2. Introduction
Leishmania causes a fatal vector-borne parasitic disease
called Leishmaniasis .
It is spread by the bite of sandflies of the genus
Phlebotomus in the Old World, and of the genus
Lutzomyia in the New World.
Leishmaniasis is the second-largest parasitic killer in the
world (after malaria) and is endemic in many parts of
Africa, Asia and South America.
3. Classification
Kingdom
Subkingdom
Phylum
Subphylum
Class
Order
Genus
Species
Protista
Sarcomastigophora
Protozoa
Mastigophora
zoomastigophora
Kinetoplastida
Leishmania
donovani, tropica,
mexicana ,
braziliensis, etc.
4. Can be classified based on
1. Clinical disease
1. Visceral
2. Cutaneous
3. mucocutaneous
2. Geographical distribution
1. Old world leishmaniasis
2. New world leishmaniasis
5. Visceral leishmaniasis
Aka., kala azar, most severe form of leishmaniasis
Caused by old world sps, L.donovani and L.infantum
and new world sps, L.chagasi
6. L.donovani
Habitat-
Are essentially the parasites of visceral organs.
Promastigote forms found in sand fly and in culture.
Amastigote forms found in man in reticuloendothelial
cells of spleen, bone marrow, liver, intestinal mucosa,
mesentric lymph node.
7. Geographical
distribution
It is estimated that visceral leishmaniasis (VL)
affects > 100 million people worldwide.
Over 90% of reported cases are from India,
Bangladesh, Nepal, Sudan and Brazil. In India it is
prevalent in the eastern region
including Bihar, West Bengal, eastern Uttar
Pradesh, Assam and foothills of Sikkim.
During the epidemic of 1984–1994 death toll was as
high as 70% in the Sudanese population.
Due to emergence of drug resistance the
prevalence is not subsiding, and in fact has spread
to central Europe. For example, during the late
1990s hundreds of cases were reported
in Switzerland.
8. Morphology
The parasite exists in 2 forms
1. Amastigotes –
aflagellar stage
2. Promastigotes –
flagellar stage
9. Morphological difference
Amastigotes Promastigotes
Aflagellar stage Flagellar stage
Occurs in the vertebrate host Occurs in the sand fly
divides by binary fission at 37oC divides by binary fission at 27oC.
There are round or oval ;2-4µm along
longitudinal axis.
They are spindle shaped ;15-20 µm in length &
1- 2µm in width.
Nucleus relatively larger and situated centrally Nucleus smaller and situated in the middle of
the cell or along the side of cell-wall
Kinetoplast situated right angle to nucleus Kinetoplast lies transversely near the anterior
end.
11. Life cycle completes in 2 different host
Sandfly – intermediate host
Humans and other vertebrates - definitive host
Source of infection:- infected sand fly
Mode of transmission:-
Bite of vector sandfly “Phlebotomus argentipus “
Blood transfusion
Congenital infection
Accidental inoculation in lab workers
Sexual intercourse
Infective form:- promastigotes
12. Leishmaniasis is transmitted by the bite of infected
female phlebotomine sand flies.
The sand flies inject the infective stage (i.e.,
promastigotes) from their proboscis during blood meals .
Promastigotes that reach the puncture wound are
phagocytized by macrophages and other types of
mononuclear phagocytic cells.
Promastigotes transform in these cells into the tissue
stage of the parasite (i.e., amastigotes) , which multiply
by simple division and proceed to infect other
mononuclear phagocytic cells .
13. Sand flies become infected by ingesting infected cells
during blood meals .
In sand flies, amastigotes transform into promastigotes,
develop in the gut (in the hindgut for leishmanial
organisms in the Viannia subgenus; in the midgut for
organisms in the Leishmania subgenus), and migrate to
the proboscis .
14. Pathogenesis
After inoculation by sandflies, the flagellate(promastigote)
form binds to macrophages in skin.
2 of the parasite surface molecules (63-kDa glycoprotein
gp63 and lipophosphoglycan- LPG) bind with complement
receptors(C3b and C3bi) present on surface of
macrophages.
Promastigotes phagocytosed by macrophages are
transformed into amastigotes and multiply by binary fission
within phagolysosome of macrophages
15. Amastigotes invade throughout the RES of spleen, liver,
bone marrow, and LNs leading to progressive
heterotrophy
Proliferation and destruction of RES of internal organs
and heavy parasitisation of skin and other organs by
parasitized cells are the characteristic pathological
changes.
17. 6. Others:- kala-azar with HIV co-infection Post kala-
azar dermal leishmaniasis(PKDL)
Complications:- pneumonia, TB, dysentery,
uncontrolled haemorrhage
Prognosis:- With an early treatment, cure rate >90% If
not treated, death occurs within 2 years.
18. Lab diagnosis
Direct
evidence
Peripheral
blood by
thick film
Blood culture
in NNN
medium
Biopsy:- bone
marrow or
spleen
Indirect
evidence
Blood count
Serum tests
Other
methods
Animal
inoculation
Leishmanin or
Montenegro
test
Adlers test
Laboratory diagnosis
20. Microscopy of splenic smear
Most sensitive method to detect LD bodies
98% positivity
average Amastigote density Grade
>100 Parasites/field 6+
10-100 Parasites/field 5+
1-10 Parasites/field 4+
1-10 Parasites/10 fields 3+
1-10 Parasites/100 fields 2+
1-10 Parasites/1000 fields 1+
0 Parasites/1000 fields 0
21. Microscopy of bone marrow
aspiration
Samples collected from
sternum or iliac crest.
LD bodies seen in stained
smear
Safer than splenic puncture
54-86% positivity
22. Culture media for axenic culture
Biphasic media
Novy & McNeal and Nicolle (NNN) medium
Rabbit blood agar that has overlay of Locke’s solution and
antibiotics
the specimen are inoculated into water of condensation
and observed for motile promastigotes for 1-4 weeks at 22˚C
23. Liquid media
Include cell culture medias such as
Schneiders
Grace’s
Mituhasi-Maramorosch
Defined media
Dulbecco’s minimum essential media supplemented with
Tween 80,haemin, biotin, bovine serum albumin fraction
V
24. Animal inoculation
IP inoculation of chineese and golden hamster.
Amastigotes can be seen in stained impression smears
of the spleen
25. Indirect evidences
Blood count
1. Leucopenia (progressive)
2. Anaemia (raised ESR)
Serum tests
1. Aldehyde test( napiers)- positive after
3 months.
2. Antimony test(chopras)- less reliable.
Not used now.
3. Complement fixation test with W.K.K.
antigen. Not used now.
4. Demonstration of antibodies (ELISA,
DAT, IHA, IFA with specific antigen
etc.)
5. Molecular diagnosis:- DNA Probes, PCR,
etc.
26. Leishmanin or Montenegro Test
It is a delayed hypersensitivity test. 0.2 ml of leishmania
antigen is injected intradermally. The test is read after 48-72
hrs.
Positive result is indicated by an induration of 5mm or more.
Positive reaction indicates prior exposure to leishmanial
parasites.
In kala-azar (visceral leishmaniasis), this test is negative.
27. Aldehyde test of Napier
Add a drop of 40% formaldehyde to 1
or 2 ml of serum in a test tube.
Positive :- jellification of milky white
opacity like egg white within 2-20
min.
Aldehyde positive Ab’s appear only
after 3 months of infection
Antimony test of chopra
Development of white flocculent
precipitate on addition of urea
stilbamine solution to patients serum
28. Epidemiology
Found on every continent except Australia and
Antarctica.
For cutaneous leishmaniasis, number of cases range
from 0.7 million to 1.2 million .
For visceral leishmaniasis, number of cases range from
0.2 million to 0.4 million.
Annual incidence of disease= 600,000 cases per year.
People infected worldwide=12 million.
29. Prevention and control
Reduction of sand fly
population
Reduction of reservoirs
Education in the
community
Prevention of exposure
33. Habitat
Parasites of Human skin
Amastigotes found in monocytes, polymorphonuclear
leucocytes and endothelial cells of capillaries of skin
Not found in peripheral blood or internal organs
Morphology
Similar to L.donovani
Lifecycle
Similar to L.donovani except that amastigotes reside in
large mononuclear cells of skin
Transmitted by P.argentipes
34. Pathogenesis
After inoculation by sandflies, the flagellate(promastigote)
form binds to macrophages in skin.
Promastigotes are phagocytosed and transformed into
amastigotes.
Papules develop at the site of bite, weeks or sometimes years
after.
It is caused by multiplication of phagocytes that contain
numerous amastigotes.
Lesion enlarges, necrosis, and ulcerates.
35. Clinical manifestations
i. Localised cutaneous leishmaniasis
Caused by L.tropica and L.aethiopica
Commonly seen in children and young adults
ii. Oriental sore
Typical ulcer found on skin
Incubation period: 2 to 4 months
Lesion begins as single red, and pruritic papule at the site of bite.
Gradually increase in size and finally ulcerates (Oriental sore).
36. iii. Diffuse cutaneous leishmaniasis
Caused by L.tropica occurs in an anergic host with poor immune
response.
Patients develop multiple, wide spreaded papules and nodules
iv. Leishmaniasis recidivans
Uncommon clinical form of leishmaniasis caused by L.tropica
Recurrence of leisions at the site of apparently healed disease years
after a localized cutaneous lesion has healed.
Manifest as enlarging papules, plaque, or coalescence of papules that
heals as central scaring at face
Destruction of nose
37. Epidemiology
1-1.5 new cases per year
Geographical distribution
World :- Iran, Afganistan, Brazil, Peru, Saudi Arabia,
and Syria.
India :- Rajasthan, Kerala, and Uttarakhand.
38. Laboratory diagnosis
Specimen : obtained from margin of ulcer by puncture of the
raised nodules or aspiration of outer edge of the ulcer.
Microscopy
Geimsa or Leishman:- demonstration of amastigotes.
Culture
NNN medium
Animal inoculation
Intradermally in hamster
Serodiagnosis
Aldehyde test is negative
Leishmanin skin test :- positive
39. Leishmania Mexicana complex
Includes
Leishmania Mexicana Mexicana
Leishmania Mexicana amazonensis,
Leishmania Mexicana venezuelensis
Leishmania Mexicana pifanoi
New world cutaneous leishmaniasis
Chiclero ulcer aka bay ulcer
41. Leishmania vianna braziliensis
Widespread in latin America
Habitat
Amastigotes are found in humans and other vertebrates.
Intracellular parasites and are found inside the
macrophages of the skin and in the mucous membrane
Promastigotes are found in sandfly and culture
42. Morphology
Similar
Lifecycle
Identical of L.donovani
Except that the amastigotes are found in reticuloendothelial cells
and lymphatic tissues of skin
Pathogenesis
Condition occurs when cutaneous lesions expand directly to mucosal
region or through metastasis.
Pathogenesis of espundia and oriental sore is similar, except that
Espundia expand rapidly, forming large and long lasting ulcers with weeping
surface
Mucosal metastasis
43. Clinical manifestations
More variable, chronic, and severe
Espundia
inital symptoms are similar to that of cutanous leishmaniasis
single or multpile lesions and ulcers develop at the mucosal regions
(nose, mouth, throat cavities) and in the adjacent tissue
The ulcerations can involve the nose, pharynx, palate and lips.
Invasion of the larynx may result in a loss of speech.
Pian bios:- single or multiple and painless dry persistent
ulcers all over the body
Uta :- single or multiple ulcers in the face
44. Geographical distribution
World : brazil, Paraguay, Ecuador, Bolivia, Peru, Colombia and
Venezuela.
India : no condition
Vector
Lutzomiya spp.
Reservoir
Rodents.
Mode of transmission
Bite of sandfly vectors
Less frequently by ticks, direct human-human transmission and
autoinfection
45. Laboratory diagnosis
Specimen :- slit skin biopsy, aspiration from edge of ulcer and
from nodules in skin
Microscopy :- amastigotes in stained smear
Culture :- grows poorly
Animal inoculation :- slowly grows in Hamster and takes
longer time to produce pathological lesions
Serodiagnosis :-
IFA
Leishmanin test :- positive
