Antimicrobial Susceptibility Testing and Rapid Diagnostics

Antibiotic course

Convential identification

- phenotypic methods

- positive blood culture -> gram stain -> biochemical tests

Rapid diagnostic identification

positive blood culture -> gram stain -> rapid diagnostic test

Goals

- identify organism

- determine antimicrobial susceptibility

- selection of appropriate antimicrobial regimen

different between conventional and rapid diagnostics

- Time

- conventional can take 2-3 days

- rapid within hours

Quicker identification leads to

- more timely antimicrobial optimization

- decrease mortality

- shortened hospital stay

- lower hospitalization costs

Conventional microorganism identification

- media inoculation & microbial growth monitoring

- gram stain

- subculture colonies to agar (identification and susceptibility testing)

When is empiric therapy used with conventional?

- up to first 72 hours

- blood culture collection

- gram stain alert

- microorganism ID

When does definitive therapy begin with conventional?

after susceptibilities are found

What is an MIC?

- Minimum inhibitory concentration

- shortens time to get actionable data

- allows for quantitative determination of in vitro antibacterial activity

MIC is the pharmacodynamic parameter most often used

to characterize antimicrobial activity (AUC/MIC)

How long does MIC take?

18-24 hours

MIC methodologies

- broth microdilution is the gold standard reference but not practical

- broth macrodilution

- agar dilution

- relegated to reference or research lab due to time-consuming nature

Technologies for MIC

- VITEK 2

- Phoenix

Automated systems advantages

- reduces time to definitive culture results

- available for most commonly isolated organisms

Automated systems disadvantages

- not for all pathogens/antibiotics (especially newer antibiotics)

- not all MICs can be tested

E-testing

utilizes a plastic strip with a pre-defined, stable gradient of antibiotic concentrations to produce a precise, drop-shaped inhibition ellipse on an inoculated agar plate

E-testing is available for

- newer drugs and expanded MIC ranges

- Eravacycline

- imipenem/ cilastatin/ relebactam

If considering eravacyclin or imipenem/cilastatin/ relebactam

ask for ETEST on front end (eg known MDR pathogen)

When should you get culture?

before giving antibiotics because they decrease yield of culture if given prior (unless in emergency)

What infections require fast admin of ABX?

- severe

- meningitis (45 min)

- sepsis (1 hour)

Issue with breakpoints

FDA and CLSI don't always agree

Who gets the final call on breakpoints?

FDA since they are the ones who make the automated testing cards for machines

Benefits of rapid diagnostics

- much faster

- easy detection of resistance mechanism

- sensitive (true positive rate)

- specific (true negative rate)

Current rapid diagnostic tests

- Polymerase chain reaction (PCR)

- Multiplex PCR

- Nanoparticle probe technology

- matrix-assisted laser desorption/ ionization time-of-flight mass spectrometry (MALDI-TOF MS)

- multiplex FISH

- peptide nucleic acid fluorescent in situ hybridization

PCR

C diff

Multiplex PCR

- blood, sputum, meninges, GI

- 1 hour

Nanoparticle probe technology

- blood

- 2.5 hours

Multiplex FISH

- accelerate

- gives antimicrobial susceptibilities in 7 hours

New microbiology timeline

PCR

- amplification of a target piece of DNA + organism detection

- fluorescently labeled probe and 1 set of primers

PCR steps

- nucleotide extraction

- PCR amplification

- pathogen detection

BD's Geneohm's Cdiff assay

- rapid detection of toxigenic clostridium difficile

- highly sensitive and specific

BD's Geneohm's Cdiff assay results

B gene results in < 2 hours

BD's Geneohm's Cdiff assay importance

C diff increases LOS nearly x3 and mortality nearly x4.5

Multiplex PCR

- amplification of a target piece of DNA + organism detection

- fluorescently labeled probe and >1 set of primers

Filmarray Multiplex PCR System

- Blood culture identification (BCID) panel

- GI panel

- respiratory panel

- meningitis/encephalitis (ME) panel

- tests for common things

BCID panel results

GI panel

- tests for common GI pathogens

- 2 minutes hands on, turn around time of 1 hour

- 22 targets

Respiratory panel

- tests for common causes of upper respiratory tract infections

- sensitive and specific

- 2 minutes hands on, turn around time of 1 hour

- 20 targets

- Flu, COVID, RSV

Meningitis panel

- tests cerebrospinal fluid for the 14 most common pathogens related to community acquired meningitis or encephalitis

- 2 minutes hands on, turn around time of 1 hour

- 14 targets

ME panel results

Nanoparticle probe technology

nucleic acid extraction and PCR amplification

Nanoparticle probe technology steps

- nucleic acid extraction

- PCR amplification

- hybridization of target DNA

- signal amplification

Nanoparticle probe technology examples

- nanosphere verigene blood culture gram-positive (BC-GP) assay

- nanosphere verigene blood culture gram- negative (BC-GN) assay

Accelerate PhenoTest

- FISH

- Fluorescent In Situ Hybridization

Mass spectrometry

MALDI-TOF

MALDI-TOF

- mass spect

- determines elemental composition of a sample

MALDI-TOF mass spectrometry steps

- ionization fo molecules

- charged molecules measured based on mass-to-charge ratio

- generation of a "molecular signature"

MALDI-TOF mass spectrometry needs what first

growth

Rapid does not always mean

new technology. You should evaluate current process, check to see if you have culture results before rounds

Optimization can happen sooner with

process changes to report results "more rapid" (VITEK2)

Role of the antimicrobial stewardship pharmacist

- rapid diagnostics can dramatically reduce the time to pathogen identification

- positively impact patient care

Rapid diagnostics have dramatically reduced the time to pathogen identification

- discontinuation of antimicrobials

- de-escalation of antimicrobials

Most clinical pharmacists are familiar with what rapid diagnostic technology

PCR

What is the most commonly used rapid diagnostic test?

Multiplex PCR

Formal ID training was associated with

Higher rates of familiarity

Antimicrobial stewardship program ASP

- appropriate use of antimicrobial

- all pharmacists should be invested in antimicrobial stewardship in order to achieve goals

What is the goal of ASP?

- best patient outcomes

- decreased risk of adverse effects

- reduce levels of resistance

- decrease cost

Rapid diagnostic test can help achieve ASP goals by offering

- decreased time to microorganism identification

- easy identification of resistance mechanisms

Receiving results faster requires

someone acting on results (de escalation is seen in 1/2 the time)

conventional microorganism identification vs multiplex PCR + pharmacist-driven reporting

- reduced time to antibiotic change

- higher selection of optimal therapy

- reduced use of vancomycin for coagulase negative staphylococci contaminants

Conventional vs. rapid time

- conventional: 48-72 hours

- rapid: 2 or less hours