With respect to coefficient of linear thermal expansion, bacterial vegetative cells and spores resemble plastics and metals, respectively
© Nakanishi et al.; licensee BioMed Central Ltd. 2013
Received: 13 August 2013
Accepted: 2 October 2013
Published: 9 October 2013
If a fixed stress is applied to the three-dimensional z-axis of a solid material, followed by heating, the amount of thermal expansion increases according to a fixed coefficient of thermal expansion. When expansion is plotted against temperature, the transition temperature at which the physical properties of the material change is at the apex of the curve. The composition of a microbial cell depends on the species and condition of the cell; consequently, the rate of thermal expansion and the transition temperature also depend on the species and condition of the cell. We have developed a method for measuring the coefficient of thermal expansion and the transition temperature of cells using a nano thermal analysis system in order to study the physical nature of the cells.
The tendency was seen that among vegetative cells, the Gram-negative Escherichia coli and Pseudomonas aeruginosa have higher coefficients of linear expansion and lower transition temperatures than the Gram-positive Staphylococcus aureus and Bacillus subtilis. On the other hand, spores, which have low water content, overall showed lower coefficients of linear expansion and higher transition temperatures than vegetative cells. Comparing these trends to non-microbial materials, vegetative cells showed phenomenon similar to plastics and spores showed behaviour similar to metals with regards to the coefficient of liner thermal expansion.
We show that vegetative cells occur phenomenon of similar to plastics and spores to metals with regard to the coefficient of liner thermal expansion. Cells may be characterized by the coefficient of linear expansion as a physical index; the coefficient of linear expansion may also characterize cells structurally since it relates to volumetric changes, surface area changes, the degree of expansion of water contained within the cell, and the intensity of the internal stress on the cellular membrane. The coefficient of linear expansion holds promise as a new index for furthering the understanding of the characteristics of cells. It is likely to be a powerful tool for investigating changes in the rate of expansion and also in understanding the physical properties of cells.
KeywordsScanning probe microscope Nano thermal analysis Coefficient of liner thermal expansion Transition temperature Microorganisms Spore
When solid materials are heated, they expand and generally exhibit a thermal creep curve. If a fixed stress is applied to the three-dimensional z-axis of a material and the material is then heated, the amount of expansion resulting from thermal stress increases according to a fixed coefficient of thermal expansion . As the temperature approaches the transition temperature, at which the physical properties of the material change, the rate of expansion decreases and the material reaches its maximum expansion. If expansion is plotted against temperature, the transition temperature is at the apex of the curve. In the case of a solid, the transition temperature may be determined as the melting point of the solid . Different materials, such as metal and plastic, differ in their melting points and their patterns of thermal expansion, so they provide different curves. Microbial cells are not made of a single solid material, but rather of various constituent materials, and the composition varies according to the species and the condition of the cell. The rate of expansion and the transition temperature determined from the amount of expansion thus differ depending on the species and the condition of the cell, and may therefore offer a new approach for the study of cellular structure. However, currently there are no valid methods for determining the rate of expansion and the transition temperature from the amount of expansion of a single cell, and to date there have been no studies conducted on this topic. To address this, we have developed a method for measuring the coefficient of thermal expansion and the transition temperature of microbial samples using a nano thermal analysis (nano-TA) system.
Results and discussion
Change in the coefficient of linear expansion and its relationship to the transition temperature, determined by nano-TA-SPM
The coefficient of linear expansion and transition temperature of bacteria and yeast
The transition temperature and the coefficient of linear expansion of different bacteria, yeast, and plastic materials
Bacteria, yeast, and plastic materials
Coefficient of linear expansion (×10-6/°C)
58 ± 0.7
190 ± 10.5
48 ± 0.5
360 ± 13.5
71 ± 1.8
105 ± 7.5
56 ± 0.9
230 ± 16.5
54 ± 1.2
280 ± 12.5
172 ± 1.2
8 ± 0.3
131 ± 2.0
11 ± 0.2
125 ± 2.5
14 ± 0.6
107 ± 1.1
19 ± 0.5
113 ± 0.9
18 ± 0.5
239 ± 6.1
5 ± 0.2
289 ± 6.7
4 ± 0.3
122 ± 3.5
102 ± 7.5
70 ± 5.5
65 ± 4.5
Comparison of the coefficient of linear expansion and transition temperature between bacteria, yeast, and materials
Cells may be characterized by the coefficient of linear expansion as a physical index, and the coefficient of linear expansion may also characterize cells structurally since it relates to volumetric changes, surface area changes, the degree of expansion of water contained within the cell, and the intensity of the internal stress on the cellular membrane. The coefficient of linear expansion holds promise as a new index for furthering the understanding of the characteristics of cells. One of the problems in the future includes a difference in the water content of the vegetative cells and the spores. Estimating the possibility to influence the difference that this difference is a coefficient of linear thermal expansion to be high. We would investigate whether the quantity of water contained in a cell effects on the coefficient of linear thermal expansion. It is likely to be a powerful tool for investigating changes in the rate of expansion and also in understanding the physical properties of cells.
Bacterial and yeast strains
The spores used were prepared from Geobacillus stearothermophilus NBRC 13737, Bacillus coagulans DSM 1, B. subtilis NBRC 13719T, B. megaterium NBRC 15308T, B. licheniformis NBRC 12200, Thermoanaerobacter mathranii DSM 11426, and Moorella thermoacetica DSM521T. The vegetative cells were Staphylococcus aureus NBRC 100910, Escherichia coli IFO 3301, B. subtilis NBRC 13719T, and Pseudomonas aeruginosa ATCC 10145, and the yeast Saccharomyces pastorianus RIB 2010.
Culture and pretreatment methods
Bacteria were cultured in nutrient broth (Difco, Becton Dickinson and Co., Franklin Lakes, NJ, USA); G. stearothermophilus was cultured at 60°C; all other bacteria were cultured at 35°C. For vegetative cells, the log phase (OD600 = 0.8 – 1.0) after 4 to 12 h of culture was used. Where spores were used, the bacteria were cultured under the same conditions for 96 h . For T. mathranii and M. thermoacetica spores, the bacteria were cultured in modified TGC culture medium (Nissui Pharmaceutical Co., Ltd., Tokyo, Japan) at 60°C for 72 h. The yeast S. pastorianus was cultured in YM Broth (Difco, Becton Dickinson) at 25°C for 48 h. Spores were collected from the culture fluid as reported previously .
The thin films of the plastic materials used were polycaprolactone (PCL; Tm = 55°C, Wako, Tokyo, Japan) and polyethylene (PE; Tm = 116°C, Wako, Tokyo, Japan), which were processed into thin films, as well as polyethylene terephtalate (PET; Tm = 235°C, Pana Chemical Co., Ltd., Tokyo, Japan) and polyamide 66 (nylon 66; Tm = 256°C, Murakami Dengyo Co., Ltd., Yokohama, Japan).
Measurement of transition temperature and coefficient of linear thermal expansion of bacteria
A Nano Search Microscope type SFT-3500 (Shimadzu Corporation, Kyoto, Japan) was combined with a nano-TA system (nano thermal analysis) (Anasys Instruments, Santa Barbara, CA, USA) [2, 3]. The cantilever was brought into contact with a single microbial cell at a constant stress of 200 nN and heated from 25°C at 10°C/s to a temperature of 100°C or 400°C, continuously. The measurement point was the highest point, determined as reported previously .
We would like to thank Dr. Daisuke Imamura of Okayama University for his invaluable advice.
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