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Chapter 92 : Techniques for Studying Uncultured and Cultured Magnetotactic Bacteria *
Uncultivated magnetotactic bacteria (MTB) can be studied most easily in microcosms set up in the laboratory. In their simplest form, microcosms consist of about one-third sediment together with about two-thirds overlying water and are placed in a flask or aquarium (0.1 to 5 liters) and incubated in dim light at room temperature. Many MTB can easily be detected and collected by taking advantage of their active, directed migration along magnetic field lines. Cell morphology and flagellar patterns of specific MTB can be determined by using negatively stained preparations of cells and transmission electron microscopy. Magnetosomes are electron-dense structures that can be visualized in cells without staining. The high abundance, conspicuous morphology, and magnetic response of the MTB allow for their easy microscopic identification and have made them an attractive subject for the early application of cultivation-independent, rRNA based techniques. Isolated cultures of MTB need to be properly preserved, as both the magnetic polarity and the capability of biomineralizing magnetosomes can be irreversibly lost upon prolonged subcultivation and storage in the lab due to genetic instability. For the establishment of a genetic system for the MTB, the ability to grow cells as colonies on agar plates is a necessity, since colony formation on the surfaces of agar plates is required for clonal selection in genetic analysis.
Key Concept Ranking
- Denaturing Gradient Gel Electrophoresis
Vertical distribution of MTB along chemical gradients in a typical freshwater microcosm. Numbers of MTB (gray bars) were determined by viable cell counts, using the hangingdrop assay as described in the text. Error bars indicate standard deviations. Various biochemical parameters (concentrations of oxygen, nitrate, and sulfide) were determined by using microelectrodes (modified as described in reference 22 ).
Scheme for magnetic collection and axenic isolation of uncultivated MTB. Details of the techniques are given in the text.
Phase-contrast light micrographs of MTB in a hanging drop at different time points after application of a magnetic field. Cells swim in random directions in the absence of a magnetic field (0 s). If a magnetic field is applied, MTB immediately align and swim along the magnetic field lines (arrows). After several minutes, large numbers of bacteria accumulate at the “northern” edge of the drop (next to the south pole of a bar magnet).