Correlation of serological markers with haematological parameters in early diagnosis of dengue infection in dengue prone areas

Background: Dengue is a serious, rapidly spreading tropical mosquito-borne viral disease caused by dengue virus belonging to the family flaviviridae and genus flavivirus in the last five decades. In many dengue endemic remote areas with limited laboratory diagnostic services their rapid diagnostic tests are simple method which provides the immediate diagnosis and early management. Materials and methods: The study includes clinically suspected 1236 cases with all haematological parameters and tested at clinical laboratory in which 500 serum samples showed positivity for one or more markers like NS1, IgM or IgG antibodies. Results: In our study among 500 seropositive cases thrombocytopenia (<1 lakh) was evident in 66.7% and in 736 seronegative cases thrombocytopenia was seen in 27.47%. Out of 323 NS1 positive cases, thrombocytopenia was evident in 56.0% and in exclusive antibody positive cases thrombocytopenia was noted in 19.2%. These associations were considered to be statistically significant. Conclusion: The problem of dengue is complex in India because large population, poor medical and diagnostic facilities and inadequate mosquito control. Therefore, studies like this will significantly contribute to the early diagnosis, management and can reduce patient’s morbidity and mortality.


Introduction
Dengue is a mosquito-borne viral disease caused by dengue virus belonging to the family flaviviridae and genus flavivirus. It is one of the most serious, rapidly spreading tropical diseases in the last five decades. The incidence has increased many folds with increasing geographic expansion from urban to rural population. Approximately 2.5 billion people live in dengue endemic countries with an estimated 50 million dengue infections at risk annually [1,2,3].
Dengue virus is small single-stranded RNA virus which is spherical in shape having 50 nm in diameter. It contains a host-derived membrane bilayer, a single copy of a positive-sense, single-stranded RNA genome and multiple copies of the three structural proteins and seven non-structural proteins [NS]. Dengue virus is Manuscript received: 10 th January 2018 Reviewed: 20 th January 2018 Author Corrected: 30 th January 2018 Accepted for Publication: 7 th February 2018 divided in five types due to its characteristic antigenicity as DEN-1 to DEN-5. Out of these DEN-2 and DEN-3 known as "Asian" genotypes which are frequently associated with severe disease accompanying secondary dengue infections [4][5][6][7][8].
Now the diagnosis and confirmation of dengue viral infection can be done by viral culture, viral RNA detection by reverse transcriptase Polymerase Chain Reaction (RTPCR) and serological tests such as an immunoglobulin M (IgM) capture enzyme-linked immunosorbent assay (MAC-ELISA) [9].
In many dengue endemic remote areas with limited laboratory diagnostic services there rapid diagnostic tests are simple method which provides the immediate diagnosis and early management. The confirmation of these results can be done later as there is no antiviral treatment or vaccine available to prevent infection. But

Original Research Article
Pathology Update: Tropical Journal of Pathology & Microbiology Available online at: www.pathologyreview.in 77 | P a g e careful fluid management and monitoring for complications is the only option available. As a result of early fluid management regimes and interventions, the case mortality rates can be decreased in many dengue endemic areas [10].

Materials and Methods
The present study was a retrospective study, conducted in the Pathology department of Dr. D. Y. Patil hospital at Pune from January 2016 to November 2016. Total 1236 serum samples were collected from clinically suspected cases of dengue out of which 500 samples were positive for either one or more of the dengue parameters. The samples were tested immediately in central laboratory for NS1 antigen, IgM and IgG antibodies by using the immunochromatography test kit-Dengue NS1 antigen and antibody using Combi Card supplied by well-known manufacturer.
The tests were performed in accordance with the manufacturer's instructions. Haematological parameters of all the dengue positive cases were recorded using haematology analyser. Statistical analysis was done by using Chi-square and Z-test wherever applicable. The clinically suspected 1236 cases are tested at clinical laboratory in which 500 serum samples showed positivity for one or more markers like NS1, IgM or IgG out of which 303 are male and 197 were female. The majority of 296 (59.2%) cases were positive for only NS1 antigen, followed by IgG and IgM with 151 (30.2%) and 21 (4.2%) cases respectively [ Table 1]. Out of 500 cases study showed 487 (97.4%) cases with haemoglobin values more than 10gm%. Only 3 cases and 10 cases show haemoglobin range <7.0gm% and between 7.1-9.9 gm% respectively. [ Table 3] The P value is less than 0.0001. The association is considered to be statistically significant. The P value is less than 0.0001. The association is considered to be statistically significant.  [11,12,16] In Primary dengue case there will be a low titre and slow rising of antibodies. IgM antibody will appear first after 3-5 day followed by IgG antibody at the end of first week of illness. IgM levels peak about two weeks after the onset of symptoms and then decline generally to undetectable levels over 2-3 months. But anti-dengue serum IgG is increasing slowly after first week and there after it remains detectable after several months, and probably even for life. In contrast during secondary infection, rapid increase and high titre of antibodies are seen, i.e., high levels of IgG can be detected even during acute phase of secondary infection and IgM response is variable [1,17].  [11,13,16] The cases with rapid decrease in platelet and white blood cell count with rising haematocrit levels are most likely to be due to dengue infection [1]. In our study out of 500 positive cases of dengue infection; leucopenia was evident in 141 (28.2%) cases and leucocytosis in 35 (7%) cases [ Table No.6]. While the study done by Manimala S there were 57(57%) cases of leucopenia which is not comparable with our study. This disparity in results may be due to difference in sample size [21].

Conclusion and future
In this country the problem of dengue is complex due to the large population, poor medical and diagnostic facilities and inadequate mosquito control. This country needs a quick effective technique and reliable diagnosis in addressing this potentially fatal, epidemic prone infection and large number of virus laboratories to confirm it and alert the public to take action. Therefore, studies like this will significantly contribute to the clinical management and can reduce patient's morbidity and mortality. To conclude this study emphasises on the fact that dengue is a life-threatening condition and there is a need for early diagnosis and treatment in order to prevent its complications.