MALDI-TOF:
Microbiology gets modern
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Peter Tilley MD FRCPC
ptilley@mail.ubc.ca
Why MALDI is better
Some limitations
Some potential changes for Micro Labs
Bruker and BioMerieux MALDI-TOF’s
now licensed in Canada
RFP now open for C&W, therefore detailed discussion
on comparative performance deferred until close. (sorry!!!)
Agar plates have been around a long
time
From Lina Hesse’s jam recipe, about 1882
So has Gram stain
Hans Christian Gram,
1884
Various enzymes
• e.g.
– carbohydrate fermentation
– amino acid metabolism
flickrhivemind.net
Panels of biochemicals
• e.g. API Strips
photo:www.jlinquist.net
Automated ID &
Susceptibilities
Misidentification
• “Based on the findings of this study, the Phoenix 100
ID/AST system NID panel falls short of being an acceptable
new method for the identification of the
Enterobacteriaceae, Vibrionaceae, and gram-negative
nonenteric isolates that are commonly encountered in
many hospital micro- biology laboratories.”
Non-fermentors and B. cepacia
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Measurements and results: Among the isolates identified as B cepacia
by referring laboratories, 11% could not be confirmed as B cepacia by
polyphasic analyses. In addition, 36% of isolates not specifically
identified by the referring laboratory or identified as a species other
than B cepacia were, in fact, found to be members of the B cepacia
complex.
Conclusions: Rates of misidentification of B cepacia remain unacceptably
high among US treatment centers.
Phenotype doesn’t always get it right
• e.g. Burkholderia cepacia used to be called
Pseudomonas cepacia
16S Ribosomal RNA
Sequencing
Carl Woese, U of Urbana
16S Ribosomal sequencing
Bacterial phyla
Closely related organisms
• E. coli and Shigella
• Bordetella pertussis, parapertussis &
bronchiseptica
• Bacillus cereus, mycoides & anthracis
• Strep. pneumoniae & mitis
allposters.com
Current methods are slow,
inaccurate or both!
theodmgroup.com
MALDI-TOF
• Koichi Tanaka
shares 2002
Nobel prize for
MALDI-TOF
• Ability to measure
large proteins
How MALDI-TOF works
Matrix-assisted laser desorption/ionization – time of flight
From Sauer, Nature Review Methods 2010
Protein patterns identify the bacteria
Degand et al JCM 2008
Bruker and BioMerieux MALDI-TOF’s
now licensed in Canada
RFP now open for C&W, therefore detailed data
on comparative performance deferred until close. (sorry!!!)
MALDI and Bugs
MALDI Plates
Inoculating the plate
Add Matrix
Load instrument
Spectrum= E. coli
The Team!
BC Children’s & Women’s
Kim Diprose
Suk Dhaliwal
Jane Margach
Laura Book
Peter Tilley
Vancouver General
Jackie O’Connell
Leane Kishu
Colleen Fletcher
Charlene Porter
Diane Roscoe
Royal Columbian
Jenifer Epp
Joy Davies
Dale Purych
BCCDC
Tazim Rahim
Ana Paccagnella
Linda Hoang
Miguel Imperial
St. Paul’s
Anna Wong
Wilson Jang
Sylvie Champagne
Marc Romney
Definitions
• Species
– High score with unique organism
– complexes accepted, e.g. Burholderia cepacia complex,
Acinetobacter baumanii complex, Enterobacter cloacae complex
• Genus
– Medium score on Bruker (1.7-1.99)
– Two species on Vitek with similar score (90-99)
• No ID
– Score too low (<1.7) or no identification provided
• Minor error
– Wrong species with high score but no clinical impact, e.g. wrong
Corynebacterium species
• Clinical Error
– Wrong species, and may affect interpretation of result, e.g.
Strep mitis vs pneumoniae
Gm neg
• Shigella, Enterobacter, Raoultella
• Acinetobacter, Roseomonas, Pseudomonas sp.
• Haemophilus, Bordetella, Legionella sp., Chryseobacteria
Gm Pos
• Strep pneumoniae vs mitis, S. anginosis
• Corynebacteria, Actinomyces
• Cryptococcus
lab type
Rapid and accurate identification of genomic species from the Acinetobacter baumannii (Ab) group by
MALDI‐TOF MS
Clinical Microbiology and Infection
pages no-no, 15 NOV 2011 DOI: 10.1111/j.1469-0691.2011.03696.x
http://onlinelibrary.wiley.com/doi/10.1111/j.1469-0691.2011.03696.x/full#f2
Costs
Average reagent cost for ID
and Susceptibility
Current methods
$8.76
MALDI-TOF
Susceptibility
$0.54
$4.64
Credit: Kris Gordos MBA
Chfi.com
Turnaround time
Time to Final ID (hr : min)
Routine methods
36:01
MALDI-TOF (avg.)
1:48
How will MALDI affect the bench?
• Few ID strips, panels and
tests
• Definitive ID known before
reporting
• Set up susceptibilities based
on definitive ID?
Aerotolerance?
• No longer needed
– M. Alfa, Winnipeg
Stool culture screens
• Just MALDI !
Quizlet.com
Odd Bugs
• Inquilinus limosus
• Alistipes onderdonkii
• Delftia acidovorans...
• We will need to write more interpretive
comments.
MALDI-TOF can be performed on
blood culture pellets
• ID in a few minutes,
almost as fast as Gram
stain
Time to direct
identification
of positive
blood cultures
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Buchan et al
JCM 2012
MALDI: <20 min.
Figure 2. Relatedness of Salmonella enterica ICMS spectra reflects serotype.
Strain Typing
Kuhns M, Zautner AE, Rabsch W, Zimmermann O, et al. (2012) Rapid Discrimination of Salmonella enterica Serovar Typhi from Other Serovars by
MALDI-TOF Mass Spectrometry. PLoS ONE 7(6): e40004. doi:10.1371/journal.pone.0040004
http://www.plosone.org/article/info:doi/10.1371/journal.pone.0040004
Fig. 2 Dendrogram representation of hierarchical cluster analysis of the recorded MALDI types and corresponding clonal complexes. The
vertical line indicates the similarity cut-off value of 2.5 chosen arbitrarily for MALDI group definition.
Manuel Wolters , Holger Rohde , Thomas Maier , Cristina Belmar-Campos , Gefion Franke , Stefanie Scherpe , Martin ...
MALDI-TOF MS fingerprinting allows for discrimination of major methicillin-resistant <ce:italic>Staphylococcus aureus</ce:italic> lineages
International Journal of Medical Microbiology Volume 301, Issue 1 2011 64 - 68
http://dx.doi.org/10.1016/j.ijmm.2010.06.002
But what will MALDI-TOF technology
do for the patient?
Pneumonia mortality in ICU
Ibrahim Chest 2000
Antibiotic Delay & Mortality in Septic
Shock
Kumar Crit Care Med 2006
Antibiotic delay in patients with
neutropenia
Lin AAC 2008
• Odds ratio for death if appropriate antibiotics
not started within 24 hours:
But antibiotic overuse leads to resistance
(Goosens, Lancet 2005)
Choosing the right antibiotic
(Antimicrobial Stewardship)
• 1. Empiric
– Clinical guidelines
• 2. By organism
– ID now well before susceptibility
• 3. By susceptibilities
– Requires growth of the isolated organism with
various antibiotics
– Usually takes a few days
Antibiogram
• Needs to be in the
pocket (or iphone)
of all your residents
and attendings!
Conclusion
• MALDI-TOF is:
– Accurate
– Fast
– Cheap
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It’s our turn to shine!
Questions? Comments?
• More specific data (in 3-4 weeks):
– ptilley@mail.ubc.ca