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Culturing aerobic and anaerobic bacteria and mammalian cells with a microfluidic differential oxygenator.

ABSTRACT: In this manuscript, we report on the culture of anaerobic and aerobic species within a disposable multilayer polydimethylsiloxane (PDMS) microfluidic device with an integrated differential oxygenator. A gas-filled microchannel network functioning as an oxygen-nitrogen mixer generates differential oxygen concentration. By controlling the relative flow rate of the oxygen and nitrogen input gases, the dissolved oxygen (DO) concentration in proximal microchannels filled with culture media are precisely regulated by molecular diffusion. Sensors consisting of an oxygen-sensitive dye embedded in the fluid channels permit dynamic fluorescence-based monitoring of the DO concentration using low-cost light-emitting diodes. To demonstrate the general utility of the platform for both aerobic and anaerobic culture, three bacteria with differential oxygen requirements (E. coli, A. viscosus, and F. nucleatum), as well as a model mammalian cell line (murine embryonic fibroblast cells (3T3)), were cultured. Growth characteristics of the selected species were analyzed as a function of eight discrete DO concentrations, ranging from 0 ppm (anaerobic) to 42 ppm (fully saturated).

SUBMITTER: Lam RH

PROVIDER: S-EPMC2710860 | biostudies-literature | 2009 Jul

REPOSITORIES: biostudies-literature

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Culturing aerobic and anaerobic bacteria and mammalian cells with a microfluidic differential oxygenator.

Lam Raymond H W RH Kim Min-Cheol MC Thorsen Todd T

Analytical chemistry 20090701 14

In this manuscript, we report on the culture of anaerobic and aerobic species within a disposable multilayer polydimethylsiloxane (PDMS) microfluidic device with an integrated differential oxygenator. A gas-filled microchannel network functioning as an oxygen-nitrogen mixer generates differential oxygen concentration. By controlling the relative flow rate of the oxygen and nitrogen input gases, the dissolved oxygen (DO) concentration in proximal microchannels filled with culture media are precis ...[more]

PMID: 19601655

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Project description:In nature, methylmercury (MeHg) is primarily generated through microbial metabolism, and the ability of bacteria to methylate Hg(II) depends on both bacterial properties and environmental factors. It is widely known that, as a metabolic analog, molybdate can inhibit the sulfate reduction process and affect the growth and methylation of sulfate-reducing bacteria (SRB). However, after it enters the cell, molybdate can be involved in various intracellular metabolic pathways as a molybdenum cofactor; whether fluctuations in its concentration affect the growth and methylation of aerobic mercury methylating strains remains unknown. To address this gap, aerobic γ-Proteobacteria strains Raoultella terrigena TGRB3 (B3) and Pseudomonas putida TGRB4 (B4), as well as an obligate anaerobic δ-Proteobacteria strain of the SRB Desulfomicrobium escambiense CGMCC 1.3481 (DE), were used as experimental strains. The growth and methylation ability of each strain were analyzed under conditions of 500 ng·L-1 Hg(II), 0 and 21% of oxygen, and 0, 0.25, 0.50, and 1 mM of MoO4 2-. In addition, in order to explore the metabolic specificity of aerobic strains, transcriptomic data of the facultative mercury-methylated strain B3 were further analyzed in an aerobic mercuric environment. The results indicated that: (a) molybdate significantly inhibited the growth of DE, while B3 and B4 exhibited normal growth. (b) Under anaerobic conditions, in DE, the MeHg content decreased significantly with increasing molybdate concentration, while in B3, MeHg production was unaffected. Furthermore, under aerobic conditions, the MeHg productions of B3 and B4 were not influenced by the molybdate concentration. (c) The transcriptomic analysis showed several genes that were annotated as members of the molybdenum oxidoreductase family of B3 and that exhibited significant differential expression. These findings suggest that the differential expression of molybdenum-binding proteins might be related to their involvement in energy metabolism pathways that utilize nitrate and dimethyl sulfoxide as electron acceptors. Aerobic bacteria, such as B3 and B4, might possess distinct Hg(II) biotransformation pathways from anaerobic SRB, rendering their growth and biomethylation abilities unaffected by molybdate.

Dataset Information

Culturing aerobic and anaerobic bacteria and mammalian cells with a microfluidic differential oxygenator.

Publications

Culturing aerobic and anaerobic bacteria and mammalian cells with a microfluidic differential oxygenator.

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