Benjamin F. Hammond

Leukotoxic activity in different strains of the bacterium Actinobacillus actinomycetemcomitans isolated from juvenile periodontitis in man

Abstract Type culture strains and dental isolates (from juvenile periodontitis) of Act. actinomycetemcomitans (Aa) were tested for their ability to kill human polymorphonuclear leukocytes (PMN) in vitro. The majority of Aa, as well as sonic extracts prepared from these organisms, rapidly destroyed PMNs, shown by the extracellular release of lactate dehydrogenase from PMNs and by degenerative ultrastructural alterations. The leukotoxic properties of Aa could be modulated by serum: normal human sera enhanced killing but juvenile periodontitis or various rabbit anti-Aa sera neutralized toxic activity. Whereas toxic and non-toxic strains of Aa shared common antigens, immunologic analyses revealed a unique antigen in sonic extracts of leukotoxic organisms. Thus Aa-derived leukotoxin may be an aetiologic vector in juvenile periodontitis.

Bacteriology of periodontal disease in the cat.

Subgingival plaque samples were obtained from 32 cats showing different stages of periodontal disease. Correlations were sought between gingival index scores and the prevalence of various microbial groups, and between microbial populations found in sites designated as most-affected and least-affected within individual cats. The tendency with higher gingival index scores, and with the most-affected sites, was toward a microbial population composed to a greater extent of anaerobic Gram-negative rods. The most common organism was an anaerobic Gram-negative rod in the black-pigmented Bacteroides group that was biochemically similar to B. gingivalis but had catalase activity. The black-pigmented Bacteroides group and Peptostreptococcus anaerobius were found in increasing numbers with increasingly severe periodontal disease. Pasteurella multocida was isolated from most samples and appeared to decrease in numbers with increasing periodontal disease.

Cytotoxicity of the bacterium Actinobacillus actinomycetemcomitans extracts in human gingival fibroblasts

Filter-sterilized sonic extracts (SE) of strains of Actinobacillus actinomycetemcomitans were shown to inhibit the proliferation of human gingival fibroblasts in vitro. The inhibition was dose-dependent: a 50 per cent inhibitory dose of 2 micrograms protein/ml was found for A. actinomycetemcomitans strain Y4. The inhibitory activity could be neutralized by homologous antiserum and was heat inactivated by temperatures of 80 degrees C or greater. The fibroblast-inhibitory activity was present in SEs of both leukotoxic-producing and non-leukotoxic strains of A. actinomycetemcomitans, suggesting that a separate agent is responsible for leukotoxicity and fibroblast inhibition. A short (10 min) exposure of the fibroblasts to the A. actinomycetemcomitans SE was sufficient to inhibit irreversibly cell proliferation, provided that serum was present at the time that the cells were exposed to the SE. SE-challenged fibroblasts exhibited a marked decrease in the rate of DNA synthesis, but no inhibition of RNA or protein synthesis. Although the SE-treated cells did not proliferate, they appeared to remain intact and viable; and displayed no gross morphological alterations.

Antigens and surface components associated with virulence of Actinomyces viscosus.

We have isolated a specific cell wall antigen of high molecular weight which appears to be unique to virulent strains of A viscosus and A naeslundii. The antigen is composed of two parts: a polysaccharide moiety containing 6-DOT as the major sugar and determinant of serologic specificity, and a small peptide bearing some resemblance to the peptidoglycan. Other data indicate a positive correlation between the presence of this antigen and an extrachromosomal piece of DNA having most of the properties of a bacterial plasmid. The specific function of the 6-DOT antigen in disease production is not known, but its clear association with virulent strains suggests the possibility of monitoring specific populations of oral actinomycetes.

TOXICITY OF LACTOBACILLUS CASEI.

With the possible exception of Lactobacillus acidophilus, most of the oral lactobacilli are generally considered harmless saprophytes.1 In view of the virulence-enhancing effect of some bacterial capsules, the finding of a heavily encapsulated strain of Lactobacillus casei2 suggested the possibility of another pathogenic lactobacillus. Preliminary serologic studies showed that when whole encapsulated cells of this organism were injected into the marginal ear veins of rabbits, large, indurated, and erythematous lesions developed at the site of injection and became progressively worse. This report describes the toxic property of the organism and its role in the production of both local and systemic lesions using whole cells and several subcellular fractions.

Presence of Polysaccharide-forming Coliforms in Human Saliva

It is becoming increasingly clear from several reports that the biosynthesis and utilization of microbial polysaccharides may play an important role in the survival and maintenance of a number of microbes commonly found in the oral cavity, including Lactobacillus casei (B. F. HAMMOND and N. B. WILLIAMS, Arch. oral Biol., 9:341-49, 1964) and Streptococcus initis (R. J. GIBBONS, J. Bact., 87:1512-20, 1964). A erobacter aerogenes has been extensively studied in this regard by Postgate and Hunter (J. Gen. Microbiol., 29:233-63, 1962) and by Strange, Dark, and Ness (J. Gen. Microbiol., 25:61-76, 1961) who observed the degradation of intracellular polysaccharide in starving cells. Other studies have shown that Aerobacter spp. remained viable in submaxillary and parotid saliva for considerably longer periods of time than did other members of the oral flora (B. F. HAMMOND, J. dent. Res., 38: 701-2, 1959 [abstract]; and N. B. WILLIAMS and D. 0. POWLEN, Arch. oral Biol., 1: 48-61, 1959). Richardson and Jones (J. dent. Res., 37:697-709, 1958) indicated that Aerobacter spp. occur in the oral cavity infrequently and in low numbers. This report presents evidence that A. aerogenes and coliform bacilli occur in saliva more frequently than previously reported and that their occurrence may be associated with the production of cellular polysaccharides. Paraffin-stimulated human saliva specimens were collected from 87 student nurses and 116 dental students; serial tenfold dilutions were plated in duplicate onto a modified Chapmans medium (N. B. WILLIAMS, B. F. HAMMOND, and H. B. HERSCOWITZ, J. dent. Res., 43:890, 1964 [abstract]) and were incubated at 370 C. under aerobic conditions for 48 hours. Mucoid coliform strains were isolated from 52

Phage Susceptibility of Encapsulated Lactobacillus casei

ONE of the principal factors which determines the susceptibility of a given bacterium to attack by bacteriophage is the availability of specific surface receptor sites. Numerous investigators have observed that encapsulated bacteria are generally phage resistant, and it has been assumed that the capsular layer, interposed as a physical barrier between receptor and phage, acts as a non-specific inhibitor of phage adsorption. In partial confirmation of this role, Maxted1 showed that although phage resistant group A streptococci possessed a hyaluronic acid capsule, treatment with hyaluronidase destroyed the capsule and rendered the cells phage-sensitive. It is not clear, however, from this and other reports2 whether the susceptibility of non-encapsulated cells was due to the availability of receptor sites permitting increased adsorption or to some other factor permitting increased virus penetration and lysis of the bacterial cell. This report presents evidence that in the related homolactic organism, L. casei, the specific surface receptor sites are present in varying amounts on both encapsulated and non-encapsulated cells but that the loss of the capsular material, either by mutation or mechanical removal, renders the previously resistant cell sensitive to lysis by phage.