Comparative studies of bacteria with an atomic force microscopy operating in different modes

Bol’shakova, A. V.; Kiselyova, Olga I.; Filonov, A.S.; Frolova, Olga; Lyubchenko, Yuri L.; Яминский, И.В.

doi:10.1016/s0304-3991(00)00075-9

Cited by 135 publications

(97 citation statements)

References 13 publications

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“…P. aeruginosa wild and cured type cultures were cultured in LB medium and incubated at 37 °C with agitation for 12 h. Cells were harvested by centrifugation at 1, 200 g for 15 min at 4 o C. The pellet was washed twice with sterile milliQ water. Samples for AFM analysis were mounted on cover glass and airdried as described earlier (Bolshakova et al, 2001) and mounted directly on the specimen metal disc using a double adhesive tape. Samples were scanned at different areas using AFM (Shimadzu SPM 9500-2J).…”

Section: Methodsmentioning

confidence: 99%

Plasmid profile and curing analysis of Pseudomonas aeruginosa as metal resistant

Raja

Selvam

2009

Int. J. Environ. Sci. Technol.

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AB STRACT:The isolate Pseudomonas aeruginosa exhibited resistance to heavy metals such as cadmium, chromium, nickel and lead. Plasmid DNA was isolated from P. aeruginosa and designated as pBC15. The size of the plasmid DNA was approximately 23 kb. Escherichia coli DH5α was transformed with plasmid pBC15 subsequired resistance to nickel and ampicillin. The same size of the plasmid was isolated from E. coli transformant and separated on 0.7 % agarose gel electrophoresis. The restriction analysis of pBC15 showed that the plasmid DNA has single site for Bam HI and Eco RI and three sites for Xho I which were compared with 1 Kb DNA and λ Hind III digest molecular markers. Therefore, the size of the plasmid DNA of pBC15 was confirmed to be 23 kb. Curing was carried out by ethidium bromide, acridine orange, novobiocin, sodium dodechyl sulphate and elevated temperature (40 °C). Transformation and curing results suggest that nickel and ampicillin resistance gene was conferred by plasmid DNA. Cadmium resistant gene was present on chromosomal DNA along with the gene for chromium resistance. Lead resistance gene was shown to be present on the chromosomal DNA rather than the plasmid DNA as the cured and uncured cultures remained similar in lead resistance. Therefore, the ability of P. aeruginosa resistant to nickel and ampicillin is plasmid mediated and transferable to other strains whereas cadmium, chromium and lead could be chromosomal encoded. The heavy metal and antibiotic resistances of P. aeruginosa can be used to exploit for clean up industrial wastewater and bioremediation of heavy metal contaminated soil.

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Section: Methodsmentioning

confidence: 99%

Plasmid profile and curing analysis of Pseudomonas aeruginosa as metal resistant

Raja

Selvam

2009

Int. J. Environ. Sci. Technol.

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“…Bacteria were attached through electrostatic interactions (physical adsorption) to both a glass slide and a tip-less cantilever, made positively charged through adsorption of poly L-lysine (Bolshakova et al, 2001). In order to coat the glass surface with poly L-lysine, the glass slide was cleaned by dipping in methanol, and rinsing with demineralized water, after which a drop of 0?01 % (w/v) poly L-lysine (Sigma) solution was added, and spread over the surface.…”

Section: Methodsmentioning

confidence: 99%

Atomic force microscopy study on specificity and non-specificity of interaction forces between Enterococcus faecalis cells with and without aggregation substance

et al. 2005

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Enterococcus faecalis is one of the leading causes of hospital-acquired infections, and indwelling medical devices are especially prone to infection. E. faecalis expressing aggregation substance (Agg) adheres to biomaterial surfaces by means of positive cooperativity, i.e. the ability of one adhering organism to stimulate adhesion of other organisms in its immediate vicinity. In this study, atomic force microscopy (AFM) was used to measure the specificity and non-specificity of interaction forces between E. faecalis cells with and without Agg. Bacteria were attached to a substratum surface and a tip-less cantilever. Two E. faecalis strains expressing different forms of Agg showed nearly twofold higher interaction forces between bacterial cells than a strain lacking Agg [adhesive force (F adh ), "1?3 nN]. The strong interaction forces between the strains with Agg were reduced after adsorption of antibodies against Agg from "2?6 and "2?3 nN to "1?2 and "1?3 nN, respectively. This suggests that the non-specific interaction force between the enterococci amounts to approximately 1?2 nN, while the specific force component is only twofold stronger. Comparison of the results of the AFM interaction forces with the positive cooperativity after adhesion to a biomaterial in a parallel-plate flow chamber showed that in the absence of strong interaction forces between the cells, positive cooperativity was also absent. In conclusion, this is believed to be the first time that the influence of specific antibodies on interaction forces between E. faecalis cells has been demonstrated by AFM, thereby experimentally distinguishing between specific and non-specific force components.

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“…Another is the drop-and-dry method, in which samples are immobilized by evaporating the solution to form a film. 11,12 However, because air-drying needs a relatively long period of time, bacterial cells are difficult to maintain their original living state. Nevertheless, the heat-fixation method can help retain bacterial morphology by coagulating the cytoplasm and enhance the adhesion of cells to glass slides via coagulation of surface proteins of bacteria onto the glass slides.…”

Section: Resultsmentioning

confidence: 99%

Heat-fixation Method Used in an Atomic Force Microscopy Study of Cell Morphology

Zhang

Shi

et al. 2008

ANAL. SCI.

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Extremely halophilic archaea are a diverse group of prokaryotes that inhabit highly saline environments. H. salinarum, a species of archaebacteria, is a photosynthetic bacteria that can survive in 4 M NaCl solutions. When the concentration of Na + is low, the cell wall will break apart and the cell will be lysed. 1 To date, the reason why H. salinarum requires a high concentration of sodium ions for growth has not been fully explained. Archaebacteria have a different morphology than other prokaryotes, such as E. coli. 2,3 As is known, the functions of the cells are related to the structure of the cell membrane. Thus, investigations of the structure of the H. salinarum surface are quite significant. Atomic force microscopy (AFM) is a nondestructive imaging technique with nanometer resolution. Since its introduction in 1986, AFM has been used more extensively in the field of life sciences. [4][5][6] AFM images can reveal the surface ultrastructure of living microbial cells with unprecedented resolution. [7][8][9] AFM can be operated in several operational modes, including contact, tapping and MAC modes. 10In the MAC mode, an alternating magnetic field, generated by a solenoid placed under the sample, is used to directly drive a magnetically coated cantilever to enable a stable operation. The advantages of the MAC mode are high resolution and easy operation in fluids for in situ studies, and little harm to soft biological samples due to intermittent contact.For AFM imaging, samples must be immobilized on a substrate. A variety of methods have been used to immobilize cells for AFM imaging, including trapping living cells in filtrate films, placing cells in agar, or immobilizing cells onto a surface such as mica or glass. 4 In microbiology, the heat-fixation method is widely used for immobilizing bacterial cells onto glass slides for observing cell morphology by optical microscopy. For low-resolution observations with an optical microscope, it seems that this method does not affect the cell morphology. However, the effect of this method on the ultrastructure of the cell morphology has not been examined.Herein, the effect of the heat-fixation method on cell morphology studied by MAC mode atomic force microscopy is reported. Escherichia coli (E. coli) and Bacillus subtilis (B. subtilis) were investigated, respectively, as representatives of Gram-negative bacteria and Gram-positive bacteria. The heatfixation method was then employed to prepare H. salinarum samples for AFM study.Details of the ultrastructural morphology of H. salinarum surface were obtained by MAC mode AFM. ExperimentalCulture of bacteria E. coli JM109 and B. subtilis DB104 were grown in LuriaBertani (LB). H. salinarum R1 was grown in a medium containing 0.25% lactalbumin hydrolysate, 0.2% yeast extract, 25% NaCl and 3% magnesium chloride heptahydrate at pH 7.0. The three species of bacteria were cultured at 37˚C with shaking and harvested in the late exponential phase of growth. Bacteria were centrifuged at 5000 rpm for 4 min, and the wet pellets were r...

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Comparative studies of bacteria with an atomic force microscopy operating in different modes

Cited by 135 publications

References 13 publications

Plasmid profile and curing analysis of Pseudomonas aeruginosa as metal resistant

Plasmid profile and curing analysis of Pseudomonas aeruginosa as metal resistant

Atomic force microscopy study on specificity and non-specificity of interaction forces between Enterococcus faecalis cells with and without aggregation substance

Heat-fixation Method Used in an Atomic Force Microscopy Study of Cell Morphology

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