Malaria life cycle, clinical features and management

treatment according to species, prevention

1. Malaria clinical features and management Chair person Dr P.V. Venkataramanappa Professor Dept .of General medicine Presenter Dr Amar Patil PG Gen Medicine

2. Introduction Epidiemology Life cycle Clinical features Severe malaria Malaria in pregnancy Relapse Treatment of uncomplicated and complicated severe malaria Chemoprophylaxis prevention

3. Introduction Malaria continues to pose a major public health threat in india , particularly due to plasmodium falciparum . Malaria occurs mostly in the tropics. India harbours both p.vivax and p.falciparum and contributes to 70% of malarial cases in south-east asian regions.

4. According to the World Malaria Report 2014 , 22% (275.5m) of India’s population live in high transmission (> 1 case per 1000 population) areas, 67%  (838.9m) live in low transmission (0–1 cases per 1000 population) areas and 11% (137.7m) live in malaria-free (0 cases) areas.  In 2013, 0.88 million cases have been recorded, with 128 million tests being conducted on the suspected cases, with  P. falciparum  causing 53% and  P. vivax  causing 47% of the infections. The incidence of malaria in India accounted for 58% of cases in the South East Asia Region of WHO WHO. World Malaria Report 2014. WHO, Geneva. 2014

5. At present, official figures for malaria in India, available at NVBDCP, indicate 0.7–1.6 million confirmed cases and 400-1,000 deaths annually. The biggest burden of malaria in India is borne by the most backward, poor and remote parts of the country, with >90-95% cases reported from rural areas and <5-10% from urban areas. Ashwani Kumar, Neena Valecha , Tanu Jain, Aditya P. Dash. Burden of Malaria in India: Retrospective and Prospective View.  Am J Trop Med Hyg .  2007;77(6_Suppl):69-78.

6.  The state of Orissa, with a population of 36.7 million (3.5%), contributes about 25% of the total annual malaria cases, more than 40% of  P. falciparum  malaria cases and nearly 20–30% of deaths caused by malaria in India. Followed by Meghalaya, Mizoram, Maharashtra, Rajasthan, Gujarat, Karnataka, Goa, southern Madhya Pradesh, Chhattisgarh, and Jharkhand that also report significant number of malaria cases and deaths. A Profile of National Institute of Malaria Research. Estimation of True Malaria Burden in India

7. The parasite The malaria parasite is a mosquito transmitted protozoan. Four species of the genus plasmodium infect the humans , they are P.falciparum , p.vivax , p.ovale , p.malariae . Although infections with the fifth parasite p.knowlesi , is an important cause of human malaria in the islands of borneo and pennisular malaysia .

8. Geographical aspects India is predominantly characterized by unstable malaria transmission, which is seasonal with increased intensity related to rain. Due to low or unstable malaria transmission dynamics, most of the population has no or little immunity toward malaria. Surveys have shown that in some people living in malarious area mainly in forested areas.

9. Epidemiology The mosquito vector Malaria is transmitted by some species of anopheline mosquites . The transmission does not occur at temperature below 16c, or above 33c and at altitudes >2000mts because development in the mosquito cannot takes place. The optimum conditions for transmission are high humidity and an ambient temperature between 20 and 30c.

10. The human host T he behaviour of man also plays an important role in the epidemiology of malaria. There must be a human reservoir of viable gametocytes to transmit the infection. In areas where there is long dry season, and malaria is highly seasonal, the reservior for malaria transmission is in people who asymptomatically harbour parasites for long periods until the next rainy season.

11. Malaria parasite life cycle Infection with human malaria begins when feeding female anopheline mosquito inoculates plasmodial sporozites at the time of feeding. The small motile sporozites are injected during the phase of probing as the mosquito searches for a vascular space before aspirating blood.

12. After injection, they enter the circulation, either directly or via lymph channels(approx 20%) and rapidly target the hepatic parenchymal cells. Each sporozite bores into the hepatocyte and there begins a phase of asexual reproduction. This stage lasts on average between 5.5( p.falciparum ) and 15 days( p.malariae ) before the hepatic schizont ruptures to release merozites .

13. Period of Pre erythrocytic cycle P.vivax 8 days P.falciparum – 6 days P.malariae - 13 – 16 days, P.ovale 9 days

14. In p.vivax and p.ovale infections a proportion of the intrahepatic parasites do not develop, but instead rest inert as sleeping forms or ‘ hypnozoites ’. To awaken weeks or months later and cause relapses which characterize infections with these two species. During the pre- erythrocytic or hepatic phase of development considerable asexual multiplication takes place and many thousands merozoites are released from each ruptured infected hepatocyte .

15. Erythrocytic Schizogony ( asexual blood stage development) The merozoites liberated in the blood stream closely resemble sporozoites . They are motile ovoid forms which invade the red cells rapidly. Once inside the erythrocyte, the parasite lies within the erythrocyte cytosol enveloped by its own plasma membrane . D uring the early stages of development(<12hrs) the small ‘ring forms’ of the four parasites appear similar under light microscope.

16. The young developing parasite looks like a signet ring or in case of p.falciparum like a pair of stereo headphones, with darkly stained chromatin in the nucleus and a circular rim of cytoplasm.

17. Erythrocytic Schizogony Parasites are freely motile in the erythrocytes as they grow they readily consume the contents within the RBC mainly haemoglobin . As the proteolysis of the haemoglobin takes place there is release of haem , it oxidizes to toxic ferric form. Intra parasitic toxicity is avoided by spontaneous dimerization to an inert substance haemozion . This brown or blakish pigment can readily be seen within the digestive vacuole of the growing parasite.

18. Erythrocytic Schizogony At approximately 12-14hrs of development, p.falciparum erythrocyte membrane protein(pfEMP1) on the exterior surface of the infected red cells which mediate attachment of the infected erythrocytes to vascular endothelium. This process is called ‘sequestration’. The other three benign human malarias do not cytoadhere in systemic blood vessels and all stages of development circulate in the bloodstream.

19. Erythrocytic Schizogony Eventually the growing parasite occupies the entire red cell which has become spherical,depleted in haemoglobin , and full of merozoites . It then ruptures so that 6 to 36 merozoites are released destroying the red cells. The released merozoites rapidly reinvade other red cells and start a new asexual cycle.

20. p.falciparum p.vivax p.ovale p.malariae Exoerythrocytic cycle (days) 5.5 8 9 15 Erythrocytic cycle 2 2 2 3 hypnozites no yes Yes No Erythrocyte preference Young can invade all ages Reticulocytes Reticulocytes Old RBCs

21. Sexual stages After a series of asexual cycles in p.falciparum , the parasites develops into sexual forms(gametocytes) which are long lived and motile. The process of gametocytogony takes about 7-10 days in p.falciparum compared to p.vivax where it takes only 4 days. Merozoites differentiate into Male and female gametocyte. One male (contains 8 microgametes ) and one female(macrogamete) are required for mosquito blood meal for infection to occur.

22. Development in the mosquito sporogony Following ingestion in the blood meal of a biting female anopheline mosquito, the male and female gametocytes become activated in the mosquito’s gut. The male and female gametocytes undergo rapid nuclear division.

23. sporogony The motile male microgametes separate and seek female macrogamete, fusion and meiosis then takes place to form a zygote. Within 24 hrs enlarging zygote becomes motile and this forms the ookinete which peneterates the wall of the mosquito gut where it encysts as an oocyst . The oocyst finally bursts to liberate myriads of sporozites into the coelomic cavity of the mosquito.

24. The sporozoites then migrate to the salivary glands to await inoculation into next human host during feeding. The development in the mosquito takes about 8 and 35 days.

25. Expansion of the blood stage infection When the hepatic schizonts rupture they liberate approximately 103- 106 merozoites into the circulation which then again infect the circulating red cells. For the first few stages the host is unware of the brewing infection. May complain of vague non-specific symptoms of malaise, headache, myalgia , weakness or anorexia. The infection increases exponentially before settles around a plateau then declines for several months before finally being eliminated.

26. Clinical features in relation to transmission intensity The clinical manifestations of malaria are dependent on the previous immune status of the host. In areas of intense p.falciparum malaria transmission, asymptomatic parasitaemia is usual in adults.( premunition ) Severe malaria never occurs in this age group it is confined to the first years of life, and becomes progressively less frequent with increasing age.

27. Clinical features in relation to transmission intensity The rate at which age specific acquisition of premunition occurs is proportional to the intensity of malaria transmission. In areas with a constant high level p.falciparum transmission, severe malaria occurs predominantly between 6 months and 3 years of age, milder symptoms are seen in older childrens , and adults are usually asymptomatic and have a low parasitaemias .

28. Mixed species infection The incidence of mixed species infection is always under estimated. In simultaneous infection with p.falciparum and p.vivax , the former suppresses the latter, and the primary vivax malaria infection may not appear until several weeks later. The converse( p.vivax malaria with undiagnosed coincident p.falciparum infection) occurs in approximately 8% of cases.

29. Pyrogenic density The parasitaemia at which fever occurs is termed the ‘ pyrogenic density’. The pryogenic density for p.vivax (<100parasites/ ul ) is generally lower than that of p.falciparum (10000/ ul ). The pyrogenic density is a marker of immunity. High progenic density indicates premunition , and a lower risk of severe disease.

30. Uncomplicated malaria The cardinal feature of malaria is fever. The clinical features of uncomplicated malaria are common to all four species. P.malariae and possibly p.ovale both have a more gradual onset than p.vivax . p.falciparum is unpredictable, onset ranges from gradual to fulminant . The first symptoms of malaria are nonspecific and resemble to influenza.

31. Headache, muscular ache, vague abdominal discomfort, lethargy and lassitude often precede fever by up to 2 days. The temperature rises erraticaly at first with shivering, mild chills worsening headache and loss of appetite.

32. Cold stage : As the temperature rises there is intense headache and muscular discomfort. The patient feels cold, clutches at blankets and curls up shivering and uncommunicative (the chill).there is peripheral vasoconstriction. Hot stage : Within minutes the limbs begin to shake and the teeth chatter and the temperature climbs rapidly to a peak(between 39 and 41.5C). The rigor usually lasts for 10-30 min. Sweating stage : By the end of rigor there is vasodilation and the skin feels hot, a profuse sweat then breaks out.

33. If the infection is left untreated the fever in p.vivax and p.ovale regularizes to a 2 day cycle(tertian) and p.falciparum remain erratic for longer. P.malariae has an 72 hour life cycle, and so in an untreated infection the paroxysm occurs on fourth day termed as ‘ quartan malaria’. P.flaciparum has a daily fever spike.(quotidian fever).

34. The blood pressure is relatively low and there may be symptomatic orthostatic hypotension. Defervescence usually takes 4-8hr. Paroxysms with rigors are more common in p.vivax and p.ovale than in p.falciparum or p.malariae . If no treatment is given the natural infection stabilizes for several weeks or months and then gradually resolves. The duration of illness is proportional to the level of immunity and differs between the parasite species.

35. Pathophysiology of Clinical features Anaemia Renal failure Fluid space and electrolyte changes. Pulmonary oedema Coagulopathy and thrombocytopenia. Blackwater fever Liver dysfunction Acidosis Hypoglycaemia Gastroinstestinal dysfunstion .

36. Anaemia The pathogenesis is multifactorial . It results from the obligatory destruction of red cells containing parasites at merogony . The shortened survival of red cells from which parasites have been extracted by the spleen, and accelerated destruction of non-parasitized red cells all compunded by bone marrow dyserythropoeisis . In severe malaria anaemia develops rapidly because of the rapid haemolysis of the red cells and decline in the haematocrit .

37. Renal failure There is renal vasoconstriction and hypoperfusion in severe falciparum malaria. The renal injury in severe malaria results from acute tubular necrosis. ATN results from renal microvascular obstruction and cellular injury consequent upon sequestration in the kidney and the filtration of nephrotoxins .

38. Fluid space and electrolyte changes. In view of the general vasodilatation and a falling haemtocrit there will be increase in the plasme renin activity, anti diuretic harmone concentrations reflecting an appropriate homeostatic mechanisms to maintain adequate circulating blood volume. Mild hyponaterima and hypochloremia are common in sever malaria. Sreum potassium concentrations are usually normal.

39. Pulmonary oedema Pulmonary oedema in malaria results from a sudden increase in pulmonary capillary permeability. The cause of this increase in the pulmonary permeability is not exactly known, although the presence of sequestered RBC and host leucocytes in pulmonary capillaries may have a role in causing pulmonar y capillary endothelial dysfunction. Acute renal failure, severe metabolic acidosis, and coma are confined mainly to the falciparum malaria. Acute pulmonary oedema may also occur in vivax malaria.

40. Coagulopathy and thrombocytopenia In acute malaria coagulation cascade activity is increased with accelerated fibrinogen turnover, consumption of antithrombin III, reduced factor XIII and increased concentrations of fribrin degardation products. In severe malaria infections the antithrombin III, protein S and protein C are further reduced and prothrombin and partial thromboplastin times may be prolonged. Thrombocytpenia is common to all the four malaria species.

41. Blackwater fever In this condition there is massive intrvascular haemolysis and the passage of ‘coco-cola’ coloured urine. Blackwater (black urine) occurs in four circumstances; 1 When the pateint with G6PD deficiency take oxidant drugs( ex.primaquine , sulphones or sulphonamides ) irrespective of wether they have malaria or not. 2 Occiasonally when G6PD deficiency patients have malaria and receive quinine treatment. 3 In patients with severe falciparum malaria who have normal erythrocyte G6PD levels irrespective of the treatment given.

42. 4 When people who are exposed to malaria self-medication frequently with quinine. Blackwater fever may be associated with acute renal failure, although in the majority of cases renal function remains normal.

43. Liver dysfunction Jaundice is common in adults with severe malaria, and there is other evidence of hepatic dysfunction with reduced metabolic clearance of the antimalarial drugs and failure of gluconeogenesis which contributes to lactic acidosis and hypoglycaemia . Jaundice in malaria appears to have haemolytic , hepatic and cholestatic components. Cholestatic jaundice may persist well into the recovery period. There is no residual liver damage following malaria.

44. Acidosis Acidosis is the major cause of death in severe falciparum malaria, both in adults and children. In severe malaria the arterial, capillary, venous and CSF concentrations of lactate rise in direct proportion to disease severity. Lactate- pyruvate ratios often exceed 30, reflecting tissue hypoxia and anaerobic glycolysis .

45. Gastroinstestinal dysfunstion Abdominal pain may be prominent in acute malaria. Minor stress ulceration of the stomach and duodenum is common in severe malaria. The spleen Is often dark or black from malaria pigment, enlarged, soft and firable . It is full of erythrocytes contaning mature and immature parasites. Soft spleen is in acute infections and hard fibrous enlargement associated with repeated malaria.

46. Hypoglycaemia Hypoglycaemia is an important manifestation of severe malaria. An increased peripheral requirement for glucose consquent upon anaerobic glycolysis , the increased maetabolic demands of febrile illness. The obligatory use of glucose by parasite and failure of hepatic gluconeogenesis and glycogenolysis .

47. Plancental dysfunction Pregnancy increases susceptibility to malaria. This is probably caused by a supression of systemic and placental cell mediated immune responses. There is intense sequestration of p.falciparum infected erythrocytes in the placenta, local activation of pro-inflammatory cytokine production and maternal anaemia . This leads to cellular infiltration and thickening of the syncytiotrophoblast and plancental insufficiency with consequent fetal growth retardation.

48. Malaria in Pregnancy : Double Trouble Malaria is more common in pregnancy compared to the general population. Immuno suppression and loss of acquired immunity to malaria could be the causes. M alaria in pregnancy being more severe, also turns out to be more fatal, the mortality being double (13 %) in pregnant compared to the non-pregnant population (6.5%). Some anti malarials are contra indicated in pregnancy and some may cause severe adverse effects. Therefore the treatment may become difficult, particularly in cases of severe  P. falciparum  malaria. Meghna Desai, Feiko O ter Kuile , François Nosten , Rose McGready , Kwame Asamoa , Bernard Brabin , Robert D Newman. Epidemiology and burden of malaria in pregnancy.  Lancet Infect Dis  2007;7:93–104 

49.  In pregnant women the morbidity due to malaria includes anemia, fever illness, hypoglycemia, cerebral malaria, pulmonary edema, puerperal sepsis and mortality can occur from severe malaria and haemorrhage . The problems in the new born include low birth weight, prematurity, IUGR, malaria illness and mortality.

50. Transfusion malaria Malaria can be transmitted by blood transfusion, needle-stick injury, sharing of needles by infected injection drug users. The incubation in this setting is often short because there is no pre erythrocytic cycle . The clinical features are same as of naturally acquired malarias. Radical therapy with primaquine is unnecessary in transfusion related malaria.

51. Cerebral Malaria A strict definition of cerebral malaria has been recommended for sake of clarity and this requires the presence of unarousable coma, exclusion of other encephalopathies and confirmation of  P. falciparum  infection.   This requires the presence of  P.falciparum   parasitemia and the patient to be unrousable . And other causes ( e.g.hypoglycemia , bacterial meningitis and viral encephalitis) ruled out.

52. To distinguish cerebral malaria from transient postictal coma, unconsciousness should persist for at least 30 min after a convulsion. The deeper the coma, the worse is the prognosis.  If necessary, a lumbar puncture should be performed to rule out bacterial meningitis. However, all patients with  P. falciparum  malaria with neurological manifestations of any degree should be treated as cases of cerebral malaria.

53. Neurological signs in cerebral malaria: As per the definition, patient should have unarousable coma, not responding to noxious stimuli with a Glasgow coma scale of <7/15. Mild neck stiffness may be seen. Retinal haemorrhages occur in about 15% of cases. Pupils are normal. Corneal reflexes are preserved. Papilloedema is unusual and is a sign of poor prognosis.

54. Fixed jaw closure and tooth grinding( bruxism ) The corneal reflexes are preserved except in case of deep coma. Motor abnormalities like decerebrate rigidity, decorticate rigidity and opisthotonus can occur. Deep jerks and plantar reflexes are variable. The patients may also have anemia, jaundice and hepatosplenomegaly .

55. Investigations Lumbar puncture and CSF analysis may have to be done in all doubtful cases and to rule out associated meningitis. EEG may show non-specific abnormalities. CT scan of the brain is usually normal. observations Results CSF Pressure Normal to elevated Fluid Clear. WBCs less than 10/µl. protein and lactic acid levels Elevated

56. Malarial Retinopathy: The malarial retinopathy consists of four main components: 1 Retinal whitening. 2 Vessel changes. 3 Retinal hemorrhages, and 4 Papilledema . The first two of these abnormalities are specific to malaria, and are not seen in other ocular or systemic conditions.

57. Algid malaria Algid malaria is characterized by hemodynamic disorders as shock with pronounced metabolic changes and hypothermia. A number of factors are involved in the development of algid malaria. These include: 1. Pathological phenomena that are associated with the changes in the state of red blood cells and lead to impaired microcirculation ( cytoadherence , sequestration, rosetting );

58. 2. Tumor necrosis factor (TNF) that provokes hypoglycemia, coagulopathy , and impaired erythropoiesis . 3. Altered acid-alkali balance with the development of metabolic acidosis. 4. Gastrointestinal lesion. - Adherence of contaminated red blood cells in the intestinal mucosal vessels induces epithelial ischemic damage. - Impaired absorption of liquid and its loss with vomiting and diarrhea result in acute hypovolemia ;

59. Gram negative septicemia has been associated as an important cause of hypotension in some cases of falciparum infection.  Gram-negative septicemia and possible sites of associated infection should be sought in such patients, e.g. lung, urinary tract (especially if there is an indwelling catheter), meninges (meningitis), intravenous injection sites, intravenous lines .

60.  The presenting feature in some cases of malaria, with a systolic blood pressure less than 80 mmHg a cold, clammy, cyanotic skin, constricted peripheral veins and rapid feeble pulse.  Severe hypotension can also develop suddenly in patients with pulmonary edema, metabolic acidosis, sepsis. massive hemorrhage due to splenic rupture or from the gastrointestinal tract.

61. Postural hypotension may be present. Hypovolumeia (due to reduced fluid intake, high grade fever, sweating, vomiting and diarrhoea ) also may contribute to the reduced pressures.. There may be reduction in visceral perfusion. Septicemia, metabolic acidosis and hypoxia may result in a drop in cardiac function.

62. Relapse Both p.vivax and p.ovale have a tendency to relapse after resolution of the primary infection. Relapse must be distuinguished from recrudescene of the primary infection because of the incomplete treatment. P.falciparum is the usual cause of recrudescent infections and tend to arise 2-4 weeks following treatment . Where as relapses occur weeks or months after the primary infection.

63. The proportion of cases relapsing and the intervals between relapses vary between strains. The pattern is determined largely on the geographical orgin of infections. Over 50% of cases of p.vivax in thailand relapse where as in India the proportion is closer to 20%. The tropical strains have shorter intervals(3-6weeks) compared to subtropical p.vivax which have long gaps.

64. In a famous experiment conducted by Patrick Manson in sept 1900, he infected his 23yr old son with p.vivax through mosquitoes sent from rail from rome to london . His son became ill with double tertian fever, but was treated with quinine and recovered fully. In june 1901, he suddenly become ill again with p.vivax malaria, a relapse interval of 9 months.

65. In recent years a relapse interval of 6 weeks has been quoted widely for p.vivax malaria. But this is an artefact of the use of chloroquine for the treatment, which suppresses the first relapse. (at 3wks) Blood chloroquine levels decline by the time of 6weeks and this is the first to manifest itself. The symptoms of a relapse start more abruptly than in primary infection. They may begin with a sudden chill or a rigor.

66. Definitions of severe malaria are useful for clinical and epidemilogical purposes. Definitions were proposed by WHO in 1990 and 2000. In severe malaria there is often evidence of multiple organ dysfunction. More than one of the criteria are fulfilled. Pateints can be treated for severe malaria even if they don’t fall clearly into any of the criteria. Severe malaria

67. Manifestation Features Cerebral malaria Unarousable coma not attributable to any other cause, with a Glasgow Coma Scale score ≤9; Coma should persist for at least 30 min after a generalized convulsion   Severe anemia Hematocrit <15% or hemoglobin < 5 g/dl in the presence of parasite count >10000/µl Renal failure Urine output <400 ml/24 hours in adults and a serum creatinine >265 µmol/l (> 3.0 mg/dl ) despite adequate volume repletion   Metabolic (Lactic) Acidosis/acidosis Metabolic acidosis is defined by an arterial blood pH of <7.35 with a plasma bicarbonate concentration of <22 mmol /L; hyperlactatemia is defined as a plasma lactate concentration of 2-5 mmol /L and lactic acidosis is characterized by a pH <7.25 and a plasma lactate >5 mmol /L. Hypoglycemia Whole blood glucose concentration of less than 2.2 mmol /l (less than 40 mg/dl). Initial World Health Organization criteria from 1990 

68. Manifestation features   Pulmonary edema or acute respiratory distress syndrome (ARDS) Breathlessness, bilateral crackles, and other features of pulmonary oedema . The acute lung injury score is calculated on the basis of radiographic densities, severity of hypoxemia, and positive end-expiratory pressure Hypotension and shock ( algid malaria) Systolic blood pressure < 70 mm Hg in patients ≥5 years ; cold and clammy skin or a core-skin temperature difference >10   Abnormal bleeding and/or disseminated intavascular coagulation Spontaneous bleeding from the gums, nose, gastrointestinal tract, retinal haemorrhages and/or laboratory evidence of disseminated intravascular coagulation. Repeated generalised convulsions ≥3 generalized seizures within 24 hours Haemoglobinuria Macroscopic black, brown or red urine; not associated with effects of oxidant drugs or enzyme defects (like G6PD deficiency)

69. Added World Health Organization criteria from 2000 Prostration Extreme weakness, needs support Hyperparasitemia 5% parasitized erythrocytes or > 250 000 parasites/µl (in nonimmune individuals

70. Investigations Microscopy : thin and thick smears Rapid diagnostic tests Polymerase Chain Reaction Immunofloroscence

71. Light microscopy of thick and thin stained blood smears remains the standard method for diagnosing malaria. It involves collection of a blood smear, its staining with Romanowsky stains and examination of the Red Blood Cells for intracellular malarial parasites. Thick smears are 20–40 times more sensitive than thin smears for screening of Plasmodium parasites, with a detection limit of 10–50 trophozoites / μl .

72. Thin smears allow one to identify malaria species (including the diagnosis of mixed infections), - quantify parasitemia , and assess for the - presence of schizonts , - gametocytes, and - malarial pigment in neutrophils and monocytes . Jaswant Singh Battacharya (JSB) Stain for thick and thin films:  This is the standard method used by the laboratories under the National Malaria Eradication Programme in India.

73. Rapid diagnostic tests The introduction of simple, rapid, sensitive and increasingly affordable dipstick or card tests for the diagnosis of malaria has been a major advance in recent years. These are based on antibody detection of malaria specific antigens in blood samples. 1 Histidine -rich protein 2 of  P. falciparum   (PfHRP2)   2 Plasmodium   aldolase 3 Parasite lactate dehydrogenase ( pLDH )

74. Histidine -rich protein 2 of  P. falciparum   (PfHRP2)   - Is a water soluble protein that is produced by the asexual stages and gametocytes of  P. falciparum ,  expressed on the red cell membrane surface, and shown to remain in the blood for at least 28 days after the initiation of antimalarial therapy. 

75. Plasmodium   aldolase  is an enzyme of the parasite glycolytic pathway expressed by the blood stages of  P. falciparum   as well as the non-fa1ciparum malaria parasites. Monoclonal antibodies against  Plasmodium  aldolase are pan-specific in their reaction and have been used in a combined ‘ P.f / P.v ’ immunochromatographic test that targets the pan malarial antigen (PMA) along with PfHRP2.

76. Parasite lactate dehydrogenase ( pLDH )  is a soluble glycolytic enzyme produced by the asexual and sexual stages of the liver parasites and it is present in and released from the parasite infected erythrocytes. It has been found in all 4 human malaria species, and different isomers of pLDH for each of the 4 species exist

77.  If the target antigen is present in the blood, a labeled antigen/antibody complex is formed and it migrates up the test strip to be captured by the pre-deposited capture antibodies specific against the antigens and against the labeled antibody

78. False Positivity:  False positive tests can occur with RDTs for many reasons. Potential causes for PfHRP2 positivity, include Persistent viable asexual-stage parasitemia below the detection limit of microscopy (possibly due to drug resistance) Persistence of antigens due to sequestration and incomplete treatment, delayed clearance of circulating antigen (free or in antigen-antibody complexes) and Cross reaction with non- falciparum malaria or rheumatoid factor.

79. False negativity:  On the other hand, false negative tests have been observed even in severe malaria with parasitemias >40000 parasites/µl. This has been attributed to possible genetic heterogeneity of PfHRP2 expression, deletion of HRP-2 gene, presence of blocking antibodies for PfHRP2 antigen or immune-complex formation.

80. Sensitivity:  RDTs for the diagnosis of   P.falciparum   malaria generally achieve a sensitivity of >90% at densities above 100 parasites per µL blood and the sensitivity decreases markedly below that level of parasite density.

81. Peripheral smear RDT’s Test duration 20-60 minutes or more 5-30 minutes Test result Direct visualization of the parasites Color changes on antibody coated lines Capability Detects and differentiates all plasmodia at different stages Detects malaria antigens (PfHRP2/ PMA/ pLDH ) from asexual and/or sexual forms of the parasite Detection threshold 5-10 parasites/µL of blood 1 00-500/µL for P.  falciparum ,  higher for non- falciparum Species differentiation Possible Cannot differentiate among non- falciparum species; mixed infections of  P.falciparum   and non- falciparum appear as  P.   falciparum Quantification Possible Not possible Comparison of Peripheral Blood Smear Examination and RDTs for Malaria

82. Approach to Malaria Diagnosis A clinician who faces a case of fever would need answers to the following questions: Is it malaria? If yes; What is the species? Is it severe? Is it new/ recurrence? Is it active?

83. Approach to Malaria Diagnosis

84. Treatment Classification: Aryl amino alcohols:  Quinine, quinidine (cinchona alkaloids), mefloquine , halofantrine . 4-aminoquinolines:  Chloroquine , amodiaquine . Folate synthesis inhibitors:  Type 1 – competitive inhibitors of dihydropteroate synthase – sulphones , sulphonamides ; Type 2 – inhibit dihydrofolate reductase – biguanides like proguanil and chloroproguanil ; diaminopyrimidine like pyrimethamine . 8-aminoquinolines:  Primaquine . Antimicrobials:  Tetracycline, doxycycline , clindamycin , azithromycin , fluoroquinolones . Peroxides:  Artemisinin ( Qinghaosu ) derivatives and analogues – artemether , arteether , artesunate , artelinic acid. Naphthoquinones :  Atovaquone .

85.  According to anti malarial activity: 1 Tissue schizonticides for causal prophylaxis:  These drugs act on the primary tissue forms of the plasmodia which grow within the liver. Ex : Pyrimethamine and Primaquine . 2 Tissue schizonticides for preventing relapse:  These drugs act on the hypnozoites of P. vivax and P. ovale in the liver that cause relapse of symptoms on reactivation. ex : Primaquine is the prototype drug.

86. 3 Blood schizonticides :  These drugs act on the blood forms of the parasite and thereby terminate clinical attacks of malaria. Ex : chloroquine , quinine, mefloquine , halofantrine , pyrimethamine , sulfadoxine , sulfones , tetracyclines etc. 4 Gametocytocides :  These drugs destroy the sexual forms of the parasite in the blood and thereby prevent transmission of the infection to the mosquito. Ex : Chloroquine and quinine have gametocytocidal activity against P. vivax and P. malariae . - Primaquine has gametocytocidal activity against all plasmodia, including P. falciparum .

87. 5 Sporontocides :  These drugs prevent the development of oocysts in the mosquito and thus ablate the transmission. Primaquine and chloroguanide have this action.

88. Treatment Aim Aims of Treatment To alleviate symptoms : Blood schizonticidal drugs Chloroquine , quinine, artemisinin combinations To prevent relapses : Tissue schizonticidal drugs Primaquine To prevent spread : Gametocytocidal drugs Primaquine for  P. falciparum , Chloroquine for all other

89. Treatment of uncomplicated malaria FIRST-LINE DRUGS IN ENDEMIC AREAS MALARIA DRUG TREATMENT Known chloroquine sensitive p.vivax , p.malariae , p.ovale , p.falciparum Chloroquine 10mg base/kg stat followed by 5mg/kg at 12, 24 and 36 hrs or Amodiaquine 10-12 mg base/kg/day for 3days. Sensitive p.falciparum malaria (Other than north eastern states) Artesunate 4mg/kg per day for 3 days+ sulphadoxine 25mg/kg + pyrimethamine 1.25mg/kg single dose. ( ACT-SP) OR Artesunate 4mg/kg per day for 3 days+ amodiaquine 10mg base/kg per day for 3 days. Multi drug resistant p.falciparum (In north eastern states) Artesunate 4mg/kg per day for 3 days+ mefloquine 25mg base/kg(8mg/kg per day for 3 days) or Artemether-lumefantrine 1.5/9 mg/kg twice daily for three days.i.e adult dose is 4tabs b.d for 3 days. (ACT-AL)

90. Radical treatment Patients with p.vivax and p.ovale infections should also be given primaquine 0.25 mg base/kg daily with food for 14 days to prevent relapse. In mild G6PD deficiency 0.75 mg base/kg should be given once weekly for 6 weeks. Primaquine should not be given in severe G6PD deficiency.

91. Treatment of mixed infections ( p.vivax + p.falciparum ) All mixed infections should be treated with full course of ACT and primaquine 0.25mg per kg body weight daily for 14 days. Mixed infections ( p.vivax + p.falciparum ) In north eastern states Artemether-lumefantrine 1.5/9 mg/kg twice daily for three days.i.e adult dose is 4tabs b.d for 3 days. (ACT-AL )+ primaquine 0.25mg/ kg body wgt daily for 14 days. In other states Artesunate 4mg/kg per day for 3 days+ sulphadoxine 25mg/kg + pyrimethamine 1.25mg/kg single dose. (ACT-SP ) + primaquine 0.25mg/ kg body wgt daily for 14 days.

92. Treatment in pregnancy Anti malarials in pregnancy: All trimesters:  Chloroquine ; Quinine; Artesunate / Artemether / Arteether 2 nd  trimester:  Mefloquine ; Pyrimethamine / sulfadoxine . 3 rd  trimester:   Mefloquine ; ? Pyrimethamine / sulfadoxine . Contra indicated:   Primaquine ; Tetracycline; Doxycycline ; Halofantrine

93. Treatment of uncomplicated p.falciparum cases in pregnancy Primaquine should be avioded in pregnancy. Treatment of uncomplicated malaria 1 st trimester Quinine salt 10mg/kg 3 times daily for 7 days. Combined with clindamycin 10mg/kg twice daily. 2 nd and 3 rd trimester In north eastern states Artemether-lumefantrine 1.5/9 mg/kg twice daily for three days.i.e adult dose is 4tabs b.d for 3 days. (ACT-AL ) In other states Artesunate 4mg/kg per day for 3 days+ sulphadoxine 25mg/kg + pyrimethamine 1.25mg/kg single dose. (ACT-SP )

94. Assesment of therapeutic response Parasite clearance time(PCT) This is the interval between beginning of antimalarial treatment and the first negative blood slide. Fever clearance time(FCT) This is the time from beginning of the antimalarial treatment until the pateint is apyrexial . Approach is to record when temp first falls below 37.5c and then when temperature falls and remains below 37.5c for 24 hours.

95. Clinical manifestations of severe malaria Manifestation/complications Coma Hyperpyrexia Convulsions Hypoglycaemia Severe anaemia (HB <5gm/dl or PCV <15%) Acute pulmonary oedema Acute renal failure Spontaneous bleeding and coagulopathy Metabolic acidosis shock

96. Chemotherapy of severe and complicated malaria Chloroquine sensitive Chloroquine dihydrochloride 10mg base/kg body weight is isotonic fluid IV over 8 hours followed 15mg/kg body weight over next 24 hours. OR Chloroquine dihydrochloride 5mg base/kg body weight in isotonic fluid IV over 6 hours followed by 5mg/kg body weight over next 30 hours. OR If IV infusion is not p ossible chloroquine dihydrochloride 3.5mg base/kg body weight every 6 hours by IM or SC route. Complete course of ACT should be administerd to all cases following a paraenteral treatment as soon as oral treatment is feasible.

97. Chemotherapy of severe and complicated malaria Initial parenteral treatment for atleast 48 hours ( Chloroquine resistant ) Follow up treatment when the patient can take oral medication following parenteral Quinine : 20mg quinine salt/kg body wgt on admission(IV infusion or IM divided doses ) followed by maintainence dose of 10mg/kg 8 th hourly. Full course of area specific ACT: In north eastern states : Artemether-lumefantrine 1.5/9 mg/kg twice daily for three days.i.e adult dose is 4tabs b.d for 3 days (ACT-AL) for 3 days+ PQ single dose on second day. In other states : Artesunate 4mg/kg per day for 3 days+ sulphadoxine 25mg/kg + pyrimethamine 1.25mg/kg single dose. (ACT-SP) for 3 days+ PQ single dose on second day. Artesunate : 2.4mg/kg IV on admission(time =0) , then at 12 and 24hrs, then once daily. Artemether : 3.2mg/kg body wgt IM given on admission then 1.6mg/kg per day.

98. Adverse effects and toxicity Drugs Effects and toxixity Artemisinin derivatives  The most common toxic effects that have been identified are nausea, vomiting, anorexia, and dizziness. More serious toxic effects , including neutropenia , anemia, hemolysis , and elevated levels of liver enzymes. Embryotoxic effects : first trimester, congenital abnormalities, stillbirths, and abortions.   Chloroquine At therapeutic doses, it can cause - Dizziness, headache, - Diplopia , disturbed visual accomodation , - Nausea, malaise, and - Pruritus of palms, soles and scalp. - It can also cause visual hallucinations.

99. Quinine Quinine is a potentially toxic drug. The typical syndrome of quinine side effects is called as cinchonism .  Mild cinchonism consists of ringing in the ears, headache, nausea and disturbed vision. Functional impairment of the eighth nerve results in tinnitus, decreased auditory acuity and vertigo. Visual symptoms consist of blurred vision, disturbed colour perception, photophobia, diplopia , night blindness. Rashes, sweating, angioedema can occur. 

100.  Quinine can also cause renal failure. Massive hemolysis and hemoglobinuria can occur, especially in pregnancy or on repeated use. Quinine stimulates insulin secretion and in therapeutic doses it can cause hypoglycemia. This can be more severe in patients with severe infection and in pregnancy.

101. Sulfadoxine+Pyrimethamine Pyrimethamine can cause skin rashes depression of hematopoiesis . Excessive doses can produce megaloblastic anemia. Sulfonamides cause : - Agranulocytosis . - Aplastic anemia - Hypersensitivity reactions like rashes, fixed drug eruptions, exfoliative dermatitis. - Serum sickness - Liver dysfunction; anorexia, vomiting and acute hemolytic anemia can also occur.

102. Mefloquine Less frequently it can cause - Nightmares - Sleeping disturbances - Dizziness - Ataxia - Sinus bradycardia , sinus arrhythmia. - Postural hypotension. Should be avioded in psychiatric diseases. And in first trimester of pregnancy.

103. Atovaquone  It may cause rash, fever, vomiting, diarrhoea and head ache. Safety in pregnancy, lactation, children, and elderly is yet to be established. Primaquine In therapeutic doses, primaquine is well tolerated. At larger doses, it may cause occasional epigastric distress and abdominal cramps. Patients with deficiency of Glucose 6-phosphate dehydrogenase will develop hemolytic anemia on taking usual doses of primaquine .  Granulocytopenia and agranulocytosis are rare complications.

104. Antimalarial treatment failure Treatment outcome Symptoms and signs Early treatment failure Development of danger signs or severe malaria on days 1-3 in presence of parasitaemia . Parasitaemia on day 2 higher than day 0 Parasitaemia on day 3 >25% of count on day 0 Late treatment failure Development of danger signs or severe malaria after day 3 in the presence of parasitaemia without previously meeting any of the criteria of early treatment failure. Late clinical failure Presence of parasitaemia on any day from 4 to day 28 and axillary temperature of >37.5c without previously meeting any of the criteria of early treatment failure. Adequate clinical response Absence of parasitaemia on day 28 irrespective of axillary temperature without previously meeting any of the criteria of early treat failure, late clinical failure.

105. Chronic complications of malaria Quartan nephropathy Hyper- reactive malarial splenomegaly lymphoma

106. Quartan nephropathy The pattern of renal involvement varies from asyptomatic proteinuria to full-blown nephrotic syndrome. Oedema , ascites or pleural effusions are usual presenting features. Anaemia and hepatosplenomegaly are common. The disease usually progresses to renal failure over 3-5 years. Antimalarial treatment does not prevent progression.

107. Hyper- reactive malarial splenomegaly (tropical splenomegaly ) Most pateints present with abdominal swelling and a dragging sensation in the abdomen. The large, hard spleen is vulnerable to trauma and has increased susceptibility to infections. HMS appears to be a premalignant condition developing into lymphoma in some patients. The enlarged spleen usually regress over a period of time with effective antimalarial prophylaxis.

108. Lymphoma In some countries Burkitt’s lymphoma is the most common malignancy of childhood. It is an uncontrolled proliferation of B lymphocytes and is associated with Epstein-Barr virus infections and malaria. The EB virus cytotoxic T cell response is decreased significantly during acute malaria and there is increased proliferation of EB virus infected lymphocytes. This may predispose to malignant transformation.

109. chemoprophylaxis The National Vector Borne Disease Control Programme (NVBDCP) recommends chemoprophylaxis for selective groups in high  P. falciparum  endemic areas Chemoprphylaxis is recommened for travellers from non-endemic areas and pregnant women in endemic areas. For longer stay of Military and Para-military forces in high  P. falciparum   endemic areas, the practice of chemoprophylaxis should be followed wherever appropriate, e.g. troops on night patrol duty,

110. Drug regimens for prophylaxis Drug Dosage Comments Atovaquone / proguanil Only in areas with chloroquine or mefloquine resistant P.f 250mg of atovaquone and 100 mg of proguanil OD Begin 1-2 days before and for 7 days after leaving such areas. Chloroquine phosphate (Sensitive p.f strains) 300mg base once a week on the same day each week Begin 1-2 days before and for 4 weeks after leaving such areas. Doxycycline ( chloroquine or mefloquine resistant P.f ) 100mg PO OD Begin 1-2 days before and for 4 weeks after leaving such areas Mefloquine ( Chloroquine resistant areas) 228mg base=250mg tablet 1 tablet once a week on the same day each week. Begin 1-2 days before and for 4 weeks after leaving such areas National Drug Policy on Malaria – 2013. Directorate of National Vector Borne Disease Control Programme . Govt. of India. New Delhi. 2013

111. Mefloquine is the only prophylactic drug advised for pregnant women. This drug is generally considered safe in the second and third trimesters of pregnancy. For pregnant women living in endemic areas( pf % >30) IPTp (intermittent preventing therapy in pregnant women) with sulphadoxine-pyrimethamine is recommeneded .

112. Vaccines for Malaria This degree of protection would be extremely difficult to achieve and might not be technically feasible with current vaccine art and science . Many vaccine developers have therefore focused their efforts on creating a vaccine that limits the ability of the parasite to successfully infect large numbers of red blood cells. This would not prevent infection but would limit the severity of the disease and help prevent malaria deaths.  

113. Current Initiatives The PATH Malaria Vaccine Initiative and partner, GlaxoSmithKline Biologicals , published recent Phase 2 trial results showing that the vaccine candidate, RTS,S , has a promising safety and tolerability profile and reduces malaria parasite infection and clinical illness due to malaria. This was the first RTS,S vaccine trial in African infants.

114. Current successful Trails In Phase IIa trials, RTS,S/AS02 protected 40–86% of malaria-naive individuals after artificial challenge. Phase IIb trials demonstrated a partial delay of infection, a 30% reduction in clinical episodes of malaria, and reduction in severe malaria by 58%. A phase III trial of RTS,S has been conducted in 11 countries of sub-Saharan Africa from March 2009 through January 2011, in 15460 children. The first results have shown a reduction in the total number of episodes of clinical malaria by 55.1% and reduction in severe malaria by 47%, both in the older group. The RTS,S Clinical Trials Partnership. First Results of Phase 3 Trial of RTS,S/AS01 Malaria Vaccine in African Children.  New Eng J Med . October 18, 2011. doi : 10.1056/NEJMoa1102287

115. Prevention Malaraia vector control measures Action For individual and family protection For community protection Reduction of human mosquito contact Insecticide treated nets, repellents, protective clothing Insecticide treated nets Destruction of adult mosquitoes Indoor residual spraying, space spraying Destruction of mosquito larvae Domestic sanitation Larviciding of water surfaces, biological control Source reduction Small scale drainage Environmental sanitation, drainage management

116. Anti-adult measures Residual spraying : the spraying of houses with residual insecticides( eg : DDT, malathion , fenitrothion ) is still most effective measure to kill the adult mosquito. Space application : it involves the application of pesticides in the form of fog or mist using special equipment. Individual protection : man vector control can be reduced by other preventive measures such as use of repellents, protective clothing, bed nets.

117. Anti larval measures Larvicides : anti larval measures such as oiling the collection of standing water. Source reduction : techniques to reduce mosquito breeding sites which include drainage, flushing, management of water level, intermittent irrigation.

118. References Parks textbook of preventive medicine. 23 rd edition. The malaria .pg 255-70. Harrisons principles of internal medicine.19 th edition Mansons tropical diseases.22 nd edition section 10,malaria.pg 1201-74. National Drug Policy on Malaria – 2013. Directorate of National Vector Borne Disease Control Programme . Govt. of India. New Delhi. 2013 Meghna Desai, Feiko O ter Kuile , François Nosten , Rose McGready , Kwame Asamoa , Bernard Brabin , Robert D Newman. Epidemiology and burden of malaria in pregnancy.  Lancet Infect Dis  2007;7:93–104 

119. Ashwani Kumar, Neena Valecha , Tanu Jain, Aditya P. Dash. Burden of Malaria in India: Retrospective and Prospective View.  Am J Trop Med Hyg .  2007;77(6_Suppl):69-78. WHO. World Malaria Report 2014. WHO, Geneva. 2014

120. Thank you

121. The recommendations for short term prophylaxis(less than 6weeks) are as follows: 1 antimalarials that have to be taken daily should be started one day before arrival in the risk area. 2 weekly chloroquine should be started 1 week before arrival. 3 weekly mefloquine should be preferbly started 2-3 weeks before departure 4 All prophylactic drugs should be taken with unfailing regularity for the duration of the stay in the malaria risk area, and should be continued for 4 weeks after the last possible exposure to infection.

122. The recommendations for long term prophylaxis(more than 6 weeks) are as follows 1 The person who has taken 300mg of chloroquine weekly for over five years and requires further prophylaxis should be screened twice yearly for early retinal changes. Mefloquine is contraindicated in cases with history of convulsions, neuropsychiatric problems and cardiac problems. Available data on long term chemoprophylaxis with doxycycline is limited.