Phylum Sporozoa (Apicomplexa) PDF

Phylum Apicomplexa (Sporozoa) Phylum Apicomplexa (Sporozoa) Apical complex  From phylum Apicomplexa we are going to study malaria parasites and Toxoplasma gondii.  At some stages in their life cycle, they possess a structure called the apical complex, by means of which they attach to and pene...

Phylum Apicomplexa (Sporozoa) Phylum Apicomplexa (Sporozoa) Apical complex  From phylum Apicomplexa we are going to study malaria parasites and Toxoplasma gondii.  At some stages in their life cycle, they possess a structure called the apical complex, by means of which they attach to and penetrate host cells.  They live intracellularly, at least during part of their life cycle Phylum Apicomplexa (Sporozoa) Intracellular parasites TYPES OF REPRODUCTION IN PHYLUM APICOMPLEXA Alternate sexually with asexual reproduction: o Schizogony: asexual reproduction o Gametogony: gametes and zygote are formed o Sporogony: asexual process in which sporozoites are formed o Endodyogeny: A form of asexual reproduction, favoured by parasites such as Toxoplasma gondii, in which two daughter cells are produced inside a mother cell, which is then consumed by the offspring prior to their separation. GENUS PLASMODIUM Genus Plasmodium includes: ▪ Plasmodium vivax ▪ Plasmodium malariae ▪ Plasmodium falciparum ▪ Plasmodium ovale ▪ ▪ ▪ ▪ Plasmodium vivax causes malaria tertiana Plasmodium malariae causes malaria quartana the most severe type of Plasmodium falciparum causes malaria tropica malaria Plasmodium ovale causes malaria ovale (malaria tertiana type)* ✓ Malaria has been known from ancient times. ✓ The name malaria (mal: bad, aria: air) was given in the 18th century in Italy. ✓ Incidence of malaria is more in poor population in rural areas, also in urban areas having bad sanitary condition. VECTORS Human malaria is transmitted by over 60 species of female Anopheles mosquito. The mosquitoes feeds exclusively on fruits and juices, but the female needs at least two blood meals more, before the first batch of eggs can be laid. There are Anopheles mosquitoes in Bulgaria. Anopheles mosquito Life cycle of malaria parasites 1. Asexual phase (occurs in man – intermediate host) – schizogony take place 2. Sexual phase (occurs in female Anopheles mosquito – definitive host) – gametogony еnds and sporogony takes place Life cycle in the definitive host 1. When a female Anopheles mosquito ingests parasitized erythrocytes along with its blood meal, the asexual forms of malaria parasite are digested, but the gametocytes are set free in the stomach of mosquito and undergo further development. 2. The nuclear material and cytoplasm of the male gametocytes divides to produce eight microgametes with long, actively motile, whip-like filaments. 3. 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. Schematic diagram showing formation of microgamete and macrogamete Life cycle in the definitive host 4. The zygote, which is initially a motionless round body, gradually elongates and becomes a vermicular motile form with an apical complex anteriorly. This is called the ookinete. 5. Ookinete penetrates the mosquito stomach wall. It becomes rounded into a sphere this stage is called the oocyst – its nucleus starts dividing, that’s why numerous sporozoites are formed in the oocyst. (It’s the sporogony). sporozoites Life cycle in the definitive host 6. The oocyst ruptures and the sporozoites via hemolymph (fluid, analogous to the blood in vertebrates) move to the salivary glands of mosquito. The mosquito is now infective and when it bites humans, the sporozoites are injected into blood (the sporozoites are infective form for humans). sporozoites LIFE CYCLE IN THE INTERMEDIATE HOST In this asexual phase, the malaria parasite multiplies by division or splitting a process designated to as schizogony*. In humans, schizogony occurs in two locations-in the red blood cell (erythrocytic schizogony) and in the liver cells (exoerythrocytic schizogony or the tissue schizogony). *schizogony – from schizo: to split, and gone: generation Tissue schizogony Erythrocytic schizogony LIFE CYCLE IN THE INTERMEDIATE HOST Exoerythrocytic (tissue) schizogony: During a blood meal, a malariainfected female Anopheles mosquito inoculates sporozoites into the human host. They pass into the bloodstream, where many are destroyed by the phagocytes, but some reach the liver and enter the parenchymal cells (hepatocytes). Liver cell (hepatocyte) Elongated spindle-shaped sporozoites enter liver cells Sporozoites become rounded. They enlarge in size and undergo repeated nuclear division to form several daughter nuclei, each of which is surrounded by cytoplasm. This stage of the parasite is called the tissue (preerythrocytic or exoerythrocytic) schizont or meront. It contains thousands of merozoites. Merozoites rupture the liver cells and enter the bloodstream. Exoerythrocytic schizogony LIFE CYCLE IN THE INTERMEDIATE HOST Exoerythrocytic schizogony: In P. vivax and P. ovale, two kinds of sporozoites are seen, some of which multiply inside hepatic cells immediately to form schizonts (tachyzoites) and others persist and remain dormant (resting phase). The resting forms are called hypnozoites (hypnos: sleep) or bradyzoites. From time to time, some are activated to become schizonts and release merozoites. In P. falciparum and P. malariae no bradyzoites are found. Tachyzoites and bradyzoites are also called tachysporozoites and bradysporozoites. LIFE CYCLE IN THE INTERMEDIATE HOST Erythrocytic schizogony: Тhe merozoiles released by preerythrocytic schizonts invade the red blood cells. The receptor for merozoites is glycophorin, which is a major glycoprotein on the red cells. In the erythrocyte, 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. These young parasites are, therefore called the ring-form trophozoites. Ring-form trophozoite Ring-form trophozoites (young trophozoites) LIFE CYCLE IN THE INTERMEDIATE HOST Erythrocytic schizogony: The parasite feeds on the hemoglobin of the erythrocyte. it does not metabolize hemoglobin completely and therefore, leaves behind a hematin-globin pigment called the malaria pigment or hemozoin pigment. hemozoin pigment Microscopic findings of spleen. Malarial pigment (hemozoin) is seen in macrophages and endothelial cells lining sinus (H&E, ×400). The malaria pigment released when the parasitized cells rupture is taken up by reticuloendothelial cells. Such pigment-laden cells in the internal organs provide histological evidence of previous malaria infection. LIFE CYCLE IN THE INTERMEDIATE HOST Erythrocytic schizogony: As the ring form develops, it enlarges in size becoming irregular in shape and shows ameboid motility. This is called the ameboid form. This trophozoite is called ameboid form (ameboid trophozoite), because produces pseudopods. LIFE CYCLE IN THE INTERMEDIATE HOST Erythrocytic schizogony: The development continues with: mature trophozoite – the plasmodium grows almost as large as the erythrocyte, rounds up, stops making pseudopods and feeding, does not move and its vacuole disappears Mature trophozoite LIFE CYCLE IN THE INTERMEDIATE HOST Erythrocytic schizogony: The development continues with: • immature schizont – the nucleus of mature trophozoite starts dividing by mitosis • mature schizont – a part of the cytoplasm is separated around each nuclear fragment and merozoites are formed. The mature schizont bursts releasing the merozoites into the circulation. The merozoites invade fresh erythrocytes within which they go through the same process of development. LIFE CYCLE IN THE INTERMEDIATE HOST Mature schizont with merozoites and hemozoin LIFE CYCLE IN THE INTERMEDIATE HOST The rupture of the erythrocytes by merozoites releases large quantities of pyrogens. This is responsible for the febrile paroxysms characterizing malaria. Pyrogens substances that increase body temperature. They reach the brain and affect the thermoregulation center. LIFE CYCLE IN THE INTERMEDIATE HOST Gametogony Micro- and macrogametes Red blood cells (RBC) After a few erythrocytic cycles, some of the merozoites that infect RBCs do not proceed to become trophozoites or schizonts but instead, develop into sexually differentiated forms, the gametocytes. LIFE CYCLE IN THE INTERMEDIATE HOST Gametogony Gametogony starts in human, but еnds in the definitive host – female Anopheles mosquito! In all species, the female gametocyte is larger (macrogametocyte) and has cytoplasm staining dark blue with a compact nucleus staining deep red. In the smaller male gametocyte (microgametocyte), the cytoplasm stains pale blue or pink and the nucleus is larger, pale stained. Pigment granules are prominent. Female gametocytes are generally more numerous than the male. A person with gametocytes in blood is a carrier or reservoir. The gametocytes do not cause any clinical illness in the host. Gametogony and sporogony: summary Schizogony: summary Erythrocytic schizogony stages ■ Ring-form trophozoite ■ Ameboid trophozoite Mononuclear forms ■ Mature trophozoite ■ Immature schizont ■ Mature schizont Мultinucleated forms Erythrocytic schizogony proceeds with significant differences in different types of plasmodia, which will be discussed in the next few slides. PLASMODIUM VIVAX  It is believed to account for 80% of all malaria infections. It causes malaria tertiana.  lt is the most common species of malaria parasite in Asia and America.  All stages of erythrocytic schizogony can be seen in peripheral smears. The duration of erythrocytic schizogony is 48 hours, therefore malaria attacks occur every 3th day. A B  The infected erythrocytes are enlarged and show red granules known as Schuffner's dots on the surface. 🔬 A. Trophozoite - often multiply infected with large chromatin dots 🔬 B. Trophozoite - Schuffner dot's can be seen (in Gimesa stained films) 🔬 C. Schizont - 12-14 merozites clustered around dark brown pigment 🔬 D. Gemetocyte - large and fill the cell, pigmented with coarse brown pigment, a microgematocyte is shown due to the diffuse pigment C D PLASMODIUM VIVAX PLASMODIUM FALCIPARUM  The name Falciparum comes from the characteristic sickle shape of the gametocytes of this species.  This is the highly pathogenic of all the plasmodia. It causes malaria tropica.  No bradyzoites occur.  In erythrocytes between 2 and 5 ring-form trophozoites are seen  Binucleate rings are common (resembling stereo headphones) PLASMODIUM FALCIPARUM  The subsequent stages of the asexual cycle – ameboid trophozoite and schizonts are not ordinarily seen in peripheral blood, except in very severe cases of malaria. The presence of P. falciparum schizonts in peripheral smears indicates a grave prognosis.  The infected erythrocytes are of normal size. They show a few coarse brick-red dots which are called Maurer’s clefts.  Its erythrocytic schizogony lasts on average about 24 h (the periodicity of febrile seizures is not clear because individual seizures last a long time and overlap) Plasmodium falciparum Ring trophozoites (more than 1) Crescent gametocyte PLASMODIUM MALARIAE It causes malaria quartana, in which febrile paroxysms occur every 4th day, because the erythrocytic schizogony lasts 72 hours. P. malariae occurs in tropical Africa, Sri Lanka, Burma and parts of lndia, but its distribution is patchy. The old trophozoites are sometimes seen stretched across the erythrocyte as a broad band. These band forms are a unique feature of P. malariae. The mature schizont has an average of eight merozoites, which usually present a rosette appearance. The infected erythrocytes may be of the normal size or slightly smaller. Band form PLASMODIUM MALARIAE Rosette appearance of mature schizont Band form PLASMODIUM OVALE  It causes malaria ovale. Malaria attacks occur every 3th day.  The infected erythrocytes are slightly enlarged. In thin films, many of them present an oval shape with fimbriated margins. This oval appearance of the infected erythrocyte is the reason for the name ovale given to this species. James’s dots in invased erythrocytes are also typical for P.ovale. Malaria – clinical features: ➢ The most typical clinical symptom in malaria is periodic fever, expressed in high temperature and profuse sweating. Anaemia and severe tiredness develop. Hepato- and splenomegaly are observed. In severe forms of malaria tropica, the kidneys and central nervous system may be affected. Coma may occur. ➢ In Bulgaria there is no local malaria. ➢ In addition to mosquito bites, malaria can be spread by blood transfusion, organ transplantations, transmission from the mother to the fetus via the placenta (during delivery), blood manipulations (syringes). RELAPSE  It is seen in inadequately treated P. vivax and P. ovale infections.  In both these species, two kinds of sporozoites are seen: tachyzoites and bradyzoites (hypnozoites).  Reactivation of hypnozoites leads to initiation of fresh erythrocytic cycles and new atacks of malarial fever. Such new attacks of malaria, caused by dormant exoerythrocytic forms, reactivated usually from 24 weeks to 5 years after the primary attack are called relapses. WHICH POPULATION GROUPS ARE RESISTANT TO MALARIA? (INNATE IMMUNITY)  Innate immunity could be due to: A) Duffy negative red blood cells (the invasion of red cells by merozoites requires the presence of specific glycoprotein receptors (Duffy antigens) on the erythrocyte surface. It has been found duffy blood group negative persons are protected from P. vivax infection) WHICH POPULATION GROUPS ARE RESISTANT TO MALARIA? (INNATE IMMUNITY)  Innate immunity could be due to: B) Nature of Hemoglobin:  Hemoglobin E provides natural protection against P. vivax.  P. falciparum does not multiply properly in sickled red cells containing HbS. Maternal antibodies protect babies  Acquired antibody-mediated immunity is transferred from mother to fetus across the placenta and is evident in endemic areas where infants below the age of 3 months are protected by passive maternal antibodies. How can malaria be diagnosed? 1. Diagnosis of malaria can be made by demonstration of malarial parasite in the blood - two types of smears are prepared from the peripheral blood (one is called thin smear and the other is called thick smear). In P. vivax, P. ovale and P. matariae all asexual forms and gametocytes can be seen in peripheral blood. In P. fatciparum infection, only ring form alone or with gametocytes can be seen. How can malaria be diagnosed? 2. Polymerase chain reaction: Polymerase chain reaction (PCR) is increasingly used now for species specification and for detection of drug resistance in malaria. 3. Serology methods: immunological techniques for the demonstration of antibodies against various stage-specific antigens are used. Treatment and prophylaxis of malaria ■ Antimalarial drugs are used for treatment of malaria. ■ All travellers to potentially dangerous areas are advised to take prophylactic antimalarial drugs and limit contact with mosquitoes. ■ it is advisable to avoid sleeping outdoors. ■ The use of clothing that covers as much as possible the body and repellents (chemical substances that repel arthropods) provides reliable protection. GENUS TOXOPLASMA Toxoplasma gondii is an obligate intracellular parasite, first described in 1908 by Nicolle and Manceaux in a small North American rodent called gundi (Ctenodactylus gundi). Toxoplasma gondii Toxoplasma is now recognized as the most common protozoan parasite globally, with the widest range of hosts spread over 200 species of birds, reptiles and mammals, including humans. Ctenodactylus gundi GENUS TOXOPLASMA Definitive host: cats and other felines, in which both sexual and asexual cycles take place. Intermediate host: Birds, rodents, man and other mammals, in which only the asexual cycle takes place. TOXOPLASMA GONDII OCCURS IN 3 FORMS: 1. Trophozoite (tachyzoite): - the trophozoite is crescent-shaped with one end pointed and the other end rounded - electron microscopy reveals an apical complex at the pointed end - It can invade any nucleated cell (without erythrocytes) and replicate within cytoplasmic vacuoles by a process called endodyogeny (internal budding), wherein two daughter trophozoites are formed, each surrounded by a membrane, while still within the parent cell. When the host cell becomes distended with the parasite, it disintegrates, releasing the trophozoites that infect other cells. The actively multiplying trophozoite is seen intracellularly in various tissues during early acute phase of infection. Extracellular trophozoites can also be seen in impression smears ENDODYOGENY TOXOPLASMA GONDII OCCURS IN 3 FORMS: 1. Trophozoite (tachyzoite): - during acute infection, the proliferating trophozoite within host cell may appear rounded and enclosed by the host cell membrane. This is called pseudocyst and can be differentiated from tissue cysts by staining reactions The rapidly proliferating trophozoites in acute infection are called tachyzoites. pseudocyst TOXOPLASMA GONDII OCCURS IN 3 FORMS: bradyzoites 2. Tissue cyst - tissue cysts are the resting form of the parasite - they are found during chronic stage of the infection and can be found in the brain (most common site), skeletal muscles and various other organs. - the cyst wall is eosinophilic and stains with silver, in contrast to the pseudocyst - the slowly multiplying parasites within the cyst are called bradyzoites. tissue cyst TOXOPLASMA GONDII OCCURS IN 3 FORMS: bradyzoites 2. Tissue cyst - the cyst is round or oval and contains numerous bradyzoites. Cysts remain viable in tissue for several years. - In immunologically normal hosts, the cysts remain silent, but in the people with low immunity, they may get reactivated, leading to clinical disease. It is relatively resistant and when the raw or undercooked meat containing the cysts is eaten, infection occurs. • The cyst wall is disrupted by peptic or tryptic digestion and the released parasites initiate infection by invading intestinal epithelial cells. • They reach various tissues and organs through blood and lymphatic dissemination. • Cysts are susceptible to desiccation, freezing, and thawing and heat above 60°C. tissue cyst PSEUDOCYST AND TISSUE CYST (COMPARISON) Pseudocyst • Contains tachyzoites • Trophozoites are enclosed by the host cell membrane • During acute infection Tissue cyst • Contains bradyzoites • Possess real wall • During chronic infection • Can be found in the brain, muscles and various other organs TOXOPLASMA GONDII OCCURS IN 3 FORMS: 3. Oocyst - oocysts develop only in definitive hosts - in the intestine of cats and other felines but not in humans. - the oocysts are formed by sexual reproduction (gametogony). - the freshly passed oocyst is not infectious - oocysts undergo sporulation in the soil with formation of two sporocysts, each containing four sporozoites. The sporulated oocyst (mature oocyst) is infective stage -oocyst is very resistant to environmental conditions and can remain infective in soil for about a year. • when the infective oocyst is ingested, it releases sporozoites in the intestine, which initiates infection. LIFE CYCLE OF TOXOPLASMA GONDII Enteric cycle – occurs in definitive hosts (cats): includes sexual reproduction (gametogony) and asexual reproduction (schizogony) Cat acquires infection by ingestion of tissue cysts in the meat of rats and other animals. The bradyzoites are released in the small intestine and they undergo asexual multiplication (schizogony) in intestine cells, leading to formation of merozoites. Merozoites transform into male and female gametocytes and sexual cycle (gametogony) begins, with the formation of microgamete and macrogamete. LIFE CYCLE OF TOXOPLASMA GONDII Enteric cycle – occurs in definitive hosts (cats): includes sexual reproduction (gametogony) and asexual reproduction (schizogony) A macrogamete is fertilized by motile microgamete resulting in the formation of zygote. After the destruction of a cell containing zygotes, the latter, now called oocysts, fall into the lumen of the intestine. Oocysts are excreted via feces in the soil and passes through maturation stages for 1-5 days (sporogony). A mature oocyst containing eight sporozoites is the infective form which may be ingested by rats or other mammals to repeat the cycle. LIFE CYCLE OF TOXOPLASMA GONDII Exoenteric cycle – occurs in intermediate hosts (human, mice, rats, pigs, birds…): includes only asexual reproduction of parasite Humans acquire infection after: 1. Eating uncooked or undercooked infected meat that contains tissue cyst with bradyzoites 2. Ingestion of mature oocysts through food, water, or fingers contaminated with cat feces directly or indirectly 3. Intrauterine infection from mother to fetus 4. Blood transfusion or transplantation from infected donors. LIFE CYCLE OF TOXOPLASMA GONDII Exoenteric cycle – occurs in intermediate hosts (human, mice, rats, pigs, birds…): includes only asexual reproduction of parasite Sporozoites from the oocysts and bradyzoites from the tissue cysts enter into the intestinal mucosa and multiply asexually and tachyzoites are formed (endodyogeny). Tachyzoites continue to multiply and spread locally by lymphatic system and blood. Some tachyzoites also spread to distant extraintestinal organs like brain, eye, liver, spleen, lung and skeletal muscles and form tissue cysts. Tachyzoites may cross the placenta too. The slowly multiplying forms inside the tissue cysts are known as bradyzoites, which remain viable for years. The dormant bradyzoites inside the cyst may be reactivated in immune suppression causing renewed infection in the host. LIFE CYCLE OF TOXOPLASMA GONDII Exoenteric cycle – occurs in intermediate hosts (human, mice, rats, pigs, birds…): includes only asexual reproduction of parasite Human infection is a dead end for the parasite. Mice eat materials contaminated with oocysts shed in cat's feces. Tissue cysts develop in mice. When such mice are eaten by cats, they get infected and again shed oocysts in feces. ENTERIC AND EXOENTERIC CYLE Pathogenicity and Clinical Features  The outcome of Toxoplasma infection depends on the immune status of the infected person.  Active progression of infection is more likely in immunocompromised individuals. Toxoplasmosis has acquired great importance as one of the major fatal complications in acquired immunodeficiency syndrome (AIDS).  Most human infections are asymptomatic.  Clinical toxoplasmosis may be congenital or acquired. Congenital toxoplasmosis  Congenital toxoplasmosis results when T. gondii is transmitted transplacentally from mother to fetus  Мanifests itself with severe clinical symptoms when the infection of the mother with T. gondii occurred for the first time during pregnancy.  if the mother has old, past (chronic) infection, she has antibodies against the parasite (the fetus is not in danger)  Congenital toxoplasmosis proceeds most commonly with fetal hydrocephalus, calcifications in the brain, and retinal involvement leading to blindness, hepatosplenomegaly. hydrocephalus calcifications in the brain Congenital toxoplasmosis Acquired toxoplasmosis Acquired toxoplasmosis is often asymptomatic. Clinically, it usually manifests with generalized enlargement of the lymph nodes, involvement of the lung, heart and brain is possible. Toxoplasmic encephalitis (inflammation of the brain) is a serious complication also leading to death. Ocular toxoplasmosis may present as uveitis, choroiditis, or chorioretinitis. Toxoplasmosis is the most serious and often fatal in immunocompromised patients, particularly in AIDS (involvement of brain is most common). Diagnosis of toxoplasmosis  Toxoplasmosis is very difficult to prove by establishing of the parasite in tissue preparations. The most common are immunological (serology) methods of diagnosis. They consist in monitoring the titer of specific antibodies against T. gondii  Microscopy: Tachyzoites and tissue cysts can be detected in various specimens like blood, sputum, bone marrow aspirate, cerebrospinal fluid (CSF), amniotic fluid, and biopsy material from lymph node, spleen and brain (Tachyzoites appear as crescent-shaped structures with blue cytoplasm and dark nucleus)  Molecular diagnosis (PCR)  Imaging: MRI and CT scan for brain envolvement Prophylaxis of human toxoplasmosis  good heat treatment of the meat  proper washing of hands and washing of vegetables and fruits before eating  individuals at risk, particularly pregnant women, children and immunocompromised persons should avoid contact with cat and its feces  blood or blood products from seropositive persons should not be given and screening for T. gondii antibody should be done in all blood banks REFERENCES: Created by: Assist. Prof. Dr. Stoyan Stoyanov