1. Tissue flagellates Mrs. Dalia Kamal Eldien MSC in microbiology Lecture NO 8

2. Objectives  Common species of Leishmania  Common disease cause by Leishmania  Cutaneous leishmaniasis  Diffuse cutaneous leishmaniasis  Mucocutaneous leishmaniasis  Visceral leishmaniasis  Morphology of the parasite (Amastigote& promastigote form)  Vector and life cycle  Laboratory diagnosis

3. Leishmaniasis  Leishmania are one of several genera within the family Trypanosomatidae, and are characterized by the possession of a kinetoplast, a unique form of mitochondrial DNA  Leishmaniasis is a parasitic infection transmitted by the bite of an infected female sandfly  A total of about 21 Leishmania species have been identified to be pathogenic to human.

4. Common species  Leishmania tropica  Leishmania major  Leishmania donovani  Leishmania braziliensis  Leishmania chagasi  Leishmania mexicana

5.  The disease has four main forms  Cutaneous leishmaniasis  Diffuse cutaneous leishmaniasis  Mucocutaneous leishmaniasis  Visceral leishmaniasis

6. Cutaneous leishmaniasis  Cutaneous leishmaniasis produces skin lesions mainly on the face, arms and legs. Although this form is often self-healing, it can create permanent scars.  After recovery or successful treatment, cutaneous leishmaniasis induces immunity to re-infection.

7. Cutaneous leishmaniasis  The main species are:-  Leishmania tropica :referred to as dry urban oriental sore, produce dry painless ulcers  Leishmania major referred to as wet rural oriental sore. The early papule is often inflamed and resembles a boil  Leishmania aethiopica  Leishmania mexicana

8. Cutaneous leishmaniasis

9. Diffuse cutaneous leishmaniasis  Diffuse cutaneous leishmaniasis is difficult to treat due to disseminated lesions that resemble leprosy and do not heal spontaneously.  This form especially is related to a defective immune system and it is often characterized by relapses after treatment.

10. Diffuse cutaneous leishmaniasis

11. Mucocutaneous leishmaniasis  Mucocutaneous leishmaniasis, causes disfiguring lesions to the face; it destroys the mucous membranes of the nose, mouth and throat.  Reconstructive surgery of deformities is an important part of therapy

12. Mucocutaneous leishmaniasis

13. Visceral leishmaniasis  Visceral leishmaniasis, also known as 'kala azar‘.  It is the most severe form of Leishmaniasis, and is usually fatal if left untreated.  As the name suggests, visceral leishmaniasis affects the visceral organs, most notably the liver, spleen and bone marrow.  Is characterized by irregular fever, weight loss, swelling of the liver and spleen and anaemia.  The incubation period can be months or years.

14. Visceral leishmaniasis  After treatment and recovery, patients may develop chronic cutaneous leishmaniasis that requires long and expensive treatment.  Main causative agent: o Leishmania donovani o Leishmania infantum

15. Morphology of the parasite  Amastigote form: Are ovoid and non flagellated form of Leishmania, measuring 3-5 µm in length9. On simple light microscopy, a central round or oval nucleus and adjacent but smaller round or rod shaped kinetoplast can be discovered.  Promastigote form: In the sandfly host the parasite is found in the promastigote form. The transformation of amastigotes to promastigotes starts within hours of ingestion of the amastigotes and occurs exclusively in the gut

16. Morphology of leishmania parasite

17. Vector and life cycle  Leishmania species are transmitted by the bite of an infected female sandfly, belonging to the genus Phlebotomus  Promastigotes injected through skin when sandfly takes a blood meal.  Promastigotes taken up by macrophages. Become amastigotes. Multiply in reticuloenothelial cells (VL) or skin macrophages (CL, MCL).  Amastigotes ingested by sandfly.  Amastigotes become promastigotes, multiply and Migrate to head and mouth parts of fly.

18. Sand fly

19. Life cycle

20. Laboratory diagnosis  The laboratory diagnosis of leishmaniasis is by Finding amastigotes, amastigote stage seen in clinical samples is commonly known as Leishman-Donovan (LD) bodies, examine in:  Lesion scraping, biopsy, impression smears (CL)  Aspirates from spleen, bone marrow, lymph nodes (VL)  Buffy coat preparations, peripheral blood monocytes and less commonly in neutrophils (VL)

21. Amastigotes in monocyte

22. a stained bone marrow specimen from a patient with visceral leishmaniasis—showing a mastigote in macrophage

23. Leishmania donovani, causes Kala- Azar, smear from the infected spleen

24.  Histopathological exam of lesion biopsies  Culture  Serological test:-  Specific humeral response in VL  Cell mediated immune response in CL and MCL Laboratory diagnosis

25. Different Serological tests Indirect fluorescence antibody test (IFAT) ELISA Direct agglutination test Rapid antibody detection test immunochromatographic strip Antigen detection test  Molecular biology is increasingly becoming relevant to the diagnosis and control of infectious diseases. Laboratory diagnosis

26. Formal gel test Formol gel (aldehyde) test  This is a non-specific screening test which detects marked increases in IgG.  Large amounts of polyclonal non-specific immunoglobulin are produced by patients with active VL. Use for detection of anti-leishmanial antibody  Collect about 5 ml of venous blood in a dry glass tube and leave to clot.  Collect 1 ml serum Add 2 drops of concentrated formalin solution and mix. Allow to stand for 30 minutes.  Most positive tests, however, can be read after a few minutes.

27. Formal gel test