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Functionalized Buckyballs for Visualizing Microbial Species in Different States and Environments.

Cheng Q, Aravind A, Buckley M, Gifford A, Parvin B - Sci Rep (2015)

Bottom Line: To date, in situ visualization of microbial density has remained an open problem.Here, functionalized buckyballs (e.g., C60-pyrrolidine tris acid) are shown to be a versatile platform that allows internalization within a microorganism without either adhering to the cell wall and cell membrane or binding to a matrix substrate such as soil.We also demonstrate that cysteine-functionalized C60-pyrrolidine tris acid can differentiate live and dead microorganisms.

View Article: PubMed Central - PubMed

Affiliation: Department of Electrical and Biomedical Engineering, University of Nevada, Reno, 1664 N Virginia Street, Reno NV, 89503, USA.

ABSTRACT
To date, in situ visualization of microbial density has remained an open problem. Here, functionalized buckyballs (e.g., C60-pyrrolidine tris acid) are shown to be a versatile platform that allows internalization within a microorganism without either adhering to the cell wall and cell membrane or binding to a matrix substrate such as soil. These molecular probes are validated via multi-scale imaging, to show association with microorganisms via fluorescence microscopy, positive cellular uptake via electron microscopy, and non-specific binding to the substrates through a combination of fluorescence and autoradiography imaging. We also demonstrate that cysteine-functionalized C60-pyrrolidine tris acid can differentiate live and dead microorganisms.

No MeSH data available.


Cellular uptake of microorganisms embedded in different matrices and monitored by Autoradiography following several washes.Left and right columns indicate uptake by 14C- and 125I- labelled C60-pyrrolidine tris acid on B. subtilis and E. coli, respectively. The data suggests a residual signal that is presumably due to the uptake of microorganisms, since Fig. 7 indicates non-stickiness to the same substrates.
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f8: Cellular uptake of microorganisms embedded in different matrices and monitored by Autoradiography following several washes.Left and right columns indicate uptake by 14C- and 125I- labelled C60-pyrrolidine tris acid on B. subtilis and E. coli, respectively. The data suggests a residual signal that is presumably due to the uptake of microorganisms, since Fig. 7 indicates non-stickiness to the same substrates.

Mentions: Natural soil is a complicated biomaterial, hosting thousands of microorganisms with intrinsic organic and inorganic matters that hinder probe delivery. 14C- and 125I-radiolabeled C60-pyrrolidine tris acid are incubated with a mixture of soil and microorganisms and then washed to remove excess probes as before. The autoradiography, shown in Fig. 8, indicates strong β- and γ− radiation from the mixture of soil and microorganisms. Comparison of this result with both (i) Fig. 4a,b, which indicated association with microorganisms, and (ii) Fig. 7a,b, which indicated non-stickiness to the matrix, suggests that radiotracers can label microbes in their native environment. Moreover, interesting observations are made when β-radiation is quantified using a liquid scintillation counter, comparing both control and treated matrices with 14C-labeled C60-pyrrolidine tris acid. All control matrices (e.g., background) show around 1,000 counts per second (Fig. 9, green columns), while 14C-labeled C60-pyrrolidine tris acid incubated with glass beads, alumina, VWR sand, wild sand, and natural soil show 2,000, 2,000, 2,000, 4,000, and 6,000 counts per second respectively (Fig. 9, red columns). These results suggest that the natural microorganisms in wild sand and natural soil have successfully taken up 14C-labeled C60-pyrrolidine tris acid, which accounts for the increased number of counts per second.


Functionalized Buckyballs for Visualizing Microbial Species in Different States and Environments.

Cheng Q, Aravind A, Buckley M, Gifford A, Parvin B - Sci Rep (2015)

Cellular uptake of microorganisms embedded in different matrices and monitored by Autoradiography following several washes.Left and right columns indicate uptake by 14C- and 125I- labelled C60-pyrrolidine tris acid on B. subtilis and E. coli, respectively. The data suggests a residual signal that is presumably due to the uptake of microorganisms, since Fig. 7 indicates non-stickiness to the same substrates.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4561912&req=5

f8: Cellular uptake of microorganisms embedded in different matrices and monitored by Autoradiography following several washes.Left and right columns indicate uptake by 14C- and 125I- labelled C60-pyrrolidine tris acid on B. subtilis and E. coli, respectively. The data suggests a residual signal that is presumably due to the uptake of microorganisms, since Fig. 7 indicates non-stickiness to the same substrates.
Mentions: Natural soil is a complicated biomaterial, hosting thousands of microorganisms with intrinsic organic and inorganic matters that hinder probe delivery. 14C- and 125I-radiolabeled C60-pyrrolidine tris acid are incubated with a mixture of soil and microorganisms and then washed to remove excess probes as before. The autoradiography, shown in Fig. 8, indicates strong β- and γ− radiation from the mixture of soil and microorganisms. Comparison of this result with both (i) Fig. 4a,b, which indicated association with microorganisms, and (ii) Fig. 7a,b, which indicated non-stickiness to the matrix, suggests that radiotracers can label microbes in their native environment. Moreover, interesting observations are made when β-radiation is quantified using a liquid scintillation counter, comparing both control and treated matrices with 14C-labeled C60-pyrrolidine tris acid. All control matrices (e.g., background) show around 1,000 counts per second (Fig. 9, green columns), while 14C-labeled C60-pyrrolidine tris acid incubated with glass beads, alumina, VWR sand, wild sand, and natural soil show 2,000, 2,000, 2,000, 4,000, and 6,000 counts per second respectively (Fig. 9, red columns). These results suggest that the natural microorganisms in wild sand and natural soil have successfully taken up 14C-labeled C60-pyrrolidine tris acid, which accounts for the increased number of counts per second.

Bottom Line: To date, in situ visualization of microbial density has remained an open problem.Here, functionalized buckyballs (e.g., C60-pyrrolidine tris acid) are shown to be a versatile platform that allows internalization within a microorganism without either adhering to the cell wall and cell membrane or binding to a matrix substrate such as soil.We also demonstrate that cysteine-functionalized C60-pyrrolidine tris acid can differentiate live and dead microorganisms.

View Article: PubMed Central - PubMed

Affiliation: Department of Electrical and Biomedical Engineering, University of Nevada, Reno, 1664 N Virginia Street, Reno NV, 89503, USA.

ABSTRACT
To date, in situ visualization of microbial density has remained an open problem. Here, functionalized buckyballs (e.g., C60-pyrrolidine tris acid) are shown to be a versatile platform that allows internalization within a microorganism without either adhering to the cell wall and cell membrane or binding to a matrix substrate such as soil. These molecular probes are validated via multi-scale imaging, to show association with microorganisms via fluorescence microscopy, positive cellular uptake via electron microscopy, and non-specific binding to the substrates through a combination of fluorescence and autoradiography imaging. We also demonstrate that cysteine-functionalized C60-pyrrolidine tris acid can differentiate live and dead microorganisms.

No MeSH data available.