Biologically produced pyocyanin was purified, and the nature of its antibacterial action was determined for several bacteria. The pigment was shown to be bactericidal for all susceptible organisms. The bactericidal effect was dependent upon pyocyanin concentration and resulted in decreases in viability ranging from 1 to 8 log viable cells ml-'. The gram-positive bacteria were more susceptible as a group to the antibiotic, action than were the gram-negative bacteria. All apyocyanogenic pseudomonads tested were totally resistant to the pigment, suggesting that resistance may be a characteristic of the genus. Pseudomonas aeruginosa, the producer organism, was also essentially unaffected by high concentrations of pyocyanin. Facultative anaerobes were twofold or more times resistant to the action of the pigment under fermentative conditions; however, the antibiotic action did not require oxygen since denitriffying bacteria were more susceptible during anaerobic respiration than during aerobic respiration.Pyocyanin is a water-soluble blue-green phenazine pigment produced in large quantities by active cultures of Pseudomonas aeruginosa. Pyocyanin has antibiotic activity against bacteria (4), fungi (8), and protozoa (9), but is of little therapeutic value because it is quite toxic to eucaryotic cells (18). The physiological significance of the pigment is not known, but because of its inhibitory action, it has been postulated that pyocyanin production may give P. aeruginosa a selective advantage in certain growth situations. The nature of bacterial inhibition by the phenazine is neither well understood nor well documented, and no information is available concerning the effect of pyocyanin on the producer organism P. aeruginosa. Recently, Hassan and Fridovich (10) proposed that the inhibitory action of pyocyanin is the result of its unique redox potential. They propose that, during respiration, pyocyanin becomes reduced and univalently reduces oxygen to the toxic superoxide radical. The resistance of various bacteria to pyocyanin would therefore be dependent upon the levels of superoxide dismutase and catalase possessed by the organism and on the presence of oxygen.The purpose of this investigation was to determine the nature of the antibacterial action of pyocyanin and to characterize its inhibitory action on a variety of bacteria, including P. aeruginosa, under different environmental and physiological conditions. The results of our study demonstrate that the antimicrobial action of pyocyanin is bactericidal in nature and that the Media and culture conditions. The semidefined medium used for most of the experimentation contained the following per liter of deionized water: K2HPO4, 7.0 g; NaH2PO4. H20, 3.0 g; MgSO4.7H20, 0.24 g; (NH4)2SO4, 1.0 g; yeast extract, 1.0 g; and Dglucose, 1.0 g. The final pH was 7.0. A combination of the first three ingredients per liter of water will be referred to as basal salts solution (pH 7.0). The entire medium as described will be referred to as medium A. When needed, medium A was supple...

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Growth of S. mutans on mixed or parotid saliva from CF individuals may be influenced by the availability of growth-supportive proteins or the inhibitory activity present in parotid saliva. A deficiency in growth-supportive proteins may explain the limited growth of S. sanguis on mixed or submandibular saliva from these individuals.

The growth responses of Streptococcus mutans VA-29R, Streptococcus sanguis ATCC 10556, and Streptococcus mitior NIH to hydrophilic and hydrophobic peptides obtained following isopropanol fractionation of Trypticase were compared. Although the two fractions contained peptides of similar molecular size, differences were observed with respect to amino acid compositions. S. mutans VA-29R showed a pronounced difference in growth response to hydrophilic vs. hydrophobic peptides. While growth of this micro-organism on hydrophilic peptides was indistinguishable from that on unfractionated Trypticase, only very slow growth occurred on the hydrophobic peptides. S. sanguis ATCC 10556 and S. mitior NIH also displayed some selectivity, as evidenced by their faster relative growth rates on hydrophilic, as compared with hydrophobic, peptides. The results of this study support the conclusion that the properties of the substrate, as defined by its amino acid composition, may be more important than molecular size as a factor influencing recognition and subsequent utilization of oligopeptides as sources of amino acids for growth by these three oral streptococci.

The growth of Streptococcus mutans and Streptococcus sanguis in the oral environment requires that these micro-organisms be able to degrade salivary proteins and to assimilate the resulting peptides as an amino nitrogen source. Our research is aimed at the definition of the proteolytic enzyme systems in these oral streptococci which allow them to utilize such substrates. In the present work, the nature of the hydrolytic activity expressed by S. mutans VA-29R and S. sanguis ATCC 10556 against X-Pro4-nitroanilide and X-Pro-Y tripeptide substrates was investigated. This activity was predominantly associated with a cytoplasmic dipeptidyl peptidase which preferentially catalyzes the release of an N-terminal dipeptide from substrates in which proline is the penultimate residue. These streptococci also possess a second cytoplasmic peptidase, pepD, which catalyzes the hydrolysis of X-Pro dipeptides. We found that Gly-Pro-Ala or Ala-Pro-Gly were transported into the bacterial cells only when an energy source such as glucose was present. Peptide uptake was time-dependent, and selective exodus of peptide-derived amino acids from the bacterial cells occurred during peptide uptake. Results from these studies provide evidence that S. mutans VA-29R and S. sanguis ATCC 10556 possess a pathway for the complete degradation of X-Pro tripeptides. Transport of the peptides into cells prior to hydrolysis provides an efficient way by which all amino acids of a peptide may be obtained at an energy expense equivalent to that associated with the transport of just one amino acid. In light of the abundance of proline in salivary polypeptides, this degradative pathway could be an important component in the proteolytic pathway for salivary polypeptide utilization in these oral streptococci.