Franca Crociani

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.

Deoxyribonucleic Acid Homology Relationships Among Species of the Genus Bifidobacterium

Genetic relatedness among 179 strains representing 13 named species and several unnamed taxa of the genus Bifidobacterium from three main habitats, i.e., (i) feces of man, (ii) feces of various other animals, and (iii) the bovine rumen, was assessed by means of deoxyribonucleic acid (DNA)—DNA hybridization by using a filter-paper technique in competition experiments. Assignment of these strains to the genus Bifidobacterium was based on the production of lactic and acetic acids as chief products from glucose, lack of gas production, fructose-6-phosphate phosphoketolase activity, and on morphology. About 180 DNA competitors were tested with 23 reference systems. Several genetically distinct groups were recognized. Little or no DNA homology was demonstrated between some of the groups, suggesting large evolutionary divergence in this genus. B. infantis, B. liberorum, and B. lactentis form one of these groups; the ecological significance of this relatedness was discussed. The DNA of the following pairs are homologous: (i) B. breve and B. parvulorum, (ii) B. thermophilum and B. ruminale, and (iii) B. pseudolongum and B. globosum. Within a number of strains assigned to B. adolescentis, many of which were isolated from waste waters, a large genetic heterogeneity was demonstrated: in addition to B. adolescentis, at least three unrelated groups were recognized and are provisionally referred to as “dentium,” “catenulatum,” and “angulatum.” Since these groups are not related genetically to any species of the genus and are phenotypically distinct, they may represent new species. The validity of the species B. bifidum, B. longum, and B. suis was confirmed at the genetic level.

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).

Bifidobacterium pullorum sp. nov.: A new species isolated from chicken feces and a related group of bifidobacteria isolated from rabbit feces

Among bifidobacteria isolated from feces of chickens 7 were distinctive in displaying (i) curved cells with tapered ends, mostly arranged in long chains, (ii) preferred alcaline reaction for good growth and need for Tween 80 as stimulating factor, (iii) guanosine plus cytosine content of deoxyribonucleic acid (% GC) exceptionally high (66–68 moles per cent) and (iiii) genetic unrelatdedness (DNA-DNA homology) to any other species or types of the genus Bifidobacterium except for 4 strains isolated from the feces of rabbit (59–66% similarity). A new species Bifidobacterium pullorum sp. nov. (type strain P145, ATCC27685) is proposed for the strains from chicken; the strains from rabbit were described as Unassigned Homology Group I (representative strain RA 161).

Bifid bacteria in bovine rumen

SummaryAbout 500 bifid isolates from 150 samples of bovine rumen liquor were examined for their morphology, physiology and biochemistry. Diagnosis as “bifid bacteria” was based upon the peculiar pathway of glucose anaerobic metabolism i.e. the fructose-6-phosphate shunt. Four phenetic types were recognized. These types can be differentiated from those found in human habitats because their cell-free extracts are aldolase and HMP dehydrogenases positive: they are potential heterofermenters; furthermore the rumen types are nutritionally different. The distinction of the rumen bifids from the Bifidobacterium species of the intestine of Apis mellifica and Apis indica is still more consistent for a lot of characters. The characters of two rumen types warranted the creation of two new species of the genus Bifidobacterium. One of these, B. globosum n. sp., has a proper morphology, is serologically distinct and has a deoxyribonucleic acid base composition, in % GC, of 64.5. The other, B. ruminale n. sp., found so far only in rumen, is characteristically lactose non fermenter, at variance with all the bifids from human habitats and has peculiar morphological traits. A third type is probably a rough variant of B. ruminale and a fourth is serologically distinct and mannitol fermenter; their taxonomic definition is still, however, premature.

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 inBifidobacterium animalis

Strains isolated from rabbit, chicken, and rat feces and from sewage and fermented milk products, all identified asBifidobacterium 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.

Isoleucine production in bifidobacteria

SummaryAbout 300 strains of bifidobacteria were examined for their capacity to release L-isoleucine in the fermentation broth. A strain of the speciesBifidobacterium ruminale was selected as the best producer. After treatment of this strain with NTG a DL-α-aminobutyric acid-resistant mutant capable of producing about 5 mg/ml of L-isoleucine in presence of 1.5 % DL-α-aminobutyric acid was obtained. Cultural conditions and acetohydroxyacid synthetase activity were studied.

Increase in isoleucine accumulation by α-aminobutyric acid-resistant mutants ofBifidobacterium Ruminale

SummaryA number of DL-α-aminobutyric acid-resistant mutants ofBifidobacterium ruminale were isolated. Several of these mutants were found to be superior to the parent strain in converting α-aminobutyric acid to L-isoleucine and in the valine accumulation. One of them accumulated over 9 mg/ml of L-isoleucine in presence of 3% DL-α-aminobutyric acid and 3 mg/ml of L-valine in absence of the precursor.

Urease production and DNA-homology in the species Bifidobacterium suis

SummaryUrease production in the species Bifidobacterium suis was studied. The strains examined were strictly homogeneous in their DNA homology relationships. Most strains (74%) possess this enzyme. The presence of urease is therefore of value as additional diagnostic character of this species.