SERUM FERRITIN AS AN EARLY INDICATOR OF SEVERITY OF DENGUE
DISSERTATION SUBMITTED FOR THE DEGREE OF M.D BRANCH VII
(PAEDIATRIC MEDICINE) REG. NO: 201717102
MAY 2020
MADURAI MEDICAL COLLEGE, MADURAI
THE TAMILNADU DR. M.G.R MEDICAL UNIVERSITY CHENNAI, TAMIL NADU
CERTIFICATE
This is to certify that the dissertation entitled “SERUM FERRITIN AS AN EARLY INDICATOR OF SEVERITY OF DENGUE” is the bonafide work of Dr. V. MOHAN RAJ in partial fulfilment of the university regulations of the Tamil Nadu Dr. M.G.R Medical University, Chennai, for M.D Degree Branch VII – PAEDIATRIC MEDICINE examination to be held in MAY 2020.
Dr. J. SANGUMANI, MD., Dean, Madurai Medical College, Government Rajaji Hospital, Madurai – 625020
BONAFIDE CERTIFICATE
This is to certify that the dissertation entitled “SERUM FERRITIN AS AN EARLY INDICATOR OF SEVERITY OF DENGUE” submitted by Dr. V MOHAN RAJ to the faculty of Pediatrics, The Tamil Nadu Dr. M.G.R Medical University, Chennai in partial fulfillment of the requirement for the award of M.D Degree Branch VII (PAEDIATRIC MEDICINE) is a bonafide research work carried out by him under our direct supervision and guidance.
Dr. S.BALASANKAR MD DCH Director I/C & Professor of Paediatrics Institute of Child Health & Research center, Madurai Medical College,
Madurai.
DECLARATION
I, Dr. V MOHAN RAJ, solemnly declare that the dissertation titled “SERUM FERRITIN AS AN EARLY INDICATOR OF SEVERITY OF DENGUE” has been conducted by me at Institute of Child Health and Research Centre, Madurai under the guidance and supervision of Prof. Dr. S. BALASANKAR M.D., DCH.,
This is submitted in part of fulfillment of the regulations for the award of M.D Degree Branch VII (Paediatric Medicine) for the May 2020 examination to be held under The Tamil Nadu Dr. M.G.R Medical University, Chennai. This has not been submitted previously by me for any Degree or Diploma from any other University.
Place: Madurai Dr. V MOHAN RAJ Date:
CERTIFICATE - II
This is to certify that this dissertation work titled “SERUM FERRITIN AS AN EARLY INDICATOR OF SEVERITY OF DENGUE” of the candidate Dr. V MOHAN RAJ with registration Number 201717102 for the award of M.D., in the branch of PAEDIATRICS personally verified the urkund.com website for the purpose of plagiarism Check. I found that the uploaded thesis file contains from introduction to conclusion pages and result shows 16%
percentage of plagiarism in the dissertation.
Guide & Supervisor sign with Seal.
ACKNOWLEDGEMENT
First, I would like to thank the almighty for giving me this opportunity. My sincere thanks to Prof. Dr. J. SANGUMANI, M.D., Dean, Government Rajaji Hospital and Madurai Medical College for permitting me to do this study and utilize the institutional facilities.
I express my sincere thanks and gratitude to Prof. Dr. S. BALASANKAR M.D., DCH., Professor and Director I/C,
Institute of Child Health & Research Centre, Madurai, for his able supervision, encouragement, valuable suggestions and support for this study. I am also greatly thankful for his able guidance, critical review, constant encouragement and full support rendered in every aspect of this study.
I would extend my sincere thanks to Prof. Dr.M.S.
Rajarajeshwaran, M.D., DCH., Prof. Dr. M. Kulandaivel, M.D.,
DCH., Prof. Dr. S. Shanmugasundaram, M.D., DCH., and Prof. Dr. Nandhini, M.D., DCH., Prof. Dr. M. Balasubramanian,
M.D., DCH., Prof Dr. D Raj Kumar, M.D., and Prof. Dr. J Ashokaraja, M.D., DM., for their valuable advice and
encouragement at every stage of this study.
I wish to express my sincere thanks to my Assistant Professors of Pediatrics, Dr.P.Ramasubramaniam, M.D., DCH., Dr. Vanitha, M.D.,
Dr.J.Balasubramanian, M.D., DCH., Dr.R.Suresh, M.D., for their constant guidance, encouragement and support throughout my study. I also extend my thanks to Dr. P. Murugalatha, M.D., Dr. P. Kannan, M.D.,
Dr. K. Ramya, M.D., Dr. P. Guna, M.D., DMRD., Dr. S. Murugesalakshmanan, M.D., Dr. Abu backer siddiq, M.D.,
DCH., Dr. Suganthi, M.D., Dr. A.N.Karthik, M.D., Dr. Venkataramanan, M.D., DCH., Dr. Sonia Rosalind Martina,
M.D., Dr. M. K. Lenin, M.D., for their guidance, supervision, valuable suggestions and support throughout this study.
I thank the Institutional Ethical Committee for granting me permission to conduct the study. I also express my gratitude to all my fellow postgraduates for their kind cooperation in carrying out this study and for their critical analysis.
I thank the Institute of Microbiology, Department of Biochemistry and Department of Radiology of Government Rajaji hospital, Madurai for their co-operation throughout my study.
Last but not the least, I submit my heartfelt thanks to the children and their parents for extending full co–operation to complete my study successfully.
CONTENTS
Sl.
No Title Page no
1. Introduction 1
2. Aims and Objectives 38
3. Review of literature 39
4. Materials and Methods 41
5. Observation and Results 46
6. Discussion 74
7. Conclusion 77
8. Limitations 78
Annexures :
• Bibliography
• Proforma
• Abbreviations
• Consent form
• Master Chart
1
INTRODUCTION
Dengue is a arthropod borne viral infection caused by flavi virus which causes simple fever to severe complications like dengue shock syndrome and dengue hemorrhagic syndrome. It is epidemic all over the world and in recent upsurge.
Dengue epidemic has resulted in significant mortality. There is no appropriate treatment and only supportive treatment can be given.
Most of the cases recover with no complications and only certain number of cases can have complications and they require careful monitoring and assessment. In order to predict the complications and to find out which case requires monitoring, we need a parameter to assess.
In this study we aimed to predict the severity in early phase of the disease, using Serum Ferritin.
SERUM FERRITN & DENGUE
In recent times a phenomenon called macrophage activation syndrome (MAS) or hemophagocytic syndrome (HS) is being frequently reported in patients with severe dengue. MAS is a severe systemic inflammatory condition due to excessive activation and proliferation of T cells and well differentiated macrophage that leads to hyperactivated but dysregulated immune responses22,23,24. This results in an overwhelming
2
inflammatory response leading to non remitting high rise of temperature, organomegaly (involving liver and spleen), hemorrhage, lymphadenopathy, and central nervous system (CNS) dysfunction.
Hyperferritinemia (levels above 10000ug/L) is a flagship sign of MAS;
however hypoalbuminemia, cytopenia, coagulopathy, abnormal liver function tests, hypertriglyceridemia, hemophagocytosis, and elevated serum sCD25 and sCD16 levels also serve as adjunct markers of MAS.
Hemophagocytic syndrome (HS) is being increasingly reported in patients with severe dengue with multiorgan complication and is observed in severe dengue involving children.25-28
Another mechanism by which ferritin can be used as a predictor is increased expression of acute phase reactants is observed in patients with severe dengue infection when compared to non-severe cases. This serves to prognosticate the dengue infected patients well ahead of the appearance of clinical warning signs. One such acute phase reactant is ferritin which is produced by reticuloendothelial cells in response to infection and inflammation. Ferritin is highly elevated in dengue infected patients than in patients with other febrile illnesses. Hyperferritinemia seen in these patients exhibit two opposite functions; Early in the phase of clinical illness, increased serum ferritin levels exert a protective effect by chelating the toxic free iron radicals at the site of inflammation15. while in severe
3
cases, raised ferritin may assume a pathogenic role by activating immune cells resulting in cytokine storm.
GEOGRAPHICAL DISTRIBUTION Global
Dengue fever is known in the tropical Southeast Asia and Western Pacific for more than a century. The haemorrhagic form was first recognized in Philippines in 1953. Subsequently DHF was recognized in Thailand, India, Singapore, malaysia and Vietnam. In 1978, a big outbreak was reported from China resulting in 22,122 cases. In 1981, a large epidemic occurred in Cuba resulting in nearly 0.35 million cases of dengue fever. Out of these, 24,000 had DHF and 10,000 had DSS. Dengue has also been noticed in temperate regions of North America, Africa, and Mediterranean Europe1,2.
As per current estimates, more than 100 countries are endemic for dengue fever and about 40% of the world populations (2.5 billion people) are at risk in tropics and subtropics. In 2010, 1.6 million cases of dengue were reported in the Americas alone, of which 49,000 cases were severe dengue3,4. Recently, dengue has also been reported from Costa Rica, France, Mexico, Croatia and Portugal. Incidence of dengue infections annually has almost doubled from 50 million to 96 million (2010) in last few years.
4
Figure 1- Global distribution of Dengue
Figure 1.1 Countries/areas at risk of dengue transmission, 2008
10.C
10.C
(As of 1 November 2008)
5
India
India alone accounted for almost 34% of global dengue burden by 2010. The National Dengue Day was observed on 16th May 2016. Disease is prevalent throughout India in most of the metropolitan cities and towns and is endemic in 18 out of 35 states. Outbreaks have also been reported from rural areas of Haryana, Maharashtra, and Karnataka. Recent trends in transmission have shown occurrence of larger and more frequent outbreaks, geographic expansion of endemic transmission, spread of the disease from urban to semiurban and rural areas and an increasing proportion of severe cases and deaths. An increased propensity to hyperendemic areas particularly in large urban areas, is also noted5.
During 1996, a severe outbreak of dengue or DHF occurred in Delhi wherein about 10,252 cases and 423 deaths were reported. In 2006, India witnessed another outbreak with 12,317 cases and 184 deaths in 21 states.
The initial epidemics in India were due to serotype 2 or 4. The dengue serotype 1 was seen as predominant serotype in Delhi during 2007-2010.
Concurrent infection of chikungunya and dengue serotype 2 has been reported from Delhi and vellore.
Cyclic epidemics are increasing in frequency and in-country geographic expansion is reported in India due to deciduous dry and wet climatic zone with circulation of multiple virus serotypes. Cyclic
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epidemics occurs once in every few years, which is usually associated with good monsoon rains. Sporadic case usually present throughout the year.
However, with improved case management, the case fatality has decreased from 3.3% in 1996 to 0.4% in 2010.
DENGUE VIRUS
Dengue virus has at least four serotypes.
DENV-1 DENV-2 DENV-3 DENV-4
These are antigenically similar but do not offer a complete cross protection after infection by any one of them to other serotypes. Infections in human by a serotype will produce lifelong immunity against reinfection only for that particular serotype but not aganist other serotype. Subsequent infection (secondary infection) by another serotype causes severe dengue along with increased mortality. The severity of epidemics mostly caused by serotype 1 followed by types 2 and 3.
EPIDEMIOLOGY Vector
Aedes aegypti is the vector for dengue virus. Female mosquito bites the man mostly during daytime. After feeding on a person with viremia,
7
the female mosquito can transmit dengue virus immediately or after a period of 10-14 days (extrinsic incubation period). The extrinsic incubation period is a important factor in successful transmission of the disease. A lower environmental temperature increases the extrinsic incubation period, which in turn, decreases the transmission. Once the mosquito becomes infective, it remains so till its death
Figure 2- Aedes mosquito
The flight range of an adult A. aegypti mosquito is usually around 25-50 metres in an urban environment. However, the vector can be transported by water, land, and air travel contributing to the speedy transmission for vast areas. For dengue transmission, the number of infected female mosquitoes per house is important. Usually this number is small, and for an Indian epidemic it was observed to be just 1 per household (house index). The minimum vector density, below which the dengue
8
transmission ceases, is not known at present. The Aedes aegypti mosquito breeding is not necessarily related to the ambient temperature. The mosquito has been found at very high altitudes like 2,200 metres above the sea level. Vectors must survive longer than the sum of the initial non- feeding period after birth (usually 2 days) and the extrinsic incubation period to be able to infect another human. Longevity under natural conditions ranges from 8 days to 42 days. The eggs of Aedes aegypti can survive without water for a year.
Host factor
People at all ages are susceptible to dengue. In Asians, disease is more severe in children. This is in contrast to America where infection mainly occurs in adults which is usually mild. Severe dengue occurs in children who have experienced a previous dengue infection. Other factors which are associated with increase host susceptibilities to severe dengue include HIV infection, bronchial asthma, certain HLA types like HLA 1(A04, A2, B0, B46), HLA 2 (DQ, DR4, DR1), TNF alpha, TGF beta, G6PD deficiency, mutation in mannose binding lectin 2 gene.
ENVIRONMENTAL FACTORS
In tropical countries, a positive association between rainfall or larval density and dengue incidence has been documented. The vector survives at hot and humid temperature (16-30 degree celsius & 60-80% humidity). But
9
dengue cases are also reported in areas with less rainfall also. The transmission of disease occurs only, if the ambient temperature is above 16 degree Celsius. In winter season the transmission rate is very low. This is due to prolongation of extrinsic incubation period beyond the longevity of mosquito.
TRANSMISSION RISK FACTORS
When a member of house hold is infected with dengue, other family members who are living with them are at risk. Dengue virus spread is mainly due to vector infestation.
Place where people gather, like offices, schools, hospitals, factories, etc… in urban places, the movement of infected people causes the spread of virus than the movement of aedes mosquitoes.
Figure 3-Transmission cycle of Dengue
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PATHOGENESIS
Dengue virus infects the peripheral blood mononuclear cells within a few days of infective mosquito bite.
Two patterns of immune response follow:
(1) Primary and (2) secondary (anamnestic).
Persons never previously infected with a flavivirus, nor immunized with a flavivirus vaccine (e.g. Yellow Fever, Japanese encephalitis), mount a primary IgM antibody response when infected with dengue virus, appearing within 2-3 days of defervescence and peaking at 2 weeks after the onset of symptoms. Antidengue IgG appears afterwards. Individuals with immunity due to previous flavivirus infection or immunization mount a secondary anamnestic antibody response when infected with dengue virus5,6. In secondary flavivirus infections, which account for most cases of severe dengue, the dominant immunoglobulin is IgG; the levels of IgM being much lower. A mechanism of immune enhancement or antibody dependent enhancement (ADE) is observed in dengue due to heterologous non-neutralizing antibodies. This is responsible for serious organ dysfunction and haemorrhagic disturbances which can occur during secondary infection by a different serotype. This mechanism promotes binding of dengue virus to surface expressed Fc gamma receptors on monocytes and macrophages, further promoting viral replication and
11
spread. Thus, sequential rather than simultaneous exposure of different serotypes of dengue virus carry a higher chance of ADE resulting in serious disease. Thus, antibody against a strain of dengue virus does not protect from a different strain of virus. Rather, it may increase its capacity to multiply in human monocytes. The infected monocytes result in activation of cross-reactive CD4+ and CD8+ cytotoxic lymphocytes. Cytotoxic lymphocytes mediate release of cytokines resulting in plasma leakage and haemorrhage and are primarily responsible for host defense in dengue mediated via interferon-gamma. Recent studies have highlighted the role of HLA linked protective role of CD8 lymphocytes. Researchers have found that certain phenotypes of HLA may cause hyporesponsiveness of interferon -7 response, thereby weekening the host response.7
12
PATHOPHYSIOLOGY
Two main pathophysiological changes occur in dengue, These are
(1) Increased vascular permeability, resulting in loss of plasma from the vascular compartment to third space accumulation, hemoconcentration, low pulse pressure, and other signs of shock8 and
(2) Disorder in the hemostasis involving thrombocytopenia, coagulopathy and vascular changes.
Secondary dengue infection results in formation of immune complexes and activation of complement system. TNF-alpha, interferon, and interleukin-2 are elevated, and C1q, C3-C8, are depressed. As a result, vasoactive amines are released from the platelets, These cause massive release of water, electrolytes, and plasma proteins from the blood vessels and lead to hypovolemic shock. Increased vascular permeability is mediated through the nitric oxide pathway
Platelet defects are both quantitative and qualitative. Thus, a patient with a normal platelet count may still have a prolonged bleeding time, maculopapular and petechial rashes are present. In these lesions, dengue antigen, IgM, and complement (C3) have been observed.
13
It may be noted that virus is usually not detectable in blood once shock manifests, though viral replication occurs in various organs.
COURSE OF DENGUE FEVER
Figure 4- Course of Dengue illness FEBRILE PHASE
Febrile phase lasts from 2-5 days. During the febrile phase child presents with fever, headache, retro orbital pain, myalgia and vomiting.
14
It is difficult to distinguish dengue fever from non dengue illnesses at this stage. Child may present with progressive decrease in leucocyte count with tender hepatomegaly.
CRITICAL PHASE
Following the febrile phase, child may enter into the critical phase, which usually lasts from 4 to 7 days. Patients with increased capillary permeability may manifest with the warning signs, mostly as a result of plasma leakage.
The warning signs are persistent vomiting and severe abdominal pain which are the early indications of plasma leakage and become increasingly worse and the patient progresses to the shock state.
The patient becomes increasingly lethargic but usually remains mentally alert. These symptoms may persist into the shock stage.
Weakness, dizziness or postural hypotension, spontaneous mucosal bleeding or bleeding at previous venepuncture sites are important haemorrhagic manifestations. Increasing liver size and a tender liver is frequently observed.
Appropriate management at this stage prevents the complication and mortality.
15
RECOVERY PHASE
As the patient survives the critical period, enters into the recovery phase, which occurs due to reabsorption of fluid from the extravascular compartment. At this stage patient develops bradycardia, electrocardiac abnormalities and pruritis.
Figure 5- Case classification of dengue DIAGNOSIS
Hematological Tests
The clinical diagnosis is corroborated by raised hematocrit (Hct) and thrombocytopenia.
16
An Hct level rise of greater than 20% is a sign of hemo concentration and precedes shock. The Hct level should be monitored at least every 24 h to facilitate early recognition of warning signs and every 3-4 h in severe dengue.
Thrombocytopenia occurs in up to 50% of children with dengue.
Platelet counts of less than 100,000 cells/uL indicate onset of critical phase and typically occur before defervescence and the onset of shock. The platelet count should be monitored at least every 24 h initially.
The white blood cell count can be normal or show leukocytosis during initial phase. Leukopenia, often with lymphopenia, precedes thrombocytopenia and is observed near the end of the febrile phase of illness.
Electrolyte abnormalities are seen in critical phase. Metabolic acidosis and elevated blood urea are observed in those with shock.
Serum glutamic pyruvic transaminase (SGPT) levels are elevated.
Low serum albumin levels are a sign of hemoconcentration10
17
TABLE 1. DIAGNOSTIC TESTS FOR DENGUE FEVER
Diagnostic method
Timing of test (after fever onset)
Validity
Virus isolation(culture) 1-5 days ++++
Genome detection (PCR) 1-5 days ++++
Antigen detection (NS1) 1-5 days +++
Antibody detection (IgM) After 5 days* ++
IgG (Paired sera) **
Acute sera 1-5 days;
convalescent sera after 15 days
+
(PCR: Polymerase chain reaction; Ig: Immunoglobulin).
*IgM positivity rates: by 3-5 days (50%), 5-7 days (80%), and 10 days (90%). IgM appears between 3 days and 10 days and disappears by 2–3 months9.
**IgG appears after 1-2 weeks and may persist for life.
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Serodiagnosis
Serum specimens should be sent to the laboratory for serodiagnosis, polymerase chain reaction (PCR), and viral isolation. Because the signs and symptoms of dengue fever are nonspecific, attempting laboratory confirmation of dengue infection is important.
Serodiagnosis is based on (1) detection of viral nonstructural protein 1 (NS1) during initial illness; (2) detection of IgM antibodies to dengue; or (3) fourfold rise in dengue IgG in paired samples11. Table 1 outlines the desired timing of these tests for confirming the diagnosis. Laboratory criteria for definitive diagnosis include one or more of the following:
Isolation of the dengue virus from serum, plasma, leukocytes, or autopsy samples
Demonstration of dengue virus antigen in serum samples via enzyme immunoassay or in autopsy tissue via immunohistochemistry or immunofluorescence can be done. NS1 is a glycoprotein produced by the virus which can be detected early, between 1st to 4th day of illness. It is specific and has a high sensitivity.
Demonstration of a fourfold or greater change in reciprocal IgG or IgM antibody titers to one or more dengue virus antigens in paired serum samples. IgM antibody appears early in disease course, requires single sample and is less cross-reactive to other flaviviruses12.
19
Thus, measurement of raised IgM appears to be most prudent when done after 5th day of illness. As per the NVBDCP, the laboratory test being followed is the IgM antibody-capture enzyme-linked immunosorbent assay (MAC-ELISA) for dengue virus, which captures the dengue specific IgM by using anti-human IgM. There are quicker and cheaper rapid diagnostic test (RDT) kits available which test the presence of anti-IgM or IgG or NS1 antigen. However, these kits carry a high false positive rate and are not recommended by the WHO or under NVBDCP.
Detection of viral genomic sequences in autopsy tissue, serum, or cerebral spinal fluid (CSF) samples via PCR.
Diagnostic kits for NS1 and MAC-ELISA can be procured from National Institute of Virology, Pune under NVBDCP. The Government of India has set up surveillance hospitals and apex referral laboratories for improved disease surveillance.
Dengue diagnostics for clinicians
The objectives of dengue laboratory diagnosis are (i) to confirm the clinical diagnosis and
(ii) to provide information for epidemiological surveillance.
Laboratory diagnosis is not necessary for clinical management except in atypical cases or when carrying out differential diagnosis with other infectious diseases.
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Laboratory diagnosis of dengue is made by detecting the virus and/or any of its components (infective virus, virus genome, dengue antigen) or by investigating the serological responses present after infection (specifically IgM and IgG levels)
Table 2. Dengue diagnostics and sample characteristics
Clinical sample Diagnostic
method Methodology Time to result
Virus detection
and its components
Acute serum (1–5 days of
fever) and
necropsy tissues
Viral isolation
Mosquito or mosquito cell
culture inoculation
One week or more Nucleic acid
detection
RT-PCR and real time
RTPCR
1 or 2 days Antigen
detection
NS1 Ag rapid
tests Minutes NS1 Ag ELISA 1 day
Immuno-
histochemistry 2-5 days
Serological response
Paired sera (acute serum from1–5 days
and second serum 15–21
days after)
IgM or IgG seroconversion
ELISA
HIA 1–2 days
Neutralization Test
Minimum 7 days
Serum after day 5 of fever
IgM detection (recent infection)
ELISA 1 or 2
days Rapid tests Minutes IgG detection IgG ELISA
HIA 1 or 2
days ELISA = enzyme-linked immunosorbent assay; HIA = haemagglutination inhibition assay; IgG = immunoglobulinG; IgM = immunoglobulin M;
NS1 Ag = non-structural protein 1 antigen; RT-PCR = reverse transcriptase polymerase chain reaction
21
Dengue viruses are RNA viruses belonging to the family flaviviridae, genus flavivirus. The four dengue viruses (DEN-[1–4]) are
serologically related but antigenically and genetically distinctive 13.
Three main aspects should be considered for an adequate dengue diagnosis:
● virological and serological markers in relation to the time of dengue infection;
● type of diagnostic method in relation to clinical illness;
● characteristics of the clinical samples.
Virological and serological markers in relation to time of dengue infection
An incubation period of 4–10 days occurs after the mosquito bites, resulting in an asymptomatic or symptomatic dengue infection. During this period the virus replicates and an antibody response is developed. In general, viraemia is detectable in most dengue cases at the same time that symptoms appear, and is no longer detectable at the time of defervescence.
The development of IgM antibody is coincident with the disappearance of fever and viraemia. Virological and serological markers differ in time evolution and titre response and according to whether the infection is primary or secondary.
22
In a primary infection (i.e. when an individual is infected for the first time with a flavivirus), viraemia develops from 1–2 days before the onset of fever until 4–5 days after. Accordingly, anti-dengue IgM specific antibodies can be detected 3−6 days after fever onset. On average, IgM is detected in 50% of cases by days 3–5 after the onset of illness, this figure increasing to 95–98% for days 6−10. Low levels of IgM are still detectable around one to three months after fever. In addition, the primary infection is characterized by slowly increasing but low levels of dengue-specific IgG, becoming elevated at days 9−10. Low IgG levels persist for decades, an indication of a past dengue infection. A totally different picture is observed during a secondary infection, with a rapid and higher increase of anti-dengue specific IgG antibodies and slower and lower levels of IgM.
High IgG levels remain for 30–40 days. A short-lasting but higher viraemia level characterizes the secondary infection compared to the primary infection.
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Figure 6. Virological and serological markers of dengue infection according to time of illness
IgG = immunoglobulin G; IgM = immunoglobulin M
Type of dengue diagnostic method in relation to time of clinical illness The diagnostic method to confirm an acute infection depends on the time of clinical illness: the febrile phase is coincident with the presence of viraemia, some viral components and replication products in blood; the critical and convalescent phases coincide with the development of antibodies, as summarized in Table 3.
Febrile phase (day 1 to days 4–5 of fever)
The infective virus can be isolated in serum by inoculation in tissue culture (mosquito cell cultures) and mosquitoes. This method allows for identification of the viral serotype.
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Virus genome detection using reverse transcriptase polymerase chain reaction (RT-PCR) and real-time RT-PCR confirms an acute dengue infection. Both methods have a high sensitivity and allow serotype identification and quantification of genome copies. Some studies suggest the presence of a higher number of copies in severe dengue cases.
NS1 Ag is a marker of acute dengue infection. Both enzyme-linked immunosorbent assay (ELISA) and rapid commercial tests are available for NS1 Ag detection. The sensitivity and specificity of commercial kits in different serotype infections and days of illness are being evaluated.14 Critical and convalescent phases (after days 4–5 of illness)
Specific IgM is the best marker of a recent dengue infection. MAC- ELISA and rapid tests are the most frequent methods for IgM detection;
however a recent evaluation of four rapid tests demonstrated a low sensitivity. In addition to IgM, high levels of specific IgG in sera collected early after fever onset as detected by ELISA and haemagglutination inhibition assay (HIA) also suggest a recent dengue infection.
Primary infections are characterized by high levels of IgM and low levels of IgG, while low levels of IgM with high levels of IgG characterize secondary infections.
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A single serum sample collected after day 5 of fever onset is useful for IgM determination. Depending on the IgG level in the sample, classification into primary or secondary infection can also be determined using the IgM/IgG optical density ratio. Ratios greater than 1.2 (using the patient’s sera at 1/100 serum dilution) or 1.4 (using serum dilution of 1/20) suggest a primary infection. In addition, IgG titres higher than 1/1280 by HIA or ELISA are also suggestive of a secondary infection.
As IgM antibodies persist for almost three months after fever onset, the detection in samples collected late after the acute phase of illness suggests a recent infection. In dengue endemic countries, acute clinical cases with a positive IgM are classified as probable dengue cases.
The study of paired sera (acute and convalescent serum samples with the second sample being collected 15–21 days after the first sample), allows for serological confirmation of dengue infection. The diagnosis depends upon the demonstration of rising titres of dengue antibodies between acute and convalescent sera
A broad cross-reactivity of ELISA and HIA with other flaviviruses has been observed. Neutralization Test is the method of choice for determination of specific serotype15.
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Table 3. Confirmed and probable dengue diagnosis, interpretation of results and sample characteristics
Method Interpretation Sample
characteristics
Confirmed dengue infection
Viral isolation Virus isolated
Serum (collected at 1–5 days of
fever) Necropsy tissues Genome
detection
Positive RT-PCR or positive real-time RT-PCR
Antigen detection
Positive NS1 Ag Positive
immunohistochemical Necropsy tissues IgM
seroconversion
From negative IgM to positive IgM in
paired sera Acute serum (days 1–5) and convalescent serum (15–21 days
after first serum) IgG
seroconversion
From negative IgG to positive IgG in
paired sera or 4-fold increase IgG levels among paired
sera Probable
dengue infection
Positive IgM Positive IgM
Single serum collected after day
5 High IgG
levels
High IgG levels by ELISA or HI (≥ 1280)
ELISA = enzyme-linked immunosorbent assay; IgG = immunoglobulin G;
IgM = immunoglobulin M; NS1 Ag = non-structural protein 1 antigen;
RT-PCR = reverse transcriptase polymerase chain reaction
27
TREATMENT OF DENGUE FEVER
Supportive therapy is the mainstay of treatment. Increased oral fluid intake is recommended to prevent dehydration. Supplementation with IV fluids may be necessary to prevent dehydration and significant hemo concentration. But unnecessary iv fluid without dehydration may lead to fluid overload. Fever is managed with paracetamol and tepid sponging.
Aspirin and other nonsteroidal anti-inflammatory drugs (NSAIDS) should be avoided as these drugs may worsen the bleeding tendency associated with some of these infections. Shock is managed with isotonic fluids.
Packed cell (PRBC) transfusion is indicated in refractory shock or if there is significant bleeding.
Patients with known or suspected dengue fever should have their platelet count and hematocrit measured daily from the third day of illness until 1-2 days after defervescence. Patients with a rising Hct level or falling platelet count should be monitored more frequently.
Management of dengue illness can be discussed in three steps:
Step 1: Overall assessment of signs and symptoms Step 2: Diagnosis and severity assessment
Step 3: Categorizing the patients into mild, moderate or severe dengue and treating accordingly.
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Overall Assessment History and Examination
Emphasis on history should be on assessment of warning signs.
Physical examination should concentrate on hemodynamic assessment, to determine the presence and extent of shock, confirming or finding the new warning signs, and checking for bleeding and shock manifestations, abdominal tenderness, mental state, and hydration. Tourniquet test should be done compulsorily.
Investigations
Initial investigations should include Hematocrit, WBC count, platelet count and tests to confirm the diagnosis, as described in the sec.
Laboratory diagnosis. In critical phase, additional tests need to be and include liver function test, renal function test, chest X-ray, serum electrolytes, ultrasound abdomen and neuroimaging as necessary.
Diagnosis and Severity Assessment
Determine the phase of disease (febrile, critical, and recovery) and severity (non-severe, severe) of dengue, as per criteria explained earlier.
The child will need admission if any of the following criteria is fulfilled.
● Presence of any of warning signs
● Bleeding from any site.
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● Signs and symptoms suggestive of hypotension
● Renal, hepatic, or/and central nervous system (CNS) involvement
● Pleural effusion or pericardial effusion, ascites
● Rising Hematocrit
● Platelet countless than 50,000/mm3
● High-risk age group-infants and old age
Categorize Patients in Mild, Moderate, and Severe Dengue (Table 2) This step is aimed to place the patient in an appropriate Group (mild, moderate and severe) to decide on future course of treatment, which is as follows:
● Mild: Patients, who may be sent home with advised to come for follow up.
● Moderate: Patients needing continuous monitoring and hospitalization
● Severe: Patients requiring tertiary level care.
Mild Dengue: Home Management
All children who are tolerating oral fluids, passing urine at least once in 6 hr, and not having any of the warning signs can be sent home.
Following managements need to be advised:
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Encourage fluid intake; can give oral rehydration salt (ORS), fresh fruit juice, tender coconut, etc. The parents should be advised to increase the amount of oral fluids in any form to be given (e.g. 3-10 kg: 100 ml/kg and 10-20 kg: 75 ml/kg) Paracetamol (15 mg/kg/dose) if the child is uncomfortable because of fever once in six hours if required. Avoid aspirin, ibuprofen, mefenamic acid, nimesulide and other NSAIDS.
Monitor at home for fluid intake, urine output, fever, obvious bleeding, and altered sensorium. Bring back if any of the above is present or the child develops any of the warning signs.
Parents of infants should be explained the danger signs before sending them home. Tepid sponging for fever may be done as febrile convulsions may occur commonly in this age group. Breastfeeding should be encouraged and continued.
Moderate Dengue:
Close Monitoring and Hospital Management is needed. Any patient who fulfill the admission criteria should be admitted and monitered (as mentioned above). They may or may not have warning signs. A baseline Hematocrit is measured and continuous monitoring is started. In cases where no warning signs are there, patients should be started on maintenance fluids with isotonic fluid. If patient shows signs of mild dehydration, a correction of 50 mL/kg (<12 months) and 30 ml/kg (>12
31
months) is added to maintainence fluid. At all the times clinical parameters are monitored closely and correlated with Hematocrit to guide further fluid therapy. For those who present with warning signs, the following is advised.
• Start isotonic IV fluids (normal saline or Ringer's lactate) at 6 ml/kg/hr for 1-2hr
• Reassess Hematocrit and clinical status:
• If improvement occurs, decrease to 3 ml/kg/hr for 2-4 h, and then continue with 1.5 ml/kg/hr for 2-4 hr,
• If clinical status worsens or Hematocrit rises, increase rate of fluids to 10ml/kg/hr for 1-2 hr
• Reassess clinical status, repeat Hematocrit, and review fluid infusion rates, till the child is better
• If the child improves, maintain minimum IV fluids at 1 ml/kg/hr for 24-48 hr. Stop fluids when child demands and accepts adequate oral fluids, and food along with adequate urine output.
• Those who worsen or develop profound shock bleeding or multisystem involvement manage as severe dengue.
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Severe Dengue:
Tertiary Level Care
Emergency treatment is required in children with severe dengue or those in critical phase as follows:
Obtain Hematocrit, blood count and other organ function tests, as indicated.
Compensated shock: This stage is characterized by low systolic blood pressure, narrow pulse pressure (<20 mmHg), and rise in hematocrit (>20%). In these children, fluid resuscitation is started at 10-20 ml kg/hr, and further directed as per flowchart below.
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Hypotensive shock: Administer isotonic fluid bolus 20 ml/kg in 15 min. For further management, follow the algorithm depicted in Flowchart below.
Colloids may be needed in refractory shock. Colloids can be gelatine or starch based. They carry theoretical risk of allergic reactions and impaired coagulation. They should be infused slowly with strict monitoring for signs of fluid overload.
Haemorrhagic complications: Suspect severe bleeding if there is an unexplained fall in Hematocrit, refractory shock not responding to 40- 60 mL/kg of fluid, and persistent or worsening metabolic acidosis. Packed cell transfusion 10 ml/kg over 2-3 hr can be lifesaving in these children.
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There is not much evidence for platelet transfusion or fresh frozen plasma (FFP) for severe bleeding. Platelet transfusions should not be used prophylactically. Its use has neither shown to prevent progression to severe bleeding nor does it shorten the bleeding time, but may be associated with severe side effects. Platelet transfusion should be restricted to cases with severe bleeding or when platelet counts are below 10,000/mm3. Platelets obtained by single donor apheresis are preferred as they raise the platelet count by 30,000-50,000 as compared to random donor platelets which result in rise by 5,000-10,000 per unit.
Monitoring: This essentially remains the basic prerequisite for treating children with severe dengue, in an emergency setting
• Monitor vital signs and peripheral perfusion, urine output 1-4 hourly till patient is out of critical phase. Monitor Hematocrit before and after fluid replacement, then 6-12 hourly.
• Monitor blood glucose and other organ dysfunction both clinically and biochemically.
• A typical monitoring chart for dengue fever should record the following: body temperature, heart rate, BP, pulse volume, capillary refill time, abdominal pain, appetite, abdominal pain, vomiting, bleeding, and sensorium.
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Treatment of Fluid Overload
A child with dengue can have fluid overload due to excessive or rapidly transfused IV fluids, use of hypotonic fluids, and inappropriate use of fresh frozen plasma (FFP) or platelets. Another important reason is continuation of IV fluids even during the phase of plasma reabsorption and recovery phase.
These children may present with features of respiratory distress, pulmonary edema or congestive heart failure.
Following management is suggested:
● Oxygen therapy
● Discontinuation of IV fluids
● Frusemide 0.1-0.5 mg/kg/dose once or twice daily
● Correction of electrolyte imbalance
● Look for occult hemorrhage and transfuse packed cells (PRBC) Management of Other Complications
Encephalopathy in dengue may result due to dengue encephalitis, intracranial bleeding, electrolyte disturbances, occlusion due to DIVC or hepatic failure (hepatic encephalopathy). Appropriate diagnosis for cause and specific management should be instituted. Cardiac involvement may
36
be seen during shock or during convalescence, which may manifest as arrhythmias or heart failure.
Criteria for Discharge
Patient should be discharged only if he has been
Afebrile for at least 48 hours
Passing urine normally and adequately
Having improved appetite
And has no respiratory distress
Laboratory parameters should show a stable Hematocrit and platelet count of more than 50,000/mm3 and in rising trend.
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VACCINES FOR DENGUE
A live attenuated tetravalent dengue vaccine (CYD-TDV) has been developed which contains four dengue virus with expression of dengue pre membrane and envelope protein and non-structural and capsid protein of yellow fever strain (YF-17D)
Dosage : 3 dose schedule against serotypes 1,3 and 4 The vaccine is still under multicentric phase III trials.
The Chimeri Vax TM system, originally developed to construct JE vaccine, has now been applied to dengue viruses. This vaccine was shown to be safe and immunogenic in a monkey study.
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AIM AND OBJECTIVES
To find out whether Serum Ferritin level during the febrile phase of dengue fever can be used as a predictor of complications.
39
REVIEW OF LITERATURE
Chaiyaratana w. et al (2008)16, 172 children with dengue infection between 2002 to 2005 were included in their study. Their study showed median duration of febrile period is 5 days. Their data showed that both patients with DF and DHF had higher serum ferritin levels during the febrile, toxic and convalescent stages than they did at follow-up. Patients with DHF had higher levels of serum ferritin than those with DF throughout the course of the illness. Those with DHF grades III and IV had higher serum ferritin levels than those with DF and DHF grades I and II.
Cornelia A.M.Van de wag (2014), conducted a study on the carribean island aruba, during the epidemic between sep 2011 to april 2012.
It stated that ferritin can be used as a clinical marker to discriminate between dengue and other febrile illnesses. The occurrence of hyperferritinaemia in dengue virus infected patients is indicative for highly active disease resulting in immune activation and coagulation disturbances.
Therefore, they recommended that patients with hyperferritinaemia should be monitered carefully, as there is a high risk for them to develop severe disease.
R.Soundravally et al (2014)17, conducted a study on 96 patients with dengue fever and other febrile illness. A steady increase in the level of serum ferritin was recorded throughout the course of illness. The
40
elevated ferritin level could predict the disease severity with highest sensitivity and specificity of 76.9 and 83.3 %, respectively, on the day of admission and the same was found to be 90 and 91.6% around defervescence. On the basis on its diagnostic efficiency, they proposed that ferritin may serve as a potential biomarker for an early prediction of disease severity.
Muhammed Nadeem et al (2016)18, conducted a study on 104 patients with dengue infection. In that study they concluded 1. Raised serum ferritin levels are significantly associated with severe dengue. Mean ferritin levels are also high in patients with severe dengue as compared to dengue fever. 2. Serum ferritin levels on the day of admission may serve as biomarker for an early prediction of disease severity in dengue virus infection.
Soumyabatra Roy Chaudri et al (2017)19 conducted a study on 358 cases of serological proven cases and selected only 30 cases of confirmed dengue and compared them with other febrile illness. Ferritin was evaluated as an adjunct marker for the diagnosis of dengue which could possibly aid their clinical judgment and prompt early fluid resuscitation which in turn could be useful in avoiding undue complications. Ferritin, as evaluated in the present study may serve as a significant marker for differentiating between dengue fever and fever of other etiology, even in the absence of a positive NS1 antigen or a positive IgM antibody for dengue.
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MATERIALS AND METHODS STUDY POPULATION:
The study was conducted on 80 dengue patients admitted in Institute of Child Health & Research Centre, Madurai during the study period of 6 months from February 2019 to august 2019.
INCLUSION CRITERIA:
All fever cases which are dengue NS1 antigen positive or which show classical signs and symptoms of dengue.
Children from 1 month to 12 years of age EXCLUSION CRITERIA:
Patients who do not have serological evidence of NS1 positivity or IgM antibody.
Patient with anemia (Hb <11g/dl) and transfusion dependent chronic disease.
DATA COLLECTION:
A previously designed proforma was used to collect the demographic and clinical details of the patients. All the patients underwent detailed clinical evaluation and appropriate investigations are noted.
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STUDY PROTOCOL:
DESIGN OF STUDY:
Prospective study PERIOD OF STUDY:
February 2019 to august 2019.
METHODOLOGY:
History was taken in detail and duration of fever and other associated signs and symptoms noted. Necessary investigations were done.
Child was then followed throughout the clinical course of the disease.
LABORATORY INVESTIGATIONS:
Total count, Platelet count, serum ferritin, USG abdomen, LFT, RFT, neuroimaging and other investigations if necessary.
SERUM FERRITIN ESTIMATION20
Ferritin is a globular protein found mainly in the liver, which can store about 2250 iron (Fe3+) ions. The ferritin molecule consists of a protein shell (Apoferritin) composed of heavy and light subunits, which surrounds a crystalline core containing iron oxide and phosphate. Ferritin is synthesized in the liver, spleen and numerous other body tissues, with major concentrations found in the liver, spleen, bone marrow, and intestinal mucosa. The ferritin levels measured have a direct correlation with the total
43
amount of iron stored in the body. If ferritin is high there is iron in excess, which would be excreted in the stool. If ferritin is low there is a risk for lack in iron, which sooner or later could lead to anaemia. In the setting of anaemia, serum ferritin is the most sensitive lab test for iron deficiency anaemia. In contrast, serum ferritin levels are normal or increased in anemia associated with chronic disease. Elevated serum ferritin levels have been observed in acute and chronic liver disease and lymphoid malignancy (leukemia and Hodgkin lymphoma).
Ferritin is an acute-phase reactant, it is often elevated in the course of disease.
In dengue serum ferritin is elevated because of macrophage activation.
METHOD AND PRINCIPLE OF SERUM FERRITIN ESTIMATION:
Immuno enzymatic colorimetric method for quantitative determination of Ferritin concentration in human serum or plasma.
Ferritin ELISA test is based on simultaneous binding of human Ferritin to two monoclonal antibodies, one immobilized on microwell plates and the other conjugated with horseradish peroxidase (HRP). After incubation the bound/free separation is performed by a simple solid-phase washing. Then the enzyme HRP in the bound-fraction reacts with the
44
Substrate (H2O2) and the TMB Substrate and develops a blue colour that changes into yellow when the Stop Solution (H2SO4) is added. The colour intensity is proportional to the Ferritin concentration in the sample. The Ferritin concentration in the sample is calculated based on a standard curve.
Typical reference interval
Children 1-9 years, 10-60 ng/ml 10-18 years male 10-300 ng/ml 10-18 years female 10-70 ng/ml Men, 18 - 60 years: 30–400 ng/ml Women, 18 - 60 years: 15‐150 ng/ml
Men and women, 60–90 years: 15–650 ng/ml COLLABORATING DEPARTMENTS:
Department of Microbiology Department of Biochemistry Department of Radiology
ETHICAL CLEARANCE: Clearance obtained
CONSENT: Individual written and informed consent obtained from the parents of their children who were included in this study.
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STATISTICAL ANALYSIS:
All data were entered in Excel 2007 and statistical analysis was performed using the statistical software SPSS 1.6.0. Data were expressed as frequency (with percentages), median values (with range (min, max). For continuous variables, Mann Whitney U-test was performed to find the differences between two groups and for categorical variables Pearson’s chi-square test was performed. Results were defined as statistically significant when the P value (2-sided) was less than 0.05.
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OBSERVATION AND RESULTS
Table 4 AGE DISTRIBUTION
Out of 80 cases 23 were less than 5 years and 57 cases were more than 5 years of age.
Age in years No. of cases Percentage
< 5 (1 - 5) 23 28.75
> 5 (5 - 12) 57 71.25
Total 80 100
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Figure 7 - Age distribution 23
57
28.75
71.25
0 10 20 30 40 50 60 70 80
< 5 (1 - 5) > 5 (5.5 - 12)
AGE DISTRIBUTION
No.of cases Percentage (5-12)
5
48
Table 5
GENDER DISTRIBUTION
Gender No.of cases Percentage
Male 43 53.75
Female 37 46.25
Total 80 100
Among 80 cases, 43 were male and 37 cases were female.
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53.75 46.25
Figure 8 - Gender distribution
Male Female
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Table 6 Age & Gender
Age in years Male Female
< 5 (1 - 5) 11 12
> 5 (5 - 12) 32 25
Total 43 37
51
0 5 10 15 20 25 30 35
< 5 (1 - 5) > 5 (5.5 - 12) 11
32
12
25
Figure 9 Age vs Gender
Male Female
(5-12)
5
52
Table 7
DENGUE CASE CLASSIFICATION
Out of 80 cases, 63 cases were non severe dengue and 17 were severe dengue.
Dengue No. of cases Percentage
Non severe 63 78.75
Severe 17 21.25
Total 80 100
53
78.75 21.25
Figure 10 - Dengue Case Classification
Non severe Severe
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Table 8
SEVERE DENGUE MANIFESTATIONS
Out of 17 severe dengue cases, 1 case had fluid accumulation with respiratory distress, 11 cases had shock, 3 cases had bleeding, 2 cases had organ involvement (1 case had liver involvement with elevated liver enzymes and 1 other case had CNS involvement in the form of encephalitis).
Complications No. of cases Percentage Fluid Accumulation With
Respiratory Distress 1 1.25
Shock 11 13.75
Bleeding 3 3.75
Organ Involvement 2 2.5
55
0 2 4 6 8 10 12
FLUID ACCUMULAT
ION WITH RESPIRATOR
Y DISTRESS
SHOCK BLEEDING ORGAN
INVOLVEME NT
No.of cases 1 11 3 2
1
11
3
2
Figure 11 - Severe Dengue Manifestations
No.of cases
56
Table 9
SERUM FERRITIN ON DAY 3 OF ILLNESS
Out of 80 cases, 5 cases had serum ferritin < 300, 54 cases had 301-600 cases, 8 cases had serum ferritin 601 - 800 and 13 cases had serum ferritin levels >800.
Sr Ferritin Day 3
(ng/ml) No. of cases Percentage
< 300 5 6.25
301 - 600 54 67.5
601 - 800 8 10
> 800 13 16.25
Total 80 100
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0 10 20 30 40 50 60 70
< 300 301 - 600 601 - 800 > 800 6.25
67.5
10
16.25
Figure 12 - SERUM FERRITIN - DAY 3
Percentage
(ng/ml)
58
Table 10
SERUM FERRITIN ON DAY 4 OF ILLNESS
Sr Ferritin Day 4
(ng/ml) No. of cases Percentage
< 300 0 0
301 - 600 47 58.75
601 - 800 13 16.25
801 - 1200 2 2.5
> 1200 18 22.5
Total 80 100
47 cases had serum ferritin levels 301-600, 13 cases had levels between 601-800 and 2 cases had levels between 801-1200. 18 cases had serum ferritin levels of more than 1200.
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0 10 20 30 40 50 60
< 300 301 - 600 601 - 800 801 - 1200 > 1200 0
58.75
16.25
2.5
22.5
Figure 13 - SERUM FERRITIN - DAY 4
PERCENTAGE
(ng/ml)
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Table 11
SERUM FERRITIN ON DAY 3 & 4 OF ILLNESS
Serum ferritin levels is elevated in 18 cases to levels more than 1200 on day 4.
Sr Ferritin
(ng/ml) Sr Ferritin Day 3 Sr Ferritin Day 4
< 300 5 0
301 - 600 54 47
601 - 800 8 13
801 - 1200 13 2
> 1200 0 18
Total 80 80
61 0
10 20 30 40 50 60
< 300 301 - 600 601 - 800 801 - 1200 > 1200 5
54
8
13
0 0
47
13
2
18
Figure 14 -SERUM FERRITIN on Day 3 & 4
Sr Ferritin Day 3 Sr Ferritin Day 4
(ng/ml)
62
Table 12 FEVER DAYS
Majority of cases had fever for 4 to 5 days. Mean fever day is 4.78.
Fever no.of cases Percentage
3 days 2 2.5
4 days 25 31.25
5 days 43 53.75
6 days 9 11.25
7 days 1 1.25
Total 80 100
Mean fever days 4.78
63 2
25
43
9
1 2.5
31.25
53.75
11.25
1.25 0
10 20 30 40 50 60
3 days 4 days 5 days 6 days 7 days
Figure 15 - FEVER DAYS
no.of cases Percentage
64
Table 13
MEAN & SD FOR SERUM FERRITIN ON DAY 3 OF ILLNESS
The mean value of serum ferritin in non severe dengue on day 3 of illness is 404.43 with standard deviation 102.82. The mean value of serum ferritin in severe dengue on day 3 of illness is 830.006 with standard deviation 160.064.
Dengue Mean SD
Non severe 404.43 102.82
Severe 830.006 160.064
65
404.43
102.82 830.006
160.064
0 100 200 300 400 500 600 700 800 900
Mean SD
Figure 16 - Day 3 Serum Ferritin Mean and SD
Non severe Severe Serum Ferritin (ng/ml)
66
Table 14
MEAN & SD FOR SERUM FERRITIN ON DAY 4 OF ILLNESS
Dengue Mean SD
Non severe 585.905 193.082
Severe 1259.824 192.461
The mean value of serum ferritin in non severe dengue on day 4 of illness is 585.905 with standard deviation 193.082. The mean value of serum ferritin in severe dengue on day 4 of illness is 1259.824 with standard deviation 192.461.
67 585.905
193.082 1259.824
192.461
0 200 400 600 800 1000 1200 1400
Mean SD
Sr. Ferritin
Figure 17 - Day 4 Serum Ferritin Mean and SD
Non severe Severe Serum Ferritin (ng/ml)
68
Table 15
CLINICAL DIAGNOSIS VS SERUM FERRITIN DAY 3
p value < 0.001 Significant
In severe dengue 12 cases out of 17 cases had serum ferritin levels of more than 800 which is statistically significant compared to non severe cases where only 1 had high ferritin levels.
Day 3 Ferritin > 800 (13) (ng/ml)
Ferritin < 800 (67) (ng/ml)
Severe dengue 12 5
Non severe dengue 1 62
69 12
5 1
62
0 10 20 30 40 50 60 70
Ferritin > 800 (13) Ferritin < 800 (67)
Figure 18 Clinical Diagnosis vs Serum Ferritin on Day 3
severe dengue non severe dengue (ng/ml)
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Table 16
SENSITIVITY & SPECIFICITY OF SERUM FERRITIN DAY 3
Value
Sensitivity 92.31%
Specificity 92.54%
Positive predictive value 70.59%
Negative predictive value 98.41%
Accuracy 92.50%
Over all the sensitivity and specificity of serum ferritin in predicting the severity on Day 3 is 92.31% and 92.54%
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Table 17
CLINICAL DIAGNOSIS VS SERUM FERRITIN DAY 4
In severe dengue 14 cases out of 17 cases had serum ferritin levels of more than 1200 which is statistically significant compared to non severe cases where only 4 had high ferritin levels.
Ferritin > 1200 (18) (ng/ml)
Ferritin < 1200 (62) (ng/ml)
Severe Dengue 14 3
Non severe Dengue 4 59
p value < 0.001 Significant
72 0
10 20 30 40 50 60
Ferritin > 1200 (18) Ferritin < 1200 (62) 14
4 3
59
Figure 19. Clinical Diagnosis vs Serum Ferritin on Day 4
severe dengue non severe dengue (ng/ml)
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Table 18- Sensitivity & Specificity of Serum Ferritin on Day 4
Serum ferritin Value
Sensitivity 77.78%
Specificity 95.16%
Positive predictive value 82.35%
Negative predictive value 93.65%
Accuracy 91.25%
Over all the sensitivity and specificity of serum ferritin in predicting the severity on Day 4 is 77.78% and 95.16%.
