Stress-induced Antibiotic Susceptibility Testing on a Chip

Maxim Kalashnikov; Jennifer Campbell; Jean C. Lee; Andre Sharon; Alexis F. Sauer-Budge

doi:10.3791/50828

Test di suscettibilità agli antibiotici indotti dallo stress su un chip

JoVE Journal
Bioengineering

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JoVE Journal Bioengineering

Stress-induced Antibiotic Susceptibility Testing on a Chip

Test di suscettibilità agli antibiotici indotti dallo stress su un chip

Full Text

6,953 Views

12:41 min

January 8, 2014

DOI: 10.3791/50828-v

Maxim Kalashnikov¹, Jennifer Campbell¹, Jean C. Lee², Andre Sharon^1,3, Alexis F. Sauer-Budge^1,4

¹Fraunhofer USA Center for Manufacturing Innovation, ²Division of Infectious Diseases, Department of Medicine, Brigham and Women's Hospital,Harvard Medical School, ³Department of Mechanical Engineering,Boston University, ⁴Department of Biomedical Engineering,Boston University

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Overview

This study presents a microfluidic platform designed for rapid antibiotic susceptibility testing of bacteria. By immobilizing bacteria in a microfluidic channel and applying antibiotics under stress conditions, the method allows for quick assessment of bacterial viability.

Key Study Components

Area of Science

Microfluidics
Antibiotic susceptibility testing
Cell viability assessment

Background

Traditional methods for testing antibiotic susceptibility can be time-consuming.
Microfluidic technologies offer advantages in speed and efficiency.
Understanding bacterial responses to antibiotics is crucial for effective treatment.
This method eliminates the need for monitoring bacterial division.

Purpose of Study

To develop a rapid method for determining bacterial susceptibility to antibiotics.
To assess bacterial viability under stress conditions.
To improve upon existing susceptibility testing techniques.

Methods Used

Immobilization of log phase bacteria in a microfluidic channel.
Application of antibiotic and fluorescent stains under high flow rates.
Automated collection of phase contrast and fluorescence images.
Calculation of normalized bacterial death percentages over time.

Main Results

Susceptible bacteria show rapid cell death in the presence of antibiotics.
Resistant bacteria can survive under stressful conditions.
Normalized death percentages provide insights into antibiotic susceptibility.
The method allows for real-time monitoring of bacterial viability.

Conclusions

The microfluidic platform offers a rapid alternative to traditional susceptibility testing.
This approach enhances the understanding of bacterial responses to antibiotics.
Future applications may include broader testing of various bacterial strains.

Frequently Asked Questions

What is the main advantage of this microfluidic method?

It allows for rapid assessment of antibiotic susceptibility without needing to monitor bacterial division.

How are bacterial responses monitored in this study?

Bacterial viability is monitored using phase contrast and fluorescence imaging techniques.

What types of bacteria can be tested using this method?

The method can be applied to various strains of bacteria, particularly those of clinical interest.

How does the flow rate affect the experiment?

High flow rates stress the bacteria, activating their antibiotic-targeted biochemical pathways.

What is the significance of calculating normalized death percentages?

Normalized death percentages help quantify the effectiveness of the antibiotic over time.

Can this method be used for other types of assays?

While primarily designed for antibiotic susceptibility testing, the platform may be adapted for other assays involving bacterial responses.

Abbiamo sviluppato una piattaforma microfluidica per test di suscettibilità agli antibiotici rapidi. Il fluido viene passato ad alta velocità sui batteri immobilizzati sul fondo di un canale microfluidico. In presenza di stress e antibiotici, i ceppi sensibili di batteri muoiono rapidamente. Tuttavia, i batteri resistenti possono sopravvivere a queste condizioni stressanti.

L'obiettivo generale del seguente esperimento è quello di determinare rapidamente la suscettibilità dei batteri agli antibiotici di interesse. Ciò si ottiene immobilizzando prima i batteri in fase logaritmica sul fondo di un canale microfluidico. Nella seconda fase, il terreno contenente l'antibiotico e una colorazione fluorescente morta viva viene quindi spinto attraverso il canale ad alta velocità, stressando i batteri e attivando i loro percorsi biochimici mirati agli antibiotici.

Le immagini di contrasto e fluorescenza della fase successiva vengono raccolte automaticamente ogni due minuti per un'ora per monitorare la vitalità batterica, le percentuali di morte batterica normalizzate vengono calcolate dalle immagini acquisite in ogni momento dell'esperimento. In definitiva, il fenotipo di suscettibilità agli antibiotici dei batteri è determinato da un aumento della percentuale di morte cellulare normalizzata nel tempo. Il vantaggio principale di questa tecnica rispetto ai metodi di test di suscettibilità fenotipica esistenti come il brodo, la diluizione o la microcoltura, è che il nostro metodo non richiede il monitoraggio della divisione batterica.

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