Serodiagnosis of Dengue Infection by Rapid Immunochromatography Test in a Hospital Setting in Delhi, India, 1999-2001 by
A Chakravarti*, R Kumaria, N Berry and VK Sharma Department of Microbiology, Maulana Azad Medical College, New Delhi – 110002, India
Abstract A hospital-based cross-sectional serodiagnostic survey was undertaken during 1999-2001 at the Lok Nayak Hospital, Delhi, using a rapid immunochromatography test aimed at detecting anti-dengue antibodies from cases experiencing a febrile illness consistent with dengue infection. Acute-phase blood samples were collected from 345 patients attending outpatient and inpatient departments with clinical suspicion of dengue infection. A total of 85 patients were found to be antibody positive, of which 15 had IgM antibodies alone, indicating primary infection, whereas 19 cases had both IgM and IgG antibodies indicating secondary infection. The remaining 51 cases were presumed to have either secondary dengue infection or cross reactivity with other flaviviruses. Males outnumbered females during this study. Adults between the ages 21-30 years were found to be the most vulnerable group as 32 (37.6%) positive cases belonged to this group. Eighty-two per cent of positive serum samples were collected between October and November, thus indicating that the post-monsoon period was the most affected. Although the number of the serologically-confirmed cases of dengue infection had decreased, more community awareness and stringent measures for vector control are desired to contain this infection. Keywords: Serodiagnosis, immunochromatography, dengue, Delhi.
Introduction Dengue infection is caused by any one of the four closely related but antigenetically distinct serotypes of dengue virus (DEN-1, DEN-2, DEN-3 and DEN-4). In the past 60 years, dengue transmission and the * For correspondence: dochak@yahoo.com
frequency of epidemics have increased dramatically in most tropical and subtropical countries, including the Americas(1,2). This disease is more prevalent now than it was before and it is predicted to increase further(3). In India, clinical disease compatible with dengue fever is known to
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have existed in an endemic form for a very long time. More than 50 outbreaks were reported during 1956-1996. All the four serotypes of dengue are prevalent almost all over the country. Due to its potential of rapid spread, dengue infection has now become a leading public health problem in India and has been declared as a notifiable disease in Delhi. Trends of the outbreak over the last few years indicate that dengue infection is now occurring with increasing frequency not only in urban areas but in rural as well(4,5,6). Even so, it should be emphasized that a probable under-reporting of this disease occurs as a large proportion of the cases are asymptomatic. A number of dengue outbreaks have been reported from the National Capital Territory of Delhi since 1967 and a major outbreak of DHF was reported in 1996. In this outbreak more than 10,000 cases and 423 deaths were reported, giving a case fatality of 4.1%(7). After this outbreak guidelines were developed for the prevention and control of dengue by the Municipal Corporation of Delhi. Although due to these efforts dengue infection is under check in Delhi, however a few cases are still reported every year. Serology is the mainstay of the diagnosis of dengue infection in most routine laboratories as it is rapid and easier to perform as compared to the conventional cell culture technique. Detection of antibodies is useful for the serological diagnosis of dengue, active surveillance and disease control. The present report on a hospital-based cross-sectional serological survey aimed at detecting anti-dengue antibodies from cases experiencing a febrile
illness consistent with dengue infection was conducted in Delhi during 1999-2001. The findings are presented below.
Methodology Study design, population and sample size Acute-phase blood samples were collected from 345 patients experiencing a febrile illness consistent with dengue infection, attending the outpatient and inpatient departments of Lok Nayak Hospital, a tertiary care hospital in Delhi, over the period of three years. The study population comprised individuals of all age groups, selected according to the following inclusion and exclusion criteria.
Case-inclusion criteria A case was included if there was high fever with clinical symptoms suggestive of dengue infection as per WHO criteria(8).
Case-exclusion criteria A case was excluded, if routine laboratory testing suggested bacterial or any viral infection other than dengue infection or any other disease. A performa containing detailed clinical history and clinical findings was maintained for every patient.
Serology Dengue Duo IgM and IgG Rapid Strip test (Pan Bio, Australia) was used for the detection of anti-dengue antibodies. The Dengue Bulletin – Vol 26, 2002
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presence of IgM antibodies alone indicated primary infection whereas IgG and IgM antibodies indicated secondary infection. IgG antibodies alone was considered as suspected secondary infection or having cross-reactivity with other flaviviruses.
22.5% suspected cases were serologically positive (Table 1). Figure 1: Year-wise distribution of total suspected and serologically positive dengue cases 140 120
Results Seropositivity Blood samples were collected from 345 patients experiencing a febrile episode consistent with dengue infection over a period of three years, 1999-2001. Eighty-five cases (25%) were confirmed as serologically positive, with 15 cases showing IgM antibodies indicating primary infection and 19 cases showing both IgM and IgG antibodies indicating secondary infection (Figure 1). IgG antibodies alone were detected in 51 cases and these cases were presumed to have either suspected secondary dengue infection or had cross reactivity with any other flaviviruses. The year-wise case distribution revealed that in 1999, 27%, in 2000, 24% and in 2001,
100 80 60 40 20 0 1999 Total Cases 2000 Positive 2001
Distribution by age and sex The maximum number of positive cases (32 out of 85) belonged to the age group 21-30 years; and males outnumbered females. In 1999, 40%, in 2000, 37.5% and in 2001, 34.8% of the cases belonged to the age group 21-30 years (Table 2).
Table1: Year-wise distribution of suspected and serologically positive dengue cases Year Total cases 111 132 102 345 Serologically positive cases 30 (27%) 32 (24%) 23 (22.5%) 85 (25%) Number of cases with anti-dengue antibodies IgM 4 (13%) 6 (18.8%) 5 (22.2%) 15 (17.6%) IgM+IgG 12 (40%) 3 (9.4%) 4 (17%) 19 (22.4%) IgG 14 (47%) 23 (71.8%) 14 (61%) 51 (60%)
1999 2000 2001 Total
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Table 2: Age-wise distribution of serologically positive dengue cases Year Total positive cases 30 32 25 85 Age groups (years) 0-10 4 (13.3%) 3 (9.4%) 6 (26.1%) 13 (15.3%) 11-20 8 (26.7%) 10 (31.2%) 5 (21.7%) 23 (27.1%) 21-30 12 (40%) 12 (37.5%) 8 (34.8%) 32 (37.6%) > 30 6 (20%) 7 (21.9%) 4 (17.4%) 17 (20)
1999 2000 2001 Total
Table 3: Month-wise distribution of serologically positive dengue cases Year 1999 2000 2001 Total August 0 0 1 1 September 2 3 4 9 October 15 9 3 27 November 12 20 11 43 December 1 0 4 5 Total cases 30 32 23 85
Distribution by month and season The maximum number of cases, 70 out of 85 serologically positive cases, were reported during October – November (Table 3). In 1999, the maximum number of cases (50%) were reported in October. During 2000 and 2001, the maximum number of cases (62.5%) and (48%) respectively, were reported in November. Thus, the seasonal occurrence of positive cases showed that the post-monsoon period was the most affected period, though the dengue vector is present throughout the year (Figure 2).
Discussion The analysis of the results in this study were based upon the interpretations mentioned in PanBio Rapid Immunochromatography test procedure manual. This rapid test had a good sensitivity and specificity comparable to other assays as reported by others and us in our previous studies(9,10). Results were available within 30 minutes, and even with a single serum sample both IgM and IgG antibodies could be detected. A total of 85 out of 345 suspected cases were found to be serologically positive Dengue Bulletin – Vol 26, 2002
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during the three-year study period. The positive cases included primary, secondary and suspected secondary infections. There was a slight increase in the number of cases with primary infection over the three-year period, though this was not significant. Sharma et al(11), in their study in 1998, reported 64% cases with recent infection, which was higher than that observed by us. This was because our study was limited to only one hospital whereas they had cases from all over Delhi, or it suggested that there was a reduction in the cases with current infection. As compared to 1999, more than Figure 2: Month-wise distribution of serologically positive dengue cases 25 20 15 10 5 0 A ug. Sep. O c t. N ov. D ec. 1999 2000 2001
females in all the age groups studied. Sharma et al(11) had reported that the maximum number of patients (69.5%) belonged to age 14 years and above. Quang et al(13) from north Viet Nam also observed the highest number of positives in the >20year age group. These findings led to the suggestion that adults are at a higher risk of contracting dengue infection. Ae. aegypti, the vector responsible for dengue infection, is present throughout the year in Delhi, but the seasonal peak of dengue infection coincides with the monsoon season, as stagnant water provides the most suitable conditions for its breeding(14). In our study the post-monsoon period seemed to be the most favorable since 82% of the samples were collected during the months of October and November. A similar pattern was observed in 1996 and 1998 in Delhi(10,11), Bangladesh(15) and Taiwan(16). Thus, it may be concluded from our study that there has been a decrease in the number of cases with dengue infection over the three-year period though fresh cases are being reported. As it may take years before a dengue vaccine is made available, measures to be considered for its containment are: improved sanitary conditions, community awareness and their participation to check and eliminate all kinds of breeding sites for the vector, and last but not the least, early serological detection of cases for active surveillance and disease control.
a four-fold decrease was observed in the number of cases with secondary infection in 2000, while the decrease was more than two-fold in 2001 (Table 1). The total number of cases with dengue infection showed a declining trend during 1999-2001. A similar declining pattern in dengue infection was reported in Delhi during 1997-1999(12). The most vulnerable age group was found to be 21-30 years (37.6%). Males outnumbered
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