Further Evidence of the Existence of Mitochondria in Bacteria

Abstract

The granules of mycobacteria, originally described by Robert Koch as spores, have been shown in a preceding communication to be sites of oxidative-reductive enzymatic activity. In respect to several enzymatic reactions, as well as in tinctorial and morphological characteristics, these mycobacterial granules were shown to correspond to mitochondria (Mudd, Winterscheid, DeLamater, and Henderson, 1951). In the present communication granules in several other species of bacteria have been examined by similar criteria. These granules also have been found to correspond in essential characteristics to mitochondria. METHODS AND MATERIALS The microorganisms used were: Escherichia coli, strain B; Bacillus megatherium, a strain originally given us by Dr. C. F. Robinow; Micrococcus cryophilus, a large gram-variable coccus isolated in the Department of Agriculture (McLean, Sulzbacher, and Mudd, 1951) and utilized for studies on nuclear mitosis (DeLamater and Woodburn, 1951); and Saccharomyces cerevisiae, a strain from baker's yeast and a "cytochromeless" variant, received through the courtesy of Dr. H. M. Davidson, which had been derived as described by Ephrussi and colleagues (Ephrussi, Hottinguer, and Chimenes, 1949; Slonimski and Ephrussi, 1949). The cells of E. coli and B. megatherium used had been grown for 3 to 4 hours at 37 C on Morton and Engley (1945) agar slants or in M. and E. broth. The micrococcus was incubated 24 hours at 20 C on similar agar slants or broth prior to use. Both strains of S. cerevisiae were incubated 24 hours at 37 C onSabouraud's agar slants. The reactions examined for were the reduction of 2,3,5-triphenyltetrazolium and neotetrazolium, probably indicating the action of flavoprotein enzymes, oxidation of the Nadi reagent, indicating cytochrome oxidase, staining with the mitochondrial stain Janus green B, the Baker acid-hematin stain for phospholipid, and the Harman mitochondrial stain. Experimental procedures were followed as in the preceding communication (Mudd, Winterscheid, DeLamater, and Henderson, 1951) with the following modifications: The reduction of triphenyltetrazolium and neotetrazolium was demonstrated I This research has been aided by a grant from the Damon Runyon Fund through the American Cancer Society as recommended by the Committee on Growth of the National Research Council.

Oxidative phosphorylation by subcellular particles from yeast

“…The presence in Sacchlaromye8 cerevisiae of refractile cytoplasmic granules with staining properties resembling those of mitochondria from animal cells has been reported by Mudd et al (1951) and Hartman & Liu (1954). On the basis of earlier evidence Lindegren (1949) had suggested that the cytoplasmic granules of yeast were mitochondrial in nature.…”

“…10-3 x Relative centrifugal force Yield* 0*63 5000 g. The 'respiratory granules' of yeast, as identified by microscopic observation (Mudd et al 1951), have been reported to range from 0-3 to 1 , in diameter. Linnane & Still (1955) estimated that the particles obtained by disintegration in sucrose were about 0-3 ,u in diameter, although the fraction used was precipitated between 3000 and 25 000 g and no fractionation studies were reported.…”

A Cytochemical Localization of Reductive Sites in a Gram-Positive Bacterium

“…With the aid of tetrazolium compounds in particular, but also with the Nadi reagent, Janus Green B, and potassium tellurite, Mudd et al (24)(25)(26)(27)(28)(29)(30)(31) described circumscribed local sites where oxidative-reductive events were found to take place. Weibull (47) pointed out for light microscopy that vital staining of bacteria with tetrazolium cannot be relied upon to reveal the intracellular localization of centres of reducing activity; on the electron microscopic level, there is even less certainty that the formazan is at the site of its formation.…”

LOCALIZATION OF THE SUCCINIC DEHYDROGENASE SYSTEM IN ESCHERICHIA COLI USING COMBINED TECHNIQUES OF CYTOCHEMISTRY AND ELECTRON MICROSCOPY

“…Results of such studies have shown that oxidative enzymes are associated with the cell Membran fraction (1) which contains both cell wall and plasma membrane fragments. Early work (2)(3)(4)(5) involved visual localization of enzyme sites with the use of tctrazolium salts which became reduced to colored formazans. More recently combined techniques of histochcmistry and electron microscopy have been applied to the specific structural localization of respiratory enzyme sites in bacteria (6)(7)(8).…”