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[Bacteriocins of Clostridium septicum (author's transl)].

17 strains of Clostridium septicum were examined for bacteriocin production. Two bacteriocin-producing strains were detected by the stab culture method and the spot test using the supernatant fluids of the cultures. The bacteriocin of Cl. septicum Ovinus was active against all other strains of Cl. septicum without inhibiting the donro strain (Table 2). This bacteriocin and one indicator strain (Wex) were chosen for further study. The bacteriocin, which was spontaneously produced during the logarithmic growth phase of the bacteriocinogenic strain Ovinus (Fig. 1) was not inducible with high temperatures or Mitomycin C. It was resistent to chloroform, but sensitive to heat (Fig. 4, 5) and proteolytic enzymes (Table 4). Its MG is about 20000 (Fig. 3).

Animals↗

Bacteriocin-like antagonism in Yersinia enterocolitica.

A total of 121 Yersinia enterocolitica O3 isolates from patients with gastroenteritis and 37 Y. enterocolitica reference strains with different O antigens were tested for bacteriocine production and sensitivity. By using cross-streaking method strains belonging to serogroups of O5; O7,8; O7,13; O11; O11,23; O13,27; O17; O19,8 and O34 produced bacteriocin-like substances. None of the Y. enterocolitica O3 strains produced bacteriocin-like material and most of them were uniformly sensitive against the bacteriocin-like material produced by strains of serogroups O7,8; O7,13; O13,27 and O19,8. By sodium dodecyl sulphate-polyacrylamide gel electrophoresis (SDS-PAGE) significant differences were demonstrated in the whole cell protein patterns of Y. enterocolitica reference strains belonging to different serogroups in the range of 33-47 kilodalton (kDa). Out of the ten examined bacteriocin-like material producer strains only one strain harboured a plasmid of about 60 megadalton (MDa).

Antibiosis↗

[Bacteriocins: criteria, classification, characteristics, methods of detection].

The review on bacteriocins of Gram negative and Gram positive bacteria. Criteria making it possible to regard antagonistic substances as bateriocins or bacteriocin-like substances and on their classification are presented. Examples of bacteriocins naming depending on the taxonomic position of the producer culture are given. Information on the physico-chemical and biological properties of bacteriocins and their purification is presented as well as on detection tools of bacteriocins in microorganisms and evaluation of the producer activity of the bacteriological culture.

Bacteria↗

Synthesis of bacteriocins in liquid cultures of Streptococcus mutans.

Strains of Streptococcus mutans synthesized bacteriocins in agar plates, but synthesis of detectable bacteriocins in liquid media took place only under certain culture conditions. The composition of the medium proved to be crucial. Trypticase Soy Broth with 4% Yeast Extract meeting the requirements. The effect of the Yeast Extract is obscure, for some strains also formed detectable bacteriocins in a special Trypticase medium without this agent. It was noted that the broth should be filter-sterilized rather than autoclaved and only a few days old. Attempts at liberating cell-bound bacteriocins from washed cells were unsuccessful, even when they were treated with ultrasound, EDTA, or various chemicals followed by ultrasound. On the basis of size and sensitivity to heat the bacteriocins could be divided into two groups, while their resistance to ether and chloroform and to trypsin did not follow this pattern. Dependence on plasmids could not be demonstrated by attempts at curing with acridine orange or ethidium bromide; and the involvement of phages was unlikely, since the inhibition was not transmissible and phage-like structures were not observed in the electron microscope.

Bacteriocins↗

[Differentiation of several species of lactobacilli of the subgenus Thermobacterium according to their bacteriocin sensitivity spectra].

The method of retarded antagonism was applied to the study of L. acidophilus (187 cultures), L. salivarius (65 cultures), L. jugurti (16 cultures), L. helveticus (4 cultures) sensitivity to the action of 39 bacteriocines produced by various lactobacilli species. By the sensitivity spectra to bacteriocines L. ècidophilus cultures were divided into 76, L. salivarius--22, L. jugurti--15, and L. helveticus into 2 bacteriocines. By bacteriocine typing it was possible to differentiate into a number of bacteriocine types L. acidophilus and L. salivarius variants with the same biochemical and physiological properties.

Bacteriocins↗

[Bacteriocin properties of lactobacilli of the subgenus Streptobacterium].

Morphology of the growth inhibition zones of Lactobacilli indicator strains, as well as cellophane diffusion, thermostability, sensitivity to bacterial proteases, trypsin, chemotrypsin, pepsin, papain, nucleases and lysosyme of 3 bacteriocins produced by L. casei and 2 bacteriocins produced by L. plantarum was studied. By differences in their properties the bacteriocins were divided into 5 types, including 3 types of bacteriocins from L. casei and 2 types of bacteriocins from L. plantarum.

Antibiosis↗

Bacteriocin typing of environmental Klebsiella isolates.

A total of 88 environmental Klebsiella pneumoniae and K. oxytoca isolates were typed according to their bacteriocin susceptibility patterns. Bacteriocin typing was performed by a modification of the scrape-and-point method, using a set of 8 producer strains. This method proved to be very applicable to environmental strains of both species; all but one strain could be typed. Twenty-one different patterns were observed. Type 1-3456-8 was most common in both species. In contrast to K. pneumoniae, more than half of the K. oxytoca strains fell into only two bacteriocin susceptibility patterns. Bacteriocin types 1 and 3 showed a very broad spectrum, more than 95% of the isolates being sensitive to one of these bacteriocins.

Bacteriocins↗

[The antimicrobial spectrum of the action of bacteriocins and bacteriophages from Klebsiella strains].

The percentage of bacteriocin-producing and phage-producing Klebsiella strains was as follows: K. pneumoniae-10%, K. ozaenae-7%, K. rhinoscleromatis-9%. The antimicrobial spectrum of the studied inducible particler was broad and was not limited by the frames of the genus and family. Bacteriocins and bacteriophages from Klebsiella were active to Klebsiella, Enterobacter, Escherichia, Shigella and Proteus representatives significant in medicine. Klebocins and Klebsiella phages exhibited antagonistic effects to phytopathogenic bacteria. Some strains of Erwinia and Pseudomonas were sensitive to phages or bacteriocins from Klebsiella. Bacteriocins protected corn and tomato seeds from contamination by erwinioses agents. All cultures of Agrobacterium, Corynebacterium, Micrococcus, Staphylococcus, Streptococcus were resistant to action of phages and klebocins. Bacteriocins from Klebsiella were assayed for their sensitivity to trypsin, chymotrypsin, lysozyme, ribonuclease, deoxyribonuclease. Action of klebocins was associated with a protein component. Proceeding from data of diffusion through the disc ultrafiltration membranes molecular weight of klebocins was in the range of 30,000 and 50,000 Da.

Animals↗

[Bacteriocin typing of lactobacilli of the Streptobacterium subgenus].

Sensitivity of 144 L. casei cultures and 23 L. plantarum cultures to the action of 39 bacteriocins produced by lactobacillae of different species was studied by the method of delayed antagonism. By the bacteriocin sensitivity the L. casei cultures were subdivided into 37 and L. plantarum cultures--into 7 bacteriocin types. The method of bacteriocin typing permitted differentiation of the L. casei and L. plantarum cultures, which had the same biochemical and physiological properties, as a number of bacteriocin types.

Bacteriocins↗

Effect of base substitutions in the colicin E1 gene on colicin E1 export and bacteriocin activity.

Base substitutions have been introduced into the segment of the colicin E1 gene corresponding to the polypeptide region between the 404th and the 502nd residues which was considered to participate in colicin E1 export and bacteriocin activity. The methods used were in vitro localized mutagenesis with sodium bisulphite and in vivo mutagenesis using either nitrosoguanidine or ethyl methane sulphonate. Cells carrying mutagenized plasmids were screened by their inability to form a clear zone on a lawn of colicin E1 sensitive cells. Mutation sites were determined from the nucleotide sequence analysis and the altered amino acid residues were reduced. The mutant proteins were analysed for their ability to be exported to the periplasmic space and for their bacteriocin activity. Out of eight mutants obtained, three had a single amino acid replacement. Mutant proteins that had Ser and Glu in place of Pro-462 and Gly-502, respectively, showed a decrease in both the export and the bacteriocin activity. A mutant protein having Arg in place of Gly-439 showed a decrease only in the bacteriocin activity. These results suggest that the target region of colicin E1 contributes to the export as well as the bacteriocin activity but the two functions are supported in part by different amino acid residues of the protein.

Amino Acid Sequence↗

Acute lymphoblastic leukemia of childhood monitored by bacteriocin and flowcytometry.

Bacteriocin and flowcytometric analysis of 106 blood samples from children with acute lymphoblastic leukemia were correlated with clinical stages of disease. Bacteriocins interacted selectively with malignant, and not with normal, lymphocytes causing cell cycle perturbation which was rapidly and objectively recorded by the flowcytometer. The patients were grouped as: (A) newly-diagnosed (15); (B) early induction (11); (C) remission with viral infection (7); (D) remission (64); (E) bone marrow relapsed (5); (F) extramedullary relapsed (3); (G) non-malignant pediatric controls (8). Bacteriocin reacted usually with groups A, B, C, E and not with groups D, F and G. Repeated testing correlated well with the clinical status. Blood from 7 patients in remission and from 3 normal individuals, each with transient viral infection, reacted with bacteriocin. A quantitative correlation between peripheral blood blasts or surface markers for ALL and bacteriocin reactivity was not established. Unexpected results were obtained only in 13% (false-positive 11% and false-negative 3%). This test can be recommended for preliminary diagnosis and possibly prognosis of lymphoblastic leukemia and provides means of monitoring progress during chemotherapy.

Antigens, Surface↗

Purification and characterization of a bacteriocin produced by Lactococcus lactis subsp. lactis H-559 isolated from kimchi.

Lactic acid bacteria were isolated from kimchi and screened for bacteriocin production. Strain H-559, identified as Lactococcus lactis subsp. lactis, exhibited the strongest antibacterial activity among them and was active against pathogenic bacteria such as Listeria monocytogenes and Staphylococcus aureus as well as many lactic acid bacteria. The antimicrobial substance produced by L. lactis subsp. lactis H-559 was inactivated by alpha-chymotrypsin, and protease type IX and XIV and was confirmed to be a bacteriocin. The bacteriocin activity was stable from pH 2.0-11.0 and up to 10 min heating at 100 degrees C. The bacteriocin was sequentially purified by ammonium sulfate precipitation, ion-exchange chromatography, and reversed-phase high-performance liquid chromatography (HPLC). Its molecular weight was determined to be 3343.7 Da by MALDI-mass spectrometry. Isoleucine was detected as the first N-terminal amino acid residue but the remaining amino acid sequence could not be determined by the Edman degradation method. It was different from other bacteriocins in terms of pH stability, molecular weight, amino acid composition, and the partial amino acid sequences of peptides obtained by acid hydrolysis.

Journal Article↗

Use of Lactobacillus plantarum LPCO10, a Bacteriocin Producer, as a Starter Culture in Spanish-Style Green Olive Fermentations.

Bacteriocin-producing Lactobacillus plantarum LPCO10 and its non-bacteriocin-producing, bacteriocinimmune derivative, L. plantarum 55-1, were evaluated separately for growth and persistence in natural Spanish-style green olive fermentations. Both strains were genetically marked and selectively enumerated using antibiotic-containing media. Plasmid profile and bacteriocin production (bac) were used as additional markers. When olive brines were inoculated at 10 CFU/ml, the parent strain, LPCO10, proliferated to dominate the epiphytic microflora, sharing high population levels with other spontaneously occurring lactobacilli and persisting throughout the fermentation (12 weeks). In contrast, the derivative strain could not be isolated after 7 weeks. Stability of both plasmid profile and bac (LPCO10 strain) or bac (55-1 strain) phenotype was shown by L. plantarum LPCO10 and L. plantarum 55-1 isolated throughout the fermentation. Bacteriocin activity could be found in the L. plantarum LPCO10-inoculated brines only after ammonium sulfate precipitation and concentration (20 times) of the final brine. Spontaneously occurring lactobacilli and lactic coccus populations, which were isolated from each of the fermenting brines studied during this investigation, were shown to be sensitive to the bacteriocins produced by L. plantarum LPCO10 when tested by the drop diffusion test. The declines in both pH and glucose levels throughout the fermentative process were similar in L. plantarum LPCO10- and in L. plantarum 55-1-inoculated brines and were comparable to the declines in the uninoculated brines. However, the final concentration of lactic acid in L. plantarum LPCO10-inoculated brines was higher than in the L. plantarum 55-1-inoculated brines and uninoculated brines. These results indicated that L. plantarum LPCO10 may be useful as a starter culture to control the lactic acid fermentation of Spanish-style green olives.

Journal Article↗

PROBABLE IDENTITY OF A GROUP D HEMOLYSIN WITH A BACTERIOCINE.

Brock, Thomas D. (Indiana University, Bloomington) and Joseph M. Davie. Probable identity of a group D hemolysin with a bacteriocine. J. Bacteriol. 86:708-712. 1963.-All strains of Streptococcus zymogenes (S. faecalis var. zymogenes) produce a bacteriocine which is active against lactic acid bacteria and most other grampositive bacteria. Mutants which have lost the hemolytic characteristic lose at the same time their bacteriocine-producing ability. A strain which was resistant to the bacteriocine but which was nonhemolytic and nonbacteriocinogenic was irradiated, and two hemolytic mutants were isolated from it. These mutants were also bacteriocinogenic. Thus, the two activities are gained or lost together by mutation. Both activities are destroyed by chloroform vapors and are antagonized by lecithin. Both activities are destroyed at the same rate by treatment at 45 C under mildly acid conditions, and both activities are stable when heated in agar. The two activities are produced in parallel during the growth cycle, and disappear in parallel. The possible ecological role of a substance which is both a hemolysin and a bacteriocine is discussed.

Bacteriolysis↗

[Components of fermentation medium regulate bacteriocin synthesis by the recombinant strain Lactococcus lactis subsp. lactis F-116].

The regulation of the synthesis of bacteriocin produced by the recombinant strain Lactococcus lactis subsp. lactis F-116 has been studied. The synthesis is regulated by the components of the fermentation medium, the content of inorganic phosphate (KH2PO4), yeast autolysate (source of amine nitrogen), and changes in carbohydrates and amino acids. The strain was obtained by fusion of protoplasts derived from two related L. lactis subsp. lactis strains, both exhibiting a weak ability to synthesize the bacteriocin nisin. Decreasing the content of KH2PO4 from 2.0 to 1.0 or 0.5% caused bacteriocin production to go down from 4100 to 2800 or 1150 IU/ml, respectively; the base fermentation medium contained 1.0% glucose, 0.2% NaCl, 0.02% MgSO4, and yeast autolysate (an amount corresponding to 35 mg % ammonium nitrogen). The substitution of sucrose for glucose (as the source of carbon) increased the antibiotic activity by 26%, and the addition of isoleucine, by 28.5%. Elevation of the concentration of yeast autolysate in the low-phosphate fermentation medium stimulated both the growth of the lactococci and the synthesis of bacteriocin. Introduction of 1% KH2PO4, yeast autolysate (in an amount corresponding to 70 mg % ammonium nitrogen), 2.0% sucrose, and 0.1% isoleucine increased the bacteriocin-producing activity of the strain by 2.4 times.

Culture Media↗

Bacteriocin diversity in Bacillus sphaericus.

Nondenaturing polyacrylamide gel electrophoresis revealed the presence of diversity among bacteriocins produced by strains of Bacillus sphaericus. Bacteriocin bands of six strains (pathogenic and non pathogenic) were found to be located just below the stacking gel. However, in two other strains (1 pathogenic and 1 collection strain) more than one protein band with bacteriocin activity were seen in the middle of resolving gel. In bacteriocin-treated cultures, electron-microscopy studies revealed the growth of lysedswollen ghost cells, and loss of viability among sensitive strains.

Animals↗

Identification of the DNA-binding sites for two response regulators involved in control of bacteriocin synthesis in Lactobacillus plantarum C11.

In Lactobacillus plantarum C11, bacteriocin production has previously been shown to be an inducible process, in which a secreted peptide, produced by the host itself, is involved. The inducing factor, designated plantaricin A (PlnA), is a bacteriocin-like peptide encoded by a gene (plnA) located on the same operon as the genes for a two-component regulatory system (plnBCD). This system consists of a histidine kinase (PlnB) and two response regulators (PlnC,D), and belongs to a recently defined subfamily of two-component regulatory systems, which are activated by secreted peptide pheromones through a quorum-sensing mechanism. We show here that the two response regulators PlnC and PlnD bind specifically to imperfect direct repeats found within the adjacent promoter of the plnABCD operon, and to similar sequences found within the promoter regions of two nearby operons containing bacteriocin structural genes (plnEFI and plnJKLR). Binding of PlnC and PlnD was increased two to three fold in the presence of acetyl phosphate. The results suggest that bacteriocin synthesis in L. plantarum C11 is regulated by the DNA-binding activity of the two response regulators PlnC and PlnD.

Bacteriocins↗

Influence of temperature and pH on production of two bacteriocins by Leuconostoc mesenteroides subsp. mesenteroides FR52 during batch fermentation.

The influence of temperature and pH on growth of Leuconostoc mesenteroides subsp. mesenteroides FR52 and production of its two bacteriocins, mesenterocin 52A and mesenterocin 52B, was studied during batch fermentation. Temperature and pH had a strong influence on the production of the two bacteriocins which was stimulated by slow growth rates. The optimal temperature was 20 degrees C for production of mesenterocin 52A and 25 degrees C for mesenterocin 52B. Optimal pH values were 5.5 and 5.0 for production of mesenterocin 52A and mesenterocin 52B respectively. Thus, by changing the culture conditions, production of one bacteriocin can be favoured in relation to the other. The relationship between growth and specific production rates of the two bacteriocins, as a function of the culture conditions, showed different kinetics of production and the presence of several peaks in the specific production rates during growth.

Animals↗