Dengue NS1 Antigen Research Articles

Exploring Potential Application of Anti-Dengue NS1 Human Polyclonal Antibodies for Detection of Dengue Virus Infection

Dengue, a vector-borne disease, affects nearly 400 million people annually. Although commercially available dengue NS1 antigen-based ELISA kits are simple and rapid, they are expensive as monoclonal antibodies are used in these tests, and also, they have short expiry dates. As an alternative, the polyclonal antibodies generated against dengue NS1 antigen from the individuals who recovered from the dengue infection (human polyclonal antibodies against dengue; HuPA-D) can be explored for the detection of NS1 antigen in the dengue virus (DENV) infected patients’ sera. In this study, blood samples were collected from the dengue-recovered patients after obtaining the IHEC approval. The anti-NS1 HuPA-D (IgG) was purified using NAb™ Spin Column kit and tested on SDS-PAGE. HuPA-D ELISA was developed to test the sensitivity and specificity of the antibodies using the recombinant NS1 antigens of dengue serotypes and flaviviruses. Further, the HuPA-D were used to detect the sera of the dengue patients. The assay was found to be sensitive to detect all the serotypes of recombinant dengue NS1 antigen and also NS1 antigen from the sera of DENV infected patients.

Simultaneous co-circulation of two genotypes of dengue virus serotype 3 causing a large outbreak in Sri Lanka in year 2023.

We observed a discrepancy between dengue NS1 antigen test and molecular diagnostics, with the emergence of (DENV) serotype 3 in Sri Lanka and sought to understand the cause for the rise in cases and high failure rates of molecular diagnostics. Whole genomic sequencing was carried out in 22 DENV-3 samples. Phylogenetic and molecular clock analysis were done for genotype assignment and to understand the rate of evolution. Mutation analysis was done to understand the reasons for PCR non-detection. We identified two DENV-3 genotypes (I and III) co-circulating. DENV-3 genotype III strains shared a common ancestor with a sequence from India collected in 2022, while DENV-3 genotype I, was found to share a common ancestor with DENV-3 sequences from China. DENV-3 genotype III was detected by the modified CDC DENV-3 primers whereas, genotype I evaded detection due to key mutations at forward and reverse primer binding sites. We identified point mutations, C744T and A756G of the forward primer binding sites and in position G795A of the reverse primer binding sites which were not identified in DENV-3 genotype III. Furthermore, our Sri Lankan DENV-3 strains demonstrated a high root to tip ratio, compared to the previous DENV-3 sequences, indicating a high mutation rate during the points of sampling (year 2017 to 2023). The co-circulation of multiple genotypes associated with an increase in cases highlights the importance of continuous surveillance of DENVs to identify mutations resulting in non-detection by diagnostics and differences in virulence.

A-261 Outcome of a survey on dengue lateral flow immunoassay test kit reading and interpretation by medical laboratory staff in a hospital laboratory

Abstract Background Lateral flow immunoassay (LFIA) tests, first introduced in 1959, are now the mainstay of clinical laboratories. Khoo Teck Puat Hospital (KTPH) is a 795-bed acute care general hospital located in the north of Singapore. The Department of Laboratory Medicine has been using dengue LFIA test kits for the detection of dengue NS1 antigen, IgM antibodies and IgG antibodies. While kits are cost-effective, user-friendly and provide rapid results, there is subjectivity in reading and interpretation of results. Methods We conducted a survey to assess test reading and interpretation by medical laboratory staff. Six serum and plasma specimens were analysed using Bioline™ Dengue IgG/IgM WB test device. Test kits at the end-point of sample incubation were photographed in both well-lit and dim conditions. A survey questionnaire was prepared using an online form builder tool using the photographs. Staff were invited to participate in the survey. They were allowed to report the result as ‘Positive’, ‘Negative’ or ‘Equivocal’. Results A total of sixteen responses were received. Results were 100% concordant for all 6 specimens in both well-lit and dim conditions. For one case, under a well-lit condition, 15 staff reported the result as ‘Positive’ while one staff reported ‘Equivocal’. Most of the reported results for samples with no consensus is split between ‘Positive’ and ‘Equivocal’. Conclusions Dengue viruses of all 4 known serotypes are endemic in Singapore. Positive results are tracked on a national level to identify areas of high dengue infection for intensive environmental and vector control measures. Hence, accurate reporting and identification of patients with dengue infection is important. Our survey shows that lighting conditions do not affect the interpretation of LFIA results. Reading and interpretation of dengue LFIA test kits however, remain dependent on user interpretation and newer test kits with optical readers may be useful in mitigating this.

First reported cases of Borrelia and dengue co-infection: A case series.

Dengue fever is one of the most common diagnoses in patients presenting with acute febrile illness in tropical countries. Borrelia, on the other hand, is the cause of vector-borne infections of relapsing fever and Lyme disease. While co-infections of dengue with chikungunya, Zika, malaria, influenza and typhoid have been reported, clinical cases of Borrelia infections have never been reported in Malaysia. Based on available evidence, this is the first case series to report dengue fever and Borrelia spp. co-infection. All three patients in this report were admitted to medical wards on day 3 to day 8 of illness for dengue fever as evidenced by a positive dengue NS1 antigen test. The clinical manifestations were typical of dengue fever, with all patients having high grade fever, myalgia, and arthralgia. All patients also had thrombocytopenia. Features of severe dengue, such as shock, hemorrhage and impaired consciousness were absent. Borrelia DNA were detected in the blood samples of the patients. However, all the patients denied having skin lesions and a history of insect bites. All the patients were given intravenous fluid therapy and were discharged after 3 to 5 days of hospitalization.

Dengue NS1 interaction with lipids alters its pathogenic effects on monocyte derived macrophages

BackgroundWhile dengue NS1 antigen has been shown to be associated with disease pathogenesis in some studies, it has not been linked in other studies, with the reasons remaining unclear. NS1 antigen levels in acute dengue are often associated with increased disease severity, but there has been a wide variation in results based on past dengue infection and infecting dengue virus (DENV) serotype. As NS1 engages with many host lipids, we hypothesize that the type of NS1-lipid interactions alters its pathogenicity.MethodsPrimary human monocyte derived macrophages (MDMs) were co-cultured with NS1 alone or with HDL, LDL, LPS and/or platelet activating factor (PAF) from individuals with a history of past dengue fever (DF = 8) or dengue haemorrhagic fever (DHF = 8). IL-1β levels were measured in culture supernatants, and gene expression analysis carried out in MDMs. Monocyte subpopulations were assessed by flow cytometry. Hierarchical cluster analysis with Euclidean distance calculations were used to differentiate clusters. Differentially expressed variables were extracted and a classifier model was developed to differentiate between past DF and DHF.ResultsSignificantly higher levels of IL-1β were seen in culture supernatants when NS1 was co-cultured with LDL (p = 0.01, median = 45.69 pg/ml), but lower levels when NS1 was co-cultured with HDL (p = 0.05, median = 4.617 pg/ml). MDMs of those with past DHF produced higher levels of IL-1β when NS1 was co-cultured with PAF (p = 0.02). MDMs of individuals with past DHF, were significantly more likely to down-regulate RPLP2 gene expression when macrophages were co-cultured with either PAF alone, or NS1 combined with PAF, or NS1 combined with LDL. When NS1 was co-cultured with PAF, HDL or LDL two clusters were detected based on IL10 expression, but these did not differentiate those with past DF or DHF.ConclusionsAs RPLP2 is important in DENV replication, regulating cellular stress responses and immune responses and IL-10 is associated with severe disease, it would be important to further explore how differential expression of RPLP2 and IL-10 could lead to disease pathogenesis based on NS1 and lipid interactions.

Beyond the Bite: Detailed findings on Chikungunya and Dengue co-detection in Punjab, North India - clinical insights and diagnostic challenges.

The co-circulation of Chikungunya virus (CHIKV) and Dengue virus (DENV) in India poses a challenge for the diagnosing clinician, as they share similar clinical signs and symptoms and geographical distribution. Both arthropod-borne viruses are maintained in the environment by the Aedes mosquito, commonly found in tropical countries including India. Here we aim to investigate the clinical and laboratory aspects of Chikungunya/Dengue suspected cases in Punjab, India during 2021-2022, focusing on the differential diagnosis of Dengue. All suspected cases were submitted to serological differential diagnosis approaches to arboviruses like Chikungunya and Dengue. For the detection of Chikungunya Infection, CHIK IgM Capture ELISA was employed. Whereas, for Dengue NS1 antigen ELISA and IgM Capture ELISA assays were employed. A total of 370 cases suspected of arboviral infection were investigated and 38.3% (142/370) were confirmed as Chikungunya. Chikungunya cases were slightly more prevalent in males (54%) and the most frequently affected age group was adults between 16 and 30 years old (45.7%). Polyarthralgia affected 79.5% of patients, 63.3% exhibited headache and 50% presented with retro-orbital pain. 28.9% (107/370) had serological evidence of DENV exposure by detection of specific anti-DENV IgM or NS1 and 9.1% (34/370) cases of co-detection of Chikungunya and Dengue were reported. Urban populations had a higher infection rate of co-detection of Chikungunya and Dengue than rural populations with 83% versus 17%, respectively. Despite an initial clinical diagnosis of Dengue, most patients with fever and arthralgia were serologically confirmed as Chikungunya cases, with a notable prevalence of CHIKV/DENV co-detection. Strengthening differential diagnosis of circulating arboviruses is crucial for improving patient care and enhancing vector control and environmental management strategies.

Commercial Enzyme-Linked Immunosorbent Assay Kit Is Useful for Detection of Recombinant and Secretory Nonstructural-1 Protein Antigen of Dengue Virus.

Background: Dengue is a mosquito-borne tropical disease, caused by the Dengue virus (DENV). It has become a severe problem and is a rising threat to public health. In this study, we have evaluated commercial Merilisa i Dengue NS1 Antigen kit (Meril LifeSciences India Pvt. Ltd.) to detect recombinant dengue virus 2 NS1 antigen (rDNS1Ag) and secreted forms of NS1 antigen (sDNS1Ag). Methods: To determine the detection limit of the kit, 100 nanogram (ng) to 0.001 ng rDNS1Ag was tested. The sensitivity and specificity of the kit was determined using recombinant NS1 antigens of all serotypes of DENV and other flaviviruses. For testing sDNS1Ag, the culture supernatant of the Vero cell lines infected with DENV-2 was tested. Further, a spiking experiment was carried out to check the sensitivity of the kit to detect rDNS1Ag in the pools of Aedes aegypti mosquitoes. Results: It was observed that the kit can detect the rDNS1Ag at 1 ng concentration. The kit was sensitive to detect NS1 antigen of DENV-1, DENV-2 and DENV-3 serotypes and specific for detection of only DNS1Ag as it did not cross-react with NS1 antigen of flaviviruses. The kit was sensitive to detect rDNS1Ag in the mosquito pools as well. In addition, the kit was able to detect the sDNS1Ag in Vero cell culture supernatant. Conclusions: Overall, we observed that the Merilisa i Dengue NS1 Ag kit is sensitive and specific for the detection of DNS1Ag both in recombinant and secretory forms.

Serodiagnosis of Secondary Dengue Infection in a Tertiary Care Hospital, Mysuru

Dengue is one among the acute viral infections with the probability of fatal complications. In 2017; NVBDCP reported 157220 positive dengue cases with 250 deaths in India; 17018 cases and 5 deaths in Karnataka. Most primary infections are uneventful. The critical illness like Dengue Haemorrhagic Fever and Dengue Shock Syndrome are generally attributed to serotype cross-reactivity. Identification of secondary dengue infection in the early onset of illness is beneficial. Therefore, methods to discrepate primary and secondary dengue infection are of significant prognostic value. The current study is a hospital based prospective analytical evaluation and was aimed to discriminate secondary from primary dengue virus infection in clinically suspected dengue cases presenting with fever and thrombocytopenia. Patients of all age groups attending Krishna Rajendra Hospital on outpatient and inpatient basis with clinically suspected dengue fever of less than 5 days associated with thrombocytopenia were included in the study. The samples were tested in the VRDL of the Microbiology Department for dengue NS1 antigen and IgM antibodies, positive for both were further subjected to IgG antibodies. IgM /IgG ratio was used to differentiate primary and secondary dengue infections. Dengue infection was categorized based on WHO guidelines. A total of 17,841 samples were tested from May 2017 to December 2023; out of 17841 samples tested 2111 (60.74%) were positive for dengue NS1 and IgM. Of the 2111 dengue NS1 and IgM positive cases, 1700 (80.5%) were having secondary dengue infection, whereas 411 (19.46%) were having primary infection. Early detection of secondary infection helps the clinician in anticipating dengue related complications with appropriate therapeutic intervention, thereby reducing further complications and mortality.

A study on evaluation and correlation of serological methods and platelet count for diagnosis of dengue virus infection.

Dengue is arthropod-borne, acute potentially fatal viral infection, endemic in many tropical and sub-tropical countries having public health threat globally in recent era. Effective and prompt diagnosis is very essential to reduce morbidity and mortality associated with severe form. Diagnosis by dengue-specific markers like Non-Structural protein 1 antigen or IgM/IgG antibody by serological method is the choice of investigation and the most widely used non-specific biomarker is platelet count. To evaluate and correlate results of two serological diagnostic methods (Dengue NS1 Antigen and Dengue IgM antibodies by Enzyme-Linked Immunosorbent Assay) and with platelet count for diagnosis of Dengue infection. Retrospective cross-sectional study for duration of 6 months from April 2022 to September 2022. Total 4063 serum samples of dengue suspected patients were tested according to duration of illness by ELISA for presence of NS1 Ag (≤5 days of illness) or IgM antibody (>5 days of illness). 393/4063 were detected positive by either method (NS1/IgM) and majority of patients were diagnosed between 3-8 days of illness. 248/2250 were tested positive for dengue NS1-Ag only, while 145/1813 were seropositive by dengue IgM only. Thrombocytopenia (platelet counts <100000/mm3) found in 301/393 seropositive cases which was also observed in NS1 (207/248 cases) and IgM (94/145 cases) seropositive patients. Dengue specific parameters (NS1/IgM detection) in combination with platelet count helps in accurate, timely diagnosis and better monitoring in clinical management of patients.

CLINICAL PROFILE OF EXPANDED DENGUE SYNDROME IN CHILDREN.

The objective is to study the clinical profile and outcome of expanded dengue syndrome (EDS) in children. We conducted a retrospective cohort study and enrolled consecutive children (1 month to 18 years) who presented with dengue fever for over 1 year. The diagnosis of dengue fever was confirmed by the presence of dengue NS1 antigen and/or dengue IgM positivity. Subsequently, we identified children exhibiting EDS according to the revised World Health Organization guidelines (2011). We compared the clinical and laboratory profiles of children diagnosed with EDS and those without EDS. Of 178 children with dengue fever, 33 (18.5%) had EDS [95% confidence interval (CI): 13.1%-25.0%]. In these 33 children, neurological involvement was the most common manifestation [n = 16/33 (49%, 95% CI: 32%-65%)], namely encephalopathy, febrile seizures and encephalitis, respectively. Gastrointestinal involvement was the second most common [n = 10/33 (30%, 95% CI: 17%-47%)], which included fulminant hepatic failure, acalculous cholecystitis and acute pancreatitis. Renal presentation [n = 7 (21%, 95% CI: 10%-37%)] was limited to acute kidney injury. Children with EDS were significantly anemic and had leukocytosis, hyperkalemia, azotemia, hyperbilirubinemia, raised serum transaminases and fluid-refractory shock than their non-EDS counterparts. Children with EDS had a trend toward higher mortality (P = 0.07) and the survivors had a longer duration of hospital stay (5 days vs. 4 days in non-EDS, P = 0.001). In conclusion, we observed a high prevalence (18.5%) of EDS among children hospitalized for dengue fever. The common manifestations of EDS include neurological, renal and gastrointestinal involvement. Children with EDS showed a trend toward higher mortality and longer duration of hospital stay than children without EDS.

Dengue NS1 interaction with lipids alters its pathogenic effects on monocyte derived macrophages.

While dengue NS1 antigen has been shown to be associated with disease pathogenesis in some studies, it has not been linked in other studies, with the reasons remaining unclear. NS1 antigen levels in acute dengue are often associated with increased disease severity, but there have been a wide variation in results based on past dengue infection and infecting dengue virus (DENV) serotype. As NS1 engages with many host lipids, we hypothesize that the type of NS1-lipid interactions alters its pathogenicity. Primary human monocyte derived macrophages (MDMs) were co-cultured with NS1 alone or with HDL, LDL, LPS and/or platelet activating factor (PAF) from individuals with a history of past dengue fever (DF=8) or dengue haemorrhagic fever (DHF=8). IL-1β levels were measured in culture supernatants, and gene expression analysis carried out in MDMs. Monocyte subpopulations were assessed by flow cytometry. Hierarchical cluster analysis with Euclidean distance calculations were used to differentiate clusters. Differentially expressed variables were extracted and a classifier model was developed to differentiate between past DF and DHF. Significantly higher levels of IL-1β were seen in culture supernatants when NS1 was co-cultured with LDL (p=0.01), but with lower levels with HDL (p=0.05). MDMs of those past DHF produced more IL-1β when NS1 with PAF (p=0.02). MDMs of individuals with past DHF, were significantly more likely to down-regulate RPLP2 gene expression when macrophages were co-cultured with either PAF alone, or NS1 combined with PAF, or NS1 combined with LDL. When NS1 was co-cultured with PAF, HDL or LDL two clusters were detected based on IL10 expression, but these did not differentiate those with past DF or DHF. As RPLP2 is important in DENV replication and in regulating cellular stress responses and immune responses and IL-10 is associated with severe disease, it would be important to further explore how differential expression of RPLP2 and IL-10 could lead to disease pathogenesis based on NS1 and lipid interactions.

Significance of Serological Markers in Diagnosis of Acute Dengue Infection: 2021 Outbreak

Objective: To highlight the significance of Dengue anti-IgM and Dengue NS1 antigen in diagnosing Acute Dengue infection. Study Design: Cross-sectional study. Place and Duration of Study: Virology Department, Armed Forces Institute of Pathology, Rawalpindi Pakistan, from Sep to Nov 2021. Methodology: One hundred and fifty Dengue NS1-positive patients of either gender with age≥5 years were included. ELISA tested patients for detection of Dengue anti-IgM antibody. A questionnaire was used to collect demographic and personal data of each patient. Results: Out of 150 dengue NS1 positive samples, 26(17%) were positive for dengue anti-IgM. In collected samples, Dengue serological markers (NS1 and anti-IgM) positivity was observed based on ‘the day of onset of symptoms. On days 1 and 2, 54 and 44 NS1 positive samples were collected, respectively, but no one was found positive for anti-IgM. Anti-IgM was detected on day-3 and onwards. 27% Dengue anti-IgM positivity was found on day-3, which increased gradually as the time of infection increased. On day four, it was 53%; on day 5, it was 55%; on day-6, it was 60%; on day 7, it was 75%; and on day 8, it was 100%. Conclusions: The findings of present study suggested that anti-Dengue IgM antibodies are less significant than the Dengue NS1 antigen in diagnosing acute dengue infection.

Investigating the Clinico-Demographic Characteristics of Dengue Fever and Its Seroprevalence at a Tertiary Care Hospital in Northern India.

Background and objective Dengue virus (DENV) is a major global health threat, causing over 50,000 deaths annually. The state of Uttar Pradesh (UP) in India faces significant challenges due to the increasing number of dengue cases detected. This study aimed to assess DENV seropositivity in the Raebareli district of UP, to offer crucial insights into the region's effective control and management strategies. Materials and methods This study, after obtaining approval from the ethics committee, analyzed blood samples of individuals suspected of having dengue at a teaching hospitalin rural UP between January and December 2022. To determine the disease's seroprevalence, both dengue NS1 antigen ELISA and dengue IgM Microlisa were conducted. Furthermore, RT-PCR was performed on NS1-positive samples to confirm the serotypes. The collected data were analyzed using Epi Info 7.0. Results Of the 589 suspected dengue cases, 86 (14.60%) tested positive for dengue NS1 and/or IgM. Our findings showed that males (n=330, 56.03%)and adolescents and young adults (n=301, 51.1%)from rural areas (n=523, 88.4%) were predominantly affected. Cases peaked post-monsoon, and platelet levels were notably low in NS1-positive cases. Dengue serotype 2 (DEN-2) was found in all RT-PCR-positive samples. Our results revealed a dengue seroprevalence of 14.60% (n=86), which peaked in post-monsoon months. The higher incidence among males and young adults from rural areas attending the outpatient department highlights the importance of targeted interventions and community surveillance. RT-PCR confirmed the circulation of a single serotype in the region. Conclusions This study contributes crucial insights into dengue's epidemiology and clinical profile and its findings are all the more significant now as India prepares for phase 3 trials of a quadrivalent dengue-virus vaccine in 2024. Adolescent and young adult males have an increased likelihood of acquiring the virus, and this demographic can be prioritized for vaccine trials.

Atypical Dengue Outbreak in Odisha: Insights from the Entomological Investigations.

An outbreak of dengue fever struck Tikarapada village, Rayagada district (April 2020). Among 117 fever cases, 49 tested positive for the dengue NS1 antigen. To identify mosquito-related factors in the outbreak, a survey of 101 houses found Aedes breeding sites in 11.6% of containers. Aedes aegypti (79%) and Aedes albopictus (21%) emerged from collected pupae. The village exhibited high (house index = 24.8), (container index = 11.6), (pupal index = 32.7), and (Breteau index = 40.6) indices. The findings confirmed the presence of Aedes and ample breeding sites, suggesting their role in the outbreak. A report recommending integrated vector control measures was submitted to district and state health authorities.

Seroprevalence Study of Dengue at a Tertiary Care Hospital in Jaipur, Rajasthan

Background and Aim: Dengue is one of the most severe mosquito-borne viral infection. In the absence of specific treatments and vaccines for dengue fever (DF), vector control is the only method by which the spread of dengue can be prevented. This study investigates the seroprevalence of DF cases by detecting NS1 antigen and immunoglobulin (Ig)-G and IgM antibody by enzyme-linked immunosorbent assay (ELISA) test. Materials and Methods: This study was carried out in the Department of Microbiology, at a Tertiary Care Hospital, Jaipur (Rajasthan), done between 6 months. Blood samples from clinically suspected cases of dengue were tested immediately for qualitative detection of NS1 Ag, IgM, and IgG antibodies by ELISA. Results: One thousand seven hundred and sixty-three serum samples were tested for dengue NS1 antigen, IgM, and IgG, out of which 1066 were positive for one or more serological parameters. Dengue cases for the NS1 infection are 52.53%, for the IgM infection is 33.39%, and for the IgG infection is 14.07%. Sixty-two percentage positive samples were of male patients and 57% of positive samples were from 19 to 40 years of age group (adult population) (59%). The seasonal trend showed a gradual increase in dengue-positive cases that started in September with a peak in October (46.24%). Conclusion: The monsoon and postmonsoon seasons are times of high virus activity and enhanced vector breeding. The population must thus be constantly monitored to prevent the virus from spreading further, and serological tests are crucial for making an early diagnosis. ELISA is advised for all suspected dengue patients to initiate therapy and decrease the morbidity and fatality rate.

APRI as a predictor of severe dengue fever.

The AST/platelet ratio index (APRI) is a well-researched indicator of liver fibrosis. Some studies have shown that APRI can be used as a predictor of severe dengue, but the data is limited. As dengue epidemics are common in our country with limited healthcare resources, we believe APRI can help emergency physicians/primary physicians in predicting the severity of dengue and plan for the appropriate use of limited healthcare resources. 1) To determine the utility of APRI as a predictor of severe dengue. 2) To determine the association of APRI with length of hospital stay and platelet requirement. A retrospective cross-sectional study was done on patients presented to the Emergency Medicine department at Travancore Medicity Medical College with a positive Dengue NS1 antigen or IgM antibody. We found from the univariate analysis results that ALT > 74.5 IU/L has a sensitivity of 59.6 and a specificity of 76.3 (AUC: 0.696; 95% CI: 0.606-0.786), AST > 160.5 IU/L has a sensitivity of 42.3 and a specificity of 93.7 (AUC: 0.747; 95% CI: 0.665-0.829), and APRI > 3.2 has a sensitivity of 69.2 and a specificity of 84.2 (AUC: 0.806; 95% CI: 0.72-0.884) to predict severe dengue. Patients with an APRI of >3.2 required a mean hospital stay of 5.47 days (P = 0.005); 27 (81.8%) requiring platelet transfusion had an APRI of > 3.2 (P = 0.00). APRI is a straightforward index that can be easily derived from AST and platelet values. APRI values of >3.2 can predict severe dengue with a sensitivity of 69.2 and a specificity of 84.2. APRI values of >3.2 are also associated with the length of hospital stay and requirement of platelet transfusion.

Portable Paper-Based Immunoassay Combined with Smartphone Application for Colorimetric and Quantitative Detection of Dengue NS1 Antigen.

Dengue virus (DENV) infection, which is transmitted by Aedes mosquitoes, is a major public health concern in tropical and subtropical countries. With an annual incidence of approximately 10 million cases and 20,000-25,000 deaths, particularly among children, there is an urgent need for practical diagnostic tools. The presence of dengue non-structural protein 1 (NS1) during early infection has been linked to cytokine release, vascular leakage, and endothelial dysfunction, making it a potential marker for severe dengue. Paper-based immunoassays such as lateral flow assays (LFAs) and microfluidic paper-based analytical devices (PADs) have gained popularity as diagnostic tests due to their simplicity, rapidity, inexpensiveness, specificity, and ease of interpretation. However, conventional paper-based immunoassays for dengue NS1 detection typically rely on visual inspection, yielding only qualitative results. To address this limitation and enhance sensitivity, we proposed a highly portable NS1 dengue detection assay on a Paper-based Analytical Device (PAD), namely, DEN-NS1-PAD, that integrates a smartphone application as a colorimetric and quantitative reader. The development system enables direct quantification of NS1 concentrations in clinical samples. Serum and blood samples obtained from patients were utilized to demonstrate the system prototype performance. The results were obtained immediately and can be employed for clinical assessment, both in well-equipped healthcare facilities and resource-limited settings. This innovative combination of a paper-based immunoassay with a smartphone application offers a promising approach for enhanced detection and quantification of dengue NS1 antigen. By augmenting sensitivity beyond the capabilities of the naked eye, this system holds great potential for improving clinical decision-making in dengue management, particularly in remote or underserved areas.

Seroprevalence and Co-infections of Different Infectious Aetiologies in Blood Culture Negative Febrile Patients Seeking Healthcare at a University Hospital in Northern India: A Cross-sectional Study

Introduction: The burden of acute febrile illness remains underestimated in many low- and middle-income countries. The rationale of this study revolves around the gaps in diagnostics for acute febrile illnesses in these regions. Aim: To highlight the seroprevalence and co-infections of different infectious aetiologies in blood culture negative febrile patients. Materials and Methods: This cross-sectional study was conducted at Sanjay Gandhi Post Graduate Institute of Medical Sciences, Lucknow, Uttar Pradesh, India from October 2021 to July 2022, involving 153 serum samples from patients presenting with fever for less than two weeks duration during outpatient and emergency visits. The blood culture negative samples were further tested for other aetiologies such as Leptospirosis, Dengue NS1 antigen, Dengue IgM antibody, Chikungunya, Scrub typhus IgM, and Widal, respectively. Convalescent sera were also tested for all positive Widal results. All statistical analyses were performed using Statistical Package for the Social Sciences (SPSS) statistical software (IBM SPSS version 26.0, Armonk, N.Y.). Results: Out of the 153 patients enrolled, 120 patients were tested for a panel of serological tests. The majority 52 (43.3%) were aged between 21 and 40 years. Fifty-five patients (45.8%) exhibited positivity, among which Scrub typhus was the most common aetiology with 21 (17.5%) case, followed by Leptospirosis 19 (15.8%), Dengue 10 (8.3%), Chikungunya 3 (2.5%), and Enteric fever 2 (1.7%), with no cases of malaria reported. Overall, co-infections reported in present study included 7 (5.8%) cases, of which scrub typhus and Leptospirosis were the most prevalent in 5 (4.16%) cases. Two patients (1.6%) presented with a co-infection of dengue and chikungunya. Conclusion: The diverse aetiologies causing acute febrile illness necessitate a syndrome-based disease surveillance approach, accompanied by strong clinical acumen, for the betterment of patient outcomes.

Isolation and molecular detection of dengue and chikungunya virus from field-collected adult mosquitoes in Kelantan, Malaysia

Background &amp; objectives: Dengue and chikungunya infections are one of the major health problems that have plagued the human population globally. All dengue virus (DENV) serotypes circulate within Malaysia with particular serotypes dominating in different years/outbreaks. In the state of Kelantan, an increasing number of DENV and chikungunya virus (CHIKV) new cases have been reported, including several deaths. This study aimed to isolate and detect these arboviruses from adult mosquitoes in Kelantan. Methods: Adult mo squito samples were collected from January to August 2019 and were identified according to gender, species and locality. The isolation of the virus was done in C6/36 cells. Dengue NS1 antigen was carried out using direct mosquito lysate and mosquito culture supernatant. Detection and serotyping of the DENV was performed using multiplex RT-PCR and CHIKV detection using a one-step RT-PCR assay. Results: Of 91 mosquito pools, four were positive for NS1 antigen comprising two pools (2.2%) of male Ae. albopictus (Pulau Melaka and Kubang Siput) and two pools (2.2%) of Ae. aegypti (Kampung Demit Sungai). DENV 1 was detected in one pool (0.9%) of female Ae. albopictus among 114 tested Aedes pools. Two pools of 114 pools (1.7%) from both male Aedes species were positive with double serotypes, DENV 1 and DENV 2 (Pulau Melaka). However, no pool was positive for CHIKV. Interpretation &amp; conclusion: The presence of DENV and the main vectors of arboviruses in Kelantan are pertinent indicators of the need to improve vector controls to reduce arbovirus infections among people in the localities.

Prognosis of Dengue Fever based on Clinical Symptoms and Complete Blood Count in children aged 2-12 years

Objective: We aim to investigate the value of Complete Blood Counts (CBCs) and clinical symptomatology of patients in determining the prognosis of dengue fever in children aged 2-12 years. Study Design: Descriptive cross-sectional study. Place and Duration: This study was conducted in Pediatric Department, HBS General Hospital Islamabad, over 3 months period, from 15th November 22, till 15th February 23. Methodology: Patients who tested positive for either NS-1 antigen or dengue IgM antibody were enrolled in study. Demographic details of patients, clinical and hematological parameters, and daily progress of fever, serial CBCs and duration of hospital stay were recorded on a proforma. Data was analyzed using SPSS 22.00. Percentages of hematological and clinical parameters, average duration of fever and hospital stay were calculated. Results: Dengue NS-1 antigen was positive in 94% of patients. Fever was seen in 88%, thrombocytopenia and leucopenia in 88% and 53% cases respectively. Thrombocytopenia and leucopenia improved within 5 days of admission in majority of patients. Average duration of improvement of symptoms and hospital stay was 3.5 days. A weak association between duration of hospital stay and lymphocyte count at admission was observed. Conclusion: Dengue fever is a mild febrile illness in our study population. Thrombocytopenia was seen in majority of patients, yet none of them developed dengue hemorrhagic fever. Clinical symptomatology and complete blood count can act as monitoring tools in these patients. Majority of patients showed improvement in their clinical symptoms and hematological abnormalities within a week time and were discharged with follow up advised. Keywords: Complete blood count, Dengue fever, Hematological abnormalities, Leucopenia, Prognosis, Thrombocytopenia. How to Cite This: Qayyum H, Rabia M, Sohail S, Zafar T. Prognosis of Dengue Fever based on Clinical Symptoms and Complete Blood Count in children aged 2-12 years. Isra Med J. 2023; 15(2): 51-55. DOI: https://doi.org/10.55282/imj.oa1363