High-resolution genotyping of Campylobacter species by use of PCR and high-throughput mass spectrometry.
In this work we report on a high-throughput mass spectrometry-based technique for the rapid high-resolution identification of Campylobacter jejuni strain types. This method readily distinguishes C. jejuni from C. coli, has a resolving power comparable to that of multilocus sequence typing (MLST), is applicable to mixtures, and is highly automated. The strain typing approach is based on high-performance mass spectrometry, which "weighs" PCR amplicons with enough mass accuracy to unambiguously determine the base composition of each amplicon (i.e., the numbers of A's, G's, C's, and T's). Amplicons are derived from PCR primers which amplify short (<140-bp) regions of the housekeeping genes used by conventional MLST strategies. The results obtained with a challenge panel that comprised 25 strain types of C. jejuni and 25 strain types of C. coli are presented. These samples were parsed and resolved with demonstrated sensitivity down to 10 genomes/PCR from pure isolates.
Hannis
James C
JC
Ibis Biosciences, Inc., 1891 Rutherford Road, Carlsbad, CA 92008, USA.
Manalili
Sheri M
SM
Hall
Thomas A
TA
Ranken
Raymond
R
White
Neill
N
Sampath
Rangarajan
R
Blyn
Lawrence B
LB
Ecker
David J
DJ
Mandrell
Robert E
RE
Fagerquist
Clifton K
CK
Bates
Anna H
AH
Miller
William G
WG
Hofstadler
Steven A
SA
D000818:Animals; D001426:Bacterial Proteins / Q000737:chemistry / Q000235:genetics; D015373:Bacterial Typing Techniques; D001482:Base Composition; D002167:Campylobacter / Q000737:chemistry / Q000145:classification* / Q000235:genetics*; D017000:Campylobacter coli / Q000145:classification / Q000235:genetics; D016123:Campylobacter jejuni / Q000145:classification / Q000235:genetics; D017931:DNA Primers; D004269:DNA, Bacterial / Q000032:analysis; D005838:Genotype; D006801:Humans; D013058:Mass Spectrometry / Q000379:methods*; D016133:Polymerase Chain Reaction / Q000379:methods*; D013045:Species Specificity
D001426:Bacterial Proteins; D017931:DNA Primers; D004269:DNA, Bacterial