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Effect of pH on the Growth and Proteolytic Activity of Porphyromonas gingivalis and Bacteroides intermedius N. Takahashi and C.F. Schachtele J DENT RES 1990 69: 1266 DOI: 10.1177/00220345900690060801 The online version of this article can be found at: http://jdr.sagepub.com/content/69/6/1266

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Effect of pH on the Growth and Proteolytic Activity of Porphyromonas gingivalis and Bacteroides intermedius N. TAKAHASHI' and C.F. SCHACHTELE2 Department of Oral Sciences and Dental Research Institute, School of Dentistry, University of Minnesota, Minneapolis, Minnesota 55455

The effect of pH on the growth and proteolytic activity of the type strain and fresh isolates of Porphyromonas gingivalis and Bacteroides intermedius was investigated. B. internedius strains grew with and without glucose at a pH as low as 5.0. These bacteria grew almost as well as Streptococcus mutans at pH 5.0 and better than Actinomyces viscosus at pH 5.5 and 5.0. Some B. intermedius strains raised the culture pH when grown at a low pH without glucose. In contrast, P. gingivalis strains grew only at pH 6.5 to 7.0. The P. gingivalis strains had proteolytic activities against azocoll, azocasein, and azoalbumin, while the B. intermedius strains degraded azocasein and azoalbumin, but not azocoll. B. internedius showed maximum proteolytic activity at pH 7.0, and high activity over a wide pH range. In contrast, the optimum pH of proteolytic activity in P. gingivalis was pH 7.5 to 8.0. The P. gingivalis activities were more sensitive than those of B. intennedius to low pH. The capacity of B. intermedius to degrade proteins to more readily metabolizable substrates at low pH might explain the growth of this bacterium in an acidic environment. These differences between B. internedius and P. gingivalis could explain their capacity to survive at different sites in the oral cavity and indicate how B. intermedius might positively influence the growth of P. gingivalis in subgingival plaque.

J Dent Res 69(6):1266-1269, June, 1990

Introduction. Various oral bacteroides and members of phenotypically similar genera have been implicated in the development of periodontal disease in humans. Porphyromonas gingivalis is usually absent in samples of plaque from healthy gingival sulci (Slots, 1977), but it can represent up to 5% of the cultivable subgingival flora from individuals with gingivitis (Slots, 1982; Slots et al., 1978; White and Mayrand, 1981; Zambon et al., 1981) and is found in increased numbers in plaque from adult subjects with periodontitis (Speigel et al., 1979; Tanner et al., 1979; White and Mayrand, 1981). Similarly, Bacteroides internedius is isolated in increasing numbers in subgingival plaque of individuals with various forms of periodontal disease (Loesche et al., 1985; Moore et al., 1987; Slots, 1982). Although these two species have many common biochemical characteristics and both can frequently be isolated from plaque obtained from specific periodontal pockets (Socransky et al., 1988), their distribution in the oral cavity appears to differ, with B. intermedius being found in significant numbers in both supra- and subgingival plaques (Moore et al., 1983, 1985). In addition, Received for publication November 30, 1989 Accepted for publication February 2, 1990 This investigation was supported in part by USPHS Research Grant P50-DE08489 from the National Institute of Dental Research, National Institutes of Health, Bethesda, MD 20892. 'Present address: Department of Oral Biochemistry, Tohoku University, School of Dentistry, Sendai, Japan 2To whom correspondence and reprint requests should be addressed 1266

results from ligature-induced periodontal disease experiments in monkeys (Kiel et al., 1983; Kornman et al., 1981) have demonstrated that although B. intermedius is present in higher concentrations than P. gingivalis during the early stages of disease, the latter bacterium becomes dominant as the disease progresses. It has been suggested (Hamilton et al., 1989) that simple environmental factors such as pH and nutrients may play a role in the subgingival shifts in these bacterial populations. Recent continuous culture studies with P. gingivalis (McDermid et al., 1988) and B. internedius (Hamilton et al., 1989) have investigated the effect of environment on the growth and metabolic properties of these bacteria. In this communication, we present the results of studies designed to compare, under identical conditions, the effect of pH on growth and production and activity of proteolytic enzymes by type strains and fresh plaque isolates of P. gingivalis and B. intermedius.

Materials and methods. Bacteria and growth conditions. -P. gingivalis ATCC 33277 and B. intermedius ATCC 25261 and ATCC 25611 were purchased from the American Type Culture Collection (Rockville, MD). Isolates from human dental plaque included two strains of P. gingivalis and six strains of B. intermedius. These bacteria were isolated, identified, and characterized by Dr. L. Wolff (University of Minnesota, Minneapolis). Actinomyces viscosus WVU 627 and Streptococcus mutans Ingbritt were kindly supplied by Dr. W. Liljemark (University of Minnesota, Minneapolis). Initially, all the bacteria were grown in BM medium (Shah et al., 1976) supplemented with 5.0 mg/L of hemin, 0.5 mg/L of menadione, and 0.2% glucose at 370C in an anaerobic chamber containing 80% N2, 10% H2, and 10% CO2. After growth, the cultures were transferred into a semichemically-defined medium (inoculum size

Effect of pH on the growth and proteolytic activity of Porphyromonas gingivalis and Bacteroides intermedius.

The effect of pH on the growth and proteolytic activity of the type strain and fresh isolates of Porphyromonas gingivalis and Bacteroides intermedius ...
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