Incomplete Protection against Dengue Virus Type 2 Re-infection in Peru

Background

Nearly half of the world’s population is at risk for dengue, yet no licensed vaccine or anti-viral drug is currently available. Dengue is caused by any of four dengue virus serotypes (DENV-1 through DENV-4), and infection by a DENV serotype is assumed to provide life-long protection against re-infection by that serotype. We investigated the validity of this fundamental assumption during a large dengue epidemic caused by DENV-2 in Iquitos, Peru, in 2010–2011, 15 years after the first outbreak of DENV-2 in the region.

Methodology/Principal Findings

We estimated the age-dependent prevalence of serotype-specific DENV antibodies from longitudinal cohort studies conducted between 1993 and 2010. During the 2010–2011 epidemic, active dengue cases were identified through active community- and clinic-based febrile surveillance studies, and acute inapparent DENV infections were identified through contact tracing studies. Based on the age-specific prevalence of DENV-2 neutralizing antibodies, the age distribution of DENV-2 cases was markedly older than expected. Homologous protection was estimated at 35.1% (95% confidence interval: 0%–65.2%). At the individual level, pre-existing DENV-2 antibodies were associated with an incomplete reduction in the frequency of symptoms. Among dengue cases, 43% (26/66) exhibited elevated DENV-2 neutralizing antibody titers for years prior to infection, compared with 76% (13/17) of inapparent infections (age-adjusted odds ratio: 4.2; 95% confidence interval: 1.1–17.7).

Conclusions/Significance

Our data indicate that protection from homologous DENV re-infection may be incomplete in some circumstances, which provides context for the limited vaccine efficacy against DENV-2 in recent trials. Further studies are warranted to confirm this phenomenon and to evaluate the potential role of incomplete homologous protection in DENV transmission dynamics.     

Long-Term and Seasonal Dynamics of Dengue in Iquitos,Peru

Introduction

Long-term disease surveillance data provide a basis for studying drivers of pathogen transmission dynamics. Dengue is a mosquito-borne disease caused by four distinct, but related, viruses (DENV-1-4) that potentially affect over half the world''s population. Dengue incidence varies seasonally and on longer time scales, presumably driven by the interaction of climate and host susceptibility. Precise understanding of dengue dynamics is constrained, however, by the relative paucity of laboratory-confirmed longitudinal data.

Methods

We studied 10 years (2000–2010) of laboratory-confirmed, clinic-based surveillance data collected in Iquitos, Peru. We characterized inter and intra-annual patterns of dengue dynamics on a weekly time scale using wavelet analysis. We explored the relationships of case counts to climatic variables with cross-correlation maps on annual and trimester bases.

Findings

Transmission was dominated by single serotypes, first DENV-3 (2001–2007) then DENV-4 (2008–2010). After 2003, incidence fluctuated inter-annually with outbreaks usually occurring between October and April. We detected a strong positive autocorrelation in case counts at a lag of ∼70 weeks, indicating a shift in the timing of peak incidence year-to-year. All climatic variables showed modest seasonality and correlated weakly with the number of reported dengue cases across a range of time lags. Cases were reduced after citywide insecticide fumigation if conducted early in the transmission season.

Conclusions

Dengue case counts peaked seasonally despite limited intra-annual variation in climate conditions. Contrary to expectations for this mosquito-borne disease, no climatic variable considered exhibited a strong relationship with transmission. Vector control operations did, however, appear to have a significant impact on transmission some years. Our results indicate that a complicated interplay of factors underlie DENV transmission in contexts such as Iquitos.     

Dengue infection in children in ratchaburi, Thailand: a cohort study. I. Epidemiology of symptomatic acute dengue infection in children, 2006-2009

Background

There is an urgent need to field test dengue vaccines to determine their role in the control of the disease. Our aims were to study dengue epidemiology and prepare the site for a dengue vaccine efficacy trial.

Methods and Findings

We performed a prospective cohort study of children in primary schools in central Thailand from 2006 through 2009. We assessed the epidemiology of dengue by active fever surveillance for acute febrile illness as detected by school absenteeism and telephone contact of parents, and dengue diagnostic testing. Dengue accounted for 394 (6.74%) of the 5,842 febrile cases identified in 2882, 3104, 2717 and 2312 student person-years over the four years, respectively. Dengue incidence was 1.77% in 2006, 3.58% in 2007, 5.74% in 2008 and 3.29% in 2009. Mean dengue incidence over the 4 years was 3.6%. Dengue virus (DENV) types were determined in 333 (84.5%) of positive specimens; DENV serotype 1 (DENV-1) was the most common (43%), followed by DENV-2 (29%), DENV-3 (20%) and DENV-4 (8%). Disease severity ranged from dengue hemorrhagic fever (DHF) in 42 (10.5%) cases, dengue fever (DF) in 142 (35.5%) cases and undifferentiated fever (UF) in 210 (52.5%) cases. All four DENV serotypes were involved in all disease severity. A majority of cases had secondary DENV infection, 95% in DHF, 88.7% in DF and 81.9% in UF. Two DHF (0.5%) cases had primary DENV-3 infection.

Conclusion

The results illustrate the high incidence of dengue with all four DENV serotypes in primary school children, with approximately 50% of disease manifesting as mild clinical symptoms of UF, not meeting the 1997 WHO criteria for dengue. Severe disease (DHF) occurred in one tenth of cases. Data of this type are required for clinical trials to evaluate the efficacy of dengue vaccines in large scale clinical trials.     

Evaluation of Dengue NS1 Antigen Rapid Tests and ELISA Kits Using Clinical Samples

Background

Early diagnosis of dengue virus (DENV) infection can improve clinical outcomes by ensuring close follow-up, initiating appropriate supportive therapies and raising awareness to the potential of hemorrhage or shock. Non-structural glycoprotein-1 (NS1) has proven to be a useful biomarker for early diagnosis of dengue. A number of rapid diagnostic tests (RDTs) and enzyme-linked immunosorbent assays (ELISAs) targeting NS1 antigen (Ag) are now commercially available. Here we evaluated these tests using a well-characterized panel of clinical samples to determine their effectiveness for early diagnosis.

Methodology/Principal Findings

Retrospective samples from South America were used to evaluate the following tests: (i) “Dengue NS1 Ag STRIP” and (ii) “Platelia Dengue NS1 Ag ELISA” (Bio-Rad, France), (iii) “Dengue NS1 Detect Rapid Test (1^st Generation)” and (iv) “DENV Detect NS1 ELISA” (InBios International, United States), (v) “Panbio Dengue Early Rapid (1^st generation)” (vi) “Panbio Dengue Early ELISA (2^nd generation)” and (vii) “SD Bioline Dengue NS1 Ag Rapid Test” (Alere, United States). Overall, the sensitivity of the RDTs ranged from 71.9%–79.1% while the sensitivity of the ELISAs varied between 85.6–95.9%, using virus isolation as the reference method. Most tests had lower sensitivity for DENV-4 relative to the other three serotypes, were less sensitive in detecting secondary infections, and appeared to be most sensitive on Day 3–4 post symptom onset. The specificity of all evaluated tests ranged from 95%–100%.

Conclusions

ELISAs had greater overall sensitivity than RDTs. In conjunction with other parameters, the performance data can help determine which dengue diagnostics should be used during the first few days of illness, when the patients are most likely to present to a clinic seeking care.     

HLA-A2 and B35 Restricted Hantaan Virus Nucleoprotein CD8+ T-Cell Epitope-Specific Immune Response Correlates with Milder Disease in Hemorrhagic Fever with Renal Syndrome

Background

Hantaan virus (HTNV) infection in humans is a serious public health concern in Asia. A potent T cell activation peptide vaccine from HTNV structure protein represents a promising immunotherapy for disease control. However, the T cell epitopes of the HTNV restricted by the HLA alleles and the role of epitope-specific T cell response after HTNV infection remain largely unexplored.

Methodology/Principal Findings

Five well-conserved novel CD8⁺ T-cell epitopes of the HTNV nucleoprotein restricted by the most popular HLA alleles in Chinese Han population were defined with interferon-γ enzyme-linked immunospot assay in 37 patients infected with HTNV during hospitalization. Two epitopes aa129–aa137 and aa131–aa139 restricted by HLA-A2 and B35, respectively, were selected to evaluate the epitope-specific CD8⁺ T-cell response. HLA-peptide pentamer complex staining showed that the frequency of single epitope-specific CD8⁺ T cell could be detected in patients (95% confidence interval for aa129–aa137: 0.080%–0.208%; for aa131–aa139: 0.030%–0.094%). The frequency of epitope-specific pentamer⁺ CD8⁺ T-cell response was much higher in mild/moderate patients than in severe/critical ones at the acute stage of the disease. Moreover, the frequency of epitope-specific CD8⁺ T cells at acute stage was inversely associated with the peak level of serum creatinine and was positively associated with the nadir platelet counts during the hospitalization. The intracellular cytokine staining and the proliferation assay showed that the effective epitope-specific CD8⁺ T cells were characterized with the production of interferon-γ, expression of CD69 and the strong capacity of proliferation.

Conclusion/Significance

The novel HLA class I restricted HTNV nucleoprotein epitopes-specific CD8⁺ T-cell responses would be closely related with the progression and the severity of the disease, which could provide the first step toward effective peptide vaccine development against HTNV infection in humans.     

The Immunodominance Change and Protection of CD4+ T-Cell Responses Elicited by an Envelope Protein Domain III-Based Tetravalent Dengue Vaccine in Mice

Dengue is the leading cause of mosquito-borne viral infections and no vaccine is available now. Envelope protein domain III (ED3) is the major target for the binding of dengue virus neutralizing antibodies; however, the ED3-specifc T-cell response is less well understood. To investigate the T-cell responses to four serotypes of dengue virus (DENV-1 to 4), we immunized mice using either a tetravalent ED3-based DNA or protein vaccine, or combined both as a DNA prime-protein boost strategy (prime-boost). A significant serotype-dependent IFN-γ or IL-4 response was observed in mice immunized with either the DNA or protein vaccine. The IFN-γ response was dominant to DENV-1 to 3, whereas the IL-4 response was dominant to DENV-4. Although the similar IgG titers for the four serotypes were observed in mice immunized with the tetravalent vaccines, the neutralizing antibody titers varied and followed the order of 2 = 3>1>4. Interestingly, the lower IFN-γ response to DENV-4 is attributable to the immunodominance change between two CD4⁺ T-cell epitopes; one T-cell epitope located at E_349-363 of DENV-1 to 3 was more immunogenic than the DENV-4 epitope E_313-327. Despite DENV-4 specific IFN-γ responses were suppressed by immunodominance change, either DENV-4-specific IFN-γ or neutralizing antibody responses were still recalled after DENV-4 challenge and contributed to virus clearance. Immunization with the prime-boost elicited both IFN-γ and neutralizing antibody responses and provided better protection than either DNA or protein immunization. Our findings shed light on how ED3-based tetravalent dengue vaccines sharpen host CD4 T-cell responses and contribute to protection against dengue virus.     

Dengue Virus Neutralizing Antibody Levels Associated with Protection from Infection in Thai Cluster Studies

Background

Long-term homologous and temporary heterologous protection from dengue virus (DENV) infection may be mediated by neutralizing antibodies. However, neutralizing antibody titers (NTs) have not been clearly associated with protection from infection.

Methodology/Principal Findings

Data from two geographic cluster studies conducted in Kamphaeng Phet, Thailand were used for this analysis. In the first study (2004–2007), cluster investigations of 100-meter radius were triggered by DENV-infected index cases from a concurrent prospective cohort. Subjects between 6 months and 15 years old were evaluated for DENV infection at days 0 and 15 by DENV PCR and IgM ELISA. In the second study (2009–2012), clusters of 200-meter radius were triggered by DENV-infected index cases admitted to the provincial hospital. Subjects of any age ≥6 months were evaluated for DENV infection at days 0 and 14. In both studies, subjects who were DENV PCR positive at day 14/15 were considered to have been “susceptible” on day 0. Comparison subjects from houses in which someone had documented DENV infection, but the subject remained DENV negative at days 0 and 14/15, were considered “non-susceptible.” Day 0 samples were presumed to be from just before virus exposure, and underwent plaque reduction neutralization testing (PRNT). Seventeen “susceptible” (six DENV-1, five DENV-2, and six DENV-4), and 32 “non-susceptible” (13 exposed to DENV-1, 10 DENV-2, and 9 DENV-4) subjects were evaluated. Comparing subjects exposed to the same serotype, receiver operating characteristic (ROC) curves identified homotypic PRNT titers of 11, 323 and 16 for DENV-1, -2 and -4, respectively, to differentiate “susceptible” from “non-susceptible” subjects.

Conclusions/Significance

PRNT titers were associated with protection from infection by DENV-1, -2 and -4. Protective NTs appeared to be serotype-dependent and may be higher for DENV-2 than other serotypes. These findings are relevant for both dengue epidemiology studies and vaccine development efforts.     

Computational Prediction and Analysis of Envelop Glycoprotein Epitopes of DENV-2 and DENV-3 Pakistani Isolates: A First Step towards Dengue Vaccine Development

Dengue fever of tropics is a mosquito transmitted devastating disease caused by dengue virus (DENV). There is no effective vaccine available, so far, against any of its four serotypes (DENV-1, DENV-2, DENV-3, and DENV-4). There is a need for the development of preventive and therapeutic vaccines against DENV to decrease the prevalence of dengue fever, especially in Pakistan. In this research, linear and conformational B-cell epitopes of envelope glycoprotein of DENV-2 and DENV-3 (the most prevalent serotypes in Pakistan) were predicted. We used Kolaskar and Tongaonkar method for linear epitope prediction, Emini’s method for surface accessibility prediction and Karplus and Schulz’s algorithm for flexibility determination. To propose three dimensional epitopes, the E proteins for both serotypes were homology modeled by using Phyre2 V 2.0 server, and ElliPro was used for the prediction of surface epitopes on their globular structure. Total 21 and 19 linear epitopes were predicted for DENV-2 and DENV-3 Pakistani isolates respectively. Whereas, 5 and 4 discontinuous epitopes were proposed for DENV-2 and DENV-3 Pakistani isolates respectively. Moreover, the values of surface accessibility, flexibility and solvent-accessibility can be helpful in analyzing vaccines against DENV-2 and DENV-3. In conclusion, the proposed continuous and discontinuous antigenic peptides can be valuable candidates for diagnostic and therapeutics of DENV.     

Rapid testing requires clinical evaluation for accurate diagnosis of dengue disease: A passive surveillance study in Southern Malaysia

BackgroundDengue fever is the most common mosquito-borne infection worldwide where an expanding surveillance and characterization of this infection are needed to better inform the healthcare system. In this surveillance-based study, we explored the prevalence and distinguishing features of dengue fever amongst febrile patients in a large community-based health facility in southern peninsular Malaysia.MethodsOver six months in 2018, we recruited 368 adults who met the WHO 2009 criteria for probable dengue infection. They underwent the following blood tests: full blood count, dengue virus (DENV) rapid diagnostic test (RDT), ELISA (dengue IgM and IgG), nested RT-PCR for dengue, multiplex qRT-PCR for Zika, Chikungunya and dengue as well as PCR tests for Leptopspira spp., Japanese encephalitis and West Nile virus.ResultsLaboratory-confirmed dengue infections (defined by positive tests in NS1, IgM, high-titre IgG or nested RT-PCR) were found in 167 (45.4%) patients. Of these 167 dengue patients, only 104 (62.3%) were positive on rapid diagnostic testing. Dengue infection was significantly associated with the following features: family or neighbours with dengue in the past week (AOR: 3.59, 95% CI:2.14–6.00, p<0.001), cutaneous rash (AOR: 3.58, 95% CI:1.77–7.23, p<0.001), increased temperature (AOR: 1.33, 95% CI:1.04–1.70, p = 0.021), leucopenia (white cell count < 4,000/μL) (AOR: 3.44, 95% CI:1.72–6.89, p<0.001) and thrombocytopenia (platelet count <150,000/μL)(AOR: 4.63, 95% CI:2.33–9.21, p<0.001). Dengue infection was negatively associated with runny nose (AOR: 0.47, 95% CI:0.29–0.78, p = 0.003) and arthralgia (AOR: 0.42, 95% CI:0.24–0.75, p = 0.004). Serotyping by nested RT-PCR revealed mostly mono-infections with DENV-2 (n = 64), DENV-1 (n = 32) and DENV-3 (n = 17); 14 co-infections occurred with DENV-1/DENV-2 (n = 13) and DENV-1/DENV-4 (n = 1). Besides dengue, none of the pathogens above were found in patients’ serum.ConclusionsAcute undifferentiated febrile infections are a diagnostic challenge for community-based clinicians. Rapid diagnostic tests are increasingly used to diagnose dengue infection but negative tests should be interpreted with caution as they fail to detect a considerable proportion of dengue infection. Certain clinical features and haematological parameters are important in the clinical diagnosis of dengue infection.     

Economic and Disease Burden of Dengue in Southeast Asia

Background

Dengue poses a substantial economic and disease burden in Southeast Asia (SEA). Quantifying this burden is critical to set policy priorities and disease-control strategies.

Methods and Findings

We estimated the economic and disease burden of dengue in 12 countries in SEA: Bhutan, Brunei, Cambodia, East-Timor, Indonesia, Laos, Malaysia, Myanmar, Philippines, Singapore, Thailand, and Viet Nam. We obtained reported cases from multiple sources—surveillance data, World Health Organization (WHO), and published studies—and adjusted for underreporting using expansion factors from previous literature. We obtained unit costs per episode through a systematic literature review, and completed missing data using linear regressions. We excluded costs such as prevention and vector control, and long-term sequelae of dengue. Over the decade of 2001–2010, we obtained an annual average of 2.9 million (m) dengue episodes and 5,906 deaths. The annual economic burden (with 95% certainty levels) was US$950m (US$610m–US$1,384m) or about US$1.65 (US$1.06–US$2.41) per capita. The annual number of disability-adjusted life years (DALYs), based on the original 1994 definition, was 214,000 (120,000–299,000), which is equivalent to 372 (210–520) DALYs per million inhabitants.

Conclusion

Dengue poses a substantial economic and disease burden in SEA with a DALY burden per million inhabitants in the region. This burden is higher than that of 17 other conditions, including Japanese encephalitis, upper respiratory infections, and hepatitis B.     

Laboratory Surveillance of Dengue in Rio Grande do Sul,Brazil, from 2007 to 2013

Background

According to official records, dengue was introduced in Brazil in the 80''s; since then several epidemics have occurred. Meanwhile, in Rio Grande do Sul (RS, Southern Brazil) the first autochthonous case occurred only in 2007.

Methodology and Principal Findings

In this study we report laboratory surveillance of dengue cases and seasonality of positive cases, describe serotypes and characterize the epidemiological pattern of dengue in RS from 2007 to 2013. A total of 9,779 serum samples from patients with suspected dengue fever were collected and submitted to molecular and/or serological analyses for dengue virus identification and serotyping, based on viral isolation, NS1 antigen detection and qRT-PCR, or Dengue IgM capture ELISA and MAC-ELISA. The first autochthonous dengue case in RS was confirmed in 2007 (DENV-3). While in 2008 and 2009 only imported cases were registered, autochthonous infection waves have been occurring since 2010. The highest number of dengue infections occurred in 2010, with DENV-1 and DENV-2 outbreaks in Northwestern RS. In 2011, another DENV-1 and DENV-2 outbreak occurred in the Northwestern region; moreover, DENV-4 was detected in travelers. In 2012, DENV-1 and DENV-4 co-circulated. DENV-2 circulation was only detected again in 2013, in high frequency (56.7%), co-circulating with DENV-4 (35%). Most infections occur in adults during summer. Differences in prevalence between genders were observed in 2007 (60% females), 2008 (60.8% males) and 2009 (77.5% males).

Conclusions

According to results of dengue surveillance, there was an increase in the number of dengue cases in RS and of cities infested with Aedes aegypti, possibly as a consequence of introduction of new serotypes and the difficulty of health programs to control the vector.     

Dengue 1 Diversity and Microevolution,French Polynesia 2001–2006: Connection with Epidemiology and Clinics

Background

Dengue fever (DF) is an emerging infectious disease in the tropics and subtropics. Determinants of DF epidemiology and factors involved in severe cases—dengue haemorrhagic fever (DHF) and dengue shock syndrome (DSS)—remain imperfectly characterized. Since 2000, serotype 1 (DENV-1) has predominated in the South Pacific. The aim of this study was (i) to determine the origin and (ii) to study the evolutionary relationships of DENV-1 viruses that have circulated in French Polynesia (FP) from the severe 2001 outbreak to the recent 2006 epidemic, and (iii) to analyse the viral intra-host genetic diversity according to clinical presentation.

Methodology/Principal Findings

Sequences of 181 envelope gene and 12 complete polyproteins of DENV-1 viruses obtained from human sera in FP during the 2001–2006 period were generated. Phylogenetic analysis showed that all DENV-1 FP strains belonged to genotype IV–“South Pacific” and derived from a single introduction event from South-East Asia followed by a 6-year in situ evolution. Although the ratio of nonsynonymous/synonymous substitutions per site indicated strong negative selection, a mutation in the envelope glycoprotein (S222T) appeared in 2002 and was subsequently fixed. It was noted that genetic diversification was very significant during the 2002–2005 period of endemic DENV-1 circulation. For nine DF sera and eight DHF/DSS sera, approximately 40 clones/serum of partial envelope gene were sequenced. Importantly, analysis revealed that the intra-host genetic diversity was significantly lower in severe cases than in classical DF.

Conclusions/Significance

First, this study showed that DENV-1 epidemiology in FP was different from that described in other South-Pacific islands, characterized by a long sustained viral circulation and the absence of new viral introduction over a 6-year period. Second, a significant part of DENV-1 evolution was observed during the endemic period characterized by the rapid fixation of S222T in the envelope protein that may reflect genetic drift or adaptation to the mosquito vector. Third, for the first time, it is suggested that clinical outcome may be correlated with intra-host genetic diversity.     

Flavonoid from Carica papaya inhibits NS2B-NS3 protease and prevents Dengue 2 viral assembly

Dengue virus belongs to the virus family Flaviviridae. Dengue hemorrhagic disease caused by dengue virus is a public health problem worldwide. The viral non structural 2B and 3 (NS2B-NS3) protease complex is crucial for virus replication and hence, it is considered to be a good anti-viral target. Leaf extracts from Carica papaya is generally prescribed for patients with dengue fever, but there are no scientific evidences for its anti-dengue activity; hence we intended to investigate the anti-viral activity of compounds present in the leaves of Carica papaya against dengue 2 virus (DENV-2). We analysed the anti-dengue activities of the extracts from Carica papaya by using bioinformatics tools. Interestingly, we find the flavonoid quercetin with highest binding energy against NS2B-NS3 protease which is evident by the formation of six hydrogen bonds with the amino acid residues at the binding site of the receptor. Our results suggest that the flavonoids from Carica papaya have significant anti-dengue activities.

Abbreviations

ADME - Absorption, distribution, metabolism and excretion, BBB - Blood brain barrier, CYP - Cytochrome P450, DENV - – Dengue virus, DHF - Dengue hemorrhagic fever, DSS - Dengue shock syndrome, GCMS - – Gas chromatography- Mass spectrometry, MOLCAD - Molecular Computer Aided Design, NS - Non structural, PDB - Protein data bank, PMF - Potential Mean Force.     

Comparison of Three Commercially Available Dengue NS1 Antigen Capture Assays for Acute Diagnosis of Dengue in Brazil

Background

Dengue is associated with explosive urban epidemics and has become a major public health problem in many tropical developing countries, including Brazil. The laboratory diagnosis of dengue can be carried out using several approaches, however sensitive and specific assays useful to diagnose in the early stage of fever are desirable. The flavivirus non-structural protein NS1, a highly conserved and secreted glycoprotein, is a candidate protein for rapid diagnosis of dengue in endemic countries.

Methodology/Principal Findings

We aimed to evaluate the potential use of 3 commercial kits in a panel of 450 serum samples for early diagnosis of dengue in Brazil. The PanBio Early ELISA (PanBio Diagnostics) showed a sensitivity of 72.3% (159/220) and a specificity of 100%, while the sensitivity of the Platelia™ NS1 assay (Biorad Laboratories) was 83.6% (184/220). However, the highest sensitivity (89.6%; 197/220) was obtained by using the NS1 Ag Strip (Biorad Laboratories). A lower sensitivity was observed in DENV-3 cases by all 3 kits. Serum positive by virus isolation were more often positive than cases positive by RT-PCR by all three assays and a higher detection rate was observed during the first four days after the onset of the symptoms. The presence or absence of IgM showed no influence in the confirmation by the pan-E Early ELISA (P = 0,6159). However, a higher confirmation by both Platelia™ NS1 (Biorad) and Dengue NS1 Ag Strip (Biorad) in the absence of IgM was statistically significant (P<0,0001 and P = 0,0008, respectively). Only the Platelia™ NS1 test showed a higher sensitivity in confirming primary infections than secondary ones.

Conclusions/Significance

The results indicate that commercial kits of dengue NS1 antigen are useful for the laboratory diagnosis of acute primary and secondary dengue. It can be used in combination with the MAC-ELISA for case detection and as screening test to complement viral isolation.     

Laboratory-Enhanced Dengue Sentinel Surveillance in Colombo District,Sri Lanka: 2012-2014

Introduction

Dengue has emerged as a significant public health problem in Sri Lanka. Historically surveillance was passive, with mandatory dengue notifications based on clinical diagnosis with only limited laboratory confirmation. To obtain more accurate data on the disease burden of dengue, we set up a laboratory-based enhanced sentinel surveillance system in Colombo District. Here we describe the study design and report our findings of enhanced surveillance in the years 2012–2014.

Methods

Three outpatient clinics and three government hospitals in Colombo District that covered most of the Colombo metropolitan area were selected for the sentinel surveillance system. Up to 60 patients per week presenting with an undifferentiated fever were enrolled. Acute blood samples from each patient were tested by dengue specific PCR, NS1 ELISA and IgM ELISA. A sub-set of samples was sent to Duke-NUS Singapore for quality assurance, virus isolation and serotyping. Trained medical research assistants used a standardized case report form to record clinical and epidemiological data. Clinical diagnoses by the clinicians-in-charge were recorded for hospitalized cases.

Results

Of 3,127 febrile cases, 43.6% were PCR and/or NS1 positive for dengue. A high proportion of lab confirmed dengue was observed from inpatients (IPD) (53.9%) compared to outpatient (clinics in hospitals and general practice) (7.6%). Dengue hemorrhagic fever (DHF) was diagnosed in 11% of patients at the time of first contact, and the median day of illness at time of presentation to the sentinel sites was 4. Dengue serotype 1 was responsible for 85% of the cases and serotype 4 for 15%. The sensitivity and specificity of the clinicians’ presumptive diagnosis of dengue was 84% and 34%, respectively.

Conclusion

DENV-1, and to a lesser degree DENV-4, infection were responsible for a high proportion of febrile illnesses in Colombo in the years 2012 to 2014. Clinicians’ diagnoses were associated with high sensitivity, but laboratory confirmation is required to enhance specificity.     

Dengue Virus Activates Membrane TRAIL Relocalization and IFN-α Production by Human Plasmacytoid Dendritic Cells In Vitro and In Vivo

Background

Dengue displays a broad spectrum of clinical manifestations that may vary from asymptomatic to severe and even fatal features. Plasma leakage/hemorrhages can be caused by a cytokine storm induced by monocytes and dendritic cells during dengue virus (DENV) replication. Plasmacytoid dendritic cells (pDCs) are innate immune cells and in response to virus exposure secrete IFN-α and express membrane TRAIL (mTRAIL). We aimed to characterize pDC activation in dengue patients and their function under DENV-2 stimulation in vitro.

Methods & Findings

Flow cytometry analysis (FCA) revealed that pDCs of mild dengue patients exhibit significantly higher frequencies of mTRAIL compared to severe cases or healthy controls. Plasma levels of IFN-α and soluble TRAIL are increased in mild compared to severe dengue patients, positively correlating with pDC activation. FCA experiments showed that in vitro exposure to DENV-2 induced mTRAIL expression on pDC. Furthermore, three dimension microscopy highlighted that TRAIL was relocalized from intracellular compartment to plasma membrane. Chloroquine treatment inhibited DENV-2-induced mTRAIL relocalization and IFN-α production by pDC. Endosomal viral degradation blockade by chloroquine allowed viral antigens detection inside pDCs. All those data are in favor of endocytosis pathway activation by DENV-2 in pDC. Coculture of pDC/DENV-2-infected monocytes revealed a dramatic decrease of antigen detection by FCA. This viral antigens reduction in monocytes was also observed after exogenous IFN-α treatment. Thus, pDC effect on viral load reduction was mainly dependent on IFN-α production

Conclusions

This investigation characterizes, during DENV-2 infection, activation of pDCs in vivo and their antiviral role in vitro. Thus, we propose TRAIL-expressing pDCs may have an important role in the outcome of disease.     

Spatial and Temporal Patterns of Dengue Transmission along a Red Sea Coastline: A Longitudinal Entomological and Serological Survey in Port Sudan City

Background

Dengue is an emerging health problem in several coastlines along the Red Sea. The objective of the present work is to elucidate spatial and temporal patterns of dengue transmission in Port Sudan.

Methods/Findings

A longitudinal study with three cross-sectional surveys was carried out in upper, middle and lower class neighborhoods, from November 2008 to October 2009. Monthly, entomological surveys were followed by serological surveys in dengue vector-positive houses. Meteorological records were obtained from two weather stations in the city during the same time. Overall, 2825 houses were inspected. Aedes aegypti represented 65% (35,714/54,944) and 68% (2526/3715) of the collected larvae and pupae, respectively. Out of 4640 drinking water containers, 2297 were positive for Ae. aegypti. Clay-pots “Zeirr” followed by plastic barrels were key productive containers for pupae of dengue vector, 63% (n = 3959) and 26% (n = 1651), respectively. A total of 791 blood samples were tested using PanBio Capture/Indirect IgM ELISA. Overall, the sero-prevalence rate of dengue ranged between 3%–8% (41/791), compared to an incidence of 29–40 new cases per 10,000 (193/54886) in the same examined population. Lower and middle class neighborhoods had higher entomological indices compared with upper class ones (p<0.001). Although, dengue incidence rate was significantly lower in the middle and lower class neighborhoods (F = 73.97, d.f. = 2, p<0.001), no difference in IgM prevalence was shown. The city is subject to two transmission peaks in the winter (i.e. November–January), and summer (i.e. June–August). The serological peaks of dengue are preceded by entomological peaks that occur before the onset of winter (November) and summer (March) respectively.

Conclusion

Dengue incidence is heterogeneously distributed across the neighborhoods of Port Sudan and exhibits a bi-cyclic intra-annual pattern. Hence, it should be feasible to carry out timely vector control measures to prevent or reduce dengue transmission.