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1.
PLoS One ; 7(4): e33443, 2012.
Article in English | MEDLINE | ID: mdl-22511922

ABSTRACT

Genotyping methods are essential to understand the transmission dynamics of Acinetobacter baumannii. We examined the representative genotypes of A. baumannii at different time periods in select locations in Ohio, using two rapid automated typing methods: PCR coupled with electrospray ionization mass spectrometry (PCR/ESI-MS), a form of multi-locus sequence typing (MLST), and repetitive-sequence-based-PCR (rep-PCR). Our analysis included 122 isolates from 4 referral hospital systems, in 2 urban areas of Ohio. These isolates were associated with outbreaks at 3 different time periods (1996, 2000 and 2005-2007). Type assignments of PCR/ESI-MS and rep-PCR were compared to each other and to worldwide (WW) clone types. The discriminatory power of each method was determined using the Simpson's index of diversity (DI). We observed that PCR/ESI-MS sequence type (ST) 14, corresponding to WW clone 3, predominated in 1996, whereas ST 12 and 14 co-existed in the intermediate period (2000) and ST 10 and 12, belonging to WW clone 2, predominated more recently in 2007. The shift from WW clone 3 to WW clone 2 was accompanied by an increase in carbapenem resistance. The DI was approximately 0.74 for PCR/ESI-MS, 0.88 for rep-PCR and 0.90 for the combination of both typing methods. We conclude that combining rapid automated typing methods such as PCR/ESI-MS and rep-PCR serves to optimally characterize the regional molecular epidemiology of A. baumannii. Our data also sheds light on the changing sequence types in an 11 year period in Northeast Ohio.


Subject(s)
Acinetobacter baumannii/genetics , Bacterial Typing Techniques , Evolution, Molecular , Acinetobacter Infections/epidemiology , Acinetobacter Infections/transmission , Acinetobacter baumannii/isolation & purification , Disease Outbreaks , Genotype , Humans , Ohio , Polymerase Chain Reaction/methods , Spectrometry, Mass, Electrospray Ionization , Time Factors
2.
Infect Control Hosp Epidemiol ; 29(6): 553-5, 2008 Jun.
Article in English | MEDLINE | ID: mdl-18510464

ABSTRACT

The Walter Reed Army Medical Center has experienced an influx of traumatically injured patients either infected or colonized with Acinetobacter baumannii. Using multilocus polymerase chain reaction (PCR) and mass spectrometry to genotype isolates, we found an atypical and evolving strain distribution, distinct from those found at nonmilitary hospitals in the United States.


Subject(s)
Acinetobacter Infections/epidemiology , Acinetobacter baumannii/classification , Disease Outbreaks , Evolution, Molecular , Hospitals, Military , Warfare , Acinetobacter Infections/microbiology , Acinetobacter baumannii/genetics , Bacterial Typing Techniques , Genotype , Humans , Mass Spectrometry , Polymerase Chain Reaction
3.
Virology ; 368(2): 286-95, 2007 Nov 25.
Article in English | MEDLINE | ID: mdl-17655905

ABSTRACT

Members of the genus Alphavirus are a diverse group of principally mosquito-borne RNA viruses. There are at least 29 species and many more subtypes of alphaviruses and some are considered potential bioweapons. We have developed a multi-locus RT-PCR followed by electrospray ionization mass spectrometry (RT-PCR/ESI-MS) assay that uses the amplicon base compositions to detect and identify alphaviruses. A small set of primer pairs targeting conserved sites in the alphavirus RNA genome were used to amplify a panel of 36 virus isolates representing characterized Old World and New World alphaviruses. Base compositions from the resulting amplicons could be used to unambiguously determine the species or subtype of 35 of the 36 isolates. The assay detected, without culture, Venezuelan equine encephalitis virus (VEEV), Eastern equine encephalitis virus (EEEV), and mixtures of both in pools consisting of laboratory-infected and -uninfected mosquitoes. Further, the assay was used to detect alphaviruses in naturally occurring mosquito vectors collected from locations in South America and Asia. Mosquito pools collected near Iquitos, Peru, were found to contain an alphavirus with a very distinct signature. Subsequent sequence analysis confirmed that the virus was a member of the Mucambo virus species (subtype IIID in the VEEV complex). The assay we have developed provides a rapid, accurate, and high-throughput assay for surveillance of alphaviruses.


Subject(s)
Aedes/virology , Alphavirus/isolation & purification , Culex/virology , Reverse Transcriptase Polymerase Chain Reaction/methods , Spectrometry, Mass, Electrospray Ionization/methods , Alphavirus/classification , Alphavirus/genetics , Animals , Base Composition , Base Sequence , DNA Primers , Humans , Molecular Sequence Data , RNA, Viral/analysis , RNA, Viral/chemistry , RNA, Viral/isolation & purification , Sensitivity and Specificity , Time Factors
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