1) Visceral leishmaniasis is caused by the protozoan Leishmania donovani and L. infantum, transmitted by the bite of infected sandflies. 2) It affects the reticuloendothelial system and causes a pentad of fever, weight loss, hepatosplenomegaly, pancytopenia, and hypergammaglobulinemia. 3) Diagnosis involves microscopy of tissue aspirates to detect amastigotes or culture to grow promastigotes, and serology to detect antibodies. Liposomal amphotericin B is the recommended treatment in most regions.

1. Essentials of Medical Microbiology by Apurba S Sastry © Jaypee Brothers Medical Publishers
VISCERAL
LEISHMANIASIS
Dr.Vaishali T
2nd year PG
Department of Microbiology

2. Essentials of Medical Microbiology by Apurba S Sastry © Jaypee Brothers Medical Publishers
INTRODUCTION
• Hemoflagellates - flagellated protozoa that are found in
peripheral blood circulation
• Examples - Leishmania and Trypanosoma - both are transmitted
by bite of insect vector.
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3. Essentials of Medical Microbiology by Apurba S Sastry © Jaypee Brothers Medical Publishers
VARIOUS MORPHOLOGICAL FORMS
OF FLAGELLATE
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A. Amastigote; B. Promastigote; C. Epimastigote; D. Trypomastigote

4. Essentials of Medical Microbiology by Apurba S Sastry © Jaypee Brothers Medical Publishers
INTRODUCTION (CONT..)
• In Leishmania - amastigote and promastigote forms are seen.
Amastigotes - diagnostic form, found in man
Promastigotes - infective stage to man, found in insect
vector
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5. Essentials of Medical Microbiology by Apurba S Sastry © Jaypee Brothers Medical Publishers
INTRODUCTION (CONT..)
• In Trypanosoma, epimastigote and
trypomastigote forms are seen.
In man, trypomastigotes - diagnostic form
In insect vector, both epimastigotes and
trypomastigotes are found, the latter being
the infective stage to man.
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6. Essentials of Medical Microbiology by Apurba S Sastry © Jaypee Brothers Medical Publishers
LEISHMANIASIS
• Caused by an obligatory intracellular protozoa
of the genus Leishmania - primarily affects the
reticulo-endothelial system of the host.
• Vector - bite of the female sandfly vector.
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7. Essentials of Medical Microbiology by Apurba S Sastry © Jaypee Brothers Medical Publishers
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8. Essentials of Medical Microbiology by Apurba S Sastry © Jaypee Brothers Medical Publishers
LEISHMANIASIS (CONT..)
• Clinical forms:
Visceral leishmaniasis (VL)
Post–kala-azar dermal leishmaniasis
(PKDL) - occurs few months to years
followingVL.
Cutaneous forms - cutaneous leishmaniasis
(CL), diffuse cutaneous leishmaniasis (DCL),
leishmaniasis recidivans (LR) and
mucocutaneous leishmaniasis (MCL).
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9. Essentials of Medical Microbiology by Apurba S Sastry © Jaypee Brothers Medical Publishers
LEISHMANIASIS (CONT..)
• Subgenera: Leishmania has two subgenera - L.
Leishmania (L.L.) and L.Viannia (L.V.).
• Both of the subgenera comprise of nearly 20
species.
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10. Essentials of Medical Microbiology by Apurba S Sastry © Jaypee Brothers Medical Publishers
OLD WORLD LEISHMANIASIS
• Seen in Asia,Africa and less frequently in
Europe; transmitted by sandfly of genus
Phlebotomus.
L.L. donovani: CausesVL and PKDL
L.L. infantum: CausesVL and PKDL
L.L. tropica complex: Causes CL
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11. Essentials of Medical Microbiology by Apurba S Sastry © Jaypee Brothers Medical Publishers
NEW WORLD LEISHMANIASIS
• Seen in Central and South America; transmitted
by sandfly of genus Lutzomyia.
L.L. chagasi: CausesVL and CL
L.L. mexicana complex: Causes CL
L.Viannia braziliensis complex: Causes MCL
and CL
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12. Essentials of Medical Microbiology by Apurba S Sastry © Jaypee Brothers Medical Publishers
VISCERAL LEISHMANIASIS
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13. Essentials of Medical Microbiology by Apurba S Sastry © Jaypee Brothers Medical Publishers
VISCERAL LEISHMANIASIS
• Mainly caused by the old world species - L. donovani
and L. infantum, together known as L. donovani
complex and also by new world species, L. chagasi.
• L. donovani - most common species causingVL.
• Found in India and Africa.
• Named after SirWilliam Leishman and Sir Donovan.
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14. Essentials of Medical Microbiology by Apurba S Sastry © Jaypee Brothers Medical Publishers
VARIOUS FORMS OF VISCERAL LEISHMANIASIS
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Characters Old World VL New World VL
Indian VL (kala-azar) Infantile VL African VL Mediterranean VL
Agent Leishmania donovani L. infantum L. donovani L. chagasi
Vector Phlebotomus argentipes P. perniciosus P. orientalis, P. martini Lutzomyia longipalpis
Epidemiology India Middle East, Central Asia,
China and Mediterranean
basin
Sudan, Ethiopia,
Kenya and Uganda
Central and South
America
Age affected Young adults Infants and children <5 years Adults Children
Reservoir Anthroponotic (human) Zoonotic (canine) Anthroponotic,
Rarely-Zoonotic
(rodents)
Zoonotic (canine)
PKDL Common Less common Common Less common
Lymph node
involvement
Less common More common, Aggravated
by poor nutrition
Less common Less common

15. Essentials of Medical Microbiology by Apurba S Sastry © Jaypee Brothers Medical Publishers
EPIDEMIOLOGY OF VL
• Leishmaniasis is endemic in 97 countries.
• Most of them are developing countries of tropical and temperate regions.
• World: About 50-90 thousand new cases ofVL occur worldwide each year - of which >90% of
cases are from three regions: (i) South-EastAsia: India, Bangladesh, and Nepal; (ii) East Africa:
Ethiopia, Sudan, and Kenya; and (iii) Brazil .
India:
• One of the worst affected country.
• In 2019, about 3,139 cases ofVL and 821 cases of PKDL - reported from India with nil death.
• Maximum cases - Bihar (76% ofVL, and 52% of PKDL) and followed by Jharkhand.
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16. Essentials of Medical Microbiology by Apurba S Sastry © Jaypee Brothers Medical Publishers
1
6
Life cycle of
L. donovani

17. Essentials of Medical Microbiology by Apurba S Sastry © Jaypee Brothers Medical Publishers
LIFE CYCLE
• Host
1. Vertebrate host – man, dog, rodents
2. Invertebrate host – female sandfly
• Infective form – promastigotes
• MOT – bite of sandfly
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18. Essentials of Medical Microbiology by Apurba S Sastry © Jaypee Brothers Medical Publishers
PATHOGENICITY
• The phagocytosis of the promastigotes - facilitated by binding of its surface
antigens - gp-63 and LPG (lipophosphogycan)to specific receptors on
macrophages.
• LPG - principle virulence factor - prevents phagosome maturation and
protects the parasite against hydrolytic enzymes secreted from the
phagolysosome.
• GPI (glycosyl-phosphatidyl-inositol) - major surface protein on amastigotes,
helps in protecting from phago-lysosomal attack inside the macrophage.
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19. Essentials of Medical Microbiology by Apurba S Sastry © Jaypee Brothers Medical Publishers
HOST IMMUNE RESPONSE
• Depending on the host immune response (TH1 orTH2) - amastigotes are either killed or allowed to
multiply inside the macrophages.
 TH I RESPONSE:
• IfTH1 cells are activated, leads to a CMI response - contain the disease spread by secreting interferon γ
(IFN- γ) to activate macrophages, which in turn kill the amastigotes.
• Observed in cutaneous leishmaniasis and patients after recovery/treated forVL.
 TH 2 RESPONSE:
• IfTH2 cells are activated, leads to a humoral immune response due to increased production of IL-
10 and IL-4; which in turn causes polyclonal B cell activation leading to hypergammaglobulinemia.
• Observed in patients developing activeVL and in diffuse CL
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20. Essentials of Medical Microbiology by Apurba S Sastry © Jaypee Brothers Medical Publishers
CLINICAL FEATURES (VL)
• VL - also called as kala-azar (a hindi term meaning “black fever”).
• Incubation period - 2–6 months.
• Hallmark ofVL - pentad of fever, progressive weight loss, hepatosplenomegaly, pancytopenia and hypergammaglobulinemia.
• Fever
• Splenomegaly
• Hepatomegaly
• Lymphadenopathy
• Hyperpigmentation
• Pedal edema and ascites
• Hematological abnormalities – pancytopenia and hypergammaglobulinemia
• Leishmanoma – Nodular skin lesions
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21. Essentials of Medical Microbiology by Apurba S Sastry © Jaypee Brothers Medical Publishers
POST-KALA-AZAR DERMAL LEISHMANIASIS (PKDL)
• Non-ulcerative lesion of skin occurs in 2–50% of patients ofVL
following treatment with antimonials.
• Aggravated by exposure to sunlight.
• Mainly seen in India and East African countries
• Develops as hypopigmented macule near mouth which spreads to face,
arms and trunk and finally becomes nodules resembling leprosy.
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22. Essentials of Medical Microbiology by Apurba S Sastry © Jaypee Brothers Medical Publishers
• Ocular lesions - conjunctivitis and uveitis - associated in some
patients.
• Diagnosis: Detection of amastigotes in the nodular lesions and by
serological tests such as direct agglutination test (DAT) and antibodies
to rK39 antigen; positive in most of the cases.
• Amphotericin B - drug of choice, given for 4 months.
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23. Essentials of Medical Microbiology by Apurba S Sastry © Jaypee Brothers Medical Publishers
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A. Splenomegaly seen in visceral leishmaniasis; B. Hypopigmented skin changes in early PKDL;
C and D. Extensive facial nodular lesions in late PKDL

24. Essentials of Medical Microbiology by Apurba S Sastry © Jaypee Brothers Medical Publishers
LEISHMANIASIS WITH HIV CO-INFECTION
• Both HIV and Leishmania affect each other’s pathogenesis
Effect on HIV: Leishmania - cause activation of latent HIV. It
expresses high level of chemokine receptors (CCR5) on
macrophages.
Effect on Leishmania: HIV causes activation of TH2 cells
response leading to disease progression
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25. Essentials of Medical Microbiology by Apurba S Sastry © Jaypee Brothers Medical Publishers
LABORATORY DIAGNOSIS OF VL
• Microscopy—Giemsa staining, detects LD bodies (i.e.
macrophage filled with amastigote forms)
Splenic aspiration: Most sensitive
Bone marrow aspiration: Most commonly preferred
Lymph node aspiration
Liver biopsy
Peripheral blood smear (in HIV-infected people)
Biopsy of various organs (in HIV-infected people)
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26. Essentials of Medical Microbiology by Apurba S Sastry © Jaypee Brothers Medical Publishers
L. DONOVANI AMASTIGOTES: SHOWING A MACROPHAGE
CONTAINING
MULTIPLE LEISHMANIA AMASTIGOTES
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A. Schematic; B. In bone marrow smear stained with Giemsa. Note that each
amastigote has a nucleus (red arrow) and a rod-shaped kinetoplast (black arrow)

27. Essentials of Medical Microbiology by Apurba S Sastry © Jaypee Brothers Medical Publishers
• Culture (detects promastigotes)—useful for
species identification and drug sensitivity testing
NNN (Novy-MacNeal-Nicolle) medium
Schneider’s liquid medium (Drosophila insect
media)
Examined weekly x 4 weeks – giemsa stain
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28. Essentials of Medical Microbiology by Apurba S Sastry © Jaypee Brothers Medical Publishers
LABORATORY DIAGNOSIS OF
VL (CONT..)
28
A. Smear made from culture fluid shows promastigote forms (Giemsa stain); B. NNN medium

29. Essentials of Medical Microbiology by Apurba S Sastry © Jaypee Brothers Medical Publishers
LABORATORY DIAGNOSIS OF
VL (CONT..)
• Antibody detection in serum
Direct agglutination test
ICT – detects antibody using rk39
or rKE16 antigens
ELISA and IFA
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Direct agglutination test

30. Essentials of Medical Microbiology by Apurba S Sastry © Jaypee Brothers Medical Publishers
• Antigen detection - carbohydrate antigen in the urine (latex
agglutination test)
• Molecular method - PCR, real-time PCR detecting specific kinetoplast
(mitochondrial) DNA
• Leishmanin test (montenegro test) –
- Intradermal injection of promastigotes.
- indicates good CMI (DTH reaction);
- positive in all stages (induration >5mm in 72 hrs), except activeVL and
diffuse CL.
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31. Essentials of Medical Microbiology by Apurba S Sastry © Jaypee Brothers Medical Publishers
• Animal inoculation—golden hamster
• Nonspecific tests to detect:
Hypergammaglobulinemia - by Napier’s aldehyde test
and Chopra’s antimony test
Pancytopenia - by complete blood count
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32. Essentials of Medical Microbiology by Apurba S Sastry © Jaypee Brothers Medical Publishers
TREATMENT OF VISCERAL
LEISHMANIASIS
• The various drugs used in the treatment ofVL are:
(i) Pentavalent antimonials,
(ii) Liposomal amphotericin B,
(iii) Miltefosine,
(iv) Paromomycin.
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33. Essentials of Medical Microbiology by Apurba S Sastry © Jaypee Brothers Medical Publishers
PENTAVALENT ANTIMONIALS
• Drug of choice and widely used in the past for several decades.
• Dosage: given as 20 mg/kg per day IM or IV for 30 days
• Mechanism of action: it induces oxidative stress in the parasite
• Resistance: Resistance to antimonials - Bihar, India
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34. Essentials of Medical Microbiology by Apurba S Sastry © Jaypee Brothers Medical Publishers
LIPOSOMAL AMPHOTERICIN B
• It has been the current drug of choice of leishmaniasis, especially in areas where
resistance to antimonials have been reported.
• Dosage: given as 3–5 mg/kg per daily dose by IV infusion for 3–5 days up to a
total dose of 15 mg/kg
• Mechanism of action: It acts by disrupting the cell membrane by forming
pores.
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35. Essentials of Medical Microbiology by Apurba S Sastry © Jaypee Brothers Medical Publishers
MILTEFOSINE
• Dosage: given as 150 mg/day; orally for 28 days
• Mechanism of action: by interacting with lipids, inhibiting cytochrome C oxidase
• Resistance: Resistance to miltefosine is rare, mainly reported from India and Nepal.
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• It is an aminoglycoside antibiotic with anti-leishmanial activity.
• Given IM -15 mg per kg per day for 21 days.
 IMMUNOTHERAPY: Interferon-γ has been used in antimonial resistant and
in selected relapse cases from Kenya, Brazil, and India.
PAROMOMYCIN

36. Essentials of Medical Microbiology by Apurba S Sastry © Jaypee Brothers Medical Publishers
WHO RECOMMENDATION FOR TREATMENT OF VL IN
DIFFERENT REGIONS OF THE WORLD
36
Region Drug regimen(s)
Indian subcontinent (ranked by preference)  Liposomal amphotericin B
 Combinations
 Liposomal amphotericin B plus miltefosine
 Liposomal amphotericin B plus paromomycin
 Miltefosine plus paromomycin
 Amphotericin B deoxycholate
 Miltefosine
 Paromomycin
 Pentavalent antimonials
East Africa and Yemen Pentavalent antimonial plus paromomycin
Mediterranean Basin, Middle East, Central Asia
and in America (due to L. infantum)
Liposomal amphotericin B

37. Essentials of Medical Microbiology by Apurba S Sastry © Jaypee Brothers Medical Publishers
PREVENTION OF VL
• NationalVector-borne Disease Control Program - It has launched the
accelerated plan for kala-azar elimination in 2017.
• Vaccine Trials
• Control Measures
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38. Essentials of Medical Microbiology by Apurba S Sastry © Jaypee Brothers Medical Publishers
CONTROL MEASURES
• Personal prophylaxis by using insect repellents or bed nets
• Control of canine or rodent reservoir
• Phlebotomus does not fly high above the ground level and it is
nocturnal in habitat. So, sleeping at top floors also can prevent
transmission.
• Early treatment of all cases (mainly anthroponoticVL and PKDL
cases).
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39. Essentials of Medical Microbiology by Apurba S Sastry © Jaypee Brothers Medical Publishers
THANK YOU
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