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Physiology and Growth |
School of Biochemistry and Molecular Genetics, University of New South Wales, Sydney 2052, Australia1
School of Biosciences, University of Wales Cardiff, Cardiff CF1 3TL, UK2
Author for correspondence: Giancarlo A. Biagini. Tel: +44 151 7053151. Fax: +44 151 7089007. e-mail: Giancarlobiagini{at}hotmail.com
Giardia intestinalis and Hexamita inflata are microaerophilic protozoa which rely on fermentative metabolism for energy generation. These organisms have developed a number of antioxidant defence strategies to cope with elevated O2 tensions which are inimical to survival. In this study, the ability of pyruvate, a central component of their energy metabolism, to act as a physiological antioxidant was investigated. The intracellular pools of 2-oxo acids in G. intestinalis were determined by HPLC. With the aid of a dichlorodihydrofluorescein diacetate-based assay, intracellular reactive oxygen species generation by G. intestinalis and H. inflata suspensions was monitored on-line. Addition of physiologically relevant concentrations of pyruvate to G. intestinalis and H. inflata cell suspensions was shown to attenuate the rate of H2O2- and menadione-induced generation of reactive oxygen species. In addition, pyruvate was also shown to decrease the generation of low-level chemiluminescence arising from the oxygenation of anaerobic suspensions of H. inflata. In contrast, addition of pyruvate to suspensions of respiring Saccharomyces cerevisiae was shown to increase the generation of reactive oxygen species. These data suggest that (i) in G. intestinalis and H. inflata, pyruvate exerts antioxidant activity at physiological levels, and (ii) it is the absence of a respiratory chain in the diplomonads which facilitates the observed antioxidant activity.
Keywords: parasite, oxidative stress, reactive oxygen species, yeast, dichlorodihydrofluorescein diacetate
Abbreviations: SOD, superoxide dismutase
a Present address: Liverpool School of Tropical Medicine, Penbroke Place, Liverpool L3 5QA, UK.
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