Introduction to Medical Parasitology and Protozoology and Nonpathogenic

Nimit Morakote, Ph.D. Human 2 21 July 2021

1 วัตถุประสงค์เชิงพฤติกรรม

1. อธิบายศัพท์เฉพาะที่ใช้ในวิชาปรสิตวิทยาได้ 2. อธิบายผลกระทบของโรคปรสิตต่อชุมชนได้ 3. ประยุกต์ความรู้เข้ากับบทเรียนต่อไปได้ 4. แสดงเกณฑ์ที่ใช้แยก ของอะมีบาของคนได้

2 Two animals living together

news.tecnozoom.it www.sevensheaven.nl 3 = Symbiosis Symbiont

= Mutualism Mutual = Commensalism

Host Commensal

=

imdb Host Parasite

4 • “Parasitology” can be defined as a study of the relationship between parasites and hosts • Parasites - animals that live in or on the hosts (for food and shelter). • The principal aim in parasitology is to acquire knowledge about parasitism in natural populations to control or eliminate parasite from unnatural populations, both of the people and of the animals on which the people depend (Beaver, 1981)

5 Parasites

• Facultative parasites – Having both free-living and parasitic phase – Examples: Strongyloides and Naegleria • Obligatory parasites – Only parasitic phase – Example: Beef tapeworm https://www.insider.com/rise-of-brain-eating-amoeba- • Incidental parasites naegleria-fowleri-in-us-waters-2020-7 – Associate with unnatural host -> Mostly pathogenic – Example: Toxocara- man

6 • Pseudoparasites (artefacts) – “Look alike”, e.g., pollen grains • Coprozoic (spurious) parasites – Organisms eaten and found in faeces

– Spurious infection; a follow-up reveals no Credit Adulsak Vijit more organism, e.g., liver fluke eggs, paramphistome eggs

7 8 9 • Opportunistic parasites – Immunocompetent host : no disease or self-limited disease – Immunocompromised host : severe disease, leading to death in many cases – Examples: Toxoplasma, Cryptosporidium

10 Hosts Terminology Parasite stage in host Definitive hosts Adult or sexual stage Intermediate hosts Larva or asexual stage- develop to infective stage Paratenic hosts Larval stage (infective)/ no development Reservoir hosts Adult (source for human infection) Accidental hosts (Dead end of parasite) Larva (mostly) Adult (rare)

11 Parasite Life Cycle

• Simple (homoxenous) – do not require intermediate host – For example, the protozoa histolytica life cycle is completed via fecal-oral transmission • Complex (heteroxenous) – Requires one or more intermediate host – For example, beef tapeworm life cycle requires cattle to complete its life cycle

12 H Fish-eating Human O mammals S T Fish tapeworm Adults In the small intestine Eggs

Fish 2 Coracidium Plerocercoid larva Procercoid larva Plerocercoid larva

Fish 1 Copepod

13 Terminology

• Human is …………………host • Fish-eating mammals are……………….host • Copepod is……………………..host • Fish 1 is…………………..host • Fish 2 is……………………….host • The tapeworm infective stage to man is………………….

14 https://www.thairath.co.th/news/local/northeast/2105852

15 16 Source of Parasite Exposure

• Environment • Contaminated water and soil (waterborne and soil-transmitted) • E.g., hookworm larva in soil • Man • Infected persons or self (direct contact, finger-to-mouth, airborne, insect borne, sexually transmitted) • E.g., Giardia in salad prepared by infected food handler

17 Source of Parasite Exposure

• Animal • Domestic or wild animals as reservoir host

• Foodborne, insect borne The prevalence of Toxoplasma gondii in free-range • E.g., Toxoplasma cysts in chickens from Campos dos Goytacazes, Rio de Janeiro State, Brazil, was examined to evaluate environmental chicken meat (food borne) contamination by oocysts. Brains and hearts of 86 of the 198 chickens were bioassayed in mice for the presence of T. gondii. Viable parasites were isolated from 61 (70.9%) of the 86 chickens.

18 Period

• Prepatent period – Parasite enters to parasite or its products detected – PP of vivax malaria = 11-13 days

• (Clinical) Incubation period – Parasite enters to appearance of signs and symptoms – IP of vivax malaria = 12-17 days

19 Pathogenesis

Can be one or more mechanisms • Destruction of tissues by parasite’s enzymes • Mechanical injury – Large worm- obstruction, e.g., Ascaris – Small worm- tissue lesion or migratory track, e.g., Gnathostoma – Ciliate- penetrating intestinal wall • Immunological reaction to parasites or their metabolites • Others

20 Significance of Parasitic Infection

• Dead e.g., cerebral malaria • Chronic illness – e.g., gnathostomiasis • Deformities – e.g., leishmaniasis • Symptomless – e.g., opisthorchiasis

Thenewsminute.com

21 Economic impact

Reduced Reduced working days income Morbidity Increase Drugs expense

Measurement of economic loss=DALY (disability- adjusted Life- Year)= Healthy years lost due to death or disability Cysticercosis 8.7 million US dollars

22 Important Parasitic Diseases

http://www.stanford.edu/class/humbio153/AntiParasiticDrugs/index.html 23 Factors influencing parasite fauna

average temp. animal fauna • Tropical • Nearctic • Subtropical, • Palearctic • Temperate, • Neotropical • Frigid Parasite • Oriental • Polar or tundra fauna • Australian • Ethiopian flora • Very wet • Humid • Equatorial forest • Subhumid average rainfall • Evergreen forest • Semi-arid • Savannah • Arid

24 Faunistic areas

Nearctic Palearctic

Oriental Ethiopian Neotropical Australian

The Macdonald encyclopedia of mamamals, 1983. 25 Zoonosis

• Zoonosis- โรคสัตว์สู่คน Rock hyrax is a natural reservoir host of leishmania parasite in Israel. – Avian flu, leishmaniasis Commonly found in arid land habitat including deserts, • Anthroponosis- โรคคนสู่สัตว์ savannas and scrub forests. หรือคนสู่คน • Sapronosis- โรคจากสิ่งแวดล้อม Flies carry protozoa in human stool via their legs. • Vector- พาหะ – Mechanical vector Reduviid bugs transmit infective stage of Trypanosoma cruzi via their feces – Biological vector

26 Zoonotic status of man’s parasite

Reptile-2% Environment Unknown Birds-3% 6% 12%

Man 26%

Mammals 51% Ashford RW. Ann Trop Med Parasitol 1991;85:189-98. Emerging and re-emerging disease

• “The leishmaniases as emerging and reemerging zoonoses” (Ashford, 2000) • อุบัติใหม่ (emerging infectious diseases) e.g., microsporidiosis, cyclosporiosis • อุบัติซ ้ำ (re-emerging infectious diseases) เคยพบแล้วหายไป บัดนี้โผล่มาใหม่ e.g., malaria, schistosomiasis • เกิดจำกกำรเปลี่ยนแปลงนิเวศวิทยำ พฤติกรรมประชำกร กำรเดินทำงเสรี- โลกำภิวัตน์

28 Parasite control, elimination and eradication

• Bajoh recorded 75 new cases of malaria in calendar year 2015 (1,010 per 100,000 population). – Propose plans of the malaria control campaign to reduce malaria incidence to no more than 300 per 100,000 population per year. • “Thailand has set the goal of eliminating malaria by 2024, which is defined as the reduction to zero of local, or indigenous, malaria incidence.”

29 Parasite (Simple classification)

• Protozoa – Single cell animal • Metazoa- Multicellular animal • Helminths- Worms (หนอนพยาธิ) • Acanthocephalans- Thorny-headed worms (พยาธิหัวหนาม) • Arthropods- แมงและแมลง

30 Parasite systematic classification

• Animalia – Subkingdom Protozoa – Subkingdom Metazoa • Phylum Platyhelminthes • Class Cestoda (members = cestodes; common name = tapeworms) • Class Trematoda (members = trematodes; common name = flukes) • Phylum Nematoda (members = nematodes; common name = roundworms) • Phylum Acanthocephala • Phylum Arthropoda • Order, Class, Family, Genus, Species

31 Composition of human parasite fauna

Ashford RW. Ann Trop Med Parasitol 1991;85:189-98. Parasite Name

– Scientific name: Binomial nomenclature • Enterbius vermicularis • Genus Enterobius • Species E. vermicularis – Common name • Pinworm, threadworm Pinworm • Absent in microscopic Credit: nejm.org parasites

33 Parasitological diagnosis

• Parasitic infections are diagnosed by demonstration of Blood parasites – parasites, e.g, worms or – their products, e.g., eggs, larva Lung parasites Tissue parasites • Clinical specimens Liver (bile ducts) – Stool parasites – Secretion – Tissues Intestinal parasites

34 PROTOZOA

35 Protozoa

• Single cell animal • Over 30,000 species • Microscopic, 1-60 micrometer in size • Single cell – + nucleus – Other organelles like eukaryotic cell, e.g., endoplasmic reticulum – Specific organelle in some species, e.g., plastid

36 Simple classification

Group Locomotory Special Reproduction organelle organelle Amoeba Pseudopodium(a) - Asexual/ binary fission Flagellate Flagellum(a) - Asexual/ binary fission- longitudinal Ciliate Cilium (a) - Asexual/ binary fission- transverse Sporozoa - Apical complex Asexual and sexual Microspoidia - Polar tube Asexual and sexual (fungus)

37 Simple classification

Amoebae (อะมีบา) Flagellates (สัตว์แส้)

38 38 Ciliates Sporozoa Microsporidia มี sexual reproduction (fungus) มี microspore, polar tube

39 Protozoa encystment

• Trophozoites = stage that feed, grow, reproduce • Cysts = infective stage – Don’t feed, grow or reproduce trophozoite – resistant to environment, waiting for new hosts • Encystment induced by host enzymes or other factors • 1-3 layers of cyst wall precyst • Some species do not have cyst stage, e.g., E. gingivalis

cyst

40 Reproduction

• Asexual (mitosis) – Binary fission (1 to 2) • Amoeba- asymmetric binary fission • Flagellate- longitudinal binary fission • Ciliate- transverse binary fission – Multiple fission (1 to many) in sporozoa – Internal budding (1 → 2 or 1 → many progeny) in sporozoa • Sexual (meiosis)- involve combination of female and male sex cells to diploid zygote followed by meiosis

41 In diagnostic laboratory

Protozoa infections

Clinical specimens Wet smear or Under the • Blood smears Stained smear microscope • Stool smears • Other specimens Morphological identification Gross specimens Helminth infections Stained specimens

Researchgate.net 42 Nonpathogenic amoeba of man

43 Pathogenic amoeba Nonpathogenic amoeba Large intestine `Large intestine • • Entamoeba coli, etc. Brain • Naegleria fowleri • Acanthamoeba spp.

Stool examination requires differentiation between pathogenic and nonpathogenic amoeba

44 Nonpathogenic amoeba

Name Habitat Stage in life cycle Infective stage Entamoeba dispar Large intestine Trophozoite & Cyst Cysts- 4 nuclei (In stool) Can not differentiate from E.histolytica by microscopy Entamoeba hartmanni Cysts- 4 nuclei <12 m (smaller than E. histolytica) Cysts-8 nuclei Entamoeba coli >12 m (15-30 m)

Endolimax nana Cysts- 4 nuclei (6-9 m) Iodamoeba butschlii Cysts- 1 nucleus (5-20 m) Entamoeba gingivalis Mouth Trophozoite only Trophozoite (In specimens from mouth) 45 • Life cycle – Fecal- oral through food or water contaminated with feces, cyst stage (Except E. gingivalis which is directly transmitted from mouth-to- mouth) • Differential diagnosis – Required for amoeba detected in stool • Differential morphology in stool smear (except E. dispar) • Molecular technique (not available in the department)

46 Staining of amoeba

Wet smear Specimens

Iodine stain

Iron hematoxylin stain (Monochrome: black or blue) Trichrome stain

(red, blue, green) 47 Criteria for microscopic diagnosis of amoeba species 1. Nucleus – Morphology • karyosome, • peripheral chromatin – Number of nuclei in cyst stage 2. Cytoplasmic content, e.g., – chromatoid bodies – glycogen vacuole

48 Nuclear morphology of intestinal amoeba

1. Entamoeba histolytica/ E. dispar/ E. hartmanni 2. Entamoeba coli 3. Endolimax nana 4. Iodamoeba butschlii

49 Entamoeba coli

1= E. histolytica 2= E. coli

Trophozoite- 15-50 µm Cyst 15-30 µm nucleus with eccentric 8 nuclei; karyosome Chromatoid bodies- splinter-like ends Slideshare.net https://www.msu.edu/course/zol/316/ameba.htm

50 IH stain

Trichrome stain

http://www.medical-labs.net/entamoeba-coli- Medical-labs.net parasite-2543/

51 Endolimax nana Smallest amoeba of man

Nucleus with large irregular Trophozoite 8-10 µm Infective cyst 6-9 µm shaped 4 small nuclei karyosome, no peripheral nuclear chromatin 52 IH stain

Trichrome stain

http://www.medical-labs.net/endolimax- E. nana trophozoites (From: Medical-labs.net) nana-parasite-trophozoite-and-cyst-2576/

53 IH stain

Trichrome stain

E. nana cysts (From: Medical-labs.net) http://www.medical-labs.net/endolimax-nana- parasite-trophozoite-and-cyst-2576/

54 Iodamoeba butschlii

Nucleus with large irregular shape Trophozoite- 8-20µm Infective cyst- 5-20µm karyosome; 1 Nucleus; 1 Nucleus; Chromatin granules Cytoplasm with large Cytoplasm with large radiating from the glycogen vacuole glycogen vacuole karyosome

55 IH stain

Trichrome stain

www.keyword-suggestions.com http://www.medical-labs.net/iodamoeba- butschlii-3091/

56 E. gingivalis • From smear of gingival pocket scraping • Trichrome stain • 1 nucleus, leukocyte and tissue debris in cytoplasm

57 Parasite image

https://w1.med.cmu.ac.th/parasite/หน้าแรก-image/

58 Book online

http://www.microbiologybook.org/book/parasit-sta.htm

59 Books in the library

• Schmidt & Roberts’ Foundations of Parasitology. 9th edition, 2013. • Bogitsh, Carter and Oeltmann. Human Parasitology. Fourth edition, 2013. • Zeibig EA. Clinical Parasitology. A practical approach. Second edition, 2013. • Ridley JW. Parasitology for Medical and Clinical Laboratory Professionals, 2012. • Gunn A and Pitt SJ. Parasitology. An integrated approach, 2012. • Elsheikha HM, Jarrol EL, editors. lIllustrated dictionary of parasitology in the post-genomic Era. 2017. • Loker ES, Hofkin BV. Parasitology : a conceptual approach, 2015. • Nopparatana C, Nopparatana C. Medical parasitology and entomology. 2015. • Lecture notes on medical parasitology. Leelayoova et al, editors. 2015.

60 Some notes

• Literatures are evolving. • Parasitological terms can sometimes deviate from original terms. • Critical appraisal is essential when social media discuss human cases- true parasites or pseudoparasites.

61