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)
12/03/0812/03/08 Dr Ekta, Microbiology, GMCADr Ekta, Microbiology, GMCA
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
12/03/0812/03/08 Dr Ekta, Microbiology, GMCADr Ekta, Microbiology, GMCA
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
12/03/0812/03/08 Dr Ekta, Microbiology, GMCADr Ekta, Microbiology, GMCA
12/03/0812/03/08 Dr Ekta, Microbiology, GMCADr Ekta, Microbiology, GMCA
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
12/03/0812/03/08 Dr Ekta, Microbiology, GMCADr Ekta, Microbiology, GMCA
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.
12/03/0812/03/08 Dr Ekta, Microbiology, GMCADr Ekta, Microbiology, GMCA
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
12/03/0812/03/08 Dr Ekta, Microbiology, GMCADr Ekta, Microbiology, GMCA
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
12/03/0812/03/08 Dr Ekta, Microbiology, GMCADr Ekta, Microbiology, GMCA
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.)
12/03/0812/03/08 Dr Ekta, Microbiology, GMCADr Ekta, Microbiology, GMCA
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
12/03/0812/03/08 Dr Ekta, Microbiology, GMCADr Ekta, Microbiology, GMCA
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
12/03/0812/03/08 Dr Ekta, Microbiology, GMCADr Ekta, Microbiology, GMCA
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. .
12/03/0812/03/08 Dr Ekta, Microbiology, GMCADr Ekta, Microbiology, GMCA
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)
12/03/0812/03/08 Dr Ekta, Microbiology, GMCADr Ekta, Microbiology, GMCA
Making of Thin & Thick filmsMaking of Thin & Thick films
12/03/0812/03/08 Dr Ekta, Microbiology, GMCADr Ekta, Microbiology, GMCA
12/03/0812/03/08 Dr Ekta, Microbiology, GMCADr Ekta, Microbiology, GMCA
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.
12/03/0812/03/08 Dr Ekta, Microbiology, GMCADr Ekta, Microbiology, GMCA
Optimal test
ParaSightF test
ICT Malaria Pf / Pv
12/03/0812/03/08 Dr Ekta, Microbiology, GMCADr Ekta, Microbiology, GMCA
12/03/0812/03/08 Dr Ekta, Microbiology, GMCADr Ekta, Microbiology, GMCA
12/03/0812/03/08 Dr Ekta, Microbiology, GMCADr Ekta, Microbiology, GMCA
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
12/03/0812/03/08 Dr Ekta, Microbiology, GMCADr Ekta, Microbiology, GMCA
12/03/0812/03/08 Dr Ekta, Microbiology, GMCADr Ekta, Microbiology, GMCA
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
12/03/0812/03/08 Dr Ekta, Microbiology, GMCADr Ekta, Microbiology, GMCA
Haemoflagellates Haemoflagellates
12/03/0812/03/08 Dr Ekta, Microbiology, GMCADr Ekta, Microbiology, GMCA
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
12/03/0812/03/08 Dr Ekta, Microbiology, GMCADr Ekta, Microbiology, GMCA
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
12/03/0812/03/08 Dr Ekta, Microbiology, GMCADr Ekta, Microbiology, GMCA
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
12/03/0812/03/08 Dr Ekta, Microbiology, GMCADr Ekta, Microbiology, GMCA
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
12/03/0812/03/08 Dr Ekta, Microbiology, GMCADr Ekta, Microbiology, GMCA
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
12/03/0812/03/08 Dr Ekta, Microbiology, GMCADr Ekta, Microbiology, GMCA
Laboratory Diagnosis – TrypanosomiasisLaboratory Diagnosis – Trypanosomiasis
Specimens Blood, CSF, Aspirates (LN)
Microscopy Trypomastigotes in blood
12/03/0812/03/08 Dr Ekta, Microbiology, GMCADr Ekta, Microbiology, GMCA
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
12/03/0812/03/08 Dr Ekta, Microbiology, GMCADr Ekta, Microbiology, GMCA
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
12/03/0812/03/08 Dr Ekta, Microbiology, GMCADr Ekta, Microbiology, GMCA
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
12/03/0812/03/08 Dr Ekta, Microbiology, GMCADr Ekta, Microbiology, GMCA
12/03/0812/03/08 Dr Ekta, Microbiology, GMCADr Ekta, Microbiology, GMCA
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
12/03/0812/03/08 Dr Ekta, Microbiology, GMCADr Ekta, Microbiology, GMCA
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
12/03/0812/03/08 Dr Ekta, Microbiology, GMCADr Ekta, Microbiology, GMCA
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.