Characterization of a new organization of the plantaricin locus in the inducible bacteriocin-producing<Emphasis Type="Italic"> Lactobacillus plantarum</Emphasis> J23 of grape must origin

Lactobacillus plantarum J23 was previously characterized as a bacteriocin-producer-strain when it was cocultured with other lactic acid bacteria. In this work, the genetic organization of the pln locus in the J23 strain was studied and compared with those of previously described L. plantarum C11, WCFS1 and NC8 strains. A new organization of the plantaricin locus was detected in the J23 strain. The sequenced fragment (20,266 bp) comprised plnJLR, plnMNOP, plnEFI, plnGHSTUVWXY, and plNC8IF-plNC8HK-plnD operons, as well as a new region that includes three new orfs (GenBank accession number DQ323671). When the J23 pln gene sequences were compared with those included in the GenBank database, the identity of the putative encoded proteins was in the range 67.1–100%. The regulatory system and the repertoire of putative bacteriocins of the J23 pln locus presented important differences with respect to the ones of C11, WCFS1 and NC8, such as the absence of plnK and the presence of a larger plnJ gene than the previously described for the other L. plantarum strains. The pln locus in L. plantarum strains seems to be a mosaic-like structure with different modules and reorganizations that presents highly conserved regions related to transport and bacteriocin maturation and variable regions related to regulation and bacteriocin production.     

Production of a Class IIb Bacteriocin with Broad-spectrum Antimicrobial Activity in Lactiplantibacillus plantarum RUB1

Bacteriocins produced by lactic acid bacteria have potential use as natural food preservatives, which may alleviate current problems associated with the overuse of antibiotics and emerging multi-drug-resistant microbes. In this work, Lactiplantibacillus plantarum RUB1 was found to produce a class IIb bacteriocin with strong antibacterial activity. Except for plnXY encoding putative proteins, L. plantarum RUB1 contains most genes in five operons (plnABCD, plnGHSTUVW, plnMNOP, plnIEF, and plnRLJK) related to bacteriocin synthesis. Adding low (100 and 500 ng/mL) and medium (1 μg/mL) concentrations of PlnA to broth promoted bacteriocin production and upregulated bacteriocin gene plnA, while high concentrations (50 and 200 μg/mL) inhibited expression of these genes. Co-culturing L. plantarum RUB1 with Enterococcus hirae 1003, Enterococcus hirae LWS, Limosilactobacillus fermentum RC4, L. plantarum B6, and even Listeria monocytogenes ATCC 19111 and Staphylococcus aureus ATCC 6538 enhanced bacteriocin activity and expression of bacteriocin-related genes. This study verifies that PlnA can indeed upregulate the expression of bacteriocin genes, and also bacteriocin production can be induced by co-culture with some specific bacteria or their cell-free supernatants. Bacteriocin production by L. plantarum RUB1 is mediated by a quorum sensing mechanism, directly influenced by autoinducing peptide or specific strains. The findings provide new methods and insight into bacteriocin production mechanisms.

     

Purification and genetic characterization of plantaricin NC8, a novel coculture-inducible two-peptide bacteriocin from Lactobacillus plantarum NC8

A new, coculture-inducible two-peptide bacteriocin named plantaricin NC8 (PLNC8) was isolated from Lactobacillus plantarum NC8 cultures which had been induced with Lactococcus lactis MG1363 or Pediococcus pentosaceus FBB63. This bacteriocin consists of two distinct peptides, named alpha and beta, which were separated by C(2)-C(18) reverse-phase chromatography and whose complementary action is necessary for full plantaricin NC8 activity. N-terminal sequencing of both purified peptides showed 28 and 34 amino acids residues for PLNC8 alpha and PLNC8 beta, respectively, which showed no sequence similarity to other known bacteriocins. Mass spectrometry analysis showed molecular masses of 3,587 Da (alpha) and 4,000 Da (beta). The corresponding genes, designated plNC8A and plNC8B, were sequenced, and their nucleotide sequences revealed that both peptides are produced as bacteriocin precursors of 47 and 55 amino acids, respectively, which include N-terminal leader sequences of the double-glycine type. The mature alpha and beta peptides contain 29 and 34 amino acids, respectively. An open reading frame, orfC, which encodes a putative immunity protein was found downstream of plNC8B and overlapping plNC8A. Upstream of the putative -35 region of plNC8B, two direct repeats of 9 bp were identified, which agrees with the consensus sequence and structure of promoters of class II bacteriocin operons whose expression is dependent on an autoinduction mechanism.     

The cost and benefit of quorum sensing‐controlled bacteriocin production in Lactobacillus plantarum

Bacteria eliminate competitors via ‘chemical warfare’ with bacteriocins. Some species appear to adjust bacteriocin production conditionally in response to the social environment. We tested whether variation in the cost and benefit of producing bacteriocins could explain such conditional behaviour, in the bacteria Lactobacillus plantarum. We found that: (a) bacterial bacteriocin production could be upregulated by either the addition of a synthetic autoinducer peptide (PLNC8IF; signalling molecule), or by a plasmid which constitutively encodes for the production of this peptide; (b) bacteriocin production is costly, leading to reduced growth when grown in poor and, to a lesser extent, in rich media; (c) bacteriocin production provides a fitness advantage, when grown in competition with sensitive strains; and (d) the fitness benefits provided by bacteriocin production are greater at higher cell densities. These results show how the costs and benefits of upregulating bacteriocin production can depend upon abiotic and biotic conditions.     

Characterization of the three selected probiotic strains for the application in food industry

Previously selected bacterial probiotic strains Enterococcus faecium L3, Lactobacillus plantarum L4 and Lactobacillus acidophilus M92 have shown their potential as functional starter cultures in silage, white cabbage and milk fermentation. Therefore, the phenotypic and genotypic characteristics important for their application in food industry were investigated. Pulsed-field gel electrophoresis (PFGE) of NotI digested genomic DNA, in combination with physiological traits determined by API tests, made a useful tool for identification of these probiotic strains and differentiation among them. Lyophilized probiotic cells remained viable during 75 days of storage at −20, +4 and +15°C, while fresh concentrated cells remained viable only at −20°C with addition of glycerol as cryoprotectant. After the lyophilization with addition of skim milk as lyoprotectant, the viability of L. acidophilus M92, L. plantarum L4 and E. faecium L3 was reduced by only 0.37, 0.44 and 0.50 log, respectively. Furthermore, probiotic strains L. acidophilus M92, L. plantarum L4, and E. faecium L3, demonstrated anti-Salmonella activity, and L. acidophilus M92 having also antilisterial activity demonstrated by in vitro competition test. Overnight cultures and cell-free supernatants of the three probiotic strains exerted also an antagonistic effect against the Gram-positive and Gram-negative test microorganisms examined, demonstrated by the agar-well diffusion test. The inhibition of Listeria monocytogenes, Salmonella typhimurium, Yersinia enterocolitica, and Acinetobacter calcoaceticus obtained, achieved by the neutralized, 5-fold concentrated supernatant of L. plantarum L4, may be the result of its bacteriocinogenic activity. On the basis of these results, the application of the three examined probiotic strains may become a point of great importance in respect of food safety.     

Quorum-sensing based bacteriocin production is down-regulated by N-terminally truncated species of gene activators

Down-regulation of quorum-sensing based pathways is an important but yet poorly understood process in bacterial gene regulation. In this study, we show that the gene regulator plnC not only acts as an activator gene in the quorum-sensing based bacteriocin production in Lactobacillus plantarum C11, but it also concurrently codes for truncated forms that were shown to repress bacteriocin production. By amino acid N-terminal sequencing and DNA sequence analysis, the truncated species of PlnC are believed to be translated from alternative start codons located in the so-called receiver domain of the regulator. To analyse the structure–function relationship of truncated species of PlnC, we performed a series of systematic truncation mutations: ten in the receiver domain, one in the hinge region and two in the C-terminal DNA-binding domain. It was revealed that any truncation mutation containing a disrupted receiver domain together with an intact DNA-binding domain displayed a repressive effect on bacteriocin production. Such a gene repression mechanism mediated by truncated regulators was also found in two other quorum-sensing based bacteriocin systems (spp in L. sakei LTH673 and NC8-pln in L. plantarum NC8), suggesting that this mode of repression might represent a common means applied by bacteria to down-regulate certain quorum-sensing based pathways. Electronic supplementary material The online version of this article (doi:) contains supplementary material, which is available to authorized users.     

Heterologous Expression of Oxalate Decarboxylase in <Emphasis Type="Italic">Lactobacillus plantarum</Emphasis> NC8

Lactic acid bacteria (LABs) are being used as a probiotic very often for various enteric problems. Many genetically modified LABs are created by different workers for various novel applications. In this study we examine the expression of heterologous oxalate decarboxylase (oxdc) in Lactobacillus plantarum NC8. Generally, this enzyme is not present in Lactobacillus spp. Oxdc gene from Bacillus subtilis was polymerase chain reaction-amplified and cloned in a shuttle vector pSIP400 series, downstream of the inducible promoter, P_orfx. In the presence of an inducing peptide, Sakacin-P, the expression of OxdC was observed in sodium dodecyl sulfate–polyacrylamide gel electrophoresis. The cell-free extract and the purified protein from the recombinant LABs showed the presence of OxdC activity. The above recombinant LABs, with desired modifications, can be used as a possible probiotic for the degradation of intestinal dietary oxalate for preventing enteric hyperoxaluria.     

Lactonase-expressing Lactobacillus plantarum NC8 attenuates the virulence factors of multiple drug resistant Pseudomonas aeruginosa in co-culturing environment

Pseudomonas aeruginosa possesses an arcade of both cell-associated and extracellular cytotoxic virulence factors which are regulated by a multi-component quorum sensing system. Many research studies report success of lactonase in combating the pathogenicity of P. aeruginosa but delivery of lactonase remains a challenge. The present study aims at developing a delivery vehicle for lactonase. Lactobacillus plantarum NC8 was used as host for aiiA (Bacillus thuringiensis 4A3 lactonase gene) using pSIP409 expression vector. pSIP409: aiiA construct was stably maintained in L. plantarum NC8. Co-culturing of multi-drug resistant (MDR) clinical isolates of P. aeruginosa and PAO1 with recombinant L. plantarum NC8 led to significant reduction (p < 0.001) in extracellular virulence factors like pyocyanin, protease, elastase and rhamnolipids in P. aeruginosa and also showed significant reduction in adhesion of P. aeruginosa strains to uroepithelial cells in vitro. This study shows the heterologous expression of AiiA lactonase in L. plantarum NC8. Co-culturing of lactonase expressing L. plantarum NC8 with MDR P. aeruginosa strains led to attenuation of their virulence significantly. These results underscore the potential application of recombinant L. plantarum NC8 with anti-quorum sensing properties to control infections caused by multidrug resistant P. aeruginosa.     

Quorum-Sensing Regulation of Constitutive Plantaricin by Lactobacillus plantarum Strains under a Model System for Vegetables and Fruits

This study aimed at investigating the regulatory system of bacteriocin synthesis by Lactobacillus plantarum strains in vegetables and fruits in a model system. Sterile and neutralized cell-free supernatant (CFS) from L. plantarum strains grown in MRS broth showed in vitro antimicrobial activities toward various indicator strains. The highest activity was that of L. plantarum C2. The antimicrobial activity was further assayed on vegetable and fruit agar plates (solid conditions) and in juices (liquid conditions). A regulatory mechanism of bacteriocin synthesis via quorum sensing was hypothesized. The synthesis of antimicrobial compounds seemed to be constitutive under solid conditions of growth on vegetable and fruit agar plates. In contrast, it depended on the size of the inoculum when L. plantarum C2 was grown in carrot juice. Only the inoculum of ca. 9.0 log CFU ml⁻¹ produced detectable activity. The genes plnA, plnEF, plnG, and plnH were found in all L. plantarum strains. The genes plnJK and plnN were detected in only three or four strains. Reverse-phase high-performance liquid chromatography purification and mass spectrometry analysis revealed the presence of a mixture of eight peptides in the most active fraction of the CFS from L. plantarum C2. Active peptides were encrypted into bacteriocin precursors, such as plantaricins PlnJ/K and PlnH and PlnG, which are involved in the ABC transport system. A real-time PCR assay showed an increase in the expression of plnJK and plnG during growth of L. plantarum C2 in carrot juice.     

Identification and functional properties of dominant lactic acid bacteria isolated at different stages of solid state fermentation of cassava during traditional <Emphasis Type="Italic">gari</Emphasis> production

Culture-based technique was used to study the population dynamics of the bacteria and determine the dominant lactic acid bacteria (LAB) during cassava fermentation. LAB was consistently isolated from the fermented mash with an initial viable count of 6.00 log c.f.u. g⁻¹ observed at 12 h. The aerobic viable count of amylolytic lactic acid bacteria (ALAB) was higher than other group of LAB throughout the fermentation up to 96 h with the highest viable count of 8.08 log c.f.u. g⁻¹. Combination of phenotypic parameters and 16S rDNA gene sequencing identified the dominant group of LAB as Lactobacillus plantarum, L. fermentum and Leuconostoc mesenteroides while the pulse field gel electrophoresis determined that the strains were genotypically heterogeneous. The sugar fermentation profile of the isolates showed that indigestible sugars such as raffinose and stachyose can be fermented by the strains. Information was also generated about the functional properties of the strains. Only strain L. plantarum 9st0 isolate at 0 h of the fermentation produced bacteriocin with antagonism against closely related indicator strains. Quantitatively, the highest amylase activity was produced by strain L. plantarum 7st12, while appreciable amylase was also produced by L. fermentum 1st96. The result of this work showed that selection of mixed starter cultures of bacteriocin- and amylase-producing L. plantarum and L. fermentum will be highly relevant as starter cultures during the intermediate and large scale gari production.     

Characterization of lactic acid bacteria isolated from sourdoughs for Cornetto, a traditional bread produced in Basilicata (Southern Italy)

A total of 41 strains of lactic acid bacteria (LAB) isolated from durum wheat sourdoughs used to produce Cornetto di Matera bread, were identified by SDS-PAGE of whole cell proteins (WCP) and screened for acid production ability, antimicrobial activity and exopolysaccharide (EPS) production. The isolates were identified as Lactobacillus plantarum (49%), Leuconostoc mesenteroides (17%), Lactobacillus curvatus (15%), Lactobacillus paraplantarum (12%), Weissella cibaria (5%) and Lactobacillus pentosus (2%). Several strains of Lb. plantarum and Leuc. mesenteroides showed a high acid production ability. The antagonistic activity was tested using an agar-spot deferred antagonism assay against a set of five indicators. The species had different profiles of inhibition. Lb. plantarum had the largest spectrum of inhibition, while no isolates of W. cibaria and Leuc. mesenteroides showed antimicrobial activity. No strains had antimicrobial activity against Bacillus cereus. The inhibitory activity of five strains was confirmed to be sensitive to proteolytic enzymes and thus potentially due to bacteriocin production. All Leuc. mesenteroides and W. cibaria strains produced EPS from sucrose. Some Lb. plantarum and Lb. paraplantarum strains produced EPS from different sugars in solid media. EPS production in liquid media was different within the species, with the highest production in liquid media containing glucose and maltose. A defined strain starter culture (W. cibaria DBPZ1006, Lb. plantarum DBPZ1015 and S. cerevisiae MTG10) was selected on the basis of technological properties and tested in model sourdough fermentations.     

Genomic Analysis for Antioxidant Property of Lactobacillus plantarum FLPL05 from Chinese Longevity People

Antioxidant activity is one of the important probiotic characteristics for lactic acid bacteria including Lactobacillus plantarum, which is used for food fermentation or as a probiotic supplement. L. plantarum FLPL05 is a novel strain originally isolated from a healthy elderly individual of longevity. The organism has been demonstrated to exhibit high antioxidant property. However, there are limited genomic insights into the antioxidant properties of this organism. In this study, we performed whole-genome analysis regarding its antioxidant property. L. plantarum FLPL05 exhibited higher antioxidant activity compared with that of L. plantarum strains ATCC14917, ATCC8014, and WCFS1. The antioxidant capacity of L. plantarum FLPL05 was genetically linked to its antioxidant system, i.e., glutathione and thioredoxin involved in global regulation of defense against hydrogen peroxide challenge. L. plantarum FLPL05 was further examined for its antioxidant potential in d-Gal-induced aging mice and exhibited a significant increase in the activity of serum glutathione peroxidase (GSH-PX) and a decrease in the level of malondialdehyde (MDA). Moreover, our analyses exhibited a complete gene cluster including plnA, plnB, plnC, plnD, plnE, plnF, plnG, plnH, plnI, plnJ, plnK, plnM, plnN, plnO, plnP, plnQ, plnST, plnU, plnV, plnW, plnX, and plnY for production of bacteriocin. Our results suggest that L. plantarum FLPL05 could be a probiotic candidate.

     

Characterization of mesentericin ST99, a bacteriocin produced by <Emphasis Type="Italic">Leuconostoc mesenteroides</Emphasis> subsp. <Emphasis Type="Italic">dextranicum</Emphasis> ST99 isolated from boza

Lactic acid bacteria isolated from Boza, a cereal-fermented beverage from Belogratchik, Bulgaria, were screened for the production of bacteriocins. With the first screening, 13 of the 52 isolates inhibited the growth of Listeria innocua and Lactobacillus plantarum. The cell-free supernatant of one of these strains, classified as Leuconostoc mesenteroides subsp. dextranicum ST99, inhibited the growth of Bacillus subtilis, Enterococcus faecalis, several Lactobacillus spp., Lactococcus lactis subsp. cremoris, Listeria innocua, Listeria monocytogenes, Pediococcus pentosaceus, Staphylococcus aureus and Streptococcus thermophilus. Clostridium spp., Carnobacterium spp., L. mesenteroides and Gram-negative bacteria were not inhibited. Maximum antimicrobial activity, i.e. 6,400 arbitrary units (AU)/ml, was recorded in MRS broth after 24 h at 30°C. Incubation in the presence of protease IV and pronase E resulted in loss of antimicrobial activity, confirming that growth inhibition was caused by a bacteriocin, designated here as mesentericin ST99. No loss in activity was recorded after treatment with -amylase, SDS, Tween 20, Tween 80, urea, Triton X-100, N-laurylsarcosin, EDTA and phenylmethylsulfonylfluoride. Mesentericin ST99 remained active after 30 min at 121°C and after 2 h of incubation at pH 2 to 12. Metabolically active cells of L. innocua treated with mesentericin ST99 did not undergo lysis. Mesentericin ST99 did not adhere to the cell surface of strain ST99. Precipitation with ammonium sulfate (70% saturation), followed by Sep-Pack C₁₈ chromatography and reverse-phase HPLC on a C₁₈ Nucleosil column yielded one antimicrobial peptide.     

Interference of <Emphasis Type="Italic">Lactobacillus plantarum</Emphasis> Strains in the In Vitro Conjugative Transfer of R-Plasmids

Probiotic compounds, which are often constituted of lactobacilli, exert a number of health benefits through maintenance of the intestinal ecosystem balance. Among the important interactions that occur in the gut microbiota, plasmid transfer by mating is an increasing cause of concern, particularly when antibiotic-resistant genes are involved. Because lactobacilli seem to be able to influence this mechanism, the aim of the present work was to investigate the in vitro capability of two Lactobacillus plantarum strains (one bacteriocin producer and one nonproducer) to interfere with the conjugation processes. For this purpose different matings were performed adding to the donor and recipient cells L. plantarum 35d bac+ and L. plantarum 396/1 bac– as agents of interference. Conjugations added with a Staphylococcus aureus strain or without any agent of interference were used as controls. The results of our experiments demonstrated that both lactobacillus strains were able to decrease mating frequency. Statistically significant differences in the viable transconjugants were obtained in the presence and in the absence of the lactobacilli. The effect was almost the same with the two L. plantarum independent of bacteriocin production. In the trial performed with S. aureus, no decrease in mating frequency was observed, confirming that the capability to interfere with R-plasmid transfer ability could be a property of the tested L. plantarum strains.     

Mathematical evaluation of plantaricin formation supports an auto-induced production mechanism

The production rate of a bacteriocin, produced by Lactobacillus plantarum TMW1.25 and previously named plantaricin1.25, was studied during pH-constant batch fermentations under various growth media conditions. The growth of L. plantarum and production of bacteriocin during the retardation phase were modelled, using 11 different empirical and mechanistic approaches. The optimal pH for bacteriocin production was 4.5. Among the different nitrogen sources tested, yeast extract was the most important, on the basis of the fact that the maximum growth rate decreased 16% without yeast extract, and only 7.2% or 8.1% without meat extract or peptone respectively. However, the change of nitrogen source did not have a significant effect on bacteriocin production. The progression of plantaricin1.25 production during the retardation phase and growth of L. plantarum TMW1.25 could be described by a structured model in which the bacteriocin concentration induces its own production. Among those models not implementing bacteriocin induction, only the one with an exponential increase of bacteriocin yield per unit biomass was suitable to describe bacteriocin production. Computer-aided evaluation of experimental data appears to be helpful in elucidating the relationship between the growth of lactic acid bacteria and bacteriocin production. Received: 22 May 1998 / Received last revision: 9 November 1998 / Accepted: 14 November 1998     

Medium components effecting bacteriocin production by two strains of Lactobacillus plantarum ST414BZ and ST664BZ isolated from boza

Bacteriocins ST414BZ and ST664BZ, produced by Lactobacillus plantarum, inhibited the growth of a number of lactic acid bacteria, Escherichia coli, Pseudomonas aeruginosa, Klebsiella pneumoniae and Enterobacter cloacae. Optimal production of bacteriocin ST664BZ (12 800 AU/mL) was recorded in MRS broth with an initial pH of 6.0 and 6.5. Bacteriocin ST414BZ was produced in MRS broth at lower pH values, ranging from 6.5 to 5.0. Low levels of bacteriocin activity were produced in BHI, M17, 10% (w/v) soy flour and 10% (w/v) molasses, suggesting that specific nutrients are required for optimal production. Bacteriocin ST414BZ production doubled (from 12 800 to 25 600 AU/mL) in MRS broth with tryptone as sole nitrogen source, or when glucose was replaced with maltose. Bacteriocin ST664BZ production, on the other hand, was less influenced by changes in nitrogen content, but increased two-fold (to 25 600 AU/mL) when glucose was replaced with sucrose, maltose or mannose, or when MRS broth was supplemented with 2.0 g/L KH₂ PO₄. Enrichment of MRS broth with vitamins B₁₂, B₁ or C did not stimulate production of the two bacteriocins. Growth in the presence of DL-6,8-thioctic acid increased bacteriocin ST664BZ production to 25 600 AU/mL. Concluded from these results, optimal levels of bacteriocins ST414BZ and ST664BZ will be produced in boza enriched with tryptone and maltose.     

Effect of hematin on the activities of nitrite reductase and catalase in lactobacilli

The effect of the addition of hematin on the activities of nitrite reductase and catalase was studied with cell suspensions of strains of lactobacillus species. In cells of Lactobacillus plantarum grown under aerobic or anaerobic conditions nitrite reductase was present. This activity was not exhibited by cells of L. curvatus and only rarely by L. sake. In addition, catalase activity was detected in aerobically grown cells only, with all strains of L. plantarum and L. sake; again L. curvatus was devoid of this activity. Ammonia was formed as the main product of nitrite reduction by L. plantarum. With lactate as the electron donor, the end products of carbohydrate catabolism were carbon dioxide, acetoin and acetate. The activities of nitrite reductase and catalase in strains of lactobacillus species may be used for optimizing the quality of starter cultures applied for the production of raw sausages.     

Effect of bacteriocinogenic Lactobacillus spp. on the shelf life of fufu, a traditional fermented cassava product

Quantitative determination of antimicrobial compounds produced by the Lactobacillus strains under test was carried out. L. plantarum F1 produced the highest quantity of lactic acid (16.4 g/l) while the lowest amount (0.3 g/l) was produced by L. jensenii F9. All the test organisms produced hydrogen peroxide, with L. brevis OG1 having the highest yield of 0.037 g/l. Diacetyl was also produced by all the organisms, with L. plantarum F1 and L. brevis OG1 having the highest yield of 1.7 g/l, while the lowest producer was L. jensenii F9 (0.86 g/l). Determination of bacteriocin activity was carried out. L. plantarum F1 exhibited 6400 AU/ml bacteriocin activity, while L. brevis OG1 had the lowest activity of 3200 AU/ml using E. coli NCTC10418 as indicator organism. However, L. fermentum F5 and L. jensenii F9 did not produce any detectable bacteriocin. The pH value in the culture supernatant of L. plantarum F1 reached 3.1 within 48 h of incubation, while that of L. jensenii F9 was 5.2. Fufu was prepared using both bacteriocin-producing (BP) L. plantarum F1 and L. brevis OG1, and non-bacteriocin producing L. fermentum F5 and L. jensenii F9. No viable cells of Salmonella typhimurium ATCC13311 and Shigella flexneri AP23498 were detected after 12 h in the cassava products fermented with mixed starter culture of L. plantarum F1 and L. brevis OG1. The rate of survival of enteropathogens in cassava fermented with mixed starter cultures of L. plantarum F1 and L. brevis OG1 was much lower when compared to cassava fermented with mixed starter culture of L. fermentum F5 and L. jensenii F9. At 12 h, the viable count of E. coli NCTC10418 in cassava fermented with mixed starter cultures of L. plantarum F1 and L. brevis OG1 was 1.1 log₁₀ c.f.u./g whereas in cassava fermented with mixed starter cultures of L. fermentum F5 and L. jensenii F9, 8.5 log₁₀ c.f.u./g was obtained The study revealed that fufu produced with BP mixed starter cultures had a better shelf life and kept for 13 days before spoilage occurred, relative to 5 days observed for fufu produced using non-bacteriocin-producing starter cultures, and 6 days for the traditional fermented fufu.     

Energy conservation in malolactic fermentation by Lactobacillus plantarum and Lactobacillus sake

A comparably poor growth medium containing 0.1% yeast extract as sole non-defined constituent was developed which allowed good reproducible growth of lactic acid bacteria. Of seven different strains of lactic acid bacteria tested, only Lactobacillus plantarum and Lactobacillus sake were found to catalyze stoichiometric conversion of l-malate to l-lactate and CO₂ concomitant with growth. The specific growth yield of malate fermentation to lactate at pH 5.0 was 2.0 g and 3.7 g per mol with L. plantarum and L. sake, respectively. Growth in batch cultures depended linearly on the malate concentration provided. Malate was decarboxylated nearly exclusively by the cytoplasmically localized malo-lactic enzyme. No other C₄-dicarboxylic acid-decarboxylating enzyme activity could be detected at significant activity in cell-free extracts. In pH-controlled continuous cultures, L. plantarum grew well with glucose as substrate, but not with malate. Addition of lactate to continuous cultures metabolizing glucose or malate decreased cell yields significantly. These results indicate that malo-lactic fermentation by these bacteria can be coupled with energy conservation, and that membrane energetization and ATP synthesis through this metabolic activity are due to malate uptake and/or lactate excretion rather than to an ion-translocating decarboxylase enzyme.