Comparison of the sensitivity and specificity of commercial anti-dengue virus IgG tests to identify persons eligible for dengue vaccination.

The Advisory Committee on Immunization Practices (ACIP) recommended that dengue pre-vaccination screening tests for Dengvaxia administration have at least 98% specificity and 75% sensitivity. This study evaluates the performance of commercial anti-DENV IgG tests to identify tests that could be used for pre-vaccination screening. First, for seven tests, we evaluated sensitivity and specificity in early convalescent dengue virus (DENV) infection, using 44 samples collected 7-30 days after symptom onset and confirmed by RT-PCR. Next, for the five best-performing tests and two additional tests (with and without an external test reader) that became available later, we evaluated performance to detect past dengue infection among a panel of 44 specimens collected in 2018-2019 from healthy 9- to 16-year-old children from Puerto Rico. Finally, a full-scale evaluation was done with the four best-performing tests using 400 specimens from the same population. We used virus focus reduction neutralization test and an in-house DENV IgG ELISA as reference standards. Of seven tests, five showed ≥75% sensitivity in detecting anti-DENV IgG in early convalescent specimens with low cross-reactivity to the Zika virus. For the detection of previous DENV infections, the tests with the highest performance were the Euroimmun NS1 IgG ELISA (sensitivity 84.5%, specificity 97.1%) and CTK Dengue IgG rapid test R0065C with the test reader (sensitivity 76.2% specificity 98.1%). There are IgG tests available that can be used to accurately classify individuals with previous DENV infection as eligible for dengue vaccination to support safe vaccine implementation. IMPORTANCE: The Advisory Committee on Immunization Practices (ACIP) has set forth recommendations that dengue pre-vaccination screening tests must exhibit at least 98% specificity and 75% sensitivity. Our research rigorously assesses the performance of various commercial tests against these benchmarks using well-characterized specimens from Puerto Rico. The findings from our study are particularly relevant given FDA approval and ACIP recommendation of Sanofi Pasteur's Dengvaxia vaccine, highlighting the need for accurate pre-vaccination screening tools.

North-south pathways, emerging variants, and high climate suitability characterize the recent spread of dengue virus serotypes 2 and 3 in the Dominican Republic.

BACKGROUND: Dengue fever remains a significant public health challenge in tropical and subtropical regions, with its transmission dynamics being influenced by both environmental factors and human mobility. The Dominican Republic, a biodiversity hotspot in the Caribbean, has experienced recurrent dengue outbreaks, yet detailed understanding of the virus's transmission pathways and the impact of climatic factors remains limited. This study aims to elucidate the recent transmission dynamics of the dengue virus (DENV) in the Dominican Republic, utilizing a combination of genomic sequencing and epidemiological data analysis, alongside an examination of historical climate patterns. METHODS: We conducted a comprehensive study involving the genomic sequencing of DENV samples collected from patients across different regions of the Dominican Republic over a two-year period. Phylogenetic analyses were performed to identify the circulation of DENV lineages and to trace transmission pathways. Epidemiological data were integrated to analyze trends in dengue incidence and distribution. Additionally, we integrated historical climate data spanning several decades to assess trends in temperature and their potential impact on DENV transmission potential. RESULTS: Our results highlight a previously unknown north-south transmission pathway within the country, with the co-circulation of multiple virus lineages. Additionally, we examine the historical climate data, revealing long-term trends towards higher theoretical potential for dengue transmission due to rising temperatures. CONCLUSION: This multidisciplinary study reveals intricate patterns of dengue virus transmission in the Dominican Republic, characterized by the co-circulation of multiple DENV lineages and a novel transmission pathway. The observed correlation between rising temperatures and increased dengue transmission potential emphasizes the need for integrated climate-informed strategies in dengue control efforts. Our findings offer critical insights for public health authorities in the Dominican Republic and similar settings, guiding resource allocation and the development of preparedness strategies to mitigate the impacts of climate change on dengue transmission.

SARS-CoV-2 Omicron Replacement of Delta as Predominant Variant, Puerto Rico.

We reconstructed the SARS-CoV-2 epidemic caused by Omicron variant in Puerto Rico by sampling genomes collected during October 2021-May 2022. Our study revealed that Omicron BA.1 emerged and replaced Delta as the predominant variant in December 2021. Increased transmission rates and a dynamic landscape of Omicron sublineage infections followed.

Dengue Outbreak Response during COVID-19 Pandemic, Key Largo, Florida, USA, 2020.

We report a dengue outbreak in Key Largo, Florida, USA, from February through August 2020, during the COVID-19 pandemic. Successful community engagement resulted in 61% of case-patients self-reporting. We also describe COVID-19 pandemic effects on the dengue outbreak investigation and the need to increase clinician awareness of dengue testing recommendations.

Reemergence of Dengue Virus Serotype 3, Brazil, 2023.

We characterized 3 autochthonous dengue virus serotype 3 cases and 1 imported case from 2 states in the North and South Regions of Brazil, 15 years after Brazil's last outbreak involving this serotype. We also identified a new Asian lineage recently introduced into the Americas, raising concerns about future outbreaks.

Predominance of Severe Plasma Leakage in Pediatric Patients With Severe Dengue in Puerto Rico.

BACKGROUND: We evaluated clinical and laboratory findings among patients with nonsevere or severe dengue in Puerto Rico to examine whether clinical manifestations vary by age. METHODS: During 2012-2014, we enrolled patients who arrived at the emergency department with fever or history of fever within 7 days of presentation. Serum samples were tested for dengue virus (DENV) by reverse transcriptase-polymerase chain reaction (RT-PCR) and IgM enzyme-linked immunosorbent assay (ELISA). Severe dengue was defined as severe plasma leakage or shock, severe bleeding, or organ involvement at presentation, during hospitalization, or follow-up. RESULTS: Of 1089 dengue patients identified, 281 (26%) were severe. Compared to those with nonsevere dengue, patients with severe dengue were more often aged 10-19 years (55% vs 40%, P < .001) and hospitalized (87% vs 30%, P < .001). Severe plasma leakage or shock was more common among children aged 0-9 (59%) or 10-19 years (86%) than adults (49%) (P < .01). Severe bleeding was less common among 10-19 year olds (24%) compared to 0-9 year olds (45%) and adults (52%; P < .01). CONCLUSIONS: Severe plasma leakage was the most common presentation among children, highlighting important differences from adults. Vaccination against dengue could help prevent severe dengue among children in Puerto Rico.

Emergence of Dengue Virus Serotype 2 Cosmopolitan Genotype, Brazil.

We used nanopore sequencing and phylogenetic analyses to identify a cosmopolitan genotype of dengue virus serotype 2 that was isolated from a 56-year-old male patient from the state of Goiás in Brazil. The emergence of a cosmopolitan genotype in Brazil will require risk assessment and surveillance to reduce epidemic potential.

Tracing the Origin, Spread, and Molecular Evolution of Zika Virus in Puerto Rico, 2016-2017.

We reconstructed the 2016-2017 Zika virus epidemic in Puerto Rico by using complete genomes to uncover the epidemic's origin, spread, and evolutionary dynamics. Our study revealed that the epidemic was propelled by multiple introductions that spread across the island, intricate evolutionary patterns, and ≈10 months of cryptic transmission.

Novel Assay to Measure Seroprevalence of Zika Virus in the Philippines.

Zika virus (ZIKV) is a member of the Flaviviridae family, which includes other clinically notable viruses such as the 4 dengue virus serotypes (DENV-1-4). Distinguishing DENVs from ZIKV using the established serologic assays widely used for monitoring DENV transmission is difficult because of antibody cross-reactivity between these closely related flaviviruses. We describe a modified and improved recombinant envelope domain III-based serologic assay for detecting ZIKV type-specific antibodies in regions with endemic DENV transmission. When the assay was used to measure ZIKV seroprevalence in 2017 among children 9-14 years of age living in a region of the Philippines with endemic DENV transmission, we observed a ZIKV seroprevalence of 18%. Investigators should consider using the ZIKV envelope domain III-based assay, which is simple and readily adaptable for use in standard clinical and public health laboratories, to assess ZIKV seroprevalence in areas with endemic DENV transmission.

Evaluating Differences in Whole Blood, Serum, and Urine Screening Tests for Zika Virus, Puerto Rico, USA, 2016.

We evaluated nucleic acid amplification testing (NAAT) for Zika virus on whole-blood specimens compared with NAAT on serum and urine specimens among asymptomatic pregnant women during the 2015-2016 Puerto Rico Zika outbreak. Using NAAT, more infections were detected in serum and urine than in whole blood specimens.

Dengue and Zika Virus Diagnostic Testing for Patients with a Clinically Compatible Illness and Risk for Infection with Both Viruses.

Dengue and Zika viruses are closely related mosquitoborne flaviviruses with similar transmission cycles, distribution throughout the tropics and subtropics, and disease manifestations including fever, rash, myalgia, and arthralgia. For patients with suspected dengue or Zika virus disease, nucleic acid amplification tests (NAATs) are the preferred method of diagnosis. Immunoglobulin M (IgM) antibody testing can identify additional infections and remains an important tool for the diagnosis of these diseases, but interpreting the results is complicated by cross-reactivity, and determining the specific timing of infection can be difficult. These limitations are a particular challenge for pregnant women in determining whether Zika virus infection occurred during or before the pregnancy.This report summarizes existing and new guidance on dengue and Zika virus diagnostic testing for patients with a clinically compatible illness who live in or recently traveled to an area where there is risk for infection with both viruses. CDC recommendations for screening of asymptomatic pregnant women with possible Zika virus exposure are unchanged. For symptomatic nonpregnant persons, dengue and Zika virus NAATs should be performed on serum collected ≤7 days after symptom onset. Dengue and Zika virus IgM antibody testing should be performed on NAAT-negative serum specimens or serum collected >7 days after onset of symptoms. For symptomatic pregnant women, serum and urine specimens should be collected as soon as possible within 12 weeks of symptom onset for concurrent dengue and Zika virus NAATs and IgM antibody testing. Positive IgM antibody test results with negative NAAT results should be confirmed by neutralizing antibody tests when clinically or epidemiologically indicated, including for all pregnant women. Data on the epidemiology of viruses known to be circulating at the location of exposure and clinical findings should be considered when deciding which tests to perform and for interpreting results.Patients with clinically suspected dengue should receive appropriate management to monitor and treat shock and hemorrhage. Women with laboratory evidence of possible Zika virus infection during pregnancy and their infants should be evaluated and managed for possible adverse outcomes. Dengue and Zika virus disease are nationally notifiable conditions, and cases should be reported to public health authorities.

Multiplexed Biomarker Panels Discriminate Zika and Dengue Virus Infection in Humans.

Zika virus (ZIKV) and dengue virus (DENV) are closely related flaviviruses that cause widespread, acute febrile illnesses, notably microcephaly for fetuses of infected pregnant women. Detecting the viral cause of these illnesses is paramount to determine risks to patients, counsel pregnant women, and help fight outbreaks. A combined diagnostic algorithm for ZIKV and DENV requires Reverse transcription polymerase chain reaction (RT-PCR) and IgM antibody detection. RT-PCR differentiates between DENV and ZIKV infections during the acute phases of infection, but differentiation based on IgM antibodies is currently nearly impossible in endemic areas. We have developed a ZIKV/DENV protein array and tested it with serum samples collected from ZIKV- and DENV-infected patients and healthy subjects in Puerto Rico. Our analyses reveal a biomarker panel that are capable of discriminating ZIKV and DENV infections with high accuracy, including Capsid protein from African ZIKV strain MR766, and other 5 pair of proteins (NS1, NS2A, NS3, NS4B and NS5) from ZIKV and DENV respectively. Both sensitivity and specificity of the test for ZIKV from DENV are around 90%. We propose that the ZIKV/DENV protein array will be used in future studies to discriminate patients infected with ZIKV from DENV.

Prevalence and Incidence of Zika Virus Infection Among Household Contacts of Patients With Zika Virus Disease, Puerto Rico, 2016-2017.

BACKGROUND: Little is known about the prevalence or incidence of Zika virus (ZIKV) infection in settings affected by the 2015-2016 Zika pandemic and associated risk factors. We assessed these factors among household contacts of patients with ZIKV disease enrolled in a cohort study in Puerto Rico during 2016-2017. METHODS: Household contacts of index case patients completed a questionnaire and gave specimens for real-time polymerase chain reaction (RT-PCR) and immunoglobulin M enzyme-linked immunosorbent assay testing to detect ZIKV infection. We measured the prevalence of ZIKV infection among contacts and associated individual and household factors, examined sexual transmission using a sexual-networks approach, and assessed incident infection among initially uninfected household contacts 2-4 months later. RESULTS: Of 366 contacts, 34.4% had evidence of ZIKV infection at enrollment, including 11.2% by RT-PCR. Having open doors and windows that were either screened (prevalence ratio [PR], 2.1 [95% confidence interval {CI}, 1.2-3.6]) or unscreened (PR, 2.5 [95% CI, 1.5-4.1]) was associated with increased prevalence. Sexual partners were more likely to both be RT-PCR positive relative to other relationships (odds ratio, 2.2 [95% CI, 1.1-4.5]). At follow-up, 6.1% of contacts had evidence of incident infection. CONCLUSIONS: This study identified sexual contact as a risk factor for ZIKV infection. Persons living with ZIKV-infected individuals should be a focus of public health efforts.

Duration of the Presence of Infectious Zika Virus in Semen and Serum.

Zika virus (ZIKV) has recently caused a large epidemic in the Americas that is associated with birth defects. Although ZIKV is primarily transmitted by Aedes mosquitoes, ZIKV RNA is detectable in blood and semen of infected individuals for weeks or months, during which sexual and other modes of transmission are possible. However, viral RNA is usually detectable longer than infectious virus is present. We determined the frequency of isolation of infectious virus from semen and serum samples prospectively obtained from a cohort of patients in Puerto Rico. We confirmed isolation of infectious virus on the basis of a tissue culture cytopathic effect, an increase in virus genome copy equivalents (GCE), and positive results of immunofluorescence analysis; virus in infected cells was quantitated by flow cytometry. These criteria confirmed the presence of infectious virus in semen specimens from 8 of 97 patients for up to 38 days after initial detection when virus loads are >1.4 × 106 genome copy equivalents/mL. Two serum isolates were obtained from 296 patients. These findings can help guide important prevention guidelines for persons that may potentially be infectious and transmit ZIKV sexually.

Development of a Standardized Sanger-Based Method for Partial Sequencing and Genotyping of Dengue Viruses.

The global expansion of dengue viruses (DENV-1 to DENV-4) has contributed to the divergence, transmission, and establishment of genetic lineages of epidemiological concern; however, tracking the phylogenetic relationships of these virus is not always possible due to the inability of standardized sequencing procedures in resource-limited public health laboratories. Consequently, public genomic data banks contain inadequate representation of geographical regions and historical periods. In order to improve detection of the DENV-1 to DENV-4 lineages, we report the development of a serotype-specific Sanger-based method standardized to sequence DENV-1 to DENV-4 directly from clinical samples using universal primers that detect most DENV genotypes. The resulting envelope protein coding sequences are analyzed for genotyping with phylogenetic methods. We evaluated the performance of this method by detecting, amplifying, and sequencing 54 contemporary DENV isolates, including 29 clinical samples, representing a variety of genotypes of epidemiological importance and global presence. All specimens were sequenced successfully and phylogenetic reconstructions resulted in the expected genotype classification. To further improve genomic surveillance in regions where dengue is endemic, this method was transferred to 16 public health laboratories in 13 Latin American countries, to date. Our objective is to provide an accessible method that facilitates the integration of genomics with dengue surveillance.

Ability To Serologically Confirm Recent Zika Virus Infection in Areas with Varying Past Incidence of Dengue Virus Infection in the United States and U.S. Territories in 2016.

Cross-reactivity within flavivirus antibody assays, produced by shared epitopes in the envelope proteins, can complicate the serological diagnosis of Zika virus (ZIKAV) infection. We assessed the utility of the plaque reduction neutralization test (PRNT) to confirm recent ZIKAV infections and rule out misleading positive immunoglobulin M (IgM) results in areas with various levels of past dengue virus (DENV) infection incidence. We reviewed PRNT results of sera collected for diagnosis of ZIKAV infection from 1 January through 31 August 2016 with positive ZIKAV IgM results, and ZIKAV and DENV PRNTs were performed. PRNT result interpretations included ZIKAV, unspecified flavivirus, DENV infection, or negative. For this analysis, ZIKAV IgM was considered false positive for samples interpreted as a DENV infection or negative. In U.S. states, 208 (27%) of 759 IgM-positive results were confirmed to be ZIKAV compared to 11 (21%) of 52 in the U.S. Virgin Islands (USVI), 15 (15%) of 103 in American Samoa, and 13 (11%) of 123 in Puerto Rico. In American Samoa and Puerto Rico, more than 80% of IgM-positive results were unspecified flavivirus infections. The false-positivity rate was 27% in U.S. states, 18% in the USVI, 2% in American Samoa, and 6% in Puerto Rico. In U.S. states, the PRNT provided a virus-specific diagnosis or ruled out infection in the majority of IgM-positive samples. Almost a third of ZIKAV IgM-positive results were not confirmed; therefore, providers and patients must understand that IgM results are preliminary. In territories with historically higher rates of DENV transmission, the PRNT usually could not differentiate between ZIKAV and DENV infections.

Diagnosis of Zika Virus Infections: Challenges and Opportunities.

Accurate diagnosis of Zika virus (ZIKV) infections has become a pressing need for the effective prevention and control of the epidemic. The findings that ZIKV infections are associated with birth defects and neurologic disease, and that the virus can be sexually transmitted, accentuate the need for accurate diagnostic testing for different applications new to the arbovirus field. Antibody response to related flaviviruses has long been known to be cross-reactive, and antibody detection of ZIKV is nonspecific in populations previously exposed to any of the four dengue viruses or West Nile virus, or vaccinated against yellow fever virus. Therefore, the diagnosis of ZIKV infections has increasingly depended on detection by nucleic acid tests. During the recent epidemic, tests authorized for emergency use have been utilized by public health laboratories and the commercial sector, but a more dependable and responsive diagnostic testing has yet to be developed.

Relative analytical sensitivity of donor nucleic acid amplification technology screening and diagnostic real-time polymerase chain reaction assays for detection of Zika virus RNA.

BACKGROUND: Zika virus (ZIKV) has spread rapidly in the Pacific and throughout the Americas and is associated with severe congenital and adult neurologic outcomes. Nucleic acid amplification technology (NAT) assays were developed for diagnostic applications and for blood donor screening on high-throughput NAT systems. We distributed blinded panels to compare the analytical performance of blood screening relative to diagnostic NAT assays. STUDY DESIGN AND METHODS: A 25-member, coded panel (11 half-log dilutions of a 2013 French Polynesia ZIKV isolate and 2015 Brazilian donor plasma implicated in transfusion transmission, and 3 negative controls) was sent to 11 laboratories that performed 17 assays with 2 to 12 replicates per panel member. Results were analyzed for the percentage reactivity at each dilution and by probit analysis to estimate the 50% and 95% limits of detection (LOD50 and LOD95 , respectively). RESULTS: Donor-screening NAT assays that process approximately 500 µL of plasma into amplification reactions were comparable in sensitivity (LOD50 and LOD95 , 2.5 and 15-18 copies/mL) and were approximately 10-fold to 100-fold more sensitive than research laboratory-developed and diagnostic reverse transcriptase-polymerase chain reaction tests that process from 10 to 30 µL of plasma per amplification. Increasing sample input volume assayed with the Centers for Disease Control and Prevention reverse transcriptase-polymerase chain reaction assays increased the LODs by 10-fold to 30-fold. CONCLUSIONS: Blood donor-screening ZIKV NAT assays demonstrate similar excellent sensitivities to assays currently used for screening for transfusion-transmitted viruses and are substantially more sensitive than most other laboratory-developed and diagnostic ZIKV reverse transcriptase-polymerase chain reaction assays. Enhancing sensitivities of laboratory-developed and diagnostic assays may be achievable by increasing sample input.