Bacteriocins of lactic acid bacteria

Spectrophotometric Assay Using o-Phthaldialdehyde for Determination of Proteolysis in Milk and Isolated Milk Proteins

Numerous prokaryote genomes contain structures known as clustered regularly interspaced short palindromic repeats (CRISPRs), composed of 25-50 bp repeats separated by unique sequence spacers of similar length. CRISPR structures are found in the vicinity of four genes named cas1 to cas4. In silico analysis revealed another cluster of three genes associated with CRISPR structures in many bacterial species, named here as cas1B, cas5 and cas6, and also revealed a certain number of spacers that have homology with extant genes, most frequently derived from phages, but also derived from other extrachromosomal elements. Sequence analysis of CRISPR structures from 24 strains of Streptococcus thermophilus and Streptococcus vestibularis confirmed the homology of spacers with extrachromosomal elements. Phage sensitivity of S. thermophilus strains appears to be correlated with the number of spacers in the CRISPR locus the strain carries. The authors suggest that the spacer elements are the traces of past invasions by extrachromosomal elements, and hypothesize that they provide the cell immunity against phage infection, and more generally foreign DNA expression, by coding an anti-sense RNA. The presence of gene fragments in CRISPR structures and the nuclease motifs in cas genes of both cluster types suggests that CRISPR formation involves a DNA degradation step.

Clustered regularly interspaced short palindrome repeats (CRISPRs) have spacers of extrachromosomal origin.

A procedure for the rapid isolation of plasmid DNA larger than 30 megadaltons from lactic streptococci is described This protocol can be used on a preparative scale to isolate sufficient quantities of plasmid DNA required for restriction analysis, cloning, or transformation experiments A scaled-down protocol is very useful for rapidly screening the plasmid content of streptococcal strains With this methodology, previously undetected large plasmids were observed

Simple and rapid method for isolating large plasmid DNA from lactic streptococci.

Lactic acid bacteria (LAB) constitute a group of gram-positive bacteria united by a constellation of morphological, metabolic, and physiological characteristics The general description of the bacteria included in the group is gram-positive, nonsporing, nonrespiring cocci or rods, which produce lactic acid as the major end product during the fermentation of carbohydrates The LAB term is intimately associated with bacteria involved in food and feed fermentation, including related bacteria normally associated with the (healthy) mucosal surfaces of humans and animals The boundaries of the group have been subject to some controversy, but historically the genera Lactobacillus, Leuconostoc, Pediococcus, and Streptococcus form the core of the group Taxonomic revisions of these genera and the description of new genera mean that LAB could, in their broad physiological definition, comprise around 20 genera However, from a practical, food-technology point of view, the following genera are considered the principal LAB: Aerococcus, Carnobacterium, Enterococcus, Lactobacillus, Lactococcus, Leuconostoc, Oenococcus, Pediococcus, Streptococcus, Tetragenococcus, Vagococcus, and Weissella The genus Bifidobacterium, often considered in the same context as the genuine lactic acid bacteria and sharing some of their typical features, is phylogenetically unrelated and has a unique mode of sugar fermentation The classification of lactic acid bacteria into different genera is largely based on morphology, mode of glucose fermentation, growth at different temperatures, configuration of the lactic acid produced, ability to grow at high salt concentrations, and acid or alkaline tolerance Chemotaxonomic markers such as fatty acid composition and constituents of the cell wall are also used in classification In addition, the present taxonomy relies partly on true phylogenetic relationships,

Lactic acid bacteria: classification and physiology.

Incorporation of 1.9% beta-disodium glycerophosphate (GP) into a complex medium resulted in improved growth by lactic streptococci at 30 C. The medium, called M17, contained: Phytone peptone, 5.0 g; polypeptone, 5.0 g; yeast extract, 2.5 g; beef extract, 5.0 g; lactose, 5.0 g; ascorbic acid, 0.5 g; GP, 19.0 g; 1.0 M MgSO(4).7H(2)O, 1.0 ml; and glass-distilled water, 1,000 ml. Based on absorbance readings and total counts, all strains of Streptococcus cremoris, S. diacetilactis, and S. lactis grew better in M17 medium than in a similar medium lacking GP or in lactic broth. Enhanced growth was probably due to the increased buffering capacity of the medium, since pH values below 5.70 were not reached after 24 h of growth at 30 C by S. lactis or S. cremoris strains. The medium also proved useful for isolation of bacterial mutants lacking the ability to ferment lactose; such mutants formed minute colonies on M17 agar plates, whereas wild-type cells formed colonies 3 to 4 mm in diameter. Incorporation of sterile GP into skim milk at 1.9% final concentration resulted in enhanced acid-producing activity by lactic streptococci when cells were inoculated from GP milk into skim milk not containing GP. M17 medium also proved superior to other media in demonstrating and distinguishing between lactic streptococcal bacteriophages. Plaques larger than 6 mm in diameter developed with some phage-host combinations, and turbid plaques, indicative of lysogeny, were also easily demonstrated for some systems.

Improved Medium for Lactic Streptococci and Their Bacteriophages

Single colony isolates of lactic streptococcal strains, most of which have been utilized for cheese-making in New Zealand, were u.v.-irradiated and their growth response was followed optically. All 45 strains showed either lysis or impairment of growth. Their concentrated, bacteria-free supernatants were examined by electron microscopy for the presence of phage particles. Intact phages were observed in most lysates, while disrupted phage parts were found in the remaining lysates. Plaque-forming ability was demonstrable in a few cases. Lysogenization of a different strain by one of the phages detected by this technique was attempted. Similarities and differences with respect to lytic phages which have been isolated for these same strains in New Zealand cheese factories are discussed in relation to the possible origin and control of phages.

/pdf/bacteriophage-production-following-exposure-of-lactic-3wv321psgv.pdf

Bacteriophage Production Following Exposure of Lactic Streptococci to Ultraviolet Radiation

The role of the collar/whisker complex in bacteriophage T4D tail fiber attachment.

Purification and characterization of the protein product of gene 11 of bacteriophage T4D.

The efficiency of plating of phages derived by ultraviolet induction of, or by lytic growth on, certain strains of Streptococcus cremoris was found to vary by as much as 105 depending on the lactose concentration of the medium in which the indicator bacteria were grown and the length of time the stationary-phase indicator cultures were aged. This effect was noted only when the culture was used as an indicator for phages that had previously grown on an apparently unrelated strain of bacteria. Conditions of culturing and aging had no detectable effect upon the ability of a strain to serve as an indicator for phage that had previously been cultured on the same strain. These observations suggest the presence of some kind of physiologically labile restriction system in strains of S. cremoris. The implications of this finding for increasing the sensitivity of the host range test in determining phage susceptibility, whether from induced lysates, whey, or lytic phage stocks, are discussed. It is recommended that, for all such testing, the concentration of lactose in buffered media be increased to such levels as required to obtain a final pH similar to that of a freshly coagulated milk culture, namely, below pH 5.0.

Betty E. Terzaghi

Papers

Improved Medium for Lactic Streptococci and Their Bacteriophages

Bacteriophage Production Following Exposure of Lactic Streptococci to Ultraviolet Radiation

The role of the collar/whisker complex in bacteriophage T4D tail fiber attachment.

Purification and characterization of the protein product of gene 11 of bacteriophage T4D.

Effect of Lactose Concentration on the Efficiency of Plating of Bacteriophages on Streptococcus cremoris