Use of a Coulter Counter to Detect Discrete Changes in Cell Numbers and Volume During Growth of Escherichia coli

SMITHER, R.

doi:10.1111/j.1365-2672.1976.tb00586.x

Cited by 5 publications

(5 citation statements)

References 8 publications

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“…Cubana 12007, and Meleagridis DR was 0.61 ,um3, which is essentially the same as that previously obtained for Escherichia coli(6,17). The lethality of the LPS was determined by the female mouse procedure of Peroni et al…”

supporting

confidence: 66%

Biological and Physicochemical Characteristics of Four Serotypes of Salmonella enteritidis

et al. 1978

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Four serotypes of Salmonella enteritidis , Anatum ATCC 9270, Newbrunswick ATCC 1608, Oranienburg 200 E, and Pullorum RM, were studied to determine biological, chemical, or physical differences which might explain variations in Salmonella virulence as previously reported by McCullough and Eisele (J. Infect. Dis. 88 :278-289, 1951; 89 :259-265, 1951). These investigators found that serotype Pullorum was significantly less virulent than serotypes Newport, Derby, Barielly, Meleagridis and Anatum when fed to healthy humans. Results of our own experiments showed that serotype Pullorum RM had a generation time approximately twice that of serotype Anatum 9270. The volume of serotype Pullorum was approximately one-half the volume of the other serotypes used (Anatum 9270, Newbrunswick 1608, Oranienburg 200 E, Cubana 12007, and Meleagridis DR). The number of cells required to yield 1 g dry weight was substantially higher for serotype Pullorum RM than for serotypes Anatum 9270, Newbrunswick 1608, and Oranienburg 200 E. The yield of endotoxin per gram dry weight for serotype Pullorum RM averaged 22 mg/g, whereas yields of endotoxin for serotypes Anatum 9270, Newbrunswick 1608, and Oranienburg 200 E averaged 32 to 35 mg/g. The relative abundance of the four major fatty acids (measured by gas chromatography) also showed distinct differences among the serotypes. Pullorum RM contained less lauric and 3-hydroxymyristic acids and more myristic and palmitic acids than the other three serotypes. The identity of 3-hydroxymyristate was confirmed by mass spectroscopy. Serotype Pullorum RM required 10 times more lipopolysaccharides (endotoxin) to obtain a 50% lethal dose in mice than the other three serotypes. When the lipid part was separated from the polysaccharide and solubilized with bovine serum, the 50% lethal dose of serotype Pullorum RM was equal to that of the other three.

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supporting

confidence: 66%

Biological and Physicochemical Characteristics of Four Serotypes of Salmonella enteritidis

et al. 1978

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“…In the LC-MS/MS assay, each sample had 2 × 10 9 cells using 1 μm 3 as the cellular volume of E. coli. 55,82 The cellular concentration was calculated on the basis of the calculation in a previous study from Cai et al 55 Thus, the total volume of E. coli 2 × 10 9 cells is 2 μL [(2 × 10 9 ) × 1 μm 3 = 2 × 10 9 μm 3 = 2 μL (1 mL = 1 × 10 12 μm 3 )]. Therefore, the cellular concentration was equal to the measured concentration using LC-MS/MS (in μM) multiplied by the extraction volume (150 μL) and divided by the total cellular volume per sample (2 μL).…”

Section: ■ Methodsmentioning

confidence: 99%

Fluorescent Trimethoprim Conjugate Probes To Assess Drug Accumulation in Wild Type and Mutant Escherichia coli

et al. 2016

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Reduced susceptibility to antimicrobials in Gram-negative bacteria may result from multiple resistance mechanisms, including increased efflux pump activity or reduced porin protein expression. Up-regulation of the efflux pump system is closely associated with multidrug resistance (MDR). To help investigate the role of efflux pumps on compound accumulation, a fluorescence-based assay was developed using fluorescent derivatives of trimethoprim (TMP), a broad-spectrum synthetic antibiotic that inhibits an intracellular target, dihydrofolate reductase (DHFR). Novel fluorescent TMP probes inhibited eDHFR activity with comparable potency to TMP, but did not kill or inhibit growth of wild type Escherichia coli. However, bactericidal activity was observed against an efflux pump deficient E. coli mutant strain (ΔtolC). A simple and quick fluorescence assay was developed to measure cellular accumulation of the TMP probe using either fluorescence spectroscopy or flow cytometry, with validation by LC-MS/MS. This fluorescence assay may provide a simple method to assess efflux pump activity with standard laboratory equipment.

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“…Assuming that 3 mL of E. coli at OD 600 = 1.00, contains 3 × 10 8 cells. [36][37][38] Example of intracellular uptake of quercetin 3-β-D-glucoside in E. coli at 1 st hour is as: 1.42 µg/1000 µL × 150 µL = 0.213 µg → 0.213 µg/3 × 10 8 Cells = 7.1 × 10 −10 µg per Cell (note: the final sample was dissolved in 150 µL methanol for the LC-MS from which 3 µl of sample was injected. 1.42 µg mL −1 is the final concentration obtained).…”

Section: Intracellular Concentration Of Flavonoids In E Colimentioning

confidence: 99%

Quantification of microbial uptake of quercetin and its derivatives using an UHPLC-ESI-QTOF mass spectrometry assay

et al. 2016

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A liquid chromatography-mass spectrometry-based method for the quantification of intracellular concentrations of dietary phenolics in bacteria was developed. Using this assay, the time-dependent uptake of quercetin and two of its glycosides into gut microbiota model organisms (Escherichia coli and Bifidobacterium bifidum) under aerobic and anaerobic conditions was studied. For the first time, quantitative data on the uptake of dietary phenols were obtained indicating a time-dependent differential uptake of the dietary compounds. The shape of the uptake curve and the comparative use of bacterial mutants lacking porins OmpFC or the multi-drug efflux pump AcrAB, respectively, along with the identification and quantification of selected bacterial metabolites provided a mechanistic insight into the uptake process.

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Use of a Coulter Counter to Detect Discrete Changes in Cell Numbers and Volume During Growth of Escherichia coli

Cited by 5 publications

References 8 publications

Biological and Physicochemical Characteristics of Four Serotypes of Salmonella enteritidis

Biological and Physicochemical Characteristics of Four Serotypes of Salmonella enteritidis

Fluorescent Trimethoprim Conjugate Probes To Assess Drug Accumulation in Wild Type and Mutant Escherichia coli

Quantification of microbial uptake of quercetin and its derivatives using an UHPLC-ESI-QTOF mass spectrometry assay

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