Intracellular Water Exchange for Measuring the Dry Mass, Water Mass and Changes in Chemical Composition of Living Cells

Delgado, Francisco Feijó; Cermak, Nathan; Hecht, Vivian; Son, Sungmin; Li, Yingzhong; Knudsen, Scott M.; Ölçüm, Selim; Higgins, John M.; Chen, Jianzhu; Grover, William H.; Manalis, Scott R.

doi:10.1371/journal.pone.0067590

Cited by 140 publications

(125 citation statements)

References 37 publications

Supporting

Mentioning

111

Contrasting

Order By: Relevance

“…The buoyant mass of a spore in solutions of two different densities can be used to calculate the mass, volume, and density of the cell. Likewise, measurements in H 2 O and D 2 O solutions can be used to separately quantify the dry and aqueous contents (31). To determine the distribution across the population, the buoyant mass of ϳ1,000 individual hydrated B. subtilis (PS533; wild-type) spores is determined by weighing them in an SMR.…”

Section: Resultsmentioning

confidence: 99%

“…1a. The chip containing the SMR is mounted on a fluidic manifold, and computer-controlled pressure regulators with pressurized glass sample vials are used to precisely control fluid flow within the SMR as previously described (31). Spores are suspended in the desired solution, allowed to equilibrate for 30 min, and loaded into the sample bypass channel.…”

Section: Methodsmentioning

confidence: 99%

See 1 more Smart Citation

Water and Small-Molecule Permeation of Dormant Bacillus subtilis Spores

et al. 2016

Self Cite

View full text Add to dashboard Cite

We use a suspended microchannel resonator to characterize the water and small-molecule permeability of Bacillus subtilis spores based on spores' buoyant mass in different solutions. Consistent with previous results, we found that the spore coat is not a significant barrier to small molecules, and the extent to which small molecules may enter the spore is size dependent. We have developed a method to directly observe the exchange kinetics of intraspore water with deuterium oxide, and we applied this method to wild-type spores and a panel of congenic mutants with deficiencies in the assembly or structure of the coat. Compared to wild-type spores, which exchange in approximately 1 s, several coat mutant spores were found to have relatively high water permeability with exchange times below the ϳ200-ms temporal resolution of our assay. In addition, we found that the water permeability of the spore correlates with the ability of spores to germinate with dodecylamine and with the ability of TbCl 3 to inhibit germination with L-valine. These results suggest that the structure of the coat may be necessary for maintaining low water permeability. IMPORTANCESpores of Bacillus species cause food spoilage and disease and are extremely resistant to standard decontamination methods. This hardiness is partly due to spores' extremely low permeability to chemicals, including water. We present a method to directly monitor the uptake of molecules into B. subtilis spores by weighing spores in fluid. The results demonstrate the exchange of core water with subsecond resolution and show a correlation between water permeability and the rate at which small molecules can initiate or inhibit germination in coat-damaged spores. The ability to directly measure the uptake of molecules in the context of spores with known structural or genetic deficiencies is expected to provide insight into the determinants of spores' extreme resistance.

show abstract

Section: Resultsmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

Water and Small-Molecule Permeation of Dormant Bacillus subtilis Spores

et al. 2016

Self Cite

View full text Add to dashboard Cite

show abstract

“…In comparison to other reports on physical measurements of E. coli, Godin et al report a density of 1.160 Ϯ 0.001g/ml for E. coli (one shot with ampicillin resistance) using the same buoyant mass measurement method in PBS and Histodenz solution without exposure to any stressor (20). Similarly, Feijó Delgado et al report 727 Ϯ 15 fg as the total cell mass value for fixed E. coli (ATCC 23725) (26). Overall, these findings illustrate how buoyant mass measurements for cell populations can be used to determine additional physical features of the cell population.…”

Section: Utilizing Archimedes To Compare Features Of Live and Deadmentioning

confidence: 94%

Mass and Density Measurements of Live and Dead Gram-Negative and Gram-Positive Bacterial Populations

Lewis

Craig

Senecal

2014

Appl Environ Microbiol

View full text Add to dashboard Cite

Monitoring cell growth and measuring physical features of food-borne pathogenic bacteria are important for better understanding the conditions under which these organisms survive and proliferate. To address this challenge, buoyant masses of live and dead Escherichia coli O157:H7 and Listeria innocua were measured using Archimedes, a commercially available suspended microchannel resonator (SMR). Cell growth was monitored with Archimedes by observing increased cell concentration and buoyant mass values of live growing bacteria. These growth data were compared to optical density measurements obtained with a Bioscreen system. We observed buoyant mass measurements with Archimedes at cell concentrations between 10 5 and 10 8 cells/ ml, while growth was not observed with optical density measurements until the concentration was 10 7 cells/ml. Buoyant mass measurements of live and dead cells with and without exposure to hydrogen peroxide stress were also compared; live cells generally had a larger buoyant mass than dead cells. Additionally, buoyant mass measurements were used to determine cell density and total mass for both live and dead cells. Dead E. coli cells were found to have a larger density and smaller total mass than live E. coli cells. In contrast, density was the same for both live and dead L. innocua cells, while the total mass was greater for live than for dead cells. These results contribute to the ongoing challenge to further develop existing technologies used to observe cell populations at low concentrations and to measure unique physical features of cells that may be useful for developing future diagnostics.

show abstract

“…A cell's buoyant mass is its total mass minus the mass of the fluid it displaces. To obtain dry mass (biomass), we combine information from paired buoyant mass measurements performed in H 2 O and D 2 O (Feijó Delgado et al, 2013). In pure H 2 O, a cell's buoyant mass is only the buoyant mass of its dry material, as its intracellular water is neutrally buoyant.…”

Section: Introductionmentioning

confidence: 99%

Direct single-cell biomass estimates for marine bacteria via Archimedes’ principle

et al. 2016

Self Cite

View full text Add to dashboard Cite

Microbes are an essential component of marine food webs and biogeochemical cycles, and therefore precise estimates of their biomass are of significant value. Here, we measured single-cell biomass distributions of isolates from several numerically abundant marine bacterial groups, including Pelagibacter (SAR11), Prochlorococcus and Vibrio using a microfluidic mass sensor known as a suspended microchannel resonator (SMR). We show that the SMR can provide biomass (dry mass) measurements for cells spanning more than two orders of magnitude and that these estimates are consistent with other independent measures. We find that Pelagibacterales strain HTCC1062 has a median biomass of 11.9 ± 0.7 fg per cell, which is five-to twelve-fold smaller than the median Prochlorococcus cell's biomass (depending upon strain) and nearly 100-fold lower than that of rapidly growing V. splendidus strain 13B01. Knowing the biomass contributions from various taxonomic groups will provide more precise estimates of total marine biomass, aiding models of nutrient flux in the ocean.

show abstract

Intracellular Water Exchange for Measuring the Dry Mass, Water Mass and Changes in Chemical Composition of Living Cells

Cited by 140 publications

References 37 publications

Water and Small-Molecule Permeation of Dormant Bacillus subtilis Spores

Water and Small-Molecule Permeation of Dormant Bacillus subtilis Spores

Mass and Density Measurements of Live and Dead Gram-Negative and Gram-Positive Bacterial Populations

Direct single-cell biomass estimates for marine bacteria via Archimedes’ principle

Contact Info

Product

Resources

About