Antibiotic Susceptibility Testing of Grown Blood Cultures by Combining Culture and Real-Time Polymerase Chain Reaction Is Rapid and Effective

Judith Beuving,Annelies Verbon,Petra F G Wolffs,Ellen E Stobberingh,Gunnar F Kaufmann,Firza A Gronthoud

doi:10.1371/journal.pone.0027689

Judith Beuving, Annelies Verbon + Show 4 more

Open Access

https://doi.org/10.1371/journal.pone.0027689

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Journal: PLoS ONE	Publication Date: Dec 14, 2011
Citations: 36	License type: CC BY 4.0

Affiliation: Maastricht University

Abstract

BackgroundEarly administration of appropriate antibiotic therapy in bacteraemia patients dramatically reduces mortality. A new method for RApid Molecular Antibiotic Susceptibility Testing (RAMAST) that can be applied directly to positive blood cultures was developed and evaluated.Methodology/Principal FindingsGrowth curves and antibiotic susceptibility of blood culture isolates (Staphylococcus aureus, enterococci and (facultative) aerobic Gram-negative rods) were determined by incubating diluted blood cultures with and without antibiotics, followed by a quantitative universal 16S PCR to detect the presence or absence of growth. Testing 114 positive blood cultures, RAMAST showed an agreement with microbroth dilution of 96.7% for Gram-negative rods, with a minor error (false-susceptibility with a intermediate resistant strain) rate of 1.9%, a major error (false resistance) rate of 0.8% and a very major error (false susceptibility) rate of 0.6%. Agreement for S.aureus was 97.9%, with a very major error rate of 2.1%. Enterococcus species showed 95.0% agreement, with a major error rate of 5.0%. These agreements are comparable with those of the Phoenix system. Starting from a positive blood culture, the test was completed within 9 hours.Conclusions/SignificanceThis new rapid method for antibiotic susceptibility testing can potentially provide accurate results for most relevant bacteria commonly isolated from positive blood cultures in less time than routine methods.

Highlights

Invasion of bacteria in the bloodstream can result in sepsis
The growth curves showed that blood culture isolates require 6 hours of incubation before sufficient growth has occurred for reliable antibiotic susceptibility testing with RApid Molecular Antibiotic Susceptibility Testing (RAMAST)
Phase II: Susceptibility testing on positive blood cultures with RAMAST

Summary

Introduction

Invasion of bacteria in the bloodstream (bacteraemia) can result in sepsis. Sepsis occurs in about two percent of all hospitalized patients and in as much as 37.4% of all ICU patients and the incidence is rising [1,2,3,4]. Administration of appropriate antibiotics reduces mortality of sepsis dramatically [7,8,9]. Broad spectrum antibiotics are active against most bacteria causing bacteraemia and are often used in combination. The empiric antibiotic therapy may not cover the causative microorganism, especially with the rising incidence of multi-drug resistant bacteria [10]. This is the case in up to 40% of all bacteraemia patients [9,11,12]. Administration of appropriate antibiotic therapy in bacteraemia patients dramatically reduces mortality. A new method for RApid Molecular Antibiotic Susceptibility Testing (RAMAST) that can be applied directly to positive blood cultures was developed and evaluated

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