blood protozoa

BLOOD PROTOZOABLOOD PROTOZOAII BPTII BPT

Dr Ekta ChourasiaDr Ekta Chourasia

MicrobiologyMicrobiology

12/03/0812/03/08 Dr Ekta, Microbiology, GMCADr Ekta, Microbiology, GMCA

Protozoa - Blood protozoaProtozoa - Blood protozoa

Plasmodium

Toxoplasma gondii

Trypanasoma

Leishmania

Babesia

Malaria

Babesiosis

Leishmaniasis (Visceral, Cutaneous)

African sleeping sickness, Chagas disease

Toxoplasmosis (congenital infections)


Genus PlasmodiumGenus Plasmodium

Consists of 4 species:Consists of 4 species:1.1. P. vivaxP. vivax

2.2. P. falciparumP. falciparum

3.3. P. malariaeP. malariae

4.4. P. ovaleP. ovale


Transmission & Life CycleTransmission & Life Cycle

Definitive host Female Anopheles mosquitoDefinitive host Female Anopheles mosquito

Intermediate host ManIntermediate host Man

Infective form SporozoitesInfective form Sporozoites

Portal of entry SkinPortal of entry Skin

Mode of transmission Bite of an infected mosquitoMode of transmission Bite of an infected mosquito

Site of localization First in liver cells & then in Site of localization First in liver cells & then in

RBCs RBCs


Morphological forms seen in HumansMorphological forms seen in Humans

In liver:In liver:1.1. SporozoitesSporozoites2.2. Pre erythrocytic schizontsPre erythrocytic schizonts

3.3. Merozoites – infect RBCsMerozoites – infect RBCs

In RBCs :In RBCs :1.1. Trophozoites – ring formTrophozoites – ring form2.2. Schizonts Schizonts 3.3. Merozoites – released by the rupture of schizonts Merozoites – released by the rupture of schizonts

– infect other RBCs– infect other RBCs4.4. Gametocytes – micro and macro gametocytesGametocytes – micro and macro gametocytes


Morphological forms seen in MosquitoMorphological forms seen in Mosquito

Further differentiation & development of Further differentiation & development of gametocytes take place in mosquitogametocytes take place in mosquito

1.1. Macro gametes (female gametes)Macro gametes (female gametes) – each – each macro gametocyte develops in to one macro macro gametocyte develops in to one macro gamete in the mid gut of mosquitogamete in the mid gut of mosquito

2.2. Micro gametes (male gametes)Micro gametes (male gametes) – one micro – one micro gametocyte produces 6 to 8 micro gametes gametocyte produces 6 to 8 micro gametes by exflagellation.by exflagellation.

3.3. Zygote Zygote – – OokineteOokinete – – OocystOocyst – rupture – – rupture – release of release of SporozoitesSporozoites – predilection to – predilection to salivary glands.salivary glands.


Incubation periodIncubation period

P. vivax P. vivax

P. ovale 10 to 14 daysP. ovale 10 to 14 days

P. falciparum P. falciparum

P. malariae 18 days to 6 P. malariae 18 days to 6

weeksweeks


Pathogenicity Pathogenicity Infection causes intermittent fever – MalariaInfection causes intermittent fever – Malaria

Each of the 4 species causes a characteristic Each of the 4 species causes a characteristic fever:fever:

P. vivax Benign tertian/ vivax malariaP. vivax Benign tertian/ vivax malaria

P. falciparum Malignant tertian/ falciparumP. falciparum Malignant tertian/ falciparum

malaria, black water fevermalaria, black water fever

P. malariae Quartan malariaP. malariae Quartan malaria

P. ovale Ovale malaria P. ovale Ovale malaria


Clinical FeaturesClinical FeaturesSeries of febrile paroxysmsSeries of febrile paroxysms – fever is caused – fever is caused by the release of merozoites & toxins from by the release of merozoites & toxins from ruptured erythrocytic schizont which in turn ruptured erythrocytic schizont which in turn causes the release of cytokines.causes the release of cytokines.

Quartan malaria – every 72 hrsQuartan malaria – every 72 hrs Tertian malaria - every 48 hrs Tertian malaria - every 48 hrs

* each paroxysm has 3 stages - * each paroxysm has 3 stages - cold stagecold stage (rigors), (rigors), hot stagehot stage (high temp., body & joint (high temp., body & joint pains, vomiting & diarrhoea) and pains, vomiting & diarrhoea) and perspiration perspiration stagestage (fall in temp.) (fall in temp.)


Clinical FeaturesClinical Features

AnaemiaAnaemia – due to breakdown of RBCs, – due to breakdown of RBCs, particularly occurs in falciparum malariaparticularly occurs in falciparum malaria

Splenomegaly Splenomegaly – all forms – all forms


Falciparum malariaFalciparum malariaSevere falciparum malaria is associated Severe falciparum malaria is associated withwith

1.1. Pernicious malaria /cerebral malariaPernicious malaria /cerebral malaria

2.2. Blackwater feverBlackwater fever

3.3. AnaemiaAnaemia

4.4. HypoglycaemiaHypoglycaemia

5.5. Complications in pregnancyComplications in pregnancy


Malaria caused by P.vivax, Malaria caused by P.vivax, P.ovale & P.malariaeP.ovale & P.malariae

Rarely life threateningRarely life threatening Relapses/ recurrences are a featureRelapses/ recurrences are a feature

Recurrences in MalariaRecurrences in Malaria May result from – reinfection or May result from – reinfection or

- due to persistence of infection - due to persistence of infection

- Occurs due to a special form - Occurs due to a special form of of parasites called parasites called hypnozoiteshypnozoites. .


Laboratory diagnosis of MalariaLaboratory diagnosis of Malaria

Specimen: peripheral blood smears

Leishman or Giemsa stain

Trophozoites (ring forms) or gametocytes are seen within RBCs

thick and thin blood smears

Quantitative Buffy Coat (QBC) examination

P falciparum antigen detection (ELISA)


Making of Thin & Thick filmsMaking of Thin & Thick films


Rapid Diagnostic testsRapid Diagnostic tests

HRP2 testsHRP2 tests detection of detection of P.falciparumP.falciparum Two types of test – ParaSight FTwo types of test – ParaSight F - ICT Malaria Pf- ICT Malaria Pf

pLDH test e.g. OptiMAL testpLDH test e.g. OptiMAL test Detection of Detection of P.falciparumP.falciparum & & P.vivaxP.vivax Produced by all human malarial parasitesProduced by all human malarial parasites Differentiation of species is based on antigenic Differentiation of species is based on antigenic

differences between pLDH isoforms. differences between pLDH isoforms.


Optimal test

ParaSightF test

ICT Malaria Pf / Pv


Toxoplasma gondiiToxoplasma gondii

Infective form Matured oocyst / tissue cyst / tachyzoites

Mode of transmission

Ingestion / intrauterine / blood transfusion/ improper handling of cat litter

Site of localization Any organs (RE system)

Definitive host Cat

Intermediate host Man, sheep, birds

Eye & Brain


Laboratory Diagnosis –T gondiiLaboratory Diagnosis –T gondii

Specimens Impression smear of LN, bone marrow, spleen, brain biopsy, blood, sputum, CSF

Microscopy Giemsa stain, tachyzoites or cysts

Serology ELISA / IFA – 16 fold rise in Ab titre: Acute infection

Sabin-Feldman dye test: inhibition by antibody of the staining of tachyzoites by alkaline methylene blue

Prenatal diagnosis Fetal blood for IgM Ab / PCR


Haemoflagellates Haemoflagellates


TrypanosomesTrypanosomes

T. brucei subspecies gambiense (Chronic)

T. brucei subspecies rhodesiense (Acute)

West African sleeping sickness

East African sleeping sickness

T. cruzi (acute and chronic) American trypanosomiasis Chagas disease


T. bruceiT. brucei

Definitive host

Intermediate host

Man

Tsetse fly

Infective form Metacyclic Trypomastigote

Mode of transmission Bite of infected tsetse fly

Site of localization CNS


Clinical features Sleeping SicknessClinical features Sleeping Sickness

Trypanosoma chancre at the site of bite

Winter bottom’s sign: prominent cervical lymphadenopathy

Meningoencephalitis - Apathetic, confused, comatose


T. cruziT. cruzi

Definitive host

Intermediate host

Man

Reduviid bug

Infective form Metacyclic Trypomastigote

Mode of transmission Feces of bug rubbed into site of bite / contamination of conjunctiva or other exposed mucous membranes with fingers

Site of localization Autonomous nervous system of heart / GIT

Infect cardiac, smooth and skeletal muscle, reticuloendothelial cells and neuroglial cells


Clinical featuresClinical features

Chagoma chancre at the site of biteChagoma chancre at the site of bite

Acute Chagas diseaseAcute Chagas disease Romana’s sign: unilateral edema of face with

conjunctivitis and swelling of upper & lower eyelids Fever, splenomegaly, anasarca, meningoencephalitis

Chronic Chagas disease Cardiomyopathy, AV block, CCF Megaesophagus / Megacolon


Laboratory Diagnosis – TrypanosomiasisLaboratory Diagnosis – Trypanosomiasis

Specimens Blood, CSF, Aspirates (LN)

Microscopy Trypomastigotes in blood


LeishmaniaLeishmania

Definitive host

Intermediate host

Man

Sand fly (Phlebotomus)

Infective form Promastigote

Mode of transmission Bite of infected sand fly

Site of localization Reticuloendothelial system


LeishmaniaLeishmania

Three major species:Three major species:1.1. L. donovaniL. donovani – kala azar/ visceral leishmaniasis – kala azar/ visceral leishmaniasis

2.2. L. majorL. major – cutaneous leishmaniasis – cutaneous leishmaniasis

3.3. L. braziliensisL. braziliensis – mucocutaneous leishmaniasis – mucocutaneous leishmaniasis


Clinical TypesClinical Types

Visceral leishmaniasis -Visceral leishmaniasis - fever, malaise, weight loss, anaemia and swelling of the spleen, liver, and lymph nodes

Cutaneous leishmaniasis - causes 1-200 simple skin lesions which self-heal within a few months but which leave unsightly scars

Mucocutaneous leishmaniasis - infection begins with skin ulcers which spread, causing dreadful and massive tissue destruction, especially of the nose and mouth


Laboratory DiagnosisLaboratory Diagnosis

1.1. Demonstration of parasite in clinical specimen:Demonstration of parasite in clinical specimen: MicroscopyMicroscopy CultureCulture Animal inoculationAnimal inoculation

2.2. Demonstartion of antibodies usingDemonstartion of antibodies using Specific leishmanial Ag – ELISA / IFA / AgglutinationSpecific leishmanial Ag – ELISA / IFA / Agglutination Non-specific Ag – CFTNon-specific Ag – CFT

3.3. Non-specific serum testsNon-specific serum tests Aldehyde test (Napier’s)Aldehyde test (Napier’s) Chopra’s Antimony testChopra’s Antimony test

4.4. Absence of hypersensitivity to leishmanial AgAbsence of hypersensitivity to leishmanial Ag5.5. Contributory lab findingsContributory lab findings – anemia, leucopenia, neutropenia – anemia, leucopenia, neutropenia


1. Demonstration of parasite in Clinical 1. Demonstration of parasite in Clinical SpecimenSpecimen

Clinical specimens:Clinical specimens: Peripheral bloodPeripheral blood Bone marrow aspirateBone marrow aspirate Spleen aspirateSpleen aspirate Lymph node aspirateLymph node aspirate

MicroscopyMicroscopy: Leishman, Giemsa or Wright’s : Leishman, Giemsa or Wright’s stain - Amastigotes within macropahgesstain - Amastigotes within macropahges

Culture:Culture: NNN (Novy, MacNeal, Nicolle) medium for 7 NNN (Novy, MacNeal, Nicolle) medium for 7 days – promastigote form days – promastigote form

Animal inoculation:Animal inoculation: Hamster - Animal kept at 23-26°C Hamster - Animal kept at 23-26°C


2. Absence of Hypersensitivity to 2. Absence of Hypersensitivity to Leishmanial AgLeishmanial Ag

Montenigro test – 0.1 ml of killed promastigote Montenigro test – 0.1 ml of killed promastigote Ag I.D. Result read after 72 hrs.Ag I.D. Result read after 72 hrs. Positive in Dermal leishmaniasis & recovered cases Positive in Dermal leishmaniasis & recovered cases

of Kala Azar.of Kala Azar.

Negative in active cases of Kala Azar.Negative in active cases of Kala Azar.

blood protozoa

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