Guidelines for Work-up of Wound Specimens

2013 Webinar Series
Guidelines for Work-up of Wound Specimens
2/12/2013
Speaker
Yvette S. McCarter, PhD, D(ABMM), Director, Clinical Microbiology Laboratory, Shands Jacksonville Medical Center, Jacksonville,
FL
Yvette S. McCarter, PhD, D(ABMM) is Professor of Pathology at the University of Florida College of Medicine-Jacksonville and the
Director of Clinical Microbiology at Shands Jacksonville in Jacksonville, FL. She received her doctorate in Clinical Microbiology
from the Medical College of Virginia and completed postdoctoral fellowship training in Public Health and Medical Microbiology at
Hartford Hospital under the direction of Raymond Bartlett, MD. Following fellowship training she assumed the position of
Associate Director of the Division of Microbiology at Hartford Hospital. She held this position from 1992-1999 and in 1999 she
became the Director of the Division of Microbiology at Hartford Hospital and Director of the Microbiology Laboratory at Clinical
Laboratory Partners, Newington, CT. In 2001 she assumed her current position at the University of Florida. Dr. McCarter is also
a Diplomate of the American Board of Medical Microbiology.
Objectives
At the conclusion of this program, participants will be able to:
•Identify methods for the evaluation and work up of bacteriology cultures.
•Discuss the advantages of reporting morphological identifications of organisms on direct specimen Gram stains.
•Design cost-effective, clinically-relevant strategies for laboratory work up of wound specimens.
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Faculty Disclosure
GUIDELINES FOR WORK-UP OF WOUND
SPECIMENS
The Association of Public Health Laboratories adheres to established standards
regarding industry support of continuing education for healthcare professionals.
The following disclosures of personal financial relationships with commercial
interests within the last 12 months as relative to this presentation have been
made by the speaker:
Yvette McCarter, PhD, D(ABMM)
Director of Clinical Microbiology, Shands Jacksonville
Professor of Pathology, UF College of Medicine-Jacksonville
Jacksonville, FL
Nothing to disclose
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Objectives
Introduction
• Discuss the use of the direct Gram stain to assess
specimen quality and the advantages of reporting
morphological identifications of organisms on direct
specimen Gram stains.
• Wound cultures are frequently contaminated with
resident flora -- difficult to determine which organisms
are potential pathogens.
• Work up can be problematic
time and
resources spent to work up cultures of little clinical
value.
• Working up mixed cultures may generate clinically
misleading results
inappropriate and
unnecessary antimicrobial therapy.
• Describe the use of direct Gram stain results to aid in
the work-up of wound cultures and review a new CAP
requirement for utilizing the Gram stain.
• Demonstrate how to design and implement clinically
relevant, timely approaches for cost-effective wound
culture work-up.
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Still striving…
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Utility of the Gram Stain
• A well-performed direct Gram stain is rapid,
inexpensive, informational
The major goal of the clinical microbiology
laboratory is to provide information of
maximal clinical usefulness as rapidly as is
consistent with acceptable accuracy and
minimal cost.
• Allows for evaluation of specimen quality
o Identification of superficially contaminated specimens
o Enhances the discrimination between samples with
potential pathogens vs. colonizing flora
Jay P. Sanford, MD (1974)
• Provides presumptive organism identification
o Guides rational selection of preliminary antibiotic
therapy
o Guides interpretation of culture results
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Squamous Epithelial Cells –
Superficial Contamination
Gram Stain Screening Criteria
• Standardized screening criteria
o Consistent smear interpretations among technologists
o Establish the quality of specimens submitted
• Optimal smear preparation and staining
o Targeted selection of grossly mucopurulent portions of
the specimen
• Use interpretive comments
o Assist clinicians in interpreting results
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Interpretation and Reporting of
Organisms in Direct Smears
Neutrophils – Inflammation
Bartlett. 1982. JAMA 247:857-59
• Designation of organism genera more useful than
description of organism morphology
Bartlett et al. 1991. Diagn Microbiol Infect Dis 14:
195-201
• Reliable differentiation of Gram negative bacilli
o Bacteroides or Haemophilus – 95%
o Enteric Gram negative bacilli – 82%
o Pseudomonas – 56%
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Enteric-like Gram negative bacilli
Plump Gram-negative rods, can have non-uniform sizes and can
sometimes show bipolar staining
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Gram negative coccobacilli suggestive of Haemophilus
or Bacteroides
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Gram-negative coccobacilli/pleomorphic rods that may be faint
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YM
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staining
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Non-enteric Gram negative bacilli
Gram positive cocci suggestive of Staphylococcus
Thin, somewhat faint staining, uniform in shape, elongated rods,
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sometimes in pairs end to end (“hot dogs”)
Gram-positive spherical Gram-positive
cocci in clusters or tetrads 14
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Gram positive cocci suggestive of Streptococcus
Gram positive bacilli suggestive
of Bacillus/Clostridium
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Gram positive cocci in pairs or chains
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Large box-car shaped Gram
positive bacilli
Gram positive bacilli suggestive
of Diphtheroids
Small Gram positive bacilli, clubshaped, Chinese letter
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16 appearance
Use of Interpretive Comments
Yeast
Wound specimens containing more squamous
epithelial cells than neutrophils
X6976 Collect D/T: 12/31/2012 1215
Receive D/T: 12/31/2012 1345
Wound Culture and Gram Stain
Specimen Description
Foot
Direct Smear Suggests
No neutrophils
Many squamous epithelial cells
No organisms seen
Squamous cells in this specimen indicate the presence of superficial material
that may contain contaminating or colonizing bacteria unrelated to infection.
Collection of another specimen is suggested avoiding superficial sources of
contamination.
Culture Pending
Gram positive/variable yeast with/without buds, or with/without
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pseudohyphae
Report Status
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Preliminary
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Use of Interpretive Comments
Use of Interpretive Comments
Squamous epithelial cells are seen in a specimen
with an anaerobic culture request
Specimens containing numerous morphotypes of
bacteria in the direct Gram stain
DIRECT SMEAR SUGGESTS:
Moderate neutrophils
H73026 Collect D/T: 01/01/2013 0900
No squamous cells
Receive D/T: 01/01/2013 1100
Gram negative rods suggestive of Enteric-like Gram negative bacilli
Anaerobe Culture
Specimen Description
Gram positive cocci in clusters suggestive of Staphylococcus
Leg
Gram positive rods suggestive of Bacillus/Clostridium
Request credited
Gram negative coccobacilli suggestive of Bacteroides/Haemophilus
Evidence of superficial material. Specimen unsuitable for anaerobic culture.
Please consult Microbiology if clinical considerations warrant complete
processing of this specimen. (Specimen will be held 5 days)
Report Status
Gram stained direct smear suggests the presence of a mixture of organisms.
Culture has not been performed because a mixed culture can be anticipated. This
information is of doubtful value for direction of therapy against mixed infections
containing this many potential pathogens. Please consult Microbiology if clinical
considerations warrant complete processing of this specimen. (Specimen will be
held 5 days).
Final 01/01/2013
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A new requirement from the College of
American Pathologists
Work up of Wound Cultures
• There are no clear guidelines for working up bacterial
cultures
o Rely on information that is available in the literature or
colleagues
o Query colleagues from same or different laboratories –
often get numerous differing opinions
• There seems to be a need for some consistency when
performing culture work up
**NEW/REVISED** 07/31/2012
MIC.21530 Direct Gram Stain Procedures
o Uniformity in work up and reporting of bacterial isolates
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MIC.21530 Direct Gram Stain Procedures
(Phase I)
**NEW/REVISED** 07/31/2012
• The laboratory has protocols in place to use Gram stain results
to provide a preliminary identification of organisms, evaluate
specimen quality when appropriate, and to guide work-up of
cultures.
NOTE: The laboratory should have guidelines for the interpretation of the Gram
stain reaction of the organism, morphology of the organism, and the
quantification of organisms and cells. The protocol should address correlation
of direct Gram stain results with final culture results.
This does not mean that interpretation of the Gram stain morphology
suggesting a specific organism identification (e.g. gram positive diplococcic
morphologically suggestive of pneumococcus) is required.
Evidence of Compliance:
 Written procedure for Gram stain (laboratories may use the correlation of
Gram stain results with the final culture results as a component of the QC
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program)
Work up of Wound Cultures
Specimen Quality
• When working up wound cultures the following are
assumed:
o Neutrophils – infection or inflammation
o Squamous epithelial cells – superficial contamination =
If a specimen contains a large amount of SEC,
superficial contamination is likely.
• Bacteria isolated from such specimens should be
minimally worked up
o Extensive testing on heavily mixed cultures should not
routinely be performed.
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Work up of Wound Cultures
Work up of Wound Cultures
Two Approaches
Q Score System (RC Bartlett, 1974)
• Determine average number or PMN and SEC/LPF on direct
gram stain and assign value (see Key)
• Add values together using table below to obtain Q score
• Q score = # of potential pathogens (PP) worked up
These systems provide the means to consistently
work up organisms from wound cultures
• Quality (Q) Score System
• Q/234 System
Cumitech 7B: Lower Respiratory Tract Infections, ASM 2004
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Key:
0 = no cells
1 = 1-9/lpf
2 = 10-24/lpf
3 = ≥25/lpf
Q0 = 0PP
Q1 = 1PP
Q2 = 2PP
Q3 = 3PP
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Work up of Wound Cultures
Work up of Wound Cultures
Q Score System
Q Score System
• # PP in culture < Q-score: work up PP with ID/AST
(2PP)
(Q3)
• Specimens without SEC are considered good quality
specimens regardless of the number of PMN
• Up to 3 organisms can be considered potential
pathogens (PP) and be worked up (ID/AST) if from a
good quality specimen (Q3)
• The lower quality of the specimen (e.g., the more SEC
present) the fewer the organisms worked up (Q2, Q1)
• # PP in culture > Q-score: Look to direct Gram stain
(3PP)
(Q2)
Work up only PP that were
seen in direct Gram stain with
ID/AST
o Q0 – provide morphologic ID of organisms present but
no work up
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MID = any rapid biochemical testing that can be performed (catalase, indole,
oxidase, strep typing, etc.)
Work up of Wound Cultures
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Example 1: Leg Wound
Q/234 System
• Evaluate specimen quality (PMN and SEC) using whichever
system you choose
• Culture work up is based on number of PP present:
≤ 2PP = Work up with ID/AST, as appropriate
3PP = Look to direct Gram stain
Work up to 2 PP if they are
seen in the direct
Gram stain
If all PP in the culture are seen
in direct Gram stain – Do not
work up; perform morphological
identification (MID) on all
isolates
GS: moderate PMN (+2), few SEC (-1), many gram positive cocci
suggestive of Staph, many gram positive cocci suggestive
of Strep
CULT: many Staph aureus, many β hemolytic strep, moderate
coagulase neg staph, few diphtheroids
WORK UP:
If all 3 PP are seen in the
direct Gram stain, perform
MID on all 3
Q Score (Q1=1PP):
4PP = Perform morphological identification
(MID) on all isolates
Q234 (2PP):
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Example 1: Leg Wound
Example 1: Leg Wound
GS: moderate PMN (+2), few SEC (-1), many gram positive cocci
suggestive of Staph, many gram positive cocci suggestive
of Strep
CULT: many Staph aureus, many β hemolytic strep, moderate
coagulase neg staph, few diphtheroids
WORK UP:
GS: moderate PMN (+2), few SEC (-1), many gram positive cocci
suggestive of Staph, many gram positive cocci suggestive
of Strep
CULT: many Staph aureus, many β hemolytic strep, moderate
coagulase neg staph, few diphtheroids
WORK UP:
Q Score (Q1=1PP):
Q Score (Q1=1PP):
MID S. aureus and β Strep; Report Mixed
flora
Q234 (2PP):
MID S. aureus and β Strep; Report Mixed
flora
Q234 (2PP): Work up S. aureus and β Strep; Report Mixed flora
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Example 2: Abdominal Wound
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Example 2: Abdominal Wound
GS: many PMN (+3), no SEC (0), many enteric-like gram
negative bacilli, many gram negative coccobacilli
CULT: moderate E. coli, moderate Bacteroides spp., few
Enterococcus spp.
GS: many PMN (+3), no SEC (0), many enteric-like gram
negative bacilli, many gram negative coccobacilli
CULT: moderate E. coli, moderate Bacteroides spp., few
Enterococcus spp.
WORK UP:
WORK UP:
Q Score (Q3=3PP):
Q Score (Q3=3PP):
Q234 (3PP):
Work up E. coli, Bacteroides spp. and
Enterococcus spp.
Q234 (3PP):
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Example 2: Abdominal Wound
Premise for Q Systems
• The more superficially contaminated the specimen, the
higher the number of colonizing organisms present
• The quality of the specimen is important in determining
acceptability of specimen and extent of culture work up
• If organisms are seen in the direct Gram stain, there is
a greater chance they are associated with an infective
process
GS: many PMN (+3), no SEC (0), many enteric-like gram
negative bacilli, many gram negative coccobacilli
CULT: moderate E. coli, moderate Bacteroides spp., few
Enterococcus spp.
WORK UP:
Q Score (Q3=3PP):
o At least 105 organisms must be present to visualize
them in direct Gram stain
Work up E. coli, Bacteroides spp. and
Enterococcus spp.
Q234 (3PP): Work up E. coli and Bacteroides spp.; MID
Enterococcus spp.
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Advantages of Q Systems
Advantages of Q Systems
• Offer a consistent approach for interpreting
cultures
• No potential pathogen is ever ignored
o All potential pathogens are reported – may not be
fully identified or have full AST performed
o The pathogens that some believe should “ALWAYS
BE WORKED UP” (S. aureus, β Strep, and P.
aeruginosa) always indicated on the report
o Either system can be modified to include screening
for MRSA, VRE, ESBLs, etc
o The systems are based on the quality of the
specimen (primarily SEC)
o Work up is based on organisms seen in the direct
Gram stain
o Limits the number of organisms worked up from
mixed cultures
reporting of misleading
information can be minimized
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Advantages of Q Systems
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The Q Systems in Practice…
• They provide guidelines
o Both Q Systems offer guidelines for a systematic
approach to culture interpretation and work up
o These Guidelines are just that = Guidelines!
Exceptions can be made if necessary
o Any concerned physician can consult with
microbiology to have further work performed on any
culture if clinically indicated
C Matkoski, SE Sharp, and DL Kiska. 2006.
Evaluation of the Q Score and Q234 Systems for
Cost-Effective and Clinically Relevant Interpretation of
Wound Cultures.
J Clin Microbiol 44:1869-1872
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Matkoski et al.
Matkoski et al.
• 305 wound cultures evaluated over 3 months
• The reagent cost savings as compared to the
routine procedure were greatest with Q score:
o 147 (48%) and 84 (28%) would have been
interpreted differently with Q-score and Q234
systems, respectively, as compared to their routine
wound procedure
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o Q-score: $4895 per year
o Q234: $1503 per year
Routine procedure = work up to 3 organisms per
culture
• The cost savings would increase for both Q systems
if technologist time were factored in as well
o The majority of these differences were the result of
unnecessary culture work-up utilizing their routine
procedure
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Matkoski et al.
Matkoski et al.
Conclusions
• Although the Q-score had a yearly savings of
$3,392 over the Q234 system and would have
eliminated more culture work-up, it had several
issues that would need to be addressed prior to
implementation:
o It would require physician approval for not culturing
specimens with a Q score of zero (Q0)
o It would require the training of all shifts to be able
to examine the Gram stain and generate a Q score

• Both the Q score and the Q234 systems showed
clinical relevance, cost effectiveness and would
allow for standardized approaches to the work up
of wound cultures.
• The Q234 system proved to be a more practical
procedure to implement in their laboratory.
o The Q234 system does not require a change in Gram
stain interpretation or physician approval for specimen
rejection.
The Q234 system did not require a different
interpretation of direct smears
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Matkoski et al.
As your mother used to say…
Conclusions
“Watch your P’s and Q’s”
• All potential pathogens are reported from a culture
with either full ID or MID, allowing for further
consultation with the ordering physician if clinically
warranted.
• The Q234 system, which is more structured and
cost-effective than their current method, was
acceptable to their Infectious Disease physicians.
• This system allowed technologists to make more
independent and consistent decisions about the
significance of organisms in a wound culture.
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Helpful References
Sharp, SE, et al. 2004. Cumitech 7B, Lower Respiratory
Tract Infections. ASM Press, Washington, DC
C Matkoski, SE Sharp, and DL Kiska. 2006. Evaluation of
the Q Score and Q234 Systems for Cost-Effective and
Clinically Relevant Interpretation of Wound Cultures. J
Clin Microbiol 44:1869-1872.
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• Gram stain of direct specimens is useful…
o
o
o
o
For specimen quality assessment
For providing rapid presumptive identification
As a guide for culture work up
Permits compliance with new CAP checklist item
• Watch your P’s and Q’s
Determine your P’s
Potential Pathogens
Pick one of the Q’s
Q Systems
Report consistent and
clinically-relevant wound
culture results
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