Malaria: Introduction, cause, mode of transmission and prevention

Medical Laboratory

1. sporozoa Plasmodium species

2. Definition Malaria definition; Malaria is a parasitic infection caused by genus Plasmodium.or An acute disease by a protozoa of genus plasmodium

3. Epidemiology Malaria is the most important tropical disease known to man. It remains a significant problem in many tropical areas, especially in sub-Saharan Africa and is spreading as a result of environmental changes, including global warming, civil disturbances, increasing travel and drug resistance Malaria can be acute or chronic There are approximately 100 million cases of malaria worldwide with about 1 million of these proving fatal.

4. Epidemiology The parasites invade red blood cells Four species are responsible for the disease; P.falciparum P.vivax P.malariae and P.ovale P.knowlesi

5. Falciparum being of more public health importance Falciparum contributes 90-98% of all infections in population Malariae is second with 1-3% infections as a mono infection Mixed infections of malariae and falciparum contribute about 16% ; Malaria constitutes 25% of child deaths in Africa

6. Pregnant mothers are not spared either Low birth weight, preterm delivery, cerebral malaria, severe malaria anaemia are the major causes of death Travellers, tourists and immigrants may be at high risk Plasmodium parasitize all kinds of vertebrates including reptiles ,avian, rodents, monkeys, buffalo and amphibians

7. Rodent plasmodia Rodent plasmodia include; P.berghei P.elongatum Monkey plasmodia include; P.simium P.simiovale P.anuli P.cynomongi

8. Classification of human malaria parasites Empire :Eukaryote Kingdom : Animalia Phylum :protozoa Sub phylum : Sporozoa ( Apicomplexa ) Class : Telosporia Sub class : Haemosporina Order : Coccidiomorphida Sub order : Haemosporidina Family : Plasmodidae ( haemosporinae ) Genus :Plasmodium Species : P.falciparum,vivax,ovale and malariae

9. Transmission Female Anopheles mosquitoes primary vector; breeds in clear, slow flowing streams Anopheles gambiae Anopheles funestus Anopheles arabiensis

10. Mode of transmission Inoculation/ Bite from an infected female Anophelese mosquito Blood transfusions Transplacental Congenital transmission in falciparum and rearly in vivax / Anophelese makes its bite between 10:00pm to 5:00am indoor Incubation period is between 1-2 weeks Environmental conditions that favour malaria transmission; -Presence of sufficient breeding sites for mosquitoes -A reservoir of parasites is in humans

11. Mode of transmission Some people are natural carriers of malaria parasites Sufficient environmental conditions for mosquitoes and parasites; - Mean temperatures of 18-20 c and relative humidity of above 60% No malaria transmission occurring below 18 c at high altitudes Humans in proximity of breeding sites NB Mosquitoes fly further than 2kms

12. Vector Plasmodium is spread by a female Anophelese mosquito Principally two species are involved; Anophelese gambiae complex Anophelese funestus Anophelese gambiae is the most prevalent Anophelese gambiae include species like Anophelese gambiae arabiesis , Anophelese gambiae quadriannuratus . Anophelese bwambae is found only near semiliki hot springs in Bundibugyo Other species seem not to transmit malaria parasites Anophelese funestus is common in dry seasons

13. Disease burden About 270 million people are infected worldwide Approximately 2 million die of the disease each year globally In sub-Saharan africa,1out of every 20 children die before their 5 th birth day due to malaria Number one cause of ill-health and death in Uganda accounting for; -25%40 % of all outpatient visits at health facilities -20% of all hospital admissions and -23% of deaths among children under five years in Uganda Malaria is endemic in Uganda covering more than 90% of the country

14. Clinical signs and symptoms Simple malaria -Malaise(General feelings of discomfort or uneasiness) -Joint pains -Intermittent fever -Nausea -Vomiting -Dizziness -Muscle aches -Blurred vision -Sweating

15. Signs and symptoms cont … Complicated malaria Severe sweating Temperature of more than 40 c Anaemia Anxiety Confusion Coma

16. Signs and symptoms cont … Miscourage in pregnant women Convulsions Drowsiness(Feeling abnormally sleepy during day) Nightmares Reddish/brown urine

17. Reproduction of plasmodium parasites Plasmodium under goes both sexual and asexual reproduction Sexual reproduction ( Sporogony ) is in an invertebrate host Asexual reproduction ( Schizogony ) is in a vertebrate host Schizogony is divided into two; Pre- erythrocytic Schizogony ( Exo-erythrocytic Schizogony ) Erythrocytic Schizogony

18. Life cycle The malaria parasite life cycle involves two hosts . During a blood meal, a malaria-infected female Anopheles mosquito inoculates sporozoites into the human host . Sporozoites infect liver cells and mature into schizonts , which rupture and release merozoites N ote in P. vivax and P. ovale a dormant stage [ hypnozoites ] can persist in the liver and cause relapses by invading the bloodstream weeks, or even years later .)

19. Life cycle cont … After this initial replication in the liver ( exo-erythrocytic schizogony ) the parasites undergo asexual multiplication in the erythrocytes ( erythrocytic schizogony ). Merozoites infect red blood cells. The ring stage trophozoites mature into schizonts , which rupture releasing merozoites . Some parasites differentiate into sexual erythrocytic stages (gametocytes). Blood stage parasites are responsible for the clinical manifestations of the disease

20. Life cycle cont … The gametocytes, male (microgametocytes) and female ( macrogametocytes ), are ingested by an Anopheles mosquito during a blood meal. The parasites multiply in the mosquito and is known as the sporogonic cycle.

21. Life cycle cont … While in the mosquito's stomach, the microgametes penetrate the macrogametes generating zygotes. The zygotes in turn become motile and elongated ( ookinetes ) which invade the midgut wall of the mosquito where they develop into oocysts . The oocysts grow, rupture, and release sporozoites , which make their way to the mosquito's salivary glands. Inoculation of the sporozoites into a new human host perpetuates the malaria life cycle.

22. Risk factors Un treated persons in the community Favourable climatic conditions for mosquitoes to survive and for the parasite Human settlements near breeding sites for mosquitoes Drug resistant strains of the parasite in the community

23. Vulnerable groups Children below 5 years of age Pregnant mothers Sicklers Those living in slum areas

24. Prevention and control Physical methods; Cutting down grass and bushes Clearing away stagnant waters Meshing windows and ventilators Sleeping in long sleeved clothing Electrocuting adult mosquitoes Screening doors Biological methods; Introducing predators in ponds

25. Prevention and control cont … Chemical methods; Sleeping under ITNs Smearing with insecticide treated substances Oiling stagnant waters Residual indoor spray with insecticides Use of insecticide coils Use of wall paints containing insecticides

26. Management of Malaria Un complicated malaria; First line treatment is; Artemether / Lumefantrine Artesunate+Amodiaquine is the alternative Second line treatment is; Oral quinine for all patients Given only when first line has failed or contraindicated

27. Management of Malaria… Complicated Malaria; Paranteral quinine for all type of patients Paranteral artemisim may be used where quinine is contraindicated Quinine is recommended in pregnancy

28. Management of Malaria… Prophylactic treatment; Sulfadoxine / Pyrimethamine in pregnancy Antipyretics like Paracetamol needed when fever is above 38.5 Aspirin must not be administered to children below 12yrs of age

29. Management of Malaria… Supportive Treatment; Plenty of nourishing fluids and water Encourage continued breast feeding Encourage plenty of rest Encourage a balanced diet Keep re-assuring the patient Cold sponging in complicated malaria Continue monitoring patient for progress

30. Laboratory Diagnosis of malaria Methods; Examination of Romanowsky stained blood smears (Microscopy) Use of Rapid diagnostic Tests (RDTs) Quantitative buffycoart concentration method (QBC) Serology(To detect the antibodies due malaria parasites) Molecular Diagnosis Indirect Florescence Antibody Test

31. Examination of Blood Smears (Microscopy) It is the gold standard technique for malaria confirmation The most commonly used in Uganda Smears must be well made Dried sufficiently at room temperature Stained well with quality controlled stains Examined adequately by knowledgeable personnel Results reported as recommended by WHO standards

32. Problems of microscopy in malaria diagnosis Not very reliable in developing countries because; Insufficient training and supervision of the microscopists Poor quality of the microscopes and reagents used Un reliable supply of electricity Laboratory staff un familiar with malaria Overworked microscopists resulting into limited concentration

33. Specimen Fresh un anticoagulated capillary blood Collected from the tip of the 3 rd or 4 th finger Must be used immediately EDTA anticoagulated venous blood is best used within 2hrs of blood collection

34. Purpose of Thick Blood Smears For easy detection of parasites Detection of WBCs and platelets Useful in situations of mild Parasitemia

35. Purpose of Thin Blood Smears Identification of species of the parasites Reveals well the morphological characteristics of parasites Reveals morphological characteristics of red blood cells Reveals the size relationship between infected and un infected red blood cells Allows detection of parasites inside red blood cells Acts as a permanent record in the lab for future references

36. Preparation of blood smears Requirements; Grease free glass slides Clean cover slips Well mixed blood sample Pasture pipette/capillary tubes Grease pencil Cotton wool Drying rack

37. Preparation of Thick Blood Smears Label the slide with the patient’s number or name Place 2-3 drops of blood in centre of clean slide Mix gently and continue stirring for about 30 seconds Spread over circular area of about 2cm in diameter For anticoagulated blood, do not continue stirring

38. Preparation of Thin Blood Smears Label the slide with patient’s number/name Place a smaller drop of blood at one end of slide Using a cover slip inclined at an angle of 45 Spread the blood quickly using a uniform motion But the film should be evenly distributed one layer thick The feathered end should be at least 2cm long The film should occupy the central area of the slide The film should be away from the margins of the slide

39. Common sources of errors in making blood smears Poor positioning of the smear Too much blood used Too little blood used Using a greasy slides Using spreader with chipped edges Badly scratched slides Thick smear drying un evenly Allowing insects, fingers or dirt to touch smear

40. Drying Blood Smears Allow free air drying of the smears Should be placed horizontally on drying rack Drying times will depend on area humidity Avoid heating smears to force them dry Quick drying produces best results Keep away from insects, dirt and dust

41. Fixing of Blood smears Thick blood smears must never be fixed Thin blood smears must be fixed before staining Absolute methanol is preferred and for 2-3 minutes Water free methanol must be used For leishman /wright’s stains ,stock solution can be used as fixative

42. Staining blood smears for parasites Requirements Stock solutions of Romanowsky stains Staining rack Draining rack Buffered water Cotton wool Washing bottle Pasture pipettes Gloves Large mouthed containers Staining bottles

43. Precautions during staining Avoid under staining of the slides Avoid over staining of the slides Use quality controlled stains Always keep control slides in your lab Use filtered stains Avoid using Romanowsky stains containing water droplets Avoid using un ripe / immature stains Maintain the pH of the water used

44. THE END THANKS