Sporozoa (Plasmodium and Babesia) PDF

Sporozoa (Plasmodium spp. and Babesia spp.) College of Medical Laboratory Science Our Lady of Fatima University-Valenzuela 1  Classification of Protozoan Parasites Phylum Sarcomastigophora Subphylum Sarcodina...

Sporozoa (Plasmodium spp. and Babesia spp.) College of Medical Laboratory Science Our Lady of Fatima University-Valenzuela 1  Classification of Protozoan Parasites Phylum Sarcomastigophora Subphylum Sarcodina Acathamoeba Entamoeba dispar Endolimax nana Entamoeba gingivalis Entamoeba coli Entamoeba histolytica Iodamoeba butschlii Naegleria fowleri Subphylum Mastigophora Chilomastix mesnili Dientamoeba fragilis Giardia lamblia Trichomonas vaginalis Trichomonas hominis Trichomonas tenax Trichomonas vaginalis Leishmania braziliensis Leishmania donovani Leishmania tropica Trypanosoma cruzi Trypanosoma brucei complex Phylum Ciliophora Balantidium coli 2  Classification of Protozoan Parasites (cont.) Phylum Apicomplexa Babesia spp. Cryptosporidium hominis Cyclospora cayetanesis Isospora belli Plasmodium spp. Toxoplasma gondii Phylum Microspora Enterocytozon bineusi Encephalitozoon spp. Vittaforma cornea Pleistophora spp. Brachiola vesicularum Microsporidium spp. 3  Plasmodium spp. Plasmodium falciparum Plasmodium vivax Plasmodium malariae Plasmodium ovale Plasmodium knowlesi Babesia spp. Babesia microti Babesia divergens Babesia bovis 4 Malaria from Italian word “mal’aria” which means “bad air” Considered to be the most important parasitic disease affecting man (Belizario, 2015) Vector: female Anopheles mosquito 1˚: Anopheles minimus var. flavirostris Others: Anopheles litoralis Anopheles maculates Anopheles mangyamus Final Host: female Anopheles mosquito Intermediate Host: Man Infective stages: sporozoites (man) gametocytes (mosquito) 5  Periodicity/ Febrile Cycle Age of Interval Species Febrile cycle Infected (hours) Erythrocytes RBC of all P. falciparum * Malignant tertian 36-48 stages P. vivax Benign tertian 48 Young RBC P. malariae Quartan 72 Aging RBC P. ovale Ovale tertian 48 Young RBC *Plasmodium knowlesi – Quotidian malaria 6  Periodicity/ Febrile Cycle New/Alternative Species Old Name Name Malignant tertian, Plasmodium falciparum Aestivoautumnal, Subtertian Falciparum malaria malaria Plasmodium vivax Benign tertian malaria Vivax malaria Plasmodium malariae Quartan malaria Malarial malaria Plasmodium ovale Benign tertian malaria Ovale tertian malaria * malaria – associated to P. falciparum 7 Source of Exposure to infection: Vector borne (Arthropod borne) Other modes of transmission: 1. Imported malaria 2. Transfusion malaria 3. Mainline malaria 4. Congenital malaria 8  Vector Biology : Anopheles flavirostris Aquatic Habitat: slow flowing streams; shaded streams Adult biting: Night biting (indoor and outdoor) Adult resting: inside walls 9 10 Microgamete Sporogony + Macrogamete Zygote Ookinete Oocyst Sporozoite 11  The Mosquito Cycle (Sporogony) Exflagellating male gametocytes: the nuclear material and cytoplasm of the male gametocytes divides to produce 8 microgametes with long, actively motile, whip-like filaments. The female gametocyte does not divide but undergoes a process of maturation to become the female gamete or macrogamete. It is fertilized by one of the microgametes to produce the zygote. Ookinete: the zygote, which is initially a motionless, round body, gradually elongates becomes a vermicular motile (traveling vermicule) form with an apical complex anteriorly. Oocyst: rounded into a sphere with an elastic membrane within which numerous sporozoites are formed. Sporozoites: when the oocysts ruptures, the sporozoites enter into the hemocele or body cavity, from where some find their way to the mosquito’s salivary glands. The mosquito is now infective and when it feeds on humans, the sporozoites are injected into skin capillaries to initiate human infection. 12 A. Pre-erythrocytic or Exo-erythrocytic cycle Schizogony Sporozoite infect liver parenchymal cells Schizont Liver cells rupture releasing the merozoites B. Erythrocytic cycle 13  Pre-erythrocytic (Tissue) Stage or Exoerythrocytic Stage The hepatocyte is distended by the enlarging schizont and the liver cell nucleus is pushed to the periphery. These normally rupture in 6–15 days and release thousands of merozoites into the blood stream. Hypnozoites (hypnos: sleep): resting forms in Plasmodium vivax and Plasmodium ovale 14 B. Erythrocytic Cycle Schizogony Merozoites invade RBC Ring Form (young trophozoite) Mature Trophozoite Schizont Rupture of RBC releasing the merozoites Develop into a micro or macrogamete 15  Erythrocytic Stage The merozoites released by pre-erythrocytic schizonts invade the red blood cell: The receptor for merozoites is glycophorin, which is a major glycoprotein on the red cells. Merozoites are pear-shaped bodies, about 1.5 μm in length, possessing an apical complex (rhoptery). They attach to the erythrocytes by their apex. Ring forms or young trophozoites: the merozoite loses its internal organelles and appears as a rounded body having a vacuole in the center with the cytoplasm pushed to the periphery and the nucleus at one pole. Malaria pigment or haemozoin pigment: parasite feeds on the hemoglobin of the erythrocyte but it does not metabolize hemoglobin completely and therefore, leaves behind a hematin- globin pigment as residue. 16 17  Erythrocytic Stage The appearance of malaria pigments varies in different species as follows: P vivax: numerous fine golden-brown dust-like particle P. falciparum: few 1–3 solid blocks of black pigment P. malariae: numerous coarse dark brown particles P ovale: numerous blackish brown particles. Amoeboid form or late trophozoite form: as the ring form develops, it enlarges in size becoming irregular in shape and shows amoeboid motility. When the amoeboid form reaches a certain stage of development, its nucleus starts dividing by mitosis followed by a division of cytoplasm to become mature schizonts or meronts. The merozoites invade fresh erythrocytes within which they go through the same process of development. The rupture of the mature schizont releases large quantities of pyrogens. This is responsible for the febrile paroxysms characterizing malaria. 18 19 20 21  Gametogony Development of gametocytes generally takes place within the internal organs and only the mature forms appear in circulation. The mature gametocytes are round in shape, except in P. falciparum, in which they are crescent-shaped. In all species, the female gametocyte is larger (macrogametocyte) than the male gametocyte (microgametocyte). The gametocytes do not cause any clinical illness in the host, but are essential for transmission of the infection. A gametocyte concentration of 12 or more per cumm of blood in the human host is necessary for mosquitoes to become infected. 22 23 24 25  Intervals Species Prepatent period Incubation period P. falciparum 11-14 days 8-15 days P. vivax 11-15 days 12-20 days P. malariae 3-4 weeks 18-40 days P. ovale 14-26 days 11-16 days 26  Clinical Feature Benign Malaria The typical picture of malaria consists of periodic bouts of fever with rigor, followed by anemia and splenomegaly. Severe headache, nausea, and vomiting are common. CLASSICAL MALARIA PAROXYSMS 1. Cold stage sudden coldness and apprehension mild shivering turns to teeth chattering and shaking of the whole body may last for 15 to 60 minutes 2. Hot stage/ flush phase : best stage to collect blood sample high temperature (40-41˚C), headache, palpitations, epigastric discomfort, thirst, nausea and vomiting patient is confused and delirious may last for 2 to 6 hours 3. Sweating stage (Defervescence or Diaphoresis) profuse sweating, temperature lowers and symptoms diminishes may last for 2 to 4 hours 27 Malignant Tertian Malaria The most serious and fatal type of malaria is malignant tertian malaria caused by P. falciparum. Pernicious malaria has been applied to a complex of life-threatening complications that sometimes supervene in acute falciparum malaria. Cerebral Malaria: is the most common cause of death in malignant malaria, capillary plugging of cerebral microvasculature, which results in anoxia, ischemia, and hemorrhage in brain. Blackwater fever: malarial hemoglobinuria, is sometimes seen in falciparum malaria. Clinical manifestation include bilious vomiting and prostration, with passage of dark red or blackish urine (black water). There is a massive intravascular hemolysis caused by anti-erythrocyte antibodies, leading to massive absorption of hemoglobin by the renal tubules (hemoglobinuric nephrosis). 28  Algid Malaria: peripheral circulatory failure, rapid thready pulse with low blood pressure, and cold clammy skin. There may be severe abdominal pain, vomiting, diarrhea, and profound shock. Septicemic malaria: high continuous fever with dissemination of the parasite to various organs, leading to multiorgan failure. Death occurs in 80% of the cases. Merozoite-induced Malaria Injection of merozoites can lead to direct infection of red cells and erythrocytic schizogony with clinical illness. Such merozoite-induced malaria may occur in transfusion malaria, congenital malaria, renal transplantation and mainline malaria. Tropical Splenomegaly Syndrome Also known as hyper-reactive malarial splenomegaly (HMS) is a chronic benign condition seen in some adults in endemic areas, mainly tropical Africa, New Guinea, and and Vietnam. Abnormal immunological response to malaria causing splenomegaly, high titers of circulating anti-malaria antibodies and absence of malaria parasites in peripheral blood smears, hypergammaglobulinemia (IgM), cryoglobulinemia reduced C3, and presence of rheumatoid factor without arthritis. 29 Recrudescence: new malarial attacks that appear after a period of latency usually within 8 weeks after the primary attack and resulting from persistence of the erythrocytic cycle of the parasites. Relapse: common to P. vivax and P. ovale infections, as result from the reactivation of hypnozoite forms of the parasite in the liver. Recrudescence Relapse Seen in P. falciparum and P. malariae Seen in P. vivax and P. ovale Due to persistence of the parasite at a Due to reactivation of hypnozoites subclinical level in circulation present in liver cells Occurs within a few weeks or months of Occurs usually 24 weeks to 5 years after a previous attack the primary attack Can be prevented by adequate drug Can be prevented by giving primaquine therapy or use of newer antimalarial to eradicate hypnozoites drugs in case of drug resistance 30  Pathological Process of the RBC 1. Poikilocytosis and Anisocytosis 2. Altered RBC membrane transport 3. RBC stiffness and cytoplasmic viscosity 31  Morphology Parameter P. falciparum P. vivax P. malariae P. ovale Normal or sl. Normal - sl. 1. Size of RBC Normal Enlarged smaller enlarged Usually not 2. Trophozoite Ameboid Band form fimbriated present 3. No. of merozoite in 6-12 in rosette 8-36 12-24 8 schizont form Maurer’s, 4. Stipplings Stephen’s, Schuffner’s Ziemman’s Schuffner’s (James) Christopher’s 5. Ring forms Single, multiple single Single Single 6. Chromatin dot Single, double Single, dense, big Single single 7. Applique or accole Present - - - Macro-crescent Large, round, oval Large, round, oval 8. Gametocyte Micro- banana, Large, round, oval sausage shape 9. Stages in peripheral Ring forms and all all all blood gametocytes 32 Trophozoite (ring form) Trophozoite (band form) P. falciparum P. malariae Mature schizont Macrogametocyte and (P. falciparum) microgametocyte (P. falciparum) 33 Schizont (P. malariae) 34  Immunity Duffy negative RBCs: It has been found that persons, who lack the Duffy blood group (Fya and Fyb alleles) antigen, are refractory to infection by P. vivax. These genetically determined blood group antigen appears to be the specific receptor for P. vivax. Nature of hemoglobin: Hemoglobin E provides natural protection against P. vivax. P. falciparum does not multiply properly in sickled red cells containing HbS. Sickle cell anemia trait is very common in Africa, where falciparum malaria is hyperendemic and offers a survival advantage. HbF present in neonates protects them against all Plasmodium species. G6PD deficiency: Innate immunity to malaria has also been related to G6PD deficiency found in Mediterranean coast, Africa, Middle East, and India. HLA-B53: HLA-B53 is associated with protection from malaria. There is some evidence that severe malnutrition and iron deficiency may confer some protection against malaria. 35  Diagnosis 1. Microscopy (Gold Standard) - “Thick and Thin Blood Smear” - stained with Giemsa or Wright’s stain - perform multiple sets of blood films (blood collected every 6 to 12 hours for up to 48 hours) Manner of Reporting A. Qualitative + = 1-10 parasite/100 thick field ++ = 11-100 parasite/100 thick field +++ = 1-10 parasite/thick field ++++ = more than 10/ thick field B. Quantitative Malaria parasite/uL = no. of parasites x 8,000 WBC 2. Quantitative Buffy Coat (QBC) – uses a special capillary tube with acridine orange (+) bright green and yellow under fluorescent microscope 36  Diagnosis 3. Rapid Diagnostic Test (RDT) – detects Plasmodium-specific antigens in finger prick sample a. Histidine-rich protein II (HRP II) – water soluble CHON produced by trophozoites and young gametocytes (e.g., Paracheck Pf test, ParaHIT f test) b. Plasmodium LDH – produced by both sexual and asexual stages and can distinguish between P. falciparum and non-P. falciparum (DiaMed OptiMAL IT) 4. Serologic Tests (IHA, IFAT, ELISA) 5. Molecular Methods through PCR (low cases and mixed infection) 37  Treatment Protective Chemoprophylaxis: Objective is to prevent infections in non-immune person visiting endemic areas (Mefloquine and Doxycycline) Curative Therapeutic: Objective is to eradicate the erythocytic cycle and clinical cure. Radical cure: Objective is to eradicate the exoerythrocytic cycle in liver to prevent relapse. Artemether-Lumefantrine (Coartem TM) – first line drug for confirmed P. falciparum cases. Not recommended in pregnancy, lactation and infants. Quinine (plus Tetracycline or Doxycycline) – second line drug for confirmed P. falciparum cases which AL fail or not available. Quinine IV drip – drug of choice for complicated or severe P. falciparum malaria. Preventive Gametocidal: Objective is to destroy gametocytes to prevent mosquito transmission and thereby reducing human reservoir. In addition to AL and Q+T,D, Primaquine is given on the 4th day as single dose to prevent transmission 38  Prevention 1. Use of mosquito repellant 2. Use of insecticide treated nets (ITN) 3. Take prophylactic medication 4. Wearing of light-colored clothing which cover most of the body Control 1. Environmental cleanliness (stream cleaning to speed up water flow and exposing to sunlight) 2. Indoor residual spraying 3. Zooprophylaxis – use of carabao to deviate mosquitoes 4. Use of biologic control methods a. Bacillus thuringiensis – larvicidal b. Larviparous fishes (e.g., Oreochromis niloticus) 39  Plasmodium knowlesi - A primate malarial parasite common in South East Asia - Causes malaria in long tailed macaques (Macaca fascicularis) - May also infect humans - The appearance of P. knowlesi is similar to that of P. malariae. - PCR assay and molecular characterization are the most reliable methods for detecting and diagnosing P. knowlesi infection *however, P. vivax appears to interfere PCR testing (cross-reactivity) 40  Babesia spp. (Babesia microti, Babesia divergens and Babesia bovis) - First described to cause “Texas cattle fever or red water fever” - Blood parasites that cause malaria-like infections - “Babesiosis” – pathology due to Babesia spp. - Parasites divide through binary fission or budding - Cycle in the tick is still uncertain Vector: Ticks (Ixodes scapularis) - Hard ticks - Scapularis – capable of carrying Borrelia burgdorferi → CA of Lyme disease 41 Definitive host: Ixodid ticks Intermediate host: Man or other mammals Infective form: Sporozoites Mode of Transmission: bite of the nymphal stage of Ixodid ticks Other modes of transmission: - Blood transfusion - Organ transplantation - Transplacental route Diagnostic stage: “Maltese cross” arrangement of the merozoites and ring-form trophozoite 42 43  Pathology - Associated with excessive pro-inflammatory cytokines such as the tumor necrosis factor (TNF) - Most cases are subclinical and may occur as self-limiting - Headache, high-grade fever, chills, vomiting, myalgia, DIC, hypotension, respiratory distress and renal insufficiency. Diagnosis 1. Microscopy of the Giemsa-stained peripheral blood smear a. Merozoites in Maltese cross arrangement b. Ring form → most frequent intraerthrocytic form found 2. PCR (gold standard) 3. Immunofluorescent assays (IFA) 4. Immunochromatographic test (ICT) 5. Hamster Intraperitoneal Inoculation 44 45 Treatment Clindamycin – Drug of choice Drug combination: Clindamycin and Quinine or Azithromycin and Atovaquone Chloroquine – former drug of choice (it only improve the symptoms but not the degree of the parasitemia) In the Philippines: human babesiosis is not yet reported however, it could be present in dogs (Babesia canis). Prevention and Control - avoidance of places where ticks are usually found - wearing of light-colored pants tucked into one’s socks - tick check (especially for children) - rodent population should be controlled 46 REFERENCES Belizario, V. and De Leon, W. (2015). Philippine Textbook of Medical Parasitology. Third Edition. University of the Philippines Manila. Ermita, Manila. Peter L. Chiodini BSc MBBS PhD MRCS FRCP FRCPath FFTMRCPS(Glas), Anthony H Moody, and D. W. Manser (2001). Atlas of Medical Helminthology and Protozoology. The London School of Hygiene & Tropical Medicine. London, United Kingdom. Mikhail A. Valdescona, RMT, MPH. PAR313 Lecture. Our Lady of Fatima University. Valenzuela City. 47