1. MEDICAL PARASITOLOGY & ENTOMOLOGY LECTURER: SR. NORAZSIDA RAMLI

2. BLOOD & TISSUE FLAGELLATES/ HAEMOFLAGELLATES

3. Morphologic forms There are 4 morphologic forms seen in hemoflagellates: 1) Amastigote 2) Promastigote 3) Epimastigote 4) Trypomastigote -they can exist in two or more of the 4 morphologic forms  depending on the species.

4. Kingdom: Protisata Phylum: Sarcomastigophora Class: Zoomastigophora Order: Kimetoplastida Family: Trypanosomatidae Genus: Leishmania Species: donovani, tropica, mexicana, braziliensis

5. Leismania sp. Can cause: 1) Cutaneous leishmaniasis: a localized infection of the capillaries of the skin. 2) Mucocutaneous leishmaniasis: cause lesions of the skin and mucous membranes, specifically of the oral and nasal mucosa. 3) Visceral/sistemic leismaniasis: more generalized symptoms leading to enlargement of the internal organs, especially the liver, lymph nodes and spleen.

6. Indistinguishable in appearance. Differentiated based on: - Geographic distribution. - Pathogenesis. - Kinetoplast DNA (kDNA) analysis - DNA hibridization - Serologic testing.

7. Geographical distribution

8. Leishmania sp. Divided into 4 groups: 1) Leishmania tropica complex – Old World Cutaneous Leismaniasis. 2) Leishmania mexicana complex – New World Cutaneous Leishmaniasis. 3) Leishmania braziliensis complex – Mucocutaneous Laishmaniasis. 4) Leishmania donovani complex – Visceral leishmaniasis.

9. Stage of life Only have 2 stages of life: 1) Amastigote 2) Promastigote

10. Amastigote Size: 5 by 3µm Shape: oval to round Nucleus: One, eccentric. Kinetoplast: Present, Consisting of dot-like blepharoplast, with small axoneme and prabasal body. Flagellum: absent

11. Promastigote Size: 9-15µm Shape: long and slender. Nucleus: one, central. Kinetoplast: Anterior end of the organism, no undulating membrane. Flagellum: Single, anterior free flagellum.

12. Leishmania tropica complex – Old World Cutaneous Leismaniasis. L. tropica - mediterranean region, middle East, Armenia, Caspian region, Afghanistan, India and Kenya (particularly in urban areas) L. aethiopica – Highlands of Ethiopia, Kenya and Southern Yemen. L. major – Desert regions of Turkmenistan, Uzbekistan and Kazakhstan, Northern Africa and the Sahara, Iran, Syria, Israel and Jordan.

13. Morphology Cause a chronic disease: cutaneous leishmanisis. Also known as Oriental sore, Delhi boil and dry or urban cutaneous leishmaniasis. Characterized by: production of dry, raised, ulcerated lesions at bite sites. Vectored by: tiny sandflies of the genera Phlebotomus.

14. L. tropica

15. Vector: Phlebotomus sandfly

16. Sandfly vs mosquito mosquito sandfly

18. Life cycle Only the female sandfly transmits the parasites. Vector draws a blood meal from an infected host  amastigotes form transform into promastigotes n multiply (within fly gut)  promastigotes migrate to the pharynx  fly feeds again, transmitted to a new host  engulfed by reticuloendothelial cells  amastigotes multiply repeatedly by binary fission.  cell ruptures  amastigotes invade new macrophages and perpetuate the cycle.

19. Life cycle

20. Transmission & Pathogenesis Incubation period vary from several weeks to as three years. First sign of cutaneous leishmaniasis= the development of a small red papule at the initial site of the insect bite. A local granulomatous response leads to the formation of a crateriform lesion  2cm or more. L. tropica n L. aethiopica  produce dry lesions. L. major  produce moist lesions with a serous exudate. Lesions can heal spontaneously but may leave serious scars.

21. Contact spread of infection also possible. Patient may produce multiple sores by scrathing and autoinoculating normal skin. Diffuse cutaneous leishmaniasis (DCL): -occur in anergic patient (who is unable to amount an adequate immune respon). -characterized by the presence of multiple nodular lesions, particularly on the face and limbs. -loaded with parasites and do not heal spontaneously.

22. DCL

25. Laboratory Diagnosis Montenegro (leishmanin) skin test -delayed hypersensitivity reaction provoked by a suspension of killed leishmanial promastigotes administered intradermally. -local inflammatory reaction appears at the site of injection within 48-72 hours. Microscopy examination Isoenzyme studies Molecular diagnostic technique- PCR Serologic test – ex: indirect fluorescent antibody assay.

26. Treatment Sodium stibogluconate (antimony sodium gluconate: Pentostom). Meglumine antimonate Glucantime Ampotericin B Ketoconazole

27. Prevention Use of bed netting Insect repellent and residental spraying Rodent control in reducing transmission. Individuals with active lesions  promptly treated, wound covered  to prevent autoinfection and further insect transmission to other individuals.

28. Leishmania mexicana complex – New World Cutaneous Leishmaniasis L. mexicana – Belize, Guatemala, and the Yucatan peninsula. L. pifanoi – Amazon river basin and parts of Brazil and Venezuela. L. amazonensis – Amazon basin of Brazil. L. venezuelensis – forests areas of Venezuela. L. garnhami – Venezuelan Andes.

29. Morphology Distribution extends from Southern Texas in the United states, through Mexico, Central and South America. L. mexicana causes chiclero ulcer or Bay sore. L. pifanoi causes DCL. L. amazonensis causes cutaneous and DCL. L. venezuelensis causes cutaneous leishmaniasis. L. garnhami causes Venezuelan Andean cutaneous leishmaniasis.

30. Life cycle Transmitted by Lutzomiya sandfly Reservoir host: rodents, opossums, domestic dog, cat etc. Life cycle same with L. mexicana complex.

31. Life cycle

32. Vector: Lutzomiya sandfly

33. Transmission & Pathogenesis L. mexicana - produces a lesion known as chiclero ulcer or Bay sore – common among workers who collect chicle gum from the Chicazapote trees in the rain forest in Nicaragua, Guatemala, Belize and the Yucatan peninsula of Mexico. Clinical manifestation: a single cutaneous papule, nodule or ulcer located on the ear or face. Lesion generally heal spontaneously but may cause cartilage destruction and gross disfigurement.

34. L. pifanoi and L. amazonensis – produce a single but more likely to progress to the DCL – the majority patient infected, clustered in the Amazon river Basin of Brazil and Venezuela. Clinical presentation of DCL may be confused with Leptomatous Leprosy. L. garnhami and L. venezuelensis – assosiated with cutaneous leishmaniasis in rural parts of Venezuela – infection with either organism present with a solitary lesion that is usually self-limiting.

35. Leptomatous Leprosy Picture: Mildly elevated indurating nodules are seen on the face and extremities (gross findings).

37. Diagnosis Giemsa stained smears –amastigotes will be seen. Cultivation – promastigotes forms can be obtained. Immunological testing methods.

38. Treatment In most cases, the infections are self- limiting and require no treatment. Treatment is paramount if: -the lesion should endure or -threaten cartilaginous structures; ear,nose. Therapeutic agents: same as the treatment of Oriental sore.

39. Prevention Same as the prevention ways of Oriental sore. applied insect repellent to the skin and garments along with aerial spraying.

40. Leishmania braziliensis complex – Mucocutaneous Leishmaniasis L. braziliensis – Mexico to Argentina L. panamensis – Panama and Columbia L. peruviana – Peruvian Andes. L. guyanensis – Guiana and parts of Brazil and Venezuela.

41. Morphology Cause infections throughout the Americas from Mexico to Argentina. The distinguishing feature of these infectious is the development of ulcers on or about the oral and nasal mucosa L. braziliensis causes espundia. L. guyanensis causes pain bois. L. peruviana causes uta. All cause considerable morbidity and mortility in the endemic areas.

42. espundia

43. Life cycle Same with L. mexicana complex Vector: Lutzomyia and Psychodopygus sandflies.

44. Pathogenesis The primary lesion-same manner as the Oriental sore: macrophage ingest the parasites  become heavy laden with replicating amastigotes  tissue damage. Invade mucous membranes of the mouth and nasopharynx. Spread by: direct extension of the primary lesion or metastasis via the bloodstream or lymphatics. Progression of disease may take years.

45. Resulting disease: may produce ulcers that erode soft tissues of the face and palate or form polyp-like appendages in the nasal cavity. Patient commonly present with enlargement of the regional lymph nodes and secondary bacterial infections. Untreated  patients generally succumb to these secondary infection or to starvation if destruction of the oral cavity is extensive.

48. Diagnosis By demonstrating amastigotes of Leishmania in Giemsa stained smears or biopsy material from the edge of an active ulcer. Cultivation. Serologic test. Montenegro skin test.

49. Treatment Sodium antimony (Pentostom). Cycloguanil pamoate (Camolar). Amphotericin B (Fungizone). Meglumine antimonate GlucantimeKetoconazole

50. Prevention Personal protective measures such as :protecting clothing, insect repellents and etc. Vector control Reservoir host control Public health educational programs. Prompt treatment of infected individuals  to break the cycle of disease transmission.

51. Leishmania donovani complex – Visceral leishmaniasis L. donovani – India, Pakistan, Thailand, parts of Africa and the Peoples Republic of China. L. infantum – Mediterranean area, Europe, Africa, the Near East, and parts of the former Soviet Union. L. chagasi – Central and South America.

56. Morphology Visceral leishmanisis also known as Kala Azar or dum-dum fever. The most severe of the Leishmaniasis. Generally a disease of juveniles and young adults. Natural reservoir: rodents and dog. In India, man appears to be the only mamalian reservoir. L. donovani complex –parasitize the reticuloendothelial cells, viscerotropic, infected macrophages remaining fixed or disseminate throughout the body.

57. In the Mediterranean are, Europe, Africa, Soviet union – Phlebotomus sandfly remains the vector. Natural reservoir: domesticated dogs, canines and porcupines. In the New World (Central and South America) – Lutzomiya sandfly remains the vector. Natural reservoir: Foxes, domestic dogs and cats.

58. Life cycle Same with L. mexicana complex. The infected mononuclear phagocytes do not remain confined to the skin or mucous membranes. Parasitized macrophages are carried by the bloodstream to lymphoid tissue throughout the body especially to the spleen, liver and bone marrow. Amastigotes multiply in great numbers in this tissues. Vectors: Phlebotomus sandfly and Lutzomiya sandfly.

60. Transmission & pathogenesis Transmitted by sandflies. Incubation period: 3 weeks to 2 years. The infected macrophages migrate by lymphatic and hematogenous  spread to distant lymphoid tissues throughout the body. Transmission via blood transfusion is also possible.

61. Early symptoms: prodome of headache, malaise, fever, possible weight loss and abdominal pain. Fever may occur in periodic intervals mimicking tertian or quartan malaria. Another symptoms: diarrhea, consistent with typhoid fever, enlargement of liver and spleen (hepatosplendomegaly) and lymph nodes (lymphadenopathy), and increase in serum globulin levels. Microscopically, parasitized macrophages may be found in the tissue of the spleen, liver, heart, kidneys, lymph nodes, intestines and bone marrow.

62. Rapid proliferation of reticuloendothelial cells within the involved organs lead to organ hypertrophy. Parasitized macrophages may crowd out other hematopoietic cells leading to various degrees of anemia and leukopenia. The infiltration of the intestinal mucosa may result in ulceration and yield malabsorption, and wasting.

63. As the patient become more emaciated, the abdominal distention from the hepatosplenomegaly becomes more pronounced. A characteristic hyperpigmentation of the forehead and hands, known in India as Kala Azar, may also be observed. The prognosis for untreated cases is poor. Mortality rate cab be as high as 95%. In chronic cases, death ususally occurs from medical complications or bacterial infections within 2 years of diagnosis.

64. With treatment, the prognosis is usually much better. Recovering leads to a lasting immunity. In some patients, a condition called post- kala azar dermal leishmaniasis, or dermal leishmoid, may develop following the treatment with antimony compounds. The clinical presentation of this condition is marked by the appearance of either erythematous or hypopigmented lesions anywhere on the body.

65. A butterfly rash is a characteristic of systemic lupus erythemous. – may develop on the malar portion of the face. The dermal lesions, whether a papule or patch, may progress to nodules and resemble lepromatous leprosy. These lesions do contain viable parasites and can serve as a reservoir of infection.

68. Diagnosis Tissues Biopsy Direct examination of stained smears. Cultivation. Serologic test. Direct agglutination test (DAT). Complement fixation test (CF). Indirect fluorescence technique. Molecular diagnostic technique. Montenegro skin test (not reactive in people with active disease).

69. Treatment Pantavalent antimony sodium gluconate (Pentostom). Pentamidine isothionate (Lomidine) Amphotericin B Allopurinol + Pentostom Gamma interferon + Pentostom.

70. Prevention Same with the others leishmania sp.

71. Promastigotes (left) and amastigotes (right) of Leishmania parasites. Parasites have been fixed in Giemsa stained.