A DISSERTATION ON
“AN ASSOCIATION BETWEEN SERUM CALCIUM LEVEL AND SEVERITY OF DENGUE VIRUS INFECTION IN GOVT. MOHAN KUMARAMANGALAM MEDICAL COLLEGE HOSPITAL, SALEM.”
Submitted to
THE TAMILNADU DR.M.G.R.MEDICAL UNIVERSITY CHENNAI – 600 032, TAMILNADU
In partial fulfillment of the regulations for the awards of the degree of
M.D GENERAL MEDICINE BRANCH – I
GOVERNMENT MOHAN KUMARAMANGALAM MEDICAL COLLEGE, SALEM
MAY– 2020
DECLARATION BY THE CANDIDATE
I hereby declare that this dissertation titled “AN ASSOCIATION BETWEEN SERUM CALCIUM LEVEL AND SEVERITY OF
DENGUE VIRUS INFECTION IN GOVT. MOHAN
KUMARAMANGALAM MEDICAL COLLEGE HOSPITAL, SALEM.”
is a bonafide and genuine research work carried out by me under the guidance Dr. D.VIJAYARAJU MD, Professor of the Department, Department of general medicine, Government Mohan Kumaramangalam Medical College Hospital, Salem, Tamil Nadu, India.
Date:
Place: Salem
Signature of the Candidate DR.S.KESAVAN
CERTIFICATE FROM THE GUIDE
This is to certify that the dissertation entitled “AN ASSOCIATION BETWEEN SERUM CALCIUM LEVEL AND SEVERITY OF DENGUE VIRUS INFECTION IN GOVERNMENT MOHAN KUMARAMANGALAM MEDICAL COLLEGE HOSPITAL, SALEM.”
is the bonafide work of Dr. KESAVAN.S in partial fulfillment of the university regulations of the Tamil Nadu Dr. M.G.R. Medical University, Chennai, for M.D General Medicine Branch I examination to be held in MAY 2020.
Date :
Place : Salem
Prof. Dr. D.VIJAYARAJU M.D., Professor,
Department of General Medicine, Govt. Mohan Kumaramangalam Medical college and hospital, Salem
CERTIFICATE FROM THE HOD
This is to certify that the dissertation entitled “AN ASSOCIATION BETWEEN SERUM CALCIUM LEVEL AND SEVERITY OF DENGUE VIRUS INFECTION IN GOVERNMENT MOHAN KUMARAMANGALAM MEDICAL COLLEGE HOSPITAL, SALEM.”
is the bonafide work of Dr. KESAVAN.S in partial fulfillment of the university regulations of the Tamil Nadu Dr. M.G.R. Medical University, Chennai, for M.D General Medicine Branch I examination to be held in MAY 2020.
Date :
Place : Salem
DR. S.SURESH KANNA M.D., Professor and HOD,
Department of General Medicine, Govt. Mohan Kumaramangalam
Medical college and hospital, Salem.
CERTIFICATE FROM THE DEAN
This is to certify that the dissertation entitled “AN ASSOCIATION BETWEEN SERUM CALCIUM LEVEL AND SEVERITY OF DENGUE VIRUS INFECTION IN GOVERNMENT MOHAN KUMARAMANGALAM MEDICAL COLLEGE HOSPITAL, SALEM.”
is the bonafide work of Dr. KESAVAN.S in partial fulfillment of the university regulations of the Tamil Nadu Dr. M.G.R. Medical University, Chennai, for M.D General Medicine Branch I examination to be held in MAY 2020.
Date :
Place : Salem
DR.K.THIRUMAL BABU,M.D., D.M., THE DEAN,
Govt. Mohan Kumaramangalam Medical College and hospital,
Salem.
GOVERNMENT MOHAN KUMARAMANGALAM MEDICAL COLLEGE
COPYRIGHT
I hereby declare that the Government Mohan Kumaramangalam Medical College Hospital, Salem, Tamil Nadu, India, shall have the rights to preserve, use and disseminate this dissertation / thesis in print or electronic format for academic / research purpose.
Date:
Place: Salem
SIGNATURE OF THE CANDIDATE Dr. KESAVAN.S
PLAGIARISM CERTIFICATE
This is to certify that this dissertation work titled “AN ASSOCIATION BETWEEN SERUM CALCIUM LEVEL AND SEVERITY OF DENGUE VIRUS INFECTION” IN GOVERNMENT MOHAN KUMARAMANGALAM MEDICAL COLLEGE HOSPITAL, SALEM of the candidate Dr.KESAVAN.S with registration Number 201711404 for the award of M.D DEGREE in the branch of GENERAL MEDICINE - I 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 10% percentage of plagiarism in the dissertation.
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ACKNOWLEDGEMENT
I owe my thanks to The Dean, Government Mohan Kumaramangalam Medical College and Hospital Prof. Dr. K.THIRUMAL BABU M.D.,D.M, for allowing me to avail the facilities needed for my dissertation work.
I am grateful to Prof. Dr. S.SURESH KANNA M.D., Professor and Head of the Department of Medicine, Government Mohan Kumaramangalam Medical College and Hospital for permitting me to do the study and for his encouragement. I have great pleasure in expressing my deep sense of gratitude and respect to Prof. Dr. D.VIJAYARAJU M.D., Professor Department of Medicine, Government Mohan Kumaramangalam Medical College and Hospital and Prof. Dr. MANJULA M.D., Professor, Department of medicine and chief of medical unit III, Government Mohan Kumaramangalam Medical College and Hospital, for approving this study and giving suggestions and guidance in preparing this dissertation.
I am extremely thankful to my unit assistant professors, Dr. P.ARUL M.D., Dr. P.SADHASIVAM M.D., and Dr. S.PALANIVELRAJAN M.D., Registrar, Department of medicine for their valuable guidance and constant encouragement.
I wish to acknowledge all those, including my other postgraduate colleagues and my wife who have directly or indirectly helped me to complete this work with great success.
Finally, I thank the patients who participated in the study with their extreme patience and co-operation without whom this project would have been impossible.
Above all, I thank the Lord Almighty for this kindness and benevolence.
CONTENTS
S.NO TITLE PAGE NO.
1. INTRODUCTION 1
2. AIMS AND OBJECTIVES 3
3. REVIEW OF LITERATURE 4
4. MATERIALS AND METHODS 49
5. OBSERVATION AND RESULTS 55
6. DISCUSSION 74
7. LIMITATIONS OF THE STUDY 78
8. CONCLUSION 79
9. ANNEXURES BIBILIOGRAPHY PROFORMA ABBREVIATIONS
ETHICAL COMMITTEE APPROVAL LETTER ANTI PLAGIARISM CERTIFICATE
MASTER CHART
81 88 90 92 93 94
1. INTRODUCTION
Dengue is a disease of major concern throught the world due to its ability to cause huge burden on public health system since it is rapidly transmitted by mosquito. Based on (WHO) World Health Organization reports, about 50 to 100 million new dengue infections are estimated to occur annually, with a steady increase in the number of countries reporting the disease. [1]
Dengue infection presented with variety of clinical manifestations ranging from asymptomatic infection or simple viral illness to dengue shock syndrome. Dengue causes severe bleeding, circulatory shock and even death.
So early diagnosis and recognition of severe form of dengue infections like dengue hemorrhagic fever, dengue shock syndrome is cornerstone in management. Though dengue infections are common in paediatric age group, adult admissions has been increased in recent years especially in India.
However, the data of adult dengue infections are limited; this study is to get additional data on dengue infections among adults.
In India, particularly in Tamilnadu state in recent years dengue has been a major health issue contributing to significant mortality and morbidity. The major factors contributing to this mortality is severe form of dengue infection and its complications like shock syndrome, hemorrhagic manifestations and severe thrombocytopenia. So we need to identify the patients who are all going to these complications.
1
In patients with severe dengue infection numerous serum bio chemical parameter changes occur with the onset of plasma leakage, these derangements are not apparent in non severe dengue patients. The various biochemical markers has been measured to identify the severe form of dengue infection like AST, ALT, platelet count, PCV and electrolytes especially calcium levels.
This study is done in Government Mohan Kumaramangalam medical college, Salem and deals with serum calcium levels in association with severity of dengue infection.
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2. AIMS AND OBJECTIVES
To find out the correlation between serum calcium levels and severity of dengue infection.
To estimate the serum calcium levels in patients with suspected dengue fever and with warning signs of dengue fever.
To suggest further studies in the following areas
(a) Role of calcium supplementation in dengue fever for reduction of severity,
(b) To use serum calcium levels as a potential biomarker to predict severe dengue infection.
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3. REVIEW OF LITERATURE
HISTORICAL REVIEW:
Dengue was also known as “Dandy fever” .The first probable case of Dengue fever was recorded in a Chinese medical encyclopedia. The term
“Break bone fever” was coined during the Philadelphia epidemic in 1780, by Benjamin Rush because of the symptoms of myalgia & arthralgia.
Severe dengue was first recognized during dengue epidemics in the Philippines and Thailand during 1950’s. Nowadays, severe dengue affects most Asian and Latin American countries .Severe dengue complicated by hemorrhages; shock and death were reported in outbreaks in Australia (1897), Greece (1928) and in Formosa (1931). Mosquito borne transmission by Aedes aegypti was demonstrated in 1903. Its viral etiology was demonstrated in 1906.
In Japan, Sabin isolated the virus in 1944 and demonstrated the existence of dengue viral serotypes, but the one isolated in Calcutta in 1944 from the blood of US soldiers was considered as a first report for a longtime.
After World War II, pandemics with intensified transmission of multiple viral serotypes started in Southeast Asia, leading to outbreaks of dengue hemorrhagic fever.
4
In India, first major epidemic illness compatible clinically with dengue occurred in Madras in 1780 followed by spread throught the country.
FIGURE 1: Areas based on suitability of dengue transmission [2]
FIGURE 2: Average annual number of DF/DHF cases (WHO 1955-2007)[9]
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EPIDEMIOLOGY:
DENGUE GLOBAL BURDEN
Around 2.5 billion people – two fifths of the world's population in tropical and subtropical countries are at risk. Around 50 million dengue infections occur worldwide every year .Almost 5, 00,000 people with DHF require hospitalization each year. Majority (approximately 90%) of them are children aged less than five years, with about
2.5% mortality. [9]
FIGURE 3: Dengue cases notified and incidence in WHO regions in 2013-2017
6
Epidemics of dengue are increasing in frequency. During epidemics, infection occurs in those who have not been previously exposed to the virus at rate of 40% to 50% but can also reach about 80% to 90%. In 2016 major outbreaks occurred throught the world. After which, a drop in the number of cases in 2017-18 occurred, followed by sharp increase in cases is being observed in 2019.
DENGUE BURDEN IN INDIA
FIGURE 4: No. of cases and no. of deaths due to dengue in 2010-2017 in India
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Major outbreaks in India occurred in 1996, 2006 and 2010 with reduction in case fatality rates with each epidemic due to better management techniques followed after launch of national dengue treatment guidelines. The disease has a seasonal pattern with peak incidence after the monsoons. However, the southern and western states of the country show a perennial transmission.
FIGURE 5: Seasonal trend of dengue cases in India in 2010-2013
The epidemics from India are those from Calcutta (1963), Vishakapattanam (1964), West Bengal (1968), Ajmer (1969), Kanpur (1969), Delhi (1970), Rajasthan (1985) and Delhi (1996).
8
Major DF outbreak in India occurred between September and October 2006 involving more than 12 thousand cases and 184 deaths, of which approximately 3366 cases and 65 deaths were from Delhi alone. In 2006 the number of cases reported as compared to 2005 showed some reduction whereas the case fatality rate remained above 1%. In 2009, in India 15,535 cases were reported with 96 deaths while in 2010 28,292 cases with 110 deaths were reported. In 2013, till August, 22092 cases were reported with 74 deaths.
FIGURE 6: Dengue cases notified and deaths notified in India in 2013-2017
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DISEASE BURDEN IN TAMIL NADU
In Tamil Nadu, there has been a rise in the number of dengue cases reporting units. In 1998, dengue cases were reported from only 4 units which increased to 33 units in 2006 due to the availability of serodiagnostic facilities at different centres. Out of 30 districts in Tamil Nadu, dengue cases have been reported from 29 districts during 1998-2005 which include DF/DHF outbreaks in Chennai in 2001, Nagercoil and Trichirapalli (2003) and DF outbreaks in Krishnagiri and Dharmapuri (2001).
FIGURE 7: State-wise comparision of no. Of cases 2013 vs 2017 A total of 128 cases and 5 deaths occurred in 1998 which increased to 1600 cases and 12 deaths in 2003. Around 1150 cases, 8 deaths occurred during 2005. After that major outbreaks in Tamilnadu occurred in 2007, 2009, 2012 and 2013.
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THE VIRUS:
The Dengue virus belongs to genus Flavivirus. There exist 4 types called as DEN1, DEN 2, DEN 3, and DEN 4. Recently a new serotype of dengue virus called DEN 5 has been identified from a 37 yrs old foreigner in Malaysia (Sarawak state). Researchers demonstrated that this serotype DEN 5 is genetically similar to other four serotypes. Non structural protein called as NS antigen is of 5 types and is surrounded by a lipoprotein envelope.
FIGURE 8: Electron microscopic picture of Dengue virus.
The function of envelope protein is to bind to host cells and
haemagglutination of RBC [3, 4] NS-1 has direct relation to viral titres, it is higher in patients with DHF compared with patients with dengue fever. The elevated levels of serum NS-1 signify the patients at risk of dengue
hemorrhagic fever. There are about 3 sub-types for DENV-1, six for DENV-2 (one of which is found in non-human primates), four for DENV-3 and four for DENV-4.
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THE VECTOR AND THE LIFE CYCLE OF DENGUE VIRUS:
Dengue viruses are transmitted from an infected person to other person by the bite of the female Aedes mosquito. In India, in most urban areas, Aedes aegypti is the main vector; Also Aedes albopictus is incriminated in many states. Also other species like Aedes polynesiensis and Aedes niveus have also been incriminated as secondary vectors in some countries. [9]. The average survival of Aedes aegypti is about 30 days and Aedes albopictus is about 8 weeks. Aedes is a daytime biter.
FIGURE 9: Aedes aegypti mosquito.
Aedes aegypti lives and multiplies in water storage containers, water reservoirs, overhead tanks, unused tyres, coconut shells, disposable cups, unused grinding stones etc… Aedes albopictus prefers natural habitats such as tree holes, latex collecting cups in rubber trees, bamboo stumps, coconut shells, etc. Aedes albopictus breeding has been reported recently in domestic habitats as well.
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FIGURE 10: Life cycle of Aedes aegypti
Aedes mosquito is a tropical and subtropical variety and is widely distributed. Also, due to lower temperatures, Aedes aegypti is relatively uncommon above 1000 metres. Aedes aegypti is highly domesticated and strongly anthropophilic and a nervous feeder and is a discordant species. Aedes albopictus feeds on both humans and animals and an aggressive feeder and also does not require a second blood meal for the completion of the gonotropic cycle. After an adult female mosquito bites a human with dengue fever it enters and multiplies in the mosquito. Viral multiplies over a period of 8-12 days (extrinsic incubation period) . There after virus can spread through bite of the mosquito. Transmission of dengue virus within mosquitoes can occur by transovarial spread.
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THE HOST:
Humans are the main amplifying host of the virus .The viraemia in humans reaches high titres two days before the onset of the fever (non-febrile) and lasts 5–7 days after the onset of the fever (febrile). It is only during these periods that the vector species gets infected on biting the viremic human.
Transmission primarily occurs via the bite of a vector. There are also reports of congenital dengue infections in neonates born to mothers infected very late in the pregnancy.
The dengue virus enters via the skin while an infected mosquito is taking a blood meal. Intrinsic incubation period is about four to ten days, followed by a wide spectrum of clinical illness, although most infections are asymptomatic or subclinical. During the acute phase of illness the virus is present in the blood and its clearance from generally coincides with defervescence. After infection, serotype specific and cross-reactive antibodies and CD4+ and CD8+ T cells remain measurable for years in the human host.
Initial infection by one serotype provides permanent immunity to that serotype. Secondary infection with another serotype or multiple infections with different serotypes leads to severe form of dengue (DHF/DSS).
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Infection with Serotype 1 followed by Serotype 2 is more dangerous on comparing of infection with Serotype 4 followed by Serotype 2. Antibody- dependent enhancement (ADE) of infection has been proposed as a mechanism to explain severe dengue in the course of a secondary infection and in infants with primary infections. Here non-neutralizing, cross-reactive antibodies produced during a primary infection, or acquired passively at birth, bind to epitopes on the surface of a heterologous infecting virus and facilitate virus entry into Fc -receptor-bearing host cells. During a secondary infection, cross- reactive memory T cells are also rapidly activated which proliferate and express cytokines.
TRANSMISSION CYCLE: [9]
Enzootic cycle: This cycle exists between monkeys and Aedes. Viruses do not cause disease in monkeys and the viraemia lasts for about 2–3 days. All the four dengue serotypes (DEN 1 to 4) have been isolated from monkey.
Epizootic cycle: This occurs from an adjoining human epidemic cycles by bridge vectors. The epizootic cycle was observed in Sri Lanka among touqe macaques (Macaca sinica) during 1986–1987 in a study area. Within 3kms of the study area 94% macaques were found affected on a serological basis.
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Epidemic cycle: Maintained by human- Aedes aegypti - human cycle with periodic/cyclical epidemics. All serotypes circulate and give rise to hyperendemicity. Strong anthrophilicity with multiple feeding behaviour and highly domesticated habitats of Aedes aegypti makes it an efficient vector. The persistence of dengue virus, therefore, depends on the development of high viral titres in the human host to maintain transmission in mosquitoes. In arid zones where rainfall is scanty during the dry season, high vector population builds up in man-made storage containers.
A number of factors that contribute to initiation and maintenance of an epidemic which includes:
THE VIRUS
The strain of the virus influences the magnitude and duration of the viraemia in humans.
THE VECTOR
The density, behaviour and vectorial capacity of the vector population.
THE HOST
The susceptibility of the human population (decided by both genetic factors and pre-existing immune profile).
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PATHOGENESIS:
Host immune responses play a major role in the pathogenesis of Dengue Fever. The most favoured mechanism is cytokine storm. These circulating cytokines plays a main role in development of haemorrhage and shock.
Coagulopathy
Mechanism of Coagulopathy associated with dengue Fever still remains unclear. Thrombocytopenia associated with coagulopathy increases the severity of hemorrhage. Release of heparin sulphate or chondroitin sulphate from the glycocalyx may also contribute to the coagulopathy.
In shock, blood levels of tumor necrosis factor receptor, IFN-γ, and IL- 2 are rised. C1q, C3, C4, C5–C8, and C3 proactivators are reduced, and C3 catabolic rates are rised. These factors may interact to increased vascular permeability through the nitric oxide final pathway. Levels of factor XII are depressed. A mild degree of disseminated intravascular coagulation, liver damage, and thrombocytopenia operates synergistically.
Capillary leakage and shock
Hypotension is caused by plasma leakage due to temporary alteration in the characteristics of the endothelial fibre matrix. Here, Anti-NS1 antibody acts as auto-antibodies that cross-react with platelets and non-infected endothelial cells, leading to disturbances in capillary permeability. Plasma leakage may
17
manifest as any combination of haemoconcentration, pleural effusion, Ascites.
It usually becomes evident on 3 to 7 days of illness [5].
FIGURE 11: Pathogenesis in dengue infection. [7]
Microscopically, there is perivascular edema in the soft tissues and widespread diapedesis of red cells on to the vessel wall.
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FIGURE 12: Pathogenesis in dengue infection [6]
CLINICAL FEATURES:
Dengue virus infected person may be asymptomatic or symptomatic and clinical manifestations vary from mild undifferentiated fever to severe haemorrhage and shock. The clinical presentations depend on various factors such as age, virus strain, immune status of the host, and primary or secondary infection. After an average intrinsic incubation period of 4–6 days (range 3–14 days), various non-specific, constitutional symptoms like fever and headache,
19
backache and generalized malaise may develop. Thereafter, there may be retro- orbital pain on eye pressure or eye movement, photophobia, pain in the muscles and joints and, backache. These symptoms may persist from several days to a few weeks. It is noteworthy that these symptoms and signs are marked in frequency and severity of DF.
Fever: The body temperature is usually between 39 °C and 40 °C, and may be biphasic, lasting 5–7 days in the majority of cases.
Rash Transient widespread rash is seen on head, limbs, back and chest regions during the first two to three days. These rashes disappear at the end of febrile period. Some patients may have itching.
FIGURE 13: Impression mark on skin of a dengue patient. [6]
Haemorrhagic manifestations:
Skin haemorrhage may be demonstrated as a positive tourniquet test and/or petechiae. Other bleeding manifestations such as massive epistaxis,
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hyper menorrhea and gastrointestinal bleeding also occur in severe DF, complicated with thrombocytopenia.
Tourniquet test: [9]
A blood pressure cuff is applied and inflated to midpoint between systolic and diastolic blood pressure. The test is positive if there are more than 10 petechiae per square inch. In DHF it is more than 20. The test may be only mildly positive or negative during the phase of profound shock (DSS).
FIGURE 14: Hemorrhagic manifestations of dengue infections. [7]
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CLINICAL CRITERIA FOR DF / DHF/DSS:
Dengue Fever (DF):
An acute febrile illness of 2-7 days duration with ≥2 of the symptoms mentioned below
Headache, pain behind the eyeball, body ache, joint pain, skin rash, bleeding tendencies.
Dengue Haemorrhagic Fever (DHF):
A). A case with clinical criteria of dengue Fever Plus
B). Haemorrhagic tendencies evidenced by one or more of these features mentioned below
1. More than 20 petechial spots on tourniquet test 2. GI and mucosal bleeding
3. Petechial rashes Plus
C) Low platelet count (<100 000 cells per cu. mm) Plus
D). Third space fluid collection manifested by one or more of the following:
1. A rise in average hematocrit for age and sex > 20%
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2. More than 20% fall in hematocrit after fluid therapy
3. Clinical/radiological evidence of third space fluid collection
Dengue Shock Syndrome:
Features of DHF mentioned in their criteria with clinical evidence circulatory shock (tachycardia with narrow of pulse pressure <20mm of hg)
EXPANDED DENGUE SYNDROME (EDS):
Mild or Severe organ involvement may be occur in DF/DHF. Unusual manifestations of DF/DHF are commonly associated with co-morbidities and with other co-infections.
Clinical manifestations observed in EDS are as follows:
1. NEUROLOGICAL:
Encephalopathy, Encephalitis, Febrile seizures, Intra cranial bleed.
2. GASTRO INTESTINAL OR HEPATIC:
Acute pancreatitis, acute hepatitis, Fulminant hepatic failure, Cholecystitis, Cholangitis
3. RENAL INVOLVEMENT:
Acute renal failure, Hemolytic uremic syndrome, acute tubular necrosis
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4. CARDIAC INVOLVEMENT:
Cardiac arrhythmia, Cardiomyopathy, Myocarditis, Pericardial effusion
5. RESPIRATORY:
Pulmonary edema, ARDS, Pulmonary haemorrhage, Pleural effusion
6. EYE INVOLVEMENT:
Conjunctival bleed, Macular haemorrhage, Visual impairment, Optic neuritis
Natural course of dengue Infection:[6]
The clinical course of illness passes through the following three phases:
• 1.Febrile phase
• 2.Critical phase
• 3.Convalescent phase
1. Febrile phase
Patients develop high grade fever suddenly and usually last 2-7 days.
Facial flushing, rash, generalised body ache, vomiting and headache.
Sore throat, injected pharynx and conjunctival injection.
The initial laboratory evidence is reduction of leucocytes followed by platelet count reduction.
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FIGURE 15: Natural course of dengue illness
2. Critical phase (Leakage phase)
The critical phase often occurs after 3rd day of fever (may occur earlier) or around defervescence indicated by a rapid drop in temperature.
In other viral infections, the patient’s condition improves as the
temperature subsides, but the contrary happens in severe dengue infection wherein the patient may deteriorate and manifest third space plasma leakage or organ dysfunction.
The period of plasma leakage usually persists for 36-48 hrs.
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3. Convalescent phase (recovery phase)
Plasma leakage stops followed by redistribution of fluids collected in third spaces such as peritoneum and pleural cavity with generalised pruritus.
The recovery of platelet count is typically preceded by recovery of white cell count. This phase usually occurs after 6-7 days of fever and last for 2-3 days.
Patient may develop pulmonary edema because of fluid overload if the fluid replacement is not optimized carefully.
CASE DEFINITION: [6]
Probable DF/DHF:
A case compatible with clinical description (Clinical Criteria) of dengue fever during outbreak.
(OR)
Non-ELISA based NS1 antigen/ IgM positive.
(A positive test by RDT will be considered as probable due to poor sensitivity and Specificity)
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Confirmed dengue Fever:
A case compatible with the clinical criteria of dengue fever with at least one of the following
Isolation of the dengue virus (culture +VE) from serum, plasma, leucocytes.
Demonstration of IgM antibody titre by ELISA positive in single serum sample.
Positive Serum NS1-ELISA.
IgG sero conversion in paired sera after 2 weeks of illness with four fold increase titre of IgG.
Detection of viral nucleic acid by PCR (polymerase chain reaction).
FIGURE 16: Criteria for dengue infections.
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GRADING OF DF/DHF:
Dengue Fever (DF):
Fever of 2-7 days with ≥2 of these clinical features Headache, pain behind eyeball, body ache, joint pain.
With or without laboratory evidence of leucopenia, thrombocytopenia and no evidence of plasma leakage.
Dengue Haemorrhagic Fever (DHF I):
Above dengue fever criteria plus positive tourniquet test with evidence of plasma leakage.
Thrombocytopenia with platelet count < 1, 00,000/ cu.mm and hematocrit rise more than 20% over baseline.
Dengue Haemorrhagic Fever II (DHFII):
Above DHF I criteria plus some evidence of spontaneous bleeding in skin or other organs (epistaxis, gum bleeds, black tarry stool) and abdominal pain.
Thrombocytopenia (platelet count less than 100000/ cu.mm) and Hct rise more than 20% over baseline
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FIGURE 17: Clinical spectrum of dengue infection [8,9]
Dengue Haemorrhagic Fever III (DHFIII / DSS):
Above DHF II criteria plus clinical evidence of circulatory
failure(tachycardia, narrowing of pulse pressure, cold peripheries)
Thrombocytopenia with platelet count less than 100000/ cu.mm and Hct rise more than 20% over baseline.
Dengue Haemorrhagic Fever IV (DHFIV / DSS):
Profound shock with undetectable blood pressure or pulse.
Thrombocytopenia with platelet count less than 100000/ cu.mm and Hct rise more than 20% over baseline.
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LABORATORY INVESTIGATIONS: [6]
In endemic areas, early symptoms of dengue fever similar to other prevalent diseases such as chikun gunya, malaria, viral infection, leptospirosis, urinary tract infection and typhoid etc... For proper management of dengue fever exclusion of these above conditions is hence very crucial.
ELISA-based NS1 antigen tests:
Dengue NS1 antigen is a highly conserved glycoprotein which is produced in both membrane-associated and secretion forms, is abundant in the serum of patients with dengue infection in the early stage. It has been found to be useful for the diagnosis of acute dengue infections. It is a simple test that is more specific and high sensitivity. NS1 antigen enables detection of the cases very early, i.e. in the viremic stage, which has epidemiological significance for containing the disease transmission. The NS1 ELISA-based antigen assay is commercially available for DENV nowadays and many investigators have evaluated this assay for its sensitivity and specificity. This NS1 assay may also useful for differential diagnosis between flavi viruses because of its high specificity. This test is usually positive for initial 5 days of illness.
IgM Antibody Capture Enzyme-Linked Immuno Sorbent Assay (MAC- ELISA):
MAC-ELISA has been used widely in the past few years. It is a simple test that requires a very little sophisticated equipment. MAC-ELISA is based
30
on detection of the dengue specific antibodies in the test serum by capturing them using anti-human IgM which was already bounded to the solid phase. An enzyme substrate is added to give a specific colour reaction for easy detection.
FIGURE 18: Serology in dengue infection.[10]
The anti-dengue IgM antibody develops a little faster than IgG and it is usually detectable from 5th day of illness. However, the rapidity varies considerably among patients. Some patients have detectable IgM antibodies on days 2 to 4 after the onset of illness, while others may not develop IgM even after seven to eight days after the onset. In some primary infections, detectable IgM antibodies may persist for ≥ 90 days, but in most patients it is undetectable
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level by 60 days. Hence MAC-ELISA has become an invaluable tool for of DF/DHF surveillance. In dengue non endemic areas, it can be used for clinical surveillance for viral illness and population-based sero surveys, with the certainty that any positives denote an evidence of recent infections. It is especially useful for hospitalized patients, those who are generally admitted in the late phase of clinical illness after detectable IgM is already present in the blood.
Polymerase chain reaction (PCR):
Molecular diagnosis based on RT-PCR (reverse transcription polymerase chain reaction), such as one-step or nested RT-PCR, real-time RT- PCR or nucleic acid sequence-based amplification (NASBA) has gradually replaced the older virus isolation methods as the new standard for the detection of dengue virus.
IgG-ELISA:
An IgG-ELISA has been developed that compares well to the haemagglutination inhibition (HI) test. This test can be used to differentiate primary and secondary dengue
infections. The test is simple and easy to perform and it indicates past infections only. Hence this test is not considered as a diagnostic test.
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Serological tests:
Apart from MAC-ELISA and IgG-ELISA, there are a few serological tests are available for the diagnosis of dengue virus infection such as
complement fixation (CF), hemagglutination-inhibition (HI) and neutralization test(NT). These are not routinely used due to various technical problems.
RDTs:
These tests would give a false negative result. Reliability on such tests to guide clinical management of DF results in an increase in the case fatality ratio.
Hence, use of RDT kit is not recommended under the national programme.
Collection of samples:
Laboratory diagnosis of dengue depends on proper collection, storage, processing and shipment of the specimens. While collecting blood samples for
33
serological studies from suspected dengue (DF/DHF) cases, all universal precautions should be taken. While sending the samples for lab confirmation, the following data includes, the day of onset of fever and day of sample collection should be mentioned to guide the laboratory for the type of test (NS1 for samples collected from day 1 to day 5 and IgM after day 5) to be performed.
NVBDCP recommended tests for laboratory diagnosis:
For confirmation of dengue virus infection, Government of India (GoI) recommended use of NS1 ELISA for initial five days of illness and IgM capture ELISA after five day of illness.Directorate of National Vector Borne Disease Control Programme (NVBDCP) has formed a network of laboratories for surveillance of dengue fever cases (sentinel surveillance hospitals and apex referral laboratories) across the country since 2007. These laboratories are also having diagnostic facilities in all endemic areas. They are linked with Apex Referral Laboratories (ARLs) for backup support and serotyping of dengue samples with advanced diagnostic facilities. For details about the laboratories, please refer to NVBDCP website www.nvbdcp.gov.in.
These laboratories receive the samples from a particular area, diagnose and send the reports (line list) regularly to districts/municipal health authorities for implementation of preventive measures at primary health care level to interrupt the transmission.
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Supply of kits:
IgM ELISA test kits (1 kit = 96 tests) are being provided by the National Institute of Virology (NIV) to the identified laboratories.
The laboratory findings of acute DF:
Leucopenia with decreasing neutrophils is seen during the febrile period.
Platelet counts are usually within normal limits
Mild reduction in platelet count (1 to 1.5 lakh cells/mm³) is common. About half of all DF patients have platelet count <1 lakh cells/mm³, but severe thrombocytopenia with platelet count of <0.5 lakh cells/mm³ is rare.
Mild rise in Hct of around 10 %.
The laboratory findings of DHF:
The WBC count may be normal, but sometimes with predominant
neutrophils in the early febrile phase. Thereafter, there is a drop in WBC’s and neutrophils towards the end of the febrile phase. The change in total WBC count (≤5000 cells/mm³) and ratio of neutrophils to lymphocytes
35
(neutrophils<lymphocytes) is useful to predict plasma leakage in the critical period of dengue.
A sudden drop in platelet count to <100 000 occurs by the end of the febrile phase before the onset of shock or end of afebrile phase. The level of platelet count is correlated with severity of DHF.
A sudden rise in hematocrit is observed shortly after the drop in platelet count. Haemo concentration or rising hematocrit by 20% from the baseline value
Thrombocytopenia (<1 lakh cells/mm3) .A rise in hematocrit occurs in almost all DHF cases, particularly in cases with shock. Sometimes the hematocrit level may be affected by early volume replacement and bleeding manifestations. Increased AST/ALT ratio (more than 2) may be seen in some cases.
Electrolyte abnormalities : Reduction of serum sodium and calcium levels
36
COMPLICATIONS OF SEVERE DENGUE:
Liver failure
In liver hepatocytes and Kupffer cells support viral replication. Levels of (AST&ALT) aspartate transaminase and alanine transaminase are significantly higher, and plasma proteins particularly albumin level is significantly lower among patients with severe grades of DHF.
Fulminant liver failure can occur due to hepatitis and progressive liver failure leading to hepatic encephalopathy. Jaundice may or may not be present.
In cases with CNS involvement (hepatic encephalopathy) neurological examination may show hyper-reflexes and extensor plantar response.
Encephalopathy
Factors contributing to the development of encephalopathy includes, hepatic dysfunction, electrolyte imbalances, hypoperfusion (due to shock), cerebral edema (due to vascular changes leading to fluid extravasation), in patients having features of encephalitis the dengue virus has been isolated from the (CSF) cerebrospinal fluid.
Cardiac complications
Varies from asymptomatic bradycardia to serious conditions like pericardial effusion and Myocarditis. Most of these cardiac manifestations are reversible with other features of dengue fever.
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MANAGEMENT:
Management of dengue Fever (DF):
Management of simple dengue fever is symptomatic and supportive, i). Bed rest is advised during the acute phase.
ii). Use cold/tepid sponging to reduce the temperature below 38.5°C.
iii). Antipyretics (paracetamol) may be used to lower the body temperature.
Aspirin and NSAIDS like Ibuprofen should be avoided because it may cause vomiting, gastritis, acidosis, platelet dysfunction and severe bleeding.
Paracetamol dose can be repeated every 6th hourly depending upon fever and body ache.
iv). Oral fluids and electrolyte therapy is recommended for patients who having excessive sweating or vomiting.
v). Patients should be closely monitored for 24 to 48 hours after the end of febrile phase for development of warning signs and complications.
Management during Febrile Phase:
Paracetamol is recommended to reduce the temperature below 39°C.
Adequate oral fluids should be advised to the extent the patient tolerates. Fruit juices or Oral rehydration solution (ORS) used for the treatment of diarrheal diseases are preferable to plain water. Intravenous fluid should be given if the patient is having diarrhea persistent vomiting or refusing to take feed. Patients
38
should be closely monitored for warning signs and signs of shock. The transition from the febrile to the afebrile stage is critical period and usually occurs after the 3rd day of illness. Serial hematocrit may be essential to guide treatment plan, as they reflect the degree of plasma leak and need for intravenous fluids. Hematocrit should be determined daily especially from the 3rd day until the temperature remains normal for one or two days.
Management of DHF Grade I and II: [6]
Hospitalization is advised in patients who have reduced platelet count and rised Hct and warning signs of dengue fever. All these candidates should be monitored closely to prevent further complications. In critical phase of illness patient may develop of shock is during transition from febrile to abferile phase, which usually occurs after third day. In spite of fluid therapy if the patient has fall in BP, reduction in urine output or other features of shock, the management for Grade III/IV DHF/DSS should be instituted. Oral rehydration along with antipyretics like Paracetamol, sponging, etc. as described should be given.
Management of Shock in DHF Grade III / IV: [6]
Immediately after hospital admission, the hematocrit, platelet count and vital signs should be done periodically.
Treatment algorithm for DHF Grades III and IV is given.
39
FIGURE 19: Volume replacement algorithm for patients with DHF I and II
40
FIGURE 20: Volume replacement algorithm for patients with DHF III
41
FIGURE 21: Volume replacement algorithm for patients with DHF IV
42
CHOICE OF INTRAVENOUS FLUIDS FOR RESUSCITATION:
Colloids may be the preferred choice if the blood pressure has to be restored quickly in patients with narrow range of pulse pressure (<10mm of hg) 0.9% saline NS- Normal plasma chloride ranges from 95 to 105 mmol/L and thus 0.9% Saline is a preferred for initial fluid therapy, however repeated administration of 0.9% saline may produce hyperchloraemic acidosis. In this situation may be a suitable alternative. Ringer’s Lactate should better be avoided in liver failure and in patients taking metformin where lactate metabolism may be abnormal.
Colloids
Colloids are gelatin-based, dextran-based or starch-based solutions. One of the concerns regarding their use is their impact on coagulation. Dextran bind to von Willebrand factor/Factor VIII complex and impair coagulation the most..
Indications for Platelet transfusion: [6]
1. Platelet count less than 10,000cells/cu.mm in the absence of bleeding manifestations (Prophylactic platelet transfusion).
2. Haemorrhage with or without thrombocytopenia. Packed cell transfusion and FFP along with platelets may be required in patients presenting with severe hemorrhage.
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Criteria for admission of a patient:
Constant fever (high grade) despite anti-pyretic therapy
Severe thrombocytopenia, Rising hematocrit
Patients with significant bleeding from any site
Signs of hypotension ,shock and organ involvement
Evidence of plasma leakage (clinical or radiological)
Criteria for discharge of patients:
Afebrile for at least 24 hours without antipyretics
No respiratory discomfort
Platelet count > 0.5 lakh cells/cu.mm
Return of appetite and Good urine output
Minimum 2 - 3 days after recovery from shock
Detection of fluid overload in patients:
Fluid overload is the most common complication in dengue fever treatment.The signs and symptoms include facial puffiness, abdominal distension, breathlessness and wheezing.
Treatment of fluid overload:
The following points should be noted:
Urinary bladder catheterization done to monitor hourly urine output.
44
Furosemide should be administered during dextran infusion because the hyperoncotic nature of dextran will maintain the intravascular volume while furosemide depletes in the intravascular compartment. After furosemide, the vital signs should be monitored every 15 minutes for one hour to look for its effects.
If there is no urine output in response to furosemide, the
intravascular volume status (CVP or lactate) is to be checked. If it is adequate, pre-renal failure is excluded, thus implying that the patient is in an acute renal failure status. These patients may require ventilatory support any time. If the intravascular volume is inadequate /the blood pressure is unstable, check the ABCS and other electrolyte imbalances.
Pleural and abdominal tapping can be life-saving in cases with severe respiratory distress and in case of failure of the above management.
Traumatic bleeding is the most serious complication of the procedure and leads to death. Hence caution needed
Vaccine for dengue infection [6]
As of now there is no licensed vaccine available for dengue. A tetravalent dengue vaccine is under trial. A live attenuated recombinant, tetravalent vaccine called CYD-TDV (Chimeric Yellow fever virus -Dengue virus Tetravalent Dengue Vaccine) may be useful in near future in preventing dengue infection.
45
DENGUE AND CALCIUM:
In dengue fever various biochemical parameters altered with the onset of plasma leakage, particularly in hematocrit, platelet count, elevation of liver enzymes, hyponatremia, and hypocalcemia Etc. These changes are not apparent in non severe dengue cases.
The relationship between serum calcium levels and dengue fever was analyzed by various studies across the world and observed that calcium level is reduced in severe form of dengue infection.
The calcium ion plays an important role in normal cellular function and myocardial contractility. [11].
Hypocalcemia may also be commonly seen in other illness like severe sepsis, traumatic injury. So, it has a relationship to mortality in severely ill patients. The exact cause for this relation is not defined. The following possible mechanisms may be the cause for hypocalcemia, parathyroid insufficiency, defective vitamin D3 synthesis, reduced dietary intake during illness [17, 28].
The measurement of serum calcium is routinely not done in clinical practice for dengue infection. [20-22].
Hypocalcemia in dengue:
Reduced calcium levels are commonly observed in dengue infections.
Sometimes it may present with feature of hypocalcemic tetany. [30]. The prevalence of hypocalcemia in dengue infection is around 80%. [27].
46
Calcium in the dengue pathogenesis:
In in-vitro studies, the presence of calcium ion is obligatory for the cytotoxic activity of the dengue virus and the cell death is associated with increased concentration of intracellular calcium ion [23]. Therefore, it’s postulated that hypocalcaemia in dengue fever could be due to the influx of calcium ions and calcium replacement could enhance the dengue virus activity by increasing the concentration of intracellular calcium ions. This can be supported by in vitro studies showing that calcium channel blockers inhibiting the activity of the influx of calcium ions in to T cells and macrophages and reducing the disease activity of dengue [24]. However, there is only very limited supportive evidence on calcium supplementation in the management of dengue fever [25].
Another study demonstrated that when the medium was calcium depleted production of cytokines by cultured spleen cells was inhibited. On adding calcium back, it was restored. CCB inhibited the transmission of the
suppressor signal and production of suppressor cytokines in a dose-dependent manner [31]Some studies also propose that, in dengue virus infection the production of nitrite is dependent on calcium and can be inhibited by CCB.[32,33] Thus, in dengue fever calcium plays a major role in the immunity.
Role of calcium on myocardium:
Calcium is essential for myocardial functioning. Cardiac involvement in dengue fever has been registered in many studies [34,35] Myocarditis due to
47
dengue may present as ECG changes like sinus bradycardia, sinus tachycardia, diastolic dysfunction , inverted T-wave and pericardial effusion, rised levels of creatine phosphokinase CPK-MB levels. Suggested mechanism is that Calcium storage abnormality in the infected myocardial cells is responsible for
occurrence of myocarditis.
Salgado et al., [36] tried to prove that alteration in calcium level in dengue is targeting myocardial cells resulting in myocarditis. They exposed human skeletal myocytes to dengue virus and studied the intracellular Calcium changes. The viral replication in myocytes was proved by confocal
fluorescence microscopy. It was proposed to be resulting in contractile dysfunction and arrhythmias.
The potential role of calcium in treatment of dengue:
Oral calcium carbonate and vitamin D3 in patients with dengue fever results in better clinical recover and lesser duration of illness. This inference was observed in a study done in Mexico, with less number of dengue fever patients [18]. In another study it was shown that oral calcium carbonate may improve the platelet count in dengue fever patients but currently no RCT’s is available to demonstrate the effectiveness of calcium in preventing dengue fever complications. Hence, oral or IV calcium therapy in the treatment of dengue infection is not routinely included in published guidelines
48
4. MATERIALS AND METHODS:
STUDY POPULATION:
This study was conducted among 100 patients who are admitted with fever in Government Mohan Kumaramangalam Medical College, Salem, in department of General Medicine, between January 2018 and December 2018.
INCLUSION CRITERIA:
• AGE: 15-50 yrs.
• Either sex.
• Patients with hypotension.
• Patients with third space fluid collection like ascites pleural effusion, gallbladder edema evidenced by USG.
• Patients with severe thrombocytopenia.
• Patients with bleeding manifestations.
EXCLUSION CRITERIA:
• Patient refusal.
• Patient with co morbid conditions like diabetes, liver diseases, cardiac failure.
• Patients with fever associated with other illness like URI, LRI, and UTI.
• Patient with co-morbidities like diabetes, chronic liver disease, chronic kidney disease etc…
49
ANTICIPATED OUTCOME:
In severe form of dengue infection there is reduction of serum calcium levels compared with those who are dengue negative or having mild form of dengue infection. It might be helpful in predicting the disease severity in dengue.
DATA COLLECTION:
Demographic and clinical details of the patient are collected using a pre designed profoma. The serum calcium level is measured at 2 days interval after admission.
STUDY PROTOCOL:
The serum calcium values are compared with disease severity . The disease severity is classified based on WHO guidelines. Severe dengue infection is identified by one of the following features including shock, plasma leakage, bleeding manifestations, severe thrombocytopenia.
LABORATORY INVESTIGATIONS:
1. BLOOD: HB, TC, DC, ESR, Hct, platelet count 2. Blood Urea
3. Serum Creatinine
50
4. LFT (SGOT, SGPT) 5. Serum electrolytes.
6. Serum calcium.
7. IgM ELISA 8. NS-1 Antigen
7. Other relevant indications, if required.
SPECIMEN COLLECTION:
An unhemolysed serum or plasma is used.
RADIOLOGICAL INVESTIGATIONS:
1. USG-Abdomen : to find out third space fluid collection
(Such as ascites, pleural effusion, Gall bladder wall edema) 2. CHEST X RAY: to find out pleural effusion.
ESTIMATION OF CALCIUM:
Method: Arsenazo Mode: Endpoint
51
Principle:
Arsenazo combines with calcium ions at PH 6.5 to form a coloured chromophore .The color produced is directly proportional to the concentration of calcium in the sample which is measured at 650nm.
Reagent composition
• Calcium standard:10mg/dl
• Arsenazo III
• Buffer
Procedure
The assay was performed after calibration.
10µl of serum was mixed with 1ml of reagent, mixed and incubated for 10 minute at room temperature and absorbance of standard and sample read against reagent blank at 650 nm.
Calculation
Serum Calcium (mg/dL) = (Abs.T) - (Abs.B) × 10 (Abs.S) - (Abs.B)
Reference Range
Serum calcium 8.5 - 10.5 mg/dl
52
Linearity
The kit is linear up to 16 mg/dl
DIAGNOSIS OF DENGUE:
1. IgM-capture MAC-ELISA testing.
2. ELISA based NS-1 antigen testing.
CLASSIFICATION OF SEVERE DENGUE INFECTION:
(WHO CLINICAL CRITERIA, 2009):
Severe dengue infection (SDI) is defined by one or more of the following:
(i) Plasma leakage that may lead to shock (dengue shock) and/or fluid accumulation, with or without respiratory distress, and/or
(ii) Severe bleeding, and/or
(iii) Severe organ impairment, of which most occur during the critical phase .
ETHICAL CLEARANCE : Obtained
DESIGN OF STUDY : Hospital based observational, prospective study.
53
FINANCIAL SUPPORT : Nil
PERIOD OF STUDY : January 2018 to December 2018.
CONSENT : Individual written and informed consent
DEPARTMENTS ASSOCIATED : Department of microbiology and bio- chemistry
CONFLICT OF INTEREST : Nil
STATISTICAL ANALYSIS:
The collected data were analysed with IBM.SPSS statistics software 23.0 Version. To describe about the data descriptive statistics frequency analysis, percentage analysis were used for categorical variables and the mean
& S.D were used for continuous variables. To find the significant difference in the multivariate analysis the one way ANOVA with Tukey's Post-Hoc test was used. To find the significance in categorical data Chi-Square test was used. In all the above statistical tools the probability value .05 is considered as significant level.
54
5. OBSERVATIONS AND RESULTS:
STUDY DESIGN:
This population is only for study included (62)
*SDI includes warning signs also
Total population under study 100
Dengue positive
(NS-1/ IgM positive/ both) 62
Dengue negative (NS-1/IgM negative/both) 38
Diagnosis NS-1 – 53 IgM - 09
DF without warning signs 21
Dengue with warning signs only
24
Severe dengue infection * 17
GROUPS
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1. AGE DISTRIBUTION
TABLE 1: Age distribution among all fever cases
FIGURE 1: Bar diagram depicting age distribution among all fever cases
;
0.0 5.0 10.0 15.0 20.0 25.0 30.0 35.0 40.0
< 20 yrs 20 - 29 yrs 30 - 39 yrs >= 40 yrs
Percentage
Age
AGE Frequency Percent
15- 20 yrs 36 36.0
20 - 29 yrs 38 38.0
30 - 39 yrs 18 18.0
>= 40 yrs 8 8.0
Total 100 100.0
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2. GENDER DISTRIBUTION
TABLE 2: Gender distribution among all fever cases
GENDER Frequency Percent
Female 42 42.0
Male 58 58.0
Total 100 100.0
FIGURE 2: Pie diagram depicting gender distribution among all fever cases
42.0%
58.0%
Gender
Female Male
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3. COMPARISON BETWEEN AGE WITH GROUPS
TABLE 3: Comparison between age with groups Comparison between Age with Groups
Groups
Total x2 -
value P- value No
Dengu e
Dengue without warnin
g
Dengue with warnin
g
Severe Dengu
e
Age
< 20 yrs
Count 12 8 9 7 36
3.155 0.958
#
% 31.6% 38.1% 37.5% 41.2% 36.0%
20 - 29 yrs
Count 14 8 11 5 38
% 36.8% 38.1% 45.8% 29.4% 38.0%
30 - 39 yrs
Count 9 3 3 3 18
% 23.7% 14.3% 12.5% 17.6% 18.0%
>=
40 yrs
Count 3 2 1 2 8
% 7.9% 9.5% 4.2% 11.8% 8.0%
Total
Count 38 21 24 17 100
% 100.0% 100.0% 100.0% 100.0% 100.0
%
# No Statistical Significance at P>0.05 level
FIGURE 3: Bar diagram depicting comparison between age with Groups INFERENCE: There is no statistical significance between age and fever
groups under study with p-value of 0.958.
0%
10%20%
30%
40%
50%60%
70%
80%90%
100%
No Dengue Dengue without
warning Dengue with
warning Severe Dengue
Percentage
Groups
Age with Groups
< 20 yrs 20 - 29 yrs 30 - 39 yrs >= 40 yrs
58
4. COMPARISON BETWEEN GENDER WITH GROUPS
TABLE 4: Comparison between gender with groups
FIGURE 4: Bar diagram depicting comparison between gender with Groups INFERENCE: There is no statistical significance between gender and fever
groups under study with p value of 0.853
0%
10%
20%
30%
40%
50%
60%
70%
80%
90%
100%
No Dengue Dengue without
warning Dengue with
warning Severe Dengue
Percentage
Groups
Gender with Groups
Female Male
Comparison between Gender with Groups
Groups
Total x2 -
value P- value No
Dengu e
Dengue without warning
Dengue with warning
Severe Dengue
Sex
Female Count 18 8 9 7 42
0.785 0.853
#
% 47.4% 38.1% 37.5% 41.2% 42.0%
Male Count 20 13 15 10 58
% 52.6% 61.9% 62.5% 58.8% 58.0%
Total
Count 38 21 24 17 100
% 100.0% 100.0% 100.0% 100.0% 100.0
%
# No Statistical Significance at P>0.05 level
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5. COMPARISON OF PLATELET COUNT WITH DENGUE STATUS TABLE 5: Comparison of platelet count with dengue status
Comparison of PC with Dengue status by One way ANOVA
N Mean S.D Minimu
m
Maxim
um F-value P- value
PC
No Dengue 38 1.47 0.34 .15 2.03
24.136 0.0005
**
Dengue without
warning 21 1.40 0.31 .99 2.03
Dengue with
warning 24 1.06 0.48 .35 1.93
Severe Dengue 17 0.54 0.49 .10 1.70
** Highly Significant at P < 0.01 level
Tukey's Post-hoc test for Multiple comparison
Dependent Variable Mean
Difference
Std.
Error
P- value
95% C.I Lower Bound
Upper Bound
PC
No Dengue
Dengue without warning
.07163 .10884 0.912
# -.2130 .3562 Dengue
with warning
.41252* .10437 0.001
** .1396 .6854 Severe
Dengue .93505* .11680 0.0005
** .6297 1.2404 Dengue
without warning
Dengue with warning
.34089* .11961 0.027
* .0282 .6536 Severe
Dengue .86342* .13060 0.0005
** .5220 1.2049 Dengue with
warning
Severe
Dengue .52252* .12689 0.0005
** .1907 .8543
** Highly Significant at P < 0.01 level * Significant and # No Statistical Significance at P >0.05 level
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FIGURE 5: Bar diagram depicting Comparison of platelet count with dengue status
INFERENCE:
There is no statistical difference in platelet counts between dengue negative and dengue positive cases without warning signs (p=0.912)
There is high statistical significance in platelet count between dengue negative and dengue with warning signs (p=0.001)/severe dengue infection (p=0.0005)
There is high statistical significance in in platelet count between dengue with warning signs and severe dengue infection (p=0.0005)
0.00 0.20 0.40 0.60 0.80 1.00 1.20 1.40 1.60
No Dengue Dengue without
warning Dengue with
warning Severe Dengue
Mean
Groups
Platelet count with Dengue status
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6. COMPARISON OF HCT WITH DENGUE STATUS
TABLE 6: Comparison of Hct with dengue status Comparison of Hct with Dengue status by Oneway ANOVA
N Mean S.D
Mini mum
Maxi mum
F- value
P- value Hct No Dengue 38 32.22 3.72 24.50 41.00
9.230 0.0005
**
Dengue without
warning 21 36.83 4.97 29.50 45.50 Dengue with
warning 24 35.40 3.11 28.50 42.00 Severe Dengue 17 38.76 7.41 25.00 50.00
** Highly Significant at P < 0.01 level
Tukey's Post-hoc test for Multiple comparison Dependent Variable Mean
Difference
Std.
Error P-value
95% C.I Lower Bound
Upper Bound Hct No Dengue Dengue without
warning -4.60965* 1.27388 0.003 ** -7.9403 -1.2790 Dengue with
warning -3.17215 1.22152 0.052# -6.3660 .0217 Severe Dengue
-6.54102* 1.36700 0.0005
**
-
10.1152 -2.9669 Dengue
without warning
Dengue with
warning 1.43750 1.39989 0.734 # -2.2227 5.0977 Severe Dengue -1.93137 1.52848 0.588 # -5.9278 2.0650 Dengue with
warning
Severe Dengue
-3.36887 1.48513 0.113 # -7.2519 .5142
** Highly Significant at P < 0.01 level and # No Statistical Significance at P >0.05 level
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FIGURE 6: Bar diagram depicting comparison of hct with dengue status
INFERENCE:
There is high statistical significance in hematocrit between dengue negative and severe dengue infection (p=0.0005)
There is no statistical significance in hematocrit between dengue with warning signs and severe dengue infection (p=0.113)
0.00 5.00 10.00 15.00 20.00 25.00 30.00 35.00 40.00 45.00
No Dengue Dengue without
warning Dengue with
warning Severe Dengue
Mean
Groups
Hematocrit with Dengue status
63
