Identification of Acinetobacter species and genotyping of Acinetobacter baumannii by multilocus PCR and mass spectrometry.
Members of the genus Acinetobacter are ubiquitous in soil and water and are an important cause of nosocomial infections. A rapid method is needed to genotype Acinetobacter isolates to determine epidemiology and clonality during infectious outbreaks. Multilocus PCR followed by electrospray ionization mass spectrometry (PCR/ESI-MS) is a method that uses the amplicon base compositions to genotype bacterial species. In order to identify regions of the Acinetobacter genome useful for this method, we sequenced regions of six housekeeping genes (trpE, adk, efp, mutY, fumC, and ppa) from 267 isolates of Acinetobacter. Isolates were collected from infected and colonized soldiers and civilians involved in an outbreak in the military health care system associated with the conflict in Iraq, from previously characterized outbreaks in European hospitals, and from culture collections. Most of the isolates from the Iraqi conflict were Acinetobacter baumannii (189 of 216 isolates). Among these, 111 isolates had genotypes identical or very similar to those associated with well-characterized A. baumannii isolates from European hospitals. Twenty-seven isolates from the conflict were found to have genotypes representing different Acinetobacter species, including 8 representatives of Acinetobacter genomospecies 13TU and 13 representatives of Acinetobacter genomospecies 3. Analysis by the PCR/ESI-MS method using nine primer pairs targeting the most information-rich regions of the trpE, adk, mutY, fumC, and ppa genes distinguished 47 of the 48 A. baumannii genotypes identified by sequencing and identified at the species level at least 18 Acinetobacter species. Results obtained with our genotyping method were essentially in agreement with those obtained by pulse-field gel electrophoresis analysis. The PCR/ESI-MS genotyping method required 4 h of analysis time to first answer with additional samples subsequently analyzed every 10 min. This rapid analysis allows tracking of transmission for the implementation of appropriate infection control measures on a time scale previously not achievable.
Ecker
Joseph A
JA
Ibis Biosciences, Isis Pharmaceuticals, Inc., 1891 Rutherford Road, Carlsbad, CA 92008, USA.
Massire
Christian
C
Hall
Thomas A
TA
Ranken
Raymond
R
Pennella
Thuy-Trang D
TT
Agasino Ivy
Cristina
C
Blyn
Lawrence B
LB
Hofstadler
Steven A
SA
Endy
Timothy P
TP
Scott
Paul T
PT
Lindler
Luther
L
Hamilton
Tacita
T
Gaddy
Charla
C
Snow
Kerry
K
Pe
Marie
M
Fishbain
Joel
J
Craft
David
D
Deye
Gregory
G
Riddell
Scott
S
Milstrey
Eric
E
Petruccelli
Bruno
B
Brisse
Sylvain
S
Harpin
Vanessa
V
Schink
Amy
A
Ecker
David J
DJ
Sampath
Rangarajan
R
Eshoo
Mark W
MW
D000150:Acinetobacter / Q000145:classification* / Q000235:genetics; D000151:Acinetobacter Infections / Q000453:epidemiology / Q000382:microbiology*; D040981:Acinetobacter baumannii / Q000145:classification* / Q000235:genetics*; D015373:Bacterial Typing Techniques; D016000:Cluster Analysis; D017931:DNA Primers; D004269:DNA, Bacterial / Q000032:analysis / Q000235:genetics; D004196:Disease Outbreaks; D005798:Genes, Bacterial; D005838:Genotype; D006801:Humans; D013058:Mass Spectrometry; D017720:Molecular Epidemiology / Q000379:methods; D010802:Phylogeny; D016133:Polymerase Chain Reaction; D017385:Sequence Homology
D017931:DNA Primers; D004269:DNA, Bacterial