Accelerated bacterial detection in blood culture by enhanced acoustic flow cytometry (AFC) following peptide nucleic acid fluorescence in situ hybridization (PNA-FISH)

Xiao Xuan Huang, Nadezda Urosevic, Timothy J. J. Inglis

Research output: Contribution to journalArticle

Abstract

Bacteraemia is a risk factor for subsequent clinical deterioration and death. Current reliance on culture-based methods for detection of bacteraemia delays identification and assessment of this risk until after the optimal period for positively impacting treatment decisions has passed. Therefore, a method for rapid detection and identification of bacterial infection in the peripheral bloodstream in acutely ill patients is crucial for improved patient survival through earlier targeted antibiotic treatment. The turnaround time for current clinical laboratory methods ranges from 12 to 48 hours, emphasizing the need for a faster diagnostic test. Here we describe a novel assay for accelerated generic detection of bacteria in blood culture (BC) using peptide nucleic acid fluorescence in situ hybridization enhanced acoustic flow cytometry (PNA-FISH-AFC). For assay development, we used simulated blood cultures (BCs) spiked with one of three bacterial species at a low starting concentration of 10 CFU/mL: Escherichia coli, Klebsiella pneumoniae or Pseudomonas aeruginosa. Under current clinical settings, it takes a minimum of 12 hours incubation to reach positivity on the BacTEC system, corresponding to a bacterial concentration of 10(7)-10(9) CFU/mL optimal for further analyses. In contrast, our PNA-FISH-AFC assay detected 10(3)-10(4) CFU/mL bacteria in BC following a much shorter culture incubation of 5 to 10 hours. Using either PCR-based FilmArray assay or MALDI-TOF for bacterial detection, it took 7-10 and 12-24 hours of incubation, respectively, to reach the positive result. These findings indicate a potential time advantage of PNA-FISH-AFC assay for rapid bacterial detection in BC with significantly improved turnaround time over currently used laboratory techniques.

Original languageEnglish
Article number0201332
Number of pages17
JournalPLoS One
Volume14
Issue number2
DOIs
Publication statusPublished - 8 Feb 2019

Cite this

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title = "Accelerated bacterial detection in blood culture by enhanced acoustic flow cytometry (AFC) following peptide nucleic acid fluorescence in situ hybridization (PNA-FISH)",
abstract = "Bacteraemia is a risk factor for subsequent clinical deterioration and death. Current reliance on culture-based methods for detection of bacteraemia delays identification and assessment of this risk until after the optimal period for positively impacting treatment decisions has passed. Therefore, a method for rapid detection and identification of bacterial infection in the peripheral bloodstream in acutely ill patients is crucial for improved patient survival through earlier targeted antibiotic treatment. The turnaround time for current clinical laboratory methods ranges from 12 to 48 hours, emphasizing the need for a faster diagnostic test. Here we describe a novel assay for accelerated generic detection of bacteria in blood culture (BC) using peptide nucleic acid fluorescence in situ hybridization enhanced acoustic flow cytometry (PNA-FISH-AFC). For assay development, we used simulated blood cultures (BCs) spiked with one of three bacterial species at a low starting concentration of 10 CFU/mL: Escherichia coli, Klebsiella pneumoniae or Pseudomonas aeruginosa. Under current clinical settings, it takes a minimum of 12 hours incubation to reach positivity on the BacTEC system, corresponding to a bacterial concentration of 10(7)-10(9) CFU/mL optimal for further analyses. In contrast, our PNA-FISH-AFC assay detected 10(3)-10(4) CFU/mL bacteria in BC following a much shorter culture incubation of 5 to 10 hours. Using either PCR-based FilmArray assay or MALDI-TOF for bacterial detection, it took 7-10 and 12-24 hours of incubation, respectively, to reach the positive result. These findings indicate a potential time advantage of PNA-FISH-AFC assay for rapid bacterial detection in BC with significantly improved turnaround time over currently used laboratory techniques.",
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Accelerated bacterial detection in blood culture by enhanced acoustic flow cytometry (AFC) following peptide nucleic acid fluorescence in situ hybridization (PNA-FISH). / Huang, Xiao Xuan; Urosevic, Nadezda; Inglis, Timothy J. J.

In: PLoS One, Vol. 14, No. 2, 0201332, 08.02.2019.

Research output: Contribution to journalArticle

TY - JOUR

T1 - Accelerated bacterial detection in blood culture by enhanced acoustic flow cytometry (AFC) following peptide nucleic acid fluorescence in situ hybridization (PNA-FISH)

AU - Huang, Xiao Xuan

AU - Urosevic, Nadezda

AU - Inglis, Timothy J. J.

PY - 2019/2/8

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AB - Bacteraemia is a risk factor for subsequent clinical deterioration and death. Current reliance on culture-based methods for detection of bacteraemia delays identification and assessment of this risk until after the optimal period for positively impacting treatment decisions has passed. Therefore, a method for rapid detection and identification of bacterial infection in the peripheral bloodstream in acutely ill patients is crucial for improved patient survival through earlier targeted antibiotic treatment. The turnaround time for current clinical laboratory methods ranges from 12 to 48 hours, emphasizing the need for a faster diagnostic test. Here we describe a novel assay for accelerated generic detection of bacteria in blood culture (BC) using peptide nucleic acid fluorescence in situ hybridization enhanced acoustic flow cytometry (PNA-FISH-AFC). For assay development, we used simulated blood cultures (BCs) spiked with one of three bacterial species at a low starting concentration of 10 CFU/mL: Escherichia coli, Klebsiella pneumoniae or Pseudomonas aeruginosa. Under current clinical settings, it takes a minimum of 12 hours incubation to reach positivity on the BacTEC system, corresponding to a bacterial concentration of 10(7)-10(9) CFU/mL optimal for further analyses. In contrast, our PNA-FISH-AFC assay detected 10(3)-10(4) CFU/mL bacteria in BC following a much shorter culture incubation of 5 to 10 hours. Using either PCR-based FilmArray assay or MALDI-TOF for bacterial detection, it took 7-10 and 12-24 hours of incubation, respectively, to reach the positive result. These findings indicate a potential time advantage of PNA-FISH-AFC assay for rapid bacterial detection in BC with significantly improved turnaround time over currently used laboratory techniques.

KW - DESORPTION IONIZATION-TIME

KW - FLIGHT MASS-SPECTROMETRY

KW - RAPID IDENTIFICATION

KW - BACTEREMIA

KW - POSITIVITY

KW - MICROORGANISMS

KW - PNEUMONIAE

KW - PREDICTOR

KW - PROGNOSIS

U2 - 10.1371/journal.pone.0201332

DO - 10.1371/journal.pone.0201332

M3 - Article

VL - 14

JO - P L o S One

JF - P L o S One

SN - 1932-6203

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M1 - 0201332

ER -