Development of the Firefly Bioluminescent Assay for the Rapid, Quantitative Detection of Microbal Contamination of Water

Levin, Gilbert V.; Chen, Chi-Sin; Davis, Gretchen

doi:10.21236/ad0659144

Cited by 11 publications

(8 citation statements)

References 3 publications

(3 reference statements)

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“…Variations in ATP level per viable unit observed in treated urine specimens may, among other things, be caused by differences in metabolic rate and bacterial cell size, affecting cellular ATP pools (3,10,12). Furthermore, cell aggregation, causing erroneous culture results, would be expected to be common in this type of material.…”

Section: Discussionmentioning

confidence: 99%

“…3, Pseudomonas sp. (4), various mixed infections (10), and unidentified bacteria (7). by quantitative culture.…”

Section: Oos I0 015mentioning

confidence: 99%

“…The luciferase assay has been shown to have a sensitivity allowing the detection of approximately 10-14 M ATP, corresponding to the ATP content of fewer than 10 bacteria/ml (1,10). This extreme sensitivity, combined with the simple and rapid assay procedure, would make the luciferase assay an attractive alternative to existing methods for quantitation of bacteria.…”

mentioning

confidence: 99%

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Detection of bacteriuria by luciferase assay of adenosine triphosphate

Thore¹,

Ånséhn²,

Lundin³

et al. 1975

J Clin Microbiol

148

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A selective method for distinguishing bacterial and nonbacterial adenosine triphosphate (ATP) in clinical bacteriological specimens was studied. The method involved incubation of samples with the detergent Triton X-100 and the ATP-hydrolyzing enzyme apyrase. The incubation selectively destroyed ATP in suspensions of various human cells while not affecting the ATP content in microbial cells. ATP remaining in the sample after incubation was extracted in boiling buffer and assayed by the firefly luciferase assay. Application of the method to 469 clinical urine specimens showed that the ATP level after treatment with Triton/apyrase was correlated to bacterial counts and that the sensitivity of the assay was sufficient for the detection of 105 bacteria/ml. The ATP levels per bacterial cell remaining in the urine specimens after treatment with Triton/apyrase were close to values observed in laboratory-grown cultures. The specificity and sensitivity of the luciferase assay for the detection of urinary bacteria and its possible use as a bacteriuria screening method are discussed.

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Section: Discussionmentioning

confidence: 99%

“…3, Pseudomonas sp. (4), various mixed infections (10), and unidentified bacteria (7). by quantitative culture.…”

Section: Oos I0 015mentioning

confidence: 99%

mentioning

confidence: 99%

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Detection of bacteriuria by luciferase assay of adenosine triphosphate

Thore¹,

Ånséhn²,

Lundin³

et al. 1975

J Clin Microbiol

148

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“…No 14Clabeled substrate was included. The mixture was aerated vigorously with water-saturated oxygen for 1 min and extracted, and the amount of ATP present was determined as described previously (10). Values were converted from milligrams of protein to grams of dry weight based on a prosthecal protein content of 45.2% (9).…”

mentioning

confidence: 99%

Glucose transport in isolated prosthecae of Asticcacaulis biprosthecum

Larson¹,

Pate²

1976

J Bacteriol

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Active transport of glucose in prosthecae isolated from cells of Asticcacaulis biprosthecum was stimulated by the non-physiological electron donor N,N,N',N'-tetramethyl-p-phenylenediamine dihydrochloride. Glucose uptake was mediated by two transport systems; the apparent Km of the high-affinity system was 1.8 ,tM and that of the low-affinity system was 34 ,uM. Free glucose accumulated within prosthecae at a concentration 60 to 200 times above that present externally, depending on the Km of the system being observed. The glucose transport system in prosthecae was stereospecific for 1-glucose, and neither methyl aD -glucopyranoside nor 2-deoxyglucose was transported. Uptake of glucose was inhibited by N-ethylmaleimide (NEM) and p-chloromercuribenzoate (PCMB), and the inhibition by PCMB but not by NEM was reversed by dithiothreitol. Glucose uptake was also inhibited by the uncoupling agents 5chloro-3-t-butyl-2'-nitrosalicylanilide (S-13), 5-chloro-3-(p-chlorophenyl)-4'-chlorosalicylanilide (S-6), and carbonyl-cyanide m-chlorophenylhydrazone (CCCP) and by the respiratory inhibitor KCN. Efflux of glucose from preloaded prosthecae was induced by PCMB and KCN, but not by S-13 or CCCP. Glucose uptake was not affected by arsenate or an inhibitor of membrane-bound adenosine triphosphatases, N,N'-dicyclohexylcarbodiimide. The lack of inhibition by these two compounds, combined with the extremely low levels of adenosine 5'-triphosphate present in prosthecae, indicates that adenosine 5'-triphosphate is not involved in the transport of glucose by prosthecae.

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“…The underlying assumption in the analysis is that the cytoplasmic volume of the average bacterial cell in a plaque sample is the same as that of S. sanguis. The specific ATP content of a wide variety of bacteria has been assessed and found proportional to the average cell size (12). We, therefore, have underestimated or overestimated the total cell numbers to the extent that the average cytoplasmic volume of plaque microorganisms is less than or greater than our index organism.…”

Section: Resultsmentioning

confidence: 95%

Use of extractable adenosine triphosphate to estimate the viable cell mass in dental plaque samples obtained from monkeys

Robrish¹,

Kemp²,

Bowen³

1978

Appl Environ Microbiol

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The viable cell mass in plaque samples obtained from monkeys was estimated by determining the concentration of extractable adenosine triphosphate (ATP), and total cell mass was estimated by measuring the protein content. The results were expressed in terms of the specific ATP and protein contents of Streptococcus sanguis. The viable counts estimated by these techniques were comparable to or exceeded viable counts obtained by other investigators using conventional bacteriological methods.

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Development of the Firefly Bioluminescent Assay for the Rapid, Quantitative Detection of Microbal Contamination of Water

Cited by 11 publications

References 3 publications

Detection of bacteriuria by luciferase assay of adenosine triphosphate

Detection of bacteriuria by luciferase assay of adenosine triphosphate

Glucose transport in isolated prosthecae of Asticcacaulis biprosthecum

Use of extractable adenosine triphosphate to estimate the viable cell mass in dental plaque samples obtained from monkeys

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