cphm-838 aspergillus species by laser light scattering...

Rapid Voriconazole Susceptibility Testing ofAspergillus Species by Laser Light Scattering

A. Fitzgerald1, W. Memon1, A. Tomaras2, S.X. Zhang1

1. Johns Hopkins Hospital, Baltimore, MD2. BacterioScan, St. Louis, MO

Introduction

In vitro antifungal susceptibility testing for Aspergillus species is currently performed using the CLSI-developed microbroth dilution

method, which takes at least 48 hours to obtain MIC results. There is an unmet need for a more rapid antifungal susceptibility testing

method for Aspergillus. A laser light scattering system (BacterioScan 216Dx) is able to detect microbial growth in broth by producing

signals based on optical density read every 5 minutes. The purpose of this study was to test the BacterioScan 216Dx system for faster

antifungal susceptibility results compared to the current CLSI microbroth dilution method.

Materials

20 Aspergillus species with previously performed Voriconazole susceptibility via

microbroth dilution method were tested on the BacterioScan 216Dx (Figure 1). The

isolates cultured on Potato Flake Agar were <7 days old at time of testing. Purity plates

(Sabouraud Dextrose) were inoculated from the test cartridge growth control well.

• B-MIC determining timepoint results were available in less than 24 hrs, with a mean of 1060 minutes (17.7

hours) and a median of 1013 minutes (16.9 hours)

• All B-MIC results were concordant to the CLSI MIC results within 2 doubling dilution ranges

• Possible need to determine clinical antifungal breakpoints unique to BacterioScan

• BacterioScan was able to detect the MICs for the 4 Aspergillus fumigatus isolates with an ECV ≥ 1 ug/mL at

a time frame of 669 to 1026 min (11 to 17 h)

• Promising for more rapid mold antifungal testing that will improve patient care and turnaround time

Summary

Acknowledgements:

Thank you to BacterioScan for providing ongoing instrument and reagent support for this project.

Thank you to Dr. Nathan Wiederhold of the Fungus Testing Laboratory, University of Texas at San Antonio for specimens and

genomic information.

Methods

Results

afitzg17@jhmi.edu410-955-6510

CPHM-8386/22/19

Figure 1

Testing for Optimal Growth Conditions

Organisms: Aspergillus fumigatus, A. niger, A.

flavus, and A. terreus

Medium: Sabouraud Dextrose broth vs. RPMI

broth

% Glucose: 0.2% vs. 2% vs. 4%

Inoculum Dilution/Concentration: 1.0 McFarland

concentration [McF], 0.5 McF, 1:50 dilution of 1.0

McF, 1:50 dilution of 0.5 McF

Best Growth Condition Determined

Medium: RPMI broth

% Glucose: 0.2%

Inoculum Dilution/Concentration: 1:50 dilution of

1.0 McF (OD range 0.18 to 0.22)

• 2 cartridges pre-loaded with Voriconazole

(ug/mL) (See Figure 2) were run for each isolate

• Pipetted 2.5 mL of inoculated RPMI in each well

• Cartridges inoculated at 37°C for 1440 min (24

hr)

• Kinetic laser scattering signals based on growth

were captured and data were converted into

figures.

Figure 2

Cartridge 2

Cartridge 1