An optimized SYBR green I/PI assay for rapid viability assessment and antibiotic susceptibility testing for Borrelia burgdorferi

Jie Feng, Ting Wang, Shuo Zhang, Wanliang Shi, Ying Zhang

Research output: Contribution to journalArticle

Abstract

Lyme disease caused by Borrelia burgdorferi is the most common tick-borne disease in the US and Europe. Unlike most bacteria, measurements of growth and viability of B. burgdorferi are challenging. The current B. burgdorferi viability assays based on microscopic counting and PCR are cumbersome and tedious and cannot be used in a high throughput format. Here, we evaluated several commonly used viability assays including MTT and XTT assays, fluorescein diacetate assay, Sytox Green/Hoechst 33342 assay, the commercially available LIVE/DEAD BacLight assay, and SYBR Green I/PI assay by microscopic counting and by automated 96-well plate reader for rapid viability assessment of B. burgdorferi. We found that the optimized SYBR Green I/PI assay based on green to red fluorescence ratio is superior to all the other assays for measuring the viability of B. burgdorferi in terms of sensitivity, accuracy, reliability, and speed in automated 96-well plate format and in comparison with microscopic counting. The BSK-H medium which produced a high background for the LIVE/ DEAD BacLight assay did not affect the SYBR Green I/PI assay, and the viability of B. burgdorferi culture could be directly measured using a microtiter plate reader. The SYBR Green I/PI assay was found to reliably assess the viability of planktonic as well as biofilm B. burgdorferi and could be used as a rapid antibiotic susceptibility test. Thus, the SYBR Green I/PI assay provides a more sensitive, rapid and convenient method for evaluating viability and antibiotic susceptibility of B. burgdorferi and can be used for high-throughput drug screens.

Original languageEnglish (US)
Article numbere111809
JournalPLoS One
Volume9
Issue number11
DOIs
StatePublished - Nov 3 2014

Fingerprint

Borrelia burgdorferi
Assays
antibiotics
viability
Anti-Bacterial Agents
Testing
assays
testing
Tick-Borne Diseases
Lyme Disease
SYBR Green I
Biofilms
Throughput
tick-borne diseases
Fluorescence
Lyme disease
Bacteria
fluorescein
Polymerase Chain Reaction
biofilm

ASJC Scopus subject areas

  • Agricultural and Biological Sciences(all)
  • Biochemistry, Genetics and Molecular Biology(all)
  • Medicine(all)

Cite this

An optimized SYBR green I/PI assay for rapid viability assessment and antibiotic susceptibility testing for Borrelia burgdorferi. / Feng, Jie; Wang, Ting; Zhang, Shuo; Shi, Wanliang; Zhang, Ying.

In: PLoS One, Vol. 9, No. 11, e111809, 03.11.2014.

Research output: Contribution to journalArticle

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