Bacteriology Chapter24

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					Chapter 24
Clinical Microbiology and
               Growth-Dep. Clinical
               Diagnostic Methods
   Isolation of pathogens from clinical specimens: infected
    tissues or fluids are collected, ex. blood, urine, feces, sputum,
    cerebrospinal fluid, pus.
   What steps should be taken to properly obtain clinical
   General purpose media: ex. blood agar, supports the growth
    of most aerobic and facultatively aerobic organisms.
   Enrichment media: the addition of growth factors allows
    metabolically fastidious to grow, ex. Neisseria gonorrhoeae.
   Selective media: enhance the growth of certain organisms
    while retarding the growth of others.
   Differential media: specialized media that allow ID of
    organisms based on their appearance on the media.
               Blood Cultures
   Bacteremia: presence of bacteria in the
    blood, uncommon in healthy individuals.
   Septicemia: blood infection resulting from
    the growth of a virulent organism entering
    the blood from a focus of infection, results
    in fever and chills and possibly septic
    shock including organ failure.
                Urine Cultures
   Common, causal agents often identical to
    normal flora, ex. E. coli. Enteric bacteria
    account for 90% of UTIs.
   Significant UTI = 105+ organisms per ml of
    clean-catch specimen.
   Dipstick tests test for nitrate production,
    etc. related to large #’s of organisms in the
    urinary tract.
               Fecal Cultures
   Samples must be collected and preserved
    properly and processed ASAP.
   Processing includes inoculation into a
    variety of selective media for isolation of
    individual bacteria.
   Intestinal parasites are ID’ed
    microscopically rather than by culture.
        Wounds and Abscesses
   Types of wounds include animal or human
    bites, burns, cuts, penetration of foreign
   Wounds must be carefully sampled and
    results carefully processed since normal
    flora are likely to be present.
             Other Specimens
   Genital specimens (ex. for gonorrhea) and
    anaerobic specimens must be specially
    processed in order to preserve the causal
    agents for proper diagnosis.
          Growth on Selective and
             Differential Media
   Unknown pathogens are first cultured on primary
    isolation media and then subcultured onto selective and
    differential media for further diagnosis.
   Eosin-methylene blue (EMB) = selective (for gram neg.
    bacteria) and differential (for diff. genera of gram neg.
   Biochemical tests measure the presence or absence of
    enzymes involved in the catabolism of substrate(s) in the
    differential medium.
   100’s of tests have been developed for clinical use,
    though only ~20 are used routinely. (see Table 24.3)
    Some commercial kits contain a battery of tests that ID
    and individual species, ex. S. aureus, M. tuberculosis.
Testing Cultures for Antimicrobial
        Drug Sensitivity
   Minimum Inhibitory Conc. (MIC) – broth
    (or antibiotic) dilution assay  highest
    dilution (lowest conc.) of antibiotic that
    completely inhibits growth.
   Kirby-Bauer disk diffusion method (see
    Fig. 24.8).
          Safety in the Clinical Lab
   1. Access to lab restricted.
   2. Decontamination procedures.
   3. Personnel must be adequately vaccinated.
   4. All clinical specimens should be considered
   5. No mouth pipetting.
   6. Animals handled only by trained lab personnel.
   7. Lab personnel must wear appropriate safety
    equipment: lab coat, sealed shoes, rubber gloves,
    masks, eye protection, etc.
   8. All clinical specimens should be treated as if they
    contain HIV.
                 Immunoassays for
                 Infectious Disease
   Immunoassays detect and measure specific immune
    responses to unique molecular portions of pathogens, and
    are used especially on those pathogens (ex. viruses) that are
    difficult to grow.
   New pathogen first infects individual  phagocytized 
    phagocytes present Ag from pathogen to T helper cells  T
    helper cells recruit and stimulate other phagocytes and B cells
     B cells produce Ab (primary Ab response), occurs within 5
    days, but Ab do not reach peak quantities for several weeks.
   After 1st Ag exposure specific immunity via T and B cells
    develops. After 2nd Ag exposure, stimulation of these specific
    immune cells causes a rapid and strong immune response,
    which quickly destroys the pathogen. Thus, the immune
    system has memory, which is characterized by and increase
    in Ab titer (used to track infections).
     Ab Titers, Skin Tests,
& Diagnosis of Infectious Disease
   Serological tests are set up by making dilutions of patient
    serum and determining the highest dilution at which the Ag-
    Ab rxn. occurs.
   2 situations: 1. Must see a rise in Ab titer to conclude active
    infection (having Ab present could mean prior infection or
    immunization and not current infection). 2. The mere
    presence of Ab is sufficient to indicate infection, ex. HIV.
   Not all infections result in production of systemic Ab, ex.
    Neisseria gonorrhoeae.
   Skin testing: ex. tuberculin text = intradermal injection of a
    soluble extract of Mycobacterium tuberculosis  pos.
    inflammatory rxn. at site of injection = current infection or
    previous exposure to M. tuberculosis.
     Polyclonal and Monoclonal Ab
   Antiserum (serum containing Ab) contains a mixture of
    different Ab directed at the numerous determinants on
    an Ag. This mixture of serum Ab is called polyclonal Ab
    and, though they protect the host, they do not provide
    reproducible results for use in clinical diagnostic testing.
   Only a few Ab are directed toward each Ag determinant.
    Ab produced by a single B cell (or in vitro clones of a
    single B cell) = monoclonal Ab. Monoclonal Ab do
    provide reproducible results for clinical testing ex. for
    immunological typing of bacteria, pregnancy testing and
    blood typing.

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