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2.laboratory-diagnosis-of-parasitic-infections.ppt
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- 2. Case diagnosis History(Age, occupation, residency, previous infection) Complaint Clinical examination Invesigations - Laboratory investigations - Radiology - Surgical intervention (Exploratory) Provisional diagnosis Confirm the diagnosis
- 3. DIAGNOSIS DIRECT INDIRECT MOLECULAR Urine Stool Sputum Biopsy Blood Aspirates PCR DNAprobes IHAT LAT IFAT ELISA CFT DEIDT
- 4. URINE EXAMINATION MACROSCOPIC MICROSCOPIC colour whitesmoky Sedimentation concentration Membrane filtration Chyluria Blood Filaria S. haematobium Acetic acid RBC haemolysis Clear ova Ether Dissolve fat M.f
- 5. URINE EXAMINATION SEDIMENTATION CONCENTRATION 15-20min Centrifuge (2 min) Clean conical glass receptacle
- 6. URINE EXAMINATION Membrane filtrationtechnique air 10 ml urine Nucleopore filter Eggs of Schistosoma + Saline
- 7. URINE EXAMINATION HELMINTHES PROTOZOAARTHROPODES • S. haem.egg • E. vermic. egg • S. mansoni egg • Mf (Ov, Wb) • H sand • T. Vag troph • Pthirus pubis • L. higher deptera
- 8. URINE EXAMINATION Egg viability Liveeggs Dead eggs •Well defined miracidium •Flickering F cells •Hatching moving miracidium •Dark colour •Granulated
- 9. STOOL EXAMINATION MACROSCOPIC MICROSCOPICOTHERS •Consistency •Colour •Composition •Culture •Cellophane tape •Baeremann tech. •Ova quantitaion (Stoll & Kato) Temprory Permanent Diect saline smear Iodine smear Concentration techniques Sedimentation Floatation Saline Formol ether Sat saline Zinc sulphate Sheather’s sugar
- 10. MACROSCOPIC EXAMINATION COLOUR CONSISTENCYCOMPOSITION Adult PARASITES Pale=Steatorrhea (G.l) -Liquid (Troph) -Formed (Cyst) -Semi formed (Cyst) ?? Blood ?? Mucus (dysentry) *Ascaris worm *E. vermicularis *T. saginata STOOL EXAMINATION
- 11. STOOL EXAMINATION MACROSCOPIC MICROSCOPICOTHERS •Consistency •Colour •Composition •Culture •Cellophane tape •Baeremann tech. •Ova quantitaion •(Stoll & Kato) Temprory Permanent Diect saline smear Iodine smear Concentration techniques Sedimentation Floatation Saline Formol ether Sat saline Zinc sulphate Sheather’s sugar
- 12. STOOL EXAMINATION Temporary Saline smearIodine smear saline Iodine 1% Huge number of: •Eggs • Protozoal troph. Motility (Amoeb, flagellates) Huge number of: •Cyst morphological details
- 13. STOOL EXAMINATION Scanty infection Concentrationtechniques Sedimentation Floatation • Heavy eggs (Ascaris egg) • Operculated eggs (Trematodes) • Larvae (Strong sterc.) • Cysts • Non Operculated eggs Trematodes ( S. m.) Cestode Nematode(Hookworms,Trichostong) • Cysts
- 14. STOOL EXAMINATION MACROSCOPIC MICROSCOPICOTHERS •Consistency •Colour •Composition •Culture •Cellophane tape •Baeremann tech. •Ova quantitaion •(Stoll & Kato) Temprory Permanent Diect saline smear Iodine smear Concentration techniques Sedimentation Floatation Saline Formol ether Sat saline Zinc sulphate Sheather’s sugar
- 15. STOOL EXAMINATION Saline sedimentation 10g stool Saline Mesh wire gauze Conical flask Sediment Emulsify
- 16. STOOL EXAMINATION Formol EtherSed. Conc. 10% Formalin 1 g stool Sediment formalin debris Ether Thorough mixing Ether • Ether adsorbs fecal debris & floats. • Formalin fixes & preserves the specimen. Conical flask centrif. tube
- 17. STOOL EXAMINATION MACROSCOPIC MICROSCOPICOTHERS •Consistency •Colour •Composition •Culture •Cellophane tape •Baeremann tech. •Ova quantitaion •(Stoll & Kato) Temprory Permanent Diect saline smear Iodine smear Concentration techniques Sedimentation Floatation Saline Formol ether Sat saline Zinc sulphate Sheather’s sugar
- 18. STOOL EXAMINATION Floatation concentration Satsaline Zn sulphate Sheather’s sugar • Cestode eggs (non op) •Nematode eggs????? •Hookworms??????? •Trichostong؟؟؟؟؟؟؟؟؟؟؟ •Egg of S.m. •Eggs of small tapeworms •Cysts • Crypto, Iso. oocysts Tin container 20 min Centrif. 2 min Seive Clean light eggs & cysts
- 19. STOOL EXAMINATION MACROSCOPIC MICROSCOPICOTHERS •Consistency •Colour •Composition •Culture •Cellophane tape •Baeremann tech. •Ova quantitaion •(Stoll & Kato) Temprory Permanent Diect saline smear Iodine smear Concentration techniques Sedimentation Floatation Saline Formol ether Sat saline Zinc sulphate Sheather’s sugar
- 20. STOOL EXAMINATION Permanent Stainedsmears Iron haematoxylin stain Trichrome stain Modified Ziehl Neelsen stain (Crypto.)
- 21. STOOL EXAMINATION MACROSCOPIC MICROSCOPICOTHERS •Consistency •Colour •Composition • Cellophane tape • Culture •Baeremann tech. •Ova quantitaion •(Stoll & Kato) Temprory Permanent Diect saline smear Iodine smear Concentration techniques Sedimentation Floatation Saline Formol ether Sat saline Zinc sulphate Sheather’s sugar
- 22. STOOL EXAMINATION Kato technique Meshscreen Template Hole Remove the template Cellophane soaked by glycerin (clears faeces( Egg count/ g stool Egg quant. Of: Ascaris, T. trich., Hookworms, S. mansoni
- 23. STOOL EXAMINATION Stoll’s technique NaOH 4g Stool Erlynmeyer flask 56 CC 60 CC Shake well 0.15 CC Egg count/ slide Eggs/1g= Eggs/slideX100 Egg/day=Eggs/1g X stool wt/g in 24 hrs 24 hr stool Egg quant. Of: Ascaris, T. trich., Hookworms, S. mansoni
- 24. STOOL EXAMINATION Baermann’s technique Warmwater Stool/soil seive Glass funnel clamp 30 min 25-50 CC centrifuge Detec. Of Nematode L. /stool, soil
- 25. STOOL EXAMINATION Cultures forNematode larvae Filter paper culture Scanty infection Larvae of: • St. stercoralis (A,L) • Hookworms • Trichostrong Water Sealed petri dish Filter paper Slide
- 26. DIAGNOSIS DIRECT INDIRECT MOLECULAR Urine Stool Sputum Biopsy Aspirates Blood PCR DNAprobes IHAT LAT IFAT ELISA CFT DEIDT
- 27. SPUTUM EXAMINATION MACROSCOPIC MICROSCOPIC AppearanceConcentration Bloody (Parag) Rusty brown (Parag) NaOH Sputum Centfifuge
- 28. Parasites/sputum P living inlung P migrating in lung P resulting from rupture of • P. westermani eggs • St. stercoralis • Ascaris • Hookworm (filariform L) • Hydatid cyst (sand) • Amoebic abcess (troph)
- 29. BIOPSY SPECIMEN SKIN SNIPMUSCLE BIOPSY RECTAL BIOPSY O. Volvulus mf T. Spiralis larvae Schistosoma egg • Raise skin by needle • Slice by scissors • Put snip in normal saline • Examine Muscle digestion with HCl + pepsin
- 30. ASPIRATES EXAMINATION CSF Duodenalaspirates BM aspirates Cutanoeus ucler • Afr. Tryp. (trypom) • FLA (troph) • M.f. of loa loa • L. of T spiralis • G. lamb troph • Crypto oocyst • St sterc. Rh L. • Fasciola eggs • L. donovani a,ast. •T. cruzi amast. •P. falciparum. •Leishmaniasis D. intubation D capsule (Enterotest) Direct stain (amast NNN (promast) •Lumbar puncutre • Centrifuge •Examine sed. floor Edge
- 31. DIAGNOSIS DIRECT INDIRECT MOLECULAR Urine Stool Sputum Biopsy Aspirates Blood PCR DNAprobes IHAT LAT IFAT ELISA CFT DEIDT
- 32. BLOOD EXAMINATION Blood filmsBuffy coat films QBC technique Knott’s conc. tech. Thin Thick
- 33. BLOOD EXAMINATION BLOOD FILMS Thin Thick Bld drop spread Air dry methyl alcohol Geimsa Air dry Geimsa Circular motion Malaria, Babesia, Filaria, Tryp.
- 34. BLOOD EXAMINATION Buffy coatfilm centrifuge RBC WBC (BC) plasma Citrated bld 30 min Air dry Fix spread Geimsa Tryp., L. donovani
- 35. BLOOD EXAMINATION QBC technique centrifuge RBC RBC+parasite Microhaematocrit tube Acridine orange Malaria, Filaria, Trypanosomes
- 36. BLOOD EXAMINATION KNOTT’S CONC.TECHNIQUE 10 ml 1 ml Air dry fix Geimsa Citrated bld Formalin 2 % sediment 2 min centrifuge Filaria
- 37. INDIRECT IMMUNOLOGICAL METHODS Scantyinfection. Tissue parasite no portal of exit (Hydatid dis.) Migratory stage (Fasciola) Chronic infection fibrosis (Bilharziasis)
- 38. INDIRECT IMMUNOLOGICAL METHODS Antigendetection Antibody detection • More specific • More accurate. • Active infection • Early • Quantitative Ab remain in serum for months even after cure
- 39. INDIRECT IMMUNOLOGICAL METHODS IHATLAT + Sensitized Sheep’s RBC (O–ve) Ag Patient’s serum (?? AB) Agglutination + Agglutination Ag Latex particle Patient’s serum (?? AB)
- 40. INDIRECT IMMUNOLOGICAL METHODS INDIRECTFLUORESCENT ANTIBODY TEST parasite Patient’s serum (?? AB) Anti human AB fluorescein
- 41. INDIRECT IMMUNOLOGICAL METHODS ELISA OPD OPD Flatbottom plastic micrititre plate Ag Patient’s serum (?? AB) Anti human AB Peroxidase E AB
- 42. INDIRECT IMMUNOLOGICAL METHODS CFT Ag Patient’sserum (?? AB) complement Anti sheep AB Sheep’s RBC Tube / microplate AB
- 43. INDIRECT IMMUNOLOGICAL METHODS DoubleElectro Immuno Diffusion Ag Ab Buffered gel Electric current Line of ppt
- 44. INDIRECT IMMUNOLOGICAL METHODS ImmunodiagnosticStrip Test (Dip Stick Test) Ag Nitrocellulose strip Monoclonal Ab Coloured dye Pt bld (?Ag) Malaria, Filaria, African tryp.
- 45. DIAGNOSIS DIRECT INDIRECT MOLECULAR Urine Stool Sputum Biopsy Blood Aspirates PCR DNAprobes IHAT LAT IFAT ELISA CFT DEIDT
- 46. MOLECULAR BIOLOGICAL TECHNIQUES DNAProbes DNA Probe Commercially prepared DNA sequence Radio active material Nitrocellulose paper Sample (Serum/ stool) ?? parasite +ve parasite Hybridization Radioactivity
- 47. MOLECULAR BIOLOGICAL TECHNIQUES PolymeraseChain Reaction (PCR) Single stranded DNA Replication Detection T cruzi, T gondii