Introduction
Dengue is an arthropod borne viral disease that is transmitted to humans through the bite of infected Aedes mosquitoes.1, 2 It is common in tropical and subtropical regions of the world and leads to approximately 50-100 million infections each year with 24,000 deaths. 3, 4
Majority of primary dengue fever cases are asymptomatic/ mild symptomatic with rare cases of fatality while secondary dengue infections often lead to severe dengue in the form of dengue hemorrhagic fever (DHF) and dengue shock syndrome (DSS) which can turn fatal. 5, 6, 7
We know that secondary dengue infection cases are more severe and have a higher mortality rate, but in this study, we found that all secondary dengue cases may not always lead to severe dengue while on the other hand, all severe dengue cases may not be due to secondary dengue infection. Therefore, early detection of Dengue IgG by ELISA often helps us predict prognosis of disease and the need for hospital monitoring, yet one should not be complacent for primary dengue infections.
There are 4 distinct serotypes of Dengue virus DENV1, DENV2, DENV3, DENV4 and the infecting DENV serotype and genotype often have a major impact on the clinical presentation and outcome of the patient. 8, 9, 10 The serotype detected region wise is diverse and changes every three to five years ,therefore, surveillance of molecular serotypes and early detection of any shift from the previously circulating serotypes will serve as an early indicator for the preparedness of handling more severe dengue cases, which may be of immense public health importance.
Aim
To find out the proportion of Primary and Secondary Dengue infection amongst Dengue fever cases based on Dengue IgG detection in NS1 reactive cases.
To delineate the differences of demographic, clinical and laboratory parameters among primary and Secondary dengue cases, if any.
To identify the prevalent serotypes circulating in the region.
Materials and Methods
The study is a hospital based, cross-sectional, observational study conducted at Virology Unit, Department of Microbiology, School of Tropical Medicine, Kolkata. The study period was from March 2021 to February 2022. All samples of acute febrile illness patients attending the Unit of Virology were included in this study.
Methods
Blood collected from patients suffering with acute febrile illness / suspected dengue was tested for Dengue NS1 (if fever < 5 days) using DENGUE NS1 Ag MICROLISA ELISA kit. Dengue NS1 reactive cases were subjected for dengue IgG by ELISA using DENGUE IgG MICROLISA ELISA kit and were subjected to serotyping using the Reverse transcription-polymerase chain reaction (RT-PCR) kit. To establish the circulating serotypes of Dengue from these NS1 positive samples, RNA was extracted using viral RNA extraction kit (Qiagen), and then the extracted RNA was amplified using RT-PCR kit. Results obtained were analyzed and interpreted.
Results
Out of 2286 patients of acute febrile illness only 134 (5.86%) patients were NS1 reactive. Amongst the 134 NS1 reactive patients, 21(16%) had secondary dengue infection showing early IgG detection in NS1 reactive samples, while the rest 113(84%) patients had primary dengue infection being only NS1 or NS1 & IgM reactive or only IgM reactive.
Among primary dengue infections 63(55.8%) cases were males and 50(44.2%) were females while among the cases with secondary dengue infection, 9(42.9 %) were males and 12(57.1%) were females. Among the 113 primary dengue cases, 68(60.17%) belonged to the age group 16 - 35 years indicating that young adults are affected the most, while among the 21 secondary dengue cases maximum patients belonged to the age group of 16 – 25years (23.8%) and 36 - 45 years (23.8%).
Dengue reactive cases were seen throughout the one-year study period of March 2021- February 2022 with an upsurge of cases during the months of October, November and December 2021.
Table 1
Clinical features of Primary and Secondary dengue infected cases in the study
The clinical features of primary and secondary dengue infection like rash, abdominal pain and bleeding manifestations were much higher in secondary dengue infection than primary dengue. (statistically significant p-value < 0.05, at 0.05 level of significance)
Table 2
Laboratory parameters and radiological findings of primary and secondary dengue cases
Figure 2 shows that out of 113 primary dengue infected cases, 66% received treatment at home, 29 % were treated in hospital wards and 5 % needed admission in CCU.
In comparison among 21 secondary dengue infected cases, 33 % were treated at home, 57 % were treated in the hospital wards and 10 % required admission in CCU.p-value:-p < 0.001
Figure 3 shows that out of 113 primary dengue infected cases, 99.1% recovered and 0.9% death was recorded whereas out of 21 secondary dengue infected cases, 95.2% had recovered with 4.76 % death. p-value: - p < 0.001
Figure 4 shows that DENV RNA was detected in 57 samples. The predominant serotype detected is DENV 3. The monotypic infections with DENV-3, DENV-4, DENV-2 and DENV-1 were detected in 70.18% (40), 10.53% (6), 7.02% (4), and 1.75% (1) cases respectively. Regarding dual infections, DENV-2 + DENV-3(7.02%), and DENV-1 + DENV-3(3.51%) were detected
Discussion
In the present study, primary infected cases had a slight male preponderance (males -55.8%, females -44.2%) whereas in secondary infection percentage of females were slightly more than males (females- 57.1%, males-42.9 %). This can be explained by more exposure of males to outdoor either due to occupational or recreational works in a conservative Indian social background. 11, 12 However, female sex is one of the risk factors for severe dengue, apart from high body mass index and high viral load. 13, 14
Upon analyzing the month-wise distribution of dengue cases a seasonal upsurge of cases was identified from the months of August to December in the year 2021 with a peak in the month of November. The correlation between the occurrence of dengue cases with monsoon and post monsoon season is clearly evident in our study which can be explained by the fact that there is increased vector density due to increase breeding of mosquitoes in the stagnant fresh water during the rainy season.
Previous studies have shown that majority of the patients of dengue belonged to the age group 16 – 35 years which was similar to our study. 11, 15 As per our study, majority of Dengue reactive cases in Primary infection (60.17%) and secondary infection (38.1%) were of age group 16 - 35 years.
In our study, both primary dengue cases and secondary dengue cases presented with fever for less than 5 days. Even though a higher percentage of rash (85.7%), retro orbital pain (38%), vomiting (52.3%), abdominal pain (28.5%) and bleeding manifestations (23.8%) were observed in secondary dengue cases, yet a smaller percentage of these clinical signs and symptoms were also observed among primary dengue cases.
Similarly, although anaemia (Hb% <10 gm/dL), leucopenia (<4000/μl), thrombocytopenia (platelet count < 1,00,000/μl), high serum AST levels (>80 IU/L), high serum ALT levels (>80 IU/L), free fluid accumulation in abdomen (23.8%) and pleural effusion (19%) were observed in higher percentages among secondary dengue cases, yet it was not exclusive. These findings were also observed in a small proportion of primary dengue cases.
In our study, among the 113 primary dengue cases, 2 cases (1.7 %) suffered from severe dengue in which one patient was diagnosed as dengue haemorrhagic fever and recovered while another patient was diagnosed with Expanded dengue syndrome and expired. Among 21 secondary dengue cases, 4 cases (19%) suffered from severe dengue in which two patients developed DSS both of whom recovered with supportive treatment; two other patients developed DHF among whom one patient survived while the other patient expired. Five patients had co-infection of dengue with malaria in which all recovered.
Therefore, even though prolonged hospital stay, critical care admission and death were observed more among secondary dengue cases, yet they were also seen in smaller numbers among primary dengue cases.
Among the 134 NS1 reactive patients, Dengue virus serotype was detected only in 57 patients. The reason could be because the window for viral RNA detection is limited to 2-3 days after onset of illness and patients often arrive at the hospital after 3 days of onset of fever. Out of the above 57 patients, four had secondary Dengue infection while the rest of the patients had primary infection. All the four serotypes were detected throughout the study with a predominance of DENV-3 serotype. 51 out of the 57 patients had monotypic infections and 6 had dual infections. The monotypic infections with DENV-3, DENV-4, DENV-2 and DENV-1 were detected in 70.18% (40), 10.53% (6), 7.02% (4), and 1.75% (1) cases respectively. Regarding dual infections, DENV-2+ DENV-3(7.02%), and DENV-1 + DENV-3(3.51%) serotype were commonly found. All the patients having co–infecting serotypes had prolonged hospital stay and had a longer recovery time. Only 1 dengue dual infected patient suffered from Dengue Shock Syndrome which was also a case of secondary dengue. This patient had recovered and was discharged home.
Conclusion
It is evident that patients who suffer from Secondary Dengue infection suffer from more complications as well as require more supervision and critical care support. This highlights the importance of Dengue IgG detection in the early phase of the disease in order to predict the complications of Dengue which can save many more lives.
However primary dengue infection can also have severe Dengue or Expanded Dengue syndrome of which clinicians should remain vigilant.
Molecular surveillance may help us predict large scale outbreaks of dengue if regional shifts in the predominantly circulating serotypes are detected during the early phase of the dengue season.
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