Inactivation of Lactobacillus plantarum WCFS1 during spray drying and storage assessed with complementary viability determination methods

Survival of Lactobacillus plantarum WCFS1 spray-dried and stored under different conditions was investigated using complementary methods. One method involved a cell membrane integrity viability-based determination, the other assessed cell growth behavior in a liquid medium by means of detection time or by conventional plating. Survival decreased below 95% when spray drying was carried out at higher outlet spray drying temperatures (T_out > 70 °C). However, the membrane integrity method provided higher residual viability values compared to the detection time and conventional plating. This suggests that loss of viability may be due to a combination of damage to intracellular components and cell membrane. Also during storage viability based on growth behavior declined faster and was more temperature dependent compared to the viability as determined by the membrane integrity method. Also here additional damage to intracellular components is expected responsible to loss of viability. Major conclusion is that one should not only rely on a cell-membrane integrity based method to assess survival during spray drying and storage of bacteria. Previous studies that did so most probably underestimated viability as critical damage to intracellular components was not assessed.     

Diversity of stress tolerance in Lactobacillus plantarum, Lactobacillus pentosus and Lactobacillus paraplantarum: A multivariate screening study

Sixty-three strains of the taxonomically related species Lactobacillus plantarum subsp. plantarum, L. plantarum subsp. argentoratensis, L. paraplantarum and L. pentosus isolated from sourdoughs and other food and non-food sources and 14 strains of other members of the genus Lactobacillus were screened for their tolerance of acid, alkaline, heat, oxidative, osmotic, detergent and starvation stresses in order to evaluate the diversity of stress response. Most strains of the L. plantarum group were highly tolerant of acid, alkaline and osmotic stress and highly sensitive to detergent stress, while a larger diversity was found for other stress. Multivariate analysis allowed grouping the strains in clusters with similar response patterns. Stress response patterns in the L. plantarum group were similar to those of species of the L. casei/L. paracasei group but clearly different from those of other mesophilic Lactobacillus. No relationship was found between grouping obtained on the basis of stress response patterns and by genotypic fingerprinting (rep-PCR), nor with the taxonomic position or isolation source of the strains. Further experiments with selected strains showed that exponential phase cells were generally but not always more sensitive than stationary phase cells. The ability to grow under stressful conditions showed a slightly better correlation with the ecological conditions prevailing in the isolation niches of the strains.This study will be the basis for further investigations to identify and exploit the basis of diversity in the stress response of lactic acid bacteria.     

Interactions between formulation and spray drying conditions related to survival of Lactobacillus plantarum WCFS1

Protective solid carriers are commonly added to probiotic cultures prior to drying. Their formulation is not trivial and depends on the drying conditions applied. In this study, we systematically investigated the influence of formulation parameters on the survival of Lactobacillus plantarum WCFS1 after drying. Low molecular weight carbohydrates (less than 2 kDa) with high glass transition temperatures provided the highest level of protection at both low (25 °C) and high (50 °C or higher) drying temperatures. Low molecular weight carbohydrates may provide stabilization by closely interacting with the lipid bilayer of the cell membranes. Meanwhile, carbohydrates with high glass transition temperatures probably provide stabilization via fixation of the cells in a glassy powder. Furthermore, adequate amounts of solid carrier are required to sufficiently stabilize the cells during drying. During drying, crystallization of solid carriers may occur. Depending on the crystal geometry, crystallization can be either beneficial (e.g. with mannitol or sorbitol) or detrimental (e.g. with lactose) to cell survival. Finally, the effect of formulation on cell viability during storage was studied. A decimal reduction time of approximately 300 days was observed when spray dried L. plantarum WCFS1 was stored at temperatures below 40 °C. The outcome of this study was used as a basis to construct a generalized diagram to indicate the combinations of formulation and drying conditions to maximally retain viability and operate dryers at high efficiency.     

Genotypic diversity of stress response in Lactobacillus plantarum, Lactobacillus paraplantarum and Lactobacillus pentosus

Lactobacillus plantarum, Lactobacillus pentosus and Lactobacillus paraplantarum are three closely related species which are widespread in food and non-food environments, and are important as starter bacteria or probiotics. In order to evaluate the phenotypic diversity of stress tolerance in the L. plantarum group and the ability to mount an adaptive heat shock response, the survival of exponential and stationary phase and of heat adapted exponential phase cells of six L. plantarum subsp. plantarum, one L. plantarum subsp. argentoratensis, one L. pentosus and two L. paraplantarum strains selected in a previous work upon exposure to oxidative, heat, detergent, starvation and acid stresses was compared to that of the L. plantarum WCFS1 strain. Furthermore, to evaluate the genotypic diversity in stress response genes, ten genes (encoding for chaperones DnaK, GroES and GroEL, regulators CtsR, HrcA and CcpA, ATPases/proteases ClpL, ClpP, ClpX and protease FtsH) were amplified using primers derived from the WCFS1 genome sequence and submitted to restriction with one or two endonucleases. The results were compared by univariate and multivariate statistical methods. In addition, the amplicons for hrcA and ctsR were sequenced and compared by multiple sequence alignment and polymorphism analysis. Although there was evidence of a generalized stress response in the stationary phase, with increase of oxidative, heat, and, to a lesser extent, starvation stress tolerance, and for adaptive heat stress response, with increased tolerance to heat, acid and detergent, different growth phases and adaptation patterns were found. Principal component analysis showed that while heat, acid and detergent stresses respond similarly to growth phase and adaptation, tolerance to oxidative and starvation stresses implies completely unrelated mechanisms. A dendrogram obtained using the data from multilocus restriction typing (MLRT) of stress response genes clearly separated two groups of L. plantarum strains from the other species but there was no correlation between genotypic grouping and grouping obtained on the basis of the stress response pattern, nor with the phylograms obtained from hrcA and ctsR sequences. Differences in sequence in L. plantarum strains were mostly due to single nucleotide polymorphisms with a high frequency of synonymous nucleotide changes and, while hrcA was characterized by an excess of low frequency polymorphism, very low diversity was found in ctsR sequences. Sequence alignment of hrcA allowed a correct discrimination of the strains at the species level, thus confirming the relevance of stress response genes for taxonomy.     

植物乳杆菌对pH、酒精浓度和SO2浓度耐受性的研究

通过调整MRS培养基的pH、酒精浓度及SO2浓度,以菌株培养前后的菌密度差(△OD600nm)为指标研究了6株植物乳杆菌(Lactobacillus plantarum)对低pH、高SO2、酒精浓度及三者相互作用的耐受性.结果表明:试验所选取的6株菌都可耐受pH 3.0、75mg/LSO2浓度,其中有5株菌可耐受16％vol酒精浓度,3个因素对菌株生长影响的次序是SO2浓度＞pH＞酒精浓度.     

High Pressure Inactivation of Lactobacillus plantarum in a Model Beer System

ABSTRACT: The effects of hop extract and ethanol on growth, high pressure inactivation, and survival of Lactobacillus plantarum were determined in model beer. Corresponding to the beer spoiling ability of this strain, levels of hops and ethanol typical for beer did not inhibit growth. Pressure death time curves determined at 300 MPa were described by an empirical model taking into account the sigmoid shape of survivor curves, sublethal injury, and the presence of baroresistant cells. Ethanol (5 and 10%) enhanced pressure effects on L. plantarum whereas hop extract (50 and 100 ppm) was less effective. In contrast, hop extracts killed pressurized cells during subsequent storage in beer but ethanol did not.     

Inactivation of Salmonella typhimurium and Lactobacillus plantarum by UV-C light in flour powder

This research evaluates the potential use of ultraviolet C light (UV-C) as a decontamination method for powdered foods, particularly of refined flour. This technology's lethal effectiveness was evaluated on Salmonella enterica subsp. Enterica serotype Typhimurium and Lactobacillus plantarum in wheat flour, and in laboratory liquid media of different a_w and turbidities to evaluate the action mechanisms of UV-C light in powdered products. Initial results showed a large variability of lethality in flour, obtaining between 0.2 and 3.0 log₁₀ cycles of inactivation. Results obtained in laboratory media and SEM analysis of contaminated flour indicated that the variability was due to a shadow effect on the efficacy of UV-C light and not due to the low water a_w of the flour or starch content. Based on these conclusions, a 2-m vertical tunnel with twelve 480 W UV-C lamps was designed to treat flour by forming a continuous cloud of dust (0.05–2.4 kg/h). Inactivation levels of 4.0 to 1.7 log₁₀ cycles of the population of L. plantarum in flour were achieved at flow rates of 0.2 and 2.4 kg/h respectively, with a maximum residence time of 4 s.Industrial relevanceThis investigation demonstrated the lethal efficacy of the application of UV–C light to inactivate microorganisms, both pathogenic and spoilage, present in flour. 4-log10 cycles of inactivation of both Salmonella Typhimurium and Lactobacillus plantarum were inactivated with UV-C treatments. A UV-C facility was built up which enabled to treat flour in continuous conditions creating a cloud of dust with treatments of 4 s and lethalities of 4-log10 reductions.     

(-)-3t,4t-Dihydroxycyclohexane-1c-carboxylate, a new quinate metabolite of Lactobacillus plantarum

Lactobacillus plantarum, isolate No. 13a, reduces (?)-quinate, shikimate and (?)-dihydroshikimate to a new metabolite identified as (?)-3^t, 4^t-dihydroxycyclohexane 1^c-carboxylate. The absolute stereochemistry of the product is given and is shown to resemble that of (?)-quinic and (?)-dihydroshikimic acids.     

Effect of inulin on growth and bacteriocin production by Lactobacillus plantarum in stationary and shaken cultures

The prebiotic effect of inulin added to MRS medium on growth and bacteriocin production by L. plantarum ST16 Pa was investigated in stationary cultures in anaerobic jars with medium containing 0.025% sodium thioglycolate or in flasks shaken at 100 rpm. In the presence of 1% inulin in anaerobic stationary cultures, this strain produced lactic acid at a level that was 36.5% higher than in the absence of the polysaccharide. In shaken cultures without inulin, cell count was 54% higher than in the stationary ones. Under stationary conditions in anaerobic jars, the addition of inulin increased the maximum specific growth rate from 0.37 to 0.49 h^?1 and reduced the generation time from 1.85 h to 1.40 h. Consequently, the exponential phase was shortened from 12 to 9 h when the cells were grown in stationary cultures with the oxygen scavenger. Despite this effect of inulin on growth rate, stationary cultures without inulin displayed higher antimicrobial activity against Listeria monocytogenes L104 (3200 AU/mL) than cultures with inulin (1600 AU/mL); therefore, inulin behaved as a compound able to accelerate growth rather than to stimulate bacteriocin production. The results presented in this study are very promising, as L. monocytogenes is a well‐known foodborne pathogenic microorganism. Moreover, L. plantarum ST16 Pa has proven to be a potential producer of a natural food preservative at an industrial level.     

Metabolism of ferulic acid during growth of Lactobacillus plantarum and Lactobacillus collinoides

BACKGROUND: Food‐isolated lactic acid bacteria can transform ferulic acid (FA) into several products. Since quantification of these metabolites during the different bacterial growth phases is lacking, the aim of this study was to identify and quantify conversion products of FA and to follow the kinetics of FA metabolism during growth of Lactobacillus plantarum and Lactobacillus collinoides. RESULTS: Lactobacillus plantarum and Lactobacillus collinoides were incubated in MRS broth, to which different amounts of FA were added (final concentrations of 0, 0.5, 1.5 and 3 mmol L^?1), at 30 °C until the late stationary phase. Lactobacillus plantarum metabolised FA into 4‐vinylguaiacol (4‐VG) and hydroferulic acid (HFA). Conversion to 4‐VG started simultaneously with the degradation of FA, while formation of HFA started in the mid‐exponential phase. Lactobacillus collinoides only formed 4‐VG, mainly in the stationary phase. No significant effect of the different amounts of FA was seen on the growth and fermentation characteristics of both bacteria. CONCLUSION: The results demonstrate that both bacteria are able to convert FA. However, start of conversion differs between the two strains. The different amounts of FA had no influence on the growth and fermentation characteristics of both bacteria. Copyright © 2012 Society of Chemical Industry     

Inactivation of Saccharomyces cerevisiae and Lactobacillus plantarum in orange juice using ultra high-pressure homogenisation

Yeasts and lactic acid bacteria are the usual contaminants in orange juice and responsible for decreasing the shelf life of the product. Ultra high-pressure homogenisation has been shown to be an alternative to the traditional thermal pasteurisation of pumpable foods. The product was pumped through a homogeniser valve at 100 MPa, 150 MPa, 200 MPa, 250 MPa and 300 MPa using two synchronized overlapped intensifiers at a flow rate of approximately 270 mL/min. The inlet temperature was kept at 10 °C, pH at 4.1 and soluble solids at 10.0 °Bx. After processing, the product was immediately cooled to 4 °C and the microbiological count was determined. The study showed that Lactobacillus plantarum and Saccharomyces cerevisiae are sensible to ultra high-pressure homogenisation treatment. The results indicated that pressures higher than 250 MPa were able to completely destroy initial loads of 1.2 × 107 UFC/mL of L. plantarum and 2.9 × 105 UFC/mL of S. cerevisiae in orange juice, making this technology a promising way to nonthermally process orange juices.Industrial relevanceThis paper deals with inactivation of microorganism contaminants of orange juice using dynamic ultra high process technology. It is of industrial interest and relevance to evaluate the use of this non-thermal emerging technology to process fluid foods that may result into better taste, optimum product functionality, safety and quality characteristics.     

Cultural conditions and nutritional components affecting the growth and bacteriocin production of Lactobacillus plantarum KC21

This study was conducted to investigate the effect of cultural conditions and nutritional components on the cell growth and bacteriocin production of Lactobacillus plantarum KC21. In cultures without pH control, the bacteriocin activity of L. plantarum KC21 was higher at 30oC, however, the cell growth rate was higher at 37°C. In MRS broth with an initial pH 6.0, the cell growth was lower, but high bacteriocin levels were recorded than at pH 7.0. The bacteriocin activity was maximal in medium containing 1.0 or 1.5% glucose or 1.0% lactose. Yeast extract (0.25 or 0.5%) added to MRS broth increased the bacteriocin activity, moreover, the bacteriocin in the presence of 1.0 or 3.0% NaCl, 0.5% NH₄PO₄, or 0.25 or 0.5% KH₂PO₄ resulted in the activity of 12,800 BU/mL, but excessively high salts concentration hindered the cell growth and bacteriocin production significantly. Besides 1.0 or 2.0 mM MgSO₄ highly increased the bacteriocin activity without affecting the growth of L. plantarum KC21, and ascorbic acid (1.0 or 3.0 ppm) enhanced the bacteriocin activity up to 2 fold.     

生物膜形成对植物乳杆菌PG3-1耐受性及功能基因表达的影响

研究了植物乳杆菌PG3-1生物膜形成后对其耐受性以及功能基因表达的影响。采用平板计数及实时荧光定量PCR对其耐受性以及功能基因相对表达量进行测定。结果表明,植物乳杆菌生物膜形成后对温度、酸碱、盐度耐受性均有不同程度的提高。植物乳杆菌生物膜形成过程中发生热应激、酸应激、盐应激等应激行为,Gro EL、Dan K热应激基因的相对表达量达到3.22和1.53。     

Conservation characteristics of corn ears and stover ensiled with the addition of Lactobacillus plantarum MTD-1, Lactobacillus plantarum 30114, or Lactobacillus buchneri 11A44

The aim of this study was to investigate the effects of inoculating 3 contrasting lactic acid bacteria on the fermentation profile, estimated nutritive value, and aerobic stability of corn ears and stover produced under marginal growing conditions. Ears and stover were separated from whole-crop corn plants obtained from 3 replicate field blocks. Representative subsamples were precision chopped and allocated to 1 of the following treatments: an uninoculated control, Lactobacillus plantarum MTD-1 (LP1), L. plantarum 30114 (LP2), or Lactobacillus buchneri 11A44 (LB). Each bacterial additive was applied at a rate of 1 × 10(6) cfu/g of fresh herbage. Triplicate samples of each treatment were ensiled in laboratory silos at 15°C for 3, 10, 35, or 130 d. No difference was observed between the dry matter recoveries of uninoculated ear or stover silages and silages made with LP1, and the aerobic stability of uninoculated ear and stover silages did not differ from silages made with LB. Stover silages made with LP2 and ensiled for 35 d had a lower proportion of lactic acid in total fermentation products compared with LP1. The aerobic stability and dry matter recovery of ear and stover silages in this study were not improved when made with LB, LP1, or LP2, due to the indigenous highly heterolactic fermentation that prevailed in the uninoculated ear and stover during 130-d ensilage.     

Combined Carbon Dioxide and High Pressure Inactivation of Pectin Methylesterase, Polyphenol Oxidase, Lactobacillus plantarum and Escherichia coli

ABSTRACT: High pressure processing (HPP) and CO₂have both been shown to increase food product shelf-life. CO₂ was added at approximately 0.2 molar % to solutions processed at 500 to 800 MPa in order to further inactivate pectin methylesterase (PME), polyphenol oxidase (PPO), L. plantarum ATCC 8014, and E. coli K12. An interaction was found between CO₂ and pressure at 25 °C and 50 °C for PME and PPO, respectively. Activity of PPO was decreased by CO₂ at all pressure treatments. The interaction between CO₂ and pressure was significant for L. plantarum with a significant decrease in survivors due to the addition of CO₂ at all pressures studied. No significant effect on E. coli survivors was seen with CO₂ addition.     

Use of bacteriocin-producing starter cultures of Lactobacillus plantarum and Lactobacillus curvatus in production of ostrich meat salami

Ostrich meat salami was produced by using Lactobacillus plantarum strain 423 and Lactobacillus curvatus strain DF126. The strains produce the bacteriocins plantaricin 423 and curvacin DF126, respectively. The specific activity of plantaricin 423 in MRS broth at 30?°C increased as the pH decreased from 6.5 to 3.5, but activity subsequently decreased. The activity of curvacin DF126 increased under the same conditions, but remained stable for the duration of the growth cycle. Maximum curvacin DF126 and plantaricin 423 activity levels were recorded at a culture pH of around 4. The spectra of antimicrobial activity recorded for plantaricin 423 and curvacin DF126 were similar. Neither of the two bacteriocins inhibited the growth of Micrococcus sp. MC50 and did not have any inhibitory effect on either of the producer strains. Curvacin DF126 and plantaricin 423 inhibited the growth of L. monocytogenes in salami meat. However, after 15 h of fermentation the viable count of L. monocytogenes LM1 increased, probably due to a decrease in activity of the bacteriocins and/or the development of resistant bacterial cells. This is the first report on the inhibition of L. monocytogenes in ostrich meat salami by using bacteriocinogenic starter cultures.     

低聚半乳糖对植物乳杆菌生长及部分代谢的影响

以一株能产细菌素的植物乳杆菌KLDS1.0391为研究对象,将MRS培养基中的葡萄糖用等量的低聚半乳糖替代,采用体外发酵方式,研究37℃培养36 h过程中活菌数、p H和滴定酸度的变化,同时分别测定培养28 h和24 h时有机酸和细菌素的产量。结果表明:随培养时间的延长,培养物的p H先迅速下降,至12 h后下降速度逐步平稳,以低聚半乳糖作为碳源时促生长作用显著高于葡萄糖(p<0.05),且菌体衰退较为缓慢。而且该菌株代谢低聚半乳糖时所产生的有机酸及抑菌活性均更高,其中有机酸主要为乳酸、乙酸和丙酸。