1887

Abstract

A manometric assay system employing ascorbate and -tetra-methyl--phenylenediamine (TMPD) was used to quantitate terminal oxidase activity in bacterial non-proliferating whole cells. A wide variety of physiologically diverse bacteria, all of which were grown heterotrophically, was tested by this assay. For this survey study, 79 bacterial strains, which represented 34 genera, were used. Turbidimetrically standardized resting (non-proliferating) cell suspensions were prepared from cells harvested at the late logarithmic growth phase; all cells were grown under identical nutritional conditions. The TMPD oxidase activity obtained quantitatively correlated exceptionally well with results of the standard Kovacs oxidase test. In fact, the increased sensitivity of this quantitative assay allowed for further reclassification within the two major divisions of Kovacs oxidase-positive and -negative groups. Groups I and II contained all of the oxidase-positive microorganisms and the bacteria listed in group I had the highest TMPD oxidase rates, the Q values (microliters of O consumed per hour per milligram [dry weight] at 30 C) ranging from 393 to 2, 164. The organisms listed in group II still had moderately high TMPD oxidase activity, the Q values ranging from 27 to 280. All oxidase-negative bacteria fell into groups III and IV. Bacteria in group III had low but still measurable TMPD oxidase rates, the Q values ranging from 3 to 33, whereas the bacteria found in group IV were inert and unable to oxidize TMPD. A grouping analysis allowed for the resolution of that point which separates oxidase-positive from oxidase-negative bacteria. This point, for non-proliferating cells, was found to be an absolute TMPD oxidation Q value of 33 (after correcting for the endogenous rate by subtraction) and a Q (TMPD/endogenous) ratio of 5; the latter parameter indicated that the uncorrected TMPD oxidation Q value had to be five times greater than the rate for endogenous respiration. All Kovacs oxidase-positive organisms were found to have TMPD oxidase Q values greater than these two metabolic parameters, whereas all Kovacs oxidase-negative organisms had lower values.

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1976-04-01
2022-12-09
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References

  1. Boswell P. A., Batstone G. F., Mitchell R. G. 1972; The oxidase reaction in the classification of the Micrococcaceae. J. Med. Microbiol 5:267–269
    [Google Scholar]
  2. Buchanan R. E., Gibbons N. E. 1974 Bergey’s manual of determinative bacteriology, 8th. Williams and Wilkins Co.; Baltimore:
    [Google Scholar]
  3. Conover T. E. 1970; On the occurrence of respiratory components in calf thymus nuclei. II. Cytochrome oxidase activity. Arch. Biochem. Biophys 136551562
    [Google Scholar]
  4. Deibel R. H., Evans J. B. 1960; Modified benzidine test for the detection of cytochrome-containing respiratory systems in micro-organisms. J. Bacteriol 79:356–360
    [Google Scholar]
  5. Dietrich W. E. Jr., Biggins J. 1971; Respiratory mechanisms in the Flexibacteriaceae: terminal oxidase systems of Saprospira grandis and Vitreoscilla species. J. Bacteriol 105:1083–1089
    [Google Scholar]
  6. Dolin M. I. 1961 Cytochrome-independent electron transport enzymes in bacteria. 425–460 In Gunsalus I. C., Stanier R. Y. The bacteria 2: Academic Press Inc.; New York:
    [Google Scholar]
  7. Jurtshuk P., Aston P. R., Old L. 1967; Enzymatic oxidation of tetramethyl-p-phenylenediamine and p-phenylenediamine by the electron transport particulate fraction of Azotobacter vinelandii. J. Bacteriol 93:1069–1078
    [Google Scholar]
  8. Jurtshuk P., Marcucci O. M., McQuitty D. N. 1975; Tetramethyl-p-phenylenediamine oxidase reaction in Azotobacter vinelandii. Appl. Microbiol 30:951–958
    [Google Scholar]
  9. Jurtshuk P., May A. D., Pope L. M., Aston P. R. 1969; Comparative studies on succinate and terminal oxidase activity in microbial and mammalian electron transport systems. Can. J. Microbiol 15797807
    [Google Scholar]
  10. Jurtshuk P., Milligan T. W. 1974; Quantitation of the tetramethyl-p-phenylenediamine oxidase reaction in Neisseria species. Appl. Microbiol 28:1079–1081
    [Google Scholar]
  11. Jurtshuk P., Milligan T. W. 1974; Preliminary characterization studies on the Neisseria catarrhalis electron transport chain. J. Bacteriol 120:552–555
    [Google Scholar]
  12. Jurtshuk P., Mueller T. J., Acord W. C. 1975; Bacterial terminal oxidases. CRC Crit. Rev. Microbiol 3:399–468
    [Google Scholar]
  13. Jurtshuk P., Old L. 1967; Cytochrome c oxidation by the electron transport fraction of Azotobacter vinelandii J. Bacteriol 95:1790–1797
    [Google Scholar]
  14. Kovacs N. 1956; Identification of Pseudomonas pyocyanea by the oxidase reaction. Nature (London) 178703
    [Google Scholar]
  15. Liu C. Y., Webster D. A. 1974; Spectral characteristics and interconversions of the reduced, oxidized and oxygenated forms of purified cytochrome o. J. Biol. Chem 249:4261–4266
    [Google Scholar]
  16. Revsin B., Marquez E. D., Brodie A. F. 1970; Cytochromes from Mycobacterium phlei. I. Isolation and spectral properties of a mixture of cytochromes (axa3) (o). Arch. Biochem. Biophys 139:114–120
    [Google Scholar]
  17. Revsin B., Marquez E. D., Brodie A. F. 1970; Cytochromes from Mycobacterium phlei. II. Ascorbate reduction of an isolated cytochrome (a+a3) (o) complex. Arch. Biochem. Biophys 136:563–573
    [Google Scholar]
  18. Shipp W. S. 1972; Cytochromes of Escherichia coli. Arch. Biochem. Biophys 150:459–472
    [Google Scholar]
  19. Stanier R. Y., Palleroni N. J., Doudoroff M. 1966; The aerobic pseudomonads: a taxonomic study. J. Gen. Microbiol 43:159–271
    [Google Scholar]
  20. Steel K. J. 1961; The oxidase reaction as a taxonomic tool. J. Gen. Microbiol 25:297–306
    [Google Scholar]
  21. Steel K. J. 1962; The oxidase activity of staphylococci. J. Appl. Bacteriol 25:445–447
    [Google Scholar]
  22. Webster D. A., Liu C. Y. 1974; Reduced nicotinamide adenine dinucleotide-cytochrome o purified from Vitreoscilla. J. Biol. Chem 249:4257–4260
    [Google Scholar]
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