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Learning Objectives Opportunistic Parasites of Humans General 06/07/64 Learning objectives Human Pathogens 2 • After class, students will be able to: Protozoa infections in • Describe morphology, life cycle, signs and symptoms, immunocompromised host epidemiology, prevention and control, laboratory diagnosis and treatment of opportunistic protozoan Nimit Morakote, Ph.D. parasites of man. 23 July 2021 1 2 General characteristics of Opportunistic parasites of humans apicomplixans • Sporozoa (Phylum • Fungi • Obligate intracellular parasite Apicomplexa) • Pneumocystis jirovecii • Apical complex organelles for • Toxoplasma gondii • Microsporidia host cell invasion • Conoid • Cystoisospora belli • Polar ring • Cryptosporidium spp. • Rhoptry • Cyclospora cayetanensis • Microneme • Subpellicular microtubules • Sexual and asexual reproduction http://www.nature.com/scitable/topicpage/the-apicoplast-an-organelle-with-a-green-14231555 3 4 1 06/07/64 Eukaryote classification, 2019 • Apicomplexa • Aconoidasida • Haemospororida (Plasmodium, etc.) • Piroplasmida • Conoidasia • Coccidia • Adeleorina • Eimeriorina (Cystoisospora, Toxoplasma, Sarcocystis, Cyclospora, etc.) • Gregarinasina • Archigregarinorida • Eugregarinorida • Neogregarinorida Drawing showing conoid structure • Cryptogregarinorida (Cryptosporidium) Anderson-White B, et al. Int Rev Cell Mol Biol 2012;298:1-31. Adl SM, et al. Revisions to the Classification, Nomenclature, and Diversity of Eukaryotes. J Eukaryot Microbiol 2019, 66, 4–119. 5 6 Host cell invasion- parasite active mechanism Asexual reproduction Internal budding • Endodyogeny (1 mother cell → 2 daughter cells) • Endopolygeny (1 mother cell → many daughter cells) Bang Shen, L David Sibley. The moving junction, a key portal to host cell invasion by apicomplexan parasites. Current Opinion in Microbiology, Volume 15, Issue 4, 2012, 449–455 7 8 2 06/07/64 Multiple fission (merogony or schizogony) 1 cell/ 1 nucleus ↓ 1 cell/ multiple nuclei (schizont or meront) ↓ many cells (merozoites) Apicomplexan asexual cell division by endodyogeny. (A–E) Show progressive steps of cell division as observed for T. gondii tachyzoites. Two daughters bud internally, while the mother’s cytoskeleton is maintained and only destabilized just before emergence of nearly mature daughters (E). Daughter buds emerge and the plasma membrane from the mother is incorporated onto the daughters. The mother falls away as a residual body. (Gubbels M-J, et al. (2020) Fussing About Fission: Defining Variety Among Mainstream and Exotic Apicomplexan Cell Division Modes. Front. Cell. Infect. Microbiol. 10:269.) 9 10 Sexual reproduction Macrogametes Macrogamonts Zoite #1 (female sex cells) Asexual schizont Microgametes Microgamonts Zoite #2 multiplication (male sex cells) 2, Disassembly of the mother’s cytoskeleton shortly following completion of host cell invasion, resulting in an amoeboid or pleomorphic cell. Fertilization 3, Several cycles of DNA replication and nuclear division. Mitotic cycle not synchronous. Sexual Zygote 4, The last cycle of nuclear division is synchronous and coupled to the (Sporogony) Sporulated synchronous budding of the daughters at the end of schizogony. oocysts Oocysts (Sporozoites Gubbels M-J, et al. Fussing About Fission: Defining Variety Among Mainstream and Exotic Apicomplexan Cell Division Modes. sporulation inside) Front. Cell. Infect. Microbiol. 2020;10:269. 11 12 3 06/07/64 Oocyst sporulation Opportunistic protozoan parasites Meiosis and Life cycle Affected organs/ Oocyst wall mitosis Disease in man Zygote sporoblasts sporocysts sporozoite or 1N Toxoplasma gondii Heteroxenous Brain-Toxoplasmic encephalitis sporont (haploid) Cat=DH Developing organs-Congenital 2N (diploid) toxoplasmosis Cystoisospora belli homoxenous Small intestine-Diarrhea Cryptospridium homoxenous Small intestine-Diarrhea Cyclospora cayetanensis homoxenous Small intestine-Diarrhea Unsporulated Sporulated oocyst oocyst 13 14 Toxoplasma gondii Intermediate hosts Asexual reproduction in cells and tissues • Disease: toxoplasmosis • Congenital toxoplasmosis Immature oocysts -> sporulated oocysts • Toxoplasmic encephalitis in AIDS • Stages in life cycle • Tachyzoites, 6 x 2 µm: during acute stage of infection carnivorism Definitive host • Bradyzoites, during chronic stage of infection Sexual reproduction in • with glycogen granules the small intestine • Pepsin-resistant Toxon = arc • Oocyst, 10 x 12 µm Gondii= gundi rodent tissue cysts containing bradyzoites Image from https://www2.bc.edu/~gubbelsj/Toxoplasma.html 15 16 4 06/07/64 Watch video Phase in cats (enteroepithelial) • Toxoplasma gondii - Life • Ingestion of tissue cysts containing cycle and Invasion bradyzoites. https://www.youtube.com/ • Bradyzoites infect the small intestine. watch?v=25SjWK9smnw • Enteroepithelial stage: schizogony, gametogony, fertilization. • Immature oocysts released in feces. Prepatent period 3-10 days. Toxoplasma cyst. Attias M, et al. Parasites Vectors (2020) 13:588. 17 18 Sporulation in environment Asexual phase in intermediate hosts • Immature oocysts undergo sporulation to mature, infective oocysts in soil or environment. • Sporulation requires 1-5 days • Ingestion of sporulated oocysts • Sporozoites enter epithelial cells, multiply by endodyogeny -> tachyzoites, hematogenous www2.bc.edu spread (acute phase) 19 20 5 06/07/64 Chronic phase of infection Bradyzoites in tissue cyst Human infection • Tissue cysts • Consumption of improperly cooked meat (bradyzoites) • Sporulated oocysts • Drinking unfiltered water (oocysts) • Toxoplasma cyst in mouse brain. Contaminated hand (oocysts) Attias M, et al. Parasites Vectors (2020) 13:588. • Tachyzoites • Organ transplant, blood transfusion (tachyzoites) From: Structures of Toxoplasma gondii Tachyzoites, Bradyzoites, and Sporozoites and Biology and Development of Tissue Cysts. Clin. Microbiol. Rev. 1998 11(2): 267-299; published 1 April 1998 21 22 Pathogenesis, Signs and Symptoms Diagnosis, prevention & treatment • Acute phase- tachyzoites invade and destroy • Isolation of parasite impractical host cells • Primarily diagnosed by serology (antibody detection) • Lymphadenopathy, flu-like symptoms • Chronic phase • Serological survey shows worldwide distribution including • Immune pressure: Tachyzoites bradyzoites Thailand (tissue cysts) in brain, muscle • Prevention by cook meat, proper handling of cat’s feces • Become asymptomatic • • Reactivate in AIDS toxoplasmic encephalitis Treatment: pyrimethamine and sulfadiazine, plus folinic acid. • Congenital toxoplasmosis: chorioretinitis, Austincc.edu hydrocephalus, intracerebral calcification 23 24 6 06/07/64 Laboratory demonstrations http://www.cdc.gov/parasites/toxoplasmosis/resources/printresources/catowners.pdf 25 26 Cystoisospora belli • Former name= Isospora belli • Diagnosis: • Only in human and primate • Fecal exam: simple smear or acid fast stain • Unsporulated oocyst, 20-23 x 10-19 m • Merogony, gametogony, oocyst formation in epithelial cell • of the small intestine Contains 1 or 2 sporoblasts • Treatment: trimethoprim and • Oocyst sporulation outside host: 1-5 days sulfamethoxazole • Acute infection: diarrhea, self-limited • Immunodeficient person: severe diarrhea 27 28 7 06/07/64 Unstained faecal smear Laboratory demonstrations Cryptosporidium (marvistavet.com) • Facultative gregarine parasite (Order Cryptogregarinorida) • Parasite of mammals, birds, reptiles, fish, amphibians: rather host-nonspecific • Human: mostly C. parvum, C. hominis • Oocyst 5 µm in diameter, smallest of all oocysts of human parasites • 4 naked sporozoites (oocyst without sporocyst) Jstor.org 29 30 • Life cycle similar to Cystoisospora belli (development occurs in GI epithelium) except: • Organism is intracellular, extracytoplasmic • Sporulation in host cells- • 80%=thick-wall oocysts • 20%=thin-wall oocysts (autoinfective) • Prepatent Period app. 2 days, Sateriale E, et al. PNAS 2021 Vol. 118 No. 2 Cunha FS, et al. New insights into the detection and molecular characterization of Cryptosporidium • Incubation Period app. 7-10 days e2007807118 with emphasis in Brazilian studies: a review. Rev Inst Med Trop Sao Paulo. 2019;61:e28. 31 32 8 06/07/64 • Emerging infectious disease • Infected persons secrete large • Impaired intestinal absorption + amount of oocysts in feces enhanced secretion • Infectious dose 10-100 oocysts • Watery diarrhea, abdominal cramp, fever • The median maximum number of stool per day is 12 • Self-limited in 2 wk. From: Davies AP, Chalmers RM. Cryptosporidiosis. BMJ | 24 october 2009 | Volume 339:963. 33 34 • An important opportunist in • Outbreak in EU: settings or • Outbreaks associated AIDS patients vehicle with untreated • Fulminant infection, 2L • Nursery school recreational water- US watery stool daily (CD4 < • Playing on floodplain 2000-2014 50/microliter) • Open farm • Veterinary students • 4,958 cases of disease • Extraintestinal dissemination and two deaths can occur, most common= • Frisée salad biliary tract • Bird contact • 12% by Cryptosporidium • Swimming pool • In population, most • Drinking water • Multiply in biofilms prevalent in children under Caccio SM and Chalmers RM. Human 5 years cryptosporidiosis in Europe. Clinical Microbiology and Infection 2016: 22(6) : 471-480. MMWR Weekly / Vol. 67 / No. 25 June 29, 2018 35 36 9 06/07/64 • Infection worldwide including Thailand • Account for 5% of patient diarrhea • Contaminated drinking water, food, direct contact • Clam • Outbreak in the past involving 403,000 persons in Milwaukee, USA, associated with drinking water • No effective, specific chemotherapy • nitazoxanide resolves
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