Abstract

Recent advances in phenotypic and chemotaxonomic methods have improved the ability of systems to resolve bacterial identities at the species level. Key to the effective use of these systems is the ability to draw upon databases which can be augmented with new data gleaned from atypical or novel isolates. In this study we compared the performance of the Biolog GEN III identification system (hereafter, GEN III) with matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS) and 16S rRNA gene sequencing in the identification of isolates of veterinary interest. The use of strains that had proven more difficult to identify by routine methods was designed to test the systems' abilities at the extremes of their performance range. Over an 18month period, 100 strains were analysed by all three methods. To highlight the importance of identification to species level, a weighted scoring system was devised to differentiate the capacity to identify at genus and species levels. The overall relative weighted scores were 0.869:0.781:0.769, achieved by 16S rRNA gene sequencing, GEN III and MALDI-TOF MS respectively, when compared to the ‘gold standard’. Performance to the genus level was significantly better using 16S rRNA gene sequencing; however, performance to the species level was similar for all three systems.

Highlights

  • Following a period of technological refinement in recent decades, automated or semi-automated metabolic methods have predominated in high-throughput microbiology laboratories processing routine clinical specimens

  • Where the presumptive identification could not be substantiated to species level, strains could only be compared at the genus level (n = 24)

  • 16S rRNA gene sequencing performed significantly better than both GEN III and MALDI-TOF MS (p = 0.0005 and 0.0034 respectively) in this respect

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Summary

Introduction

Following a period of technological refinement in recent decades, automated or semi-automated metabolic methods have predominated in high-throughput microbiology laboratories processing routine clinical specimens. Intra-species variation, together with the frequent recognition of new species, began to undermine the accuracy of less-sophisticated phenotypic systems (O'Hara, 2005). Such systems were found to be limited either by virtue of a fundamental incompatibility with particular groups or by a lack of flexibility within the reference database to recognise novel profiles and offer ‘intuitive’ solutions. 16S rRNA gene sequencing offered improved resolution at the genus and, in many cases, species level. P. Wragg et al / Journal of Microbiological Methods 105 (2014) 16–21

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