrolled and followed up for 12 months. ZIKV RNA was present in all body fluid specimens and detectable for extended periods in urine, sweat, rectal swabs, and semen. The longest detection duration was found in semen, with high viral loads in the specimens. ZIKV RNA clearance was associated with several factors, including age, sex, education level, body mass index, non-purulent conjunctivitis, joint pain, and whether the participant had a history of yellow fever vaccination. The influence of each of these factors on the low or fast viral clearance varied according to the specific body fluid under investigation. Recurrent ZIKV detection events after total viral clearance were observed in the cohort. Our findings provide valuable insights into the persistence and potential recurrence of ZIKV infection, highlighting the need for continued monitoring and follow-up of individuals infected with ZIKV and for effective prevention measures to reduce the risk of transmission.

Rapid On-Site Detection of Arboviruses by a Direct RT-qPCR Assay.

Arthropod-borne diseases currently constitute a source of major health concerns worldwide. They account for about 50% of global infectious diseases and cause nearly 700,000 deaths every year. Their rapid increase and spread constitute a huge challenge for public health, highlighting the need for early detection during epidemics, to curtail the virus spread, and to enhance outbreak management. Here, we compared a standard quantitative polymerase chain reaction (RT-qPCR) and a direct RT-qPCR assay for the detection of Zika (ZIKV), Chikungunya (CHIKV), and Rift Valley Fever (RVFV) viruses from experimentally infected-mosquitoes. The direct RT-qPCR could be completed within 1.5 h and required 1 µL of viral supernatant from homogenized mosquito body pools. Results showed that the direct RT-qPCR can detect 85.71%, 89%, and 100% of CHIKV, RVFV, and ZIKV samples by direct amplifications compared to the standard method. The use of 1:10 diluted supernatant is suggested for CHIKV and RVFV direct RT-qPCR. Despite a slight drop in sensitivity for direct PCR, our technique is more affordable, less time-consuming, and provides a better option for qualitative field diagnosis during outbreak management. It represents an alternative when extraction and purification steps are not possible because of insufficient sample volume or biosecurity issues.

Review of dengue, zika and chikungunya infections in nervous system in endemic areas.

Dengue, zika, and chikungunya are arboviruses of great epidemiological relevance worldwide. The emergence and re-emergence of viral infections transmitted by mosquitoes constitute a serious human public health problem. The neurological manifestations caused by these viruses have a high potential for death or sequelae. The complications that occur in the nervous system associated with arboviruses can be a challenge for diagnosis and treatment. In endemic areas, suspected cases should include acute encephalitis, myelitis, encephalomyelitis, polyradiculoneuritis, and/or other syndromes of the central or peripheral nervous system, in the absence of a known explanation. The confirmation diagnosis is based on viral (isolation or RT-PCR) or antigens detection in tissues, blood, cerebrospinal fluid, or other body fluids, increase in IgG antibody titers between paired serum samples, specific IgM antibody in cerebrospinal fluid and serological conversion to IgM between paired serum samples (non-reactive in the acute phase and reactive in the convalescent). The cerebrospinal fluid examination can demonstrate: 1. etiological agent; 2. inflammatory reaction or protein-cytological dissociation depending on the neurological condition; 3. specific IgM, 4. intrathecal synthesis of specific IgG (dengue and chikungunya); 5. exclusion of other infectious agents. The treatment of neurological complications aims to improve the symptoms, while the vaccine represents the great hope for the control and prevention of neuroinvasive arboviruses. This narrative review summarizes the updated epidemiology, general features, neuropathogenesis, and neurological manifestations associated with dengue, zika, and chikungunya infection.

Dengue, zika e chikungunya são arboviroses de grande relevância epidemiológica em todo o mundo. A emergência e reemergência dessas infecções virais transmitidas por mosquitos constituem um grave problema de saúde pública humana. As manifestações neurológicas causadas por esses vírus têm alto potencial de morte ou sequelas. As complicações que ocorrem no sistema nervoso associadas às arboviroses podem representar um desafio diagnóstico e de tratamento. Em áreas endêmicas, casos suspeitos devem incluir encefalite, mielite, encefalomielite, polirradiculoneurite e/ou outras síndromes do sistema nervoso central ou periférico, na ausência de explicação conhecida. Caso confirmado de arbovirose neuroinvasivo é baseado na detecção viral (isolamento ou RT-PCR) ou de antígenos em tecidos, sangue, líquido cefalorraquidiano ou outros fluidos corporais, aumento dos títulos de anticorpos IgG entre amostras de soro pareadas, anticorpo IgM específico no líquido cefalorraquidiano e conversão sorológica para IgM entre amostras de soro pareadas. O exame do líquido cefalorraquidiano pode demonstrar: 1. agente etiológico; 2. reação inflamatória ou dissociação proteico-citológica, dependendo do quadro neurológico; 3. valor absoluto de IgM específica; 4. síntese intratecal de anticorpos IgG específicos (dengue e chikungunya); 5. exclusão de outros agentes infecciosos. O tratamento das complicações neurológicas visa melhorar os sintomas, enquanto a vacina representa a grande esperança para o controle e a prevenção das arboviroses neuroinvasivas. Esta revisão narrativa resume a atualização da epidemiologia, características gerais, neuropatogênese e manifestações neurológicas associadas à infecção pelos vírus da dengue, zika e chikungunya.

High Seroprevalence of IgG Antibodies to Multiple Arboviruses in People Living with HIV (PLWHIV) in Madagascar.

To estimate the prevalence of IgG antibodies against six arboviruses in people living with HIV-1 (PLWHIV) in Madagascar, we tested samples collected between January 2018 and June 2021. We used a Luminex-based serological assay to detect IgG antibodies against antigens from Dengue virus serotypes 1-4 (DENV1-4), Zika virus (ZIKV), West Nile virus (WNV), Usutu virus (USUV), Chikungunya virus (CHIKV), and O'nyong nyong virus (ONNV). Of the 1036 samples tested, IgG antibody prevalence was highest for ONNV (28.4%), CHIKV (26.7%), WNV-NS1 (27.1%), DENV1 (12.4%), USUV (9.9%), and DENV3 (8.9%). ZIKV (4.9%), DENV2 (4.6%), WNV-D3 (5.1%), and DENV4 (1.4%) were lower. These rates varied by province of origin, with the highest rates observed in Toamasina, on the eastern coast (50.5% and 56.8%, for CHIKV and ONNV, respectively). The seroprevalence increased with age for DENV1 and 3 (p = 0.006 and 0.038, respectively) and WNV DIII (p = 0.041). The prevalence of IgG antibodies against any given arborvirus varied over the year and significantly correlated with rainfalls in the different areas (r = 0.61, p = 0.036). Finally, we found a significant correlation between the seroprevalence of antibodies against CHIKV and ONNV and the HIV-1 RNA plasma viral load. Thus, PLWHIV in Madagascar are highly exposed to various arboviruses. Further studies are needed to explain some of our findings.