1. What is the difference between vancomycin-susceptible S. aureus,
VISA and VRSA?
Most isolates of S. aureus are susceptible
to vancomycin. The concentration of vancomycin required to inhibit
these strains (called the minimal inhibitory concentration or MIC)
is typically between 0.5 and 2 micrograms/mL (μg/mL). In contrast, S. aureus isolates
for which vancomycin MICs are 8-16 μg/mL are classified as vancomycin
intermediate, and isolates for which vancomycin MICs are ≥32 μg/mL
are classified as vancomycin resistant. The definitions for classifying
isolates of S. aureus are based on the interpretive criteria
published by Clinical and Laboratory Standards Institute (CLSI, formerly
CLSI lists only susceptible disk diffusion interpretive criteria (in
mm) for vancomycin and Staphylococcus spp. There has not been
a sufficient number of non-susceptible isolates to develop resistant
and intermediate breakpoints. Organisms for which the vancomycin zone
diameters are ≥15mm are considered susceptible, although several studies
show that this breakpoint is unreliable for detecting VISA strains.
2. What is CLSI (formerly, NCCLS)?
is a global, interdisciplinary, nonprofit, standards-developing and
educational organization that promotes the development and use of voluntary
consensus standards and guidelines within the healthcare community.
3. What are glycopeptide-intermediate S. aureus (GISA)?
The term glycopeptide refers to a group of antimicrobial agents that
includes vancomycin and teicoplanin. Since the first two VISA isolates
in the United States were also resistant to teicoplanin, the term glycopeptide-intermediate S.
aureus (GISA) was used to indicate this broader resistance profile.
While GISA may be a more specific term for strains intermediate to both
vancomycin and teicoplanin, not all VISA strains are intermediate to
teicoplanin; therefore, VISA is a more accurate and more widely used
4. Why are VISA and VRSA isolates important?
Vancomycin continues to be a critical antimicrobial
agent for treating infections caused by S. aureus strains that
are resistant to oxacillin (MRSA; www.cdc.gov/ncidod/hip/aresist/mrsa.htm)
and other antimicrobial agents. The decreased susceptibility of VISA
and VRSA strains to vancomycin leaves clinicians with relatively few
therapeutic options for treating these infections.
5. Can routine susceptibility tests detect VISA and
Not all susceptibility testing methods detect
VISA and VRSA isolates. Two out of three confirmed VRSA isolates were
not reliably detected by automated testing systems. In addition, VISA
isolates are not detected by disk diffusion. Methods that do detect
VISA and VRSA are the vancomycin screen agar plate brain heart infusion
(BHI) agar with 6 µg/ml
of vancomycin) and non-automated MIC methods [reference broth microdilution,
agar dilution, and Etest® using a 0.5 McFarland standard to prepare
inoculum (AB Biodisk, Piscatway, NJ)]. Laboratories that use automated
methods or disk diffusion should also include a vancomycin screen plate
for enhanced detection of VISA and VRSA. If possible, laboratories should
incorporate the vancomycin agar screen plate for testing all S. aureus.
Alternatively, the screening may be limited to MRSA isolates, since nearly
all VISA and all VRSA were also MRSA. S. aureus isolates that
grow on the vancomycin agar screen plate should be checked for purity
and the species identification of the organism confirmed. Vancomycin
susceptibility should be retested by a validated MIC method incubated
for a full 24 hours.
6. What is the vancomycin agar screen test?
The vancomycin agar screen test uses commercially
prepared plates to screen pure cultures of bacteria for vancomycin
resistance. These plates contain BHI agar and 6 µg/ml of vancomycin. A 10µl
inoculum of a 0.5 McFarland suspension should be spotted on the agar
using a micropipette (final concentration=106 CFU/ml). Alternatively,
a swab may be dipped in the McFarland suspension, the excess liquid
expressed, and used to inoculate the vancomycin agar plate. For quality
control, laboratories should use Enterococcus faecalis ATCC 29212 as the susceptible
control and E. faecalis ATCC 51299 as the resistant control.
Up to eight isolates can be tested per plate; quality control should
be performed each day of testing. Growth of more than one colony is considered
a positive result. All staphylococci that grow on these plates should
be inspected for pure culture, and the original clinical isolates should
be tested by a validated MIC method for confirmation. Plates prepared
in-house using various lots of media performed inconsistently and were
inferior to those obtained commercially. Performance of commercially
prepared plates varies by individual manufacturer (CDC unpublished data).
7. Are any modifications required of routine disk
diffusion and MIC methods when testing vancomycin and staphylococci?
Testing should incorporate the following CLSI (formerly NCCLS) recommendations:
Inoculum: Use direct colony suspension
Incubation: 35°C, ambient air, for a full
Endpoint: Examine very closely for any indication of growth
According to the newest CLSI (formerly NCCLS)
standards, a vancomycin-intermediate or –resistant result for
staphylococci isolate should be verified by repeating a validated MIC
method and the organism identification.
8. What special steps should be taken when an S. aureus
is suspected of being a VISA or VRSA?
Please refer to the VISA/VRSA
testing algorithm (www.cdc.gov/ncidod/hip/lab/visa_vrsa_algo.htm),
which presents a strategy of detecting and confirming VISA and VRSA
using appropriate test methods.
9. Should repeat testing include any specific
antimicrobial agents that might not be included in the routine panel?
Yes. The following additional antimicrobial agents
should be testing against VISA and VRSA isolates. Clindamycin, daptomycin,
linezolid, quinupristin/dalfopristin, rifampin, and trimethoprim-sulfamethoxazole.
The laboratories of the Division of Healthcare Quality Promotion of the
Centers for Disease Control and Prevention will perform tests for these
additional antimicrobial agents. It is essential to send probable VISA
and VRSA as quickly as possible, even if the laboratory has the capability
to test additional agents in-house to facilitate confirmation and enhance
infection control efforts. Isolates should be sent to CDC via your local
or state health department. CDC may be notified of presumptive VISA/VRSA
by sending an email to SEARCH@cdc.gov. [S.E.A.R.C.H.](www.cdc.gov/ncidod/hip/ncidod/aresist/search.htm)
10. How should local clinical laboratories save presumptive
VISA or VRSA isolates?
It is best to freeze isolates at -60°C or lower in standard stock
culture medium, such as a broth containing 15% glycerol, or in defibrinated
sheep blood. However, if a laboratory cannot do this, the isolate should
be subcultured to a non-selective agar slant (e.g., trypticase soy agar)
and incubated overnight at 35°C. The following day, the caps should
be tightened and the slants store at 2-8°C. Repeated subcultures
should be kept to a minimum prior to storage.
11. Are VISA and VRSA isolates resistant to
All VRSA isolates to date were oxacillin-resistant (i.e., MRSA) and
contained mecA. Most VISA isolates were also oxacillin-resistant.
However, two VISA isolates became oxacillin-susceptible upon repeat isolation
from the patient and one was oxacillin-susceptible but contained mecA.
12. What are the mechanisms of resistance
for VRSA and VISA?
All VRSA isolates to date contained the vanA vancomycin resistance
gene. The vanA gene is usually found in enterococci and typically
confers high-level vancomycin resistance (MICs= 512-1024µg/ml)
to these organisms. Vancomycin-resistant enterococci containing vanA
were isolated from all patients in addition to MRSA. It is likely that
the vanA determinant was transferred via plasmids or transposons from
enterococci to a resident MRSA strain, resulting in the VRSA.
The mechanism of decreased vancomycin susceptibility in VISA strains
is not fully understood. VISA cells have thicker cell walls that contain
many cell wall subunits capable of binding vancomycin extracellularly.
Vancomycin must reach the cell membrane and bind to the growing cell
wall complex to inhibit cell growth.
13. Should VISA and VRSA be reported to the
infection control team?
Yes. There is significant concern about the spread of VISA and VRSA
among patients because of limited treatment options. If a VISA or VRSA
is suspected, specific infection control precautions need to be initiated
by infection control personnel to decrease the risk of transmission to
others. It is critical for laboratory workers to contact the infection
control team immediately when a VISA or VRSA is suspected. Additionally,
laboratories should notify the local and/or state health department and
the Division of Healthcare Quality Promotion, National Center for Infectious
Diseases, CDC, by telephone 800-893-0485 or by sending an email to SEARCH@cdc.gov.
The isolate should be saved and sent to the health department and CDC
for confirmatory testing.
14. Where can I learn more about antimicrobial
CDC developed a training tool for laboratorians to
enhance their understanding and improve their proficiency in performing
antimicrobial susceptibility testing
15. How do I report a VISA or VRSA to the
To report or request testing of suspected VISA or VRSA isolates, send
an email to SEARCH@cdc.gov with your contact information (i.e., name,
title, telephone number, laboratory or facility name, and a description
of your testing methods and results facility and/or laboratory name,
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