A new depsipeptide, malevamide E (1), was isolated from field-collected colonies of the filamentous cyanobacterium Symploca laete-viridis. The gross structure of 1 was determined by spectroscopic analyses, including one- and two-dimensional NMR and accurately measured MS/MS. Chiral HPLC analyses of an acid hydrolysate of 1 allowed the stereochemical assignments of its amino acid residues, which include N-methyl-L-alanine, alpha-N,gamma-N-dimethyl-L-asparagine, N-methyl-L-phenylalanine, L-proline, D-valine, and N-methyl-L-valine. LC-MS/MS analysis of S. laete-viridis fractions established the co-occurrence of malevamide E (1) and its homologue dolastatin 14 (2), which was previously reported in low yield from the sea hare Dolabella auricularia. Malevamide E (1) demonstrated a dose-dependent (2-45 microM) inhibition of store-operated Ca(2+) entry in thapsigargin-treated human embryonic kidney (HEK) cells, indicating an inhibitory effect on Ca(2+) release-activated Ca(2+) (CRAC) channels.

A lactate dehydrogenase-deficient (Ldh-) mutant of a human isolate of Streptococcus mutans serotype c was tested in a gnotobiotic rat caries model. Compared with the wild-type Ldh-positive (Ldh+) strains, it was significantly (a s 0.005) less cariogenic in experiments with two different sublines of Sprague-Dawley rats. The Ldh-mutant strain 044 colonized the oral cavity of the test animals to the same extent as its parent strain 041, although its initial implantation was slightly but not significantly (P -0.2) less. Multiple oral or fecal samples plated on 2,3,5-triphenyltetrazolium indicator medium revealed no evidence of back mutation from Ldhto Ldh+ in vivo. Both Ldh+ strain 041 and Ldh-strain 044 demonstrated bacteriocinlike activity in vitro against a number of human strains of mutans streptococci representing serotype a (S. cricetus) and serotypes c and e (S. mutans). Serotypes b (S. rattus) and f (S. mutans) and strains of S. mitior, S. sanguis, and S. salivarius were not inhibited. Thus, Ldh mutant strain 044 possesses a number of desirable traits that suggest it should be investigated further as a possible effector strain for replacement therapy of dental caries. These traits include its stability and low cariogenicity in the sensitive gnotobiotic rat caries model, its bacteriocinlike activity against certain other cariogenic S. mutans (but not against more inocuous indigenous oral streptococci), and the fact that it is a member of the most prevalent human serotype of cariogenic streptococci.

The prevalence of mutans streptococci (Streptococcus mutans, Streptococcus cricetus, Streptococcus sobrinus, and Streptococcus rattus) was determined in the salivas of 169 elderly individuals ranging in age from 60 to 87 years. Approximately 40% of these individuals were edentulous and wore full upper and lower dentures. With the exception of a higher proportion of saliva counts below 1,000 CFU/ml in the full-denture wearers, the prevalence and the serotype and species distributions of the mutans streptococci were similar in the denture wearers and individuals with natural teeth only. The species and serotype distributions of mutans streptococci in this elderly population were also consistent with reported observations of other workers on younger, more caries-prone populations. A total of 87 representative isolates of the mutans streptococci were tested for cariogenic potential in a hamster model system. A considerable degree of variation in virulence between different strains was observed. However, these differences were not relatable to individual species or serotypes or to whether the organisms were isolated from denture wearers or naturally dentate subjects. The results of our studies indicate that elderly individuals with either natural or artificial dentitions may be a hitherto unrecognized reservoir of mutans streptococci having varying degrees of potential cariogenicity. Hence, in close family situations they could serve, along with parents and siblings, as vectors in the initial transmission of cariogenic microorganisms to young children.

The growth response of Streptococcus sanguis groups 1:A and 1:B in a complete chemically defined medium was not influenced by the oxygen concentration of the growth atmosphere. All of the cultures required cysteine and arginine; tyrosine and branched-chain amino acids were frequently required. Proteolysis of casein, mucin, and the anionic proteins of germfree rat saliva by S. sanguis was demonstrated. Hydrolytic activity toward casein was found in the soluble contents of the cells and in the cellular debris after disruption of the cells, with the soluble fractions exhibiting greater proteolytic activity toward casein. The soluble fractions from S. sanguis did not hydrolyze mucin, but this substrate was hydrolyzed by the cell debris fraction. When the amino acid requirements and proteolytic activity of S. sanguis and S. mutans were compared, these two oral streptococcal species exhibited distinct and characteristic differences.

A cross-sectional microbiological study of recurrent (secondary) dentinal caries was conducted. Freshly extracted human teeth containing amalgam restorations judged to be clinically intact were scrubbed with chlorhexidine soap and soaked in povidone-iodine solution. The teeth were then split to remove the fillings and examined for dentinal caries. Of 54 teeth examined, 22 (40%) were free of dentinal caries in the restored site. The remainder were assigned to arbitrary groups of questionable (15), initial (8) and active caries (9), based on the amount of dentinal decay observed. Comparison of pre-extraction bite-wing radiographs with the in situ findings revealed the sensitivity for secondary caries detection to be only about 50%. In contrast, the specificity for absence of caries was 73%. There was considerable variation in the numbers and types of micro-organisms found in dentine samples from lesions of comparable severity. Facultatively, anaerobic streptococci were the most numerous and most prevalent micro-organisms in the affected dentine, but no single species was significantly associated with recurrent caries. Mutans streptococci were found in 40% of sites with any degree of caries and in only 3 of the 9 sites with the most caries. Homofermentative lactobacilli were present in 18 of the 54 dentine samples, including 4 from caries-free sites. While the prevalence and numbers of lactobacilli increased with the degree of caries, they occurred in less than half of the affected dentine samples. Actinomyces occurred in 15 of 32 affected sites but only in 2 of the 9 most active sites. Their numbers never exceeded 3×10³ CFU/mg. The observed variations in the numbers and types of organisms present in the dentinal lesions, coupled with the relative scarcity of lactobacilli and mutans streptococci, raised the possibility that other, less acidogenic, microorganisms could be involved in the causation of recurrent dentinal caries. Support for this view came from observations that most of the types of bacteria recovered from the lesions were capable of lowering the pH below the threshold of pH 5.7 for demineralization of dentine. More focussed studies to determine the issue of the etiology of recurrent dentinal caries are needed.