H. P. Fleming

Sensitivity and resistance of Listeria monocytogenes ATCC 19115, Scott A, and UAL500 to nisin

Listeria monocytogenes ATCC 19115, Scott A, and UAL500 were evaluated for sensitivity to nisin (0 to 50 μg/ml) using a direct plating method. Nisin (10 μg/ml) decreased an initial population of L. monocytogenes (109 CFU per ml) by 6- to 7-log cycles. Sensitivity to nisin was enhanced by addition of 2% NaCl or by reduction of the medium pH from 6.5 to 5.5 with either hydrochloric or lactic acid. Mutants resistant to 50 μg/ml nisin were detected at frequencies of 10-6 to 10-8. Nisin-resistant L. monocytogenes mutants should be expected to arise when nisin is used as an antimicrobial in food systems.

Isolation and characterization of a Lactobacillus plantarum bacteriophage, ΦJL-1, from a cucumber fermentation

A virulent Lactobacillus plantarum bacteriophage, PhiJL-1, was isolated from a commercial cucumber fermentation. The phage was specific for two related strains of L. plantarum, BI7 and its mutant (deficient in malolactate fermenting ability) MU45, which have been evaluated as starter cultures for controlled cucumber fermentation and as biocontrol microorganisms for minimally processed vegetable products. The phage genome of PhiJL-1 was sequenced to reveal a linear, double-stranded DNA (36.7 kbp). Sodium dodecyl sulfate-polyacryamide gel electrophoresis (SDS-PAGE) profiles indicated that PhiJL-1 contains six structural proteins (28, 34, 45, 50, 61, and 76 kDa). Electron microscopy revealed that the phage has an isometric head (59 nm in diameter), a long non-contractile tail (182 nm in length and 11 nm in width), and a complex base plate. The phage belongs to the Bradley group B1 or Siphoviridae family. One-step growth kinetics of the phage showed that the latent period was 35 min, the rise period was 40 min, and the average burst size was 22 phage particles/infected cell. Phage particles (90%) adsorbed to the host cells 20 min after infection. Calcium supplementation (up to 30 mM CaCl(2)) in MRS media did not affect the first cycle of phage adsorption, but promoted rapid phage propagation and cell lysis in the infection cycle subsequent to adsorption. The D values of PhiJL-1 at pH 6.5 were estimated to be 2.7 min at 70 degrees C and 0.2 min at 80 degrees C by a thermal inactivation experiment. Knowledge of the properties of L. plantarum bacteriophage PhiJL-1 may be important for the development of controlled vegetable fermentations.

DNA Fingerprinting of Lactic Acid Bacteria in Sauerkraut Fermentations

ABSTRACT Previous studies using traditional biochemical identification methods to study the ecology of commercial sauerkraut fermentations revealed that four species of lactic acid bacteria, Leuconostoc mesenteroides, Lactobacillus plantarum, Pediococcus pentosaceus, and Lactobacillus brevis, were the primary microorganisms in these fermentations. In this study, 686 isolates were collected from four commercial fermentations and analyzed by DNA fingerprinting. The results indicate that the species of lactic acid bacteria present in sauerkraut fermentations are more diverse than previously reported and include Leuconostoc citreum, Leuconostoc argentinum, Lactobacillus paraplantarum, Lactobacillus coryniformis, and Weissella sp. The newly identified species Leuconostoc fallax was also found. Unexpectedly, only two isolates of P. pentosaceus and 15 isolates of L. brevis were recovered during this study. A better understanding of the microbiota may aid in the development of low-salt fermentations, which may have altered microflora and altered sensory characteristics.

Microbiological Control of Cucumber Hydrocooling Water with Chlorine Dioxide

The time required to cool size 2B (3.43 to 3.75-cm-diameter) pickling cucumbers by a commercial spray-type hydrocooler to less than 9°C was about 18 min at typical initial fruit temperatures of 25 to 29°C. During this period, the fruit was exposed to the recycled water, which reached relatively high populations of bacteria (106 to 107 colony forming unites [CFU]/g total aerobes and 105 to 106 CFU/g total Enterobacteriaceae) during a typical days operation. These numbers exceeded those present on the unwashed fruit, depending upon the volume of fruit previously cooled. Residual chlorine dioxide at 1.3 ppm was found to optimally control (2 to 6 log-cycles reduction) the numbers of bacteria. At 0.95 ppm chlorine dioxide, the numbers of bacteria in the water were relatively static, while at 2.8 and 5.1 ppm the odor of chlorine dioxide became excessive. The bacterial populations in/on the cucumbers were not greatly influenced by chlorine dioxide, even at 5.1 ppm. Apparently, microorganisms on or in the fruit were protected from the chlorine dioxide. Thus, the use of chlorine dioxide in hydrocooling water of cucumbers seems to be an effective means of controlling microbial build-up in the water, but has little effect upon microorganisms on or in the fruit.

Antimicrobial activity of sulfur compounds derived from cabbage.

Selected sulfur compounds found in cabbage and its fermentation product, sauerkraut, were tested for minimum inhibitory concentration (MIC) against growth of 15 species of bacteria and 4 species of yeasts. S-Methyl-L-cysteine sulfoxide, sinigrin, and dimethyl sulfide at 500 ppm were not inhibitory to any of the bacteria and yeasts tested. Dimethyl disulfide at 500 ppm retarded some, but did not prevent growth of any of the test microorganisms. Dimethyl trisulfide had an MIC to bacteria of 200 ppm and to yeast of 20 ppm. Methyl methanethiosulfinate had an MIC between 50 and 200 ppm for all bacteria, and between 6 and 10 ppm for all yeasts tested. Methyl methanethiosulfonate had an MIC between 20 and 100 ppm for bacteria and between 50 and 500 ppm for yeasts. Allyl isothiocyanate had an MIC between 50 and 500 ppm for bacteria and between 1 and 4 ppm for yeasts. Methyl methanethiosulfinate was 10 to 100 times more inhibitory against Listeria monocytogenes at pH values of 5, 6, and 7 and was much less influenced by pH than was sodium benzoate.

Bacteriophage ecology in commercial sauerkraut fermentations.

ABSTRACT Knowledge of bacteriophage ecology in vegetable fermentations is essential for developing phage control strategies for consistent and high quality of fermented vegetable products. The ecology of phages infecting lactic acid bacteria (LAB) in commercial sauerkraut fermentations was investigated. Brine samples were taken from four commercial sauerkraut fermentation tanks over a 60- or 100-day period in 2000 and 2001. A total of 171 phage isolates, including at least 26 distinct phages, were obtained. In addition, 28 distinct host strains were isolated and identified as LAB by restriction analysis of the intergenic transcribed spacer region and 16S rRNA sequence analysis. These host strains included Leuconostoc, Weissella, and Lactobacillus species. It was found that there were two phage-host systems in the fermentations corresponding to the population shift from heterofermentative to homofermentative LAB between 3 and 7 days after the start of the fermentations. The data suggested that phages may play an important role in the microbial ecology and succession of LAB species in vegetable fermentations. Eight phage isolates, which were independently obtained two or more times, were further characterized. They belonged to the family Myoviridae or Siphoviridae and showed distinct host ranges and DNA fingerprints. Two of the phage isolates were found to be capable of infecting two Lactobacillus species. The results from this study demonstrated for the first time the complex phage ecology present in commercial sauerkraut fermentations, providing new insights into the bioprocess of vegetable fermentations.

Developments in nisin research

Abstract Nisin, produced by some strains of Lactococcus lactis , subsp. lactis was originally described in 1928 and is the most highly characterized bacteriocin produced by lactic acid bacteria. Nisin has been permitted as a food additive in the UK since the early 1960s and is currently an accepted food additive in at least 45 other countries. Technological advances have resulted in a tremendous increase in new information on nisin within the past decade. This review summarizes the recent developments in understanding the structure of nisin, the genetics of its production, and its mode of action.

Identification and Characterization of Leuconostoc fallax Strains Isolated from an Industrial Sauerkraut Fermentation

ABSTRACT Lactic acid bacterial strains were isolated from brines sampled after 7 days of an industrial sauerkraut fermentation, and six strains were selected on the basis of susceptibility to bacteriophages. Bacterial growth in cabbage juice was monitored, and the fermentation end products were identified, quantified, and compared to those of Leuconostoc mesenteroides. Identification by biochemical fingerprinting, endonuclease digestion of the 16S-23S intergenic transcribed spacer region, and sequencing of variable regions V1 and V2 of the 16S rRNA gene indicated that the six selected sauerkraut isolates were Leuconostoc fallax strains. Random amplification of polymorphic DNA fingerprints indicated that the strains were distinct from one another. The growth and fermentation patterns of the L. fallax isolates were highly similar to those of L. mesenteroides. The final pH of cabbage juice fermentation was 3.6, and the main fermentation end products were lactic acid, acetic acid, and mannitol for both species. However, none of the L. fallax strains exhibited the malolactic reaction, which is characteristic of most L. mesenteroides strains. These results indicated that in addition to L. mesenteroides, a variety of L. fallax strains may be present in the heterofermentative stage of sauerkraut fermentation. The microbial ecology of sauerkraut fermentation appears to be more complex than previously indicated, and the prevalence and roles of L. fallax require further investigation.

Modeling the specific growth rate of Lactobacillus plantarum in cucumber extract

Extensive empirical research has been published on the fermentation of vegetables, but little predictive modeling of the process is available. The objectives of this study were to assess the effects of key variables involved in cucumber fermentation and to develop models for predicting the growth of Lactobacillus plantarum in pure and mixed culture fermentations. The growth medium for the studies was cucumber juice. The effects of various concentrations of lactic, acetic, and hydochloric acids and sodium chloride on growth at 30° C were determined in batch culture. Limiting conditions for growth were pH 3.37 (lower limit), 69 mm undissociated lactic acid, 150 mm undissociated acetic acid, or 11.8% NaCl. Acetic acid was stimulatory to growth at low concentrations (up to 40 mm) but inhibitory at higher concentrations. Lactic acid was more inhibitory than acetic acid, whether total or undissociated concentrations were used as the basis of comparison. A predictive equation for specific growth rate was developed, tested, and shown to predict growth of L. plantarum in batch processes reasonably well.

Selection of lactic acid bacteria for use in vegetable fermentations

Abstract The fermentation of brined vegetables traditionally has depended upon growth of naturally occurring lactic acid bacteria to metabolize the vegetable sugars to organic acids which, together with added salt, results in preservation. Starter cultures have been used only to a limited extent commercially. However, recent efforts to improve fermentation vessels and to develop controlled fermentation methods for fermented vegetables has resulted in an increased interest in developing cultures suitable for application in such methods. Rapid and dominant growth, type and extent of acid production, salt tolerance, temperature range, CO 2 production, cell sedimentation, bacteriophage resistance, nutritional value, and ability to survive as concentrated cultures are factors to consider in developing lactic acid bacterial cultures for use in controlled fermentation of vegetables. Recent examples of efforts to improve cultures include development of nonmalate-decarboxylating strains of lactobacilli for use in cucumber fermentation and isolation of a new species of lactobacillus that produces only the L-isomer of lactic acid from the glucose for use in sauerkraut fermentation. Further improvements in starter cultures for fermented vegetables are likely when suitable genetic transfer systems are developed for selective incorporation or deletion of specific traits.