Blood and Tissue

flagellates:
Trypanosomiasis
BY
DR B.O. OLOPADE
DEPARTMENT OF MEDICAL MICROBIOLOGY/PARASITOLOGY, CHS, OAU, ILE-
IFE
OUTLINE

 INTRODUCTION
 OBJECTIVES
 CONCLUSION
 REFERENCES
INTRODUCTION

 Vector‐borne disease caused by parasitic, flagellated

protozoans.
 Two main groups: African and South American
trypanosomiasis (Chagas’disease)
 Significant cause of morbidity and mortality in endemic
areas.
OBJECTIVES

 At the end of this lecture, you should be able to:

 List the agents of African and American
Trypanosomiasis.
 Classify and describe the morphology, life cycle,
pathogenesis and clinical course of infection with the
trypanosomes
 Outline the management modalities of Trypanosomiasis.
INTRODUCTION
(Classification)
 Phylum : Sarcomastigophora
 Subphylum : Mastigophora
 Class : Kinetoplastidea
 Order : Trypanosomatida
 Family : Trypanosomatidae
INTRODUCTION(General
characteristics)
 They live in the blood and tissues of man and other
vertebrate hosts and in the gut of the insect vectors.
 Members of this family have a single nucleus, a
kinetoplast and a single flagellum.
 Nucleus is round or oval and is situated in the central
part of the body.
INTRODUCTION (General
characteristics)
 They are motile by the aid of flagellum
 Trypanosomes pass their life cycle in two hosts
vertebrate hosts (definitive hosts) and insect vectors
(intermediate hosts).
 In the vector, the trypanosomes follow one or two
modes of development and are accordingly classified
into 2 groups- Salivaria and Stercoraria.
INTRODUCTION(General
characteristics)
 Salivaria (anterior station): In the salivaria group,the
trypanosomes migrate to mouth parts of the vectors, so
that infection is transmitted by their bite (inoculative
transmission) e.g Trypanosoma brucei gambiense and
Trypanosoma brucei rhodesiense causing African
trypanosomiasis which are transmitted by the bite of
tsetse flies.
INTRODUCTION(General
characteristics)
 Stercoraria (posterior station): In the stercoraria
group, the trypanosomes migrate to the hindgut and are
passed in faeces (stercorian transmission), e.g.
Trypanosoma cruzi causing Chagas’ disease, which is
acquired by rubbing the faeces of the vector bug into
the wound caused by its bite.
African Trypanosomiasis

 Trypanosomes were first isolated from the blood of a

steamboat captain on the Gambia river in 1901 (hence,
the name gambiense) by Forde.
 Dulton, in 1902, proposed the name Trypanosoma
gambiense.
 It is endemic in scattered foci in West and Central
Africa between 15°N and 18°S latitudes.
AFRICAN TRYPANOSOMIASIS
(Causative agent)
 Three trypanosome subspecies, Trypanosoma brucei
brucei, Trypanosoma brucei rhodesiense, and
Trypanosoma brucei gambiense are the causative
agents. They are indistinguishable morphologically, and
as a group they are often referred to as the
Trypanosoma brucei complex. Trypanosoma brucei
brucei is a parasite of wild and domestic animals that is
not infectious to humans. Trypanosoma brucei brucei,
causes the disease ‘nagana’ in African cattle.
Trypanosoma rangeli is non-pathogenic.
AFRICAN TRYPANOSOMIASIS

 This disease is restricted to Sub-Saharan Africa the

range of Glossina, the tsetse fly vector.
 Two geographically and clinically distinct forms:
Trypanosoma brucei gambiense (West African sleeping
sickness/Gambian trypanosomiasis).
 Trypanosoma brucei rhodesiense (East African sleeping
sickness/Rhodesian trypanosomiasis)
Geographical distribution of
African Trypanosomiasis
Tsetse fly (Insect vector for
Trypanosoma brucei complex)
African Trypanosomiasis

 The members of the Trypanosoma brucei complex are

transmitted by various species of tsetse flies that
belong to the genus Glossina. The parasites undergo a
developmental cycle in the insect vectors. Tsetse flies
of both sexes become infected with trypanosomes when
they ingest blood from infected mammalian hosts that
contains trypomastigotes, the form of the parasite that
circulates in the bloodstream.
Epidemiology of African
Trypanosomiasis
 Sleeping sickness was a much greater problem in the
past than it is at present. Major epidemics have
occurred in the Sudan, Democratic Republic of Congo,
Uganda, Angola and several other endemic countries. In
some areas wars and the resulting lack of control
programs are the important factors underlying the
outbreaks.
West African Trypanosomiasis
 West African trypanosomiasis is caused by Trypanosoma brucei
gambiense, which is transmitted primarily by tsetse flies belonging to the
palpalis group: Glossina palpalis, Glossina tachinoides, and Glossina
fuscipes. These vectors inhabit forests and wooded areas along rivers.
West African Trypanosomiasis

 West African trypanosomiasis is primarily a problem in

rural populations. The course of the illness caused by T.
b. gambiense is less severe than that caused by T. b.
rhodesiense, although both forms eventually lead to
death if not treated.
Trypanosoma brucei
gambiense (Morphology)
 In the blood of the vertebrate host, T. brucei
gambiense exists as trypomastigote form, which is
highly pleomorphic.
 It occurs as a long slender form, a stumpy short broad
form and an intermediate form.
 The trypomastigotes are about 15–40 μm long and 1.5–
3.5 μm broad.
Trypomastigote forms
T.b.gambiese (Morphology)

 In insects, it occurs in 2 forms:

 Epimastigotes
 Metacyclic trypomastigote forms (infective stage)
Life cycle of T.b.gambiense

 T. brucei gambiense passes its life cycle in 2 hosts.

 Vertebrate host: Man, game animals, and other
domestic animals.
 Invertebrate host: Tsetse fly.
Life cycle

 Infective form: Metacyclic trypomastigote forms are

infective to humans.
 Mode of transmission:
By bite of tsetse fly
Congenital transmission has also been documented.
Life Cycle
Pathogenesis

 Dissemination of the parasites from localized chancre

(trypanosomal chancre) to the lymphatics and blood and
finally the CNS. There is an intense mononuclear cell
reaction to the parasites as well as oedema and local
tissue destruction.
 Immune evasion facilitated by antigenic variation using
variant surface glycoproteins (VSG’s)
Pathogenesis

 Hemolymphatic illness as the parasites disseminate

widely through the lymphatics and the bloodstream.
Systemic African trypanosomiasis without central
nervous system (CNS) involvement is generally referred
to as stage I disease. The parasites first travel from the
site of inoculation to regional lymph nodes where they
proliferate and bring about an inflammatory response.
Pathogenesis

 In stage I trypanosomiasis, there is widespread

lymphadenopathy and histiocytic proliferation, which
may be followed by fibrosis.
 Morular cells (Mott cells) are also often present in
tissue. The heart is frequently involved in this stage of
the disease especially with T. b. rhodesiense infections.
A pancarditis may develop involving all layers of the
heart.
Pathogenesis

 Stage II African trypanosomiasis involves invasion of the

CNS. Parasites reach the brain and meninges via the
bloodstream and cause meningoencephalitis . In the
brain, they are found mainly in the frontal lobes, the
pons and the medulla, but other areas may be
parasitized as well. Edema and hemorrhages may be
evident on gross examination of affected areas at
autopsy.
Clinical features (Gambiesan
Trypanosomiasis)
The illness is chronic and can last for many years.
Incubation period is usually 2-3 weeks.
 Three clinical stages:
Trypanosomal chancre
Hemolymphatic stage
Meningoencephalitic stage.
Clinical features

 The chancre - This lesion usually appears 1 to 2 weeks

after the bite of the infected fly and resolves
spontaneously over several weeks. The chancre may
ulcerate and reach a diameter of several centimeters;
regional lymphadenopathy may also develop. It is
painful and indurated.
Clinical features

 Stage I disease
Intermittent fever, chills, myalgia, fatigue, general
malaise, rash, pallor, weight loss, and headache.
There is also transient oedema of the face,
hepatosplenomegaly and lymphadenopathy, particularly in
the posterior cervical region (Winterbottom’s sign)- 85%
of hospitalized patients.
Clinical features

 Stage II disease involves invasion of central nervous

system.
This is marked by increasing headache, mental dullness,
apathy and day time sleepiness, insomnia and restlessness
in the night, abnormal movements, tremors and
behavioural changes. The patient falls into profound coma
followed by death.
Clinical features

 The meningoencephalitic stage is seen in 3 to 6 weeks

in T. b. rhodesiense and up to two years in T.b.
gambiense.
Stage II disease
East African Trypanosomiasis

 It is found in Eastern and Central Africa

(Uganda,Tanzania, Zambia, and Mozambique)
 Stephans and Fanthan discovered T. brucei rhodsiense
in 1910 from the blood of a patient in Rhodesia
suffering from sleeping sickness.
East African Trypanosomiasis

 The etiologic agent of East African trypanosomiasis is T.

brucei. rhodesiense.
 It is transmitted by tsetse flies of the morsitans group,
principally Glossina morsitans, Glossina pallidipes and
Glossina swynnertoni.
 These vectors are widely distributed in savanna and
woodland areas of Central and East Africa. Wild animals
are the reservoir of this organism, principally antelopes.
East African Trypanosomiasis

 Although the disease is usually transmitted by the

vector from man to man, the disease is actually a
zoonosis, with the reservoir being wild game animals
like bushbuck, antelope and domestic animals like
cattle.
 Its morphology, habitat and life cycle is similar to T.
brucei gambiense.
East African Trypanosomiasis

 East African trypanosomiasis is more acute than the

Gambian form .
 It may end fatally within a year of onset before central
nervous system involvement develops.
Pathogenesis/clinical features of
East African Trypanosomiasis

 Pathological features are similar in both diseases with

some variations
 € Oedema, myocarditis, and weakness are more
prominent in East African sickness.
 € Lymphadenitis is less prominent.
 € Febrile paroxysms are more frequent and severe.
 € There is a larger quantity of parasite in the peripheral
blood.
 € Central nervous system involvement occurs early.
Management of African
Trypanosomiasis
 History
 Physical examination
 Investigations
 Treatment
Diagnosis

 Definitive diagnosis of sleeping sickness is established

by the demonstration of trypanosomes in peripheral
blood, bone marrow, lymph node, CSF and chancre
fluid.
 Blood films- (Wet, thin and thick). Wet blood films are
for detection of motile trypanosomes whereas thin and
thick blood films are fixed in methanol and stained in
Giemsa.
Blood smear showing
trypanosomes
Diagnosis

 Concentration methods increase the chances for the

detection of the parasite in cases of low parasitaemia.
 Serodiagnosis- Antibody and antigen detection. Almost
all patients with African tryponosomiasis have very high
levels of total serum IgM antibodies and later, CSF IgM
antibodies.
Diagnosis

 Various serological methods have been developed to

detect these antibodies and are as follows:
Indirect hemagglutination (IHA)
Indirect immunofluroscence (IIF)
Enzyme linked immunosorbent assay (ELISA)
Card agglutination trypanosomiasis test (CATT)
Complement fixation test (CFT)
Diagnosis

 Antigens from serum and CSF can be detected by ELISA.

 CSF examination- Increased cerebrospinal fluid (CSF)

pressure and raised cell count and proteins in CSF. The
criteria for second stage infection are: either
trypanosomes in the CSF and/or a raised leukocyte count
of > 5 cells/mm³

 Other investigations- Full blood count (FBC) will reveal

anaemia and monocytosis, Erythrocyte sedimentation rate
(ESR) will be increased.
Diagnosis

 Animal Inoculation:
Inoculation of specimens from suspected cases to white rat
or white mice is a highly sensitive procedure for detection
T.brucei rhodesiense infection.

 Radiological investigations: Late stages of CNS

involvement.
 CT scan- cerebral oedema
 MRI- white matter enhancement
Treatment
CAUSATIVE AGENT STAGE I STAGE II
T.brucei.gambiens 1) Pentamidine Eflornithine
e isoethionate(Lomid (difluoromethylorn
ine) at a dosage of ithine [DFMO],
4 mg/kg, up to Ornidyl) For adults
300mg/day, the recommended
intravenously (IV) dosage is 400
or intramuscularly mg/kg per day IV
(IM) for 10 days for in four divided
both adults and doses for 14 days,
children. and for children it
is 500 to 600mg/kg
per
2) Suramin day

2) Melarsoprol
3) Nifurtimoxl
Treatment

 Suramin ( Naphuride, Antrypol) is the first-choice drug

for stage I rhodesiense trypanosomiasis. A 100-200mg
test dose is recommended. The treatment regimen for
both adults and children, beginning 24 hours after the
test dose, is 5mg/kg on day 1, 10mg/kg on day 3, and
20mg/kg on days 5, 11, 17, 23, and 30. The drug is
administered by slow intravenous infusion and the most
important side effect of suramin is renal damage.
Treatment

 Melarsoprol:
 In adults the drug is given in three courses of 3 days
each. The recommended dosage is 2-3.6mg/kg per day
intravenously in three divided doses for 3 days, followed
1 week later by 3.6mg/kg per day, also in three divided
doses for 3 days. This latter course is then repeated 10
to 21 days later. Side effect is reactive encephalopathy
and the risk is reduced if prednisolone is administered
with the drug.
Treatment

 Fexinidaole is an oral treatment for Gambiense human

African Trypanosomiasis. It was included in 2019 in the
WHO Essential medicines list and WHO human African
Trypanosomiasis treatment guidelines. It is indicated as
first line for first stage and non-severe second stage.
Prevention of African
Trypanosomiasis.
 Control is based on early diagnosis and treatment of
cases to reduce the reservoir of infection.
 Control of tsetse fly population (most important
preventive measure) by wide spraying of insecticides,
traps, and baits impregnated with insecticides.
South American trypanosomiasis
(Chagas’disease)

 It is caused by Trypanosoma cruzi transmitted by the

bite of the triatomine bugs. When the bug takes a blood
meal, metacyclic trypomastigotes are released with the
faeces and can establish infection in the mammalian
host either by crossing mucous membranes, such as the
conjunctiva, nasal or oral mucosa, or through abraded
skin such as the wound made by the bite of the bug.
South American trypanosomiasis
(Chagas’disease)

 Transmission also vertical and through blood products.

It is a zoonotic disease and is limited to South and
Central America.
American trypanosomiasis
(Chagas’disease)
 Vector-borne
 Reduviid insects,
Kissing bugs, Triatomine bugs

 Multiple species
 Triatoma
 Rhodnius
 Panstrongylus
Triatomine bug
Morphology of T.cruzi

 In humans, T. cruzi exists in both amastigote and

trypomastigote forms.
 € Amastigotes are the intracellular parasites. They are
found in muscular tissue, nervous tissue, and
reticuloendothelial system.
 € Trypomastigotes are found in the peripheral blood.
Morphology of T.cruzi

 Amastigote
 Amastigotes are oval bodies measuring 2–4 μm in
diameter having a nucleus and kinetoplast. Flagellum is
absent.
 Morphologically, it resembles the amastigote of
Leishmania spp., hence it is frequently referred to as
the leishmanial form.
 Multiplication of the parasite occurs in this stage.
 This form is found in muscles, nerve cells, and
reticuloenodothelial systems.
Morphology of T.cruzi

 Trypomastigote
 Trypomastigotes are non multiplying forms found in the
peripheral blood of man and other mammalian hosts. In
the blood, they appear either as long, thin flagellates
about (20 μm long) or short stumpy form (15 μm long).
In stained blood smears, they are shaped like
alphabet‘C’, ‘U’, or ‘S’, having a free flagellum of
about one third the length of the body.
Trypomastigotes of T.cruzi
Morphology of T.cruzi

 Epimastigote
 Epimastigote forms are found in the insect vector. It
has a kinetoplast adjacent to the nucleus. An undulating
membrane runs along the anterior half of the parasite.
 Epimastigotes divide by binary fission in the hindgut of
the vector.
Epidemiology

 Human T. cruzi infection is confined to the Americas. It

extends from approximately South in Argentina into the
southern states of the USA.
Life cycle of T,cruzi

 T. cruzi passes its life cycle in 2 hosts

 Definitive host: Man
 Intermediate host (vector): Reduviid bug or triatomine
bugs.
 Reservoir host: Armadillo, cat, dog, and pigs.
 Infective form: Metacyclic trypomastigotes are the
infective forms found in faeces of reduviid bugs.
Life cycle of Trypanosoma
cruzi
Life cycle

 The vectors important in human infection are the

reduviid bugs adapted to living in human habitations,
mainly:Triatoma infestans, Rhodnius prolixus, and
Panstrongylus megistus. These are large (up to 3 cm
long) night-biting bugs, which typically defecate while
feeding. The faeces of infected bugs contain the
metacyclic trypomastigote.
Life cycle

 The infective metacyclic trypomastigotes in bug faeces

are slender and highly motile and they can penetrate a
wide variety of cells, particularly muscle cells, but also
phagocytic cells. Inside the cell, T. cruzi escapes from
the cell vacuole (phagolysosome) to lie free in the
cytoplasm.
 There it loses its flagellum and transforms to a small
oval amastigote stage which multiplies by binary fission,
forming a pseudocyst. After about 5 days, the
amastigotes in the mature pseudocyst transform to
small C-shaped trypomastigotes which are released
when the pseudocyst ruptures.
Life cycle

 Trypomastigotes released from pseudocysts are of two

types: a slender, highly active trypomastigote, which
can be found in the blood only in severe acute
infections, and a smaller, broader, less motile
trypomastigote, which is the most common form found
circulating in the blood.
Life cycle

 If the host is attacked by bugs, trypomastigotes

transform in the bug foregut to the epimastigote stage
which then transforms into metacyclic trypomastigotes
and migrate to the hindgut of the bug thus completing
the cycle.
 Bugs become infective 10–15 days after taking the
infected blood meal.
Pathogenesis

 Local histologic changes include intracellular parasitism

of muscle and other subcutaneous tissues, interstitial
oedema, lymphocytic infiltration, and reactive
hyperplasia of adjacent lymph nodes. Muscles,
including the myocardium, are the most heavily
parasitized tissues. A lymphocytosis accompanies the
high parasitemia of the acute stage.
CLINICAL FEATURES

 Acute stage
 Chronic stage

 ACUTE STAGE- Acute phase occurs soon after infection and

may last for 1–4 months.
 It may be asymptomatic.
 Chagoma is the typical subcutaneous lesion occuring at the
site of inoculation. Inoculation of the parasite in
conjunctiva causes unilateral, painless oedema of
periocular tissues in the eye referred to as Romana’s sign.
 This is a classical finding of the acute Chagas' disease.
Romana’s sign
CLINICAL FEATURES

 Fever, lymphadenopathy, and hepatosplenomegaly.

 Features of myocarditis e.g breathlesness, chest pain,
fatigue, palpitations.
 Features of meningoencephalitis- headache, neck
stiffness, fever, altered mental state
CLINICAL FEATURES

 CHRONIC STAGE: The chronic form is found in adults

and older children and becomes apparent years or even
decades after the initial infection.
 Features of cardiomyopathy: The cardiomyopathy that
develops insidiously often primarily affects the right
ventricle, and the classic signs of right-sided heart
failure are frequently present ( pedal oedema, tender
hepatomegaly).
 Features of megaoesophagus and megacolon: dysphagia,
odynophagia, chest pain, cough, and regurgitation and
abdominal pain and constipation for megacolon.
DIAGNOSIS OF CHAGAS’
DISEASE
 Microscopy: Demonstration of T. cruzi in blood or
tissues. Trypomastigotes can also be seen in thick and
thin peripheral blood smear stained with Giemsa stain.

 Culture: Novy, Neal, and Nicolle (NNN) medium or its

modifications are used for growing T. cruzi.
DIAGNOSIS OF CHAGAS’
DISEASE
 Xenodiagnosis: This is the method of choice in
suspected Chagas’ disease and it’s usually carried out if
other examinations are negative, especially during the
early phase of the disease onset.
 Histopathology: Biopsy examination of lymph nodes and
skeletal muscles and aspirate from chagoma may reveal
amastigotes of T. cruzi.
DIAGNOSIS OF CHAGAS’
DISEASE
 Serology:
 Antigen detection-T. cruzi antigen can be detected in
urine and serum in patients with chronic Chagas’
disease.
 Antibody detection- Antibodies (IgG) against T. cruzi
may be detected.
DIAGNOSIS OF CHAGAS’
DISEASE
 Molecular diagnosis: Polymerase chain reaction.

 Other investigations: Electrocardiography (ECG) and

Chest Xray are useful for diagnosis of cardiomyopathy
seen in chronic Chagas’ disease.
DIAGNOSIS OF CHAGAS’
DISEASE
 The combination of right bundle branch block (RBBB)
and left anterior fascicular block is a typical feature of
Chagas’ heart disease.

 Endoscopy: helps in visualization of megaoesophagus in

Chagas’ disease.
TREATMENT

 Nifutrimox and benznidazole have been used with some

success in both acute and chronic Chagas’ disease.
 Surgery is a vital part of the treatment of chronic
Chagas’disease, particularly for megaoesophagus and
megacolon.
PREVENTION

 Application of insecticide to control the vector bug.

 Personal protection using insect repellent and mosquito
net.
 Improvement in rural housing and environment to
eliminate breeding places of bugs.
Trypanosoma rangeli

 It is nonpathogenic.
 T. rangeli infections are encountered in most areas
whereT. cruzi infection also occurs (Mexico, Central
America, and northern South America).
 It is commonly found in dogs, cats and humans.
CONCLUSION

 During this lecture we have been able to:

 List the agents of African and American
Trypanosomiasis.
 Classify and describe the morphology, life cycle,
pathogenesis and clinical features of infection with the
trypanosomes.
 Outline the management modalities of Trypanosomiasis.
REFERENCES

 Mandell, Douglas and Bennett’s Principles and Practice

of infectious Diseases 7th Edition.
 Medical Parasitology 2nd Edition by Arora
 THANK YOU