Appropriate utilization of the microbiology laboratory. 11 April 2013

Transcription

1 Appropriate utilization of the microbiology laboratory 11 April 2013

2 Lecture Plan Revision of infectious disease Triad of infectious disease Interaction between host and infectious agent Pathogenesis Phases of laboratory testing Conclusion

3 Introduction Infectious disease predominant cause of illness and death in many countries Improved sanitation Improved medical standards Poor socioeconomic conditions of majority quagmire of infectious disease Introduction of antimicrobials early 20 th century erroneously thought all infectious disease could be controlled / eradicated

4 Introduction (2) Negative consequences of progress Prolong human life Success premature survival Correct and incorrect use of antimicrobials Human incursion into environments not well adapted to Free movements of animals and plants across the globe Need to utilize the laboratory correctly understanding the inter connections between humans and infectious agents

5 Triad of infectious diseases infectious agent affected host environment

6 Interactions between hosts and infectious agents Commensalistic Infectious agent and host do not affect each other Mutualistic / Symbiotic Both host and infectious agent benefit from each other Saprophytic Infectious agent benefits from host but does no harm Parasitic Infectious agent benefits from host and causes harm Colonizing bacteria Exists on surface as commensal, saprophyte or parasite Opportunistic agents Potential pathogen causing disease in immuno compromised host

7 Relationships to hosts Relationship Infectious agent Example Free living Facultatively intracellular Obligatory intracellular most bacteria fungi parasites some bacteria some fungi mycobacteria some bacteria some parasites viruses S. aureus, E coli, K. pneumoniae Candida sp, Cryptococcus sp. Taenia sp, Trypanosoma sp. Legionella sp, Brucella sp Histoplasma sp Mycobacterium tuberculosis Rickettsia, Chlamydia sp Toxoplasma sp, Plasmodium sp Influenza virus, measles virus

8 Pathogenesis of infection Attachment to the surface epithelium fimbria, adhesins Multiply on the surface (respiratory, gastrointestinal tract, endocardium ) Express virulence factors exotoxins, endotoxins Invade the surface Dissemination to other organs

9 Classes of infectious agents Bacteria Gram positive bacilli Clostridium difficile Listeria monoytogenes Gram positive cocci Staphylococcus aureus Streptococcus pneumoniae Gram negative bacilli Klebsiella pneumoniae Escherichia coli Gram negative cocci Neisseria meningitidis

10 Classes of infectious agents Yeasts Candida albicans Cryptococcus neoformans Moulds Mucorales Aspergillus fumigatus Dermatophytes Microsporum canus Trychophyton rubrum Epidermophyton floccosum Dimorphic fungi Histoplasma capsulatum Sporothrix schenkii Pneumocystis jiroveci Fungi

11 Classes of infectious agents Intestinal protozoa Entamoeba histolytica Cryptosporidium parvum Nematodes Ascaris lumbricoides Enterobius vermicularis Strongyloides stercoralis Cestodes Taenia sp Trematodes Schistosoma Blood and tissue parasites Plasmodium sp Trypanosoma sp Parasites

12 Classes of infectious agents Viruses DNA viruses Herpes viruses herpes, cytomegalovirus, varicella zoster, Epstein Barr virus Parvoviruses Adenoviruses Hepadnaviruses hepatitis B RNA viruses Orthomyxoviruses influenza Paramyxoviruses measles, mumps, respiratory syncytial Retroviruses human immunodeficiency Reoviruses rotavirus Piconaviruses polio

13 Phases of laboratory testing Before analysis (Pre analytical) During analysis (Analytical) After analysis,before receipt of results (Post analytical )

14 Pre analytical Specimen collection Specimen transport Specimen reception Specimen rejection Specimen analysis

15 Specimen collection Understanding of how disease occurs and the likely organisms (pathogenesis) Collection from ideal site with least contamination Best timing to increase chances of recovery Sufficient quantity Appropriate collection device Container correctly labelled

16 Specimen transport Aim to maintain specimen to as near original as possible Appropriate collection device, sterile Transported in a safe way (Occupational Health and Safety Act) Transport as rapidly as possible Appropriate temperature, refrigeration Transport medium e.g. Stewart s transport medium

17 Specimen reception Safe environment, protective clothing Discard leaking containers Preliminary observation (sputum vs urine)

18 Rejection of specimens Criteria must be established and communicated Reason for rejection must be given Specimens received in formalin for culture Old (24hr) sputum specimen Specimen in improper container If specimen too little, incorrect, incorrectly transported communicate with clinician

19 Specimen analysis Visual observation Microscopic specimen analysis Culture and sensitivity Serological analysis Molecular diagnosis

20 Specimen analysis Microscopic examination (differential staining) Gram stain (bacteria, scoring of sputum Bartlett s) India ink (capsule e.g. cryptococcosis) Ziehl Neelson stain (tuberculosis, leprosy) Auramine stain(tuberculosis, leprosy) Geimsa stain (malaria, sleeping sickness) Fluorescent microscopy (parasites) Dark field microscopy (syphilis) Electron microscopy(viruses)

21 Specimen analysis Culture and susceptibility testing / sensitivity (utilization of media and inhibition by antibiotics susceptible to) Select primary culture media enriched/selective Isolate microorganism Identification and antimicrobial susceptibility testing (manual or automated) Interpretation of results

22 Specimen analysis Serological diagnoses (antigen and antibody reaction) Detection of antigen (organism) or antibody Visualise reaction charcoal, fluorescent and non fluorescent linked reagents (ELISA) Measure titres Prevalence of disease important for interpretation Increase sensitivity and specificity Rapid onsite, automated immunoassays, incubation

23 Specimen analysis Molecular diagnosis Identification of sequences in genetic material Amplification by PCR detects small numbers Identification of organism Identification of resistant mutations Rapid Sensitive and specific

24 Post analytical Review of results Quality confirmed Pre determine whether routine, urgent or panic values? Reflex testing Interpretation will be given Report sent to clinicians, hard copy or electronic Correct demographic data essential Ensure turn around time (TAT) achieved

25 Conclusion Importance of comprehensive clinical examination Understanding of infectious disease pathogenesis Understand health worker role in specimen analysis Available laboratory tests for presumptive and definitive diagnosis How you can use laboratory to ensure quality health care