Effect of growth temperature on the biosynthesis of cell wall proteins from Bifidobacterium globosum.

Seventy strains of Bifidobacterium globosum isolated from gastrointestinal tracts of different animals were studied. Strains were grown at temperatures ranging from 25 to 46.5 degrees C in order to examine changes both in the expression of bifidobacterial outer proteins (BIFOPs) and in their hydrophobic properties. It was observed that the expression of BIFOPs found on the cell-surface changes according to growth temperature, with quantitative and/or qualitative variations. Generally speaking, it was observed that BIFOP expression at low-growth temperature was considerably attenuated, while at medium- and high-growth temperature it increased. Furthermore, at high-growth temperatures, the presence of a new common protein was detected in all the strains studied. Cells from B. globosum strains grown under different temperature conditions were studied in terms of their cellular hydrophobicity properties. At medium-growth temperature, the cell hydrophobicity was strictly correlated with BIFOP expression, while at low and high-growth temperatures, the presence of BIFOP only partially influenced the hydrophobic features.

Bifidobacterium inopinatum sp. nov. and Bifidobacterium denticolens sp. nov., two new species isolated from human dental caries.

In a previous investigation of bifidobacteria isolated from human dental caries (V. Scardovi and F. Crociani, Int. J. Syst. Bacteriol. 24:6-20, 1974), 40 strains were assigned to the new species Bifidobacterium dentium. In this study we examined 70 new strains of bifidobacteria isolated from dental caries. The morphological characteristics, biochemical reactions, fermentation patterns, end products from glucose metabolism, protein electrophoretic patterns, levels of DNA hybridization, and DNA G+C contents of these organisms revealed that they belong to three different taxa. One of these taxa was identified as B. dentium. The other two are described as the following new Bifidobacterium species in this paper: Bifidobacterium inopinatum (type strain, DSM 10107) and Bifidobacterium denticolens (type strain, DSM 10105). The two new species differ from other Bifidobacterium species in their morphological characteristics (especially B. inopinatum, with its very small coccoid cells), in their carbohydrate fermentation patterns (most strains ferment dextran, and B. inopinatum does not ferment galactose), and in their DNA base compositions (especially B. inopinatum).

Degradation of complex carbohydrates by Bifidobacterium spp.

Two hundred and ninety strains of 29 species of bifidobacteria from human and animal origin were surveyed for their ability to ferment complex carbohydrates. The substrates fermented by the largest number of species were D-galactosamine, D-glucosamine, amylose and amylopectin. Many of the species isolated from animal habitats showed reduced fermentation activity. Bifidobacterium dentium strains fermented gum guar and gum locust bean; porcine gastric mucin was fermented only by B. bifidum, B. infantis was the only species to ferment D-glucuronic acid; strains of B. longum fermented arabinogalactan and the gums arabic, ghatti and tragacanth; alpha-L-fucose was fermented by strains of B. breve, B. infantis and B. pseudocatenulatum. A key to the differentiation of Bifidobacterium species of human origin is provided.

Characterization of the plasmid pVS809 from Bifidobacterium globosum.

A plasmid from a B. globosum strain was cut with 38 restriction enzymes and a physical map was constructed. Out of a total of 121 clones from curing experiments, plasmid was lost in 58% and 100% for acridine orange and ethidium bromide curing agent respectively. The plasmid does not exist as a chromosomal integrated form. An attempt to determine phenotypic characters encoded by the plasmid was made by electrophoretic analyses of the total proteins.

Regulation of dihydrodipicolinate synthase and diaminopimelate decarboxylase activity in Bacillus stearothermophilus.

The feedback inhibition of the enzymes dihydrodipicolinate (DHDPS) and diaminopimelate decarboxylase (DAPD) in the wild strain Zu 183 of Bacillus stearothermophilus and in its S-(2-aminoethyl)-cysteine resistant L-lysine overproducing strain AEC 12 was studied. The optimum temperature and pH of both enzymes were also evaluated. No inhibition of DHDPS by L-lysine, L-threonine, L-methionine and L-isoleucine was observed either in the wild strain or in the AEC 12 mutant. DAPD was completely inhibited by L-lysine and only partially by L-threonine and L-methionine in Zu 183 and AEC 12 strains, but the concentration required was found to be much higher in the AEC 12 strain. The regulation mechanism of L-lysine biosynthesis in Bacillus stearothermophilus Zu 183 was also discussed.

Bifidobacterial cell wall proteins (BIFOP) in Bifidobacterium globosum.

Nearly 150 strains of Bifidobacterium globosum were isolated from faeces of calf, chicken, lamb, rabbit and rat, from sewage, from rumen content and from human infant faeces between 1962 and 1973 and scored by SDS-PAGE for the presence of cell-wall-related proteins, i.e. BIFOP (bifid outer proteins); their apparent molecular masses ranged from 94.5 to 34 kDa and were designated A to L. Purified preparations from six of these ten proteins were employed to produce polyclonal rabbit antisera for use in immunoblots to investigate the interrelationships of the major antigens, A, B and C (94.5-85.5 kDa) and their distribution in strains of various origin. Two antigens differently migrating (or polymorphic forms) reacted with anti-BIFOP F serum (called F- and F+); the identity of BIFOP E with respect to these antigens was studied with anti-E serum. Only one antigen in all strain preparations reacted to anti-BIFOP H serum, which was raised against an antigen purified from a 13.5-MDa plasmid-bearing strain from rumen.

Phase variations in Bifidobacterium animalis.

Strains isolated from rabbit, chicken, and rat feces and from sewage and fermented milk products, all identified as Bifidobacterium animalis, were found to show phase variations in colony appearance and in cellular morphology. The rate of transition in a switching system from opaque to transparent colonies and vice versa was determined. Differences in protein components and in penicillin-binding proteins (PBPs) of the cells from different colony types are shown.

Identification of bifidobacteria from fermented milk products.

Six samples of fermented milk preparations were examined for the presence of bifidobacteria. Identification was based on fermentation tests, genetic relatedness studies and electrophoretic analysis. Contrary to label information, Bifidobacterium animalis was the only species present.

Different electrophoretic patterns of cellular soluble proteins in Bifidobacterium animalis.

Twenty-nine strains of Bifidobacterium animalis, a species found only in animal habitats, were studied. Strains from known origins such as rat, rabbit and chicken feces and strains isolated from extrabody environments such as sewage and fermented milk products were examined. The intestinal origins of strains isolated from sewage and fermented milk products were determined by means of the comparison of electrophoretograms of cellular soluble proteins. Unknown origins of strains were recognized as being rabbit or chicken intestinal tract.

Urease activity in gastrointestinal tract of rabbit and electrophoretic behaviour of urease.

Urease activity in the stomach (fundus and antrum), caecal content and soft faeces of rabbit was studied. Significant differences between fundus and antral content (P less than 0.01) and between caecum and soft faeces (P less than 0.05) were observed. The urease zymograms from caecal content and soft faeces of rabbit presented two different bands. The fundus content and the caecal ureolytic Clostridium innocuum bacterium exhibited only one band. Band B of caecal content was not evident at pH 4, whereas band A, also present in the stomach, was observed both at pH 4 and at pH 6. The optimal pH of urease activity of stomach and caecal content was in the range of 4-5 and 5-6, respectively. A comparison of intestinal urease zymograms with those of the single ureolytic bacterial species was suggested in order to clarify their role in urea metabolism.

Protoplast formation and regeneration in Bifidobacterium.

Conditions for protoplast formation and cell wall regeneration in the genus Bifidobacterium are described. The ability to form protoplast in high percentage varied according to the species considered. High reversion levels were obtained with the species B. bifidum var. pennsylvanicus, B. thermophilum and B. boum.

Species of the Bifidobacterium in the feces of infants.

A total of 907 strains of bifidobacteria isolated from the feces of breast-fed and artificial milk-fed infants were studied. Their assignment to known species of the genus Bifidobacterium was primarily based on DNA homology relationships. The strains were classified as B. bifidum, B. breve, B. catenulatum, B. dentium, B. infantis, B. longum and B. pseudocatenulatum. The distribution of these species is reported.

Anaerobic ureolytic bacteria from caecal content and soft faeces of rabbit.

Forty strains of ureolytic bacteria were isolated from the caecal content and soft faeces of seven rabbits by the anaerobic roll tube method and were characterized. The isolates were identified with Clostridium coccoides, Cl. innocuum, Peptostreptococcus productus, P. micros, Peptococcus magnus, Fusobacterium russii and Fusobacterium sp. Urease activity of representative strains of the various species was also determined. The study indicated that strongly-ureolytic anaerobic bacteria are present in the caecum of the rabbit.

Antimicrobial susceptibility of Bifidobacterium.

The susceptibility pattern of 459 strains of bifidobacteria, representing 15 species, to 16 antimicrobial agents was determined by the broth dilution method. The majority of the strains derived from human faeces. Penicillin G, erythromycin, clindamycin, vancomycin and bacitracin were the most active compounds; they inhibited 90% of the strains at less than 1.6 micrograms/ml. All strains were susceptible to chloramphenicol (MIC90 = 2.0-5.8 micrograms/ml) and also to lincomycin (MIC50 = 0.64-1.5 micrograms/ml). Neomycin, streptomycin and tetracycline presented a great variability in their activity. Most strains were resistant to polymyxin B, nalidixic acid, kanamycin, gentamicin and metronidazole. The only variation in susceptibility which was observed among the different species concerned Bifidobacterium suis, which generally appeared to be more resistant than other species.

Urease activity in the genus Bifidobacterium.

The urease activity of 414 strains representing 21 species of the genus Bifidobacterium was surveyed. The strongest ureolytic strains belong mostly to the species B. suis and only a few to B. breve, B. magnum and "subtile" homology group. The study of some strongly ureolytic strains showed that urea and organic nitrogen concentration did not influence urease production. The high urease activity found also in the absence of urea suggested that this enzyme is not inducible. An ammonia concentration of 14 mM did not repress urease activity.

Amino acids produced by bifidobacteria and some Clostridia.

A lot of 121 strains of bifidobacteria and 9 strains of clostridia were examined for their ability to release free amino acids in the culture broth. The bifidobacteria studied belong to 18 species or "homology group" and the clostridia to 8 species. The growth in a synthetic medium with ammonium salts as sole nitrogen source was also studied. All the clostridia and the majority of the bifidobacteria produce various amino acids. The possible ecological significance of these findings is suggested.