Roberto Reniero

Specific identification and targeted characterization of Bifidobacterium lactis from different environmental isolates by a combined multiplex-PCR approach.

ABSTRACT The species Bifidobacterium lactis, with its main representative strain Bb12 (DSM 10140), is a yoghurt isolate used as a probiotic strain and is commercially applied in different types of yoghurts and infant formulas. In order to ensure the genetic identity and safety of this bacterial isolate, species- and strain-specific molecular tools for genetic fingerprinting must be available to identify isolated bifidobacteria or lactic acid bacteria from, e.g., various clinical environments of relevance in medical microbiology. Two opposing rRNA gene-targeted primers have been developed for specific detection of this microorganism by PCR. The specificity of this approach was evaluated and verified with DNA samples isolated from single and mixed cultures of bifidobacteria and lactobacilli (48 isolates, including the type strains of 29Bifidobacterium and 9 Lactobacillusspecies). Furthermore, we performed a Multiplex-PCR using oligonucleotide primers targeting a specific region of the 16S rRNA gene for the genus Bifidobacterium and a conserved eubacterial 16S rDNA sequence. The specificity and sensitivity of this detection with a pure culture of B. lactis were, respectively, 100 bacteria/ml after 25 cycles of PCR and 1 to 10 bacteria/ml after a 50-cycle nested-PCR approach.

High frequency of conjugation in Lactobacillus mediated by an aggregation-promoting factor

Summary: Lactobacillus plantarum strain 4B2, which exhibits a strong autoaggregating phenotype, receives the broad-host-range plasmid pAMβ1 with conjugation efficiencies as high as 10-2 transconjugants per donor using solid matings; broth matings also occur, but at low transfer frequencies. Filter-sterilized spent supernatant of this strain contains a 32 kDa protein that promotes aggregation, and consequently a high frequency of conjugation, in lactic acid bacteria containing α-1,2-glucose-substituted lipoteichoic or teichoic acids. It appears, therefore, that the substituted lipoteichoic or teichoic acids act as receptors for the aggregation-promoting protein.

Iron requirement of Lactobacillus spp. in completely chemically defined growth media.

A completely chemically‐defined growth medium, containing guanine, thymine, cytidine, 2′‐deoxyadenosine and 2′‐deoxyuridine as DNA precursors, was developed for Lactobacillus johnsonii, on the basis of statistically designed techniques suitable for other lactobacilli. Particular focus was given to the nucleotide composition of different defined media, and to the specific nucleotide requirements of Lact. johnsonii. Most of the lactobacilli tested grew in a medium containing five free bases, four ribonucleosides or five deoxyribonucleosides. Adenine and guanine were replaceable by inosine. The requirement for thymine and cytosine was satisfied with uracil. The presence of inosine and uracil was identified as being essential for the growth of different Lactobacillus species, displaying their inability to synthesize purines and pyrimidines de novo. Defined recipes with different nucleotide composition were used to investigate iron requirements of lactobacilli. Only marginal differences in growth were observed in iron‐depleted media supplemented with five free bases, four ribonucleosides or five deoxyribonucleosides; iron depletion had a greater effect on growth when inosine and uracil were supplied as the only nucleotide sources. The results suggest that iron plays a role in the pyrimidine and purine metabolism of lactobacilli. Lactobacillus spp., particularly Lact. johnsonii, require iron under particular environmental conditions with limited or specific nucleotide sources.

The development of functional foods: lessons from the gut

Functional foods have resulted from the gradual recognition that healthy diets result from eating nutritious foods and from the identification of the mechanisms by which foods modulate metabolism and health. After initial successes with foods that reduce blood cholesterol level, probiotic bacteria and prebiotic carbohydrates have now also demonstrated added health benefits. As ingredients become more complex, the need to stabilize such ingredients in foods become increasingly important to the success of functional foods. Modern biotechnologies such as genomics, genetic expression and biomarkers of health and performance will be applied to this increasingly visible portion of human diets.

Growth requirements of Lactobacillus johnsonii in skim and UHT milk

Abstract Chemically undefined substances, like yeast extract or peptones, are commonly used as milk supplements in industrial fermentations to allow the growth of bacterial strains unable to grow in pure milk. In order to look for substances to replace their positive effects on the bacterial growth, the requirements of the probiotic strain L. johnsonii NCC 533 (La1) in skim and whole fat UHT milk were investigated. A mixture of 20 amino acids was added to skim milk and the removal of single amino acids identified alanine, serine, isoleucine and cysteine as the most important amino acids for growing L. johnsonii in milk. The requirements of L. johnsonii NCC 533 (La1) for nucleotide precursors were also investigated. Supplementation of milk with the four identified amino acids, four ribonucleosides and one source of iron was able to support the growth of the tested strain in milk with cell counts and final pH as obtained by the addition of yeast extract. The impact of milk supplementation was also investigated for additional 10 lactobacilli and a positive effect on the growth could be demonstrated for other L. johnsonii strains, a probiotic L. acidophilus strain and the L. gallinarum type strain.