Microbial Ecology of Watery Kimchi

© 2015 Institute of Food Technologists®. This article has been contributed by US Government employees and their work is in the public domain in the USA. The biochemistry and microbial ecology of 2 similar types of watery (mul) kimchi, containing sliced and unsliced radish and vegetables (nabak and dongchimi, respectively), were investigated. Samples from kimchi were fermented at 4, 10, and 20 °C were analyzed by plating on differential and selective media, high-performance liquid chromatography, and high-throughput DNA sequencing of 16S rDNA. Nabak kimchi showed similar trends as dongchimi, with increasing lactic and acetic acids and decreasing pH for each temperature, but differences in microbiota were apparent. Interestingly, bacteria from the Proteobacterium phylum, including Enterobacteriaceae, decreased more rapidly during fermentation at 4 °C in nabak cabbage fermentations compared with dongchimi. Although changes for Proteobacterium and Enterobacteriaceae populations were similar during fermentation at 10 and 20 °C, the homolactic stage of fermentation did not develop for the 4 and 10 °C samples of both nabak and dongchimi during the experiment. These data show the differences in biochemistry and microbial ecology that can result from preparation method and fermentation conditions of the kimchi, which may impact safety (Enterobacteriaceae populations may include pathogenic bacteria) and quality (homolactic fermentation can be undesirable, if too much acid is produced) of the product. In addition, the data also illustrate the need for improved methods for identifying and differentiating closely related lactic acid bacteria species using high-throughput sequencing methods.This work was carried out as part of the international collaborative R&D program funded by the Agency for Korea Natl. Food Cluster (2013), and supported in part by a grant from Pickle Packers Intl. Inc., Washington, D.C., U.S.A. The authors thank the Spanish Government (MECD) for the postdoctoral fellowship support for Dr. E. Medina-Pradas.Peer Reviewe

Multifunctional ZnO/Nylon 6 nanofiber mats by an electrospinning–electrospraying hybrid process for use in protective applications

ZnO/Nylon 6 nanofiber mats were prepared by an electrospinning–electrospraying hybrid process in which ZnO nanoparticles were dispersed on the surface of Nylon 6 nanofibers without becoming completely embedded. The prepared ZnO/Nylon 6 nanofiber mats were evaluated for their abilities to kill bacteria or inhibit their growth and to catalytically detoxify chemicals. Results showed that these ZnO/Nylon 6 nanofiber mats had excellent antibacterial efficiency (99.99%) against both the Gram-negative Escherichia coli and Gram-positive Bacillus cereus bacteria. In addition, they exhibited good detoxifying efficiency (95%) against paraoxon, a simulant of highly toxic chemicals. ZnO/Nylon 6 nanofiber mats were also deposited onto nylon/cotton woven fabrics and the nanofiber mats did not significantly affect the moisture vapor transmission rates and air permeability values of the fabrics. Therefore, ZnO/Nylon 6 nanofiber mats prepared by the electrospinning–electrospraying hybrid process are promising material candidates for protective applications

Use of Linear Models for Thermal Processing of Acidified Foods

Acidified vegetable products with a pH above 3.3 must be heat processed to assure the destruction of Escherichia coli O157:H7, Salmonella enterica, Listeria monocytogenes, and other pathogenic bacteria that might be present in the product. Recently, the Food and Drug Administration has required that linear models for heat process data be used with electronic process filing forms. Existing recommendations for heat processing acidified vegetables are based on non-linear (Weibull and exponential decay) models. We report here the parameters for a linear model that meets or exceeds the established heat processing conditions needed to assure safety

Synergistic Effects of Sodium Chloride, Glucose, and Temperature on Biofilm Formation by Listeria monocytogenes Serotype 1/2a and 4b Strains ▿ † ‡

Biofilm formation by Listeria monocytogenes is generally associated with its persistence in the food-processing environment. Serotype 1/2a strains make up more than 50% of the total isolates recovered from food and the environment, while serotype 4b strains are most often associated with major outbreaks of human listeriosis. Using a microplate assay with crystal violet staining, we examined biofilm formation by 18 strains of each serotype in tryptic soy broth with varying concentrations of glucose (from 0.25% to 10.0%, wt/vol), sodium chloride (from 0.5% to 7.0%, wt/vol) and ethanol (from 1% to 5.0%, vol/vol), and at different temperatures (22.5°C, 30°C, and 37°C). A synergistic effect on biofilm formation was observed for glucose, sodium chloride, and temperature. The serotype 1/2a strains generally formed higher-density biofilms than the 4b strains under most conditions tested. Interestingly, most serotype 4b strains had a higher growth rate than the 1/2a strains, suggesting that the growth rate may not be directly related to the capacity for biofilm formation. Crystal violet was found to stain both bacterial cells and biofilm matrix material. The enhancement in biofilm formation by environmental factors was apparently due to the production of extracellular polymeric substances instead of the accumulation of viable biofilm cells

Competition of Listeria monocytogenes Serotype 1/2a and 4b Strains in Mixed-Culture Biofilms▿ †

The majority of Listeria monocytogenes isolates recovered from foods and the environment are strains of serogroup 1/2, especially serotypes 1/2a and 1/2b. However, serotype 4b strains cause the majority of human listeriosis outbreaks. Our investigation of L. monocytogenes biofilms used a simulated food-processing system that consisted of repeated cycles of growth, sanitation treatment, and starvation to determine the competitive fitness of strains of serotypes 1/2a and 4b in pure and mixed-culture biofilms. Selective enumeration of strains of a certain serotype in mixed-culture biofilms on stainless steel coupons was accomplished by using serotype-specific quantitative PCR and propidium monoazide treatment to prevent amplification of extracellular DNA or DNA from dead cells. The results showed that the serotype 1/2a strains tested were generally more efficient at forming biofilms and predominated in the mixed-culture biofilms. The growth and survival of strains of one serotype were not inhibited by strains of the other serotype in mixed-culture biofilms. However, we found that a cocktail of serotype 4b strains survived and grew significantly better in mixed-culture biofilms containing a specific strain of serotype 1/2a (strain SK1387), with final cell densities averaging 0.5 log10 CFU/cm2 higher than without the serotype 1/2a strain. The methodology used in this study contributed to our understanding of how environmental stresses and microbial competition influence the survival and growth of L. monocytogenes in pure and mixed-culture biofilms

Sequence Analysis of Leuconostoc Mesenteroides Bacteriophage Φ1-A4 Isolated from an Industrial Vegetable Fermentation

Vegetable fermentations rely on the proper succession of a variety of lactic acid bacteria (LAB) including Leuconostoc mesenteroides. L. mesenteroides initiates the fermentation, producing lactic and acetic acids, CO2, and many flavor compounds. As the fermentation proceeds, L. mesenteroides dies off and other LAB complete the fermentation. Phage infecting L. mesenteroides may significantly influence the die-off of L. mesenteroides. However, no L. mesenteroides phages have been previously sequenced and genetically characterized. Knowledge of more phage genome sequences may provide new insights into phage genomics and phage-host interactions. We have determined the complete genome sequence of L. mesenteroides phage (phi)1-A4, which was isolated from an industrial sauerkraut fermentation. The phage possesses a linear, double-stranded, DNA genome consisting of 29,508 bp with a G+C content of 36%. Fifty open reading frames (ORFs) were predicted. Based on N-terminal amino acid sequencing and bioinformatic analyses, putative functions were assigned to 26 ORFs (52%), including 5 ORFs of structural proteins. The phage genome was found to be modularly organized and consist of DNA replication, DNA packaging, head-and-tail morphogenesis, cell lysis, and DNA regulation/modification modules. In silico analyses supported the observation that (phi)1-A4 is a unique lytic phage. A large scale genome inversion (~30% of the genome) was identified by comparison with other phages. The genome inversion encompassed the lysis module, part of the structural protein module and a putative cos-site. Promoter structures were identified that may initiate the transcription of the inverted genome region. Interestingly, the lysin gene was found to be flanked by two holin genes. The tail morphogenesis module was interspersed by cell lysis genes and other genes with unknown functions. The predicted amino acid sequences of the phage proteins showed little similarity with other phages, but functional analyses showed that (phi)1-A4 clusters with several Lactococcus phages. To our knowledge, (phi)1-A4 is the first genetically characterized Leuconostoc mesenteroides phage

Energy-Based Dynamic Model for Variable Temperature Batch Fermentation by Lactococcus lactis

We developed a mechanistic mathematical model for predicting the progression of batch fermentation of cucumber juice by Lactococcus lactis under variable environmental conditions. In order to overcome the deficiencies of presently available models, we use a dynamic energy budget approach to model the dependence of growth on present as well as past environmental conditions. When parameter estimates from independent experimental data are used, our model is able to predict the outcomes of three different temperature shift scenarios. Sensitivity analyses elucidate how temperature affects the metabolism and growth of cells through all four stages of fermentation and reveal that there is a qualitative reversal in the factors limiting growth between low and high temperatures. Our model has an applied use as a predictive tool in batch culture growth. It has the added advantage of being able to suggest plausible and testable mechanistic assumptions about the interplay between cellular energetics and the modes of inhibition by temperature and end product accumulation

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

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