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INTRODUCTION
The clinical microbiology
rotation at HCMC consists of a total of 8 weeks of self-instructional
and practical laboratory experiences. The various disciplines studied,
including clinical microbiology, are: anaerobic microbiology, virology,
mycology, mycobacteriology, and parasitology.
During the course of the rotation the student will be directly involved
in the daily procedures of a large urban hospital's clinical microbiology
department. Our personnel routinely evaluate an average of 7,500 specimens
per month from a variety of body sites and a diverse patient population.
We also perform identification in specialized areas of clinical microbiology,
such as parasitology, mycology, anaerobic bacteriology, virology, and
mycobacteriology.
In order for the students to attain career-entry competencies, we ask
that they come to us with specific "pre-clinical" competencies.
Following is an outline of major academic areas we wish to be covered
before entering the 9 month program.
Attached is an expansion of that outline that includes listings of specific
objectives to be met for each category before the beginning of the 9 month
clinical study and references that we feel will be of value in teaching
and review.
NOTE: Each student is required to write out
answers for all of the objectives. If the objective involves the performance
of some activity, describe what you did in your college courses in order
to meet that objective. These answers must be turned in to the HCMC program
director two weeks before the start of the internship. They will be evaluated for completeness
by the HCMC education techs. Students will be required to review and know
this material when they enter each of the rotations.
Diane K. Berke, MT (ASCP)
Medical Technologist Specialist
Clinical Microbiology/Parasitology
Reviewed 2-01, 1-02, 12-02, 1-04, 1-07, 1-08
The isolation of bacteria from clinical material as etiologic agents of
infection is dependent upon selecting media suitable for bacterial growth.
1. Define enriched, enrichment, selective, and differential.
2. List the following media and identify if it is enriched, enrichment,
selective or differential, and name at least one organism that will grow
on this media.
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Sheep blood agar
CNA or PEA
Chocolate
Chromagar™
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Selenite-F broth
Thayer-Martin
Mac Conkey's
Regan-Lowe
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XLD agar
Hoektoen-Enteric Agar
Fastidious Broth
BCYE - alpha agar |
1. Describe aseptic technique.
2. Draw the standard isolation streaking pattern for inoculating an agar
plate.
3. Describe "safe" techniques (Universal Blood and Body Fluid
Precautions) when working with microbiologic materials.
4. Describe appropriate temperature and atmospheric conditions to incubate bacteriologic
cultures.
5. Describe the difference between disinfection and sterilization.
The most important stain used in the clinical microbiology lab is the
Gram stain. The
acid-fast
stain is also an important tool in alerting physicians to the possible
presence of Mycobacterium sp. in patient specimens.
1. Write a short discussion of bacterial cell wall content in relation
to the Gram stain reaction of the bacterial cell.
2. Write a description of how to perform a Gram stain.
3. Describe basic cellular components that may be found in a Gram stain
of clinical material, such as PMN's, mononuclear WBC's, epithelial cells,
tissue cells.
4. Describe the steps in a Ziehl-Neelson acid fast
stain and an auramine-rhodamine fluorescent stain.
Write a short explanation of each of the following tests (principles,
reactions, media used, reagents) and list an organism or group of organisms
that the test is used to identify. The test listings are common and should
be practical at the university level.
Beta lactamase
Bile solubility
Butyrate test
CAMP test
Catalase production
Citrate utilization
Deamination
Decarboxylation
D'nase
D-Test
Esculin hydrolysis
Fluorescent antibody (FA) tests
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Hydrogen sulfide production
Indole
Latex Agglutination Tests
Neisseria carbohydrate utilization
Nitrate reduction
Optochin
Oxidase
Oxidation-Fermentation tests
ONPG
PYR test
Triple sugar iron agar
Urease |
List at least 2 organisms classified as pathogens from the following sites.
If the site is normally "sterile" you will indicate it as "sterile."
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Respiratory tract
Ear
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Skin
Eye
Genito-urinary tract
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Blood
Gastro-intestinal tract
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Body Fluids: pleural,
peritoneal, pericardial
Joint, Cerebrospinal |
A. Staphylococci
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1. List the growth and media
requirements of Staphylococcus aureus, epidermidis, and saprophyticus.
2. Discuss the identification of Staphylococcus on the basis
of:
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a) Gram stain morphology
b) glucose utilization
c) hemolytic activity
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d) catalase reaction
e) coagulase test or rapid latex test
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3. List a test that will separate and identify
Staph aureus from Staph epidermidis.
4. Write a short discussion of the pathogenicity of Staph aureus
in relation to specific toxin and enzyme production.
5. Write an explanation of the relationship of coagulase, capsular
polysaccharide and beta lactamase production to the virulence of
Staph.
6. List at least 5 diseases caused by Staphylococci and a
drug of choice used in treating staphylococcal infections.
7. List at least 1 biochemical characteristic that separates the
Staph from the Micrococci and Aerococcus.
8. List at least 3 clinical sites and the species of Staph
that can be found as "normal flora."
9. Discuss the pathogenicity of Staphylococcus saprophyticus
and a test to confirm its identification.
10. Discuss the pathogenicity of Staphylococci in terms of cell
wall composition and toxin and enzyme production. In your discussion
you should be able to define:
exotoxin
enterotoxin
exfoliative toxin
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coagulase
proteinase
TSS toxin
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leukocidin
penicillinase
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11. Discuss the mechanism of antibiotic
resistance in staphylococci, ie: MRSA (methicillin resistant Staph
aureus), VISA (vancomycin intermediate Staph aureus) and how these organisms are detected in the Microbiology Lab.
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B. Streptococci
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1. List the growth and media
requirements of Streptococci.
2. Define the differences between alpha, beta, gamma, and alpha-prime
hemolysis.
3. Discuss the criteria used in categorizing streptococcus by Lancefield
grouping.
4. Discuss the use of the following media and tests and how they are
used to identify streptococci (also see section IV):
FA
Bile solubility
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6.5%
NaCl broth
CAMP test |
Optochin
Esculin medium
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PYR
test
LAP test |
5. Discuss the pathogenicity and host immunity of the following Strep,
identifying at least one disease process with each:
S. pneumoniae
S. viridans
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S. agalactiae
Enterococcus group
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S. pyogenes
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6. Describe the streptococci on the basis of:
Gram
stain
Colony morphology |
Hemolytic
activity
Catalase reaction |
7. List at least one test that will separate
the strep from the staph.
8. Discuss the virulence of certain streptococci in relationship to
the presence of antigenic substances and enzymes or toxins. You should
use and define the following terms in your discussion:
C carbohydrate
M protein
Erythrogenic toxins
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Hemolysins
Hyaluronidase |
9. Define streptolysin O and S.
10. List 2 post-Streptococcal diseases.
11. Define VRE and VSE as related to the
enterococci.
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C. Neisseria
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1. Discuss media, growth requirements
and common site of isolation of the Neisseria/Moraxella.
2. Discuss the characteristic morphology and oxidase reaction of the
Neisseria species.
3. Discuss the function and composition of Thayer-Martin agar.
4. Discuss the 4 main antigenic groups of Neisseria meningitidis
and how they are classified.
5. Discuss test used to speciate Neisseria and Moraxella.
6. Discuss host immunity to N. gonorrhoeae and N.
meningitidis.
7. List the species of Neisseria implicated in meningitis and
a venereal disease and the specimens of choice to collect for isolation.
8. Define the term "carrier" as associated with N. meningitidis.
9. Discuss the significance of beta-lactamase testing of Neisseria
in determining drug therapy.
10. List at least one "non-cultural" method of identifying
an infection with Neisseria gonorrhoeae. |
D. Yeasts
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1. Describe yeast and/or arthrospores
on a Gram stain.
2. List a yeast commonly associated with meningitis in the compromised
host.
3. Describe the germ tube test and its use.
4. Describe the cryptococcal antigen test and its use.
5. List at least 3 common laboratory media (from section I) that support
the growth of yeast.
6. List at least 1 mycologic media that supports the growth of yeast.
7. List at least 3 sites where C. albicans is endogenous and
3 sites where it may be pathogenic. |
E. Enterobacteriaceae
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1. Define the Enterobacteriaceae
on the basis of:
Gram stain
Oxidase reaction
Growth requirements
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Motility
Nitrate reduction
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Glucose utilization
Gas production
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2. List important biochemical characteristics
and implicated pathogenicity of the following:
E. coli
0157:H-7
Enterobacter
Escherichia
Klebsiella
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Morganella
Proteus
Providencia
Salmonella
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Serratia
Shigella
Yersinia |
3. Define K, Vi, O, and
H antigens.
4. Describe the composition of gram negative bacterial endotoxins
and their effects on the host.
5. List one genus-species of Enterobacteriaceae that produce endotoxins
and two that produce enterotoxins.
6. Discuss enteric serotyping and why it is used.
7. Define the term ESBL (extended spectrum beta lactamase production).
Name one genera of enterobacteriaceae where this enzyme is found.
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F. Miscellaneous Gram Negative Coccobacillary Bacteria
including:
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Pasturella
Francisella
Bordetella
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Brucella
Haemophilus
Yersinia
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Campylobacter
Legionella
Helicobacter pylori |
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1. Discuss the pathogenicity
of Pasturella multocida and how it is transmitted to man.
2. Name the etiologic agent of tularemia and describe how it is transmitted.
How is it isolated in the laboratory?
3. Name the bacteria that causes whooping cough and why Regan-Lowe
media is used to isolate this organism.
4. Discuss the value of serologic procedures in confirming cases of
tularemia and molecular procedures in confirming whooping cough.
5. Name the bacteria that causes "plague" and how it is
transmitted to man.
6. Name the bacteria that causes brucellosis. How does man acquire
this disease?
How would you isolate the bacteria?
7. Name the Gram negative coccobacillary
bacteria of the above group that is an "intracellular" obligate
parasite.
8. Discuss the special growth and media requirements of the Haemophilus,
defining the terms X and V factor in the discussion, and ALA.
9. Describe "satelliting."
10. List the species and serologic type of Haemophilus associated
with meningitis.
11. Discuss the role of beta lactamase testing in the treatment of
Haemophilus infections.
12. Name the species of Haemophilus associated with an ulcerative
venereal disease. Where is this disease endemic?
13. Describe the Gram's stain, oxidase reaction, media, and growth
temperature for isolating Campylobacter sp. vs. Helicobacter
pylori.
14. Describe the Gram's stain reaction and morphology of Legionella
species, the bacteria associated with Legionnaire's disease. List
its growth requirements.
15. List the media and growth requirements
of Yersinia sp. |
G. Aerobic and Anaerobic Spore Forming and Non-spore
Forming Gram Positive Bacilli including:
Cornebacterium
sp.
Listeria
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Nocardia
Clostridium |
JK-like
organisms
Erysipelothrix
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Bacillus Anthracis
Bacillus Cereus |
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1. Discuss the mechanisms of toxin, antitoxin, and
toxoid production of Corynebacterium diptheriae.
2. Describe the colonial morphology and gram staining characteristics
of Listeria monocytogenes.
3. Describe the Gram stain and colonial morphology of Erysipelothrix
rhusiopathiae.
4. Describe the acid fast stain reaction of Nocardia sp.
5. Discuss the mechanisms of toxin production in Clostridium
perfringens, botulinum, difficile,
and tetani.
6. Discuss the cellular antigen and exotoxin components of Bacillus
anthracis.
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H. Mycobacteria
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1. Define the term "acid fast,"
and discuss the cell wall composition of mycobacteria that contributes
to their acid fast properties.
2. List common body sites for isolation
of Mycobacterium tuberculosis and Mycobacterium leprae.
3. Name a stain of choice used to detect the presence of acid-fast
bacteria in clinical material.
4. Name the usual causative agent of tuberculosis and leprosy.
5. List a group of tests used to separate M. tuberculosisfrom
other Mycobacterium sp.
6. Name a mycobacteria that is a "rapid grower" and also
a marker for HIV.
7. List at least one species of saprophytic
mycobacteria, not associated with clinical disease.
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I. Non-Enterobacteriaceae and Other Unusual Gram
Neg Rods
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1. Define the term
"non-fermentor" using the terms glucose utilization, oxidase
reaction, Gram stain reaction and growth on MacConkey agar.
2. Describe the reaction of non-fermentors on TSI.
3. Describe the colonial and biochemical characteristics of Ps.
aeruginosa using the following terms:
Odor
Pyocyanin
Pyoverdin
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Fluorescein
Oxidase reaction
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Pyorubin
Pyomelanin
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4. Define the term "opportunist." |
J. Chlamydia
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1. List the characteristics
that make chlamydia different from viruses and bacteria.
2. List the species of chlamydia that
causes a common sexually transmitted disease.
3. Discuss a cultural and non-culture method to identify chlamydia.
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In a short paragraph, describe
the Kirby-Bauer procedure, the MIC procedure and the
E-Test and the advantages of each in testing.
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1. Define the following
terms:
Anaerobic
Microaerophilic
Strict anaerobe
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Facultative
anaerobe
Aerotolerant |
2. Describe how an anaerobe jar works.
3. List the anaerobic Gram-positive rod
that causes gas gangrene.
4. Describe, in a paragraph, how you would isolate an anaerobe.
List the media that you would use and what equipment you would need.
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1.
Describe how to make and view a wet prep.
2. Define the following terms:
parasite
endoparasite
ectoparasite
commensal
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pseudoparasite
host
definitive host
vector host
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intermediate host
1st & 2nd interm. host
reservoir host |
3. State at least 3 factors involved in the transmission
of parasitic diseases.
4. Discuss the 3 major classifications of host immunity associated
with parasitism.
5. List the five major groups of medically important
parasites and list one genus-species representative of each.
6. List at least one common fecal concentration,
preservation, and staining procedure.
7. List at least 3 parasites
found in the immunocompromised host.
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A college level course in medical mycology is highly
recommended. At HCMC, a 5 session workshop will be provided to enhance
skills acquired at the college level.
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1. Discuss how viruses differ from
bacteria.
2. List viral classifications (DNA or RNA) and list at least 2 viruses
in each group that cause human disease.
3. Discuss how viral load values affect and assist the treatment of
AIDS and Hepatitis |
XII. LABORATORY SAFETY
You should be able to discuss and practice laboratory safety and universal
blood and body fluid precautions standards.
Write a short discussion on principles of
DNA probe methods, LCR (ligase chain reaction) and PCR (polymerase chain
reaction).
Brown, Harold W.:Basic Clinical Parasitology, 5th Ed., New York
City, 1975, Appleton- Century-Crofts.
** Forbes, Betty A., et al: Bailey and Scott's Diagnostic Microbiology,
12th Ed., St Louis, 1998, C.V. Mosby Company.
Garcia, et al.: Diagnostic Medical Parasitology, 5th ed., 2007.
ASM, Washington DC.
Koneman, E.W., Allen, S.D., Dowell, V.R., Jr. Sommers, H.J.: Color
Atlas and Text booK of Diagnostic Microbiology, 5th Ed., Philadelphia,
1997, J.B. Lippincott Co.
Balows, A., Hausler, Wm. J., et al: Manual of Clinical Microbiology,
7th Ed., Washington, D.C., 1999, ASM
McFaddin, Jean F.: Biochemical Tests for Identification of Medical
Bacteria, 2nd Ed., Baltimore, 1980, Williams and Wilkins.
** Leventhal and Cheadle, Medical Parasitology Self-Instructional Unit,
5th Ed., 2002, F.A. Davis Co.
** Kern, Martha E., Medical Mycology, A Self-Instructional Unit,
2nd Ed., 1997, F.A. Davis Co.
* Recommended for student to purchase
** Required for student to purchase
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