The family Bifidobacteriaceae constitutes an important phylogenetic group that particularly includes bifidobacterial taxa demonstrating proven or debated positive effects on host health. The increasingly widespread application of probiotic cultures in the twenty-first century requires detailed classification to the level of particular strains. This study aimed to apply the glutamine synthetase class I (glnAI) gene region (717 bp representing approximately 50% of the entire gene sequence) using specific PCR primers for the classification, typing, and phylogenetic analysis of bifidobacteria and closely related scardovial genera. In the family Bifidobacteriaceae, this is the first report on the use of this gene for such purposes. To achieve high-value results, almost all valid Bifidobacteriaceae type strains (75) and 15 strains isolated from various environments were evaluated. The threshold value of the glnAI gene identity among Bifidobacterium species (86.9%) was comparable to that of other phylogenetic/identification markers proposed for bifidobacteria and was much lower compared to the 16S rRNA gene. Further statistical and phylogenetic analyses suggest that the glnAI gene can be applied as a novel genetic marker in the classification, genotyping, and phylogenetic analysis of isolates belonging to the family Bifidobacteriaceae.

• MeSH
• Genes, Bacterial MeSH
• Bifidobacterium classification   enzymology   MeSH
• DNA, Bacterial genetics   MeSH
• DNA Primers MeSH
• Phylogeny * MeSH
• Genetic Markers MeSH
• Genotype MeSH
• Glutamate-Ammonia Ligase genetics   MeSH
• RNA, Ribosomal, 16S genetics   MeSH
• Sequence Analysis, DNA MeSH
• Bacterial Typing Techniques MeSH
• Publication type
• Journal Article MeSH
• Names of Substances
• DNA, Bacterial MeSH
• DNA Primers MeSH
• Genetic Markers MeSH
• glutamine synthetase I MeSH Browser
• Glutamate-Ammonia Ligase MeSH
• RNA, Ribosomal, 16S MeSH

An alternative molecular marker with respect to the 16S rRNA gene demonstrating better identification and phylogenetic parameters has not been designed for the whole Bifidobacteriaceae family, which includes the genus Bifidobacterium and scardovial genera. Therefore, the aim of the study was to find such a gene in available genomic sequences, suggest appropriate means and conditions for asmplification and sequencing of the desired region of the selected gene in various strains of the bacterial family and verify the importance in classification and phylogeny. Specific primers flanking the variable region (~800 pb) within the pyrG gene encoding the CTP synthetase were designed by means of gene sequences retrieved from the genomes of strains belonging to the family Bifidobacteriaceae. The functionality and specificity of the primers were subsequently tested on the wild (7) and type strains of bifidobacteria (36) and scardovia (7). Comparative and phylogenetic studies based on obtained sequences revealed actual significance in classification and phylogeny of the Bifidobacteriaceae family. Gene statistics (percentages of mean sequence similarities and identical sites, mean number of nucleotide differences, P- and K-distances) and phylogenetic analyses (congruence between tree topologies, percentages of bootstrap values >50 and 70%) indicate that the pyrG gene represents an alternative identification and phylogenetic marker exhibiting higher discriminatory power among strains, (sub)species, and genera than the 16S rRNA gene. Sequences of the particular gene fragment, simply achieved through specific primers, enable more precisely to classify and evaluate phylogeny of the family Bifidobacteriaceae including, with some exceptions, health-promoting probiotic bacteria.

• Keywords
• Bifidobacteriaceae, Bifidobacterium, CTP synthetase, classification, phylogenetics, scardovia,
• MeSH
• Actinobacteria classification   enzymology   genetics   isolation & purification   MeSH
• Bacterial Proteins chemistry   genetics   metabolism   MeSH
• DNA, Bacterial genetics   MeSH
• DNA Primers genetics   MeSH
• Phylogeny * MeSH
• Carbon-Nitrogen Ligases chemistry   genetics   metabolism   MeSH
• RNA, Ribosomal, 16S genetics   MeSH
• Bacterial Typing Techniques methods   MeSH
• Publication type
• Journal Article MeSH
• Evaluation Study MeSH
• Research Support, Non-U.S. Gov't MeSH
• Names of Substances
• Bacterial Proteins MeSH
• CTP synthetase MeSH Browser
• DNA, Bacterial MeSH
• DNA Primers MeSH
• Carbon-Nitrogen Ligases MeSH
• RNA, Ribosomal, 16S MeSH

Gram-stain-positive, catalase and oxidase-negative and short rod-shaped bacterium C10 with occasional branching was isolated under strictly anaerobic conditions from the rumen fluid of a red deer (Cervus elaphus) in the course of study attempting to uncover new xylanolytic and cellulolytic rumen bacteria inhabiting the digestive tract of wild ruminants in the Czech Republic. The anaerobic M10 medium containing bovine rumen fluid and carboxymethylcellulose as a defined source of organic carbon was used in the process of bacterial isolation. The 16S rRNA gene similarity revealed recently characterized new species Actinomyces succiniciruminis Am4T (GenBank accession number of the gene retrieved from the complete genome: LK995506) and Actinomyces glycerinitolerans G10T (GenBank accession number from the complete genome: NZFQTT01000017) as the closest relatives (99.7 and 99.6% gene pairwise identity, respectively), followed by the Actinomyces ruminicola DSM 27982T (97.2%, in all compared fragment of 41468 pb). Due to the taxonomic affinity of the examined strain to both species A. succiniciruminis and A. glycerinitolerans, its taxonomic status towards these species was evaluated using variable regions of rpsA (length of 519 bp) and rplB (597 bp) gene sequences amplified based on specific primers designed so as to be applicable in differentiation, classification, and phylogeny of Actinomyces species/strains. Comparative analyses using rpsA and rplB showed 98.5 and 97.9% similarities of C10 to A. succiniciruminis, respectively, and 97.5 and 97.6% similarities to A. glycerinitolerans, respectively. Thus, gene identities revealed that the evaluated isolate C10 (=DSM 100236 = LMG 28777) is a little more related to the species A. succiniciruminis isolated from the rumen of a Holstein-Friesian cow than A. glycerinitolerans. Phylogenetic analyses confirmed affinity of strain C10 to both recently characterized species. Unfortunately, they did not allow the bacterial strain to be classified into a particular species. Phenotypic characterization suggested similar conclusions. This brief contribution is aimed at classification and detailed phenotypic characterization of bacterial strain C10 isolated from the rumen of a wild red deer exhibiting, from the point of view of Actinomyces species, noteworthy cellulolytic and xylanolytic activities.

• MeSH
• Actinomyces classification   genetics   isolation & purification   metabolism   MeSH
• Rumen microbiology   MeSH
• Genes, Bacterial genetics   MeSH
• Cellulose metabolism   MeSH
• DNA, Bacterial genetics   MeSH
• Phenotype MeSH
• Phylogeny MeSH
• Fatty Acids analysis   MeSH
• Peptidoglycan analysis   MeSH
• RNA, Ribosomal, 16S genetics   MeSH
• Deer microbiology   MeSH
• Xylans metabolism   MeSH
• Base Composition MeSH
• Animals MeSH
• Check Tag
• Animals MeSH
• Publication type
• Journal Article MeSH
• Names of Substances
• Cellulose MeSH
• DNA, Bacterial MeSH
• Fatty Acids MeSH
• Peptidoglycan MeSH
• RNA, Ribosomal, 16S MeSH
• Xylans MeSH

Occurrence of bifidobacteria, known as health-promoting probiotic microorganisms, in the digestive tract of wild pigs (Sus scrofa) has not been examined yet. One hundred forty-nine fructose-6-phosphate phosphoketolase positive bacterial strains were isolated from colonic content of twenty-two individuals of wild pigs originated from four localities in the Czechia. Based on PCR-DGGE technique targeting the variable V3 region of the 16S rRNA genes, strains were initially differentiated into four groups represented by: (i) probably a new Bifidobacterium species (89 strains), (ii) B. boum/B. thermophilum/B. thermacidophilum subsp. porcinum/B. thermacidophilum subsp. thermacidophilum (sub)species (49 strains), (iii) Pseudoscardovia suis (7 strains), and (iv) B. pseudolongum subsp. globosum/B. pseudolongum subsp. pseudolongum (4 strains), respectively. Given the fact that DGGE technique did not allow to differentiate the representatives of thermophilic bifidobacteria and B. pseudolongum subspecies, strains were further classified by the 16S rRNA and thrS gene sequences. Primers targeting the variable regions of the latter gene were designed to be applicable in identification and phylogeny of Bifidobacteriaceae family. The 16S rRNA-derived phylogenetic study classified members of the first group into five subgroups in a separated cluster of thermophilic bifidobacteria. Comparable results were obtained by the thrS-derived phylogenetic analysis. Remarkably, variability among thrS sequences was higher compared with 16S rRNA gene sequences. Overall, molecular genetic techniques application allowed to identify a new Bifidobacterium phylotype which is predominant in the digestive tract of examined wild pigs.

• MeSH
• Genes, Bacterial MeSH
• Bifidobacterium chemistry   classification   genetics   isolation & purification   MeSH
• Animals, Wild * MeSH
• Phylogeny MeSH
• Gastrointestinal Tract microbiology   MeSH
• Molecular Typing * methods   MeSH
• Swine MeSH
• RNA, Ribosomal, 16S genetics   MeSH
• Sequence Analysis, DNA MeSH
• Sus scrofa microbiology   MeSH
• Animals MeSH
• Check Tag
• Animals MeSH
• Publication type
• Journal Article MeSH
• Names of Substances
• RNA, Ribosomal, 16S MeSH