Antimicrobial susceptibility assays in paper-based portable culture devices.

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Abstract

To detect antibiotic-resistant bacteria in areas remote from microbiology laboratories, we designed portable culture devices performing an analogue of the Kirby-Bauer disk diffusion test inside patterned papers embedded in tape. We quantified the antibiotic susceptibility of several strains of Escherichia coli and Salmonella typhimurium by measuring blue-colored zones of inhibited growth.

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Detection and fate of antibiotic resistant bacteria in wastewater treatment plants: a review.

Chryssa Bouki, Danae Venieri, Evan Diamadopoulos (2013)

Antibiotics are among the most successful group of pharmaceuticals used for human and veterinary therapy. However, large amounts of antibiotics are released into municipal wastewater due to incomplete metabolism in humans or due to disposal of unused antibiotics, which finally find their ways into different natural environmental compartments. The emergence and rapid spread of antibiotic resistant bacteria (ARB) has led to an increasing concern about the potential environmental and public health risks. ARB and antibiotic resistant genes (ARGs) have been detected extensively in wastewater samples. Available data show significantly higher proportion of antibiotic resistant bacteria contained in raw and treated wastewater relative to surface water. According to these studies, the conditions in wastewater treatment plants (WWTPs) are favourable for the proliferation of ARB. Moreover, another concern with regards to the presence of ARB and ARGs is their effective removal from sewage. This review gives an overview of the available data on the occurrence of ARB and ARGs and their fate in WWTPs, on the biological methods dealing with the detection of bacterial populations and their resistance genes, and highlights areas in need for further research studies.

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Detecting bacteria and determining their susceptibility to antibiotics by stochastic confinement in nanoliter droplets using plug-based microfluidics.

Liang Li, Rustem Ismagilov, James Q. Boedicker … (2008)

This article describes plug-based microfluidic technology that enables rapid detection and drug susceptibility screening of bacteria in samples, including complex biological matrices, without pre-incubation. Unlike conventional bacterial culture and detection methods, which rely on incubation of a sample to increase the concentration of bacteria to detectable levels, this method confines individual bacteria into droplets nanoliters in volume. When single cells are confined into plugs of small volume such that the loading is less than one bacterium per plug, the detection time is proportional to plug volume. Confinement increases cell density and allows released molecules to accumulate around the cell, eliminating the pre-incubation step and reducing the time required to detect the bacteria. We refer to this approach as 'stochastic confinement'. Using the microfluidic hybrid method, this technology was used to determine the antibiogram - or chart of antibiotic sensitivity - of methicillin-resistant Staphylococcus aureus (MRSA) to many antibiotics in a single experiment and to measure the minimal inhibitory concentration (MIC) of the drug cefoxitin (CFX) against this strain. In addition, this technology was used to distinguish between sensitive and resistant strains of S. aureus in samples of human blood plasma. High-throughput microfluidic techniques combined with single-cell measurements also enable multiple tests to be performed simultaneously on a single sample containing bacteria. This technology may provide a method of rapid and effective patient-specific treatment of bacterial infections and could be extended to a variety of applications that require multiple functional tests of bacterial samples on reduced timescales.

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Multizone Paper Platform for 3D Cell Cultures

Ratmir Derda, Sindy Tang, Anna Laromaine … (2011)

In vitro 3D culture is an important model for tissues in vivo. Cells in different locations of 3D tissues are physiologically different, because they are exposed to different concentrations of oxygen, nutrients, and signaling molecules, and to other environmental factors (temperature, mechanical stress, etc). The majority of high-throughput assays based on 3D cultures, however, can only detect the average behavior of cells in the whole 3D construct. Isolation of cells from specific regions of 3D cultures is possible, but relies on low-throughput techniques such as tissue sectioning and micromanipulation. Based on a procedure reported previously (“cells-in-gels-in-paper” or CiGiP), this paper describes a simple method for culture of arrays of thin planar sections of tissues, either alone or stacked to create more complex 3D tissue structures. This procedure starts with sheets of paper patterned with hydrophobic regions that form 96 hydrophilic zones. Serial spotting of cells suspended in extracellular matrix (ECM) gel onto the patterned paper creates an array of 200 micron-thick slabs of ECM gel (supported mechanically by cellulose fibers) containing cells. Stacking the sheets with zones aligned on top of one another assembles 96 3D multilayer constructs. De-stacking the layers of the 3D culture, by peeling apart the sheets of paper, “sections” all 96 cultures at once. It is, thus, simple to isolate 200-micron-thick cell-containing slabs from each 3D culture in the 96-zone array. Because the 3D cultures are assembled from multiple layers, the number of cells plated initially in each layer determines the spatial distribution of cells in the stacked 3D cultures. This capability made it possible to compare the growth of 3D tumor models of different spatial composition, and to examine the migration of cells in these structures.