Background The raw goat milk microbiota is considered a good source of novel bacteriocinogenic lactic acid bacteria (LAB) strains that can be exploited as an alternative for use as biopreservatives in foods. The constant demand for such alternative tools justifies studies that investigate the antimicrobial potential of such strains. Results The obtained data identified a predominance of Lactococcus and Enterococcus strains in raw goat milk microbiota with antimicrobial activity against Listeria monocytogenes ATCC 7644. Enzymatic assays confirmed the bacteriocinogenic nature of the antimicrobial substances produced by the isolated strains, and PCR reactions detected a variety of bacteriocin-related genes in their genomes. Rep-PCR identified broad genetic variability among the Enterococcus isolates, and close relations between the Lactococcus strains. The sequencing of PCR products from nis-positive Lactococcus allowed the identification of a predicted nisin variant not previously described and possessing a wide inhibitory spectrum. Conclusions Raw goat milk was confirmed as a good source of novel bacteriocinogenic LAB strains, having identified Lactococcus isolates possessing variations in their genomes that suggest the production of a nisin variant not yet described and with potential for use as biopreservatives in food due to its broad spectrum of action. PMID:24521354
Luiz, L M P; Chuat, V; Madec, M N; Araújo, E A; de Carvalho, A F; Valence, F
The milk produced in regions with different traditions in Brazil is used for artisanal product production, which is characterized by different sensorial characteristics. This study aimed to identify the bacterial ecosystem of farms located in a traditional dairy region in the state of Minas Gerais and to characterize Lactococcus lactis strains, the species of interest in this study, using a multilocus sequence typing (MLST) protocol and pulsed-field gel electrophoresis (PFGE) technique. Samples were collected from raw milk and dairy environment from six farms. A total of 50 isolates were analyzed using 16S rRNA sequencing and species-specific PCR. Five genera were identified: Lactobacillus, Leuconostoc, Lactococcus, Enterococcus, and Staphylococcus, from ten different species. MLST (with six housekeeping genes) and PFGE (with SmaI endonuclease) were used for the characterization of 20 isolates of Lactococcus lactis from a dairy collection in this study. Both methods revealed a high clonal diversity of strains with a higher discriminatory level for PFGE (15 pulsotypes), compared to MLST (12 ST). This study contributes to the preservation of the Brazilian dairy heritage and provides insights into a part of the LAB population found in raw milk and dairy environment.
Spoilage potential of psychrotrophic lactic acid bacteria (LAB) species: Leuconostoc gelidum subsp. gasicomitatum and Lactococcus piscium, on sweet bell pepper (SBP) simulation medium under different gas compositions.
Pothakos, Vasileios; Nyambi, Clarice; Zhang, Bao-Yu; Papastergiadis, Antonios; De Meulenaer, Bruno; Devlieghere, Frank
Sweet bell peppers are a significant constituent of retail, chilled-stored and packaged food products like fresh salads, marinades and ready-to-eat (RTE) meals. Previously, through general screening of the Belgian market and by means of source tracking analysis in a plant manufacturing minimally processed, vegetable salads the susceptibility of fresh-cut sweet bell peppers to lactic acid bacterium (LAB) contamination was substantiated. The determination of the metabolic profiles of Leuconostoc gelidum subsp. gasicomitatum and Lactococcus piscium, two major psychrotrophic, spoilage-related LAB species, on sweet bell pepper (SBP) simulation medium under different packaging conditions - 1.) vacuum: 100% N2, 2.) air: 21% O2, 79% N2, 3.) MAP1: 30% CO2, 70% N2 and 4.) MAP2: 50% O2, 50% CO2 - facilitated a better understanding of the spoilage potential of these microbes as well as the presumptive contribution of O2 in the spectrum of produced volatile organic compounds (VOCs) associated with poor organoleptic properties of food products. Generally, none of the applied gas compositions inhibited the growth of the 4 L. gelidum subsp. gasicomitatum isolates, however the presence of O2 resulted in buttery off-odors by inducing primarily the accumulation of diacetyl and pungent "vinegar" smell due to acetic acid. The 3 tested isolates of L. piscium varied greatly among their growth dynamics and inhibition at MAP2. They exhibited either weak spoilage profile or very offensive metabolism confirming significant intraspecies diversity.
Oh, Euhlim; Lu, Mingshou; Park, Changhun; Park, Changhun; Oh, Han Bin; Lee, Sang Yup; Lee, Jinwon
A dynamic model of lactic acid fermentation using Lactococcus lactis was constructed, and a metabolic flux analysis (MFA) and metabolic control analysis (MCA) were performed to reveal an intensive metabolic understanding of lactic acid bacteria (LAB). The parameter estimation was conducted with COPASI software to construct a more accurate metabolic model. The experimental data used in the parameter estimation were obtained from an LC-MS/ MS analysis and time-course simulation study. The MFA results were a reasonable explanation of the experimental data. Through the parameter estimation, the metabolic system of lactic acid bacteria can be thoroughly understood through comparisons with the original parameters. The coefficients derived from the MCA indicated that the reaction rate of L-lactate dehydrogenase was activated by fructose 1,6-bisphosphate and pyruvate, and pyruvate appeared to be a stronger activator of L-lactate dehydrogenase than fructose 1,6-bisphosphate. Additionally, pyruvate acted as an inhibitor to pyruvate kinase and the phosphotransferase system. Glucose 6-phosphate and phosphoenolpyruvate showed activation effects on pyruvate kinase. Hexose transporter was the strongest effector on the flux through L-lactate dehydrogenase. The concentration control coefficient (CCC) showed similar results to the flux control coefficient (FCC).
Wegmann, Udo; O'Connell-Motherway, Mary; Zomer, Aldert; Buist, Girbe; Shearman, Claire; Canchaya, Carlos; Ventura, Marco; Goesmann, Alexander; Gasson, Michael J.; Kuipers, Oscar P.; van Sinderen, Douwe; Kok, Jan
Lactococcus lactis is of great importance for the nutrition of hundreds of millions of people worldwide. This paper describes the genome sequence of Lactococcus lactis subsp. cremoris MG1363, the lactococcal strain most intensively studied throughout the world. The 2,529,478-bp genome contains 81 pseudogenes and encodes 2,436 proteins. Of the 530 unique proteins, 47 belong to the COG (clusters of orthologous groups) functional category “carbohydrate metabolism and transport,” by far the largest category of novel proteins in comparison with L. lactis subsp. lactis IL1403. Nearly one-fifth of the 71 insertion elements are concentrated in a specific 56-kb region. This integration hot-spot region carries genes that are typically associated with lactococcal plasmids and a repeat sequence specifically found on plasmids and in the “lateral gene transfer hot spot” in the genome of Streptococcus thermophilus. Although the parent of L. lactis MG1363 was used to demonstrate lysogeny in Lactococcus, L. lactis MG1363 carries four remnant/satellite phages and two apparently complete prophages. The availability of the L. lactis MG1363 genome sequence will reinforce its status as the prototype among lactic acid bacteria through facilitation of further applied and fundamental research. PMID:17307855
Bolotin, Alexander; Wincker, Patrick; Mauger, Stéphane; Jaillon, Olivier; Malarme, Karine; Weissenbach, Jean; Ehrlich, S. Dusko; Sorokin, Alexei
Lactococcus lactis is a nonpathogenic AT-rich gram-positive bacterium closely related to the genus Streptococcus and is the most commonly used cheese starter. It is also the best-characterized lactic acid bacterium. We sequenced the genome of the laboratory strain IL1403, using a novel two-step strategy that comprises diagnostic sequencing of the entire genome and a shotgun polishing step. The genome contains 2,365,589 base pairs and encodes 2310 proteins, including 293 protein-coding genes belonging to six prophages and 43 insertion sequence (IS) elements. Nonrandom distribution of IS elements indicates that the chromosome of the sequenced strain may be a product of recent recombination between two closely related genomes. A complete set of late competence genes is present, indicating the ability of L. lactis to undergo DNA transformation. Genomic sequence revealed new possibilities for fermentation pathways and for aerobic respiration. It also indicated a horizontal transfer of genetic information from Lactococcus to gram-negative enteric bacteria of Salmonella-Escherichia group. [The sequence data described in this paper has been submitted to the GenBank data library under accession no. AE005176.] PMID:11337471
Biscola, Vanessa; Abriouel, Hikmate; Todorov, Svetoslav Dimitrov; Capuano, Verena Sant'Anna Cabral; Gálvez, Antonio; Franco, Bernadette Dora Gombossy de Melo
Charqui is a fermented, salted and sun-dried meat product, widely consumed in Brazil and exported to several countries. Growth of microorganisms in this product is unlikely due to reduced Aw, but halophilic and halotolerant bacteria may grow and cause spoilage. Charqui is a good source of lactic acid bacteria able to produce antimicrobial bacteriocins. In this study, an autochthonous bacteriocinogenic strain (Lactococcus lactis subsp. lactis 69), isolated from charqui, was added to the meat used for charqui manufacture and evaluated for its capability to prevent the growth of spoilage bacteria during storage up to 45 days. The influence of L. lactis 69 on the bacterial diversity during the manufacturing of the product was also studied, using denaturing gradient gel electrophoresis (DGGE). L. lactis 69 did not affect the counts and diversity of lactic acid bacteria during manufacturing and storage, but influenced negatively the populations of halotolerant microorganisms, reducing the spoilage potential. The majority of tested virulence genes was absent, evidencing the safety and potential technological application of this strain as an additional hurdle to inhibit undesirable microbial growth in this and similar fermented meat products.
Short communication: Presence of Lactococcus and lactococcal exopolysaccharide operons on the leaves of Pinguicula vulgaris supports the traditional source of bacteria present in Scandinavian ropy fermented milk.
Porcellato, Davide; Tranvåg, Malena; Narvhus, Judith
Some traditional Scandinavian fermented milk products have a pronounced ropy consistency due to the presence of exopolysaccharide-producing strains of Lactococcus lactis ssp. cremoris. Norwegian food folklore describes how leaves from the carnivorous plant Pinguicula vulgaris (common butterwort) may be added to milk to initiate the fermentation of the traditional fermented milk product tettemelk. However, scientific confirmation of the link between the plant and the milk product has not been previously published. In the present study, the microbiome on 20 samples of P. vulgaris leaves collected from 5 different rural geographical locations in Norway and from 4 samples of commercial tettemelk was analyzed using high-throughput sequencing methods. The leaf microbiota of P. vulgaris was dominated by Proteobacteria and Firmicutes and the genus Lactococcus was demonstrated in all leaf samples. In addition, DNA extracted from the leaf microbiome contained genes identical to those responsible for exopolysaccharide production in Lactococcus. These results confirm the traditional use of P. vulgaris as a source of bacteria for the Norwegian ropy fermented milk product tettemelk and indicate that P. vulgaris microbiomes can be a potential source of lactic acid bacteria with interesting dairy technological features.
Saraoui, T; Leroi, F; Björkroth, J; Pilet, M F
The genus Lactococcus comprises 12 species, some known for decades and others more recently described. Lactococcus piscium, isolated in 1990 from rainbow trout, is a psychrotrophic lactic acid bacterium, probably disregarded because most of the strains are unable to grow at 30°C. During the last 10 years, this species has been isolated from a large variety of food: meat, seafood and vegetables, mostly packed under vacuum (VP) or modified atmosphere (MAP) and stored at chilled temperature. Recently, culture-independent techniques used for characterization of microbial ecosystems have highlighted the importance of Lc. piscium in food. Its role in food spoilage varies according to the strain and the food matrix. However, most studies have indicated that Lc. piscium spoils meat, whereas it does not degrade the sensory properties of seafood. Lactococcus piscium strains have a large antimicrobial spectrum, including Gram-positive and negative bacteria. In various seafoods, some strains have a protective effect against spoilage and can extend the sensory shelf-life of the products. They can also inhibit the growth of Listeria monocytogenes, by a cell-to-cell contact-dependent. This article reviews the physiological and genomic characteristics of Lc. piscium and discusses its spoilage or protective activities in food.
Pérez-Sánchez, Tania; Balcázar, José Luis; Merrifield, Daniel L; Carnevali, Oliana; Gioacchini, Giorgia; de Blas, Ignacio; Ruiz-Zarzuela, Imanol
The aim of the present study was to investigate the effect of lactic acid bacteria (LAB) on the control of lactococcosis as well as to assess the impact of probiotics on the expression of immune-related genes in the head kidney and intestine of rainbow trout (Oncorhynchus mykiss). Lactobacillus plantarum, Lactococcus lactis and Leuconostoc mesenteroides, were administered orally at 10⁶ CFU g⁻¹ feed to fish for 36 days. Twenty-one days after the start of the feeding period, fish were challenged with Lactococcus garvieae. Only the fish fed the diet containing Lb. plantarum showed significantly (P < 0.05) improved protection against L. garvieae compared to the control. Subsequently, real-time PCR was employed to determine the mRNA levels of IL-1β, IL-8, IL-10 and TNF-α in the head kidney, and IL-8, Tlr5 and IgT in the intestine of the control and Lb. plantarum groups. IL-1β, IL-10 and TNF-α gene expression were significantly up-regulated by Lb. plantarum. Moreover, the mRNA levels of IL-10, IL-8 and IgT were significantly higher in the Lb. plantarum group after L. garvieae infection, suggesting that Lb. plantarum can stimulate the immune response of rainbow trout. PCR-DGGE revealed no detectable levels of the probiotics or the pathogen present on the distal intestinal mucosa. These findings demonstrate that direct probiotic-host interactions with the intestine are not always necessary to induce host stimulatory responses which ultimately enhance disease resistance. Furthermore, as L. garvieae did not colonise the intestinal tract, and therefore likely did not infect via this route, the antagonistic properties of the probiotic candidate towards L. garvieae were likely of little influence in mediating the improved disease resistance which could be attributed to the elevated immunological response.
To, Thi Mai Huong; Grandvalet, Cosette; Tourdot-Maréchal, Raphaëlle
Cyclopropane fatty acids (CFAs) are synthetized in situ by the transfer of a methylene group from S-adenosyl-l-methionine to a double bond of unsaturated fatty acid chains of membrane phospholipids. This conversion, catalyzed by the Cfa synthase enzyme, occurs in many bacteria and is recognized to play a key role in the adaptation of bacteria in response to a drastic perturbation of the environment. The role of CFAs in the acid tolerance response was investigated in the lactic acid bacterium Lactococcus lactis MG1363. A mutant of the cfa gene was constructed by allelic exchange. The cfa gene encoding the Cfa synthase was cloned and introduced into the mutant to obtain the complemented strain for homologous system studies. Data obtained by gas chromatography (GC) and GC-mass spectrometry (GC-MS) validated that the mutant could not produce CFA. The CFA levels in both the wild-type and complemented strains increased upon their entry to stationary phase, especially with acid-adapted cells or, more surprisingly, with ethanol-adapted cells. The results obtained by performing quantitative reverse transcription-PCR (qRT-PCR) experiments showed that transcription of the cfa gene was highly induced by acidity (by 10-fold with cells grown at pH 5.0) and by ethanol (by 9-fold with cells grown with 6% ethanol) in comparison with that in stationary phase. Cell viability experiments were performed after an acidic shock on the mutant strain, the wild-type strain, and the complemented strain, as a control. The higher viability level of the acid-adapted cells of the three strains after 3 h of shock proved that the cyclopropanation of unsaturated fatty acids is not essential for L. lactis subsp. cremoris survival under acidic conditions. Moreover, fluorescence anisotropy data showed that CFA itself could not maintain the membrane fluidity level, particularly with ethanol-grown cells. PMID:21421775
To, Thi Mai Huong; Grandvalet, Cosette; Tourdot-Maréchal, Raphaëlle
Cyclopropane fatty acids (CFAs) are synthetized in situ by the transfer of a methylene group from S-adenosyl-L-methionine to a double bond of unsaturated fatty acid chains of membrane phospholipids. This conversion, catalyzed by the Cfa synthase enzyme, occurs in many bacteria and is recognized to play a key role in the adaptation of bacteria in response to a drastic perturbation of the environment. The role of CFAs in the acid tolerance response was investigated in the lactic acid bacterium Lactococcus lactis MG1363. A mutant of the cfa gene was constructed by allelic exchange. The cfa gene encoding the Cfa synthase was cloned and introduced into the mutant to obtain the complemented strain for homologous system studies. Data obtained by gas chromatography (GC) and GC-mass spectrometry (GC-MS) validated that the mutant could not produce CFA. The CFA levels in both the wild-type and complemented strains increased upon their entry to stationary phase, especially with acid-adapted cells or, more surprisingly, with ethanol-adapted cells. The results obtained by performing quantitative reverse transcription-PCR (qRT-PCR) experiments showed that transcription of the cfa gene was highly induced by acidity (by 10-fold with cells grown at pH 5.0) and by ethanol (by 9-fold with cells grown with 6% ethanol) in comparison with that in stationary phase. Cell viability experiments were performed after an acidic shock on the mutant strain, the wild-type strain, and the complemented strain, as a control. The higher viability level of the acid-adapted cells of the three strains after 3 h of shock proved that the cyclopropanation of unsaturated fatty acids is not essential for L. lactis subsp. cremoris survival under acidic conditions. Moreover, fluorescence anisotropy data showed that CFA itself could not maintain the membrane fluidity level, particularly with ethanol-grown cells.
Sperandio, Brice; Polard, Patrice; Ehrlich, Dusko S.; Renault, Pierre; Guédon, Eric
Cysteine and methionine availability influences many processes in the cell. In bacteria, transcription of the specific genes involved in the synthesis of these two amino acids is usually regulated by different mechanisms or regulators. Pathways for the synthesis of cysteine and methionine and their interconversion were experimentally determined for Lactococcus lactis, a lactic acid bacterium commonly found in food. A new gene, yhcE, was shown to be involved in methionine recycling to cysteine. Surprisingly, 18 genes, representing almost all genes of these pathways, are under the control of a LysR-type activator, FhuR, also named CmbR. DNA microarray experiments showed that FhuR targets are restricted to this set of 18 genes clustered in seven transcriptional units, while cysteine starvation modifies the transcription level of several other genes potentially involved in oxidoreduction processes. Purified FhuR binds a 13-bp box centered 46 to 53 bp upstream of the transcriptional starts from the seven regulated promoters, while a second box with the same consensus is present upstream of the first binding box, separated by 8 to 10 bp. O-Acetyl serine increases FhuR binding affinity to its binding boxes. The overall view of sulfur amino acid metabolism and its regulation in L. lactis indicates that CysE could be a master enzyme controlling the activity of FhuR by providing its effector, while other controls at the enzymatic level appear to be necessary to compensate the absence of differential regulation of the genes involved in the interconversion of methionine and cysteine and other biosynthesis genes. PMID:15901700
van Pijkeren, Jan Peter; Britton, Robert A
Innovative new genome engineering technologies for manipulating chromosomes have appeared in the last decade. One of these technologies, recombination mediated genetic engineering (recombineering) allows for precision DNA engineering of chromosomes and plasmids in Escherichia coli. Single-stranded DNA recombineering (SSDR) allows for the generation of subtle mutations without the need for selection and without leaving behind any foreign DNA. In this review we discuss the application of SSDR technology in lactic acid bacteria, with an emphasis on key factors that were critical to move this technology from E. coli into Lactobacillus reuteri and Lactococcus lactis. We also provide a blueprint for how to proceed if one is attempting to establish SSDR technology in a lactic acid bacterium. The emergence of CRISPR-Cas technology in genome engineering and its potential application to enhancing SSDR in lactic acid bacteria is discussed. The ability to perform precision genome engineering in medically and industrially important lactic acid bacteria will allow for the genetic improvement of strains without compromising safety.
Innovative new genome engineering technologies for manipulating chromosomes have appeared in the last decade. One of these technologies, recombination mediated genetic engineering (recombineering) allows for precision DNA engineering of chromosomes and plasmids in Escherichia coli. Single-stranded DNA recombineering (SSDR) allows for the generation of subtle mutations without the need for selection and without leaving behind any foreign DNA. In this review we discuss the application of SSDR technology in lactic acid bacteria, with an emphasis on key factors that were critical to move this technology from E. coli into Lactobacillus reuteri and Lactococcus lactis. We also provide a blueprint for how to proceed if one is attempting to establish SSDR technology in a lactic acid bacterium. The emergence of CRISPR-Cas technology in genome engineering and its potential application to enhancing SSDR in lactic acid bacteria is discussed. The ability to perform precision genome engineering in medically and industrially important lactic acid bacteria will allow for the genetic improvement of strains without compromising safety. PMID:25185700
Saraoui, Taous; Fall, Papa Abdoulaye; Leroi, Françoise; Antignac, Jean-Philippe; Chéreau, Sylvain; Pilet, Marie France
Listeria monocytogenes is a pathogenic Gram positive bacterium and the etiologic agent of listeriosis, a severe food-borne disease. Lactococcus piscium CNCM I-4031 has the capacity to prevent the growth of L. monocytogenes in contaminated peeled and cooked shrimp. To investigate the inhibititory mechanism, a chemically defined medium (MSMA) based on shrimp composition and reproducing the inhibition observed in shrimp was developed. In co-culture at 26 °C, L. monocytogenes was reduced by 3-4 log CFU g(-1) after 24 h. We have demonstrated that the inhibition was not due to secretion of extracellular antimicrobial compounds as bacteriocins, organic acids and hydrogen peroxide. Global metabolomic fingerprints of these strains in pure culture were assessed by liquid chromatography coupled with high resolution mass spectrometry. Consumption of glucose, amino-acids, vitamins, nitrogen bases, iron and magnesium was measured and competition for some molecules could be hypothesized. However, after 24 h of co-culture, when inhibition of L. monocytogenes occurred, supplementation of the medium with these compounds did not restore its growth. The inhibition was observed in co-culture but not in diffusion chamber when species were separated by a filter membrane. Taken together, these data indicate that the inhibition mechanism of L. monocytogenes by L. piscium is cell-to-cell contact-dependent.
Wu, Chongde; Zhang, Juan; Du, Guocheng; Chen, Jian
The aim of this study was to investigate the effect of nisin-inducible RecO expression on the stress tolerance of Lactococcus lactis NZ9000. RecO protein from Lactobacillus casei Zhang was introduced into Lactococcus lactis NZ9000 by using a nisin-inducible expression system. The recombinant strain (NZ-RecO) exhibited higher growth performances and survival rate compared with the control strain (NZ-Vector) under stress conditions. In addition, the NZ-RecO strain exhibited 1.37-, 1.41-, and 1.42-fold higher biomass, lactate production, lactate productivity, compared with the corresponding values for NZ-Vector during NaCl-stressed condition. Analysis of lactate dehydrogenase (LDH) activity showed that the production of RecO maintained the stability of LDH during salt stress. These results suggest that overproduction of RecO improved the multiple-stress tolerance and lactic acid production in Lactococcus lactis NZ9000 during salt stress. Results presented in this study may help to enhance the industrial utility of lactic acid bacteria.
Amiri-Jami, Mitra; Lapointe, Gisele; Griffiths, Mansel W
Eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA) have been shown to be of major importance in human health. Therefore, these essential polyunsaturated fatty acids have received considerable attention in both human and farm animal nutrition. Currently, fish and fish oils are the main dietary sources of EPA/DHA. To generate sustainable novel sources for EPA and DHA, the 35-kb EPA/DHA synthesis gene cluster was isolated from a marine bacterium, Shewanella baltica MAC1. To streamline the introduction of the genes into food-grade microorganisms such as lactic acid bacteria, unnecessary genes located upstream and downstream of the EPA/DHA gene cluster were deleted. Recombinant Escherichia coli harboring the 20-kb gene cluster produced 3.5- to 6.1-fold more EPA than those carrying the 35-kb DNA fragment coding for EPA/DHA synthesis. The 20-kb EPA/DHA gene cluster was cloned into a modified broad-host-range low copy number vector, pIL252m (4.7 kb, Ery) and expressed in Lactococcus lactis subsp. cremoris MG1363. Recombinant L. lactis produced DHA (1.35 ± 0.5 mg g(-1) cell dry weight) and EPA (0.12 ± 0.04 mg g(-1) cell dry weight). This is believed to be the first successful cloning and expression of EPA/DHA synthesis gene cluster in lactic acid bacteria. Our findings advance the future use of EPA/DHA-producing lactic acid bacteria in such applications as dairy starters, silage adjuncts, and animal feed supplements.
Hmar, Rothangmawi Victoria; Prasad, Shashi Bala; Jayaraman, Guhan; Ramachandran, Kadathur B
Microbial production of hyaluronic acid (HA) is an attractive substitute for extraction of this biopolymer from animal tissues. Natural producers such as Streptococcus zooepidemicus are potential pathogens; therefore, production of HA by recombinant bacteria that are generally recognized as safe (GRAS) organisms is a viable alternative that is being extensively explored. However, plasmid-based expression systems for HA production by recombinant bacteria have the inherent disadvantage of reduced productivity because of plasmid instability. To overcome this problem, the HA synthesis genes (hasA-hasB and hasA-hasB-hasC) from has-operon of S. zooepidemicus were integrated into the chromosome of Lactococcus lactis by site-directed, double-homologous recombination developing strains VRJ2AB and VRJ3ABC. The chromosomal integration stabilized the genes and obviated the instability observed in plasmid-expressed recombinant strains. The genome-integrated strains produced higher molecular weight (3.5-4 million Dalton [MDa]) HA compared to the plasmid-expressed strains (2 MDa). High molecular weight HA was produced when the intracellular concentration of uridine diphosphate N-acetylglucosamine (UDP-GlcNAc) and uridine diphosphate-glucuronic acid (UDP-GlcUA) was almost equal and hasA to hasB ratio was low. This work suggests an optimal approach to obtain high molecular weight HA in recombinant strains.
Sarika, A R; Lipton, A P; Aishwarya, M S; Dhivya, R S
The bacteriocins of lactic acid bacteria have considerable potential for biopreservation. The Lactococcus lactis strain PSY2 (GenBank account no. JF703669) isolated from the surface of marine perch Perca flavescens produced antibacterial activity against pathogenic and spoilage-causing Gram-positive and Gram-negative bacteria viz. Arthrobacter sp., Acinetobacter sp., Bacillus subtilis, Escherichia coli, Listeria monocytogenes, Pseudomonas aeruginosa and Staphylococcus aureus and possessed broad inhibitory spectrum. The biopreservative efficacy of the bacteriocin PSY2 was evaluated using fillets of reef cod, Epinephelus diacanthus. The fillets (10 g) were sprayed with 2.0 ml of 1,600 AU/ml bacteriocin, wrapped and kept under different storage temperatures viz., 4, 0 and -18 °C. The biopreservative extended the shelf-life of fillets stored at 4 °C to >21 days as against <14 days observed in the untreated samples. The total count of spoilage bacteria was reduced by 2.5 logarithmic units in the treated sample during the 14th day of storage as against the control. Chemical analysis revealed a significant change (P < 0.05) in the pH value, free fatty acid (as % oleic acid), total volatile base nitrogen and total methyl amine content in the treated samples. The overall acceptability in terms of sensory attributes was significantly higher in the bacteriocin-treated samples stored for 21 days at 4 °C while the untreated samples became unacceptable by the 14th day. The biopreservative gave no significant effect at -18 °C. Thus, the bacteriocin derived from L. lactis PSY2 gave increased protection against spoilage bacteria and offers an alternative for the preservation of high-value sea foods.
Machii, Miki; Watanabe, Satoru; Zendo, Takeshi; Chibazakura, Taku; Sonomoto, Kenji; Shimizu-Kadota, Mariko; Yoshikawa, Hirofumi
Two chemically defined media, CDM-1G and CDM-1X, that use glucose and xylose as carbon sources, respectively, were prepared for Lactococcus lactis strain IO-1. The maximal cell density at 600 nm in CDM-1G exceeded 2. Omission growth experiments indicated that IO-1 is auxotrophic for 2 vitamins and 6 amino acids.
Velly, H.; Abraham, A.-L.; Loux, V.; Delacroix-Buchet, A.; Fonseca, F.; Bouix, M.
Lactococcus lactis is a lactic acid bacterium used in the production of many fermented foods, such as dairy products. Here, we report the genome sequence of L. lactis subsp. lactis TOMSC161, isolated from nonscalded curd pressed cheese. This genome sequence provides information in relation to dairy environment adaptation. PMID:25377704
Livins'ka, O P; Harmasheva, I L; Kovalenko, N K
The current view of the structural diversity of teichoic acids and their involvement in the biological activity of lactobacilli has been reviewed. The mechanisms of effects of probiotic lactic acid bacteria, in particular adhesive and immunostimulating functions have been described. The prospects of the use of structure data of teichoic acid in the assessment of intraspecific diversity of lactic acid bacteria have been also reflected.
Yvon, M; Thirouin, S; Rijnen, L; Fromentier, D; Gripon, J C
The enzymatic degradation of amino acids in cheese is believed to generate aroma compounds and therefore to be involved in the complex process of cheese flavor development. In lactococci, transamination is the first step in the degradation of aromatic and branched-chain amino acids which are precursors of aroma compounds. Here, the major aromatic amino acid aminotransferase of a Lactococcus lactis subsp. cremoris strain was purified and characterized. The enzyme transaminates the aromatic amino acids, leucine, and methionine. It uses the ketoacids corresponding to these amino acids and alpha-ketoglutarate as amino group acceptors. In contrast to most bacterial aromatic aminotransferases, it does not act on aspartate and does not use oxaloacetate as second substrate. It is essential for the transformation of aromatic amino acids to flavor compounds. It is a pyridoxal 5'-phosphate-dependent enzyme and is composed of two identical subunits of 43.5 kDa. The activity of the enzyme is optimal between pH 6.5 and 8 and between 35 and 45 degrees C, but it is still active under cheese-ripening conditions. PMID:9023921
Trip, Hein; Mulder, Niels L.; Lolkema, Juke S.
Degradative amino acid decarboxylation pathways in bacteria generate secondary metabolic energy and provide resistance against acid stress. The histidine decarboxylation pathway of Streptococcus thermophilus CHCC1524 was functionally expressed in the heterologous host Lactococcus lactis NZ9000, and the benefits of the newly acquired pathway for the host were analyzed. During growth in M17 medium in the pH range of 5–6.5, a small positive effect was observed on the biomass yield in batch culture, whereas no growth rate enhancement was evident. In contrast, a strong benefit for the engineered L. lactis strain was observed in acid stress survival. In the presence of histidine, the pathway enabled cells to survive at pH values as low as 3 for at least 2 h, conditions under which the host cells were rapidly dying. The flux through the histidine decarboxylation pathway in cells grown at physiological pH was under strict control of the electrochemical proton gradient (pmf) across the membrane. Ionophores that dissipated the membrane potential (ΔΨ) and/or the pH gradient (ΔpH) strongly increased the flux, whereas the presence of glucose almost completely inhibited the flux. Control of the pmf over the flux was exerted by both ΔΨ and ΔpH and was distributed over the transporter HdcP and the decarboxylase HdcA. The control allowed for a synergistic effect between the histidine decarboxylation and glycolytic pathways in acid stress survival. In a narrow pH range around 2.5 the synergism resulted in a 10-fold higher survival rate. PMID:22351775
Stiles, M E
Biopreservation refers to extended storage life and enhanced safety of foods using the natural microflora and (or) their antibacterial products. Lactic acid bacteria have a major potential for use in biopreservation because they are safe to consume and during storage they naturally dominate the microflora of many foods. In milk, brined vegetables, many cereal products and meats with added carbohydrate, the growth of lactic acid bacteria produces a new food product. In raw meats and fish that are chill stored under vacuum or in an environment with elevated carbon dioxide concentration, the lactic acid bacteria become the dominant population and preserve the meat with a "hidden' fermentation. The same applies to processed meats provided that the lactic acid bacteria survive the heat treatment or they are inoculated onto the product after heat treatment. This paper reviews the current status and potential for controlled biopreservation of foods.
Kato, Hiroaki; Shiwa, Yuh; Oshima, Kenshiro; Machii, Miki; Araya-Kojima, Tomoko; Zendo, Takeshi; Shimizu-Kadota, Mariko; Hattori, Masahira; Sonomoto, Kenji; Yoshikawa, Hirofumi
We report the complete genome sequence of Lactococcus lactis IO-1 (= JCM7638). It is a nondairy lactic acid bacterium, produces nisin Z, ferments xylose, and produces predominantly L-lactic acid at high xylose concentrations. From ortholog analysis with other five L. lactis strains, IO-1 was identified as L. lactis subsp. lactis.
Vitiñi, E; Alvarez, S; Medina, M; Medici, M; de Budeguer, M V; Perdigón, G
The beneficial properties of lactic acid bacteria (LAB) on human health have been frequently demonstrated. The interaction of LAB with the lymphoid cells associated to the gut to activate the mucosal immune system and the mechanisms by which they can exert an adjuvant effect is still unclear, as well as if this property is common for all the LAB. We studied the influence of the oral administration of different geneous of LAB such as Lactobacillus casei, L. acidophilus, L. rhamnosus, L. delbrueckii subsp. bulgaricus, L. plantarum, Lactococcus lactis and Streptococcus thermophilus. We determined if the LAB assayed were able to stimulate the specific, the non-specific immune response (inflammatory response), or both. We demonstrated that all the bacteria assayed were able to increase the number of IgA producing cells associated to the lamina propria of small intestine. This effect was dose dependent. The increase in IgA+ producing cells was not always correlated with an increase in the CD4+ T cell number, indicating that some LAB assayed only induced clonal expansion of B cells triggered to produce IgA. Most of them, induced an increase in the number of cells involved in the inflammatory immune response. CD8+ T cell were diminished or not affected, with exception of L. plantarum that induced an increase at low dose. This fact would mean that LAB are unable to induce cytotoxicity mechanisms. We demonstrated the importance in the selection of LAB to be used as gut mucosal adjuvant. The different behaviours observed among them on the gut mucosal immune response, specially those that induce inflammatory immune response, show that not all the LAB can be used as oral adjuvant and that the beneficial effect of them can not generalized to genous or specie. The immunoadjuvant capacity would be a property of the strain assayed.
Zhang, Jian; Caiyin, Qinggele; Feng, Wenjing; Zhao, Xiuli; Qiao, Bin; Zhao, Guangrong; Qiao, Jianjun
Traditionally, nisin was produced industrially by using Lactococcus lactis in the neutral fermentation process. However, nisin showed higher activity in the acidic environment. How to balance the pH value for bacterial normal growth and nisin activity might be the key problem. In this study, 17 acid-tolerant genes and 6 lactic acid synthetic genes were introduced in L. lactis F44, respectively. Comparing to the 2810 IU/mL nisin yield of the original strain F44, the nisin titer of the engineered strains over-expressing hdeAB, ldh and murG, increased to 3850, 3979 and 4377 IU/mL, respectively. These engineered strains showed more stable intracellular pH value during the fermentation process. Improvement of lactate production could partly provide the extra energy for the expression of acid tolerance genes during growth. Co-overexpression of hdeAB, murG, and ldh(Z) in strain F44 resulted in the nisin titer of 4913 IU/mL. The engineered strain (ABGL) could grow on plates with pH 4.2, comparing to the surviving pH 4.6 of strain F44. The fed-batch fermentation showed nisin titer of the co-expression L. lactis strain could reach 5563 IU/mL with lower pH condition and longer cultivation time. This work provides a novel strategy of constructing robust strains for use in industry process. PMID:27306587
Stecchini, M L; Del Torre, M; Munari, M
Responses of lactic acid bacteria (LAB) to peroxy radicals generated via thermal (40 degrees C) decomposition of the diazocompound 2,2,-azo-bis (2-amidinopropane) dihydrochloride (ABAP), were studied. In general, LAB displayed survival curves with shoulders and tails indicative of 'multihit' killing by exposure to peroxy radicals. One strain, Lactococcus lactis subsp. lactis DIP15, producing a slope of 0.0105 in the kinetic analysis when exposed to 4 mM ABAP, exhibited a measurable antioxidant capacity. The other LAB failed to show any significant antioxidant capacity. The antioxidant capacity of strain DIP15 remained constant after cells have been heat-treated, suggesting that compounds bearing free radical scavenging capacity are rather stable.
Marché, Laurent; Saraoui, Taous; Remenant, Benoit; Zagorec, Monique; Prévost, Hervé; Delbarre-Ladrat, Christine; Leroi, Françoise
ABSTRACT Lactococcus piscium CNCM I-4031 is a psychotrophic foodborne lactic acid bacterium showing potential interest for the biopreservation of seafood products due to its inhibition properties toward pathogenic and spoilage bacteria. The analysis of its genome will provide a better understanding of the mechanisms of interaction between these bacteria. PMID:28126939
Yang, J; Cao, Y; Cai, Y; Terada, F
Natural populations of lactic acid bacteria (LAB) and silage fermentation of vegetable residues were studied. Fifty-two strains of LAB isolated from cabbage, Chinese cabbage, and lettuce residues were identified and characterized. The LAB strains were gram-positive and catalase-negative bacteria, which were divided into 6 groups (A to F) according to morphological and biochemical characteristics. The strains in group A were rods that did not produce gas from glucose and formed the d and l isomers of lactate. Groups B and C were homofermentative cocci that formed l-lactic acid. Groups D, E, and F were heterofermentative cocci that formed d-lactic acid. Based on 16S rDNA gene sequence analysis, group A to F strains were identified as Lactobacillus plantarum, Lactococcus piscium, Lactococcus lactis, Leuconostoc citreum, Weissella soli and Leuconostoc gelidum, respectively. The prevalent LAB, predominantly homofermentative lactobacilli, consisted of Lactobacillus plantarum (34.6%), Weissella soli (19.2%), Leuconostoc gelidum (15.4%), Leuconostoc citreum (13.5%), Lactococcus lactis (9.6%), and Lactococcus piscium (7.7%). Lactobacillus plantarum was the dominant member of the LAB population in 3 types of vegetable residues. These vegetable residues contained a high level of crude protein (20.2 to 28.4% of dry matter). These silages prepared by using a small-scale fermentation system were well preserved, with low pH and a relatively high content of lactate. This study suggests that the vegetable residues contain abundant LAB species and nutrients, and that they could be well preserved by making silage, which is a potentially good vegetable protein source for livestock diets.
Halldórsdóttir, Halla D; Haraldsdóttir, Vilhelmina; Bödvarsson, Asgeir; Thorgeirsson, Gestur; Kristjánsson, Már
We describe a case of subacute endocarditis due to Lactococcus cremoris associated with consumption of unpasteurized milk. Treatment with amoxicillin-clavulanic acid and subsequently penicillin resulted in prompt sterilization of this patient's bloodstream and full recovery.
Zagorec, Monique; Anba-Mondoloni, Jamila; Coq, Anne-Marie Crutz-Le; Champomier-Vergès, Marie-Christine
Many meat (or fish) products, obtained by the fermentation of meat originating from various animals by the flora that naturally contaminates it, are part of the human diet since millenaries. Historically, the use of bacteria as starters for the fermentation of meat, to produce dry sausages, was thus performed empirically through the endogenous micro-biota, then, by a volunteer addition of starters, often performed by back-slopping, without knowing precisely the microbial species involved. It is only since about 50 years that well defined bacterial cultures have been used as starters for the fermentation of dry sausages. Nowadays, the indigenous micro-biota of fermented meat products is well identified, and the literature is rich of reports on the identification of lactic acid bacteria (LAB) present in many traditional fermented products from various geographical origin, obtained without the addition of commercial starters (See Talon, Leroy, & Lebert, 2007, and references therein).
Colombo, E; Franzetti, L; Frusca, M; Scarpellini, M
The lactic acid bacteria community in traditional goat cheese produced in three dairies in Valsesia (Piemonte, Italy) was studied at different steps of the manufacturing process. These cheeses were produced from raw milk without starter bacteria, and no protocol was followed during the manufacturing process. Three hundred thirty-two isolates were characterized and grouped by results of both morphophysiological tests and random amplification of polymorphic DNA plus PCR analysis. Bacteria were identified by partial sequencing of the 16S rRNA gene. Lactococci were the dominant lactic acid bacteria in raw milk. Their initial numbers ranged from 5 to 7 log CFU ml(-1). Their levels increased during manufacturing and decreased during ripening. The growth trend for enterococci was comparable to that of lactococci, although enterococci counts were lower. Lactococcus lactis subsp. cremoris, Lactococcus garviae, and Enterococcus faecalis were the most frequently isolated species during goat cheese manufacturing, whereas the highest numbers of Enterococcus (E. faecium, E. durans, E. gilvus, and E. casseliflavus) were isolated with the greatest frequency from ripened cheese samples. Occasionally, Leuconostoc mesenteroides, Leuconostoc lactis, and Lactobacillus paraplantarum also were isolated.
BELHADJ, Hani; HARZALLAH, Daoud; BOUAMRA, Dalila; KHENNOUF, Seddik; Dahamna, Saliha; GHADBANE, Mouloud
In the present work, five hundred and sixty-seven isolates of lactic acid bacteria were recovered from raw bee pollen grains. All isolates were screened for their antagonistic activity against both Gram-positive and Gram-negative pathogenic bacteria. Neutralized supernatants of 54 lactic acid bacteria (LAB) cultures from 216 active isolates inhibited the growth of indicator bacteria. They were phenotypically characterized, based on the fermentation of 39 carbohydrates. Using the simple matching coefficient and unweighted pair group algorithm with arithmetic averages (UPGMA), seven clusters with other two members were defined at the 79% similarity level. The following species were characterized: Lactobacillus plantarum, Lactobacillus fermentum, Lactococcus lactis, Pediococcus acidilactici, Pediococcus pentosaceus, and unidentified lactobacilli. Phenotypic characteristics of major and minor clusters were also identified. Partial sequencing of the 16S rRNA gene of representative isolates from each cluster was performed, and ten strains were assigned to seven species: Lactobacillus plantarum, Lactobacillus fermentum, Lactococcus lactis, Lactobacillus ingluviei, Pediococcus pentosaceus, Lactobacillus acidipiscis and Weissella cibaria. The molecular method used failed to determine the exact taxonomic status of BH0900 and AH3133. PMID:24936378
Reynolds, Jackie; Moyes, Rita B; Breakwell, Donald P
Acid-fastness is an uncommon characteristic shared by the genera Mycobacterium (Section 10A) and Nocardia. Because of this feature, this stain is extremely helpful in identification of these bacteria. Although Gram positive, acid-fast bacteria do not take the crystal violet into the wall well, appearing very light purple rather than the deep purple of normal Gram-positive bacteria.
Bunthof, Christine J.; Bloemen, Karen; Breeuwer, Pieter; Rombouts, Frank M.; Abee, Tjakko
The viability of lactic acid bacteria is crucial for their applications as dairy starters and as probiotics. We investigated the usefulness of flow cytometry (FCM) for viability assessment of lactic acid bacteria. The esterase substrate carboxyfluorescein diacetate (cFDA) and the dye exclusion DNA binding probes propidium iodide (PI) and TOTO-1 were tested for live/dead discrimination using a Lactococcus, a Streptococcus, three Lactobacillus, two Leuconostoc, an Enterococcus, and a Pediococcus species. Plate count experiments were performed to validate the results of the FCM assays. The results showed that cFDA was an accurate stain for live cells; in exponential-phase cultures almost all cells were labeled, while 70°C heat-killed cultures were left unstained. PI did not give clear live/dead discrimination for some of the species. TOTO-1, on the other hand, gave clear discrimination between live and dead cells. The combination of cFDA and TOTO-1 gave the best results. Well-separated subpopulations of live and dead cells could be detected with FCM. Cell sorting of the subpopulations and subsequent plating on agar medium provided direct evidence that cFDA labels the culturable subpopulation and that TOTO-1 labels the nonculturable subpopulation. Applied to cultures exposed to deconjugated bile salts or to acid, cFDA and TOTO-1 proved to be accurate indicators of culturability. Our experiments with lactic acid bacteria demonstrated that the combination of cFDA and TOTO-1 makes an excellent live/dead assay with versatile applications. PMID:11319119
Gallegos, Janneth; Arce, Cristina; Jordano, Rafael; Arce, Lourdes; Medina, Luis M
The purpose of this work was to study the potential of gas chromatography-ion mobility spectrometry (GC-IMS) to differentiate lactic acid bacteria (LAB) through target identification and fingerprints of volatile metabolites. The LAB selected were used as reference strains for their influence in the flavour of cheese. The four strains of LAB can be distinguished by the fingerprints generated by the volatile organic compounds (VOCs) emitted. 2-butanone, 2-pentanone, 2-heptanone and 3-methyl-1-butanol were identified as relevant VOCs for Lactobacillus casei and Lactobacillus paracasei subsp. paracasei. 2-Butanone and 3-methyl-1-butanol were identified in Lactococcus lactis subsp. lactis and Lactococcus cremoris subsp. cremoris. The IMS signals monitoring during a 24-30h period showed the growth of the LAB in vitro. The results demonstrated that GC-IMS is a useful technology for bacteria recognition and also for screening the aromatic potential of new isolates of LAB.
Yao, Jiangwei; Rock, Charles O.
Membrane phospholipid synthesis is a vital facet of bacterial physiology. Although the spectrum of phospholipid headgroup structures produced by bacteria is large, the key precursor to all of these molecules is phosphatidic acid (PtdOH). Glycerol-3-phosphate derived from the glycolysis via glycerol-phosphate synthase is the universal source for the glycerol backbone of PtdOH. There are two distinct families of enzymes responsible for the acylation of the 1-position of glycerol-3-phosphate. The PlsB acyltransferase was discovered in Escherichia coli, and homologs are present in many eukaryotes. This protein family primarily uses acyl-acyl carrier protein (ACP) endproducts of fatty acid synthesis as acyl donors, but may also use acyl-CoA derived from exogenous fatty acids. The second protein family, PlsY, is more widely distributed in bacteria and utilizes the unique acyl donor, acyl-phosphate, which is produced from acyl-ACP by the enzyme PlsX. The acylation of the 2-position is carried out by members of the PlsC protein family. All PlsCs use acyl-ACP as the acyl donor, although the PlsCs of the γ-proteobacteria also may use acyl-CoA. Phospholipid headgroups are precursors in the biosynthesis of other membrane-associated molecules and the diacylglycerol product of these reactions is converted to PtdOH by one of two distinct families of lipid kinases. The central importance of the de novo and recycling pathways to PtdOH in cell physiology suggest these enzymes are suitable targets for the development of antibacterial therapeutics in Gram-positive pathogens. This article is part of a Special Issue entitled Phospholipids and Phospholipid Metabolism. PMID:22981714
Shimizu-Kadota, Mariko; Kato, Hiroaki; Shiwa, Yuh; Oshima, Kenshiro; Machii, Miki; Araya-Kojima, Tomoko; Zendo, Takeshi; Hattori, Masahira; Sonomoto, Kenji; Yoshikawa, Hirofumi
Lactococcus lactis IO-1 (JCM7638) produces L-lactic acid predominantly when grown at high xylose concentrations, and its utilization is highly desired in the green plastics industry. Therefore it is worthwhile studying its genomic traits. In this study, we focused on (i) genes of possible horizontal transfer derivation (prophages, the nisin-sucrose transposon, and several restriction-modification systems), and (ii) genes for the synthetic pathways of amino acids and vitamins in the IO-1 genome. In view of the results of this analysis, we consider their meanings in strain IO-1.
Ganesan, Balasubramanian; Dobrowolski, Piotr; Weimer, Bart C.
Nutrient starvation and nonculturability in bacteria lead to changes in metabolism not found during the logarithmic phase. Substrates alternate to those used during growth are metabolized in these physiological states, yielding secondary metabolites. In firmicutes and actinobacteria, amino acid catabolic pathways are induced during starvation and nonculturability. Examination of lactococci showed that the population entered a nonculturable state after carbohydrate depletion and was incapable of growth on solid media; however, the cells gained the ability to produce branched-chain fatty acids from amino acids. Gene expression profiling and in silico pathway analysis coupled with nuclear magnetic resonance spectroscopy were used to delineate the leucine catabolic pathway. Lactococci produced acetic and propionic acid during logarithmic growth and starvation. At the onset of nonculturability, 2-methylbutyric acid was produced via hydroxymethyl-glutaryl-coenzyme A (CoA) and acetyl-CoA, along with ATP and oxidation/reduction precursors. Gene expression profiling and genome sequence analysis showed that lactococci contained redundant genes for branched-chain fatty acid production that were regulated by an unknown mechanism linked to carbon metabolism. This work demonstrated the ability of a firmicute to induce new metabolic capabilities in the nonculturable state for producing energy and intermediates needed for transcription and translation. Phylogenetic analyses showed that homologues of these enzymes and their functional motifs were widespread across the domains of life. PMID:16751541
Carter, R N; Schmidt, J M
The cellular fatty acid composition of 14 strains of Caulobacter speices and types, two species of Prosthecomicrobium, and two species of Asticcacaulis was determined by gas-liquid chromatography. In most of these bacteria, the major fatty acids were octadecenoic acid (C18:1), hexadecenoic acid (C16:1) and hexadecanoic acid (C16:0). Some cyclopropane and branched chain fatty acids were detected in addition to the straight chained acids. Hydroxytetradecanoic acid was an important component of P.enhydrum but significant amounts of hydroxy acids were not detected in other prosthecate bacteria examined.
Ljungh, Asa; Wadström, Torkel
A number of Lactobacillus species, Bifidobacterium sp, Saccharomyces boulardii, and some other microbes have been proposed as and are used as probiotic strains, i.e. live microorganisms as food supplement in order to benefit health. The health claims range from rather vague as regulation of bowel activity and increasing of well-being to more specific, such as exerting antagonistic effect on the gastroenteric pathogens Clostridium difficile, Campylobacter jejuni, Helicobacter pylori and rotavirus, neutralising food mutagens produced in colon, shifting the immune response towards a Th2 response, and thereby alleviating allergic reactions, and lowering serum cholesterol (Tannock, 2002). Unfortunately, most publications are case reports, uncontrolled studies in humans, or reports of animal or in vitro studies. Whether or not the probiotic strains employed shall be of human origin is a matter of debate but this is not a matter of concern, as long as the strains can be shown to survive the transport in the human gastrointestinal (GI) tract and to colonise the human large intestine. This includes survival in the stressful environment of the stomach - acidic pH and bile - with induction of new genes encoding a number of stress proteins. Since the availability of antioxidants decreases rostrally in the GI tract production of antioxidants by colonic bacteria provides a beneficial effect in scavenging free radicals. LAB strains commonly produce antimicrobial substance(s) with activity against the homologous strain, but LAB strains also often produce microbicidal substances with effect against gastric and intestinal pathogens and other microbes, or compete for cell surface and mucin binding sites. This could be the mechanism behind reports that some probiotic strains inhibit or decrease translocation of bacteria from the gut to the liver. A protective effect against cancer development can be ascribed to binding of mutagens by intestinal bacteria, reduction of the enzymes beta
Velly, H; Bouix, M; Passot, S; Penicaud, C; Beinsteiner, H; Ghorbal, S; Lieben, P; Fonseca, F
This work aimed at characterizing the biochemical and biophysical properties of the membrane of Lactococcus lactis TOMSC161 cells during fermentation at different temperatures, in relation to their freeze-drying and storage resistance. Cells were cultivated at two different temperatures (22 and 30 °C) and were harvested at different growth phases (from the middle exponential phase to the late stationary phase). Bacterial membranes were characterized by determining the fatty acid composition, the lipid phase transition, and the membrane fluidity. Cultivability and acidification activity losses of L. lactis were quantified after freezing, drying, and 3 months of storage. The direct measurement of membrane fluidity by fluorescence anisotropy was linked to lipid composition, and it was established that the cyclopropanation of unsaturated fatty acids with concomitant membrane rigidification during growth led to an increase in the freeze-drying and storage resistance of L. lactis. As expected, cultivating cells at a lower fermentation temperature than the optimum growth temperature induced a homeoviscous adaptation that was demonstrated by a lowered lipid phase transition temperature but that was not related to any improvement in freeze-drying resistance. L. lactis TOMSC161 was therefore able to develop a combined biochemical and biophysical response at the membrane level during fermentation. The ratio of cyclic fatty acids to unsaturated fatty acids (CFA/UFA) appeared to be the most relevant parameter associated with membrane rigidification and cell resistance to freeze-drying and storage. This study increased our knowledge about the physiological mechanisms that explain the resistance of lactic acid bacteria (LAB) to freeze-drying and storage stresses and demonstrated the relevance of complementary methods of membrane characterization.
Rajendran, Vivek; Puvendran, Kirubhakaran; Guru, Bharath Raja; Jayaraman, Guhan
Hyaluronic acid has a wide range of biomedical applications and its commercial value is highly dependent on its purity and molecular weight. This study highlights the utility of aqueous two-phase separation as a primary recovery step for hyaluronic acid and for removal of major protein impurities from fermentation broths. Metabolically engineered cultures of a lactate dehydrogenase mutant strain of Lactococcus lactis (L. lactis NZ9020) were used to produce high-molecular-weight hyaluronic acid. The cell-free fermentation broth was partially purified using a polyethylene glycol/potassium phosphate system, resulting in nearly 100% recovery of hyaluronic acid in the salt-rich bottom phase in all the aqueous two-phase separation experiments. These experiments were optimized for maximum removal of protein impurities in the polyethylene glycol rich top phase. The removal of protein impurities resulted in substantial reduction of membrane fouling in the subsequent diafiltration process, carried out with a 300 kDa polyether sulfone membrane. This step resulted in considerable purification of hyaluronic acid, without any loss in recovery and molecular weight. Diafiltration was followed by an adsorption step to remove minor impurities and achieve nearly 100% purity. The final hyaluronic acid product was characterized by Fourier-transform IR and NMR spectroscopy, confirming its purity.
Qiao, Jian-Jun; Zhang, Yan-Fei; Sun, Li-Fan; Liu, Wei-Wei; Zhu, Hong-Ji; Zhang, Zhijun
Spent mushroom substrate (SMS) was treated with dilute sulfuric acid followed by cellulase and xylanase treatment to produce hydrolysates that could be used as the basis for media for the production of value added products. A L9 (3(4)) orthogonal experiment was performed to optimize the acid treatment process. Pretreatment with 6% (w/w) dilute sulfuric acid at 120°C for 120 min provided the highest reducing sugar yield of 267.57 g/kg SMS. No furfural was detected in the hydrolysates. Exposure to 20PFU of cellulase and 200 XU of xylanase per gram of pretreated SMS at 40°C resulted in the release of 79.85 g/kg or reducing sugars per kg acid pretreated SMS. The dilute sulfuric acid could be recycled to process fresh SMS four times. SMS hydrolysates neutralized with ammonium hydroxide, sodium hydroxide, or calcium hydroxide could be used as the carbon source for cultivation of Lactococcus lactis subsp. lactis W28 and a cell density of 2.9×10(11)CFU/mL could be obtained. The results provide a foundation for the development of value-added products based on SMS.
Soto Del Rio, Maria de Los Dolores; Andrighetto, Christian; Dalmasso, Alessandra; Lombardi, Angiolella; Civera, Tiziana; Bottero, Maria Teresa
During the last years the interest in donkey milk has increased significantly mainly because of its compelling functional elements. Even if the composition and nutritional properties of donkey milk are known, its microbiota is less studied. This Research Communication aimed to provide a comprehensive characterisation of the lactic acid bacteria in raw donkey milk. RAPD-PCR assay combined with 16S rDNA sequencing analysis were used to describe the microbial diversity of several donkey farms in the North West part of Italy. The more frequently detected species were: Lactobacillus paracasei, Lactococcus lactis and Carnobacterium maltaromaticum. Less abundant genera were Leuconostoc, Enterococcus and Streptococcus. The yeast Kluyveromyces marxianus was also isolated. The bacterial and biotype distribution notably diverged among the farms. Several of the found species, not previously detected in donkey milk, could have an important probiotic activity and biotechnological potential. This study represents an important insight to the ample diversity of the microorganisms present in the highly selective ecosystem of raw donkey milk.
Stuart, M R; Chou, L S; Weimer, B C
Two strains of Lactococcus lactis subsp. lactis were used to determine the influence of lactose and arginine on viability and amino acid use during carbohydrate starvation. Lactose provided energy for logarithmic-phase growth, and amino acids such as arginine provided energy after carbohydrate exhaustion. Survival time, cell numbers, and ATP concentrations increased with the addition of arginine to the basal medium. By the onset of lactose exhaustion, the concentrations of glycine-valine and glutamate had decreased by as much as 67% in L. lactis ML3, whereas the serine concentration increased by 97% during the same period. When no lactose was added, the concentrations of these amino acids remained constant. Similar trends were observed for L. lactis 11454. Without lactose or arginine, L. lactis ML3 was nonculturable on agar but was viable after 2 days, as measured by fluorescent viability stains and intracellular ATP levels. However, L. lactis 11454 without lactose or arginine remained culturable for at least 14 days. These data suggest that lactococci become viable but nonculturable in response to carbohydrate depletion. Additionally, these data indicate that amino acids other than arginine facilitate the survival of L. lactis during carbohydrate starvation.
Cvrtila, Jelena; Topić, Ivana; Delaš, Frane; Markov, Ksenija
Summary The aim of this study is to identify and characterise potential autochthonous functional starter cultures in homemade horsemeat sausage. The dominant microflora in the samples of horsemeat sausage were lactic acid bacteria (LAB), followed by micrococci. Among the LAB, Lactococcus lactis ssp. lactis and Lactobacillus plantarum were the dominant species, and since the first is not common in fermented sausages, we characterised it as a potential functional starter culture. Lactococcus lactis ssp. lactis produced a significant amount of lactic acid, displayed good growth capability at 12, 18 and 22 °C, growth in the presence of 5% NaCl, good viability after lyophilisation and in simulated gastric and small intestinal juice, antimicrobial activity against test pathogens, and good adhesive properties in vitro. PMID:27904322
Dong, Xiangrong; Tian, Bing; Dai, Shang; Li, Tao; Guo, Linna; Tan, Zhongfang; Jiao, Zhen; Jin, Qingsheng; Wang, Yanping; Hua, Yuejin
PprI is a general switch protein that regulates the expression of certain proteins involved in pathways of cellular resistance in the extremophilic bacterium Deinococcus radiodurans. In this study, we transformed pprI into Lactococcus lactis strain MG1363 using the lactococcal shuttle vector pMG36e and investigated its effects on the tolerance and lactic acid production of L. lactis while under stress. PprI was stably expressed in L. lactis as confirmed by western blot assays. L. lactis expressing PprI exhibited significantly improved resistance to oxidative stress and high osmotic pressure. This enhanced cellular tolerance to stressors might be due to the regulation of resistance-related genes (e.g., recA, recO, sodA, and nah) by pprI. Moreover, transformed L. lactis demonstrated increased lactic acid production, attributed to enhanced lactate dehydrogenase activity. These results suggest that pprI can improve the tolerance of L. lactis to environmental stresses, and this transformed bacterial strain is a promising candidate for industrial applications of lactic acid production.
Dong, Xiangrong; Tian, Bing; Dai, Shang; Li, Tao; Guo, Linna; Tan, Zhongfang; Jiao, Zhen; Jin, Qingsheng; Wang, Yanping; Hua, Yuejin
PprI is a general switch protein that regulates the expression of certain proteins involved in pathways of cellular resistance in the extremophilic bacterium Deinococcus radiodurans. In this study, we transformed pprI into Lactococcus lactis strain MG1363 using the lactococcal shuttle vector pMG36e and investigated its effects on the tolerance and lactic acid production of L. lactis while under stress. PprI was stably expressed in L. lactis as confirmed by western blot assays. L. lactis expressing PprI exhibited significantly improved resistance to oxidative stress and high osmotic pressure. This enhanced cellular tolerance to stressors might be due to the regulation of resistance-related genes (e.g., recA, recO, sodA, and nah) by pprI. Moreover, transformed L. lactis demonstrated increased lactic acid production, attributed to enhanced lactate dehydrogenase activity. These results suggest that pprI can improve the tolerance of L. lactis to environmental stresses, and this transformed bacterial strain is a promising candidate for industrial applications of lactic acid production. PMID:26562776
Wyszyńska, Agnieszka; Kobierecka, Patrycja; Bardowski, Jacek; Jagusztyn-Krynicka, Elżbieta Katarzyna
Lactic acid bacteria (LAB) are a diverse group of Gram-positive, nonsporulating, low G + C content bacteria. Many of them have been given generally regarded as safe status. Over the past two decades, intensive genetic and molecular research carried out on LAB, mainly Lactococcus lactis and some species of the Lactobacillus genus, has revealed new, potential biomedical LAB applications, including the use of LAB as adjuvants, immunostimulators, or therapeutic drug delivery systems, or as factories to produce therapeutic molecules. LAB enable immunization via the mucosal route, which increases effectiveness against pathogens that use the mucosa as the major route of entry into the human body. In this review, we concentrate on the encouraging application of Lactococcus and Lactobacillus genera for the development of live mucosal vaccines. First, we present the progress that has recently been made in the field of developing tools for LAB genetic manipulations, which has resulted in the successful expression of many bacterial, parasitic, and viral antigens in LAB strains. Next, we discuss the factors influencing the efficacy of the constructed vaccine prototypes that have been tested in various animal models. Apart from the research focused on an application of live LABs as carriers of foreign antigens, a lot of work has been recently done on the potential usage of nonliving, nonrecombinant L. lactis designated as Gram-positive enhancer matrix (GEM), as a delivery system for mucosal vaccination. The advantages and disadvantages of both strategies are also presented.
Klaenhammer, Todd R; Azcarate-Peril, M Andrea; Altermann, Eric; Barrangou, Rodolphe
Lactic acid bacteria (LAB) are widely used for the industrial production of fermented dairy products and form a group of related low-GC-content gram-positive bacteria. The major species used in dairy manufacturing are Lactobacillus, Lactococcus, Streptococcus, and Leuconostoc. Traditionally most are applied as starter cultures for dairy fermentations or used as probiotic cultures, delivered in dairy vehicles. The appearance of the genomes of Lactococcus lactis, Bidifobacterium longum, Lactobacillus plantarum, L. johnsonii, L. acidophilus, 2 strains of Streptococcus thermophilus, and pending completion of many draft genomic sequences, is now promoting in-depth investigation into the comparative genetic content of LAB. Moreover, whole-genome transcriptional arrays are quickly revealing critical genes/operons that are coordinately expressed and the impact of environmental factors on expression of multiple gene sets. Comparative genomics between multiple genomes is providing insights into genes that are important in metabolic, physiological, and functional roles for different LAB in the environments they inhabit, ranging from the gastrointestinal tract to milk and acidified dairy products.
Hamasaki, Yoshikatsu; Ayaki, Mitsuko; Fuchu, Hidetaka; Sugiyama, Masaaki; Morita, Hidetoshi
Three kinds of lactic acid bacteria were isolated from spoiling cooked meat products stored below 10°C. They were identified as Leuconostoc mesenteroides subsp. mesenteroides, Lactococcus lactis subsp. lactis, and Leuconostoc citreum. All three strains grew well in MRS broth at 10°C. In particular, L. mesenteroides subsp. mesenteroides and L. citreum grew even at 4°C, and their doubling times were 23.6 and 51.5 h, respectively. On the other hand, although the bacteria were initially below the detection limit (<10 CFU/g) in model cooked meat products, the bacterial counts increased to 108 CFU/g at 10°C after 7 to 12 days. PMID:12788779
Tormo, Hélène; Ali Haimoud Lekhal, Djamila; Roques, C
Lactic acid bacteria, in particular Lactococcus lactis, play a decisive role in the cheese making process and more particularly in lactic cheeses which are primarily produced on goat dairy farms. The objective of this study was therefore to identify the main lactic acid bacteria found in raw goats' milk from three different regions in France and evaluate if certain farming practices have an effect on the distribution of species of lactic acid bacteria in the various milk samples. Identification at genus or species level was carried out using phenotypic tests and genotypic methods including repetitive element REP-PCR, species-specific PCR and 16S rRNA gene sequencing. The distribution of the main bacterial species in the milk samples varied depending on farms and their characteristics. Out of the 146 strains identified, L. lactis was the dominant species (60% of strains), followed by Enterococcus (38%) of which Enterococcus faecalis and Enterococcus faecium. Within the species L. lactis, L. lactis subsp lactis was detected more frequently than L. lactis subsp cremoris (74% vs. 26%). The predominance of L. lactis subsp cremoris was linked to geographical area studied. It appears that the animals' environment plays a role in the balance between the dominance of L. lactis and enterococci in raw goats' milk. The separation between the milking parlor and the goat shed (vs no separation) and only straw in the bedding (vs straw and hay) seems to promote L. lactis in the milk (vs enterococci).
Makarova, K.; Slesarev, A.; Wolf, Y.; Sorokin, A.; Mirkin, B.; Koonin, E.; Pavlov, A.; Pavlova, N.; Karamychev, V.; Polouchine, N.; Shakhova, V.; Grigoriev, I.; Lou, Y.; Rokhsar, D.; Lucas, S.; Huang, K.; Goodstein, D. M.; Hawkins, T.; Plengvidhya, V.; Welker, D.; Hughes, J.; Goh, Y.; Benson, A.; Baldwin, K.; Lee, J. -H.; Diaz-Muniz, I.; Dosti, B.; Smeianov, V; Wechter, W.; Barabote, R.; Lorca, G.; Altermann, E.; Barrangou, R.; Ganesan, B.; Xie, Y.; Rawsthorne, H.; Tamir, D.; Parker, C.; Breidt, F.; Broadbent, J.; Hutkins, R.; O'Sullivan, D.; Steele, J.; Unlu, G.; Saier, M.; Klaenhammer, T.; Richardson, P.; Kozyavkin, S.; Weimer, B.; Mills, D.
Lactic acid-producing bacteria are associated with various plant and animal niches and play a key role in the production of fermented foods and beverages. We report nine genome sequences representing the phylogenetic and functional diversity of these bacteria. The small genomes of lactic acid bacteria encode a broad repertoire of transporters for efficient carbon and nitrogen acquisition from the nutritionally rich environments they inhabit and reflect a limited range of biosynthetic capabilities that indicate both prototrophic and auxotrophic strains. Phylogenetic analyses, comparison of gene content across the group, and reconstruction of ancestral gene sets indicate a combination of extensive gene loss and key gene acquisitions via horizontal gene transfer during the coevolution of lactic acid bacteria with their habitats.
Okano, Kenji; Kimura, Sakurako; Narita, Junya; Fukuda, Hideki; Kondo, Akihiko
To achieve direct and efficient lactic acid production from starch, a genetically modified Lactococcus lactis IL 1403 secreting alpha-amylase, which was obtained from Streptococcus bovis 148, was constructed. Using this strain, the fermentation of soluble starch was achieved, although its rate was far from efficient (0.09 g l(-1) h(-1) lactate). High-performance liquid chromatography revealed that maltose accumulated during fermentation, and this was thought to lead to inefficient fermentation. To accelerate maltose consumption, starch fermentation was examined using L. lactis cells adapted to maltose instead of glucose. This led to a decrease in the amount of maltose accumulation in the culture, and, as a result, a more rapid fermentation was accomplished (1.31 g l(-1) h(-1) lactate). Maximum volumetric lactate productivity was further increased (1.57 g l(-1) h(-1) lactate) using cells adapted to starch, and a high yield of lactate (0.89 g of lactate per gram of consumed sugar) of high optical purity (99.2% of L: -lactate) was achieved. In this study, we propose a new approach to lactate production by alpha-amylase-secreting L. lactis that allows efficient fermentation from starch using cells adapted to maltose or starch before fermentation.
Vinderola, C G; Mocchiutti, P; Reinheimer, J A
Interactions among lactic acid starter and probiotic bacteria were investigated to establish adequate combinations of strains to manufacture probiotic dairy products. For this aim, a total of 48 strains of Streptococcus thermophilus, Lactobacillus delbrueckii subsp. bulgaricus, Lactococcus lactis, Lactobacillus acidophilus, Lactobacillus casei, and Bifidobacterium spp. (eight of each) were used. The detection of bacterial interactions was carried out using the well-diffusion agar assay, and the interactions found were further characterized by growth kinetics. A variety of interactions was demonstrated. Lb. delbrueckii subsp. bulgaricus was found to be able to inhibit S. thermophilus strains. Among probiotic cultures, Lb. acidophilus was the sole species that was inhibited by the others (Lb. casei and Bifidobacterium). In general, probiotic bacteria proved to be more inhibitory towards lactic acid bacteria than vice versa since the latter did not exert any effect on the growth of the former, with some exceptions. The study of interactions by growth kinetics allowed the setting of four different kinds of behaviors between species of lactic acid starter and probiotic bacteria (stimulation, delay, complete inhibition of growth, and no effects among them). The possible interactions among the strains selected to manufacture a probiotic fermented dairy product should be taken into account when choosing the best combination/s to optimize their performance in the process and their survival in the products during cold storage.
Ishibashi, Naoki; Seto, Hiromi; Koga, Shoko; Zendo, Takeshi; Sonomoto, Kenji
Lactic acid bacteria that produce Lactococcus-specific bacteriocins were isolated and identified as Lactococcus lactis from fresh corn or lettuce. Among them, four isolates were identified as lactococcin Q producers. Seven isolates showed antimicrobial activity against a lactococcin Q producer, L. lactis QU 4, as well as against nisin Z and lacticin Q producers belonging to L. lactis. Strain QU 7 was selected as a standard strain and showed no cross-immunity to lactococcin Q or other lactococcal bacteriocins. The bacteriocin produced by strain QU 7 was purified in three chromatographic steps, and its molecular mass was determined to be 5041.35 Da. The amino acid sequence analysis revealed that it is a novel class IId bacteriocin, referred to as lactococcin Z. It consisted of 45 amino acid residues. The lczA gene encoding the prepeptide of lactococcin Z showed homology to lactococcins A, B, and M. Thus, this report demonstrates a new example of Lactococcus-specific bacteriocins.
de Souza, Geany Targino; de Carvalho, Rayssa Julliane; de Sousa, Jossana Pereira; Tavares, Josean Fechine; Schaffner, Donald; de Souza, Evandro Leite; Magnani, Marciane
This study assessed the inhibitory effects of the essential oil from Origanum vulgare L. (OVEO) on Staphylococcus aureus, Listeria monocytogenes, and a mesophilic starter coculture composed of lactic acid bacteria (Lactococcus lactis subsp. lactis and L. lactis subsp. cremoris) in Brazilian coalho cheese systems. The MIC of OVEO was 2.5 μl/ml against both S. aureus and L. monocytogenes and 0.6 μl/ml against the tested starter coculture. In cheese broth containing OVEO at 0.6 μl/ml, no decrease in viable cell counts (VCC) of both pathogenic bacteria was observed, whereas the initial VCC of the starter coculture decreased approximately 1.0 log CFU/ml after 24 h of exposure at 10°C. OVEO at 1.25 and 2.5 μl/ml caused reductions of up to 2.0 and 2.5 log CFU/ml in S. aureus and L. monocytogenes, respectively, after 24 h of exposure in cheese broth. At these same concentrations, OVEO caused a greater decrease of initial VCC of the starter coculture following 4 h of exposure. Higher concentrations of OVEO were required to decrease the VCC of all target bacteria in semisolid coalho cheese slurry compared with cheese broth. The VCC of Lactococcus spp. in coalho cheese slurry containing OVEO were always lower than those of pathogenic bacteria under the same conditions. These results suggest that the concentrations of OVEO used to control pathogenic bacteria in semihard cheese should be carefully evaluated because of its inhibitory effects on the growth of starter lactic acid cultures used during the production of the product.
Terzic-Vidojevic, Amarela; Mihajlovic, Sanja; Uzelac, Gordana; Veljovic, Katarina; Tolinacki, Maja; Nikolic, Milica; Topisirovic, Ljubisa; Kojic, Milan
The aim of this study was to investigate the composition of lactic acid bacteria (LAB) in autochthonous young cheeses, sweet creams and sweet kajmaks produced in the Vlašić mountain region of central Bosnia and Herzegovina near the town of Travnik over a four season period. These three products were made from cow's milk by a traditional method without the addition of a starter culture. Preliminary characterization with phenotype-based assays and identification using rep-PCR with a (GTG)5 primer and 16S rDNA sequence analysis were undertaken for 460 LAB isolates obtained from all the examined samples. Fifteen species were identified as follows: Lactococcus lactis, Lactococcus raffinolactis, Lactococcus garviae, Lactobacillus casei, Lactobacillus plantarum, Lactobacillus helveticus, Enterococcus faecium, Enterococcus durans, Enterococcus faecalis, Enterococcus italicus, Leuconostoc mesenteroides, Leuconostoc pseudomesenteroides, Leuconostoc lactis, Streptococcus thermophilus and Streptococcus mitis. A wide genotypic and phenotypic heterogeneity of the species was observed, particularly within the Lc. lactis strains. In all of the tested dairy products across four seasons, a significantly positive correlation (r = 0.690) between the presence of lactococci and enterococci and a negative correlation (r = 0.722) between the presence of lactococci and leuconostocs were recorded. Forty-five percent of the lactobacilli and 54.4% of the lactococci exhibited proteolytic activity, whereas 18.7% of the total LAB isolates exhibited antimicrobial activity.
González, Leticia; Sacristán, Noelia; Arenas, Ricardo; Fresno, José M; Eugenia Tornadijo, M
Twenty-four strains of lactic acid bacteria (LAB) isolated from a traditional Spanish cheese (Genestoso cheese) were evaluated for their enzymatic activities (acidifying and proteolytic abilities and carboxypeptidase, aminopeptidase, dipeptidase, caseinolytic and esterase activities), in order to select indigenous strains of technical interest for the manufacture of cheese. These strains were selected on the basis of their antimicrobial activity relative to five reference strains and were identified as Lactococcus lactis subsp. lactis (thirteen strains), Leuconostoc mesenteroides (two strains), Leuconostoc pseudomesenteroides (one strain), Lactobacillus paracasei (two strains), Lactobacillus plantarum (one strain) and Enterococcus faecalis (five strains). Lactococcus strains were those that showed the greatest degree of acidifying and proteolytic activity. The cell-free extracts (CFE) of L. paracasei exhibited the highest level of aminopeptidase activity. The highest level of caseinolytic activity was shown by the CFE of one strain of L. lactis. High values were also obtained with the CFE of Lactobacillus and of several Leuconostoc. The highest level of dipeptidase activity was found amongst the strains of L. lactis. Carboxypeptidase activity was generally very low or undetectable for the majority of strains. The greatest degree of esterolytic activity was detected for Enterococcus.
Leong, Kun-hon; Chen, Yi-sheng; Pan, Shwu-fen; Chen, Jen-jye; Wu, Hui-chung; Chang, Yu-chung; Yanagida, Fujitoshi
A total of 102 lactic acid bacteria (LAB) were isolated from three different coffee farms in Taiwan. These isolates were classified and identified by the restriction fragment length polymorphism analysis and sequencing of 16S ribosomal DNA. Heterofermentative Leuconostoc, and Weissella species were the most common LAB found in two farms located at an approximate altitude of 800 m. Lactococcus lactis subsp. lactis was the most common LAB found in the remaining farm was located at an approximate altitude of 1,200 m. It is therefore suggested that the altitude and climate may affect the distribution of LAB. On the basis of phylogenetic analysis, two strains included in the genera Enterococcus were considered as two potential novel species or subspecies. In addition, a total of 34 isolates showed the antifungal activity against Aspergillus flavus. Moreover, seven Lactococcus lactis subsp. lactis strains and one Enterococcus faecalis strain were found to have bacteriocin-like inhibitory substance-producing capability. These results suggest that various LAB are associated with fresh coffee cherries in Taiwan. Some of the isolates found in this study showed potential as antifungal agents.
Cavicchioli, Valéria Quintana; Dornellas, Wesley Dos Santos; Perin, Luana Martins; Pieri, Fábio Alessandro; Franco, Bernadette Dora Gombossy de Melo; Todorov, Svetoslav Dimitrov; Nero, Luís Augusto
Lactic acid bacteria (LAB, n = 57) were previously obtained from raw goat milk, identified as Lactococcus spp. (n = 24) and Enterococcus spp. (n = 33), and characterized as bacteriocinogenic. Fingerprinting by pulsed field gel electrophoresis (PFGE) demonstrated high genetic diversity, and 30 strains were selected and exhibited strong antimicrobial activity against 46 target strains (LAB, spoilage, and foodborne pathogens). Six strains (Lactococcus lactis: GLc03 and GLc05; and Enterococcus durans: GEn09, GEn12, GEn14, and GEn17) were selected to characterize their bacteriocinogenic features, using Listeria monocytogenes ATCC 7644 as the target. The six strains produced bacteriocins at higher titer when incubated in MRS at 37 °C up to 12 h, when compared to growth at 25 and 30 °C. The produced bacteriocins kept their antimicrobial activity after exposure to 100 °C for 2 h and 121 °C for 20 min; the antimicrobial activity was also observed after treatment at pH 2.0 to 10.0, except for GLc03. L. monocytogenes populations were reduced approximately two logs after treatment with cell-free supernatants from the selected strains. These data show that goat milk can contain a diverse microbiota able to inhibit L. monocytogenes, a common pathogen found in dairy products, and can be potentially employed in biopreservation of food produced under different processing conditions.
Kuley, Esmeray; Ozogul, Fatih
The effect of lactic acid bacteria (LAB) strains on tyramine (TYR) and also other biogenic amines (BA) production by eight common food-borne pathogen (FBP) in tyrosine decarboxylase broth (TDB) was investigated by using a rapid HPLC method. Significant differences were observed among the FBP strains in ammonia (AMN) and BA production apart from tryptamine, histamine (HIS) and spermine formation (p<0.05). Salmonella paratyphi A was characterised as the main amine producer. LAB had an important synergetic role in some BA production by food-borne pathogenic bacteria, although the effect of some LAB strains on BA production was strain-dependent. Lactococcus spp. and Streptococcus spp. resulted in significantly higher TYR accumulation by Aeromonas hydrophila and Enterococcus faecalis in TDB. The presence of Lactococcus and/or Lactobacillus in TDB significantly increased HIS production by A. hydrophila, Escherichia coli, Ent. faecalis, Klebsiella pneumoniae and Pseudomonas aeruginosa, whereas HIS accumulation was significantly reduced by Staphylococcus aureus, S. paratyphi A and Listeria monocytogenes.
Minervini, Fabio; Celano, Giuseppe; Lattanzi, Anna; Tedone, Luigi; De Mastro, Giuseppe; Gobbetti, Marco; De Angelis, Maria
This study aimed at assessing the dynamics of lactic acid bacteria and other Firmicutes associated with durum wheat organs and processed products. 16S rRNA gene-based high-throughput sequencing showed that Lactobacillus, Streptococcus, Enterococcus, and Lactococcus were the main epiphytic and endophytic genera among lactic acid bacteria. Bacillus, Exiguobacterium, Paenibacillus, and Staphylococcus completed the picture of the core genus microbiome. The relative abundance of each lactic acid bacterium genus was affected by cultivars, phenological stages, other Firmicutes genera, environmental temperature, and water activity (aw) of plant organs. Lactobacilli, showing the highest sensitivity to aw, markedly decreased during milk development (Odisseo) and physiological maturity (Saragolla). At these stages, Lactobacillus was mainly replaced by Streptococcus, Lactococcus, and Enterococcus. However, a key sourdough species, Lactobacillus plantarum, was associated with plant organs during the life cycle of Odisseo and Saragolla wheat. The composition of the sourdough microbiota and the overall quality of leavened baked goods are also determined throughout the phenological stages of wheat cultivation, with variations depending on environmental and agronomic factors.
Minervini, Fabio; Celano, Giuseppe; Lattanzi, Anna; Tedone, Luigi; De Mastro, Giuseppe; De Angelis, Maria
This study aimed at assessing the dynamics of lactic acid bacteria and other Firmicutes associated with durum wheat organs and processed products. 16S rRNA gene-based high-throughput sequencing showed that Lactobacillus, Streptococcus, Enterococcus, and Lactococcus were the main epiphytic and endophytic genera among lactic acid bacteria. Bacillus, Exiguobacterium, Paenibacillus, and Staphylococcus completed the picture of the core genus microbiome. The relative abundance of each lactic acid bacterium genus was affected by cultivars, phenological stages, other Firmicutes genera, environmental temperature, and water activity (aw) of plant organs. Lactobacilli, showing the highest sensitivity to aw, markedly decreased during milk development (Odisseo) and physiological maturity (Saragolla). At these stages, Lactobacillus was mainly replaced by Streptococcus, Lactococcus, and Enterococcus. However, a key sourdough species, Lactobacillus plantarum, was associated with plant organs during the life cycle of Odisseo and Saragolla wheat. The composition of the sourdough microbiota and the overall quality of leavened baked goods are also determined throughout the phenological stages of wheat cultivation, with variations depending on environmental and agronomic factors. PMID:26187970
Bilková, Andrea; Sepova, Hana Kinová; Bilka, Frantisek; Balázová, Andrea
Lactic acid bacteria comprise several genera of gram-positive bacteria that are known for the production of structurally different antimicrobial substances. Among them, bacteriocins are nowadays in the centre of scientific interest. Bacteriocins, proteinaceous antimicrobial substances, are produced ribosomally and have usually a narrow spectrum of bacterial growth inhibition. According to their structure and the target of their activity, they are divided into four classes, although there are some suggestions for a renewed classification. The most interesting and usable class are lantibiotics. They comprise the most widely commercially used and well examined bacteriocin, nisin. The non-pathogenic character of lactic acid bacteria is advantageous for using their bacteriocins in food preservation as well as in feed supplements or in veterinary medicine.
Johansson, Per; Laine, Pia; Smolander, Olli-Pekka; Sonck, Matti; Rahkila, Riitta; Jääskeläinen, Elina; Paulin, Lars; Auvinen, Petri; Björkroth, Johanna
Lactococcus piscium is a psychrotrophic lactic acid bacterium and is known to be one of the predominant species within spoilage microbial communities in cold-stored packaged foods, particularly in meat products. Its presence in such products has been associated with the formation of buttery and sour off-odors. Nevertheless, the spoilage potential of L. piscium varies dramatically depending on the strain and growth conditions. Additional knowledge about the genome is required to explain such variation, understand its phylogeny, and study gene functions. Here, we present the complete and annotated genomic sequence of L. piscium MKFS47, combined with a time course analysis of the glucose catabolism-based transcriptome. In addition, a comparative analysis of gene contents was done for L. piscium MKFS47 and 29 other lactococci, revealing three distinct clades within the genus. The genome of L. piscium MKFS47 consists of one chromosome, carrying 2,289 genes, and two plasmids. A wide range of carbohydrates was predicted to be fermented, and growth on glycerol was observed. Both carbohydrate and glycerol catabolic pathways were significantly upregulated in the course of time as a result of glucose exhaustion. At the same time, differential expression of the pyruvate utilization pathways, implicated in the formation of spoilage substances, switched the metabolism toward a heterofermentative mode. In agreement with data from previous inoculation studies, L. piscium MKFS47 was identified as an efficient producer of buttery-odor compounds under aerobic conditions. Finally, genes and pathways that may contribute to increased survival in meat environments were considered. PMID:25819958
The Gram-positive Lactic acid bacteria (LAB) are considered attractive biocatalysts for biomass to biofuels for several reasons. They have GRAS (Generally Recognized As Safe) status that are acceptable in food, feed, and medical applications. LAB are fermentative: selected strains are capable of f...
Mendoza, Gabriela Montel; Pasteris, Sergio E; Ale, Cesar E; Otero, María C; Bühler, Marta I; Nader-Macías, María E Fátima
The cultivable microbiota of skin and cloaca of captive Lithobates catesbeianus includes microorganisms generally accepted as beneficial and potentially pathogenic bacteria. In order to select a group of potentially probiotic bacteria, 136 isolates were evaluated for their surface properties and production of antagonistic metabolites. Then, 11 lactic acid bacteria (LAB) strains were selected and identified as Lactobacillus plantarum, Lb. brevis, Pediococcus pentosaceus, Lactococcus lactis, L. garvieae and Enterococcus gallinarum. Studies of compatibility indicate that all the strains could be included in a multi-strain probiotic, with the exception of Ent. gallinarum CRL 1826 which inhibited LAB species through a bacteriocin-like metabolite. These results contribute to the design of a probiotic product to improve the sanitary status of bullfrogs in intensive culture systems, to avoid the use of antibiotics and thus to reduce production costs. It could also be an alternative to prevent infectious diseases during the ex situ breeding of amphibian species under threat of extinction.
Michel, Christian; Pelletier, Claire; Boussaha, Mekki; Douet, Diane-Gaëlle; Lautraite, Armand; Tailliez, Patrick
Lactic acid bacteria have become a major source of concern for aquaculture in recent decades. In addition to true pathogenic species of worldwide significance, such as Streptococcus iniae and Lactococcus garvieae, several species have been reported to produce occasional fish mortalities in limited geographic areas, and many unidentifiable or ill-defined isolates are regularly isolated from fish or fish products. To clarify the nature and prevalence of different fish-associated bacteria belonging to the lactic acid bacterium group, a collection of 57 isolates of different origins was studied and compared with a set of 22 type strains, using amplified rRNA gene restriction analysis (ARDRA). Twelve distinct clusters were delineated on the basis of ARDRA profiles and were confirmed by sequencing of sodA and 16S rRNA genes. These clusters included the following: Lactococcus raffinolactis, L. garvieae, Lactococcus l., S. iniae, S. dysgalactiae, S. parauberis, S. agalactiae, Carnobacterium spp., the Enterococcus “faecium” group, a heterogeneous Enterococcus-like cluster comprising indiscernible representatives of Vagococcus fluvialis or the recently recognized V. carniphilus, V. salmoninarum, and Aerococcus spp. Interestingly, the L. lactis and L. raffinolactis clusters appeared to include many commensals of fish, so opportunistic infections caused by these species cannot be disregarded. The significance for fish populations and fish food processing of three or four genetic clusters of uncertain or complex definition, namely, Aerococcus and Enterococcus clusters, should be established more accurately. PMID:17337536
Ho, Chun-Hoong; Stanton-Cook, Mitchell; Beatson, Scott A; Bansal, Nidhi; Turner, Mark S
Lactococcus lactis is a starter bacterium commonly used in cheese making where it has an important role in acid-mediated curd formation as well as the development of flavour compounds. Industrial L. lactis strains can harbour one or more inducible prophages which when induced can affect cell growth and possibly lead to cell lysis. This is undesirable during growth and fermentation, but can beneficially lead to faster release of enzymes during cheese ripening. Lactococci can encounter multiple stress inducing conditions during the production of cheese, such as low and high temperatures, low pH, high osmotic pressure and long-term incubation. In this study, we tested the effect of these industrial stressors on prophage induction in two cheese making L. lactis subsp. cremoris strains (ASCC890049 and ASCC890310) as well as the laboratory strain L. lactis MG1363. Firstly, in order to identify inducible prophages in these strains we exposed them to the prophage inducing chemical mitomycin C (MMC) for 1 and 2h and then subjected the total genomic DNA to next-generation Illumina sequencing. Mapping of sequence reads back to the genome sequences revealed regions which contained a much higher fold coverage indicating DNA replication. These regions were amplified by up to 332-fold per cell (relative to the control tufA gene) and were identified as having similarities to different subgroups of P335 phages including MG-5, TP901-1, ul36.k1, bIL286, TP712 and BK5-T. Next, quantitative PCR was used to confirm the strong induction of prophages by MMC and then determine the copy number of the inducible prophages following exposure to various growth inhibitory levels of HCl, lactic acid, high temperature, NaCl, hydrogen peroxide and bacitracin. With the exception of a slight induction (2 to 4-fold) with hydrogen peroxide and long-term incubation after 21days in one industrial strain, none of the other stressors induced prophage DNA replication. These findings show that the repression
Balcázar, José Luis; de Blas, Ignacio; Ruiz-Zarzuela, Imanol; Vendrell, Daniel; Gironés, Olivia; Muzquiz, José Luis
The aim of this study was to identify lactic acid bacteria (LAB) using polymerase chain reaction (PCR) amplification of variable regions of the 16S rRNA gene. Thirteen LAB strains were isolated from the intestinal microbiota of healthy salmonids. A approximately 500-bp region of the highly conserved 16S rRNA gene was PCR-amplified and following this, a portion of the amplicon (272-bp) including the V1 and V2 variable regions was sequenced. The sequence containing both the V1 and V2 region provided strong evidence for the identification of LAB. The LAB strains were identified as Carnobacterium maltaromaticum, Lactobacillus curvatus, Lactobacillus sakei, Lactobacillus plantarum, Lactococcus lactis subsp. cremoris, Lactococcus lactis subsp. lactis, and Leuconostoc mesenteroides. The method described was found to be a very simple, rapid, specific, and low-cost tool for the identification of unknown strains of LAB.
Wu, Chongde; Huang, Jun; Zhou, Rongqing
Lactic acid bacteria (LAB) are widely used for the production of a variety of fermented foods, and are considered as probiotic due to their health-promoting effect. However, LAB encounter various environmental stresses both in industrial fermentation and application, among which acid stress is one of the most important survival challenges. Improving the acid stress resistance may contribute to the application and function of probiotic action to the host. Recently, the advent of genomics, functional genomics and high-throughput technologies have allowed for the understanding of acid tolerance mechanisms at a systems level, and many method to improve acid tolerance have been developed. This review describes the current progress in engineering acid stress resistance of LAB. Special emphasis is placed on engineering cellular microenvironment (engineering amino acid metabolism, introduction of exogenous biosynthetic capacity, and overproduction of stress response proteins) and maintaining cell membrane functionality. Moreover, strategies to improve acid tolerance and the related physiological mechanisms are also discussed.
Muyanja, C M B K; Narvhus, J A; Treimo, J; Langsrud, T
One hundred and thirteen strains of lactic acid bacteria (LAB) were selected from 351 isolates from 15 samples of traditionally fermented household bushera from Uganda and also from laboratory-prepared bushera. Isolates were phenotypically characterised by their ability to ferment 49 carbohydrates using API 50 CHL kits and additional biochemical tests. Coliforms, yeasts and LAB were enumerated in bushera. The pH, volatile organic compounds and organic acids were also determined. The LAB counts in household bushera varied between 7.1 and 9.4 log cfu ml(-1). The coliform counts varied between < 1 and 5.2 log cfu ml(-1). The pH of bushera ranged from 3.7 to 4.5. Ethanol (max, 0.27%) was the major volatile organic compound while lactic acid (max, 0.52%) was identified as the dominant organic acid in household bushera. The initial numbers of LAB and coliforms in laboratory-fermented bushera were similar; however, the LAB numbers increased faster during the first 24 h. LAB counts increased from 5.5 to 9.0 log cfu ml(-1) during the laboratory fermentation. Coliform counts increased from 5.9 to 7.8 log cfu ml(-1) at 24 h, but after 48 h, counts were less 4 log cfu ml(-1). Yeasts increased from 4.3 to 7.7 log cfu ml(-1) at 48 h, but thereafter decreased slightly. The pH declined from 7.0 to around 4.0. Lactic acid and ethanol increased from zero to 0.75% and 0.20%, respectively. Lactic acid bacteria isolated from household bushera belonged to Lactobacillus, Streptococcus and Enterococcus genera. Tentatively, Lactobacillus isolates were identified as Lactobacillus plantarum, L. paracasei subsp. paracasei, L. fermentum, L. brevis and L. delbrueckii subsp. delbrueckii. Streptococcus thermophilus strains were also identified in household bushera. LAB isolated from bushera produced in the laboratory belonged to five genera (Lactococcus, Leuconostoc, Lactobacillus, Weissella and Enterococcus. Eight isolates were able to produce acid from starch and were identified as Lactococcus
Dedysh, S N
Acid sphagnum bogs cover a considerable part of the territory of Russia and are an important natural source of biogenic methane, which is formed in their anaerobic layers. A considerable portion of this methane is consumed in the aerobic part of the bog profile by acidophilic methanotrophic bacteria, which comprise the methane filter of sphagnum bogs and decrease CH4 emission to the atmosphere. For a long time, these bacteria escaped isolation, which became possible only after the elucidation of the optimal conditions of their functioning in situ: pH 4.5 to 5.5; temperature, from 15 to 20 degrees C; and low salt concentration in the solution. Reproduction of these conditions and rejection of earlier used media with a high content of biogenic elements allowed methanotrophic bacteria of two new genera and species--Methylocella palustris and Methylocapsa acidophila--to be isolated from the peat of sphagnum bogs of the northern part of European Russia and West Siberia. These bacteria are well adapted to the conditions in cold, acid, oligotrophic sphagnum bogs. They grow in a pH range of 4.2-7.5 with an optimum at 5.0-5.5, prefer moderate temperatures (15-25 degrees C) and media with a low content of mineral salts (200-500 mg/l), and are capable of active nitrogen fixation. Design of fluorescently labeled 16S rRNA-targeted oligonucleotide probes for the detection of Methylocella palustris and Methylocapsa acidophila and their application to the analysis of sphagnum peat samples showed that these bacteria represent dominant populations of methanotrophs with a density of 10(5)-10(6) cells/g peat. In addition to Methylocella and Methylocapsa populations, one more abundant population of methanotrophs was revealed (10(6) cells/g peat), which were phylogenetically close to the genus Methylocystis.
Leroi, Françoise; Fall, Papa Abdoulaye; Pilet, Marie France; Chevalier, Frédérique; Baron, Régis
The maximum specific growth rate (μ(max)) of Brochothrix thermosphacta, a spoilage bacteria of cooked peeled shrimp, and Lactococcus piscium CNCM I-4031, a bioprotective strain, was investigated under different conditions of temperature, NaCl concentrations and pH. The basic modelling approach used was the Gamma concept (γ-concept) and the model developed was then adapted to shrimp. Cardinal growth parameters were quite similar for the two strains, except for NaCl. No NaCl was required for growth and the NaCl(max) was three-times higher for B. thermosphacta than for L. piscium (62 and 23 g l(-1) respectively). However, tolerance to NaCl was higher in seafood than in liquid broth, possibly due to presence of osmoltically active molecules. L. piscium and B. thermosphacta were psychrotolerant, with T(min) = -4.8 and -3.4 °C, T(opt) = 23.4 and 27.0 °C and T(max) = 27.2 and 30.8 °C respectively. The optimal pH was neutral and growth possible till pH = 4.8 for the two strains, assuming possible applications of the bioprotective strain in lightly marinated seafood. The μ(max) of B. thermosphacta in shrimp was a little higher than in L. piscium whatever the environmental conditions. Validation of the model showed that the γ-concept was suitable for predicting μ(max) of B. thermosphacta in shrimp. Data generated in this study can be used to adapt the model to other foods with few additional experiments and the effect of different parameters may be added in the future. The model was less accurate for the bioprotective strain and the effect of NaCl must be studied in more detail directly in the matrix.
Ghahremani, Enayat; Mardani, Mahnaz; Rezapour, Sadegh
Lactic acid bacteria (LAB) with proteolitic activity are used as aromatic and antibacterial substances, cholesterol reduces, bile salt hydrolyses, and probiotic. The aims of this project were to isolate and identify natural LAB flora involved in traditional fermentation in cheeses of Khoramabad city and also to survey their probiotic potential. In order to achieve this goal, LAB were isolated and characterized using phenotypic and genotypic methods (PCR-sequencing); in the next stage, they were analyzed lowering cholesterol medium, hydrolysis of the bile, resistance to bile-resistant PH acidic stomach. At the end of the study, 88 cocci and 3 bacill were found: 58 Enterococcus faecium, 16 Enterococcus hirae, 5 Lactococcus lactis, 3 Lactobacillus plantarum, and 9 undetermined. The probiotic results of the bacteria had effects on the reduction of cholesterol, resistance to stomach acid, had relative antibacterial effects, and some strains had effects on hydrolyzing the bile. For further identification, the PCR method and the application of 16s-DNA-ITS genes and its sequencing were found useful. This study showed that lactic acid bacteria in the traditional cheese of the Khorramabad city have relative probiotic effect and that these lactic acid bacteria in fermented milk are suitable.
Drinan, D F; Robin, S; Cogan, T M
The effect of citrate on production of diacetyl and acetoin by four strains each of heterofermentative and homofermentative lactic acid bacteria capable of utilizing citrate was studied. Acetoin was quantitatively the more important compound. The heterofermentative bacteria produced no acetoin or diacetyl in the absence of citrate, and two strains produced traces of acetoin in its presence. Citrate stimulated the growth rate of the heterofermentative lactobacilli. Acidification of all heterofermentative cultures with citric acid resulted in acetoin production. Destruction of accumulated acetoin appeared to coincide with the disappearance of citrate. All homofermentative bacteria produced more acetoin and diacetyl in the presence of citrate than in its absence. Citrate utilization was begun immediately by the streptococci but was delayed until at least the middle of the exponential phase in the case of the lactobacilli. PMID:5054
Franciosi, Elena; Carafa, Ilaria; Nardin, Tiziana; Schiavon, Silvia; Poznanski, Elisa; Cavazza, Agostino; Larcher, Roberto; Tuohy, Kieran M
"Nostrano-cheeses" are traditional alpine cheeses made from raw cow's milk in Trentino-Alto Adige, Italy. This study identified lactic acid bacteria (LAB) developing during maturation of "Nostrano-cheeses" and evaluated their potential to produce γ-aminobutyric acid (GABA), an immunologically active compound and neurotransmitter. Cheese samples were collected on six cheese-making days, in three dairy factories located in different areas of Trentino and at different stages of cheese ripening (24 h, 15 days, and 1, 2, 3, 6, and 8 months). A total of 1,059 LAB isolates were screened using Random Amplified Polymorphic DNA-PCR (RAPD-PCR) and differentiated into 583 clusters. LAB strains from dominant clusters (n = 97) were genetically identified to species level by partial 16S rRNA gene sequencing. LAB species most frequently isolated were Lactobacillus paracasei, Streptococcus thermophilus, and Leuconostoc mesenteroides. The 97 dominant clusters were also characterized for their ability in producing GABA by high-performance liquid chromatography (HPLC). About 71% of the dominant bacteria clusters evolving during cheeses ripening were able to produce GABA. Most GABA producers were Lactobacillus paracasei but other GABA producing species included Lactococcus lactis, Lactobacillus plantarum, Lactobacillus rhamnosus, Pediococcus pentosaceus, and Streptococcus thermophilus. No Enterococcus faecalis or Sc. macedonicus isolates produced GABA. The isolate producing the highest amount of GABA (80.0±2.7 mg/kg) was a Sc. thermophilus.
Nardin, Tiziana; Schiavon, Silvia; Cavazza, Agostino; Larcher, Roberto; Tuohy, Kieran M.
“Nostrano-cheeses” are traditional alpine cheeses made from raw cow's milk in Trentino-Alto Adige, Italy. This study identified lactic acid bacteria (LAB) developing during maturation of “Nostrano-cheeses” and evaluated their potential to produce γ-aminobutyric acid (GABA), an immunologically active compound and neurotransmitter. Cheese samples were collected on six cheese-making days, in three dairy factories located in different areas of Trentino and at different stages of cheese ripening (24 h, 15 days, and 1, 2, 3, 6, and 8 months). A total of 1,059 LAB isolates were screened using Random Amplified Polymorphic DNA-PCR (RAPD-PCR) and differentiated into 583 clusters. LAB strains from dominant clusters (n = 97) were genetically identified to species level by partial 16S rRNA gene sequencing. LAB species most frequently isolated were Lactobacillus paracasei, Streptococcus thermophilus, and Leuconostoc mesenteroides. The 97 dominant clusters were also characterized for their ability in producing GABA by high-performance liquid chromatography (HPLC). About 71% of the dominant bacteria clusters evolving during cheeses ripening were able to produce GABA. Most GABA producers were Lactobacillus paracasei but other GABA producing species included Lactococcus lactis, Lactobacillus plantarum, Lactobacillus rhamnosus, Pediococcus pentosaceus, and Streptococcus thermophilus. No Enterococcus faecalis or Sc. macedonicus isolates produced GABA. The isolate producing the highest amount of GABA (80.0±2.7 mg/kg) was a Sc. thermophilus. PMID:25802859
Grosu-Tudor, Silvia-Simona; Stancu, Mihaela-Marilena; Pelinescu, Diana; Zamfir, Medana
Lactic acid bacteria (LAB) isolated from different sources (dairy products, fruits, fresh and fermented vegetables, fermented cereals) were screened for antimicrobial activity against other bacteria, including potential pathogens and food spoiling bacteria. Six strains have been shown to produce bacteriocins: Lactococcus lactis 19.3, Lactobacillus plantarum 26.1, Enterococcus durans 41.2, isolated from dairy products and Lactobacillus amylolyticus P40 and P50, and Lactobacillus oris P49, isolated from bors. Among the six bacteriocins, there were both heat stable, low molecular mass polypeptides, with a broad inhibitory spectrum, probably belonging to class II bacteriocins, and heat labile, high molecular mass proteins, with a very narrow inhibitory spectrum, most probably belonging to class III bacteriocins. A synergistic effect of some bacteriocins mixtures was observed. We can conclude that fermented foods are still important sources of new functional LAB. Among the six characterized bacteriocins, there might be some novel compounds with interesting features. Moreover, the bacteriocin-producing strains isolated in our study may find applications as protective cultures.
Hegazi, F Z; Abo-Elnaga, I G
One hundred and twelve different strains of lactic acid bacteria, belonging to the genera Leuconostoc, Streptococcus, and Lactobacillus, were examined for the ability to degrade 10 organic acids by detecting gas production, using the conventional Durham tube method. All the strains did not break down succinate, glutarate, 2-oxo-glutarate, and mucate. Malate, citrate, pyruvate, fumarate, tartrate, and gluconate were variably attacked. Streptococcus cremoiris AM2, ML8, and SK11 required glucose to produce gas from citrate, whereas Leuconostoc citrovorum and Streptococcus faecalis did not. Streptococcus cremoris differed from the other streptococci in not producing gas from gluconate. From all lactic acid bacteria examined, only Lactobacillus plantarum formed gas from tartarate. Determination of acetoin and diacetyl proved to be a more reliable evidence for assessing the degradation of pyruvate, compared with detection of gas production. Homofermentative lactobacilli and Leuconostoc citrovorum produced acetoin and diacetyl from pyruvate, whereas beta-bacteria did not, a character that would be of taxonomic value. Streptobacteria degraded pyruvate in the presence of glucose with lactate as the major product together with a mean acetate of 4.1%, ethanol 7.9%, acetoin 1.7%, and diacetyl 2.6% yield on a molar basis after 60 days at 30 degrees C. L. brevis produced acetate and lactate. Formation of diacetyl from pyruvate by lactic acid bacteria may play an important role in flavour development in fermenting dairy products, especially in cheese, where lactic acid bacteria usually predominate, and pyruvate is probably excreted in the breaking down of lactose and in the oxidative deamination of alanine by the accompanying microflora.
González-Arenzana, Lucía; López, Rosa; Santamaría, Pilar; López-Alfaro, Isabel
Lactic acid bacteria populations of red wine samples from industrial fermentations, including two different vinification methods were studied. For this investigation, polymerase chain reaction-denaturing gradient gel electrophoresis (PCR-DGGE) analysis was employed to supplement previous results that were obtained by culture-dependent methods. PCR-DGGE was aimed to study two targeted genes, 16S ribosomal DNA (rDNA) and rpoB, and the results were useful to evaluate the microbial populations in wine samples. Moreover, an improvement of a detection limit determined so far for DGGE analysis was obtained with the method described in this study, what made possible to identify lactic acid bacteria populations below 10(1) colony-forming unit/mL. The species Oenococcus oeni was the most frequently detected bacterium, but identifications close to species Oenococcus kitaharae and Lactococcus lactis that are not often found in wine were firstly identified in samples of this research. PCR-DGGE allowed to detect 9 out of 11 lactic acid bacteria species identified in this study (nine by PCR-16S rDNA/DGGE and four by PCR-rpoB/DGGE), while five species were detected using the modified de Man, Rogosa and Sharpe agar. Therefore, the two methods were demonstrated to be complementary. This finding suggests that analysis of the lactic acid bacteria population structure in wine should be carried out using both culture-dependent and culture-independent techniques with more than one primer pair.
Kumari, Archana; Akkoç, Nefise; Akçelik, Mustafa
Lactic acid bacteria (LAB) are possessing ability to synthesize antimicrobial compounds (like bacteriocin) during their growth. In this regard, novel bacteriocin compound secreting capability of LAB isolated from Tulum Cheese in Turkey was demonstrated. The synthesized bacteriocin was purified by ammonium sulphate precipitation, dialysis and gel filtration. The molecular weight (≈3.4 kDa) of obtained bacteriocin was confirmed by SDS-PAGE, which revealed single peptide band. Molecular identification of LAB strain isolated from Tulum Cheese was conducted using 16S rDNA gene sequencing as Lactococcus lactis ssp. lactis LL171. The amino acid sequences (KKIDTRTGKTMEKTEKKIELSLKNMKTAT) of the bacteriocin from Lactococcus lactis ssp. lactis LL171 was found unique and novel than reported bacteriocins. Further, the bacteriocin was possessed the thermostable property and active at wide range of pH values from 1 to 11. Thus, bacteriocin reported in this study has the potential applications property as food preservative agent.
Simova, Emilina; Simov, Zhelyasko; Beshkova, Dora; Frengova, Ginka; Dimitrov, Zhechko; Spasov, Zdravko
The characteristics of cell growth, lactic acid production, amino acid release and consumption by single-strain cultures of lactic acid bacteria (isolated from kefir grains), and by a multiple-strain kefir starter prepared from them, were studied. The change in the levels of free amino acids was followed throughout the kefir process: single-strain kefir bacteria and the kefir starter (Lactococcus lactis C15-1%+Lactobacillus helveticus MP12-3%+(Streptococcus thermophilus T15+Lactobacillus bulgaricus HP1 = 1:1)-3%) were cultivated in pasteurized (92 degrees C for 20 min) cow's milk (3% fat content) at 28 degrees C for 5 h (the kefir starter reached pH 4.7) and subsequently grown at 20 degrees C for 16 h; storage was at 4 degrees C for 168 h. The strain L. helveticus MP12 was unrivaled with respect to free amino acid production (53.38 mg (100 g)(-1)) and cell growth (17.8 x 10(8) CFU ml(-1)); however, it manifested the lowest acidification activity. L. bulgaricus HP1 released approximately 3.7 times less amino acids, nearly 5 times lower cell growth, and produced about 1.2 times more lactic acid. S. thermophilus T15 demonstrated dramatically complex amino acid necessities for growth and metabolism. With L. lactis C15, the highest levels of growth and lactic acid synthesis were recorded (18.3 x 10(8) CFU ml(-1) and 7.8 g l(-1) lactic acid at the 21st hour), and as for free amino acid production, it approximated L. bulgaricus HP1 (17.03 mg (100 g)(-1) maximum concentration). In the L. lactis C15 culture, the amino acids were used more actively throughout the first exponential growth phase (by the 10th hour) than during the second growth phase. The unique properties of the L. helveticus MP12 strain to produce amino acids were employed to create a symbiotic bioconsortium kefir culture, which, under conditions of kefir formation, enhanced lactic acid production and shortened the time required to reach pH 4.7; intensified cell growth activity, resulting in a respective 90
Ho, Van Thi Thuy; Zhao, Jian; Fleet, Graham
Cocoa beans (Theobroma cacao L.) are the raw material for chocolate production. Fermentation of cocoa pulp by microorganisms is crucial for developing chocolate flavor precursors. Yeasts conduct an alcoholic fermentation within the bean pulp that is essential for the production of good quality beans, giving typical chocolate characters. However, the roles of bacteria such as lactic acid bacteria and acetic acid bacteria in contributing to the quality of cocoa bean and chocolate are not fully understood. Using controlled laboratory fermentations, this study investigated the contribution of lactic acid bacteria to cocoa bean fermentation. Cocoa beans were fermented under conditions where the growth of lactic acid bacteria was restricted by the use of nisin and lysozyme. The resultant microbial ecology, chemistry and chocolate quality of beans from these fermentations were compared with those of indigenous (control) fermentations. The yeasts Hanseniaspora guilliermondii, Pichia kudriavzevii, Kluyveromyces marxianus and Saccharomyces cerevisiae, the lactic acid bacteria Lactobacillus plantarum, Lactobacillus pentosus and Lactobacillus fermentum and the acetic acid bacteria Acetobacter pasteurianus and Gluconobacter frateurii were the major species found in control fermentations. In fermentations with the presence of nisin and lysozyme, the same species of yeasts and acetic acid bacteria grew but the growth of lactic acid bacteria was prevented or restricted. These beans underwent characteristic alcoholic fermentation where the utilization of sugars and the production of ethanol, organic acids and volatile compounds in the bean pulp and nibs were similar for beans fermented in the presence of lactic acid bacteria. Lactic acid was produced during both fermentations but more so when lactic acid bacteria grew. Beans fermented in the presence or absence of lactic acid bacteria were fully fermented, had similar shell weights and gave acceptable chocolates with no differences
Alfonzo, Antonio; Miceli, Claudia; Nasca, Anna; Franciosi, Elena; Ventimiglia, Giusi; Di Gerlando, Rosalia; Tuohy, Kieran; Francesca, Nicola; Moschetti, Giancarlo; Settanni, Luca
The present work was carried out to retrieve the origin of lactic acid bacteria (LAB) in sourdough. To this purpose, wheat LAB were monitored from ear harvest until the first step of fermentation for sourdough development. The influence of the geographical area and variety on LAB species/strain composition was also determined. The ears of four Triticum durum varieties (Duilio, Iride, Saragolla and Simeto) were collected from several fields located within the Palermo province (Sicily, Italy) and microbiologically investigated. In order to trace the transfer of LAB during the consecutive steps of manipulation, ears were transformed aseptically and, after threshing, milling and fermentation, samples of kernels, semolinas and doughs, respectively, were analysed. LAB were not found to dominate the microbial communities of the raw materials. In general, kernels harboured lower levels of microorganisms than ears and ears than semolinas. Several samples showing no development of LAB colonies acidified the enrichment broth suggesting the presence of LAB below the detection limit. After fermentation, LAB loads increased consistently for all doughs, reaching levels of 7.0-7.5 Log CFU/g on M17. The values of pH (5.0) and TTA (5.6 mL NaOH/10 g of dough) indicated the occurrence of the acidification process for several doughs. LAB were phenotypically and genotypically differentiated by randomly amplified polymorphic DNA (RAPD)-PCR into eight groups including 51 strains belonging to the species Lactobacillus brevis, Lactobacillus coryniformis, Lactobacillus plantarum, Lactococcus lactis, Lactococcus garvieae, Enterococcus casseliflavus, Enterococcus faecium, Leuconostoc citreum, and Pediococcus pentosaceus. Lactobacilli constituted a minority the LAB community, while lactococci represented more than 50% of strains. Lower LAB complexity was found on kernels, while a richer biodiversity was observed in semolinas and fermented doughs. For broader microbiota characterisation in
Nami, Yousef; Abdullah, Norhafizah; Haghshenas, Babak; Radiah, Dayang; Rosli, Rozita; Khosroushahi, Ahmad Yari
Forty-five lactic acid bacteria (LAB) were isolated from the vaginal specimens of healthy fertile women, and the identities of the bacteria were confirmed by sequencing of their 16S rDNA genes. Among these bacteria, only four isolates were able to resist and survive in low pH, bile salts and simulated in vitro digestion conditions. Lactococcus lactis 2HL, Enterococcus durans 6HL, Lactobacillus acidophilus 36YL and Lactobacillus plantarum 5BL showed the best resistance to these conditions. These strains were evaluated further to assess their ability to adhere to human intestinal Caco-2 cells. Lactococcus lactis 2HL and E. durans 6HL were the most adherent strains. In vitro tests under neutralized pH proved the antimicrobial activity of both strains. Results revealed that the growth of Escherichia coli O26, Staphylococcus aureus and Shigella flexneri was suppressed by both LAB strains. The antibiotic susceptibility tests showed that these strains were sensitive to all nine antibiotics: vancomycin, tetracycline, ampicillin, penicillin, gentamicin, erythromycin, clindamycin, sulfamethoxazole and chloramphenicol. These data suggest that E. durans 6HL and Lactococcus lactis 2HL could be examined further for their useful properties and could be developed as new probiotics.
Šalomskienė, Joana; Abraitienė, Asta; Jonkuvienė, Dovilė; Mačionienė, Irena; Repečkienė, Jūratė
The aim of the study was a detail evaluation of genetic diversity among the lactic acid bacteria (LAB) strains having an advantage of a starter culture in order to select genotypically diverse strains with enhanced antimicrobial effect on some harmfull and pathogenic microorganisms. Antimicrobial activity of LAB was performed by the agar well diffusion method and was examined against the reference strains and foodborne isolates of Bacillus cereus, Listeria monocytogenes, Escherichia coli, Staphylococcus aureus and Salmonella Typhimurium. Antifungal activity was tested against the foodborne isolates of Candida parapsilosis, Debaromyces hansenii, Kluyveromyces marxianus, Pichia guilliermondii, Yarowia lipolytica, Aspergillus brasiliensis, Aspergillus versicolor, Cladosporium herbarum, Penicillium chrysogenum and Scopulariopsis brevicaulis. A total 40 LAB strains representing Lactobacillus (23 strains), Lactococcus (13 strains) and Streptococcus spp. (4 strains) were characterised by repetitive sequence based polymerase chain reaction fingerprinting which generated highly discriminatory profiles, confirmed the identity and revealed high genotypic heterogeneity among the strains. Many of tested LAB demonstrated strong antimicrobial activity specialised against one or few indicator strains. Twelve LAB strains were superior in suppressing growth of the whole complex of pathogenic bacteria and fungi. These results demonstrated that separate taxonomic units offered different possibilities of selection for novel LAB strains could be used as starter cultures enhancing food preservation.
Volzing, Katherine; Borrero, Juan; Sadowsky, Michael J.; Kaznessis, Yiannis N.
We present results of tests with recombinant Lactococcus lactis that produce and secrete heterologous antimicrobial peptides with activity against Gram-negative pathogenic Escherichia coli and Salmonella. In an initial screening, the activities of numerous candidate antimicrobial peptides, made by solid state synthesis, were assessed against several indicator pathogenic E. coli and Salmonella strains. Peptides A3APO and Alyteserin were selected as top performers based on high antimicrobial activity against the pathogens tested and on significantly lower antimicrobial activity against L. lactis. Expression cassettes containing the signal peptide of the protein Usp45 fused to the codon optimized sequence of mature A3APO and Alyteserin were cloned under the control of a nisin-inducible promoter nisA and transformed into L. lactis IL1403. The resulting recombinant strains were induced to express and secrete both peptides. A3APO- and Alyteserin-containing supernatants from these recombinant L. lactis inhibited the growth of pathogenic E. coli and Salmonella by up to 20-fold, while maintaining the host’s viability. This system may serve as a model for the production and delivery of antimicrobial peptides by lactic acid bacteria to target Gram-negative pathogenic bacteria populations. PMID:23808914
Rodríguez, Héctor; Curiel, José Antonio; Landete, José María; de las Rivas, Blanca; López de Felipe, Félix; Gómez-Cordovés, Carmen; Mancheño, José Miguel; Muñoz, Rosario
Phenolic compounds are important constituents of food products of plant origin. These compounds are directly related to sensory characteristics of foods such as flavour, astringency, and colour. In addition, the presence of phenolic compounds on the diet is beneficial to health due to their chemopreventive activities against carcinogenesis and mutagenesis, mainly due to their antioxidant activities. Lactic acid bacteria (LAB) are autochthonous microbiota of raw vegetables. To get desirable properties on fermented plant-derived food products, LAB has to be adapted to the characteristics of the plant raw materials where phenolic compounds are abundant. Lactobacillus plantarum is the commercial starter most frequently used in the fermentation of food products of plant origin. However, scarce information is still available on the influence of phenolic compounds on the growth and viability of L. plantarum and other LAB species. Moreover, metabolic pathways of biosynthesis or degradation of phenolic compounds in LAB have not been completely described. Results obtained in L. plantarum showed that L. plantarum was able to degrade some food phenolic compounds giving compounds influencing food aroma as well as compounds presenting increased antioxidant activity. Recently, several L. plantarum proteins involved in the metabolism of phenolic compounds have been genetically and biochemically characterized. The aim of this review is to give a complete and updated overview of the current knowledge among LAB and food phenolics interaction, which could facilitate the possible application of selected bacteria or their enzymes in the elaboration of food products with improved characteristics.
Carvalho, Ana Lúcia; Turner, David L; Fonseca, Luís L; Solopova, Ana; Catarino, Teresa; Kuipers, Oscar P; Voit, Eberhard O; Neves, Ana Rute; Santos, Helena
The effect of pH on the glucose metabolism of non-growing cells of L. lactis MG1363 was studied by in vivo NMR in the range 4.8 to 6.5. Immediate pH effects on glucose transporters and/or enzyme activities were distinguished from transcriptional/translational effects by using cells grown at the optimal pH of 6.5 or pre-adjusted to low pH by growth at 5.1. In cells grown at pH 5.1, glucose metabolism proceeds at a rate 35% higher than in non-adjusted cells at the same pH. Besides the upregulation of stress-related genes (such as dnaK and groEL), cells adjusted to low pH overexpressed H(+)-ATPase subunits as well as glycolytic genes. At sub-optimal pHs, the total intracellular pool of lactic acid reached approximately 500 mM in cells grown at optimal pH and about 700 mM in cells grown at pH 5.1. These high levels, together with good pH homeostasis (internal pH always above 6), imply intracellular accumulation of the ionized form of lactic acid (lactate anion), and the concomitant export of the equivalent protons. The average number, n, of protons exported with each lactate anion was determined directly from the kinetics of accumulation of intra- and extracellular lactic acid as monitored online by (13)C-NMR. In cells non-adjusted to low pH, n varies between 2 and 1 during glucose consumption, suggesting an inhibitory effect of intracellular lactate on proton export. We confirmed that extracellular lactate did not affect the lactate: proton stoichiometry. In adjusted cells, n was lower and varied less, indicating a different mix of lactic acid exporters less affected by the high level of intracellular lactate. A qualitative model for pH effects and acid stress adaptation is proposed on the basis of these results.
Masood, Muhammad Irfan; Qadir, Muhammad Imran; Shirazi, Jafir Hussain; Khan, Ikram Ullah
Lactic acid bacteria are a diverse group of bacteria that produce lactic acid as their major fermented product. Most of them are normal flora of human being and animals and produce myriad beneficial effects for human beings include, alleviation of lactose intolerance, diarrhea, peptic ulcer, stimulation of immune system, antiallergic effects, antifungal actions, preservation of food, and prevention of colon cancer. This review highlights the potential species of Lactic acid bacteria responsible for producing these effects. It has been concluded that lactic acid bacteria are highly beneficial microorganisms for human beings and are present abundantly in dairy products so their use should be promoted for good human health.
Wu, Chongde; Huang, Jun; Zhou, Rongqing
As cell factories, lactic acid bacteria are widely used in food, agriculture, pharmaceutical and other industries. Acid stress is one the important survival challenges encountered by lactic acid bacteria both in fermentation process and in the gastrointestinal tract. Recently, the development of systems biology and metabolic engineering brings unprecedented opportunity for further elucidating the acid tolerance mechanisms and improving the acid stress resistance of lactic acid bacteria. This review addresses physiological mechanisms of lactic acid bacteria during acid stress. Moreover, strategies to improve the acid stress resistance of lactic acid were proposed.
van de Guchte, Maarten; Serror, Pascale; Chervaux, Christian; Smokvina, Tamara; Ehrlich, Stanislav D; Maguin, Emmanuelle
Lactic acid bacteria (LAB) constitute a heterogeneous group of bacteria that are traditionally used to produce fermented foods. The industrialization of food bio-transformations increased the economical importance of LAB, as they play a crucial role in the development of the organoleptique and hygienic quality of fermented products. Therefore, the reliability of starter strains in terms of quality and functional properties (important for the development of aroma and texture), but also in terms of growth performance and robustness has become essential. These strains should resist to adverse conditions encountered in industrial processes, for example during starter handling and storage (freeze-drying, freezing or spray-drying). The development of new applications such as life vaccines and probiotic foods reinforces the need for robust LAB since they may have to survive in the digestive tract, resist the intestinal flora, maybe colonize the digestive or uro-genital mucosa and express specific functions under conditions that are unfavorable to growth (for example, during stationary phase or storage). Also in nature, the ability to quickly respond to stress is essential for survival and it is now well established that LAB, like other bacteria, evolved defense mechanisms against stress that allow them to withstand harsh conditions and sudden environmental changes. While genes implicated in stress responses are numerous, in LAB the levels of characterization of their actual role and regulation differ widely between species. The functional conservation of several stress proteins (for example, HS proteins, Csp, etc) and of some of their regulators (for example, HrcA, CtsR) renders even more striking the differences that exist between LAB and the classical model micro-organisms. Among the differences observed between LAB species and B. subtilis, one of the most striking is the absence of a sigma B orthologue in L. lactis ssp. lactis as well as in at least two streptococci
Tao, Lin; Pavlova, Sylvia I; Ji, Xin; Jin, Ling; Spear, Gregory
Using food and commensal lactic acid bacteria (LAB) as vehicles for DNA delivery into epithelial cells is a new strategy for vaccine delivery or gene therapy. However, present methods for DNA delivery with LAB have suffered low efficiency. Our goal was to develop a new system to deliver DNA into epithelial cells with high efficiency using food and commensal LAB. An Escherichia coli-LAB shuttle plasmid, pLKV1, for DNA delivery into eukaryotic cells was constructed. Two reporter plasmids with green and red fluorescent protein genes were also constructed to monitor the uptake of protein and DNA, respectively. Bacteria delivering these reporter plasmids into Caco-2 cells were monitored by fluorescence microscopy. Several methods that weaken the bacterial cell wall prior to co-culture with Caco-2 cells were evaluated for their role in the improvement of gene transfer efficiency. Treating Streptococcus gordonii with penicillin and lysozyme greatly increased its rate of gene delivery to mammalian cells compared to untreated control bacteria, while glycine pretreatment promoted the highest gene transfer rate for Lactococcus lactis. Uptake of green fluorescent bacteria by Caco-2 cells showed that the cell wall-weakening treatment promoted the internalization of the noninvasive bacteria into Caco-2 cells. In conclusion, we have developed a noninvasive system using LAB as a vehicle for vaccine delivery or gene therapy, and tested this system in vitro with Caco-2 cells.
El-Shafei, H A; Abd-El-Sabour, H; Ibrahim, N; Mostafa, Y A
100 lactic acid bacterial strains isolated from traditional fermented foods (yoghurt, milk cream, sour dough and milk) were screened for bacteriocin production. Twenty six strains producing a nisin-like bacteriocin were selected. Most of these isolates gave only a narrow inhibitory spectrum, although one showed a broad inhibitory spectrum against the indicator strains tested, this strain was determined as Lactococcus lactis. The influence of several parameters on the fermentative production of nisin by Lactococcus lactis was studied. Production of nisin was optimal at 30 degrees C and in the pH range 5.5-6.3. The effect of different sulphur and nitrogen sources on Lactococcus lactis growth and nisin production was studied. Magnesium sulfate and manganese sulfate were found to be the best sulphur sources while triammonium citrate was the best inorganic nitrogen source and meat extract, peptone and yeast extract were the best organic nitrogen source for nisin production.
Sheng, Juzheng; Ling, Peixue; Wang, Fengshan
Hyaluronic acid (HA), a natural high molecular weight polysaccharide, is produced by Streptococcus zooepidemicus. However, Streptococcus has several drawbacks including its potential to produce exotoxins, so there is demand for an alternative HA source. Here, a recombinant HA biosynthesis operon, as well as the HA biosynthesis operon of S. zooepidemicus were introduced into L. lactis using the nisin-controlled expression system, respectively. HA was successfully synthesized by recombinant L. lactis. Furthermore, overexpression of the endogenous enzymes directing the synthesis of precursor sugars was effective at increasing HA production, and increasing the supply of UDP-activated monosaccharide donors aided synthesis of monodisperse HA polysaccharides. Besides GRAS host strain (L. lactis) and NICE system, the selecting marker (lacF gene) of the recombinant strain is also food grade. Therefore, HA produced by recombinant L. lactis overcomes the problems associated with Streptococcus and provides a source of food-grading HA appropriate for widespread biotechnological applications.
Mazzoli, Roberto; Bosco, Francesca; Mizrahi, Itzhak; Bayer, Edward A; Pessione, Enrica
Lactic acid bacteria (LAB) have long been used in industrial applications mainly as starters for food fermentation or as biocontrol agents or as probiotics. However, LAB possess several characteristics that render them among the most promising candidates for use in future biorefineries in converting plant-derived biomass-either from dedicated crops or from municipal/industrial solid wastes-into biofuels and high value-added products. Lactic acid, their main fermentation product, is an attractive building block extensively used by the chemical industry, owing to the potential for production of polylactides as biodegradable and biocompatible plastic alternative to polymers derived from petrochemicals. LA is but one of many high-value compounds which can be produced by LAB fermentation, which also include biofuels such as ethanol and butanol, biodegradable plastic polymers, exopolysaccharides, antimicrobial agents, health-promoting substances and nutraceuticals. Furthermore, several LAB strains have ascertained probiotic properties, and their biomass can be considered a high-value product. The present contribution aims to provide an extensive overview of the main industrial applications of LAB and future perspectives concerning their utilization in biorefineries. Strategies will be described in detail for developing LAB strains with broader substrate metabolic capacity for fermentation of cheaper biomass.
Mondragón-Parada, María Elena; Nájera-Martínez, Minerva; Juárez-Ramírez, Cleotilde; Galíndez-Mayer, Juvencio; Ruiz-Ordaz, Nora; Cristiani-Urbina, Eliseo
The main purpose of this work was to isolate and characterize lactic acid bacteria (LAB) strains to be used for biomass production using a whey-based medium supplemented with an ammonium salt and with very low levels of yeast extract (0.25 g/L). Five strains of LAB were isolated from naturally soured milk after enrichment in whey-based medium. One bacterial isolate, designated MNM2, exhibited a remarkable capability to utilize whey lactose and give a high biomass yield on lactose. This strain was identified as Lactobacillus casei by its 16S rDNA sequence. A kinetic study of cell growth, lactose consumption, and titratable acidity production of this bacterial strain was performed in a bioreactor. The biomass yield on lactose, the percentage of lactose consumption, and the maximum increase in cell mass obtained in the bioreactor were 0.165 g of biomass/g of lactose, 100%, and 2.0 g/L, respectively, which were 1.44, 1.11, and 2.35 times higher than those found in flask cultures. The results suggest that it is possible to produce LAB biomass from a whey-based medium supplemented with minimal amounts of yeast extract.
Alfonzo, Antonio; Ventimiglia, Giusi; Corona, Onofrio; Di Gerlando, Rosalia; Gaglio, Raimondo; Francesca, Nicola; Moschetti, Giancarlo; Settanni, Luca
Lactic acid bacteria (LAB) were analysed from wheat flours used in traditional bread making throughout Sicily (southern Italy). Plate counts, carried out in three different media commonly used to detect food and sourdough LAB, revealed a maximal LAB concentration of approximately 4.75 Log CFU g(-1). Colonies representing various morphological appearances were isolated and differentiated based on phenotypic characteristics and genetic analysis by randomly amplified polymorphic DNA (RAPD)-PCR. Fifty unique strains were identified. Analysis by 16S rRNA gene sequencing grouped the strains into 11 LAB species, which belonged to six genera: Enterococcus, Lactobacillus, Lactococcus, Leuconostoc, Pediococcus and Weissella. Weissella cibaria, Lactobacillus plantarum, Leuconostoc pseudomesenteroides and Leuconostoc citreum were the most prevalent species. The strains were not geographically related. Denaturing gradient gel electrophoresis (DGGE) analysis of total DNA of flour was used to provide a more complete understanding of the LAB population; it confirmed the presence of species identified with the culture-dependent approach, but did not reveal the presence of any additional LAB species. Finally, the technological characteristics (acidifying capacity, antimicrobial production, proteolytic activity, organic acid, and volatile organic compound generation) of the 50 LAB strains were investigated. Eleven strains were selected for future in situ applications.
Hoyle Parks, A R; Brashears, M M; Woerner, W D; Martin, J N; Thompson, L D; Brooks, J C
Growth of pathogenic organisms such as Escherichia coli O157:H7 and Salmonella spp. can be inhibited in ground beef through the addition of certain lactic acid-producing bacteria (LAB; Lactobacillus acidophilus NP51, Lactobacillus crispatus NP35, Pediococcus acidilactici, and Lactococcus lactis ssp. lactis). This study evaluated the effects of LAB inclusion on the organoleptic and biochemical properties typically associated with spoilage in traditionally packaged ground beef displayed at abusive (10°C) temperatures for 36 h. Trained and untrained panelist evaluations of lean color and off-odor, as well as instrumental color analyses, did not indicate an effect on spoilage traits due to LAB utilization (P > 0.05). However, display length affected each variable independently and was indicative of decreased stability and acceptability as display time (h) increased (P < 0.05). Thiobarbituric acid values were decreased for ground beef with added LAB (P < 0.05), but likely can be related to bacterial degradation of lipid oxidation by-products because no reduction in organoleptic traits due to oxidation was noted between treatments. Overall, LAB did not adversely influence the spoilage characteristics of traditionally packaged ground beef displayed at abusive temperatures for up to 36 h. Furthermore, biochemical and sensory indicators of spoilage were present for all treatments at the conclusion of display. Therefore, LAB can be added to ground beef in traditional packaging as a processing intervention without masking or delaying the expected spoilage characteristics.
Oliveira, Roseane B. P.; de L. Oliveira, Afonso; Glória, M. Beatriz A.
The objective of this study was to isolate lactic acid bacteria (LAB) from vacuum packaged beef and to investigate their antagonist activity. LAB mean counts of 5.19 log cfu/cm2 were obtained from five samples of vacuum packaged beef. Two hundred isolates were selected and screened for the inhibitory effect on five ATCC reference Lactobacillus strains. Thirty six isolates showed activity in the agar spot test against at least two of the indicator strains. However, only six cell free supernatants (CFS) from these isolates exhibited activity against the indicator strains using the well-diffusion test and conditions that eliminated the effects of organic acids and hydrogen peroxide. L. acidophilus was the most sensitive indicator tested, whereas L. plantarum and L. fermentum were the most resistant ones. Identification by MIDI system indicated that these LAB isolates were Lactococcus lactis subsp. cremoris, Pediococcus acidilactici, Lactobacillus delbrueckii subsp. bulgaricus and Lactobacillus casei GC subgroup A. The antagonistic factors produced by most of these LAB against L. acidophilus were resistant to heat treatment (100°C for 10 min) and stable over a wide pH range (4.0 to 9.0). These data suggest that these isolates could be used as promising hurdles aiming increased safety and extended shelf life of meat products. PMID:24031232
Chen, Yi-sheng; Wu, Hui-chung; Yanagida, Fujitoshi
The objective of this study was to isolate, characterize, and identify lactic acid bacteria (LAB) from ripe mulberries collected in Taiwan. Ripe mulberry samples were collected at five mulberry farms, located in different counties of Taiwan. Eighty-eight acid-producing cultures were isolated from these samples, and isolates were divided into classes first by phenotype, then into groups by restriction fragment length polymorphism (RFLP) analysis and sequencing of 16S ribosomal DNA (rDNA). Phenotypic and biochemical characteristics led to identification of four bacterial groups (A to D). Weissella cibaria was the most abundant type of LAB distributed in four mulberry farms, and Lactobacillus plantarum was the most abundant LAB found in the remaining farm. Ten W. cibaria and one Lactococcus lactis subsp. lactis isolate produced bacteriocins against the indicator strain Lactobacillus sakei JCM 1157T. These results suggest that various LAB are distributed in ripe mulberries and W. cibaria was the most abundant LAB found in this study. PMID:24031571
Dieye, Y.; Usai, S.; Clier, F.; Gruss, A.; Piard, J.-C.
We designed an expression and export system that enabled the targeting of a reporter protein (the staphylococcal nuclease Nuc) to specific locations in Lactococcus lactis cells, i.e., cytoplasm, cell wall, or medium. Optimization of protein secretion and of protein cell wall anchoring was performed with L. lactis cells by modifying the signals located at the N and C termini, respectively, of the reporter protein. Efficient translocation of precursor (∼95%) is obtained using the signal peptide from the lactococcal Usp45 protein and provided that the mature protein is fused to overall anionic amino acids at its N terminus; those residues prevented interactions of Nuc with the cell envelope. Nuc could be covalently anchored to the peptidoglycan by using the cell wall anchor motif of the Streptococcus pyogenes M6 protein. However, the anchoring step proved to not be totally efficient in L. lactis, as considerable amounts of protein remained membrane associated. Our results may suggest that the defect is due to limiting sortase in the cell. The optimized expression and export vectors also allowed secretion and cell wall anchoring of Nuc in food-fermenting and commensal strains of Lactobacillus. In all strains tested, both secreted and cell wall-anchored Nuc was enzymatically active, suggesting proper enzyme folding in the different locations. These results provide the first report of a targeting system in lactic acid bacteria in which the final location of a protein is controlled and biological activity is maintained. PMID:11418555
Jones, Rhys J; Hussein, Hassan M; Zagorec, Monique; Brightwell, Gale; Tagg, John R
The use of lactic acid bacteria (LAB) as protective cultures in vacuum-packed chill-stored meat has potential application for assuring and improving food quality, safety and market access. In a study to identify candidate strains suitable for evaluation in a meat model, agar-based methods were employed to screen 181 chilled meat and meat process-related LAB for strains inhibitory to pathogens and spoilage organisms of importance to the meat industry. Six meat-derived strains, including Lactobacillus sakei and Lactococcus lactis, were found to be inhibitory to one or more of the target strains Listeria monocytogenes, Brochothrix thermosphacta, Campylobacter jejuni and Clostridium estertheticum. The inhibitory agents appeared to be either cell-associated or molecules released extracellularly with bacteriocin-like properties. Variations detected in the antimicrobial activity of LAB associated with changes to test parameters such as substrate composition underlined the importance of further in situ evaluation of the inhibitory strains in stored meat trials.
Kelly, W J; Asmundson, R V; Huang, C M
Lactic acid bacteria isolated from a range of foods sold in ready-to-eat form were screened for bacteriocin production. Twenty-two bacteriocin-producing cultures were isolated from 14 of the 41 foods sampled. Bacteriocin-producing isolates from meat, fish and dairy products were Lactobacillus and Leuconostoc species typically found associated with these products. Most of these isolates gave only a narrow inhibitory spectrum although two showed activity against Listeria monocytogenes. Fruit and vegetable products gave a broader range of organisms but most of the bacteriocin-producing cultures were found to be strains of Lactococcus. Several lactococci produced a nisin-like activity, and showed a broad inhibitory spectrum against the indicator strains tested. The ease with which bacteriocin-producing strains could be isolated implies that they are already being safely consumed in food, and highlights the potential for using bacteriocin-producing cultures for biopreservation, especially in association with minimally processed products.
Dàvila, Eduard; Zamora, Lucero M; Pla, Maria; Carretero, Carmen; Parés, Dolors
Ninety-seven lactic acid bacteria (LAB) were isolated from slaughterhouse porcine blood in order to select autochthonous LAB strains for use as biopreservatives of this by-product. They were identified by 16S rDNA sequencing; and their inhibition capacity was determined against four bacterial species frequently found in contaminated blood, i.e. Staphylococcus aureus, Escherichia coli, Pseudomonas fluorescens and Bacillus spp. The taxonomic study showed an unexpected low diversity of LAB in blood, i.e. only 8 different species were found, from which just 4, i.e. Enterococcus raffinosus, Lactobacillus murinus, Lactobacillus reuteri and Lactococcus garvieae, amounted to more than 90% of all isolates. Inhibition tests in solid culture media proved that S. aureus and Bacillus spp. were inhibited by most LAB strains obtained from porcine blood. E. coli was the indicator less affected by the isolated LAB species. Several isolates efficiently inhibited the growth of all tested indicators.
Dehli, Tore; Jensen, Peter Ruhdal
Lactic acid bacteria (LAB) are known for their high tolerance toward organic acids and alcohols (R. S. Gold, M. M. Meagher, R. Hutkins, and T. Conway, J. Ind. Microbiol. 10:45–54, 1992) and could potentially serve as platform organisms for production of these compounds. In this study, we attempted to redirect the metabolism of LAB model organism Lactococcus lactis toward ethanol production. Codon-optimized Zymomonas mobilis pyruvate decarboxylase (PDC) was introduced and expressed from synthetic promoters in different strain backgrounds. In the wild-type L. lactis strain MG1363 growing on glucose, only small amounts of ethanol were obtained after introducing PDC, probably due to a low native alcohol dehydrogenase activity. When the same strains were grown on maltose, ethanol was the major product and lesser amounts of lactate, formate, and acetate were formed. Inactivating the lactate dehydrogenase genes ldhX, ldhB, and ldh and introducing codon-optimized Z. mobilis alcohol dehydrogenase (ADHB) in addition to PDC resulted in high-yield ethanol formation when strains were grown on glucose, with only minor amounts of by-products formed. Finally, a strain with ethanol as the sole observed fermentation product was obtained by further inactivating the phosphotransacetylase (PTA) and the native alcohol dehydrogenase (ADHE). PMID:23377945
Solem, Christian; Dehli, Tore; Jensen, Peter Ruhdal
Lactic acid bacteria (LAB) are known for their high tolerance toward organic acids and alcohols (R. S. Gold, M. M. Meagher, R. Hutkins, and T. Conway, J. Ind. Microbiol. 10:45-54, 1992) and could potentially serve as platform organisms for production of these compounds. In this study, we attempted to redirect the metabolism of LAB model organism Lactococcus lactis toward ethanol production. Codon-optimized Zymomonas mobilis pyruvate decarboxylase (PDC) was introduced and expressed from synthetic promoters in different strain backgrounds. In the wild-type L. lactis strain MG1363 growing on glucose, only small amounts of ethanol were obtained after introducing PDC, probably due to a low native alcohol dehydrogenase activity. When the same strains were grown on maltose, ethanol was the major product and lesser amounts of lactate, formate, and acetate were formed. Inactivating the lactate dehydrogenase genes ldhX, ldhB, and ldh and introducing codon-optimized Z. mobilis alcohol dehydrogenase (ADHB) in addition to PDC resulted in high-yield ethanol formation when strains were grown on glucose, with only minor amounts of by-products formed. Finally, a strain with ethanol as the sole observed fermentation product was obtained by further inactivating the phosphotransacetylase (PTA) and the native alcohol dehydrogenase (ADHE).
Backus, Lennart; Boekhorst, Jos; Dijkstra, Annereinou; Beerthuyzen, Marke; Siezen, Roland J.; Bachmann, Herwig; van Hijum, Sacha A. F. T.
ABSTRACT The lactic acid bacterium Lactococcus lactis is widely used for the fermentation of dairy products. Here, we present the draft genome sequences of 11 L. lactis subsp. cremoris strains isolated from different environments. PMID:28302789
Backus, Lennart; Wels, Michiel; Boekhorst, Jos; Dijkstra, Annereinou R.; Beerthuyzen, Marke; Kelly, William J.; Siezen, Roland J.; van Hijum, Sacha A. F. T.
ABSTRACT The lactic acid bacterium Lactococcus lactis is widely used for the production of fermented dairy products. Here, we present the draft genome sequences of 24 L. lactis strains isolated from different environments and geographic locations. PMID:28360177
de las Rivas, Blanca; Rodríguez, Héctor; Curiel, José Antonio; Landete, José María; Muñoz, Rosario
The potential to produce volatile phenols from hydroxycinnamic acids was investigated for lactic acid bacteria (LAB) isolated from Spanish grape must and wine. A PCR assay was developed for the detection of LAB that potentially produce volatile phenols. Synthetic degenerate oligonucleotides for the specific detection of the pdc gene encoding a phenolic acid decarboxylase were designed. The pdc PCR assay amplifies a 321 bp DNA fragment from phenolic acid decarboxylase. The pdc PCR method was applied to 85 strains belonging to the 6 main wine LAB species. Lactobacillus plantarum, Lactobacillus brevis, and Pediococcus pentosaceus strains produce a positive response in the pdc PCR assay, whereas Oenococcus oeni, Lactobacillus hilgardii, and Leuconostoc mesenteroides strains did not produce the expected PCR product. The production of vinyl and ethyl derivatives from hydroxycinnamic acids in culture media was determined by high-performance liquid chromatography. A relationship was found between pdc PCR amplification and volatile phenol production, so that the LAB strains that gave a positive pdc PCR response produce volatile phenols, whereas strains that did not produce a PCR amplicon did not produce volatile phenols. The proposed method could be useful for a preliminary identification of LAB strains able to produce volatile phenols in wine.
Zaidi, Arsalan Haseeb; Bakkes, Patrick J.; Lubelski, Jacek; Agustiandari, Herfita; Kuipers, Oscar P.; Driessen, Arnold J. M.
Upon prolonged exposure to cholate and other toxic compounds, Lactococcus lactis develops a multidrug resistance phenotype that has been attributed to an elevated expression of the heterodimeric ABC-type multidrug transporter LmrCD. To investigate the molecular basis of bile acid resistance in L. lactis and to evaluate the contribution of efflux-based mechanisms in this process, the drug-sensitive L. lactis NZ9000 ΔlmrCD strain was challenged with cholate. A resistant strain was obtained that, compared to the parental strain, showed (i) significantly improved resistance toward several bile acids but not to drugs, (ii) morphological changes, and (iii) an altered susceptibility to antimicrobial peptides. Transcriptome and transport analyses suggest that the acquired resistance is unrelated to elevated transport activity but, instead, results from a multitude of stress responses, changes to the cell envelope, and metabolic changes. In contrast, wild-type cells induce the expression of lmrCD upon exposure to cholate, whereupon the cholate is actively extruded from the cells. Together, these data suggest a central role for an efflux-based mechanism in bile acid resistance and implicate LmrCD as the main system responsible in L. lactis. PMID:18790870
Toomey, Niamh; Monaghan, Áine; Fanning, Séamus; Bolton, Declan
Three wild-type dairy isolates of lactic acid bacteria (LAB) and one Lactococcus lactis control strain were analyzed for their ability to transfer antibiotic resistance determinants (plasmid or transposon located) to two LAB recipients using both in vitro methods and in vivo models. In vitro transfer experiments were carried out with the donors and recipients using the filter mating method. In vivo mating examined transfer in two natural environments, a rumen model and an alfalfa sprout model. All transconjugants were confirmed by Etest, PCR, pulsed-field gel electrophoresis, and Southern blotting. The in vitro filter mating method demonstrated high transfer frequencies between all LAB pairs, ranging from 1.8 × 10−5 to 2.2 × 10−2 transconjugants per recipient. Transconjugants were detected in the rumen model for all mating pairs tested; however, the frequencies of transfer were low and inconsistent over 48 h (ranging from 1.0 × 10−9 to 8.0 × 10−6 transconjugants per recipient). The plant model provided an environment that appeared to promote comparatively higher transfer frequencies between all LAB pairs tested over the 9-day period (transfer frequencies ranged from 4.7 × 10−4 to 3.9 × 10−1 transconjugants per recipient). In our test models, dairy cultures of LAB can act as a source of mobile genetic elements encoding antibiotic resistance that can spread to other LAB. This observation could have food safety and public health implications. PMID:19270126
Nikolic, Milica; Terzic-Vidojevic, Amarela; Jovcic, Branko; Begovic, Jelena; Golic, Natasa; Topisirovic, Ljubisa
The Bukuljac cheese is traditionally homemade cheese, produced from heat-treated goat's milk without the addition of any bacterial starter culture. The presence of lactic acid bacteria (LAB) in Bukuljac cheese has been analyzed by using a polyphasic approach including microbiological and molecular methods such as rep-PCR with (GTG)5 primer. Lactobacillus paracasei subsp. paracasei represents a dominant strain in the microflora of analyzed cheese. Out of 55 Gram-positive and catalase-negative isolates, 48 belonged to L. paracasei subsp. paracasei species. Besides lactobacilli, five Lactococcus lactis subsp. lactis and two Enterococcus faecalis were found. Results of PCR-denaturing gradient gel electrophoresis (DGGE) of DNA extracted directly from the fresh cheese revealed the presence of Leuconostoc mesenteroides. Only lactobacilli showed a high proteolytic activity and hydrolyzed alpha(s1)- and beta-caseins. They are also producers of diacetyl. In addition, 34 out of 55 isolates, all determined as lactobacilli, showed the ability of auto-aggregation. Among 55 isolates, 50 also exhibited antimicrobial activity.
Migaw, Sarra; Ghrairi, Taoufik; Belguesmia, Yanath; Choiset, Yvan; Berjeaud, Jean-Marc; Chobert, Jean-Marc; Hani, Khaled; Haertlé, Thomas
Nine lactic acid bacteria strains showing bacteriocin-like activity were isolated from various fresh fish viscera. The following species were identified based on 16S rDNA sequences: Enterococcus durans (7 isolates), Lactococcus lactis (1) and Enterococcus faecium (1). These strains were active against Listeria innocua and other LAB. Random amplified polymorphic DNA analyses showed four major patterns for the E. durans species. PCR analyses revealed a nisin gene in the genome of the Lc. lactis strain. Genes coding enterocins A, B and P were found in the genome of the E. faecium isolate. Enterocins A and B genes were also present in the genome of E. durans GM19. Hence, this is the first report describing E. durans strains producing enterocins A and B. Electrospray ionization mass spectrometry revealed that the purified bacteriocin produced by the E. durans GMT18 strain had an exact molecular mass of 6,316.89 Da. This bacteriocin was designated as durancin GMT18. Edman sequencing failed to proceed; suggesting that durancin GTM18 may contain terminal lanthionine residues. Overall, the results obtained revealed the presence of a variety of enterococci in Mediterranean fish viscera, as evidenced by their genetic profiles and abilities to produce different bacteriocins. These strains could be useful for food biopreservation or as probiotics.
Lactic acid bacteria play important roles in various fermented foods in Asia. Besides being the main component in kimchi and other fermented foods, they are used to preserve edible food materials through fermentation of other raw-materials such as rice wine/beer, rice cakes, and fish by producing organic acids to control putrefactive microorganisms and pathogens. These bacteria also provide a selective environment favoring fermentative microorganisms and produce desirable flavors in various fermented foods. This paper discusses the role of lactic acid bacteria in various non-dairy fermented food products in Asia and their nutritional and physiological functions in the Asian diet. PMID:21995342
Siragusa, S.; De Angelis, M.; Di Cagno, R.; Rizzello, C. G.; Coda, R.; Gobbetti, M.
The concentrations of γ-aminobutyric acid (GABA) in 22 Italian cheese varieties that differ in several technological traits markedly varied from 0.26 to 391 mg kg−1. Presumptive lactic acid bacteria were isolated from each cheese variety (total of 440 isolates) and screened for the capacity to synthesize GABA. Only 61 isolates showed this activity and were identified by partial sequencing of the 16S rRNA gene. Twelve species were found. Lactobacillus paracasei PF6, Lactobacillus delbrueckii subsp. bulgaricus PR1, Lactococcus lactis PU1, Lactobacillus plantarum C48, and Lactobacillus brevis PM17 were the best GABA-producing strains during fermentation of reconstituted skimmed milk. Except for L. plantarum C48, all these strains were isolated from cheeses with the highest concentrations of GABA. A core fragment of glutamate decarboxylase (GAD) DNA was isolated from L. paracasei PF6, L. delbrueckii subsp. bulgaricus PR1, L. lactis PU1, and L. plantarum C48 by using primers based on two highly conserved regions of GAD. A PCR product of ca. 540 bp was found for all the strains. The amino acid sequences deduced from nucleotide sequence analysis showed 98, 99, 90, and 85% identity to GadB of L. plantarum WCFS1 for L. paracasei PF6, L. delbrueckii subsp. bulgaricus PR1, L. lactis PU1, and L. plantarum C48, respectively. Except for L. lactis PU1, the three lactobacillus strains survived and synthesized GABA under simulated gastrointestinal conditions. The findings of this study provide a potential basis for exploiting selected cheese-related lactobacilli to develop health-promoting dairy products enriched in GABA. PMID:17890341
Lactic acid bacteria (LAB) are a group of facultative anaerobic, catalase negative, nonmotile and nonsporeforming–Gram positive bacteria. Most LAB utilize high energy C sources including monomer sugars to produce energy to maintain cellular structure and function. This anaerobic fermentation proce...
Divya, Jayakumar Beena; Nampoothiri, Kesavan Madhavan
Two lactic acid bacteria (LAB) isolated from cow's milk were identified as Lactococcus lactis strains and designated as L. lactis CM22 and L. lactis CM28. They were immobilised by co-encapsulation using alginate and mannitol and by hybrid entrapment with skim milk, glycerol, CaCO3 and alginate. The encapsulated cells survived better in simulated gastrointestinal conditions compared to the free cells. The percentage survival of probiotics encapsulated by hybrid entrapment method was 62.74% for L. lactis CM22 and 68% for L. lactis CM28. Studies to check their efficacy in fermentative fortification of skim milk and ice cream revealed an enhancement in folate level.
Faye, T; Tamburello, A; Vegarud, G E; Skeie, S
In the present study, the survival of 9 lactic acid bacteria (5 Lactococcus strains, 3 Lactobacillus strains, and 1 strain of Enterococcus hirae), was investigated in vitro under conditions similar to human digestion using human gastric and duodenal juices. The tolerance of the bacteria was also tested with traditional methods using acidic conditions and bile salts. The strains were subjected to a model digestive system comprising sequential incubation in human gastric and duodenal juices, in a 2-step digestion assay at 37°C, simulating the human upper gastrointestinal tract with human gastric juices at pH 2.5 and human duodenal juices at pH 7. The bacterial strains were tested either as washed cells from culture media or in fermented milk. The initial in vitro testing in acid and bile salts showed that Lactobacillus strains and the E. hirae strain displayed a significantly higher acid tolerance than the lactococci. The lactobacilli and the Enterococcus numbers increased, whereas the lactococci decreased at least 1 log during the bile salt treatment. The Lactobacillus strains showed the highest survival rate in the model digestive system when washed bacterial cultures were used with a minor log reduction, whereas the lactococci numbers were reduced by at least log 4. However, when using fermented milks in the model digestion system it was demonstrated that the Enterococcus strain and 2 strains of Lactococcus lactis ssp. cremoris benefited significantly from the presence of the fermented milk as food matrix, with log numbers >log 7 and 5, respectively, after digestion of the fermented milk. The analyses reported comprise a comprehensive in vitro testing regimen suitable for evaluation of the survival of candidate probiotic bacteria in human digestion as an initial prescreen to clinical trials.
Campos, Francisco M; Figueiredo, Ana R; Hogg, Tim A; Couto, José A
The influence of phenolic (p-coumaric, caffeic, ferulic, gallic and protocatechuic) acids on glucose and organic acid metabolism by two strains of wine lactic acid bacteria (Oenococcus oeni VF and Lactobacillus hilgardii 5) was investigated. Cultures were grown in modified MRS medium supplemented with different phenolic acids. Cellular growth was monitored and metabolite concentrations were determined by HPLC-RI. Despite the strong inhibitory effect of most tested phenolic acids on the growth of O. oeni VF, the malolactic activity of this strain was not considerably affected by these compounds. While less affected in its growth, the capacity of L. hilgardii 5 to degrade malic acid was clearly diminished. Except for gallic acid, the addition of phenolic acids delayed the metabolism of glucose and citric acid in both strains tested. It was also found that the presence of hydroxycinnamic acids (p-coumaric, caffeic and ferulic) increased the yield of lactic and acetic acid production from glucose by O. oeni VF and not by L. hilgardii 5. The results show that important oenological characteristics of wine lactic acid bacteria, such as the malolactic activity and the production of volatile organic acids, may be differently affected by the presence of phenolic acids, depending on the bacterial species or strain.
Najjari, Afef; Turki, Yousra; Jaballah, Sana; Boudabous, Abdelatif; Ouzari, Hadda
A total of 119 lactic acid bacteria (LAB) were isolated, by culture-dependant method, from rhizosphere samples of olive trees and desert truffles and evaluated for different biotechnological properties. Using the variability of the intergenic spacer 16S-23S and 16S rRNA gene sequences, the isolates were identified as the genera Lactococcus, Pediococcus, Lactobacillus, Weissella, and Enterococcus. All the strains showed proteolytic activity with variable rates 42% were EPS producers, while only 10% showed the ability to grow in 9% NaCl. In addition, a low rate of antibiotic resistance was detected among rhizospheric enterococci. Furthermore, a strong antibacterial activity against plant and/or pathogenic bacteria of Stenotrophomonas maltophilia, Pantoea agglomerans, Pseudomonas savastanoi, the food-borne Staphylococcus aureus, and Listeria monocytogenes was recorded. Antifungal activity evaluation showed that Botrytis cinerea was the most inhibited fungus followed by Penicillium expansum, Verticillium dahliae, and Aspergillus niger. Most of the active strains belonged to the genera Enterococcus and Weissella. This study led to suggest that environmental-derived LAB strains could be selected for technological application to control pathogenic bacteria and to protect food safety from postharvest deleterious microbiota. PMID:24151598
Macaluso, Giusi; Fiorenza, Gerlando; Gaglio, Raimondo; Mancuso, Isabella
Bacteriocins are antimicrobial proteins produced by bacteria that inhibit the growth of other bacteria with a bactericidal or bacteriostatic mode of action. Many lactic acid bacteria (LAB) produce a high diversity of different bacteriocins. Bacteriocinogenic LAB are generally recognised as safe (GRAS) and useful to control the frequent development of pathogens and spoilage microorganisms. For this reason they are commonly used as starter cultures in food fermentations. In this study, the authors describe the results of a screening on 699 LAB isolated from wooden vat surfaces, raw milk and traditional Sicilian cheeses, for the production of bacteriocin-like inhibitory substances, by comparing two alternative methods. The antagonistic activity of LAB and its proteinaceous nature were evaluated using the spot-on-the-lawn and the well-diffusion assay (WDA) and the sensitivity to proteolytic (proteinase K, protease B and trypsin), amylolytic (a-amylase) and lipolytic (lipase) enzymes. The indicator strains used were: Listeria monocytogenes, Staphylococcus aureus, Escherichia coli, Salmonella enteritidis. A total of 223 strains (belonging to the species Enterococcus spp., Lactobacillus spp., Pediococcus spp., Streptococcus spp., Leuconostoc spp. and Lactococcus lactis) were found to inhibit the growth of Listeria monocytogenes by using the spot-on-the-lawn method; only 37 of these were confirmed by using the WDA. The direct addition of bacteriocin-producing cultures into dairy products can be a more practical and economic option for the improvement of the safety and quality of the final product. PMID:27800430
Ni, Kuikui; Wang, Yanping; Li, Dongxia; Cai, Yimin; Pang, Huili
There has been growing interest to develop forage rice as a new feed resource for livestock. This study was to characterize the natural population of lactic acid bacteria (LAB) and select potentially excellent strains for paddy rice silage preparation in China. One hundred and twenty-six strains were isolated and screened from paddy rice silage prepared using a small-scale fermentation system, and ninety-nine of these isolates were considered to be LAB based on their Gram-positive and catalase-negative morphology and the production of most of their metabolic products as lactic acid. These isolates were divided into eight groups (A-H) on the basis of their morphological and biochemical characteristics. The Group A to H strains were identified as Lactobacillus (L.) plantarum subsp. plantarum (species ratio: 8.1%), L. casei (5.1%), Leuconostoc (Ln.) pseudomesenteroides (11.1%), Pediococcus (P.) pentosaceus (24.2%), Enterococcus (E.) mundtii (12.1%), Lactococcus (Lc.) garvieae (15.2%), E. faecium (9.1%) and Lc. lactis subsp. lactis (15.2%) based on sequence analyses of their 16S rRNA and recA genes. P. pentosaceus was the most abundant member of the LAB population in the paddy rice silage. A selected strain, namely L. casei R 465, was found to be able to grow under low pH conditions and to improve the silage quality with low pH and a relatively high content of lactic acid. This study demonstrated that forage paddy rice silage contains abundant LAB species and its silage can be well preserved by inoculation with LAB, and that strain R 465 can be a potentially excellent inoculant for paddy rice silage.
Ni, Kuikui; Wang, Yanping; Li, Dongxia; Cai, Yimin; Pang, Huili
There has been growing interest to develop forage rice as a new feed resource for livestock. This study was to characterize the natural population of lactic acid bacteria (LAB) and select potentially excellent strains for paddy rice silage preparation in China. One hundred and twenty-six strains were isolated and screened from paddy rice silage prepared using a small-scale fermentation system, and ninety-nine of these isolates were considered to be LAB based on their Gram-positive and catalase-negative morphology and the production of most of their metabolic products as lactic acid. These isolates were divided into eight groups (A-H) on the basis of their morphological and biochemical characteristics. The Group A to H strains were identified as Lactobacillus (L.) plantarum subsp. plantarum (species ratio: 8.1%), L. casei (5.1%), Leuconostoc (Ln.) pseudomesenteroides (11.1%), Pediococcus (P.) pentosaceus (24.2%), Enterococcus (E.) mundtii (12.1%), Lactococcus (Lc.) garvieae (15.2%), E. faecium (9.1%) and Lc. lactis subsp. lactis (15.2%) based on sequence analyses of their 16S rRNA and recA genes. P. pentosaceus was the most abundant member of the LAB population in the paddy rice silage. A selected strain, namely L. casei R 465, was found to be able to grow under low pH conditions and to improve the silage quality with low pH and a relatively high content of lactic acid. This study demonstrated that forage paddy rice silage contains abundant LAB species and its silage can be well preserved by inoculation with LAB, and that strain R 465 can be a potentially excellent inoculant for paddy rice silage. PMID:25803578
Tikhonov, V. V.; Yakushev, A. V.; Zavgorodnyaya, Yu. A.; Byzov, B. A.; Demin, V. V.
The influence of humic acids of different origins on the growth of bacterial cultures of different taxa isolated from the soil and the digestive tracts of earthworms ( Aporrectodea caliginosa)—habitats with contrasting conditions—was studied. More than half of the soil and intestinal isolates from the 170 tested strains grew on the humic acid of brown coal as the only carbon source. The specific growth rate of the bacteria isolated from the intestines of the earthworms was higher than that of the soil bacteria. The use of humic acids by intestinal bacteria confirms the possibility of symbiotic digestion by earthworms with the participation of bacterial symbionts. Humic acids at a concentration of 0.1 g/l stimulated the growth of the soil and intestinal bacteria strains (66 strains out of 161) on Czapek’s medium with glucose (1 g/l), probably, acting as a regulator of the cell metabolism. On the medium with the humic acid, the intestinal bacteria grew faster than the soil isolates did. The most active growth of the intestinal isolates was observed by Paenibacillus sp., Pseudomonas putida, Delftia acidovorans, Microbacterium terregens, and Aeromonas sp.; among the soil ones were the representatives of the Pseudomonas genus. A response of the bacteria to the influence of humic acids was shown at the strain level using the example of Pseudomonas representatives. The Flexom humin preparation stimulated the growth of the hydrocarbon-oxidizing Acinetobacter sp. bacteria. This effect can be used for creating a new compound with the elevated activity of bacteria that are destroyers of oil and oil products.
Cho, Young-Hee; Shin, Il-Seung; Hong, Sung-Moon; Kim, Cheol-Hyun
The aim of this study was to manufacture functional high protein fermented beverage, using whey protein concentrate (WPC) and Lactobacillus plantarum DK211 isolated from kimchi, and to evaluate the physicochemical, functional, and sensory properties of the resulting product. The fermented whey beverage (FWB) was formulated with whey protein concentrate 80 (WPC 80), skim milk powder, and sucrose; and fermented with Lactobacillus plantarum DK211 as single, or mixed with Lactococcus lactis R704, a commercial starter culture. The pH, titratable acidity, and viable cell counts during fermentation and storage were evaluated. It was found that the mixed culture showed faster acid development than the single culture. The resulting FWB had high protein (9%) and low fat content (0.2%). Increased viscosity, and antioxidant and antimicrobial activity were observed after fermentation. A viable cell count of 10(9) CFU/mL in FWB was achieved within 10 h fermentation, and it remained throughout storage at 15℃ for 28 d. Sensory analysis was also conducted, and compared to that of a commercial protein drink. The sensory scores of FWB were similar to those of the commercial protein drink in most attributes, except sourness. The sourness was highly related with the high lactic acid content produced during fermentation. The results showed that WPC and vegetable origin lactic acid bacteria isolated from kimchi might be used for the development of a high protein fermented beverage, with improved functionality and organoleptic properties.
The aim of this study was to manufacture functional high protein fermented beverage, using whey protein concentrate (WPC) and Lactobacillus plantarum DK211 isolated from kimchi, and to evaluate the physicochemical, functional, and sensory properties of the resulting product. The fermented whey beverage (FWB) was formulated with whey protein concentrate 80 (WPC 80), skim milk powder, and sucrose; and fermented with Lactobacillus plantarum DK211 as single, or mixed with Lactococcus lactis R704, a commercial starter culture. The pH, titratable acidity, and viable cell counts during fermentation and storage were evaluated. It was found that the mixed culture showed faster acid development than the single culture. The resulting FWB had high protein (9%) and low fat content (0.2%). Increased viscosity, and antioxidant and antimicrobial activity were observed after fermentation. A viable cell count of 109 CFU/mL in FWB was achieved within 10 h fermentation, and it remained throughout storage at 15℃ for 28 d. Sensory analysis was also conducted, and compared to that of a commercial protein drink. The sensory scores of FWB were similar to those of the commercial protein drink in most attributes, except sourness. The sourness was highly related with the high lactic acid content produced during fermentation. The results showed that WPC and vegetable origin lactic acid bacteria isolated from kimchi might be used for the development of a high protein fermented beverage, with improved functionality and organoleptic properties. PMID:26761827
Bulut, Cisem; Gunes, Hatice; Okuklu, Burcu; Harsa, Sebnem; Kilic, Sevda; Coban, Hatice Sevgi; Yenidunya, Ali Fazil
Comlek peyniri is a typical artisanal cheese in Central Anatolia. This type of cheese was made by using the indigenous lactic acid bacteria (LAB) flora of cow or ewes' milk. Majority of the samples were taken from fresh cheese because the aim was to isolate homofermentative LAB. Initially 661 microbial isolates were obtained from 17 cheese samples. Only 107 were found to be homofermentative LAB. These isolates were selected and identified by using both phenotypic and molecular methods. Phenotypic identification included curd formation from skim milk, catalase test, Gram staining and light microscopy, growth at different temperatures and salt concentrations, arginine hydrolysis, gas production from glucose, and carbohydrate fermentation. Molecular identification was based on the polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) of the 16S rRNA gene-ITS (internally transcribed spacer) region. By combining the phenotypic and molecular identification results, isolates belonging to each of the following genera were determined at species or subspecies level: 54 Lactococcus lactis subsp. lactis, 21 Enterococcus faecium, 3 Ec. faecalis, 2 Ec. durans, 10 Ec. sp., 15 Lactobacillus paracasei subsp. paracasei, and 2 Lb. casei strains. Technological characterisation was also performed by culturing each of the strains in UHT skim milk, and by monitoring pH change and lactic acid production at certain time intervals through the 24 h incubation. Results of the technological characterisation indicated that 33% of the isolates (35 strains) were capable of lowering the pH of UHT milk below 5.3 after 6 h incubation at 30 degrees C. Thirty four of these strains were Lc. lactis subsp. lactis, and only one was an Ec. faecium strain.
Ni, Kuikui; Wang, Yanping; Cai, Yimin; Pang, Huili
Winter wheat is a suitable crop to be ensiled for animal feed and China has the largest planting area of this crop in the world. During the ensiling process, lactic acid bacteria (LAB) play the most important role in the fermentation. We investigated the natural population of LAB in whole-crop wheat (WCW) and examined the quality of whole-crop wheat silage (WCWS) with and without LAB inoculants. Two Lactobacillus plantarum subsp. plantarum strains, Zhengzhou University 1 (ZZU 1) selected from corn and forage and grass 1 (FG 1) from a commercial inoculant, were used as additives. The silages inoculated with LAB strains (ZZU 1 and FG 1) were better preserved than the control, with lower pH values (3.5 and 3.6, respectively) (p<0.05) and higher contents of lactic acid (37.5 and 34.0 g/kg of fresh matter (FM), respectively) (p<0.05) than the control. Sixty LAB strains were isolated from fresh material and WCWS without any LAB inoculation. These LAB strains were divided into the following four genera and six species based on their phenotypic, biochemical and phylogenetic characteristics: Leuconostoc pseudomesenteroides, Leuconostoc citreum, Weissella cibaria, Lactococcus lactis subsp. lactis, Lactobacillus buchneri, and Lactobacillus plantarum subsp. plantarum. However, the prevalent LAB, which was predominantly heterofermentative (66.7%), consisted of Leuconostoc pseudomesenteroides, Leuconostoc citreum, Weissella cibaria, and Lactobacillus buchneri. This study revealed that most of isolated LAB strains from control WCWS were heterofermentative and could not grow well at low pH condition; the selective inoculants of Lactobacillus strains, especially ZZU 1, could improve WCWS quality significantly. PMID:26104520
Taniguchi, Masayuki; Ishiyama, Yohei; Takata, Takeomi; Nakanishi, Toshihiro; Kaneoke, Mitsuoki; Watanabe, Ken-ichi; Yanagida, Fujitoshi; Chen, Yi-sheng; Kouya, Tomoaki; Tanaka, Takaaki
The bacteriocins produced by Lactococcus lactis subsp. lactis C101910 (C101910) and NBRC 12007 (NBRC 12007) were used to prevent the growth of sake spoiling hiochi bacteria (Lactobacillus hilgardii, Lactobacillus fructivorans, and Lactobacillus paracasei) in namazake, which is raw (unpasteurized) sake. The bacteriocin concentrations required for decreasing the viable cell concentrations of L. hilgardii and L. fructivorans below the detection limit (1.0 x 10(2) cells/ml) in 24 h from the initial concentration of 4.0-9.5 x 10(5) cells/ml in the namazake at pH 4.5 and at 4 degrees C, were 18-35 U/ml and 5.6 U/ml for the bacteriocin from C101910 and NBRC 12007, respectively. To decrease the viable cell concentration of L. paracasei from the initial concentration of 7.5 x 10(5) cells/ml to below the detection limit (1.0 x 10(2) cells/ml) in 24 h, 350 U/ml bacteriocin from C101910 and 140 U/ml bacteriocin from NBRC 12007 were required. In experiments using McIlvaine buffer (pH 4.5) with 15% ethanol instead of namazake as the medium, the viable cell concentrations of L. hilgardii and L. paracasei decreased to less than 1.0 x 10(2) cells/ml, whereas those of L. fructivorans decreased to less than 1.0 x 10(3) cells/ml, when bacteriocins were added at the concentrations that had proven effective in namazake. The membrane depolarization assay using a fluorescent probe showed that the presence of ethanol stimulated the collapse of the membrane potential induced by bacteriocins. The ethanol induced collapse of the membrane potential suggests that the application of bacteriocins at the storage stage of namazake is more beneficial than when used in other stages of the sake brewing process.
Hauri, James F.; Schaider, Laurel A.
Sulfate reducing bacteria have been shown to be effective at treating acid mine drainage through sulfide production and subsequent precipitation of metal sulfides. In this laboratory experiment for undergraduate environmental chemistry courses, students design and implement a set of bioreactors to remediate acid mine drainage and explain observed…
Shiraishi, Tsukasa; Yokota, Shinichi; Fukiya, Satoru; Yokota, Atsushi
Bacterial cell surface molecules are at the forefront of host-bacterium interactions. Teichoic acids are observed only in Gram-positive bacteria, and they are one of the main cell surface components. Teichoic acids play important physiological roles and contribute to the bacterial interaction with their host. In particular, lipoteichoic acid (LTA) anchored to the cell membrane has attracted attention as a host immunomodulator. Chemical and biological characteristics of LTA from various bacteria have been described. However, most of the information concerns pathogenic bacteria, and information on beneficial bacteria, including probiotic lactic acid bacteria, is insufficient. LTA is structurally diverse. Strain-level structural diversity of LTA is suggested to underpin its immunomodulatory activities. Thus, the structural information on LTA in probiotics, in particular strain-associated diversity, is important for understanding its beneficial roles associated with the modulation of immune response. Continued accumulation of structural information is necessary to elucidate the detailed physiological roles and significance of LTA. In this review article, we summarize the current state of knowledge on LTA structure, in particular the structure of LTA from lactic acid bacteria. We also describe the significance of structural diversity and biological roles of LTA.
SHIRAISHI, Tsukasa; YOKOTA, Shinichi; FUKIYA, Satoru; YOKOTA, Atsushi
Bacterial cell surface molecules are at the forefront of host-bacterium interactions. Teichoic acids are observed only in Gram-positive bacteria, and they are one of the main cell surface components. Teichoic acids play important physiological roles and contribute to the bacterial interaction with their host. In particular, lipoteichoic acid (LTA) anchored to the cell membrane has attracted attention as a host immunomodulator. Chemical and biological characteristics of LTA from various bacteria have been described. However, most of the information concerns pathogenic bacteria, and information on beneficial bacteria, including probiotic lactic acid bacteria, is insufficient. LTA is structurally diverse. Strain-level structural diversity of LTA is suggested to underpin its immunomodulatory activities. Thus, the structural information on LTA in probiotics, in particular strain-associated diversity, is important for understanding its beneficial roles associated with the modulation of immune response. Continued accumulation of structural information is necessary to elucidate the detailed physiological roles and significance of LTA. In this review article, we summarize the current state of knowledge on LTA structure, in particular the structure of LTA from lactic acid bacteria. We also describe the significance of structural diversity and biological roles of LTA. PMID:27867802
Furtado, Danielle N; Todorov, Svetoslav D; Landgraf, Mariza; Destro, Maria T; Franco, Bernadette D G M
Lactic acid bacteria capable of producing bacteriocins and presenting probiotic potential open innovative technological applications in the dairy industry. In this study, a bacteriocinogenic strain (Lactococcus lactis subsp. lactis DF4Mi) was isolated from goat milk, and studied for its probiotic potential. Lc. lactis DF4Mi was resistant to acidic pH and oxbile, presented co-aggregation with Listeria monocytogenes, and was not affected by several drugs from different generic groups, being sensitive to most tested antibiotics. These properties indicate that this Lc. lactis strain can be used for enhancement of dairy foods safety and quality, in combination with potential probiotic properties.
Furtado, Danielle N.; Todorov, Svetoslav D.; Landgraf, Mariza; Destro, Maria T.; Franco, Bernadette D.G.M.
Lactic acid bacteria capable of producing bacteriocins and presenting probiotic potential open innovative technological applications in the dairy industry. In this study, a bacteriocinogenic strain (Lactococcus lactis subsp. lactis DF4Mi) was isolated from goat milk, and studied for its probiotic potential. Lc. lactis DF4Mi was resistant to acidic pH and oxbile, presented co-aggregation with Listeria monocytogenes, and was not affected by several drugs from different generic groups, being sensitive to most tested antibiotics. These properties indicate that this Lc. lactis strain can be used for enhancement of dairy foods safety and quality, in combination with potential probiotic properties. PMID:25477942
Coelho, M C; Silva, C C G; Ribeiro, S C; Dapkevicius, M L N E; Rosa, H J D
In the past years, there has been a particular focus on the application of bacteriocins produced by lactic acid bacteria (LAB) in controlling the growth of pathogenic bacteria in foods. The aim of this study was to select LAB strains with antimicrobial activity, previously isolated from a traditional Azorean artisanal cheese (Pico cheese), in order to identify those with the greatest potential in reducing Listeria monocytogenes in fresh cheese. Eight bacteriocin producer strains identified as Lactococcus lactis (1) and Enterococcus faecalis (7) were tested. In general, the bacteriocin-producing strains presented a moderate growth in fresh cheese at refrigeration temperatures (4 °C), increasing one log count in three days. They exhibited slow acidification capacity, despite the increased production of lactic acid displayed by some strains after 24h. Bacteriocin activity was only detected in the whey of fresh cheese inoculated with two Enterococcus strains, but all cheeses made with bacteriocin-producing strains inhibited L. monocytogenes growth in the agar diffusion bioassay. No significant differences were found in overall sensory evaluation made by a non-trained panel of 50-52 tasters using the isolates as adjunct culture in fresh cheese, with the exception of one Enterococcus strain. To test the effect of in situ bacteriocin production against L. monocytogenes, fresh cheese was made from pasteurized cows' milk inoculated with bacteriocin-producing LAB and artificially contaminated with approximately 10(6) CFU/mL of L. monocytogenes. The numbers of L. monocytogenes were monitored during storage of fresh cheese at refrigeration temperature (4 °C) for up to 15 days. All strains controlled the growth of L. monocytogenes, although some Enterococcus were more effective in reducing the pathogen counts. After 7 days, this reduction was of approximately 4 log units compared to the positive control. In comparison, an increase of 4 log CFU/mL in pathogen numbers was
Genome analysis using next generation sequencing technologies has revolutionized the characterization of lactic acid bacteria and complete genomes of all major groups are now available. Comparative genomics has provided new insights into the natural and laboratory evolution of lactic acid bacteria and their environmental interactions. Moreover, functional genomics approaches have been used to understand the response of lactic acid bacteria to their environment. The results have been instrumental in understanding the adaptation of lactic acid bacteria in artisanal and industrial food fermentations as well as their interactions with the human host. Collectively, this has led to a detailed analysis of genes involved in colonization, persistence, interaction and signaling towards to the human host and its health. Finally, massive parallel genome re-sequencing has provided new opportunities in applied genomics, specifically in the characterization of novel non-GMO strains that have potential to be used in the food industry. Here, we provide an overview of the state of the art of these functional genomics approaches and their impact in understanding, applying and designing lactic acid bacteria for food and health. PMID:25186768
Douillard, François P; de Vos, Willem M
Genome analysis using next generation sequencing technologies has revolutionized the characterization of lactic acid bacteria and complete genomes of all major groups are now available. Comparative genomics has provided new insights into the natural and laboratory evolution of lactic acid bacteria and their environmental interactions. Moreover, functional genomics approaches have been used to understand the response of lactic acid bacteria to their environment. The results have been instrumental in understanding the adaptation of lactic acid bacteria in artisanal and industrial food fermentations as well as their interactions with the human host. Collectively, this has led to a detailed analysis of genes involved in colonization, persistence, interaction and signaling towards to the human host and its health. Finally, massive parallel genome re-sequencing has provided new opportunities in applied genomics, specifically in the characterization of novel non-GMO strains that have potential to be used in the food industry. Here, we provide an overview of the state of the art of these functional genomics approaches and their impact in understanding, applying and designing lactic acid bacteria for food and health.
Beshkova, D M; Simova, E D; Frengova, G I; Simov, Z I; Adilov, E F
The dynamics of free amino acid production by the selected strains Streptococcus thermophilus 13a and Lactobacillus bulgaricus 2-11 were studied in pure and mixed cultivations during yogurt starter culture manufacture. L. bulgaricus 2-11 showed the highest activity for producing free amino acids with high individual concentrations over the first hour of growth (50% of the total amount). By the end of milk's full coagulation (4.5 h), 70% of the total amount of amino acids was released. S. thermophilus 13a showed poor proteolytic properties and consumed up to 70% of the free amino acids produced by L. bulgaricus 2-11 in the process of coagulation of milk with the mixed culture.
Hauri, J.F.; Schaider, L.A.
Sulfate reducing bacteria have been shown to be effective at treating acid mine drainage through sulfide production and subsequent precipitation of metal sulfides. In this laboratory experiment for undergraduate environmental chemistry courses, students design and implement a set of bioreactors to remediate acid mine drainage and explain observed changes in dissolved metal concentrations and pH. Using synthetic acid mine drainage and combinations of inputs, students monitor their bioreactors for decreases in dissolved copper and iron concentrations.
Mihara, Y; Utagawa, T; Yamada, H; Asano, Y
We have investigated the enzymatic phosphorylation of nucleosides and found that Morganella morganii phoC acid phosphatase exhibits regioselective pyrophosphate (PP(i))-nucleoside phosphotransferase activity. In this study, we isolated genes encoding an acid phosphatase with regioselective phosphotransferase activity (AP/PTase) from Providencia stuartii, Enterobacter aerogenes, Escherichia blattae and Klebsiella planticola, and compared the primary structures and enzymatic characteristics of these enzymes with those of AP/PTase (PhoC acid phosphatase) from M. morganii. The enzymes were highly homologous in primary structure with M. morganii AP/PTase, and are classified as class A1 acid phosphatases. The synthesis of inosine-5'-monophosphate (5'-IMP) by E. coli overproducing each acid phosphatase was investigated. The P. stuartii enzyme, which is most closely related to the M. morganii enzyme, exhibited high 5'-IMP productivity, similar to the M. morganii enzyme. The 5'-IMP productivities of the E. aerogenes, E. blattae and K. planticola enzymes were inferior to those of the former two enzymes. This result underlines the importance of lower K(m) values for efficient nucleotide production. As these enzymes exhibited a very high degree of homology at the amino acid sequence level, it is likely that local sequence differences in the binding pocket are responsible for the differences in the nucleoside-PP(i) phosphotransferase reaction.
Varadaraj, M C; Devi, N; Keshava, N; Manjrekar, S P
Neutralized extracellular culture filtrate obtained from isolates of Lactobacillus acidophilus, Lactobacillus delbruecki ssp. bulgaricus, Lactobacillus salivarius and Lactococcus lactis ssp. lactis from 'dahi' showed weak to moderate inhibition of Staphylococcus aureus, Bacillus cereus, Escherichia coli, Bacillus brevis, Bacillus circulans, Bacillus coagulans, Bacillus laterosporus, Bacillus subtilis and Pseudomonas aeruginosa when tested by the diffusion agar well assay method. The effective minimum quantity of lactic culture filtrates required to obtain complete inhibition of an inoculum of 10(3) cfu/ml of the bacteria tested was between 20 and 26% (vol/vol), as determined by the agar incorporation method. Neutralized extracellular culture filtrate of these lactic cultures added at a level of 10% in sterile, 10% reconstituted non-fat dry milk was able to either suppress or retard growth of selected bacterial cultures when incubated at 37 degrees C for 24 h. This study indicated the antimicrobial activity of dahi and the potential of using neutralized extracellular culture filtrate of lactic acid bacteria in the biopreservation of foods.
Gullo, Maria; Caggia, Cinzia; De Vero, Luciana; Giudici, Paolo
This study evaluated the glucose tolerance of acetic acid bacteria strains isolated from Traditional Balsamic Vinegar. The results showed that the greatest hurdle to acetic acid bacteria growth is the high sugar concentration, since the majority of the isolated strains are inhibited by 25% of glucose. Sugar tolerance is an important technological trait because Traditional Balsamic Vinegar is made with concentrated cooked must. On the contrary, ethanol concentration of the cooked and fermented must is less significant for acetic acid bacteria growth. A tentative identification of the isolated strains was done by 16S-23S-5S rDNA PCR/RFLP technique and the isolated strains were clustered: 32 strains belong to Gluconacetobacter xylinus group, two strains to Acetobacter pasteurianus group and one to Acetobacter aceti.
Glenn, A R; Reeve, W G; Tiwari, R P; Dilworth, M J
Biological nitrogen fixation, especially via the legume Rhizobium symbiosis, is important for world agriculture. The productivity of legume crops and pastures is significantly affected by soil acidity; in some cases it is the prokaryotic partner that is pH sensitive. Growth of Rhizobium is adversely affected by low pH, especially in the 'acid stress zone'. Rhizobia exhibit an adaptive acid tolerance response (ATR) that is influenced by calcium concentration. Using Tn5-mutagenesis, gusA fusions and 'proteome' analysis, we have identified a range of genes that are essential for growth at low pH (such as actA, actP, exoR, actR and actS). At least three regulatory systems exist. The two-component sensor-regulator system, actSR, is essential for induction of the adaptive ATR. Two other regulatory circuits exist that are independent of ActR. One system involves the low pH-induced regulator gene, phrR, which may control other low pH-regulated genes. The other circuit, involving a regulator that is yet unidentified, controls the expression of a pH-regulated structural gene (lpiA). We have used pH-responsive gusA fusions to identify acid-inducible genes (such as lpiA), and then attempted to identify the regulators of these genes. The emerging picture is of a relatively complex set of systems that respond to external pH.
In this study, traditional culture method and 16S rRNA gene analysis were applied to reveal the composition and diversity of lactic acid bacteria (LAB) of fermented cow milk, huruud and urum from Baotou and Bayannur of midwestern Inner Mongolia. Also, the quantitative results of dominant LAB species in three different types of dairy products from Baotou and Bayannur were gained by quantitative polymerase chain reaction (q-PCR) technology. Two hundred and two LAB strains isolated from sixty-six samples were identified and classified into four genera, namely Enterococcus, Lactococcus, Lactobacillus, Leuconostoc, and twenty-one species and subspecies. From these isolates, Lactococcus lactis subsp. lactis (32.18%), Lactobacillus plantarum (12.38%) and Leuconosto mesenteroides (11.39%) were considered as the dominated LAB species under the condition of cultivating in MRS and M17 medium. And the q-PCR results revealed that the number of dominant species varied from samples to samples and from region to region. This study clearly shows the composition and diversity of LAB existing in fermented cow milk, huruud and urum, which could be considered as valuable resources for LAB isolation and further probiotic selection. PMID:27621691
Allegretti, L; Revolledo, L; Astolfi-Ferreira, C S; Chacón, J L; Martins, L M; Seixas, G H F; Ferreira, A J P
In Brazil, the blue-fronted Amazon parrot (Amazona aestiva) is a common pet. The faecal microbiota of these birds include a wide variety of bacterial species, the majority of which belong to the Gram-positive lactic acid bacteria (LAB) clade. The aim of this study was to investigate differences in the diversity and abundance of LAB and Bifidobacterium spp. in the cloacae between wild and captive birds and to select, identify and characterise LAB for consideration as a parrot probiotic. Cloacal swabs were collected from 26 wild and 26 captive birds. Bacterial DNA was extracted, and the 16S rRNA genes were amplified. The numbers of PCR-positive Enterococcus, Pediococcus, and Lactobacillus species isolated from wild and captive birds were significantly different (P<0.05). Enterococcus was the most frequently isolated genus, followed by Pediococcus, Lactobacillus, Lactococcus and Bifidobacterium. Enterococcus faecium, Pediococcus pentosaceus, Lactococcus lactis, Lactobacillus coryniformis, Lactobacillus sanfranciscensis and Bifidobacterium bifidum were the most frequently isolated species from all birds. This study increases our understanding of the faecal microbiota, and may help to improve the nutrition and habitat management of captive and wild parrots. The bacterial population identified in the faecal microbiota of clinically healthy wild and captive parrots can serve as a database to analyse variations in the gut microbiota of pathogen-infected parrots and to develop probiotics specific to these genera.
Saraoui, Taous; Rault, Lucie; Germon, Pierre; Gonzalez-Moreno, Candelaria; Nader-Macias, Fatima M. E.; Baud, Damien; François, Patrice; Chuat, Victoria; Chain, Florian; Langella, Philippe; Nicoli, Jacques; Le Loir, Yves; Even, Sergine
Bovine mastitis is a costly disease in dairy cattle worldwide. As of yet, the control of bovine mastitis is mostly based on prevention by thorough hygienic procedures during milking. Additional strategies include vaccination and utilization of antibiotics. Despite these measures, mastitis is not fully under control, thus prompting the need for alternative strategies. The goal of this study was to isolate autochthonous lactic acid bacteria (LAB) from bovine mammary microbiota that exhibit beneficial properties that could be used for mastitis prevention and/or treatment. Sampling of the teat canal led to the isolation of 165 isolates, among which a selection of ten non-redundant LAB strains belonging to the genera Lactobacillus and Lactococcus were further characterized with regard to several properties: surface properties (hydrophobicity, autoaggregation); inhibition potential of three main mastitis pathogens, Staphylococcus aureus, Escherichia coli and Streptococcus uberis; colonization capacities of bovine mammary epithelial cells (bMEC); and immunomodulation properties. Three strains, Lactobacillus brevis 1595 and 1597 and Lactobacillus plantarum 1610, showed high colonization capacities and a medium surface hydrophobicity. These strains are good candidates to compete with pathogens for mammary gland colonization. Moreover, nine strains exhibited anti-inflammatory properties, as illustrated by the lower IL-8 secretion by E. coli-stimulated bMEC in the presence of these LAB. Full genome sequencing of five candidate strains allowed to check for undesirable genetic elements such as antibiotic resistance genes and to identify potential bacterial determinants involved in the beneficial properties. This large screening of beneficial properties while checking for undesirable genetic markers allowed the selection of promising candidate LAB strains from bovine mammary microbiota for the prevention and/or treatment of bovine mastitis. PMID:26713450
Feldman-Salit, Anna; Hering, Silvio; Messiha, Hanan L; Veith, Nadine; Cojocaru, Vlad; Sieg, Antje; Westerhoff, Hans V; Kreikemeyer, Bernd; Wade, Rebecca C; Fiedler, Tomas
Despite high similarity in sequence and catalytic properties, the l-lactate dehydrogenases (LDHs) in lactic acid bacteria (LAB) display differences in their regulation that may arise from their adaptation to different habitats. We combined experimental and computational approaches to investigate the effects of fructose 1,6-bisphosphate (FBP), phosphate (Pi), and ionic strength (NaCl concentration) on six LDHs from four LABs studied at pH 6 and pH 7. We found that 1) the extent of activation by FBP (Kact) differs. Lactobacillus plantarum LDH is not regulated by FBP, but the other LDHs are activated with increasing sensitivity in the following order: Enterococcus faecalis LDH2 ≤ Lactococcus lactis LDH2 < E. faecalis LDH1 < L. lactis LDH1 ≤ Streptococcus pyogenes LDH. This trend reflects the electrostatic properties in the allosteric binding site of the LDH enzymes. 2) For L. plantarum, S. pyogenes, and E. faecalis, the effects of Pi are distinguishable from the effect of changing ionic strength by adding NaCl. 3) Addition of Pi inhibits E. faecalis LDH2, whereas in the absence of FBP, Pi is an activator of S. pyogenes LDH, E. faecalis LDH1, and L. lactis LDH1 and LDH2 at pH 6. These effects can be interpreted by considering the computed binding affinities of Pi to the catalytic and allosteric binding sites of the enzymes modeled in protonation states corresponding to pH 6 and pH 7. Overall, the results show a subtle interplay among the effects of Pi, FBP, and pH that results in different regulatory effects on the LDHs of different LABs.
Atanasova, Jivka; Moncheva, Penka; Ivanova, Iskra
We examined 62 strains and 21 trade starter cultures from the collection of LB Bulgaricum PLC for proteolytic and antimicrobial activity of lactic acid bacteria (LAB) grown in goat milk. The aim of this study was to investigate the fermentation of caseins, α-lactalbumin and β-lactoglobulin by LAB, using the o-phthaldialdehyde (OPA) spectrophotometric assay and sodium dodecyl sulphate polyacrylamide gel electrophoresis (SDS-PAGE). The proteolysis targeted mainly caseins, especially β-casein. Whey proteins were proteolyzed, essentially β-lactoglobulin. The proteolytic activity of Lactococcus lactis l598, Streptococcus thermophilus t3D1, Dt1, Lactobacillus lactis 1043 and L. delbrueckii subsp. bulgaricus b38, b122 and b24 was notably high. The proteolysis process gave rise to medium-sized peptide populations. Most of the examined strains showed antimicrobial activity against some food pathogens, such as Escherichia coli, Staphylococcus aureus, Salmonella cholere enteridis, Listeria monocytogenes, Listeria innocua and Enterobacter aerogenes. The most active producers of antimicrobial-active peptides were strains of L. delbrueckii subsp. bulgaricus and S. thermophilus, which are of practical importance. The starter cultures containing the examined species showed high proteolytic and antimicrobial activity in skimmed goat milk. The greatest antimicrobial activity of the cultures was detected against E. aerogenes. The obtained results demonstrated the significant proteolytic potential of the examined strains in goat milk and their potential for application in the production of dairy products from goat's milk. The present results could be considered as the first data on the proteolytic capacity of strains and starter cultures in goat milk for the purposes of trade interest of LB Bulgaricum PLC.
Guarcello, Rosa; Carpino, Stefania; Gaglio, Raimondo; Pino, Alessandra; Rapisarda, Teresa; Caggia, Cinzia; Marino, Giovanni; Randazzo, Cinzia L; Settanni, Luca; Todaro, Massimo
The main hypothesis of this study was that the autochthonous lactic acid bacteria (LAB) selected for their dairy traits are able to stabilize the production of PDO (Protected Denomination of Origin) Pecorino Siciliano cheese, preserving its typicality. The experimental plan included the application of a multi-strain lactic acid bacteria (LAB) culture, composed of starter (Lactococcus lactis subsp. lactis CAG4 and CAG37) and non starter (Enterococcus faecalis PSL71, Lactococcus garviae PSL67 and Streptococcus macedonicus PSL72) strains, during the traditional production of cheese at large scale level in six factories located in different areas of Sicily. The cheese making processes were followed from milk to ripened cheeses and the effects of the added LAB were evaluated on the microbiological, chemico-physical and sensorial characteristics of the final products. Results highlighted a high variability for all investigated parameters and the dominance of LAB cocci in bulk milk samples. The experimental curds showed a faster pH drop than control curds and the levels of LAB estimated in 5-month ripened experimental cheeses (7.59 and 7.27 Log CFU/g for rods and cocci, respectively) were higher than those of control cheeses (7.02 and 6.61 Log CFU/g for rods and cocci, respectively). The comparison of the bacterial isolates by randomly amplified polymorphic DNA (RAPD)-PCR evidenced the dominance of the added starter lactococci over native milk and vat LAB, while the added non starter LAB were found at almost the same levels of the indigenous strains. The sensory evaluation showed that the mixed LAB culture did not influence the majority of the sensory attributes of the cheeses and that each factory produced cheeses with unique characteristics. Finally, the multivariate statistical analysis based on all parameters evaluated on the ripened cheeses showed the dissimilarities and the relationships among cheeses. Thus, the main hypothesis of the work was accepted since the
Saichana, Natsaran; Matsushita, Kazunobu; Adachi, Osao; Frébort, Ivo; Frebortova, Jitka
Acetic acid bacteria are gram-negative obligate aerobic bacteria assigned to the family Acetobacteraceae of Alphaproteobacteria. They are members of the genera Acetobacter, Gluconobacter, Gluconacetobacter, Acidomonas, Asaia, Kozakia, Swaminathania, Saccharibacter, Neoasaia, Granulibacter, Tanticharoenia, Ameyamaea, Neokomagataea, and Komagataeibacter. Many strains of Acetobacter and Komagataeibacter have been known to possess high acetic acid fermentation ability as well as the acetic acid and ethanol resistance, which are considered to be useful features for industrial production of acetic acid and vinegar, the commercial product. On the other hand, Gluconobacter strains have the ability to perform oxidative fermentation of various sugars, sugar alcohols, and sugar acids leading to the formation of several valuable products. Thermotolerant strains of acetic acid bacteria were isolated in order to serve as the new strains of choice for industrial fermentations, in which the cooling costs for maintaining optimum growth and production temperature in the fermentation vessels could be significantly reduced. Genetic modifications by adaptation and genetic engineering were also applied to improve their properties, such as productivity and heat resistance.
Bancalari, Elena; Bernini, Valentina; Bottari, Benedetta; Neviani, Erasmo; Gatti, Monica
Impedance microbiology is a method that enables tracing microbial growth by measuring the change in the electrical conductivity. Different systems, able to perform this measurement, are available in commerce and are commonly used for food control analysis by mean of measuring a point of the impedance curve, defined "time of detection." With this work we wanted to find an objective way to interpret the metabolic significance of impedance curves and propose it as a valid approach to evaluate the potential acidifying performances of starter lactic acid bacteria to be employed in milk transformation. To do this it was firstly investigated the possibility to use the Gompertz equation to describe the data coming from the impedance curve obtained by mean of BacTrac 4300®. Lag time (λ), maximum specific M% rate (μmax), and maximum value of M% (Yend) have been calculated and, given the similarity of the impedance fitted curve to the bacterial growth curve, their meaning has been interpreted. Potential acidifying performances of eighty strains belonging to Lactobacillus helveticus, Lactobacillus delbrueckii subsp. bulgaricus, Lactococcus lactis, and Streptococcus thermophilus species have been evaluated by using the kinetics parameters, obtained from Excel add-in DMFit version 2.1. The novelty and importance of our findings, obtained by means of BacTrac 4300®, is that they can also be applied to data obtained from other devices. Moreover, the meaning of λ, μmax, and Yend that we have extrapolated from Modified Gompertz equation and discussed for lactic acid bacteria in milk, can be exploited also to other food environment or other bacteria, assuming that they can give a curve and that curve is properly fitted with Gompertz equation.
Zanirati, Débora Ferreira; Abatemarco, Mário; Sandes, Sávio Henrique de Cicco; Nicoli, Jacques Robert; Nunes, Álvaro Cantini; Neumann, Elisabeth
Brazilian kefir is a homemade fermented beverage that is obtained by incubating milk or a brown sugar solution with kefir grains that contribute their different microbiological compositions. It is highly important to isolate and characterize microorganisms from Brazilian kefir grains to obtain starter cultures for the industrial production of a standardized commercial kefir. Thus, the present study aimed to isolate lactic acid bacteria from eight kefir grains that were propagated in milk or sugar solutions from five different locations in Brazil and to select Lactobacillus isolates based on desirable in vitro probiotic properties. One hundred eight isolates from both substrates were identified by amplified ribosomal DNA restriction analysis and/or 16S rRNA gene sequencing and were determined to belong to the following 11 species from the genera: Lactococcus, Leuconostoc, Lactobacillus (L.), and Oenococcus. Leuconostoc mesenteroides, Lactobacillus kefiri, and Lactobacillus kefiranofaciens were isolated only from milk grains, whereas Lactobacillus perolens, Lactobacillus parafarraginis, Lactobacillus diolivorans, and Oenococcus oeni were isolated exclusively from sugar water grains. When the microbial compositions of four kefir grains were evaluated with culture-independent analyses, L. kefiranofaciens was observed to predominant in milk grains, whereas Lactobacillus hilgardii was most abundant in sugar water kefir. Unfortunately, L. hilgardii was not isolated from any grain, although this bacteria was detected with a culture-independent methodology. Fifty-two isolated Lactobacilli were tested for gastric juice and bile salt tolerance, antagonism against pathogens, antimicrobial resistance, and surface hydrophobicity. Three Lactobacillus strains (L. kefiranofaciens 8U, L. diolivorans 1Z, and Lactobacillus casei 17U) could be classified as potential probiotics. In conclusion, several lactic acid bacteria that could be used in combination with yeasts as starter
Bancalari, Elena; Bernini, Valentina; Bottari, Benedetta; Neviani, Erasmo; Gatti, Monica
Impedance microbiology is a method that enables tracing microbial growth by measuring the change in the electrical conductivity. Different systems, able to perform this measurement, are available in commerce and are commonly used for food control analysis by mean of measuring a point of the impedance curve, defined “time of detection.” With this work we wanted to find an objective way to interpret the metabolic significance of impedance curves and propose it as a valid approach to evaluate the potential acidifying performances of starter lactic acid bacteria to be employed in milk transformation. To do this it was firstly investigated the possibility to use the Gompertz equation to describe the data coming from the impedance curve obtained by mean of BacTrac 4300®. Lag time (λ), maximum specific M% rate (μmax), and maximum value of M% (Yend) have been calculated and, given the similarity of the impedance fitted curve to the bacterial growth curve, their meaning has been interpreted. Potential acidifying performances of eighty strains belonging to Lactobacillus helveticus, Lactobacillus delbrueckii subsp. bulgaricus, Lactococcus lactis, and Streptococcus thermophilus species have been evaluated by using the kinetics parameters, obtained from Excel add-in DMFit version 2.1. The novelty and importance of our findings, obtained by means of BacTrac 4300®, is that they can also be applied to data obtained from other devices. Moreover, the meaning of λ, μmax, and Yend that we have extrapolated from Modified Gompertz equation and discussed for lactic acid bacteria in milk, can be exploited also to other food environment or other bacteria, assuming that they can give a curve and that curve is properly fitted with Gompertz equation. PMID:27799925
Sun, H. J.; Zhang, G.
Recently, many groups reported D-amino acid uptake by plant roots, raising the question of whether soil D-amino acids represent a source of nitrogen or a source of toxicity. The discussion needs to be placed in the context of competition with rhizosphere bacteria. To provide this context, we followed the concentrations of D- and L-enantiomers of alanine, glutamic acid, aspartic acid, and leucine after they were added to soils in the laboratory. In all cases, the uptake of L-enantiomer began immediately and proceeded rapidly until exhausted. In contrast, the uptake of D-enantiomer required induction: an initial period of inactivity followed by rapid consumption comparable in rate to L-enantiomer. The induced nature of the D activity was confirmed by the addition of rifampicin, an mRNA synthesis inhibitor. Preventing the synthesis of new enzymes abolished soil flora's ability to consume D-amino acids, but not L-amino acids. These results suggest that inducible special racemase enzymes, which can convert D-amino acids back to their native L-forms, are widespread among soil microorganisms. This finding does not rule out the possibility that some plants may out-compete microorganisms and be able to access D-amino acids. It does suggest, however, that rhizosphere bacteria can shield plants from the toxic effect of D-amino acids.
Characterization and Determination of Origin of Lactic Acid Bacteria from a Sorghum-Based Fermented Weaning Food by Analysis of Soluble Proteins and Amplified Fragment Length Polymorphism Fingerprinting
Kunene, Nokuthula F.; Geornaras, Ifigenia; von Holy, Alexander; Hastings, John W.
The group that includes the lactic acid bacteria is one of the most diverse groups of bacteria known, and these organisms have been characterized extensively by using different techniques. In this study, 180 lactic acid bacterial strains isolated from sorghum powder (44 strains) and from corresponding fermented (93 strains) and cooked fermented (43 strains) porridge samples that were prepared in 15 households were characterized by using biochemical and physiological methods, as well as by analyzing the electrophoretic profiles of total soluble proteins. A total of 58 of the 180 strains were Lactobacillus plantarum strains, 47 were Leuconostoc mesenteroides strains, 25 were Lactobacillus sake-Lactobacillus curvatus strains, 17 were Pediococcus pentosaceus strains, 13 were Pediococcus acidilactici strains, and 7 were Lactococcus lactis strains. L. plantarum and L. mesenteroides strains were the dominant strains during the fermentation process and were recovered from 87 and 73% of the households, respectively. The potential origins of these groups of lactic acid bacteria were assessed by amplified fragment length polymorphism fingerprint analysis. PMID:10698775
Slower fermentation rates, mixed sugar compositions, and lower sugar concentrations may make lignocellulosic fermentations more susceptible to contamination by lactic acid bacteria (LAB), which is a common and costly problem to the corn-based fuel ethanol industry. To examine the effects of LAB con...
Lactic acid bacteria are among the powerhouses of the food industry, colonize the surfaces of plants and animals, and contribute to our health and well-being. The genomic characterization of LAB has rocketed and presently over 100 complete or nearly complete genomes are available, many of which serve as scientific paradigms. Moreover, functional and comparative metagenomic studies are taking off and provide a wealth of insight in the activity of lactic acid bacteria used in a variety of applications, ranging from starters in complex fermentations to their marketing as probiotics. In this new era of high throughput analysis, biology has become big science. Hence, there is a need to systematically store the generated information, apply this in an intelligent way, and provide modalities for constructing self-learning systems that can be used for future improvements. This review addresses these systems solutions with a state of the art overview of the present paradigms that relate to the use of lactic acid bacteria in industrial applications. Moreover, an outlook is presented of the future developments that include the transition into practice as well as the use of lactic acid bacteria in synthetic biology and other next generation applications. PMID:21995776
Quantitative analyses of the bacterial microbiota of rearing environment, tilapia and common carp cultured in earthen ponds and inhibitory activity of its lactic acid bacteria on fish spoilage and pathogenic bacteria.
Kaktcham, Pierre Marie; Temgoua, Jules-Bocamdé; Ngoufack Zambou, François; Diaz-Ruiz, Gloria; Wacher, Carmen; Pérez-Chabela, María de Lourdes
The present study aimed to evaluate the bacterial load of water, Nile Tilapia and common Carp intestines from earthen ponds, isolate lactic acid bacteria (LAB) and assess their antimicrobial activity against fish spoilage and pathogenic bacteria. Following enumeration and isolation of microorganisms the antimicrobial activity of the LAB isolates was evaluated. Taxonomic identification of selected antagonistic LAB strains was assessed, followed by partial characterisation of their antimicrobial metabolites. Results showed that high counts (>4 log c.f.u ml(-1) or 8 log c.f.u g(-1)) of total aerobic bacteria were recorded in pond waters and fish intestines. The microbiota were also found to be dominated by Salmonella spp., Vibrio spp., Staphylococcus spp. and Escherichia coli. LAB isolates (5.60%) exhibited potent direct and extracellular antimicrobial activity against the host-derived and non host-derived spoilage and pathogenic bacteria. These antagonistic isolates were identified and Lactococcus lactis subsp. lactis was found as the predominant (42.85%) specie. The strains displayed the ability to produce lactic, acetic, butyric, propionic and valeric acids. Bacteriocin-like inhibitory substances with activity against Gram-positive and Gram-negative (Vibrio spp. and Pseudomonas aeruginosa) bacteria were produced by three L. lactis subsp. lactis strains. In this study, the LAB from the microbiota of fish and pond water showed potent antimicrobial activity against fish spoilage or pathogenic bacteria from the same host or ecological niche. The studied Cameroonian aquatic niche is an ideal source of antagonistic LAB that could be appropriate as new fish biopreservatives or disease control agents in aquaculture under tropical conditions in particular or worldwide in general.
Balamurugan, Ramadass; Chandragunasekaran, Aarthi Sophia; Chellappan, Gowri; Rajaram, Krithika; Ramamoorthi, Gayathri; Ramakrishna, Balakrishnan S.
Background & objectives: The human gut microbiota play a significant role in nutritional processes. The concept of probiotics has led to widespread consumption of food preparations containing probiotic microbes such as curd and yogurt. Curd prepared at home is consumed every day in most homes in southern India. In this study the home-made curd was evaluated for lactic acid bacteria (LAB) with probiotic potential. Methods: Fifteen LAB (12 lactobacilli, 1 Lactococcus, 2 Leuconostoc) and one yeast isolated from home-made curd were evaluated for resistance to acid, pepsin, pancreatin and bile salts; antimicrobial resistance; intrinsic antimicrobial activity; adherence to Caco-2 epithelial cells; ability to block pathogen adherence to Caco-2 cells; ability to inhibit interleukin (IL)-8 secretion from HT-29 epithelial cells in response to Vibrio cholerae; and ability to induce anti-inflammatory cytokine expression in THP-1 monocyte cells. Results: Lactobacillus abundance in fermenting curd peaked sharply at 12 h. Nine of the strains survived exposure to acid (pH 3.0) for at least one hour, and all strains survived in the presence of pancreatin or bile salts for 3 h. None showed haemolytic activity. All were resistant to most antimicrobials tested, but were sensitive to imipenem. Most strains inhibited the growth of Salmonella Typhimurium while five inhibited growth of V. cholerae O139. Seven strains showed adherence to Caco-2 cells ranging from 20-104 per cent of adherence of an adherent strain of Escherichia coli, but all inhibited V. cholerae adherence to Caco-2 cells by 20-100 per cent. They inhibited interleukin-8 secretion from HT-29 cells, in response to V. cholerae, by 50-80 per cent. Two strains induced IL-10 and IL-12 messenger ribonucleic acid (mRNA) expression in THP-1 cells. Interpretation & conclusions: LAB in curd had properties consistent with probiotic potential, but these were not consistent across species. LAB abundance in curd increased rapidly at 12 h
Bartowsky, Eveline J; Henschke, Paul A
Acetic acid bacteria (AAB) are ubiquitous organisms that are well adapted to sugar and ethanol rich environments. This family of Gram-positive bacteria are well known for their ability to produce acetic acid, the main constituent in vinegar. The oxidation of ethanol through acetaldehyde to acetic acid is well understood and characterised. AAB form part of the complex natural microbial flora of grapes and wine, however their presence is less desirable than the lactic acid bacteria and yeast. Even though AAB were described by Pasteur in the 1850s, wine associated AAB are still difficult to cultivate on artificial laboratory media and until more recently, their taxonomy has not been well characterised. Wine is at most risk of spoilage during production and the presence of these strictly aerobic bacteria in grape must and during wine maturation can be controlled by eliminating, or at least limiting oxygen, an essential growth factor. However, a new risk, spoilage of wine by AAB after packaging, has only recently been reported. As wine is not always sterile filtered prior to bottling, especially red wine, it often has a small resident bacterial population (<10(3) cfu/mL), which under conducive conditions might proliferate. Bottled red wines, sealed with natural cork closures, and stored in a vertical upright position may develop spoilage by acetic acid bacteria. This spoilage is evident as a distinct deposit of bacterial biofilm in the neck of the bottle at the interface of the wine and the headspace of air, and is accompanied with vinegar, sherry, bruised apple, nutty, and solvent like off-aromas, depending on the degree of spoilage. This review focuses on the wine associated AAB species, the aroma and flavour changes in wine due to AAB metabolism, discusses the importance of oxygen ingress into the bottle and presents a hypothesis for the mechanism of spoilage of bottled red wine.
Zuljan, Federico; Espariz, Martín; Blancato, Victor S.; Esteban, Luis; Alarcón, Sergio
We report the draft genome sequence of Lactococcus lactis subsp. lactis bv. diacetylactis CRL264, a natural strain isolated from artisanal cheese from northwest Argentina. L. lactis subsp. lactis bv. diacetylactis is one of the most important microorganisms used as starter culture around the world. The CRL264 strain constitutes a model microorganism in the studies on the generation of aroma compounds (diacetyl, acetoin, and 2,3-butanediol) by lactic acid bacteria. Our genome analysis shows similar genetic organization to other available genomes of L. lactis bv. diacetylactis strains. PMID:26847906
Zuljan, Federico; Espariz, Martín; Blancato, Victor S; Esteban, Luis; Alarcón, Sergio; Magni, Christian
We report the draft genome sequence of Lactococcus lactis subsp. lactis bv. diacetylactis CRL264, a natural strain isolated from artisanal cheese from northwest Argentina. L. lactis subsp. lactis bv. diacetylactis is one of the most important microorganisms used as starter culture around the world. The CRL264 strain constitutes a model microorganism in the studies on the generation of aroma compounds (diacetyl, acetoin, and 2,3-butanediol) by lactic acid bacteria. Our genome analysis shows similar genetic organization to other available genomes of L. lactis bv. diacetylactis strains.
Bohn, Jelena; Yüksel-Dadak, Aytül; Dröge, Stefan; König, Helmut
Direct molecular approaches provide hints that lactic acid bacteria play an important role in the degradation process of organic material to methanogenetic substrates in biogas plants. However, their diversity in biogas fermenter samples has not been analyzed in detail yet. For that reason, five different biogas fermenters, which were fed mainly with maize silage and manure from cattle or pigs, were examined for the occurrence of lactic acid-forming bacteria. A total of 197 lactic acid-forming bacterial strains were isolated, which we assigned to 21 species, belonging to the genera Bacillus, Clostridium, Lactobacillus, Pediococcus, Streptococcus and Pseudoramibacter-related. A qualitative multiplex system and a real-time quantitative PCR could be developed for most isolates, realized by the selection of specific primers. Their role in biogas plants was discussed on the basis of the quantitative results and on physiological data of the isolates.
Yang, Yiling; M. Pollard, Abiola; Höfler, Carolin; Poschet, Gernot; Wirtz, Markus; Hell, Rüdiger
Summary Chemotaxis enables bacteria to navigate chemical gradients in their environment, accumulating toward high concentrations of attractants and avoiding high concentrations of repellents. Although finding nutrients is likely to be an important function of bacterial chemotaxis, not all characterized attractants are nutrients. Moreover, even for potential nutrients, the exact relation between the metabolic value of chemicals and their efficiency as chemoattractants has not been systematically explored. Here we compare the chemotactic response of amino acids with their use by bacteria for two well‐established models of chemotactic behavior, E scherichia coli and B acillus subtilis. We demonstrate that in E . coli chemotaxis toward amino acids indeed strongly correlates with their utilization. However, no such correlation is observed for B . subtilis, suggesting that in this case, the amino acids are not followed because of their nutritional value but rather as environmental cues. PMID:25807888
Thakur, Kiran; De, Sachinandan
Summary Consumers are increasingly becoming aware of their health and nutritional requirements, and in this context, vitamins produced in situ by microbes may suit their needs and expectations. B groups vitamins are essential components of cellular metabolism and among them riboflavin is one of the vital vitamins required by bacteria, plants, animals and humans. Here, we focus on the importance of microbial production of riboflavin over chemical synthesis. In addition, genetic abilities for riboflavin biosynthesis by lactic acid bacteria are discussed. Genetically modified strains by employing genetic engineering and chemical analogues have been developed to enhance riboflavin production. The present review attempts to collect the currently available information on riboflavin production by microbes in general, while placing greater emphasis on food grade lactic acid bacteria and human gut commensals. For designing riboflavin‐enriched functional foods, proper selection and exploitation of riboflavin‐producing lactic acid bacteria is essential. Moreover, eliminating the in situ vitamin fortification step will decrease the cost of food production. PMID:26686515
Trček, Janja; Mira, Nuno Pereira; Jarboe, Laura R
Acetic acid is a weak organic acid exerting a toxic effect to most microorganisms at concentrations as low as 0.5 wt%. This toxic effect results mostly from acetic acid dissociation inside microbial cells, causing a decrease of intracellular pH and metabolic disturbance by the anion, among other deleterious effects. These microbial inhibition mechanisms enable acetic acid to be used as a preservative, although its usefulness is limited by the emergence of highly tolerant spoilage strains. Several biotechnological processes are also inhibited by the accumulation of acetic acid in the growth medium including production of bioethanol from lignocellulosics, wine making, and microbe-based production of acetic acid itself. To design better preservation strategies based on acetic acid and to improve the robustness of industrial biotechnological processes limited by this acid's toxicity, it is essential to deepen the understanding of the underlying toxicity mechanisms. In this sense, adaptive responses that improve tolerance to acetic acid have been well studied in Escherichia coli and Saccharomyces cerevisiae. Strains highly tolerant to acetic acid, either isolated from natural environments or specifically engineered for this effect, represent a unique reservoir of information that could increase our understanding of acetic acid tolerance and contribute to the design of additional tolerance mechanisms. In this article, the mechanisms underlying the acetic acid tolerance exhibited by several bacterial strains are reviewed, with emphasis on the knowledge gathered in acetic acid bacteria and E. coli. A comparison of how these bacterial adaptive responses to acetic acid stress fit to those described in the yeast Saccharomyces cerevisiae is also performed. A systematic comparison of the similarities and dissimilarities of the ways by which different microbial systems surpass the deleterious effects of acetic acid toxicity has not been performed so far, although such exchange
Rattanachaikunsopon, Pongsak; Phumkhachorn, Parichat
Lactic acid bacteria isolated from various Thai fermented foods were screened for the presence of nisin gene by using PCR with primers specific to nisin A structural gene. Only one strain, Lactococcus lactis subsp. lactis TFF 221, isolated from kung jom, a traditional shrimp paste, was found to carry a nisin gene. The TFF 221 nisin had antimicrobial activity against not only closely related lactic acid bacteria but also some foodborne pathogens. It was heat stable and inactivated by alpha-chymotrypsin and proteinase K. Some characteristics of TFF 221 nisin were found to be very similar to those of nisin A produced by Lactococcus lactis subsp. lactis NCDO 2111. Both of them had the same antimicrobial spectrum and MICs against all indicator bacteria. However, when assayed with indicator organisms, in all cases the TFF 221 nisin produced larger zones of inhibition in agar diffusion assays than the nisin A did. Sequencing of the TFF 221 nisin gene showed that it was the natural nisin variant, nisin Z, as indicated by the substitution of asparagine residue instead of histidine at position 27. The nisin determinant in strain TFF 221 was found to be located on a conjugative transposon residing in the chromosome. The ability of the nisin produced by L. lactis subsp. lactis TFF 221 to inhibit a wide range of foodborne pathogens may be useful in improving the food safety of the fermented product, especially in the Thai environment, which suffers from perennial problems of poor food hygiene.
Zareian, Mohsen; Ebrahimpour, Afshin; Bakar, Fatimah Abu; Mohamed, Abdul Karim Sabo; Forghani, Bita; Ab-Kadir, Mohd Safuan B; Saari, Nazamid
l-glutamaic acid is the principal excitatory neurotransmitter in the brain and an important intermediate in metabolism. In the present study, lactic acid bacteria (218) were isolated from six different fermented foods as potent sources of glutamic acid producers. The presumptive bacteria were tested for their ability to synthesize glutamic acid. Out of the 35 strains showing this capability, strain MNZ was determined as the highest glutamic-acid producer. Identification tests including 16S rRNA gene sequencing and sugar assimilation ability identified the strain MNZ as Lactobacillus plantarum. The characteristics of this microorganism related to its glutamic acid-producing ability, growth rate, glucose consumption and pH profile were studied. Results revealed that glutamic acid was formed inside the cell and excreted into the extracellular medium. Glutamic acid production was found to be growth-associated and glucose significantly enhanced glutamic acid production (1.032 mmol/L) compared to other carbon sources. A concentration of 0.7% ammonium nitrate as a nitrogen source effectively enhanced glutamic acid production. To the best of our knowledge this is the first report of glutamic acid production by lactic acid bacteria. The results of this study can be further applied for developing functional foods enriched in glutamic acid and subsequently γ-amino butyric acid (GABA) as a bioactive compound.
Khassehkhan, Hassan; Eberl, Hermann J.
We study a previously introduced mathematical model of amensalistic control of the foodborne pathogen Listeria monocytogenes by the generally regarded as safe lactic acid bacteria Lactococcus lactis in a chemostat setting under nutrient rich growth conditions. The control agent produces lactic acids and thus affects pH in the environment such that it becomes detrimental to the pathogen while it is much more tolerant to these self-inflicted environmental changes itself. The mathematical model consists of five nonlinear ordinary differential equations for both bacterial species, the concentration of lactic acids, the pH and malate. The model is algebraically too involved to allow a comprehensive, rigorous qualitative analysis. Therefore, we conduct a computational study. Our results imply that depending on the growth characteristics of the medium in which the bacteria are cultured, the pathogen can survive in an intermediate flow regime but will be eradicated for slower flow rates and washed out for higher flow rates. PMID:28231156
The cell wall of Gram-positive bacteria is a complex assemblage of glycopolymers and proteins. It consists of a thick peptidoglycan sacculus that surrounds the cytoplasmic membrane and that is decorated with teichoic acids, polysaccharides, and proteins. It plays a major role in bacterial physiology since it maintains cell shape and integrity during growth and division; in addition, it acts as the interface between the bacterium and its environment. Lactic acid bacteria (LAB) are traditionally and widely used to ferment food, and they are also the subject of more and more research because of their potential health-related benefits. It is now recognized that understanding the composition, structure, and properties of LAB cell walls is a crucial part of developing technological and health applications using these bacteria. In this review, we examine the different components of the Gram-positive cell wall: peptidoglycan, teichoic acids, polysaccharides, and proteins. We present recent findings regarding the structure and function of these complex compounds, results that have emerged thanks to the tandem development of structural analysis and whole genome sequencing. Although general structures and biosynthesis pathways are conserved among Gram-positive bacteria, studies have revealed that LAB cell walls demonstrate unique properties; these studies have yielded some notable, fundamental, and novel findings. Given the potential of this research to contribute to future applied strategies, in our discussion of the role played by cell wall components in LAB physiology, we pay special attention to the mechanisms controlling bacterial autolysis, bacterial sensitivity to bacteriophages and the mechanisms underlying interactions between probiotic bacteria and their hosts. PMID:25186919
Shangpliang, H. N. J.; Sharma, Sharmila; Rai, Ranjita; Tamang, Jyoti P.
Dahi and datshi are common naturally fermented milk (NFM) products of Bhutan. Population of lactic acid bacteria (LAB) in dahi (pH 3.7) and datshi (pH 5.2) was 1.4 × 107 and 3.9 × 108 cfu/ml, respectively. Based on 16S rRNA gene sequencing isolates of LAB from dahi and datshi were identified as Enterococcus faecalis, E. faecium, Lactococcus lactis subsp. lactis. LAB strains were tested for some technological properties. All LAB strains except E. faecalis CH2:17 caused coagulation of milk at both 30°C for 48 h. Only E. faecium DH4:05 strain was resistant to pH 3. No significant difference (P > 0.05) of viable counts was observed in MRS broth with and without lysozyme. All LAB strains grew well in 0.3% bile showing their ability to tolerate bile salt. None of the LAB strains showed >70% hydrophobicity. This study, being the first of its microbiological analysis of the NFM of Bhutan, has opened up to an extent of research work that gives a new insight to the products. PMID:28203227
Kaur, Baljinder; Kumar, Balvir
Vanillin is widely used as food additive and as a masking agent in various pharmaceutical formulations. Ferulic acid is an important precursor of vanillin that is available in abundance in cell walls of cereals like wheat, corn, and rice. Phenolic biotransformations can occur during growth of lactic acid bacteria (LAB), and their production can be made feasible using specialized LAB strains that have been reported to produce ferulic acid esterases. The present study aimed at screening a panel of LAB isolates for their ability to release phenolics from agrowaste materials like rice bran and their biotransformation to industrially important compounds such as ferulic acid, 4-ethyl phenol, vanillic acid, vanillin, and vanillyl alcohol. Bacterial isolates were evaluated using ferulic acid esterase, ferulic acid decarboxylase, and vanillin dehydrogenase assays. This work highlights the importance of lactic acid bacteria in phenolic biotransformations for the development of food grade flavours and additives. PMID:24066293
Alvarez-Sieiro, Patricia; Montalbán-López, Manuel; Mu, Dongdong; Kuipers, Oscar P
Lactic acid bacteria (LAB) constitute a heterogeneous group of microorganisms that produce lactic acid as the major product during the fermentation process. LAB are Gram-positive bacteria with great biotechnological potential in the food industry. They can produce bacteriocins, which are proteinaceous antimicrobial molecules with a diverse genetic origin, posttranslationally modified or not, that can help the producer organism to outcompete other bacterial species. In this review, we focus on the various types of bacteriocins that can be found in LAB and the organization and regulation of the gene clusters responsible for their production and biosynthesis, and consider the food applications of the prototype bacteriocins from LAB. Furthermore, we propose a revised classification of bacteriocins that can accommodate the increasing number of classes reported over the last years.
Sonar, Nitin R; Halami, Prakash M
Fermented bamboo shoots such as Soibum, Soidon, Eup, Hirring, Hecche and Ekung etc. are non-salted acidic products obtained by natural fermentation predominantly with lactic acid bacteria (LAB). In this study, we have characterized 11-representative LAB that includes, Lactobacillus sp. (n = 2), Lactobacillus plantarum (n = 3), and one each of Lactobacillus fermentum, Lactococcus sp., Lactobacillus brevis, Lactobacillus curvatus, Leuconostoc sp. and Lactobacillus xylosus. Subsequently, these cultures were studied for their technological and functional properties. Different isolates exhibited variation in their activities. L. brevis showed maximum phytic acid degradation ability (19.33 U ml (-1) ). L. xylosus had highest protease activity (64.2 nmol/ml) and also exhibited lipolytic activity. In addition, degree of cell hydrophobicity among these cultures ranged between 12.5 and 93.48 % with L. plantarum (SM2) showing highest degree of activity. Lactobacillus plantarum was the most common species found in the product studied. Results indicated that most of the LABs showed putative probiotic as well as antagonistic properties against the selected pathogenic bacteria. Characteristic aroma, flavour and texture in the fermented bamboo shoot products could be attributed to presence of these new LAB isolates.
Fajardo, Paula; Pastrana, Lorenzo; Méndez, Jesús; Rodríguez, Isabel; Fuciños, Clara; Guerra, Nelson P.
The aim of this study was to evaluate the potential of two probiotic preparations, containing live lactic acid bacteria (Lactococcus lactis CECT 539 and Lactobacillus casei CECT 4043) and their products of fermentation (organic acids and bacteriocins), as a replacement for antibiotics in stimulating health and growth of broiler chickens. The effects of the supplementation of both preparations (with proven probiotic effect in weaned piglets) and an antibiotic (avilamycin) on body weight gain (BWG), feed intake (FI), feed consumption efficiency (FCE), relative intestinal weight, and intestinal microbiota counts were studied in 1-day posthatch chickens. The experiments were conducted with medium-growth Sasso X44 chickens housed in cages and with nutritional stressed Ross 308 broiler distributed in pens. Consumption of the different diets did not affect significantly the final coliform counts in Sasso X44 chickens. However, counts of lactic acid bacteria and mesophilic microorganisms were higher in the animals receiving the two probiotic preparations (P < 0.05). In the second experiment, although no differences in BWG were observed between treatments, Ross 308 broilers receiving the probiotic Lactobacillus preparation exhibited the lowest FCE values and were considered the most efficient at converting feed into live weight. PMID:22666137
Konkit, Maytiya; Choi, Woo Jin; Kim, Wonyong
Many human gastrointestinal facultative anaerobic and aerobic bacteria possess alcohol dehydrogenase (ADH) activity and are therefore capable of oxidizing ethanol to acetaldehyde. However, the ADH activity of Lactococcus spp., except Lactococcus lactis ssp. lactis, has not been widely determined, though they play an important role as the starter for most cheesemaking technologies. Cheese is a functional food recognized as an aid to digestion. In the current study, the ADH activity of Lactococcus chungangensis CAU 28(T) and 11 reference strains from the genus Lactococcus was determined. Only 5 strains, 3 of dairy origin, L. lactis ssp. lactis KCTC 3769(T), L. lactis ssp. cremoris KCCM 40699(T), and Lactococcus raffinolactis DSM 20443(T), and 2 of nondairy origin, Lactococcus fujiensis NJ317(T) and Lactococcus chungangensis CAU 28(T) KCTC 13185(T), showed ADH activity and possessed the ADH gene. All these strains were capable of making cheese, but the highest level of ADH activity was found in L. chungangensis, with 45.9nmol/min per gram in tryptic soy broth and 65.8nmol/min per gram in cream cheese. The extent that consumption of cheese, following imbibing alcohol, reduced alcohol uptake was observed by following the level of alcohol in the serum of mice. The results show a potential novel benefit of cheese as a dairy functional food.
Konkit, Maytiya; Choi, Woo Jin; Kim, Wonyong
Previous studies have shown that the metabolic capability of colonic microflora may be at least as high as that of the liver or higher than that of the whole human body. Aldehyde dehydrogenase (ALDH) is an enzyme produced by these bacteria that can metabolize acetaldehyde, produce from ethanol to acetate. Lactococcus species, which is commonly used as a starter in dairy products, was recently found to possess the ALDH gene, and the activity of this enzyme was determined. In this study, the ALDH activity of Lactococcus chungangensis CAU 28(T) and 11 other type strains in the genus Lactococcus was studied. Only 5 species, 3 of dairy origin (Lactococcus lactis ssp. lactis KCTC 3769(T), Lactococcus lactis ssp. cremoris KCCM 40699(T), and Lactococcus raffinolactis DSM 20443(T)) and 2 of nondairy origin (Lactococcus fujiensis NJ317(T) and L. chungangensis CAU 28(T)), showed ALDH activity and possessed a gene encoding ALDH. All of these strains were capable of making cream cheese. Among the strains, L. chungangensis produced cream cheese that contained the highest level of ALDH and was found to reduce the level of acetaldehyde in the serum of mice. These results predict a promising role for L. chungangensis CAU28(T) to be used in cheese that can be developed as functional food.
Ahlberg, Sara H; Joutsjoki, Vesa; Korhonen, Hannu J
Aflatoxins (AF) are ubiquitous mycotoxins contaminating food and feed. Consumption of contaminated food and feed can cause a severe health risk to humans and animals. A novel biological method could reduce the health risks of aflatoxins through inhibiting mold growth and binding aflatoxins. Lactic acid bacteria (LAB) are commonly used in fermented food production. LAB are known to inhibit mold growth and, to some extent, to bind aflatoxins in different matrices. Reduced mold growth and aflatoxin production may be caused by competition for nutrients between bacterial cells and fungi. Most likely, binding of aflatoxins depends on environmental conditions and is strain-specific. Killed bacteria cells possess consistently better binding abilities for aflatoxin B1 (AFB1) than viable cells. Lactobacilli especially are relatively well studied and provide noticeable possibilities in binding of aflatoxin B1 and M1 in food. It seems that binding is reversible and that bound aflatoxins are released later on (Haskard et al., 2001; Peltonen et al., 2001). This literature review suggests that novel biological methods, such as lactic acid bacteria, show potential in mitigating toxic effects of aflatoxins in food and feed.
In vivo pulse-label studies have demonstrated that luminescent bacteria can provide myritic acid (14:0) required for the synthesis of the luciferase substrate myristyl aldehyde. Luminescent wild type Vibrio harveyi incubated with (/sup 14/C) acetate in a nutrient-depleted medium accumulated substantial tree (/sup 14/C)fatty acid (up to 20% of the total lipid label). Radio-gas chromatography revealed that > 75% of the labeled fatty acid is 14:0. No free fatty acid was detected in wild type cells labeled prior to the development of bioluminescence in the exponential growth phase, or in a dark mutant of V. harveyi (mutant M17) that requires exogenous 14:0 for light emission. The preferential accumulation of 14:0 was not observed when wild type cells were labeled with (/sup 14/C)acetate in regular growth medium. Moreover, all V. harveyi strains exhibited similar fatty acid mass compositions regardless of the state of bioluminescence. Since earlier work has shown that a luminescence-related acyltransferase (defective in the M17 mutant) can catalyze the deacylation of fatty acyl-acyl carrier protein in vitro, the present results are consistent with a model in which this enzyme diverts 14:0 to the luminescence system during fatty acid biosynthesis. Under normal conditions, the supply of 14:0 by this pathway is tightly regulated such that bioluminescence development does not significantly alter the total fatty acid composition.
In the 1950s Corynebacterium glutamicum was found to be a very efficient producer of L-glutamic acid. Since this time biotechnological processes with bacteria of the species Corynebacterium developed to be among the most important in terms of tonnage and economical value. L-Glutamic acid and L-lysine are bulk products nowadays. L-Valine, L-isoleucine, L-threonine, L-aspartic acid and L-alanine are among other amino acids produced by Corynebacteria. Applications range from feed to food and pharmaceutical products. The growing market for amino acids produced with Corynebacteria led to significant improvements in bioprocess and downstream technology as well as in molecular biology. During the last decade big efforts were made to increase the productivity and to decrease the production costs. This review gives an overview of the world market for amino acids produced by Corynebacteria. Significant improvements in bioprocess technology, i.e. repeated fed batch or continuous production are summarised. Bioprocess technology itself was improved furthermore by application of more sophisticated feeding and automatisation strategies. Even though several amino acids developed towards commodities in the last decade, side aspects of the production process like sterility or detection of contaminants still have increasing relevance. Finally one focus of this review is on recent developments in downstream technology.
Gaglio, Raimondo; Scatassa, Maria Luisa; Cruciata, Margherita; Miraglia, Viviana; Corona, Onofrio; Di Gerlando, Rosalia; Portolano, Baldassare; Moschetti, Giancarlo; Settanni, Luca
Twelve lactic acid bacteria (LAB), previously selected in vitro (Gaglio et al., 2014), were evaluated in situ for their potential to act as starter cultures for the continuous four-season production of Vastedda-like cheese, made with raw ewes' milk. The strains belonged to Lactobacillus delbrueckii, Lactococcus lactis subsp. cremoris, Leuconostoc mesenteroides subsp. mesenteroides and Streptococcus thermophilus. LAB were first inoculated in multiple-strain combinations on the basis of their optimal growth temperatures in three process conditions which differed for milk treatment and medium for strain development: process 1, growth of strains in the optimal synthetic media and pasteurised milk; process 2, growth of strains in whey based medium (WBM) and pasteurised milk; and process 3, growth of strains in WBM and raw milk. The strains that acidified the curds in short time, as shown by a pH drop, were all mesophilic and were then tested in a single inoculum through process 3. Randomly amplified polymorphic DNA (RAPD)-PCR analysis applied to the colonies isolated from the highest dilutions of samples confirmed the dominance of the added strains after curd acidification, stretching and storage. After 15days of refrigerated storage, the decrease in pH values showed an activity of the mesophilic strains at low temperatures, but only Lc. lactis subsp. cremoris PON153, Ln. mesenteroides subsp. mesenteroides PON259 and PON559 increased their number during the 15days at 7°C. A sensory evaluation indicated that the cheeses obtained by applying protocol 3 and by inoculation with lactococci are the most similar to the protected denomination of origin (PDO) cheese and received the best scores by the judges. Thus, the experimental cheeses obtained with raw milk and inoculated with single and multiple combinations of lactococci were subjected to the analysis of the volatile organic compounds (VOCs) carried out by a headspace solid phase microextraction (SPME) technique coupled
Bermúdez-Humarán, Luis G.; Innocentin, Silvia; Lefèvre, Francois; Chatel, Jean-Marc; Langella, Philippe
Today, sufficient data are available to support the use of lactic acid bacteria (LAB), notably lactococci and lactobacilli, as delivery vehicles for the development of new mucosal vaccines. These non-pathogenic Gram-positive bacteria have been safely consumed by humans for centuries in fermented foods. They thus constitute an attractive alternative to the attenuated pathogens (most popular live vectors actually studied) which could recover their pathogenic potential and are thus not totally safe for use in humans. This chapter reviews the current research and advances in the use of LAB as live delivery vectors of proteins of interest for the development of new safe mucosal vaccines. The use of LAB as DNA vaccine vehicles to deliver DNA directly to antigen-presenting cells of the immune system is also discussed.
Mahony, Jennifer; van Sinderen, Douwe
Phages infecting lactic acid bacteria have been the focus of significant research attention over the past three decades. Through the isolation and characterization of hundreds of phage isolates, it has been possible to classify phages of the dairy starter and adjunct bacteria Lactococus lactis, Streptococcus thermophilus, Leuconostoc spp., and Lactobacillus spp. Among these, phages of L. lactis have been most thoroughly scrutinized and serve as an excellent model system to address issues that arise when attempting taxonomic classification of phages infecting other LAB species. Here, we present an overview of the current taxonomy of phages infecting LAB genera of industrial significance, the methods employed in these taxonomic efforts and how these may be employed for the taxonomy of phages of currently underrepresented and emerging phage species. PMID:24478767
Bacteriocins produced by lactic acid bacteria (LAB) are expected to be safe antimicrobial agents. While the best studied LAB bacteriocin, nisin A, is widely utilized as a food preservative, various novel ones are required to control undesirable bacteria more effectively. To discover novel bacteriocins at the early step of the screening process, we developed a rapid screening system that evaluates bacteriocins produced by newly isolated LAB based on their antibacterial spectra and molecular masses. By means of this system, various novel bacteriocins were identified, including a nisin variant, nisin Q, a two-peptide bacteriocin, lactococcin Q, a leaderless bacteriocin, lacticin Q, and a circular bacteriocin, lactocyclicin Q. Moreover, some LAB isolates were found to produce multiple bacteriocins. They were characterized as to their structures, mechanisms of action, and biosynthetic mechanisms. Novel LAB bacteriocins and their biosynthetic mechanisms are expected for applications such as food preservation and peptide engineering.
Desfossés-Foucault, Émilie; LaPointe, Gisèle; Roy, Denis
The starter cultures (Lactococcus sp.) and non-starter lactic acid bacteria (mostly Lactobacillus spp.) are essential to flavor development of Cheddar cheese. The aim of this study was to elucidate the transcriptional interaction between Lactococcus lactis subsp. cremoris SK11 and Lactobacillus paracasei ATCC 334 in mixed cultures during simulated Cheddar cheese manufacture (Pearce activity test) and ripening (slurry). Reverse transcription quantitative PCR (RT-qPCR) was used to quantify the expression of 34 genes common to both bacteria and for eight genes specific to either L. lactis subsp. cremoris SK11 or L. paracasei ATCC 334. The multifactorial analysis (MFA) performed on fold change results for each gene revealed that the genes linked to stress, protein and peptide degradation as well as carbohydrate metabolism of L. paracasei ATCC 334 were especially overexpressed in mixed culture with L. lactis subsp. cremoris SK11 during the ripening simulation. For L. lactis subsp. cremoris SK11, genes coding for amino acid metabolism were more expressed during the cheese manufacture simulation, especially in single culture. These results show how complementary functions of starter and NSLAB contribute to activities useful for flavor development.
Geissler, Andreas J; Behr, Jürgen; von Kamp, Kristina; Vogel, Rudi F
Beer contains only limited amounts of readily fermentable carbohydrates and amino acids. Beer spoilage lactic acid bacteria (LAB) have to come up with metabolic strategies in order to deal with selective nutrient content, high energy demand of hop tolerance mechanisms and a low pH. The metabolism of 26 LAB strains of 6 species and varying spoilage potentialwas investigated in order to define and compare their metabolic capabilities using multivariate statistics and outline possible metabolic strategies. Metabolic capabilities of beer spoilage LAB regarding carbohydrate and amino acids did not correlate with spoilage potential, but with fermentation type (heterofermentative/homofermentative) and species. A shift to mixed acid fermentation by homofermentative (hof) Pediococcus claussenii and Lactobacillus backii was observed as a specific feature of their growth in beer. For heterofermentative (hef) LAB a mostly versatile carbohydrate metabolism could be demonstrated, supplementing the known relevance of organic acids for their growth in beer. For hef LAB a distinct amino acid metabolism, resulting in biogenic amine production, was observed, presumably contributing to energy supply and pH homeostasis.
Kudoyarova, Guzel R; Melentiev, Alexander I; Martynenko, Elena V; Timergalina, Leila N; Arkhipova, Tatiana N; Shendel, Galina V; Kuz'mina, Ludmila Yu; Dodd, Ian C; Veselov, Stanislav Yu
Phytohormone production is one mechanism by which rhizobacteria can stimulate plant growth, but it is not clear whether the bacteria gain from this mechanism. The hypothesis that microbial-derived cytokinin phytohormones stimulate root exudation of amino acids was tested. The rhizosphere of wheat plants was drenched with the synthetic cytokinin trans-zeatin or inoculated with Bacillus subtilis IB-22 (which produces zeatin type cytokinins) or B. subtilis IB-21 (which failed to accumulate cytokinins). Growing plants in a split root system allowed spatial separation of zeatin application or rhizobacterial inoculation to one compartment and analyses of amino acid release from roots (rhizodeposition) into the other compartment (without either microbial inoculation or treatment with exogenous hormone). Supplying B. subtilis IB-22 or zeatin to either the whole root system or half of the roots increased concentrations of amino acids in the soil solution although the magnitude of the increase was greater when whole roots were treated. There was some similarity in amino acid concentrations induced by either bacterial or zeatin treatment. Thus B. subtilis IB-22 increased amino acid rhizodeposition, likely due to its ability to produce cytokinins. Furthermore, B. subtilis strain IB-21, which failed to accumulate cytokinins in culture media, did not significantly affect amino acid concentrations in the wheat rhizosphere. The ability of rhizobacteria to produce cytokinins and thereby stimulate rhizodeposition may be important in enhancing rhizobacterial colonization of the rhizoplane.
Schafner, D.W.; Beuchat, R.L.
The effects of lactic acid bacterial fermentation on chemical and physical changes in aqueous extracts of cowpea (Vigna unguiculata), peanut (Arachis hypogea), soybean (Glycine max), and sorghum (Sorghum vulgare) were studied. The bacteria investigated were Lactobacillus helveticus, L. delbrueckii, L. casei, L. bulgaricus, L. acidophilus, and Streptococcus thermophilus. Organisms were inoculated individually into all of the seed extracts; L. bulgaricus and S. thermophilus were also evaluated together as inocula for fermenting the legume extracts. During fermentation, bacterial population and changes in titratable acidity, pH, viscosity, and color were measured over a 72 h period at 37 degrees C. Maximum bacterial populations, titratable acidity, pH, and viscosity varied depending upon the type of extract and bacterial strain. The maximum population of each organism was influenced by fermentable carbohydrates, which, in turn, influenced acid production and change in pH. Change in viscosity was correlated with the amount of protein and titratable acidity of products. Color was affected by pasteurization treatment and fermentation as well as the source of extract. In the extracts inoculated simultaneously with L. bulgaricus and S. thermophilus, a synergistic effect resulted in increased bacterial populations, titratable acidity, and viscosity, and decreased pH in all the legume extracts when compared to the extracts fermented with either of these organisms individually. Fermented extracts offer potential as substitutes for cultured dairy products. 24 references.
Genotypic identification of some lactic acid bacteria by amplified fragment length polymorphism analysis and investigation of their potential usage as starter culture combinations in Beyaz cheese manufacture.
Karahan, A G; Başyiğit Kiliç, G; Kart, A; Sanlidere Aloğlu, H; Oner, Z; Aydemir, S; Erkuş, O; Harsa, S
In this study, 2 different starter culture combinations were prepared for cheesemaking. Starter culture combinations were formed from 8 strains of lactic acid bacteria. They were identified as Lactococcus lactis ssp. lactis (2 strains), Lactobacillus plantarum (5 strains), and Lactobacillus paraplantarum (1 strain) by amplified fragment length polymorphism analysis. The effects of these combinations on the physicochemical and microbiological properties of Beyaz cheeses were investigated. These cheeses were compared with Beyaz cheeses that were produced with a commercial starter culture containing Lc. lactis ssp. lactis and Lc. lactis ssp. cremoris as control. All cheeses were ripened in brine at 4 degrees C for 90 d. Dry matter, fat in dry matter, titratable acidity, pH, salt in dry matter, total N, water-soluble N, and ripening index were determined. Sodium dodecyl sulfate-PAGE patterns of cheeses showed that alpha(S)-casein and beta-casein degraded slightly during the ripening period. Lactic acid bacteria, total mesophilic aerobic bacteria, yeast, molds, and coliforms were also counted. All analyses were repeated twice during d 7, 30, 60, and 90. The starter culture combinations were found to be significantly different from the control group in pH, salt content, and lactobacilli, lactococci, and total mesophilic aerobic bacteria counts, whereas the cheeses were similar in fat, dry matter content, and coliform, yeast, and mold counts. The sensory analysis of cheeses indicated that textural properties of control cheeses presented somewhat lower scores than those of the test groups. The panelists preferred the tastes of treatment cheeses, whereas cheeses with starter culture combinations and control cheeses had similar scores for appearance and flavor. These results indicated that both starter culture combinations are suitable for Beyaz cheese production.
Lactic acid bacteria (LAB) in fish flesh has long been disregarded because the high post-mortem pH, the low percentage of sugars, the high content of low molecular weight nitrogenous molecules and the low temperature of temperate waters favor the rapid growth of pH-sensitive psychrotolerant marine Gram-negative bacteria like Pseudomonas, Shewanella and Photobacterium. In seafood packed in both vacuum (VP) and modified atmosphere (MAP) packaging commonly CO(2) enriched, the growth of the Gram-negative aerobic bacteria group (predominantly pseudomonads) is effectively inhibited and the number reached by LAB during storage is higher than that achieved in air but always several log units lower than the trimethylamine oxide (TMA-O) reducing and CO(2)-resistant organisms (Shewanella putrefaciens and Photobacterium phosphoreum). Accordingly, LAB are not of much concern in seafood neither aerobically stored nor VP and MAP. However, they may acquire great relevance in lightly preserved fish products (LPFP), including those VP or MAP. Fresh fish presents a very high water activity (aw) value (0.99). However, aw is reduced to about 0.96 when salt (typically 6% WP) is added to the product. As a result, aerobic Gram-negative bacteria are inhibited, which allows the growth of other organisms more resistant to reduced aw, i.e. LAB, and then they may acquire a central role in the microbial events occurring in the product. Changes in consumers' habits have led to an increase of convenient LPFP with a relative long shelf-life (at least 3 weeks) which, on the other hand, may constitute a serious problem from a safety perspective since Listeria monocytogenes and sometimes Clostridium botulinum (mainly type E) may able to grow. In any case the LAB function in marine products is complex, depending on species, strains, interaction with other bacteria and the food matrix. They may have no particular effect or they may be responsible for spoilage and, in certain cases, they may even exert
Background Genome scale annotation of regulatory interactions and reconstruction of regulatory networks are the crucial problems in bacterial genomics. The Lactobacillales order of bacteria collates various microorganisms having a large economic impact, including both human and animal pathogens and strains used in the food industry. Nonetheless, no systematic genome-wide analysis of transcriptional regulation has been previously made for this taxonomic group. Results A comparative genomics approach was used for reconstruction of transcriptional regulatory networks in 30 selected genomes of lactic acid bacteria. The inferred networks comprise regulons for 102 orthologous transcription factors (TFs), including 47 novel regulons for previously uncharacterized TFs. Numerous differences between regulatory networks of the Streptococcaceae and Lactobacillaceae groups were described on several levels. The two groups are characterized by substantially different sets of TFs encoded in their genomes. Content of the inferred regulons and structure of their cognate TF binding motifs differ for many orthologous TFs between the two groups. Multiple cases of non-orthologous displacements of TFs that control specific metabolic pathways were reported. Conclusions The reconstructed regulatory networks substantially expand the existing knowledge of transcriptional regulation in lactic acid bacteria. In each of 30 studied genomes the obtained regulatory network contains on average 36 TFs and 250 target genes that are mostly involved in carbohydrate metabolism, stress response, metal homeostasis and amino acids biosynthesis. The inferred networks can be used for genetic experiments, functional annotations of genes, metabolic reconstruction and evolutionary analysis. All reconstructed regulons are captured within the Streptococcaceae and Lactobacillaceae collections in the RegPrecise database (http://regprecise.lbl.gov). PMID:23398941
Wang, Bin; Shao, Yanchun; Chen, Fusheng
Acetic acid bacteria (AAB) are a group of gram-negative or gram-variable bacteria which possess an obligate aerobic property with oxygen as the terminal electron acceptor, meanwhile transform ethanol and sugar to corresponding aldehydes, ketones and organic acids. Since the first genus Acetobacter of AAB was established in 1898, 16 AAB genera have been recorded so far. As the main producer of a world-wide condiment, vinegar, AAB have evolved an elegant adaptive system that enables them to survive and produce a high concentration of acetic acid. Some researches and reviews focused on mechanisms of acid resistance in enteric bacteria and made the mechanisms thoroughly understood, while a few investigations did in AAB. As the related technologies with proteome, transcriptome and genome were rapidly developed and applied to AAB research, some plausible mechanisms conferring acetic acid resistance in some AAB strains have been published. In this review, the related mechanisms of AAB against acetic acid with acetic acid assimilation, transportation systems, cell morphology and membrane compositions, adaptation response, and fermentation conditions will be described. Finally, a framework for future research for anti-acid AAB will be provided.
Fritsch, Caroline; Heinrich, Veronika; Vogel, Rudi F; Toelstede, Simone
Sunflower flour provides a high content of protein with a well-balanced amino acid composition and is therefore regarded as an attractive source for protein. The use for human nutrition is hindered by phenolic compounds, mainly chlorogenic acid, which can lead under specific circumstances to undesirable discolorations. In this study, growth behavior and degradation ability of chlorogenic acid of four lactic acid bacteria were explored. Data suggested that significant higher fermentation performances on sunflower flour as compared to sunflower protein concentrate were reached by Lactobacillus plantarum, Pediococcus pentosaceus, Lactobacillus gasseri and Bifidobacterium animalis subsp. lactis. In fermentation with the latter two strains reduced amounts of chlorogenic acid were observed in sunflower flour (-11.4% and -19.8%, respectively), which were more pronounced in the protein concentrate (-50.7% and -95.6%, respectively). High tolerances against chlorogenic acid and the cleavage product quinic acid with a minimum inhibitory concentration (MIC) of ≥20.48 mg/ml after 48 h were recorded for all strains except Bifidobacterium animalis subsp. lactis, which was more sensitive. The second cleavage compound, caffeic acid revealed a higher antimicrobial potential with MIC values of 0.64-5.12 mg/ml. In this proof of concept study, degradation versus inhibitory effect suggest the existence of basic mechanisms of interaction between phenolic acids in sunflower and lactic acid bacteria and a feasible way to reduce the chlorogenic acid content, which may help to avoid undesired color changes.
Nowak, Adriana; Kuberski, Sławomir; Libudzisz, Zdzisława
Humans can be exposed to N-nitroso compounds (NOCs) due to many environmental sources, as well as endogenous formation. The main nitrosamine found in food products and also synthesised in vivo by intestinal microbiota is N-nitrosodimethylamine (NDMA). It can cause cancer of the stomach, kidney and colon. The effect of four probiotic Lactobacillus strains on NDMA was studied under different culture conditions (24 h in MRS, 168 h in modified MRS N, and 168 h in phosphate buffer). HPLC and GC-TEA methods were used for NDMA determination in supernatants. The influence of lactic acid bacteria on NDMA genotoxicity was investigated by means of the comet assay. Additionally, the effect of NDMA (2-100 µg ml⁻¹) on the growth and survival of the probiotic strains was studied. The results indicate that the bacteria decreased NDMA concentration by up to 50%, depending on the culture conditions, time of incubation, NDMA concentration, pH and bacterial strain. Lb. brevis 0945 lowered the concentration and genotoxicity of NDMA most effectively by up to 50%. This could be due to either adsorption or metabolism. The growth and survival of the bacteria was not affected by any of the tested NDMA concentrations.
Singh, Neetigyata Pratap; Tiwari, Abhay; Bansal, Ankiti; Thakur, Shruti; Sharma, Garima; Gabrani, Reema
Bacteriocins are antimicrobial peptides which are ribosomally synthesized by mainly all bacterial species. LABs (lactic acid bacteria) are a diverse group of bacteria that include around 20 genera of various species. Though LABs have a tremendous potential for production of anti-microbial peptides, this group of bacteria is still underexplored for bacteriocins. To study the diversity among bacteriocin encoding clusters and the putative bacteriocin precursors, genome mining was performed on 20 different species of LAB not reported to be bacteriocin producers. The phylogenetic tree of gyrB, rpoB, and 16S rRNA were constructed using MEGA6 software to analyze the diversity among strains. Putative bacteriocins operons identified were found to be diverse and were further characterized on the basis of physiochemical properties and the secondary structure. The presence of at least two cysteine residues in most of the observed putative bacteriocins leads to disulphide bond formation and provide stability. Our data suggests that LABs are prolific source of low molecular weight non modified peptides.
Rudrappa, Thimmaraju; Czymmek, Kirk J; Paré, Paul W; Bais, Harsh P
Beneficial soil bacteria confer immunity against a wide range of foliar diseases by activating plant defenses, thereby reducing a plant's susceptibility to pathogen attack. Although bacterial signals have been identified that activate these plant defenses, plant metabolites that elicit rhizobacterial responses have not been demonstrated. Here, we provide biochemical evidence that the tricarboxylic acid cycle intermediate L-malic acid (MA) secreted from roots of Arabidopsis (Arabidopsis thaliana) selectively signals and recruits the beneficial rhizobacterium Bacillus subtilis FB17 in a dose-dependent manner. Root secretions of L-MA are induced by the foliar pathogen Pseudomonas syringae pv tomato (Pst DC3000) and elevated levels of L-MA promote binding and biofilm formation of FB17 on Arabidopsis roots. The demonstration that roots selectively secrete L-MA and effectively signal beneficial rhizobacteria establishes a regulatory role of root metabolites in recruitment of beneficial microbes, as well as underscores the breadth and sophistication of plant-microbial interactions.
Robert, Hervé; Gabriel, Valérie; Fontagné-Faucher, Catherine
The lactic acid microflora of nine traditional wheat sourdoughs from the Midi-Pyrénées area (South western France) was previously isolated and preliminary characterized using conventional morphological and biochemical analysis. However, such phenotypic methods alone are not always reliable and have a low taxonomic resolution for identification of lactic acid bacteria species. In the present study, a total of 290 LAB isolates were identified by PCR amplification using different sets of specific primers in order to provide a thorough characterization of the lactic flora from these traditional French sourdoughs. Overall, the LAB isolates belonged to 6 genera: Lactobacillus (39%, 8 species), Pediococcus (38%, 1 species), Leuconostoc (17%, 2 species), Weissella (4%, 2 species), Lactococcus (1%, 1 species) and Enterococcus (<1%, 1 species) and 15 different species were detected: L. plantarum, L. curvatus, L. paracasei, L. sanfranciscensis, L. pentosus, L. paraplantarum, L. sakei, L. brevis, P. pentosaceus, L. mesenteroides, L. citreum, W. cibaria, W. confusa, L. lactis and E. hirae. Facultative heterofermentative LAB represent more than 76% of the total isolates, the main species isolated herein correspond to L. plantarum and P. pentosaceus. Obligate heterofermentative lactobacilli (L. sanfranciscencis, L. brevis) represent less than 3% of the total isolates whereas Leuconostoc and Weissella species represent 21% of the total isolates and have been detected in eight of the nine samples. Detection of some LAB species was preferentially observed depending on the isolation culture medium. The number of different species within a sourdough varies from 3 to 7 and original associations of hetero- and homofermentative LAB species have been revealed. Results from this study clearly confirm the diversity encountered in the microbial community of traditional sourdough and highlight the importance of LAB cocci in the sourdough ecosystem, along with lactobacilli.
van Beek, Sylvie; Priest, Fergus G.
Seven strains of Lactobacillus isolated from malt whisky fermentations and representing Lactobacillus brevis, L. crispatus, L. fermentum, L. hilgardii, L. paracasei, L. pentosus, and L. plantarum contained genes for hydroxycinnamic acid (p-coumaric acid) decarboxylase. With the exception of L. hilgardii, these bacteria decarboxylated p-coumaric acid and/or ferulic acid, with the production of 4-vinylphenol and/or 4-vinylguaiacol, respectively, although the relative activities on the two substrates varied between strains. The addition of p-coumaric acid or ferulic acid to cultures of L. pentosus in MRS broth induced hydroxycinnamic acid decarboxylase mRNA within 5 min, and the gene was also induced by the indigenous components of malt wort. In a simulated distillery fermentation, a mixed culture of L. crispatus and L. pentosus in the presence of Saccharomyces cerevisiae decarboxylated added p-coumaric acid more rapidly than the yeast alone but had little activity on added ferulic acid. Moreover, we were able to demonstrate the induction of hydroxycinnamic acid decarboxylase mRNA under these conditions. However, in fermentations with no additional hydroxycinnamic acid, the bacteria lowered the final concentration of 4-vinylphenol in the fermented wort compared to the level seen in a pure-yeast fermentation. It seems likely that the combined activities of bacteria and yeast decarboxylate p-coumaric acid and then reduce 4-vinylphenol to 4-ethylphenol more effectively than either microorganism alone in pure cultures. Although we have shown that lactobacilli participate in the metabolism of phenolic compounds during malt whisky fermentations, the net result is a reduction in the concentrations of 4-vinylphenol and 4-vinylguaiacol prior to distillation. PMID:11097909
Wu, Jing-jing; Du, Rui-ping; Gao, Min; Sui, Yao-qiang; Xiu, Lei; Wang, Xiao
Silage making has become a significant method of forage conservation worldwide. To determine how tomato pomace (TP) may be used effectively as animal feed, it was ensilaged for 90 days and microbiology counts, fermentation characteristics and chemical composition of tomato pomace silage (TPS) were evaluated at the 30th, 60th, and 90th days, respectively. In addition, 103 lactic acid bacteria were isolated from TPS. Based on the phenotypic and chemotaxonomic characteristics, 16S rDNA sequence and carbohydrate fermentation tests, the isolates were identified as 17 species namely: Lactobacillus coryniformis subsp. torquens (0.97%), Lactobacillus pontis (0.97%), Lactobacillus hilgardii (0.97%), Lactobacillus pantheris (0.97%), Lactobacillus amylovorus (1.9%), Lactobacillus panis (1.9%), Lactobacillus vaginalis (1.9%), Lactobacillus rapi (1.9%), Lactobacillus buchneri (2.9%), Lactobacillus parafarraginis (2.9%), Lactobacillus helveticus (3.9%), Lactobacillus camelliae (3.9%), Lactobacillus fermentum (5.8%), Lactobacillus manihotivorans (6.8%), Lactobacillus plantarum (10.7%), Lactobacillus harbinensis (16.5%) and Lactobacillus paracasei subsp. paracasei (35.0%). This study has shown that TP can be well preserved for 90 days by ensilaging and that TPS is not only rich in essential nutrients, but that physiological and biochemical properties of the isolates could provide a platform for future design of lactic acid bacteria (LAB) inoculants aimed at improving the fermentation quality of silage. PMID:25049999
Macwana, Sunita; Muriana, Peter M
A practical system was devised for grouping bacteriocins of lactic acid bacteria (LAB) based on mode of action as determined by changes in inhibitory activity to spontaneously-acquired bacteriocin resistance (Bac(R)). Wild type Listeria monocytogenes 39-2 was sensitive to five bacteriocins produced by 3 genera of LAB: pediocin PA-1 and pediocin Bac3 (Pediococcus), lacticin FS97 and lacticin FS56 (Lactococcus), and curvaticin FS47 (Lactobacillus). A spontaneous Bac(R) derivative of L. monocytogenes 39-2 obtained by selective recovery against lacticin FS56 provided complete resistance to the bacteriocin made by Lactococcus lactis FS56. The lacticin FS56-resistant strain of L. monocyotgenes 39-2 was also cross-resistant to curvaticin FS47 and pediocin PA-1, but not to lacticin FS97 or pediocin Bac3. The same pattern of cross-resistance was also observed with Bac(R) isolates obtained with L. monocytogenes Scott A-2. A spontaneous mutation that renders a strain cross-resistant to different bacteriocins indicates that they share a common mechanism of resistance due to similar modes of action of the bacteriocins. Spontaneous resistance was acquired to other bacteriocins (in aggregate) by following the same procedure against which the Bac(R) strain was still sensitive. In subsequent challenge assays, mixtures of bacteriocins of different modes of action provided greater inhibition than mixtures of bacteriocins of the same mode of action (as determined by our screening method). This study identifies a methodical approach to classify bacteriocins into functional groups based on mechanism of resistance (i.e., mode of action) that could be used for identifying the best mixture of bacteriocins for use as biopreservatives.
Li, Yang; He, Dongwei; Niu, Dongjie; Zhao, Youcai
In this study, yeast and acetic acid bacteria strains were adopted to enhance the ethanol-type fermentation resulting to a volatile fatty acids yield of 30.22 g/L, and improve acetic acid production to 25.88 g/L, with food wastes as substrate. In contrast, only 12.81 g/L acetic acid can be obtained in the absence of strains. The parameters such as pH, oxidation reduction potential and volatile fatty acids were tested and the microbial diversity of different strains and activity of hydrolytic ferment were investigated to reveal the mechanism. The optimum pH and oxidation reduction potential for the acetic acid production were determined to be at 3.0-3.5 and -500 mV, respectively. Yeast can convert organic matters into ethanol, which is used by acetic acid bacteria to convert the organic wastes into acetic acid. The acetic acid thus obtained from food wastes micro-aerobic fermentation liquid could be extracted by distillation to get high-pure acetic acid.
Furtado, Danielle N; Todorov, Svetoslav D; Landgraf, Mariza; Destro, Maria T; Franco, Bernadette D G M
Lactic acid bacteria capable of producing bacteriocins and presenting probiotic potential open innovative technological applications in the dairy industry. In this study, a bacteriocinogenic strain (Lactococcus lactis subsp. lactis DF4Mi) was isolated from goat milk, and studied for its antimicrobial activity. The bacteriocin presented a broad spectrum of activity, was sensitive to proteolytic enzymes, resistant to heat and pH extremes, and not affected by the presence of SDS, Tween 20, Tween 80, EDTA or NaCl. Bacteriocin production was dependent on the components of the culture media, especially nitrogen source and salts. When tested by PCR, the bacteriocin gene presented 100% homology to nisin Z gene. These properties indicate that this L. lactis subsp. lactis DF4Mi can be used for enhancement of dairy foods safety and quality.
Furtado, Danielle N.; Todorov, Svetoslav D.; Landgraf, Mariza; Destro, Maria T.; Franco, Bernadette D.G.M.
Lactic acid bacteria capable of producing bacteriocins and presenting probiotic potential open innovative technological applications in the dairy industry. In this study, a bacteriocinogenic strain (Lactococcus lactis subsp. lactis DF4Mi) was isolated from goat milk, and studied for its antimicrobial activity. The bacteriocin presented a broad spectrum of activity, was sensitive to proteolytic enzymes, resistant to heat and pH extremes, and not affected by the presence of SDS, Tween 20, Tween 80, EDTA or NaCl. Bacteriocin production was dependent on the components of the culture media, especially nitrogen source and salts. When tested by PCR, the bacteriocin gene presented 100% homology to nisin Z gene. These properties indicate that this L. lactis subsp. lactis DF4Mi can be used for enhancement of dairy foods safety and quality. PMID:25763065
Larsen, Nadja; Werner, Birgit Brøsted; Vogensen, Finn Kvist; Jespersen, Lene
Milk acidification by DL-starter cultures [cultures containing Lactococcus lactis diacetylactis (D) and Leuconostoc (L) species] depends on the oxidation-reduction (redox) potential in milk; however, the mechanisms behind this effect are not completely clear. The objective of this study was to investigate the effect of dissolved oxygen on acidification kinetics and redox potential during milk fermentation by lactic acid bacteria (LAB). Fermentations were conducted by single strains isolated from mixed DL-starter culture, including Lactococcus lactis ssp. lactis, Lactococcus lactis ssp. cremoris, and Leuconostoc mesenteroides ssp. cremoris, by the DL-starter culture, and by the type strains. High and low levels of oxygen were produced by flushing milk with oxygen or nitrogen, respectively. The kinetics of milk acidification was characterized by the maximum rate and time of acidification (Vamax and Tamax), the maximum rate and time of reduction (Vrmax and Trmax), the minimum redox potential (Eh7 final), and time of reaching Eh7 final (Trfinal). Variations in kinetic parameters were observed at both the species and strain levels. Two of the Lc. lactis ssp. lactis strains were not able to lower redox potential to negative values. Kinetic parameters of the DL-starter culture were comparable with the best acidifying and reducing strains, indicating their additive effects. Acidification curves were mostly diauxic at all oxygen levels, displaying 2 maxima of acidification rate: before (aerobic maximum) and after (anaerobic maximum) oxygen depletion. The redox potential decreased concurrently with oxygen consumption and continued to decrease at slower rate until reaching the final values, indicating involvement of both oxygen and microbiological activity in the redox state of milk. Oxygen flushing had a negative effect on reduction and acidification capacity of tested LAB. Reduction was significantly delayed at high initial oxygen, exhibiting longer Trmax, Trfinal, or both
Khota, Waroon; Pholsen, Suradej; Higgs, David; Cai, Yimin
Natural lactic acid bacteria (LAB) populations in tropical grasses and their fermentation characteristics on silage prepared with cellulase enzyme and LAB inoculants were studied. A commercial inoculant Lactobacillus plantarum Chikuso 1 (CH), a local selected strain Lactobacillus casei TH14 (TH14), and 2 cellulases, Acremonium cellulase (AC) and Maicelase (MC; Meiji Seika Pharma Co. Ltd., Tokyo, Japan), were used as additives to silage preparation with fresh and wilted (6 h) Guinea grass and Napier grass. Silage was prepared using a laboratory-scale fermentation system. Treatments were CH, TH14, AC at 0.01% fresh matter, AC 0.1%, MC 0.01%, MC 0.1%, CH+AC 0.01%, CH+AC 0.1%, CH+MC 0.01%, CH+MC 0.1%, TH14+AC 0.1%, TH14+AC 0.01%, TH14+MC 0.1%, and TH14+MC 0.01%. Microorganism counts of Guinea grass and Napier grass before ensiling were 10(2) LAB and 10(6) aerobic bacteria; these increased during wilting. Based on morphological and biochemical characteristics, and 16S rRNA gene sequence analysis, natural strains from both grasses were identified as L. plantarum, L. casei, Lactobacillus acidipiscis, Leuconostoc pseudomesenteroides, Leuconostoc garlicum, Weissella confusa, and Lactococcus lactis. Lactobacillus plantarum and L. casei are the dominant species and could grow at lower pH and produce more lactic acid than the other isolates. Crude protein and neutral detergent fiber were 5.8 and 83.7% of dry matter (DM) for Guinea grass, and 7.5 and 77.1% of DM for Napier grass. Guinea grass had a low level of water-soluble carbohydrates (0.39% of DM). Guinea grass silage treated with cellulase had a lower pH and higher lactic acid content than control and LAB treatments. The 0.1% AC and MC treatments had the best result for fermentation quality. All high water-soluble carbohydrate (2.38% DM) Napier grass silages showed good fermentation quality. Compared with control and LAB-inoculated silage, the cellulase-treated silages had significantly higher crude protein content and
Sgarbi, E; Lazzi, C; Tabanelli, G; Gatti, M; Neviani, E; Gardini, F
In long-ripened cheese, flavor formation occurs during ripening. The metabolism of lactic acid bacteria (LAB) leads to the production of different compounds that contribute to the flavor of cheese. The contribution of LAB to the formation of cheese flavor has previously been studied. However, the specific nonstarter LAB (NSLAB) metabolic reactions in ripened cheese that lead to the formation of flavor compounds remain unclear. In ripened cheese, the nutrient sources available include small peptides or amino acids, citrate, lactate, free fatty acids, and starter LAB cell lysis products. Thus, the aim of this study was to evaluate the ability of NSLAB to produce volatile flavor compounds by using an in vitro system that used only the nutrients available in ripened cheese as the energy source. Moreover, the potential contribution of the NSLAB volatilome on total cheese flavor is discussed. For this purpose, the production of volatile compounds on cheese-based medium (CBM) and on starter LAB lysed cell medium (LCM) by 2 Lactobacillus casei and 2 Lactobacillus rhamnosus strains, previously isolated from ripened Parmigiano Reggiano cheese, was investigated. The generated volatile compounds were analyzed with head-space gas chromatography mass spectrometry. Overall, ketones, aldehydes, alcohols, and acids were the most abundant compounds produced. Differences in volatilome production were found between NSLAB grown in LCM and CBM. The catabolic metabolism of amino acids and fatty acids were required for NSLAB growth on LCM. Conversely, pyruvate metabolism was the main catabolic pathway that supported growth of NSLAB in CBM. This study can be considered a first step toward a better understanding of how microbiota involved in the long ripening of cheese may contribute to the development of cheese flavor.
Pedraza, Raúl O
Nitrogen is an essential plant nutrient, widely applied as N-fertilizer to improve yield of agriculturally important crops. An interesting alternative to avoid or reduce the use of N-fertilizers could be the exploitation of plant growth-promoting bacteria (PGPB), capable of enhancing growth and yield of many plant species, several of agronomic and ecological significance. PGPB belong to diverse genera, including Azospirillum, Azotobacter, Herbaspirillum, Bacillus, Burkholderia, Pseudomonas, Rhizobium, and Gluconacetobacter, among others. They are capable of promoting plant growth through different mechanisms including (in some cases), the biological nitrogen fixation (BNF), the enzymatic reduction of the atmospheric dinitrogen (N(2)) to ammonia, catalyzed by nitrogenase. Aerobic bacteria able to oxidize ethanol to acetic acid in neutral or acid media are candidates of belonging to the family Acetobacteraceae. At present, this family has been divided into ten genera: Acetobacter, Gluconacetobacter, Gluconobacter, Acidomonas, Asaia, Kozakia, Saccharibacter, Swaminathania, Neoasaia, and Granulibacter. Among them, only three genera include N(2)-fixing species: Gluconacetobacter, Swaminathania and Acetobacter. The first N(2)-fixing acetic acid bacterium (AAB) was described in Brazil. It was found inside tissues of the sugarcane plant, and first named as Acetobacter diazotrophicus, but then renamed as Gluconacetobacter diazotrophicus. Later, two new species within the genus Gluconacetobacter, associated to coffee plants, were described in Mexico: G. johannae and G. azotocaptans. A salt-tolerant bacterium named Swaminathania salitolerans was found associated to wild rice plants. Recently, N(2)-fixing Acetobacter peroxydans and Acetobacter nitrogenifigens, associated with rice plants and Kombucha tea, respectively, were described in India. In this paper, recent advances involving nitrogen-fixing AAB are presented. Their natural habitats, physiological and genetic aspects
Pérez-Díaz, I M; Hayes, J; Medina, E; Anekella, K; Daughtry, K; Dieck, S; Levi, M; Price, R; Butz, N; Lu, Z; Azcarate-Peril, M A
A compositional re-assessment of the microbiota present in commercial cucumber fermentation using culture independent and dependent methods was conducted, with emphasis on lactic acid bacteria (LAB). Two commercial cucumber fermentation tanks were monitored by measuring pH, dissolved oxygen and temperature, and used as sources of samples for microbial plating, genomic DNA extraction and measurement of organic acids and carbohydrates by HPLC. Six additional commercial tanks were included to identify the dominant microorganisms using molecular methods. A comparative analysis of the publically available genome sequences corresponding to the LAB found in cucumber fermentations was completed to gain an understanding of genomic features possibly enabling dominance. Analyses of the microbiota suggest Lactobacillales prevail in cucumber fermentations, including in order of prevalence Lactobacillus pentosus, Lb. plantarum, Lb. brevis, Weissella spp., Pediococcus ethanolidurans, Leuconostoc spp. and Lactococcus spp. It was observed that Lb. pentosus and Lb. plantarum have comparatively larger genomes, higher gene counts, uniquely distribute the ribosomal clusters across the genome as opposed to close to the origin of replication, and possess more predicted amino acids prototrophies and selected biosynthesis related genes. It is theorized that Lb. pentosus and Lb. plantarum dominance in cucumber fermentations is the result of their genetic make-up.
Saavedra, Lucila; Sesma, Fernando
The search for new antimicrobial peptides produced by lactic acid bacteria and other Gram-positive microorganisms has become an interesting field of research in the past decades. The fact that bacteriocins are active against numerous foodborne and human pathogens, are produced by generally regarded as safe (GRAS) microorganisms, and are readily degraded by proteolytic host systems makes them attractive candidates for biotechnological applications. However, before suggesting or choosing a new bacteriocin for future technology developments, it is necessary to elucidate its biochemical structure and its mode of action, which may be carried out once the bacteriocin is purified to homogeneity. This chapter focuses on describing the main strategies used for the purification of numerous bacteriocins.
Ozogul, Fatih; Tabanelli, Giulia; Toy, Nurten; Gardini, Fausto
Conversion of ornithine to putrescine by Salmonella Paratyphi A, Listeria monocytogenes, Staphylococcus aureus, and Escherichia coli was investigated in ornithine decarboxylase broth (ODB) using cell-free supernatants (CFSs) obtained from Leuconostoc mesenterodies subsp. cremoris, Pediococcus acidilactici, Lactococcus lactis subsp. lactis, Streptococcus thermophilus. Two groups of cell-free supernatants (25 or 50%) and control (only ODB) were prepared to investigate putrescine (PUT) and other polyamine formation by foodborne pathogens (FBPs). Significant differences (p < 0.05) were observed among the species for each amine. All of the CFSs reduced the formation of PUT by ≥65%. The production of cadaverine (CAD) was scarcely affected by the presence of CFSs, with the exception of the samples inoculated with L. monocytogenes. The variation in polyamine was found with respect to the control samples. Spermidine (SPD) was produced in lower amount in many samples, especially in Gram-negative FBPs, whereas spermine (SPN) increased drastically in the major part of the samples concerning the control. Histamine (HIS) was characterized by a marked concentration decrease in all of the samples, and tyramine (TYR) was accumulated in very low concentrations in the controls. Therefore, the ability of bacteria to produce certain biogenic amines such as HIS, TYR, PUT, and CAD has been studied to assess their risk and prevent their formation in food products. The results obtained from this study concluded that the lactic acid bacteria (LAB) strains with non-decarboxylase activity are capable of avoiding or limiting biogenic amine formation by FBP.
Zhou, N; Zhang, J X; Fan, M T; Wang, J; Guo, G; Wei, X Y
The aim of this study was to evaluate the susceptibility of 43 strains of lactic acid bacteria, isolated from Chinese yogurts made in different geographical areas, to 11 antibiotics (ampicillin, penicillin G, roxithromycin, chloramphenicol, tetracycline, chlortetracycline, lincomycin, kanamycin, streptomycin, neomycin, and gentamycin). The 43 isolates (18 Lactobacillus bulgaricus and 25 Streptococcus thermophilus) were identified at species level and were typed by random amplified polymorphic DNA analysis. Thirty-five genotypically different strains were detected and their antimicrobial resistance to 11 antibiotics was determined using the agar dilution method. Widespread resistance to ampicillin, chloramphenicol, chlortetracycline, tetracyclines, lincomycin, streptomycin, neomycin, and gentamycin was found among the 35 strains tested. All of the Strep. thermophilus strains tested were susceptible to penicillin G and roxithromycin, whereas 23.5 and 64.7% of Lb. bulgaricus strains, respectively, were resistant. All of the Strep. thermophilus and Lb. bulgaricus strains were found to be resistant to kanamycin. The presence of the corresponding resistance genes in the resistant isolates was investigated through PCR, with the following genes detected: tet(M) in 1 Lb. bulgaricus and 2 Strep. thermophilus isolates, ant(6) in 2 Lb. bulgaricus and 2 Strep. thermophilus isolates, and aph(3')-IIIa in 5 Lb. bulgaricus and 2 Strep. thermophilus isolates. The main threat associated with these bacteria is that they may transfer resistance genes to pathogenic bacteria, which has been a major cause of concern to human and animal health. To our knowledge, the aph(3')-IIIa and ant(6) genes were found in Lb. bulgaricus and Strep. thermophilus for the first time. Further investigations are required to analyze whether the genes identified in Lb. bulgaricus and Strep. thermophilus isolates might be horizontally transferred to other species.
Rokop, Z P; Horton, M A; Newton, I L G
In contrast to the honey bee gut, which is colonized by a few characteristic bacterial clades, the hive of the honey bee is home to a diverse array of microbes, including many lactic acid bacteria (LAB). In this study, we used culture, combined with sequencing, to sample the LAB communities found across hive environments. Specifically, we sought to use network analysis to identify microbial hubs sharing nearly identical operational taxonomic units, evidence which may indicate cooccurrence of bacteria between environments. In the process, we identified interactions between noncore bacterial members (Fructobacillus and Lactobacillaceae) and honey bee-specific "core" members. Both Fructobacillus and Lactobacillaceae colonize brood cells, bee bread, and nectar and may serve the role of pioneering species, establishing an environment conducive to the inoculation by honey bee core bacteria. Coculture assays showed that these noncore bacterial members promote the growth of honey bee-specific bacterial species. Specifically, Fructobacillus by-products in spent medium supported the growth of the Firm-5 honey bee-specific clade in vitro. Metabolic characterization of Fructobacillus using carbohydrate utilization assays revealed that this strain is capable of utilizing the simple sugars fructose and glucose, as well as the complex plant carbohydrate lignin. We tested Fructobacillus for antibiotic sensitivity and found that this bacterium, which may be important for establishment of the microbiome, is sensitive to the commonly used antibiotic tetracycline. Our results point to the possible significance of "noncore" and environmental microbial community members in the modulation of honey bee microbiome dynamics and suggest that tetracycline use by beekeepers should be limited.
Nishiyama, Keita; Sugiyama, Makoto; Mukai, Takao
Lactic acid bacteria (LAB) are Gram-positive bacteria that are natural inhabitants of the gastrointestinal (GI) tracts of mammals, including humans. Since Mechnikov first proposed that yogurt could prevent intestinal putrefaction and aging, the beneficial effects of LAB have been widely demonstrated. The region between the duodenum and the terminal of the ileum is the primary region colonized by LAB, particularly the Lactobacillus species, and this region is covered by a mucus layer composed mainly of mucin-type glycoproteins. The mucus layer plays a role in protecting the intestinal epithelial cells against damage, but is also considered to be critical for the adhesion of Lactobacillus in the GI tract. Consequently, the adhesion exhibited by lactobacilli on mucin has attracted attention as one of the critical factors contributing to the persistent beneficial effects of Lactobacillus in a constantly changing intestinal environment. Thus, understanding the interactions between Lactobacillus and mucin is crucial for elucidating the survival strategies of LAB in the GI tract. This review highlights the properties of the interactions between Lactobacillus and mucin, while concomitantly considering the structure of the GI tract from a histochemical perspective. PMID:27681930
Monedero, Vicente; Pérez-Martínez, Gaspar; Yebra, María J
Polyols are sugar alcohols largely used as sweeteners and they are claimed to have several health-promoting effects (low-caloric, low-glycemic, low-insulinemic, anticariogenic, and prebiotic). While at present chemical synthesis is the only strategy able to assure the polyol market demand, the biotechnological production of polyols has been implemented in yeasts, fungi, and bacteria. Lactic acid bacteria (LAB) are a group of microorganisms particularly suited for polyol production as they display a fermentative metabolism associated with an important redox modulation and a limited biosynthetic capacity. In addition, LAB participate in food fermentation processes, where in situ production of polyols during fermentation may be useful in the development of novel functional foods. Here, we review the polyol production by LAB, focusing on metabolic engineering strategies aimed to redirect sugar fermentation pathways towards the synthesis of biotechnologically important sugar alcohols such as sorbitol, mannitol, and xylitol. Furthermore, possible approaches are presented for engineering new fermentation routes in LAB for production of arabitol, ribitol, and erythritol.
Demakov, V A; Vasil'ev, D M; Maksimova, Yu G; Pavlova, Yu A; Ovechkina, G V; Maksimov, A Yu
Species diversity of bacteria from the activated sludge of Perm biological waste treatment facilities capable of transformation of cyanopyridines and amides of pyridinecarboxylic acids was investigated. Enrichment cultures in mineral media with 3-cyanopyridine as the sole carbon and nitrogen source were used to obtain 32 clones of gram-negative heterotrophic bacteria exhibiting moderate growth on solid and liquid media with 3- and 4-cyanopyridine. Sequencing of the 16S rRNA gene fragments revealed that the clones with homology of at least 99% belonged to the genera Acinetobacte, Alcaligenes, Delftia, Ochrobactrum, Pseudomonas, Stenotrophomonas, and Xanthobacter. PCR analysis showed that 13 out of 32 isolates contained the sequences (-1070 bp) homologous to the nitrilase genes reported previously in Alcaligenes faecalis JM3 (GenBank, D13419.1). Nine clones were capable of nitrile and amide transformation in minimal salt medium. Acinetobacter sp. 11 h and Alcaligenes sp. osv transformed 3-cyanopyridine to nicotinamide, while most of the clones possessed amidase activity (0.5 to 46.3 mmol/(g h) for acetamide and 0.1 to 5.6 mmol/(g h) for nicotinamide). Nicotinamide utilization by strain A. faecalis 2 was shown to result in excretion of a secondary metabolite, which was identified as dodecyl acrylate at 91% probability.
Van Overtvelt, Laurence; Moussu, Helene; Horiot, Stéphane; Samson, Sandrine; Lombardi, Vincent; Mascarell, Laurent; van de Moer, Ariane; Bourdet-Sicard, Raphaëlle; Moingeon, Philippe
We compared immunomodulatory properties of 11 strains of lactic acid bacteria as well as their capacity to enhance sublingual immunotherapy efficacy in a murine asthma model. Two types of bacterial strains were identified, including: (i) potent inducers of IL-12p70 and IL-10 in dendritic cells, supporting IFN-gamma and IL-10 production in CD4+ T cells such as Lactobacillus helveticus; (ii) pure Th1 inducers such as L. casei. Sublingual administration in ovalbumin-sensitized mice of L. helveticus, but not L. casei, reduced airways hyperresponsiveness, bronchial inflammation and proliferation of specific T cells in cervical lymph nodes. Thus, probiotics acting as a Th1/possibly Treg, but not Th1 adjuvant, potentiate tolerance induction via the sublingual route.
Endo, Akihito; Nakamura, Saki; Konishi, Kenta; Nakagawa, Junichi; Tochio, Takumi
Prebiotic oligosaccharides confer health benefits on the host by modulating the gut microbiota. Intestinal lactic acid bacteria (LAB) are potential targets of prebiotics; however, the metabolism of oligosaccharides by LAB has not been fully characterized. Here, we studied the metabolism of eight oligosaccharides by 19 strains of intestinal LAB. Among the eight oligosaccharides used, 1-kestose, lactosucrose and galactooligosaccharides (GOSs) led to the greatest increases in the numbers of the strains tested. However, mono- and disaccharides accounted for more than half of the GOSs used, and several strains only metabolized the mono- and di-saccharides in GOSs. End product profiles indicated that the amounts of lactate produced were generally consistent with the bacterial growth recorded. Oligosaccharide profiling revealed the interesting metabolic manner in Lactobacillus paracasei strains, which metabolized all oligosaccharides, but left sucrose when cultured with fructooligosaccharides. The present study clearly indicated that the prebiotic potential of each oligosaccharide differs.
Al Kassaa, I; Hober, D; Hamze, M; Chihib, N E; Drider, D
Emerging resistance to antiviral agents is a growing public health concern worldwide as it was reported for respiratory, sexually transmitted and enteric viruses. Therefore, there is a growing demand for new, unconventional antiviral agents which may serve as an alternative to the currently used drugs. Meanwhile, published literature continues shedding the light on the potency of lactic acid bacteria (LAB) and their bacteriocins as antiviral agents. Health-promoting LAB probiotics may exert their antiviral activity by (1) direct probiotic-virus interaction; (2) production of antiviral inhibitory metabolites; and/or (3) via stimulation of the immune system. The aim of this review was to highlight the antiviral activity of LAB and substances they produce with antiviral activity.
Upadhyay, Abhinav; Upadhyaya, Indu; Mooyottu, Shankumar; Venkitanarayanan, Kumar
Listeria monocytogenes is a human enteric pathogen that causes severe foodborne illness in high-risk populations. Crossing the intestinal barrier is the first critical step for Listeria monocytogenes infection. Therefore, reducing L. monocytogenes colonization and invasion of intestinal epithelium and production of virulence factors could potentially control listeriosis in humans. This study investigated the efficacy of sub-inhibitory concentration (SIC) of the plant-derived antimicrobial eugenol, either alone, or in combination with five lactic acid bacteria (LAB), namely Bifidobacterium bifidum (NRRL-B41410), Lactobacillus reuteri (B-14172), Lactobacillus fermentum (B-1840), Lactobacillus plantarum (B-4496) and Lactococcus lactis subspecies lactis (B-633) in reducing Listeria monocytogenes adhesion to and invasion of human intestinal epithelial cells (Caco-2). Additionally, the effect of the aforementioned treatments on Listeria monocytogenes listeriolysin production, epithelial E-cadherin binding and expression of virulence genes was investigated. Moreover, the in vivo efficacy of eugenol-LAB treatments in reducing Listeria monocytogenes virulence in the invertebrate model Galleria mellonella was studied. Eugenol and LAB, either alone or in combination, significantly reduced Listeria monocytogenes adhesion to and invasion of intestinal cells (P < 0.05). Moreover, eugenol-LAB treatments decreased Listeria monocytogenes haemolysin production, E-cadherin binding and virulence gene expression (P < 0.05). In addition, the eugenol-LAB treatments significantly enhanced the survival rates of G. mellonella infected with lethal doses of Listeria monocytogenes (P < 0.05). The results highlight the antilisterial effect of eugenol either alone or in combination with LAB, and justify further investigations in a mammalian model.
Garai-Ibabe, G; Ibarburu, I; Berregi, I; Claisse, O; Lonvaud-Funel, A; Irastorza, A; Dueñas, M T
Several lactic acid bacteria were isolated from bitter tasting ciders in which glycerol was partially removed. The degradation of glycerol via glycerol dehydratase pathway was found in 22 out of 67 isolates. The confirmation of glycerol degradation by this pathway was twofold: showing their glycerol dehydratase activity and detecting the presence of the corresponding gene by a PCR method. 1,3-propanediol (1,3-PDL) and 3-hydroxypropionic acid (3-HP) were the metabolic end-products of glycerol utilization, and the accumulation of the acrolein precursor 3-hydroxypropionaldehyde (3-HPA) was also detected in most of them. The strain identification by PCR-DGGE rpoB showed that Lactobacillus collinoides was the predominant species and only 2 belonged to Lactobacillus diolivorans. Environmental conditions conducting to 3-HPA accumulation in cidermaking were studied by varying the fructose concentration, pH and incubation temperature in L. collinoides 17. This strain failed to grow with glycerol as sole carbon source and the addition of fructose enhanced both growth and glycerol degradation. Regarding end-products of glycerol metabolism, 1,3-PDL was always the main end-product in all environmental conditions assayed, the only exception being the culture with 5.55 mM fructose, where equimolar amounts of 1,3-PDL and 3-HP were found. The 3-HPA was transitorily accumulated in the culture medium under almost all culture conditions, the degradation rate being notably slower at 15 degrees C. However, no disappearance of 3-HPA was found at pH 3.6, a usual value in cider making. After sugar exhaustion, L. collinoides 17 oxidated lactic acid and/or mannitol to obtain energy and these oxidations were accompanied by the removal of the toxic 3-HPA increasing the 1,3-PDL, 3-HP and acetic acid contents.
Frampton, E. W.
Frampton, E. W. (The University of Texas M. D. Anderson Hospital and Tumor Institute, Houston). Synthesis of ribonucleic acid by X-irradiated bacteria. J. Bacteriol. 87:1369–1376. 1964.—Postirradiation synthesis of total ribonucleic acid (RNA) and of RNA components was measured after exposure of Escherichia coli B/r to X rays. Net synthesis of RNA measured by the orcinol reaction and by the incorporation of uridine-2-C14 was depressed in irradiated cells, but paralleled the period of postirradiation growth (30 to 40 min). Incorporation of uridine-2-C14, added after net synthesis of RNA had ceased, detected an apparent turnover in a portion of the RNA. Irradiated cells retained their ability to adjust RNA synthesis to growth rate. After a shift-down in growth rate, irradiated cells incorporated radioactive uridine, while the net synthesis of RNA ceased—presumptive evidence for a continued synthesis of messenger RNA. Chloramphenicol addition (100 μg/ml) did not influence the total amount of RNA synthesized. Synthesis of ribosomes and transfer RNA preceded by 0, 5, 10, and 15 min of postirradiation incubation was observed by the resolution of cell-free extracts on sucrose density gradients. Little immediate influence of irradiation could be detected on the synthesis of 50S and 30S ribosomes. A decline was observed in the synthesis of 50S ribosomes with continued postirradiation incubation; 30S ribosomes, ribosomal precursors, and 4S RNA continued to be synthesized. PMID:14188715
Kim, Dong-Hoon; Lee, Ji-Hye; Hwang, Yuhoon; Kang, Seoktae; Kim, Mi-Sun
In the present work, we introduced a novel approach for microbial fatty acids (FA) production. Photosynthetic bacteria, Rhodobacter sphaeroides KD131, were cultivated in a continuous-flow, stirred-tank reactor (CFSTR) at various substrate (lactate) concentrations. At hydraulic retention time (HRT) 4d, cell concentration continuously increased from 0.97 g dcw/L to 2.05 g dcw/L as lactate concentration increased from 30 mM to 60mM. At 70 mM, however, cell concentration fluctuated with incomplete substrate degradation. By installing a membrane unit to CFSTR, a stable performance was observed under much higher substrate loading (lactate 100mM and HRT 1.5d). A maximum cell concentration of 16.2g dcw/L, cell productivity of 1.9 g dcw/L/d, and FA productivity of 665 mg FA/L/d were attained, and these values were comparable with those achieved using microalgae. The FA content of R. sphaeroides was around 35% of dry cell weight, mainly composed of vaccenic acid (C18:1, omega-7).
Pang, Huili; Zhang, Meng; Qin, Guangyong; Tan, Zhongfang; Li, Zongwei; Wang, Yanping; Cai, Yimin
One hundred and twenty-six strains were isolated from corn stover in Henan Province, China, of which 105 isolates were considered to be lactic acid bacteria (LAB) according to Gram-positive, catalase-negative and mainly metabolic lactic acid product. Analysis of the 16S ribosomal DNA sequence of 21 representative strains was used to confirm the presence of the predominant groups and to determine the phylogenetic affiliation of isolates. The sequences from the various LAB isolates showed high degrees of similarity to those of the GenBank type strains between 99.4% and 100%. The prevalent LAB, predominantly Lactobacillus (85.6%), consisted of L. plantarum (33.3%), L. pentosus (28.6%) and L. brevis (23.7%). Other LAB species as Leuconostoc lactis (4.8%), Weissella cibaria (4.8%) and Enterococcus mundtii (4.8%) also presented in corn stover. The present study is the first to fully document corn stover-associated LAB involved in the silage fermentation. The identification results revealed LAB composition inhabiting corn stover and enabling the future design of appropriate inoculants aimed at improving the fermentation quality of silage.
Kishino, Shigenobu; Ogawa, Jun; Yokozeki, Kenzo; Shimizu, Sakayu
Lactobacillus plantarum AKU 1009a effectively transforms linoleic acid to conjugated linoleic acids of cis-9,trans-11-octadecadienoic acid (18:2) and trans-9,trans-11-18:2. The transformation of various polyunsaturated fatty acids by washed cells of L. plantarum AKU 1009a was investigated. Besides linoleic acid, alpha-linolenic acid [cis-9,cis-12,cis-15-octadecatrienoic acid (18:3)], gamma-linolenic acid (cis-6,cis-9,cis-12-18:3), columbinic acid (trans-5,cis-9,cis-12-18:3), and stearidonic acid [cis-6,cis-9,cis-12,cis-15-octadecatetraenoic acid (18:4)] were found to be transformed. The fatty acids transformed by the strain had the common structure of a C18 fatty acid with the cis-9,cis-12 diene system. Three major fatty acids were produced from alpha-linolenic acid, which were identified as cis-9,trans-11,cis-15-18:3, trans-9,trans-11,cis-15-18:3, and trans-10,cis-15-18:2. Four major fatty acids were produced from gamma-linolenic acid, which were identified as cis-6,cis-9,trans-11-18:3, cis-6,trans-9,trans-11-18:3, cis-6,trans-10-18:2, and trans-10-octadecenoic acid. The strain transformed the cis-9,cis-12 diene system of C18 fatty acids into conjugated diene systems of cis-9,trans-11 and trans-9,trans-11. These conjugated dienes were further saturated into the trans-10 monoene system by the strain. The results provide valuable information for understanding the pathway of biohydrogenation by anaerobic bacteria and for establishing microbial processes for the practical production of conjugated fatty acids, especially those produced from alpha-linolenic acid and gamma-linolenic acid.
Cruciata, Margherita; Sannino, Ciro; Ercolini, Danilo; Scatassa, Maria L.; De Filippis, Francesca; Mancuso, Isabella; La Storia, Antonietta; Moschetti, Giancarlo
The microbial composition of artisan and industrial animal rennet pastes was studied by using both culture-dependent and -independent approaches. Pyrosequencing targeting the 16S rRNA gene allowed to identify 361 operational taxonomic units (OTUs) to the genus/species level. Among lactic acid bacteria (LAB), Streptococcus thermophilus and some lactobacilli, mainly Lactobacillus crispatus and Lactobacillus reuteri, were the most abundant species, with differences among the samples. Twelve groups of microorganisms were targeted by viable plate counts revealing a dominance of mesophilic cocci. All rennets were able to acidify ultrahigh-temperature-processed (UHT) milk as shown by pH and total titratable acidity (TTA). Presumptive LAB isolated at the highest dilutions of acidified milks were phenotypically characterized, grouped, differentiated at the strain level by randomly amplified polymorphic DNA (RAPD)-PCR analysis, and subjected to 16S rRNA gene sequencing. Only 18 strains were clearly identified at the species level, as Enterococcus casseliflavus, Enterococcus faecium, Enterococcus faecalis, Enterococcus lactis, Lactobacillus delbrueckii, and Streptococcus thermophilus, while the other strains, all belonging to the genus Enterococcus, could not be allotted into any previously described species. The phylogenetic analysis showed that these strains might represent different unknown species. All strains were evaluated for their dairy technological performances. All isolates produced diacetyl, and 10 of them produced a rapid pH drop in milk, but only 3 isolates were also autolytic. This work showed that animal rennet pastes can be sources of LAB, mainly enterococci, that might contribute to the microbial diversity associated with dairy productions. PMID:24441167
Cruciata, Margherita; Sannino, Ciro; Ercolini, Danilo; Scatassa, Maria L; De Filippis, Francesca; Mancuso, Isabella; La Storia, Antonietta; Moschetti, Giancarlo; Settanni, Luca
The microbial composition of artisan and industrial animal rennet pastes was studied by using both culture-dependent and -independent approaches. Pyrosequencing targeting the 16S rRNA gene allowed to identify 361 operational taxonomic units (OTUs) to the genus/species level. Among lactic acid bacteria (LAB), Streptococcus thermophilus and some lactobacilli, mainly Lactobacillus crispatus and Lactobacillus reuteri, were the most abundant species, with differences among the samples. Twelve groups of microorganisms were targeted by viable plate counts revealing a dominance of mesophilic cocci. All rennets were able to acidify ultrahigh-temperature-processed (UHT) milk as shown by pH and total titratable acidity (TTA). Presumptive LAB isolated at the highest dilutions of acidified milks were phenotypically characterized, grouped, differentiated at the strain level by randomly amplified polymorphic DNA (RAPD)-PCR analysis, and subjected to 16S rRNA gene sequencing. Only 18 strains were clearly identified at the species level, as Enterococcus casseliflavus, Enterococcus faecium, Enterococcus faecalis, Enterococcus lactis, Lactobacillus delbrueckii, and Streptococcus thermophilus, while the other strains, all belonging to the genus Enterococcus, could not be allotted into any previously described species. The phylogenetic analysis showed that these strains might represent different unknown species. All strains were evaluated for their dairy technological performances. All isolates produced diacetyl, and 10 of them produced a rapid pH drop in milk, but only 3 isolates were also autolytic. This work showed that animal rennet pastes can be sources of LAB, mainly enterococci, that might contribute to the microbial diversity associated with dairy productions.
Crotti, Elena; Rizzi, Aurora; Chouaia, Bessem; Ricci, Irene; Favia, Guido; Alma, Alberto; Sacchi, Luciano; Bourtzis, Kostas; Mandrioli, Mauro; Cherif, Ameur; Bandi, Claudio; Daffonchio, Daniele
Recent research in microbe-insect symbiosis has shown that acetic acid bacteria (AAB) establish symbiotic relationships with several insects of the orders Diptera, Hymenoptera, Hemiptera, and Homoptera, all relying on sugar-based diets, such as nectars, fruit sugars, or phloem sap. To date, the fruit flies Drosophila melanogaster and Bactrocera oleae, mosquitoes of the genera Anopheles and Aedes, the honey bee Apis mellifera, the leafhopper Scaphoideus titanus, and the mealybug Saccharicoccus sacchari have been found to be associated with the bacterial genera Acetobacter, Gluconacetobacter, Gluconobacter, Asaia, and Saccharibacter and the novel genus Commensalibacter. AAB establish symbiotic associations with the insect midgut, a niche characterized by the availability of diet-derived carbohydrates and oxygen and by an acidic pH, selective factors that support AAB growth. AAB have been shown to actively colonize different insect tissues and organs, such as the epithelia of male and female reproductive organs, the Malpighian tubules, and the salivary glands. This complex topology of the symbiosis indicates that AAB possess the keys for passing through body barriers, allowing them to migrate to different organs of the host. Recently, AAB involvement in the regulation of innate immune system homeostasis of Drosophila has been shown, indicating a functional role in host survival. All of these lines of evidence indicate that AAB can play different roles in insect biology, not being restricted to the feeding habit of the host. The close association of AAB and their insect hosts has been confirmed by the demonstration of multiple modes of transmission between individuals and to their progeny that include vertical and horizontal transmission routes, comprising a venereal one. Taken together, the data indicate that AAB represent novel secondary symbionts of insects. PMID:20851977
Säde, Elina; Lassila, Elisa; Björkroth, Johanna
Spices and dried vegetable seasonings are potential sources of bacterial contamination for foods. However, little is known about lactic acid bacteria (LAB) in spices and dried vegetables, even though certain LAB may cause food spoilage. In this study, we enumerated LAB in 104 spices and dried vegetables products aimed for the food manufacturing industry. The products were obtained from a spice wholesaler operating in Finland, and were sampled during a one-year period. We picked isolates (n = 343) for species identification based on numerical analysis of their ribotyping patterns and comparing them with the corresponding patterns of LAB type strains. We found LAB at levels >2 log CFU/g in 68 (65%) of the samples, with the highest counts detected from dried onion products and garlic powder with counts ranging from 4.24 to 6.64 log CFU/g. The LAB identified were predominantly Weissella spp. (61%) and Pediococcus spp. (15%) with Weissella confusa, Weissella cibaria, Weissella paramesenteroides, Pediococcus acidilactici and Pediococcus pentosaceus being the species identified. Other species identified belonged to the genera of Enterococcus spp. (8%), Leuconostoc spp. (6%) and Lactobacillus spp. (2%). Among the LAB identified, Leuconostoc citreum, Leuconostoc mesenteroides and W. confusa have been associated with food spoilage. Our findings suggest that spices and dried vegetables are potential sources of LAB contamination in the food industry.
Kong, Wentao; Kapuganti, Venkata S; Lu, Ting
Recent developments in synthetic biology have positioned lactic acid bacteria (LAB) as a major class of cellular chassis for applications. To achieve the full potential of LAB, one fundamental prerequisite is the capacity for rapid engineering of complex gene networks, such as natural biosynthetic pathways and multicomponent synthetic circuits, into which cellular functions are encoded. Here, we present a synthetic biology platform for rapid construction and optimization of large-scale gene networks in LAB. The platform involves a copy-controlled shuttle for hosting target networks and two associated strategies that enable efficient genetic editing and phenotypic validation. By using a nisin biosynthesis pathway and its variants as examples, we demonstrated multiplex, continuous editing of small DNA parts, such as ribosome-binding sites, as well as efficient manipulation of large building blocks such as genes and operons. To showcase the platform, we applied it to expand the phenotypic diversity of the nisin pathway by quickly generating a library of 63 pathway variants. We further demonstrated its utility by altering the regulatory topology of the nisin pathway for constitutive bacteriocin biosynthesis. This work demonstrates the feasibility of rapid and advanced engineering of gene networks in LAB, fostering their applications in biomedicine and other areas.
Nakano, Shigeru; Matsumura, Atsushi; Yamada, Toshihiro
A PCR assay for the detection of acetic acid-tolerant lactic acid bacteria in the genera of Lactobacillus and Pediococcus was developed in this study. Primers targeting the bacterial 16S rRNA gene were newly designed and used in this PCR assay. To determine the specificity of the assay, 56 different bacterial strains (of 33 genera), 2 fungi, 3 animals, and 4 plants were tested. Results were positive for most tested bacterial members of 16S rRNA gene-based phylogenetic groups (classified in the Lactobacillus casei and Pediococcus group), including Lactobacillus fructivorans, Lactobacillus brevis, Lactobacillus buchneri, Lactobacillus plantarum, and Lactobacillus paracasei. For all other bacterial strains and eukaryote tested, results were negative. Bacterial DNA for PCR was prepared with a simple procedure with the use of Chelex 100 resin from culture after growth in deMan Rogosa Sharpe broth (pH 6.0). To test this PCR assay for the monitoring of the acetic acid-tolerant lactic acid bacteria, L. fructivorans was inoculated into several acidic food as an indicator. Before the PCR, the inoculation of 10 to 50 CFU of bacteria per g of food was followed by a 28-h enrichment culture step, and the PCR assay allowed the detection of bacterial cells. Including the enrichment culture step, the entire PCR detection process can be completed within 30 h.
Branched-chain fatty acids of the iso and anteiso series occur in many bacteria as the major acyl constituents of membrane lipids. In addition, omega-cyclohexyl and omega-cycloheptyl fatty acids are present in several bacterial species. These two types of fatty acids are synthesized by the repeated condensation of malonyl coenzyme A with one of the branched-chain and cyclic primers by the same enzyme system. The pathway of de novo branched-chain fatty acid synthesis differs only in initial steps of synthesis from that of the common straight-chain fatty acid (palmitic acid) present in most organisms. The cell membranes composed largely of iso-, anteiso-, and omega-alicyclic acids support growth of bacteria, which inhabit normal as well as extreme environments. The occurrence of these types of fatty acids as major cellular fatty acids is an important criterion used to aid identification and classification of bacteria. PMID:1886522
Ellis, J L; Hindrichsen, I K; Klop, G; Kinley, R D; Milora, N; Bannink, A; Dijkstra, J
Inoculants of lactic acid bacteria (LAB) are used to improve silage quality and prevent spoilage via increased production of lactic acid and other organic acids and a rapid decline in silage pH. The addition of LAB inoculants to silage has been associated with increases in silage digestibility, dry matter intake (DMI), and milk yield. Given the potential change in silage and rumen fermentation conditions accompanying these silage additives, the aim of this study was to investigate the effect of LAB silage inoculants on DMI, digestibility, milk yield, milk composition, and methane (CH4) production from dairy cows in vivo. Eight mid-lactation Holstein-Friesian dairy cows were grouped into 2 blocks of 4 cows (multiparous and primiparous) and used in a 4×4 double Latin square design with 21-d periods. Methane emissions were measured by indirect calorimetry. Treatments were grass silage (mainly ryegrass) with no inoculant (GS), with a long-term inoculant (applied at harvest; GS+L), with a short-term inoculant (applied 16h before feeding; GS+S), or with both long and short-term inoculants (GS+L+S). All diets consisted of grass silage and concentrate (75:25 on a dry matter basis). The long-term inoculant consisted of a 10:20:70 mixture of Lactobacillus plantarum, Lactococcus lactis, and Lactobacillus buchneri, and the short-term inoculant was a preparation of Lc. lactis. Dry matter intake was not affected by long-term or short-term silage inoculation, nor was dietary neutral detergent fiber or fat digestibility, or N or energy balance. Milk composition (except milk urea) and fat and protein-corrected milk yield were not affected by long- or short-term silage inoculation, nor was milk microbial count. However, milk yield tended to be greater with long-term silage inoculation. Methane expressed in units of grams per day, grams per kilogram of DMI, grams per kilogram of milk, or grams per kilogram of fat and protein-corrected milk yield was not affected by long- or short
We designed Lactococcus lactis to detect Enterococcus faecalis. Upon detection, L. lactis produce and secrete antienterococcal peptides. The peptides inhibit enterococcal growth and reduce viability of enterococci in the vicinity of L. lactis. The enterococcal sex pheromone cCF10 serves as the signal for detection. Expression vectors derived from pCF10, a cCF10-responsive E. faecalis sex-pheromone conjugative plasmid, were engineered in L. lactis for the detection system. Recombinant host strains were engineered to express genes for three bacteriocins, enterocin A, hiracin JM79 and enterocin P, each with potent antimicrobial activity against E. faecalis. Sensitive detection and specific inhibition occur both in agar and liquid media. The engineered L. lactis also inhibited growth of multidrug-resistant E. faecium strains, when induced by cCF10. The presented vectors and strains can be components of a toolbox for the development of alternative antibiotic technologies targeting enterococci at the site of infection. PMID:24896372
Saraoui, Taous; Cornet, Josiane; Guillouet, Emilie; Pilet, Marie France; Chevalier, Frédérique; Joffraud, Jean-Jacques; Leroi, Françoise
Tropical shrimp is of considerable economic importance in the world but is highly perishable due to microbial and chemical degradation. Biopreservation is a food preservation technology based on the addition of "positive" bacteria able to kill or prevent the growth of undesirable microorganisms. Two strains of lactic acid bacteria (LAB) have previously been selected for a biopreservation strategy: Lactococcus piscium CNCM I-4031, for its ability to prevent the sensory deterioration of seafood and Carnobacterium divergens V41, which inhibits growth of Listeria monocytogenes. The objective was to test the association of the two strains to improve both the quality and safety of shrimp. In a first trial, the two LAB were inoculated alone or in a cocktail in cooked and peeled shrimp (CPS) Penaeus vannamei at 5×10(5)CFU/g. Chemical, sensory and microbiological analyses by culture-dependent and -independent methods were performed during storage under modified atmosphere packaging (MAP) at 8°C. The results were compared to a non-inoculated batch. In a second trial, the same experiments were repeated in the presence of 10(2)CFU/g of L. monocytogenes RF191. The microbiota of CPS was composed of LAB, Shewanella spp. and Enterobacteriaceae. Brochothrix thermosphacta was not detected. L. piscium and C. divergens reached 10(8) and 10(9)CFU/g, respectively, in 7days and did not inhibit each other when co-inoculated. L. piscium reduced L. monocytogenes by 1Log (CFU/g) for 28days. C. divergens had an immediate listericidal effect lasting 7days. A regrowth of L. monocytogenes was then observed but the count was always 2 to 5Log (CFU/g) lower than in the control. No additional or synergic effect between protective strains was observed and the cocktail had the same inhibitory effect as C. divergens alone. C. divergens was very effective at preventing the sensory deterioration of CPS. This may be related to the inhibition of Shewanella and Enterobacteriaceae. However, the panelists
Nguyen, Nguyen Khoi; Dong, Ngan Thi Ngoc; Nguyen, Huong Thuy; Le, Phu Hong
Kombucha is sweetened black tea that is fermented by a symbiosis of bacteria and yeast embedded within a cellulose membrane. It is considered a health drink in many countries because it is a rich source of vitamins and may have other health benefits. It has previously been reported that adding lactic acid bacteria (Lactobacillus) strains to kombucha can enhance its biological functions, but in that study only lactic acid bacteria isolated from kefir grains were tested. There are many other natural sources of lactic acid bacteria. In this study, we examined the effects of lactic acid bacteria from various fermented Vietnamese food sources (pickled cabbage, kefir and kombucha) on kombucha's three main biological functions: glucuronic acid production, antibacterial activity and antioxidant ability. Glucuronic acid production was determined by high-performance liquid chromatography-mass spectrometry, antibacterial activity was assessed by the agar-well diffusion method and antioxidant ability was evaluated by determining the 2,2-diphenyl-1-picrylhydrazyl radical scavenging capacity. Four strains of food-borne pathogenic bacteria were used in our antibacterial experiments: Listeria monocytogenes ATCC 19111, Escherichia coli ATCC 8739, Salmonella typhimurium ATCC 14028 and Bacillus cereus ATCC 11778. Our findings showed that lactic acid bacteria strains isolated from kefir are superior to those from other sources for improving glucuronic acid production and enhancing the antibacterial and antioxidant activities of kombucha. This study illustrates the potential of Lactobacillus casei and Lactobacillus plantarum isolated from kefir as biosupplements for enhancing the bioactivities of kombucha.
Watanabe, Yuko; Hayakawa, Kiyoshi; Ueno, Hiroshi
Lactic acid bacteria was searched for producing termented tea that contained a lot of γ-amino butyric acid(GABA). Also examined were the growth condition, GABA production and changes in catechin contents in the tea leaves. Lactobacillus brevis L12 was found to be suitable for the production of fermented tea since it gave as much GABA as gabaron tea when tea leaves being suspended with water at 10% and incubated for 4 days at 25°C. The amount of GABA produced was more than calculated based upon the content of glutamic acid in tea leaves. It is probable to assume that glutamate derived from glutamine and theanine is converted into GABA.
Zigangirova, N A; Tokarskaia, E A; Narodnitskiĭ, B S; Gintsburg, A L; Tugel'ian, V A
The wide use of antibiotics in livestock raising has contributed to the selection and accumulation of representatives of commensal microflora, as well as pathogenic bacteria, colonizing livestock and poultry. For this reason the problem of the possible transfer of antibiotic-resistance genes along the chain from bacteria, autochthonous for agricultural animals, to bacteria used for the production of foodstuffs, which are incorporated into normal microflora and may thus participate in the exchange of these genes with bacteria, enteropathogenic for humans, is a highly important task of medical microbiology. The article deals with the review of experimental data, indicative the possibility of the appearance of antibiotic-resistant pathogenic bacteria due to the transfer of antibiotic-resistance genes via alimentary chains.
Mahony, Jennifer; Ainsworth, Stuart; Stockdale, Stephen; Sinderen, Douwe van
Dairy fermentations are among the oldest food processing applications, aimed at preservation and shelf-life extension through the use of lactic acid bacteria (LAB) starter cultures, in particular strains of Lactococcus lactis, Streptococcus thermophilus, Lactobacillus spp. and Leuconostoc spp. Traditionally this was performed by continuous passaging of undefined cultures from a finished fermentation to initiate the next fermentation. More recently, consumer demands on consistent and desired flavours and textures of dairy products have led to a more defined approach to such processes. Dairy (starter) companies have responded to the need to define the nature and complexity of the starter culture mixes, and dairy fermentations are now frequently based on defined starter cultures of low complexity, where each starter component imparts specific technological properties that are desirable to the product. Both mixed and defined starter culture approaches create the perfect environment for the proliferation of (bacterio)phages capable of infecting these LAB. The repeated use of the same starter cultures in a single plant, coupled to the drive towards higher and consistent production levels, increases the risk and negative impact of phage infection. In this review we will discuss recent advances in tracking the adaptation of phages to the dairy industry, the advances in understanding LAB phage-host interactions, including evolutionary and genomic aspects.
Léonard, Lucie; Husson, Florence; Langella, Philippe; Châtel, Jean-Marc; Saurel, Rémi
The present study aimed to entrap probiotic lactic acid bacteria (LAB) in a sodium alginate and sodium caseinate aqueous two-phase gel system. The natural acidifying properties of two therapeutic probiotic LAB were exploited to liberate calcium ions progressively from calcium carbonate (CaCO3), which caused the gelation of the co-existing phases. Bi-biopolymeric matrix gelation of GDL/CaCO3 or LAB/CaCO3 was monitored by dynamic rheological measurements, and the final gels were characterized by frequency dependence measurements and confocal laser scanning microscopy. Weak to strong gels were formed with an elastic modulus G' from 10 to 1.000Pa, respectively. After cold-set gelation of our system, confocal laser scanning microscopy showed spherical protein microdomains trapped within a calcium alginate network. LAB cells were stained to study their partition in the self-gelling matrices. Our LAB strains showed two different behaviors, which may relate to the exopolysaccharide production: (i) Lactobacillus plantarum CNRZ1997 cells were found mainly in continuous alginate networks, whereas (ii) Lactococcus lactis cells were localized in protein microdomains. This alginate-caseinate phase-separated system that was self-gelled by LAB cells may be an innovative approach for immobilizing and protecting LAB cells.
Stevenson, David M; Muck, Richard E; Shinners, Kevin J; Weimer, Paul J
Real-time polymerase chain reaction (RT-PCR) was used to quantify seven species of lactic acid bacteria (LAB) in alfalfa silage prepared in the presence or absence of four commercial inoculants and in uninoculated corn stover harvested and stored under a variety of field conditions. Species-specific PCR primers were designed based on recA gene sequences. Commercial inoculants improved the quality of alfalfa silage, but species corresponding to those in the inoculants displayed variations in persistence over the next 96 h. Lactobacillus brevis was the most abundant LAB (12 to 32% of total sample DNA) in all of the alfalfa silages by 96 h. Modest populations (up to 10%) of Lactobacillus plantarum were also observed in inoculated silages. Pediococcus pentosaceus populations increased over time but did not exceed 2% of the total. Small populations (0.1 to 1%) of Lactobacillus buchneri and Lactococcus lactis were observed in all silages, while Lactobacillus pentosus and Enterococcus faecium were near or below detection limits. Corn stover generally displayed higher populations of L. plantarum and L. brevis and lower populations of other LAB species. The data illustrate the utility of RT-PCR for quantifying individual species of LAB in conserved forages prepared under a wide variety of conditions.
Muñoz-Atienza, Estefanía; Araújo, Carlos; Lluch, Nuria; Hernández, Pablo E; Herranz, Carmen; Cintas, Luis M; Magadán, Susana
In aquaculture, several criteria should be considered to select an appropriate probiotic, including the aquatic origin and safety of the strain and its ability to modulate the host immune response. The properties and effects of probiotics are strain-specific and some factors such as viability, dose and duration of diet supplementation may regulate their immunomodulatory activities. In this study, we assessed the in vitro effect of eight heat-inactivated and viable lactic acid bacteria (LAB) of aquatic origin belonging to the genera Enterococcus, Lactobacillus, Lactococcus, Leuconostoc, Pediococcus and Weissella on the viability and innate immune response of turbot (Scophthalmus maximus L.) leucocytes. Head-kidney leucocytes were incubated with viable and heat-inactivated LAB at different concentrations. After incubation, the viability of leucocytes was evaluated using colorimetric assays (MTT and LDH) and flow cytometry (annexin V/propidium iodide). Heat-inactivated LAB showed no cytotoxic effect while viable LAB exerted variable influence on apoptosis of turbot phagocytes and lymphocytes. Leucocyte respiratory burst activity and phagocytosis were also differentially activated, as viable LAB stimulated leucocytes more efficiently than the heat-inactivated LAB. Our results suggest diverse strain-specific mechanisms of interaction between the evaluated LAB and turbot leucocytes. Furthermore, our work sets up in vitro systems to evaluate the effect of LAB as potential probiotics, which will be useful to develop efficient screening.
Toy, Nurten; Özogul, Fatih; Özogul, Yesim
The function of cell-free solutions (CFSs) of lactic acid bacteria (LAB) on tyramine and other biogenic amine production by different food borne-pathogens (FBPs) was investigated in tyrosine decarboxylase broth (TDB) using HPLC. Cell free solutions were prepared from four LAB strains. Two different concentrations which were 50% (5 ml CFS+5 ml medium/1:1) and 25% (2.5 ml CFS+7.5 ml medium/1:3) CFS and the control without CFS were prepared. Both concentration of CFS of Streptococcus thermophilus and 50% CFS of Pediococcus acidophilus inhibited tyramine production up to 98% by Salmonella paratyphi A. Tyramine production by Escherichia coli was also inhibited by 50% CFS of Lactococcus lactis subsp. lactis and 25% CFS of Leuconostoc lactis. subsp. cremoris. The inhibitor effect of 50% CFS of P. acidophilus was the highest on tyramine production (55%) by Listeria monocytogenes, following Lc. lactis subsp. lactis and Leuconostoc mesenteroides subsp. cremoris (20%) whilst 25% CFS of Leu. mes. subsp. cremoris and Lc. lactis subsp. lactis showed stimulator effects (160%). The stimulation effects of 50% CFS of S. thermophilus and Lc. lactis subsp. lactis were more than 70% by Staphylococcus aureus comparing to the control. CFS of LAB strains showed statistically inhibitor effect since lactic acid inhibited microbial growth, decreased pH quickly and reduced the formation of AMN and BAs. Consequently, in order to avoid the formation of high concentrations of biogenic amines in fermented food by bacteria, it is advisable to use CFS for food and food products.
Wacher-Rodarte, Maria Del Carmen; Trejo-Muñúzuri, Tanya Paulina; Montiel-Aguirre, Jesús Fernando; Drago-Serrano, Maria Elisa; Gutiérrez-Lucas, Raúl L; Castañeda-Sánchez, Jorge Ismael; Sainz-Espuñes, Teresita
Pozol is a handcrafted nonalcoholic Mayan beverage produced by the spontaneous fermentation of maize dough by lactic acid bacteria. Lactic acid bacteria (LAB) are carriers of chromosomal encoded multidrug-resistant efflux pumps genes that can be transferred to pathogens and/or confer resistance to compounds released during the fermentation process causing food spoiling. The aim of this study was to evaluate the antibiotic sensibility and the transcriptional expression of ABC-type efflux pumps in LAB isolated from pozol that contributes to multidrug resistance. Analysis of LAB and Staphylococcus (S.) aureus ATCC 29213 and ATCC 6538 control strains to antibiotic susceptibility, minimal inhibitory concentration (MIC), and minimal bactericidal concentration (MBC) to ethidium bromide were based in "standard methods" whereas the ethidium bromide efflux assay was done by fluorometric assay. Transcriptional expression of efflux pumps was analyzed by RT-PCR. LAB showed antibiotic multiresistance profiles, moreover, Lactococcus (L.) lactis and Lactobacillus (L.) plantarum displayed higher ethidium bromide efflux phenotype than S. aureus control strains. Ethidium bromide resistance and ethidium bromide efflux phenotypes were unrelated with the overexpression of lmrD in L. lactics, or the underexpression of lmrA in L. plantarum and norA in S. aureus. These findings suggest that, moreover, the analyzed efflux pumps genes, other unknown redundant mechanisms may underlie the antibiotic resistance and the ethidium bromide efflux phenotype in L. lactis and L. plantarum. Phenotypic and molecular drug multiresistance assessment in LAB may improve a better selection of the fermentation starter cultures used in pozol, and to control the antibiotic resistance widespread and food spoiling for health safety.
Güngör, Ö E; Kırzıoğlu, Z; Dinçer, E; Kıvanç, M
Adhesion to oral soft and hard tissue is crucial for bacterial colonisation in the mouth. The aim of this work was to select strains of oral lactic acid bacteria that could be used as probiotics for oral health. To this end, the adhesive properties of some lactic acid bacteria were investigated. Seventeen lactic acid bacteria including two Streptococcus mutans strains were isolated from the oral cavity of healthy children, while other strains were isolated from fermented meat products. The bacterial strains were applied to teeth surfaces covered with saliva or without saliva. A significant diversity in adhesion capacity to teeth surfaces among the lactic acid bacteria was observed. Lactic acid bacteria isolated from the oral cavity adhered the best to teeth surfaces covered with saliva, whereas lactic acid bacteria isolated from fermented meat samples adhered the best to tooth surface without saliva. All strains of lactic acid bacteria were able to reduce the number of S. mutans cells, in particular on saliva-coated tooth surface. Therefore, they might have potential as probiotics for the oral cavity.
The naturally occurring compound, fumaric acid, was evaluated as a potential preservative for the long-term storage of cucumbers. Fumaric acid inhibited growth of lactic acid bacteria (LAB) in an acidified cucumber juice medium model system resembling conditions that could allow preservation of cucu...
Nomura, Toshiyuki; Kuriyama, Yuta; Tokumoto, Hayato; Konishi, Yasuhiro
The cytotoxicity and colloidal behavior of surface-functionalized polystyrene latex (PSL) nanoparticles (NPs) (nominal diameter: 100 nm) toward a model gram positive bacterium Lactococcus lactis JCM 5805 were examined. Nearly all the L. lactis cells exposed to the negatively charged PSL NPs survived because the surface of the bacterial cell was charged negatively, and the NPs therefore hardly adhere to the cell surface. In contrast, the positively charged PSL NPs adhered to the L. lactis cell surface but were not entrapped within the cell, and cell death subsequently occurred. The bacterial growth curves after the toxic NP exposure suggested that NP toxicity did not affect the specific growth phase, but did affect lag time. These results indicated that the cells were damaged by the cell disruption that resulted from the adhesion of the NPs to the cell surface. Finally, the cytotoxicity of the toxic, positively charged PSL NPs toward L. lactis was compared with that displayed toward a model gram negative bacterium Escherichia coli and a model eukaryote Saccharomyces cerevisiae. The cytotoxic behaviors of NPs on L. lactis and E. coli were similar, and depended not on the bacterial surface structure, but rather the environmental ionic strength. In contrast, the cytotoxicity of the prokaryote bacteria was higher than that toward the model eukaryote S. cerevisiae. The difference between the NP sensitivities of the prokaryote and eukaryote resulted from the prokaryote's lack of an endocytotic pathway.
Kimoto-Nira, Hiromi; Moriya, Naoko; Yamasaki, Seishi; Takenaka, Akio; Suzuki, Chise
The association of lactic acid bacteria with mucosal surfaces plays important roles in the beneficial effects of these bacteria on human health, such as colonization of the gastrointestinal tract for pathogen antagonism. Previously, we found that the adhesion of Lactococcus lactis 7-1 to porcine gastric mucin was higher with fructose than with lactose, galactose or xylose as the carbon source. In this study, we examined the effect of growth conditions on the adhesion of strain 7-1 grown on fructose. Medium components affect the adhesion: the adhesion of strain 7-1 grown with sodium acetate was higher than that without it. The enhancement of adhesion by sodium acetate was not observed under aerobic conditions. Cellular properties grown with or without sodium acetate were characterized: strain 7-1 grown with sodium acetate had similar sugar contents, and different fatty acid composition to those grown without it. Strain 7-1 grown with sodium acetate showed significantly lower cell yield and significantly higher hydrophobicity than those grown without it, which is associated with higher adhesion. Fructose and sodium acetate are frequently used in the food industry; this study may reveal a simple way to enhance the adhesion of lactic acid bacteria by growing them with these substances.
Narushima, Seiko; Itoha, Kikuji; Miyamoto, Yukiko; Park, Sang-Hee; Nagata, Keiko; Kuruma, Kazuo; Uchida, Kiyohisa
In humans and animals, intestinal flora is indispensable for bile acid transformation. The goal of our study was to establish gnotobiotic mice with intestinal bacteria of human origin in order to examine the role of intestinal bacteria in the transformation of bile acids in vivo using the technique of gnotobiology. Eight strains of bile acid-deconjugating bacteria were isolated from ex-germ-free mice inoculated with a human fecal dilution of 10(-6), and five strains of 7alpha-dehydroxylating bacteria were isolated from the intestine of limited human flora mice inoculated only with clostridia. The results of biochemical tests and 16S rDNA sequence analysis showed that seven out of eight bile acid-deconjugating strains belong to a bacteroides cluster (Bacteroides vulgatus, B. distasonis, and B. uniformis), and one strain had high similarity with Bilophila wadsworthia. All five strains that converted cholic acid to deoxycholic acid had greatest similarity with Clostridium hylemonae. A combination of 10 isolated strains converted taurocholic acid into deoxycholic acid both in vitro and in the mouse intestine. These results indicate that the predominant bacteria, mainly Bacteroides, in human feces comprise one of the main bacterial groups for the deconjugation of bile acids, and clostridia may play an important role in 7aplha-dehydroxylation of free-form primary bile acids in the intestine although these strains are not predominant. The gnotobiotic mouse with bacteria of human origin could be a useful model in studies of bile acid metabolism by human intestinal bacteria in vivo.
Du, Huihui; Chen, Wenli; Cai, Peng; Rong, Xingmin; Dai, Ke; Peacock, Caroline L.; Huang, Qiaoyun
Soil components (e.g., clays, bacteria and humic substances) are known to produce mineral-organic composites in natural systems. Herein, batch sorption isotherms, isothermal titration calorimetry (ITC), and Cd K-edge EXAFS spectroscopy were applied to investigate the binding characteristics of Cd on montmorillonite(Mont)-humic acid(HA)-bacteria composites. Additive sorption and non-additive Cd(II) sorption behaviour is observed for the binary Mont-bacteria and ternary Mont-HA-bacteria composite, respectively. Specifically, in the ternary composite, the coexistence of HA and bacteria inhibits Cd adsorption, suggesting a “blocking effect” between humic acid and bacterial cells. Large positive entropies (68.1 ~ 114.4 J/mol/K), and linear combination fitting of the EXAFS spectra for Cd adsorbed onto Mont-bacteria and Mont-HA-bacteria composites, demonstrate that Cd is mostly bound to bacterial surface functional groups by forming inner-sphere complexes. All our results together support the assertion that there is a degree of site masking in the ternary clay mineral-humic acid-bacteria composite. Because of this, in the ternary composite, Cd preferentially binds to the higher affinity components-i.e., the bacteria.
Du, Huihui; Chen, Wenli; Cai, Peng; Rong, Xingmin; Dai, Ke; Peacock, Caroline L.; Huang, Qiaoyun
Soil components (e.g., clays, bacteria and humic substances) are known to produce mineral-organic composites in natural systems. Herein, batch sorption isotherms, isothermal titration calorimetry (ITC), and Cd K-edge EXAFS spectroscopy were applied to investigate the binding characteristics of Cd on montmorillonite(Mont)-humic acid(HA)-bacteria composites. Additive sorption and non-additive Cd(II) sorption behaviour is observed for the binary Mont-bacteria and ternary Mont-HA-bacteria composite, respectively. Specifically, in the ternary composite, the coexistence of HA and bacteria inhibits Cd adsorption, suggesting a “blocking effect” between humic acid and bacterial cells. Large positive entropies (68.1 ~ 114.4 J/mol/K), and linear combination fitting of the EXAFS spectra for Cd adsorbed onto Mont-bacteria and Mont-HA-bacteria composites, demonstrate that Cd is mostly bound to bacterial surface functional groups by forming inner-sphere complexes. All our results together support the assertion that there is a degree of site masking in the ternary clay mineral-humic acid-bacteria composite. Because of this, in the ternary composite, Cd preferentially binds to the higher affinity components-i.e., the bacteria. PMID:26792640
HAKIM, Hakimullah; ALAM, Md. Shahin; SANGSRIRATANAKUL, Natthanan; NAKAJIMA, Katsuhiro; KITAZAWA, Minori; OTA, Mari; TOYOFUKU, Chiharu; YAMADA, Masashi; THAMMAKARN, Chanathip; SHOHAM, Dany; TAKEHARA, Kazuaki
The capacity of slightly acidic hypochlorous acid water (SAHW), in both liquid and spray form, to inactivate bacteria was evaluated as a potential candidate for biosecurity enhancement in poultry production. SAHW (containing 50 or 100 ppm chlorine, pH 6) was able to inactivate Escherichia coli and Salmonella Infantis in liquid to below detectable levels (≤2.6 log10 CFU/ml) within 5 sec of exposure. In addition, SAHW antibacterial capacity was evaluated by spraying it using a nebulizer into a box containing these bacteria, which were present on the surfaces of glass plates and rayon sheets. SAHW was able to inactivate both bacterial species on the glass plates (dry condition) and rayon sheets within 5 min spraying and 5 min contact times, with the exception of 50 ppm SAHW on the rayon sheets. Furthermore, a corrosivity test determined that SAHW does not corrode metallic objects, even at the longest exposure times (83 days). Our findings demonstrate that SAHW is a good candidate for biosecurity enhancement in the poultry industry. Spraying it on the surfaces of objects, eggshells, egg incubators and transport cages could reduce the chances of contamination and disease transmission. These results augment previous findings demonstrating the competence of SAHW as an anti-viral disinfectant. PMID:27052464
Lactic acid bacteria (LAB) frequently contaminate commercial fuel ethanol fermentations, reducing yields and decreasing profitability of biofuel production. Microorganisms from environmental sources in different geographic regions of Thailand were tested for antibacterial activity against LAB. Fou...
Raghavendra, Ponnala; Halami, Prakash M
This study was undertaken to screen and select potent phytate degrading lactic acid bacteria and to evaluate their additional characteristic features. Forty lactic acid bacterial strains were isolated from different sources and screened for their ability to degrade myo-inositol hexaphosphate or IP(6) by cobalt chloride staining (plate assay) method, using calcium or sodium salt of phytic acid as substrate. All the forty isolates were able to degrade calcium phytate. However, only two Pediococcus pentosaceus strains (CFR R38 and CFR R35) were found to degrade sodium phytate. These strains showed phytase activity of 213 and 89 U at 50 degrees C, respectively and poor acid phosphatase activity. These strains were further evaluated for additional characteristic features. At pH 2, P. pentosaceus strains CFR R38 and CFR R35 showed 50.7 and 48.5 percentage survivability after 2 h of incubation respectively and they could also withstand 0.3% ox-bile. These cultures exhibited 54.6 and 44.8% of hydrophobicity to xylene, antibacterial activity against food borne pathogens and possessed beta-galactosidase activity. The resistance pattern to several antibiotics was also analyzed. The present study indicates that these strains, having phytate degrading ability and other characteristic features can be exploited as starter cultures in fermented foods to improve the mineral bioavailability.
Alvarado, C; García Almendárez, B E; Martin, S E; Regalado, C
This work was conducted to identify indigenous LAB capable of antimicrobial activity, present in traditional Mexican-foods with potential as natural preservatives. A total of 27 artisan unlabeled Mexican products were evaluated, from which 94 LAB strains were isolated, and only 25 strains showed antimicrobial activity against at least one pathogen indicator microorganism. Most of the inhibitory activity showed by the isolated LAB strains was attributed to pH reduction by organic acids. Lactobacillus and Lactococcus strains were good acid producers, depending on the substrate, and may enhance the safety of food products. Cell free cultures of Leuconostoc mesenteroides CH210, and PT8 (from chorizo and pulque, respectively) reduced the number of viable cells of enteropathogenic E. coli in broth system. Lb. plantarum CC10 (from "madre" of vinegar) showed significant inhibitory effect against S. aureus 8943. E. faecium QPII (from panela cheese) produced a bacteriocin with wide anti-L. monocytogenes activity. Selected LAB from traditional Mexican foods showed good potential as bio-preservatives.
Broadbent, J R; Brighton, C; McMahon, D J; Farkye, N Y; Johnson, M E; Steele, J L
Flavor development in low-fat Cheddar cheese is typified by delayed or muted evolution of desirable flavor and aroma, and a propensity to acquire undesirable meaty-brothy or burnt-brothy off-flavor notes early in ripening. The biochemical basis for these flavor deficiencies is unclear, but flavor production in bacterial-ripened cheese is known to rely on microorganisms and enzymes present in the cheese matrix. Lipid removal fundamentally alters cheese composition, which can modify the cheese microenvironment in ways that may affect growth and enzymatic activity of starter or nonstarter lactic acid bacteria (NSLAB). Additionally, manufacture of low-fat cheeses often involves changes to processing protocols that may substantially alter cheese redox potential, salt-in-moisture content, acid content, water activity, or pH. However, the consequences of these changes on microbial ecology and metabolism remain obscure. The objective of this study was to investigate the influence of fat content on population dynamics of starter bacteria and NSLAB over 9 mo of aging. Duplicate vats of full fat, 50% reduced-fat, and low-fat (containing <6% fat) Cheddar cheeses were manufactured at 3 different locations with a single-strain Lactococcus lactis starter culture using standardized procedures. Cheeses were ripened at 8°C and sampled periodically for microbiological attributes. Microbiological counts indicated that initial populations of nonstarter bacteria were much lower in full-fat compared with low-fat cheeses made at all 3 sites, and starter viability also declined at a more rapid rate during ripening in full-fat compared with 50% reduced-fat and low-fat cheeses. Denaturing gradient gel electrophoresis of cheese bacteria showed that the NSLAB fraction of all cheeses was dominated by Lactobacillus curvatus, but a few other species of bacteria were sporadically detected. Thus, changes in fat level were correlated with populations of different bacteria, but did not appear to
Rodríguez, J M; Cintas, L M; Casaus, P; Horn, N; Dodd, H M; Hernández, P E; Gasson, M J
A total of 4608 lactic acid bacteria (LAB) were isolated from 24 Spanish fermented sausages and screened for bacteriocin production. Two strains, BB24 and G18, produced bacteriocins that inhibited a broad spectrum of Gram-positive bacteria. BB24 and G18 were tentatively identified as Lactococcus lactis by carbohydrate fermentation patterns and other biochemical characteristics. The characterization of their bacteriocins suggested that both could be the well-known lantibiotic nisin. This was confirmed by PCR analysis of their genomic DNA. Nucleotide sequencing revealed that they produced nisin A. The fact that BB24 and G18 were isolated from sausages produced in two different regions of Spain suggests that nisin-producing L. lactis strains may be more widespread in meat products than previously thought. Nisin produced by L. lactis BB24 has been purified to homogeneity by a procedure that included ammonium sulphate precipitation and cation-exchange, hydrophobic-interaction and reverse-phase chromatography. The purification procedure was simple, rapid and reproducible.
Kalschne, Daneysa Lahis; Womer, Rute; Mattana, Ademir; Sarmento, Cleonice Mendes Pereira; Colla, Luciane Maria; Colla, Eliane
The lactic acid bacteria are involved with food fermentation and in such cases with food spoilage. Considering the need to reduce the lactic acid bacteria growth in meat products, the aim of this work was to enumerated and investigated the lactic acid bacteria present on sliced vacuum-packed cooked ham stored at 4 °C and 8 °C for 45 days by phenotypic and molecular techniques. The quantification showed that the lactic acid bacteria were present from the first day with mean count of 1.98 log cfu/g for the four batches analyzed. The lactic acid bacteria grew rapidly on the samples, and plate counts around 7.59 log cfu/g and 8.25 log cfu/g were detected after 45 days of storage at 4 °C and 8 °C, respectively; storage temperatures studied showed significant influence on the microorganism in study growth. The predominant lactic acid bacteria associated with the spoilage samples at one day of storage includes Lactobacillus sp., the phenotypic overlap Leuconostoc / Weissella sp. and Enterococcus sp. At 45 days of storage at 4 and 8 °C the mainly specie was Lactobacillus curvatus , following by Lactobacillus sakei and Leuconostoc mesentereoides ; the Enterococcus sp. was not present in the samples.
Kalschne, Daneysa Lahis; Womer, Rute; Mattana, Ademir; Sarmento, Cleonice Mendes Pereira; Colla, Luciane Maria; Colla, Eliane
The lactic acid bacteria are involved with food fermentation and in such cases with food spoilage. Considering the need to reduce the lactic acid bacteria growth in meat products, the aim of this work was to enumerated and investigated the lactic acid bacteria present on sliced vacuum-packed cooked ham stored at 4 °C and 8 °C for 45 days by phenotypic and molecular techniques. The quantification showed that the lactic acid bacteria were present from the first day with mean count of 1.98 log cfu/g for the four batches analyzed. The lactic acid bacteria grew rapidly on the samples, and plate counts around 7.59 log cfu/g and 8.25 log cfu/g were detected after 45 days of storage at 4 °C and 8 °C, respectively; storage temperatures studied showed significant influence on the microorganism in study growth. The predominant lactic acid bacteria associated with the spoilage samples at one day of storage includes Lactobacillus sp., the phenotypic overlap Leuconostoc / Weissella sp. and Enterococcus sp. At 45 days of storage at 4 and 8 °C the mainly specie was Lactobacillus curvatus , following by Lactobacillus sakei and Leuconostoc mesentereoides ; the Enterococcus sp. was not present in the samples. PMID:26221105
Milanowski, Maciej; Pomastowski, Paweł; Railean-Plugaru, Viorica; Rafińska, Katarzyna; Ligor, Tomasz; Buszewski, Bogusław
The current work deals with the phenomenon of silver cations uptake by two kinds of bacteria isolated from dairy products. The mechanism of sorption of silver cations by Lactococcus lactis and Lactobacillus casei bacteria was investigated. Inductively coupled plasma–mass spectrometry (ICP-MS) was used for determination of silver concentration sorbed by bacteria. Analysis of charge distribution was conducted by diffraction light scattering method. Changes in the ultrastructure of Lactococcus lactis and Lactobacillus casei cells after treatment with silver cations were investigated using transmission electron microscopy observation. Molecular spectroscopy methods, namely Fourier transform-infrared spectroscopy (FT-IR) and matrix-assisted laser desorption/ionization mass spectrometry (MALDI MS) were employed for description of the sorption mechanism. Moreover, an analysis of volatile organic compounds (VOCs) extracted from bacterial cells was performed. PMID:28362838
Berstad, Arnold; Raa, Jan; Midtvedt, Tore; Valeur, Jørgen
It is tempting to look at bacteria from our human egocentric point of view and label them as either ‘good’ or ‘bad’. However, a microbial society has its own system of government – ‘microcracy’ – and its own rules of play. Lactic acid bacteria are often referred to as representatives of the good ones, and there is little doubt that those belonging to the normal intestinal flora are beneficial for human health. But we should stop thinking of lactic acid bacteria as always being ‘friendly’ – they may instead behave like fledgling cuckoos. PMID:27235098
Pudlik, Agata M.
Transamination is the first step in the conversion of amino acids into aroma compounds by lactic acid bacteria (LAB) used in food fermentations. The process is limited by the availability of α-ketoglutarate, which is the best α-keto donor for transaminases in LAB. Here, uptake of α-ketoglutarate by the citrate transporter CitP is reported. Cells of Lactococcus lactis IL1403 expressing CitP showed significant levels of transamination activity in the presence of α-ketoglutarate and one of the amino acids Ile, Leu, Val, Phe, or Met, while the same cells lacking CitP showed transamination activity only after permeabilization of the cell membrane. Moreover, the transamination activity of the cells followed the levels of CitP in a controlled expression system. The involvement of CitP in the uptake of the α-keto donor was further demonstrated by the increased consumption rate in the presence of l-lactate, which drives CitP in the fast exchange mode of transport. Transamination is the only active pathway for the conversion of α-ketoglutarate in IL1403; a stoichiometric conversion to glutamate and the corresponding α-keto acid from the amino acids was observed. The transamination activity by both the cells and the cytoplasmic fraction showed a remarkably flat pH profile over the range from pH 5 to pH 8, especially with the branched-chain amino acids. Further metabolism of the produced α-keto acids into α-hydroxy acids and other flavor compounds required the coupling of transamination to glycolysis. The results suggest a much broader role of the citrate transporter CitP in LAB than citrate uptake in the citrate fermentation pathway alone. PMID:23204417
Gullo, Maria; Giudici, Paolo
This review focuses on acetic acid bacteria in traditional balsamic vinegar process. Although several studies are available on acetic acid bacteria ecology, metabolism and nutritional requirements, their activity as well as their technological traits in homemade vinegars as traditional balsamic vinegar is not well known. The basic technology to oxidise cooked grape must to produce traditional balsamic vinegar is performed by the so called "seed-vinegar" that is a microbiologically undefined starter culture obtained from spontaneous acetification of previous raw material. Selected starter cultures are the main technological improvement in order to innovate traditional balsamic vinegar production but until now they are rarely applied. To develop acetic acid bacteria starter cultures, selection criteria have to take in account composition of raw material, acetic acid bacteria metabolic activities, applied technology and desired characteristics of the final product. For traditional balsamic vinegar, significative phenotypical traits of acetic acid bacteria have been highlighted. Basic traits are: ethanol preferred and efficient oxidation, fast rate of acetic acid production, tolerance to high concentration of acetic acid, no overoxidation and low pH resistance. Specific traits are tolerance to high sugar concentration and to a wide temperature range. Gluconacetobacter europaeus and Acetobacter malorum strains can be evaluated to develop selected starter cultures since they show one or more suitable characters.
Riaz Rajoka, Muhammad Shahid; Shi, Junling; Zhu, Jing; Shao, Dongyan; Huang, Qingsheng; Yang, Hui; Jin, Mingliang
Lactic acid bacteria are associated with the human gastrointestinal tract. They are important for maintaining the balance of microflora in the human gut. An increasing number of published research reports in recent years have denoted the importance of producing interferon-gamma and IgA for treatment of disease. These agents can enhance the specific and nonspecific immune systems that are dependent on specific bacterial strains. The mechanisms of these effects were revealed in this investigation, where the cell walls of these bacteria were modulated by the cytokine pathways, while the whole bacterial cell mediated the host cell immune system and regulated the production of tumor necrosis factors and interleukins. A supplement of highly active lactic acid bacteria strains provided significant potential to enhance host's immunity, offering prevention from many diseases including some cancers. This review summarizes the current understanding of the function of lactic acid bacteria immunity enhancement and cancer prevention.
Hashim, Nurul Huda; Sani, Norrakiah Abdullah
Lactic acid bacteria are also known as beneficial microorganisms abundantly found in fermented food products. In this study, lactic acid bacteria were isolated from fresh cut fruits obtained from local markets. Throughout the isolation process from 11 samples of fruits, 225 presumptive lactic acid bacteria were isolated on MRS agar medium. After catalase and oxidase tests, 149 resulted to fit the characteristics of lactic acid bacteria. Further identification using Gram staining was conducted to identify the Gram positive bacteria. After this confirmation, the fermentation characteristics of these isolates were identified. It was found that 87 (58.4%) isolates were heterofermentative, while the rest of 62 (41.6%) are homofermentative lactic acid bacteria. Later, all these isolates were investigated for the ability to inhibit growth of Staphylococcus aureus using agar spot assay method. Seven (4.7%) isolates showed strong antagonistic capacity, while 127 (85.2%) and 8 (5.4%) isolates have medium and weak antagonistic capacity, respectively. The other 7 (4.7%) isolates indicated to have no antagonistic effect on S. aureus. Results support the potential of LAB isolated in this study which showed strong antagonistic activity against S. aureus may be manipulated to become protective cultures in food products. While the homofermentative or heterofermentative LAB can be utilized in fermentation of food and non-food products depending on the by-products required during the fermentation.
Costantini, Antonella; Pietroniro, Roberta; Doria, Francesca; Pessione, Enrica; Garcia-Moruno, Emilia
The aim of this work was to study the production of biogenic amines and particularly putrescine in lactic acid bacteria (LAB) related to wine and cider. We applied an analytical protocol that involves the use of PCR and TLC techniques to determine the production of putrescine from different precursors. Moreover, we also studied the ability of the Lactobacillus and Pediococcus tested to produce histamine and tyramine. The results showed that the majority of the Lactobacillus brevis analyzed harbour both AgDI and tdc genes and are tyramine and putrescine producers. Conversely, among the other LAB tested, only one Lactobacillus hilgardii and one Pediococcus pentosaceus produced putrescine. The AgDI gene was also detected in two other LAB (Lactobacillus mali and Pediococcus parvulus), but no putrescine production was observed. Finally, hdc gene and histamine production were found in strains (L. hilgardii 5211, isolated from wine, and Lactobacillus casei 18, isolated from cider) that were not putrescine producers.
Wu, Jia Jia; Ma, Ying Kun; Zhang, Fen Fen; Chen, Fu Sheng
Shanxi aged vinegar is a famous traditional Chinese vinegar made from several kinds of cereal by spontaneous solid-state fermentation techniques. In order to get a comprehensive understanding of culturable microorganism's diversity present in its fermentation, the indigenous microorganisms including 47 yeast isolates, 28 lactic acid bacteria isolates and 58 acetic acid bacteria isolates were recovered in different fermenting time and characterized based on a combination of phenotypic and genotypic approaches including inter-delta/PCR, PCR-RFLP, ERIC/PCR analysis, as well as 16S rRNA and 26S rRNA partial gene sequencing. In the alcoholic fermentation, the dominant yeast species Saccharomyces (S.) cerevisiae (96%) exhibited low phenotypic and genotypic diversity among the isolates, while Lactobacillus (Lb.) fermentum together with Lb. plantarum, Lb. buchneri, Lb. casei, Pediococcus (P.) acidilactici, P. pentosaceus and Weissella confusa were predominated in the bacterial population at the same stage. Acetobacter (A.) pasteurianus showing great variety both in genotypic and phenotypic tests was the dominant species (76%) in the acetic acid fermentation stage, while the other acetic acid bacteria species including A. senegalensis, A. indonesiensis, A. malorum and A. orientalis, as well as Gluconobacter (G.) oxydans were detected at initial point of alcoholic and acetic acid fermentation stage respectively.
Achilleos, Christine; Berthier, Françoise
The first objective of this work was to develop real-time quantitative PCR (qPCR) assays to quantify two species of mesophilic lactic acid bacteria technologically active in food fermentation, including cheese making: Lactococcus lactis and Lactobacillus paracasei. The second objective was to compare qPCR and plate counts of these two species in cheese samples. Newly designed primers efficiently amplified a region of the tuf gene from the target species. Sixty-three DNA samples from twenty different bacterial species, phylogenetically related or commonly found in raw milk and dairy products, were selected as positive and negative controls. Target DNA was successfully amplified showing a single peak on the amplicon melting curve; non-target DNA was not amplified. Quantification was linear over 5 log units (R(2) > 0.990), down to 22 gene copies/μL per well for Lc. lactis and 73 gene copies/μL per well for Lb. paracasei. qPCR efficiency ranged from 82.9% to 93.7% for Lc. lactis and from 81.1% to 99.5% for Lb. paracasei. At two stages of growth, Lc. lactis was quantified in 12 soft cheeses and Lb. paracasei in 24 hard cooked cheeses. qPCR proved to be useful for quantifying Lc. lactis, but not Lb. paracasei.
The winemaking process includes two main steps: lactic acid bacteria are responsible for the malolactic fermentation which follows the alcoholic fermentation by yeasts. Both types of microorganisms are present on grapes and on cellar equipment. Yeasts are better adapted to growth in grape must than lactic acid bacteria, so the alcoholic fermentation starts quickly. In must, up to ten lactic acid bacteria species can be identified. They belong to the Lactobacillus, Pediococcus, Leuconostoc and Oenococcus genera. Throughout alcoholic fermentation, a natural selection occurs and finally the dominant species is O. oeni, due to interactions between yeasts and bacteria and between bacteria themselves. After bacterial growth, when the population is over 10(6) CFU/ml, malolactic transformation is the obvious change in wine composition. However, many other substrates can be metabolized. Some like remaining sugars and citric acid are always assimilated by lactic acid bacteria, thus providing them with energy and carbon. Other substrates such as some amino acids may be used following pathways restricted to strains carrying the adequate enzymes. Some strains can also produce exopolysaccharides. All these transformations greatly influence the sensory and hygienic quality of wine. Malic acid transformation is encouraged because it induces deacidification. Diacetyl produced from citric acid is also helpful to some extent. Sensory analyses show that many other reactions change the aromas and make malolactic fermentation beneficial, but they are as yet unknown. On the contrary, an excess of acetic acid, the synthesis of glucane, biogenic amines and precursors of ethylcarbamate are undesirable. Fortunately, lactic acid bacteria normally multiply in dry wines; moreover some of these activities are not widespread. Moreover, the most striking trait of wine lactic acid bacteria is their capacity to adapt to a hostile environment. The mechanisms for this are not yet completely elucidated
Without the addition of preservative compounds cucumbers acidified with 150 mM acetic acid with pH adjusted to 3.5 typically undergo fermentation by lactic acid bacteria. Fumaric acid (20 mM) inhibited growth of Lactobacillus plantarum and the lactic acid bacteria present on fresh cucumbers, but sp...
Bezerra, Taliana Kênia Alves; de Araujo, Ana Rita Ribeiro; do Nascimento, Edilza Santos; de Matos Paz, José Eduardo; Gadelha, Carlos Alberto; Gadelha, Tatiane Santi; Pacheco, Maria Teresa Bertoldo; do Egypto Queiroga, Rita de Cássia Ramos; de Oliveira, Maria Elieidy Gomes; Madruga, Marta Suely
This study aimed to analyse the proteolytic effects of adding isolated and combined probiotic strains to goat "coalho" cheese. The cheeses were: QS - with culture Start, composed by Lactococcus lactis subsp. lactis and L. lactis subsp. cremoris (R704); QLA - with Lactobacillus acidophilus (LA-5); QLP - with Lactobacillus paracasei subsp. paracasei (L. casei 01); QB - with Bifidobacterium animalis subsp. lactis (BB 12); and QC, co-culture with the three probiotic microorganisms. The cheeses were analysed during 28 days of storage at 10°C. The probiotic cell count was higher than 6.5 and 7 log colony-forming units (CFU) g(-1) of cheese at the 1st and 28th days of storage, respectively. The addition of co-culture influenced (p<0.01) proteolysis in the cheese and resulted in a higher content of soluble protein and release of amino acids at the 1st day after processing. However, over all 28 days, the cheese supplemented with Bifidobacterium lactis in its isolated form showed the highest proteolytic activity, particularly in the hydrolysis of the alpha-s2 and kappa-casein fractions.
Valerio, Francesca; Favilla, Mara; De Bellis, Palmira; Sisto, Angelo; de Candia, Silvia; Lavermicocca, Paola
Thirty samples of Italian durum wheat semolina and whole durum wheat semolina, generally used for the production of Southern Italy's traditional breads, were subjected to microbiological analysis in order to explore their lactic acid bacteria (LAB) diversity and to find strains with antifungal activity. A total of 125 presumptive LAB isolates (Gram-positive and catalase-negative) were characterized by repetitive extragenic palindromic-PCR (REP-PCR) and sequence analysis of the 16S rRNA gene, leading to the identification of the following species: Weissella confusa, Weissella cibaria, Leuconostoc citreum, Leuconostoc mesenteroides, Lactococcus lactis, Lactobacillus rossiae and Lactobacillus plantarum. The REP-PCR results delineated 17 different patterns whose cluster analysis clearly differentiated W. cibaria from W. confusa isolates. Seventeen strains, each characterized by a different REP-PCR pattern, were screened for their antifungal properties. They were grown in a flour-based medium, comparable to a real food system, and the resulting fermentation products (FPs) were tested against fungal species generally contaminating bakery products, Aspergillus niger, Penicillium roqueforti and Endomyces fibuliger. The results of the study indicated a strong inhibitory activity - comparable to that obtained with the common preservative calcium propionate (0.3% w/v) - of ten LAB strains against the most widespread contaminant of bakery products, P. roqueforti. The screening also highlighted the unexplored antifungal activity of L. citreum, L. rossiae and W. cibaria (1 strain), which inhibited all fungal strains to the same or a higher extent compared with calcium propionate. The fermentation products of these three strains were characterized by low pH values, and a high content of lactic and acetic acids.
Kobierecka, Patrycja A.; Olech, Barbara; Książek, Monika; Derlatka, Katarzyna; Adamska, Iwona; Majewski, Paweł M.; Jagusztyn-Krynicka, Elżbieta K.; Wyszyńska, Agnieszka K.
Campylobacter jejuni is the most frequent cause of human food-borne gastroenteritis and chicken meat is the main source of infection. Recent studies showed that broiler chicken immunization against Campylobacter should be the most efficient way to lower the number of human infections by this pathogen. Induction of the mucosal immune system after oral antigen administration should provide protective immunity to chickens. In this work we tested the usefulness of Lactococcus lactis, the most extensively studied lactic acid bacterium, as a delivery vector for Campylobacter antigens. First we constructed hybrid protein – CjaA antigen presenting CjaD peptide epitopes on its surface. We showed that specific rabbit anti-rCjaAD serum reacted strongly with both CjaA and CjaD produced by a wild type C. jejuni strain. Next, rCjaAD and CjaA were fused to the C-terminus of the L. lactis YndF containing the LPTXG motif. The genes expressing these proteins were transcribed under control of the L. lactis Usp45 promoter and their products contain the Usp45 signal sequences. This strategy ensures a cell surface location of both analyzed proteins, which was confirmed by immunofluorescence assay. In order to evaluate the impact of antigen location on vaccine prototype efficacy, a L. lactis strain producing cytoplasm-located rCjaAD was also generated. Animal experiments showed a decrease of Campylobacter cecal load in vaccinated birds as compared with the control group and showed that the L. lactis harboring the surface-exposed rCjaAD antigen afforded greater protection than the L. lactis producing cytoplasm-located rCjaAD. To the best of our knowledge, this is the first attempt to employ Lactic Acid Bacteria (LAB) strains as a mucosal delivery vehicle for chicken immunization. Although the observed reduction of chicken colonization by Campylobacter resulting from vaccination was rather moderate, the experiments showed that LAB strains can be considered as an alternative vector to
Romi, Wahengbam; Ahmed, Giasuddin; Jeyaram, Kumaraswamy
Microbial community structure and population dynamics during spontaneous bamboo shoot fermentation for production of 'soidon' (indigenous fermented food) in North-east India were studied using cultivation-dependent and cultivation-independent molecular approaches. Cultivation-dependent analyses (PCR-amplified ribosomal DNA restriction analysis and rRNA gene sequencing) and cultivation-independent analyses (PCR-DGGE, qPCR and Illumina amplicon sequencing) were conducted on the time series samples collected from three independent indigenous soidon fermentation batches. The current findings revealed three-phase succession of autochthonous lactic acid bacteria to attain a stable ecosystem within 7 days natural fermentation of bamboo shoots. Weissella spp. (Weissella cibaria, uncultured Weissella ghanensis) and Lactococcus lactis subsp. cremoris predominated the early phase (1-2 days) which was joined by Leuconostoc citreum during the mid-phase (3 days), while Lactobacillus brevis and Lactobacillus plantarum emerged and became dominant in the late phase (5-7 days) with concurrent disappearance of W. cibaria and L. lactis subsp. cremoris. Lactococcus lactis subsp. lactis and uncultured Lactobacillus acetotolerans were predominantly present throughout the fermentation with no visible dynamics. The above identified dominant bacterial species along with their dynamics can be effectively utilized for designing a starter culture for industrialization of soidon production. Our results showed that a more realistic view on the microbial ecology of soidon fermentation could be obtained by cultivation-dependent studies complemented with cultivation-independent molecular approaches. Moreover, the critical issues to be considered for reducing methodological biases while studying the microbial ecology of traditional food fermentation were also highlighted with this soidon fermentation model.
Breidt, Frederick; Fleming, Henry P.
Current mathematical models used by food microbiologists do not address the issue of competitive growth in mixed cultures of bacteria. We developed a mathematical model which consists of a system of nonlinear differential equations describing the growth of competing bacterial cell cultures. In this model, bacterial cell growth is limited by the accumulation of protonated lactic acid and decreasing pH. In our experimental system, pure and mixed cultures of Lactococcus lactis and Listeria monocytogenes were grown in a vegetable broth medium. Predictions of the model indicate that pH is the primary factor that limits the growth of L. monocytogenes in competition with a strain of L. lactis which does not produce the bacteriocin nisin. The model also predicts the values of parameters that affect the growth and death of the competing populations. Further development of this model will incorporate the effects of additional inhibitors, such as bacteriocins, and may aid in the selection of lactic acid bacterium cultures for use in competitive inhibition of pathogens in minimally processed foods. PMID:9726854
Goto, Tsuyoshi; Kim, Young-Il; Furuzono, Tomoya; Takahashi, Nobuyuki; Yamakuni, Kanae; Yang, Ha-Eun; Li, Yongjia; Ohue, Ryuji; Nomura, Wataru; Sugawara, Tatsuya; Yu, Rina; Kitamura, Nahoko; Park, Si-Bum; Kishino, Shigenobu; Ogawa, Jun; Kawada, Teruo
Our previous study has shown that gut lactic acid bacteria generate various kinds of fatty acids from polyunsaturated fatty acids such as linoleic acid (LA). In this study, we investigated the effects of LA and LA-derived fatty acids on the activation of peroxisome proliferator-activated receptors (PPARs) which regulate whole-body energy metabolism. None of the fatty acids activated PPARδ, whereas almost all activated PPARα in luciferase assays. Two fatty acids potently activated PPARγ, a master regulator of adipocyte differentiation, with 10-oxo-12(Z)-octadecenoic acid (KetoA) having the most potency. In 3T3-L1 cells, KetoA induced adipocyte differentiation via the activation of PPARγ, and increased adiponectin production and insulin-stimulated glucose uptake. These findings suggest that fatty acids, including KetoA, generated in gut by lactic acid bacteria may be involved in the regulation of host energy metabolism.
Le Boucher, Clémentine; Gagnaire, Valérie; Briard-Bion, Valérie; Jardin, Julien; Maillard, Marie-Bernadette; Dervilly-Pinel, Gaud; Le Bizec, Bruno; Lortal, Sylvie; Jeanson, Sophie
In cheese, lactic acid bacteria are immobilized at the coagulation step and grow as colonies. The spatial distribution of bacterial colonies is characterized by the size and number of colonies for a given bacterial population within cheese. Our objective was to demonstrate that different spatial distributions, which lead to differences in the exchange surface between the colonies and the cheese matrix, can influence the ripening process. The strategy was to generate cheeses with the same growth and acidification of a Lactococcus lactis strain with two different spatial distributions, big and small colonies, to monitor the production of the major ripening metabolites, including sugars, organic acids, peptides, free amino acids, and volatile metabolites, over 1 month of ripening. The monitored metabolites were qualitatively the same for both cheeses, but many of them were more abundant in the small-colony cheeses than in the big-colony cheeses over 1 month of ripening. Therefore, the results obtained showed that two different spatial distributions of L. lactis modulated the ripening time course by generating moderate but significant differences in the rates of production or consumption for many of the metabolites commonly monitored throughout ripening. The present work further explores the immobilization of bacteria as colonies within cheese and highlights the consequences of this immobilization on cheese ripening. PMID:26497453
Sun, Mengjun; Zhou, Zichao; Dong, Jiachen; Zhang, Jichun; Xia, Yiru; Shu, Rong
Docosahexaenoic acid (DHA) and eicosapentaenoic acid (EPA) are two major omega-3 polyunsaturated fatty acids (n-3 PUFAs) with antimicrobial properties. In this study, we evaluated the potential antibacterial and antibiofilm activities of DHA and EPA against two periodontal pathogens, Porphyromonas gingivalis (P. gingivalis) and Fusobacterium nucleatum (F. nucleatum). MTT assay showed that DHA and EPA still exhibited no cytotoxicity to human oral tissue cells when the concentration came to 100 μM and 200 μM, respectively. Against P. gingivalis, DHA and EPA showed the same minimum inhibitory concentration (MIC) of 12.5 μM, and a respective minimum bactericidal concentration (MBC) of 12.5 μM and 25 μM. However, the MIC and MBC values of DHA or EPA against F. nucleatum were both greater than 100 μM. For early-stage bacteria, DHA or EPA displayed complete inhibition on the planktonic growth and biofilm formation of P. gingivalis from the lowest concentration of 12.5 μM. And the planktonic growth of F. nucleatum was slightly but not completely inhibited by DHA or EPA even at the concentration of 100 μM, however, the biofilm formation of F. nucleatum at 24 h was significantly restrained by 100 μM EPA. For exponential-phase bacteria, 100 μM DHA or EPA completely killed P. gingivalis and significantly decreased the viable counts of F. nucleatum. Meanwhile, the morphology of P. gingivalis was apparently damaged, and the virulence factor gene expression of P. gingivalis and F. nucleatum was strongly downregulated. Besides, the viability and the thickness of mature P. gingivalis biofilm, together with the viability of mature F. nucleatum biofilm were both significantly decreased in the presence of 100 μM DHA or EPA. In conclusion, DHA and EPA possessed antibacterial activities against planktonic and biofilm forms of periodontal pathogens, which suggested that DHA and EPA might be potentially supplementary therapeutic agents for prevention
Horisaki, Tadafumi; Yoshida, Eiko; Sumiya, Kaori; Takemura, Tetsuo; Yamane, Hisakazu; Nojiri, Hideaki
Five Burkholderia strains (CL-1, CL-2, CL-3, CL-4, and CL-5) capable of degrading monochloroacetic acid (MCA) were isolated from activated sludge or soil samples gathered from several parts of Japan. All five isolates were able to grow on MCA as the sole source of carbon and energy, and argentometry and gas chromatography-mass spectroscopy analyses showed that these five strains consumed MCA completely and released chloride ions stoichiometrically within 25 h. The five isolates also grew on monobromoacetic acid, monoiodoacetic acid, and L-2-monochloropropionic acid as sole sources of carbon and energy. In addition, the five isolates could not grow with DCA but dehalogenate single chlorine from DCA. Because PCR analyses revealed that all five isolates have an identical group II dehalogenase gene fragment and no group I deh gene, only strain CL-1 was analyzed further. The partial amino acid sequence of the group II dehalogenase of strain CL-1, named DehCL1, showed 74.6% and 65.2% identities to corresponding regions of the two MCA dehalogenases, DehCI from Pseudomonas sp. strain CBS-3 and Hdl IVa from Burkholderia cepacia strain MBA4, respectively. The secondary-structure motifs of the haloacid dehalogenase (HAD) superfamily and the amino acid residues involved in substrate binding, catalysis, and hydrophobic pocket formation were conserved in the partial amino acid sequence of DehCL1.
Lactoferricin is a 25 amino acid antimicrobial peptide domain that is liberated by pepsin digestion of lactoferrin in bovine milk. Along with its antibacterial properties, lactoferricin has also been reported to have immunostimulatory, antiviral, and anticarcinogenic effects. There is substantial ...
This invention relates to nucleic acid probes and a method for the rapid detection of Shigella and enteroinvasive Escherichia coli, the causative agents of bacterial dysentery, by use of a nucleic acid hybridization probe, equivalent to a plasmid DNA region encoding one of 4 specific invasion-associated, peptides of all strains of Shigella and enterinvasive E . coli , in a nucleic acid hybridization reaction with a clinical specimen containing dysentery bacteria.
Laroute, Valérie; Yasaro, Chonthicha; Narin, Waranya; Mazzoli, Roberto; Pessione, Enrica; Cocaign-Bousquet, Muriel; Loubière, Pascal
Lactococcus lactis NCDO 2118 was previously selected for its ability to decarboxylate glutamate to γ-aminobutyric acid (GABA), an interesting nutritional supplement able to improve mood and relaxation. Amino acid decarboxylation is generally considered as among the biochemical systems allowing lactic acid bacteria to counteracting acidic stress and obtaining metabolic energy. These strategies also include arginine deiminase pathway and malolactic fermentation but little is known about their possible interactions of with GABA production. In the present study, the effects of glutamate, arginine, and malate (i.e., the substrates of these acid-resistance pathways) on L. lactis NCDO 2118 growth and GABA production performances were analyzed. Both malate and arginine supplementation resulted in an efficient reduction of acidity and improvement of bacterial biomass compared to glutamate supplementation. Glutamate decarboxylation was limited to narrow environmental conditions (pH < 5.1) and physiological state (stationary phase). However, some conditions were able to improve GABA production or activate glutamate decarboxylation system even outside of this compass. Arginine clearly stimulated glutamate decarboxylation: the highest GABA production (8.6 mM) was observed in cultures supplemented with both arginine and glutamate. The simultaneous addition of arginine, malate, and glutamate enabled earlier GABA production (i.e., during exponential growth) at relatively high pH (6.5). As far as we know, no previous study has reported GABA production in such conditions. Although further studies are needed to understand the molecular basis of these phenomena, these results represent important keys suitable of application in GABA production processes. PMID:27458444
Every biotechnology process that relies on the use of bacteria to make a product or to overproduce a molecule may, at some time, struggle with the presence of virulent phages. For example, phages are the primary cause of fermentation failure in the milk transformation industry. This review focuses on the recent scientific advances in the field of lactic acid bacteria phage research. Three specific topics, namely, the sources of contamination, the detection methods and the control procedures will be discussed. PMID:21995802
Filannino, Pasquale; Gobbetti, Marco; De Angelis, Maria
The metabolism of hydroxycinnamic acids by strictly heterofermentative lactic acid bacteria (19 strains) was investigated as a potential alternative energy route. Lactobacillus curvatus PE5 was the most tolerant to hydroxycinnamic acids, followed by strains of Weissella spp., Lactobacillus brevis, Lactobacillus fermentum, and Leuconostoc mesenteroides, for which the MIC values were the same. The highest sensitivity was found for Lactobacillus rossiae strains. During growth in MRS broth, lactic acid bacteria reduced caffeic, p-coumaric, and ferulic acids into dihydrocaffeic, phloretic, and dihydroferulic acids, respectively, or decarboxylated hydroxycinnamic acids into the corresponding vinyl derivatives and then reduced the latter compounds to ethyl compounds. Reductase activities mainly emerged, and the activities of selected strains were further investigated in chemically defined basal medium (CDM) under anaerobic conditions. The end products of carbon metabolism were quantified, as were the levels of intracellular ATP and the NAD+/NADH ratio. Electron and carbon balances and theoretical ATP/glucose yields were also estimated. When CDM was supplemented with hydroxycinnamic acids, the synthesis of ethanol decreased and the concentration of acetic acid increased. The levels of these metabolites reflected on the alcohol dehydrogenase and acetate kinase activities. Overall, some biochemical traits distinguished the common metabolism of strictly heterofermentative strains: main reductase activity toward hydroxycinnamic acids, a shift from alcohol dehydrogenase to acetate kinase activities, an increase in the NAD+/NADH ratio, and the accumulation of supplementary intracellular ATP. Taken together, the above-described metabolic responses suggest that strictly heterofermentative lactic acid bacteria mainly use hydroxycinnamic acids as external acceptors of electrons. PMID:25261518
Filannino, Pasquale; Gobbetti, Marco; De Angelis, Maria; Di Cagno, Raffaella
The metabolism of hydroxycinnamic acids by strictly heterofermentative lactic acid bacteria (19 strains) was investigated as a potential alternative energy route. Lactobacillus curvatus PE5 was the most tolerant to hydroxycinnamic acids, followed by strains of Weissella spp., Lactobacillus brevis, Lactobacillus fermentum, and Leuconostoc mesenteroides, for which the MIC values were the same. The highest sensitivity was found for Lactobacillus rossiae strains. During growth in MRS broth, lactic acid bacteria reduced caffeic, p-coumaric, and ferulic acids into dihydrocaffeic, phloretic, and dihydroferulic acids, respectively, or decarboxylated hydroxycinnamic acids into the corresponding vinyl derivatives and then reduced the latter compounds to ethyl compounds. Reductase activities mainly emerged, and the activities of selected strains were further investigated in chemically defined basal medium (CDM) under anaerobic conditions. The end products of carbon metabolism were quantified, as were the levels of intracellular ATP and the NAD(+)/NADH ratio. Electron and carbon balances and theoretical ATP/glucose yields were also estimated. When CDM was supplemented with hydroxycinnamic acids, the synthesis of ethanol decreased and the concentration of acetic acid increased. The levels of these metabolites reflected on the alcohol dehydrogenase and acetate kinase activities. Overall, some biochemical traits distinguished the common metabolism of strictly heterofermentative strains: main reductase activity toward hydroxycinnamic acids, a shift from alcohol dehydrogenase to acetate kinase activities, an increase in the NAD(+)/NADH ratio, and the accumulation of supplementary intracellular ATP. Taken together, the above-described metabolic responses suggest that strictly heterofermentative lactic acid bacteria mainly use hydroxycinnamic acids as external acceptors of electrons.
García-Hernández, Yaneisy; Pérez-Sánchez, Tania; Boucourt, Ramón; Balcázar, José L; Nicoli, Jacques R; Moreira-Silva, João; Rodríguez, Zoraya; Fuertes, Héctor; Nuñez, Odalys; Albelo, Nereyda; Halaihel, Nabil
In livestock production, lactic acid bacteria (LAB) are the most common microorganisms used as probiotics. For such use, these bacteria must be correctly identified and characterized to ensure their safety and efficiency. In the present study, LAB were isolated from broiler excreta, where a fermentation process was used. Nine among sixteen isolates were identified by biochemical and molecular (sequencing of the 16S rRNA gene) methods as Lactobacillus crispatus (n=1), Lactobacillus pentosus (n=1), Weissella cibaria (n=1), Pediococcus pentosaceus (n=2) and Enterococcus hirae (n=4). Subsequently, these bacteria were characterized for their growth capabilities, lactic acid production, acidic pH and bile salts tolerance, cell surface hydrophobicity, antimicrobial susceptibility and antagonistic activity. Lactobacillus pentosus strain LB-31, which showed the best characteristics, was selected for further analysis. This strain was administered to broilers and showed the ability of modulating the immune response and producing beneficial effects on morpho-physiological, productive and health indicators of the animals.
Antimicrobial susceptibilities and random amplified polymorphic DNA-PCR fingerprint characterization of Lactococcus lactis ssp. lactis and Lactococcus garvieae isolated from bovine intramammary infections.
Plumed-Ferrer, C; Barberio, A; Franklin-Guild, R; Werner, B; McDonough, P; Bennett, J; Gioia, G; Rota, N; Welcome, F; Nydam, D V; Moroni, P
In total, 181 streptococci-like bacteria isolated from intramammary infections (IMI) were submitted by a veterinary clinic to Quality Milk Production Services (QMPS, Cornell University, Ithaca, NY). The isolates were characterized by sequence analysis, and 46 Lactococcus lactis ssp. lactis and 47 Lactococcus garvieae were tested for susceptibility to 17 antibiotics. No resistant strains were found for β-lactam antibiotics widely used in clinical practice (penicillin, ampicillin, and amoxicillin), and all minimum inhibitory concentrations (MIC) were far from the resistance breakpoints. Eight strains had MIC intermediate to cefazolin. The random amplification of polymorphic DNA (RAPD)-PCR fingerprint patterns showed a slightly higher heterogeneity for Lc. lactis ssp. lactis isolates than for Lc. garvieae isolates.
Desiere, Frank; Lucchini, Sacha; Canchaya, Carlos; Ventura, Marco; Brüssow, Harald
Comparative phage genomics has become possible due to the availability of more than 100 complete phage genome sequences and the development of powerful bioinformatics tools. This technology, profiting from classical molecular-biology knowledge, has opened avenues of research for topics, which were difficult to address in the past. Now, it is possible to retrace part of the evolutionary history of phage modules by comparative genomics. The diagnosis of relatedness is hereby not uniquely based on sequence similarity alone, but includes topological considerations of genome organization. Detailed transcription maps have allowed in silico predictions of genome organization to be verified and refined. This comparative knowledge is providing the basis for a new taxonomic classification concept for bacteriophages infecting low G + C-content Gram-positive bacteria based on the genetic organization of the structural gene module. An Sfi21-like and an Sfi11-like genus of Siphoviridae is proposed. The gene maps of many phages show remarkable synteny in their structural genes defining a lambda super-group within Siphoviridae. A hierarchy of relatedness within the lambda super-group suggests elements of vertical evolution in Siphoviridae. Tailed phages are the result of both vertical and horizontal evolution and are thus fascinating objects for the study of molecular evolution. Prophage sequences integrated into the genomes of their bacterial host present theoretical challenges for evolutionary biologists. Prophages represent up to 10% of the genome in some LAB. In pathogenic streptococci prophages confer genes of selective value for the lysogenic cell. The lysogenic conversion genes are located between the lysin gene and the right phage attachment site. Non-attributed genes were found at the same genome position of prophages from lactic streptococci. These genes belong to the few prophage genes transcribed in the lysogen. Prophages from dairy bacteria might therefore also
Visintin, Simonetta; Alessandria, Valentina; Valente, Antonio; Dolci, Paola; Cocolin, Luca
Yeast, lactic acid bacteria (LAB) and acetic acid bacteria (AAB) populations, isolated from cocoa bean heap and box fermentations in West Africa, have been investigated. The fermentation dynamicswere determined by viable counts, and 106 yeasts, 105 LAB and 82 AAB isolateswere identified by means of rep-PCR grouping and sequencing of the rRNA genes. During the box fermentations, the most abundant species were Saccharomyces cerevisiae, Candida ethanolica, Lactobacillus fermentum, Lactobacillus plantarum, Acetobacter pasteurianus and Acetobacter syzygii, while S. cerevisiae, Schizosaccharomyces pombe, Hanseniaspora guilliermondii, Pichia manshurica, C. ethanolica, Hanseniaspora uvarum, Lb. fermentum, Lb. plantarum, A. pasteurianus and Acetobacter lovaniensis were identified in the heap fermentations. Furthermore, the most abundant species were molecularly characterized by analyzing the rep-PCR profiles. Strains grouped according to the type of fermentations and their progression during the transformation process were also highlighted. The yeast, LAB and AAB isolates were physiologically characterized to determine their ability to grow at different temperatures, as well as at different pH, and ethanol concentrations, tolerance to osmotic stress, and lactic acid and acetic acid inhibition. Temperatures of 45 °C, a pH of 2.5 to 3.5, 12% (v/v) ethanol and high concentrations of lactic and acetic acid have a significant influence on the growth of yeasts, LAB and AAB. Finally, the yeastswere screened for enzymatic activity, and the S. cerevisiae, H. guilliermondii, H. uvarumand C. ethanolica species were shown to possess several enzymes that may impact the quality of the final product.
Fan, Xuetong; Wagner, Karen; Sokorai, Kimberly J B; Ngo, Helen
Novel phenolic branched-chain fatty acids (PBC-FAs) were evaluated for their antimicrobial properties against both gram-positive ( Listeria innocua , Bacillus subtilis , Enterococcus faecium ) and gram-negative ( Escherichia coli , Salmonella Typhimurium, and Pseudomonas tolaasii ) bacteria. In addition, PBC-FA derivatives, such as PBC-FA methyl ester mixture, methyl-branched fatty acid mixtures, and trimethylsilyl-PBC-FA methyl esters, were synthesized to study the structure activity relationship. Results showed that PBC-FAs were a potent antimicrobial against gram-positive bacteria with MICs of 1.8 to 3.6 μg/ml. The compounds were less effective against gram-negative bacteria. Derivatives of PBC-FAs and an equimolar mixture of oleic acid and phenol all had MICs above 233 μg/ml against both gram-positive and gram-negative bacteria. Comparison of antimicrobial activities of the PBC-FAs with those of the derivatives suggests that the carboxylic group in the fatty acid moiety and the hydroxyl group on the phenol moiety were responsible for the antimicrobial efficacy. Growth curves of L. innocua revealed that PBC-FAs prevented bacterial growth, while MBC-FAs only delayed the onset of rapid growth of L. innocua . Our results demonstrated that the novel PBC-FAs have potential for use as antimicrobials against gram-positive bacteria.
Kai, Xia; Xinle, Liang; Yudong, Li
The clustered regularly interspaced short palindromic repeat (CRISPR) is a widespread adaptive immunity system that exists in most archaea and many bacteria against foreign DNA, such as phages, viruses and plasmids. In general, CRISPR system consists of direct repeat, leader, spacer and CRISPR-associated sequences. Acetic acid bacteria (AAB) play an important role in industrial fermentation of vinegar and bioelectrochemistry. To investigate the polymorphism and evolution pattern of CRISPR loci in acetic acid bacteria, bioinformatic analyses were performed on 48 species from three main genera (Acetobacter, Gluconacetobacter and Gluconobacter) with whole genome sequences available from the NCBI database. The results showed that the CRISPR system existed in 32 species of the 48 strains studied. Most of the CRISPR-Cas system in AAB belonged to type I CRISPR-Cas system (subtype E and C), but type II CRISPR-Cas system which contain cas9 gene was only found in the genus Acetobacter and Gluconacetobacter. The repeat sequences of some CRISPR were highly conserved among species from different genera, and the leader sequences of some CRISPR possessed conservative motif, which was associated with regulated promoters. Moreover, phylogenetic analysis of cas1 demonstrated that they were suitable for classification of species. The conservation of cas1 genes was associated with that of repeat sequences among different strains, suggesting they were subjected to similar functional constraints. Moreover, the number of spacer was positively correlated with the number of prophages and insertion sequences, indicating the acetic acid bacteria were continually invaded by new foreign DNA. The comparative analysis of CRISR loci in acetic acid bacteria provided the basis for investigating the molecular mechanism of different acetic acid tolerance and genome stability in acetic acid bacteria.
Mansour, S.; Bailly, J.; Landaud, S.; Monnet, C.; Sarthou, A. S.; Cocaign-Bousquet, M.; Leroy, S.; Irlinger, F.; Bonnarme, P.
The interactions that may occur between microorganisms in different ecosystems have not been adequately studied yet. We investigated yeast-bacterium interactions in a synthetic medium using different culture associations involving the yeast Yarrowia lipolytica 1E07 and two bacteria, Staphylococcus xylosus C2a and Lactococcus lactis LD61. The growth and biochemical characteristics of each microorganism in the different culture associations were studied. The expression of genes related to glucose, lactate, and amino acid catabolism was analyzed by reverse transcription followed by quantitative PCR. Our results show that the growth of Y. lipolytica 1E07 is dramatically reduced by the presence of S. xylosus C2a. As a result of a low amino acid concentration in the medium, the expression of Y. lipolytica genes involved in amino acid catabolism was downregulated in the presence of S. xylosus C2a, even when L. lactis was present in the culture. Furthermore, the production of lactate by both bacteria had an impact on the lactate dehydrogenase gene expression of the yeast, which increased up to 30-fold in the three-species culture compared to the Y. lipolytica 1E07 pure culture. S. xylosus C2a growth dramatically decreased in the presence of Y. lipolytica 1E07. The growth of lactic acid bacteria was not affected by the presence of S. xylosus C2a or Y. lipolytica 1E07, although the study of gene expression showed significant variations. PMID:19684166
Goto, Tsuyoshi; Kim, Young-Il; Furuzono, Tomoya; Takahashi, Nobuyuki; Yamakuni, Kanae; Yang, Ha-Eun; Li, Yongjia; Ohue, Ryuji; Nomura, Wataru; Sugawara, Tatsuya; Yu, Rina; Kitamura, Nahoko; and others
Our previous study has shown that gut lactic acid bacteria generate various kinds of fatty acids from polyunsaturated fatty acids such as linoleic acid (LA). In this study, we investigated the effects of LA and LA-derived fatty acids on the activation of peroxisome proliferator-activated receptors (PPARs) which regulate whole-body energy metabolism. None of the fatty acids activated PPARδ, whereas almost all activated PPARα in luciferase assays. Two fatty acids potently activated PPARγ, a master regulator of adipocyte differentiation, with 10-oxo-12(Z)-octadecenoic acid (KetoA) having the most potency. In 3T3-L1 cells, KetoA induced adipocyte differentiation via the activation of PPARγ, and increased adiponectin production and insulin-stimulated glucose uptake. These findings suggest that fatty acids, including KetoA, generated in gut by lactic acid bacteria may be involved in the regulation of host energy metabolism. - Highlights: • Most LA-derived fatty acids from gut lactic acid bacteria potently activated PPARα. • Among tested fatty acids, KetoA and KetoC significantly activated PPARγ. • KetoA induced adipocyte differentiation via the activation of PPARγ. • KetoA enhanced adiponectin production and glucose uptake during adipogenesis.
Cui, Yanhua; Hu, Tong; Qu, Xiaojun; Zhang, Lanwei; Ding, Zhongqing; Dong, Aijun
Plasmids are widely distributed in different sources of lactic acid bacteria (LAB) as self-replicating extrachromosomal genetic materials, and have received considerable attention due to their close relationship with many important functions as well as some industrially relevant characteristics of the LAB species. They are interesting with regard to the development of food-grade cloning vectors. This review summarizes new developments in the area of lactic acid bacteria plasmids and aims to provide up to date information that can be used in related future research.
Torija, María Jesús; García-Parrilla, María del Carmen; Troncoso, Ana María
The production of vinegar depends on an oxidation process that is mainly performed by acetic acid bacteria. Despite the different methods of vinegar production (more or less designated as either “fast” or “traditional”), the use of pure starter cultures remains far from being a reality. Uncontrolled mixed cultures are normally used, but this review proposes the use of controlled mixed cultures. The acetic acid bacteria species determine the quality of vinegar, although the final quality is a combined result of technological process, wood contact, and aging. This discussion centers on wine vinegar and evaluates the effects of these different processes on its chemical and sensory properties. PMID:24574887
Cui, Yanhua; Hu, Tong; Qu, Xiaojun; Zhang, Lanwei; Ding, Zhongqing; Dong, Aijun
Plasmids are widely distributed in different sources of lactic acid bacteria (LAB) as self-replicating extrachromosomal genetic materials, and have received considerable attention due to their close relationship with many important functions as well as some industrially relevant characteristics of the LAB species. They are interesting with regard to the development of food-grade cloning vectors. This review summarizes new developments in the area of lactic acid bacteria plasmids and aims to provide up to date information that can be used in related future research. PMID:26068451
Mas, Albert; Torija, María Jesús; García-Parrilla, María del Carmen; Troncoso, Ana María
The production of vinegar depends on an oxidation process that is mainly performed by acetic acid bacteria. Despite the different methods of vinegar production (more or less designated as either "fast" or "traditional"), the use of pure starter cultures remains far from being a reality. Uncontrolled mixed cultures are normally used, but this review proposes the use of controlled mixed cultures. The acetic acid bacteria species determine the quality of vinegar, although the final quality is a combined result of technological process, wood contact, and aging. This discussion centers on wine vinegar and evaluates the effects of these different processes on its chemical and sensory properties.
Maeda, M; Shibata, A; Biswas, G; Korenaga, H; Kono, T; Itami, T; Sakai, M
Fifty-one lactic acid bacteria (LAB) strains were isolated and identified based on 16S ribosomal DNA sequence from the intestinal tracts of 142 kuruma shrimps (Marsupenaeus japonicus) collected from Kanmon Strait, Fukuoka and Tachibana Bay, Nagasaki, Japan. Cellular immunomodulatory function of 51 isolated LAB strains was assessed by measuring the level of interferon (IFN)-γ induction in mouse spleen cell culture. The strain Lactococcus lactis D1813 exhibited the highest amount of IFN-γ production and also bactericidal activity and was selected for testing its immunomodulatory role as a probiotic in kuruma shrimp. We also assessed the effect of dietary incorporation of this probiotic on resistance to Vibrio penaeicida infection in the kuruma shrimp. Our results demonstrate that probiotic L. lactis D1813-containing diet-fed (10⁵ cfu g⁻¹) shrimps displayed a significant up-regulation of lysozyme gene expressions in the intestine and hepatopancreas. However, insignificantly higher expression of anti-lipopolysaccharide factor, super oxide dismutase, prophenoloxidase, and toll-like receptor 1 was recorded in the intestine of shrimps fed the probiotic diet. Moreover, significantly increased (P < 0.01) resistance to the bacterial pathogen in term of better post-infection survival (61.7 %) was observed in the shrimps fed with the probiotic-incorporated diet compared with the control diet-fed group (28.3 %). The present study indicates the immunomodulatory role of the LAB L. lactis D1813 on the kuruma shrimp immune system and supports its potential use as an effective probiotic in shrimp aquaculture.
Guarcello, Rosa; De Angelis, Maria; Settanni, Luca; Formiglio, Sabino; Gaglio, Raimondo; Minervini, Fabio; Moschetti, Giancarlo; Gobbetti, Marco
Accumulation of biogenic amines (BAs) in cheese and other foods is a matter of public health concern. The aim of this study was to identify the enzyme activities responsible for BA degradation in lactic acid bacteria which were previously isolated from traditional Sicilian and Apulian cheeses. The selected strains would control the concentration of BAs during cheese manufacture. First, 431 isolates not showing genes encoding the decarboxylases responsible for BA formation were selected using PCR-based methods. Ninety-four out of the 431 isolates degraded BAs (2-phenylethylamine, cadaverine, histamine, putrescine, spermine, spermidine, tyramine, or tryptamine) during cultivation on chemically defined medium. As shown by random amplification of polymorphic DNA-PCR and partial sequencing of the 16S rRNA gene, 78 of the 94 strains were Lactobacillus species (Lactobacillus casei, Lb. fermentum, Lb. parabuchneri, Lb. paracasei, Lb. paraplantarum, and Lb. rhamnosus), Leuconostoc species (Leuconostoc lactis and Ln. mesenteroides), Pediococcus pentosaceus, Lactococcus lactis, Streptococcus species (Streptococcus gallolyticus and S. thermophilus), Enterococcus lactis, and Weissella paramesenteroides A multicopper oxidase-hydrolyzing BA was purified from the most active strain, Lb. paracasei subsp. paracasei CB9CT. The gene encoding the multicopper oxidase was sequenced and was also detected in other amine-degrading strains of Lb. fermentum, Lb. paraplantarum, and P. pentosaceus Lb. paracasei subsp. paracasei CB9CT and another strain (CACIO6CT) of the same species that was able to degrade all the BAs were singly used as adjunct starters for decreasing the concentration of histamine and tyramine in industrial Caciocavallo cheese. The results of this study disclose a feasible strategy for increasing the safety of traditional cheeses while maintaining their typical sensorial traits.
Sánchez, Jorge; Borrero, Juan; Gómez-Sala, Beatriz; Basanta, Antonio; Herranz, Carmen; Cintas, Luis M.; Hernández, Pablo E.
Hiracin JM79 (HirJM79), a Sec-dependent bacteriocin produced by Enterococcus hirae DCH5, was cloned and produced in Lactococcus lactis, Lactobacillus sakei, Enterococcus faecium, Enterococcus faecalis, and Pichia pastoris. For heterologous production of HirJM79 in lactic acid bacteria (LAB), the HirJM79 structural gene (hirJM79), with or without the HirJM79 immunity gene (hiriJM79), was cloned into the plasmid pMG36c under the control of the constitutive promoter P32 and into the plasmid pNZ8048 under the control of the inducible PNisA promoter. For the production of HirJM79 in P. pastoris, the gene encoding the mature HirJM79 protein was cloned into the pPICZαA expression vector. The recombinant plasmids permitted the production of biologically active HirJM79 in the supernatants of L. lactis IL1403, L. lactis NZ9000, L. sakei Lb790, E. faecalis JH2-2, and P. pastoris X-33, the coproduction of HirJM79 and nisin A in L. lactis DPC5598, and the coproduction of HirJM79 and enterocin P in E. faecium L50/14-2. All recombinant LAB produced larger quantities of HirJM79 than E. hirae DCH5, although the antimicrobial activities of most transformants were lower than that predicted from their production of HirJM79. The synthesis, processing, and secretion of HirJM79 proceed efficiently in recombinant LAB strains and P. pastoris. PMID:18310424
Vido, Karin; Diemer, Hélène; Van Dorsselaer, Alain; Leize, Emmanuelle; Juillard, Vincent; Gruss, Alexandra; Gaudu, Philippe
Thiol-disulfide bond balance is generally maintained in bacteria by thioredoxin reductase-thioredoxin and/or glutathione-glutaredoxin systems. Some gram-positive bacteria, including Lactococcus lactis, do not produce glutathione, and the thioredoxin system is presumed to be essential. We constructed an L. lactis trxB1 mutant. The mutant was obtained under anaerobic conditions in the presence of dithiothreitol (DTT). Unexpectedly, the trxB1 mutant was viable without DTT and under aerated static conditions, thus disproving the essentiality of this system. Aerobic growth of the trxB1 mutant did not require glutathione, also ruling out the need for this redox maintenance system. Proteomic analyses showed that known oxidative stress defense proteins are induced in the trxB1 mutant. Two additional effects of trxB1 were not previously reported in other bacteria: (i) induction of proteins involved in fatty acid or menaquinone biosynthesis, indicating that membrane synthesis is part of the cellular response to a redox imbalance, and (ii) alteration of the isoforms of the glycolytic enzyme glyceraldehyde-3-phosphate dehydrogenase (GapB). We determined that the two GapB isoforms in L. lactis differed by the oxidation state of catalytic-site cysteine C152. Unexpectedly, a decrease specific to the oxidized, inactive form was observed in the trxB1 mutant, possibly because of proteolysis of oxidized GapB. This study showed that thioredoxin reductase is not essential in L. lactis and that its inactivation triggers induction of several mechanisms acting at the membrane and metabolic levels. The existence of a novel redox function that compensates for trxB1 deficiency is suggested. PMID:15629931
Szczepankowska, Agnieszka K; Szatraj, Katarzyna; Sałański, Przemysław; Rózga, Agnieszka; Górecki, Roman K; Bardowski, Jacek K
Lactic acid bacteria (LAB) are Gram-positive, nonpathogenic microorganisms that are gaining much interest as antigen producers for development of live vaccine vectors. Heterologous proteins of different origin have been successfully expressed in various LAB species, including Lactococcus lactis. Recombinant L. lactis strains have been shown to induce specific local and systemic immune responses against various antigens. Our study aimed at constructing a L. lactis strain expressing haemagglutinin of a Polish avian H5H1 influenza isolate and examining its effect on animals. Expression of the cloned H5 gene was achieved using the nisin-controlled gene expression system. Detection of the intracellular H5 antigen produced in L. lactis was performed by Western blot analysis and confirmed using mass spectrometry. The potential of L. lactis recombinant cells to induce an immune response was examined by setting up preliminary immunization trials on chickens and mice. Obtained sera were tested for specific antibodies by ELISA assays. The results of these studies are a promising step toward developing a vaccine against the bird flu using Lactococcus lactis cells as bioreactors for efficient antigen production and delivery to the mucosal surface.
Szatraj, Katarzyna; Szczepankowska, Agnieszka K; Sączyńska, Violetta; Florys, Katarzyna; Gromadzka, Beata; Łepek, Krzysztof; Płucienniczak, Grażyna; Szewczyk, Bogusław; Zagórski-Ostoja, Włodzimierz; Bardowski, Jacek
Gram-positive and nonpathogenic lactic acid bacteria (LAB) are considered to be promising candidates for the development of new, safe systems of heterologous protein expression. Recombinant LAB has been shown to induce specific local and systemic immune response against selected pathogens, and could be a good alternative to classical attenuated carriers. The main goal of our study was to express the avian influenza haemagglutinin (H5) and chicken interleukin 2 (chIL-2) in Lactococcus lactis. Results of this study were anticipated to lead to construction of lactococcal strain(s) with potential vaccine properties against the avian influenza A (H5N1) virus. Expression of the cloned H5 gene, its His-tagged variant and chIL-2 gene, under the control of the ptcB gene promoter was attested by RT-PCR on transcriptional level and Western or dot blot analysis on translational level, demonstrating that system can be an attractive solution for production of heterologous proteins. The results of the preliminary animal trial conducted in mice are a promising step toward development of a vaccine against avian bird flu using Lactococcus lactis cells as antigen carriers.
Szatraj, Katarzyna; Sałański, Przemysław; Rózga, Agnieszka; Górecki, Roman K.; Bardowski, Jacek K.
Lactic acid bacteria (LAB) are Gram-positive, nonpathogenic microorganisms that are gaining much interest as antigen producers for development of live vaccine vectors. Heterologous proteins of different origin have been successfully expressed in various LAB species, including Lactococcus lactis. Recombinant L. lactis strains have been shown to induce specific local and systemic immune responses against various antigens. Our study aimed at constructing a L. lactis strain expressing haemagglutinin of a Polish avian H5H1 influenza isolate and examining its effect on animals. Expression of the cloned H5 gene was achieved using the nisin-controlled gene expression system. Detection of the intracellular H5 antigen produced in L. lactis was performed by Western blot analysis and confirmed using mass spectrometry. The potential of L. lactis recombinant cells to induce an immune response was examined by setting up preliminary immunization trials on chickens and mice. Obtained sera were tested for specific antibodies by ELISA assays. The results of these studies are a promising step toward developing a vaccine against the bird flu using Lactococcus lactis cells as bioreactors for efficient antigen production and delivery to the mucosal surface. PMID:28321412
Zadravec, Petra; Marečková, Lucie; Petroková, Hana; Hodnik, Vesna; Perišić Nanut, Milica; Anderluh, Gregor; Štrukelj, Borut; Malý, Petr; Berlec, Aleš
Infections with shiga toxin-producing bacteria, like enterohemorrhagic Escherichia coli and Shigella dysenteriae, represent a serious medical problem. No specific and effective treatment is available for patients with these infections, creating a need for the development of new therapies. Recombinant lactic acid bacterium Lactococcus lactis was engineered to bind Shiga toxin by displaying novel designed albumin binding domains (ABD) against Shiga toxin 1 B subunit (Stx1B) on their surface. Functional recombinant Stx1B was produced in Escherichia coli and used as a target for selection of 17 different ABD variants (named S1B) from the ABD scaffold-derived high-complex combinatorial library in combination with a five-round ribosome display. Two most promising S1Bs (S1B22 and S1B26) were characterized into more details by ELISA, surface plasmon resonance and microscale thermophoresis. Addition of S1Bs changed the subcellular distribution of Stx1B, completely eliminating it from Golgi apparatus most likely by interfering with its retrograde transport. All ABD variants were successfully displayed on the surface of L. lactis by fusing to the Usp45 secretion signal and to the peptidoglycan-binding C terminus of AcmA. Binding of Stx1B by engineered lactococcal cells was confirmed using flow cytometry and whole cell ELISA. Lactic acid bacteria prepared in this study are potentially useful for the removal of Shiga toxin from human intestine. PMID:27606705
Meadow, James F.; Liebert, Melissa A.; Cepon-Robins, Tara J.; Gildner, Theresa E.; Urlacher, Samuel S.; Bohannan, Brendan J.M.; Snodgrass, J. Josh; Sugiyama, Lawrence S.
Cassava beer, or chicha, is typically consumed daily by the indigenous Shuar people of the Ecuadorian Amazon. This traditional beverage made from cassava tuber (Manihot esculenta) is thought to improve nutritional quality and flavor while extending shelf life in a tropical climate. Bacteria responsible for chicha fermentation could be a source of microbes for the human microbiome, but little is known regarding the microbiology of chicha. We investigated bacterial community composition of chicha batches using Illumina high-throughput sequencing. Fermented chicha samples were collected from seven Shuar households in two neighboring villages in the Morona-Santiago region of Ecuador, and the composition of the bacterial communities within each chicha sample was determined by sequencing a region of the 16S ribosomal gene. Members of the genus Lactobacillus dominated all samples. Significantly greater phylogenetic similarity was observed among chicha samples taken within a village than those from different villages. Community composition varied among chicha samples, even those separated by short geographic distances, suggesting that ecological and/or evolutionary processes, including human-mediated factors, may be responsible for creating locally distinct ferments. Our results add to evidence from other fermentation systems suggesting that traditional fermentation may be a form of domestication, providing endemic beneficial inocula for consumers, but additional research is needed to identify the mechanisms and extent of microbial dispersal. PMID:25071997
Itoi, Shiro; Abe, Takeshi; Washio, Sayaka; Ikuno, Erika; Kanomata, Yuna; Sugita, Haruo
We isolated lactic acid bacteria from the intestinal tract of the pufferfish Takifugu niphobles caught in Shimoda, Shizuoka, Japan by using MRS broth prepared with 50% seawater. Additional screening was carried out using phenotypic tests such as Gram staining, cell morphology, catalase, oxidase and fermentation of glucose. Subsequently 227 isolates screened by the phenotypic tests were subjected to species-specific PCR for Lactococcus lactis, resulting in four positive isolates. The 16S rRNA gene sequences from three isolates were highly similar to that of L. lactis subsp. lactis (DNA database accession number M58837), while that of one isolate was identical to that of Leuconostoc mesenteroides (AB023246). These isolates were characterized by API 50 CH for carbohydrate fermentation and other phenotypic criteria for salt tolerance, and the characteristics were compared with those of L. lactis subsp. lactis from a cheese starter culture. The carbohydrate fermentation profiles of these isolates were characteristic of L. lactis subsp. lactis strains, whereas the tolerance of these isolates to salt was higher than that of L. lactis subsp. lactis from the cheese starter culture: the new L. lactis isolates showed high salt tolerance in MRS-agar plates containing 200% seawater or 6% sodium chloride. This is the first report of the isolation of halotolerant strains of L. lactis subsp. lactis from a marine environment.
Ladero, Victor; Rattray, Fergal P; Mayo, Baltasar; Martín, María Cruz; Fernández, María; Alvarez, Miguel A
Lactococcus lactis is a prokaryotic microorganism with great importance as a culture starter and has become the model species among the lactic acid bacteria. The long and safe history of use of L. lactis in dairy fermentations has resulted in the classification of this species as GRAS (General Regarded As Safe) or QPS (Qualified Presumption of Safety). However, our group has identified several strains of L. lactis subsp. lactis and L. lactis subsp. cremoris that are able to produce putrescine from agmatine via the agmatine deiminase (AGDI) pathway. Putrescine is a biogenic amine that confers undesirable flavor characteristics and may even have toxic effects. The AGDI cluster of L. lactis is composed of a putative regulatory gene, aguR, followed by the genes (aguB, aguD, aguA, and aguC) encoding the catabolic enzymes. These genes are transcribed as an operon that is induced in the presence of agmatine. In some strains, an insertion (IS) element interrupts the transcription of the cluster, which results in a non-putrescine-producing phenotype. Based on this knowledge, a PCR-based test was developed in order to differentiate nonproducing L. lactis strains from those with a functional AGDI cluster. The analysis of the AGDI cluster and their flanking regions revealed that the capacity to produce putrescine via the AGDI pathway could be a specific characteristic that was lost during the adaptation to the milk environment by a process of reductive genome evolution.
Ustiugova, E A; Timofeeva, A V; Stoianova, L G; Netrusov, A I; Katrukha, G S
The Lactococcus lactis subsp. lactis 194-K strain has been established to be able to produce two bacteriocins, one of which was identified as the known lantibiotic nisin A, and the other 194-D bacteriocin represents a polypeptide with a 2589-Da molecular mass and comprises 20 amino acid residues. Both bacteriocins were produced in varying proportions in all of the studied nutrient media, which support the growth of the producer. Depending on the cultivation medium, the nisin A content was 380- to 1123-fold lower in the 194-K stain culture fluid than that of the 194-D peptide. In comparision to to nisin A Bacteriocin 194-D possessed a wide range of antibacterial activity and suppressed the growth of both Gram-positive and Gram-negative bacteria. An optimal medium for 194-D bacteriocin synthesis was shown to be a fermentation medium which contained yeast extract, casein hydrolysate, and potassium phosphate. The biosynthesis ofbacteriocin 194-D by the 194-K strain in these media occurred parallel to producer growth, and its maximal accumulation in the culture fluid was observed at 14-20 h of the strain's growth.
Enan, Gamal; Abdel-Shafi, Seham; Ouda, Sahar; Negm, Sally
The purpose of this study was to select and characterize a probiotic bacterium with distinctive antimicrobial activities. In this respect, Lactococcus lactis subspecies lactis Z11 (L. lactis Z11) isolated from Zabady (Arabian yoghurt) inhibited other strains of lactic acid bacteria and some food-born pathogens including Listeria monocytogenes, Bacillus cereus and staphylococcus aureus. The inhibitory activity of cell free supernatant (CFS) of L. lactis Z11 isolated from zabady was lost by proteolytic enzymes, heat resistant. Consequently, the active substance(s) of CFS was characterized as a bacteriocin. This bacteriocin has been shown to consist of protein but has no lipidic or glucidic moieties in its active molecule. Its activity was stable in the pH range 2.0 to 7.0 and was not affected by organic solvents. The L. lactis Z11 bacteriocin was produced in CFS throughout the mide to the late exponential phase of growth of the producer organism and maximum bacteriocin production was obtained at initial pH 6.5 at incubation temperature of about 30°C. PMID:24151453
Burgain, J; Scher, J; Francius, G; Borges, F; Corgneau, M; Revol-Junelles, A M; Cailliez-Grimal, C; Gaiani, C
This review gives an overview of the importance of interactions occurring in dairy matrices between Lactic Acid Bacteria and milk components. Dairy products are important sources of biological active compounds of particular relevance to human health. These compounds include immunoglobulins, whey proteins and peptides, polar lipids, and lactic acid bacteria including probiotics. A better understanding of interactions between bioactive components and their delivery matrix may successfully improve their transport to their target site of action. Pioneering research on probiotic lactic acid bacteria has mainly focused on their host effects. However, very little is known about their interaction with dairy ingredients. Such knowledge could contribute to designing new and more efficient dairy food, and to better understand relationships between milk constituents. The purpose of this review is first to provide an overview of the current knowledge about the biomolecules produced on bacterial surface and the composition of the dairy matter. In order to understand how bacteria interact with dairy molecules, adhesion mechanisms are subsequently reviewed with a special focus on the environmental conditions affecting bacterial adhesion. Methods dedicated to investigate the bacterial surface and to decipher interactions between bacteria and abiotic dairy components are also detailed. Finally, relevant industrial implications of these interactions are presented and discussed.
Okada, S; Ishikawa, M; Yoshida, I; Uchimura, T; Ohara, N; Kozaki, M
Lactic acid bacteria in four samples of sour dough sponges were studied quantitatively and qualitatively. In each sponge, there were one or two species of the genus Lactobacillus: L. reuteri and L. curvatus in San Francisco sour dough sponge, L. brevis and L. hilgardii in panettone sour dough sponge produced in Italy, L. sanfrancisco from a rye sour dough sponge produced in Germany, and L. casei and L. curvatus from a rye sour dough sponge produced in Switzerland. For all isolates except the L. reuteri strains oleic acid, a component of the Tween 80 added to the medium, was essential for growth. It was of interest that lactobacilli requiring oleic acid were the predominant flora of lactic acid bacteria in the microbial environment of sour dough sponges.
Fernández de Palencia, Pilar; de la Plaza, Marta; Mohedano, M Luz; Martínez-Cuesta, M Carmen; Requena, Teresa; López, Paloma; Peláez, Carmen
The amino acid conversion to volatile compounds by lactic acid bacteria is important for aroma formation in cheese. In this work, we analyzed the effect of the lytic bacteriocin Lacticin 3147 on transamination of isoleucine and further formation of the volatile compound 2-methylbutanal in cheese. The Lacticin 3147 producing strain Lactococcus lactis IFPL3593 was fluorescently tagged (IFPL3593-GFP) by conjugative transfer of the plasmid pMV158GFP from Streptococcus pneumoniae, and used as starter in cheese manufacture. Starter adjuncts were the bacteriocin-sensitive strains L. lactis T1 and L. lactis IFPL730, showing branched chain amino acid aminotransferase and alpha-keto acid decarboxylase activity, respectively. Adjunct strains were selected to complete the isoleucine conversion pathway and, hence, increase formation of 2-methylbutanal conferring aroma to the cheese. The non-bacteriocin-producing strain L. lactis IFPL359-GFP was included as starter in the control batch. Fluorescent tagging of the starter strains allowed their tracing in cheese during ripening by fluorescence microscopy and confocal scanning laser microscopy. The bacteriocin produced by L. lactis IFPL3593-GFP enhanced lysis of the adjuncts with a concomitant increase in isoleucine transamination and about a two-fold increase of the derived volatile compound 2-methylbutanal. This led to an enhancement of the cheese aroma detected by a sensory panel. The improvement of cheese flavour and aroma may be of significant importance for the dairy industry.
Zadravec, Petra; Štrukelj, Borut; Berlec, Aleš
Lactic acid bacteria (LAB) are food-grade hosts for surface display with potential applications in food and therapy. Alternative approaches to surface display on LAB would avoid the use of recombinant DNA technology and genetically-modified organism (GMO)-related regulatory requirements. Non-covalent surface display of proteins can be achieved by fusing them to various cell-wall binding domains, of which the Lysine motif domain (LysM) is particularly well studied. Fusion proteins have been isolated from recombinant bacteria or from their growth medium and displayed on unmodified bacteria, enabling heterologous surface display. This was demonstrated on non-viable cells devoid of protein content, termed bacteria-like particles, and on various species of genus Lactobacillus. Of the latter, Lactobacillus salivarius ATCC 11741 was recently shown to be particularly amenable for LysM-mediated display. Possible regulatory implications of heterologous surface display are discussed, particularly those relevant for the European Union.
Zadravec, Petra; Štrukelj, Borut; Berlec, Aleš
Lactic acid bacteria (LAB) are food-grade hosts for surface display with potential applications in food and therapy. Alternative approaches to surface display on LAB would avoid the use of recombinant DNA technology and genetically-modified organism (GMO)-related regulatory requirements. Non-covalent surface display of proteins can be achieved by fusing them to various cell-wall binding domains, of which the Lysine motif domain (LysM) is particularly well studied. Fusion proteins have been isolated from recombinant bacteria or from their growth medium and displayed on unmodified bacteria, enabling heterologous surface display. This was demonstrated on non-viable cells devoid of protein content, termed bacteria-like particles, and on various species of genus Lactobacillus. Of the latter, Lactobacillus salivarius ATCC 11741 was recently shown to be particularly amenable for LysM-mediated display. Possible regulatory implications of heterologous surface display are discussed, particularly those relevant for the European Union. PMID:25880164
Gómez, Natacha C; Ramiro, Juan M P; Quecan, Beatriz X V; de Melo Franco, Bernadette D G
Use of probiotic biofilms can be an alternative approach for reducing the formation of pathogenic biofilms in food industries. The aims of this study were (i) to evaluate the probiotic properties of bacteriocinogenic (Lactococcus lactis VB69, L. lactis VB94, Lactobacillus sakei MBSa1, and Lactobacillus curvatus MBSa3) and non-bacteriocinogenic (L. lactis 368, Lactobacillus helveticus 354, Lactobacillus casei 40, and Weissela viridescens 113) lactic acid bacteria (LAB) isolated from Brazilian's foods and (ii) to develop protective biofilms with these strains and test them for exclusion of Listeria monocytogenes, Escherichia coli O157:H7, and Salmonella Typhimurium. LAB were tested for survival in acid and bile salt conditions, surface properties, biosurfactant production, β-galactosidase and gelatinase activity, antibiotic resistance and presence of virulence genes. Most strains survived exposure to pH 2 and 4% bile salts. The highest percentages of auto-aggregation were obtained after 24 h of incubation. Sixty-seven percentage auto-aggregation value was observed in W. viridescens 113 and Lactobacillus curvatus MBSa3 exhibited the highest co-aggregation (69% with Listeria monocytogenes and 74.6% with E. coli O157:H7), while the lowest co-aggregation was exhibited by W. viridescens 113 (53.4% with Listeria monocytogenes and 38% with E. coli O157:H7). Tests for hemolytic activity, bacterial cell adherence with xylene, and drop collapse confirmed the biosurfactant-producing ability of most strains. Only one strain (L. lactis 368) produced β-galactosidase. All strains were negative for virulence genes cob, ccf, cylLL, cylLs, cyllM, cylB, cylA and efaAfs and gelatinase production. The antibiotic susceptibility tests indicated that the MIC for ciprofloxacin, clindamycin, gentamicin, kanamycin, and streptomycin did not exceed the epidemiological cut-off suggested by the European Food Safety Authority. Some strains were resistant to one or more antibiotics and resistance
Gómez, Natacha C.; Ramiro, Juan M. P.; Quecan, Beatriz X. V.; de Melo Franco, Bernadette D. G.
Use of probiotic biofilms can be an alternative approach for reducing the formation of pathogenic biofilms in food industries. The aims of this study were (i) to evaluate the probiotic properties of bacteriocinogenic (Lactococcus lactis VB69, L. lactis VB94, Lactobacillus sakei MBSa1, and Lactobacillus curvatus MBSa3) and non-bacteriocinogenic (L. lactis 368, Lactobacillus helveticus 354, Lactobacillus casei 40, and Weissela viridescens 113) lactic acid bacteria (LAB) isolated from Brazilian’s foods and (ii) to develop protective biofilms with these strains and test them for exclusion of Listeria monocytogenes, Escherichia coli O157:H7, and Salmonella Typhimurium. LAB were tested for survival in acid and bile salt conditions, surface properties, biosurfactant production, β-galactosidase and gelatinase activity, antibiotic resistance and presence of virulence genes. Most strains survived exposure to pH 2 and 4% bile salts. The highest percentages of auto-aggregation were obtained after 24 h of incubation. Sixty-seven percentage auto-aggregation value was observed in W. viridescens 113 and Lactobacillus curvatus MBSa3 exhibited the highest co-aggregation (69% with Listeria monocytogenes and 74.6% with E. coli O157:H7), while the lowest co-aggregation was exhibited by W. viridescens 113 (53.4% with Listeria monocytogenes and 38% with E. coli O157:H7). Tests for hemolytic activity, bacterial cell adherence with xylene, and drop collapse confirmed the biosurfactant-producing ability of most strains. Only one strain (L. lactis 368) produced β-galactosidase. All strains were negative for virulence genes cob, ccf, cylLL, cylLs, cyllM, cylB, cylA and efaAfs and gelatinase production. The antibiotic susceptibility tests indicated that the MIC for ciprofloxacin, clindamycin, gentamicin, kanamycin, and streptomycin did not exceed the epidemiological cut-off suggested by the European Food Safety Authority. Some strains were resistant to one or more antibiotics and
Dugan, P.R.; Apel, W.A.
Studies have shown that the application of an aqueous solution of sodium lauryl sulphate and sodium benzoate to the surface of high-sulphur coal refuse inhibits the activity of iron- and sulphur-oxidising chemo-autotrophic bacteria and reduces the amount of acid drainage from the refuse. Further studies are recommended to assess the usefulness of this method for controlling formation of acid mine drainage in the field.
Dextrans and related glucans produced from sucrose by lactic acid bacteria have been studied for many years and are used in numerous commercial applications and products. Most of these glucans are water-soluble, except for a few notable exceptions from cariogenic Streptococcus spp. and a very small ...
Gobbetti, Marco; Minervini, Fabio; Pontonio, Erica; Di Cagno, Raffaella; De Angelis, Maria
The drivers for the establishment and composition of the sourdough microbiota, with particular emphasis on lactic acid bacteria, are reviewed and discussed. More than 60 different species of lactobacilli were identified from sourdoughs, showing the main overlapping between sourdough and human intestine ecosystems. The microbial kinetics during sourdough preparation was described by several studies using various methodological approaches, including culture-dependent and -independent (e.g., high throughput sequencing), and metabolite and meta-transcriptome analyses. Although the abundant microbial diversity harbored by flours, a succession of dominating and sub-dominating populations of lactic acid bacteria suddenly occurred during sourdough propagation, leading to the progressive assembly of the bacterial community. The contribution of all the potential sources (house microbiota, flour, types of flours and additional ingredients) for contaminating lactic acid bacteria was compared with the aim to find overlapping or specific routes that affect the sourdough microbiota. Once established and mature, pros and cons regarding the stability of the sourdough lactic acid bacteria biota were also reviewed, showing contradictory results, which were mainly dependent on the species/strains. Probably, the future research efforts should be dedicated to decrease the sources/drivers of noticeable variation rather than to full standardization of the process for sourdough preparation and use.
Geissler, Andreas J.; Vogel, Rudi F.
Seven strains of important beer-spoiling lactic acid bacteria were sequenced using single-molecule real-time sequencing. Complete genomes were obtained for strains of Lactobacillus paracollinoides, Lactobacillus lindneri, and Pediococcus claussenii. The analysis of these genomes emphasizes the role of plasmids as the genomic foundation of beer-spoiling ability. PMID:27795248
We examined 10 lactic acid bacteria that have been previously characterized for commercial use as probiotic cultures, mostly for dairy products, including 1 Pediococcus and 9 Lactobacilli. Our objectives were to develop a rapid procedure for determining the long-term survivability of these cultures ...
This report summarizes the results of the Mine Waste Technology Program (MWTP) Activity III, Project 3, In Situ Source Control of Acid Generation Using Sulfate-Reducing Bacteria, funded by the U.S. Environmental Protection Agency (EPA) and jointly administered by EPA and the U.S....
Natural and anthropogenic acidic environments are dominated by bacteria and Archaea. As many as 86 genera or species have been identified or isolated from pH <4.5 environments. This paper reviews the worldwide literature and provide tables of morphological characteristics, habitat information and a key for light microscope identification for the non-microbiologist.
Mirkovic, Nemanja; Radulovic, Zorica; Uzelac, Gordana; Lozo, Jelena; Obradovic, Dragojlo; Topisirovic, Ljubisa
Summary Lactococcus lactis ssp. lactis BGBM50, a producer of lactococcin G and aggregation-promoting factor, was isolated from selected lactic acid bacteria taken from semi-hard cheese traditionally produced in the village Žanjic, Montenegro. Strain BGBM50 harbours a number of plasmids of different sizes. Plasmid curing experiments showed that genes for bacteriocin production are located on pBM140, a plasmid 140 kb in length. PCR analysis with primers specific for lactococcin Q and G genes gave fragment of the expected size. In addition, after plasmid curing of strain BGBM50, different derivatives with altered phenotypes were obtained, among them BGBM50-34 strain, which retained bacteriocin synthesis but had enhanced aggregation ability. PMID:27904354
Lasta, Samar; Fajloun, Ziad; Darbon, Hervé; Mansuelle, Pascal; Andreotti, Nicolas; Sabatier, Jean-Marc; Boudabous, Abdellatif; Sampieri, François
Bacteriocin J46 is a 27-residue polypeptide produced by Lactococcus lactis subsp. cremoris J46 in fermented milk. The natural form of J46 (nJ46) exhibits a broad antimicrobial spectrum. Herein, we produced the synthetic form of J46 (sJ46) by solid-phase chemical synthesis. The biochemical and physico-chemical properties of sJ46, as well as its antimicrobial activity, were found to be identical to those of its natural counterpart nJ46. It showed a potent antimicrobial activity against both lactic acid bacteria and other Gram-positive microorganisms. (1)H-NMR conformational analysis of sJ46 indicates that it adopts a flexible random coil structure.
Yokaryo, Hiroto; Tokiwa, Yutaka
Lactic acid bacteria that grow under alkaline conditions (pH 10) were isolated from various sources in Okinawa (Japan). These alkali-tolerant and alkaliphilic bacteria were classified as follows: Microbacterium sp. (1 strain), Enterococcus spp. (9 strains), Alkalibacterium spp. (3 strains), Exiguobacterium spp. (5 strains), Oceanobacillus spp. (3 strains) and Bacillus spp. (7 strains) by 16S rRNA gene sequencing. By fermentation, many strains were able to convert glucose into mainly L-(+)-lactic acid of high optical purity in alkaline broth. This result indicated that valuable L-(+)-lactic acid-producing bacteria could be isolated efficiently by screening under alkaline conditions. Six strains were selected and their ability to produce lactic acid at different initial pH was compared. Enterococcus casseliflavus strain 79w3 gave the highest lactic acid concentration. Lactic acid concentration and productivity were 103 g L(-1) (optical purity of 99.5% as L-isomer) and 2.2 g L(-1) h(-1), respectively when 129 g L(-1) of glucose was used by batch fermentation.
Pholsen, Suradej; Khota, Waroon; Pang, Huili; Higgs, David; Cai, Yimin
Strains TH 14, TH 21 and TH 64 were isolated from tropical silages, namely corn stover, sugar cane top and rice straw, respectively, prepared in Thailand. These strains were selected by low pH growth range and high lactic acid-producing ability, similar to some commercial inoculants. Based on the analysis of 16S ribosomal RNA gene sequence and DNA-DNA relatedness, strain TH 14 was identified as Lactobacillus casei, and strains TH 21 and TH 64 were identified as L. plantarum. Strains TH 14, TH 21, TH 64 and two commercial inoculants, CH (L. plantarum) and SN (L. rhamnosus), were used as additives to fresh and wilted purple Guinea and sorghum silages prepared using a small-scale fermentation method. The number of epiphytic lactic acid bacteria (LAB) in the forages before ensilage was relatively low but the numbers of coliform and aerobic bacteria were higher. Sorghum silages at 30 days of fermentation were all well preserved with low pH (3.56) and high lactic acid production (72.86 g/kg dry matter). Purple Guinea silage inoculated with LAB exhibited reduced count levels of aerobic and coliform bacteria, lower pH, butyric acid and ammonia nitrogen and increased lactic acid concentration, compared with the control. Strain TH 14 more effectively improved lactic acid production compared with inoculants and other strains. © 2016 Japanese Society of Animal Science.
Fernández, Leonides; Beerthuyzen, Marke M.; Brown, Julie; Siezen, Roland J.; Coolbear, Tim; Holland, Ross; Kuipers, Oscar P.
The gene encoding the major intracellular tributyrin esterase of Lactococcus lactis was cloned using degenerate DNA probes based on 19 known N-terminal amino acid residues of the purified enzyme. The gene, named estA, was sequenced and found to encode a protein of 258 amino acid residues. The transcription start site was mapped 233 nucleotides upstream of the start codon, and a canonical promoter sequence was identified. The deduced amino acid sequence of the estA product contained the typical GXSXG motif found in most lipases and esterases. The protein was overproduced up to 170-fold in L. lactis by use of the nisin-controlled expression system recently developed for lactic acid bacteria. The estA gene was inactivated by chromosomal integration of a temperature-sensitive integration vector. This resulted in the complete loss of esterase activity, which could then be recovered after complementation of the constructed esterase-deficient strain with the wild-type estA gene. This confirms that EstA is the main enzyme responsible for esterase activity in L. lactis. Purified recombinant enzyme showed a preference for short-chain acyl esters, surprisingly also including phospholipids. Medium- and long-acyl-chain lipids were also hydrolyzed, albeit less efficiently. Intermediate characteristics between esterases and lipases make intracellular lactococcal EstA difficult to classify in either of these two groups of esterolytic enzymes. We suggest that, in vivo, EstA could be involved in (phospho)lipid metabolism or cellular detoxification or both, as its sequence showed significant similarity to S-formylglutathione hydrolase (FGH) of Paracoccus denitrificans and human EstD (or FGH), which are part of a universal formaldehyde detoxification pathway. PMID:10742212
Zhang, Gaosen; Sun, Henry J.
D-amino acids are toxic for life on Earth. Yet, they form constantly due to geochemical racemization and bacterial growth (the cell walls of which contain D-amino acids), raising the fundamental question of how they ultimately are recycled. This study provides evidence that bacteria use D-amino acids as a source of nitrogen by running enzymatic racemization in reverse. Consequently, when soils are inundated with racemic amino acids, resident bacteria consume D- as well as L-enantiomers, either simultaneously or sequentially depending on the level of their racemase activity. Bacteria thus protect life on Earth by keeping environments D-amino acid free. PMID:24647559
Zhang, Gaosen; Sun, Henry J
D-amino acids are toxic for life on Earth. Yet, they form constantly due to geochemical racemization and bacterial growth (the cell walls of which contain D-amino acids), raising the fundamental question of how they ultimately are recycled. This study provides evidence that bacteria use D-amino acids as a source of nitrogen by running enzymatic racemization in reverse. Consequently, when soils are inundated with racemic amino acids, resident bacteria consume D- as well as L-enantiomers, either simultaneously or sequentially depending on the level of their racemase activity. Bacteria thus protect life on Earth by keeping environments D-amino acid free.
Machielsen, Ronnie; Siezen, Roland J.; van Hijum, Sacha A. F. T.; van Hylckama Vlieg, Johan E. T.
A novel 51-kb conjugative transposon of Lactococcus lactis, designated Tn6098, encoding the capacity to utilize α-galactosides such as raffinose and stachyose, was identified and characterized. Alpha-galactosides are a dominant carbon source in many plant-derived foods. Most dairy lactococcus strains are unable to use α-galactosides as a growth substrate, yet many of these strains are known to have beneficial industrial traits. Conjugal transfer of Tn6098 was demonstrated from the plant-derived donor strain L. lactis KF147 to the recipient L. lactis NZ4501, a derivative of the dairy model strain L. lactis MG1363. The integration of Tn6098 into the genome of the recipient strain was confirmed by Illumina sequencing of the transconjugant L. lactis NIZO3921. The molecular structure of the integration site was confirmed by a PCR product spanning the insertion site. A 15-bp direct repeat sequence (TTATACCATAATTAC) is present on either side of Tn6098 in the chromosome of L. lactis KF147. One copy of this sequence is also present in the L. lactis MG1363 chromosome and represents the sole integration site. Phenotypic characterization of all strains showed that the transconjugant has not only acquired the ability to grow well in soy milk, a substrate rich in α-galactosides, but also has retained the flavor-forming capabilities of the recipient strain L. lactis MG1363. This study demonstrates how (induced) conjugation can be used to exploit the beneficial industrial traits of industrial dairy lactic acid bacteria in fermentation of plant-derived substrates. PMID:21115709
Fischer, Maren M; Egli, Ines M; Aeberli, Isabelle; Hurrell, Richard F; Meile, Leo
Ethiopian injera, a soft pancake, baked from fermented batter, is preferentially prepared from tef (Eragrostis tef) flour. The phytic acid (PA) content of tef is high and is only partly degraded during the fermentation step. PA chelates with iron and zinc in the human digestive tract and strongly inhibits their absorption. With the aim to formulate a starter culture that would substantially degrade PA during injera preparation, we assessed the potential of microorganisms isolated from Ethiopian household-tef fermentations to degrade PA. Lactic acid bacteria (LAB) were found to be among the dominating microorganisms. Seventy-six isolates from thirteen different tef fermentations were analyzed for phytase activity and thirteen different isolates of seven different species were detected to be positive in a phytase screening assay. In 20-mL model tef fermentations, out of these thirteen isolates, the use of Lactobacillus (L.) buchneri strain MF58 and Pediococcus pentosaceus strain MF35 resulted in lowest PA contents in the fermented tef of 41% and 42%, respectively of its initial content. In comparison 59% of PA remained when spontaneously fermented. Full scale tef fermentation (0.6L) and injera production using L. buchneri MF58 as culture additive decreased PA in cooked injera from 1.05 to 0.34±0.02 g/100 g, representing a degradation of 68% compared to 42% in injera from non-inoculated traditional fermentation. The visual appearance of the pancakes was similar. The final molar ratios of PA to iron of 4 and to zinc of 12 achieved with L. buchneri MF58 were decreased by ca. 50% compared to the traditional fermentation. In conclusion, selected LAB strains in tef fermentations can degrade PA, with L. buchneri MF58 displaying the highest PA degrading potential. The 68% PA degradation achieved by the application of L. buchneri MF58 would be expected to improve human zinc absorption from tef-injera, but further PA degradation is probably necessary if iron absorption has to
Zendo, Takeshi; Visessanguan, Wonnop; Roytrakul, Sittiruk; Pumpuang, Laphaslada; Jaresitthikunchai, Janthima; Sonomoto, Kenji
Lactococcus garvieae BCC 43578 produces a novel class II bacteriocin, garvieacin Q (GarQ), 70 amino acids in length and containing a 20-amino-acid N-terminal leader peptide. It is cleaved at the Gly-Gly site to generate the mature GarQ (5,339 Da), which is especially inhibitory against Listeria monocytogenes ATCC 19115 and other L. garvieae strains. PMID:22210221
Ma, Lili; Ji, Yurui; Bartlam, Mark; Wang, Yingying
Bacteria with high nucleic acid (HNA) and low nucleic acid (LNA) content are commonly observed in aquatic environments. To date, limited knowledge is available on their temporal and spatial variations in freshwater environments. Here an investigation of HNA and LNA bacterial abundance and their flow cytometric characteristics was conducted in an exorheic river (Haihe River, Northern China) over a one year period covering September (autumn) 2011, December (winter) 2011, April (spring) 2012, and July (summer) 2012. The results showed that LNA and HNA bacteria contributed similarly to the total bacterial abundance on both the spatial and temporal scale. The variability of HNA on abundance, fluorescence intensity (FL1) and side scatter (SSC) were more sensitive to environmental factors than that of LNA bacteria. Meanwhile, the relative distance of SSC between HNA and LNA was more variable than that of FL1. Multivariate analysis further demonstrated that the influence of geographical distance (reflected by the salinity gradient along river to ocean) and temporal changes (as temperature variation due to seasonal succession) on the patterns of LNA and HNA were stronger than the effects of nutrient conditions. Furthermore, the results demonstrated that the distribution of LNA and HNA bacteria, including the abundance, FL1 and SSC, was controlled by different variables. The results suggested that LNA and HNA bacteria might play different ecological roles in the exorheic river. PMID:27082986
Liu, Jie; Hao, Zhenyu; Ma, Lili; Ji, Yurui; Bartlam, Mark; Wang, Yingying
Bacteria with high nucleic acid (HNA) and low nucleic acid (LNA) content are commonly observed in aquatic environments. To date, limited knowledge is available on their temporal and spatial variations in freshwater environments. Here an investigation of HNA and LNA bacterial abundance and their flow cytometric characteristics was conducted in an exorheic river (Haihe River, Northern China) over a one year period covering September (autumn) 2011, December (winter) 2011, April (spring) 2012, and July (summer) 2012. The results showed that LNA and HNA bacteria contributed similarly to the total bacterial abundance on both the spatial and temporal scale. The variability of HNA on abundance, fluorescence intensity (FL1) and side scatter (SSC) were more sensitive to environmental factors than that of LNA bacteria. Meanwhile, the relative distance of SSC between HNA and LNA was more variable than that of FL1. Multivariate analysis further demonstrated that the influence of geographical distance (reflected by the salinity gradient along river to ocean) and temporal changes (as temperature variation due to seasonal succession) on the patterns of LNA and HNA were stronger than the effects of nutrient conditions. Furthermore, the results demonstrated that the distribution of LNA and HNA bacteria, including the abundance, FL1 and SSC, was controlled by different variables. The results suggested that LNA and HNA bacteria might play different ecological roles in the exorheic river.
Reale, Anna; Konietzny, Ursula; Coppola, Raffaele; Sorrentino, Elena; Greiner, Ralf
Lactic acid fermentation of cereal flours resulted in a 100 (rye), 95-100 (wheat), and 39-47% (oat) reduction in phytate content within 24 h. The extent of phytate degradation was shown to be independent from the lactic acid bacteria strain used for fermentation. However, phytate degradation during cereal dough fermentation was positively correlated with endogenous plant phytase activity (rye, 6750 mU g(-1); wheat, 2930 mU g(-1); and oat, 23 mU g(-1)), and heat inactivation of the endogenous cereal phytases prior to lactic acid fermentation resulted in a complete loss of phytate degradation. Phytate degradation was restored after addition of a purified phytase to the liquid dough. Incubation of the cereal flours in buffered solutions resulted in a pH-dependent phytate degradation. The optimum of phytate degradation was shown to be around pH 5.5. Studies on phytase production of 50 lactic acid bacteria strains, previously isolated from sourdoughs, did not result in a significant production of intra- as well as extracellular phytase activity. Therefore, lactic acid bacteria do not participate directly in phytate degradation but provide favorable conditions for the endogenous cereal phytase activity by lowering the pH value.
Onaka, Hiroyasu; Mori, Yukiko; Igarashi, Yasuhiro; Furumai, Tamotsu
Natural products produced by microorganisms are important starting compounds for drug discovery. Secondary metabolites, including antibiotics, have been isolated from different Streptomyces species. The production of these metabolites depends on the culture conditions. Therefore, the development of a new culture method can facilitate the discovery of new natural products. Here, we show that mycolic acid-containing bacteria can influence the biosynthesis of cryptic natural products in Streptomyces species. The production of red pigment by Streptomyces lividans TK23 was induced by coculture with Tsukamurella pulmonis TP-B0596, which is a mycolic acid-containing bacterium. Only living cells induced this pigment production, which was not mediated by any substances. T. pulmonis could induce natural-product synthesis in other Streptomyces strains too: it altered natural-product biosynthesis in 88.4% of the Streptomyces strains isolated from soil. The other mycolic acid-containing bacteria, Rhodococcus erythropolis and Corynebacterium glutamicum, altered biosynthesis in 87.5 and 90.2% of the Streptomyces strains, respectively. The coculture broth of T. pulmonis and Streptomyces endus S-522 contained a novel antibiotic, which we named alchivemycin A. We concluded that the mycolic acid localized in the outer cell layer of the inducer bacterium influences secondary metabolism in Streptomyces, and this activity is a result of the direct interaction between the mycolic acid-containing bacteria and Streptomyces. We used these results to develop a new coculture method, called the combined-culture method, which facilitates the screening of natural products.
Liao, Xue-Yi; Guo, Li-Qiong; Ye, Zhi-Wei; Qiu, Ling-Yan; Gu, Feng-Wei; Lin, Jun-Fang
Strains of Leuconostoc mesenteroides, Pediococcus pentosaceus, and Lactobacillus brevis were identified from mango fruits by partial 16S rDNA gene sequence. Based on the ability of producing mannitol and diacetyl, Leuconostoc mesenteroides MPL18 and MPL39 were selected within the lactic acid bacteria isolates, and used as mixed starters to ferment mango juice (MJ). Both the autochthonous strains grew well in fermented mango juice (FMJ) and remained viable at 9.81 log cfu mL(-1) during 30 days of storage at 4°C. The content of total sugar of FMJ was lower than that of MJ, while the concentration of mannitol was higher than that of MJ, and the concentration of diacetyl was 3.29 ± 0.12 mg L(-1). Among detected organic acids including citric acid, gallic acid, lactic acid, and acetic acid, only citric acid and gallic acid were found in MJ, while all detected organic acids were found in FMJ. The concentration of lactic acid of FMJ was the highest (78.62 ± 13.66 mM) among all detected organic acids. The DPPH radical scavenging capacity of FMJ was higher than that of MJ. Total phenolic compounds were better preserved in FMJ. The acidity and sweetness had a noticeable impact on the overall acceptance of the treated sample.
Könneke, Martin; Widdel, Friedrich
The effect of growth temperature on the cellular fatty acid composition of sulphate-reducing bacteria (SRB) was studied in 12 species belonging to eight genera including psychrophiles and mesophiles. Most of these species were of marine origin. The investigated SRB with the exception of four Desulfobacter species exhibited only a minor increase in the proportion of cis-unsaturated fatty acids (by < or = 5% per 10 degrees C) when the growth temperature was decreased; psychrophiles maintained their typically high content of cis-unsaturated fatty acids (around 75% of total fatty acids) nearly constant. The four Desulfobacter species, however, increased the proportion of cis-unsaturated among total fatty acids significantly (by > or =14% per 10 degrees C; measured in late growth phase) with decreasing growth temperature. The ratio between unsaturated and saturated fatty acids in Desulfobacter species changed not only with the growth temperature, but also with the growth state in batch cultures at constant temperature. Changes of cellular fatty acids were studied in detail with D. hydrogenophilus, the most psychrotolerant (growth range 0-35 degrees C) among the mesophilic SRB examined. Desulfobacter hydrogenophilus also formed cis-9,10-methylenehexadecanoic acid (a cyclopropane fatty acid) and 10-methylhexadecanoic acid. At low growth temperature (12 degrees C), the relative amount of these fatty acids was at least threefold lower; this questions the usefulness of 10-methylhexadecanoic acid as a reliable biomarker of Desulfobacter in cold sediments.
Polen, Tino; Wendisch, Volker F
DNA microarray technology has become an important research tool for biotechnology and microbiology. It is now possible to characterize genetic diversity and gene expression in a genomewide manner. DNA microarrays have been applied extensively to study the biology of many bacteria including Escherichia coli, but only recently have they been developed for the Gram-positive Corynebacterium glutamicum. Both bacteria are widely used for biotechnological amino acid production. In this article, in addition to the design and generation of microarrays as well as their use in hybridization experiments and subsequent data analysis, we describe recent applications of DNA microarray technology regarding amino acid production in C. glutamicum and E. coli. We also discuss the impact of functional genomics studies on fundamental as well as applied aspects of amino acid production with C. glutamicum and E. coli.
Spicher, G; Nierle, W
Acidification of the dough by the use of sourdough or acidifiers is necessary not only for good baking quality of rye flour but it is also very important for development of the typical sensory characteristics of rye bread. We confirmed that the lactic acid bacteria of sour dough are proteolytic. Proteolytic effects are observed in the increase of the amino acid content during fermentation. A marked increase was found in the content of leucine, alanine, valine, isoleucine, glutamic acid, glutamine, arginine, lysine, methionine, phenylalanine, tyrosine and serine. Lactobacillus plantarum showed a higher proteolytic activity than L. brevis ssp. lindneri or L. fructivorans.
Yang, Haiyan; Ge, Zhi; Wu, Dan; Tong, Meiping; Ni, Jinren
This study investigated the influence of multiple colloids (hematite and humic acid) on the transport and deposition of bacteria (Escherichia coli) in packed porous media in both NaCl (5 mM) and CaCl2 (1 mM) solutions at pH 6. Due to the alteration of cell physicochemical properties, the presence of hematite and humic acid in cell suspensions significantly affected bacterial transport and deposition in quartz sand. Specifically, the presence of hematite (5 mg/L) decreased cell transport (increased cell deposition) in quartz sand in both NaCl and CaCl2 solutions, which could be attributed to the less negative overall zeta potentials of bacteria induced by the adsorption of positively charged hematite onto cell surfaces. The presence of a low concentration (0.1 mg/L) of humic acid in bacteria and hematite mixed suspensions reduced the adsorption of hematite onto cell surfaces, leading to increased cell transport in quartz sand in NaCl solutions, whereas, in CaCl2 solutions, the presence of 0.1 mg/L humic acid increased the formation of hematite-cell aggregates and thus decreased cell transport in quartz sand. When the concentration of humic acid was increased to 1 mg/L, enhanced cell transport was observed in both NaCl and CaCl2 solutions. The decreased adsorption of hematite onto cell surfaces as well as the competition of deposition sites on quartz sand with bacteria by the suspended humic acid contributed to the increased cell transport.
Eisenberg, E.; Padova, R.; Kirsch, E.; Weissman, Sh.; Hirshfeld, T.; Shenfeld, A.
APT agar (APT) used for enumeration of lactic acid bacteria and Plate Count agar (PCA) applied for total count were sterilized by gamma radiation using radiation dose of 10-15 kGy. Radiosterilized PCA and APT modified by adding catalase prior to irradiation, or APT with increased content of yeast extract performed, as well as, the heat sterilized commercial media. Growth performance was evaluated on several strains of microorganisms, as well as, by enumeration of bacteria in food products. Radiosterilization of culture media in final packaging, can be applied to produce dip slide kits containing PCA or APT.
Scott, M G; Gold, M R; Hancock, R E
Compounds with antiendotoxin properties have been extensively studied for their potential as therapeutic agents for sepsis attributable to gram-negative bacteria. However, with the increasing incidence of gram-positive sepsis, there is interest in identifying compounds with a broad spectrum of action against both gram-positive and gram-negative bacteria. A series of synthetic alpha-helical cationic peptides related to bee melittin and silk moth cecropin have previously been shown to bind lipopolysaccharide (LPS) with high affinity, inhibit LPS-induced tumor necrosis factor alpha (TNF-alpha) production in vitro and in vivo, and kill gram-negative bacteria. In this study, we analyzed whether these peptides were active against gram-positive bacteria; whether they could bind to lipoteichoic acid (LTA), the major proinflammatory structure on gram-positive bacteria; and whether they could block the ability of LTA to promote the release of cytokines by the RAW 264.7 murine macrophage cell line. We found that the cationic peptides demonstrated moderate growth-inhibitory activity toward gram-positive bacteria. In addition, the peptides bound LTA with high affinity. This correlated with the ability of the peptides to block LTA-induced production of TNF and interleukin-6 by RAW 264.7 cells but did not correlate with their ability to kill the bacteria. The peptides also effectively inhibited LTA-induced TNF production in a whole human blood assay. The peptides were also able to partly block the ability of heat-killed Staphylococcus aureus, as well as soluble products of live S. aureus, to stimulate cytokine production by macrophages. Our results indicate that these cationic peptides may be useful to prevent sepsis and inflammation caused by both gram-negative and gram-positive bacteria.
Cibis, Katharina Gabriela; Gneipel, Armin; König, Helmut
In this study, acetic, propionic and butyric acid-forming bacteria were isolated from thermophilic and mesophilic biogas plants (BGP) located in Germany. The fermenters were fed with maize silage and cattle or swine manure. Furthermore, pressurized laboratory fermenters digesting maize silage were sampled. Enrichment cultures for the isolation of acid-forming bacteria were grown in minimal medium supplemented with one of the following carbon sources: Na(+)-dl-lactate, succinate, ethanol, glycerol, glucose or a mixture of amino acids. These substrates could be converted by the isolates to acetic, propionic or butyric acid. In total, 49 isolates were obtained, which belonged to the phyla Firmicutes, Tenericutes or Thermotogae. According to 16S rRNA gene sequences, most isolates were related to Clostridium sporosphaeroides, Defluviitoga tunisiensis and Dendrosporobacter quercicolus. Acetic, propionic or butyric acid were produced in cultures of isolates affiliated to Bacillus thermoamylovorans, Clostridium aminovalericum, Clostridium cochlearium/Clostridium tetani, C. sporosphaeroides, D. quercicolus, Proteiniborus ethanoligenes, Selenomonas bovis and Tepidanaerobacter sp. Isolates related to Thermoanaerobacterium thermosaccharolyticum produced acetic, butyric and lactic acid, and isolates related to D. tunisiensis formed acetic acid. Specific primer sets targeting 16S rRNA gene sequences were designed and used for real-time quantitative PCR (qPCR). The isolates were physiologically characterized and their role in BGP discussed.
Jørgensen, L.; Lechtenfeld, O. J.; Benner, R.; Middelboe, M.; Stedmon, C. A.
Dissolved organic matter (DOM) in the ocean consists of a heterogeneous mixture of molecules, most of which are of unknown origin. Neutral sugars and amino acids are among the few recognizable biomolecules in DOM, and the molecular composition of these biomolecules is shaped primarily by biological production and degradation processes. This study provides insight into the bioavailability of biomolecules as well as the chemical composition of DOM produced by bacteria. The molecular compositions of combined neutral sugars and amino acids were investigated in DOM produced by bacteria and in DOM remaining after 32 days of bacterial degradation. Results from bioassay incubations with natural seawater (sampled from water masses originating from the surface waters of the Arctic Ocean and the North Atlantic Ocean) and artificial seawater indicate that the molecular compositions following bacterial degradation are not strongly influenced by the initial substrate or bacterial community. The molecular composition of neutral sugars released by bacteria was characterized by a high glucose content (47 mol %) and heterogeneous contributions from other neutral sugars (3-14 mol %). DOM remaining after bacterial degradation was characterized by a high galactose content (33 mol %), followed by glucose (22 mol %) and the remaining neutral sugars (7-11 mol %). The ratio of D-amino acids to L-amino acids increased during the experiments as a response to bacterial degradation, and after 32 days, the D/L ratios of aspartic acid, glutamic acid, serine and alanine reached around 0.79, 0.32, 0.30 and 0.51 in all treatments, respectively. The striking similarity in neutral sugar and amino acid compositions between natural (representing marine semi-labile and refractory DOM) and artificial (representing bacterially produced DOM) seawater samples, suggests that microbes transform bioavailable neutral sugars and amino acids into a common, more persistent form.
This paper gives an overview of the recent advances in engineering the central carbon metabolism of the industrially important bacteria Escherichia coli, Bacillus subtilis, Corynobacterium glutamicum, Streptomyces spp., Lactococcus lactis and other lactic acid bacteria. All of them are established producers of important classes of products, e.g. proteins, amino acids, organic acids, antibiotics, high-value metabolites for the food industry and also, promising producers of a large number of industrially or therapeutically important chemicals. Optimization of existing or introduction of new cellular processes in these microorganisms is often achieved through manipulation of targets that reside at major points of central metabolic pathways, such as glycolysis, gluconeogenesis, the pentose phosphate pathway and the tricarboxylic acid cycle with the glyoxylate shunt. Based on the huge progress made in recent years in biochemical, genetic and regulatory studies, new fascinating engineering approaches aim at ensuring an optimal carbon and energy flow within central metabolism in order to achieve optimized metabolite production. PMID:22545791
Golomb, Benjamin L; Marco, Maria L
Lactic acid bacteria have been isolated from living, harvested, and fermented plant materials; however, the adaptations these bacteria possess for growth on plant tissues are largely unknown. In this study, we investigated plant habitat-specific traits of Lactococcus lactis during growth in an Arabidopsis thaliana leaf tissue lysate (ATL). L. lactis KF147, a strain originally isolated from plants, exhibited a higher growth rate and reached 7.9-fold-greater cell densities during growth in ATL than the dairy-associated strain L. lactis IL1403. Transcriptome profiling (RNA-seq) of KF147 identified 853 induced and 264 repressed genes during growth in ATL compared to that in GM17 laboratory culture medium. Genes induced in ATL included those involved in the arginine deiminase pathway and a total of 140 carbohydrate transport and metabolism genes, many of which are involved in xylose, arabinose, cellobiose, and hemicellulose metabolism. The induction of those genes corresponded with L. lactis KF147 nutrient consumption and production of metabolic end products in ATL as measured by gas chromatography-time of flight mass spectrometry (GC-TOF/MS) untargeted metabolomic profiling. To assess the importance of specific plant-inducible genes for L. lactis growth in ATL, xylose metabolism was targeted for gene knockout mutagenesis. Wild-type L. lactis strain KF147 but not an xylA deletion mutant was able to grow using xylose as the sole carbon source. However, both strains grew to similarly high levels in ATL, indicating redundancy in L. lactis carbohydrate metabolism on plant tissues. These findings show that certain strains of L. lactis are well adapted for growth on plants and possess specific traits relevant for plant-based food, fuel, and feed fermentations.
Golomb, Benjamin L.
Lactic acid bacteria have been isolated from living, harvested, and fermented plant materials; however, the adaptations these bacteria possess for growth on plant tissues are largely unknown. In this study, we investigated plant habitat-specific traits of Lactococcus lactis during growth in an Arabidopsis thaliana leaf tissue lysate (ATL). L. lactis KF147, a strain originally isolated from plants, exhibited a higher growth rate and reached 7.9-fold-greater cell densities during growth in ATL than the dairy-associated strain L. lactis IL1403. Transcriptome profiling (RNA-seq) of KF147 identified 853 induced and 264 repressed genes during growth in ATL compared to that in GM17 laboratory culture medium. Genes induced in ATL included those involved in the arginine deiminase pathway and a total of 140 carbohydrate transport and metabolism genes, many of which are involved in xylose, arabinose, cellobiose, and hemicellulose metabolism. The induction of those genes corresponded with L. lactis KF147 nutrient consumption and production of metabolic end products in ATL as measured by gas chromatography-time of flight mass spectrometry (GC-TOF/MS) untargeted metabolomic profiling. To assess the importance of specific plant-inducible genes for L. lactis growth in ATL, xylose metabolism was targeted for gene knockout mutagenesis. Wild-type L. lactis strain KF147 but not an xylA deletion mutant was able to grow using xylose as the sole carbon source. However, both strains grew to similarly high levels in ATL, indicating redundancy in L. lactis carbohydrate metabolism on plant tissues. These findings show that certain strains of L. lactis are well adapted for growth on plants and possess specific traits relevant for plant-based food, fuel, and feed fermentations. PMID:25384484
Liu, S.; Pritchard, G. G.; Hardman, M. J.; Pilone, G. J.
l-Arginine, an amino acid found in significant quantities in grape juice and wine, is known to be catabolized by some wine lactic acid bacteria. The correlation between the occurrence of arginine deiminase pathway enzymes and the ability to catabolize arginine was examined in this study. The activities of the three arginine deiminase pathway enzymes, arginine deiminase, ornithine transcarbamylase, and carbamate kinase, were measured in cell extracts of 35 strains of wine lactic acid bacteria. These enzymes were present in all heterofermentative lactobacilli and most leuconostocs but were absent in all the homofermentative lactobacilli and pediococci examined. There was a good correlation among arginine degradation, formation of ammonia and citrulline, and the occurrence of arginine deiminase pathway enzymes. Urea was not detected during arginine degradation, suggesting that the catabolism of arginine did not proceed via the arginase-catalyzed reaction, as has been suggested in some earlier studies. Detection of ammonia with Nessler's reagent was shown to be a simple, rapid test to assess the ability of wine lactic acid bacteria to degrade arginine, although in media containing relatively high concentrations (>0.5%) of fructose, ammonia formation is inhibited. PMID:16534912
Ordóñez, J L; Sainz, F; Callejón, R M; Troncoso, A M; Torija, M J; García-Parrilla, M C
This paper studies the amino acid profile of beverages obtained through the fermentation of strawberry purée by a surface culture using three strains belonging to different acetic acid bacteria species (one of Gluconobacter japonicus, one of Gluconobacter oxydans and one of Acetobacter malorum). An HPLC-UV method involving diethyl ethoxymethylenemalonate (DEEMM) was adapted and validated. From the entire set of 21 amino acids, multiple linear regressions showed that glutamine, alanine, arginine, tryptophan, GABA and proline were significantly related to the fermentation process. Furthermore, linear discriminant analysis classified 100% of the samples correctly in accordance with the microorganism involved. G. japonicus consumed glucose most quickly and achieved the greatest decrease in amino acid concentration. None of the 8 biogenic amines were detected in the final products, which could serve as a safety guarantee for these strawberry gluconic fermentation beverages, in this regard.
Hamidi, Adel; Mirnejad, Reza; Yahaghi, Emad; Behnod, Vahid; Mirhosseini, Ali; Amani, Sajad; Sattari, Sara; Darian, Ebrahim Khodaverdi
Objective To determine lactic acid bacteria's capability to enhance the process of binding and isolating aflatoxin B1 and to utilize such lactic acid bacteria as a food supplement or probiotic products for preventing absorption of aflatoxin B1 in human and animal bodies. Methods In the present research, the bacteria were isolated from five different sources. For surveying the capability of the bacteria in isolating aflatoxin B1, ELISA method was implemented, and for identifying the resultant strains through 16S rRNA sequencing method, universal primers were applied. Results Among the strains which were isolated, two strains of Lactobacillus pentosus and Lactobacillus beveris exhibited the capability of absorbing and isolating aflatoxin B1 by respectively absorbing and discharging 17.4% and 34.7% of the aforementioned toxin existing in the experiment solution. Conclusions Strains of Lactobacillus pentosus and Lactobacillus beveris were isolated from human feces and local milk samples, respectively. And both strains has the ability to isolate or bind with aflatoxin B1. PMID:23998015
Alvarez-Calatayud, Guillermo; Margolles, Abelardo
Probiotics are living micro-organisms that do not naturally have shelf life, and normally are weakly protected against the digestive action of the GI tract. A new dual coating technology has been developed in an effort to maximize survival, that is, to be able to reach the intestine alive and in sufficient numbers to confer the beneficial health effects on the host. Dual-coating of lactic acid bacteria (LAB) is the result of fourth-generation coating technology for the protection of these bacteria at least 100-fold or greater than the uncoated LAB. This innovative technique involves a first pH-dependent protein layer that protects bacteria from gastric acid and bile salt, and a second polysaccharide matrix that protects bacteria from external factors, such as humidity, temperature and pressure, as well as the digestive action during the passage through the GI tract. Dual-coated probiotic formulation is applicable to different therapeutic areas, including irritable bowel syndrome, atopic dermatitis, acute diarrhea, chronic constipation, Helicobacter pylori eradication, and prevention of antibiotic-associated diarrhea. An updated review of the efficacy of doubly coated probiotic strains for improving bacterial survival in the intestinal tract and its consequent clinical benefits in humans is here presented.
Sugimoto, Shinya; Abdullah-Al-Mahin; Sonomoto, Kenji
Recently, lactic acid bacteria (LAB) have attracted much attention because of their potential application to probiotics and industrial applications as starters for dairy products or lactic acid fermentation. Additional emphasis is also being paid to them as commensal bacteria in gastrointestinal tract. Since LAB exhibit a stress response, insight into the relationship between stress proteins such as molecular chaperones and stress tolerance or adaptation is increasing gradually along with current research examining these important bacteria. Similar to other bacteria, one of the major stress-response systems in LAB is the expression of molecular chaperones. The recently completed genome sequencing of various LAB strains, combined with the development of advanced molecular techniques, have enabled us to identify molecular chaperones and to understand their regulation systems in response to various stresses. Furthermore, recent biochemical studies provided novel insight into the molecular mechanisms of LAB chaperone systems. This review highlights the physiological consequences and biochemical properties of molecular chaperones (especially sHsps, Hsp70, and Hsp100) in LAB and their use in biotechnological applications.
Cao, Rong; Liu, Qi; Chen, Shengjun; Yang, Xianqing; Li, Laihao
Aquatic products are extremely perishable food commodities. Developing methods to keep the freshness of fish represents a major task of the fishery processing industry. Application of Lactic Acid Bacteria (LAB) as food preservative is a novel approach. In the present study, the possibility of using lactic acid bacteria in freshness keeping of tilapia fillets as sashimi was examined. Fish fillets were dipped in Lactobacillus plantarum 1.19 (obtained from China General Microbiological Culture Collection Center) suspension as LAB-treated group. Changes in K-value, APC, sensory properties and microbial flora were analyzed. Results showed that LAB treatment slowed the increase of K-value and APC in the earlier storage, and caused a smooth decrease in sensory score. Gram-negative bacteria dominated during refrigerated storage, with Pseudomonas and Aeromonas being relatively abundant. Lactobacillus plantarum 1.19 had no obvious inhibitory effect against these Gram-negatives. However, Lactobacillus plantarum 1.19 changed the composition of Gram-positive bacteria. No Micrococcus were detected and the proportion of Staphylococcus decreased in the spoiled LAB-treated samples. The period that tilapia fillets could be used as sashimi material extended from 24 h to 48 h after LAB treatment. The potential of using LAB in sashimi processing was confirmed.
Sánchez, Borja; Chaignepain, Sthéphane; Schmitter, Jean-Marie; Urdaci, María C
Lactic acid bacteria (LAB) are known for their special nutritional requirements, being usually cultured in complex media to achieve optimal growth. In this paper, a protocol based on trichloroacetic acid precipitation of peptides and proteins is presented. The method has been tested on four probiotic LAB strains grown in De Man Rogosa Sharpe (MRS) broth, a complex medium that is often used for the culture of such bacteria. This protocol allowed the detection of 19 proteins after sodium dodecyl sulfate-polyacrylamide gel electrophoresis, 10 of them being successfully identified by tandem MS. Thereafter, the 10 were found to be secreted or surface associated by bioinformatic means. In conclusion, this work supplies a method for the identification of proteins secreted by LAB, allowing discrimination between the proteins present in the MRS and those produced by probiotic LAB.
Asano, Shizuka; Suzuki, Koji; Iijima, Kazumaru; Motoyama, Yasuo; Kuriyama, Hidetoshi; Kitagawa, Yasushi
Membrane filter performance was investigated using beer-spoilage lactic acid bacteria (LAB). As a result, beer-adapted LAB strains showed considerably increased penetration rate through filters, as compared with non-adapted strains. Further statistical analyses demonstrated the significant shifts in cell size distribution towards shorter rods, when Lactobacillus brevis and L. lindneri strains were precultured in beer. These results indicate that diminished cell size is responsible for the deteriorated filter performance and, therefore, beer-adapted lactic acid bacteria are regarded as a serious threat to the production of unpasteurized beers. In addition, the selection of test strains and preculture conditions are suggested to be important for the rigorous and standardized evaluation of membrane filter performance in the brewing industry.
El Bassi, Leila; Hassouna, Mnasser; Shinzato, Naoya; Matsui, Toru
Two lactic acid bacteria (LAB) were selected from 100 LAB isolated from various sea products to examine their use in Dicentrarchus labrax preservation. The isolates, tentatively named strain nr 3 and 7, were identified as Lactobacillus plantarum and L. pentosus, respectively. They showed antagonistic activity against psychrotroph, pathogenic, and coliform bacteria. The antagonistic activity of strain 3 was suggested to be by bacteriocins since activity was abolished by protease treatment, while that of strain 7 was due to the effect of pH decrease caused by the produced organic acids. Their use prevented total volatile basic nitrogen contents (TVB-N) and trimethylamine (TMA) to some extent, suggesting that inoculation could extend the period of storage.
Martin, F; Cachon, R; Pernin, K; De Coninck, J; Gervais, P; Guichard, E; Cayot, N
The aim of this study was to investigate the effect of oxidoreduction potential (Eh) on the biosynthesis of aroma compounds by lactic acid bacteria in non-fat yogurt. The study was done with yogurts fermented by Lactobacillus bulgaricus and Streptococcus thermophilus. The Eh was modified by the application of different gaseous conditions (air, nitrogen, and nitrogen/hydrogen). Acetaldehyde, dimethyl sulfide, diacetyl, and pentane-2,3-dione, as the major endogenous odorant compounds of yogurt, were chosen as tracers for the biosynthesis of aroma compounds by lactic acid bacteria. Oxidative conditions favored the production of acetaldehyde, dimethyl sulfide, and diketones (diacetyl and pentane-2,3-dione). The Eh of the medium influences aroma production in yogurt by modifying the metabolic pathways of Lb. bulgaricus and Strep. thermophilus. The use of Eh as a control parameter during yogurt production could permit the control of aroma formation.
Shcherbanovsky, L R; Kapelev, I G
The antimicrobial activity of 25 volatile oils from aerial parts and seeds of dill (Anethum graveolens L.) of different geographical origin towards yeast Saccharomyces vini and lactic acid bacteria Lactobacterium buchneri was measured by serial dilutions. Volatile oils from mature seeds and green parts of the plants harvested at late vegetation phases showed the highest activity. The geographical origin of plants influenced insignificantly the antimicrobial activity of volatile oil.
Background Many probiotic bacteria have been described as promising tools for the treatment and prevention of inflammatory bowel diseases (IBDs). Most of these bacteria are lactic acid bacteria, which are part of the healthy human microbiota. However, little is known about the effects of transient bacteria present in normal diets, including Lactococcus lactis. Methods In the present study, we analysed the immunomodulatory effects of three L. lactis strains in vitro using intestinal epithelial cells. L. lactis NCDO 2118 was administered for 4 days to C57BL/6 mice during the remission period of colitis induced by dextran sodium sulphate (DSS). Results Only one strain, L. lactis NCDO 2118, was able to reduce IL-1β-induced IL-8 secretion in Caco-2 cells, suggesting a potential anti-inflammatory effect. Oral treatment using L. lactis NCDO 2118 resulted in a milder form of recurrent colitis than that observed in control diseased mice. This protective effect was not attributable to changes in secretory IgA (sIgA); however, NCDO 2118 administration was associated with an early increase in IL-6 production and sustained IL-10 production in colonic tissue. Mice fed L. lactis NCDO 2118 had an increased number of regulatory CD4+ T cells (Tregs) bearing surface TGF-β in its latent form (Latency-associated peptide-LAP) in the mesenteric lymph nodes and spleen. Conclusions Here, we identified a new probiotic strain with a potential role in the treatment of IBD, and we elucidated some of the mechanisms underlying its anti-inflammatory effect. PMID:25110521
Furukawa, Soichi; Akiyoshi, Yuko; O'Toole, George A; Ogihara, Hirokazu; Morinaga, Yasushi
Effects of food additives on biofilm formation by food-borne pathogenic bacteria were investigated. Thirty-three potential food additives and 3 related compounds were added to the culture medium at concentrations from 0.001 to 0.1% (w/w), followed by inoculation and cultivation of five biofilm-forming bacterial strains for the evaluation of biofilm formation. Among the tested food additives, 21 showed inhibitory effects of biofilm formation by Staphylococcus aureus and Escherichia coli, and in particular, sugar fatty acid esters showed significant anti-biofilm activity. Sugar fatty acid esters with long chain fatty acid residues (C14-16) exerted their inhibitory effect at the concentration of 0.001% (w/w), but bacterial growth was not affected at this low concentration. Activities of the sugar fatty acid esters positively correlated with the increase of the chain length of the fatty acid residues. Sugar fatty acid esters inhibited the initial attachment of the S. aureus cells to the abiotic surface. Sugar fatty acid esters with long chain fatty acid residues (C14-16) also inhibited biofilm formation by Streptococcus mutans and Listeria monocytogenes at 0.01% (w/w), while the inhibition of biofilm formation by Pseudomonas aeruginosa required the addition of a far higher concentration (0.1% (w/w)) of the sugar fatty acid esters.
Sebastià, Natividad; Meca, Giuseppe; Soriano, José Miguel; Mañes, Jordi
Enniatins (ENs) are N-methylated cyclohexadepsipeptides, secondary metabolites produced by various species of the genus Fusarium. They are known to act as antifungal, antiyeast and antibacterial and to possess antiinsecticidal and phytotoxic properties. In this study we evaluated for the first time the antibiotic effect of pure fractions of EN J(1) and J(3) on several pathogenic strains and lactic acid bacteria. The ENs J(1) and J(3) were purified from the fermentation extract of Fusarium solani growth on solid medium of wheat kamut, using the technique of the low pressure liquid chromatography (LPLC) followed by a semipreparative liquid chromatography (LC). The purity and the structure of the isolated compound were confirmed by electrospray ionization-mass spectrometry study-linear ion trap (ESI-MS-LIT). The use of both chromatographic techniques have permitted to produce and purify 47mg of the En J(1) and 50mg of the EN J(3) with a mean purity of 98% completely characterized with the technique of the ESI-MS-LIT. Microbial bioassay analyses were carried out by incubation in MRSA and TSA for acid lactic and pathogenic bacteria, respectively during 24h at 37°C. None of the tested strains were inhibited by a 1ng dose of EN J(1) and J(3). These compounds were only not effective against Listeria monocytogenes, Pseudomonas aeruginosa and Salmonella enteric. This study highlight ENs J(1) and J(3) could be potentially effective antibacterial agents against several pathogenic and lactic acid bacteria.
MENCONI, Anita; KALLAPURA, Gopala; LATORRE, Juan D.; MORGAN, Marion J.; PUMFORD, Neil R.; HARGIS, Billy M.; TELLEZ, Guillermo
The aim of the present study was to describe the identification and characterization (physiological properties) of two strains of lactic acid bacteria (LAB 18 and 48) present in a commercial probiotic culture, FloraMax®-B11. Isolates were characterized morphologically, and identified biochemically. In addition, the MIDI System ID, the Biolog ID System, and 16S rRNA sequence analyses for identification of LAB 18 and LAB 48 strains were used to compare the identification results. Tolerance and resistance to acidic pH, high osmotic concentration of NaCl, and bile salts were tested in broth medium. In vitro assessment of antimicrobial activity against enteropathogenic bacteria and susceptibility to antibiotics were also tested. The results obtained in this study showed tolerance of LAB 18 and LAB 48 to pH 3.0, 6.5% NaCl and a high bile salt concentration (0.6%). Both strains evaluated showed in vitro antibacterial activity against Salmonella enterica serovar Enteritidis, Escherichia coli (O157:H7), and Campylobacter jejuni. These are important characteristics of lactic acid bacteria that should be evaluated when selecting strains to be used as probiotics. Antimicrobial activity of these effective isolates may contribute to efficacy, possibly by direct antimicrobial activity in vivo. PMID:24936379
Kormin, Salasiah; Rusul, Gulam; Radu, Son; Ling, Foo Hooi
Lactic Acid Bacteria (LAB) isolated from several traditional fermented foods such as “tempeh”, “tempoyak” and “tapai” were screened for the production of bacteriocin. One strain isolated from “tempeh” gives an inhibitory activity against several LAB. The strain was later identified as Lactobacillus plantarum BS2. Study shows that the inhibitory activity was not caused by hydrogen peroxide, organic acids or bacteriophage. The bacteriocin production was maximum after 10 hours of incubation with an activity of 200 AU/ml. The bacteriocin was found to be sensitive towards trypsin, α-chymotrypsin, β-chymotrypsin, α-amylase and lysozyme. PMID:22973159
Mutaguchi, Yuta; Ohmori, Taketo; Akano, Hirofumi; Doi, Katsumi; Ohshima, Toshihisa
Levels of free D-amino acids were compared in 11 vinegars produced from different sources or through different manufacturing processes. To analyze the D- and L-amino acids, the enantiomers were initially converted into diastereomers using pre-column derivatization with o-phthaldialdehyde plus N-acethyl-L-cysteine or N-tert-butyloxycarbonyl-L-cysteine. This was followed by separation of the resultant fluorescent isoindol derivatives on an octadecylsilyl stationary phase using ultra-performance liquid chromatography. The analyses showed that the total D-amino acid level in lactic fermented tomato vinegar was very high. Furthermore, analysis of the amino acids in tomato juice samples collected after alcoholic, lactic and acetic fermentation during the production of lactic fermented tomato vinegar showed clearly that lactic fermentation is responsible for the D-amino acids production; marked increases in D-amino acids were seen during lactic fermentation, but not during alcoholic or acetic fermentation. This suggests lactic acid bacteria have a greater ability to produce D-amino acids than yeast or acetic acid bacteria.
Lasta, Samar; Ouzari, Hadda; Andreotti, Nicolas; Fajloun, Ziad; Mansuelle, Pascal; Boudabous, Abdellatif; Sampieri, Francois; Sabatier, Jean Marc
A new bacteriocin, lacticin LC14, produced by Lactococcus lactis BMG6.14, was isolated and characterized. It was purified to homogeneity from overnight broth culture by ammonium sulfate precipitation, Sep-Pak chromatography, and two steps of reversed-phase HPLC. Lacticin LC14 showed bactericidal-type antimicrobial activity against several lactic acid bacteria and pathogenic strains including Listeria monocytogenes. It was inactivated by proteinase K and pronase E, but was resistant to papain, lysozyme, lipase and catalase. Lacticin LC14 was heat resistant, stable over a wide range of pH (2-10) and after treatment by solvents and detergents. Its N-terminal end was found unreactive towards Edman sequencing. Based on MALDI-TOF mass spectrometry, its molecular mass was 3333.7 Da. LC14 amino acid composition revealed a high proportion of hydrophobic residues, but no modified ones. LC14 may be able to challenge other well known other bacteriocins in probiotic and therapeutic applications.
Herranz, Carmen; Driessen, Arnold J. M.
Most lactic acid bacterium bacteriocins utilize specific leader peptides and dedicated machineries for secretion. In contrast, the enterococcal bacteriocin enterocin P (EntP) contains a typical signal peptide that directs its secretion when heterologously expressed in Lactococcus lactis. Signal peptide mutations and the SecA inhibitor azide blocked secretion. These observations demonstrate that EntP is secreted by the Sec translocase. PMID:15812026
Thompson, A. S.; Owens, N.; Murrell, J. C.
Two methylotrophic bacterial strains, TR3 and PSCH4, capable of growth on methanesulfonic acid as the sole carbon source were isolated from the marine environment. Methanesulfonic acid metabolism in these strains was initiated by an inducible NADH-dependent monooxygenase, which cleaved methanesulfonic acid into formaldehyde and sulfite. The presence of hydroxypyruvate reductase and the absence of ribulose monophosphate-dependent hexulose monophosphate synthase indicated the presence of the serine pathway for formaldehyde assimilation. Cell suspensions of bacteria grown on methanesulfonic acid completely oxidized methanesulfonic acid to carbon dioxide and sulfite with a methanesulfonic acid/oxygen stoichiometry of 1.0:2.0. Oxygen electrode-substrate studies indicated the dissimilation of formaldehyde to formate and carbon dioxide for energy generation. Carbon dioxide was not fixed by ribulose bisphosphate carboxylase. It was shown that methanol is not an intermediate in methanesulfonic acid metabolism, although these strains grew on methanol and other one-carbon compounds, as well as a variety of heterotrophic carbon sources. These two novel marine facultative methylotrophs have the ability to mineralize methanesulfonic acid and may play a role in the cycling of global organic sulfur. PMID:16535055
Nsogning Dongmo, Sorelle; Sacher, Bertram; Kollmannsberger, Hubert; Becker, Thomas
This study aims to define the aroma composition and key aroma compounds of barley malt wort beverages produced from fermentation using six lactic acid bacteria (LAB) strains. Gas chromatography mass spectrometry-olfactometry and flame ionization detection was employed; key aroma compounds were determined by means of aroma extract dilution analysis. Fifty-six detected volatile compounds were similar among beverages. However, significant differences were observed in the concentration of individual compounds. Key aroma compounds (flavor dilution (FD) factors ≥16) were β-damascenone, furaneol, phenylacetic acid, 2-phenylethanol, 4-vinylguaiacol, sotolon, methional, vanillin, acetic acid, nor-furaneol, guaiacol and ethyl 2-methylbutanoate. Furthermore, acetaldehyde had the greatest odor activity value of up to 4266. Sensory analyses revealed large differences in the flavor profile. Beverage from L. plantarum Lp. 758 showed the highest FD factors in key aroma compounds and was correlated to fruity flavors. Therefore, we suggest that suitable LAB strain selection may improve the flavor of malt based beverages.
Le Lay, Céline; Coton, Emmanuel; Le Blay, Gwenaëlle; Chobert, Jean-Marc; Haertlé, Thomas; Choiset, Yvan; Van Long, Nicolas Nguyen; Meslet-Cladière, Laurence; Mounier, Jérôme
Fungal growth in bakery products represents the most frequent cause of spoilage and leads to economic losses for industrials and consumers. Bacteria, such as lactic acid bacteria and propionibacteria, are commonly known to play an active role in preservation of fermented food, producing a large range of antifungal metabolites. In a previous study (Le Lay et al., 2016), an extensive screening performed both in vitro and in situ allowed for the selection of bacteria exhibiting an antifungal activity. In the present study, active supernatants against Penicillium corylophilum and Aspergillus niger were analyzed to identify and quantify the antifungal compounds associated with the observed activity. Supernatant treatments (pH neutralization, heating and addition of proteinase K) suggested that organic acids played the most important role in the antifungal activity of each tested supernatant. Different methods (HPLC, mass spectrometry, colorimetric and enzymatic assays) were then applied to analyze the supernatants and it was shown that the main antifungal compounds corresponded to lactic, acetic and propionic acids, ethanol and hydrogen peroxide, as well as other compounds present at low levels such as phenyllactic, hydroxyphenyllactic, azelaic and caproic acids. Based on these results, various combinations of the identified compounds were used to evaluate their effect on conidial germination and fungal growth of P. corylophilum and Eurotium repens. Some combinations presented the same activity than the bacterial culture supernatant thus confirming the involvement of the identified molecules in the antifungal activity. The obtained results suggested that acetic acid was mainly responsible for the antifungal activity against P. corylophilum and played an important role in E. repens inhibition.
Tizzano, Marco; Sbarbati, Andrea
Bacteria are known to regulate diverse physiological processes through a mechanism called quorum sensing (QS). Prokaryotes communicate by extracellular signalling compounds, i.e. autoinducers (acyl homoserine lactone, AHL of Gram negative bacteria) or pheromones (post-translationally modified peptides of Gram positive bacteria), which activate genetic pathways when they reach a sufficient concentration (QS). A large number of Gram-negative quorum-sensing systems studied so far utilize N-acyl homoserine lactones as signal molecules. In vertebrates small synthetic molecules called growth hormone secretagogues (GHSs) stimulate the release of growth hormone (GH) from the pituitary. GH release is stimulated by hypothalamic GH-releasing hormone (GHRH) and ghrelin (endogenous ligand of the GHS-receptor, GHS-R). Ghrelin is a 28-amino acid peptide, in which the serine-3 (Ser3) is n-octanoylated, and this modification is essential for ghrelin's activity. Ghrelin is the first known case of a peptide hormone modified by a fatty acid. The major active form of ghrelin is a 28-amino acid peptide with octanoylated Ser3; one of the more represented bacterial autoinducers is the N-Octanoyl-DL-homoserine lactone (C8-HL) molecule. The authors hypothesize that Gram-negative bacteria and vertebrates have a functional similarity in the search of food and an important structural homology of AHL and ghrelin for the highly conserved Serine-acylated motive in both molecules. Our suggestions could help one to understand the convergent origin and the biologic meaning of the Serine-acylated group in these organisms, a biologic meaning very important due to the high conservation in two kingdoms which are so different.
Carpenter, C E; Broadbent, J R
Although the mechanisms by which organic acids inhibit growth of bacteria in mildly acidic foods are not fully understood, it is clear that intracellular accumulation of anions is a primary contributor to inhibition of bacterial growth. We hypothesize that intracellular accumulation of anions is driven by 2 factors, external anion concentration and external acidity. This hypothesis follows from basic chemistry principles that heretofore have not been fully applied to studies in the field, and it has led us to develop a novel approach for predicting internal anion concentration by controlling the external concentration of anions and pH. This approach overcomes critical flaws in contemporary experimental design that invariably target concentration of either protonated acid or total acid in the growth media thereby leaving anion concentration to vary depending on the pK(a) of the acids involved. Failure to control external concentration of anions has undoubtedly confounded results, and it has likely led to misleading conclusions regarding the antimicrobial action of organic acids. In summary, we advocate an approach for directing internal anion levels by controlling external concentration of anions and pH because it presents an additional opportunity to study the mechanisms by which organic acids inhibit bacterial growth. Knowledge gained from such studies would have important application in the control of important foodborne pathogens such as Listeria monocytogenes, and may also facilitate efforts to promote the survival in foods or beverages of desirable probiotic bacteria.
Costa, Stefania; Maldonado Rodriguez, Maria Elena; Rugiero, Irene; De Bastiani, Morena; Medici, Alessandro; Tamburini, Elena; Pedrini, Paola
The biotransformations of cholic acid (1a), deoxycholic acid (1b), and hyodeoxycholic acid (1c) to bendigoles and other metabolites with bacteria isolated from the rural slaughterhouse of Cayambe (Pichincha Province, Ecuador) were reported. The more active strains were characterized, and belong to the genera Pseudomonas and Rhodococcus. Various biotransformation products were obtained depending on bacteria and substrates. Cholic acid (1a) afforded the 3-oxo and 3-oxo-4-ene derivatives 2a and 3a (45% and 45%, resp.) with P. mendocina ECS10, 3,12-dioxo-4-ene derivative 4a (60%) with Rh. erythropolis ECS25, and 9,10-secosteroid 6 (15%) with Rh. erythropolis ECS12. Bendigole F (5a) was obtained in 20% with P. fragi ECS22. Deoxycholic acid (1b) gave 3-oxo derivative 2b with P. prosekii ECS1 and Rh. erythropolis ECS25 (20% and 61%, resp.), while 3-oxo-4-ene derivative 3b was obtained with P. prosekii ECS1 and P. mendocina ECS10 (22% and 95%, resp.). Moreover, P. fragi ECS9 afforded bendigole A (8b; 80%). Finally, P. mendocina ECS10 biotransformed hyodeoxycholic acid (1c) to 3-oxo derivative 2c (50%) and Rh. erythropolis ECS12 to 6α-hydroxy-3-oxo-23,24-dinor-5β-cholan-22-oic acid (9c, 66%). Bendigole G (5c; 13%) with P. prosekii ECS1 and bendigole H (8c) with P. prosekii ECS1 and Rh. erythropolis ECS12 (20% and 16%, resp.) were obtained.
Blobel, F; Eberspächer, J; Haug, S; Lingens, F
The cis,cis-muconic acid derivative of pyrazon, which was formerly isolated from the medium of pyrazon-degrading bacteria, was formed enzymatically by incubation of the catechol derivative of pyrazon with partially purified ortho pyrocatechase from pyrazon-degrading bacteria.
Axel, Claudia; Brosnan, Brid; Zannini, Emanuele; Furey, Ambrose; Coffey, Aidan; Arendt, Elke K
The use of sourdough fermented with specific strains of antifungal lactic acid bacteria can reduce chemical preservatives in bakery products. The main objective of this study was to investigate the production of antifungal carboxylic acids after sourdough fermentation of quinoa and rice flour using the antifungal strains Lactobacillus reuteri R29 and Lactobacillus brevis R2Δ as bioprotective cultures and the non-antifungal L. brevis L1105 as a negative control strain. The impact of the fermentation substrate was evaluated in terms of metabolic activity, acidification pattern and quantity of antifungal carboxylic acids. These in situ produced compounds (n=20) were extracted from the sourdough using a QuEChERS method and detected by a new UHPLC-MS/MS chromatography. Furthermore, the sourdough was applied in situ using durability tests against environmental moulds to investigate the biopreservative potential to prolong the shelf life of bread. Organic acid production and TTA values were lowest in rice sourdough. The sourdough fermentation of the different flour substrates generated a complex and significantly different profile of carboxylic acids. Extracted quinoa sourdough detected the greatest number of carboxylic acids (n=11) at a much higher concentration than what was detected from rice sourdough (n=9). Comparing the lactic acid bacteria strains, L. reuteri R29 fermented sourdoughs contained generally higher concentrations of acetic and lactic acid but also the carboxylic acids. Among them, 3-phenyllactic acid and 2-hydroxyisocaproic acid were present at a significant concentration. This was correlated with the superior protein content of quinoa flour and its high protease activity. With the addition of L. reuteri R29 inoculated sourdough, the shelf life was extended by 2 days for quinoa (+100%) and rice bread (+67%) when compared to the non-acidified controls. The L. brevis R2Δ fermented sourdough bread reached a shelf life of 4 days for quinoa (+100%) and
Bover-Cid, S; Holzapfel, W H
An improved screening plate method for the detection of amino acid decarboxylase-positive microorganisms (especially lactic acid bacteria) was developed. The suitability and detection level of the designed medium were quantitatively evaluated by confirmation of amine-forming capacity using an HPLC procedure. The potential to produce the biogenic amines (BA) tyramine, histamine, putrescine, and cadaverine, was investigated in a wide number of lactic acid bacteria (LAB) of different origin, including starter cultures, protective cultures, type strains and strains isolated from different food products. Also, several strains of Enterobacteriaceae were examined. Modifications to previously described methods included lowering glucose and sodium chloride concentrations, and increasing the buffer effect with calcium carbonate and potassium phosphate. In addition, pyridoxal-5-phosphate was included as a codecarboxylase factor for its enhancing effect on the amino acid decarboxylase activity. The screening plate method showed a good correlation with the chemical analysis and due to its simplicity it is presented as a suitable and sensitive method to investigate the capacity of biogenic amine production by LAB. Tyramine was the main amine formed by the LAB strains investigated. Enterococci, carnobacteria and some strains of lactobacilli, particularly of Lb. curvatus. Lb. brevis and Lb. buchneri, were the most intensive tyramine formers. Several strains of lactobacilli, Leuconostoc spp., Weissella spp. and pediococci did not show any potential to produce amines. Enterobacteriaceae were associated with cadaverine and putrescine formation. No significant histamine production could be detected for any of the strains tested.
Coda, Rossana; Pinto, Daniela; Gobbetti, Marco
A pool of selected lactic acid bacteria was used for the sourdough fermentation of various cereal flours with the aim of synthesizing antioxidant peptides. The radical-scavenging activity of water/salt-soluble extracts (WSE) from sourdoughs was significantly (P < 0.05) higher than that of chemically acidified doughs. The highest activity was found for whole wheat, spelt, rye, and kamut sourdoughs. Almost the same results were found for the inhibition of linoleic acid autoxidation. WSE were subjected to reverse-phase fast protein liquid chromatography. Thirty-seven fractions were collected and assayed in vitro. The most active fractions were resistant to further hydrolysis by digestive enzymes. Twenty-five peptides of 8 to 57 amino acid residues were identified by nano-liquid chromatography-electrospray ionization-tandem mass spectrometry. Almost all of the sequences shared compositional features which are typical of antioxidant peptides. All of the purified fractions showed ex vivo antioxidant activity on mouse fibroblasts artificially subjected to oxidative stress. This study demonstrates the capacity of sourdough lactic acid bacteria to release peptides with antioxidant activity through the proteolysis of native cereal proteins. PMID:22156436
Rizzello, Carlo Giuseppe; Lorusso, Anna; Russo, Vito; Pinto, Daniela; Marzani, Barbara; Gobbetti, Marco
Lactic acid bacteria strains, previously isolated from the same matrix, were used to ferment quinoa flour aiming at exploiting the antioxidant potential. As in vitro determined on DPPH and ABTS radicals, the scavenging activity of water/salt-soluble extracts (WSE) from fermented doughs was significantly (P<0.05) higher than that of non-inoculated doughs. The highest inhibition of linoleic acid autoxidation was found for the quinoa dough fermented with Lactobacillus plantarum T0A10. The corresponding WSE was subjected to Reverse Phase Fast Protein Liquid Chromatography, and 32 fractions were collected and subjected to in vitro assays. The most active fraction was resistant to further hydrolysis by digestive enzymes. Five peptides, having sizes from 5 to 9 amino acid residues, were identified by nano-Liquid Chromatography-Electrospray Ionisation-Mass Spectra/Mass Spectra. The sequences shared compositional features which are typical of antioxidant peptides. As shown by determining cell viability and radical scavenging activity (MTT and DCFH-DA assays, respectively), the purified fraction showed antioxidant activity on human keratinocytes NCTC 2544 artificially subjected to oxidative stress. This study demonstrated the capacity of autochthonous lactic acid bacteria to release peptides with antioxidant activity through proteolysis of native quinoa proteins. Fermentation of the quinoa flour with a selected starter might be considered suitable for novel applications as functional food ingredient, dietary supplement or pharmaceutical preparations.
Wang, Chung-Yi; Ng, Chang-Chai; Su, Hsuan; Tzeng, Wen-Sheng; Shyu, Yuan-Tay
The present study assesses the feasibility of noni as a raw substrate for the production of probiotic noni juice by lactic acid bacteria (Lactobacilluscasei and Lactobacillus plantarum) and bifidobacteria (Bifidobacteriumlongum). Changes in pH, acidity, sugar content, cell survival and antioxidant properties during fermentation were monitored. All tested strains grew well on noni juice, reaching nearly 10⁹ colony-forming units/ml after 48 h fermentation. L.casei produced less lactic acid than B.longum and L. plantarum. After 4 weeks of cold storage at 4°C, B.longum and L. plantarum survived under low-pH conditions in fermented noni juice. In contrast, L.casei exhibited no cell viability after 3 weeks. Moreover, noni juice fermented with B.longum had a high antioxidant capacity that did not differ significantly (P <0.05) from that of lactic acid bacteria. Finally, we found that B.longum and L. plantarum are optimal probiotics for fermentation with noni juice.
Barreau, C; Bimet, F; Kiredjian, M; Rouillon, N; Bizet, C
Forty-two clinical isolates were classified as Corynebacterium minutissimum, Corynebacterium striatum, and Corynebacterium CDC group I by the API Coryne system. The chemotaxonomic characteristics of the isolates were determined by thin-layer chromatographic analysis. Twenty-six isolates were found to have a type IV cell wall (meso-di-aminopimelic acid arabinose, galactose) but did not contain mycolic acids. These 26 isolates shared chemotaxonomic characteristics with those of mycolic acid-free reference strains (including the Corynebacterium amycolatum NCFB 2768 type strain, "Corynebacterium asperum," and coryneform CDC groups I2 and F2). The total protein profiles of the isolates determined by sodium dodecyl sulfate-polyacrylamide gel electrophoresis were similar to each other and to that of the C. amycolatum type strain. The profiles of the reference strains "Corynebacterium asperum" (CIP 100836, CIP 80.54, CIP 79.37, CIP 52.13), coryneform bacteria CDC groups I2 and F2 (CDC F5771, F5890, G723, G1970), and C. amycolatum were closely related. Thus, the mycolic acid-negative strains with a chemotype IV wall may belong to a single taxon. DNA hybridization studies could confirm this hypothesis. The present study shows the importance of chemotaxonomic analysis for verifying strain identifications and completing results from biochemical tests, particularly for coryneform bacteria. Images PMID:8370733
Vegas, Carlos; Mateo, Estibaliz; González, Angel; Jara, Carla; Guillamón, José Manuel; Poblet, Montse; Torija, Ma Jesús; Mas, Albert
The population dynamics of acetic acid bacteria in traditional vinegar production was determined in two independent vinegar plants at both the species and strain level. The effect of barrels made of four different woods upon the population dynamics was also determined. Acetic acid bacteria were isolated on solid media and the species were identified by RFLP-PCR of 16S rRNA genes and confirmed by 16S rRNA gene sequencing, while strains were typed by ERIC-PCR and (GTG)(5)-rep-PCR. The most widely isolated species was Acetobacter pasteurianus, which accounted for 100% of all the isolates during most of the acetification. Gluconacetobacter europaeus only appeared at any notable level at the end of the process in oak barrels from one vinegar plant. The various A. pasteurianus strains showed a clear succession as the concentration of acetic acid increased. In both vinegar plants the relative dominance of different strains was modified as the concentrations of acetic acid increased, and strain diversity tended to reduce at the end of the process.
Garofalo, Cristiana; Osimani, Andrea; Milanović, Vesna; Taccari, Manuela; Aquilanti, Lucia; Clementi, Francesca
Beer is one of the world's most ancient and widely consumed fermented alcoholic beverages produced with water, malted cereal grains (generally barley and wheat), hops, and yeast. Beer is considered an unfavorable substrate of growth for many microorganisms, however, there are a limited number of bacteria and yeasts, which are capable of growth and may spoil beer especially if it is not pasteurized or sterile-filtered as craft beer. The aim of this research study was to track beer spoilage lactic acid bacteria (LAB) inside a brewery and during the craft beer production process. To that end, indoor air and work surface samples, collected in the brewery under study, together with commercial active dry yeasts, exhausted yeasts, yeast pellet (obtained after mature beer centrifugation), and spoiled beers were analyzed through culture-dependent methods and PCR-DGGE in order to identify the contaminant LAB species and the source of contamination. Lactobacillus brevis was detected in a spoiled beer and in a commercial active dry yeast. Other LAB species and bacteria ascribed to Staphylococcus sp., Enterobaceriaceae, and Acetobacter sp. were found in the brewery. In conclusion, the PCR-DGGE technique coupled with the culture-dependent method was found to be a useful tool for identifying the beer spoilage bacteria and the source of contamination. The analyses carried out on raw materials, by-products, final products, and the brewery were useful for implementing a sanitization plan to be adopted in the production plant.
Lactic acid bacteria (LAB) have taken centre stage in perspectives of modern fermented food industry and probiotic based therapeutics. These bacteria encounter various stress conditions during industrial processing or in the gastrointestinal environment. Such conditions are overcome by complex molecular assemblies capable of synthesizing and/or metabolizing molecules that play a specific role in stress adaptation. Thiols are important class of molecules which contribute towards stress management in cell. Glutathione, a low molecular weight thiol antioxidant distributed widely in eukaryotes and Gram negative organisms, is present sporadically in Gram positive bacteria. However, new insights on its occurrence and role in the latter group are coming to light. Some LAB and closely related Gram positive organisms are proposed to possess glutathione synthesis and/or utilization machinery. Also, supplementation of glutathione in food grade LAB is gaining attention for its role in stress protection and as a nutrient and sulfur source. Owing to the immense benefits of glutathione, its release by probiotic bacteria could also find important applications in health improvement. This review presents our current understanding about the status of glutathione and its role as an exogenously added molecule in food grade LAB and closely related organisms. PMID:22920585
Schellenberger, Stefanie; Drake, Harold L; Kolb, Steffen
Herbicides have the potential to impair the metabolism of soil microorganisms. The current study addressed the toxic effect of bentazon and 4-chloro-2-methylphenoxyacetic acid on aerobic and anaerobic Bacteria that are involved in cellulose and cellobiose degradation in an agricultural soil. Aerobic saccharide degradation was reduced at concentrations of herbicides above environmental values. Microbial processes (e.g. fermentations, ferric iron reduction) that were linked to anaerobic cellulose and cellobiose degradation were reduced in the presence of both herbicides at concentrations above and at those that occur in crop field soil. 16S rRNA gene transcript numbers of total Bacteria, and selected bacterial taxa (Clostridia [Group I], Planctomycetaceae, and two uncultivated taxa of Bacteroidetes) decreased more in anoxic than in oxic cellulose-supplemented soil microcosms in the presence of both herbicides. Collectively, the results suggested that the metabolism of anaerobic cellulose-degrading Bacteria was impaired by typical in situ herbicide concentrations, whereas in situ concentrations did not impair metabolism of aerobic cellulose- and cellobiose-degrading soil Bacteria.
Stellato, Giuseppina; De Filippis, Francesca; La Storia, Antonietta; Ercolini, Danilo
Microbial contamination in food processing plants can play a fundamental role in food quality and safety. In this study, the microbiota in a dairy plant was studied by both 16S rRNA- and 26S rRNA-based culture-independent high-throughput amplicon sequencing. Environmental samples from surfaces and tools were studied along with the different types of cheese produced in the same plant. The microbiota of environmental swabs was very complex, including more than 200 operational taxonomic units with extremely variable relative abundances (0.01 to 99%) depending on the species and sample. A core microbiota shared by 70% of the samples indicated a coexistence of lactic acid bacteria with a remarkable level of Streptococcus thermophilus and possible spoilage-associated bacteria, including Pseudomonas, Acinetobacter, and Psychrobacter, with a relative abundance above 50%. The most abundant yeasts were Kluyveromyces marxianus, Yamadazyma triangularis, Trichosporon faecale, and Debaryomyces hansenii. Beta-diversity analyses showed a clear separation of environmental and cheese samples based on both yeast and bacterial community structure. In addition, predicted metagenomes also indicated differential distribution of metabolic pathways between the two categories of samples. Cooccurrence and coexclusion pattern analyses indicated that the occurrence of potential spoilers was excluded by lactic acid bacteria. In addition, their persistence in the environment can be helpful to counter the development of potential spoilers that may contaminate the cheeses, with possible negative effects on their microbiological quality.
Wu, Dan; Tong, Meiping; Kim, Hyunjung
The significance of perfluorooctanoic acid (PFOA) on the transport and deposition behaviors of bacteria (Gram-negative Escherichia coli and Gram-positive Bacillus subtilis) in quartz sand is examined in both NaCl and CaCl2 solutions at pH 5.6 by comparing both breakthrough curves and retained profiles with PFOA in solutions versus those without PFOA. All test conditions are found to be highly unfavorable for cell deposition regardless of the presence of PFOA; however, 7%-46% cell deposition is observed depending on the conditions. The cell deposition may be attributed to micro- or nanoscale roughness and/or to chemical heterogeneity of the sand surface. The results show that, under all examined conditions, PFOA in suspensions increases cell transport and decreases cell deposition in porous media regardless of cell type, presence or absence of extracellular polymeric substances, ionic strength, and ion valence. We find that the additional repulsion between bacteria and quartz sand caused by both acid-base interaction and steric repulsion as well as the competition for deposition sites on quartz sand surfaces by PFOA are responsible for the enhanced transport and decreased deposition of bacteria with PFOA in solutions.
Stellato, Giuseppina; De Filippis, Francesca; La Storia, Antonietta
Microbial contamination in food processing plants can play a fundamental role in food quality and safety. In this study, the microbiota in a dairy plant was studied by both 16S rRNA- and 26S rRNA-based culture-independent high-throughput amplicon sequencing. Environmental samples from surfaces and tools were studied along with the different types of cheese produced in the same plant. The microbiota of environmental swabs was very complex, including more than 200 operational taxonomic units with extremely variable relative abundances (0.01 to 99%) depending on the species and sample. A core microbiota shared by 70% of the samples indicated a coexistence of lactic acid bacteria with a remarkable level of Streptococcus thermophilus and possible spoilage-associated bacteria, including Pseudomonas, Acinetobacter, and Psychrobacter, with a relative abundance above 50%. The most abundant yeasts were Kluyveromyces marxianus, Yamadazyma triangularis, Trichosporon faecale, and Debaryomyces hansenii. Beta-diversity analyses showed a clear separation of environmental and cheese samples based on both yeast and bacterial community structure. In addition, predicted metagenomes also indicated differential distribution of metabolic pathways between the two categories of samples. Cooccurrence and coexclusion pattern analyses indicated that the occurrence of potential spoilers was excluded by lactic acid bacteria. In addition, their persistence in the environment can be helpful to counter the development of potential spoilers that may contaminate the cheeses, with possible negative effects on their microbiological quality. PMID:26341209
Riefler, R Guy; Krohn, Jeremy; Stuart, Ben; Socotch, Cheryl
This report describes a twenty month case study of a successive alkalinity producing system (SAPS) treating a strong acid mine drainage (AMD) source in Coshocton County, Ohio. Prior to the commencement of the project, a large volume of black amorphous sludge had accumulated in several of the constructed wetlands. The sludge was found to be 43% organic, with very high concentrations of sulfur, iron, aluminum, and acidity. Based on several biological, physical, and chemical analyses, the sludge was determined to be an anaerobic biofilm with a large population of sulfur-reducing bacteria and a high mineral content due to the formation of iron sulfide and aluminum precipitates. On average the system performed well, generating 26 kg CaCO3/d of alkalinity and capturing 5.0 kg/d of iron and 1.7 kg/d of aluminum. Several simple performance analysis tools were presented in this work. By comparing the pollutant influent and effluent loading, it was determined that the SAPS was performing at capacity and over the past year increased effluent concentrations were due to increased influent loadings and not system deterioration. Further, by performing a detailed cell-by-cell loading analysis of multiple chemical components, the alkalinity generated by limestone dissolution and by sulfate reduction was determined. Interestingly, 61% of the alkalinity generation in the vertical flow wetlands was due to sulfur-reducing bacteria activity, indicating that sulfur-reducing bacteria may play a more significant role in SAPS than expected.
Ripamonti, Barbara; Agazzi, Alessandro; Bersani, Carla; De Dea, Paola; Pecorini, Chiara; Pirani, Silvia; Rebucci, Raffaella; Savoini, Giovanni; Stella, Simone; Stenico, Alberta; Tirloni, Erica; Domeneghini, Cinzia
The selection of promising specific species of lactic acid bacteria with potential probiotic characteristics is of particular interest in producing multi species-specific probiotic adjuncts in veal calves rearing. The aim of the present work was to select and evaluate in vitro the functional activity of lactic acid bacteria, Bifidobacterium longum and Bacillus coagulans strains isolated from veal calves in order to assess their potential use as multi species-specific probiotics for veal calves. For this purpose, bacterial strains isolated from faeces collected from 40 healthy 50-day-calves, were identified by RiboPrinter and 16s rRNA gene sequence. The most frequent strains belonged to the species B. longum, Streptococcus bovis, Lactobacillus animalis and Streptococcus macedonicus. Among these, 7 strains were chosen for testing their probiotic characteristics in vitro. Three strains, namely L. animalis SB310, Lactobacillus paracasei subsp. paracasei SB137 and B. coagulans SB117 showed varying individual but promising capabilities to survive in the gastrointestinal tract, to adhere, to produce antimicrobial compounds. These three selected species-specific bacteria demonstrated in vitro, both singularly and mixed, the functional properties needed for their use as potential probiotics in veal calves.
Borges, Anabela; Saavedra, Maria J; Simões, Manuel
The activity of two phenolic acids, gallic acid (GA) and ferulic acid (FA) at 1000 μg ml(-1), was evaluated on the prevention and control of biofilms formed by Escherichia coli, Pseudomonas aeruginosa, Staphylococcus aureus and Listeria monocytogenes. In addition, the effect of the two phenolic acids was tested on planktonic cell susceptibility, bacterial motility and adhesion. Biofilm prevention and control were tested using a microtiter plate assay and the effect of the phenolic acids was assessed on biofilm mass (crystal violet staining) and on the quantification of metabolic activity (alamar blue assay). The minimum bactericidal concentration for P. aeruginosa was 500 μg ml(-1) (for both phenolic acids), whilst for E. coli it was 2500 μg ml(-1) (FA) and 5000 μg ml(-1) (GA), for L. monocytogenes it was >5000 μg ml(-1) (for both phenolic acids), and for S. aureus it was 5000 μg ml(-1) (FA) and >5000 μg ml(-1) (GA). GA caused total inhibition of swimming (L. monocytogenes) and swarming (L. monocytogenes and E. coli) motilities. FA caused total inhibition of swimming (L. monocytogenes) and swarming (L. monocytogenes and E. coli) motilities. Colony spreading of S. aureus was completely inhibited by FA. The interference of GA and FA with bacterial adhesion was evaluated by the determination of the free energy of adhesion. Adhesion was less favorable when the bacteria were exposed to GA (P. aeruginosa, S. aureus and L. monocytogenes) and FA (P. aeruginosa and S. aureus). Both phenolics had preventive action on biofilm formation and showed a higher potential to reduce the mass of biofilms formed by the Gram-negative bacteria. GA and FA promoted reductions in biofilm activity >70% for all the biofilms tested. The two phenolic acids demonstrated the potential to inhibit bacterial motility and to prevent and control biofilms of four important human pathogenic bacteria. This study also emphasizes the potential of phytochemicals as an emergent source of biofilm
Kim, Jungkyu; Elsnab, John; Johnson, Michael; Gale, Bruce K.
A fully integrated microfluidic system was developed and incorporates an EC-MWCNT (electrochemical multiwalled carbon nanotube) sensor for the detection of bacteria. Sample metering, reagent metering and delivery was implemented with microvalves and pumps embedded inside the microfluidic system. The nucleic acid extraction was performed using microchannels controlled using automated platforms and a disposable microfluidic silica cartridge. The target samples were flowed and hybridized with probe ssDNA (single strand DNA) across the MWCNT-EC sensor (built on a silicon chip), which was embedded in a microfluidic cell. The 9-pad sensor was scanned before and after hybridization to measure the quantity of RNA (Ribonucleic acid) bound to the array surface. A rapid and accurate sample-in answer-out nucleic acid system was realized with automated volume metering, microfluidic sample preparation, and integrated nano-biosensors.
Hronská, Helena; Tokošová, Silvia; Pilniková, Anna; Krištofíková, Ľudmila; Rosenberg, Michal
The bacterial strains of the genus Nocardia were used for the bioconversion of fumaric acid to L-malic acid. The ability of the bacterial strain Nocardia sp. CCM 4837/A to produce L-malic acid from fumaric acid was investigated under various conditions. The optimal temperature for the bioconversion was approximately 37 °C, and the optimal pH was around 8.0. The addition of an inductor (fumarate salt) to the fermentation medium was necessary to enhance enzyme activity. The presence of detergent Triton X-100 (0.02-0.1 %) in the reaction mixture rapidly increased the enzyme activity of fumarase. The specific fumarase activity of intact cells Nocardia sp. CCM 4837/A increased from 2.8 to 75 U/mg after optimising the experimental conditions described here. Pretreatment of the Nocardia cells with malonate was not necessary because succinate was not detected as a by-product under our experimental conditions.
Blasco, Lucía; Ferrer, Sergi; Pardo, Isabel
A rapid method for the identification of lactic acid bacteria (LAB) from wine has been developed. This method is based on fluorescence in situ hybridisation (FISH), using fluorescent oligonucleotide probes, homologous to 16S rDNA of those species of LAB commonly found in wines. The protocol for the specific detection of these bacteria was established through the hybridisation of 36 reference strains. The specificity of the probes was evaluated by using pure cultures. Probes were used to identify species in different wines, making it evident that direct identification and quantification from natural samples without culturing is also possible. The results show that FISH is a promising technique for the rapid identification of LAB, allowing positive identification in a few hours (4-16 h).
van Zyl, Winschau F; Deane, Shelly M; Dicks, Leon M T
Bioluminescence (BLI) and fluorescence imaging (FI) allow for non-invasive detection of viable microorganisms from within living tissue and are thus ideally suited for in vivo probiotic studies. Highly sensitive optical imaging techniques detect signals from the excitation of fluorescent proteins, or luciferase-catalyzed oxidation reactions. The excellent relation between microbial numbers and photon emission allow for quantification of tagged bacteria in vivo with extreme accuracy. More information is gained over a shorter period compared to traditional pre-clinical animal studies. The review summarizes the latest advances in in vivo bioluminescence and fluorescence imaging and points out the advantages and limitations of different techniques. The practical application of BLI and FI in the tracking of lactic acid bacteria in animal models is addressed. PMID:26516656
Ortiz, Maria Eugenia; Bleckwedel, Juliana; Raya, Raúl R; Mozzi, Fernanda
Polyols such as mannitol, erythritol, sorbitol, and xylitol are naturally found in fruits and vegetables and are produced by certain bacteria, fungi, yeasts, and algae. These sugar alcohols are widely used in food and pharmaceutical industries and in medicine because of their interesting physicochemical properties. In the food industry, polyols are employed as natural sweeteners applicable in light and diabetic food products. In the last decade, biotechnological production of polyols by lactic acid bacteria (LAB) has been investigated as an alternative to their current industrial production. While heterofermentative LAB may naturally produce mannitol and erythritol under certain culture conditions, sorbitol and xylitol have been only synthesized through metabolic engineering processes. This review deals with the spontaneous formation of mannitol and erythritol in fermented foods and their biotechnological production by heterofermentative LAB and briefly presented the metabolic engineering processes applied for polyol formation.
Perez, Rodney H; Zendo, Takeshi; Sonomoto, Kenji
Bacteriocins are heat-stable ribosomally synthesized antimicrobial peptides produced by various bacteria, including food-grade lactic acid bacteria (LAB). These antimicrobial peptides have huge potential as both food preservatives, and as next-generation antibiotics targeting the multiple-drug resistant pathogens. The increasing number of reports of new bacteriocins with unique properties indicates that there is still a lot to learn about this family of peptide antibiotics. In this review, we highlight our system of fast tracking the discovery of novel bacteriocins, belonging to different classes, and isolated from various sources. This system employs molecular mass analysis of supernatant from the candidate strain, coupled with a statistical analysis of their antimicrobial spectra that can even discriminate novel variants of known bacteriocins. This review also discusses current updates regarding the structural characterization, mode of antimicrobial action, and biosynthetic mechanisms of various novel bacteriocins. Future perspectives and potential applications of these novel bacteriocins are also discussed.
Bacteriocins are heat-stable ribosomally synthesized antimicrobial peptides produced by various bacteria, including food-grade lactic acid bacteria (LAB). These antimicrobial peptides have huge potential as both food preservatives, and as next-generation antibiotics targeting the multiple-drug resistant pathogens. The increasing number of reports of new bacteriocins with unique properties indicates that there is still a lot to learn about this family of peptide antibiotics. In this review, we highlight our system of fast tracking the discovery of novel bacteriocins, belonging to different classes, and isolated from various sources. This system employs molecular mass analysis of supernatant from the candidate strain, coupled with a statistical analysis of their antimicrobial spectra that can even discriminate novel variants of known bacteriocins. This review also discusses current updates regarding the structural characterization, mode of antimicrobial action, and biosynthetic mechanisms of various novel bacteriocins. Future perspectives and potential applications of these novel bacteriocins are also discussed. PMID:25186038
Andújar-Ortiz, Inmaculada; Pozo-Bayón, Maria Angeles; García-Ruiz, Almudena; Moreno-Arribas, M Victoria
The role of specific components from inactive dry yeast preparations widely used in winemaking on the growth of three representative wine lactic acid bacteria (Oenococcus oeni, Lactobacillus hilgardii and Pediococcus pentosaceus) has been studied. A pressure liquid extraction technique using solvents of different polarity was employed to obtain extracts with different chemical composition from the inactive dry yeast preparations. Each of the extracts was assayed against the three lactic acid bacteria. Important differences in the effect of the extracts on the growth of the bacteria were observed, which depended on the solvent employed during the extraction, on the type of commercial preparations and on the lactic acid bacteria species. The extracts that exhibited the most different activity were chemically characterized in amino acids, free monosaccharides, monosaccharides from polysaccharides, fatty acids and volatile compounds. In general, specific amino acids and monosaccharides were related to a stimulating effect whereas fatty acid composition and likely some volatile compounds seemed to show an inhibitory effect on the growth of the lactic acid bacteria. These results may provide novel and useful information in trying to obtain better and more specific formulations of winemaking inactive dry yeast preparations.
Elizaquível, Patricia; Pérez-Cataluña, Alba; Yépez, Alba; Aristimuño, Cecilia; Jiménez, Eugenia; Cocconcelli, Pier Sandro; Vignolo, Graciela; Aznar, Rosa
The diversity of lactic acid bacteria (LAB) associated with chicha, a traditional maize-based fermented alcoholic beverage from Northwestern Argentina, was analyzed using culture-dependent and culture-independent approaches. Samples corresponding to 10 production steps were obtained from two local producers at Maimará (chicha M) and Tumbaya (chicha T). Whereas by culture-dependent approach a few number of species (Lactobacillus plantarum and Weissella viridescens in chicha M, and Enterococcus faecium and Leuconostoc mesenteroides in chicha T) were identified, a higher quantitative distribution of taxa was found in both beverages by pyrosequencing. The relative abundance of OTUs was higher in chicha M than in chicha T; six LAB genera were common for chicha M and T: Enterococcus, Lactococcus, Streptococcus, Weissella, Leuconostoc and Lactobacillus while Pediococcus only was detected in chicha M. Among the 46 identified LAB species, those of Lactobacillus were dominant in both chicha samples, exhibiting the highest diversity, whereas Enterococcus and Leuconostoc were recorded as the second dominant genera in chicha T and M, respectively. Identification at species level showed the predominance of Lb. plantarum, Lactobacillus rossiae, Leuconostoc lactis and W. viridescens in chicha M while Enterococcus hirae, E. faecium, Lc. mesenteroides and Weissella confusa predominated in chicha T samples. In parallel, when presumptive LAB isolates (chicha M: 146; chicha T: 246) recovered from the same samples were identified by ISR-PCR and RAPD-PCR profiles, species-specific PCR and 16S rRNA gene sequencing, most of them were assigned to the Leuconostoc genus (Lc. mesenteroides and Lc. lactis) in chicha M, Lactobacillus, Weissella and Enterococcus being also present. In contrast, chicha T exhibited the presence of Enterococcus and Leuconostoc, E. faecium being the most representative species. Massive sequencing approach was applied for the first time to study the diversity and
Nowak, Adriana; Śliżewska, Katarzyna; Otlewska, Anna
Dietary components such as lactic acid bacteria (LAB) and prebiotics can modulate the intestinal microbiota and are thought to be involved in the reduction of colorectal cancer risk. The presented study measured, using the comet assay, the antigenotoxic activity of both probiotic and non-probiotic LAB, as well as some prebiotics and the end-products of their fermentation, against fecal water (FW). The production of short chain fatty acids by the bacteria was quantified using HPLC. Seven out of the ten tested viable strains significantly decreased DNA damage induced by FW. The most effective of them were Lactobacillus mucosae 0988 and Bifidobacterium animalis ssp. lactis Bb-12, leading to a 76% and 80% decrease in genotoxicity, respectively. The end-products of fermentation of seven prebiotics by Lactobacillus casei DN 114-001 exhibited the strongest antigenotoxic activity against FW, with fermented inulin reducing genotoxicity by 75%. Among the tested bacteria, this strain produced the highest amounts of butyrate in the process of prebiotic fermentation, and especially from resistant dextrin (4.09 μM/mL). Fermented resistant dextrin improved DNA repair by 78% in cells pre-treated with 6.8 μM methylnitronitrosoguanidine (MNNG). Fermented inulin induced stronger DNA repair in cells pre-treated with mutagens (FW, 25 μM hydrogen peroxide, or MNNG) than non-fermented inulin, and the efficiency of DNA repair after 120 min of incubation decreased by 71%, 50% and 70%, respectively. The different degrees of genotoxicity inhibition observed for the various combinations of bacteria and prebiotics suggest that this effect may be attributable to carbohydrate type, SCFA yield, and the ratio of the end-products of prebiotic fermentation.
Manzoor, Asma; Ul-Haq, Ikram; Baig, Shahjhan; Qazi, Javed Iqbal; Seratlic, Sanja
Background: Antibiotic resistance represents a serious global health threat to public health, so infections such as pneumonia and urinary tract infection (UTI) are becoming harder to treat. Therefore, it is necessary to develop an action plan to restrain the problem of antibiotic resistance. One approach in UTI control could be the use of lactobacilli because these indigenous inhabitants in human intestine have been found to play an important role in protecting the host from various infections. Objectives: We sought to check the efficacy of locally isolated Lactobacillus species to eradicate antibiotic-resistant pathogenic bacteria causing UTI. Materials and Methods: Lactic acid bacteria isolated from spoiled fruits and vegetables and grown in MRS medium were screened against multi-drug-resistant Candida albicans, Klebsiella pneumoniae, Pseudomonas aeruginosa, Escherichia coli, and Enterococcus fecalis. Results: Fifty-four lactic acid bacteria were isolated from spoiled fruits and vegetables, of which 11 Gram-positive and catalase-negative Lactobacillus isolates were identified by carbohydrate assimilation profiles as Lactobacillus acidophilus, L. paracasei, L. delbrueckii, L. casei, L. helveticus, L. brevis, L. salivarius, L. fermentum, L. rhamnosus, L. animalis, and L. plantarum. The latter organism had the highest abundance of all the samples, so its isolates were also verified through 16S rRNA gene sequencing. The isolated Lactobacilli were screened against multi-drug-resistant uropathogens, viz. C. albicans, P. aeruginosa, K. pneumoniae, E. fecalis, and E. coli. The growth inhibition zone (GIZ) was over 10 mm against all the uropathogenic test organisms, where L. fermentum and L. plantarum strains demonstrated remarkable inhibitory activities against E. coli and E. faecalis, with a GIZ up to 28 mm. The susceptibility test to 16 antibiotics showed multidrug resistance (3 to 5 antibiotics) among all the tested uropathogens. Conclusions: The obtained results
Russell, James B.
Bacteria from the bovine rumen capable of reducing trans-aconitate to tricarballylate were enriched in an anaerobic chemostat containing rumen fluid medium and aconitate. After 9 days at a dilution rate of 0.07 h−1, the medium was diluted and plated in an anaerobic glove box. Three types of isolates were obtained from the plates (a crescent-shaped organism, a pleomorphic rod, and a spiral-shaped organism), and all three produced tricarballylate in batch cultures that contained glucose and trans-aconitate. In glucose-limited chemostats (0.10 h−1), trans-aconitate reduction was associated with a decrease in the amount of reduced products formed from glucose. The crescent-shaped organism produced less propionate, the pleomorphic rod produced less ethanol, and the spiral made less succinate and possibly H2. Aconitate reduction by the pleomorphic rod and the spiral organism was associated with a significant increase in cellular dry matter. Experiments with stock cultures of predominant rumen bacteria indicated that Selenomonas ruminantium, a species taxonomically similar to the crescent-shaped isolate, was an active reducer of trans-aconitate. Strains of Bacteroides ruminicola, Butyrivibrio fibrisolvens, and Megasphaera elsdenii produced little if any tricarballylate. Wolinella succinogenes produced some tricarballylate. Based on its stability constant for magnesium (Keq = 115), tricarballylate could be a factor in the hypomagnesemia that leads to grass tetany. Images PMID:16346691
Khalil, Maha A.; El-Sheekh, Mostafa M.; El-Adawi, Hala I.; El-Deeb, Nehal M.; Hussein, Mohamed Z.
Background: Probiotic delivery systems are widely used nutraceutical products for the supplementation of natural intestinal flora. These delivery systems vary greatly in the effectiveness to exert health benefits for a patient. This study focuses on providing probiotic living cells with a physical barrier against adverse environmental conditions. Materials and Methods: Microencapsulation of the selected lactic acid bacteria (LAB) using chitosan and alginate was performed. Physical examination of the formulated LAB microcapsules was observed using phase contrast inverted microscope and scanning electron microscope (SEM). Finally, the survival of microencapsulated and noncapsulated bacteria was cheeked in the simulated human gastric tract (GT). The potential antimicrobial activity of the most potent microencapsulated LAB strain was in vivo evaluated in rabbit models. Results: Microencapsulated L. plantarum, L. acidophilus, and L. bulgaricus DSMZ 20080 were loaded with 1.03 × 1010 CFU viable bacteria/g, 1.9 × 1010 CFU viable bacteria/g, and 5.5 × 109 CFU viable bacteria/g, respectively. The survival of microencapsulated cells was significantly higher than that of the free cells after exposure to simulated gastric juice (SGJ) at pH 2. Additionally, in simulated small intestine juice (SSJ), larger amounts of the selected LAB cells were found, whereas in simulated colon juice (SCJ), the released LAB reached the maximum counts. In vivo results pointed out that an 8-week supplementation with a triple therapy of a microencapsulated L. plantarum, L. acidophilus, and L. bulgaricus DSMZ 20080 might be able to reduce H. pylori. Conclusion: Microencapsulated probiotics could possibly compete with and downregulate H. pylori infection in humans. PMID:26929759
Clementi, Francesca; Aquilanti, Lucia
The worldwide use, and misuse, of antibiotics for about sixty years in the so-called antibiotic era, has been estimated in some one to ten million tons, a relevant part of which destined for non-therapeutic purposes such as growth promoting treatments for livestock or crop protection. As highly adaptable organisms, bacteria have reacted to this dramatic change in their environment by developing several well-known mechanisms of antibiotic resistance and are becoming increasingly resistant to conventional antibiotics. In recent years, commensal bacteria have become a cause of concern since they may act as reservoirs for the antibiotic resistance genes found in human pathogens. In particular, the food chain has been considered the main route for the introduction of animal and environment associated antibiotic resistant bacteria into the human gastrointestinal tract (GIT) where these genes may be transferred to pathogenic and opportunistic bacteria. As fundamental microbial communities in a large variety of fermented foods and feed, the anaerobe facultative, aerotolerant lactic acid bacteria (LAB) are likely to play a pivotal role in the resistance gene exchange occurring in the environment, food, feed and animal and human GIT. Therefore their antibiotic resistance features and their genetic basis have recently received increasing attention. The present article summarises the results of the latest studies on the most typical genera belonging to the low G + C branch of LAB. The evolution of the criteria established by European regulatory bodies to ensure a safe use of microorganisms in food and feed, including the assessment of their antibiotic resistance is also reviewed.
Rudnick, M B; van Veen, J A; de Boer, W
Mycophagous (=fungus feeding) soil bacteria of the genus Collimonas have been shown to colonize and grow on hyphae of different fungal hosts as the only source of energy and carbon. The ability to exploit fungal nutrient resources might require a strategy for collimonads to sense fungi in the soil matrix. Oxalic acid is ubiquitously secreted by soil fungi, serving different purposes. In this study, we investigated the possibility that collimonads might use oxalic acid secretion to localize a fungal host and move towards it. We first confirmed earlier indications that collimonads have a very limited ability to use oxalic acid as growth substrate. In a second step, with using different assays, we show that oxalic acid triggers bacterial movement in such a way that accumulation of cells can be expected at micro-sites with high free oxalic acid concentrations. Based on these observations we propose that oxalic acid functions as a signal molecule to guide collimonads to hyphal tips, the mycelial zones that are most sensitive for mycophagous bacterial attack.
Rasimus, Stiina; Kolari, Marko; Rita, Hannu; Hoornstra, Douwe; Salkinoja-Salonen, Mirja
Biofilms cause runnability problems in paper machines and are therefore controlled with biocides. Peracetic acid is usually effective in preventing bulky biofilms. This study investigated the microbiological status of a paper machine where low concentrations (≤ 15 ppm active ingredient) of peracetic acid had been used for several years. The paper machine contained a low amount of biofilms. Biofilm-forming bacteria from this environment were isolated and characterized by 16S rRNA gene sequencing, whole-cell fatty acid analysis, biochemical tests, and DNA fingerprinting. Seventy-five percent of the isolates were identified as members of the subclades Sphingomonas trueperi and S. aquatilis, and the others as species of the genera Burkholderia (B. cepacia complex), Methylobacterium, and Rhizobium. Although the isolation media were suitable for the common paper machine biofoulers Deinococcus, Meiothermus, and Pseudoxanthomonas, none of these were found, indicating that peracetic acid had prevented their growth. Spontaneous, irreversible loss of the ability to form biofilm was observed during subculturing of certain isolates of the subclade S. trueperi. The Sphingomonas isolates formed monoculture biofilms that tolerated peracetic acid at concentrations (10 ppm active ingredient) used for antifouling in paper machines. High pH and low conductivity of the process waters favored the peracetic acid tolerance of Sphingomonas sp. biofilms. This appears to be the first report on sphingomonads as biofilm formers in warm water using industries.
Araque, Isabel; Reguant, Cristina; Rozès, Nicolas; Bordons, Albert
Wine can contain trace amounts of ethyl carbamate (EC), a carcinogen formed when ethanol reacts with carbamyl compounds such as citrulline. EC is produced from arginine by lactic acid bacteria (LAB), e.g., Lactobacillus and Pediococcus. Although the amounts of EC in wine are usually negligible, over the last few years there has been a slight but steady increase, as climate change has increased temperatures and alcohol levels have become proportionately higher, both of which favor EC formation. In this study, resting cells of LAB were used to evaluate the effects of ethanol, glucose, malic acid, and low pH on the ability of non-oenococcal strains of these bacteria to degrade arginine and excrete citrulline. Malic acid was found to clearly inhibit arginine consumption in all strains. The relation between citrulline produced and arginine consumed was clearly higher in the presence of ethanol (10-12%) and at low pH (3.0), which is consistent with both the decreased amount of ornithine produced from arginine and the reduction in arginine degradation. In L. brevis and L. buchneri strains isolated from wine and beer, respectively, the synthesis of citrulline from arginine was highest.
Petri, A; Pfannebecker, J; Fröhlich, J; König, H
The microflora of must and wine consists of yeasts, acetic acid bacteria and lactic acid bacteria (LAB). The latter group plays an important role for wine quality. The malolactic fermentation carried out by LAB leads to deacidification and stabilisation of wines. Nevertheless, LAB are often associated with wine spoilage. They are mainly responsible for the formation of biogenic amines. Furthermore, some strains produce exopolysaccharide slimes, acetic acid, diacetyl and other off-flavours. In this context a better monitoring of the vinification process is crucial to improve wine quality. Moreover, a lot of biodiversity studies would also profit from a fast and reliable identification method. In this study, we propose a species-specific multiplex PCR system for a rapid and simultaneous detection of 13 LAB species, frequently occurring in must or wine: Lactobacillus brevis, Lb. buchneri, Lb. curvatus, Lb. hilgardii, Lb. plantarum, Leuconostoc mesenteroides, Oenococcus oeni, Pediococcus acidilactici, P. damnosus, P. inopinatus, P. parvulus, P. pentosaceus and Weissella paramesenteroides.
Chen, Yang; Bai, Ye; Li, Dongsheng; Wang, Chao; Xu, Ning; Hu, Yong
Acetic acid bacteria (AAB) are important microorganisms in the vinegar industry. However, AAB have to tolerate the presence of ethanol and high temperatures, especially in submerged fermentation (SF), which inhibits AAB growth and acid yield. In this study, seven AAB that are tolerant to temperatures above 40 °C and ethanol concentrations above 10% (v/v) were isolated from Chinese vinegar Pei. All the isolated AAB belong to Acetobacter pasteurianus according to 16S rDNA analysis. Among all AAB, AAB4 produced the highest acid yield under high temperature and ethanol test conditions. At 4% ethanol and 30-40 °C temperatures, AAB4 maintained an alcohol-acid transform ratio of more than 90.5 %. High alcohol-acid transform ratio was still maintained even at higher temperatures, namely, 87.2, 77.1, 14.5 and 2.9% at 41, 42, 43 and 44 °C, respectively. At 30 °C and different initial ethanol concentrations (4-10%), the acid yield by AAB4 increased gradually, although the alcohol-acid transform ratio decreased to some extent. However, 46.5, 8.7 and 0.9% ratios were retained at ethanol concentrations of 11, 12 and 13%, respectively. When compared with AS1.41 (an AAB widely used in China) using a 10 L fermentor, AAB4 produced 42.0 g/L acetic acid at 37 °C with 10% ethanol, whereas AS1.41 almost stopped producing acetic acid. In conclusion, these traits suggest that AAB4 is a valuable strain for vinegar production in SF.
Elshaghabee, Fouad M. F.; Bockelmann, Wilhelm; Meske, Diana; de Vrese, Michael; Walte, Hans-Georg; Schrezenmeir, Juergen; Heller, Knut J.
To gain some specific insight into the roles microorganisms might play in non-alcoholic fatty liver disease (NAFLD), some intestinal and lactic acid bacteria and one yeast (Anaerostipes caccae, Bacteroides thetaiotaomicron, Bifidobacterium longum, Enterococcus fecalis, Escherichia coli, Lactobacillus acidophilus, Lactobacillus fermentum, Lactobacillus plantarum, Weissella confusa, Saccharomyces cerevisiae) were characterized by high performance liquid chromatography for production of ethanol when grown on different carbohydrates: hexoses (glucose and fructose), pentoses (arabinose and ribose), disaccharides (lactose and lactulose), and inulin. Highest amounts of ethanol were produced by S. cerevisiae, L. fermentum, and W. confusa on glucose and by S. cerevisiae and W. confusa on fructose. Due to mannitol-dehydrogenase expressed in L. fermentum, ethanol production on fructose was significantly (P < 0.05) reduced. Pyruvate and citrate, two potential electron acceptors for regeneration of NAD+/NADP+, drastically reduced ethanol production with acetate produced instead in L. fermentum grown on glucose and W. confusa grown on glucose and fructose, respectively. In fecal slurries prepared from feces of four overweight volunteers, ethanol was found to be produced upon addition of fructose. Addition of A. caccae, L. acidophilus, L. fermentum, as well as citrate and pyruvate, respectively, abolished ethanol production. However, addition of W. confusa resulted in significantly (P < 0.05) increased production of ethanol. These results indicate that microorganisms like W. confusa, a hetero-fermentative, mannitol-dehydrogenase negative lactic acid bacterium, may promote NAFLD through ethanol produced from sugar fermentation, while other intestinal bacteria and homo- and hetero-fermentative but mannitol-dehydrogenase positive lactic acid bacteria may not promote NAFLD. Also, our studies indicate that dietary factors interfering with gastrointestinal microbiota and microbial
Biscola, V; Todorov, S D; Capuano, V S C; Abriouel, H; Gálvez, A; Franco, B D G M
A Lactococcus lactis subsp. lactis strain (L. lactis 69) capable to produce a heat-stable bacteriocin was isolated from charqui, a Brazilian fermented, salted and sun-dried meat product. The bacteriocin inhibited, in vitro, Listeria monocytogenes, Staphylococcus aureus, several lactic acid bacteria isolated from foods and spoilage halotolerant bacteria isolated from charqui. The activity of the bacteriocin was not affected by pH (2.0-10.0), heating (100 °C), and chemical agents (1% w/v). Treatment of growing cells of L. monocytogenes ScottA with the cell-free supernatant of L. lactis 69 resulted in complete cell inactivation. L. lactis 69 harbored the gene for the production of a nisin-like bacteriocin, and the amino acid sequence of the active peptide was identical to sequences previously described for nisin Z. However, differences were observed regarding the leader peptide. Besides, the isolate was able to survive and produce bacteriocins in culture medium with NaCl content up to 20%, evidencing a potential application as an additional hurdle in the preservation of charqui.
Miao, Yuzhi; Xu, Hui; Fei, Baojin; Qiao, Dairong; Cao, Yi
The major objective of this study was to engineer lactic acid bacteria to produce the enzyme phytase from a gene native to Bacillus subtilis GYPB04. The phytase gene (phyC) of B. subtilis GYPB04 was cloned into the plasmid pMG36e for expression in Lactococcus lactis. The enzyme activity in L. lactis cultured in GM17 broth was 20.25 U/mL at 36°C. The expressed phytase was characterized as active in a pH range of 2.0-9.0 at a temperature range of 20-80°C, with an optimum pH of 5.5-6.5 and temperature of 60°C. When cultured in food-grade milk broth, the transformed L. lactis grew to an OD(600 nm) value of 1.05 and had a phytase yield of 13.58 U/mL. In same broth under optimized conditions for cell growth and phytase production, the transformant reached an OD(600 nm) value of 1.68 and a phytase yield of 42.12 U/mL, representing approximately 1.6-fold and 3.1-fold increases, respectively, compared to growth in natural milk broth. Fermentation was scaled to 5 L under optimized conditions, and product analysis revealed a final OD(600 nm) value of 1.89 and an extracellular enzyme activity of 24.23 U/mL. The results of this study may be used in the dairy fermentation industry for the development of functional, healthy yogurts and other fermented dairy foods that provide both active phytase and viable probiotics to the consumer.
Blanco, M. Mar; López-Campos, Guillermo H.; Cutuli, M. Teresa; Fernández-Garayzábal, José F.
The present work describes the molecular characterization of five circular plasmids found in the human clinical strain Lactococcus garvieae 21881. The plasmids were designated pGL1-pGL5, with molecular sizes of 4,536 bp, 4,572 bp, 12,948 bp, 14,006 bp and 68,798 bp, respectively. Based on detailed sequence analysis, some of these plasmids appear to be mosaics composed of DNA obtained by modular exchange between different species of lactic acid bacteria. Based on sequence data and the derived presence of certain genes and proteins, the plasmid pGL2 appears to replicate via a rolling-circle mechanism, while the other four plasmids appear to belong to the group of lactococcal theta-type replicons. The plasmids pGL1, pGL2 and pGL5 encode putative proteins related with bacteriocin synthesis and bacteriocin secretion and immunity. The plasmid pGL5 harbors genes (txn, orf5 and orf25) encoding proteins that could be considered putative virulence factors. The gene txn encodes a protein with an enzymatic domain corresponding to the family actin-ADP-ribosyltransferases toxins, which are known to play a key role in pathogenesis of a variety of bacterial pathogens. The genes orf5 and orf25 encode two putative surface proteins containing the cell wall-sorting motif LPXTG, with mucin-binding and collagen-binding protein domains, respectively. These proteins could be involved in the adherence of L. garvieae to mucus from the intestine, facilitating further interaction with intestinal epithelial cells and to collagenous tissues such as the collagen-rich heart valves. To our knowledge, this is the first report on the characterization of plasmids in a human clinical strain of this pathogen. PMID:22768237
Lahtvee, Petri-Jaan; Seiman, Andrus; Arike, Liisa; Adamberg, Kaarel; Vilu, Raivo
Protein turnover plays an important role in cell metabolism by regulating metabolic fluxes. Furthermore, the energy costs for protein turnover have been estimated to account for up to a third of the total energy production during cell replication and hence may represent a major limiting factor in achieving either higher biomass or production yields. This work aimed to measure the specific growth rate (μ)-dependent abundance and turnover rate of individual proteins, estimate the ATP cost for protein production and turnover, and compare this with the total energy balance and other maintenance costs. The lactic acid bacteria model organism Lactococcus lactis was used to measure protein turnover rates at μ = 0.1 and 0.5 h(-1) in chemostat experiments. Individual turnover rates were measured for ~75% of the total proteome. On average, protein turnover increased by sevenfold with a fivefold increase in growth rate, whilst biomass yield increased by 35%. The median turnover rates found were higher than the specific growth rate of the bacterium, which suggests relatively high energy consumption for protein turnover. We found that protein turnover costs alone account for 38 and 47% of the total energy produced at μ = 0.1 and 0.5 h(-1), respectively, and gene ontology groups Energy metabolism and Translation dominated synthesis costs at both growth rates studied. These results reflect the complexity of metabolic changes that occur in response to changes in environmental conditions, and signify the trade-off between biomass yield and the need to produce ATP for maintenance processes.
Landete, José María; Pardo, Isabel; Ferrer, Sergi
The ability of wine lactic acid bacteria to produce tyramine and phenylethylamine was investigated by biochemical and genetic methods. An easy and accurate plate medium was developed to detect tyramine-producer strains, and a specific PCR assay that detects the presence of tdc gene was employed. All strains possessing the tdc gene were shown to produce tyramine and phenylethylamine. Wines containing high quantities of tyramine and phenylethylamine were found to contain Lactobacillus brevis or Lactobacillus hilgardii. The main tyramine producer was L. brevis. The ability to produce tyramine was absent or infrequent in the rest of the analysed wine species.
Pothakos, Vasileios; Devlieghere, Frank; Villani, Francesco; Björkroth, Johanna; Ercolini, Danilo
Lactic acid bacteria (LAB) constitute a heterogeneous group that has been widely associated with fresh meat and cooked meat products. They represent a controversial cohort of microbial species that either contribute to spoilage through generation of offensive metabolites and the subsequent organoleptic downgrading of meat or serve as bioprotective agents with strains of certain species causing unperceivable or no alterations. Therefore, significant distinction among biotypes is substantiated by studies determining spoilage potential as a strain-specific trait corroborating the need to revisit the concept of spoilage.
Bai, He; Kang, Yong; Quan, Hongen; Han, Yang; Sun, Jiao; Feng, Ying
In order to treat acid mine drainage (AMD) effectively using sulfate-reducing bacteria (SRB) at high concentration of sulfate and heavy metals, Fe(0) was added to enhance the activity of SRB. When AMD was treated by SRB and Fe(0) at 25 °C, more than 61% of sulfate was removed and the effluent pH was improved from 2.75 to 6.20 during the operation. Cu(2+) was removed effectively with the removal efficiency at 99%, while only 86% of Fe(2+) was removed during the AMD treatment, without conspicuous change of Mn(2+) in the effluent in the process.
Ohirchuk, K S; Poltavs'ka, O A; Kovalenko, N K
Antagonistic activity of 74 cultures of lactic acid bacteria, isolated from healthy and osteoporotic women-patients aged 50-79 years, has been studied. It has been shown that the inhibitory effect of the strain studied was independent of the health of women (control group of women or patients with osteoporosis), but had strain specificity. Seventeen most active strains of lactobacilli, which showed the highest inhibitory activity against B. cereus, P. aeruginosa, P. vulgaris were selected. Only 6 strains of lactobacillus demonstrated specific antagonistic activity against the test-strains.
Terahara, M; Kurama, S; Takemoto, N
Ether extracts of lactic acid bacteria were analyzed for prevention of the oxidation of erythrocyte membrane and human low-density lipoprotein in vivo. Streptococcus thermophilus 1131 and Lactobacillus delbrueckii subsp. bulgaricus 2038, yogurt starters, were chosen as test-strains, and ether extracts of these cultures were used as samples. Both strain 1131 and strain 2038 produced radical scavengers and inhibited oxidation of erythrocyte membranes and low-density lipoproteins. The antioxidative activity of strain 2038 was higher than that of strain 1131.
Lu, W W; Wang, T; Wang, Y; Xin, M; Kong, J
Enterotoxigenic Escherichia coli (ETEC) infection is the major cause of diarrhea in neonatal piglets. The fimbriae as colonizing factor in the pathogenesis of ETEC constitute a primary target for vaccination against ETEC. Lactic acid bacteria (LAB) are attractive tools to deliver antigens at the mucosal level. With the safety of genetically modified LAB in mind, a food-grade secretion vector (pALRc or pALRb) was constructed with DNA entirely from LAB, including the replicon, promoter, signal peptide, and selection marker alanine racemase gene (alr). To evaluate the feasibility of the system, the nuclease gene (nuc) from Staphylococcus aureus was used as a reporter to be expressed in both Lactococcus lactis and Lactobacillus casei. Subsequently, the extracellular secretion of the fimbrial adhesin FaeG of ETEC was confirmed by Western blot analysis. These results showed that this food-grade expression system has potential as the delivery vehicle for the safe use of genetically modified LAB for the development of vaccines against ETEC infection.
Ladero, Victor; Rattray, Fergal P.; Mayo, Baltasar; Martín, María Cruz; Fernández, María; Alvarez, Miguel A.
Lactococcus lactis is a prokaryotic microorganism with great importance as a culture starter and has become the model species among the lactic acid bacteria. The long and safe history of use of L. lactis in dairy fermentations has resulted in the classification of this species as GRAS (General Regarded As Safe) or QPS (Qualified Presumption of Safety). However, our group has identified several strains of L. lactis subsp. lactis and L. lactis subsp. cremoris that are able to produce putrescine from agmatine via the agmatine deiminase (AGDI) pathway. Putrescine is a biogenic amine that confers undesirable flavor characteristics and may even have toxic effects. The AGDI cluster of L. lactis is composed of a putative regulatory gene, aguR, followed by the genes (aguB, aguD, aguA, and aguC) encoding the catabolic enzymes. These genes are transcribed as an operon that is induced in the presence of agmatine. In some strains, an insertion (IS) element interrupts the transcription of the cluster, which results in a non-putrescine-producing phenotype. Based on this knowledge, a PCR-based test was developed in order to differentiate nonproducing L. lactis strains from those with a functional AGDI cluster. The analysis of the AGDI cluster and their flanking regions revealed that the capacity to produce putrescine via the AGDI pathway could be a specific characteristic that was lost during the adaptation to the milk environment by a process of reductive genome evolution. PMID:21803900
Yagnik, Bhrugu; Padh, Harish; Desai, Priti
Use of food grade Lactococcus lactis (L. lactis) is fast emerging as a safe alternative for delivery of DNA vaccine. To attain efficient DNA delivery, L. lactis, a non-invasive bacterium is converted to invasive strain either by expressing proteins like Internalin A (InlA) or Fibronectin binding protein A (FnBPA) or through chemical treatments. However the safety status of invasive L. lactis is questionable. In the present report, we have shown that non-invasive L. lactis efficiently delivered the newly constructed reporter plasmid pPERDBY to mammalian cells without any chemical enhancers. The salient features of the vector are; I) Ability to replicate in two different hosts; Escherichia coli (E. coli) and Lactic Acid Bacteria (LAB), II) One of the smallest reporter plasmid for DNA vaccine, III) Enhanced Green Fluorescence Protein (EGFP) linked to Multiple Cloning Site (MCS), IV) Immunostimulatory CpG motifs functioning as an adjuvant. Expression of EGFP in pPERDBY transfected CHO-K1 and Caco-2 cells demonstrates its functionality. Non-invasive r-L. lactis was found efficient in delivering pPERDBY to Caco-2 cells. The in vitro data presented in this article supports the hypothesis that in the absence of invasive proteins or relevant chemical treatment, L. lactis was found efficient in delivering DNA to mammalian cells.
Hanniffy, Sean B; Philo, Mark; Peláez, Carmen; Gasson, Michael J; Requena, Teresa; Martínez-Cuesta, M C
The conversion of methionine to volatile sulfur compounds (VSCs) is of great importance in flavor formation during cheese ripening and is the focus of biotechnological approaches toward flavor improvement. A synthetic mgl gene encoding methionine-gamma-lyase (MGL) from Brevibacterium linens BL2 was cloned into a Lactococcus lactis expression plasmid under the control of the nisin-inducible promoter PnisA. When expressed in L. lactis and purified as a recombinant protein, MGL was shown to degrade L-methionine as well as other sulfur-containing compounds such as L-cysteine, L-cystathionine, and L-cystine. Overproduction of MGL in recombinant L. lactis also resulted in an increase in the degradation of these compounds compared to the wild-type strain. Importantly, gas chromatography-mass spectrometry analysis identified considerably higher formation of methanethiol (and its oxidized derivatives dimethyl disulfide and dimethyl trisulfide) in reactions containing either purified protein, whole cells, or cell extracts from the heterologous L. lactis strain. This is the first report of production of MGL from B. linens in L. lactis. Given their significance in cheese flavor development, the use of lactic acid bacteria with enhanced VSC-producing abilities could be an efficient way to enhance cheese flavor development.
Background Hydrogen peroxide (H2O2) produced by vaginal lactobacilli is generally believed to protect against bacteria associated with bacterial vaginosis (BV), and strains of lactobacilli that can produce H2O2 are being developed as vaginal probiotics. However, evidence that led to this belief was based in part on non-physiological conditions, antioxidant-free aerobic conditions selected to maximize both production and microbicidal activity of H2O2. Here we used conditions more like those in vivo to compare the effects of physiologically plausible concentrations of H2O2 and lactic acid on a broad range of BV-associated bacteria and vaginal lactobacilli. Methods Anaerobic cultures of seventeen species of BV-associated bacteria and four species of vaginal lactobacilli were exposed to H2O2, lactic acid, or acetic acid at pH 7.0 and pH 4.5. After two hours, the remaining viable bacteria were enumerated by growth on agar media plates. The effect of vaginal fluid (VF) on the microbicidal activities of H2O2 and lactic acid was also measured. Results Physiological concentrations of H2O2 (< 100 μM) failed to inactivate any of the BV-associated bacteria tested, even in the presence of human myeloperoxidase (MPO) that increases the microbicidal activity of H2O2. At 10 mM, H2O2 inactivated all four species of vaginal lactobacilli but only one of seventeen species of BV-associated bacteria. Moreover, the addition of just 1% vaginal fluid (VF) blocked the microbicidal activity of 1 M H2O2. In contrast, lactic acid at physiological concentrations (55-111 mM) and pH (4.5) inactivated all the BV-associated bacteria tested, and had no detectable effect on the vaginal lactobacilli. Also, the addition of 10% VF did not block the microbicidal activity of lactic acid. Conclusions Under optimal, anaerobic growth conditions, physiological concentrations of lactic acid inactivated BV-associated bacteria without affecting vaginal lactobacilli, whereas physiological concentrations of H2O2
Pontes, Daniela; Innocentin, Silvia; Del Carmen, Silvina; Almeida, Juliana Franco; Leblanc, Jean-Guy; de Moreno de Leblanc, Alejandra; Blugeon, Sébastien; Cherbuy, Claire; Lefèvre, François; Azevedo, Vasco; Miyoshi, Anderson; Langella, Philippe; Chatel, Jean-Marc
Lactococci are noninvasive lactic acid bacteria frequently used as protein delivery vectors and, more recently, as DNA delivery vehicles. We previously showed that Lactococcus lactis (LL) expressing the Fibronectin-Binding Protein A of Staphylococcus aureus (LL-FnBPA+) showed higher internalization rates in vitro in Caco-2 cells than the native (wt) lactococci and were able to deliver a eukaryotic Green Fluorescent Protein (GFP) expression plasmid in 1% of human Caco-2 cells. Here, using the bovine beta-lactoglobulin (BLG), one of the major cow's milk allergen, and GFP we characterized the potential of LL-FnBPA+ as an in vivo DNA vaccine delivery vehicle. We first showed that the invasive strain LL-FnBPA+ carrying the plasmid pValac:BLG (LL-FnBPA+ BLG) was more invasive than LL-BLG and showed the same invasivity as LL-FnBPA+. Then we demonstrated that the Caco-2 cells, co-incubated with LL-FnBPA+ BLG produced up to 30 times more BLG than the Caco-2 cells co-incubated with the non invasive LL-BLG. Using two different gene reporters, BLG and GFP, and two different methods of detection, EIA and fluorescence microscopy, we showed in vivo that: i) in order to be effective, LL-FnBPA+ required a pre-coating with Fetal Calf Serum before oral administration; ii) plasmid transfer occurred in enterocytes without regard to the strains used (invasive or not); iii) the use of LL-FnBPA+ increased the number of mice producing BLG, but not the level of BLG produced. We thus confirmed the good potential of invasive recombinant lactic acid bacteria as DNA delivery vector in vivo.
Ringø, E; Bendiksen, H R; Wesmajervi, M S; Olsen, R E; Jansen, P A; Mikkelsen, H
The present study reports the effect of excessive handling stress and starvation on the lactic acid bacteria associated with the digestive tract of Atlantic salmon (Salmo salar L.). A relatively low population level (approximately 2 x 103 bacteria per gram wet tissue) of viable adherent heterotrophic bacteria was associated with the digestive tract (foregut, midgut and hindgut). Of the 752 bacterial isolates isolated from diet, water and the digestive tract, 201 isolates belonged to the carnobacteria. Of these isolates, one from the diet, one from the rearing water and 80 from the gastrointestinal tract, were further identified on the basis of 16S rDNA sequence analysis. All these isolates were identified as being Carnobacterium piscicola-like. Daily repeated stress and starvation of the fish over 11 d had no influence on the total culturable bacterial numbers or population level of C. piscicola associated with the digestive tract. C. piscicola-like isolates colonizing the various intestinal regions (foregut, midgut and hindgut) were also screened for their ability to produce growth inhibitory compounds active against the fish pathogen Aeromonas salmonicida. Of the 199 C. piscicola isolates tested, 139 inhibited growth of the pathogen.
Grimoud, Julien; Durand, Henri; Courtin, Céline; Monsan, Pierre; Ouarné, Françoise; Theodorou, Vassilia; Roques, Christine
Probiotics and prebiotics have been demonstrated to positively modulate the intestinal microflora and could promote host health. Although some studies have been performed on combinations of probiotics and prebiotics, constituting synbiotics, results on the synergistic effects tend to be discordant in the published works. The first aim of our study was to screen some lactic acid bacteria on the basis of probiotic characteristics (resistance to intestinal conditions, inhibition of pathogenic strains). Bifidobacterium was the most resistant genus whereas Lactobacillus farciminis was strongly inhibited. The inhibitory effect on pathogen growth was strain dependent but lactobacilli were the most effective, especially L. farciminis. The second aim of the work was to select glucooligosaccharides for their ability to support the growth of the probiotics tested. We demonstrated the selective fermentability of oligodextran and oligoalternan by probiotic bacteria, especially the bifidobacteria, for shorter degrees of polymerisation and absence of metabolism by pathogenic bacteria. Thus, the observed characteristics confer potential prebiotic properties on these glucooligosaccharides, to be further confirmed in vivo, and suggest some possible applications in synbiotic combinations with the selected probiotics. Furthermore, the distinctive patterns of the different genera suggest a combination of lactobacilli and bifidobacteria with complementary probiotic effects in addition to the prebiotic ones. These associations should be further evaluated for their synbiotic effects through in vitro and in vivo models.
Ren, Chengcheng; Zhang, Qiuxiang; de Haan, Bart J.; Zhang, Hao; Faas, Marijke M.; de Vos, Paul
Although many lactic acid bacteria (LAB) influence the consumer’s immune status it is not completely understood how this is established. Bacteria-host interactions between bacterial cell-wall components and toll-like receptors (TLRs) have been suggested to play an essential role. Here we investigated the interaction between LABs with reported health effects and TLRs. By using cell-lines expressing single or combination of TLRs, we show that LABs can signal via TLR-dependent and independent pathways. The strains only stimulated and did not inhibit TLRs. We found that several strains such as L. plantarum CCFM634, L. plantarum CCFM734, L. fermentum CCFM381, L. acidophilus CCFM137, and S. thermophilus CCFM218 stimulated TLR2/TLR6. TLR2/TLR6 is essential in immune regulatory processes and of interest for prevention of diseases. Specificity of the TLR2/TLR6 stimulation was confirmed with blocking antibodies. Immunomodulatory properties of LABs were also studied by assessing IL-10 and IL-6 secretion patterns in bacteria-stimulated THP1-derived macrophages, which confirmed species and strain specific effects of the LABs. With this study we provide novel insight in LAB specific host-microbe interactions. Our data demonstrates that interactions between pattern recognition receptors such as TLRs is species and strain specific and underpins the importance of selecting specific strains for promoting specific health effects. PMID:27708357
Zhou, J S; Shu, Q; Rutherfurd, K J; Prasad, J; Gopal, P K; Gill, H S
Three potentially probiotic lactic acid bacteria (LAB) strains, Lactobacillus rhamnosus HN001 (DR20(TM)), Lb. acidophilus HN017 and Bifidobacterium lactis HN019 (DR10()), have recently been identified and characterized. The present study was designed to evaluate the acute oral toxicity of these strains to mice, and also to investigate bacterial translocation and gut mucosal pathology in BALB/c mice fed HN019, HN001 or HN017 for 8 consecutive days at a high dose of 10(11)cfu/mouse/day. Results showed that these probiotic strains had no adverse effect on general health status, feed intake, body weight gain and intestinal mucosal morphology (villus height, crypt depth, epithelial cell height and mucosal thickness). No viable bacteria were recovered from blood and tissue samples (mesenteric lymph nodes, liver and spleen) of mice, and no treatment-associated illness or death was observed. According to these results, the oral LD(50) of HN019, HN001 and HN017 is more than 50g/kg/day for mice, and their acceptable daily intake (ADI) value is 35g dry bacteria per day for a 70-kg person. This suggests that the probiotic strains HN019, HN001 and HN017 are non-pathogenic and likely to be safe for human consumption.
Aswathy, Ravindran Girija; Ismail, Bindhumol; John, Rojan Pappy; Nampoothiri, Kesavan Madhavan
Lactic acid bacteria were isolated from fermented vegetables, sour dough, milk products, sheep and human excreta. The newly isolated cultures were evaluated for a number of probiotic characteristics like bile salt resistance, salt tolerance in general, survival in low pH, hydrophobicity of the cell surface, resistance to low phenol concentration, antimicrobial activity and susceptibility pattern against vancomycin and erythromycin. The selected cultures were further screened for their ability to produce the nutraceticals such as folic acid and exopolysaccharide (EPS). Two potent isolates, CB2 (from cabbage) and SD2 (from sour dough) were found to produce both extracellular and intracellular folate. One of the isolates from yogurt (MC-1) and the one from whey (W3) produced significant amount of EPS with a maximum production of 8.79 +/- 0.05 g/l by MC-1.
Santos, A O; Ávila, C L S; Schwan, R F
The objective of this study was to select lactic acid bacteria (LAB) strains isolated from silage and assess their effect on the quality of maize silage. The LAB strains were inoculated into aqueous extract obtained from maize to evaluate their production of metabolites and pH reduction. The ability to inhibit the pathogenic and silage-spoilage microorganisms' growth was evaluated. Nine LAB strains that showed the best results were assessed in polyvinyl chloride experimental silos. The inoculation of the LAB strains influenced the concentration of lactic and acetic acids and the diversity of Listeria. The inoculation of silages with Lactobacillus buchneri (UFLA SLM11 and UFLA SLM103 strains) resulted in silages with greater LAB populations and improvements after aerobic exposure. The UFLA SLM11 and SLM103 strains identified as L. buchneri showed to be promising in the treatment of maize silage.
Wang, Zhenyu; Zong, Haiying; Zheng, Hao; Liu, Guocheng; Chen, Lei; Xing, Baoshan
Adding biochar into soils has potential to manipulate soil nitrification process due to its impacts on nitrogen (N) cycling, however, the exact mechanisms underlying the alteration of nitrification process in soils are still not clear. Nitrification in an acidic orchard soil amended with peanut shell biochar (PBC) produced at 400 °C was investigated. Nitrification was weakened by PBC addition due to the decreased NH4(+)-N content and reduced ammonia-oxidizing bacteria (AOB) abundance in PBC-amended soils. Adding phenolic compounds (PHCs) free biochar (PBC-P) increased the AOB abundance and the DGGE band number, indicating that PHCs remaining in the PBC likely reduced AOB abundance and diversity. However, PBC addition stimulated rape growth and increased N bioavailability. Overall, adding PBC could suppress the nitrification process and improve N bioavailability in the agricultural soils, and thus possibly mitigate the environmental negative impacts and improving N use efficiency in the acidic soils added with N fertilizer.
Ji, Keunho; Jang, Na Young; Kim, Young Tae
The purpose of this study was to investigate lactic acid bacteria with antioxidative and probiotic activities isolated from Korean healthy infant feces and kimchi. Isolates A1, A2, S1, S2, and S3 were assigned to Lactobacillus sp. and isolates A3, A4, E1, E2, E3, and E4 were assigned to Leuconostoc sp. on the basis of their physiological properties and 16S ribosomal DNA sequence analysis. Most strains were confirmed as safe bioresources through nonhemolytic activities and non-production of harmful enzymes such as β-glucosidase, β- glucuronidase and tryptophanase. The 11 isolates showed different resistance to acid and bile acids. In addition, they exhibited antibacterial activity against foodborne bacteria, especially Bacillus cereus, Listeria monocytogenes, and Escherichia coli. Furthermore, all strains showed significantly high levels of hydrophobicity. The antioxidant effects of culture filtrates of the 11 strains included 2,2-diphenyl-1-picryl-hydrazyl (DPPH) radical scavenging capacity, 2.2'- azino-bis (2-ethylbenzthiazoline-6-sulfonic acid) (ABTS) radical cation scavenging activity, and superoxide dismutase activity. The results revealed that most of the culture filtrates have effective scavenging activity for DPPH and ABTS radicals. All strains appeared to have effective superoxide dismutase activity. In conclusion, the isolated strains A1, A3, S1, and S3 have significant probiotic activities applicable to the development of functional foods and health-related products. These strains might also contribute to preventing and controlling several diseases associated with oxidative stress, when used as probiotics.
Damayanti, Ema; Ratisiwi, Febiyani Ndaru; Istiqomah, Lusty; Sembiring, Langkah; Febrisiantosa, Andi
The objective of this study was to determine the potential of LAB with phytate degrading activity from fermented traditional food grain-based and legume-based. Lactic acid bacteria were isolated from different sources of traditional fermented food from Gunungkidul Yogyakarta Indonesia such as gembus tempeh (tofu waste), soybean tempeh, lamtoro tempeh (Leucaena bean) and kara tempeh. Isolation of LAB was performed using Total Plate Count (TPC) on de Man Rogosa Sharpe Agar (MRSA) medium supplemented with CaCO3. They were screened for their ability to degrade myo-inositol hexaphosphate or IP6 by using qualitative streak platemethod with modified de Man Rogosa-MorpholinoPropanesulfonic Acid Sharpe (MRS-MOPS) medium contained sodium salt of phytic acid as substrate and cobalt chloride staining (plate assay) method. The selected isolates were further assayed for phytase activities using quantitative method with spectrophotometer and the two selected isolates growth were optimized. Furthermore, thhe isolates that shown the highest phytase activity was characterized and identified using API 50 CH kitand 16S rRNA gene sequencing. The results showed that there were 18 LAB isolates obtained from samplesand 13 isolates were able to degrade sodium phytate based on qualitative screening. According to quantitative assay, the highest phytate degrading activities were found in TG-2(23.562 U/mL) and TG-1 (19.641 U/mL) isolated from gembus tempeh. The phytate activity of TG-2 was optimum at 37 °C with agitation, while the phytate activity of TG-1 was optimum at 45 °C without agitation. Characterization and identification of TG-2 isolate with the highest phytate degrading activity using API 50 CH and 16S rRNA showed that TG-2had homology with Lactobacillus fermentum. It could be concluded that LAB from from fermented traditional food grain-based and legume-based produced the extracellular phytase. Keywords: lactic acid bacteria, tempeh, phytatedegrading activity
Martin, S A; Jenkins, T C
The objective of this study was to identify environmental factors that influence conjugated linoleic acid (CLA) and trans-C18:1 fatty acid production by mixed ruminal bacteria. Ruminal contents were collected from a 600-kg ruminally fistulated Hereford steer maintained on pasture. Mixed ruminal bacteria were obtained by differential centrifugation under anaerobic conditions and added to a basal medium that contained a commercial emulsified preparation of soybean oil and a mixture of soluble carbohydrates (cellobiose, glucose, maltose, and xylose). Culture samples were collected from batch culture incubations at 0, 2, 4, 6, 8, 12, 24, 26, 28, 30, 32, and 48 h. Continuous culture incubations were conducted at dilution rates of 0.05 and 0.10 h(-1) with extracellular pH values of 5.5 and 6.5, and 0.5 and 1.0 g/L of mixed soluble carbohydrates. Culture samples were obtained from the culture vessel once steady-state conditions had been achieved. In batch culture, trans-C18:1 concentrations increased over time and reached a maximum at 48 h. Little CLA was produced during the first 8 h, but cis-9, trans-11 CLA concentrations remained high between 24 and 30 h. When mixed ruminal bacteria were maintained in continuous culture on 0.5 g/L of mixed soluble carbohydrates, concentrations of trans-C18:1 and cis-9, trans-11 CLA were reduced (P < 0.05) at a dilution rate of 0.05 h(-1) and an extracellular pH of 5.5. Similar effects were also observed when 1.0 g/L of mixed soluble carbohydrates was used. When extracellular pH was lowered to 5.0, neither trans-C18:1 or CLA isomers were detected. In conclusion, our results suggest that culture pH appears to have the most influence on the production of trans-C18:1 and CLA isomers by mixed ruminal bacteria.
The agar diffusion assay was used to examine the effect of pH on the ability of alkaline salts of three fatty acids (FA) to inhibit growth of bacteria associated with poultry processing. FA solutions were prepared by dissolving 0.5 M concentrations of caprylic, capric, or lauric acid in separate ali...
Salma, Mohammad; Rousseaux, Sandrine; Sequeira-Le Grand, Anabelle; Alexandre, Hervé
In this study we report for the first time a rapid, efficient and cost-effective method for the enumeration of lactic acid bacteria (LAB) in wine. Indeed, up to now, detection of LAB in wine, especially red wine, was not possible. Wines contain debris that cannot be separated from bacteria using flow cytometry (FCM). Furthermore, the dyes tested in previous reports did not allow an efficient staining of bacteria. Using FCM and a combination of BOX/PI dyes, we were able to count bacteria in wines. The study was performed in wine inoculated with Oenococcus oeni (10(6) CFU ml(-1)) stained with either FDA or BOX/PI and analyzed by FCM during the malolactic fermentation (MLF). The analysis show a strong correlation between the numbers of BOX/PI-stained cells determined by FCM and the cell numbers determined by plate counts (red wine: R (2) ≥ 0.97, white wine R (2) ≥ 0.965). On the other hand, we found that the enumeration of O. oeni labeled with FDA was only possible in white wine (R (2) ≥ 0.97). Viable yeast and LAB populations can be rapidly discriminated and quantified in simultaneous malolactic-alcoholic wine fermentations using BOX/PI and scatter parameters in a one single measurement. This rapid procedure is therefore a suitable method for monitoring O. oeni populations during winemaking, offers a detection limit of <10(4) CFU ml(-1) and can be considered a useful method for investigating the dynamics of microbial growth in wine and applied for microbiological quality control in wineries.
Drozdz, Iwona; Makarewicz, Malgorzata; Tuszyński, Tadeusz
The aim of this study was to identify various bacteria isolated from grapes and their wines. Additionally we investigated the capacity of lactic acid bacteria for microbiological deacidification of wines produced in Poland. We have identified Oenococcus oeni, Lactobacillus acidophilus and Lactobacillus delbrueckii. During the microbial deacidification process, we observed decreases of total acidity and increases of volatile acidity, with statistically significant changes noted for O. oeni in Marechal Foch and Seyval Blanc, and for Lb. acidophilus in Frontenac. On the other hand, a statistically significant increase in pH was observed in Marechal Foch and Seyval Blanc following deacidification by O. oeni.
Endo, Akihito; Salminen, Seppo
Fructophilic lactic acid bacteria (FLAB) are a specific group of lactic acid bacteria (LAB) characterized and described only recently. They prefer fructose as growth substrate and inhabit only fructose-rich niches. Honeybees are high-fructose-consuming insects and important pollinators in nature, but reported to be decreasing in the wild. In the present study, we analyzed FLAB microbiota in honeybees, larvae, fresh honey and bee pollen. A total of 66 strains of LAB were isolated from samples using a selective isolation technique for FLAB. Surprisingly, all strains showed fructophilic characteristics. The 66 strains and ten FLAB strains isolated from flowers in a separate study were genotypically separated into six groups, four of which being identified as Lactobacillus kunkeei and two as Fructobacillus fructosus. One of the L. kunkeei isolates showed antibacterial activity against Melissococcus plutonius, a causative pathogen of European foulbrood, this protection being attributable to production of an antibacterial peptide or protein. Culture-independent analysis suggested that bee products and larvae contained simple Lactobacillus-group microbiota, dominated by L. kunkeei, although adult bees carried a more complex microbiota. The findings clearly demonstrate that honeybees and their products are rich sources of FLAB, and FLAB are potential candidates for future bee probiotics.
Ňancucheo, Ivan; Johnson, D. Barrie
Summary Two continuous‐flow bench‐scale bioreactor systems populated by mixed communities of acidophilic sulfate‐reducing bacteria were constructed and tested for their abilities to promote the selective precipitation of transition metals (as sulfides) present in synthetic mine waters, using glycerol as electron donor. The objective with the first system (selective precipitation of copper from acidic mine water containing a variety of soluble metals) was achieved by maintaining a bioreactor pH of ∼2.2–2.5. The second system was fed with acidic (pH 2.5) synthetic mine water containing 3 mM of both zinc and ferrous iron, and varying concentrations (0.5–30 mM) of aluminium. Selective precipitation of zinc sulfide was possible by operating the bioreactor at pH 4.0 and supplementing the synthetic mine water with 4 mM glycerol. Analysis of the microbial populations in the bioreactors showed that they changed with varying operational parameters, and novel acidophilic bacteria (including one sulfidogen) were isolated from the bioreactors. The acidophilic sulfidogenic bioreactors provided ‘proof of principle’ that segregation of metals present in mine waters is possible using simple online systems within which controlled pH conditions are maintained. The modular units are versatile and robust, and involve minimum engineering complexity. PMID:21895996
Capozzi, Vittorio; Russo, Pasquale; Dueñas, María Teresa; López, Paloma; Spano, Giuseppe
Wheat contains various essential nutrients including the B group of vitamins. However, B group vitamins, normally present in cereals-derived products, are easily removed or destroyed during milling, food processing or cooking. Lactic acid bacteria (LAB) are widely used as starter cultures for the fermentation of a large variety of foods and can improve the safety, shelf life, nutritional value, flavor and overall quality of the fermented products. In this regard, the identification and application of strains delivering health-promoting compounds is a fascinating field. Besides their key role in food fermentations, several LAB found in the gastrointestinal tract of humans and animals are commercially used as probiotics and possess generally recognized as safe status. LAB are usually auxotrophic for several vitamins although certain strains of LAB have the capability to synthesize water-soluble vitamins such as those included in the B group. In recent years, a number of biotechnological processes have been explored to perform a more economical and sustainable vitamin production than that obtained via chemical synthesis. This review article will briefly report the current knowledge on lactic acid bacteria synthesis of vitamins B2, B11 and B12 and the potential strategies to increase B-group vitamin content in cereals-based products, where vitamins-producing LAB have been leading to the elaboration of novel fermented functional foods. In addition, the use of genetic strategies to increase vitamin production or to create novel vitamin-producing strains will be also discussed.
Guan, Ling; Shiiya, Ayaka; Hisatomi, Shihoko; Fujii, Kunihiko; Nonaka, Masanori; Harada, Naoki
Diphenylarsinic acid (DPAA) is often found as a toxic intermediate metabolite of diphenylchloroarsine or diphenylcyanoarsine that were produced as chemical warfare agents and were buried in soil after the World Wars. In our previous study Guan et al. (J Hazard Mater 241-242:355-362, 2012), after application of sulfate and carbon sources, anaerobic transformation of DPAA in soil was enhanced with the production of diphenylthioarsinic acid (DPTAA) as a main metabolite. This study aimed to isolate and characterize anaerobic soil microorganisms responsible for the metabolism of DPAA. First, we obtained four microbial consortia capable of transforming DPAA to DPTAA at a high transformation rate of more than 80% after 4 weeks of incubation. Sequencing for the bacterial 16S rRNA gene clone libraries constructed from the consortia revealed that all the positive consortia contained Desulfotomaculum acetoxidans species. In contrast, the absence of dissimilatory sulfite reductase gene (dsrAB) which is unique to sulfate-reducing bacteria was confirmed in the negative consortia showing no DPAA reduction. Finally, strain DEA14 showing transformation of DPAA to DPTAA was isolated from one of the positive consortia. The isolate was assigned to D. acetoxidans based on the partial 16S rDNA sequence analysis. Thionation of DPAA was also carried out in a pure culture of a known sulfate-reducing bacterial strain, Desulfovibrio aerotolerans JCM 12613(T). These facts indicate that sulfate-reducing bacteria are microorganisms responsible for the transformation of DPAA to DPTAA under anaerobic conditions.
Zhang, Mingliang; Wang, Haixia; Han, Xuemei
Novel immobilized sulfate-reducing bacteria (SRB) beads were prepared for the treatment of synthetic acid mine drainage (AMD) containing high concentrations of Fe, Cu, Cd and Zn using up-flow anaerobic packed-bed bioreactor. The tolerance of immobilized SRB beads to heavy metals was significantly enhanced compared with that of suspended SRB. High removal efficiencies of sulfate (61-88%) and heavy metals (>99.9%) as well as slightly alkaline effluent pH (7.3-7.8) were achieved when the bioreactor was fed with acidic influent (pH 2.7) containing high concentrations of multiple metals (Fe 469 mg/L, Cu 88 mg/L, Cd 92 mg/L and Zn 128 mg/L), which showed that the bioreactor filled with immobilized SRB beads had tolerance to AMD containing high concentrations of heavy metals. Partially decomposed maize straw was a carbon source and stabilizing agent in the initial phase of bioreactor operation but later had to be supplemented by a soluble carbon source such as sodium lactate. The microbial community in the bioreactor was characterized by denaturing gradient gel electrophoresis (DGGE) and sequencing of partial 16S rDNA genes. Synergistic interaction between SRB (Desulfovibrio desulfuricans) and co-existing fermentative bacteria could be the key factor for the utilization of complex organic substrate (maize straw) as carbon and nutrients source for sulfate reduction.
Mundt, J. Orvin; Graham, Wanda F.; McCarty, I. E.
The frequency and levels of population of the spherical lactic acid-producing bacteria were determined on raw and processed yellow summer and zucchini squash, a variety of greens, green beans, okra, southern peas, and butter and lima beans, and on fresh cucumbers and corn flowers. Six taxa occurred consistently: Leuconostoc mesenteroides, yellow-pigmented streptococci, Streptococcus faecium, Aerococcus viridans, and S. faecalis and S. faecalis var. liquefaciens. The same taxa occurred with the same order of frequency on processed, frozen vegetables, but with a marked decrease in the occurrence of S. faecalis var. liquefaciens. S. lactis, S. cremoris, S. equinus, S. bovis, and pediococci were isolated infrequently. No other member of the viridans group of the streptococci and no member of the pyogenic group was isolated. Approximately 88% of the cultures were identified. Total counts of the lactic-acid-producing bacteria rarely exceeded 105 per gram of sample, and there was a reduction by 90% during the second year of study, probably because of drought. Only one bacterial species was found on 40% of the raw and 34% of the processed vegetable samples. Two or more species or taxa were present on the remainder of 153 raw and 56 processed vegetable samples. A. viridans was present on squash, greens, okra, and southern peas, and its frequency of occurrence on vegetables suggests that plants are its natural habitat. PMID:16349739
Kubota, Hiromi; Senda, Shouko; Nomura, Nobuhiko; Tokuda, Hajime; Uchiyama, Hiroo
We investigated the formation of biofilms by 3 type strains of lactic acid bacteria (LAB), Lactobacillus plantarum, Lactobacillus brevis and Lactobacillus fructivorans, as representatives of LAB that cause food deterioration or contamination. Lactobacillus plantarum subsp. plantarum JCM1149 and Lactobacillus brevis JCM1059 appeared to adhere and accumulate on glass cover slips. Lactobacillus fructivorans JCM1117 cells made thin cellophane-like biofilms, and most of the biofilm cells became longer than the planktonic cells. We tested the resistance of biofilm and planktonic L. plantarum subsp. plantarum JCM1149 cells to acetic acid and ethanol, which strongly inhibit the growth of bacteria and are important in food preservation. The biofilm cells were more resistant than the planktonic cells and the surfaces of the treated planktonic cells were badly damaged, whereas those of the biofilm cells were only slightly damaged. We isolated 43 LAB from onions and the biofolm cells of an isolate, L. plantarum M606 also had high resistance. These results demonstrate the significance of studying biofilms of LAB in the food industry.
Mathias, S.P.; Rosenthal, A.; Gaspar, A.; Aragão, G.M.F.; Slongo-Marcusi, A.
High hydrostatic pressure (HHP) has been investigated and industrially applied to extend shelf life of meat-based products. Traditional ham packaged under microaerophilic conditions may sometimes present high lactic acid bacteria population during refrigerated storage, which limits shelf life due to development of unpleasant odor and greenish and sticky appearance. This study aimed at evaluating the shelf life of turkey ham pressurized at 400 MPa for 15 min and stored at 4, 8 and 12 °C, in comparison to the non pressurized product. The lactic acid bacteria population up to 107 CFU/g of product was set as the criteria to determine the limiting shelf life According to such parameter the pressurized sample achieved a commercial viability within 75 days when stored at 4 °C while the control lasted only 45 days. Predictive microbiology using Gompertz and Baranyi and Roberts models fitted well both for the pressurized and control samples. The results indicated that the high hydrostatic pressure treatment greatly increased the turkey ham commercial viability in comparison to the usual length, by slowing down the growth of microorganisms in the product. PMID:24159279
Mancini, Andrea; Lazzi, Camilla; Bernini, Valentina; Neviani, Erasmo; Gatti, Monica
The aim of this study was to evaluate the potential of target tRNA(Ala)-23S ribosomal DNA for identification of lactic acid bacteria strains associated with dairy ecosystem. For this purpose tRNA(Ala)-23S ribosomal DNA Restriction Fragment Length Polymorphism (tRNA(Ala)-23S rDNA-RFLP) was compared with two widely used DNA fingerprinting methods - P1 Random Amplified Polymorphic DNA (RAPD), (GTG)5 repetitive extragenic palindromic PCR (rep-PCR) - for their ability to identify different species on a set of 10 type and 34 reference strains. Moreover, 75 unknown isolates collected during different stages of Grana Padano cheese production and ripening were identified using tRNA(Ala)-23S rDNA-RFLP and compared to the RFLP profiles of the strains in the reference database. This study demonstrated that the target tRNA(Ala)-23S rDNA has high potential in bacterial identification and tRNA(Ala)-23S rDNA-RFLP is a promising method for reliable species-level identification of lactic acid bacteria (LAB) in dairy products.
Wang, Yi-Chieh; Yu, Roch-Chui; Chou, Cheng-Chun
To further the goal of developing a probiotic dietary adjunct using soymilk, soymilk is fermented with lactic acid bacteria (Lactobacillus acidophilus CCRC 14079 or Streptococcus thermophilus CCRC 14085) and bifidobacteria (Bifidobacterium infantis CCRC 14633 or Bifidobacterium longum B6) individually, and in conjunction. We investigate several antioxidative activities including the inhibition of ascorbate autoxidation, the scavenging effect of superoxide anion radicals and hydrogen peroxide, and the reducing activity exerted by different varieties of fermented soymilks. In addition, the effect of spray-drying and freeze-drying on changes in antioxidative activity is examined. We find that in fermented soymilk both the inhibition of ascorbate autoxidation, and the reducing activity and scavenging effect of superoxide anion radicals varied with the starters used, but nevertheless are significantly higher than those found in unfermented soymilk. In general, antioxidative activity in soymilk fermented with lactic acid bacteria and bifidobacteria simultaneously is significantly higher (P < 0.05) than that fermented with either individually. Moreover, antioxidative activity increases as the fermentation period is extended. However, unfermented soymilk shows an H2O2-scavenging effect, while there is no scavenging effect except for the accumulation of H2O2 in fermented soymilk. Finally, we find that freeze-drying causes a significantly lesser (P < 0.05) reduction in the antioxidative activity of soymilk than does spray-drying. Irrespective of the drying method and the starters used for fermentation. The antioxidative activity of fermented soymilk reduces after drying yet remains higher than that of dried unfermented soymilk.
Nancucheo, Ivan; Johnson, D Barrie
Two continuous-flow bench-scale bioreactor systems populated by mixed communities of acidophilic sulfate-reducing bacteria were constructed and tested for their abilities to promote the selective precipitation of transition metals (as sulfides) present in synthetic mine waters, using glycerol as electron donor. The objective with the first system (selective precipitation of copper from acidic mine water containing a variety of soluble metals) was achieved by maintaining a bioreactor pH of ≈ 2.2-2.5. The second system was fed with acidic (pH 2.5) synthetic mine water containing 3 mM of both zinc and ferrous iron, and varying concentrations (0.5-30 mM) of aluminium. Selective precipitation of zinc sulfide was possible by operating the bioreactor at pH 4.0 and supplementing the synthetic mine water with 4 mM glycerol. Analysis of the microbial populations in the bioreactors showed that they changed with varying operational parameters, and novel acidophilic bacteria (including one sulfidogen) were isolated from the bioreactors. The acidophilic sulfidogenic bioreactors provided 'proof of principle' that segregation of metals present in mine waters is possible using simple online systems within which controlled pH conditions are maintained. The modular units are versatile and robust, and involve minimum engineering complexity.
Gala, Elisabetta; Landi, Sara; Solieri, Lisa; Nocetti, Marco; Pulvirenti, Andrea; Giudici, Paolo
The diversity of dominant lactic acid bacteria population in 12 months ripened Parmigiano Reggiano cheeses was investigated by a polyphasic approach including culture-dependent and independent methods. Traditional plating, isolation of LAB and identification by 16S rDNA analysis showed that strains belonging to Lactobacillus casei group were the most frequently isolated. Lactobacillus helveticus, Lactobacillus delbrueckii subsp. lactis, Lactobacillus parabuchneri, and Lactobacillus buchneri species were detected with lower frequency. PCR-denaturing gradient gel electrophoresis (DGGE) applied to DNA extracted directly from cheese samples and sequencing of rDNA amplicons confirmed the complex microbiological pattern of LAB in ripened Parmigiano Reggiano cheeses, with the significant exception of the Lactobacillus fermentum species, which dominated in several samples, but was not detected by cultivation. The present combination of different approaches can effectively describe the lactic acid bacteria population of Parmigiano Reggiano cheese in advanced stages of ripening, giving useful information for elucidating the role of LAB in determining the final cheese quality.
Lopez de Victoria, G. . Dept. of Biology)
The chemotactic behavior of deep terrestrial subsurface bacteria toward amino acids, carbohydrates and trichloroethylene was assayed using a modification of the capillary method and bacterial enumeration by acridine orange direct counts. Eleven isolates of bacteria isolated from six different geological formations were investigated. A bimodal response rather than an absolute positive or negative response was observed in most assays. Most of the isolates were positively chemotactic to low concentrations of substrates and were repelled by high concentrations of the same substrate. However, this was not the case for trichloroethylene (TCE) which was mostly an attractant and elicited the highest responses in all the isolates when compared with amino acids and carbohydrates. The movement rates of these isolates in aseptic subsurface sediments in the absence and presence of TCE were also determined using a laboratory model. All of the isolates showed distinct response range, peak, and threshold concentrations when exposed to the same substrates suggesting that they are possibly different species as has been inferred from DNA homology studies. 101 refs., 4 figs., 57 tabs.
SIGAL, N; SENEZ, J C; LEGALL, J; SEBALD, M
Sigal, Nicole (Laboratoire de Chimie Bactérienne du CNRS, Marseille, France), Jacques C. Senez, Jean Le Gall, and Madeleine Sebald. Base composition of the deoxyribonucleic acid of sulfate-reducing bacteria. J. Bacteriol. 85:1315-1318. 1963-The deoxyribonucleic acid constitution of several strains of sulfate-reducing bacteria has been analytically determined. The results of these studies show that this group of microorganisms includes at least four subgroups characterized by significantly different values of the adenine plus thymine to guanine plus cytosine ratio. The nonsporulated forms with polar flagellation, containing both cytochrome c(3) and desulfoviridin, are divided into two subgroups. One includes the fresh-water, nonhalophilic strains with base ratio from 0.54 to 0.59, and the other includes the halophilic or halotolerant strains with base ratio from 0.74 to 0.77. The sporulated, peritrichous strains without cytochrome and desulfoviridin ("nigrificans" and "orientis") are distinct from the above two types and differ from each other, having base ratios of 1.20 and 1.43, respectively.
Patra, F; Tomar, S K; Arora, S
Lactic acid bacteria (LAB) have been extensively used for centuries as starter cultures to carry out food fermentations and are looked upon as burgeoning "cell factories" for production of host of functional biomolecules and food ingredients. Low-calorie sugars have been a recent addition and have attracted a great deal of interest of researchers, manufacturers, and consumers for varied reasons. These sweeteners also getting popularized as low-carb sugars have been granted generally recommended as safe (GRAS) status by the U.S. Federal Drug Administration (USFDA) and include both sugars and sugar alcohols (polyols) which in addition to their technological attributes (sugar replacer, bulking agent, texturiser, humectant, cryoprotectant) have been observed to exert a number of health benefits (low calories, low glycemic index, anticariogenic, osmotic diuretics, obesity control, prebiotic). Some of these sweeteners successfully produced by lactic acid bacteria include mannitol, sorbitol, tagatose, and trehalose and there is a potential to further enhance their production with the help of metabolic engineering. These safe sweeteners can be exploited as vital food ingredients for development of low-calorie foods with added functional values especially for children, diabetic patients, and weight watchers.