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Lab diagnosis of bacterial infections


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  • 1.  Advantages in making a specific diagnosesbetter patient careappropriate antibioticsparing of expensespreventive measures can be initiated
  • 2. Specimen collection &transport Most important Protect from contamination Selected media
  • 3. Approch for identification ofinfectious agents5 different approaches detection of antibodies- identify NA using dna probes
  • 4.  Gram stain: pneumococci, gonococci,staphylococci,meningococci Acid fast stain: mycobacteriumtuberculosis,leprae, nocordia, cryptosporidia
  • 5.  Other new stainsAura mine rhodamine for mycobacteriaWright stains – malarial,& microfilarialIndian ink – capsular organismsSilver stains- fungi & pneumocystis
  • 6.  Immune electron microscopy Direct detection of antigen Non cultivables like Rota virus, hepatitis A&NORWALK VIRUS
  • 7.  In bacteremiaacute febrile illnesstyphoid,meningitis,pneumonia,osteomylitis,PUO 1% total blood volume At onset of chills
  • 8.  Sputum cultureEarly morning samples(pooled overnight secretions likely tocontain pathogenic organisms)Heated or ultrasonic nebulised saline inducesputum production
  • 9.  mid stream sample Reach lab by ½ hr Presence of more than 10000 colonies/mlsignifies true infection
  • 10.  Gastric aspirates- TB Endoscopic biopsies-helicobacter pylori Stool samples
  • 11.  BACTEC TB-460 : rapid,specific,&rapidculture method Both respiratory & non respiratory specimens 13-14 days 200 viable bacilli 14-17 days 20 viable bacilli
  • 12.  Mycobacterial growth indicator tube(MGIT)960 Art fluorescent technology Rapidly 7-10 daysBased on O2 quenching of mycobacteria withfluorescent dye
  • 13.  Uses specific Mycobacterial phages to detectviable bacteria within 48 hrs
  • 14.  Serology helps in detecting either thespecific or non specific immune responsesof the host or the presence of the antigen inthe host
  • 15. Specific Ig G &Ig M can be detected usingimmunologic techniques against virusesVarious types includeprecipitationAgglutination widal testComplement fixationimmunofluorescenceElisaWestern blot
  • 16. Specific,sensitive,simple,inexpensible,& reproducibleUsed extensively to detect either Ag or AbAlso detects small quantities of AgUsed to diagnose TORCH,HIV,MEASLES,HEPATITIS(A,B,C,D,E)…….
  • 17. Tag Ab with fluorescein isothiocynateAb-virus complex or viralantigensmicroscopically by UV illuminationDetects viruses which are uncultivable
  • 18. Ω Assays are available for rapid detection ofbacterial Ags in various body fluidsΩ Useful when prior antibiotic therapy hasbeen initiated and cultures are negative after24 hrs of incubation
  • 19. Detection of microbialantigens/products The following fluid/samples can be assayed CSF: Latex agglutination & counter-currentimmuno electrophoresis are used todemonstrate the soluble polysaccharide Agof cryptococcus &….
  • 20. Detection of microbialantigens/products Serum- Pl.falciparum/vivax are detected atlevels of 100-200 parasite/űl Urine-strep.pneumonia legionella, Stool :helicobacter pylori,clostridiumdifficile,giardia
  • 21. Detection of microbialantigens/products GLC : anaerobes HPLC : mycobacteria
  • 22.  New developments Answer for organisms with growthcharacteristicsslow – mycobacteriadifficult – viruses,chlamydiafastidious- mycoplasma
  • 23. Highly sensitive – detects low pathogen no’sas in meningitisUsed to monitor response to treatment(viral load assays for hepatitis B,C&HIV)
  • 24. Nucleic acid probes :NA hybridization is powerful & widely usedtechniqueUsed to detect and locate specificDNA&RNA sequence in tissues orchromosomes by making use ofradioactive/fluorescent labelled DNA/RNAprobes complementary to the requiredsequence
  • 25. Commercially available for the available foridentification of M.tuberculosiscomplex,avium,intracellulaire,kansasi,gordonae
  • 26. Nucleic acid amplication3 typestarget amplificationsignal amplificationprobe amplification
  • 27. Nucleic acid amplication- Target amplificationPCR : An in vitro method for amplifyingspecific DNA sequenceextremely minute amounts of NAgenerates billions of exact copiesmaking genetic analysis a simpleprocess
  • 28. Target DNA acts as templatenucleotidesprimers&DNA polymerasegenerates copies by alternate heating &cooling for denturation, annealing, &extension
  • 29. Nucleic acid amplication- Target amplificationReal time PCR : rapida fluorescent signal is used forreal time monitoring of amplificationTranscription mediated amplification Uses ribosomal rna as the target for reversetranscriptase
  • 30. Nucleic acid amplication-signalamplification Increases the signal generated fromhybridized probe moleculeProbe amplificationEnd product is an amplified version of originalproduct includes LCR, cycling probetechnologyIn LCR phenotypic change in the organism suchas virulence or drug resistance
  • 31. Disadvantages of molecularmethodsAmplification can amplify even minutequantities of contaminating DNA – false (+)Do not differentiate dead from livingorganismsFalse(-) results may due to low copy no’s ofmicroorganisims at site of infection