Contemporary Issues in Antimicrobial
Susceptibility Testing (AST) for
Public Health Laboratories!
Janet Hindler, MCLS MT(ASCP)
UCLA Medical Center
jhindler@ucla.edu
….and working as a consultant with the
Association of Public Health Laboratories
with support from CDC
At the conclusion of this talk,
you will be able to……
♦ Describe what is currently being done in PHLs re:
antimicrobial susceptibility testing (AST).
♦ List features, benefits, and costs of commonly used
automated and non-automated AST systems.
♦ Discuss the role of the PHL in assisting clinical
laboratories with AST problems including emerging
resistance (e.g., CA-MRSA, VISA, VRSA).
♦ List resources available for performance of AST and
ways in which PHLs may help to provide access to these
resources.
What are PHLs doing
now re: AST?
AST State PHL
Training Needs Survey 2005
♦ 36/52 responses (69%)
– 2 states >1 lab responded
♦ 29/36 labs do some type of AST
– 16 referral testing
– 10 diagnostic testing
– 25 surveillance
♦ Desire more training in AST
– 29/36 for surveillance
– 13/36 for diagnostic testing
Number of State PHLs Performing
AST on Respective Organisms*
For
Surveillance
For
Diagnostics
Campylobacter
4
2
Salmonella spp.
19
6
N. gonorrhoeae
14
7
N. meningitidis
4
1
S. aureus
21
11
S. pneumoniae
12
5
Group B streptococci
2
6
Organism
* of 36 labs that responded
Number of State PHLs Using
Test Methods*
Disk Diff
Etest
Broth
Dilution
Campylobacter
3
3
-
Salmonella spp. 1
12
2
3
N. gonorrhoeae
10
7
1
N. meningitidis
1
3
1
S. aureus
6
15
8
S. pneumoniae
7
10
2
Group B streptococci
2
1
2
Organism
* of 36 labs that responded; 1 Vitek (2)
Additional Data from
AST State PHL Survey
♦ 9/36 said that their epidemiology section collects
cumulative antibiogram data from clinical
laboratories
♦ Additional AST Methods Utilization
– Vitek – 3
– MicroScan – 2
– Sensititre – 1
♦ 23/36 labs use Etest for one or more organisms
AST State PHL Survey
Spring 2005
AK
VT NH
WA
MT
OR
MN
CA
No Response
AZ
CO
IA
IL
KS
OK
NM
TX
HA
RI
MI
NE
UT
MA
NY
WY
NV
Do No AST
WI
SD
ID
Do Some AST
ME
ND
OH
IN
KY
MO
WV VA
AR
SC
AL
NJ
DE
NC
TN
MS
CT
PA
MD
GA
LA
FL
PR
What methods/systems
are available for AST?
Who makes the rules for AST and
reporting of clinical isolates?
♦ The Clinical and Laboratory Standards Institute
(CLSI, formerly NCCLS) subcommittee on AST
– Virtually all labs in USA follow CLSI standards
– Each lab must customize testing and reporting to meet the
needs of the facilities they serve
– Accrediting agency requirements primarily based on CLSI
recommendations
♦ FDA has rules for manufacturers of AST
systems
Current CLSI AST Standards
♦ M100-S16 Tables (2006)*
…..to be used with text documents
explaining how to perform the tests….
M2-A9 Disk Diffusion (2006)**
M7-A7 MIC (2006)**
♦ M45-P Infrequently Isolated/Fastidious
Bacteria (NEW, 2005)
* M100 updated yearly
**M2, M7 updated every 3 years
What drugs should we test / report?
CLSI M100-S16 Table 1
Drugs to Test/Report
How should we interpret results?
CLSI M100-S16 Table 2C
Interpretive Criteria
(Breakpoints)
How can we ensure accurate results?
CLSI M100-S16 Table 3
QC Strain Ranges
Clinical lab can use a CLSI reference
method…..
Broth microdilution MIC
- +
0.5
1
2
4
8
16
32
64
Disk diffusion
(Kirby Bauer)
Drug
>64 >64
A B C D
…or an FDA-cleared commercial system
Etest
Vitek 2
MicroScan
Sensititre
BIOMIC
Phoenix
Comparison of AST Systems
CLSI
Disk Diff
BIO
MIC
Etest Micro Phoenix Sensititre
Scan
Identification of
bacteria
-
X
-
X
X
X
X
AST fastidious
bacteria
X
X
X
X
X
X
X
Automated read
-
X
-
X
X
X
X
Rapid result
option
-
-
-
X
X
X
X
Manual read
option
X
-*
X
X
-*
X
-*
Cost/panel of 10
drugs (approx)
$2
$2
$22
$7 - 10
$7 - 10
$7 - 10
$7 - 10
-*, significant capital equipment cost
Vitek
Why would a PHL do AST?
– Emerging antimicrobial resistance (AR) is a
PH concern
– Clinical labs sometimes need help with
detection of emerging resistance
Why would a PHL NOT do AST?
– Limited resources
– Clinical labs have expertise in AST
“PHLs: Potential Leaders in the Fight
Against Antimicrobial Resistance”
(APHL Minute, May-June, 2006)
Approaches to AR & AST in a
PHL…
♦ Test isolates “referred” from clinical labs
(“referral” isolates)
– To verify noteworthy results from clinical lab
– To test supplemental drugs on highly resistant bacteria
– To do tests not done in smaller clinical lab [e.g., extendedspectrum β-lactamase (ESBL) confirmatory test; mecA,
etc.]
– Clinical labs on “front line”
♦ Surveillance for AR
♦ Test isolates from diagnostic specimens
♦ QA of AR detection / reporting
CLSI M100-S16 (2006) Table 8 (M7)
What isolates should be “verified”
and possibly referred?
Excerpt from:
“Suggestions for Verification of AST Results
and Confirmation of Organism Identification”
Organism or
Group
Staphylococcus
aureus
Streptococcus
pneumoniae
NS, not susceptible
Category I
Verify at all labs
linezolid – NS
vancomycin – I or R
vancomycin – NS
Category II
Verify – institution
specific
oxacillin – R
penicillin – R
CLSI M100-S16 Tables 4 (M2) and 8 (M7)
What is the basis of the
“suggestions for verification” list?
♦List includes results that..
– have never been documented*
– have rarely been documented
– can easily result from technical errors
*critical PH importance!
Algorithm for Testing S. aureus with Vancomycin (VA)
Acceptable
Primary Test Methods
Include:
MIC method plus VA screen plate1
(BHIA with 6 µg/ml of VA)
VA MIC <2 µg/ml
AND NO growth on
VA screen plate
VA MIC = 4 µg/ml
AND/OR
GROWTH on VA screen plate
Report VSSA3
Possible VISA/VRSA
Disk diffusion plus VA screen plate2
(BHIA with 6 µg/ml of VA)
VA zone <15 mm
AND/OR GROWTH on
VA screen plate
VA zone >15 mm
AND GROWTH on
VA screen plate
Possible VISA/VRSA
VA zone >15 mm
AND NO growth on
VA screen plate
Report Probable VSSA3
CHECK for purity
Clinical and Laboratory Standards Institute
S. aureus/Vancomycin Breakpoints
(M100-S16; Jan. 2006)
Susceptible: ¡Â2 µg/ml (VSSA)
Intermediate: 4-8 µg/ml (VISA)
Resistant: ¡Ã16 µg/ml (VRSA)
CONFIRM isolate ID
Algorithm Revised:
March 31, 2006
RETEST using a validated MIC method4
SAVE ISOLATE
NOTIFY infection control, physician, local health department and CDC5 of “possible VISA/VRSA”
SEND to reference laboratory for confirmation
Important Footnotes
1 Laboratories
using automated MIC methods that have not been validated for VRSA detection should add a commercial VA agar screen plate (6 µg/ml).
2
Disk diffusion will not differentiate VISA (MICs 4-8) from susceptible strains (MICs 0.5-2). VA screen plate will not reliably detect strains for which MIC=4.
3 If concerned about a result based on a patient’s history, send to a reference lab for MIC testing.
4 Validated methods: reference broth microdilution, agar dilution, Etest® (0.5 McFarland inoculum, Mueller-Hinton agar), MicroScan® overnight and Synergies plus™; BD
Phoenix™ system. For other automated methods, check with the manufacturer about FDA-clearance to detect MICs =4 (i.e., VISA/VRSA).
5 Report to CDC by email: SEARCH@cdc.gov
More VISA/VRSA info: http://www.cdc.gov/ncidod/dhqp/ar_visavrsa.html
Streptococcus pneumoniae
Vancomycin
Susc
Int
Res
DD (mm)
≥17
-
-
MIC (µg/ml)
≤1
-
-
*CLSI states: investigate non- susceptible (NS) result
..Repeat identification and AST
..Save isolate
..Send to reference lab
Where do Clinical Labs need help
with “referral” isolates?
Resistance
VISA, VRSA
Clinical Labs need help to rapidly and
accurately….
Characterize CA-MRSA vs. HA-MRSA
(?outbreaks) – rapid result not always
essential
Verify
VRE
Test broad spectrum drugs
DRSP
Test certain drugs by MIC (systemic isolates)
MRSA
Where do Clinical Labs need help
with “referral” isolates? (con’t)?
Resistance
ESBL
Multidrug-R
GNR
Other
Clinical Labs need help to rapidly and
accurately….
Perform confirmatory test (smaller labs often
don’t do confirmatory test)
Test broad spectrum drugs
Verify unusual results (e.g., vancomycin-NS S.
pneumoniae; imipenem-R Klebsiella spp.)
AST of Referral Isolates
♦ Preferred AST method = CLSI broth
microdilution MIC reference method …or
♦ Method proven reliable by CDC
♦ Etest sometimes an option
♦ Must be aware of pitfalls of certain AST
methods to reliably resolve equivocal
results or identify emerging resistance
♦ Often requires expertise
PHL Surveillance for AR
♦ Compile AST data to answer specific question, e.g.,
– How many MRSA from EMC patients are susceptible to
trimethoprim-sulfamethoxazole? clindamycin?
♦ Collect isolates from the community and test
♦ Alternative, collect data from hospital laboratories
♦ Must consider all factors that might impact data,
e.g.,
–
–
–
–
Isolates or drugs selectively tested?
Inclusion / exclusion of duplicate isolates on a patient?
Hospitalization history?
Clonal isolates from a given facility?
CLSI M39-A2 Guideline
“Analysis and Presentation of
Cumulative Antimicrobial
Susceptibility Test Data”
Purpose: aid in preparation of
cumulative antibiogram reports to
guide empiric therapy of initial
infections.
Relative importance of requirements of AST
system depending on purpose of testing
(Hindler opinion!)…..
Requirement
Referral
Surveillance
Accuracy
5
4
Turn around time
5
1
MIC result
(vs. S I R only)
5
3 (depends on
question posed)
Cost
3
5
AST expertise
5
4
Scale = 1 (least important) to 5 (extremely important)
What resources (in addition to
CLSI standards) are available for
keeping appraised of new
developments in AST?
http://www.phppo.cdc.gov/nltn/default.aspx
NLTN
To view Jan 2006 teleconference details of M100-S16
CDC AST CD-ROM
♦ Modes of action /
mechanisms of resistance
♦ Methods
♦ Gram-positive bacteria
♦ Gram-negative bacteria
If your lab does NOT have a copy of this, please email
jhindler@ucla.edu.
www.phppo.cdc.gov/dls/master/default.asp
American Society for
Microbiology Online
Features!
http://www.asm.org/division/c/greatfeatures.htm
http://www.cdc.gov/ncidod/dhqp/pdf/ar/CAMRSA_ExpMtgStrategies.pdf
Summary of Expectations…..
PHL expects Clin Lab to
provide:
Clin Lab expects PHL to
provide:
• Isolates w/ unusual “R” (e.g.,
VRSA)
• Strategy for verification of
unusual isolates
• “New” Technical information
from CDC / APHL / Others
• Notice of training events
….maybe
• Cumulative antibiogram data
…maybe
• Regional antibiogram data
• CLSI AST standards
How can PHLs help Clinical Labs
now…for minimal $$$$?
♦ Inform them of resources for verification of
“referral” isolates
– PHL? (test in house or send to CDC)
– Local university lab or reference lab – partner???
♦ Inform them which isolates with specific R
should be reported to PH department
♦ Inform them of any emerging R bug
problem in their state rapidly!
– e.g., ceftriaxone-R Salmonella spp.
How can PHLs help Clinical Labs
now…for minimal $$$$ (con’t)?
♦Inform them of what the state PHL (or
Epi) is doing re: AR
♦Provide education on new AR and
AST issues – Many materials provided by National
Laboratory Training Network (NLTN)
– CLSI standards for AST
Meeting Antimicrobial Resistance
Challenges
PHLs (& Epi)
State Training
Coordinators
NLTN
APHL
Clin Labs
NLTN Ongoing Efforts re: AST
♦PHL-directed webcasts on AST
– March 2006 (AST systems)
– May 2006 (VISA, VRSA, MRSA)
– More planned!
♦Programs for clinical labs
♦Assist individual PHLs with specific
needs
“I would ask PHLs to look for ways that
they can become more integrated with
clinical labs and develop strategies for
ensuring accurate AST. Good
surveillance data is the lynchpin for all
our work.”
Todd Weber, MD;
Director of CDCs Office or AR
(APHL Minute, May-June, 2006)