Abstract
Magnetotactic bacteria (MTB) are a diverse group of bacteria that synthesise magnetosomes, magnetic membrane-bound nanoparticles that have a variety of diagnostic, clinical and biotechnological applications. We present the development of rapid methods using flow cytometry to characterize several aspects of the physiology of the commonly-used MTB Magnetospirillum gryphiswaldense MSR-1. Flow cytometry is an optical technique that rapidly measures characteristics of individual bacteria within a culture, thereby allowing determination of population heterogeneity and also permitting direct analysis of bacteria. Scatter measurements were used to measure and compare bacterial size, shape and morphology. Membrane permeability and polarization were measured using the dyes propidium iodide and bis-(1,3-dibutylbarbituric acid) trimethine oxonol to determine the viability and ‘health’ of bacteria. Dyes were also used to determine changes in concentration of intracellular free iron and polyhydroxylakanoate (PHA), a bacterial energy storage polymer. These tools were then used to characterize the responses of MTB to different O2 concentrations and iron-sufficient or iron-limited growth. Rapid analysis of MTB physiology will allow development of bioprocesses for the production of magnetosomes, and will increase understanding of this fascinating and useful group of bacteria.
Highlights
Magnetic nanomaterials are increasingly important products with myriad applications in diverse settings including but not limited to environmental pollution control, information and energy storage[1], catalysis[2], biotechnological[1,3,4] and especially biomedical research[1,5,6,7]
Qualitative evidence of magnetosome production within Magnetotactic bacteria (MTB) can be obtained by observing a shimmering effect in cell suspensions mounted on magnetic stirrer plates, and black coloration of cell suspensions and/or colonies on agar plates, while magneto-spectrophotometric assay of cellular magnetism (Cmag) of suspended cells provides a rapid indirect measure of cellular magnetosome content[24,25]
Reports on the application of flow cytometry (FCM) to MTB are few in number[34,35] and the full power of the technique has not exploited in any case
Summary
Received: 7 July 2017 Accepted: 21 September 2017 Published: xx xx xxxx responses to changing O2 and iron concentration by Magnetospirillum gryphiswaldense strain MSR-1. It is recognised that future widespread application of magnetosomes will, to a large extent, depend on the development of intensified high yielding manufacturing platforms for magnetosomes[10,12,16] Fundamental to this are appropriate means for analysing MTB growth, viability, physiology and biomineralization of magnetic iron minerals, in order to understand and optimise magnetosome formation at any scale, from initial small (millilitre) studies on strain isolation and cultivability in the laboratory, and pilot scale manufacture (10–100 L), to fully fledged commercial production in cubic metre scale bioreactors. We describe methods for determination of cellular concentration, cell size distribution, single-cell physiology and relative changes over time of intracellular contents of PHA and the chelatable iron pool
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