Scanning electron microscopy and transmission electron microscopy have been used to visualize the morphology and ultrastructure of two types of microorganisms in the hindgut of the cockroach Blaberus posticus. Both organisms, designated as either short or long rods, are attached to chitinous projections from the gut wall. Micrographs suggest that the organisms are prokaryotic with a cell wall complex characteristic of gram-negative bacteria. However, certain differences were noted between the cell wall complex of the two types. Two forms of the long-rod type were noted, with one form appearing to be a "degenerate" or "transitional" cell. In the degenerate cells, vesicles are observed that often are contiguous with the cytoplasmic membrane. There are indications that the long-rod type may divide by longitudinal fission. Neither the short- nor long-rod type has been cultivated in its respective recognizable form.

Fusobacteria are commonly isolated from the hindgut of the cockroach Eublaberus posticus . Eleven strains isolated from E. posticus by us were keyed to four species, Fusobacterium necrophorum, F. varium , F. gonidiaformans , and F. prausnitzii , using current taxonomic criteria. With the exception of F. gonidiaformis , all species showed rods with swollen centers and large bodies. The pleomorphism of F. varium was examined by phase microscopy and scanning and transmission electron microscopy. The pleomorphic process begins with a gradual swelling at the center of the rod until a large round body is formed. Some of these round bodies then fragment, giving rise to rod-shaped cells. When 10% yeast extract was added to growth media, pleomorphism was not induced. A dialyzable factor was found to account for this observation. Fermentation of [1-14C]glutamic acid gives rise to butyrate labeled in the carboxyl carbon, indicating that butyrate is formed by the hydroxyglutarate pathway which may be characteristic for the genus Fusobacterium.

Cefaclor solutions in pH 2.5 and 4.5 buffers contained at least 90% of their initial activity after 72 h at 40C. Samples in pH 6.0, 7.0, and 8.0 buffers contained 70, 46, and 34%, respectively, of their initial activity after 72 h at 4C. After 72 h at 250C, samples prepared with pH 2.5, 4.5, 6.0, 7.0, and 8.0 buffers contained 95, 69, 16, 5, and 3%, respectively, of their initial activity. After 72 h at 370C, cefaclor solutions in pH 2.5 buffer contained 80% of the initial activity, whereas samples prepared in pH 4.5, 6.0, 7.0, and 8.0 buffers contained less than 20%. Laboratory-prepared plasma and serum samples showed an 8% loss in activity when incubated for 6 h at 40C, a 51% loss when incubated for 6 h at 250C, and a 48% loss when incubated for 2 h at 370C. Clinical samples demonstrated a similar stability pattern. Degradation rates for cefaclor in commercially prepared serum increased from 4-to 10-fold in comparison to rates obtained when samples were made in human serum freshly prepared in our laboratory. Consequently, serum standards should be made in freshly prepared human serum.Cefaclor, 3-chloro-7-D-(2-phenylglycinamido)-3-cephem-4-carboxylic acid, is an orally effective, broad-spectrum cephalosporin derivative that has been shown by Sullivan et al. (3) to be quantitatively absorbed as the intact antibiotic from the gastrointestinal tract of mice, rats, and dogs. Cefaclor is structurally related to other cephalosporins, in particular cephalexin, but possesses stability characteristics different from those of cephalexin. This study describes the stability characteristics of cefaclor and outlines procedures for the proper handling and evaluation of clinical samples. MATERIALS AND METHODSStability of cefaclor in buffer. The stability of cefaclor in the following 0.1 M potassium phosphate buffers was examined: pH 2.5, 4.5, 6.0, 7.5, and 8.0. Samples were prepared at 1 mg of cefaclor activity per ml in the appropriate buffer and incubated at 37, 25, and 40C. Samples were assayed at 0-, 24-, 48-, and 72-h intervals by removing an appropriate amount of the sample, diluting it 100-fold with 0.1 M pH 4.5 potassium phosphate buffer, and assaying by the AUTOTURB System (2) against a standard curve prepared in 0.1 M pH 4.5 potassium phosphate buffer at concentrations of 2.0, 5.0, 10.0, 20.0, and 40.0 itg of cefaclor activity per ml. Staphylococcus aureus ATCC 9144 was used as the test organism.In vitro stability of cefaclor in plasma and serum. Samples were prepared at 10 Zg of cefaclor activity per ml in either pooled human serum obtained commercially or serum and plasma obtained from the same healthy volunteers. The fluids were tested before use to ensure that they were devoid of antimicrobial activity. The pH was adjusted to 7.45 by the addition of 2 N HCl. Samples were incubated at 4, 25, and 3700, and the pH was maintained at 7.45 by overlaying the sample with an atmosphere of 95% 02-5% C02. A 1-ml sample was removed at appropriate intervals up to 6 h, diluted 100-fold with pH 4.5 buffer, stored at 40C, and a...

Washed cells from 72-h cultures of Streptomyces fradiae GS14 were used to examine the distribution of radiolabel from 14C-amino acids and related compounds into tylactone, C02, and cells. Test compounds were categorized according to products of their oxidative degradation. Those compounds known to produce propionyl-coenzyme A by direct catabolic oxidation were designated as group I. Group II included those compounds oxidized to methylmalonyl-coenzyme A via succinyl-coenzyme A and the tricarboxylic acid cycle. Group III contained compounds known to be oxidized to acetoacetyl-coenzyme A. The total amount of label recovered after 60 min ranged from 3 to 65%. Although label from all test compounds except proline (group II) and lysine (group III) was incorporated into tylactone after 60 min, label from group I and group III compounds was incorporated at levels five times greater than label from group II compounds. From 55 to 75% of the recovered label from propionate (I), asparagine (II), glutamine (II), glutamate (II), aketoglutarate (II), and succinate (II) was recovered as 14 C02. From 75 to 95% of the recovered label from the remaining compounds tested was located in the cells. Based on the data, a pathway for the role of amino acids in the biosynthesis of tylactone is proposed.Tylactone (Fig. 1) is the initial lactone precursor to tylosin, an antibiotic produced commercially by Streptomyces fradiae (1-3). Omura et al. (6), using 13C enrichment, showed that tylactone was formed from five propionate molecules, two acetate molecules, and one butyrate molecule. Subsequent inhibition studies with cerulenin, an inhibitor of fatty acid synthetase, showed that tylactone and related macrolide aglycone molecules were assembled in a manner similar to fatty acid biosynthesis (5, 7-9). The data of suggested that the precursors for tylactone biosynthesis were incorporated in the following manner: propionate as one propionyl-coenzyme A (CoA) (primer) and four methylmalonyl-CoA molecules, acetate as two malonyl-CoA molecules, and butyrate as one ethylmalonylCoA molecule.In this study, the role of amino acids as sources for the tylactone precursors was examined by using a tylA mutant of S. fradiae (1). This strain was blocked in the production of the tylosin sugars and therefore produced tylactone as its product (1, were grown in a complex medium as described previously (1).Preparation of washed cell suspensions. After 72 h of incubation, the cells from 300 ml of broth were removed by centrifugation at 16,000 x g for 5 min in a Sorvall RC-5 centrifuge. Cell pellets were washed twice with 300 ml of 0.1 M potassium phosphate buffer, pH 7.0. Approximately 6.5 g of wet cells were suspended in the buffer to a volume of 50 ml, and portions were diluted 10-fold with buffer to yield final cell suspensions containing 1.2 to 1.7 mg of dry cells per ml.Uptake and distribution reactions. Reactions were carried out in 35-ml flint glass bottles (Kimble, Toledo, Ohio) that were closed with gray butyl rubber stoppers (West Rubber Co., Phoenix...

The morphology of bacteriophage-like particles from the strict anaerobe Fusobacterium symbiosum is described. Attempts to demonstrate plaque formation on the host strain of F. symbiosum and other related species were unsuccessful.