Blood & Tissue Protozoa-II

Prof.Dr.Shaheen Sharafat
MBBS.M.Phil Ph.D
Dept.Of Pathology
LNH&MC
 Objectives
• - Discuss the important properties
pathogenesis , clinical findings,laboratory
diagnosis, treatment and prevention of
Leishmania and Toxoplama
 Leishmaniasis
Leishmaniasis is a disorder resulting from
infection by protozoan parasites called
Leishmania
Four major human pathogens of the genus
Leishmania
• L.donovani (Visceral leishmaniasis or Kala-azar
• L.tropica (cutaneous.L, oriental sore, Dehli boil)
• L.mexicana ( Espundia)
• L.braziliensis (Mucocutaneous leishmaniasis in
Central and South America)
 Visceral leishmaniasis

Occurs in Mediterranean basin, Middle East,

southern Russia, and parts of China,where the
reservoir hosts are primarily dogs and foxes
In subSaharan Africa, rats and small carnivores
(e.g., civets) are the main reservoirs
A third pattern is seen in India and neighboring
countries (and Kenya), in which humans appear
to be the only reservoir
Pathogenesis of Visceral Leishmaniasis
- Reticuloendothelial system (liver, spleen, and bone
marrow) is most severely affected.
- Reduced bone marrow activity
- Cellular destruction in the spleen
- Anemia, leukopenia, and thrombocytopenia.
- Tendency to bleed
Striking enlargement of the spleen due to combination of
proliferating macrophages and sequestered blood cells
The marked increase in IgG is neither specific nor
protective
The majority of VL cases occur in just six countries —
Bangladesh, Brazil, Ethiopia, India, Nepal and Sudan1.
1. Desjeux, P. Disease Watch Focus: Leishmaniasis. Nature Rev. Microbiol. 2, 692–693
(2004).
 Cutaneous leishmaniasis
1-Leishmania tropica found in the Old World (Oriental sore,
Delhi boil)
2- L. mexicana found only in the Americas (Chicle ulcer /Bay
sore)
3- Mucocutaneous leishmaniasis :- L.braziliensis occurs only in
Central and South America (espundia),
 Life cycle
Female Sandfly is the vector and a variety of mammals
such as dogs, foxes, and rodents are reservoirs
When the sandfly sucks blood from an infected host, it
ingests macrophages containing amastigotes
Amastigotes differentiate into promastigotes in the gut,
multiply and migrate to the pharynx and proboscis,
where they can be transmitted during the next bite.
The cycle in the sandfly takes approximately 10 day
• The organisms are transmitted to humans by the bite
of female sandflies (Phlebotomus in the Eastern
Hemisphere; Lutzomyia in the Western Hemisphere).
• In endemic areas, nonhuman mammalian species
serve as reservoir hosts.
• The vector sand fly breeds in forest areas, caves, or
the burrows of small rodents.
• There are more than twenty species of Leishmania
that can infect human beings.
 Leishmania-The parasite
The Parasite exists in two forms:
Amastigote (LD body, aflagellate form):
• Occurs in vertebrate host (humans, dog, hamster)
• It is found in macrophages, monocytes,
neutrophils or endothelial cells.
Promastigote (Leptomonad, Flagellated form):
• Occurs in culture
• Single flagellum
 Promastigotes

Amastigotes
Morphological forms of Leishmania
 Visceral Leishmaniasis
Kala-azar (L.donovani)
• It spreads into the spleen, bone marrow and liver and
attacks and destroys the immune system.
• This form occurs 2 - 8 months after a person is bitten by
the female sandfly.
• Most people do not remember having a skin sore.
• This form can lead to deadly complications.
• The organs of reticuloendothelial system (liver, spleen,
bone marrow) are severely affected.
• Consequently there is anaemia, leukopenia, and
thrombocytopenia.
 Clinical findings
Visceral leishmaniasis:-Intermittent fever, weakness, and
weight loss. Massive enlargement of the spleen is characteristic.
-Hyperpigmentation of the skin is seen in light skinned patients
(kala-azar means black sickness).
The course of the disease runs for months to years. Initially,
patients feel reasonably well despite persistent fever.
As anemia, leukopenia, and thrombocytopenia become more
profound, weakness, infection, and gastrointestinal bleeding
occur.
Untreated severe disease is nearly always fatal as a result of
secondary infections
 Clinical Presentation

• Cutaneous Leishmaniasis :-The most common clinical

presentation is single or multiple localized ulcers or
nodules.
• Typically, ulcerative CL starts as a small, erythematous
papule at the site where promastigotes, the infective
form of the parasite, are inoculated by the bite of a
sandfly.
• The incubation period is usually several weeks or
months, but lesions can appear within a few days or
several years after leaving an endemic area.
 Cutaneous Leishmaniasis
Clinical Presentation
• In most cases, within a few weeks the papule
enlarges, crusts over, and breaks down into a
slow-growing ulcer up to several centimeters in
diameter.
• The ulcer is shallow and well-defined, with a
raised erythematous border and central
granulation tissue.
• The ulcer heals slowly, leaving a depressed,
atrophic scar.
 Symptoms
• Cutaneous leishmaniasis affects the skin and sometimes
the mucus membranes. Symptoms may include:
• Skin sores, which may become a skin ulcer that heals
very slowly.
• Ulcers and wearing away (erosion) in the mouth,
tongue, gums, lips, nose, and inner nose.
• Stuffy nose, runny nose, and nosebleeds.
• Breathing difficulty.
• Swallowing difficulty.
There are about 1.5 million cases of cutaneous
Leishmaniasis each year worldwide, with the
bulk reported from Afghanistan, Iran, Iraq,
Algeria, Saudi Arabia, Peru, and Pakistan
The most common clinical presentation is
localized ulcer or nodule
A= Cutaneous
leishmaniasis

B= Mucocutaneous
A B
leishmaniasis

C D

C= Visceral D= nodular post-kala-azar

leishmaniasis dermal leishmaniasis (PKDL).
 Clinical Manifestations
Type Pathogen Location
Visceral leishmaniasis Caused exclusively by Found in tropical and
infections are often species of the L. subtropical areas of all
recognized by fever, donovani complex (L. continents except
swelling of the liver and donovani, Australia, most
spleen, and anemia. L. infantum common in Bangladesh,
Many local names, the Brazil, India, Nepal and
most comm Sudan. Also found in
Kala azar part of China,
 Clinical Manifestations
Type Pathogen Location
Cutaneous leishmaniasis The most common is the Cutaneous infections are
(localized and diffuse) Oriental Sore (caused by most common in
infections appear as Old World species L. Afghanistan, Brazil, Iran,
obvious skin lesions. major, L. tropica, and L. Peru, Saudi Arabia and
aethiopica). In the New Syria.
World, the most
common culprits is L.
mexicana.

Mucocutaneous L. braziliensis Mucocutaneous

leishmaniasis (Espundia infections are most
or Uta) infections start common in Bolivia,
off as a lesion at the Brazil and Peru.
bite, and can Mucocutaneous
metastasise into the infections are also found
mucous membrane and in Karamay, China
become fatal. Xinjiang Uygur
Autonomous Region.
 Pathogenesis of Cutaneous and
Mucocutaneous Leishmaniasis
• Lesions are confined to the skin in cutaneous leishmaniasis and
to the mucous membranes, cartilage, and skin in mucocutaneous
leishmaniasis.
The initial lesion is a red papule at the bite site, usually on an
exposed extremity that forms a granulomatous necrotic ulcer
• This enlarges slowly to form multiple satellite nodules that
coalesce and ulcerate
• Secondary bacterial infection is common that can be fatal
• Mucocutaneous leishmaniasis begins with a papule at the bite
site, but then metastatic lesions form, usually at the
mucocutaneous junction of the nose and mouth if cell mediated
immunity is deficient
Disfiguring granulomatous, ulcerating lesions destroy nasal cartilage but not
adjacent bone
 Lab.Diagnosis
1-Microscopic detection of amastigotes (LD Bodies) in a bone
marrow, spleen, or lymph node biopsy or “touch” preparation or
skin biopsy/smear taken by scrapping the periphery of skin
lesion
2- Culture
3-Serologic (indirect immunofluorescence) tests are positive in
most patients.
4- A skin test using a crude homogenate of promastigotes
(leishmanin) as the antigen is available
skin test is negative during active disease but positive in patients
who have recovered
 Lab diagnosis
For Visceral Leishmaniasis:
• Demonstration of amastigote forms in smears of
blood, bone marrow, spleen aspirate or biopsy
tissue by Romanowsky’s stain.
For cutaneous and mucocutaneous forms:
• Demonstration of amastigote forms, by
Romanowsky’s or Leishman’s stain, in samples
taken from lesions on skin and mucous
membranes.
• Culture: NNN medium is used to culture
leishmania from clinical samples.
Leishmania donovani in bone marrow cell
Cases of
leishmaniasis from
Pakistan
Treatment: Liposomal amphotericin B or sodium
stibogluconate ( the mortality rate is reduced to almost
5% with proper thrapy)
Recovery results in permanent immunity
Prevention: Prevention involves protection from sandfly
bites by using netting, window screens, protective
clothing, and insect repellents
 Toxoplasmosis
Toxoplasma gondii a unicellular parasite
Causes toxoplasmosis, including congenital toxoplasmosis
Domestic cat is the definitive host along with other felines;
humans and other mammals are intermediate hosts
Humans get Infected by ingestion of cysts in undercooked meat
or from accidental contact with cysts in cat feces
The cysts rupture in the small intestine and release forms that
invade the gut wall, where they are ingested by macrophages
and differentiate into rapidly multiplying trophozoites
(tachyzoites), which kill the cells and infect other cells
Ingestion of cysts in raw meat
Cell-mediated immunity usually limits
the spread of tachyzoites
Parasites enter host cells in the brain,
muscle, and other tissues, where they
develop into cysts in which the parasites
multiply slowly in the form of
Bradyzoites
• Recent or acute infection with T gondii can be
evaluated
• A suspected diagnosis of acute toxoplasmosis should
be confirmed by
Serology:- TXM / Toxoplasma gondii Antibody, IgM in
serum
• Molecular Detection :-Polymerase chain reaction
(PCR) detection of Toxoplasma gondii DNA by analysis
of cerebrospinal fluid or amniotic fluid specimens by
amniocentesis