Application of a Tetrazolium Salt with a Water-Soluble Formazan as an Indicator of Viability in Respiring Bacteria

Abstract

The tetrazolium salt sodium 3'4{1-[(phenylamino)-carbonyll-3,4-tetrazolium}-bis (4-methoxy-6-nitro)benzenesulfonic acid hydrate (XTT) was examined for use as a colorimetric indicator of viability in respiring bacteria. XTT was reduced to an orange, water-soluble formazan product by Methylosinus trichosporium OB3b, Pseudomonas putida, Escherichia coli, and Bacilus subtilis. Formazan production was proportional to live cell biomass, and XTT was reduced by all cultures in the absence of added electron-coupling agents. XTT reduction by M. trichosporium OB3b was linear over several hours and was stimulated by the presence of an exogenous substrate (methanol). Addition of cyanide to cultures incubated under oxic conditions gave an initial 10-fold increase in XTT reduction. Viability of bacteria incubated in the absence of exogenous carbon substrates was measured as XTT reduction and compared with viability estimates from plate counts. Results obtained with the two methods were generally comparable, but the XTI7 assay was superior when cell recovery on plates was low. Incubation of E. coli for 7 days in the absence of exogenous carbon substrates decreased viability by 90%, whereas the corresponding decreases for cultures ofM. trichosporium OB3b, P. putida, and B. subtilis were less than 4%o.

Ultramicrocells form by reductive division in macroscopic Pseudomonas aerial structures

“…The cellular metabolic activity change in cells during the incubation was investigated using tetrazolium blue chloride as an artificial electron acceptor (Roslev and King, 1993). The metabolic activity of cells from the 1-month-old aerial structure was maintained at around 47% of that of the 1-week sample but no activity with 10-month-old samples, indicating that cells of 10-month-old aerial structures are almost metabolically inactive.…”

“…The metabolic activity of live cells was determined by tetrazolium blue chloride (Aldrich-Sigma) reduction method (Roslev and King, 1993) with modifications. Cells were thoroughly suspended in saline and diluted with saline at an optical density of 0.1 (660 nm).…”

Assessment of metabolic activity of single bacterial cells — new developments in microcolony and dehydrogenase assays

“…For more than a decade, the reduction of tetrazolium salts has been used to determine dehydroge-nase activity in individual bacterial cells [15]. Within the cells the water-soluble, colourless tetrazolium salts are reduced by dehydrogenases coupled to the electron transport system, and hence converted to insoluble, coloured or fluorescent formazans (soluble, coloured formazans are known [16] but obviously cannot provide information on individual cells and thus cannot yield information on heterogeneity within populations).…”

The enhancement of conjugal plasmid pBHR1 transfer between bacteria in the presence of extracellular metabolic products produced by Microcystis aeruginosa

“…1). In our preliminary experiment using tetrazolium salt, sodium 3 0 -{1-[(phenylamino)-car-bonyl]-3,4-tetrazolium}-bis(4-methoxy-6-nitro)benzenesulfonic acid hydrate (XTT) [27,28], the electron transport system (ETS) activity of E. coli S17-1 (pBHR1) in CBY100 medium with EMPs was significantly higher than in CBY100 medium alone (data not shown). It has been suggested that bacterial metabolic activity correlates positively with the conjugal plasmid transfer frequency [3], so the increased ETS activity that was observed in CBY100 medium with EMPs might be a key mechanism in the high conjugal plasmid transfer in the flasks containing either M. aeruginosa or its EMPs.…”