Comparison of rpoB gene sequencing, 16S rRNA gene sequencing, gyrB multiplex PCR, and the VITEK2 system for identification of Acinetobacter clinical isolates

Lee, Min Jung; Jang, Sook Jin; Li, Xue Min; Park, Geon; Kook, Joong-Ki; Kim, Min Jung; Chang, Young-Hyo; Shin, Jong Hee; Kim, Soo Hyun; Kim, Dong-Min; Kang, Seong‐Ho; Moon, Dae-Soo

doi:10.1016/j.diagmicrobio.2013.07.013

Cited by 51 publications

(43 citation statements)

References 25 publications

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“…Five isolates gave poor-quality sequencing assemblies, despite repeated attempts, and were also excluded, leaving 148 isolates for analysis. We identified the species of these 148 isolates by rpoB gene sequence analysis, an accepted method of parsing ACB complex bacteria into species (45,71). This analysis indicated the following Acinetobacter species distribution: A. baumannii (n ϭ 116), A. pittii (n ϭ 28), A. nosocomialis (n ϭ 3), and A. soli (n ϭ 1).…”

Section: Resultsmentioning

confidence: 99%

Utility of Whole-Genome Sequencing in Characterizing Acinetobacter Epidemiology and Analyzing Hospital Outbreaks

2016

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bAcinetobacter baumannii frequently causes nosocomial infections and outbreaks. Whole-genome sequencing (WGS) is a promising technique for strain typing and outbreak investigations. We compared the performance of conventional methods with WGS for strain typing clinical Acinetobacter isolates and analyzing a carbapenem-resistant A. baumannii (CRAB) outbreak. We performed two band-based typing techniques (pulsed-field gel electrophoresis and repetitive extragenic palindromic-PCR), multilocus sequence type (MLST) analysis, and WGS on 148 Acinetobacter calcoaceticus-A. baumannii complex bloodstream isolates collected from a single hospital from 2005 to 2012. Phylogenetic trees inferred from core-genome single nucleotide polymorphisms (SNPs) confirmed three Acinetobacter species within this collection. Four major A. baumannii clonal lineages (as defined by MLST) circulated during the study, three of which are globally distributed and one of which is novel. WGS indicated that a threshold of 2,500 core SNPs accurately distinguished A. baumannii isolates from different clonal lineages. The bandbased techniques performed poorly in assigning isolates to clonal lineages and exhibited little agreement with sequence-based techniques. After applying WGS to a CRAB outbreak that occurred during the study, we identified a threshold of 2.5 core SNPs that distinguished nonoutbreak from outbreak strains. WGS was more discriminatory than the band-based techniques and was used to construct a more accurate transmission map that resolved many of the plausible transmission routes suggested by epidemiologic links. Our study demonstrates that WGS is superior to conventional techniques for A. baumannii strain typing and outbreak analysis. These findings support the incorporation of WGS into health care infection prevention efforts.A n important role of clinical microbiology is to identify relationships between bacterial isolates. At a broad level, phenotypic and genotypic tests are used to categorize bacterial isolates into the same or different species. Within a bacterial species, techniques are used to group isolates into clonal lineages, which are groups of closely related bacteria that share a recent common ancestor but have spread regionally or globally. At a more local level, infection control practitioners must determine whether a group of isolates constitutes a hospital outbreak by ascertaining whether the isolates have a degree of similarity consistent with a common source within the hospital. Once isolates belonging to a hospital outbreak have been identified, similarities and differences between these isolates can be exploited to generate a transmission map to aid in finding the source of the outbreak and in disrupting ongoing pathways of transmission. For some groups of bacteria, such as Acinetobacter, discernment at each level has medically important consequences and therefore must be accomplished by hospital-associated clinical microbiology laboratories.Within the Acinetobacter genus, the Acinetobacter calcoaceticusAcinetobac...

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Section: Resultsmentioning

confidence: 99%

Utility of Whole-Genome Sequencing in Characterizing Acinetobacter Epidemiology and Analyzing Hospital Outbreaks

2016

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“…VITEK 2, a routine laboratory method, can help to discriminate among genera but not among species. MALDI-TOF MS and 16S rDNA sequencing/16S rRNA sequencing can be used to identify species and are more frequently used for research purposes (Lee et al, 2014;Schröttner et al, 2014). According to Schröttner et al (2014), the genus Myroides was reliably identified in tests of 22 isolates using VITEK 2.…”

Section: Present Diagnostic Methods Do Not Clarify the Antibiotic Resmentioning

confidence: 99%

“…However, as the antibiotic treatment history was not described and the antibiotic resistance status of this strain was not given, the antibiotic resistance mechanisms could not be analyzed from the data. Yoon et al, 2006;Lee et al, 2014;Schröttner et al, 2014 To study these aspects in our M. odoratimimus strain PR63039, we extracted its genomic DNA. Agarose gel electrophoresis results from several experiments revealed that it might harbor at least six different types of plasmids which could be related to antibiotic resistance (Fig.…”

Section: Whole Genome Sequencing Is a Feasible Way To Investigate Thementioning

confidence: 99%

Antibiotic resistance mechanisms of Myroides sp.

Yuan

et al. 2016

J. Zhejiang Univ. Sci. B

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Bacteria of the genus Myroides (Myroides spp.) are rare opportunistic pathogens. Myroides sp. infections have been reported mainly in China. Myroides sp. is highly resistant to most available antibiotics, but the resistance mechanisms are not fully elucidated. Current strain identification methods based on biochemical traits are unable to identify strains accurately at the species level. While 16S ribosomal RNA (rRNA) gene sequencing can accurately achieve this, it fails to give information on the status and mechanisms of antibiotic resistance, because the 16S rRNA sequence contains no information on resistance genes, resistance islands or enzymes. We hypothesized that obtaining the whole genome sequence of Myroides sp., using next generation sequencing methods, would help to clarify the mechanisms of pathogenesis and antibiotic resistance, and guide antibiotic selection to treat Myroides sp. infections. As Myroides sp. can survive in hospitals and the environment, there is a risk of nosocomial infections and pandemics. For better management of Myroides sp. infections, it is imperative to apply next generation sequencing technologies to clarify the antibiotic resistance mechanisms in these bacteria.

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“…A recent paper has described increased severity and mortality in bacteraemia patients infected with A. baumannii compared with those infected with A. pitii and A. nosocomialis, emphasising the need for accurate differentiation (Fitzpatrick et al, 2015). However, these four species which together make up the A. calcoaceticus -A. baumannii (Acb) complex are indistinguishable by phenotypic based tests (Peleg et al, 2008;Wang et al, 2013;Lee et al, 2014).…”

Section: Introductionmentioning

confidence: 99%

Species identification within Acinetobacter calcoaceticus–baumannii complex using MALDI-TOF MS

Toh

Paterson

Kamolvit

et al. 2015

Journal of Microbiological Methods

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Acinetobacter baumannii, one of the more clinically relevant species in the Acinetobacter genus is well known to be multi-drug resistant and associated with bacteremia, urinary tract infection, pneumonia, wound infection and meningitis. However, it cannot be differentiated from closely related species such as A. calcoaceticus, A. pittii and A. nosocomialis by most phenotypic tests and can only be differentiated by specific, time consuming genotypic tests with very limited use in clinical microbiological laboratories. As a result, these species are grouped into the A.calcoaceticus -A. baumannii (Acb) complex. Herein we investigated the mass spectra of 73 Acinetobacter spp., representing ten different species, using an AB SCIEX 5800 MALDI -TOF MS to differentiate members of the Acinetobacter genus, including the species of the Acb complex. RpoB gene sequencing, 16S rRNA sequencing, and gyrB multiplex PCR were also evaluated as orthogonal methods to identify the organisms used in this study. We found that whilst 16S rRNA and rpoB gene sequencing could not differentiate A. pittii or A. calcoaceticus, they can be differentiated using gyrB multiplex PCR and MALDI -TOF MS. All ten Acinetobacter species investigated could be differentiated by their MALDI -TOF mass spectra.

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Comparison of rpoB gene sequencing, 16S rRNA gene sequencing, gyrB multiplex PCR, and the VITEK2 system for identification of Acinetobacter clinical isolates

Cited by 51 publications

References 25 publications

Utility of Whole-Genome Sequencing in Characterizing Acinetobacter Epidemiology and Analyzing Hospital Outbreaks

Utility of Whole-Genome Sequencing in Characterizing Acinetobacter Epidemiology and Analyzing Hospital Outbreaks

Antibiotic resistance mechanisms of Myroides sp.

Species identification within Acinetobacter calcoaceticus–baumannii complex using MALDI-TOF MS

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