Seroprevalence and clinical profile of dengue infection among acute febrile patients at a rural tertiary care hospital

Abstract

BACKGROUND

Dengue fever remains a major cause of acute febrile illness in India, particularly in rural areas where diagnostic facilities are limited. Early detection using point-of-care tests helps guide timely management and reduce unnecessary antibiotic use.

AIM

To determine the seroprevalence and clinical profile of dengue infection among acute febrile patients attending a rural tertiary care hospital in Maharashtra.

METHODS

A prospective cross-sectional study was conducted at Ashwini Rural Medical College Hospital and Research Centre, Solapur from May to October 2025. A total of 320 consecutive patients presenting with acute fever of ≤ 7 days duration prior to hospital presentation were enrolled. Patients of all age groups were included. Serum samples were tested for dengue NS1 antigen and anti-dengue IgM and IgG antibodies using a commercial rapid test (Manufacturer: Meril diagnostics; Meriscreen dengue NS1 antigen and IgM + IgG antibodies). A 10% subset (n = 32) was tested by enzyme-linked immunosorbent assay (ELISA) for quality assurance. Patients were classified as recent dengue (NS1+ and/or IgM+), probable secondary infection (IgM+ IgG+), past exposure (IgG+ IgM-), or negative (NS1- IgM-). Clinical data and outcomes were recorded. Statistical analysis included χ², Mann-Whitney U, and logistic regression (α = 0.05), adjusting for age, sex, platelet count, total leukocyte count, and presence of rash.

RESULTS

Overall, 28% (90/320) had evidence of recent dengue, while 35% (112/320) showed IgG positivity, and 10% (32/320) had probable secondary infection. Median platelet count was significantly lower in the recent dengue group [78000/mm³; interquartile range (IQR): 55000-110000] compared with non-dengue patients (210000/mm³; IQR: 160000-240000; P < 0.001). The most common symptoms were headache (68%), myalgia (61%), and retro-orbital pain (44%). Antibiotic use prior to testing was 60%; among those diagnosed with dengue, a relative 25% reduction in antibiotic prescriptions was observed following laboratory confirmation. Quality assurance analysis showed rapid test sensitivity of 92%, specificity of 95%, and Cohen’s kappa of 0.85 vs ELISA. Logistic regression identified thrombocytopenia (< 100000/mm³) [adjust odds ratio (aOR) 5.2; 95%CI: 2.4-10.9] and rash (aOR 3.6; 95%CI: 1.8-7.0) as independent predictors of dengue positivity.

CONCLUSION

A substantial proportion of acute febrile cases in this rural setting were due to dengue, highlighting the utility of NS1/IgM rapid tests for early diagnosis where molecular facilities are unavailable. Integrating rapid dengue testing into routine fever evaluation can improve case detection and rationalize antibiotic use.

Key Words: Dengue fever; Seroprevalence; NS1 antigen; IgM/IgG antibodies; Rural hospital; Rapid test; Platelet count

Core Tip: This prospective study from a rural tertiary hospital in Maharashtra found that nearly one-third of acute febrile illnesses were due to dengue infection. Rapid NS1/IgM testing showed high diagnostic accuracy (sensitivity 92%, specificity 95%) and led to a clinically meaningful reduction in unnecessary antibiotic use. Incorporating such point-of-care diagnostics in rural healthcare can enable early dengue detection, improve patient management, and strengthen antimicrobial stewardship.

INTRODUCTION

Dengue fever, caused by Dengue virus (DENV) and transmitted by Aedes mosquitoes, continues to pose a significant public health burden in India[1,2]. The disease manifests as a wide clinical spectrum ranging from self-limiting fever to severe dengue hemorrhagic fever and shock[3,4]. Early and accurate diagnosis is critical for patient management and outbreak control.

In rural India, limited access to advanced molecular diagnostics such as reverse transcription polymerase chain reaction (RT-PCR) challenges early case confirmation. Point-of-care rapid tests detecting NS1 antigen and anti-dengue IgM/IgG antibodies are valuable tools in these settings[5,6]. The NS1 antigen is detectable from day 1 to 5 of fever, while IgM and IgG appear after day 4, facilitating differentiation of acute and secondary infections[7,8].

Previous studies in urban tertiary hospitals report seroprevalence ranging between 20%-40% among febrile patients[9,10]. Most of these studies relied primarily on serological assays, including NS1 antigen detection and IgM/IgG enzyme-linked immunosorbent assay (ELISA), with limited use of molecular confirmation. However, data from rural tertiary care hospitals remain scarce. Assessing local seroprevalence and correlating it with clinical and hematological parameters is crucial for improving diagnostic strategies and antibiotic stewardship in such areas.

This study aimed to estimate the seroprevalence of dengue infection using rapid diagnostic tests among acute febrile patients at a rural tertiary care hospital and to analyze associated clinical and laboratory profiles.

MATERIALS AND METHODS

Study design and setting

A prospective cross-sectional study was carried out at Ashwini Rural Medical College Hospital and Research Centre, Solapur, Maharashtra, from May 2025 to October 2025. This 800-bed rural tertiary hospital caters to semi-urban and village populations in western Maharashtra. During the study period, dengue testing was not routinely performed for all febrile patients and was generally requested at the discretion of the treating physician; the present study implemented systematic screening of all eligible acute febrile cases.

Sample size calculation

Based on an expected dengue prevalence of 25% among acute febrile patients, with 95% confidence level (z = 1.96) and 5% absolute precision, the required sample size was 289. Allowing 10% for incomplete data, the final sample size was 320 patients.

Inclusion criteria

Patients of any age presenting with acute fever of ≤ 7 days duration prior to hospital presentation. Willingness to provide written informed consent.

Exclusion criteria

Fever of known bacterial or non-infectious etiology. Refusal to consent.

Specimen collection and testing

Venous blood (3-5 mL) was collected aseptically. Serum was separated and tested immediately using a commercial dengue rapid combo test (NS1/IgM/IgG; Manufacturer Meril diagnostics, Meriscreen Dengue NS1 Antigen and IgM + IgG Antibodies) as per the manufacturer’s instructions. Ten percent of samples (n = 32) were randomly selected and retested using ELISA for NS1 and IgM/IgG antibodies at the State Public Health Laboratory for quality assurance.

Serostatus definitions

Recent infection: NS1+ and/or IgM+.

Probable secondary infection: IgM+ and IgG+.

Past exposure: IgG+ IgM-.

Negative: NS1- IgM-.

Data collection

Clinical and demographic data were recorded in a structured proforma, including age, sex, residence, duration of fever, symptoms, laboratory parameters (complete blood cell count, platelet count, liver function tests), and outcome. Patients were also assessed for clinical or laboratory evidence suggestive of concurrent bacterial infection during hospitalization.

Statistical analysis

Data was analyzed using SPSS version 26. Descriptive statistics summarized demographic and clinical characteristics. Continuous variables were expressed as median [interquartile range (IQR)]; categorical variables as n (%). Comparisons between dengue-positive and dengue-negative groups used χ²/Fisher’s exact test for categorical variables and Mann-Whitney U test for continuous data. Logistic regression identified independent predictors of recent dengue infection, adjusting for age, sex, platelet count, total leukocyte count, and presence of rash. Statistical significance was set at P < 0.05.

Ethical considerations

The study was conducted in accordance with the ethical principles of the Declaration of Helsinki. As this was an observational study involving investigations performed as part of routine clinical care, formal Institutional Ethics Committee approval was not required. Written informed consent was obtained from all participants or their legally authorized representatives, and patient confidentiality was strictly maintained

RESULTS

Study population

A total of 320 patients were enrolled. The median age was 31 years (IQR: 22-44), and 56% were male. Most patients (62%) resided in rural villages within a 30 km radius of the hospital.

Serological profile

Among 320 patients, 90 (28%) had evidence of recent dengue infection, 32 (10%) probable secondary infection, and 112 (35%) past exposure (Table 1). NS1 antigen was detected in 64 patients (20%), and IgM antibodies in 58 (18%). Co-positivity (NS1+IgM+) occurred in 32 patients (10%).

Table 1 Serological profile of patients (n = 320), n (%).

Serological category	Criteria	n (%)
Recent dengue infection	NS1+ and/or IgM+	90 (28.1)
Probable secondary infection	IgM+ IgG+	32 (10.0)
Past exposure	IgG+ IgM-	112 (35.0)
Negative	NS1- IgM-	86 (26.9)

Clinical features

The most frequent symptoms among dengue-positive patients were headache (68%), myalgia (61%), retro-orbital pain (44%), and rash (26%). Bleeding manifestations were seen in 8%. Median platelet count was significantly lower among dengue-positive cases (78000/mm³, IQR: 55000-110000) compared to negatives (210000/mm³, IQR: 160000-240000; P < 0.001) (Table 2).

Table 2 Clinical and laboratory features by serostatus, median (25^th-75^th percentiles).

Parameter	Recent dengue (n = 90)	Non-dengue (n = 230)	P value
Median age (years)	30 (20-42)	33 (24-45)	0.18
Male sex (%)	55.6	57.0	0.82
Headache (%)	68.0	42.0	< 0.001
Rash (%)	26.0	8.0	0.002
Bleeding signs (%)	8.0	2.0	0.04
Median platelet count (/mm³)	78000 (55000-110000)	210000 (160000-240000)	< 0.001

Antibiotic use

Overall, 60% of patients received empirical antibiotics before dengue testing. Among confirmed dengue cases, antibiotic usage declined by 25% following laboratory confirmation, indicating partial but clinically relevant rationalization of antibiotic prescribing.

Quality assurance results

Of the 32 samples tested by ELISA, the rapid test demonstrated a sensitivity of 92% and specificity of 95%, with Cohen’s kappa of 0.85, indicating excellent agreement.

Predictors of dengue infection

On multivariable logistic regression, two independent predictors of recent dengue infection were identified: Thrombocytopenia (< 100000/mm³): Adjust odds ratio (aOR) = 5.2; 95%CI: 2.4-10.9; P < 0.001. Rash: AOR = 3.6; 95%CI: 1.8-7.0; P = 0.001 (Figure 1).

Figure 1 Of 350 patients screened for eligibility, 30 were excluded due to fever duration > 7 days (n = 14), identified non-dengue etiology at presentation (n = 9), or refusal to provide consent (n = 7). The remaining 320 eligible patients were enrolled and underwent NS1/IgM/IgG testing. A quality-assurance subset (n = 32) underwent enzyme-linked immunosorbent assay confirmation. ELISA: Enzyme-linked immunosorbent assay; QA: Quality assurance.

DISCUSSION

This study demonstrates that nearly one-third of acute febrile illnesses in a rural tertiary hospital were attributable to dengue, consistent with regional studies reporting 20%-35% prevalence[11,12]. The use of NS1 and IgM rapid tests effectively identified cases during the acute phase, supporting their value where PCR is unavailable.

The predominance of young adults and males parallels findings from other Indian cohorts[13,14]. Common clinical features-headache, myalgia, retro-orbital pain-remain characteristic[15,16]. Platelet counts were significantly reduced in dengue-positive patients, reinforcing thrombocytopenia as a strong predictor. The observed association of rash and thrombocytopenia with seropositivity was statistically significant, consistent with previous literature[17,18]. Although antibiotic therapy is inherently inappropriate in uncomplicated dengue infection, this reduction reflects the real-world impact of early diagnostic confirmation on prescribing practices in resource-limited rural settings. Similar trends have been reported in endemic regions, where point-of-care dengue diagnostics contributed to reductions in unnecessary antibiotic use without compromising patient outcomes[19,20].

No laboratory-confirmed bacterial co-infections were identified among dengue-positive patients in this study. Antibiotics were continued only in a small subset of patients with clinical features suggestive of secondary bacterial infection, pending further evaluation. This underscores the importance of diagnostic stewardship alongside antimicrobial stewardship in acute febrile illness management.

Rapid tests demonstrated high sensitivity (92%) and specificity (95%) with excellent agreement (κ = 0.85) against ELISA, validating their operational reliability. However, cross-reactivity with other flaviviruses remains a potential limitation[21,22].

Strengths and limitations

Strengths include prospective design, consecutive sampling, and quality assurance via ELISA. Limitations are the lack of molecular confirmation and potential cross-reactivity in serology. Nevertheless, findings provide critical local epidemiological data and demonstrate how rapid testing can enhance case detection and antibiotic stewardship in resource-limited settings.

Limitations include the absence of molecular confirmation by RT-PCR and the inherent possibility of serological cross-reactivity. Although microbiological culture facilities were available, cultures were not systematically performed in all patients with suspected dengue, as clinical management was guided primarily by laboratory-confirmed dengue diagnosis and clinical judgment. Nevertheless, the findings offer important real-world evidence supporting the role of rapid diagnostic testing in dengue-endemic, resource-constrained settings.

Public health implications

Routine integration of NS1/IgM rapid tests into fever clinics in rural hospitals can expedite diagnosis, enable timely supportive management, and reduce antibiotic misuse. Periodic serosurveys can aid in tracking local transmission trends and guiding vector-control efforts.

CONCLUSION

A significant proportion of acute febrile illnesses in this rural tertiary hospital were due to dengue infection. NS1/IgM rapid diagnostic tests proved reliable for early diagnosis, with strong correlation to clinical and hematological findings. Implementation of point-of-care dengue testing in similar rural settings can improve diagnostic accuracy, support more rational antibiotic prescribing, and strengthen dengue surveillance and control efforts.

ACKNOWLEDGEMENTS

The authors thank the Department of Medicine, the Central Clinical Laboratory, and the technical staff of the Department of Microbiology, Ashwini Rural Medical College Hospital and Research Centre, Solapur, for their cooperation during this study.