M.C. Johnson

Direct detection of an antimicrobial peptide ofPediococcus acidilactici in sodium dodecyl sulfate-polyacrylamide gel electrophoresis

SummaryAn SDS-PAGE technique is described that allows identification of the antimicrobial activity of a peptide secreted by a strain ofPediococcus acidilactici. This peptide has an antimicrobial property against several baeteria associated with food. This technique enables detection of the specific peptide (or protein) band(s) associated with the inhibitory effect which can then be eluted from the gel for further studies.

Cellular Damage in Dried Lactobacillus acidophilus

Lactobacillus acidophilus cells surviving freeze drying and vacuum drying became sensitive to oxgall and lysozyme probably from damage to the cell wall. The dried cells also became sensitive to NaCl and permeable to orthonitrophenol β-galactoside from damage to the cytoplasmic membrane. Scanning electron microscopy indicated loss of some surface material from the damaged cells. Transmission electron microscopy (TEM) revealed partial loss of wall and membrane material, but these losses seemed to have resulted from the treatments given during fixation of cells for TEM and as a consequence of damage to the wall and membrane that occurred during drying. A surface protein of 46-kilodalton molecular weight, that is bound to the wall by hydrogen bonding, was also lost from the dried cells. It is postulated that drying adversely affects some weak bonds of the cellular macromolecules probably from the loss of bound water.

Spoilage of vacuum-packaged refrigerated beef by Clostridium

A motile, gram-positive, spore forming, anaerobic, psychrotrophic bacterial species, probably from the genus Clostridium , was involved in spoilage of vacuum-packaged refrigerated fresh beef. The spoilage was associated with accumulation of large quantities of foul smelling gas and purge in the bag and loss of color and texture of the meat. Attempts to grow the organism in several laboratory media were not yet successful; however, inoculation of purge from a spoiled sample into a fresh beef, vacuum-packaging and refrigeration storage facilitated growth of this species and produced characteristic spoilage of beef.

Inhibition of Listeria spp. in sterile food systems by pediocin AcH, a bacteriocin produced by Pediococcus acidilactici H

The effectiveness of pediocin AcH, a bacteriocin produced by Pediococcus acidilactici H, in reducing population levels and growth of Listeria monocytogenes strains and Listeria ivanovii in sterile ground beef, sausage mix, cottage cheese, ice cream, and reconstituted dry milk was examined. Predetermined numbers of Listeria cells and concentrations of pediocin AcH were added to the foods suspended in water and the final volumes made to 10 ml. Listeria in the pediocin AcH treated and control samples during storage at 4 and 10°C were enumerated as CFU on tryptic soy agar. Results indicated that: (a) the maximum bactericidal action of pediocin AcH occurred within 1 h and was not interfered with by the foods; (b) Listeria strains differed in sensitivity to pediocin AcH; (c) reduction in CFU was greater with higher levels of pediocin AcH and as the initial cell number of Listeria spp. decreased; and (d) the surviving cells in pediocin AcH treated food samples multiplied in the presence of residual pediocin AcH during storage at 4 and 10°C.

Bacteriocin plasmids ofPediococcus acidilactici

SummaryPediococcus acidilactici strains E, F and H isolated from fermented sausages produced bacteriocins which were protein in nature and inhibitory to a variety of spoilage and pathogenic microorganisms often encountered in foods. These strains harbored two to three plasmids ranging in size from 7.4 to 40.2 megadaltons. Curing experiments and plasmid profile analysis indicated the involvement of plasmid DNA with bacteriocin activity in all three strains. Carbohydrate fermentation and antibiotic resistance phenotypes did not appear to be associated with bacteriocin plasmids. Both bacteriocin activity and resistance determinants were linked in strain H and mediated by a 7.4-megadalton plasmid, whereas in strains E and F these two traits were not linked.

Viability loss of foodborne pathogens by starter culture metabolites

Antibacterial metabolites produced by starter culture bacteria were tested for inhibitory properties against foodborne pathogens capable of growing in foods at refrigeration temperature. Three culture preparations containing the bacteriocins pediocin AcH, nisin, and sakacin A produced by Pediococcus acidilactici H, Lactococcus lactis ssp. lactis DL 16, and Lactobacillus sake 706, respectively, were studied in addition to four commercial preparations: Nisaplin (similar to nisin), Na-lactate, diacetyl and Microgard. All bacteriocin preparations were bactericidal to the Listeria strains tested including nine strains of L. monocytogenes , but not to any of the gram-negative pathogens studied. Some strains of Listeria were lysed by pediocin AcH. Storage studies at 4°C indicated that the bacteriocin preparations did not have bacteriostatic properties against the surviving cells. Diacetyl produced some bactericidal effect against strains of Yersinia enterocolitica , Aeromonas hydrophila , pathogenic Escherichia coli , and Salmonella anatum , but not against Listeria . Lactate had limited bacteriostatic effect against gram-negative pathogens and none against the gram-positive bacteria. Microgard had neither bactericidal nor bacteriostatic action against the bacteria used in this study.

Selection of Lactobacillus acidophilus strains for use in «acidophilus products»

AbstractRecent studies by DNA-DNA hybridization revealed that strains now designated as L. acidophilus, can be divided into several groups and only one group should be classified as L. acidophilus. We studied several phenotypic characteristics in representative strains from the six DNA-homology groups of L. acidophilus. No group specific pattern was observed among the strains for fermentation of eight carbohydrates, growth at 15 and 45°C, resistance to 0.2% oxgall, lysis by lysozyme or sensitivity to 17 antibiotics. However, some differences among groups were observed in β-galactosidase (β-gal) activity and surface layer (s-layer) protein. Strains in B1 do not have a s-layer or β-gal while B2 strains also lack a s-layer but do possess β-gal. All strains in groups A1, A2, A3 and A4, capable of growing in lactose, have β-gal activity and also have a s-layer composed of protein subunits of different molecular weights (MW). Strains in A1 homology group have a s-layer with 46 Kd protein subunits while strains in other A groups have s-layer protein subunits that varied in MW within each group. On the basis of these two traits several isolates of unknown homology groups have been tentatively placed in A1, B1 or B2 groups. L. acidophilus from A1 group showed strain variation in β-gal specific activity and rate of acid production and growth. For use in dietary adjuncts, L. acidophilus strains should be selected for these three and other desirable traits. They should be maintained and grown in media containing lactose.

Lactose hydrolyzing enzymes in Lactobacillus acidophilus strains

Abstract Five Lactobacillus acidophilus strains were examined for lactose hydrolyzing enzymes under several growth conditions. All have both β-galactosidase (β-gal) and phospho-β-galactosidase (P-β-gal) activities. Strains W, ATCC 4356 and NCDO 1748 had different amounts of β-gal activity, but all had 100-fold or more higher than P-β-gal activity. In contrast, in strains ATCC 4962 and ATCC 19992, activities of both enzymes were similar. The β-gal synthesis in strains W, 4356 and 1748 was induced by lactose and galactose, while in strains 4962 and 19992 it appeared not to be inducible. At 45°C in the presence of lactose synthesis of only β-gal occurred in strains 4356, 1748 and W. These differences in lactose hydrolyzing enzymes between the strains could be used, along with other characteristics, for taxonomic differentiation between L. acidophilus strains. Studies with L. acidophilus W indicated that cells have maximum β-gal activity at their early stationary phase of growth and when grown in the presence of an inducer and in the absence of oxgall. Selection of strains, type of carbohydrate in growth medium, growth temperature, culture age, and the absence of oxgall during cell growth should be considered in producing L. acidophilus cultures containing high β-gal activity for use as a dietary adjunct by lactose-intolerant individuals and in the manufacture of acidophilus-fermented dairy products.

Factors influencing synthesis and activity of β-galactosidase inLactobacillus acidophilus

SummaryIn the type-strainLactobacillus acidophilus ATCC 4356 β-galactosidase (β-gal) was inducible; lactose, galactose, melibiose and probably maltose, but not glucose, fructose, mannose, sucrose and cellobiose, induced β-gal synthesis. Glucose partially inhibited β-gal-induction by lactose but not by isopropyl-β-D-thiogalactoside. β-gal synthesis during cell growth was maximal at 0.4% lactose, stimulated by Ca2+ but inhibited by Mg2+ and Mn2+. β-gal in the cell-free extract had optimum activity at pH 6.5 and at 45°C. The enzyme activity was stimulated by Mg2+, inhibited by Ca2+, destroyed by oxidizing agents and protected by reducing agents.

Isolation and partial characterization of the surface protein of Lactobacillus acidophilus strains

A surface protein layer was found to be present in some Lactobacillus acidophilus strains. it could be visualized with electron microscopy and extracted with sodium dodecyl sulfate or guanidine hydrochloride. The protein was resistant to several proteolytic enzymes, but sensitive to pronase. Amino acid composition analysis revealed about 30% hydrophobic amino acids. Hydrophobicity study with a hexadecane partition assay demonstrated that the cells were hydrophobic and removal of the surface layer with pronase destroyed the hydrophobic nature of the cells. Several growth conditions studies were found to have little effect on synthesis of the surface protein.