CLSI AST Update Gram Positive Bacteria_1

Susan E. Sharp, Ph.D., DABMM, FAAM Director - Regional and Sunnyside Medical Center Laboratories

Director - Regional Clinical Microbiology Kaiser Permanente

Associate Professor - Department of Pathology Oregon Health & Sciences University

Portland, OR

Update on the CLSI Standards for

Antimicrobial Susceptibility Testing:

What’s New with the Gram Positive Organisms?

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http://www.swacm.org/index.htm

OBJECTIVES

•Review significant changes made in 2012.

•Review this year’s 2013 CLSI M100-S23 document.

•Describe any specific susceptibility testing

methodologies.

•Present new antimicrobials.

•Discuss new developments for the January 2014 CLSI

M100-S24 guideline.

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CLSI AST Standards – January 2013

M100-S23 Tables (2013)*

M02-A11 Disk Diffusion Method (2012)^

M07-A9 MIC Method (2012)^

* M100 updated every year

^ M02, M07 updated every 3 years

Summary of Major Changes

Changes to CLSI documents are summarized in the front of each document.

Information listed in boldface type is new or modified since the previous edition of M100 document.

Recent breakpoint addition/revision dates are listed in the front of M100-S23.

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Today’s Review:

2012 – Significant changes from last year

2013 – Changes/New items in these guidelines

2014 – What’s on the horizon

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Significant changes in 2012

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Staphylococcus species -2012

Penicillin testing

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Staphylococcus spp. – Penicillin - 2012

The story…..

> 90% of staphylococci are penicillin “R”

Penicillin rarely considered for treatment of staphylococcal infections

– …BUT - Penicillin might be considered for infections requiring lengthy therapy (e.g., endocarditis, osteomyelitis) - IF penicillin were known to be “S”

Some Staphylococcus spp. that test “S” to penicillin by MIC or disk diffusion may actually possess a β-lactamase (BL) that may cause the patient to fail penicillin therapy

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Staphylococcus spp. – Penicillin - 2012

CLSI Previous recommendation (2011):

– Perform induced nitrocefin BL test before reporting penicillin as “S” if:

zone diameter ≥29 mm

MIC ≤0.12 µg/ml

– PCR for the blaZ BL gene may be considered

Penicillin Breakpoints

MIC (µg/ml) Zone (mm)

S I R S I R

≤0.12 - ≥0.25 ≥29 - ≤28

9 Reference: M100-S21 (2011) - Table 2C. Page 70

Induced ß-lactamase (BL) Test - 2012

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-Subculture isolate to blood agar

-Drop disk to induce BL production

(e.g., oxacillin or cefoxitin)

-Incubate overnight

-Test cells from periphery of zone

-If BL positive, report penicillin R

Pos Neg


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-Incubate overnight



Pos Neg


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-Incubate overnight



Pos Neg

Staphylococcus aureus (BL) - 2012

Induced nitrocefin BL test usually, but not always, detects staphylococcal BLs

Other BL tests are more sensitive for the detection of BL:

– Cloverleaf test

– Penicillin disk zone edge test

blaZ gene PCR not optimal for BL

– blaZ codes for BL production

– Several types of blaZ genes

all types may not be detected by a single PCR assay

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Staphylococcus aureus β-lactamase (BL) Study* - 2012

348 MSSA (<4 g/ml PEN MICs) characterized for blaZ by PCR:

– 303 PCR negative

– 45 PCR positive (~13%)

Methods:

– Phenotypic BL tests

Nitrocefin - Cefinase

Nitrocefin - Dryslide

Cloverleaf assay

Penicillin disk zone edge

14 *Statens Serum Institut (Denmark), CDC (Atlanta), MGH (Boston)

Staphylococcus aureus

BL Study - 2012

Pen MIC

(µg/ml)

blaZ functional

Neg Pos

0.008 2

0.016 15

0.032 180 1

0.06 90 5

0.12 15 17

0.25 1 14

0.5 4

1.0

2.0 2

4.0 1

303 45

• 1 blaZ neg and penicillin “R”

• 23 blaZ pos and penicillin “S”

Reference: CLSI Agenda Book January 2011.

S

R

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Cloverleaf Assay

for BL in S. aureus

• 5% sheep blood agar

• S. aureus ATCC 25923 as the indicator organism

• 1 unit penicillin disk

• Negative (pen-S) strain QC

• Some difficulties reading

Reference: CLSI Agenda Book January 2011.

Isolates A-D are all

BL positive

A

B

C

D

BL negative

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β-lactamase positive

β-lactamase negative

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Staphylococcus aureus - 2012 Disk Zone Edge Test (10 U penicillin disk and standard disk diffusion method)

Fuzzy / “beach” =

β-lactamase negative,

Penicillin - S

Sharp / “cliff” =

β-lactamase positive,

Penicillin - R

S. aureus

supplemental QC:

Neg - ATCC 25923

Pos - ATCC 29213

Reference: M100-S22. Table 2C Supplemental Table 1. Page 83

Staphylococcus aureus 3 Lab BL Study Results (N=348), 2012

Test Sensitivity Specificity

Cefinase 77% 100%

Dryslide 88% 100%

Cloverleaf 100% 100%

Penicillin disk zone edge 96% 100%

19 Reference: CLSI Agenda Book January, 2011

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Penicillin MIC ≤0.12 µg/ml

Nitrocefin β-lactamase

positive

Report penicillin “R”

Nitrocefin β-

lactamase negative

Perform penicillin disk zone-edge test

≥ 29 mm fuzzy

Report penicillin “S”

≥ 29 mm sharp

Report penicillin “R”

Staphylococcus aureus

Penicillin (MIC ≤0.12 µg/ml) Reporting, 2012

Note: If doing disk diffusion routinely, just examine zone edge for those with zone sizes of > 29mm.

M100-S22. Table 2C Supplemental Table 1. Page 80

A A B

– 2012 NEW RECOMMENDATION:

Added ‘penicillin disk zone edge test’ for BL

production in S. aureus

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Staphylococcus spp. –

Penicillin 2012

Staphylococcus spp. –

Penicillin Optional Strategy - 2012

Report penicillin if “R”

Suppress penicillin if “S” and consider a note: “Contact laboratory if penicillin results needed.”

– If penicillin “S” and penicillin results needed, perform:

Nitrocefin BL test , and if negative

– Penicillin zone edge test

» Beach = PEN S

» Cliff = PEN R

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S. aureus

Isolates where penicillin zones are ≥29 mm or penicillin MICs are ≤0.12 µg/ml, perform a penicillin ‘disk zone edge test’ before reporting as penicillin susceptible.

NOTE:

– S.lugdunensis isolates where penicillin zones are ≥29 mm or penicillin MICs are ≤0.12 µg/ml, perform an induced nitrocefin assay or other CLSI reference method on isolates before reporting as penicillin susceptible.

– The penicillin disk zone edge test was shown to be inferior as compared to the induced nitrocefin assay and should not be used with S.lugdunensis.

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Action Items - 2012

Staphylococcus – Oxacillin 2012

Intermediate

–Table 2C / Note (13)

If oxacillin-I results (disk diffusion testing) are obtained for S.aureus, perform testing for mecA or PBP 2a, the cefoxitin MIC or cefoxitin disk test, an oxacillin MIC test, or the oxacillin-salt agar screening test. Report the result of the alternative test rather than the oxacillin-I result.

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Staphylococcus – Oxacillin 2012

Resistance

–Table 2C / Note (12)

If oxacillin-R staphylococci report penicillin as resistant or do not report.

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Staphylococcus - 2012

Disks per plate – clarification

– 12 disks only on a 150mm plate


– Do not measure zone of inhibition of hemolysis

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Enterococcus – Vancomycin - 2012

For isolates with MICs of 8-16 g/ml (I)

– Perform tests listed in 2D-Supplemental Table 1

2D-Supplemental Table 1

– Motility

– Pigment

E.gallinarum (non-pigmented, motility +)

E.casseliflavus (pigmented yellow, motility +)

Other Enterococcus (non-pigmented, non-motile)

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Enterococcus – Vancomycin 2012

Alternative inoculum method provided for vancomycin resistance screen test

– 2D-Supplemental Table 1

OLD: 1-10 L of a 0.5 McFarland suspension spotted onto agar surface (agar = 6 g/ml vanco in BHI agar)

NEW (added): Alternatively, using a swab dipped in the suspension and the excess liquid expressed, spot an area 10-15mm in diameter or streak a portion of the plate.

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Streptococcus pneumoniae - 2012

Predicting susceptibility to Fluoroquinolones

– Isolates susceptible to levofloxacin are predictably susceptible to gemifloxacin and moxifloxacin.

– Isolates susceptible to gemifloxacin or moxifloxacin can not be assumed to be susceptible to levofloxacin.

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Streptococcus pneumoniae & -Streptococcus – 2012

Disks per plate – clarification



– Do not measure zone of inhibition of hemolysis

(for Viridans streptococci as well)

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-Streptococcus - 2012

Table 2H-1 Supplemental Table 1

(inducible clindamycin resistance)

– Included new comment regarding CDC recommendations:

“The 2010 CDC guidelines on prevention of group B streptococcal disease in neonates recommend that colonization isolates from pregnant women with severe penicillin allergy (high risk for anaphylaxis) should be tested for inducible clindamycin resistance.”

Use erythromycin & clindamycin for D zone testing, but do not report erythromcyin.

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** If an isolate is resistant to erythromycin, it might have inducible resistance to clindamycin, even if it appears susceptible to clindamycin. If a GBS isolate is susceptible to clindamycin, resistant to erythromycin, and testing for inducible clindamycin resistance is negative, then clindamycin can be used for GBS intrapartum prophylaxis instead of vancomycin.

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2010 CDC Guidelines

Check for inducible

clindamycin R if:

erythromycin-R

clindamycin-S

But only report

clindamycin!

www.cdc.gov

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ASM’s Clinical Microbiology Portal : http://clinmicro.asm.org/

36 | © 2011 Kaiser Foundation Health Plan, Inc. For internal use only. January 8, 2014

http://clinmicro.asm.org/



GPB Protocols jointly developed by ASM (CLP) and CDC.


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protocols



Prenatal Screening for Group B Streptococcus: Non-pigmented broth, subculture only


-Streptococcus - 2012

Table 2H-1

– Daptomycin

Disk diffusion testing is not reliable

(previously indicated for the staphylococci)

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New antibiotics - 2012

Doripenem

Ceftaroline

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Doripenem (Doribax)

A broad spectrum injectable antibiotic

A -lactam drug

Belongs to the carbapenem group (imipenem, ertapenem, meropenem)

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Doripenem

Complicated Intra-Abdominal Infections

Indicated as a single agent for the treatment of complicated intra-abdominal infections caused by Escherichia coli, Klebsiella pneumoniae, Pseudomonas aeruginosa, Bacteroides caccae, Bacteroides fragilis, Bacteroides thetaiotaomicron, Bacteroides uniformis, Bacteroides vulgatus, Streptococcus intermedius, Streptococcus constellatus and Peptostreptococcus micros.

Complicated Urinary Tract Infections, Including Pyelonephritis

Indicated as a single agent for the treatment of complicated urinary tract infections, including pyelonephritis caused by Escherichia coli including cases with concurrent bacteremia, Klebsiella pneumoniae, Proteus mirabilis, Pseudomonas aeruginosa, and Acinetobacter baumannii.

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Doripenem

Exerts its bactericidal activity by inhibiting bacterial cell wall biosynthesis.

Inactivates multiple essential penicillin-binding proteins (PBPs) resulting in inhibition of cell wall synthesis with subsequent cell death.

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Doripenem

Bacterial resistance mechanisms that affect doripenem include:

– Inactivation by carbapenem-hydrolyzing enzymes

KPC, NDM-1, etc.

– Mutant or altered PBPs

– Decreased outer membrane permeability

– Active efflux

Doripenem is stable to hydrolysis by most BL, including penicillinases and cephalosporinases produced by GP &GN bacteria

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Doripenem (Gram positive’s)

Staphylococcus aureus (MSSA only)

Streptococcus agalactiae

Streptococcus pyogenes

Streptococcus Viridans group

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S ( g/ml) I ( g/ml) R ( g/ml)

Streptococcus Viridans group (O) < 1.0

- -

-Streptococcus (O) < 0.12 - -

S.pneumoniae (O)

< 1.0 - -

S.aureus*

- - -

Doripenem (Gram positives)

*Remember -

Only penicillin, oxacillin (cefoxitin), ceftaroline for staph with the -lactam drugs in 2013.

(No disk diffusion criteria)

Ceftaroline (Teflaro)

Ceftaroline is a cephalosporin with in vitro activity against GP and GN bacteria.

Bactericidal action is mediated through binding to essential penicillin-binding proteins (PBPs).

Bactericidal against S. aureus due to its affinity for PBP2a and against Streptococcus pneumoniae due to its affinity for PBP2x.

Ceftaroline is not active against Gram negative bacteria which produce ESBLs or carbapenemases.

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Ceftaroline

Acute Bacterial Skin and Skin Structure Infections

Indicated for the treatment of acute bacterial skin and skin structure infections caused by susceptible isolates of Staphylococcus aureus (including methicillin-susceptible and -resistant isolates), Streptococcus pyogenes, S.agalactiae, E.coli, K.pneumoniae, and K.oxytoca.

Community-Acquired Bacterial Pneumonia

Indicated for the treatment of community-acquired bacterial pneumonia caused by susceptible isolates of the following microorganisms: Streptococcus pneumoniae, Staphylococcus aureus (MSSA only), Haemophilus influenzae, Klebsiella pneumoniae, K.oxytoca, and E.coli.

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Ceftaroline

Staphylococcus aureus (MSSA & MRSA)

Streptococcus pyogenes

Streptococcus agalactiae

Streptococcus pneumoniae

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Ceftaroline vs. Other β-lactams agents % Susceptible

Organism N Ceftaroline Ceftriaxone Imipenem

MSSA 186 100 100 100

MRSA 215 100 0 0

S.pneumoniae 894 100 90.8 NA

NA - not available

REF: Jones et al. 2011. J Antimicrob Chemother. 66 (Suppl 3):iii69–iii80.

Ceftaroline

S

( g/ml) / mm

I

( g/ml) / mm

R

( g/ml) -mm

S.aureus

(B)

< 1 / > 24 2 / 21-23 > 4 / < 20

-Streptococcus

(C)

< 0.5 / > 26 - -

S.pneumoniae

(nonmeningitis)

(C)

< 0. 5 / > 26 - -

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NEW FOR 2013 !

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All cephalosporins/many penicillins in the 2012 Table 2C were removed.

– Deleted all β-lactam breakpoints except penicillin, oxacillin [cefoxitin], and ceftaroline.

A statement is provided to indicate that results for cephalosporins and other -lactam antibiotics can be predicted from the results of penicillin, oxacillin MIC, cefoxitin MIC, or cefoxitin disk diffusion testing.

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Staphylococcus spp.

M100-S22. Table 2C.

Staphylococcus spp.

2013 - Eliminated breakpoints for:

Penicillins

BL/BL inhibitor combos

Cephalosporins

Carbapenems

M100-S22. Table 2C.


Rationale for deleting breakpoints for -lactams (except pencilllin, oxacillin [cefoxitin], & ceftaroline) from the CLSI M100 tables for staphylococci:

– Current breakpoints are most likely inaccurate

They were ‘Grandfathered’ into the staphylococcal tables with other major table over-hauls in the early 2000’s.

Can deduce all anti-staphylococcal -lactam results from penicillin and oxacillin [cefoxitin] results.

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Oxacillin disk diffusion testing has been removed form the staphylococci charts.

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“Oxacillin disk testing is not reliable. For disk

testing see cefoxitin for reporting oxacillin…”

Staphylococcus spp. - 2013 β-Lactam Breakpoints Remaining

Penicillin

– Represents penicillinase-labile penicillins

Oxacillin (MIC only; no more DD)

– Represents penicillinase-stable penicillins

Cefoxitin

– Surrogate for oxacillin

Ceftaroline (added 2013)

– Cephem with anti-MRSA activity

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Staphylococcus spp. - 2013 Penicillins and Penicillinases

Penicillinase-labile Penicillins

(penicillin represents)

Penicillinase-stable Penicillins

(oxacillin represents)

amoxicillin

ampicillin

carbenicillin

mezlocillin

penicillin

piperacillin

ticarcillin

cloxacillin

dicloxacillin

flucloxacillin

methicillin

nafcillin

oxacillin

Staphylococcus - β-Lactams - 2013 Use Pen and Oxa (FOX) results to predict results for other ß-lactams

Test Results

Predicts Penicillin

Oxacillin (cefoxitin)

S S

Susceptible to: All penicillins ß-lac / ß-lactamase inhibitor combos Cephems Carbapenems

R S

Resistant to: Penicillinase-labile penicillins Susceptible to: Penicillinase-stable penicillins ß-lac / ß-lactamase inhibitor combos Cephems Carbapenems

R R Resistant to: All ß-lactams (except cephems with anti- MRSA activity, e.g., ceftaroline)


Detection of oxacillin resistance:

In most staphylococcal isolates, oxacillin resistance is mediated by mecA-encoding the penicillin-binding protein 2a (PBP 2a, also called PBP2‘).

Other mechanisms of oxacillin resistance are rare and include a novel mecA homologue (eg, mecC)REF which may not be detected by tests for mecA or PBP 2a.

Isolates that test positive for mecA or PBP 2a should be reported as oxacillin resistant.

If both cefoxitin and oxacillin are tested against S.aureus or S.lugdunensis, and either result is resistant, the organism should be reported as oxacillin resistant.

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REF: Stegger M, Andersen PS, Kearns A, Pichon B, Holmes MA, Edwards G, Laurent F, Teale C, Skov R, Larsen AR.

Rapid detection, differentiation and typing of methicillin-resistant Staphylococcus aureus harboring either mecA or the

new mecA homologue mecA(LGA251). Clin Microbiol Infect. 2012;18(4):395-400 .


Table 1A (Test and Report) and Table 2C (Interpretive Standards): 2013

For doxycycline and minocycline: added not to report on organisms isolated from the urinary tract.

Removed telithromycin from Table 1A due to black box warning from FDA; and changing Test/Report Group from ‘B’ to ‘O’ in Table 2C.

Added footnote to indicate that daptomycin should not be reported for isolates from the lower respiratory tract.

Removed quinupristin/dalfopristin from Table 1A as it is not FDA cleared for MRSA or CoNS; stating that for isolates of MSSA there are much better drugs to use for treatment with less toxicity. Changing Test/Report Group from ‘C’ to ‘O’ in Table 2C.

Removed amikacin, kanamcin, netilimicin and tobramycin from Table 1A. Only gentamicin will remain as ‘C’ in Table 2C (others changed to ‘O’).

– Added note to aminoglycoside-S isolates stating that they are to be used only in combination with other active agents.

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Streptococcus pneumoniae - 2013

New (revised) tetracycline disk diffusion and MIC interpretive criteria.

New doxycycline disk diffusion and MIC interpretive criteria.

Clarified that isolates of S. pneumoniae from CSF can also be tested against vancomycin using either a MIC or disk method.

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Inducible clindamycin resistance - S.pneumoniae: 2013

The clinical significance of this mechanism of clindamycin resistance is not known for S.pneumoniae;

– but inducible clindamycin resistance can be detected using the D-zone test and is now be included in the 2013 CLSI documents.

The 2013 document includes: “IF testing for clindamycin resistance in S.pneumoniae is performed, it should include screening for inducible clindamycin resistance.”

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NEW AST QC Guidance - 2013

Table 3C. Disk Diffusion: Reference Guide to QC Frequency

Conversion from Daily to Weekly QC

Routine QC is performed each day the test is performed unless an alternative quality control plan has been established.

CLSI document M02-A11 Section 15.7 describes a QC plan using a 20-30 day protocol that if successfully completed allows a user to convert from daily to weekly quality control.

– < 1/20 or < 3/30 weekly QC testing

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NEW AST QC: 3x5 (15) Plan - 2013

An alternate QC plan using a two-phase, 15 replicate (3 X 5 day) plan is described in Table 3C as follows:

– 15 replicate (3 X 5 day) plan

Test three replicates using individual inocula preparations of the appropriate QC strains for 5 consecutive test days to perform 15 replicates (3 x 5 day) plan.

Each QC strain tested is evaluated separately according to the acceptance criteria and recommended action described below (e.g., pass / test another 3 replicates for 5 days / fail).

Upon successful completion of the QC plan the laboratory can convert from daily to weekly QC testing.

If unsuccessful investigate, take corrective action as appropriate and continue daily QC testing.

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NEW AST QC 3x5 (15) - 2013

Table 3C*

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Number out of range

with initial testing

(based on 15 replicates)

Conclusion from initial

testing

Number out of range

after repeat testing

(based on all 30

replicates)

Conclusion after repeat

testing

0-1

QC plan successful.

Convert to weekly QC

testing.

NA

NA

2-3

Test another 3

replicates for 5 days.

2-3

QC plan successful.

Convert to weekly QC

testing.

4 or greater

QC plan fails.

Investigate and take

corrective action as

appropriate. Continue

QC each test day.

4 or greater

QC plan fails. Investigate

and take corrective action

as appropriate. Continue

QC each test day.

*Assess each QC strain individually

NEW AST QC 3x5 (15) - 2013

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Test 3 replicated of each QC

strain for 5 days using individually

prepared inoculum

Fail.

Continue to

include QC

each test

day. Take

corrective

action. Test another 3

replicates for 5

days

Pass. Convert

to weekly QC.

Pass. Convert

to weekly

QC.

0-1 of 15 out

of range?

2-3 of 15 out

of range?

> 4 of 15 out

of range?

2-3 of 30 out

of range?

> 4 of 30 out

of range?


Statistician’s comments:

– 3x5 Plan

Similar to manufactured product releases

– ‘Go’ or ‘No-Go’ based on mathematical considerations

– Two-Stage sampling plan:

May be completed in first stage or proceed to a second stage

Two new plans were considered:

– 0-1 error allowed in first stage of Plan 1

– 0 errors allowed in first stage of Plan 2

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Statistician’s comments

– Out-of-control results could be due to either systemic or random errors

– Systemic errors = likely to get >2 outliers out of 15 results

– Random (allowable) errors = very high probability of getting 1 outlier of 15 results due to random error

Plan 1: 0-1 errors allowed:

– Deemed likely to pick up systematic errors (>2/15)

Plan 2: 0 errors allowed:

– Deemed likely to be problematic and unlikely to improve quality of results (no allowance for random errors @ <1/15)

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Conversion from Daily to Weekly QC - 2013

In addition, this testing is required for the following modifications of existing antimicrobial susceptibility test system:

– Addition of new antimicrobial agent to existing system

ceftraoline, doripenem

Can use 20-30 day consecutive days of QC testing or the 3x5 QC plan

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QC Recommendations:

‘Routine’

– Test negative (susceptible) QC strain:

With each new lot/shipment of testing materials (eg, disks, or agar plates used for agar dilution, or single wells or tubes used with broth dilution methods).

Weekly if the screening test is performed at least once a week and criteria for converting from daily to weekly QC testing have been met (see Section 15.7.2.1 in M02 or Section 16.7.2.1 in M07).

Daily if the screening test is performed less than once per week and/or if criteria for converting from daily to weekly QC testing have not been met (see bullet above).

‘Lot/shipment’

– Test positive (resistant) QC strain at minimum of at least once with each new lot/shipment of testing materials.

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NEW QC Testing Frequency: Screening Tests (Table 2D-Supplemental Table 1) – 2013 (ex:HLAR)

NEW QC Testing Frequency: Screening Tests - 2013

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QC Strain Previous New 2013

Positive (resistant) Daily; convert to weekly

after 20-30 days/3x5 plan

Each new

batch/lot/shipment of

testing materials*

Negative (susceptible) Daily; convert to weekly


Daily; convert to weekly


*Applies to disks, agar plates used for agar dilution, or single wells or tubes

used with broth dilution methods.

Intrinsic Resistance Table – 2013

Intrinsic Resistance (Appendix B):

Split out to four appendixes as follows:

B.1 Enterobacteriaceae

– Added imipenem with note that Proteus species, Providencia species and Morganella species may have elevated MICs by mechanisms other than by production of carbapenemases. Isolates that test “S” should be reported as “S”.

– Added in a Note 2 information that Enterobacteriaceae are also intrinsically resistant to clindamycin, daptomycin, fusidic acid, glycopeptides (vancomycin, teicoplanin), linezolid, macrolides (erythromycin, clarithromycin, azithromycin), quinupristin-dalfopristin, and rifampin.

New Appendix B.2 Non-Enterobacteriaceae

New Appendix B.3 Staphylococci

New Appendix B.4 Enterococcus spp.

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Intrinsic Resistance Tables – 2013 Staphylococcus (Appendix B) – 2013

Organism / Drug Novobiocin Fosfomycin Fusidic Acid

S.aureus/S.lugdunensis There is no intrinsic resistance in these species.

S.epidermidis There is no intrinsic resistance in this species.

S.haemolyticus There is no intrinsic resistance in this species.

S.saprophyticus R R R

S.capitis R

S.cohnii R

S.xylosus R

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Note 1: Gram-positive bacteria are also intrinsically resistant to aztreonam, polymyxin B/colistin and

naladixic acid.

Note 2: Oxa-R SA and CoNS are considered R to other BL agents, ie, pen, BL/BL inhibitors, cephems

(exc. ceftraoline), and carbapenems.

Organism/drug Cephalo-

sporins

Vanco-

mycin

Teico-

planin

Amino-

glycosides

Clinda-

mycin Q/D

Trimetho-

prim SXT

Fusidic

acid

E.faecalis R* R* R* R R* R R

E.faecium R* R* R* R* R R

E.gallinarum/c

asseliflavus

R* R R* R* R R* R R

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Intrinsic Resistance Tables – 2013 Enterococcus (Appendix B) – 2013

*Warning: For Enterococcus spp., cephalosporins, aminoglycosides (except

for high-level resistance screening), clindamycin, and SXT may

appear active in vivo, but are not effective clinically and should

not be reported as susceptible.

NOTE 1: Gram-positive bacteria are also intrinsically resistant to aztreonam,

polymyxin B/colistin and naladixic acid.

NEW FOR 2014 !

I think……

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2014

Not a lot in the way of new Gram Positive guidelines - but we’ll review the following:

– Vancomycin DD with staphylococci

– New drugs added

– New QC recommendations

(you’re gonna like these!)

– “S-DD”

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Vancomycin / Staphylococci - 2014

Table 2C:

– Vancomycin disk diffusion

– Previously you could use it for detecting “R”

VRSA due to the vanA gene will show no zone of inhibition around a 30- g vancomycin disk

Any isolate with > 7mm zone must be tested with a MIC method before reporting as “S”

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- Staphylococcus species

No DD criteria !

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NEW DRUGS - 2014

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Ceftazidime/avibactam - 2014

Avibactam is a synthetic β-lactamase (BL) inhibitor

– Addition of avibactam greatly improves the activity of ceftazidime (4-1024x MIC) against most of the Enterobacteriaceae.

– Avibactam also improves the activity of ceftazidime to P.aeruginosa (~4x MIC).

– Avibactam does not improve the activity of ceftazidime against Acinetobacter spp. or to most anaerobic bacteria (exceptions: B.fragilis, C.perfringens, Prevotella and Porphyromonas spp.).

– Data suggest that ceftazidime-avibactam is rapidly active against BL-producing GNRs that are not inhibited by ceftazidime alone.

– Ceftazidime-avibactam is as effective as carbapenems in complicated intra-abdominal infection and complicated urinary tract infection.

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Ceftazidime/avibactam - 2014

– Avibactam serves to broaden the spectrum of ceftazidime against BL-producing GNRs.

– Potential future roles include the treatment of infections caused by GNR producing ESBLs, KPCs, and/or AmpC BL.

– It may be used in combination (with metronidazole) for suspected polymicrobial infections.

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NEW DRUGS - 2014

Ceftazidime/avibactam

Ceftaroline/avibactam

Aztreonam/avibactam

Biapenem (a new carbapemen drug)

– Ceftolozane/tazobactam

– Developing for the treatment of certain serious GNR infections, including those caused by MDR-P.aeruginosa.

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Quality Control - 2014

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Quality Control - 2014

Continuing to look at ways to decrease routine QC when adding a new drug:

– < 1/20

– < 3/30

– 3x15

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Decreasing daily QC testing - 2014

CLSI QC DD Working Group Project

Background – CLSI guidance recommends that prior to initiating testing with a new antibiotic, 20-30 day QC studies must be performed (and now we have the 3x5 testing as well).

The intent of this current project is to evaluate the necessity of performing a full 20 or 30 days of testing.

The goal is to assimilate a large amount of 20-30 day QC study data from different institutions with a variety of drug/bug combinations.

– Analyze the data to determine when, during the course of the 20-30 day study, QC failures occur.

– Hypothesize that the systemic problems with QC will reveal themselves early and that extended testing (ie., 20-30 days) is not necessary.

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Preliminary Data* – 63 total preliminary QC studies have been accumulated thus far from 2 institutions (with the majority of the data coming from a single institution).

– 5 of these studies were conducted for 30 days, 25 for 21 days, and 33 for 19 days.

– A total of 6 failures over 1,302 data points have been identified.

day 1 (n=3), day 2 (n=1), day 14 (n=1) and day 22 (n=1).

(*From Chris D. Doern, PhD, D(ABMM) on behalf of ASM’s Committee on Laboratory Practices)

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Day of Testing

QC Failures by Day of Testing



– More data is required - but these findings suggest that it may be possible to reduce the number of days required for QC studies.

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Currently = The M2/M7 text and flowchart clearly indicates that you must perform 5 consecutive days of repeat QC “if” you can not attribute the out-of-range result to a specific issue.

– If you can contribute it to a particular issue (e.g., contamination, used the wrong QC strain, etc.), you only have to do one repeat and if ‘in’, you may continue weekly QC testing.

However, the above does not address the 0-5% of out-of-range results which would be expected due to random error (as ranges are established based on > 95% agreement).

– One suggestion is to use retrospective data from the same lot of materials to satisfy the 5 replicate repeat. This would not be a "statistical" change, only a change in approach.

Also considering to allow for multiple replicates in one day (up to 3) to determine more quickly if there is a problem (as laboratories may still be testing patient specimens while troubleshooting).

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Out-of-Range QC – 2014


Add a new section to 3C and 4F (QC frequency) - Troubleshooting out-of-range results:

If the cause can be identified –

– Correct the issue, document the reason, and retest the strain on the day the error is observed (or as soon as possible if need to subculture a new QC strain first).

– If the repeated result is within range, no further corrective action is required.

If the cause can not be identified -

– If the 4 previous QC results have been acceptable using the same lots of materials (e.g., agar plates, disks, MIC trays/plates/panels), these 4 results, along with the current repeat, may be used to retrospectively satisfy the requirement for 5 repeats.

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If the cause can not be identified. Example :

– If the 4 previous QC results were acceptable and you now obtain an out-of-range result, repeat the QC as soon as possible and if this repeat is within range, the corrective action was successful and you can resume weekly testing.

If the repeat is out-of-range you must take additional corrective action.

Daily QC must be continued until the problem is resolved.

– If 4 previous QC results were not acceptable or not available (e.g., new lot recently put into use), test sufficient QC replicates to satisfy the requirement for a total of 5 results.

Up to 3 QC replicates can be tested / day if individual inocula are used.

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Week Day Lot # Results Action

1 1 1234 4

2 1 1234 8

3 1 1234 8

4 1 1234 4

5 1 1234 16 Out of range. Repeat QC testing same day.

5 2 1234 8 In range. 5/6 acceptable in range QC tests for

E.coli ATCC 25922 and ampicillin with lot 1234.

Resume weekly QC testing.

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Using retrospective QC to correct a random error if no error is readily apparent:

If you have an out of range QC result upon weekly QC testing, you will be able to

repeat the QC once if in range and if the last 4 times you previously tested were also

in range you can go back to weekly QC testing. You must have used the same lot of

all materials for all test results (example below).

Example 1 - E.coli ATCC 25922 ampicillin: range 2-8 ug/ml


1 1 4567 4

2 1 4567 8

3 1 4567 16 Out of range. Repeat QC testing same day.

3 2 4567 4 In range. 3 acceptable in range QC tests for E.coli

ATCC 25922 and ampicillin with lot 4567. Repeat QC for

2 more consecutive days.

3 3 4567 8 In range.

3 4 4567 8 In range. 5/6 acceptable in range QC tests for E.coli

ATCC 25922 and ampicillin with lot 4567. Resume

weekly QC testing.

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You may also repeat the QC testing prospectively to do as many as you need to

get the 1/6 to be in range using same lots # of all testing materials (e.g.; 2

retrospective and 3 prospective). If all within range, you and can go back to weekly

QC. You must use the same lot of all materials (example below).



1 1 7891 4

2 1 7891 8

3 1 7891 16 Out of range. Repeat QC testing x3 same day

from different starting dilutions.

3 2x1 7891 4 3 replicates from different starting dilutions

are in range. 5/6 acceptable in range QC tests

for E.coli ATCC 25922 and ampicillin with lot

7891. Resume weekly QC testing.

3 2x2 7891 8

3 2x3 7891 8

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You may also do up to 3 replicates per day with same lots of

materials using different starting dilutions (example below).


Using retrospective data to go to weekly testing:

– If you have been performing QC testing of a drug each time you test patient organisms, you will be able to use those contemporary (within the last year) retrospective results in order to collect data that can be used for going to weekly QC testing.

– For example, if you have QC tested a drug approximately 2x/month for the last year, you can use the most recent 20 QC tests to document your ability to go to weekly QC testing for the drug.

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Streamlined QC – 2014

Streamlined QC 2014

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Date

tested

LOT # disk/MHA

Results OK

(Y/N)

11/13/12 123/258 Y

12/12/12 123/456 Y

12/22/12 123/456 Y

1/1/13 123/003 Y

1/15/13 456/003 Y

1/29/13 456/003 Y

2/5/13 456/225 Y

2.26.13 456/225 Y

3/3/13 456/258 Y

3/26/13 456/258 Y

4/16/13 456/258 Y

4/20/13 788/459 Y

5/1/13 788/459 Y

5/17/13 788/560 Y

6/6/13 788/560 Y

6/18/13 788/777 Y

7/3/13 788/777 Y

7/30/13 788/889 Y

8/16/13 788/889 Y

8/25/13 799/889 Y

20 days consecutive QC testing days

= all acceptable document and

begin weekly QC

Routine QC recommendations/organism:

– Those that appear for each organism group will be streamlined.

– These new guidelines can be used for both weekly QC as well as individual drug QC.

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Streamlined QC – 2014

P.aeruginosa : Table 2B-1

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CURRENT:

Routine QC Recommendations (See Tables 3A and 4A for acceptable QC ranges.)

Escherichia coli ATCC® 25922

Pseudomonas aeruginosa ATCC® 27853

Escherichia coli ATCC® 35218 (for -lactam/ -lactamase inhibitor combinations)

NEW for 2014:





Acinetobacter : Table 2B-2

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CURRENT:




Escherichia coli ATCC® 35218 (for -lactam/ lactamase inhibitor combinations)

NEW for 2014:



Escherichia coli ATCC® 25922 (for tetracyclines and SXT)

Escherichia coli ATCC® 35218 (for -lactam/ lactamase inhibitor combinations)

Burkholderia : Table 2B-3 Stenotrophomonas : Table 2B-4

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CURRENT:





NEW for 2014:



Escherichia coli ATCC® 25922 (for chloramphenicol, minocycline, and SXT)


Other non-Enterobacteriaceae : Table 2B-5

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CURRENT:





NEW for 2014:



Escherichia coli ATCC® 25922 (for chloramphenicol, tetrayclines, sulfonamide and SXT)


Enterobacteriaceae : Table 2A

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CURRENT:




NEW for 2014:




Pseudomonas aeruginosa ATCC® 27853 (carbapenems)

“S-DD”

Enterobacteriaceae and cefepime

ISSUE: When the cephalosporin breakpoints to the Enterobacteriaceae were change (lowered) in 2010, cefepime was also reviewed but the committee at that time left the breakpoint unchanged at < 8 / 16 / > 32 ug/ml.

RESULT: The committee after MUCH discussion voted to change cefepime

breakpoints for the Enterobacteriaceae from current to the below:

S < 2 ug/ml

SDD 4-8 ug/ml (SDD = susceptible, dose-dependent*)

R > 16 ug/ml


“S-DD”

Enterobacteriaceae and cefepime

This new breakpoints will be published in the 2014 guidelines (I think). The committee will also attempt to get disk diffusion changes done before end of year so these new zone sizes can also be published in 2014 as well.

*A ‘SDD’ indicates that approved, higher therapeutic levels of this drug can be used to treat serious infections with organisms having an MIC in this range.

– A definition of SDD will be included in the 2014 M100 document; it will read something like: “SDD indicates that this drug may be used in high doses to achieve the maximum achievable drug doses”.

– They will also add drug dosages for SDD. For example:

“The SDD breakpoint was derived based on a dosage regimens of 1 g Q 8 hr or 2 g Q 12 hr for organisms with a MIC of 4 ug/ml, and 2 g Q 8 hr for those with a MIC of 8 ug/ml.”


Summary -

CLSI updates AST tables (M100) each January.

CLSI updates documents that describe how to perform reference disk diffusion (M02) and reference MIC (M07) tests every 3 years.

Changes to CLSI documents are summarized in the front of each document.

Information listed in boldface type is new or modified since the previous edition of M100.

Recent breakpoint addition/revision dates are listed in the front of M100.

Minutes of CLSI AST Subcommittee meetings and other materials are available at www.clsi.org.

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http://www.clsi.org/

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CLSI

Watch for the January 2014 M100-S24 document!

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Documents

CLSI AST Update Gram Positive Bacteria_1