1. AMERICAN TRYPANOSOMIASIS Prof. of Medical Parasitology
(CHAGAS DISEASE)
Dr. Hala El Deeb
Prof. of Medical Parasitology

2. Lecture Topics (ILOs)
Lecture: American Trypanosomiasis (Chagas Disease)
1. List main Trypanosoma spp., habitat and associated disease
2. Discuss epidemiology (G.D, vector Triatoma, transmission & R.H)
3. Compare similarities and differences between life cycle of trypanosomes in both vertebrate & invertebrate hosts (morphologic forms, infective & diagnostic stages)
4. Explain pathogenesis & clinical picture
5. Outline management (diagnosis & treatment)
6. Discuss differential diagnosis
7. Design prevention and control plan

3. Why WHO consider American Trypanosomiasis a Major Parasitic Disease?* *

4. Family Trypanosomatidae (Hemoflagellates = Kinetoplastids)
Overview
Hemoflagellates are pathogenic flagellated protozoa found in the blood
Two genera are of medical importance, Trypanosoma & Leishmania
Life cycle includes both a vertebrate and an invertebrate hosts (= heteroxenous L.C)
African trypanosomes (T. brucei complex)
African trypanosomiasis (Sleeping sickness)
Trypanosoma cruzi
American trypanosomiasis (Chagas’ disease)
(named after the Brazilian physician Carlos Chagas, who discovered the disease in 1909)
Leishmania species
Visceral Leishmaniasis

5. American Trypanosomiasis (Chagas Disease)
- Chagas disease is a zoonosis caused by Trypanosoma cruzi
- It is a vector-borne disease (VBD). Life cycle includes both a vertebrate and an invertebrate hosts (= heteroxenous L.C) *
Parasite - Trypanosoma cruzi
Vector
Reduviid or triatomine bugs (kissing bugs) * ** **

6. Geographical Distribution
Epidemiology
Geographical Distribution *
Latin America (21 countries in: Mexico, Central America, and South America), affecting an estimated million people
Is considered a Major Parasitic Disease by WHO
Mainly, in rural areas (poverty)
In the past few decades it has been increasingly detected in USA, Canada, many European and some Western Pacific countries. This is due mainly to population mobility
Urban transmission associated with blood transfusions
Leading cause of cardiac disease
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7. Epidemiology
Chagas disease is a zoonosis. Originally (>9000 years ago), T. cruzi only affected wild animals. It later spread to domestic animals and people. The large reservoir of T. cruzi parasites in wild animals of the Americas means that the parasite cannot be eradicated. Instead, the control targets are elimination of the transmission and health care access for the infected and ill population

8. Reservoir Host
The primary wildlife reservoirs for Trypanosoma cruzi include opossums, raccoons, armadillos, squirrels, woodrats, and mice
Armadillos
Opossum
Raccoon
Squirrel

9. Vector * Reduviid or triatomine bugs (Triatoma or kissing bugs)
Large (2-3 cm)
Incomplete metamorphosis (Egg, Nymph, Adult. Cycle: months)
Normally they hide during the day and become active at night. Both sexes & nymph feed on human blood. They usually bite an exposed area of skin such as the face, and the bug defecates close to the bite
Releases metacyclic trypomastigotes in its feces near the site of the bite & parasites enter host through the wound
Have earned their name because their preferred site to take a blood meal is on face, often near the mouth
Bite while we are asleep, and take up to 25 min for their meal
Reduviid bites usually presents as papular urticaria ** * **
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10. Kissing bug habitat Undeveloped Housing !!
Note: Triatomine bugs typically live in the cracks of poorly-constructed homes in rural or suburban areas = rural poverty!!

11. Transmission
Contamination of bite wound by triatomine bug feces containing metacyclic trypomastigotes (I.S); after they bite and ingest blood, they defecate on the person. When the human or animal host scratches the bite area, penetration of the infected feces is facilitated (VBD)
Blood transfusions
Mother-to-baby (congenital)
Organ, tissue or cell transplanted from an infected donor
Laboratory accident (rare)
Contaminated food or drink (rare)

12. The parasite
Three morphological forms:

13. The parasite *
Trypomastigote Slender shaped (20µ) – C or U-shaped, one form (monomorphic ), central nucleus, free flagellum 1/3 body, large peripheral kinetoplast - Extracellular in the blood and tissue fluids
Amastigote 2-6 µ, round or oval devoid of external flagellum
- Intracellular, mainly in cardiac & skeletal muscles, brain meninges, nerve ganglia & cells of GIT & RE system **
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Epimastigote Spindle shape– kinetoplast anterior to central nucleus, undulating membrane is short, terminal free flagellum
- Vector (hindgut) & Culture
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14. Life Cycle
Includes both a vertebrate and an invertebrate hosts (=heteroxenous life cycle)
Parasite found intracellularly & in the blood and tissue fluids of vertebrate hosts

15. Life Cycle * Vertebrate host
Reduviid bug takes a blood meal and releases metacyclic trypomastigotes in its feces.  Trypomastigotes enter the host through the wound or through intact mucosal membranes, such as the conjunctiva
Trypomastigotes invade host cells &differentiate into intracellular amastigotes *
Amastigotes undergo asexual reproduction and differentiate into trypomastigotes & are released into the circulation as trypomastigotes
Trypomastigotes infect cells (prefer cells of mesenchymal origin, e.g., heart & GI) and transform into intracellular amastigotes
Bloodstream trypomastigotes DO NOT replicate (different from the African trypanosomes) *

16. * Life Cycle Invertebrate host
The “kissing” bug becomes infected by feeding on human or animal blood that contains circulating trypomasigotes
The ingested trypomastigotes transform into epimastigotes in the vector’s midgut
The parasites multiply and differentiate in the midgut and finally differentiate into infective metacyclic trypomastigotes in the hindgut (=posterior station development) *
Metacyclic trypomastigotes leave insects with feces & infect new hosts through the skin or mucous membranes (Stercoraria), i.e., mode of infection is contaminative in type
Transmission of monomorphic trypanosomes in reduviid bug is cyclopropagative *

17. Pathogenesis & Clinical Features
Chagas disease has an acute and a chronic phase. If untreated, it is lifelong
Acute Stage
Acute Chagas disease occurs immediately after infection, may last up to a few wks or months (1-4 months), and parasites may be found in the circulating blood. Children & infants are mostly affected
Infection may be mild or asymptomatic. There may be fever, headache, lymphadenopathy, mild hepatosplenomegaly. Children most likely to be symptomatic
In less than 50% , characteristic first visible signs can be a skin lesion or a purplish swelling of the lids of one eye (= chagoma & Romana’s sign), where the parasite entered into the skin or mucous membrane
Most acute cases resolve over a few wks or months into an asymptomatic chronic form
Rarely, acute infection may result in severe myocarditis or meningoencephalitis * ** **

18. Pathogenesis & Clinical Features
Acute Stage
Chagoma
Upon introduction into the human, the parasites invade cells of the subcutaneous tissue at the site of the infection, causing an acute inflammatory reaction or swelling “erythematous indurated area” often called a chagoma

19. Pathogenesis & Clinical Features
Acute Stage
Romana’s sign
When parasites enter through the conjunctiva of the eye, they cause a unilateral edema of the eyelid and conjunctivitis, a syndrome known as Romana’s sign (pathgnomonic)
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Romana’s sign
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20. Pathogenesis & Clinical Features
Chronic Stage
A. Chronic indeterminate “asymptomatic” stage
Following the acute phase, most infected people (60–80%) enter into a prolonged asymptomatic form of disease called "chronic indeterminate“(10-30 yrs of latency) and never develop Chagas-related symptoms, i.e., completely free of cardiac, GI and neurological symptoms
Infected people of chronic indeterminate stage are seropositive with no detectable parasitemia; as parasites are hidden (mainly in the heart and digestive muscle)

21. Pathogenesis & Clinical Features
B. Chronic determinate stage
An estimated 20–40% of infected people will develop debilitating and sometimes life-threatening medical complications over the course of their lives. The complications of chronic Chagas disease may include cardiac, GI and neurologic disorders, and can be fatal
The most serious manifestations include:
Dilated cardiomyopathy: Occur in 10-30% of cases; with arrhythmias, conduction defects & congestive heart failure
Damage of the GI system and disturbances of peristalsis lead to dilatation with megaesophagus and megacolon (mega-organ syndrome), and weight loss.  Difficulty in swallowing, regurgitation & aspiration may cause pneumonia and death. Chronic constipation, fecal compacting causes perforation of the colon

22. Pathogenesis & Clinical Features
The clinical manifestations of Chagas disease are due to cell death in the target tissues that occurs during the infective cycle, by sequentially inducing an inflammatory response, cellular lesions, and fibrosis
For example, intracellular amastigotes destroy the intramural neurons of the autonomic nervous system in the intestine and heart, leading to mega-intestine and heart aneurysms, respectively *

23. Cardiomegaly Megacolon Megaoesophagus * ** *** ***** **
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Megacolon

24. Differential Diagnosis
Acute Chagas'disease
Brucellosis, endocarditis, salmonellosis, schistosomiasis, toxoplasmosis (toxoplasmic encephalitis & congenital toxoplasmosis), visceral leishmaniasis, malaria, tuberculosis, connective tissue diseases and leukemia
Chronic Chagas'disease with cardiomyopathy Endocarditis & ischemic heart disease

25. Diagnosis A. Clinical Diagnosis
History whether patient has lived in or visited an endemic area
Clinical signs and symptoms
B. Laboratory Diagnosis

26. Laboratory Diagnosis MICROSCOPY ON*
DIRECT METHODS
MICROSCOPY ON
Giemsa stained thin and thick blood smears:
Direct blood film or concentration techniques (e.g., microhaematocrit centrifugation technique or buffy coat technique) in acute stage (examined for trypomastigotes). Trypomastigotes may be easily detected in blood during acute stages, however, in chronic disease, this stage are rare or absent
Histopathological examination of biopsy from tissues, e.g., L.Ns & skeletal muscles, liver or spleen (Amasigotes) ** * **
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27. Laboratory Diagnosis DIRECT METHODS B. CULTURE
Inoculation in NNN culture (examined for Epimastigotes)
C. ANIMAL INOCULATION
Inoculation into mice

28. D. Xenodiagnosis
Highly efficient – demonstrate low level of parasite in blood
Method:
A Laboratory bred winged bug is starved for 2 wks then fed on suspected patient’s blood for 3 consecutive days–
30 days later, faeces & gut examined for trypanosomes

29. Laboratory Diagnosis
II. INDIRECT METHODS
Serological tests
Complement fixation "GUERREIRO-MACHADO TEST", IFA, IHA & ELISA. Diagnosis is generally made by testing with at least two different serologic tests
Immunoassays
ELISA is highly sensitive and specific to detect antigens in urine and sera in patients with congenital infections and those with chronic disease and for blood screening

30. III. POLYMERASE CHAIN REACTION (PCR) IV. IMAGING Chest x-ray
Laboratory Diagnosis
III. POLYMERASE CHAIN REACTION (PCR)
IV. IMAGING
Chest x-ray
Echocardiogram
Electrocardiogram (ECG)

31. Trypomastigote
1 Form
Nucleus
C-shaped
Subterminal, large kinetoplast

32. T. cruzi trypomastigotes in thin blood smears stained with Giemsa
Note the typical C-shape of the trypomastigote

33. Polymorphic trypanosomes Monomorphic trypanosomes
T. cruzi trypomastigotes in thin blood smears should be differentiated from polymorphic trypanosomes
Polymorphic trypanosomes
Monomorphic trypanosomes

34. Trypanosoma cruzi pseudocyst in cardiac muscle
Amastigotes

35. Treatment
Treatment for Chagas disease is recommended for all people diagnosed with an acute infection, congenital infection, and for those with suppressed immune systems, and for all children with chronic infection
No medications are given to patients with the chronic phase
Chronic-phase patients are usually treated using treatments directed at the specific symptoms or organ damage
The two drugs used for treatment of Chagas disease are Benznidazole and Nifurtimox
Almost 100% effective if given soon after infection
These drugs are fairly toxic, expensive. They should not be taken by pregnant women or by people with kidney or liver failure

36. Prevention & Control Endemic areas
A. Vector control is the most useful method to prevent Chagas disease in Latin America
House improvements to prevent vector infestation (building structures that discourage the bug’s habitation, e.g., homes with palm roofs and cracks)
Spraying with residual insecticides
Mechanical elimination of the vector
B. Personal preventive measures such as bed nets
C. Health education regarding possible transmission routes
D. Avoid pets in the home environment to limit attraction

37. Prevention & Control E. Screening of blood donors*
E. Screening of blood donors
F. Testing of organ, tissue or cell donors
G. Mother-to-baby screening of newborns and other children of infected mothers to provide early diagnosis and treatment ** * **
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38. Prevention &Control
Non-endemic areas
These are other regions where Chagas disease is found but is not endemic (e.g., US)
Screening of blood donors
Testing of organ, tissue or cell donors
Mother-to-baby screening of newborns and other children of infected mothers to provide early diagnosis and treatment

39. Prevention & Control Problems
There is no vaccine for Chagas disease !!
The large reservoir of T. cruzi parasites in wild animals!!
Vector control is the most effective method of prevention
in Latin America

42. Which form? Promastigote Answer: anterior Flagellum blepharoblast
Parabasal body
Nucleus
posterior

43. Which form? Epimastigote Answer: anterior Undulating membrane
posterior

44. Which form? Trypomastigote Answer: anterior Undulating membrane
posterior

45. Which form? Amastigote Answer: anterior Flagellum blepharoblast
Parabasal body
Nucleus
posterior

46. Answer: A Protozoa causing Romans sign is: A- Trypanosoma cruzi
Choose the Correct Answer
Protozoa causing Romans sign is:
A- Trypanosoma cruzi
B- T. brucei gambiense
C- T. brucei rhodesiense
D- Leishmania donovani
E- Leishmania infantum
Answer: A

47. The parasite changes in shape (morphology) and increases in number
Interpret the following statement
Transmission of Trypanosoma cruzi in Triatoma megista is cyclopropagative
Answer:
The parasite changes in shape (morphology) and increases in number

48. Xenodaignosis is a method used for diagnosis of…………………
Complete
Xenodaignosis is a method used for diagnosis of…………………
Answer: Chagas’ disease

49. Answer: C Cardiac muscle is especially affected in:
Choose the Correct Answer
Cardiac muscle is especially affected in:
A- Sleeping sickness
B- Kala azar
C- Chagas’ disease
D- Espundia
E- All of the above
Answer: C

50. Choose the Correct Answer
The metacyclic forms of Trypanosoma cruzi are found in:
A- Reticuloendothelial cells
B- Cardiac muscle
C- CSF
D- Hind gut of infected insect
E- Salivary gland of infected insect
Answer: D

51. Answer: C A- Digestive and respiratory tracts B- Heart and liver
Choose the Correct Answer
In chronic Chagas’ disease, the main lesions are in:
A- Digestive and respiratory tracts
B- Heart and liver
C- Heart and digestive tract
D- Liver and spleen
E- CNS
Answer: C

52. Answer: C A- Epimastigote form B- Promastigote form C- Amastigote form
Choose the Correct Answer
The diagnostic stage that may be found in lymph node biopsy in Chagas’ disease is:
A- Epimastigote form
B- Promastigote form
C- Amastigote form
D- Trypomasigote form
E- Metacyclic trypomastigote
Answer: C

53. Interpret
Although African & American trypanosomes belong to the same genus, they have different types of transmission and infection
Answer:
African trypanosomes are transmitted to man by the bite of infected Glossina (anterior station development)
American trypanosomes are transmitted to man through contamination of the bite wound with the feces of the infected Triatoma megista (posterior station development)

54. Choose the Correct Answer
In American trypanosomiasis, the infective stage is found in:
A- Saliva of Triatoma
B- Saliva of Glossina
C- Stool of Triatoma
D- Stool of Glossina
E- Saliva of Simulium
Answer: C

55. Answer: C Metacyclic trypomastigotes are the:
Choose the Correct Answer
Metacyclic trypomastigotes are the:
A. Infective stage of Leishmania spp..
B. Diagnostic stage of African trypanosomes
C. Infective stage of African trypanosomes
D. Diagnostic stage of Leishmania spp..
E. Diagnostic stage of American trypanosomes
Answer: C

56. Choose the Correct Answer
The intracellular form of Trypansoma cruzi in cardiac tissue is the:
A. Epimastigote.
B. Amastigote.
C. Promastigote.
D. Trypamastigote.
E. Metacyclic trypomastigote.
Answer: B

57. References
- Study guide “Blood module” by staff members of Parasitology Dept., Faculty of Medicine, Ain Shams University
- Diagnostic Medical Parasitology, (Garcia LS), 6thed, 2016
Foundations of Parasitology, (Roberts), 9th ed, 2013
- Human Parasitology, (Bogitsh), 4th ed, 2013

58. Thank you for your attention!