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Microbio and Body Fluid Analysis
clin. Skills
49
Medical
Graduate
06/10/2009

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Term
define fungus
Definition
plant-like org-m --- yeast and mold
Term
define arthropod
Definition
invertebrate oranisms; may serve as Dz vector
Term
define Helminthes
Definition
classification of parasitic worms
Term
define NML flora
Definition
microorganisms residing in healthy host
Term
define pathogens
Definition
organisms capable of producing Dz or illness
Term
define Opportunistic infection –
Definition
causes Dz in host w/weaken immune system
Term
define Protozoa
Definition
motile, animal-like single celled org-ms, classified by locomotion
Term
define Parasites
Definition
depend upon another organism for survival which it receives at the expense of host
Term
define Virus
Definition
non-living organism dependent on another cell for replication
Term
areas of microbiology (list 5):
Definition
Bacteriology –
Virology –
Mycology –
Parasitology –
Serology (+/-) –
Term
what technique is used for specimen collection?
Definition
- Aseptic or “sterile” technique --- Avoid contamination
- Specimen site
o Site appropriate for type of study
o Proper medium and collection apparatus
Term
specimen collection: what is important about transport, specimen processing and interpretation results?
Definition
Transport media --- support microbial viability during transit
Anaerobic specimens require special transport and maintenance media
Specimen processing: label properly (inoculation  incubation ,temperature, CO2)
Dx-ic relevant test --- proper Dx test based upon S/S
Interpretation of results -record correctly – 80% lab error d/t clerical error!!!
Term
What is Gram stain?
Definition
- helps Preliminary measure of org-ms
- Method of chemically staining specimens, ientifying org-ms by staining results along w/morphology
Term
what are the major steps in gram stain?
Definition
- Prep slide w/body fluid/specimen
- Heat for seconds
- Apply 1’ stain crystal violet for 1 minute
- Apply mordant; Gram’s iodine for 1 minute
- Wash w/ acid EtOH
- Apply counterstain; saffarin for 1 min
- Wash with H2O (~5 seconds)
- View under microscope
Term
what can you examin under low power (10x)?
Definition
- Note PMNs, epithelial cells
- Note quality of smear
Term
what can you examin under high power - oil immersion(100x)?
Definition
- Note microbial elements
- Scan several fields
- Quantities (few, many, or numeral grade)
- Patterns
- Results are subjective
Term
what is culture media? list some properties:
Definition
- Nutrient media of various composites and additives
- Selective or isolative
- Enrichment
- Maintenance
- Liquid, solid or semisolid consistency
- Transport
Term
list Bacterial Culture Media
Definition
Blood Agar Medi: Sheep’s Blood Agar (SBA), Chocholate Agar - Thayer Martin: GC Agar/plate Agar - MacConkey Agar:Inhibits Gram(+)s, pH color indicator - Tryptic Soy Agar/Broth: grow on pure Cx
Term
What are sources of error in gram stain prep?
Definition
- Sources of error: poor slide making ; failure to heat fix slide; overheating; expired reagents; over- or under-decolorizing slide; contaminated reagents w/microorganisms; operator interpretation
Term
list other methods to differentiate bacteria (not gram stain)
Definition
o Biochemical and Enzyme Testing: Catalase, Oxidase, Coagulase (clot=”+”), Citrate
o Automated
o Serological methods: PCR
o Immunoassays
Term
Serological methods: PCR allow the following:
Definition
 DNA probe (nucleic acid)
 Genetic id-n of microbes
 Rapid multiplication of DNA sequence
 GC/Chlamydia probe
Term
Describe Immunoassays:
Definition
 Biochemical test that measures the concentrationn of a substance in a biological liquid: e.g. serum or urine
 Specific binding of an Ab --> antigen
 Reference lab test
 Expensive !!!
 Enzyme-linked immunosorbent assay (ELISA) --> RMSF --- Scarlet Fever
 Fluorescent Ab test
 Antigen-Ab test --> rapid Strep
 Latex agglutination --> Syphillis Fast (new)
Term
Microbial Classification - Taxonomy:
Definition
classification, nomenclature, laboratory, identification
Term
Microbial Classification - Gram Positive
Definition
Aerobic bacilli and aerobic cocci
Term
Microbial Classification - Gram Negative
Definition
Aerobic cocci, aerobic rods, Nonfermentors
Term
Microbial Classification - anaerobes
Definition
Gram (+) and Gram (-)
Term
classify as G(+)/(-):
o Aerobic cocci: Streptococcus, Staphylococcus, Enterococcus (prev. gr D Streptococcal)
Definition
Gram (+)
Term
classify as G(+)/(-):
Rods or bacilli: Corynebacteria, Listeria monocytogenes, Nocardia, ; Bacillus anthracis (Anthrax)
Definition
Gram (+)
Term
classify as G(+)/(-):
anaerobs (survive in GI) - Actinomycetes – branching organism; Lactobacillus; Clostrodium species (C. tetani, C. difficile, C. perfringens, C. botulinum)
Definition
Gram(+)
Term
classify as G(+)/(-):
o Aerobes bacilli or rodes ---
 Pseudoonadacae  Pseudomonas aeruginosa
 Bordetella pertussis
 Helicobacter pylori
 Legionella pneumophila
Definition
Gram (-)
Term
classify as G(+)/(-):
o Rods – Facultative anaerobes – Enterobacteriaceae: large Gram(-) rods, assoc-d w/GI infections
 Escherichia coli – common encountered pathogen, NML flora in gut
 Klebsiella – mucoid colonies, capsule !!! (hard to treat)
 Proteus species – stinks badly
 Obligate aerobe – Vibrionaceae (Vibrio cholerae)
 Facultative anaerobic - pasteurellae: coccobacilli, Haemophilus influenza, P. multocida
Definition
Gram (-)
Term
classify as G(+)/(-):
Cocci: Neisseriaeae – Neisseria meningitides – aerobic
Definition
Gram(-)
Term
classify as G(+)/(-):
o Obligate IC parasites: Richettsiae, Rocky Mountain Spotted Fever
Definition
G(-)
Term
Classify as G(+)/(-):
o Anaerobes: Bacteroidaceae (Bacteriodes fragilis); Fusobacterium (F. nucleatum)
Definition
G(-)
Term
Classify as G(+)/(-):
o Spirochetes (Treponema pallidum - syphillis); Mycobacteria (Mycobacterium avium, M. tuberculosis)
Definition
G(-)
Term
Classify as G(+)/(-):
o Chlamydia (bacteria):
 Obligate IC org-m: class-d as bacteria (phylum Chlamydiae); replicates IC-ly, smaller than viruses
 Respiratory and sexual transmitted infections
Definition
G(-)
Term
is Cryptococcus neoforma G(+) or G(-)?
Definition
Gram (-)
Term
list common microbial cultures specimen types:
Definition
-Bloos Cx - Respiratory Cx - Tissue Cx - Body fluid Cx (pleural fluid, CSF, peritoneal fluid) - urine Cx)
Term
Blood Cx: indications, collection, process...
Definition
o Indications: detect presences of bacteria in blood; suspicion of bacteremia, sepsis or endocarditis; assess after antimicrobial Tx initiated
o NML ranges: NO growth after 5 Days
o Collection: 2 sets (aerobic/anaerobic) from 2 different sites or times
 Peripheral access preferred
 Volume dependent
 Skinn prepped with Povidine-iodine
 Blood Cx bottle or test tube also cleansed
o Process: Automated sys-m of detection; id-n & antimicrobial sensitivities takes longer time;
 False Positives --- Contaminants
Term
Respiratory Cx
Definition
o Indications: Dx & eval pulmonary related illness suspected to be of infxs etiology; pts w/risk factors
o Specimen culturing methodology & testing divided by location in pulmonary tree: UR, LR
o NML ranges: no growth or NML flora
o UR Cx or rapid test: nasopharyngeal swab or aspirate (wash)
 Specimen from secretions of the upper-most part of the throat
 Dacron or rayon swab
 Wash w/sterile saline in < 2 yo
 Immediate transport  LAB
 Id viruses or bacteria: RSV, Influenza A/B, Bordetella, Staphylococcus carriers
 Throat swab --- specimen often obtained by provider --- rapid streptest:
• Commercially available immunoassay
• Highly specific & sensitive
• Group A β-hemolytic streptococcus or Streptococcus pyogenes
• Cx still gold std for strep Dx: SBA, id-ing β-hemolysis on agar
o LR Cx
 Sputum /Tracheal aspirate (sputum is worthless!!! Per Ferguson)
 Gram stain screening for “acceptability”:
• Contamination  10-25 squamous epithelial cells
• Low yield  specimen rejection or report or NML flora
 Other common analysis:
• Acid fast stain: TB smear; incl-d w/TB Cx; Carbol Fuschin Stain
• Acid fast bacillus (AFB) Cx:
o Mycobacterium tuberculosis
o 3 respiratory specimens
o 2-4 wks for init, res-ts
o Id-n of AFB w/DNA probes faster yield
Term
Urine Culture: indications, colection, ranges, sensitivity...
Definition
- Indications: Dx or confirm UTI, screening in early pregnancy, pts w/risk factors
- Collection: clean catch specimen or sterile collection; sterile container
- Bacteria in suprapubic catheterized specimen is significant
- Urine Cx and Sensitivity: results measured over 48 hrs
 Id-n of microorganisms
 ~Lab will test up to 2 org-ms: E. coli causes ~80% of infections (non indwelling catherized pts)
 Susceptibility or sensitivity testing takes longer --- guide to antimicrobial Tx
Term
What is Antimicrobial Susceptibility Testing(aka sensitivity testing or drug resistance testing)and what are indications?
Definition
- Determines the likelihood that an antimicrobial will be effective in eliminating or inhibiting the growth of the infection
- Indications: guide antimicrobial Tx; nrrow antimicrobial Tx to appropriate agents; decrease med. costs
Term
Waht is the methodology of Antimicrobial susceptibility testing?
Definition
o Kirby Bauer disk agar diffusion
o Broth serial dilution or broth microdilution
o Automation --- rapid results 12-20 hrs
Term
How are results of Antimicrobial Suseptibility testing reported?
Definition
o Zones of inhibition are interpreted for a microbe based upon when exposed in vitro to a antimicrobial agent at specific conc-n
o Susceptible, Indeterminate and Resistance
o Qualitative and Quantitative measurements
o MIC
o MBC
Term
What is MIC?
Definition
– Min. Inhibitory Conc-n:
 Lowest antimicrobial conc-n of antimicrobial agent req-d to inhibit a microorganism
 Methods: (1) serial broth dilution; (2) Agar dilution
Term
what is MBC?
Definition
– Min. Bactericidal Conc-n --- aka min. lethal conc-n (MLC)
 Max dilution of the product that will kill a microbe can be determined by subculturing last clear MIC tube onto growth medium and examining for bacterial growth
Term
How viruses can be detected?
Definition
- Viruses can detected by cytopathic effect, Ab staining, antigen detection, DNA probes, electron microscopy
Term
what is Monospot?
Definition
- Serologic Dx-ic testing for viral Dz: Monospot:
o Heterophile Ab test
o “mono” – EBV
o < Sensitivity in young children
o Blood specimen
Term
what is "cytopathic effect"?
Definition
o Shut down of cell fxn
o Cell transformation: changes in IC structure; viral inclusion bodies; malignant cell changes
o SARS (Conovirus) vs unaffected –
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