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1

[Microbiology method validation: illustration with Bact'Alert 3D].  

PubMed

As far as laboratories accreditation according to ISO 15189 is concerned, validation of each method used has to be achieved. Manufacturer's recommendations are not always helpful in this context. That's why each laboratory must proceed systematically to a logical risk analysis. Few publications are available on this subject concerning microbiology. We propose to illustrate it with the example of a blood culture automate: the Bact'Alert 3D commercialised by Biomérieux(®). PMID:23747676

Védy, Serge

2013-01-01

2

Differentiation of Enterobacter aerogenes from Klebsiellae by Deoxyribonucleic Acid Reassociation  

Microsoft Academic Search

Polynucleotide sequence relatedness tests were carried out to determine the extent of deoxyribonucleic acid (DNA) divergence among species of Klebsiella and Enterobacter aerogenes strains. Labeled, denatured DNA fragments from K. pneumoniae type 2 and E. aerogenes 1627-66 were each incubated with an excess of unlabeled DNA fragments from Klebsielia species and strains of E. aerogenes. Reassociated DNA duplexes were separated

DON J. BRENNER; A. G. STEIGERWALT; G. R. FANNING

1972-01-01

3

VetBact - culturing bacteriological knowledge for veterinarians.  

PubMed

In 2004, Karl-Erik Johansson, then professor of veterinary bacteriology at the veterinary school at Uppsala in Sweden, was asked by his students for a list of the most important bacteria and the diseases that they cause. So began the development of VetBact, an online database giving details of the bacterial species with most relevance to veterinary medicine. The non-commercial database, www.vetbact.org, has since grown and can now be accessed by veterinarians and others worldwide. PMID:24526535

Johansson, Karl-Erik

2014-02-15

4

Biodegradation Kinetics of Four Substituted Chlorobenzoic Acids by Enterobacter aerogenes  

Microsoft Academic Search

Enterobacter aerogenes is generally found in soil, sewage plants, and human gastrointestinal tract. Thus, this study was conducted to evaluate the ability of Enterobacter aerogenes to degrade four chlorobenzoic acid compounds (2-chlorobenzoic acid (2-CBA), 3-chlorobenzoic acid (3-CBA), 4-chlorobenzoic acid (4-CBA), and 3,4-dichlorobenzoic acid (3,4-dCBA)) in minimal salt medium. Enterobacter aerogenes was partially able to degrade and dechlorinate these CBAs at

Khaled A. Tarawneh; Fawzi Irshaid; Isam H. Ajlundi; Muayad M. Abboud; Noor A. Mohammed; Ali M. Khleifat

2010-01-01

5

mar Operon Involved in Multidrug Resistance of Enterobacter aerogenes  

Microsoft Academic Search

We determined the sequence of the entire marRAB operon in Enterobacter aerogenes. It is functionally and structurally analogous to the Escherichia coli operon. The overexpression of E. aerogenes MarA induces a multidrug resistance phenotype in a susceptible strain, demonstrated by a noticeable resistance to various antibiotics, a decrease in immunodetected porins, and active efflux of norfloxacin.

Renaud Chollet; Claude Bollet; Jacqueline Chevalier; M. Mallea; J.-M. Pages; Anne Davin-Regli

2002-01-01

6

Robust Multivariable Flutter Suppression for the Benchmark Active Control Technology (BACT) Wind-Tunnel Model  

NASA Technical Reports Server (NTRS)

The Benchmark Active Controls Technology (BACT) project is part of NASA Langley Research Center s Benchmark Models Program for studying transonic aeroelastic phenomena. In January of 1996 the BACT wind-tunnel model was used to successfully demonstrate the application of robust multivariable control design methods (H and -synthesis) to flutter suppression. This paper addresses the design and experimental evaluation of robust multivariable flutter suppression control laws with particular attention paid to the degree to which stability and performance robustness was achieved.

Waszak, Martin R.

1997-01-01

7

Amino acid utilization by Aerobacter aerogenes and Escherichia coli  

E-print Network

A considerable amount of work has been done on the growth of A. aerogenes and E. coli in synthetic media, but little work has been undertaken on the utilization by these organisms of amino acids as comparative sources of ...

Herrera, Rodolfo Eduardo

1938-01-01

8

Effects of Formate on Fermentative Hydrogen Production by Enterobacter aerogenes  

Microsoft Academic Search

This paper describes the effects of formate on fermentative hydrogen production by Enterobacter aerogenes by way of batch culture. When 20 mM formate was added to pH 6.3 and pH 5.8 E. aerogenes glucose cultures (formate culture) at the beginning of cultivation, hydrogen evolution through both glucose consumption and decomposition of the extrinsic formate occurred together, while hydrogen evolution occurred

Tatsuo Kurokawa; Shigeharu Tanisho

2005-01-01

9

Dissolution of Xylose Metabolism in Lactococcus lactis  

PubMed Central

Xylose metabolism, a variable phenotype in strains of Lactococcus lactis, was studied and evidence was obtained for the accumulation of mutations that inactivate the xyl operon. The xylose metabolism operon (xylRAB) was sequenced from three strains of lactococci. Fragments of 4.2, 4.2, and 5.4 kb that included the xyl locus were sequenced from L. lactis subsp. lactis B-4449 (formerly Lactobacillus xylosus), L. lactis subsp. lactis IO-1, and L. lactis subsp. lactis 210, respectively. The two environmental isolates, L. lactis B-4449 and L. lactis IO-1, produce active xylose isomerases and xylulokinases and can metabolize xylose. L. lactis 210, a dairy starter culture strain, has neither xylose isomerase nor xylulokinase activity and is Xyl?. Xylose isomerase and xylulokinase activities are induced by xylose and repressed by glucose in the two Xyl+ strains. Sequence comparisons revealed a number of point mutations in the xylA, xylB, and xylR genes in L. lactis 210, IO-1, and B-4449. None of these mutations, with the exception of a premature stop codon in xylB, are obviously lethal, since they lie outside of regions recognized as critical for activity. Nevertheless, either cumulatively or because of indirect affects on the structures of catalytic sites, these mutations render some strains of L. lactis unable to metabolize xylose. PMID:10966417

Erlandson, Karn A.; Park, Joo-Heon; Wissam; El Khal; Kao, Hsin-Hsin; Basaran, Pervin; Brydges, Susannah; Batt, Carl A.

2000-01-01

10

Natural immune systems protect animals from dangerous foreign pathogens, including bacte-  

E-print Network

Natural immune systems protect animals from dangerous foreign pathogens, including bacte- ria of Naval Research (N00014-95-1-0364), Defense Advanced Research Projects Agency (N00014 infections are all highly parallel and distributed. This is one reason immune system mechanisms are so

Garlan, David

11

TEM Derivative-Producing Enterobacter aerogenes Strains: Dissemination of a Prevalent Clone  

Microsoft Academic Search

TEM-24 (CAZ-6) extended-spectrum -lactamase (ESBL) was detected in 1988 in Clermont-Ferrand, France, in Klebsiella pneumoniae (blaTEM-24) and Enterobacter aerogenes (blaTEM-24b), and since 1994, a TEM- 24-producing E. aerogenes clonal strain has been observed elsewhere in the country. To determine if the spread of this clonal strain was restricted to TEM-24-producing E. aerogenes strains, 84 E. aerogenes strains (non- TEM\\/SHV-producing strains,

P. Dumarche; C. De Champs; D. Sirot; C. Chanal; R. Bonnet; J. Sirot

2002-01-01

12

Lactococcus lactis subsp. lactis infection in waterfowl: first confirmation in animals.  

PubMed Central

We report the first description, confirmed by bacteriologic and molecular (polymerase chain reaction and pulsed-field gel electrophoresis) analysis, of an infection in animals caused by Lactococcus lactis subsp. lactis, affecting waterfowl. PMID:11747704

Goyache, J.; Vela, A. I.; Gibello, A.; Blanco, M. M.; Briones, V.; González, S.; Téllez, S.; Ballesteros, C.; Domínguez, L.; Fernández-Garayzábal, J. F.

2001-01-01

13

Exploring the Genome of Cheese Starter Lactic Acid Bacterium Lactococcus lactis subsp. lactis CECT 4433.  

PubMed

Here, we present the draft genome sequences of Lactococcus lactis subsp. lactis CECT 4433, a cheese fermentation starter strain. The genome provides further insight into the genomic plasticity, biocomplexity (including gene strain specifics), and evolution of these genera. PMID:25395632

Tschoeke, Diogo Antonio; Moreira, Ana Paula B; Chimetto Tonon, Luciane A; de Mesquita, Milene Miranda A; Gregoracci, Gustavo B; Gomez-Gil, Bruno; Valle, Rogério; Thompson, Cristiane C; Thompson, Fabiano L

2014-01-01

14

Inhibitors of antibiotic efflux pump in resistant Enterobacter aerogenes strains  

Microsoft Academic Search

Enterobacter aerogenes, a nosocomial pathogen, is frequently exhibiting multidrug resistance mechanisms associated with a change in membrane permeability. In clinical isolates, active efflux plays a prominent role in antibiotic resistance. We report here the effect of three unrelated compounds that are able to restore a noticeable antibiotic susceptibility to resistant strains. The targeting of various parameters which contribute to the

Monique Malléa; Jacqueline Chevalier; Annie Eyraud; Jean-Marie Pagès

2002-01-01

15

Chloramphenicol and expression of multidrug efflux pump in Enterobacter aerogenes  

Microsoft Academic Search

Chloramphenicol has been reported to act as an inducer of the multidrug resistance in Escherichia coli. A resistant variant able to grow on plates containing 64?g\\/ml chloramphenicol was obtained from the Enterobacter aerogenes ATCC 13048-type strain. Chloramphenicol resistance was due to an active efflux of this antibiotic and it was associated with resistance to fluoroquinolones and tetracycline, but not to

Didier Ghisalberti; Muriel Masi; Jean-Marie Pagès; Jacqueline Chevalier

2005-01-01

16

Carbapenem Resistance in Enterobacter aerogenes is due to Lipopolysaccharide Alterations  

Microsoft Academic Search

The extensive characterization of 2 clinical Enterobacter aerogenes isolates resistant to all ?-lactam antibiotics including imipenem revealed that imipenem resistance could not be attributed to overproduction of the chromosomal ?-lactamase; moreover, it was lost after subcultivation and can be thus considered as unstable. The comparison of sensitive and resistant clones revealed that the ?-lactamase in the resistant clones was less

Hermann Leying; Wolfgang Cullmann; Wolfgang Dick

1991-01-01

17

Imipenem and expression of multidrug efflux pump in Enterobacter aerogenes  

Microsoft Academic Search

Imipenem is often used to treat intensive care unit patients infected by Enterobacter aerogenes, but it is leading to an increasing number of antibiotic resistant strains. Clinical isolates and imipenem resistant variants presented a high level of resistance to ?-lactam antibiotic group and to chemically unrelated drugs. We report here that imipenem selects strains which contain active efflux pumps ejecting

Charléric Bornet; Renaud Chollet; Monique Malléa; Jacqueline Chevalier; Anne Davin-Regli; Jean-Marie Pagès; Claude Bollet

2003-01-01

18

Interrelations Between Two Pathways of Methionine Biosynthesis in Aerobacter aerogenes  

Microsoft Academic Search

SUMMARY Two pathways for methionine methyl formation, one cobalamin- dependent and one cobalamin-independent, corresponding to those observed in Escherichia coli PA 15 have been found in Aerobacter aerogenes. An initial difficulty in showing the cobalamin-dependent pathway in cell- free extracts proved to be due to the presence of enzymes which caused the removal of adenosylmethionine, a cofactor required for this

J. F. MORNINGSTAR; R. L. KISLIUK

1965-01-01

19

Energy Production During Nitrate Respiration by Aerobacter aerogenes  

Microsoft Academic Search

SUMMARY The molar growth yield of Aerobacter aerogenes growing anaerobically with glucose in a mineral medium was almost doubled when NO3- was added as hydrogen acceptor. About half a mole of NO,- was reduced to NH,+ per mole of glucose. The amount of ATP produced from glucose fermentation calculated from the molar growth yield and the acetate production was about

LIGERI P. HADJIPETROU; A. H. STOUTHAMER

1965-01-01

20

The Citrate Transport System of Lactococcus lactis subsp. lactis biovar diacetylactis Is Induced by Acid Stress  

Microsoft Academic Search

Citrate transport in Lactococcus lactis subsp. lactis biovar diacetylactis is catalyzed by citrate permease P (CitP), which is encoded by the plasmidic citP gene. We have shown previously that citP is included in the citQRP operon, which is mainly transcribed from the P1 promoter in L. lactis subsp. lactis biovar diacetylactis. Furthermore, transcription of citQRP and citrate transport are not

NIEVES GARCIA-QUINTANS; CHRISTIAN MAGNI; DIEGO DE MENDOZA; PALOMA LOPEZ

1998-01-01

21

Genome Sequence of Lactococcus lactis subsp. lactis NCDO 2118, a GABA-Producing Strain  

PubMed Central

Lactococcus lactis subsp. lactis NCDO 2118 is a nondairy lactic acid bacterium, a xylose fermenter, and a gamma-aminobutyric acid (GABA) producer isolated from frozen peas. Here, we report the complete genome sequence of L. lactis NCDO 2118, a strain with probiotic potential activity. PMID:25278529

Oliveira, Leticia C.; Saraiva, Tessalia D. L.; Soares, Siomar C.; Ramos, Rommel T. J.; Sa, Pablo H. C. G.; Carneiro, Adriana R.; Miranda, Fabio; Freire, Matheus; Renan, Wendel; Junior, Alberto F. O.; Santos, Anderson R.; Pinto, Anne C.; Souza, Bianca M.; Castro, Camila P.; Diniz, Carlos A. A.; Rocha, Clarissa S.; Mariano, Diego C. B.; de Aguiar, Edgar L.; Folador, Edson L.; Barbosa, Eudes G. V.; Aburjaile, Flavia F.; Goncalves, Lucas A.; Guimaraes, Luis C.; Azevedo, Marcela; Agresti, Pamela C. M.; Silva, Renata F.; Tiwari, Sandeep; Almeida, Sintia S.; Hassan, Syed S.; Pereira, Vanessa B.; Abreu, Vinicius A. C.; Pereira, Ulisses P.; Dorella, Fernanda A.; Carvalho, Alex F.; Pereira, Felipe L.; Leal, Carlos A. G.; Figueiredo, Henrique C. P.; Silva, Artur; Miyoshi, Anderson

2014-01-01

22

Isolation of halotolerant Lactococcus lactis subsp. lactis from intestinal tract of coastal fish.  

PubMed

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. PMID:18068256

Itoi, Shiro; Abe, Takeshi; Washio, Sayaka; Ikuno, Erika; Kanomata, Yuna; Sugita, Haruo

2008-01-15

23

NEW STRAIN KLUYVEROMYCES LACTIS 3  

Microsoft Academic Search

ß-Galactosidase (EC.3.2.1.23 is an important enzyme industrially used for the hydrolysis of lactose from milk and milk whey for several applications.Lately, the importance of this enzyme was enhanced by its galaktosyltransferase activity, which is responsible for synthesis of transgalactosylated oligosaccharides that act as prebiotics, with several beneficial effects on consumers. ß-Galactosidase production by K. lactis 3 was studied in shake

Ilia Iliev; Tonka Vassileva

24

Functionality of Sortase A in Lactococcus lactis?  

PubMed Central

Lactococcus lactis IL1403 harbors a putative sortase A (SrtA) and 11 putative sortase substrates that carry the canonical LPXTG signature of such substrates. We report here on the functionality of SrtA to anchor five LPXTG substrates to the cell wall, thus suggesting that SrtA is the housekeeping sortase in L. lactis IL1403. PMID:20851967

Dieye, Yakhya; Oxaran, Virginie; Ledue-Clier, Florence; Alkhalaf, Walid; Buist, Girbe; Juillard, Vincent; Lee, Chang Won; Piard, Jean-Christophe

2010-01-01

25

Ribitol dehydrogenase from Klebsiella aerogenes. Purification and subunit structure  

PubMed Central

Ribitol dehydrogenase has been purified to homogeneity from several strains of Klebsiella aerogenes. One strain yields 3–6g of pure enzyme from 1kg of cells. The enzyme is a tetramer of four subunits, mol.wt. 27000. Preliminary studies of the activity of the enzyme are reported. Peptide `maps' together with the amino acid composition indicate that the subunits are identical. ImagesPLATE 2PLATE 1 PMID:4618776

Taylor, Susan S.; Rigby, Peter W. J.; Hartley, Brian S.

1974-01-01

26

Thermodynamic study and optimization of hydrogen production by Enterobacter aerogenes  

Microsoft Academic Search

This paper investigates the influence of pH and temperature on hydrogen bioproduction by Enterobacter aerogenes (NCIMB 10102) utilizing starch hydrolysate as substrate. An optimum pH range corresponding to 6.1–6.6 is the main evidence of batch runs carried out at different pHs. An optimum value of temperature corresponding to 40°C is experimentally determined by means of batch fermentation runs carried out

B. Fabiano; P. Perego

2002-01-01

27

Molecular Analysis of Tetracycline Resistance in Pasteurella aerogenes  

PubMed Central

Tetracycline-resistant Pasteurella aerogenes isolates obtained from the intestinal tract of swine were investigated for their tet genes by PCR analysis and hybridization experiments. In contrast to Pasteurella isolates from the respiratory tract, tet(H) genes were detected in the chromosomal DNA of only 2 of the 24 isolates, one of which also carried two copies of a tet(B) gene. All other P. aerogenes isolates carried tet(B) genes, which are the predominant tet genes among Enterobacteriaceae. A single isolate harbored a tet(B) gene as part of a truncated Tn10 element on the 4.8-kb plasmid pPAT2. Comparative analysis of the pPAT2 sequence suggested that the Tn10 relic on plasmid pPAT2 is the result of several illegitimate recombination events. The remaining 21 P. aerogenes isolates carried one or two copies of the tet(B) gene in their chromosomal DNA. In the majority of the cases, these tet(B) genes were associated with copies of Tn10 as confirmed by their SfuI and BamHI hybridization patterns. No correlation between the number of tet gene copies and the MICs of tetracycline, doxycyline and minocycline was observed. PMID:11557485

Kehrenberg, Corinna; Schwarz, Stefan

2001-01-01

28

Biological Conversion of Glycerol to Ethanol by Enterobacter aerogenes  

NASA Astrophysics Data System (ADS)

In a search to turn the economically and environmentally non-valuable "waste" streams of biodiesel production into a profitable byproduct, a mutant strain of Enterobacter aerogenes ATCC 13048 was developed by six-tube subculturing technique. This technique is based on the principle of adaptive evolution, and involved subculturing the bacterium in a tryptic soy broth without dextrose (TSB) containing specific glycerol and ethanol concentration for six consecutive times. Then, the six consecutive subculturing was repeated in a fresh TSB of higher glycerol and ethanol concentrations. A new mutant strain, E. aerogenes S012, which could withstand a combination of 200 g/l glycerol and 30 g/l ethanol concentrations, was developed. The wild and mutant strains were used for the fermentation of pure (P-) and recovered (R-) glycerol. Taguchi and full factorial methods of design of experiments were used to screen and optimize the important process factors that influence the microbial production of ethanol. A statistically sound regression model was used to establish the mathematical relationship between the process variables and ethanol production. Temperature of 38°C, agitation speed of 200 rpm, pH of 6.3-6.6, and microaerobic condition were the optimum process conditions. Different pretreatment methods to recover glycerol from the crude glycerol and the subsequent fermentation method showed that direct acidification using 85% H3PO4 was the best. The R-glycerol contained 51% pure glycerol and 21% methanol. The wild strain, E. aerogenes ATCC 13048, produced only 12 g/l and 12.8 g/l ethanol from 20 g/l P- and R-glycerol respectively, and could not utilize higher glycerol concentrations. The mutant, E. aerogenes S012, produced ethanol amount and yield of 43 g/l and 1.12 mol/mol-glycerol from P-glycerol, respectively within 96 h. It also produced ethanol amount and yield of 26.8 g/l and 1.07 mol/mol-glycerol, respectively, from R-glycerol within the same duration. In a fermentation to estimate hydrogen production using a respirometer, the hydrogen yield and volumetric rate of 1.06 mol/mol-glycerol and 217 ml/l/h, respectively were obtained from 6% P-glycerol in 72 h by E. aerogenes S012. The result was higher from R-glycerol, which produced hydrogen yield and productivity of 1.83 mol/mol-glycerol and 326 ml/l/h, respectively.

Nwachukwu, Raymond E. S.

29

Molecular Epidemiological Study of Nosocomial Enterobacter aerogenes Isolates in a Belgian Hospital  

Microsoft Academic Search

In 1995, the rate of isolation of Enterobacter aerogenes in the Saint-Pierre University Hospital in Brussels, Belgium, was higher than that in the preceding years. A total of 45 nosocomial E. aerogenes strains were collected from 33 patients of different units during that year, and they were isolated from 19 respiratory specimens, 13 pus specimens, 7 blood specimens, 4 urinary

SHEIKH JALALUDDIN; JEANNE-MARIE DEVASTER; ROBERT SCHEEN; MICHELE GERARD; JEAN-PAUL BUTZLER

1998-01-01

30

Regulation of hydrogen production by Enterobacter aerogenes by external NADH and NAD +  

Microsoft Academic Search

Experiments involving the addition of external nicotinamide adenine dinucleotide, reduced form (NADH) or nicotinamide adenine dinucleotide (NAD+) have been designed to examine how the hydrogen in Enterobacter aerogenes is liberated by NADH or NAD+. The addition of external NADH or NAD+ was found to regulate hydrogen production by E. aerogenes in resting cells, batch cultures, and chemostat cultures. Particularly in chemostat

Chong Zhang; Kun Ma; Xin-Hui Xing

2009-01-01

31

Kinetic Behaviour of Lactococcus lactis ssp. lactis bv. diacetylactis Immobilized in Calcium Alginate Gel Beads  

Microsoft Academic Search

Alginate beads with entrapped Lactococcus lactis spp. lactis bv. diacetylactis were used as biocatalysts in continuous fermentation and the dynamics of the system analysed. Colonization by cells changed during fermentation, concentrating at the periphery (where cell densities reach 350 g litre?1). In the steady state, colonization of the gel was interrupted while growth continued, producing cells that were released into

R. Cachon; M. Catté; R. Nommé; H. Prévost; C. Diviès

1995-01-01

32

Uridine diphosphate D-glucose dehydrogenase of Aerobacter aerogenes.  

PubMed

Uridine diphosphate d-glucose dehydrogenase (EC 1.1.1.22) from Aerobacter aerogenes has been partially purified and its properties have been investigated. The molecular weight of the enzyme is between 70,000 and 100,000. Uridine diphosphate d-glucose is a substrate; the diphosphoglucose derivatives of adenosine, cytidine, guanosine, and thymidine are not substrates. Nicotinamide adenine dinucleotide (NAD), but not nicotinamide adenine dinucleotide phosphate, is active as hydrogen acceptor. The pH optimum is between 9.4 and 9.7; the K(m) is 0.6 mm for uridine diphosphate d-glucose and 0.06 mm for NAD. Inhibition of the enzyme by uridine diphosphate d-xylose is noncooperative and of mixed type; the K(i) is 0.08 mm. Thus, uridine diphosphate d-glucose dehydrogenase from A. aerogenes differs from the enzyme from mammalian liver, higher plants, and Cryptococcus laurentii, in which uridine diphosphate d-xylose functions as a cooperative, allosteric feedback inhibitor. PMID:4387161

Bdolah, A; Feingold, D S

1968-10-01

33

Regulation of tyramine oxidase synthesis in Klebsiella aerogenes.  

PubMed Central

Tyramine oxidase in Klebsiella aerogenes is highly specific for tyramine, dopamine, octopamine, and norepinephrine, and its synthesis is induced specifically by these compounds. The enzyme is present in a membrane-bound form. The Km value for tyramine is 9 X 10(-4) M. Tyramine oxidase synthesis was subjected to catabolite repression by glucose in the presence of ammonium salts. Addition of cyclic adenosine 3',5'-monophosphate (cAMP) overcame the catabolite repression. A mutant strain, K711, which can produce a high level of beta-galactosidase in the presence of glucose and ammonium chloride, can also synthesize tyramine oxidase and histidase in the presence of inducer in glucose ammonium medium. Catabolite repression of tyramine oxidase synthesis was relieved when the cells were grown under conditions of nitrogen limitation, whereas beta-galactosidase was strongly repressed under these conditions. A cAMP-requiring mutant, MK54, synthesized tyramine oxidase rapidly when tyramine was used as the sole source of nitrogen in the absence of cAMP. However, a glutamine synthetase-constitutive mutant, MK94, failed to synthesize tyramine oxidase in the presence of glucose and ammonium chloride, although it synthesized histidase rapidly under these conditions. These results suggest that catabolite repression of tyramine oxidase synthesis in K. aerogenes is regulated by the intracellular level of cAMP and an unknown cytoplasmic factor that acts independently of cAMP and is formed under conditions of nitrogen limitation. PMID:179974

Okamura, H; Murooka, Y; Harada, T

1976-01-01

34

Comparative Phenotypic and Molecular Genetic Profiling of Wild Lactococcus lactis subsp. lactis Strains of the L. lactis subsp. lactis and L. lactis subsp. cremoris Genotypes, Isolated from Starter-Free Cheeses Made of Raw Milk?  

PubMed Central

Twenty Lactococcus lactis strains with an L. lactis subsp. lactis phenotype isolated from five traditional cheeses made of raw milk with no added starters belonging to the L. lactis subsp. lactis and L. lactis subsp. cremoris genotypes (lactis and cremoris genotypes, respectively; 10 strains each) were subjected to a series of phenotypic and genetic typing methods, with the aims of determining their phylogenetic relationships and suitability as starters. Pulsed-field gel electrophoresis (PFGE) analysis of intact genomes digested with SalI and SmaI proved that all strains were different except for three isolates of the cremoris genotype, which showed identical PFGE profiles. Multilocus sequence typing (MLST) analysis using internal sequences of seven loci (namely, atpA, rpoA, pheS, pepN, bcaT, pepX, and 16S rRNA gene) revealed considerable intergenotype nucleotide polymorphism, although deduced amino acid changes were scarce. Analysis of the MLST data for the present strains and others from other dairy and nondairy sources showed that all of them clustered into the cremoris or lactis genotype group, by using both independent and combined gene sequences. These two groups of strains also showed distinctive carbohydrate fermentation and enzyme activity profiles, with the strains in the cremoris group showing broader profiles. However, the profiles of resistance/susceptibility to 16 antibiotics were very similar, showing no atypical resistance, except for tetracycline resistance in three identical cremoris genotype isolates. The numbers and concentrations of volatile compounds produced in milk by the strains belonging to these two groups were clearly different, with the cremoris genotype strains producing higher concentrations of more branched-chain, derived compounds. Together, the present results support the idea that the lactis and cremoris genotypes of phenotypic Lactococcus lactis subsp. lactis actually represent true subspecies. Some strains of the two subspecies in this study appear to be good starter candidates. PMID:21666023

Fernandez, Elena; Alegria, Angel; Delgado, Susana; Martin, M. Cruz; Mayo, Baltasar

2011-01-01

35

Insertion and amplification of foreign genes in the Lactococcus lactis subsp. lactis chromosome.  

PubMed Central

The plasmid pE194 is unable to replicate in Lactococcus lactis subsp. lactis (formerly Streptococcus lactis). When linked to resident bacteriophage sequences, pE194 was able to integrate into the L. lactis subsp. lactis chromosome either by Campbell-like recombination or by double crossing over with deletion. Integration occurred into the DNA of the prophage and prevented its multiplication. When a selective pressure was applied to an integrant in which pE194 was flanked by two direct repeats of prophage fragment, amplification of pE194 and the prophage fragment was observed. The pE194 copy number was assessed at six to nine, and amplification was stable upon growth under nonselective conditions. Images PMID:2504115

Chopin, M C; Chopin, A; Rouault, A; Galleron, N

1989-01-01

36

Genome Sequence of the Lactic Acid Bacterium Lactococcus lactis subsp. lactis TOMSC161, Isolated from a Nonscalded Curd Pressed Cheese  

PubMed Central

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

Velly, H.; Abraham, A.-L.; Loux, V.; Delacroix-Buchet, A.; Fonseca, F.; Bouix, M.

2014-01-01

37

Genome Sequence of the Lactic Acid Bacterium Lactococcus lactis subsp. lactis TOMSC161, Isolated from a Nonscalded Curd Pressed Cheese.  

PubMed

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

Velly, H; Renault, P; Abraham, A-L; Loux, V; Delacroix-Buchet, A; Fonseca, F; Bouix, M

2014-01-01

38

[Purification and properties of chitinase from Enterobacter aerogenes].  

PubMed

A bacterium producing chitinase was isolated from the dead body of Gymephorap ruoergensis. A chitinase was isolated from the culture of E. aerogenes and purified by means of ammonium sulfate precipitation, DEAE-cellulose column chromatography, and Sephadex G-100 column gel filtration. The purified chitinase showed homogeneity on the native polyacrylamide gel electrophoresis. Its molecular weight was estimated to be about 42.5 kD by SDS-PAGE. The optimum pH and temperature for hydrolysis of chitin were 6.0 and 55 degrees C respectively. Michaelis constant was 2.88 mg/mL. Different metal ions showed different effects on the chitinase activity, The chitinase activity was enhanced by Zn2+, Ba2+, Ca2+, Mn2+ and was strongly inhibited by Hg2+, Co2+, Mg2+. PMID:12549194

Tang, Y; Zhao, J; Ding, S; Liu, S; Yang, Z

2001-02-01

39

Bioconversion of glycerol to ethanol by a mutant Enterobacter aerogenes  

PubMed Central

The main objective of this research is to develop, by adaptive evolution, mutant strains of Enterobacter aerogenes ATCC 13048 that are capable of withstanding high glycerol concentration as well as resisting ethanol-inhibition. The mutant will be used for high ethanol fermentation from glycerol feedstock. Ethanol production from pure (P-) and recovered (R-) glycerol using the stock was evaluated. A six-tube-subculture-generations method was used for developing the mutant. This involved subculturing the organism six consecutive times in tubes containing the same glycerol and ethanol concentrations at the same culture conditions. Then, the glycerol and/or ethanol concentration was increased and the six subculture generations were repeated. A strain capable of growing in 200 g/L glycerol and 30 g/L ethanol was obtained. The ability of this mutant, vis-à-vis the original strain, in utilizing glycerol in a high glycerol containing medium, with the concomitant ethanol yield, was assessed. Tryptic soy broth without dextrose (TSB) was used as the fermentation medium. Fermentation products were analyzed using HPLC. In a 20 g/L glycerol TSB, E. aerogenes ATCC 13048 converted 18.5 g/L P-glycerol and 17.8 g/L R-glycerol into 12 and 12.8 g/L ethanol, respectively. In a 50 g/L P-glycerol TSB, it utilized only 15.6 g/L glycerol; but the new strain used up 39 g/L, yielding 20 g/L ethanol after 120 h, an equivalence of 1.02 mol ethanol/mol-glycerol. This is the highest ethanol yield reported from glycerol bioconversion. The result of this P-glycerol fermentation can be duplicated using the R-glycerol from biodiesel production. PMID:22455837

2012-01-01

40

Evaluation and application of an improved bacteriocin typing method for Klebsiella aerogenes.  

PubMed Central

A bacteriocin typing method was evaluated using 200 strains of Klebsiella aerogenes, 93% of which fell into 11 distinct types. The typing technique was successfully applied to the monitoring and control of hospital cross-infection. PMID:342545

Heddell, G W; Mitchell, A A

1978-01-01

41

Possible role of membrane proteins in mercury resistance of Enterobacter aerogenes  

Microsoft Academic Search

Mercury resistance shown by a strain of Enterobacter aerogenes was found to be determined by a plasmid. The resistance appeared to be not due to enzymatic volatilization of mercury, but due to the alteration in cellular permeability to mercury.

Hidemitsu S Pan-Hou; Masayo Nishimoto; Nobumasa Imura

1981-01-01

42

A model for multiproduct-inhibited growth of Enterobacter aerogenes in 2,3-butanediol fermentation  

Microsoft Academic Search

Ethanol is identified as a strongly inhibitory metabolite in addition to acetic acid and 2,3-butanediol in 2,3-butanediol production by Enterobacter aerogenes. A model is proposed to describe the multiproduct-inhibited growth of E. aerogenes in 2,3-butanediol fermentation. The model is verified with data from anaerobic and microaerobic continuous culture. On the basis of this model the difference in biomass production and

An-Ping Zeng; Wolf-Dieter Deckwer

1991-01-01

43

Plasmid-Mediated Resistance to Expanded-Spectrum Cephalosporins among Enterobacter aerogenes Strains  

Microsoft Academic Search

Resistance to expanded-spectrum cephalosporins commonly develops in Enterobacter aerogenes during ther- apy due to selection of mutants producing high levels of the chromosomal Bush group 1 b-lactamase. Recently, resistant strains producing plasmid-mediated extended-spectrum b-lactamases (ESBLs) have been isolated as well. A study was designed to investigate ESBL production among 31 clinical isolates of E. aerogenes from Richmond, Va., with decreased

JOHANN D. D. PITOUT; KENNETH S. THOMSON; NANCY D. HANSON; ANTON F. EHRHARDT; PHILIP COUDRON; CHRISTINE C. SANDERS

1998-01-01

44

RamA Is an Alternate Activator of the Multidrug Resistance Cascade in Enterobacter aerogenes  

Microsoft Academic Search

Multidrug resistance (MDR) in Enterobacter aerogenes can be mediated by induction of MarA, which is triggered by certain antibiotics and phenolic compounds. In this study, we identified the gene encoding RamA, a 113-amino-acid regulatory protein belonging to the AraC-XylS transcriptional activator family, in the Enterobacter aerogenes ATCC 13048 type strain and in a clinical multiresistant isolate. Overexpression of RamA induced

Renaud Chollet; Jacqueline Chevalier; Claude Bollet; Jean-Marie Pages; Anne Davin-Regli

2004-01-01

45

Resistance to imipenem, cefepime, and cefpirome associated with mutation in Omp36 osmoporin of Enterobacter aerogenes  

Microsoft Academic Search

Enterobacter aerogenes develops increased multidrug resistance via a functional alteration of outer-membrane permeability associated with a decrease in porin function. We have sequenced the gene coding the major porin of Enterobacter aerogenes, omp36. The sequence shows a high similarity with the Klebsiella pneumoniae ompK36 gene and is closely related to the enterobacterial OmpC family. Sequence analysis of several Omp36 issued

Aurélie Thiolas; Charléric Bornet; Anne Davin-Régli; Jean-Marie Pagès; Claude Bollet

2004-01-01

46

BactPepDB: a database of predicted peptides from a exhaustive survey of complete prokaryote genomes  

PubMed Central

With the recent progress in complete genome sequencing, mining the increasing amount of genomic information available should in theory provide the means to discover new classes of peptides. However, annotation pipelines often do not consider small reading frames likely to be expressed. BactPepDB, available online at http://bactpepdb.rpbs.univ-paris-diderot.fr, is a database that aims at providing an exhaustive re-annotation of all complete prokaryotic genomes—chromosomal and plasmid DNA—available in RefSeq for coding sequences ranging between 10 and 80 amino acids. The identified peptides are classified as (i) previously identified in RefSeq, (ii) entity-overlapping (intragenic) or intergenic, and (iii) potential pseudogenes—intergenic sequences corresponding to a portion of a previously annotated larger gene. Additional information is related to homologs within order, predicted signal sequence, transmembrane segments, disulfide bonds, secondary structure, and the existence of a related 3D structure in the Protein Databank. As a result, BactPepDB provides insights about candidate peptides, and provides information about their conservation, together with some of their expected biological/structural features. The BactPepDB interface allows to search for candidate peptides in the database, or to search for peptides similar to a query, according to the multiple properties predicted or related to genomic localization. Database URL: http://www.yeastgenome.org/ PMID:25377257

Rey, Julien; Deschavanne, Patrick; Tuffery, Pierre

2014-01-01

47

A descriptive model for citrate utilization by Lactococcus lactis ssp lactis bv diacetylactis  

Microsoft Academic Search

A model for the use of citrate by Lactococcus lactis ssp lactis bv diacetylactis CNRZ 125 is proposed. Citrate metabolism by this strain leads to the production of acetate, CO2 and C4 compounds (diacetyl, acetoin, 2,3-butylene glycol). The model furnishes correct simulations, consistent with published results on the pathways used and on lactose-citrate co-metabolism. Citric acid is incorporated independently of

R. Cachon; C. Divdies

1993-01-01

48

Complete Genome Sequence of Lactococcus lactis subsp. lactis KLDS4.0325  

PubMed Central

We report the complete genome sequence of Lactococcus lactis subsp. lactis KLDS4.0325, a probiotic bacterium isolated from homemade koumiss in Xinjiang, China. We have determined the complete genome sequence of strain KLDS4.0325, which consists of a chromosome and three plasmids and reveals genes that are likely to be involved in dairy fermentation and that have probiotic qualities. PMID:24285665

Yang, Xiaochun; Wang, Yutang

2013-01-01

49

PCR Identification of Lysogenic Lactococcus lactis Strains  

Microsoft Academic Search

Lactococcus lactis plays an essential role as a starter in the dairy industry. Unfortunately this species is susceptible to bacteriophage infections that result in fermentation failures and subsequent economic losses. This paper reports a PCR screening method that detects prophages in the genomes of wild lactococcal strains isolated from Cabrales cheese, a traditional, Spanish, blue-veined cheese, which is not inoculated.

M. Cruz Martín; V. Ladero; M. A. Alvarez

2006-01-01

50

Best available control technology (BACT) equivalent for the control of volatile organic emissions from paint dipping operations  

SciTech Connect

This paper provides details of a study conducted to demonstrate an equivalent method of Best Available Control Technology (BACT) compliance for volatile organic emissions from dip coating of certain miscellaneous metal parts. The study was proposed to show that the total volatile organic compound (VOC) emissions from 3.8 lb of VOC/gallon coating formulations were no greater than the total VOC emissions from 3.5 lb/gallon formulations used under the same conditions for coating steel joists. The presumptive BACT standard enforced by the Virginia Department of Environmental Quality (DEQ) for dip coating of steel joists is 3.5 lb/gallon. The requirement of 3.5 lb/gallon was derived from the US Environmental Protection Agency Guideline Series Control of Volatile Organic Emissions from Existing Stationary Sources--Volume 6: Surface Coating of Miscellaneous Metal Parts and Products. On June 5, 1998 the source completed a 12 month, full scale comparison study under a consent order with the Virginia DEQ. During the study period, the source made daily measurements of product produced, paint used, and emissions from the control and test paint tanks, and reported data to EPA and the DEQ every two months. The study concluded that a 26 percent reduction in paint usage and a 20 percent reduction in emissions was achieved in the test tanks using a 3.8 lb/gal coating compared to the control tanks using a 3.5 lb/gal coating. This study enables the source to achieve greater emission reductions than the presumptive BACT level and at the same time reduce painting costs by 34%. This study provides positive results for the environment, the steel joist industry, and the construction industry. This study could impact EPA's current Maximum Achievable Control Technology (MACT) rule development for Miscellaneous Metal Parts and Products and national VOC rules for this source category under Section 183(e) of the Clean Air Act.

Blankenship, W.R.; Pugh, C.W. Jr.

1999-07-01

51

The AcrAB-TolC Efflux Pump Contributes to Multidrug Resistance in the Nosocomial Pathogen Enterobacter aerogenes  

Microsoft Academic Search

For the last decade, Enterobacter aerogenes, a commensal gram-negative bacterium of human intestinal flora, has been rapidly emerging as an important nosocomial pathogen (14, 18). Of concern is the increasing frequency of E. aerogenes isolates that are resistant to antibiotics and antiseptics (3). Several types of systems have evolved in gram-negative bac- teria to pump deleterious molecules out of the

Elizabeth Pradel; Jean-Marie Pages

2002-01-01

52

Physical and genetic map of the chromosome of Lactococcus lactis subsp. lactis IL1403.  

PubMed Central

A combined physical and genetic map of the chromosome of Lactococcus lactis subsp. lactis IL1403 was determined. We constructed a restriction map for the NotI, ApaI, and SmaI enzymes. The order of the restriction fragments was determined by using the randomly integrative plasmid pRL1 and by performing indirect end-labeling experiments. The strain IL1403 chromosome was found to be circular and 2,420 kb in size. A total of 24 chromosomal markers were mapped on the chromosome by performing hybridization experiments with gene probes for L. lactis and various other bacteria. Integration of pRC1-derived plasmids via homologous recombination allowed more precise location of some lactococcal genes and allowed us to determine the orientation of these genes on the chromosome. Recurrent sequences, such as insertion elements and rRNA gene (rrn) clusters, were also mapped. At least seven copies of IS1076 were present and were located on 50% of the chromosome. In contrast, no copy of ISS1RS was detected. Six ribosomal operons were found on the strain IL1403 chromosome; five were located on 16% of the chromosome and were transcribed in the same direction. A comparison of the physical maps of L. lactis subsp. lactis IL1403 and DL11 showed that these two strains are closely related and that the variable regions are located mainly near the rrn gene clusters. In contrast, despite major restriction pattern dissimilarities between L. lactis IL1403 and MG1363, the overall genetic organization of the genome seems to be conserved between these two strains. Images PMID:1328163

Le Bourgeois, P; Lautier, M; Mata, M; Ritzenthaler, P

1992-01-01

53

Regulation of primary carbon metabolism in Kluyveromyces lactis  

Microsoft Academic Search

In the recent past, through advances in development of genetic tools, the budding yeast Kluyveromyces lactis has become a model system for studies on molecular physiology of so-called “Nonconventional Yeasts.” The regulation of primary carbon metabolism in K. lactis differs markedly from Saccharomyces cerevisiae and reflects the dominance of respiration over fermentation typical for the majority of yeasts. The absence

K. D Breunig; M. M Bianchi; D Bourgarel; C Falcone; I Ferrero; L Frontali; P Goffrini; J. J Krijger; C Mazzoni; C Milkowski; H. Y Steensma; A. M Zeeman

2000-01-01

54

Multivitamin production in Lactococcus lactis using metabolic engineering  

Microsoft Academic Search

The dairy starter bacterium Lactococcus lactis has the potential to synthesize both folate (vitamin B11) and riboflavin (vitamin B2). By directed mutagenesis followed by selection and metabolic engineering we have modified two complicated biosynthetic pathways in L. lactis resulting in simultaneous overproduction of both folate and riboflavin: Following exposure to the riboflavin analogue roseoflavin we have isolated a spontaneous mutant

Wilbert Sybesma; Catherine Burgess; Marjo Starrenburg; Douwe van Sinderen; Jeroen Hugenholtza

2004-01-01

55

Optimization of key process variables for enhanced hydrogen production by Enterobacter aerogenes using statistical methods  

Microsoft Academic Search

The individual and mutual effects of glucose concentration, temperature and pH on the hydrogen production by Enterobacter aerogenes were investigated in a batch system. A Box–Behnken design and response surface methodology (RSM) were employed to determine the optimum condition for enhanced hydrogen production. The hydrogen production rate was investigated by simultaneously changing the three independent variables, which all had significant

Ji Hye Jo; Dae Sung Lee; Donghee Park; Woo-Seok Choe; Jong Moon Park

2008-01-01

56

Repeated cadmium biosorption by regenerated Enterobacter aerogenes biofilm attached to activated carbon  

Microsoft Academic Search

Summary The bacteriumEnterobacter aerogenes has been used to develop a biofilm over activated carbon for biosorption from various strength cadmium solutions (25–500ppm). High bacterial resistance to metal poisoning allowed biofilm regeneration to raise the net loading of cadmium over the carbon by repeated biosorption runs.

J. A. Scott; A. M. Karanjkar

1992-01-01

57

Hydrogen production of Enterobacter aerogenes altered by extracellular and intracellular redox states  

Microsoft Academic Search

Enterobacter aerogenes HU-101, tested for its hydrogen production in batch cultures on various substrates, produced the highest amount of hydrogen when the substrate was glycerol. The yield of hydrogen is a function of the degree to which the substrates are reduced. To examine the effect of intracellular redox state on hydrogen yield, glucose-limiting chemostat cultures were carried out at various

Y. Nakashimada; M. A. Rachman; T. Kakizono; N. Nishio

2002-01-01

58

In Vivo Modification of Porin Activity Conferring Antibiotic Resistance to Enterobacter aerogenes  

Microsoft Academic Search

Cephalosporins are widely used in chemotherapy of bacterial infections and resistance mechanisms seriously impair their antibacterial activity. Several resistant strains of Enterobacter aerogenes, a frequently isolated nosocomial pathogen, were analyzed. One isolate exhibited a strong modification of the porin antigenic pattern, especially with an immunological probe directed against an epitope located inside the pore lumen. A strong decrease of cefepime

Jacqueline Chevalier; Jean-Marie Pagès; Monique Malléa

1999-01-01

59

Porin alteration and active efflux: two in vivo drug resistance strategies used by Enterobacter aerogenes  

Microsoft Academic Search

Entembacter aemgenes is among the five most frequently isolated nosocomial pathogens in France, and this bacterium also shows increasing multidrug resistance. In this study, various E. aerogenes strains isolated from hospital units were characterized for their outer-membrane proteins, antibiotic susceptibilities (inhibition diameters and MICs) and resistance mechanisms associated with modification of envelope permeability (porin alteration and active efflux). Diminished outer-membrane

Monique Mallea; Jacqueline Chevalier; Charleric Bornet; Annie Eyraud; Anne Davin-Regli; Claude Bollet; Jean-Marie Pages

1998-01-01

60

Hydrogen production by immobilized cells of aciduric Enterobacter aerogenes strain HO39  

Microsoft Academic Search

Cell immobilization of Enterobacter aerogenes strain HO-39 in agar gel or on porous glass beads was effective for hydrogen production in batch cultures of cells immobilized by the entrapment and adsorption methods. Stirring of the culture was indispensable for effective hydrogen production using cells immobilized in agar gel. However, relatively good hydrogen-production performance was obtained with cells immobilized on porous

Haruhiko Yokoi; Tadafumi Tokushige; Jun Hirose; Sachio Hayashi; Yoshiyuki Takasaki

1997-01-01

61

Rewiring Lactococcus lactis for Ethanol Production  

PubMed Central

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

Dehli, Tore; Jensen, Peter Ruhdal

2013-01-01

62

Ornithine transport and exchange in Streptococcus lactis.  

PubMed Central

Resting cells of Streptococcus lactis 133 appeared to accumulate [14C]ornithine to a high concentration in the absence of an exogenous energy source. However, analysis of intracellular amino acid pool constituents and results of transport experiments revealed that the accumulation of ornithine represented a homoexchange between extracellular [14C]ornithine and unlabeled ornithine in the cell. The energy-independent exchange of ornithine was not inhibited by proton-conducting uncouplers or by metabolic inhibitors. Intracellular [14C]ornithine was retained by resting cells after suspension in a buffered medium. However, addition of unlabeled ornithine to the suspension elicited rapid exit of labeled amino acid. The initial rate of exit of [14C]ornithine was dependent on the concentration of unlabeled ornithine in the medium, but this accelerative exchange diffusion process caused no net loss of amino acid. By contrast, the presence of a fermentable energy source caused a rapid expulsion of and net decrease in the concentration of intracellular ornithine. Kinetic analyses of amino acid transport demonstrated competitive inhibition between lysine and ornithine, and data obtained by two-dimensional thin-layer chromatography established the heteroexchange of these basic amino acids. The effects of amino acids and of ornithine analogs on both entry and exit of [14C]ornithine have been examined. The data suggest that a common carrier mediates the entry and exchange of lysine, arginine, and ornithine in cells of S. lactis. Images PMID:3114235

Thompson, J

1987-01-01

63

Citrate can partially replace carbon dioxide required for growth of Lactococcus lactis subsp. lactis biovar diacetylactis.  

PubMed

Lactococcus lactis subsp. lactis biovar diacetylactis was grown as batch cultures on a chemically defined medium. No growth was observed when the cultures were sparged with pure nitrogen (1.3 l l-1 min-1) whereas the cultures displayed exponential growth in the presence of minute amounts of carbon dioxide (0.035 mol-% of the inlet gas). However, in the former case, the addition of citrate restored growth. This suggested that oxaloacetate required for aspartate biosynthesis can be formed by the carboxylation of pyruvate or by citrate catabolism. When the cultures were heavily sparged with nitrogen (2.6 l l-1 min-1), no growth was observed even in the presence of citrate. This indicated that growth in these conditions was repressed by the absence of carbon dioxide required in some other biosynthetic reaction than in the carboxylation of pyruvate leading to oxaloacetate/aspartate biosynthesis. PMID:10792674

Henriksen, C M; Curic, M; Nilsson, D

2000-05-01

64

Production of adenine arabinoside by gel-entrapped cells of Enterobacter aerogenes in water-organic cosolvent system  

Microsoft Academic Search

Gel-entrapped whole cells of Enterobacter aerogenes, which has a transglycosylation activity, were used to produce adenine arabinoside from uracil arabinoside and adenine, in an appropriate water-organic cosolvent system. Cells of E. aerogenes entrapped with a hydrophilic photo-crosslinkable resin prepolymer, ENT-4000, or a urethane prepolymer, PU-6, had a high and stable transglycosylation activity. To improve the poor solubility in water of

Kenzo Yokozekil; Shigeru Yamanaka; Takashi Utagawa; Koichi Takinami; Yoshio Hirose; Atsuo Tanaka; Kenji Sonomoto; Saburo Fukui

1982-01-01

65

Bifidobacterium animalis subsp. lactis strains isolated from dog faeces.  

PubMed

The aim of the study was to identify and characterize dog bifidobacterial isolates and compare them with commercial probiotic strains. Sixteen isolates of Bifidobacterium animalis ssp. lactis from dog faeces (German Shepherd Dog) were identified by subspecies-specific PCR, MALDI-TOF MS and sequencing. This study is the first describing B. animalis ssp. lactis occurring within the intestinal tract of dogs. Our dog isolates showed slightly different fingerprinting profiles obtained by RAPD-PCR and REP-PCR from those isolated from yogurt and type strains of B. animalis ssp. lactis. Both, dog and yogurt origin strains indicated survival in the simulated in vitro digestion assay and were resistant to low pH and bile salts. Moreover, strong auto-aggregation activity was observed only in dog origin B. animalis ssp. lactis strains. Dog strains showed good properties predicting their survival ability in GIT and could be tested as a potential new probiotics for dogs or other hosts. PMID:22749610

Bunešová, V?ra; Vlková, Eva; Rada, Vojt?ch; Ro?ková, Sárka; Svobodová, Ivona; Jebavý, Lukáš; Kme?, Vladimír

2012-12-01

66

Microencapsulation of Bifidobacterium lactis for incorporation into soft foods  

Microsoft Academic Search

Summary Micro-encapsulation of the probiotic micro-organism Bifidobacterium lactis isolated from a bio-yoghurt starter culture, was carried out using a mixture of hydrated gellan and xanthan gums. Rheological studies showed that the gum mix was suitable for encapsulation of B. lactis, for incorporation into soft foods\\/beverages. The gel behaved as a non-Newtonian material, and the flow curve fitted well to the

L. D. McMaster; S. A. Kokott

2005-01-01

67

Optimization of key process variables for enhanced hydrogen production by Enterobacter aerogenes using statistical methods.  

PubMed

The individual and mutual effects of glucose concentration, temperature and pH on the hydrogen production by Enterobacter aerogenes were investigated in a batch system. A Box-Behnken design and response surface methodology (RSM) were employed to determine the optimum condition for enhanced hydrogen production. The hydrogen production rate was investigated by simultaneously changing the three independent variables, which all had significant influences on the hydrogen production rate. The maximum hydrogen production rate of 425.8 ml H(2)(g dry cell h)(-1) was obtained under the optimum condition of glucose concentration 118.06 mM, temperature 38 degrees C and pH 6.13. The experimental results showed that the RSM with the Box-Behnken design was a useful tool for achieving high rate of hydrogen production by E. aerogenes. PMID:17582761

Jo, Ji Hye; Lee, Dae Sung; Park, Donghee; Choe, Woo-Seok; Park, Jong Moon

2008-04-01

68

Engineered Enterobacter aerogenes for efficient utilization of sugarcane molasses in 2,3-butanediol production.  

PubMed

Sugarcane molasses is considered to be a good carbon source for biorefinery due to its high sugar content and low price. Sucrose occupies more than half of the sugar in the molasses. Enterobacter aerogenes is a good host strain for 2,3-butanediol production, but its utilization of sucrose is not very efficient. To improve sucrose utilization in E. aerogenes, a sucrose regulator (ScrR) was disrupted from the genomic DNA. The deletion mutation increased the sucrose consumption rate significantly when sucrose or sugarcane molasses was used as a carbon source. The 2,3-butanediol production from sugarcane molasses by the mutant was enhanced by 60% in batch fermentation compared to that by the wild type strain. In fed-batch fermentation, 98.69 g/L of 2,3-butanediol production was achieved at 36 h. PMID:23644066

Jung, Moo-Young; Park, Bu-Soo; Lee, Jinwon; Oh, Min-Kyu

2013-07-01

69

[Evaluation of aerogenic occupational health risk for workers engaged into periclase-carbon refractories production].  

PubMed

The work is aimed to evaluate aerogenic occupational health risk for workers engaged into preparation and formation of technologic mass in periclase-carbon refractories production, using organic binding agent according to criteria R 2.2.2006-05 and R 2.2.1716-03. Occupational dust is a complicated chemical mixture containing manganum oxide, phenol, formaldehyde, aerosols containing silicon, benzpyrene (if "Carbores" binding agent used). Hygienic evaluation revealed occupational health risk due to occupational dust at workplaces of runners operator, press operator, batching feeder, crane operator. Aerogenic occupational risk at workplace of grinder operator is assessed as negligibly small (tolerable). Experimental and epidemiologic studies prove probable (proof category 1B) occupational risk of respiratory disease at the studied production. PMID:25282807

Drugova, O G; Rosly?, O F

2014-01-01

70

Control of the activity and synthesis of aspartate transcarbamylase in Aerobacter aerogenes  

E-print Network

Zi and A. aerogenes. Inhibition by CTP and a sigmoidal velocity- substrate plot were demonstrated when aspartate concentration was varied. The effects of heat-treatment of ATCase resulted in in- f creased inhibition by CIP, a phenomenon unlike that seen... CONTROL OF ENZYME SYNTHESIS. DISCUSSION. . ~ ~ LITERATURE CITED. VITA. 11 20 26 32 34 vii LIST OF TABLES Tab le l. Inhibition of ATCase. Page 19 2. Levels of ATCase specific activity of wild type strain PRLR3 and mutant strain HA14...

Deutsch, Walter Andrew

2012-06-07

71

Aerogenic vaccination with a Burkholderia mallei auxotroph protects against aerosol-initiated glanders in mice  

Microsoft Academic Search

Burkholderia mallei is an obligate mammalian pathogen that causes the zoonotic disease glanders. Two live attenuated B. mallei strains, a capsule mutant and a branched-chain amino acid auxotroph, were evaluated for use as vaccines against aerosol-initiated glanders in mice. Animals were aerogenically vaccinated and serum samples were obtained before aerosol challenge with a high-dose (>300 times the LD50) of B.

Ricky L. Ulrich; Kei Amemiya; David M. Waag; Chad J. Roy; David DeShazer

2005-01-01

72

Synergistic effect of Candida maltosa HY-35 and Enterobacter aerogenes W-23 on hydrogen production  

Microsoft Academic Search

An aciduric high-yielding hydrogen yeast named Candida maltosa HY-35 was screened, which can grow and produce hydrogen at pH 1.3. Further research was carried out batchwise to measure the hydrogen-producing ability of a mixed culture of C. maltosa HY-35 and a facultative anaerobe Enterobacter aerogenes W-23. In this method, with the mixed culture of these two strains at 35?C for

Wenyu Lu; Jianping Wen; Yu Chen; Bing Sun; Xiaoqiang Jia; Minghui Liu; Qinggele Caiyin

2007-01-01

73

Characteristics of hydrogen production by aciduric Enterobacter aerogenes strain HO39  

Microsoft Academic Search

An aciduric facultative anaerobe with a hydrogen-producing ability was isolated and identified as Enterobacter aerogenes strain HO-39. The bacterium was able to grow at acidic pH of 3.3 aerobically and at 4.0 anaerobically. Although the optimum pH for hydrogen production was 6.0 to 7.0, hydrogen could be produced at acidic pH of 4.0. The optimum temperature for hydrogen production and

Haruhiko Yokoi; Takanobu Ohkawara; Jun Hirose; Satio Hayashi; Yoshiyuki Takasaki

1995-01-01

74

Process development of continuous hydrogen production by Enterobacter aerogenes in a packed column reactor  

Microsoft Academic Search

Hydrogen bioproduction from agro-industrial residues by Enterobacter aerogenes in a continuous packed column has been investigated and a complete reactor characterization is presented. Experimental runs carried out at different residence time, liable of interest for industrial application, showed hydrogen yields ranging from 1.36 to 3.02 mmolH2mmolуglucose or, in other words, from 37.5% to 75% of the theoretical hydrogen yield. A

E. Palazzi; B. Fabiano; P. Perego

2000-01-01

75

Production of l-asparaginase in Enterobacter aerogenes expressing Vitreoscilla hemoglobin for efficient oxygen uptake  

Microsoft Academic Search

This study is the first utilizing Vitreoscilla hemoglobin in a heterologous bacterium, Enterobacter aerogenes, to determine the effect of such a highly efficient oxygen-uptake system on the production of l-asparaginase, an enzyme that has attracted considerable attention due to its anti-tumor activity. Here, we show that the Vitreoscilla hemoglobin expressing strain has from 10-fold to more than two orders of

H. Geckil; S. Gencer

2004-01-01

76

A novel bioflocculant produced by Enterobacter aerogenes and its use in defecating the trona suspension  

Microsoft Academic Search

In this paper, a bioflocculant-producing bacterium, named W-23, was isolated from soil and identified as Enterobacter aerogenes. The bioflocculant (named WF-1) produced by W-23 was an acidic polysaccharide composed mainly of uronic acid (13.2%), pyruvic acid (7.4%) and acetic acid (1.6%). The three components sugars of WF-1 were glucose, fructose and manose, and the molar ratio was 10.3:5.4:1 for glucose:fructose:manose.

Wen-Yu Lu; Tong Zhang; Dong-Yan Zhang; Cai-Hong Li; Jian-Ping Wen; Lian-Xiang Du

2005-01-01

77

Enterobacter aerogenes OmpX, a cation-selective channel mar- and osmo-regulated  

Microsoft Academic Search

The ompX gene of Enterobacter aerogenes was cloned. Its overexpression induced a decrease in the major porin Omp36 production and consequently a ?-lactam resistance was noted. Purified outer membrane protein X (OmpX) was reconstituted into artificial membranes and formed ion channels with a conductance of 20 pS in 1 M NaCl and a cationic selectivity. Both MarA expression and high

Myrielle Dupont; Emmanuelle Dé; Renaud Chollet; Jacqueline Chevalier; Jean-Marie Pagès

2004-01-01

78

2,3Butanediol production by Enterobacter aerogenes in continuous culture: role of oxygen supply  

Microsoft Academic Search

The influence of oxygen on growth and production of 2,3-butanediol and acetoin by Enterobacter aerogenes was studied in continuous culture. At all dilution rates (D) studied cell mass increased steadily with increasing oxygen uptake rate (OUR), hence the micro-aerobic cultivation was energy-limited. The biomass yield on oxygen increased with D which suggests that cells need more energy for maintenance functions

An-Ping Zeng; Hanno Biebl; Wolf-Dieter Deckwer

1990-01-01

79

Biodegradation of 2-methylquinoline by Enterobacter aerogenes TJ-D isolated from activated sludge.  

PubMed

Bacterial strain Enterobacter aerogenes TJ-D capable of utilizing 2-methylquinoline as the sole carbon and energy source was isolated from acclimated activated sludge under denitrifying conditions. The ability to degrade 2-methylquinoline by E. aerogenes TJ-D was investigated under denitrifying conditions. Under optimal conditions of temperature (35 degrees C) and initial pH 7, 2-methylquinoline of 100 mg/L was degraded within 176 hr. The degradation of 2-methylquinoline by E. aerogenes TJ-D could be well described by the Haldane model (R2 > 0.91). During the degradation period of 2-methylquinoline (initial concentration 100 mg/L), nitrate was almost completely consumed (the removal efficiency was 98.5%), while nitrite remained at low concentration (< 0.62 mg/L) during the whole denitrification period. 1,2,3,4-Tetrahydro-2-methylquinoline, 4-ethyl-benzenamine, N-butyl-benzenamine, N-ethyl-benzenamine and 2,6-diethyl-benzenamine were metabolites produced during the degradation. The degradation pathway of 2-methylquinoline by E. aerogenes TJ-D was proposed. 2-Methylquinoline is initially hydroxylated at C-4 to form 2-methyl-4-hydroxy-quinoline, and then forms 2-methyl-4-quinolinol as a result of tautomerism. Hydrogenation of the heterocyclic ring at positions 2 and 3 produces 2,3-dihydro-2-methyl-4-quinolinol. The carbon-carbon bond at position 2 and 3 in the heterocyclic ring may cleave and form 2-ethyl-N-ethyl-benzenamine. Tautomerism may result in the formation of 2,6-diethyl-benzenamine and N-butyl-benzenamine. 4-Ethyl-benzenamine and N-ethyl-benzenamine were produced as a result of losing one ethyl group from the above molecules. PMID:24218841

Wang, Lin; Li, Yongmei; Duan, Jingyuan

2013-07-01

80

Rapid detection of a gfp-marked Enterobacter aerogenes under anaerobic conditions by aerobic fluorescence recovery  

Microsoft Academic Search

A gfp- and kanamycin-resistance gene-containing plasmid pUCGK was successfully constructed and transformed into Enterobacter aerogenes to develop a rapid GFP-based method for quantifying the bacterial concentration under anaerobic conditions for production of biohydrogen. Since the use of GFP as a molecular reporter is restricted by its requirement for oxygen in the development of the fluorophore, fluorescence detection for the fluorescent

Chong Zhang; Xin-Hui Xing; Kai Lou

2005-01-01

81

H2 production from starch by a mixed culture of Clostridium butyricum and Enterobacter aerogenes  

Microsoft Academic Search

A mixed continuous culture of Clostridium butyricum and Enterobacter aerogenes removed O2 in a reactor and produced H2 from starch with yield of more than 2 mol H2\\/mol glucose without any reducing agents in the medium. Co-immobilized cells of the bacteria on porous glass beads evolved H2 from starch at 1.3 l\\/l.h, with H2 yield of 2.6 mol H2\\/ mol

Haruhiko Yokoi; Tadafumi Tokushige; Jun Hirose; Sachio Hayashi; Yoshiyuki Takasaki

1998-01-01

82

Nucleotide Sequence of the Chromosomal ampC Gene of Enterobacter aerogenes  

Microsoft Academic Search

The AmpC b-lactamase gene and a small portion of the regulatory ampR sequence of Enterobacter aerogenes 97B were cloned and sequenced. The b-lactamase had an isoelectric point of 8 and conferred cephalosporin and cephamycin resistance on the host. The sequence of the cloned gene is most closely related to those of the ampC genes of E. cloacae and C. freundii.

KAREN E. PRESTON; CHRISTOPHER C. A. RADOMSKI; RICHARD A. VENEZIA

2000-01-01

83

Fluorescence monitoring during cultivation of Enterobacter aerogenes at different oxygen levels  

Microsoft Academic Search

On-line monitoring of NAD(P)H fluorescence and 2D fluorescence spectroscopy was performed with Enterobacter aerogenes, a bacterium sensitive to oxygen availability. The organism was grown in a reactor under low and high dissolved oxygen concentrations\\u000a and circulated through a bypass attached to the reactor. Under low dissolved oxygen concentration in the reactor, NAD(P)H\\u000a fluorescence in the reactor and the bypass showed

J. Mukherjee; C. Lindemann; T. Scheper

1999-01-01

84

Studies on nutritional and oxygen requirements for production of L-asparaginase by Enterobacter aerogenes  

Microsoft Academic Search

The carbon and nitrogen sources most suitable for L-asparaginase production by Enterobacter aerogenes were selected and their concentrations optimized in shake-flask cultures. Sodium citrate (1.0%) and diammonium hydrogen phosphate\\u000a (0.16%) proved to be the best sources of carbon and nitrogen, respectively. Nitrogen catabolite repression of enzyme formation\\u000a was absent in this bacterium. Cultivation in a reactor showed that the dissolved

J. Mukherjee; S. Majumdar; T. Scheper

2000-01-01

85

Effects of Carbon Source and Vitreoscilla Hemoglobin (VHb) on the Production of ?-Galactosidase in Enterobacter aerogenes  

Microsoft Academic Search

At fixed concentration (0.5%), lactose and galactose acted as inducers while glucose and other tested carbon sugars showed\\u000a repression effects on ?-galactosidase production in Enterobacter aerogenes strain. The expression of Vitreoscilla hemoglobin gene (vgb) in this bacterial strain managed to overcome the repression effects as well as improving the induction of ?-galactosidase\\u000a formation by carbon sources. In parallel, the bacterial

Khaled M. Khleifat; Muayad M. Abboud; Ahmed H. Al-Mustafa; Khalid Y. Al-Sharafa

2006-01-01

86

21 CFR 184.1388 - Lactase enzyme preparation from Kluyveromyces lactis.  

Code of Federal Regulations, 2010 CFR

...2010-04-01 2009-04-01 true Lactase enzyme preparation from Kluyveromyces lactis...Affirmed as GRAS § 184.1388 Lactase enzyme preparation from Kluyveromyces lactis. (a) This enzyme preparation is derived from the...

2010-04-01

87

21 CFR 184.1388 - Lactase enzyme preparation from Kluyveromyces lactis.  

Code of Federal Regulations, 2011 CFR

...2011-04-01 2011-04-01 false Lactase enzyme preparation from Kluyveromyces lactis...Affirmed as GRAS § 184.1388 Lactase enzyme preparation from Kluyveromyces lactis. (a) This enzyme preparation is derived from the...

2011-04-01

88

21 CFR 184.1388 - Lactase enzyme preparation from Kluyveromyces lactis.  

...2014-04-01 2014-04-01 false Lactase enzyme preparation from Kluyveromyces lactis...Affirmed as GRAS § 184.1388 Lactase enzyme preparation from Kluyveromyces lactis. (a) This enzyme preparation is derived from the...

2014-04-01

89

21 CFR 184.1388 - Lactase enzyme preparation from Kluyveromyces lactis.  

Code of Federal Regulations, 2013 CFR

...2013-04-01 2013-04-01 false Lactase enzyme preparation from Kluyveromyces lactis...Affirmed as GRAS § 184.1388 Lactase enzyme preparation from Kluyveromyces lactis. (a) This enzyme preparation is derived from the...

2013-04-01

90

21 CFR 184.1388 - Lactase enzyme preparation from Kluyveromyces lactis.  

Code of Federal Regulations, 2012 CFR

...2012-04-01 2012-04-01 false Lactase enzyme preparation from Kluyveromyces lactis...Affirmed as GRAS § 184.1388 Lactase enzyme preparation from Kluyveromyces lactis. (a) This enzyme preparation is derived from the...

2012-04-01

91

Parameter Estimation of Actuators for Benchmark Active Control Technology (BACT) Wind Tunnel Model with Analysis of Wear and Aerodynamic Loading Effects  

NASA Technical Reports Server (NTRS)

This report describes the development of transfer function models for the trailing-edge and upper and lower spoiler actuators of the Benchmark Active Control Technology (BACT) wind tunnel model for application to control system analysis and design. A simple nonlinear least-squares parameter estimation approach is applied to determine transfer function parameters from frequency response data. Unconstrained quasi-Newton minimization of weighted frequency response error was employed to estimate the transfer function parameters. An analysis of the behavior of the actuators over time to assess the effects of wear and aerodynamic load by using the transfer function models is also presented. The frequency responses indicate consistent actuator behavior throughout the wind tunnel test and only slight degradation in effectiveness due to aerodynamic hinge loading. The resulting actuator models have been used in design, analysis, and simulation of controllers for the BACT to successfully suppress flutter over a wide range of conditions.

Waszak, Martin R.; Fung, Jimmy

1998-01-01

92

Ubiquity and diversity of multidrug resistance genes in Lactococcus lactis strains isolated between1936 and1995  

Microsoft Academic Search

The presence and the nucleotide sequence of four multidrug resistance genes, lmrA, lmrP, lmrC, and lmrD,were investigated in 13 strains of Lactococcus lactis ssp. lactis, four strains of Lactococcus lactis ssp. cremoris, two strains of Lactococcus plantarum, and two strains of Lactococcus raffinolactis. Multidrug resistance genes were present in all L. lactis isolates tested. However, none of them could be

Ana Bel; Anette Wind; Baltasar Mayo; Abelardo Margolles; Consejo Superior de Investigaciones; Cient ´ õficas

93

Protein secretion in Lactococcus lactis : an efficient way to increase the overall heterologous protein production  

Microsoft Academic Search

Lactococcus lactis, the model lactic acid bacterium (LAB), is a food grade and well-characterized Gram positive bacterium. It is a good candidate for heterologous protein delivery in foodstuff or in the digestive tract. L. lactis can also be used as a protein producer in fermentor. Many heterologous proteins have already been produced in L. lactis but only few reports allow

Yves Le Loir; Vasco Azevedo; Sergio C Oliveira; Daniela A Freitas; Anderson Miyoshi; Luis G Bermúdez-Humarán; Sébastien Nouaille; Luciana A Ribeiro; Sophie Leclercq; Jane E Gabriel; Maricê N Oliveira; Cathy Charlier; Michel Gautier; Philippe Langella

2005-01-01

94

Impact of osmotic stress on protein diffusion in Lactococcus lactis.  

PubMed

We measured translational diffusion of proteins in the cytoplasm and plasma membrane of the Gram-positive bacterium Lactococcus lactis and probed the effect of osmotic upshift. For cells in standard growth medium the diffusion coefficients for cytosolic proteins (27 and 582?kDa) and 12-transmembrane helix membrane proteins are similar to those in Escherichia coli. The translational diffusion of GFP in L. lactis drops by two orders of magnitude when the medium osmolality is increased by ??1.9 Osm, and the decrease in mobility is partly reversed in the presence of osmoprotectants. We find a large spread in diffusion coefficients over the full population of cells but a smaller spread if only sister cells are compared. While in general the diffusion coefficients we measure under normal osmotic conditions in L. lactis are similar to those reported in E. coli, the decrease in translational diffusion upon osmotic challenge in L. lactis is smaller than in E. coli. An even more striking difference is that in L. lactis the GFP diffusion coefficient drops much more rapidly with volume than in E. coli. We discuss these findings in the light of differences in turgor, cell volume, crowding and cytoplasmic structure of Gram-positive and Gram-negative bacteria. PMID:25244659

Mika, Jacek T; Schavemaker, Paul E; Krasnikov, Victor; Poolman, Bert

2014-11-01

95

Physical and genetic map of the Lactococcus lactis subsp. cremoris MG1363 chromosome: comparison with that of Lactococcus lactis subsp. lactis IL 1403 reveals a large genome inversion.  

PubMed Central

A physical and genetic map of the chromosome of the Lactococcus lactis subsp. cremoris reference strain MG1363 was established. The physical map was constructed for NotI, ApaI, and SmaI enzymes by using a strategy that combines creation of new rare restriction sites by the random-integration vector pRL1 and ordering of restriction fragments by indirect end-labeling experiments. The MG1363 chromosome appeared to be circular and 2,560 kb long. Seventy-seven chromosomal markers were located on the physical map by hybridization experiments. Integration via homologous recombination of pRC1-derived plasmids allowed a more precise location of some lactococcal genes and determination of their orientation on the chromosome. The MG1363 chromosome contains six rRNA operons; five are clustered within 15% of the chromosome and transcribed in the same direction. Comparison of the L. lactis subsp. cremoris MG1363 physical map with those of the two L. lactis subsp. lactis strains IL1403 and DL11 revealed a high degree of restriction polymorphism. At the genetic organization level, despite an overall conservation of gene organization, strain MG1363 presents a large inversion of half of the genome in the region containing the rRNA operons. PMID:7751295

Le Bourgeois, P; Lautier, M; van den Berghe, L; Gasson, M J; Ritzenthaler, P

1995-01-01

96

The Complete Genome Sequence of the Lactic Acid Bacterium Lactococcus lactis ssp. lactis IL1403  

PubMed Central

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

Bolotin, Alexander; Wincker, Patrick; Mauger, Stephane; Jaillon, Olivier; Malarme, Karine; Weissenbach, Jean; Ehrlich, S. Dusko; Sorokin, Alexei

2001-01-01

97

Fermentative hydrogen production by two novel strains of Enterobacter aerogenes HGN-2 and HT 34 isolated from sea buried crude oil pipelines  

Microsoft Academic Search

Present study investigated fermentative hydrogen production of two novel isolates of Enterobacter aerogenes HGN-2 and HT 34 isolated from oil water mixtures. The two isolates were identified as novel strains of E. aerogenes based on 16S rRNA gene. The batch fermentations of two strains from glucose and xylose were carried out using economical culture medium under various conditions such as

H. S. Jayasinghearachchi; Priyangshu M. Sarma; Sneha Singh; Anil Aginihotri; Ajay K. Mandal; Banwari Lal

2009-01-01

98

Mineral nutrition of Aerobacter aerogenes for valine production in a synthetic medium.  

PubMed

The effect of a number of mineral salts, like dipotassium hydrogen phosphate, magnesium sulphate, and sodium chloride, and of some trace elements including iron, copper, cobalt, nickel, molybdenum, and calcium, on the production of valine by Aerobacter aerogenes in a synthetic medium was investigated. It was found that all the mineral salts were necessary for valine formation. Among the trace elements, iron and molybdenum were found to be necessary in minute concentrations for the optimum yield of the amino acid, while all the others had an adverse effect on valine production, even at lower levels. PMID:4013530

Mukhopadhyay, A K; Majumdar, S K

1985-01-01

99

Hospital clonal dissemination of Enterobacter aerogenes producing carbapenemase KPC-2 in a Chinese teaching hospital.  

PubMed

Carbapenems are first-line agents for the treatment of serious nosocomial infections caused by multidrug-resistant Enterobacteriaceae. However, resistance to carbapenems has increased dramatically among Enterobacteriaceae in our hospital. In this study, we report clonal dissemination caused by carbapenem-resistant Enterobacter aerogenes (CREA). In 2011, CREA was identified from 12 patients admitted to the neurosurgical ward. All 12 clinical isolates were non-susceptible to cefotaxime, ceftazidime, cefoxitin, ertapenem, imipenem or meropenem. All isolates carried the gene encoding Klebsiella pneumoniae carbapenemase-2 (KPC-2), except for the isolate E4. However, a remarkably lower expression level of the porin OmpF was detected in the non-KPC-2-producing isolate E4 on SDS-PAGE compared with the carbapenem-susceptible isolate. Epidemiological and molecular investigations showed that a single E. aerogenes strain (PFGE type A), including seven KPC-2-producing clinical isolates, was primarily responsible for the first isolation and subsequent dissemination. In a case-control study, we identified risk factors for infection/colonization with CREA. Mechanical ventilation, the changing of sickbeds and previous use of broad-spectrum antibiotics were identified as potential risk factors. Our findings suggest that further studies should focus on judicious use of available antibiotics, implementation of active antibiotic resistance surveillance and strict implementation of infection-control measures to avoid the rapid spread or clonal dissemination caused by carbapenem-resistant Enterobacteriaceae in healthcare facilities. PMID:24273320

Qin, Xiaohua; Yang, Yang; Hu, Fupin; Zhu, Demei

2014-02-01

100

Study of the role of anaerobic metabolism in succinate production by Enterobacter aerogenes.  

PubMed

Succinate is a core biochemical building block; optimizing succinate production from biomass by microbial fermentation is a focus of basic and applied biotechnology research. Lowering pH in anaerobic succinate fermentation culture is a cost-effective and environmentally friendly approach to reducing the use of sub-raw materials such as alkali, which are needed for neutralization. To evaluate the potential of bacteria-based succinate fermentation under weak acidic (pH <6.2) and anaerobic conditions, we characterized the anaerobic metabolism of Enterobacter aerogenes AJ110637, which rapidly assimilates glucose at pH 5.0. Based on the profile of anaerobic products, we constructed single-gene knockout mutants to eliminate the main anaerobic metabolic pathways involved in NADH re-oxidation. These single-gene knockout studies showed that the ethanol synthesis pathway serves as the dominant NADH re-oxidation pathway in this organism. To generate a metabolically engineered strain for succinate production, we eliminated ethanol formation and introduced a heterogeneous carboxylation enzyme, yielding E. aerogenes strain ?adhE/PCK. The strain produced succinate from glucose with a 60.5% yield (grams of succinate produced per gram of glucose consumed) at pH <6.2 and anaerobic conditions. Thus, we showed the potential of bacteria-based succinate fermentation under weak acidic conditions. PMID:24962116

Tajima, Yoshinori; Kaida, Kenichi; Hayakawa, Atsushi; Fukui, Keita; Nishio, Yousuke; Hashiguchi, Kenichi; Fudou, Ryosuke; Matsui, Kazuhiko; Usuda, Yoshihiro; Sode, Koji

2014-09-01

101

Regulation of nitrogen metabolism in Escherichia coli and Klebsiella aerogenes: studies with the continuous-culture technique.  

PubMed Central

Ammonia-nitrogen-limited continuous cultures of Escherichia coli and Klebsiella aerogenes contain induced levels of glutamine synthetase that is deadenylyated (i.e., fully active). In the presence of excess ammonia or glutamate in glucose-limited cultures of E. coli, glutamine synthetase is repressed and adenylylated (inactive). The average state of adenylylation (n) is a linear function of the specific growth rate. At low specific growth rates, glutamine synthetase is adenylylated; as the specific growth rate increases, n decreases, approaching 0 to 2 at rapid growth rates. The average state of adenylylation correlates well with the intracellular concentrations and ratios of alpha-ketoglutarate and glutamine, which are key effectors in the adenylylation-deadenylylation systems. E. coli and K. aerogenes differ markedly in their growth yields, growth rates, and enzymatic composition during nitrogen limitation. The data suggest that, unlike K. aerogenes, E. coli W uses glutamate dehydrogenase to incorporate ammonia during nitrogen limitation. In E. coli, glutamate dehydrogenase is progressively induced during nitrogen limitation when mu (growth rate) approaches mumax. In contrast, in K. aerogenes glutamate dehydrogenase is repressed during nitrogen limitation, whereas glutamate synthase, an alternative supplier of glutamate to the cell, is induced. Data are presented that support the regulatory schemes proposed for the control of glutamine synthetase activity by induction-repression phenomena and adenylylation-deadenylylation reaction. We propose that the intracellular ratio of alpha-ketoglutarate to glutamine may be the most important physiological parameter in determining the activity of glutamine synthetase. PMID:238954

Senior, P J

1975-01-01

102

The Influence of Dissolved Oxygen Concentration on the Respiration and Glucose Metabolism of Klebsiella aerogenes during Growth  

Microsoft Academic Search

SUMMARY The influence of dissolved oxygen concentration on the metabolism and respiration of growing Klebsiella aerogenes NCTC 8017 was studied by means of a continuous-flow culture technique. Different dissolved oxygen tensions (equivalent partial pressures) were obtained by varying the partial pressure of oxygen in the gas phase. The respiration rate (oxygen uptake rate per unit mass organism) was independent of

D. E. F. HARRISON; S. J. PIRT

1967-01-01

103

Modification of Outer Membrane Protein Profile and Evidence Suggesting an Active Drug Pump in Enterobacter aerogenes Clinical Strains  

Microsoft Academic Search

Two clinical strains of Enterobacter aerogenes that exhibited phenotypes of multiresistance to -lactam antibiotics, fluoroquinolones, chloramphenicol, tetracycline, and kanamycin were investigated. Both strains showed a porin pattern different from that of a susceptible strain, with a drastic reduction in the amount of the major porin but with an apparently conserved normal structure (size and immunogenicity), together with overproduction of two

Stephane Gayet; Renaud Chollet; Gerard Molle; Jean-Marie Pages

2003-01-01

104

Cholate Resistance in Lactococcus lactis Is Mediated by an ATP-Dependent Multispecific Organic Anion Transporter  

Microsoft Academic Search

The cholate-resistant Lactococcus lactis strain C41-2, derived from wild-type L. lactis MG1363 through selection for growth on cholate-containing medium, displayed a reduced accumulation of cholate due to an enhanced active efflux. However, L. lactis C41-2 was not cross resistant to deoxycholate or cationic drugs, such as ethidium and rhodamine 6G, which are typical substrates of the multidrug transporters LmrP and

ATSUSHI YOKOTA; MARLOES VEENSTRA; PETER KURDI; HENDRIK W. VAN VEEN; WIL N. KONINGS

2000-01-01

105

Biological containment of genetically modified Lactococcus lactis for intestinal delivery of human interleukin 10  

Microsoft Academic Search

Genetically modified Lactococcus lactis secreting interleukin 10 provides a therapeutic approach for inflammatory bowel disease. However, the release of such genetically modified organisms through clinical use raises safety concerns. In an effort to address this problem, we replaced the thymidylate synthase gene thyA of L. lactis with a synthetic human IL10 gene. This thyA?hIL10+L. lactis strain produced human IL-10 (hIL-10),

Sabine Neirynck; Nathalie Huyghebaert; Veerle Snoeck; An Vermeire; Bruno Goddeeris; Eric Cox; Jean Paul Remon; Erik Remaut; Lothar Steidler

2003-01-01

106

Generation of Dipeptidyl Peptidase-IV-Inhibiting Peptides from ?-Lactoglobulin Secreted by Lactococcus lactis  

PubMed Central

Previous studies showed that hydrolysates of ?-lactoglobulin (BLG) prepared using gastrointestinal proteases strongly inhibit dipeptidyl peptidase-IV (DPP-IV) activity in vitro. In this study, we developed a BLG-secreting Lactococcus lactis strain as a delivery vehicle and in situ expression system. Interestingly, trypsin-digested recombinant BLG from L. lactis inhibited DPP-IV activity, suggesting that BLG-secreting L. lactis may be useful in the treatment of type 2 diabetes mellitus. PMID:25157356

Shigemori, Suguru; Oshiro, Kazushi; Wang, Pengfei; Yamamoto, Yoshinari; Wang, Yeqin; Sato, Takashi; Uyeno, Yutaka; Shimosato, Takeshi

2014-01-01

107

Effect of Methionine Sulfoximine and Methionine Sulfone on Glutamate Synthesis in Klebsiella aerogenes  

PubMed Central

At least two pathways exist in Klebsiella aerogenes for glutamate synthesis. A mutant blocked in one pathway due to the loss of glutamate dehydrogenase (gltD) does not require glutamate and has the same growth characteristics as the parent strain in most media; however, its growth is inhibited by the analogues methionine sulfoximine and methionine sulfone. Wild-type Klebsiella is resistant to 0.1 M methionine sulfoximine or methionine sulfone, whereas the gltD mutant is sensitive to 1 mM concentrations. Either glutamate or glutamine is effective in overcoming this inhibition. Activities of both glutamine synthetase and glutamate synthetase, two enzymes involved in the second pathway of glutamate synthesis, are inhibited by methionine sulfoximine and methionine sulfone. The primary effect of methionine sulfoximine appears to be the prevention of glutamine production necessary for subsequent glutamate synthesis via glutamate synthetase enzyme. PMID:4145197

Brenchley, Jean E.

1973-01-01

108

Biosensing and bioremediation of Cr(VI) by cell free extract of Enterobacter aerogenes T2.  

PubMed

Hexavalent chromium or Cr(VI) enters the environment through several anthropogenic activities and it is highly toxic and carcinogenic. Hence it is required to be detected and remediated from the environment. In this study, low-cost and environment-friendly methods of biosensing and bioremediation of Cr(VI) have been proposed. Crude cell free extract (CFE) of previously isolated Enterobacter aerogenes T2 (GU265554; NII 1111) was prepared and exploited to develop a stable biosensor for direct estimation of Cr(VI) in waste water, by using three electrodes via cyclic voltammetry. For bioremediation studies, a homogeneous solution of commercially available sodium alginate and CFE was added dropwise in a continuously stirred calcium chloride solution. Biologically modified calcium alginate beads were produced and these were further utilized for bioremediation studies. The proposed sensor showed linear response in the range of 10-40 ?g L(-1) Cr(VI) and the limit of detection was found to be 6.6 ?g L(-1) Cr(VI). No interference was observed in presence of metal ions, e.g., lead, cadmium, arsenic, tin etc., except for insignificant interference with molybdenum and manganese. In bioremediation studies, modified calcium alginate beads showed encouraging removal rate 900 mg Cr(VI)/m(3) water per day with a removal efficiency of 90%, much above than reported in literature. The proposed sensing system could be a viable alternative to costly measurement procedures. Calcium alginate beads, modified with CFE of E. aerogenes, could be used in bioremediation of Cr(VI) since it could work in real conditions with extraordinarily high capacity. PMID:24410691

Panda, Jigisha; Sarkar, Priyabrata

2014-01-01

109

Irreproducible and uninterpretable Polymyxin B MICs for Enterobacter cloacae and Enterobacter aerogenes.  

PubMed

Carbapenem-resistant Enterobacter species are emerging nosocomial pathogens. As with most multidrug-resistant Gram-negative pathogens, the polymyxins are often the only therapeutic option. In this study involving clinical isolates of E. cloacae and E. aerogenes, susceptibility testing methods with polymyxin B were analyzed. All isolates underwent testing by the broth microdilution (in duplicate) and agar dilution (in duplicate) methods, and select isolates were examined by the Etest method. Selected isolates were also examined for heteroresistance by population analysis profiling. Using a susceptibility breakpoint of ?2 ?g/ml, categorical agreement by all four dilution tests (two broth microdilution and two agar dilution) was achieved in only 76/114 (67%) of E. cloacae isolates (65 susceptible, 11 resistant). Thirty-eight (33%) had either conflicting or uninterpretable results (multiple skip wells, i.e., wells that exhibit no growth although growth does occur at higher concentrations). Of the 11 consistently resistant isolates, five had susceptible MICs as determined by Etest. Heteroresistant subpopulations were detected in eight of eight isolates tested, with greater percentages in isolates with uninterpretable MICs. For E. aerogenes, categorical agreement between the four dilution tests was obtained in 48/56 (86%), with conflicting and/or uninterpretable results in 8/56 (14%). For polymyxin susceptibility testing of Enterobacter species, close attention must be paid to the presence of multiple skip wells, leading to uninterpretable results. Susceptibility also should not be assumed based on the results of a single test. Until the clinical relevance of skip wells is defined, interpretation of polymyxin susceptibility tests for Enterobacter species should be undertaken with extreme caution. PMID:24088860

Landman, David; Salamera, Julius; Quale, John

2013-12-01

110

Treatment of Murine Colitis by Lactococcus lactis Secreting Interleukin10  

Microsoft Academic Search

The cytokine interleukin-10 (IL-10) has shown promise in clinical trials for treatment of inflammatory bowel disease (IBD). Using two mouse models, we show that the therapeutic dose of IL-10 can be reduced by localized delivery of a bacterium genetically engineered to secrete the cytokine. Intragastric administration of IL-10-secreting Lactococcus lactis caused a 50% reduction in colitis in mice treated with

Lothar Steidler; Wolfgang Hans; Lieven Schotte; Sabine Neirynck; Florian Obermeier; Werner Falk; Walter Fiers; Erik Remaut

2000-01-01

111

Mobile CRISPR/Cas-Mediated Bacteriophage Resistance in Lactococcus lactis  

PubMed Central

Lactococcus lactis is a biotechnological workhorse for food fermentations and potentially therapeutic products and is therefore widely consumed by humans. It is predominantly used as a starter microbe for fermented dairy products, and specialized strains have adapted from a plant environment through reductive evolution and horizontal gene transfer as evidenced by the association of adventitious traits with mobile elements. Specifically, L. lactis has armed itself with a myriad of plasmid-encoded bacteriophage defensive systems to protect against viral predation. This known arsenal had not included CRISPR/Cas (clustered regularly interspaced short palindromic repeats/CRISPR-associated proteins), which forms a remarkable microbial immunity system against invading DNA. Although CRISPR/Cas systems are common in the genomes of closely related lactic acid bacteria (LAB), none was identified within the eight published lactococcal genomes. Furthermore, a PCR-based search of the common LAB CRISPR/Cas systems (Types I and II) in 383 industrial L. lactis strains proved unsuccessful. Here we describe a novel, Type III, self-transmissible, plasmid-encoded, phage-interfering CRISPR/Cas discovered in L. lactis. The native CRISPR spacers confer resistance based on sequence identity to corresponding lactococcal phage. The interference is directed at phages problematic to the dairy industry, indicative of a responsive system. Moreover, targeting could be modified by engineering the spacer content. The 62.8-kb plasmid was shown to be conjugally transferrable to various strains. Its mobility should facilitate dissemination within microbial communities and provide a readily applicable system to naturally introduce CRISPR/Cas to industrially relevant strains for enhanced phage resistance and prevention against acquisition of undesirable genes. PMID:23240053

Millen, Anne M.; Horvath, Philippe; Boyaval, Patrick; Romero, Dennis A.

2012-01-01

112

Exploration of sulfur metabolism in the yeast Kluyveromyces lactis  

Microsoft Academic Search

Hemiascomycetes are separated by considerable evolutionary distances and, as a consequence, the mechanisms involved in sulfur\\u000a metabolism in the extensively studied yeast, Saccharomyces cerevisiae, could be different from those of other species of the phylum. This is the first time that a global view of sulfur metabolism\\u000a is reported in the biotechnological yeast Kluyveromyces lactis. We used combined approaches based

Agnès Hébert; Marie-Pierre Forquin-Gomez; Aurélie Roux; Julie Aubert; Christophe Junot; Valentin Loux; Jean-François Heilier; Pascal Bonnarme; Jean-Marie Beckerich; Sophie Landaud

113

The AcrAB-TolC Pump Is Involved in Macrolide Resistance but Not in Telithromycin Efflux in Enterobacter aerogenes and Escherichia coli  

Microsoft Academic Search

The role of the AcrAB-TolC pump in macrolide and ketolide susceptibility in Escherichia coli and Enterobacter aerogenes was studied. Efflux pump inhibitor restored erythromycin, clarithromycin, and telithromycin sus- ceptibilities to multidrug-resistant isolates. No modification of telithromycin accumulation was detected in E. aerogenes acrAB or tolC derivatives compared to that in the parental strain. Two independent efflux pumps, inhibited by phenylalanine

Renaud Chollet; Jacqueline Chevalier; A. Bryskier; J.-M. Pages

2004-01-01

114

Selection during Cefepime Treatment of a New Cephalosporinase Variant with Extended-Spectrum Resistance to Cefepime in an Enterobacter aerogenes Clinical Isolate  

Microsoft Academic Search

Enterobacter aerogenes resistant to cefepime (MIC, 32 g\\/ml) was isolated from a patient treated with cefepime for an infection caused by a strain of E. aerogenes overproducing its AmpC -lactamase (MIC of cefepime, 0.5 g\\/ml). The AmpC -lactamase of the resistant strain had an L-293-P amino acid substitution and a high kcat\\/Km ratio for cefepime. Both of these modifications were

G. Barnaud; Y. Benzerara; J. Gravisse; L. Raskine; M. J. Sanson-Le Pors; R. Labia; G. Arlet

2004-01-01

115

Vitreoscilla hemoglobin expressing Enterobacter aerogenes and Pseudomonas aeruginosa respond differently to carbon catabolite and oxygen repression for production of l-asparaginase, an enzyme used in cancer therapy  

Microsoft Academic Search

The production of antileukemic enzyme l-asparaginase in two distinctly related bacteria, Enterobacter aerogenes, Pseudomonas aeruginosa, and in their recombinants expressing the Vitresocilla hemoglobin (VHb) has been studied. Both bacteria showed a substantially different degree of carbon catabolite repression of the enzyme production. E. aerogenes grown under catabolite repression had more than 20-fold lower l-asparaginase activity than the controls. This figure

Hikmet Geckil; Salih Gencer; Mirac Uckun

2004-01-01

116

Phosphate/hexose 6-phosphate antiport in Streptococcus lactis.  

PubMed

After growth in appropriate media, resting cells of Streptococcus lactis 7962 showed a rapid exchange between external and internal pools of inorganic phosphate. This exchange was not found in other strains of S. lactis (ML3, 133, or K1) or in Streptococcus faecalis. Phosphate exchange in S. lactis 7962 did not require other anions or cations in the assay medium, nor was phosphate influx affected by the membrane potential and pH gradient formed during glycolysis. Thus, the exchange reaction was independent of known ionic drivers (H+, Na+, OH-, etc.). Experiments testing inhibitions of phosphate entry suggested that alternative substrates for exchange included arsenate, as well as the 6-phosphates of glucose, 2-deoxyglucose, fructose, mannose, or glucosamine, and direct studies with 2-deoxyglucose 6-phosphate verified that resting cells could accumulate this sugar phosphate to levels expected for exchange with internal phosphate. Two other observations supported the idea of an exchange between phosphate and sugar phosphate. First, early addition of the heterologous substrate blocked entry of the test compound, whereas later addition caused efflux of preaccumulated material. Second, expression of phosphate exchange and 2-deoxyglucose 6-phosphate transport varied in parallel. Both activities were found at high levels after growth in medium supplemented with rhamnose or arabinose, at intermediate levels with addition of galactose, and at low levels after growth with glucose, fructose, or mannose. We conclude that these findings describe a novel anion antiporter that mediates the exchange of phosphate (arsenate) and sugar 6-phosphates. PMID:6325388

Maloney, P C; Ambudkar, S V; Thomas, J; Schiller, L

1984-04-01

117

Chromosomal stabilization of the proteinase genes in Lactococcus lactis.  

PubMed Central

The plasmid-encoded proteinase genes prtP and prtM of Lactococcus lactis subsp. cremoris Wg2 were integrated by a Campbell-like mechanism into the L. lactis subsp. lactis MG1363 chromosome by using the insertion vector pKLG610. Two transformants were obtained that differed in the number of amplified pKLG610 copies in head-to-tail arrangements on their chromosomes; MG610 contained approximately two copies, and MG611 contained about eight copies. The amplifications were stably maintained during growth in milk in the absence of antibiotics. The proteolytic activity of strain MG611 was approximately 11-fold higher than that of strain MG610 and about 1.5 times higher than that of strain MG1363(pGKV552), which carried the proteinase genes on an autonomously replicating plasmid with a copy number of approximately 5. All three strains showed rapid growth in milk with concomitant rapid production of acid. The results suggest that a limited number of copies of the proteinase genes prtP and prtM per genome is sufficient for good growth in milk. Images PMID:1768129

Leenhouts, K J; Gietema, J; Kok, J; Venema, G

1991-01-01

118

KPC-2 carbapenemase and DHA-1 AmpC determinants carried on the same plasmid in Enterobacter aerogenes.  

PubMed

This study was conducted to analyse the presence of a plasmid-mediated carbapenem resistance mechanism in a clinical Enterobacter aerogenes isolate from a patient from Jiangsu province, People's Republic of China. PCR and sequencing confirmed that the isolate harboured Klebsiella pneumoniae carbapenemase (KPC)-2, DHA-1 and TEM-1 ?-lactamase genes. Both the KPC-2 and DHA-1 genes were transferred to Escherichia coli C600 by transconjugation, and Southern blotting confirmed that these two genes were located on the same plasmid, which was of approximately 56 kb in size. The Enterobacter aerogenes isolate was resistant to carbapenems and other tested antimicrobial agents. The Escherichia coli transconjugant showed reduced susceptibility but not resistance to carbapenems and other ?-lactams, indicating the presence of another, possibly permeability-related, resistance mechanism in the clinical isolate. PMID:24173427

Kuai, Shougang; Shao, Haifeng; Huang, Lihua; Pei, Hao; Lu, Zhonghua; Wang, Weiping; Liu, Jun

2014-03-01

119

Alteration of hydrogen metabolism of ldh- deleted Enterobacter aerogenes by overexpression of NAD(+)-dependent formate dehydrogenase  

Microsoft Academic Search

The NAD+-dependent formate dehydrogenase FDH1 gene (fdh1), cloned from Candida boidinii, was expressed in the ldh-deleted mutant of Enterobacter aerogenes IAM1183 strain. The plasmid of pCom10 driven by the PalkB promoter was used to construct the fdh1 expression system and thus introduce a new dihydronicotinamide adenine dinucleotide (NADH) regeneration pathway from formate\\u000a in the ldh-deleted mutant. The knockout of NADH-consuming

Yuan Lu; Hongxin Zhao; Chong Zhang; Qiheng Lai; Xi Wu; Xin-Hui Xing

2010-01-01

120

Expression of NAD + -dependent formate dehydrogenase in Enterobacter aerogenes and its involvement in anaerobic metabolism and H 2 production  

Microsoft Academic Search

An expression system for NAD+-dependent formate dehydrogenase gene (fdh1), from Candida boidinii, was constructed and cloned into Enterobacter aerogenes IAM1183. With the fdh1 expression, the total H2 yield was attributed to a decrease in activity of the lactate pathway and an increase of the formate pathway flux due to\\u000a the NADH regeneration. Analysis of the redox state balance and ethanol-to-acetate

Yuan Lu; Hongxin Zhao; Chong Zhang; Qiheng Lai; Xi Wu; Xin-Hui Xing

2009-01-01

121

2,3Butanediol production by Enterobacter aerogenes : selection of the optimal conditions and application to food industry residues  

Microsoft Academic Search

Optimum values of temperature, pH, and starting substrate concentration are experimentally determined for 2,3-butanediol production by Enterobacter aerogenes through three set of batch fermentations of synthetic glucose solutions. The results of tests carried out at variable temperature show an optimum of 39 °C and are used to estimate, for both fermentation and thermal inactivation, the activation enthalpies (7.19 and 23.6

P. Perego; A. Converti; A. Del Borghi; P. Canepa

2000-01-01

122

Overflow metabolism during anaerobic growth of Klebsiella aerogenes NCTC 418 on glycerol and dihydroxyacetone in chemostat culture  

Microsoft Academic Search

Klebsiella aerogenes NCTC 418 was grown anaerobically in chemostat culture with glycerol as source of carbon and energy. Glycerol-limited cultures did not ferment the carbon source with maximal efficiency but produced considerable amounts of 1,3-propanediol. The fraction of glycerol converted to this product depended on the growth rate and on the limitation: faster growing cells produced relatively more of this

H. Streekstra; M. J. Teixeira de Mattos; O. M. Neijssel; D. W. Tempest

1987-01-01

123

Detection of Extended-Spectrum b-Lactamases in Clinical Isolates of Enterobacter cloacae and Enterobacter aerogenes  

Microsoft Academic Search

The aim of the present study was to investigate the frequency of extended-spectrum b-lactamases (ESBLs) in a consecutive collection of clinical isolates of Enterobacter spp. The abilities of various screening methods to detect ESBLs in enterobacters were simultaneously tested. Among the 68 consecutive isolates (56 Enterobacter cloacae and 12 Enterobacter aerogenes isolates) that were analyzed for b-lactamase content, 21 (25

EVA TZELEPI; PANAGIOTA GIAKKOUPI; DANAI SOFIANOU; VENETA LOUKOVA; ANASTASSIA KEMEROGLOU; ATHANASSIOS TSAKRIS

2000-01-01

124

Successive Emergence of Extended-Spectrum  -Lactamase-Producing and Carbapenemase-Producing Enterobacter aerogenes Isolates in a University Hospital  

Microsoft Academic Search

Sixty-two clinical isolates of Enterobacter aerogenes resistant to expanded-spectrum cephalosporins were collected between July 2003 and May 2005. Among these isolates, 23 (37.1%) were imipenem (IPM) susceptible, and 39 (62.9%) were IPM insusceptible, of which 89.7% (35\\/39) were resistant and 10.3% (4\\/39) were inter- mediate. Isolate genotypes were compared by pulsed-field gel electrophoresis. Of 62 isolates, 48 belonged to epidemic

M. Biendo; B. Canarelli; D. Thomas; F. Rousseau; F. Hamdad; C. Adjide; G. Laurans; F. Eb

2008-01-01

125

Isolation of Enterobacter aerogenes susceptible to beta-lactam antibiotics despite high level beta-lactamase production  

Microsoft Academic Search

This report describes a patient with nosocomial meningitis from whom four distinct isolates ofEnterobacter aerogenes were recovered over a complicated course of chemotherapy. The initial isolate was susceptible to expanded spectrum ?-lactams despite constitutive production of high levels of ?-lactamase. Resistant isolates recovered during antibiotic therapy had lost a 42,000 outer membrane protein. These data suggest that b-lactam susceptibility in

M. A. Mellencamp; J. S. Roccaforte; L. C. Preheim; C. C. Sanders; C. A. Anene; M. J. Bittner

1990-01-01

126

Co-fermentation of carbon sources by Enterobacter aerogenes ATCC 29007 to enhance the production of bioethanol.  

PubMed

We investigated the enhancement of bioethanol production in Enterobacter aerogenes ATCC 29007 by co-fermentation of carbon sources such as glycerol, glucose, galactose, sucrose, fructose, xylose, starch, mannitol and citric acid. Biofuel production increases with increasing growth rate of microorganisms; that is why we investigated the optimal growth rate of E. aerogenes ATCC 29007, using mixtures of different carbon sources with glycerol. E. aerogenes ATCC 29007 was incubated in media containing each carbon source and glycerol; growth rate and bioethanol production improved in all cases compared to those in medium containing glycerol alone. The growth rate and bioethanol production were highest with mannitol. Fermentation was carried out at 37 °C for 18 h, pH 7, using 50 mL defined production medium in 100 mL serum bottles at 200 rpm. Bioethanol production under optimized conditions in medium containing 16 g/L mannitol and 20 g/L glycerol increased sixfold (32.10 g/L) than that containing glycerol alone (5.23 g/L) as the carbon source in anaerobic conditions. Similarly, bioethanol production using free cells in continuous co-fermentation also improved (27.28 g/L) when 90.37 % of 16 g/L mannitol and 67.15 % of 20 g/L glycerol were used. Although naturally existing or engineered microorganisms can ferment mixed sugars sequentially, the preferential utilization of glucose to non-glucose sugars often results in lower overall yield and productivity of ethanol. Here, we present new findings in E. aerogenes ATCC 29007 that can be used to improve bioethanol production by simultaneous co-fermentation of glycerol and mannitol. PMID:24185706

Thapa, Laxmi Prasad; Lee, Sang Jun; Yang, Xiao Guang; Yoo, Hah Young; Kim, Sung Bong; Park, Chulhwan; Kim, Seung Wook

2014-06-01

127

Inactivation of the panE Gene in Lactococcus lactis Enhances Formation of Cheese Aroma Compounds  

PubMed Central

Hydroxyacid dehydrogenases limit the conversion of ?-keto acids into aroma compounds. Here we report that inactivation of the panE gene, encoding the ?-hydroxyacid dehydrogenase activity in Lactococcus lactis, enhanced the formation of 3-methylbutanal and 3-methylbutanol. L. lactis IFPL953?panE was an efficient strain producing volatile compounds related to cheese aroma. PMID:23524675

de Cadinanos, Luz P. Gomez; Garcia-Cayuela, Tomas; Yvon, Mireille; Martinez-Cuesta, M. Carmen; Pelaez, Carmen

2013-01-01

128

Use of traditional African fermented beverages as delivery vehicles for Bifidobacterium lactis DSM 10140  

Microsoft Academic Search

A microencapsulation delivery system for Bifidobacterium lactis, a possible probiotic suited for use by the rural population of South Africa, was evaluated using two existing traditional fermented foods, amasi and mahewu. Gellan\\/xanthan microcapsules containing viable B. lactis, were tested under simulated physiological conditions, and added to pasteurized beverages. The capsules protected the organism under simulated low pH conditions associated with

L. D. McMaster; S. A. Kokott; S. J. Reid; V. R. Abratt

2005-01-01

129

Mutant telomeres inhibit transcriptional silencing at native telomeres of the yeast Kluyveromyces lactis  

Microsoft Academic Search

We report the identification and characterization of transcriptional silencing at native telomeres in the budding yeast Kluyveromyces lactis. We show that K. lactis telomeres are able to repress the transcription of a gene located at the junction between the telomeric repeat tract and the subtelomeric domain. As in Saccharomyces cerevisiae, switching between the repressed and derepressed transcriptional states occurs. C-terminal

R. Gurevich; S. Smolikov; H. Maddar; A. Krauskopf

2003-01-01

130

Engineering of carbon distribution between glycolysis and sugar nucleobiosynthesis in Lactococcus lactis  

Microsoft Academic Search

We describe the effects of modulating the activities of glucokinase, phosphofructokinase, and phosphoglu- comutase on the branching point between sugar degradation and the biosynthesis of sugar nucleotides involved in the production of exopolysaccharide biosynthesis by Lactococcus lactis. This was realized by using a described isogenic L. lactis mutant with reduced enzyme activities or by controlled expression of the well-characterized genes

Ingeborg C. Boels; Michiel Kleerebezem; Vos de W. M

2003-01-01

131

moaR, a gene that encodes a positive regulator of the monoamine regulon in Klebsiella aerogenes.  

PubMed Central

We cloned and sequenced a Klebsiella aerogenes gene (moaR) for activation of arylsulfatase synthesis by tyramine. This gene was cloned by complementation of a K. aerogenes mutant in which tyramine fails to relieve the arylsulfatase repression caused by sulfur compounds. The moaR gene also activated induction of the synthesis of both tyramine oxidase and the 30-kDa protein that is specifically induced by high concentrations of tyramine or catecholamines. The moaR gene on the chromosome of the wild-type strain of K. aerogenes was disrupted by homologous recombination with a plasmid containing the inactivated moaR. The resultant mutant showed the same phenotype as previously isolated atsT mutant strains that are negative for the derepressed synthesis of arylsulfatase. In this mutant strain, tyramine also failed to induce the synthesis of tyramine oxidase or the production of a 30-kDa protein. The moaR gene is capable of encoding a protein of 26,238 Da. The putative MoaR protein has a helix-turn-helix motif in its C terminus. Thus, it seems likely that the MoaR protein regulates the operons by binding to the regulatory region of the monoamine regulon. The MoaR protein is subject to autogenous control, which was shown by use of a moaR'-lacZ transcriptional fusion. Images PMID:8407801

Azakami, H; Sugino, H; Yokoro, N; Iwata, N; Murooka, Y

1993-01-01

132

Generalized model of the effect of pH on lactate fermentation and citrate bioconversion in Lactococcus lactis ssp. Lactis biovar. diacetylactis  

Microsoft Academic Search

An aroma-imparting mesophilic lactic starter (Lactococcus lactis ssp. lactis biovar. diacetylactis) was studied in batch culture in medium with 50 g·l-1 lactose and 2 g·l-1 citrate. The effect of pH on the physiology of growth and the production of flavour compounds was investigated with a mathematical model. The specific rates of growth and of lactose fermentation obeyed a law of

R. Cachon; C. Diviés

1994-01-01

133

Characterization of an insertion sequence-like element identified in plasmid pCIT264 from Lactococcus lactis subsp. lactis biovar diacetylactis  

Microsoft Academic Search

Plasmid pCIT264 from Lactococcus lactis subsp. lactis biovar diacetylactis (L. diacetylactis) contains an insertion sequence (IS)-like element located in the citrate utilization (citQRP) cluster. This 967-nucleotide long element is bounded by 17 bp perfect inverted repeats and contains an open reading frame (ORF1) composed of 296 codons, which could encode a transposase. Expression of the IS from pCIT264 generates two

Christian Magni; Félix López de Felipe; Paloma López; Diego de Mendoza

1996-01-01

134

Generalized model of the effect of pH on lactate fermentation and citrate bioconversion in Lactococcus lactis ssp. lactis biovar. diacetylactis  

Microsoft Academic Search

An aroma-imparting mesophilic lactic starter ( Lactococcus lactis ssp. lactis biovar. diacetylactis) was studied in batch culture in medium with 50?g·l –-1 lactose and 2?g·l –-1 citrate. The effect of pH on the physiology of growth and the production of flavour compounds was investigated with a mathematical model. The specific rates of growth and of lactose fermentation obeyed a law

R. Cachon; C. Diviès

1994-01-01

135

Influence of growth conditions on the production of a nisin-like bacteriocin by Lactococcus lactis subsp. lactis A164 isolated from kimchi  

Microsoft Academic Search

The influence of growth parameters on the fermentative production of a nisin-like bacteriocin by Lactococcus lactis subsp. lactis A164 isolated from kimchi was studied. The bacteriocin production was greatly affected by carbon and nitrogen sources. Strain A164 produced at least 4-fold greater bacteriocin in M17 broth supplemented with lactose than other carbon sources. The amount of 3% yeast extract was

Chan-Ick Cheigh; Hak-Jong Choi; Hoon Park; Seong-Bo Kim; Moo-Chang Kook; Tae-Seok Kim; Jae-Kwan Hwang; Yu-Ryang Pyun

2002-01-01

136

Optimization of cultural conditions for conversion of glycerol to ethanol by Enterobacter aerogenes S012.  

PubMed

The aim of this research is to optimize the cultural conditions for the conversion of glycerol to ethanol by Enterobacter aerogenes S012. Taguchi method was used to screen the cultural conditions based on their signal to noise ratio (SN). Temperature (°C), agitation speed (rpm) and time (h) were found to have the highest influence on both glycerol utilization and ethanol production by the organism while pH had the lowest. Full factorial design, statistical analysis, and regression model equation were used to optimize the selected cultural parameters for maximum ethanol production. The result showed that fermentation at 38°C and 200 rpm for 48 h would be ideal for the bacteria to produce maximum amount of ethanol from glycerol. At these optimum conditions, ethanol production, yield and productivity were 25.4 g/l, 0.53 g/l/h, and 1.12 mol/mol-glycerol, repectively. Ethanol production increased to 26.5 g/l while yield and productivity decreased to 1.04 mol/mol-glycerol and 0.37 g/l/h, respectively, after 72 h. Analysis of the fermentation products was performed using HPLC, while anaerobic condition was created by purging the fermentation vessel with nitrogen gas. PMID:23388539

Nwachukwu, Raymond E S; Shahbazi, Abolghasem; Wang, Lijun; Worku, Mulumebet; Ibrahim, Salam; Schimmel, Keith

2013-01-01

137

Optimization of cultural conditions for conversion of glycerol to ethanol by Enterobacter aerogenes S012  

PubMed Central

The aim of this research is to optimize the cultural conditions for the conversion of glycerol to ethanol by Enterobacter aerogenes S012. Taguchi method was used to screen the cultural conditions based on their signal to noise ratio (SN). Temperature (°C), agitation speed (rpm) and time (h) were found to have the highest influence on both glycerol utilization and ethanol production by the organism while pH had the lowest. Full factorial design, statistical analysis, and regression model equation were used to optimize the selected cultural parameters for maximum ethanol production. The result showed that fermentation at 38°C and 200 rpm for 48 h would be ideal for the bacteria to produce maximum amount of ethanol from glycerol. At these optimum conditions, ethanol production, yield and productivity were 25.4 g/l, 0.53 g/l/h, and 1.12 mol/mol-glycerol, repectively. Ethanol production increased to 26.5 g/l while yield and productivity decreased to 1.04 mol/mol-glycerol and 0.37 g/l/h, respectively, after 72 h. Analysis of the fermentation products was performed using HPLC, while anaerobic condition was created by purging the fermentation vessel with nitrogen gas. PMID:23388539

2013-01-01

138

Engineering Signal Peptides for Enhanced Protein Secretion from Lactococcus lactis  

PubMed Central

Lactococcus lactis is an attractive vehicle for biotechnological production of proteins and clinical delivery of therapeutics. In many such applications using this host, it is desirable to maximize secretion of recombinant proteins into the extracellular space, which is typically achieved by using the native signal peptide from a major secreted lactococcal protein, Usp45. In order to further increase protein secretion from L. lactis, inherent limitations of the Usp45 signal peptide (Usp45sp) must be elucidated. Here, we performed extensive mutagenesis on Usp45sp to probe the effects of both the mRNA sequence (silent mutations) and the peptide sequence (amino acid substitutions) on secretion. We screened signal peptides based on their resulting secretion levels of Staphylococcus aureus nuclease and further evaluated them for secretion of Bacillus subtilis ?-amylase. Silent mutations alone gave an increase of up to 16% in the secretion of ?-amylase through a mechanism consistent with relaxed mRNA folding around the ribosome binding site and enhanced translation. Targeted amino acid mutagenesis in Usp45sp, combined with additional silent mutations from the best clone in the initial screen, yielded an increase of up to 51% in maximum secretion of ?-amylase while maintaining secretion at lower induction levels. The best sequence from our screen preserves the tripartite structure of the native signal peptide but increases the positive charge of the n-region. Our study presents the first example of an engineered L. lactis signal peptide with a higher secretion yield than Usp45sp and, more generally, provides strategies for further enhancing protein secretion in bacterial hosts. PMID:23124224

Ng, Daphne T. W.

2013-01-01

139

Complete Genome Sequence of the Prototype Lactic Acid Bacterium Lactococcus lactis subsp. cremoris MG1363?  

PubMed Central

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

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

2007-01-01

140

Staphylococcus aureus virulence and metabolism are dramatically affected by Lactococcus lactis in cheese matrix.  

PubMed

In complex environments such as cheeses, the lack of relevant information on the physiology and virulence expression of pathogenic bacteria and the impact of endogenous microbiota has hindered progress in risk assessment and control. Here, we investigated the behaviour of Staphylococcus aureus, a major foodborne pathogen, in a cheese matrix, either alone or in the presence of Lactococcus lactis, as a dominant species of cheese ecosystems. The dynamics of S. aureus was explored in situ by coupling a microbiological and, for the first time, a transcriptomic approach. Lactococcus lactis affected the carbohydrate and nitrogen metabolisms and the stress response of S. aureus by acidifying, proteolysing and decreasing the redox potential of the cheese matrix. Enterotoxin expression was positively or negatively modulated by both L. lactis and the cheese matrix itself, depending on the enterotoxin type. Among the main enterotoxins involved in staphylococcal food poisoning, sea expression was slightly favoured in the presence of L. lactis, whereas a strong repression of sec4 was observed in cheese matrix, even in the absence of L. lactis, and correlated with a reduced saeRS expression. Remarkably, the agr system was downregulated by the presence of L. lactis, in part because of the decrease in pH. This study highlights the intimate link between environment, metabolism and virulence, as illustrated by the influence of the cheese matrix context, including the presence of L. lactis, on two major virulence regulators, the agr system and saeRS. PMID:23761280

Cretenet, Marina; Nouaille, Sébastien; Thouin, Jennifer; Rault, Lucie; Stenz, Ludwig; François, Patrice; Hennekinne, Jacques-Antoine; Piot, Michel; Maillard, Marie Bernadette; Fauquant, Jacques; Loubière, Pascal; Loir, Yves Le; Even, Sergine

2011-06-01

141

Sequencing and transcriptional analysis of the biosynthesis gene cluster of putrescine-producing Lactococcus lactis.  

PubMed

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

Ladero, Victor; Rattray, Fergal P; Mayo, Baltasar; Martín, María Cruz; Fernández, María; Alvarez, Miguel A

2011-09-01

142

Sequencing and Transcriptional Analysis of the Biosynthesis Gene Cluster of Putrescine-Producing Lactococcus lactis ? †  

PubMed Central

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

Ladero, Victor; Rattray, Fergal P.; Mayo, Baltasar; Martin, Maria Cruz; Fernandez, Maria; Alvarez, Miguel A.

2011-01-01

143

Heterologous expression and characterization of an N-acetyl-?-D-hexosaminidase from Lactococcus lactis ssp. lactis IL1403.  

PubMed

The lnbA gene of Lactococcus lactis ssp. lactis IL1403 encodes a polypeptide with similarity to lacto-N-biosidases and N-acetyl-?-D-hexosaminidases. The gene was cloned into the expression vector pET-21d and overexpressed in Escherichia coli BL21* (DE3). The recombinant purified enzyme (LnbA) was a monomer with a molecular weight of approximately 37 kDa. Studies with chromogenic substrates including p-nitrophenyl N-acetyl-?-D-glucosamine (pNP-GlcNAc) and p-nitrophenyl N-acetyl-?-D-galactosamine (pNP-GalNAc) showed that the enzyme had both N-acetyl-?-D-glucosaminidase and N-acetyl-?-D-galactosaminidase activity, thus indicating that the enzyme is an N-acetyl-?-D-hexosaminidase. K(m) and k(cat) for pNP-GlcNAc were 2.56 mM and 26.7 s(-1), respectively, whereas kinetic parameters for pNP-GalNAc could not be determined due to the K(m) being very high (>10 mM). The optimal temperature and pH of the enzyme were 37 °C and 5.5, respectively, for both substrates. The half-life of activity at 37 °C and pH 6.0 was 53 h, but activity was completely abolished after 30 min at 50 °C, meaning that the enzyme has relatively low temperature stability. The enzyme was stable in the pH 5.5-8 range and was unstable at pH below 5.5. Studies with natural substrates showed hydrolytic activity on chito-oligosaccharides but not on colloidal chitin or chitosan. Transglycosylation products were not detected. In all, the data suggest that LnbA's role may be to degrade chito-oligosaccharides that are produced by the previously described chitinolytic system of L. lactis. PMID:22356128

Nguyen, Hoang Anh; Nguyen, Thu-Ha; K?en, Vladimír; Eijsink, Vincent G H; Haltrich, Dietmar; Peterbauer, Clemens K

2012-03-28

144

Transcriptional analysis of oligosaccharide utilization by Bifidobacterium lactis Bl-04  

PubMed Central

Background Probiotic bifidobacteria in combination with prebiotic carbohydrates have documented positive effects on human health regarding gastrointestinal disorders and improved immunity, however the selective routes of uptake remain unknown for most candidate prebiotics. The differential transcriptomes of Bifidobacterium animalis subsp. lactis Bl-04, induced by 11 potential prebiotic oligosaccharides were analyzed to identify the genetic loci involved in the uptake and catabolism of ?- and ?-linked hexoses, and ?-xylosides. Results The overall transcriptome was modulated dependent on the type of glycoside (galactosides, glucosides or xylosides) utilized. Carbohydrate transporters of the major facilitator superfamily (induced by gentiobiose and ?-galacto-oligosaccharides (GOS)) and ATP-binding cassette (ABC) transporters (upregulated by cellobiose, GOS, isomaltose, maltotriose, melibiose, panose, raffinose, stachyose, xylobiose and ?-xylo-oligosaccharides) were differentially upregulated, together with glycoside hydrolases from families 1, 2, 13, 36, 42, 43 and 77. Sequence analysis of the identified solute-binding proteins that determine the specificity of ABC transporters revealed similarities in the breadth and selectivity of prebiotic utilization by bifidobacteria. Conclusion This study identified the differential gene expression for utilization of potential prebiotics highlighting the extensive capabilities of Bifidobacterium lactis Bl-04 to utilize oligosaccharides. Results provide insights into the ability of this probiotic microbe to utilize indigestible carbohydrates in the human gastrointestinal tract. PMID:23663691

2013-01-01

145

Nutritional requirements and media development for Lactococcus lactis IL1403.  

PubMed

Lactic acid bacteria are extensively used in food technology and for the production of various compounds, but they are fastidious in nutrient requirements. In order to elucidate the role of each component precisely, defined multicomponent media are required. This study focuses on determining nutrient auxotrophies and minimizing media components (amino acids, vitamins, metal ions, buffers and additional compounds) for the cultivation of Lactococcus lactis subsp. lactis IL1403, using microtitre plates and test tubes. It was shown that glutamine and asparagine were the most important media components for achieving higher biomass yields while the branched-chain amino acids were necessary to increase specific growth rate. The amino acid and glucose ratio was reduced to achieve minimal residual concentration of amino acids in the medium after the growth of cells, whereas the specific growth rate and biomass yield of cells were not considerably affected. As the percentage of each consumed amino acid compared to initial amount is larger than measurement error, these optimized media are important for achieving more precise data about amino acid utilization and metabolism. PMID:24626960

Aller, Kadri; Adamberg, Kaarel; Timarova, Veronica; Seiman, Andrus; Feštšenko, Darja; Vilu, Raivo

2014-07-01

146

An evaluation and partial characterization of a bacteriocin produced by Lactococcus lactis subsp lactis ST1 isolated from goat milk  

PubMed Central

A bacteriocin-like inhibitory substance producing Lactococcus lactis subsp lactis strain, ST1, isolated from goat milk of Iranian origin and with broad spectrum of activity and desirable technical properties was used for evaluating some futures of bacteriocin inhibitory activity. Cell growth and bacteriocin production studies were carried out in MRS medium incubated statically under uncontrolled pH condition. The antibacterial activity presented a primary metabolite pattern and showed a rapid decrease at the stationary phase. Microaerobiosis and capnophily growth conditions resulted in higher bacteriocin production while aerobiosis showed negative effect on both cell growth and bacteriocin production. Bacteriocin production, on the other hand, was favored in MRS broth (pH; 6.5) inoculated with 0.1 ml l-1 fresh culture when incubation was carried out at 30 °C. This indicated that the conditions resulted in higher levels of growth were frequently favoring bacteriocin production by ST1 as well. Decrease in activity, at the stationary growth phase, was much pronounced in favored growth condition. Nutrient depletion, deferent effect of low pH on bacteriocin production and/or protein degradation seemed more responsible for this phenomenon. The study also provided further data on new method for bacteriocin release from the cell wall of producer. It was clearly shown that both heating and ultrasound shock for 5 min at pH 2 could increase bacteriocin activity significantly. The release was more pronounced in the presence of 0.5% Tween80. PMID:24031976

Taheri, Parinaz; Samadi?, Nasrin; Ehsani, Mohammad Reza; Khoshayand, Mohammad Reza; Jamalifar, Hossein

2012-01-01

147

Characterization of bacteriocin HV219, produced by Lactococcus lactis subsp. lactis HV219 isolated from human vaginal secretions.  

PubMed

Bacteriocin HV219, produced by Lactococcus lactis subsp. lactis HV219, is active against Gram-positive and Gram-negative bacteria. Activity was lost when treated with proteolytic enzymes, SDS, Triton X-114 and Triton X-100, but not at pH 2.0 to 10.0 or after 20 min at 121 degrees C. Growth in the presence of yeast extract as sole nitrogen source yielded 3200 AU/ml. No bacHV219 activity was recorded in MRS broth with maltose, mannose, lactose or sucrose as sole carbohydrate, but fructose yielded 1600 AU/ml. K(2)HPO(4) at 10.0 g/l yielded 3200 AU/ml. Addition of 1.0 mg/l cyanocobalamin, l-ascorbic acid and thiamine to MRS broth yielded 3200 AU/ml, 1600 AU/ml and 1600 AU/ml, respectively. The mode of activity is bacteriolytic, as confirmed by atomic force microscopy. PMID:16721875

Todorov, Svetoslav D; Danova, Svetla T; Van Reenen, Carol A; Meincken, Martina; Dinkova, Galina; Ivanova, Iskra V; Dicks, Leon M T

2006-01-01

148

Glucose/citrate cometabolism in Lactococcus lactis subsp. lactis biovar diacetylactis with impaired alpha-acetolactate decarboxylase.  

PubMed

The pyruvate metabolism of a Lactococcus lactis subsp. lactis biovar diacetylactis mutant deficient in alpha-acetolactate decarboxylase and its wild-type strain was studied during batch cultivations. A chemically defined medium was used containing glucose as carbon- and energy-source. The alpha-acetolactate decarboxylase deficiency had no effect on the specific growth rate. Addition of citrate was found to increase the specific growth rate of both strains under aerobic and anaerobic conditions. The product formation was monitored throughout the cultivations. The carbon- and redox-balances were within the accuracy of the experimental data. When citrate was added, alpha-acetolactate, diacetyl, and acetoin were formed, and aeration was shown to have a positive effect on the formation of these metabolites. By omitting lipoic acid (required for a functional pyruvate dehydrogenase complex) from the growth medium, a similar stimulatory effect on alpha-acetolactate, diacetyl, and acetoin formation was observed under aerobic conditions. The strain with impaired alpha-acetolactate decarboxylase activity accumulated alpha-acetolactate which resulted in an increased diacetyl formation compared to the wild-type strain, under aerobic and anaerobic conditions. PMID:10937822

Curic, M; de Richelieu, M; Henriksen, C M; Jochumsen, K V; Villadsen, J; Nilsson, D

1999-10-01

149

Genetic analysis of regions of the Lactococcus lactis subsp. lactis plasmid pRS01 involved in conjugative transfer.  

PubMed

The genes responsible for conjugative transfer of the 48.4-kb Lactococcus lactis subsp. lactis ML3 plasmid pRS01 were localized by insertional mutagenesis. Integration of the IS946-containing plasmid pTRK28 into pRS01 generated a pool of stable cointegrates, including a number of plasmids altered in conjugative proficiency. Mapping of pTRK28 insertions and phenotypic analysis of cointegrate plasmids identified four distinct regions (Tra1, Tra2, Tra3, and Tra4) involved in pRS01 conjugative transfer. Tra3 corresponds closely to a region previously identified (D. G. Anderson and L. L. McKay, J. Bacteriol. 158:954-962, 1984). Another region (Tra4) was localized within an inversion sequence shown to correlate with a cell aggregation phenotype. Tra1 and Tra2, two previously unidentified regions, were located at a distance of 9 kb from Tra3. When provided in trans, a cloned portion of the Tra3 region complemented Tra3 mutants. PMID:7811081

Mills, D A; Choi, C K; Dunny, G M; McKay, L L

1994-12-01

150

Effect of selenium-enriched exopolysaccharide produced by Lactococcus lactis subsp. lactis on signaling molecules in mouse spleen lymphocytes.  

PubMed

Selenium-enriched exopolysaccharides (Se-EPS) produced by Lactococcus lactis subsp. lactis can be used as a safe and effective selenium (Se) supplement. Analysis of the Se content and monosaccharide components of Se-EPS demonstrated that it consisted of mannose, fucose, ribose, glucose, galactose, and arabinose with a molar ratio of 5.48 : 0.39 : 9.77 : 4.03 : 1.00 : 1.92, and an Se content of 183.263 ?g g(-1). The differing effects of Se-EPS and EPS on calcium channels and certain key secondary messengers in spleen lymphocytes were examined and compared. Results showed that low-dose Se-EPS, but not EPS, increased the intracellular calcium ([Ca(2+)]i) levels of mouse spleen lymphocytes. Se-EPS also increased the expression and phosphorylation of Ca(2+)-calmodulin-dependent kinase (CaMK) II in lymphocytes. In addition, increased intracellular Ca(2+) levels were also found in the cells with blocked Ca(2+) channels. We speculated that Se-EPS enhanced the activation and proliferation of lymphocytes through calcium signaling by drawing from extracellular and intracellular stores. Low-dose Se-EPS also enhanced NO production, cAMP levels and PKA activity. We speculated that low-dose Se-EPS may activate certain pathways, including the calcium channel, NO, cAMP, and PKA related pathways. PMID:24056684

Liu, Lu; Pan, Daodong; Zeng, Xiaoqun; Li, Hua

2013-10-01

151

Antilisterial Activity of Nisin-Like Bacteriocin-Producing Lactococcus lactis subsp. lactis Isolated from Traditional Sardinian Dairy Products  

PubMed Central

With the aim of selecting LAB strains with antilisterial activity to be used as protective cultures to enhance the safety of dairy products, the antimicrobial properties of 117 Lactococcus lactis subsp. lactis isolated from artisanal Sardinian dairy products were evaluated, and six strains were found to produce bacteriocin-like substances. The capacity of these strains to antagonize Listeria monocytogenes during cocultivation in skimmed milk was evaluated, showing a reduction of L. monocytogenes counts of approximately 4 log units compared to the positive control after 24?h of incubation. In order for a strain to be used as bioprotective culture, it should be carefully evaluated for the presence of virulence factors, to determine what potential risks might be involved in its use. None of the strains tested was found to produce biogenic amines or to possess haemolytic activity. In addition, all strains were sensitive to clinically important antibiotics such as ampicillin, tetracycline, and vancomycin. Our results suggest that these bac+ strains could be potentially applied in cheese manufacturing to control the growth of L. monocytogenes. PMID:22536018

Cosentino, Sofia; Fadda, Maria Elisabetta; Deplano, Maura; Melis, Roberta; Pomata, Rita; Pisano, Maria Barbara

2012-01-01

152

21 CFR 184.1985 - Aminopeptidase enzyme preparation derived from lactococcus lactis.  

Code of Federal Regulations, 2013 CFR

...2013-04-01 false Aminopeptidase enzyme preparation derived from lactococcus...GRAS § 184.1985 Aminopeptidase enzyme preparation derived from lactococcus lactis. (a) Aminopeptidase enzyme preparation is derived from the...

2013-04-01

153

21 CFR 184.1985 - Aminopeptidase enzyme preparation derived from lactococcus lactis.  

Code of Federal Regulations, 2012 CFR

...2012-04-01 false Aminopeptidase enzyme preparation derived from lactococcus...GRAS § 184.1985 Aminopeptidase enzyme preparation derived from lactococcus lactis. (a) Aminopeptidase enzyme preparation is derived from the...

2012-04-01

154

21 CFR 184.1985 - Aminopeptidase enzyme preparation derived from lactococcus lactis.  

Code of Federal Regulations, 2011 CFR

...2011-04-01 false Aminopeptidase enzyme preparation derived from lactococcus...GRAS § 184.1985 Aminopeptidase enzyme preparation derived from lactococcus lactis. (a) Aminopeptidase enzyme preparation is derived from the...

2011-04-01

155

21 CFR 184.1985 - Aminopeptidase enzyme preparation derived from lactococcus lactis.  

...2014-04-01 false Aminopeptidase enzyme preparation derived from lactococcus...GRAS § 184.1985 Aminopeptidase enzyme preparation derived from lactococcus lactis. (a) Aminopeptidase enzyme preparation is derived from the...

2014-04-01

156

Comparative Genomics of Bifidobacterium animalis subsp. lactis Reveals a Strict Monophyletic Bifidobacterial Taxon  

PubMed Central

Strains of Bifidobacterium animalis subsp. lactis are extensively exploited by the food industry as health-promoting bacteria, although the genetic variability of members belonging to this taxon has so far not received much scientific attention. In this article, we describe the complete genetic makeup of the B. animalis subsp. lactis Bl12 genome and discuss the genetic relatedness of this strain with other sequenced strains belonging to this taxon. Moreover, a detailed comparative genomic analysis of B. animalis subsp. lactis genomes was performed, which revealed a closely related and isogenic nature of all currently available B. animalis subsp. lactis strains, thus strongly suggesting a closed pan-genome structure of this bacterial group. PMID:23645200

Milani, Christian; Duranti, Sabrina; Lugli, Gabriele Andrea; Bottacini, Francesca; Strati, Francesco; Arioli, Stefania; Foroni, Elena; Turroni, Francesca; van Sinderen, Douwe

2013-01-01

157

Lactococcus lactis M4, a potential host for the expression of heterologous proteins  

Microsoft Academic Search

Background  Many plasmid-harbouring strains of Lactococcus lactis have been isolated from milk and other sources. Plasmids of Lactococcus have been shown to harbour antibiotic resistance genes and those that express some important proteins. The generally regarded\\u000a as safe (GRAS) status of L. lactis also makes it an attractive host for the production of proteins that are beneficial in numerous applications such

Nanyan Noreen; Wei Yeng Hooi; Ali Baradaran; Mohamad Rosfarizan; Chin Chin Sieo; Khatijah Yusoff; Abdul Rahim Raha

2011-01-01

158

Adaptation and response of Bifidobacterium animalis subsp. lactis to bile: a proteomic and physiological approach.  

PubMed

Bile salts are natural detergents that facilitate the digestion and absorption of the hydrophobic components of the diet. However, their amphiphilic nature makes them very inhibitory for bacteria and strongly influences bacterial survival in the gastrointestinal tract. Adaptation to and tolerance of bile stress is therefore crucial for the persistence of bacteria in the human colonic niche. Bifidobacterium animalis subsp. lactis, a probiotic bacterium with documented health benefits, is applied largely in fermented dairy products. In this study, the effect of bile salts on proteomes of B. animalis subsp. lactis IPLA 4549 and its bile-resistant derivative B. animalis subsp. lactis 4549dOx was analyzed, leading to the identification of proteins which may represent the targets of bile salt response and adaptation in B. animalis subsp. lactis. The comparison of the wild-type and the bile-resistant strain responses allowed us to hypothesize about the resistance mechanisms acquired by the derivative resistant strain and about the bile salt response in B. animalis subsp. lactis. In addition, significant differences in the levels of metabolic end products of the bifid shunt and in the redox status of the cells were also detected, which correlate with some differences observed between the proteomes. These results indicate that adaptation and response to bile in B. animalis subsp. lactis involve several physiological mechanisms that are jointly dedicated to reduce the deleterious impact of bile on the cell's physiology. PMID:17827318

Sánchez, Borja; Champomier-Vergès, Marie-Christine; Stuer-Lauridsen, Birgitte; Ruas-Madiedo, Patricia; Anglade, Patricia; Baraige, Fabienne; de los Reyes-Gavilán, Clara G; Johansen, Eric; Zagorec, Monique; Margolles, Abelardo

2007-11-01

159

Adaptation and Response of Bifidobacterium animalis subsp. lactis to Bile: a Proteomic and Physiological Approach?  

PubMed Central

Bile salts are natural detergents that facilitate the digestion and absorption of the hydrophobic components of the diet. However, their amphiphilic nature makes them very inhibitory for bacteria and strongly influences bacterial survival in the gastrointestinal tract. Adaptation to and tolerance of bile stress is therefore crucial for the persistence of bacteria in the human colonic niche. Bifidobacterium animalis subsp. lactis, a probiotic bacterium with documented health benefits, is applied largely in fermented dairy products. In this study, the effect of bile salts on proteomes of B. animalis subsp. lactis IPLA 4549 and its bile-resistant derivative B. animalis subsp. lactis 4549dOx was analyzed, leading to the identification of proteins which may represent the targets of bile salt response and adaptation in B. animalis subsp. lactis. The comparison of the wild-type and the bile-resistant strain responses allowed us to hypothesize about the resistance mechanisms acquired by the derivative resistant strain and about the bile salt response in B. animalis subsp. lactis. In addition, significant differences in the levels of metabolic end products of the bifid shunt and in the redox status of the cells were also detected, which correlate with some differences observed between the proteomes. These results indicate that adaptation and response to bile in B. animalis subsp. lactis involve several physiological mechanisms that are jointly dedicated to reduce the deleterious impact of bile on the cell's physiology. PMID:17827318

Sanchez, Borja; Champomier-Verges, Marie-Christine; Stuer-Lauridsen, Birgitte; Ruas-Madiedo, Patricia; Anglade, Patricia; Baraige, Fabienne; de los Reyes-Gavilan, Clara G.; Johansen, Eric; Zagorec, Monique; Margolles, Abelardo

2007-01-01

160

Variations in bile tolerance among Lactococcus lactis strains derived from different sources.  

PubMed

Lactococcus lactis subsp. lactis has been isolated from the intestines of marine fish and is a candidate probiotic for aquaculture. In order to use the bacterium as a probiotic, properties such as bile tolerance need to be assessed. Here, we compared bile tolerance in L. lactis strains derived from different sources. Three L. lactis subsp. lactis strains from marine fish (MFL), freshwater fish (FFL), and cheese starter (CSL) were used along with an Lactococcus lactis subsp. cremoris strain from cheese starter (CSC). The four strains were grown under various culture conditions: deMan-Rogosa-Sharpe (MRS) broth containing bile salts/acids, MRS agar containing oxgall, and phosphate-buffered saline (PBS) containing fish bile. Survival/growth of the strains in the presence of sodium cholate and sodium deoxycholate varied in the order MFL, CSL > CSC > FFL; in the presence of sodium taurocholate, the order was MFL > CSL > CSC > FFL. In liquid media containing various concentrations of oxgall, survival of the strains was observed in the order MFL > CSL > FFL and CSC. The survival of MFL was not affected by bile collected from the goldfish (Carassius auratus subsp. auratus) or the puffer fish (Takifugu niphobles), although the other strains showed significant inhibition of growth. It is a novel and beneficial finding that MFL has the highest resistance to bile acid. PMID:24395331

Takanashi, Shihori; Miura, Ai; Abe, Koko; Uchida, Junya; Itoi, Shiro; Sugita, Haruo

2014-07-01

161

Antigenicity and Immunogenicity of Rotavirus VP6 Protein Expressed on the Surface of Lactococcus lactis  

PubMed Central

Group A rotaviruses are the major etiologic agents of acute gastroenteritis worldwide in children and young animals. Among its structural proteins, VP6 is the most immunogenic and is highly conserved within this group. Lactococcus lactis is a food-grade, Gram-positive, and nonpathogenic lactic acid bacteria that has already been explored as a mucosal delivery system of heterologous antigens. In this work, the nisin-controlled expression system was used to display the VP6 protein at the cell surface of L. lactis. Conditions for optimal gene expression were established by testing different nisin concentrations, cell density at induction, and incubation times after induction. Cytoplasmic and cell wall protein extracts were analyzed by Western blot and surface expression was confirmed by flow cytometry. Both analysis provided evidence that VP6 was efficiently expressed and displayed on the cell surface of L. lactis. Furthermore, the humoral response of mice immunized with recombinant L. lactis was evaluated and the displayed recombinant VP6 protein proved to be immunogenic. In conclusion, this is the first report of displaying VP6 protein on the surface of L. lactis to induce a specific immune response against rotavirus. These results provide the basis for further evaluation of this VP6-displaying L. lactis as a mucosal delivery vector in a mouse model of rotavirus infection. PMID:23984337

Esteban, L. E.; Temprana, C. F.; Arguelles, M. H.; Glikmann, G.; Castello, A. A.

2013-01-01

162

Effects of carbon source and Vitreoscilla hemoglobin (VHb) on the production of beta-galactosidase in Enterobacter aerogenes.  

PubMed

At fixed concentration (0.5%), lactose and galactose acted as inducers while glucose and other tested carbon sugars showed repression effects on beta-galactosidase production in Enterobacter aerogenes strain. The expression of Vitreoscilla hemoglobin gene (vgb) in this bacterial strain managed to overcome the repression effects as well as improving the induction of beta-galactosidase formation by carbon sources. In parallel, the bacterial O(2) consumption was increased correspondingly to the vgb induction of beta-galactosidase synthesis. When Enterobacter aerogenes strains were grown at the incubation temperature 42 degrees C, about 5-fold higher enzyme productivity was obtained than with a similar incubation at 37 degrees C. The bacterial growth expressed as biomass yield had a different optimum temperature and was not influenced to the same extent by variations in the carbon sources. These data are discussed in terms of proposed enhancement in beta-galactosidase productivity by vgb expression as well as its significance to improve the technology of whey processing. PMID:16972134

Khleifat, Khaled M; Abboud, Muayad M; Al-Mustafa, Ahmed H; Al-Sharafa, Khalid Y

2006-10-01

163

Biohydrogen and polyhydroxyalkanoate co-production by Enterobacter aerogenes and Rhodobacter sphaeroides from Calophyllum inophyllum oil cake.  

PubMed

The feasibility of coupled biohydrogen and polyhydroxyalkanoate production by Enterobacter aerogenes and Rhodobacter sphaeroides using Calophyllum inophyllum oil cake was studied under dark and photo fermentation conditions. The utilization of a non-edible acidic oil cake (C. inophyllum), and exploitation of a modified minimal salt media led to reduction in the cost of media. Cost of fermentation is reduced by implementation of alternate dark-photo fermentative periods and through the use of a co-culture consisting of a dark fermentative (E. aerogenes) and a photo fermentative (R. sphaeroides) bacterium. The biohydrogen and polyhydroxyalkanoate produced were 7.95 L H2/L media and 10.73 g/L media, respectively, under alternate dark and photo fermentation and were 3.23 L H2/L media and 5.6g/L media, respectively under complete dark fermentation. The characteristics of the oil cake and alternate dark (16 h) and photo (8h) fermentative conditions were found to be supportive in producing high biohydrogen and polyhydroxyalkanoate (PHA) yield. PMID:24859207

Arumugam, A; Sandhya, M; Ponnusami, V

2014-07-01

164

Characterization of a bacteriocin produced by Lactococcus lactis subsp. lactis CRL 1584 isolated from a Lithobates catesbeianus hatchery.  

PubMed

Lactococcus lactis CRL 1584 isolated from a Lithobates catesbeianus hatchery inhibits the growth of Citrobacter freundii (a bullfrog pathogen) and Listeria monocytogenes by a synergistic effect between lactic acid, hydrogen peroxide and a bacteriocin-like molecule. The chemical characterization of the bacteriocin in cell-free supernatants indicates that it has a proteinaceous nature. Hexadecane and ethyl acetate did not modify the bacteriocin activity, while 10 and 20 % (v/v) chloroform decreased the activity by 29 and 43 %, respectively. The antimicrobial peptide was heat stable since 85 % of residual activity was detected when neutralized supernatants were heated at 80 °C for 30 min. Moreover, no bacteriocin inactivation was observed when supernatants were kept at -20 °C for 3 months. The synthesis of the bacteriocin was associated with bacterial growth, highest production (2,100 AU/ml) being detected at the end of the exponential growth phase. At pH ranges of 5-6.5 and 5.0-5.5 the inhibitory molecule was stable when stored for 2 days at 4 and 25 °C, respectively. Moreover, it had a bactericidal effect on L. monocytogenes and the ultrastructural studies of pathogenic cells revealed clumping of the cytoplasmic material, increased periplasmic space and cell wall modifications. The deduced amino acid sequence of the bacteriocin was identical to nisin Z and the genetic determinants for its production are harbored in the chromosome. These results, described for the first time in L. lactis from a bullfrog hatchery, will increase knowledge of the bacteriocin under study with a view to its potential inclusion in probiotics for raniculture or biopreservatives. PMID:24150985

Pasteris, Sergio E; Vera Pingitore, Esteban; Ale, Cesar E; Nader-Macías, María E Fatima

2014-03-01

165

Use of carbon and energy balances in the study of the anaerobic metabolism of Enterobacter aerogenes at variable starting glucose concentrations  

Microsoft Academic Search

The anaerobic metabolism of Enterobacter aerogenes was studied in batch culture at increasing initial glucose levels (9.0So -1). The ultimate concentrations of fermentation products were utilized to check a metabolic flux analysis based on simple carbon mass and energy balances that promise to be suitable for the study of different fermentation processes, either under aerobic or anaerobic conditions. The stoichiometric

A. Converti; P. Perego

2002-01-01

166

Lactose metabolism in Streptococcus lactis: studies with a mutant lacking glucokinase and mannose-phosphotransferase activities  

SciTech Connect

A mutant of Streptococcus lactis 133 has been isolated that lacks both glucokinase and phosphoenolpyruvate-dependent mannose- phosphotransferase (mannose-PTS) activities. The double mutant S. lactis 133 mannose-PTSd GK- is unable to utilize either exogenously supplied or intracellularly generated glucose for growth. Fluorographic analyses of metabolites formed during the metabolism of (/sup 14/C)lactose labeled specifically in the glucose or galactosyl moiety established that the cells were unable to phosphorylate intracellular glucose. However, cells of S. lactis 133 mannose-PTSd GK- readily metabolized intracellular glucose 6-phosphate, and the growth rates and cell yield of the mutant and parental strains on sucrose were the same. During growth on lactose, S. lactis 133 mannose-PTSd GK- fermented only the galactose moiety of the disaccharide, and 1 mol of glucose was generated per mol of lactose consumed. For an equivalent concentration of lactose, the cell yield of the mutant was 50% that of the wild type. The specific rate of lactose utilization by growing cells of S. lactis 133 mannose-PTSd GK- was ca. 50% greater than that of the wild type, but the cell doubling times were 70 and 47 min, respectively. High-resolution /sup 31/P nuclear magnetic resonance studies of lactose transport by starved cells of S. lactis 133 and S. lactis 133 mannose-PTSd GK- showed that the latter cells contained elevated lactose-PTS activity. Throughout exponential growth on lactose, the mutant maintained an intracellular steady-state glucose concentration of 100 mM.

Thompson, J.; Chassy, B.M.; Egan, W.

1985-04-01

167

Cloning and nucleotide sequence of a negative regulator gene for Klebsiella aerogenes arylsulfatase synthesis and identification of the gene as folA.  

PubMed Central

A negative regulator gene for synthesis of arylsulfatase in Klebsiella aerogenes was cloned. Deletion analysis showed that the regulator gene was located within a 1.6-kb cloned segment. Transfer of the plasmid, which contains the cloned fragment, into constitutive atsR mutant strains of K. aerogenes resulted in complementation of atsR; the synthesis of arylsulfatase was repressed in the presence of inorganic sulfate or cysteine, and this repression was relieved, in each case, by the addition of tyramine. The nucleotide sequence of the 1.6-kb fragment was determined. From the amino acid sequence deduced from the DNA sequence, we found two open reading frames. One of them lacked the N-terminal region but was highly homologous to the gene which codes for diadenosine tetraphosphatase (apaH) in Escherichia coli. The other open reading frame was located counterclockwise to the apaH-like gene. This gene was highly homologous to the gene which codes for dihydrofolate reductase (folA) in E. coli. We detected 30 times more activity of dihydrofolate reductase in the K. aerogenes strains carrying the plasmid, which contains the arylsulfatase regulator gene, than in the strains without plasmid. Further deletion analysis showed that the K. aerogenes folA gene is consistent with the essential region required for the repression of arylsulfatase synthesis. Transfer of a plasmid containing the E. coli folA gene into atsR mutant cells of K. aerogenes resulted in repression of the arylsulfatase synthesis. Thus, we conclude that the folA gene codes a negative regulator for the ats operon. PMID:1551851

Azakami, H; Sugino, H; Murooka, Y

1992-01-01

168

Complete Genome Sequences of Four Novel Lactococcus lactis Phages Distantly Related to the Rare 1706 Phage Species  

PubMed Central

Lactoccocus lactis is a Gram-positive bacterium widely used in the dairy industry in the production of an array of cheeses and other fermented milk products. Here, we describe the sequencing and genome annotations of a set of four phages virulent to L. lactis and exhibiting similarities to phage 1706. PMID:25013130

Vogensen, Finn K.; Heller, Knut J.; S?rensen, S?ren J.; Hansen, Lars H.

2014-01-01

169

Transcriptomic response of Lactococcus lactis in mixed culture with Staphylococcus aureus.  

PubMed

The mechanisms of interaction between Lactococcus lactis and the food pathogen Staphylococcus aureus are of crucial importance, as one major role of lactic acid bacteria (LAB) in fermented foods is to inhibit undesirable and pathogenic flora. It was never questioned if the presence of a pathogen can actively modify the gene expression patterns of LAB in a shared environment. In this study, transcriptome and biochemical analyses were combined to assess the dynamic response of L. lactis in a mixed culture with S. aureus. The presence of S. aureus hardly affected the growth of L. lactis but dramatically modified its gene expression profile. The main effect was related to earlier carbon limitation and a concomitantly lower growth rate in the mixed culture due to the consumption of glucose by both species. More specific responses involved diverse cellular functions. Genes associated with amino acid metabolism, ion transport, oxygen response, menaquinone metabolism, and cell surface and phage expression were differentially expressed in the mixed culture. This study led to new insights into possible mechanisms of interaction between L. lactis and S. aureus. Moreover, new and unexpected effects of L. lactis on the virulence of S. aureus were discovered, as described elsewhere (S. Even, C. Charlier, S. Nouaille, N. L. Ben Zakour, M. Cretenet, F. J. Cousin, M. Gautier, M. Cocaign-Bousquet, P. Loubière, and Y. Le Loir, Appl. Environ. Microbiol. 75:4459-4472, 2009). PMID:19429566

Nouaille, Sébastien; Even, Sergine; Charlier, Cathy; Le Loir, Yves; Cocaign-Bousquet, Muriel; Loubière, Pascal

2009-07-01

170

Nisin Production by a Mixed-Culture System Consisting of Lactococcus lactis and Kluyveromyces marxianus  

PubMed Central

To control the pH during antimicrobial peptide (nisin) production by a lactic acid bacterium, Lactococcus lactis subsp. lactis (ATCC11454), a novel method involving neither addition of alkali nor a separation system such as a ceramic membrane filter and electrodialyzer was developed. A mixed culture of L. lactis and Kluyveromyces marxianus, which was isolated from kefir grains, was utilized in the developed system. The interaction between lactate production by L. lactis and its assimilation by K. marxianus was used to control the pH. To utilize the interaction of these microorganisms to maintain high-level production of nisin, the kinetics of growth of, and production of lactate, acetate, and nisin by, L. lactis were investigated. The kinetics of growth of and lactic acid consumption by K. marxianus were also investigated. Because the pH of the medium could be controlled by the lactate consumption of K. marxianus and the specific lactate consumption rate of K. marxianus could be controlled by changing the dissolved oxygen (DO) concentration, a cascade pH controller coupled with DO control was developed. As a result, the pH was kept constant because the lactate level was kept low and nisin accumulated in the medium to a high level compared with that attained using other pH control strategies, such as with processes lacking pH control and those in which pH is controlled by addition of alkali. PMID:10388714

Shimizu, Hiroshi; Mizuguchi, Taiji; Tanaka, Eiji; Shioya, Suteaki

1999-01-01

171

Transcriptomic Response of Lactococcus lactis in Mixed Culture with Staphylococcus aureus? †  

PubMed Central

The mechanisms of interaction between Lactococcus lactis and the food pathogen Staphylococcus aureus are of crucial importance, as one major role of lactic acid bacteria (LAB) in fermented foods is to inhibit undesirable and pathogenic flora. It was never questioned if the presence of a pathogen can actively modify the gene expression patterns of LAB in a shared environment. In this study, transcriptome and biochemical analyses were combined to assess the dynamic response of L. lactis in a mixed culture with S. aureus. The presence of S. aureus hardly affected the growth of L. lactis but dramatically modified its gene expression profile. The main effect was related to earlier carbon limitation and a concomitantly lower growth rate in the mixed culture due to the consumption of glucose by both species. More specific responses involved diverse cellular functions. Genes associated with amino acid metabolism, ion transport, oxygen response, menaquinone metabolism, and cell surface and phage expression were differentially expressed in the mixed culture. This study led to new insights into possible mechanisms of interaction between L. lactis and S. aureus. Moreover, new and unexpected effects of L. lactis on the virulence of S. aureus were discovered, as described elsewhere (S. Even, C. Charlier, S. Nouaille, N. L. Ben Zakour, M. Cretenet, F. J. Cousin, M. Gautier, M. Cocaign-Bousquet, P. Loubière, and Y. Le Loir, Appl. Environ. Microbiol. 75:4459-4472, 2009). PMID:19429566

Nouaille, Sebastien; Even, Sergine; Charlier, Cathy; Le Loir, Yves; Cocaign-Bousquet, Muriel; Loubiere, Pascal

2009-01-01

172

Immunomodulatory effect of Lactococcus lactis JCM5805 on human plasmacytoid dendritic cells.  

PubMed

Plasmacytoid dendritic cells (pDCs) play a crucial role in anti-viral immunity through production of large amounts of interferons (IFNs). A previous study revealed the existence of lactic acid bacteria that directly stimulate pDCs in mice. In this study, we demonstrated that Lactococcus lactis JCM5805 activates human pDCs and induces IFN production in vitro. In addition, our randomized, placebo-controlled, double blind test showed that yogurt fermented with L. lactis JCM5805 activated pDC activity in vivo. This effect was greater in low pDC subjects, and their ability to produce IFNs was increased from the beginning. Furthermore, the risk of morbidity from the common cold was suppressed in the L. lactis JCM5805 group compared with the placebo group. In conclusion, intake of L. lactis JCM5805 can directly activate pDCs and increase the ability to produce IFNs in vivo. Therefore, L. lactis JCM5805 may be a beneficial tool to enhance anti-viral immunity in humans. PMID:24239838

Sugimura, Tetsu; Jounai, Kenta; Ohshio, Konomi; Tanaka, Takaaki; Suwa, Masahiro; Fujiwara, Daisuke

2013-12-01

173

Lactococcus lactis subsp. tructae subsp. nov. isolated from the intestinal mucus of brown trout (Salmo trutta) and rainbow trout (Oncorhynchus mykiss).  

PubMed

The species Lactococcus lactis currently includes three subspecies; L. lactis subsp. lactis and L. lactis subsp. cremoris, isolated from milk sources, and L. lactis subsp. hordniae, isolated from the leafhopper Hordnia circellata. In this study, three strains, designated L105(T), I3 and L101, were isolated from the intestinal mucus of brown trout (Salmo trutta) and rainbow trout (Oncorhynchus mykiss). These strains were closely related to members of the species Lactococcus lactis. Strain L105(T) showed 99.4 % 16S rRNA gene sequence similarity to that of the type strains L. lactis subsp. lactis NCDO 604(T) and L. lactis subsp. hordniae NCDO 2181(T) and showed 99.9 % similarity to the type strain Lactococcus lactis subsp. cremoris NCDO 607(T). Analysis of two housekeeping genes, rpoB and recA, confirmed the close relationship between the novel strains and L. lactis subsp. cremoris with similarities of 99.3 and 99.7 %, respectively. The three strains could, however, be differentiated from their closest relatives on the basis of several phenotypic characteristics, as was the case for L. lactis subsp. lactis and L. lactis subsp. hordniae, which were also closely related on the basis of 16S rRNA, rpoB and recA gene sequence similarities. The strains isolated in this study represent a new subspecies, for which the name Lactococcus lactis subsp. tructae subsp. nov. is proposed. The type strain is L105(T) (?= LMG 24662(T) ?= DSM 21502(T)). PMID:20833888

Pérez, Tania; Balcázar, José Luis; Peix, Alvaro; Valverde, Angel; Velázquez, Encarna; de Blas, Ignacio; Ruiz-Zarzuela, Imanol

2011-08-01

174

In vitro fermentation of prebiotic oligosaccharides by Bifidobacterium lactis HN019 and Lactobacillus spp.  

PubMed

The utilisation of various prebiotic oligosaccharides by probiotic strains of Bifidobacterium lactis, Lactobacillus rhamnosus and Lactobacillus acidophilus was investigated in order to determine the synbiotic potential of various prebiotic/probiotic combinations. Analysis by HPLC and high-performance anion-exchange chromatography of the cell-free medium taken during growth of the three probiotic bacteria showed differences in the consumption of the various oligosaccharides. Analysis of galactooligosaccharides showed that both L. rhamnosus and B. lactis consumed mostly mono- and di-saccharide, while L. acidophilus consumed oligosaccharides up to trisaccharide. Both B. lactis and L. acidophilus utilised fructooligosaccharides and inulin, but showed different patterns of oligosaccharide consumption. Only L. rhamnosus grew on ?-glucan oligosaccharides and preferentially consumed the trisaccharide. The results indicate the synbiotic potential of the various probiotic/prebiotic combinations, particularly L. acidophilus/galactooligosaccharides, L. acidophilus/fructooligosaccharides or inulin and L. rhamnosus/?-glucan oligosaccharides. PMID:24239979

Sims, Ian M; Ryan, Jason L J; Kim, Sang H

2014-02-01

175

[The humoral immune response in mice induced by recombinant Lactococcus lactis expressing HIV-1 gag].  

PubMed

Objective To analyze the humoral immune response induced by recombinant Lactococcus lactis expressing HIV-1 gag in mice immunized orally, intranasally, subcutaneously or in the combined way of above three. Methods Fifty BALB/c mice were randomly divided into 5 groups, 10 mice per group. The mice were immunized consecutively three times at two week intervals with 10(9) CFU of recombinant Lactococcus lactis expressing gag through oral, intranasal, subcutaneous administration or the mix of them. The mice that were immunized orally with Lactococcus lactis containing PMG36e served as a control group. The sera of mice were collected before primary immunization and 2 weeks after each immunization to detect the gag specific IgG by ELISA. Results Compared with the control group, the higher titer of serum gag specific IgG was detected in the four groups immunized with recombinant Lactococcus lactis expressing gag, and it was the highest in the mixed immunization group (P<0.01). The titer of serum gag specific IgG from the oral and subcutaneous immunization groups was significantly higher than that from the intranasal immunization group (P<0.01) 6 weeks after primary immunization . The serum antibody positive rates of the oral immunization group after the first, the second, the third immunization were 40%, 40%, 90%, respectively; the positive rates of the intranasal immunization group were 10%, 20%, 20%; the positive rates of the subcutaneous immunization group were 10%, 60%, 90%; the positive rate of the combined immunization group reached 100% 2 weeks after primary immunization. Conclusion Recombinant Lactococcus lactis expressing gag can induce humoral immune response in mice by oral, intranasal, subcutaneous injection or the mix of them, and the mixed immunization can enhance the immune effects of Lactococcus lactis vector vaccine. PMID:25374076

Zhao, Xiaofei; Zhang, Cairong; Liu, Xiaojuan; Ma, Zhenghai

2014-11-01

176

Bifidobacterium animalis subsp. lactis ATCC 27673 Is a Genomically Unique Strain within Its Conserved Subspecies  

PubMed Central

Many strains of Bifidobacterium animalis subsp. lactis are considered health-promoting probiotic microorganisms and are commonly formulated into fermented dairy foods. Analyses of previously sequenced genomes of B. animalis subsp. lactis have revealed little genetic diversity, suggesting that it is a monomorphic subspecies. However, during a multilocus sequence typing survey of Bifidobacterium, it was revealed that B. animalis subsp. lactis ATCC 27673 gave a profile distinct from that of the other strains of the subspecies. As part of an ongoing study designed to understand the genetic diversity of this subspecies, the genome of this strain was sequenced and compared to other sequenced genomes of B. animalis subsp. lactis and B. animalis subsp. animalis. The complete genome of ATCC 27673 was 1,963,012 bp, contained 1,616 genes and 4 rRNA operons, and had a G+C content of 61.55%. Comparative analyses revealed that the genome of ATCC 27673 contained six distinct genomic islands encoding 83 open reading frames not found in other strains of the same subspecies. In four islands, either phage or mobile genetic elements were identified. In island 6, a novel clustered regularly interspaced short palindromic repeat (CRISPR) locus which contained 81 unique spacers was identified. This type I-E CRISPR-cas system differs from the type I-C systems previously identified in this subspecies, representing the first identification of a different system in B. animalis subsp. lactis. This study revealed that ATCC 27673 is a strain of B. animalis subsp. lactis with novel genetic content and suggests that the lack of genetic variability observed is likely due to the repeated sequencing of a limited number of widely distributed commercial strains. PMID:23995933

Loquasto, Joseph R.; Barrangou, Rodolphe; Dudley, Edward G.; Stahl, Buffy; Chen, Chun

2013-01-01

177

Comparison of sequences from the malB regions of Salmonella typhimurium and Enterobacter aerogenes with Escherichia coli K12: A potential new regulatory site in the interoperonic region  

Microsoft Academic Search

The malE and malK genes from Salmonella typhimurium, and the MalEFG operon and a portion of malK from Enterobacter aerogenes were cloned and sequenced. Plasmid-borne malE genes from both species and the malF and malG genes from E. aerogenes were expressed normally in Escherichia coli, and their products function in maltose transport. This shows that the malB products from the

Michael K. Dahl; Eric Francoz; William Saurin; Winfried Boos; Michael D. Manson; Maurice Hofnung

1989-01-01

178

Comparative In Vitro Activities of Ciprofloxacin, Clinafloxacin, Gatifloxacin, Levofloxacin, Moxifloxacin, and Trovafloxacin against Klebsiella pneumoniae, Klebsiella oxytoca, Enterobacter cloacae, and Enterobacter aerogenes Clinical Isolates with Alterations in GyrA and ParC Proteins  

Microsoft Academic Search

The in vitro activities of ciprofloxacin, clinafloxacin, gatifloxacin, levofloxacin, moxifloxacin, and trovafloxa- cin were tested against 72 ciprofloxacin-resistant and 28 ciprofloxacin-susceptible isolates of Klebsiella pneu- moniae, Klebsiella oxytoca, Enterobacter cloacae, and Enterobacter aerogenes. Irrespective of the alterations in GyrA and ParC proteins, clinafloxacin exhibited greater activity than all other fluoroquinolones tested against K. pneumoniae and E. aerogenes.

SYLVAIN BRISSE; DANA MILATOVIC; AD C. FLUIT; JAN VERHOEF; NELE MARTIN; SYBILLE SCHEURING; KARL KOHRER; FRANZ-JOSEF SCHMITZ

1999-01-01

179

Hydrogen production with high yield and high evolution rate by self-flocculated cells of Enterobacter aerogenes in a packed-bed reactor  

Microsoft Academic Search

Continuous hydrogen gas evolution by self-flocculated cells of Enterobacter aerogenes, a natural isolate HU-101 and its mutant AY-2, was performed in a packed-bed reactor under glucose-limiting conditions in\\u000a a minimal medium. The flocs that formed during the continuous culture were retained even when the dilution rate was increased\\u000a to 0.9?h?1. The H2 production rate increased linearly with increases in the

M. A. Rachman; Y. Nakashimada; T. Kakizono; N. Nishio

1998-01-01

180

Bioproduction of a novel sugar 1-deoxy- l-fructose by Enterobacter aerogenes IK7; isomerization of a 6-deoxyhexose to a 1-deoxyhexose  

Microsoft Academic Search

1-Deoxy-l-fructose, a very rare monosaccharide, was produced by hydrogenation of 6-deoxy-l-mannose (l-rhamnose)—the only cheaply available deoxy sugar—to 1-deoxy-l-mannitol (l-rhamnitol) followed by oxidation with Enterobacter aerogenes IK7. The entire procedure was conducted in water and shows the power of green environmentally friendly chemistry combined with biotechnology in the preparation of new monosaccharides with potential for novel bioactive properties or alternative foodstuffs;

Pushpakiran Gullapalli; Takayuki Shiji; Devendar Rao; Akihide Yoshihara; Kenji Morimoto; Goro Takata; George W. J. Fleet; Ken Izumori

2007-01-01

181

Treatment of a meningitis due to an Enterobacter aerogenes producing a derepressed cephalosporinase and a klebsiella pneumoniae producing an extended-spectrum ?-lactamase  

Microsoft Academic Search

Summary A case of nosocomial meningitis due to aKlebsiella pneumoniae producing a CAZ-5 extendedspectrum ß-lactamase and anEnterobacter aerogenes producing a derepressed cephalosporinase is reported. The intrathecal catheter incriminated was removed and a treatment with ceftazidime (4 g\\/24 h) and amikacin (1.5 g\\/24 h) was started. After 24 h ceftazidime was replaced by imipenem (2 then 4 g\\/24 h). This treatment

C. de Champs; D. Sirot; M. Chanal; J. Sirot; D. Guelon; D. Joyon

1991-01-01

182

Coexistence of SHV-4- and TEM24-Producing Enterobacter aerogenes Strains before a Large Outbreak of TEM24-Producing Strains in a French Hospital  

Microsoft Academic Search

In 1996, a monitoring program was initiated at the teaching hospital of Amiens, France, and carried out for 3 years. All extended-spectrum b-lactamase (ESBL)-producing Enterobacter aerogenes isolates recovered from clinical specimens were collected for investigation of their epidemiological relatedness by pulsed-field gel electrophoresis and enterobacterial repetitive intergenic consensus PCR (ERIC-PCR) and determination of the type of ESBL harbored by isoelectric

H. Mammeri; G. Laurans; M. Eveillard; S. Castelain; F. Eb

2001-01-01

183

Draft Genome Assemblies of Enterobacter aerogenes CDC 6003-71, Enterobacter cloacae CDC 442-68, and Pantoea agglomerans UA 0804-01.  

PubMed

The Enterobacteriaceae are environmental and enteric microbes. We sequenced the genomes of two Enterobacter reference strains, E. aerogenes CDC 6003-71 and E. cloacae CDC 442-68, as well as one near neighbor used as an exclusionary reference for diagnostics, Pantoea agglomerans CDC UA0804-01. The genome sizes range from 4.72 to 5.55 Mbp and have G+C contents from 54.6 to 55.1%. PMID:25342683

Minogue, T D; Daligault, H E; Davenport, K W; Bishop-Lilly, K A; Bruce, D C; Chain, P S; Coyne, S R; Chertkov, O; Freitas, T; Frey, K G; Jaissle, J; Koroleva, G I; Ladner, J T; Palacios, G F; Redden, C L; Xu, Y; Johnson, S L

2014-01-01

184

Carbapenem Resistance in a Clinical Isolate of Enterobacter aerogenes Is Associated with Decreased Expression of OmpF and OmpC Porin Analogs  

Microsoft Academic Search

We investigated the mechanism of imipenem resistance in Enterobacter aerogenes strain 810, a clinical isolate from the United States for which the imipenem MIC was 16 g\\/ml and the meropenem MIC was 8 g\\/ml. An imipenem-susceptible revertant, strain 810-REV, was obtained after multiple passages of the strain on non- selective media. For the revertant, the imipenem MIC was <1 g\\/ml

Hesna Yigit; Gregory J. Anderson; James W. Biddle; Christine D. Steward; J. Kamile Rasheed; Lourdes L. Valera; John E. McGowan; F. C. Tenover

2002-01-01

185

Reactor comparison and scale-up for the microaerobic production of 2,3-butanediol by Enterobacter aerogenes at constant oxygen transfer rate  

Microsoft Academic Search

Stirred tank (STR), bubble column (BCR) and airlift (ALR) bioreactors of 0.05 and 1.5 m3 total volume were compared for the production of 2,3-butanediol using Enterobacter aerogenes under microaerobic conditions. Batch fermentations were carried out at constant oxygen transfer rate (OTR=35 mmol\\/lh). At 0.05 m3 scale, the STR reactor achieved much higher biomass and product concentrations than the BCR and

T.-G. Byun; A.-P. Zeng; W.-D. Deckwer

1994-01-01

186

Short-chain organic acids produced on glucose, lactose, and citrate media by Enterococcus faecalis, Lactobacillus casei, and Enterobacter aerogenes strains  

Microsoft Academic Search

Three strains of Enterococcus faecalis, three of Lactobacillus casei and two of Enterobacter aerogenes, isolated from commercial Palmita-type cheese were cultured in peptone-yeast extract broth with glucose (PYG), lactose (PYL), or citrate (PYC) added as the main carbon sources. The short-chain volatile and non-volatile organic acids were extracted and their concentration determined by GC with a FID detector. The identity

D. Urdaneta; D. Raffe; A. Ferrer; B. Sulbarán de Ferrer; L. Cabrera; M. Pérez

1995-01-01

187

Draft Genome Assemblies of Enterobacter aerogenes CDC 6003-71, Enterobacter cloacae CDC 442-68, and Pantoea agglomerans UA 0804-01  

PubMed Central

The Enterobacteriaceae are environmental and enteric microbes. We sequenced the genomes of two Enterobacter reference strains, E. aerogenes CDC 6003-71 and E. cloacae CDC 442-68, as well as one near neighbor used as an exclusionary reference for diagnostics, Pantoea agglomerans CDC UA0804-01. The genome sizes range from 4.72 to 5.55 Mbp and have G+C contents from 54.6 to 55.1%. PMID:25342683

Minogue, T. D.; Daligault, H. E.; Davenport, K. W.; Bishop-Lilly, K. A.; Bruce, D. C.; Chain, P. S.; Coyne, S. R.; Chertkov, O.; Freitas, T.; Frey, K. G.; Jaissle, J.; Koroleva, G. I.; Ladner, J. T.; Palacios, G. F.; Redden, C. L.; Xu, Y.

2014-01-01

188

Integrated evaluation of aerogenic pollution by air-transported heavy metals (Pb, Cd, Ni, Zn, Mn and Cu) in the analysis of the main deposit media.  

PubMed

The composition of the ambient air is constantly changing; therefore, the monitoring of ambient air quality to detect the changes caused by aerogenic pollutants makes the essential part of general environmental monitoring. To achieve more effective improvement of the ambient air quality, the Directive 2008/50/EC on 'Ambient Air Quality and Cleaner Air for Europe' was adopted by the European Parliament and the European Council. It informed the public and enterprises about a negative effect of pollution on humans, animals and plants, as well as about the need for monitoring aerogenic pollutants not only at the continuous monitoring stations but also by using indicator methods, i.e. by analysing natural deposit media. The problem of determining the relationship between the accumulation level of pollutants by a deposit medium and the level of air pollution and its risks is constantly growing in importance. The paper presents a comprehensive analysis of the response of the main four deposit media, i.e. snow cover, soil, pine bark and epigeic mosses, to the long-term pollution by aerogenic pollutants which can be observed in the area of oil refinery influence. Based on the quantitative expressions of the amounts of the accumulated pollutants in the deposit media, the territory of the oil refinery investigated in this paper has been referred to the areas of mild or moderate pollution. PMID:23933956

Baltr?nait?, Edita; Baltr?nas, Pranas; Lietuvninkas, Arvydas; Serevi?ien?, Vaida; Zuokait?, Egl?

2014-01-01

189

Fine Tuning of the Lactate and Diacetyl Production through Promoter Engineering in Lactococcus lactis  

Microsoft Academic Search

Lactococcus lactis is a well-studied bacterium widely used in dairy fermentation and capable of producing metabolites with organoleptic and nutritional characteristics. For fine tuning of the distribution of glycolytic flux at the pyruvate branch from lactate to diacetyl and balancing the production of the two metabolites under aerobic conditions, a constitutive promoter library was constructed by randomizing the promoter sequence

Tingting Guo; Jian Kong; Li Zhang; Chenchen Zhang; Shumin Hu

2012-01-01

190

Complete Genome Sequence of Lactococcus lactis Strain AI06, an Endophyte of the Amazonian Açaí Palm  

PubMed Central

We report the genome, in a single chromosome, of Lactococcus lactis strain AI06, isolated from the mesocarp of the açaí fruit (Euterpe oleracea) in eastern Amazonia, Brazil. This strain is an endophyte of the açaí palm and also a component of the microbiota of the edible food product. PMID:25414513

de Oliveira, Viviane Matoso; de Almeida Pina, André Vicioli; Pérez-Chaparro, Paula Juliana; de Almeida, Lara Mendes; de Vasconcelos, Janaina Mota; de Oliveira, Layanna Freitas; da Silva, Daisy Elaine Andrade; Rogez, Hervé Louis Ghislain; Cretenet, Marina; Mamizuka, Elsa Masae; Nunes, Marcio Roberto Teixeira

2014-01-01

191

Lactococcus lactis as a Cell Factory for High-Level Diacetyl Production  

PubMed Central

We report the engineering of Lactococcus lactis for the efficient conversion of sugar into diacetyl by combining NADH-oxidase overproduction and ?-acetolactate decarboxylase inactivation. Eighty percent of the carbon flux was found to be rerouted via ?-acetolactate to the production of diacetyl by preloading the cells with NADH-oxidase before their use as a cell factory. PMID:10966436

Hugenholtz, Jeroen; Kleerebezem, Michiel; Starrenburg, Marjo; Delcour, Jean; de Vos, Willem; Hols, Pascal

2000-01-01

192

Diversity in robustness of Lactococcus lactis strains during heat stress, oxidative stress, and spray drying stress.  

PubMed

In this study we tested 39 Lactococcus lactis strains isolated from diverse habitats for their robustness under heat and oxidative stress, demonstrating high diversity in survival (up to 4 log units). Strains with an L. lactis subsp. lactis phenotype generally displayed more-robust phenotypes than strains with an L. lactis subsp. cremoris phenotype, whereas the habitat from which the strains had been isolated did not appear to influence stress survival. Comparison of the stress survival phenotypes with already available comparative genomic data sets revealed that the absence or presence of specific genes, including genes encoding a GntR family transcriptional regulator, a manganese ABC transporter permease, a cellobiose phosphotransferase system (PTS) component, the FtsY protein, and hypothetical proteins, was associated with heat or oxidative stress survival. Finally, 14 selected strains also displayed diversity in survival after spray drying, ranging from 20% survival for the most robust strains, which appears acceptable for industrial application, to 0.1% survival for the least-tolerant strains. The high and low levels of survival upon spray drying correlated clearly with the combined robustness under heat and oxidative stress. These results demonstrate the relevance of screening culture collections for robustness under heat and oxidative stress on top of the typical screening for acidifying and flavor-forming properties. PMID:24212574

Dijkstra, Annereinou R; Setyawati, Meily C; Bayjanov, Jumamurat R; Alkema, Wynand; van Hijum, Sacha A F T; Bron, Peter A; Hugenholtz, Jeroen

2014-01-01

193

Sugar metabolism, redox balance and oxidative stress response in the respiratory yeast Kluyveromyces lactis  

Microsoft Academic Search

A lot of studies have been carried out on Saccharomyces cerevisiae, an yeast with a predominant fermentative metabolism under aerobic conditions, which allows exploring the complex response induced by oxidative stress. S. cerevisiae is considered a eukaryote model for these studies. We propose Kluyveromyces lactis as a good alternative model to analyse variants in the oxidative stress response, since the

M. Isabel Gonzalez-Siso; Ana García-Leiro; Nuria Tarrío; M. Esperanza Cerdan

2009-01-01

194

Bifidobacterium animalis ssp. lactis 420 Protects against Indomethacin-Induced Gastric Permeability in Rats  

PubMed Central

Gastrointestinal (GI) adverse effects such as erosion and increased permeability are common during the use of nonsteroidal anti-inflammatory drugs (NSAIDs). Our objective was to assess whether Bifidobacterium animalis ssp. lactis 420 protects against NSAID-induced GI side effects in a rat model. A total of 120 male Wistar rats were allocated into groups designated as control, NSAID, and probiotic. The NSAID and probiotic groups were challenged with indomethacin (10?mg/kg?1; single dose). The probiotic group was also supplemented daily with 1010?CFU of B. lactis 420 for seven days prior to the indomethacin administration. The control group rats received no indomethacin or probiotic. The permeability of the rat intestine was analysed using carbohydrate probes and the visual damage of the rat stomach mucosa was graded according to severity. B. lactis 420 significantly reduced the indomethacin-induced increase in stomach permeability. However, the protective effect on the visual mucosal damage was not significant. The incidence of severe NSAID-induced lesions was, nevertheless, reduced from 50% to 33% with the probiotic treatment. To conclude, the B. lactis 420 supplementation protected the rats from an NSAID-induced increase in stomach permeability and may reduce the formation of more serious GI mucosal damage and/or enhance the recovery rate of the stomach mucosa. PMID:22848210

Lyra, Anna; Saarinen, Markku; Putaala, Heli; Olli, Kaisa; Lahtinen, Sampo J.; Ouwehand, Arthur C.; Madetoja, Mari; Tiihonen, Kirsti

2012-01-01

195

Diversity in Robustness of Lactococcus lactis Strains during Heat Stress, Oxidative Stress, and Spray Drying Stress  

PubMed Central

In this study we tested 39 Lactococcus lactis strains isolated from diverse habitats for their robustness under heat and oxidative stress, demonstrating high diversity in survival (up to 4 log units). Strains with an L. lactis subsp. lactis phenotype generally displayed more-robust phenotypes than strains with an L. lactis subsp. cremoris phenotype, whereas the habitat from which the strains had been isolated did not appear to influence stress survival. Comparison of the stress survival phenotypes with already available comparative genomic data sets revealed that the absence or presence of specific genes, including genes encoding a GntR family transcriptional regulator, a manganese ABC transporter permease, a cellobiose phosphotransferase system (PTS) component, the FtsY protein, and hypothetical proteins, was associated with heat or oxidative stress survival. Finally, 14 selected strains also displayed diversity in survival after spray drying, ranging from 20% survival for the most robust strains, which appears acceptable for industrial application, to 0.1% survival for the least-tolerant strains. The high and low levels of survival upon spray drying correlated clearly with the combined robustness under heat and oxidative stress. These results demonstrate the relevance of screening culture collections for robustness under heat and oxidative stress on top of the typical screening for acidifying and flavor-forming properties. PMID:24212574

Dijkstra, Annereinou R.; Setyawati, Meily C.; Bayjanov, Jumamurat R.; Alkema, Wynand; van Hijum, Sacha A. F. T.; Hugenholtz, Jeroen

2014-01-01

196

Standardized Assay Medium To Measure Lactococcus lactis Enzyme Activities while Mimicking Intracellular Conditions  

PubMed Central

Knowledge of how the activity of enzymes is affected under in vivo conditions is essential for analyzing their regulation and constructing models that yield an integrated understanding of cell behavior. Current kinetic parameters for Lactococcus lactis are scattered through different studies and performed under different assay conditions. Furthermore, assay conditions often diverge from conditions prevailing in the intracellular environment. To establish uniform assay conditions that resemble intracellular conditions, we analyzed the intracellular composition of anaerobic glucose-limited chemostat cultures of L. lactis subsp. cremoris MG 1363. Based on this, we designed a new assay medium for enzyme activity measurements of growing cells of L. lactis, mimicking as closely as practically possible its intracellular environment. Procedures were optimized to be carried out in 96-well plates, and the reproducibility and dynamic range were checked for all enzyme activity measurements. The effects of freezing and the carryover of ammonium sulfate from the addition of coupling enzymes were also established. Activities of all 10 glycolytic and 4 fermentative enzymes were measured. Remarkably, most in vivo-like activities were lower than previously published data. Yet, the ratios of Vmax over measured in vivo fluxes were above 1. With this work, we have developed and extensively validated standard protocols for enzyme activity measurements for L. lactis. PMID:22020503

Goel, Anisha; Santos, Filipe; de Vos, Willem M.; Teusink, Bas

2012-01-01

197

Characterization of the Role of para-Aminobenzoic Acid Biosynthesis in Folate Production by Lactococcus lactis?  

PubMed Central

The pab genes for para-aminobenzoic acid (pABA) biosynthesis in Lactococcus lactis were identified and characterized. In L. lactis NZ9000, only two of the three genes needed for pABA production were initially found. No gene coding for 4-amino-4-deoxychorismate lyase (pabC) was initially annotated, but detailed analysis revealed that pabC was fused with the 3? end of the gene coding for chorismate synthetase component II (pabB). Therefore, we hypothesize that all three enzyme activities needed for pABA production are present in L. lactis, allowing for the production of pABA. Indeed, the overexpression of the pABA gene cluster in L. lactis resulted in elevated pABA pools, demonstrating that the genes are involved in the biosynthesis of pABA. Moreover, a pABA knockout (KO) strain lacking pabA and pabBC was constructed and shown to be unable to produce folate when cultivated in the absence of pABA. This KO strain was unable to grow in chemically defined medium lacking glycine, serine, nucleobases/nucleosides, and pABA. The addition of the purine guanine, adenine, xanthine, or inosine restored growth but not the production of folate. This suggests that, in the presence of purines, folate is not essential for the growth of L. lactis. It also shows that folate is not strictly required for the pyrimidine biosynthesis pathway. L. lactis strain NZ7024, overexpressing both the folate and pABA gene clusters, was found to produce 2.7 mg of folate/liter per optical density unit at 600 nm when the strain was grown on chemically defined medium without pABA. This is in sharp contrast to L. lactis strains overexpressing only one of the two gene clusters. Therefore, we conclude that elevated folate levels can be obtained only by the overexpression of folate combined with the overexpression of the pABA biosynthesis gene cluster, suggesting the need for a balanced carbon flux through the folate and pABA biosynthesis pathway in the wild-type strain. PMID:17308179

Wegkamp, Arno; van Oorschot, Wietske; de Vos, Willem M.; Smid, Eddy J.

2007-01-01

198

An extended dynamic model of Lactococcus lactis metabolism for mannitol and 2,3-butanediol production.  

PubMed

Biomedical research and biotechnological production are greatly benefiting from the results provided by the development of dynamic models of microbial metabolism. Although several kinetic models of Lactococcus lactis (a Lactic Acid Bacterium (LAB) commonly used in the dairy industry) have been developed so far, most of them are simplified and focus only on specific metabolic pathways. Therefore, the application of mathematical models in the design of an engineering strategy for the production of industrially important products by L. lactis has been very limited. In this work, we extend the existing kinetic model of L. lactis central metabolism to include industrially relevant production pathways such as mannitol and 2,3-butanediol. In this way, we expect to study the dynamics of metabolite production and make predictive simulations in L. lactis. We used a system of ordinary differential equations (ODEs) with approximate Michaelis-Menten-like kinetics for each reaction, where the parameters were estimated from multivariate time-series metabolite concentrations obtained by our team through in vivo Nuclear Magnetic Resonance (NMR). The results show that the model captures observed transient dynamics when validated under a wide range of experimental conditions. Furthermore, we analyzed the model using global perturbations, which corroborate experimental evidence about metabolic responses upon enzymatic changes. These include that mannitol production is very sensitive to lactate dehydrogenase (LDH) in the wild type (W.T.) strain, and to mannitol phosphoenolpyruvate: a phosphotransferase system (PTS(Mtl)) in a LDH mutant strain. LDH reduction has also a positive control on 2,3-butanediol levels. Furthermore, it was found that overproduction of mannitol-1-phosphate dehydrogenase (MPD) in a LDH/PTS(Mtl) deficient strain can increase the mannitol levels. The results show that this model has prediction capability over new experimental conditions and offers promising possibilities to elucidate the effect of alterations in the main metabolism of L. lactis, with application in strain optimization. PMID:24413179

Costa, Rafael S; Hartmann, Andras; Gaspar, Paula; Neves, Ana R; Vinga, Susana

2014-03-01

199

Chromosomal Diversity in Lactococcus lactis and the Origin of Dairy Starter Cultures  

PubMed Central

A large collection of Lactococcus lactis strains, including wild-type isolates and dairy starter cultures, were screened on the basis of their phenotype and the macrorestriction patterns produced from pulsed-field gel electrophoresis (PFGE) analysis of SmaI digests of genomic DNA. Three groups of dairy starter cultures, used for different purposes in the dairy industry, and a fourth group made up of strains isolated from the environment were selected for analysis of their chromosomal diversity using the endonuclease I-CeuI. Chromosome architecture was largely conserved with each strain having six copies of the rRNA genes, and the chromosome size of individual strains ranged between 2,240 and 2,688 kb. The origin of L. lactis strains showed the greatest correlation with chromosome size, and dairy strains, particularly those with the cremoris phenotype, had smaller chromosomes than wild-type strains. Overall, this study, coupled with analysis of the sequenced L. lactis genomes, provides evidence that defined strain dairy starter cultures have arisen from plant L. lactis strains. Adaptation of these strains to the dairy environment has involved loss of functions resulting in smaller chromosomes and acquisition of genes (usually plasmid associated) that facilitate growth in milk. We conclude that dairy starter cultures generally and the industrially used cremoris and diacetylactis phenotype strains in particular comprise a specialized group of L. lactis strains that have been selected to become an essential component of industrial processes and have evolved accordingly, so that they are no longer fit to survive outside the dairy environment. PMID:20847124

Kelly, William J.; Ward, Lawrence J. H.; Leahy, Sinead C.

2010-01-01

200

L-2,4-diaminobutyric acid decarboxylase activity responsible for the formation of 1,3-diaminopropane in Enterobacter aerogenes.  

PubMed

High content of 1,3-diaminopropane (DAP), a normally minor derivative of polyamine metabolism, have been observed in cells of Enterobacter aerogenes. Supplementation of the growth medium with L-2,4-diaminobutyric acid (L-DABA) resulted in increased production of DAP, but not if supplemented with spermidine. On the basis of these observations, the biosynthetic route for DAP was evaluated. It has appeared that this bacterium possesses a novel enzyme activity catalysing the formation of DAP from L-DABA. Lack of the activity for oxidative cleavage of spermidine yielding DAP suggests that the enzyme termed DABA decarboxylase is responsible for the formation of DAP in this bacterium. The enzyme was partially purified 360-fold and some properties were examined. The pH optimum for the activity was 7.75-8.0, and the enzyme showed an absolute requirement for pyridoxal 5'-phosphate with the Km value of 41 microM. The Km value for L-DABA was 0.32 mM, and neither L-2,3-diaminopropionic acid, L-ornithine nor L-lysine showed detectable substrate activity towards the partially purified enzyme. Mg2+ and dithiothreitol greatly activated the enzyme. PMID:2397881

Nakao, H; Takeuchi, K; Shinoda, S; Yamamoto, S

1990-06-15

201

The existence of three types of acetohydroxy acid synthetase in an isoleucine-requiring mutant of Aerobacter aerogenes.  

PubMed

The synthesis of the three types of acetolactate synthase (EC 4.1.3.18) which are responsible for the biosynthesis os isoleucine and valine, was observed in Aerobacter aerogenes I-12, an isoleucine-requiring mutant, when grown on the four kinds of media. When the cells were grown on isoleucine-rich medium, acetolactate synthase sensitive to feedback inhibition and having an optimum pH at 8.0 was formed. By increasing the amount of potassium phosphate in the medium, the catabolite repression of the enzyme having an optimum pH at 6.0 and which is insensitive to feedback inhibition, was released. In contrast, acetolactate synthase having an optimum pH at 8.0 and insensitive to feedback inhibition was formd when isoleucine was limited, irrespective of phosphate concentrations. Two insensitive enzymes were not regulated by isoleucine, leucine and valine, although sensitive pH 8.0 enzyme was repressed by them. Thus, it may be assumed that the synthesis of insensitive pH 8.0 enzyme were repressed by limiting the amount of isoleucine is still open. PMID:5136

Asada, Y; Okuzawa, Y; Yamaguchi, K

1976-05-13

202

Characterization of the genes of the 2,3-butanediol operons from Klebsiella terrigena and Enterobacter aerogenes.  

PubMed Central

The genes involved in the 2,3-butanediol pathway coding for alpha-acetolactate decarboxylase, alpha-acetolactate synthase (alpha-ALS), and acetoin (diacetyl) reductase were isolated from Klebsiella terrigena and shown to be located in one operon. This operon was also shown to exist in Enterobacter aerogenes. The budA gene, coding for alpha-acetolactate decarboxylase, gives in both organisms a protein of 259 amino acids. The amino acid similarity between these proteins is 87%. The K. terrigena genes budB and budC, coding for alpha-ALS and acetoin reductase, respectively, were sequenced. The 559-amino-acid-long alpha-ALS enzyme shows similarities to the large subunits of the Escherichia coli anabolic alpha-ALS enzymes encoded by the genes ilvB, ilvG, and ilvI. The K. terrigena alpha-ALS is also shown to complement an anabolic alpha-ALS-deficient E. coli strain for valine synthesis. The 243-amino-acid-long acetoin reductase has the consensus amino acid sequence for the insect-type alcohol dehydrogenase/ribitol dehydrogenase family and has extensive similarities with the N-terminal and internal regions of three known dehydrogenases and one oxidoreductase. Images PMID:8444801

Blomqvist, K; Nikkola, M; Lehtovaara, P; Suihko, M L; Airaksinen, U; Straby, K B; Knowles, J K; Penttila, M E

1993-01-01

203

Changes in transcription profiles reflect strain contributions to defined cultures of Lactococcus lactis subsp. cremoris during milk fermentation  

Microsoft Academic Search

Cheddar cheese production uses mixed starters composed of Lactococcus lactis subsp. cremoris strains with complementary or competing enzymatic activity. However, strain interactions within the same subspecies are difficult\\u000a to investigate by conventional microbiological methods. This study uses fluorescent RNA arbitrarily primed PCR (FRAP-PCR)\\u000a to analyze the association of three L. lactis subsp. cremoris strains (LL074, LL225, and LL390 with proteinase

Fabien Dachet; Denis Roy; Gisèle LaPointe

204

Induction by glucose of an antimycin-insensitive, azide-sensitive respiration in the yeast Kluyveromyces lactis  

Microsoft Academic Search

Increasing the glucose concentration from 0.1 to 10% in exponentially growing cultures of Kluyveromyces lactis CBS 2359 does not repress the antimycin-sensitive respiration (QO2 of 80 µl O2·h-1·mg-1 dry weight) but raises the antimycin-insensitive respiration from 3 to 12 µl O2·h-1·mg-1 dry weight. Antimycin A inhibits the growth of K. lactis on a variety of substrates with the exception of

Iliana Ferrero; Anna-Maria Viola; A. Goffeau

1981-01-01

205

Genome-scale diversity and niche adaptation analysis of Lactococcus lactis by comparative genome hybridization using multi-strain arrays.  

PubMed

Lactococcus lactis produces lactic acid and is widely used in the manufacturing of various fermented dairy products. However, the species is also frequently isolated from non-dairy niches, such as fermented plant material. Recently, these non-dairy strains have gained increasing interest, as they have been described to possess flavour-forming activities that are rarely found in dairy isolates and have diverse metabolic properties. We performed an extensive whole-genome diversity analysis on 39 L. lactis strains, isolated from dairy and plant sources. Comparative genome hybridization analysis with multi-strain microarrays was used to assess presence or absence of genes and gene clusters in these strains, relative to all L. lactis sequences in public databases, whereby chromosomal and plasmid-encoded genes were computationally analysed separately. Nearly 3900 chromosomal orthologous groups (chrOGs) were defined on basis of four sequenced chromosomes of L. lactis strains (IL1403, KF147, SK11, MG1363). Of these, 1268 chrOGs are present in at least 35 strains and represent the presently known core genome of L. lactis, and 72 chrOGs appear to be unique for L. lactis. Nearly 600 and 400 chrOGs were found to be specific for either the subspecies lactis or subspecies cremoris respectively. Strain variability was found in presence or absence of gene clusters related to growth on plant substrates, such as genes involved in the consumption of arabinose, xylan, ?-galactosides and galacturonate. Further niche-specific differences were found in gene clusters for exopolysaccharides biosynthesis, stress response (iron transport, osmotolerance) and bacterial defence mechanisms (nisin biosynthesis). Strain variability of functions encoded on known plasmids included proteolysis, lactose fermentation, citrate uptake, metal ion resistance and exopolysaccharides biosynthesis. The present study supports the view of L. lactis as a species with a very flexible genome. PMID:21338475

Siezen, Roland J; Bayjanov, Jumamurat R; Felis, Giovanna E; van der Sijde, Marijke R; Starrenburg, Marjo; Molenaar, Douwe; Wels, Michiel; van Hijum, Sacha A F T; van Hylckama Vlieg, Johan E T

2011-05-01

206

Proton Motive Force-Driven and ATP-Dependent Drug Extrusion Systems in Multidrug-Resistant Lactococcus lactis  

Microsoft Academic Search

Three mutants of Lactococcus lactis subsp. lactis MG1363, termed EthR, DauR, and RhoR, were selected for resistance to high concentrations of ethidium bromide, daunomycin, and rhodamine 6G, respectively. These mutants were found to be cross resistant to a number of structurally and functionally unrelated drugs, among which were typical substrates of the mammalian multidrug transporter (P-glycoprotein) such as daunomycin, quinine,

Hendrik W. van Veen; Gerrit Poelarends; Douwe Molenaar; Henk Bolhuis; Bert Poolman; Arnold J. M. Driessen; Wil N. Konings

1994-01-01

207

Identification and functional characterization of the Lactococcus lactis rfb operon, required for dTDP-rhamnose biosynthesis  

Microsoft Academic Search

dTDP-rhamnose is an important precursor of cell wall polysaccharides and rhamnose-containing exopolysaccharides (EPS) in Lactococcus lactis. We cloned the rfbACBD operon from L. lactis MG1363, which comprises four genes involved in dTDP-rhamnose biosynthesis. When expressed in Escherichia coli, the lactococcal rfbACBD genes could sustain heterologous production of the Shigella flexneri O antigen, providing evidence of their functionality. Overproduction of the

Ingeborg C. Boels; Marke M. Beerthuyzen; Marit H. W. Kosters; Kaauwen van M. P. W; M. Kleerebezem; Vos de W. M

2004-01-01

208

Contribution of Citrate Metabolism to the Growth of Lactococcus lactis CRL264 at Low pH  

Microsoft Academic Search

Lactococcus lactis subsp. lactis biovar diacetylactis CRL264 is a natural strain isolated from cheese (F. Sesma, D. Gardiol, A. P. de Ruiz Holgado, and D. de Mendoza, Appl. Environ. Microbiol. 56:2099-2103, 1990). The effect of citrate on the growth parameters at a very acidic pH value was studied with this strain and with derivatives whose citrate uptake capacity was genetically

Claudia Sanchez; Ana Rute Neves; Joao Cavalheiro; Margarida Moreira dos Santos; N. Garcia-Quintans; Paloma Lopez; Helena Santos

2008-01-01

209

Genome-scale diversity and niche adaptation analysis of Lactococcus lactis by comparative genome hybridization using multi-strain arrays  

PubMed Central

Summary Lactococcus lactis produces lactic acid and is widely used in the manufacturing of various fermented dairy products. However, the species is also frequently isolated from non?dairy niches, such as fermented plant material. Recently, these non?dairy strains have gained increasing interest, as they have been described to possess flavour?forming activities that are rarely found in dairy isolates and have diverse metabolic properties. We performed an extensive whole?genome diversity analysis on 39 L. lactis strains, isolated from dairy and plant sources. Comparative genome hybridization analysis with multi?strain microarrays was used to assess presence or absence of genes and gene clusters in these strains, relative to all L. lactis sequences in public databases, whereby chromosomal and plasmid?encoded genes were computationally analysed separately. Nearly 3900 chromosomal orthologous groups (chrOGs) were defined on basis of four sequenced chromosomes of L. lactis strains (IL1403, KF147, SK11, MG1363). Of these, 1268 chrOGs are present in at least 35 strains and represent the presently known core genome of L. lactis, and 72 chrOGs appear to be unique for L. lactis. Nearly 600 and 400 chrOGs were found to be specific for either the subspecies lactis or subspecies cremoris respectively. Strain variability was found in presence or absence of gene clusters related to growth on plant substrates, such as genes involved in the consumption of arabinose, xylan, ??galactosides and galacturonate. Further niche?specific differences were found in gene clusters for exopolysaccharides biosynthesis, stress response (iron transport, osmotolerance) and bacterial defence mechanisms (nisin biosynthesis). Strain variability of functions encoded on known plasmids included proteolysis, lactose fermentation, citrate uptake, metal ion resistance and exopolysaccharides biosynthesis. The present study supports the view of L. lactis as a species with a very flexible genome. PMID:21338475

Siezen, Roland J.; Bayjanov, Jumamurat R.; Felis, Giovanna E.; van der Sijde, Marijke R.; Starrenburg, Marjo; Molenaar, Douwe; Wels, Michiel; van Hijum, Sacha A. F. T.; van Hylckama Vlieg, Johan E. T.

2011-01-01

210

Effects of Strains of Lactococcus lactis on the Production of Nitric Oxide and Cytokines in Murine Macrophages.  

PubMed

Nitric oxide (NO) is a multifunctional mediator that is involved in a variety of pathologic and physiologic processes. Few studies have addressed the effect of lactic acid bacteria (LAB), especially Lactococcus lactis strains used in dairy products, on inducible nitric oxide synthase (iNOS) induction as a component of the host's gastrointestinal immune response. We investigated the ability of L. lactis strains to induce NO synthesis in the murine macrophage-like cell line J774.1 and in peritoneal macrophages from mice. The degree of NO induction was specific to the L. lactis strain used. Compared with the no-treatment control, heat treatment of L. lactis cells decreased NO and TNF-? levels but further stimulated interleukin (IL)-12 production. Adding L. lactis cells to peritoneal macrophages dose-dependently increased the production of NO and IL-10 but decreased that of IL-12p70. Adding L. lactis cells to interferon-?-stimulated J774.1 cells enhanced cell death and the production of NO and IL-12p40, whereas addition of 1400W, a specific inhibitor of iNOS, decreased NO production and cell death. Conversely, adding 1400W to J774.1 cells further enhanced IL-12p40 production, suggesting that IL-12 production is perturbed by excess endogenous NO. IL-12 production is thought to be a marker of improved immunostimulation. Our results suggest that IL-12 production could be increased by limiting endogenous NO production. PMID:24818707

Suzuki, Chise; Aoki-Yoshida, Ayako; Kimoto-Nira, Hiromi; Kobayashi, Miho; Sasaki, Keisuke; Mizumachi, Koko

2014-10-01

211

Identification of genomic heterogeneity among Lactococcus lactis strains by plasmid profiling, PFGE and 16S rDNA sequence analysis.  

PubMed

Lactococcus lactis strains are used commonly as starters, which contribute to desirable flavour and texture properties known as strain-specific, in dairy industry. Genomic heterogeneity of 30 L. lactis strains originating from Turkey and characterized phenotypically were investigated in this study. Plasmid profiling, PFGE and 16S rDNA sequence analyses were performed to determine the genetic variability of strains. High degree of heterogeneity was detected among the L. lactis strains. Plasmid profiles of strains showed that compared to the plasmid free control strains, namely; L. lactis subsp. lactis IL1403 and L. lactis subsp. cremoris MG1614, all tested strains carried one to ten plasmids with molecular size ranging from 1.5 to 41.5kb. The fingerprints of strains obtained by PFGE from digestion with ApaI, SmaI and I-CeuI restriction endonucleases of chromosomal DNA's were compared with each other. All strains out of four were grouped into a large cluster A with at least 44% similarity level. The other four strains formed a minor duster B, distinctively different from major cluster A. PFGE results were confirmed by 16S rDNA sequence analysis and strains included in cluster B were identified as members of different species. These results suggested that morphologic and biochemical methods should be verified by reliable molecular approaches for the purpose of strain typing. Also, PFGE was found suitable to determine genomic differentiations among inter- and intra species. PMID:25115109

Gunay-Esiyok, Ozlem; Akcelik, Nefise; Akcelik, Mustafa

2014-01-01

212

Metabolic engineering of Kluyveromyces lactis for L-ascorbic acid (vitamin C) biosynthesis  

PubMed Central

Background L-ascorbic acid (L-AA) is naturally synthesized in plants from D-glucose by 10 steps pathway. The pathway branch to synthesize L-galactose, the key intermediate for L-ascorbic acid biosynthesis, has been recently elucidated. Budding yeast produces an 5-carbon ascorbic acid analogue Dehydro-D-arabinono 1,4-lactone (D-DAL), which is synthesized from D-arabinose. Yeast is able to synthesize L-ascorbic acid only if it is cultivated in the presence of one of its precursors: L-galactose, L-galactono 1,4-lactone, or L-gulono 1,4-lactone extracted from plants or animals. To avoid feeding the yeast culture with this “L” enantiomer, we engineered Kluyveromyces lactis with L-galactose biosynthesis pathway genes: GDP-mannose 3,5-epimerase (GME), GDP-L-galactose phosphorylase (VTC2) and L-galactose-1-phosphate phosphatase (VTC4) isolated from Arabidopsis thaliana. Results Plasmids were constructed and modified such that the cloned plant genes were targeted to the K. lactis LAC4 Locus by homologous recombination and that the expression was associated to the growth on D-galactose or lactose. Upon K. lactis transformation, GME was under the control of the native LAC4 promoter whereas VTC2 and VTC4 were expressed from the S. cerevisiae promoters GPD1 and ADH1 respectively. The expression in K. lactis, of the L-galactose biosynthesis genes was determined by Reverse Transcriptase-PCR and western blotting. The recombinant yeasts were capable to produce about 30 mg.L-1 of L-ascorbic acid in 48 hours of cultivation when cultured on rich medium with 2% (w/v) D-galactose. We also evaluated the L-AA production culturing recombinant recombinant strains in cheese whey, a waste product during cheese production, as an alternative source of lactose. Conclusions This work is the first attempt to engineer K. lactis cells for L-ascorbic acid biosynthesis by a fermentation process without any trace of “L” isomers precursors in the culture medium. We have engineered K. lactis strains capable of converting lactose and D-galactose into L-galactose, by the integration of the genes from the A. thaliana L-galactose pathway. L-galactose is a rare sugar, which is one of the main precursors for L-AA production. PMID:23799937

2013-01-01

213

Production of Recombinant Peanut Allergen Ara h 2 using Lactococcus lactis  

PubMed Central

Background Natural allergen sources can supply large quantities of authentic allergen mixtures for use as immunotherapeutics. However, such extracts are complex, difficult to define, vary from batch to batch, which may lead to unpredictable efficacy and/or unacceptable levels of side effects. The use of recombinant expression systems for allergen production can alleviate some of these issues. Several allergens have been tested in high-level expression systems and in most cases show immunereactivity comparable to their natural counterparts. The gram positive lactic acid bacterium Lactococcus lactis is an attractive microorganism for use in the production of protein therapeutics. L. lactis is considered food grade, free of endotoxins, and is able to secrete the heterologous product together with few other native proteins. Hypersensitivity to peanut represents a serious allergic problem. Some of the major allergens in peanut have been described. However, for therapeutic usage more information about the individual allergenic components is needed. In this paper we report recombinant production of the Ara h 2 peanut allergen using L. lactis. Results A synthetic ara h 2 gene was cloned into an L. lactis expression plasmid containing the P170 promoter and the SP310mut2 signal sequence. Flask cultures grown overnight showed secretion of the 17 kDa Ara h 2 protein. A batch fermentation resulted in 40 mg/L recombinant Ara h 2. Purification of Ara h 2 from the culture supernatant was done by hydrophobic exclusion and size separation. Mass spectrometry and N-terminal analysis showed a recombinant Ara h 2 of full length and correctly processed by the signal peptidase. The immunological activity of recombinant Ara h 2 was analysed by ELISA using antibodies specific for native Ara h 2. The recombinant Ara h 2 showed comparable immunereactivity to that of native Ara h 2. Conclusion Recombinant production of Ara h 2 using L. lactis can offer high yields of secreted, full length and immunologically active allergen. The L. lactis expression system can support recombinant allergen material for immunotherapy and component resolved allergen diagnostics. PMID:17711578

Glenting, Jacob; Poulsen, Lars K; Kato, Kentaro; Madsen, Søren M; Frøkiær, Hanne; Wendt, Camilla; Sørensen, Helle W

2007-01-01

214

Intranasal immunization with live recombinant Lactococcus lactis combined with heat-labile toxin B subunit protects chickens from highly pathogenic avian influenza H5N1 virus.  

PubMed

Development of safe and effective vaccines to prevent highly pathogenic avian influenza H5N1 virus infection is a challenging goal. Lactococcus lactis (L. lactis) is an ideal delivery vector for vaccine development, and it has been shown previously that oral immunization of encapsulated secretory L. lactis-hemagglutinin (HA) could provide complete protection against homologous H5N1 virus challenge in the mice model. While intranasal immunization is an appealing approach, it is now reported that secretory L. lactis-HA combined with mucosal adjuvant heat-labile toxin B subunit (LTB) could provide protective immunity in the chicken model. As compared to intranasal immunization with L. lactis-HA alone, L. lactis-HA combined with LTB (L. lactis-HA?+?LTB) could elicit robust neutralizing antibody responses and mucosal IgA responses, as well as strong cellular immune responses in the vaccinated chickens. Importantly, intranasal immunization with L. lactis-HA?+?LTB could provide 100% protection against H5N1 virus challenge. Taken together, these results suggest that intranasal immunization with L. lactis-HA?+?LTB can be considered as an effective approach for preventing and controlling infection of H5N1 virus in poultry during an avian influenza A/H5N1 pandemic. J. Med. Virol. 87: 39-44, 2015. © 2014 Wiley Periodicals, Inc. PMID:24861477

Lei, Han; Peng, Xiaojue; Shu, Handing; Zhao, Daxian

2015-01-01

215

Plasmids of Raw Milk Cheese Isolate Lactococcus lactis subsp. lactis Biovar diacetylactis DPC3901 Suggest a Plant-Based Origin for the Strain ? †  

PubMed Central

The four-plasmid complement of the raw milk cheese isolate Lactococcus lactis subsp. lactis biovar diacetylactis DPC3901 was sequenced, and some genetic features were functionally analyzed. The complete sequences of pVF18 (18,977 bp), pVF21 (21,739 bp), pVF22 (22,166 bp), and pVF50 (53,876 bp) were obtained. Each plasmid contained genes not previously described for Lactococcus, in addition to genes associated with plant-derived lactococcal strains. Most of the novel genes were found on pVF18 and encoded functions typical of bacteria associated with plants, such as activities of plant cell wall modification (orf11 and orf25). In addition, a predicted high-affinity regulated system for the uptake of cobalt was identified (orf19 to orf21 [orf19-21]), which has a single database homolog on a plant-derived Leuconostoc plasmid and whose functionality was demonstrated following curing of pVF18. pVF21 and pVF22 encode additional metal transporters, which, along with orf19-21 of pVF18, could enhance host ability to uptake growth-limiting amounts of biologically essential ions within the soil. In addition, vast regions from pVF50 and pVF21 share significant homology with the plant-derived lactococcal plasmid pGdh442, which is indicative of extensive horizontal gene transfer and recombination between these plasmids and suggests a common plant niche for their hosts. Phenotypes associated with these regions include glutamate dehydrogenase activity and Na+ and K+ transport. The presence of numerous plant-associated markers in L. lactis DPC3901 suggests a plant origin for the raw milk cheese isolate and provides for the first time the genetic basis to support the concept of the plant-milk transition for Lactococcus strains. PMID:21803914

Fallico, Vincenzo; McAuliffe, Olivia; Fitzgerald, Gerald F.; Ross, R. Paul

2011-01-01

216

Secretion of biologically active human interleukin 22 (IL-22) by Lactococcus lactis.  

PubMed

Interleukin-22 (IL-22) participates in the modulation of innate immunity and inflammation. This cytokine has important therapeutic potential, such as with ulcerative colitis, liver and lung injury, and infection, in different animal models. We generated a Lactococcus lactis strain that secretes human IL-22 under the regulation of the nisin-inducible promoter. Identification and secretion of this cytokine was demonstrated using western blots of culture supernatants from IL-22-expressing bacteria. The recombinant IL-22 protein produced by L. lactis was biologically active as determined by its ability to induce IL-10 secretion when co-cultured with a colon epithelial cell line in vitro. We consider this novel strain a promising live vaccine for various therapeutic applications. PMID:25214209

Loera-Arias, María J; Villatoro-Hernández, Julio; Parga-Castillo, Miguel A; Salcido-Montenegro, Alejandro; Barboza-Quintana, Oralia; Muñoz-Maldonado, Gerardo E; Montes-de-Oca-Luna, Roberto; Saucedo-Cárdenas, Odila

2014-12-01

217

Prevalence and diversity of qnr alleles in AmpC-producing Enterobacter cloacae, Enterobacter aerogenes, Citrobacter freundii and Serratia marcescens: a multicentre study from Korea  

Microsoft Academic Search

Methods: A total of 644 consecutive, non-duplicate isolates of Enterobacter cloacae (186), Enterobacter aerogenes (154), Citrobacter freundii (138) and Serratia marcescens (166) were examined. We performed antimicrobial susceptibility testing and PCR for qnr determinants (qnrA, qnrB and qnrS), extended-spectrum b-lactamase (ESBL) (blaTEM, blaSHV and blaCTX-M), orf513, orf1005 and blaDHA-1. To differentiate qnr subtypes, restriction enzyme analysis and sequencing was performed.

Yeon-Joon Park; Jin Kyung Yu; Seungok Lee; Eun-Jee Oh; Gun-Jo Woo

2007-01-01

218

Characterization of the Highly Autolytic Lactococcus lactis subsp. cremoris Strains CO and 2250  

PubMed Central

Two highly autolytic Lactococcus lactis subsp. cremoris strains (CO and 2250) were selected and analyzed for their autolytic properties. Both strains showed maximum lysis when grown in M17 broth containing a limiting concentration of glucose (0.4 to 0.5%) as the carbohydrate source. Lysis did not vary greatly with pH or temperature but was reduced when strains were grown on lactose or galactose. Growth in M17 containing excess glucose (1%) prevented autolysis, although rapid lysis of L. lactis subsp. cremoris CO did occur in the presence of 1% glucose if sodium fluoride (an inhibitor of glycolysis) was added to the medium. Maximum cell lysis in a buffer system was observed early in the stationary phase, and for CO, two pH optima were observed for log-phase and stationary-phase cells (6.5 and 8.5, respectively). Autolysins were extracted from the cell wall fraction of each strain by using either 4% sodium dodecyl sulfate (SDS), 6 M guanidine hydrochloride, or 4 M lithium chloride, and their activities were analyzed by renaturing SDS-polyacrylamide gel electrophoresis on gels containing Micrococcus luteus or L. lactis subsp. cremoris CO cells as the substrate. More than one lytic band was observed on each substrate, with the major band having an apparent molecular mass of 48 kDa for CO. Each lytic band was present throughout growth and lysis. These results suggest that at least two different autolytic enzymes are present in the autolytic L. lactis subsp. cremoris strains. The presence of the lactococcal cell wall hydrolase gene, acmA (G. Buist, J. Kok, K. J. Leenhouts, M. Dabrowska, G. Venema, and A. J. Haandrikman, J. Bacteriol. 177:1554-1563, 1995), in strains 2250 and CO was confirmed by Southern hybridization. Analysis of an acmA deletion mutant of 2250 confirmed that the gene was involved in cell separation and had a role in cell lysis. PMID:16535702

Riepe, H. R.; Pillidge, C. J.; Gopal, P. K.; Mckay, L. L.

1997-01-01

219

Glycolic acid production in the engineered yeasts Saccharomyces cerevisiae and Kluyveromyces lactis  

PubMed Central

Background Glycolic acid is a C2 hydroxy acid that is a widely used chemical compound. It can be polymerised to produce biodegradable polymers with excellent gas barrier properties. Currently, glycolic acid is produced in a chemical process using fossil resources and toxic chemicals. Biotechnological production of glycolic acid using renewable resources is a desirable alternative. Results The yeasts Saccharomyces cerevisiae and Kluyveromyces lactis are suitable organisms for glycolic acid production since they are acid tolerant and can grow in the presence of up to 50 g l-1 glycolic acid. We engineered S. cerevisiae and K. lactis for glycolic acid production using the reactions of the glyoxylate cycle to produce glyoxylic acid and then reducing it to glycolic acid. The expression of a high affinity glyoxylate reductase alone already led to glycolic acid production. The production was further improved by deleting genes encoding malate synthase and the cytosolic form of isocitrate dehydrogenase. The engineered S. cerevisiae strain produced up to about 1 g l-1 of glycolic acid in a medium containing d-xylose and ethanol. Similar modifications in K. lactis resulted in a much higher glycolic acid titer. In a bioreactor cultivation with d-xylose and ethanol up to 15 g l-1 of glycolic acid was obtained. Conclusions This is the first demonstration of engineering yeast to produce glycolic acid. Prior to this work glycolic acid production through the glyoxylate cycle has only been reported in bacteria. The benefit of a yeast host is the possibility for glycolic acid production also at low pH, which was demonstrated in flask cultivations. Production of glycolic acid was first shown in S. cerevisiae. To test whether a Crabtree negative yeast would be better suited for glycolic acid production we engineered K. lactis in the same way and demonstrated it to be a better host for glycolic acid production. PMID:24053654

2013-01-01

220

Characterization of the Highly Autolytic Lactococcus lactis subsp. cremoris Strains CO and 2250  

Microsoft Academic Search

Two highly autolytic Lactococcus lactis subsp. cremoris strains (CO and 2250) were selected and analyzed for their autolytic properties. Both strains showed maximum lysis when grown in M17 broth containing a limiting concentration of glucose (0.4 to 0.5%) as the carbohydrate source. Lysis did not vary greatly with pH or temperature but was reduced when strains were grown on lactose

HEIDI R. RIEPE; CHRISTOPHER J. PILLIDGE; PRAMOD K. GOPAL; LARRY L. MCKAY

221

Optimization of a cultural medium for bacteriocin production by Lactococcus lactis using response surface methodology  

Microsoft Academic Search

The medium composition for bacteriocin production by Lactococcus lactis ATCC 11454 was optimized using response surface methodology. The selected six factors based on CM medium were sucrose, soybean peptone, yeast extract, KH2PO4, NaCl, and MgSO4·7H2O. Fractional factorial designs (FFD) and the path of steepest ascent were effective in searching for the main factors and approaching the optimum region of the

Chan Li; Jinghua Bai; Zhaoling Cai; Fan Ouyang

2002-01-01

222

Genotype-phenotype matching analysis of 38 Lactococcus lactis strains using random forest methods  

PubMed Central

Background Lactococcus lactis is used in dairy food fermentation and for the efficient production of industrially relevant enzymes. The genome content and different phenotypes have been determined for multiple L. lactis strains in order to understand intra-species genotype and phenotype diversity and annotate gene functions. In this study, we identified relations between gene presence and a collection of 207 phenotypes across 38 L. lactis strains of dairy and plant origin. Gene occurrence and phenotype data were used in an iterative gene selection procedure, based on the Random Forest algorithm, to identify genotype-phenotype relations. Results A total of 1388 gene-phenotype relations were found, of which some confirmed known gene-phenotype relations, such as the importance of arabinose utilization genes only for strains of plant origin. We also identified a gene cluster related to growth on melibiose, a plant disaccharide; this cluster is present only in melibiose-positive strains and can be used as a genetic marker in trait improvement. Additionally, several novel gene-phenotype relations were uncovered, for instance, genes related to arsenite resistance or arginine metabolism. Conclusions Our results indicate that genotype-phenotype matching by integrating large data sets provides the possibility to identify gene-phenotype relations, possibly improve gene function annotation and identified relations can be used for screening bacterial culture collections for desired phenotypes. In addition to all gene-phenotype relations, we also provide coherent phenotype data for 38 Lactococcus strains assessed in 207 different phenotyping experiments, which to our knowledge is the largest to date for the Lactococcus lactis species. PMID:23530958

2013-01-01

223

Abrupt disruption of capping and a single source for recombinationally elongated telomeres in Kluyveromyces lactis  

Microsoft Academic Search

Eukaryotic cells, including some human cancers, that lack telomerase can sometimes maintain telomeres by using recombination. It was recently proposed that recombinational telomere elongation (RTE) in a telomerase-deletion mutant of the yeast Kluyveromyces lactis occurs through a roll-and-spread mechanism as described in our previous work. According to this model, a tiny circle of telomeric DNA is copied by a rolling-circle

Zeki Topcu; Kristy Nickles; Charity Davis; Michael J. McEachern

2005-01-01

224

Citrate utilization gene cluster of the Lactococcus lactis biovar diacetylactis : organization and regulation of expression  

Microsoft Academic Search

The transport of citrate in Lactococcus lactis biovar diacetylactis is mediated by the citrate permease P. This polypeptide is encoded by the citP gene carried by plasmid pCIT264. In this report, we characterize the citP transcript, identify a cluster of two genes cotranscribed with citP and describe their post-transcriptional regulation. The transcriptional promoter is located 1500 nucleotides upstream of the

Felix López Felipe; Christian Magni; Diego Mendoza; Paloma López

1995-01-01

225

Genetic Marking ofLactococcus lactisShows Its Survival in the Human Gastrointestinal Tract  

Microsoft Academic Search

A human feeding study was performed with Lactococcus lactis TC165.5, which is genetically marked by insertion of the sucrose-nisin conjugative transposon Tn5276 and chromosomal resistance to rifampin and streptomycin. The fate of strain TC165.5 and its nucleic acids was monitored by conventional plating methods and by molecular detection techniques based on specific PCR amplification of the nisin (nisA) gene from

NICOLETTE KLIJN; ANTON H. WEERKAMP; ANDWILLEM M. DEVOS

1995-01-01

226

Interrelationships of Microorganisms in Cream. II. Lactobacillus casei, Bacillus subtilis, and Streptococcus lactis1  

Microsoft Academic Search

Bacillus subtilis and Lactobacillus casei were inoculated individually and in combination with Streptococcus lactis into sterile cream which was incubated at different temperatures and analyzed for protein and fat de- composition, pH, growth and flavor changes. Editor. Measurements of proteolysis by the tryptophan determination of Duggan (8) and water-insoluble acids (W.I.A.) by the Hillig (11) method have been used by

L. G. Harmon; F. E. Nelson

1957-01-01

227

Characterization of the Lactococcus lactis lactose genes and regulation of their expression  

Microsoft Academic Search

An important trait of the lactic acid bacterium Lactococcus lactis , that is used in industrial dairy fermentations, is the conversion of lactose into lactic acid. The enzymatic steps involved in the breakdown of lactose, that is transported into the cell via a phosphoenolpyruvate-dependent lactose phosphotransferase system (PEP-PTS lac<\\/SUP>), have been well established (Fig. 1). However, except for the molecular

Rooijen van R. J

1993-01-01

228

Genetic and biochemical characterization of the oligopeptide transport system of Lactococcus lactis.  

PubMed Central

The nucleotide sequence of a chromosomal DNA fragment of Lactococcus lactis subsp. lactis SSL135, previously implicated in peptide utilization, has been determined. The genes oppDFBCA, encoding the oligopeptide transport system (Opp), and that encoding the endopeptidase PepO were located on this 8.9-kb DNA fragment. The oppDFBCA and pepO genes are probably organized in an operon. Analysis of the deduced amino acid sequences of the genes indicated that the oligopeptide transport system consists of two ATP-binding proteins OppD and OppF, two integral membrane proteins OppB and OppC, and a substrate-binding protein OppA. On the basis of the homology of OppF and OppD of L. lactis with other ABC (ATP-binding cassette) transporter proteins, the L. lactis Opp system can be classified as a member of this group. Two integration mutants, one defective in OppA and the other defective in PepO, were constructed. Growth of these mutants in a chemically defined medium with oligopeptides showed that the transport system, but not the endopeptidase, is essential for the utilization of peptides longer than three residues. Uptake of the pentapeptide Leu-enkephalin in glycolyzing lactococcal cells was followed by rapid hydrolysis of the peptide intracellularly. Importantly, extracellular hydrolysis of Leu-enkephalin is not observed. The OppA-deficient mutant was unable to transport Leu-enkephalin. Growth experiments with pasteurized milk revealed that transport of oligopeptides forms an essential part of the proteolytic system in lactococci. Images PMID:8244921

Tynkkynen, S; Buist, G; Kunji, E; Kok, J; Poolman, B; Venema, G; Haandrikman, A

1993-01-01

229

The Membrane-Bound H+ATPase Complex Is Essential for Growth of Lactococcus lactis  

Microsoft Academic Search

The eight genes which encode the (F1Fo) H+-ATPase in Lactococcus lactis subsp. cremoris MG1363 were cloned and sequenced. The genes were organized in an operon with the gene order atpEBFHAGDC; i.e., the order of atpE and atpB is reversed with respect to the more typical bacterial organization. The deduced amino acid sequences of the corresponding H+-ATPase subunits showed significant homology

Brian J. Koebmann; Dan Nilsson; Oscar P. Kuipers; Peter R. Jensen

2000-01-01

230

HrtBA and menaquinones control haem homeostasis in Lactococcus lactis.  

PubMed

Lactococcus lactis is a fermenting Gram-positive bacterium widely used for production of dairy products. Lacking haem biosynthesis genes, L. lactis can still shift to an energetically favourable respiratory metabolism by activating a terminal cytochrome bd oxidase when haem is added to an aerated culture. Haem intracellular homeostasis is mediated by the hrtRBA operon encoding the conserved membrane HrtBA haem efflux permease and the unique intracellular haem sensor and regulator, HrtR. Here we report that membrane-associated menaquinones (MK) favour the accumulation of reduced haem in membranes. An oxidative environment, provided by oxygen, prevents and reverses haemin reduction by MK and thus limits haem accumulation in membranes. HrtBA counteracts MK-dependent membrane retention of excess haem in membrane, suggesting direct efflux from this compartment. Moreover, both HrtBA and MK-mediated reduction have a strong impact on haem intracellular pools, as determined via HrtR haem sensor induction, suggesting that intracellular haem acquisition is controlled at the membrane level without the need for dedicated import systems. Our conclusions lead to a new hypothesis of haem acquisition and regulation in which HrtBA and the bacterial membrane have central roles in L. lactis. PMID:25040434

Joubert, Laetitia; Derré-Bobillot, Aurélie; Gaudu, Philippe; Gruss, Alexandra; Lechardeur, Delphine

2014-08-01

231

Microencapsulation of Bifidobacterium animalis subsp. lactis and Lactobacillus acidophilus in cocoa butter using spray chilling technology  

PubMed Central

In the present study, the cells of Bifidobacterium animalis subsp. lactis (BI-01) and Lactobacillus acidophilus (LAC-04) were encapsulated in cocoa butter using spray-chilling technology. Survival assays were conducted to evaluate the resistance of the probiotics to the spray-chilling process, their resistance to the simulated gastric and intestinal fluids (SGF and SIF), and their stability during 90 days of storage. The viability of the cells was not affected by microencapsulation. The free and encapsulated cells of B. animalis subsp. lactis were resistant to both SGF and SIF. The micro-encapsulated cells of L. acidophilus were more resistant to SGF and SIF than the free cells; the viability of the encapsulated cells was enhanced by 67%, while the free cells reached the detection limit of the method (103 CFU/g). The encapsulated probiotics were unstable when they were stored at 20 °C. The population of encapsulated L. acidophilus decreased drastically when they were stored at 7 °C; only 20% of cells were viable after 90 days of storage. The percentage of viable cells of the encapsulated B. animalis subsp.lactis, however, was 72% after the same period of storage. Promising results were obtained when the microparticles were stored at ?18 °C; the freeze granted 90 days of shelf life to the encapsulated cells. These results suggest that the spray-chilling process using cocoa butter as carrier protects L. acidophilus from gastrointestinal fluids. However, the viability of the cells during storage must be improved. PMID:24516445

Pedroso, D.L.; Dogenski, M.; Thomazini, M.; Heinemann, R.J.B.; Favaro-Trindade, C.S.

2013-01-01

232

Regulation of Acetate Kinase Isozymes and Its Importance for Mixed-Acid Fermentation in Lactococcus lactis  

PubMed Central

Acetate kinase (ACK) converts acetyl phosphate to acetate along with the generation of ATP in the pathway for mixed-acid fermentation in Lactococcus lactis. The reverse reaction yields acetyl phosphate for assimilation purposes. Remarkably, L. lactis has two ACK isozymes, and the corresponding genes are present in an operon. We purified both enzymes (AckA1 and AckA2) from L. lactis MG1363 and determined their oligomeric state, specific activities, and allosteric regulation. Both proteins form homodimeric complexes, as shown by size exclusion chromatography and static light-scattering measurements. The turnover number of AckA1 is about an order of magnitude higher than that of AckA2 for the reaction in either direction. The Km values for acetyl phosphate, ATP, and ADP are similar for both enzymes. However, AckA2 has a higher affinity for acetate than does AckA1, suggesting an important role under acetate-limiting conditions despite the lower activity. Fructose-1,6-bisphosphate, glyceraldehyde-3-phosphate, and phospho-enol-pyruvate inhibit the activities of AckA1 and AckA2 to different extents. The allosteric regulation of AckA1 and AckA2 and the pool sizes of the glycolytic intermediates are consistent with a switch from homolactic to mixed-acid fermentation upon slowing of the growth rate. PMID:24464460

Puri, Pranav; Goel, Anisha; Bochynska, Agnieszka

2014-01-01

233

Biotechnological and safety characterization of Enterococcus lactis, a recently described species of dairy origin.  

PubMed

The biotechnological and safety properties of a recently described enterococcal species, Enterococcus lactis, were investigated. With regard to the technological properties, in milk all the strains tested had weak acidifying and proteolytic activities, generally medium reduction activity over 24 h (-102 mV < Eh < -2 mV) and low lipolytic activity on tributyrin agar. The isolates were tested for resistance against 14 antibiotics and none of the studied strains were classified as resistant to clinically important antibiotics such as ampicillin, erythromycin, penicillin G, tetracycline and vancomycin. Furthermore, PCR-based detection did not identify any of the common genetic determinants for vancomycin, tetracycline and erythromycin resistance. The E. lactis strains showed good survival in simulated in vitro digestion and were able to inhibit the growth of Enterococcus durans, Enterococcus faecalis, Enterococcus faecium, Clostridium sporogenes, Clostridium tyrobutyricum and Pseudomonas syringae. Screening for enterocin structural genes showed that all isolates harboured the entP gene. The presence of nine virulence factor genes (cylA, asa1, gelE, hyl, esp, ace, efaA, hdc and tdc) was investigated by PCR and no virulence determinants were detected. This study highlights that the recently described E. lactis may be a potential source of novel strains with interesting features that could be used for fermented dairy foods. PMID:22961639

Morandi, Stefano; Silvetti, Tiziana; Brasca, Milena

2013-01-01

234

Probiotic assessment of Enterococcus durans 6HL and Lactococcus lactis 2HL isolated from vaginal microflora.  

PubMed

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. PMID:24913559

Nami, Yousef; Abdullah, Norhafizah; Haghshenas, Babak; Radiah, Dayang; Rosli, Rozita; Khosroushahi, Ahmad Yari

2014-08-01

235

Non-enzymic copper reduction by menaquinone enhances copper toxicity in Lactococcus lactis IL1403.  

PubMed

Lactococcus lactis possesses a pronounced extracellular Cu(2+)-reduction activity which leads to the accumulation of Cu(+) in the medium. The kinetics of this reaction were not saturable by increasing copper concentrations, suggesting a non-enzymic reaction. A copper-reductase-deficient mutant, isolated by random transposon mutagenesis, had an insertion in the menE gene, which encodes O-succinylbenzoic acid CoA ligase. This is a key enzyme in menaquinone biosynthesis. The ?menE mutant was deficient in short-chain menaquinones, and exogenously added menaquinone complemented the copper-reductase-deficient phenotype. Haem-induced respiration of wild-type L. lactis efficiently suppressed copper reduction, presumably by competition by the bd-type quinol oxidase for menaquinone. As expected, the ?menE mutant was respiration-deficient, but could be made respiration-proficient by supplementation with menaquinone. Growth of wild-type cells was more copper-sensitive than that of the ?menE mutant, due to the production of Cu(+) ions by the wild-type. This growth inhibition of the wild-type was strongly attenuated if Cu(+) was scavenged with the Cu(I) chelator bicinchoninic acid. These findings support a model whereby copper is non-enzymically reduced at the membrane by menaquinones. Respiration effectively competes for reduced quinones, which suppresses copper reduction. These findings highlight novel links between copper reduction, respiration and Cu(+) toxicity in L. lactis. PMID:23579688

Abicht, Helge K; Gonskikh, Yulia; Gerber, Simon D; Solioz, Marc

2013-06-01

236

Measuring kinetic dissociation/association constants between Lactococcus lactis bacteria and mucins using living cell probes.  

PubMed

In this work we focused on quantifying adhesion between Lactococcus lactis, the model for lactic acid bacteria (LAB) and mucins. Interactions between two strains of L. lactis (IBB477 and MG1820 as control) and pig gastric mucin-based coating were measured and compared with the use of atomic force microscopy. Analysis of retraction force-distance curves shed light on the differential contributions of nonspecific and specific forces. An increased proportion of specific adhesive events was obtained for IBB477 (20% vs. 5% for the control). Blocking assays with free pig gastric mucin and its O-glycan moiety showed that oligosaccharides play a major (but not exclusive) role in L. lactis-mucins interactions. Specific interactions were analyzed in terms of kinetic constants. An increase in the loading rate of atomic force microscope tip led to a higher force between interacting biological entities, which was directly linked to the kinetic dissociation constant (K(off)). Enhancing the contact time between the tip and the sample allowed an increase in the interaction probability, which can be related to the kinetic association constant (K(on)). Variations in the loading rate and contact time enabled us to determine K(on) (3.3 × 10(2) M(-1)·s(-1)) and K(off) (0.46 s(-1)), and the latter was consistent with values given in the literature for sugar-protein interactions. PMID:22261074

Le, Doan Thanh Lam; Guérardel, Yann; Loubière, Pascal; Mercier-Bonin, Muriel; Dague, Etienne

2011-12-01

237

Production of galactooligosaccharides using a hyperthermophilic ?-galactosidase in permeabilized whole cells of Lactococcus lactis.  

PubMed

Galactooligosaccharides (GOS) are novel prebiotic food ingredients that can be produced from lactose using ?-galactosidase, but the process is more efficient at higher temperatures. To efficiently express the lacS gene from the hyperthermophile Sulfolobus solfataricus, in Lactococcus lactis a synthetic gene (lacSt) with optimized codon usage for Lc. lactis was designed and synthesized. This hyperthermostable ?-galactosidase enzyme was successfully overexpressed in Lc. lactis LM0230 using a nisin-controlled gene expression system. Enzyme-containing cells were then killed and permeabilized using 50% ethanol and were used to determine both hydrolysis and transgalactosylation activity. The optimum conditions for GOS synthesis was found to be at pH 6.0 and 85 °C. A maximum production of 197 g/L of GOS tri- and tetrasaccharides was obtained from 40% initial lactose, after 55 h of incubation. The total GOS yield increased with the initial lactose concentration, whereas the highest lactose conversion rate (72%) was achieved from a low lactose solution (5%). Given that a significant proportion of the remaining lactose would be expected to be converted into disaccharide GOS, this should enable the future development of a cost-effective approach for the conversion of whey-based substrates into GOS-enriched food ingredients using this cell-based technology. PMID:24359820

Yu, L; O'Sullivan, D J

2014-02-01

238

Oxidative Stress at High Temperatures in Lactococcus lactis Due to an Insufficient Supply of Riboflavin  

PubMed Central

Lactococcus lactis MG1363 was found to be unable to grow at temperatures above 37°C in a defined medium without riboflavin, and the cause was identified to be dissolved oxygen introduced during preparation of the medium. At 30°C, growth was unaffected by dissolved oxygen and oxygen was consumed quickly. Raising the temperature to 37°C resulted in severe growth inhibition and only slow removal of dissolved oxygen. Under these conditions, an abnormally low intracellular ratio of [ATP] to [ADP] (1.4) was found (normally around 5), which indicates that the cells are energy limited. By adding riboflavin to the medium, it was possible to improve growth and oxygen consumption at 37°C, and this also normalized the [ATP]-to-[ADP] ratio. A codon-optimized redox-sensitive green fluorescent protein (GFP) was introduced into L. lactis and revealed a more oxidized cytoplasm at 37°C than at 30°C. These results indicate that L. lactis suffers from heat-induced oxidative stress at increased temperatures. A decrease in intracellular flavin adenine dinucleotide (FAD), which is derived from riboflavin, was observed with increasing growth temperature, but the presence of riboflavin made the decrease smaller. The drop was accompanied by a decrease in NADH oxidase and pyruvate dehydrogenase activities, both of which depend on FAD as a cofactor. By overexpressing the riboflavin transporter, it was possible to improve FAD biosynthesis, which resulted in increased NADH oxidase and pyruvate dehydrogenase activities and improved fitness at high temperatures in the presence of oxygen. PMID:23913422

Chen, Jun; Shen, Jing

2013-01-01

239

Metabolic and Transcriptomic Adaptation of Lactococcus lactis subsp. lactis Biovar diacetylactis in Response to Autoacidification and Temperature Downshift in Skim Milk  

PubMed Central

For the first time, a combined genome-wide transcriptome and metabolic analysis was performed with a dairy Lactococcus lactis subsp. lactis biovar diacetylactis strain under dynamic conditions similar to the conditions encountered during the cheese-making process. A culture was grown in skim milk in an anaerobic environment without pH regulation and with a controlled temperature downshift. Fermentation kinetics, as well as central metabolism enzyme activities, were determined throughout the culture. Based on the enzymatic analysis, a type of glycolytic control was postulated, which was shared by most of the enzymes during the growth phase; in particular, the phosphofructokinase and some enzymes of the phosphoglycerate pathway during the postacidification phase were implicated. These conclusions were reinforced by whole-genome transcriptomic data. First, limited enzyme activities relative to the carbon flux were measured for most of the glycolytic enzymes; second, transcripts and enzyme activities exhibited similar changes during the culture; and third, genes involved in alternative metabolic pathways derived from some glycolytic metabolites were induced just upstream of the postulated glycolytic bottlenecks, as a consequence of accumulation of these metabolites. Other transcriptional responses to autoacidification and a decrease in temperature were induced at the end of the growth phase and were partially maintained during the stationary phase. If specific responses to acid and cold stresses were identified, this exhaustive analysis also enabled induction of unexpected pathways to be shown. PMID:16332781

Raynaud, Sandy; Perrin, Rémi; Cocaign-Bousquet, Muriel; Loubiere, Pascal

2005-01-01

240

Recombinant porcine epidermal growth factor-secreting Lactococcus lactis promotes the growth performance of early-weaned piglets  

PubMed Central

Background Epidermal growth factor (EGF) is an important growth factor in regulation of cell proliferation, differentiation, survival and apoptosis. Studies showed that food-grade Lactococcus lactis (L. lactis) and NICE expression system have superior performance in exogenous protein expression. This study aimed to construct and express porcine EGF (pEGF), and use L. lactis as vehicle for producing and delivering pEGF. Furthermore, investigating biological activity of pEGF and exploring applications feasibility of combination effects of L. lactis and pEGF on early weaned piglets’ production. Results A recombinant Lactococcus lactis which produced and secreted pEGF at 1000 ng/ml in culture supernatant was generated. Secreted pEGF was a fully biologically active protein, as demonstrated by its capacity to stimulate L929 mouse fibroblast cell line proliferation in vitro. For in vivo study, forty piglets were randomly allocated to control, antibiotic control, empty vector-expressing L. lactis (LL-EV) and pEGF-secreting L. lactis (LL-pEGF). After 14 d of rearing, final body weight and average daily gain in LL-pEGF were greater (P < 0.05, 8.95 vs. 8.37 kg, 206.1 vs. 157.7 g/day, respectively) than those in control, but no significant differences between LL-pEGF, LL-EV and antibiotic control. Overall period average daily feed intake was higher in LL-pEGF, LL-EV and antibiotic control than in control (P < 0.05, 252.9, 255.6, 250.0, 207.3 g/day, respectively). No significant difference was observed on ADFI/ADG. LL-pEGF increased villous height in the duodenum, jejunum and ileum than in control and LL-EV (P < 0.05). Sucrase in the 3 intestinal segments, aminopeptidase A in the duodenum and Jejunum, aminopeptidase N and dipeptidase IV in the duodenum in LL-pEGF were higher than those in control (P < 0.05). Furthermore, Escherichia coli and Enterococcus counts decreased in the ileum and Lactobacillus increased in the ileum and cecum digesta in LL-pEGF compare with the control (P < 0.05). Lactobacillus increased in the cecum in LL-EV compared with control and antibiotic control (P < 0.05). Conclusion We have generated a recombinant Lactococcus lactis which produced and secreted fully biologically active porcine EGF. Oral administration of pEGF-secreting L. lactis had beneficial effects on intestinal health and performance of early-weaned piglets. PMID:25142032

2014-01-01

241

A comparative study on phosphotransferase activity of acid phosphatases from Raoultella planticola and Enterobacter aerogenes on nucleosides, sugars, and related compounds.  

PubMed

Natural and modified nucleoside-5'-monophosphates and their precursors are valuable compounds widely used in biochemical studies. Bacterial nonspecific acid phosphatases (NSAPs) are a group of enzymes involved in the hydrolysis of phosphoester bonds, and some of them exhibit phosphotransferase activity. NSAP containing Enterobacter aerogenes and Raoultella planticola whole cells were evaluated in the phosphorylation of a wide range of nucleosides and nucleoside precursors using pyrophosphate as phosphate donor. To increase the productivity of the process, we developed two genetically modified strains of Escherichia coli which overexpressed NSAPs of E. aerogenes and R. planticola. These new recombinant microorganisms (E. coli BL21 pET22b-phoEa and E. coli BL21 pET22b-phoRp) showed higher activity than the corresponding wild-type strains. Reductions in the reaction times from 21 h to 60 min, from 4 h to 15 min, and from 24 h to 40 min in cases of dihydroxyacetone, inosine, and fludarabine, respectively, were obtained. PMID:23995227

Médici, Rosario; Garaycoechea, Juan I; Valino, Ana L; Pereira, Claudio A; Lewkowicz, Elizabeth S; Iribarren, Adolfo M

2014-04-01

242

Functional Display of a Heterologous Protein on the Surface of Lactococcus lactis by Means of the Cell Wall Anchor of Staphylococcus aureus Protein A  

Microsoft Academic Search

In this study, we showed that the cell wall anchor of protein A from Staphylococcus aureus is functional in the food-grade organism Lactococcus lactis. A fusion protein composed of the lactococcal Usp45 secretion signal peptide, streptavidin monomer, and the S. aureus protein A anchor became covalently attached to the pepti- doglycan when expressed in L. lactis. The streptavidin moiety of

LOTHAR STEIDLER; JASMINE VIAENE; WALTER FIERS; ERIK REMAUT

1998-01-01

243

Study of the influence of yeast inoculum concentration (Yarrowia lipolytica and Kluyveromyces lactis) on blue cheese aroma development using microbiological models.  

PubMed

Yarrowia lipolytica and Kluyveromyces lactis occur as part of Stilton cheese microflora yet are not controlled during production. This study investigated the influence of their inoculum concentration on aroma production. Models of Y. lipolytica and K. lactis, with Penicillium roqueforti, were analysed using instrumental and sensory analysis. Different concentrations of Y. lipolytica produced important changes in the aroma profiles of microbiological models, analysed by solid-phase microextraction (SPME GC-MS). Sensory analysis with discrimination tests showed differences were detectable via human perception but did not concern the similarity to blue cheese odour. Increasing the inoculum concentration of K. lactis resulted in decreased variation between replicates. Partial least squares (PLS) regression on Flash profile data showed models inoculated with low concentrations of K. lactis exhibited blue cheese-related attributes, associated with increased ketone production. Results suggest that controlling the amount of Y. lipolytica and K. lactis during production offers potential to manipulate blue cheese aroma development. PMID:24128502

Price, Elliott J; Linforth, Robert S T; Dodd, Christine E R; Phillips, Carol A; Hewson, Louise; Hort, Joanne; Gkatzionis, Konstantinos

2014-02-15

244

Anti-inflammatory effects of Lactococcus lactis NCDO 2118 during the remission period of chemically induced colitis  

PubMed Central

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-1?-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-? 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

2014-01-01

245

Genetically Engineered Lactococcus lactis Protect against House Dust Mite Allergy in a BALB/c Mouse Model  

PubMed Central

Background Mucosal vaccine based on lactic acid bacteria is an attractive concept for the prevention and treatment of allergic diseases, but their mechanisms of action in vivo are poorly understood. Therefore, we sought to investigate how recombinant major dust mite allergen Der p2-expressing Lactococcus lactis as a mucosal vaccine induced the immune tolerance against house dust mite allergy in a mouse model. Methods Three strains of recombinant L. lactis producing Der p2 in different cell components (extracellular, intracellular and cell wall) were firstly constructed. Their prophylactic potential was evaluated in a Der p2-sensitised mouse model, and immunomodulation properties at the cellular level were determined by measuring cytokine production in vitro. Results Der p2 expressed in the different recombinant L. lactis strains was recognized by a polyclonal anti-Der p2 antibody. Oral treatment with the recombinant L. lactis prior sensitization significantly prevented the development of airway inflammation in the Der p2-sensitized mice, as determined by the attenuation of inflammatory cells infiltration in the lung tissues and decrease of Th2 cytokines IL-4 and IL-5 levels in bronchoalveolar lavage. In addition, the serum allergen-specific IgE levels were significantly reduced, and the levels of IL-4 in the spleen and mesenteric lymph nodes cell cultures were also markedly decreased upon allergen stimulation in the mice fed with the recombinant L. lactis strains. These protective effects correlated with a significant up-regulation of regulatory T cells in the mesenteric lymph nodes. Conclusion Oral pretreatment with live recombinant L. lactis prevented the development of allergen-induced airway inflammation primarily by the induction of specific mucosal immune tolerance. PMID:25290938

Ai, Chunqing; Zhang, Qiuxiang; Ren, Chengcheng; Wang, Gang; Liu, Xiaoming; Tian, Fengwei; Zhao, Jianxin; Zhang, Hao; Chen, Yong Q.; Chen, Wei

2014-01-01

246

A Zn-Dependent Metallopeptidase Is Responsible for Sensitivity to LsbB, a Class II Leaderless Bacteriocin of Lactococcus lactis subsp. lactis BGMN1-5  

PubMed Central

Lactococcus lactis subsp. lactis BGMN1-5 produces a leaderless class II bacteriocin called LsbB. To identify the receptor for LsbB, a cosmid library of the LsbB-sensitive strain BGMN1-596 was constructed. About 150 cosmid clones were individually isolated and transferred to LsbB-resistant mutants of BGMN1-596. Cosmid pAZILcos/MN2, carrying a 40-kb insert, was found to restore LsbB sensitivity in LsbB-resistant mutants. Further subcloning revealed that a 1.9-kb fragment, containing only one open reading frame, was sufficient to restore sensitivity. The fragment contains the gene yvjB coding for a Zn-dependent membrane-bound metallopeptidase, suggesting that this gene may serve as the receptor for LsbB. Further support for this notion derives from several independent experiments: (i) whole-genome sequencing confirmed that all LsbB-resistant mutants contain mutations in yvjB; (ii) disruption of yvjB by direct gene knockout rendered sensitive strains BGMN1-596 and IL1403 resistant to LsbB; and (iii) most compellingly, heterologous expression of yvjB in naturally resistant strains of other species, such as Lactobacillus paracasei and Enterococcus faecalis, also rendered them sensitive to the bacteriocin. To our knowledge, this is the first time a membrane-bound peptidase gene has been shown to be involved in bacteriocin sensitivity in target cells. We also demonstrated a novel successful approach for identifying bacteriocin receptors. PMID:24123824

Uzelac, Gordana; Lozo, Jelena; Aleksandrzak-Piekarczyk, Tamara; Gabrielsen, Christina; Kristensen, Tom; Nes, Ingolf F.; Diep, Dzung B.; Topisirovic, Ljubisa

2013-01-01

247

Enzymatic synthesis and characterization of hydroquinone galactoside using Kluyveromyces lactis lactase.  

PubMed

Hydroquinone galactoside (HQ-Gal) as a potential skin whitening agent was synthesized by the reaction of lactase (beta-galactosidase) from Kluyveromyces lactis, Aspergillus oryzae, Bacillus circulans, and Thermus sp. with lactose as a donor and HQ as an acceptor. Among these lactases, the acceptor reaction involving HQ and lactose with K. lactis lactase showed a higher conversion ratio to HQ-Gal (60.27%). HQ-Gal was purified using butanol partitioning and silica gel column chromatography. The structure of the purified HQ-Gal was determined by nuclear magnetic resonance, and the ionic product was observed at m/z 295 (C12H16O7Na)+ using matrix assisted laser desorption ionization time-of-flight mass spectrometry. HQ-Gal was identified as 4-hydroxyphenyl-beta-d-galactopyranoside. The optimum conditions for HQ-Gal synthesis by K. lactis determined using response surface methodology were 50 mM HQ, 60 mM lactose, and 20 U mL(-1) lactase. These conditions produced a yield of 2.01 g L(-1) HQ-Gal. The half maximal inhibitory concentration (IC50) of diphenylpicrylhydrazyl scavenging activity was 3.31 mM, indicating a similar antioxidant activity compared to beta-arbutin (IC50=3.95 mM). The Ki value of HQ-Gal (0.75 mM) against tyrosinase was smaller than that of beta-arbutin (Ki=1.97 mM), indicating its superiority as an inhibitor. HQ-Gal inhibited (23%) melanin synthesis without being significantly toxic to the cells, while beta-arbutin exhibited only 8% reduction of melanin synthesis in B16 melanoma cells compared with the control. These results indicate that HQ-Gal may be a suitable functional component in the cosmetics industry. PMID:20687552

Kim, Go-Eun; Lee, Jin-Ha; Jung, Sun-Hwa; Seo, Eun-Seong; Jin, Sheng-De; Kim, Ghahyun J; Cha, Jaeho; Kim, Eui-Joong; Park, Ki-Deok; Kim, Doman

2010-09-01

248

Altered superoxide dismutase activity by carbohydrate utilization in a Lactococcus lactis strain.  

PubMed

Reactive oxygen species, such as superoxide, can damage cellular components, such as proteins, lipids, and DNA. Superoxide dismutase (SOD) enzymes catalyze the conversion of superoxide anions to hydrogen peroxide and dioxygen. SOD is present in most lactococcal bacteria, which are commonly used as starters for manufacturing fermented dairy products and may have health benefits when taken orally. We assessed the effects of carbohydrate use on SOD activity in lactococci. In Lactococcus lactis ssp. lactis G50, the SOD activity of cells grown on lactose and galactose was higher than that on glucose; in Lactococcus lactis ssp. cremoris H61, SOD activity was independent of the type of carbohydrate used. We also investigated the activity of NADH oxidase, which is related to the production of superoxide in strains G50 and H61. Activity was highest in G50 cells grown on lactose, lower on galactose, and lowest on glucose, whereas activity in H61 cells did not differ with the carbohydrate source used. The SOD and NADH oxidase activities of strain G50 in three carbohydrates were linked. Strain G50 fermented lactose and galactose to lactate, acetate, formate, and ethanol (mixed-acid fermentation) and fermented glucose to mainly lactate (homolactic fermentation). Strain H61 fermented glucose, lactose, and galactose to mainly lactate (homolactic fermentation). In strain G50, when growth efficiency was reduced by adding a metabolic inhibitor to the growth medium, SOD activity was higher than in the control; however, the metabolism was homofermentative. Aerobic conditions, but not glucose-limited conditions, increased SOD activity, and mixed-acid fermentation occurred. We conclude that the effect of carbohydrate on SOD activity in lactococci is strain dependent and that the activity of commercial lactococci can be enhanced through carbohydrate selection for mixed-acid fermentation or by changing the energy distribution, thus enhancing the value of the starter and the resulting dairy products. PMID:24988023

Kimoto-Nira, H; Moriya, N; Ohmori, H; Suzuki, C

2014-07-01

249

Expression of food-grade phytase in Lactococcus lactis from optimized conditions in milk broth.  

PubMed

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. PMID:23453854

Miao, Yuzhi; Xu, Hui; Fei, Baojin; Qiao, Dairong; Cao, Yi

2013-07-01

250

Production of Human Papillomavirus Type 16 E7 Protein in Lactococcus lactis  

PubMed Central

The E7 protein of human papillomavirus type 16 was produced in Lactococcus lactis. Secretion allowed higher production yields than cytoplasmic production. In stationary phase, amounts of cytoplasmic E7 were reduced, while amounts of secreted E7 increased, suggesting a phase-dependent intracellular proteolysis. Fusion of E7 to the staphylococcal nuclease, a stable protein, resulted in a highly stable cytoplasmic protein. This work provides new candidates for development of viral screening systems and for oral vaccine against cervical cancer. PMID:11823236

Bermudez-Humaran, L. G.; Langella, P.; Miyoshi, A.; Gruss, A.; Tamez Guerra, R.; Montes de Oca-Luna, R.; Le Loir, Y.

2002-01-01

251

Permeabilization and lysis induced by bacteriocins and its effect on aldehyde formation by Lactococcus lactis.  

PubMed

Permeabilization induced by lacticin 3147, lactococcins A, B and M, enterocin AS-48 and nisin, bacteriocins described as cell membrane-pore forming and lytic agents, enhanced in all cases aldehyde formation by Lactococcus lactis IFPL730. Nevertheless, the conversion of isoleucine into 2-methylbutyraldehyde depended not only on the degree of permeabilization but also on the bacteriocin that caused the cell membrane damage. The highest values of 2-methylbutyraldehyde corresponded to cell suspensions containing lacticin 3147 and lactococcins, treatments that provoked further lysis in addition to induced permeabilization. PMID:16900333

Martínez-Cuesta, M Carmen; Requena, Teresa; Peláez, Carmen

2006-10-01

252

[Protoplast isolation from a nisin-forming culture of Streptococcus lactis strain MGU].  

PubMed

A procedure for protoplasts of Str. lactis producing nisin was developed. The following lysing factors were used for obtaining the protoplasts: lycozyme, LE enzyme, lysosubtilin and an enzyme isolated from the culture fluid A. levoris. The use of the above factors in different combinations provided an increase in the number of the formed protoplasts from 1-2 in the field of the microscope vision to 15-20, which amounted to 25 per cent of the total number of the cells visible under microscope. PMID:6428305

Baranova, I P; Kozlova, Iu I; Kozhevin, P A; Egorov, N S

1984-03-01

253

Characterization of KPC-2-producing Escherichia coli, Citrobacter freundii, Enterobacter cloacae, Enterobacter aerogenes, and Klebsiella oxytoca isolates from a Chinese Hospital.  

PubMed

Twelve nonduplicated KPC-2-producing enterobacterial isolates, including three Escherichia coli, two Citrobacter freundii, two Enterobacter cloacae, four Enterobacter aerogenes, and one Klebsiella oxytoca, were collected from various clinical samples within 18 months (March 2011 to September 2012). Two of the 12 patients died from infections caused by KPC-2-producing pathogens, while the rest of the patients with KPC-2-producing pathogens improved or were cured. The majority of the clinical isolates exhibited a high-level of resistance to oxyimino-cephalosporins and carbapenems, and possessed self-transferable bla(KPC-2)-carrying plasmids with sizes ranging from 20 to 120?kb. Most isolates carried bla(CTX-M) and plasmid-mediated quinolone resistance genes, while some isolates produced 16S rRNA methylases (ArmA or RmtB). The genetic environment of bla(KPC-2) of most clinical strains was consistent with the genetic structure surrounding bla(KPC-2) on the plasmid pKP048, which contains an integration structure of a Tn3-based transposon and partial Tn4401 segment. Inserted fragments (truncated bla(TEM)) were detected upstream of the bla(KPC-2) gene for two E. aerogenes strains. In conclusion, the enterobacterial isolates exhibited sporadic emergence and did not arise by clonal spread at our hospital. The outcome of infections caused by KPC-producing enterobacterial isolates and their mortality were closely associated with the baseline condition of patients. The spread of bla(KPC-2) gene between different enterobacterial species in China was mainly mediated by horizontal transfer of the Tn3-based transposons and not the bla(KPC-2)-carrying plasmids. PMID:24433026

Luo, Yanping; Yang, Jiyong; Ye, Liyan; Guo, Lin; Zhao, Qiang; Chen, Rong; Chen, Yong; Han, Xuelin; Zhao, Jingya; Tian, Shuguang; Han, Li

2014-08-01

254

A sulfur- and tyramine-regulated Klebsiella aerogenes operon containing the arylsulfatase (atsA) gene and the atsB gene.  

PubMed Central

The structural gene for arylsulfatase (atsA) of Klebsiella aerogenes was cloned into a pKI212 vector in Escherichia coli. Deletion analysis showed that the atsA gene with the promoter region was located within a 3.2-kilobase cloned segment. In E. coli cells which carried the plasmid, the synthesis of arylsulfatase was repressed by various sources of sulfur; the repression was relieved, in each case, by tyramine. Transfer of the plasmid into atsA or constitutive atsR mutant strains of K. aerogenes resulted in complementation of atsA but not of atsR. The nucleotide sequence of the 3.2-kilobase fragment was determined. Two open reading frames, the atsA gene and an unknown gene (atsB), were found. These are located between a potential promoter and a transcriptional terminator sequence. Deletion analysis suggests that atsB is a potential positive factor for the regulation of arylsulfatase. Analysis of the amino acid sequences of the first 13 amino acids from the N terminus of the purified secreted arysulfatase agrees with that of the nucleotide sequence of atsA. The leader peptide extends over 20 amino acids and has the characteristics of a signal sequence. Primer extension mapping of transcripts generated in vivo suggests that the synthesis of mRNA starts at a site 31 or 32 bases upstream from the ATG initiation codon of the atsB gene. By Northern (RNA) blot analysis of the transcripts induced by tyramine, we found a 2.7-kilobase transcript which is identical in size to the total sequence of the atsB and atsA genes. Thus, the ats operon is composed of two cistrons and is regulated by sulfur and tyramine. Images FIG. 5 FIG. 6 PMID:2180918

Murooka, Y; Ishibashi, K; Yasumoto, M; Sasaki, M; Sugino, H; Azakami, H; Yamashita, M

1990-01-01

255

Effect of signal peptides on the secretion of ?-cyclodextrin glucanotransferase in Lactococcus lactis NZ9000.  

PubMed

Cyclodextrin glucanotransferase (CGTase) is an extracellular enzyme which catalyzes the formation of cyclodextrin from starch. The production of CGTase using lactic acid bacterium is an attractive alternative and safer strategy to produce CGTase. In this study, we report the construction of genetically modified Lactococcus lactis strains harboring plasmids that secrete the Bacillus sp. G1 ?-CGTase, with the aid of the signal peptides (SPs) SPK1, USP45 and native SP (NSP). Three constructed vectors, pNZ:NSP:CGT, pNZ:USP:CGT and pNZ:SPK1:CGT, were developed in this study. Each vector harbored a different SP fused to the CGTase. The formation of halo zones on starch plates indicated the production and secretion of ?-CGTase by the recombinants. The expression of this enzyme is shown by sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) and zymogram analysis. A band size of ?75 kDa corresponding to ?-CGTase is identified in the intracellular and the extracellular environments of the host after medium modification. The replacement of glucose by starch in the medium was shown to induce ?-CGTase production in L. lactis. Although ?-CGTase production is comparatively low in NZ:SPK1:CGT, the SP SPK1 was shown to have higher secretion efficiency compared to the other SPs used in this study. PMID:23295307

Subramaniam, Menaga; Baradaran, Ali; Rosli, Md Illias; Rosfarizan, Mohamad; Khatijah, Yusoff; Raha, Abdul Rahim

2012-01-01

256

Biotransformation of monoterpene alcohols by Saccharomyces cerevisiae, Torulaspora delbrueckii and Kluyveromyces lactis.  

PubMed

Monoterpenoids are important flavour compounds produced by many plant species, including grapes (Vitis vinifera) and hops (Humulus lupulus). Biotransformation reactions involving monoterpenoids have been characterized in filamentous fungi, but few examples have been observed in yeasts. As monoterpenoids are in contact with yeasts during beer and wine production, biotransformation reactions may occur during the fermentation of these beverages. This paper describes the biotransformation of monoterpene alcohols, of significance in the alcoholic beverage industries, by three yeast species. All three species analysed had the ability to convert monoterpenoids. Saccharomyces cerevisiae and Kluyveromyces lactis reduced geraniol into citronellol, whilst all three yeasts produced linalool from both geraniol and nerol. Monocyclic alpha-terpineol was formed from both linalool and nerol, by all three yeasts. alpha-Terpineol was then converted into the diol cis-terpin hydrate. K. lactis and Torulaspora delbrueckii also had the ability to form geraniol from nerol. Finally, the stereospecificity of terpenoid formation was analysed. Both (+) and (-) enantiomers of linalool and alpha-terpineol were formed in roughly equal quantities, from either geraniol or nerol. PMID:10790686

King, A; Richard Dickinson, J

2000-04-01

257

A Food-Grade Cloning System for Industrial Strains of Lactococcus lactis  

PubMed Central

We have previously reported the construction of a food-grade cloning vector for Lactococcus using the ochre suppressor, supB, as the selective marker. This vector, pFG1, causes only a slight growth inhibition in the laboratory strain MG1363 but is unstable in the industrial strains tested. As supB suppresses both amber and ochre stop codons, which are present in 82% of all known lactococcal genes, this undesirable finding may result from the accumulation of elongated mistranslated polypeptides. Here, we report the development of a new food-grade cloning vector, pFG200, which is suitable for overexpressing a variety of genes in industrial strains of Lactococcus lactis. The vector uses an amber suppressor, supD, as selectable marker and consists entirely of Lactococcus DNA, with the exception of a small polylinker region. Using suppressible pyrimidine auxotrophs, selection and maintenance are efficient in any pyrimidine-free medium including milk. Importantly, the presence of this vector in a variety of industrial strains has no significant effect on the growth rate or the rate of acidification in milk, making this an ideal system for food-grade modification of industrially relevant L. lactis strains. The usefulness of this system is demonstrated by overexpressing the pepN gene in a number of industrial backgrounds. PMID:10742196

S?rensen, Kim I.; Larsen, Rasmus; Kibenich, Annette; Junge, Mette P.; Johansen, Eric

2000-01-01

258

The carbohydrate metabolism signature of lactococcus lactis strain A12 reveals its sourdough ecosystem origin.  

PubMed

Lactococcus lactis subsp. lactis strain A12 was isolated from sourdough. Combined genomic, transcriptomic, and phenotypic analyses were performed to understand its survival capacity in the complex sourdough ecosystem and its role in the microbial community. The genome sequence comparison of strain A12 with strain IL1403 (a derivative of an industrial dairy strain) revealed 78 strain-specific regions representing 23% of the total genome size. Most of the strain-specific genes were involved in carbohydrate metabolism and are potentially required for its persistence in sourdough. Phenotype microarray, growth tests, and analysis of glycoside hydrolase content showed that strain A12 fermented plant-derived carbohydrates, such as arabinose and ?-galactosides. Strain A12 exhibited specific growth rates on raffinose that were as high as they were on glucose and was able to release sucrose and galactose outside the cell, providing soluble carbohydrates for sourdough microflora. Transcriptomic analysis identified genes specifically induced during growth on raffinose and arabinose and reveals an alternative pathway for raffinose assimilation to that used by other lactococci. PMID:23872564

Passerini, Delphine; Coddeville, Michèle; Le Bourgeois, Pascal; Loubière, Pascal; Ritzenthaler, Paul; Fontagné-Faucher, Catherine; Daveran-Mingot, Marie-Line; Cocaign-Bousquet, Muriel

2013-10-01

259

Purification and characterization of a novel glucansucrase from Leuconostoc lactis EG001.  

PubMed

A gene encoding glucansucrase was identified in Leuconostoc lactis EG001 isolated from lactic acid bacteria (LAB) in Kimchi, a traditional Korean fermented food. The L. lactis EG001 glucansucrase gene consists of 4503 bp open reading frame (ORF) and encodes an enzyme of 1500 amino acids with an apparent molecular mass of 165 kDa. The deduced amino-acid sequence showed the highest amino-acid sequence identity (75%) to that of dextransucrase of L. mesenteroides. The gene was cloned and over-expressed in Escherichia coli strain. The recombinant enzyme was purified via Ni-NTA affinity chromatography and its enzymatic properties were characterized. The enzyme exhibited optimum activity at 30 degrees C and pH 5.0. In addition, the enzyme was able to catalyze the glycosylation of l-ascorbic acid to l-ascorbic acid 2-glucoside. The glycosylated product via EG001 glucansucrase has the potential as an antioxidant in industrial applications. PMID:19853426

Kim, Yong-Mo; Yeon, Min Ji; Choi, Nack-Shick; Chang, Young-Hyo; Jung, Min Young; Song, Jae Jun; Kim, Joong Su

2010-07-20

260

The Carbohydrate Metabolism Signature of Lactococcus lactis Strain A12 Reveals Its Sourdough Ecosystem Origin  

PubMed Central

Lactococcus lactis subsp. lactis strain A12 was isolated from sourdough. Combined genomic, transcriptomic, and phenotypic analyses were performed to understand its survival capacity in the complex sourdough ecosystem and its role in the microbial community. The genome sequence comparison of strain A12 with strain IL1403 (a derivative of an industrial dairy strain) revealed 78 strain-specific regions representing 23% of the total genome size. Most of the strain-specific genes were involved in carbohydrate metabolism and are potentially required for its persistence in sourdough. Phenotype microarray, growth tests, and analysis of glycoside hydrolase content showed that strain A12 fermented plant-derived carbohydrates, such as arabinose and ?-galactosides. Strain A12 exhibited specific growth rates on raffinose that were as high as they were on glucose and was able to release sucrose and galactose outside the cell, providing soluble carbohydrates for sourdough microflora. Transcriptomic analysis identified genes specifically induced during growth on raffinose and arabinose and reveals an alternative pathway for raffinose assimilation to that used by other lactococci. PMID:23872564

Passerini, Delphine; Coddeville, Michele; Le Bourgeois, Pascal; Loubiere, Pascal; Ritzenthaler, Paul; Fontagne-Faucher, Catherine; Cocaign-Bousquet, Muriel

2013-01-01

261

Exploring optimization parameters to increase ssDNA recombineering in Lactococcus lactis and Lactobacillus reuteri.  

PubMed

Single-stranded DNA (ssDNA) recombineering is a technology which is used to make subtle changes in the chromosome of several bacterial genera. Cells which express a single-stranded DNA binding protein (RecT or Bet) are transformed with an oligonucleotide which is incorporated via an annealing and replication-dependent mechanism. By in silico analysis we identified ssDNA binding protein homologs in the genus Lactobacillus and Lactococcus lactis. To assess whether we could further improve the recombineering efficiency in Lactobacillus reuteri ATCC PTA 6475 we expressed several RecT homologs in this strain. RecT derived from Enterococcus faecalis CRMEN 19 yielded comparable efficiencies compared with a native RecT protein, but none of the other proteins further increased the recombineering efficiency. We successfully improved recombineering efficiency 10-fold in L. lactis by increasing oligonucleotide concentration combined with the use of oligonucleotides containing phosphorothioate-linkages (PTOs). Surprisingly, neither increased oligonucleotide concentration nor PTO linkages enhanced recombineering in L. reuteri 6475. To emphasize the utility of this technology in improving probiotic features we modified six bases in a transcriptional regulatory element region of the pdu-operon of L. reuteri 6475, yielding a 3-fold increase in the production of the antimicrobial compound reuterin. Directed genetic modification of lactic acid bacteria through ssDNA recombineering will simplify strain improvement in a way that, when mutating a single base, is genetically indistinguishable from strains obtained through directed evolution. PMID:22750793

Van Pijkeren, Jan-Peter; Neoh, Kar Mun; Sirias, Denise; Findley, Anthony S; Britton, Robert A

2012-01-01

262

Bifidobacterium animalis subsp. lactis fermented milk product reduces inflammation by altering a niche for colitogenic microbes  

PubMed Central

Intestinal health requires the coexistence of eukaryotic self with the gut microbiota and dysregulated host-microbial interactions can result in intestinal inflammation. Here, we show that colitis improved in T-bet?/?Rag2?/? mice that consumed a fermented milk product containing Bifidobacterium animalis subsp. lactis DN-173 010 strain. A decrease in cecal pH and alterations in short chain fatty acid profiles occurred with consumption, and there were concomitant increases in the abundance of select lactate-consuming and butyrate-producing bacteria. These metabolic shifts created a nonpermissive environment for the Enterobacteriaceae recently identified as colitogenic in a T-bet?/?Rag2?/? ulcerative colitis mouse model. In addition, 16S rRNA-based analysis of the T-bet?/?Rag2?/?fecal microbiota suggest that the structure of the endogenous gut microbiota played a key role in shaping the host response to the bacterial strains studied herein. We have identified features of the gut microbiota, at the membership and functional level, associated with response to this B. lactis-containing fermented milk product, and therefore this model provides a framework for evaluating and optimizing probiotic-based functional foods. PMID:20921388

Veiga, Patrick; Gallini, Carey Ann; Beal, Chloe; Michaud, Monia; Delaney, Mary L.; DuBois, Andrea; Khlebnikov, Artem; van Hylckama Vlieg, Johan E.T.; Punit, Shivesh; Glickman, Jonathan N.; Onderdonk, Andrew; Glimcher, Laurie H.; Garrett, Wendy S.

2010-01-01

263

Statistical investigation of Kluyveromyces lactis cells permeabilization with ethanol by response surface methodology  

PubMed Central

The aim of our study was to select the optimal operating conditions to permeabilize Kluyveromyces lactis cells using ethanol as a solvent as an alternative to cell disruption and extraction. Cell permeabilization was carried out by a non-mechanical method consisting of chemical treatment with ethanol, and the results were expressed as ?-galactosidase activity. Experiments were conducted under different conditions of ethanol concentration, treatment time and temperature according to a central composite rotatable design (CCRD), and the collected results were then worked out by response surface methodology (RSM). Cell permeabilization was improved by an increase in ethanol concentration and simultaneous decreases in the incubation temperature and treatment time. Such an approach allowed us to identify an optimal range of the independent variables within which the ?-galactosidase activity was optimized. A maximum permeabilization of 2,816 mmol L?1 oNP min?1 g?1 was obtained by treating cells with 75.0% v/v of ethanol at 20.0 °C for 15.0 min. The proposed methodology resulted to be effective and suited for K. lactis cells permeabilization at a lab-scale and promises to be of possible interest for future applications mainly in the food industry. PMID:24688494

de Faria, Janaina T.; Rocha, Pollyana F.; Converti, Attilio; Passos, Flavia M.L.; Minim, Luis A.; Sampaio, Fabio C.

2013-01-01

264

Modeling of the Competitive Growth of Listeria monocytogenes and Lactococcus lactis in Vegetable Broth  

PubMed Central

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

Breidt, Frederick; Fleming, Henry P.

1998-01-01

265

Assessing the effects of exposure to environmental stress on some functional properties of Bifidobacterium animalis ssp. lactis.  

PubMed

This study assessed the effects of exposing a strain of Bifidobacterium animalis ssp. lactis to acid, bile and osmotic stresses on antagonistic properties, biofilm formation and antibiotic susceptibility/resistance profile. Exposure to each stress factor appeared to have no significant effect on the antagonism against Escherichia coli NCTC 12900 and Salmonella enterica serovar Enteritidis PT4. No suppression in biofilm formation due to exposure to stress was observed. Bile and osmotic stresses resulted in significantly higher biofilm formation. Expression of an exopolysaccharide synthesis gene, gtf 01207, was significantly higher when the B. animalis ssp. lactis strain was exposed to osmotic stress. Susceptibility of the B. animalis ssp. lactis strain to chloramphenicol, erythromycin, ampicillin and vancomycin, and resistance to tetracycline remained unchanged when exposed to each stress. The expression of a tetracycline resistance gene, tet(W), was significantly higher when exposed to each stress. These results may suggest that the potential for the B. animalis ssp. lactis strain to provide probiotic benefit, after exposure to the stressful conditions of the gastrointestinal tract, remains intact. PMID:25097108

Amund, O D; Ouoba, L I I; Sutherland, J P; Ghoddusi, H B

2014-12-01

266

Genome Sequence of Lactobacillus delbrueckii subsp. lactis CNRZ327, a Dairy Bacterium with Anti-Inflammatory Properties  

PubMed Central

Lactobacillus delbrueckii subsp. lactis CNRZ327 is a dairy bacterium with anti-inflammatory properties both in vitro and in vivo. Here, we report the genome sequence of this bacterium, which appears to contain no less than 215 insertion sequence (IS) elements, an exceptionally high number regarding the small genome size of the strain. PMID:25035318

El Kafsi, Hela; Binesse, Johan; Loux, Valentin; Buratti, Julien; Boudebbouze, Samira; Dervyn, Rozenn; Hammani, Amal; Maguin, Emmanuelle

2014-01-01

267

Genotypic and Phenotypic Analysis of Dairy Lactococcus lactis Biodiversity in Milk: Volatile Organic Compounds as Discriminating Markers  

PubMed Central

The diversity of nine dairy strains of Lactococcus lactis subsp. lactis in fermented milk was investigated by both genotypic and phenotypic analyses. Pulsed-field gel electrophoresis and multilocus sequence typing were used to establish an integrated genotypic classification. This classification was coherent with discrimination of the L. lactis subsp. lactis bv. diacetylactis lineage and reflected clonal complex phylogeny and the uniqueness of the genomes of these strains. To assess phenotypic diversity, 82 variables were selected as important dairy features; they included physiological descriptors and the production of metabolites and volatile organic compounds (VOCs). Principal-component analysis (PCA) demonstrated the phenotypic uniqueness of each of these genetically closely related strains, allowing strain discrimination. A method of variable selection was developed to reduce the time-consuming experimentation. We therefore identified 20 variables, all associated with VOCs, as phenotypic markers allowing discrimination between strain groups. These markers are representative of the three metabolic pathways involved in flavor: lipolysis, proteolysis, and glycolysis. Despite great phenotypic diversity, the strains could be divided into four robust phenotypic clusters based on their metabolic orientations. Inclusion of genotypic diversity in addition to phenotypic characters in the classification led to five clusters rather than four being defined. However, genotypic characters make a smaller contribution than phenotypic variables (no genetic distances selected among the most contributory variables). This work proposes an original method for the phenotypic differentiation of closely related strains in milk and may be the first step toward a predictive classification for the manufacture of starters. PMID:23709512

Dhaisne, Amandine; Guellerin, Maeva; Laroute, Valerie; Laguerre, Sandrine; Le Bourgeois, Pascal; Loubiere, Pascal

2013-01-01

268

Kluyveromyces lactis and Saccharomyces cerevisiae , two potent deacidifying and volatile-sulphur-aroma-producing microorganisms of the cheese ecosystem  

Microsoft Academic Search

Cheese flavour is the result of complex biochemical transformations attributed to bacteria and yeasts grown on the curd of smear-ripened cheeses. Volatile sulphur compounds (VSCs) are responsible for the characteristic aromatic notes of several cheeses. In the present study, we have assessed the ability of Kluyveromyces lactis, Kluyveromyces marxianus and Saccharomyces cerevisiae strains, which are frequently isolated from smear-ripened cheeses,

Dafni-Maria Kagkli; Roselyne Tâche; Timothy M. Cogan; Colin Hill; Serge Casaregola; Pascal Bonnarme

2006-01-01

269

An alternative, arginase-independent pathway for arginine metabolism in Kluyveromyces lactis involves guanidinobutyrase as a key enzyme.  

PubMed

Most available knowledge on fungal arginine metabolism is derived from studies on Saccharomyces cerevisiae, in which arginine catabolism is initiated by releasing urea via the arginase reaction. Orthologues of the S. cerevisiae genes encoding the first three enzymes in the arginase pathway were cloned from Kluyveromyces lactis and shown to functionally complement the corresponding deletion in S. cerevisiae. Surprisingly, deletion of the single K. lactis arginase gene KlCAR1 did not completely abolish growth on arginine as nitrogen source. Growth rate of the deletion mutant strongly increased during serial transfer in shake-flask cultures. A combination of RNAseq-based transcriptome analysis and (13)C-(15)N-based flux analysis was used to elucidate the arginase-independent pathway. Isotopic (13)C(15)N-enrichment in ?-aminobutyrate revealed succinate as the entry point in the TCA cycle of the alternative pathway. Transcript analysis combined with enzyme activity measurements indicated increased expression in the Klcar1? mutant of a guanidinobutyrase (EC.3.5.3.7), a key enzyme in a new pathway for arginine degradation. Expression of the K. lactis?KLLA0F27995g (renamed KlGBU1) encoding guanidinobutyrase enabled S. cerevisiae to use guanidinobutyrate as sole nitrogen source and its deletion in K. lactis almost completely abolish growth on this nitrogen source. Phylogenetic analysis suggests that this enzyme activity is widespread in fungi. PMID:24912400

Romagnoli, G; Verhoeven, M D; Mans, R; Fleury Rey, Y; Bel-Rhlid, R; van den Broek, M; Seifar, R Maleki; Ten Pierick, A; Thompson, M; Müller, V; Wahl, S A; Pronk, J T; Daran, J M

2014-07-01

270

Identification, Cloning, and Characterization of a Lactococcus lactis Branched-Chain ?-Keto Acid Decarboxylase Involved in Flavor Formation  

PubMed Central

The biochemical pathway for formation of branched-chain aldehydes, which are important flavor compounds derived from proteins in fermented dairy products, consists of a protease, peptidases, a transaminase, and a branched-chain ?-keto acid decarboxylase (KdcA). The activity of the latter enzyme has been found only in a limited number of Lactococcus lactis strains. By using a random mutagenesis approach, the gene encoding KdcA in L. lactis B1157 was identified. The gene for this enzyme is highly homologous to the gene annotated ipd, which encodes a putative indole pyruvate decarboxylase, in L. lactis IL1403. Strain IL1403 does not produce KdcA, which could be explained by a 270-nucleotide deletion at the 3? terminus of the ipd gene encoding a truncated nonfunctional decarboxylase. The kdcA gene was overexpressed in L. lactis for further characterization of the decarboxylase enzyme. Of all of the potential substrates tested, the highest activity was observed with branched-chain ?-keto acids. Moreover, the enzyme activity was hardly affected by high salinity, and optimal activity was found at pH 6.3, indicating that the enzyme might be active under cheese ripening conditions. PMID:15640202

Smit, Bart A.; van Hylckama Vlieg, Johan E. T.; Engels, Wim J. M.; Meijer, Laura; Wouters, Jan T. M.; Smit, Gerrit

2005-01-01

271

Contribution of Lactococcus lactis Reducing Properties to the Downregulation of a Major Virulence Regulator in Staphylococcus aureus, the agr System.  

PubMed

Staphylococcus aureus is a major cause of food poisoning outbreaks associated with dairy products, because of the ingestion of preformed enterotoxins. The biocontrol of S. aureus using lactic acid bacteria (LAB) offers a promising opportunity to fight this pathogen while respecting the product ecosystem. We had previously established the ability of Lactococcus lactis, a lactic acid bacterium widely used in the dairy industry, to downregulate a major staphylococcal virulence regulator, the accessory gene regulator (agr) system, and, as a consequence, agr-controlled enterotoxins. In the present paper, we have shown that the oxygen-independent reducing properties of L. lactis contribute to agr downregulation. Neutralizing lactococcal reduction by adding potassium ferricyanide or maintaining the oxygen pressure constant at 50% released agr downregulation in the presence of L. lactis. This downregulation still occurred in an S. aureus srrA mutant, indicating that the staphylococcal respiratory response regulator SrrAB was not the only component in the signaling pathway. Therefore, this study clearly demonstrates the ability of L. lactis reducing properties to interfere with the expression of S. aureus virulence, thus highlighting this general property of LAB as a lever to control the virulence expression of this major pathogen in a food context and beyond. PMID:25192992

Nouaille, Sébastien; Rault, Lucie; Jeanson, Sophie; Loubière, Pascal; Le Loir, Yves; Even, Sergine

2014-11-15

272

Effect of autochthonous bacteriocin-producing Lactococcus lactis on bacterial population dynamics and growth of halotolerant bacteria in Brazilian charqui.  

PubMed

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. PMID:25084676

Biscola, Vanessa; Abriouel, Hikmate; Todorov, Svetoslav Dimitrov; Capuano, Verena Sant'Anna Cabral; Gálvez, Antonio; Franco, Bernadette Dora Gombossy de Melo

2014-12-01

273

Potential probiotic Bifidobacterium animalis ssp. lactis 420 prevents weight gain and glucose intolerance in diet-induced obese mice.  

PubMed

Alterations of the gut microbiota and mucosal barrier are linked with metabolic diseases. Our aim was to investigate the potential benefit of the potential probiotic Bifidobacterium animalis ssp. lactis 420 in reducing high-fat diet-induced body weight gain and diabetes in mice. In the obesity model, C57Bl/6J mice were fed a high-fat diet (60 energy %) for 12 weeks, and gavaged daily with B. lactis 420 (109 cfu) or vehicle. In the diabetes model, mice were fed a high-fat, ketogenic diet (72 energy % fat) for 4 weeks, with a 6-week subsequent treatment with B. lactis 420 (108-1010 cfu/day) or vehicle, after which they were analysed for body composition. We also analysed glucose tolerance, plasma lipopolysaccharide and target tissue inflammation using only one of the B. lactis 420 groups (109 cfu/day). Intestinal bacterial translocation and adhesion were analysed in a separate experiment using an Escherichia coli gavage. Body fat mass was increased in both obese (10.7±0.8 g (mean ± standard error of mean) vs. 1.86±0.21 g, P<0.001) and diabetic mice (3.01±0.4 g vs. 1.14±0.15 g, P<0.001) compared to healthy controls. Treatment with B. lactis 420 significantly decreased fat mass in obese (7.83 ± 0.67 g, P=0.007 compared to obese with vehicle) and diabetic mice (1.89 ± 0.16 g, P=0.02 for highest dose). This was reflected as reduced weight gain and improved glucose tolerance. Furthermore, B. lactis 420 decreased plasma lipopolysaccharide levels (P<0.001), liver inflammation (P=0.04), and E. coli adhesion in the distal gut (P<0.05). In conclusion, B. lactis 420 reduces fat mass and glucose intolerance in both obese and diabetic mice. Reduced intestinal mucosal adherence and plasma lipopolysaccharide suggest a mechanism related to reduced translocation of gut microbes. PMID:25062610

Stenman, L K; Waget, A; Garret, C; Klopp, P; Burcelin, R; Lahtinen, S

2014-12-01

274

Oral immunization with Lactococcus lactis secreting attenuated recombinant staphylococcal enterotoxin B induces a protective immune response in a murine model  

PubMed Central

Background Staphylococcus aureus is unrestrictedly found in humans and in animal species that maintain thermal homeostasis. Inadequate cleaning of processing equipment or inappropriate handling can contaminate processed food and cause severe food poisoning. Staphylococcal enterotoxin B (SEB), a potent superantigenic exotoxin, is produced by 50% of clinical isolates of S. aureus and is associated with massive food poisoning and with the induction of toxic shock syndrome. Results A gene sequence encoding a recombinant SEB (rSEB), devoid of superantigenic activity, was successfully cloned and expressed in a cytoplasmic or a secreted form in the food-grade lactic acid bacterium Lactococcus lactis. The recombinant protein detected in the cytoplasm or in the culture medium exhibited the expected molecular mass and was recognized by a SEB-polyclonal antibody. Oral immunization with the recombinant L. lactis strains induced a protective immune response in a murine model of S. aureus infection. Immunized mice survived intraperitoneal challenge with an S. aureus SEB-producer strain. Counts of S. aureus in the spleen of rSEB-immunized mice were significantly reduced. The rSEB-immunized mice showed significant titers of anti-SEB IgA and IgG in stools and serum, respectively. Both recombinant L. lactis strains were able to elicit cellular or systemic immune responses in mice, with no significant difference if rSEB was produced in its cytoplasmic or secreted form. However, recombinant L. lactis expressing the cytoplasmic rSEB increased the survival rate of the challenged mice by 43%. Conclusions These findings show the vaccine efficacy of L. lactis carrying an attenuated SEB, in a murine model, following lethal S. aureus challenge. PMID:23561053

2013-01-01

275

Functional analysis of the Lactococcus lactis galU and galE genes and their impact on sugar nucleotide and exopolysaccharide biosynthesis  

Microsoft Academic Search

We studied the UDP-glucose pyrophosphorylase (galU) and UDP-galactose epimerase (galE) genes of Lactococcus lactis MG1363 to investigate their involvement in biosynthesis of UDP-glucose and UDP-galactose, which are precursors of glucose- and galactose-containing exopolysaccharides (EPS) in L. lactis. The lactococcal galU gene was identified by a PCR approach using degenerate primers and was found by Northern blot analysis to be transcribed

INGEBORG C. BOELS; ANA RAMOS; MICHIEL KLEEREBEZEM; Vos de W. M

2001-01-01

276

Bifidobacterium animalis ssp. lactis BI07 modulates the tumor necrosis factor alpha-dependent imbalances of the enterocyte-associated intestinal microbiota fraction.  

PubMed

Using a previously developed in vitro model to characterize the enterocyte-adherent microbiota fraction, we explored the potential of the probiotic strain Bifidobacterium animalis ssp. lactis BI07 to modulate the inflammation-dependent dysbioses of the enterocyte-adherent microbiota from 12 healthy human donors. According to our findings, B. animalis ssp. lactis BI07 is effective in limiting the increase of pro-inflammatory pathobionts on the inflamed mucosal site, supporting the recovery of a mutualistic community. PMID:24964713

Centanni, Manuela; Turroni, Silvia; Rampelli, Simone; Biagi, Elena; Quercia, Sara; Consolandi, Clarissa; Severgnini, Marco; Brigidi, Patrizia; Candela, Marco

2014-08-01

277

Increased D-alanylation of lipoteichoic acid and a thickened septum are main determinants in the nisin resistance mechanism of Lactococcus lactis  

Microsoft Academic Search

Nisin is a post-translationally modified antimicrobial peptide produced by Lactococcus lactis\\u000awhich binds to lipid II in the membrane to form pores and inhibit cell-wall synthesis. A nisinresistant\\u000a(NisR) strain of L. lactis, which is able to grow at a 75-fold higher nisin concentration than\\u000aits parent strain, was investigated with respect to changes in the cell wall. Direct binding

Naomi E. Kramer; Hester E. Hasper; Siegfried Morath; Ben de Kruijff; Thomas Hartung; Eddy J. Smid; Eefjan Breukink; Jan Kok; Oscar P. Kuipers

2008-01-01

278

Expression and purification of a truncated macrophage colony stimulating factor in Kluyveromyces lactis.  

PubMed

A truncated human macrophage colony stimulating factor (M-CSF) cDNA encoding amino acid residues from 3 to 149 of the native M-CSF was obtained by using polymerase chain reaction. When inserted into plasmid pCXJ1 and psPHO5 and introduced into Kluyveromyces lactis, it directs the the secretory expression of the biologically active dimeric form of M-CSF. Through a four-step purification protocol, i.e. ammonium sulfate salting out, DEAE-cellulose column chromatography, hydrophobic chromatography on phenyl-sepharose and Mono Q fast protein liquid chromatography, the recombinant truncated M-CSF was purified to homogenerity and show its apparent molecular mass at 21KDa on reduced SDS-PAGE, with a specific activity of 1.21 x 10(7) units/mg protein. PMID:7849653

Hua, Z; Liang, X; Zhu, D

1994-09-01

279

Structural Basis for the Transcriptional Regulation of Heme Homeostasis in Lactococcus lactis*  

PubMed Central

Although heme is a crucial element for many biological processes including respiration, heme homeostasis should be regulated strictly due to the cytotoxicity of free heme molecules. Numerous lactic acid bacteria, including Lactococcus lactis, acquire heme molecules exogenously to establish an aerobic respiratory chain. A heme efflux system plays an important role for heme homeostasis to avoid cytotoxicity of acquired free heme, but its regulatory mechanism is not clear. Here, we report that the transcriptional regulator heme-regulated transporter regulator (HrtR) senses and binds a heme molecule as its physiological effector to regulate the expression of the heme-efflux system responsible for heme homeostasis in L. lactis. To elucidate the molecular mechanisms of how HrtR senses a heme molecule and regulates gene expression for the heme efflux system, we determined the crystal structures of the apo-HrtR·DNA complex, apo-HrtR, and holo-HrtR at a resolution of 2.0, 3.1, and 1.9 ?, respectively. These structures revealed that HrtR is a member of the TetR family of transcriptional regulators. The residue pair Arg-46 and Tyr-50 plays a crucial role for specific DNA binding through hydrogen bonding and a CH-? interaction with the DNA bases. HrtR adopts a unique mechanism for its functional regulation upon heme sensing. Heme binding to HrtR causes a coil-to-helix transition of the ?4 helix in the heme-sensing domain, which triggers a structural change of HrtR, causing it to dissociate from the target DNA for derepression of the genes encoding the heme efflux system. HrtR uses a unique heme-sensing motif with bis-His (His-72 and His-149) ligation to the heme, which is essential for the coil-to-helix transition of the ?4 helix upon heme sensing. PMID:22798069

Sawai, Hitomi; Yamanaka, Masaru; Sugimoto, Hiroshi; Shiro, Yoshitsugu; Aono, Shigetoshi

2012-01-01

280

Toxicity of Nalidixic Acid on Candida albicans, Saccharomyces cerevisiae, and Kluyveromyces lactis  

PubMed Central

The antibacterial drug nalidixic acid (Nal) can suppress the growth of Candida albicans at levels of the drug normally found in urine. Growth suppression increases as drug levels are increased, and Nal also causes a similar proportional inhibition of the synthesis of all cellular macromolecules. However, growth temperature (25 versus 37 C) and the divalent cations Mg2+ and Mn2+ can increase C. albicans resistance to Nal. Also, nitrogen depletion of Candida shows that Nal-treated and untreated cells exhibit no difference in leucine uptake during readaptation to nitrogen. In Nal-treated, nitrogen-starved cells, ribonucleic acid and deoxyribonucleic acid (DNA) biosynthesis are less affected than in unstarved Nal-treated cells, but of the two nucleic acids DNA synthesis is the most affected. Nal-resistant strains of C. albicans exhibit a slight toxicity for macromolecular synthesis. Nal treatment of a synchronized population of Saccharomyces cerevisiae results in an increase in the culture mean doubling time of, at most, 20%, but Nal causes the loss of synchronous cell division. With a synchronized population of Kluyveromyces lactis, Nal causes an increase in the mean doubling time of upwards of 300%, with synchrony of cell division being maintained. It is known that S. cerevisiae asynchronously synthesizes mitochondrial DNA during the cell cycle, whereas with K. lactis it is synchronous. Thus, with C. albicans Nal toxicity is dependent both on the dose and the physiological state of the cell. Furthermore, Nal inhibits growth of yeast with synchronous mitochondrial DNA synthesis more adversely than yeast with asynchronous mitochondrial DNA synthesis. PMID:769680

Sobieski, Rodney J.; Brewer, Alan R.

1976-01-01

281

Genetic and Physiological Responses of Bifidobacterium animalis subsp. lactis to Hydrogen Peroxide Stress  

PubMed Central

Consumer interest in probiotic bifidobacteria is increasing, but industry efforts to secure high cell viability in foods is undermined by these anaerobes' sensitivity to oxidative stress. To address this limitation, we investigated genetic and physiological responses of two fully sequenced Bifidobacterium animalis subsp. lactis strains, BL-04 and DSM 10140, to hydrogen peroxide (H2O2) stress. Although the genome sequences for these strains are highly clonal, prior work showed that they differ in both intrinsic and inducible H2O2 resistance. Transcriptome analysis of early-stationary-phase cells exposed to a sublethal H2O2 concentration detected significant (P < 0.05) changes in expression of 138 genes in strain BL-04 after 5 min and 27 genes after 20 min. Surprisingly, no significant changes in gene expression were detected in DSM 10140 at either time. Genomic data suggested that differences in H2O2 stress resistance might be due to a mutation in a BL-04 gene encoding long-chain fatty acid coenzyme A (CoA) ligase. To explore this possibility, membrane fatty acids were isolated and analyzed by gas chromatography-mass spectrometry (GC-MS). Results confirmed that the strains had significantly different lipid profiles: the BL-04 membrane contained higher percentages of C14:0 and C16:0 and lower percentages of C18:1n9. Alteration of the DSM 10140 membrane lipid composition using modified growth medium to more closely mimic that of BL-04 yielded cells that showed increased intrinsic resistance to lethal H2O2 challenge but did not display an inducible H2O2 stress response. The results show that deliberate stress induction or membrane lipid modification can be employed to significantly improve H2O2 resistance in B. animalis subsp. lactis strains. PMID:23772066

Oberg, Taylor S.; Ward, Robert E.; Steele, James L.

2013-01-01

282

Structural and kinetic studies of sugar binding to galactose mutarotase from Lactococcus lactis.  

PubMed

Galactose mutarotase catalyzes the conversion of beta-D-galactose to alpha-D-galactose in the Leloir pathway for galactose metabolism. The high resolution x-ray structure of the dimeric enzyme from Lactococcus lactis was recently solved and shown to be topologically similar to the 18-stranded, anti-parallel beta-motif observed for domain 5 of beta-galactosidase. In addition to determining the overall molecular fold of galactose mutarotase, this initial investigation also provided a detailed description of the electrostatic interactions between the enzyme and its physiologically relevant substrate, galactose. Specifically, the side chains of His-96 and His-170 were shown to be located within hydrogen bonding distance to the C-5 oxygen of the substrate, while the carboxylate of Glu-304 was positioned near the C-1 hydroxyl group of the sugar. On the basis of this initial study, a possible role for Glu-304 as the general acid/base group in catalysis was put forth. Here we describe the combined x-ray crystallographic and kinetic analyses of L. lactis galactose mutarotase complexed with D-glucose, D-fucose, D-quinovose, L-arabinose, or D-xylose. These investigations have revealed that there are several distinct binding modes for these sugars, which are dependent upon the spatial orientation of the C-4 hydroxyl group. In those sugars with the same C-4 hydroxyl group orientation as galactose, their C-1 hydroxyl groups are invariably located near Glu-304. For those sugars, which have the same C-4 hydroxyl group configuration as glucose, the C-1 hydroxyls are typically located near Asp-243. These different binding modes correlate with both the observed kinetic parameters and the presence or absence of a hydrogen bond between the guanidinium group of Arg-71 and the C-4 hydroxyl group of the sugar ligand. PMID:12218067

Thoden, James B; Kim, Jungwook; Raushel, Frank M; Holden, Hazel M

2002-11-22

283

6-Phosphogluconate dehydrogenase from Lactococcus lactis: a role for arginine residues in binding substrate and coenzyme.  

PubMed Central

A gene encoding 6-phosphogluconate dehydrogenase (6-PGDH, EC 1.1.1. 44) was identified from the homofermentative lactic acid bacterium Lactococcus lactis, by complementation of Escherichia coli mutants. The cloned gene was then expressed to high levels in E. coli and the protein purified for kinetic analysis. The enzyme had a Km for 6-phosphogluconate of 15.4+/-1.4 microM and for NADP of 1.9+/-0.2 microM at pH 7.5. Sequence comparison of the L. lactis 6-PGDH with the corresponding enzyme derived from the pathogenic protozoan Trypanosoma brucei and sheep liver revealed the substrate-binding residues to be identical in all three species, although the three coenzyme-binding pockets differed slightly. A totally conserved arginine residue (Arg-447), believed to bind the 6-phosphate of substrate, was mutated to lysine, aspartate, alanine or tryptophan. In each case enzyme activity was lost, confirming an essential role for this residue on activity. A second arginine (Arg-34), believed to be critical in binding the 2'-phosphate of cofactor NADP+, was mutated to a tyrosine residue, as found in one atypical isoform of the enzyme in Bacillus subtilis. This alteration led to decrease in affinity for NADP+ of nearly three orders of magnitude. A second 6-PGDH gene has been identified from the genome of B. subtilis. This second isoform contains an arginine (Arg-34) in this position, suggesting that B. subtilis has two 6-PGDHs with different coenzyme specificities. PMID:9931298

Tetaud, E; Hanau, S; Wells, J M; Le Page, R W; Adams, M J; Arkison, S; Barrett, M P

1999-01-01

284

Metabolic characterization of Lactococcus lactis deficient in lactate dehydrogenase using in vivo 13C-NMR.  

PubMed

The metabolism of glucose by nongrowing cells of Lactococcus lactis strain FI7851, constructed from the wild-type L. lactis strain MG1363 by disruption of the lactate dehydrogenase (ldh) gene [Gasson, M.J., Benson, K., Swindel, S. & Griffin, H. (1996) Lait 76, 33-40] was studied in a noninvasive manner by 13C-NMR. The kinetics of the build-up and consumption of the pools of intracellular intermediates mannitol 1-phosphate, fructose 1,6-bisphosphate, 3-phosphoglycerate, and phosphoenolpyruvate as well as the utilization of [1-13C]glucose and formation of products (lactate, acetate, mannitol, ethanol, acetoin, 2,3-butanediol) were monitored in vivo with a time resolution of 30 s. The metabolism of glucose by the parental wild-type strain was also examined for comparison. A clear shift from typical homolactic fermentation (parental strain) to a mixed acid fermentation (lactate dehdydrogenase deficient; LDHd strain) was observed. Furthermore, high levels of mannitol were transiently produced and metabolized once glucose was depleted. Mannitol 1-phosphate accumulated intracellularly up to 76 mM concentration. Mannitol was formed from fructose 6-phosphate by the combined action of mannitol-1-phosphate dehydrogenase and phosphatase. The results show that the formation of mannitol 1-phosphate by the LDHd strain during glucose catabolism is a consequence of impairment in NADH oxidation caused by a highly reduced LDH activity, the transient production of mannitol 1-phosphate serving as a regeneration pathway for NAD+ regeneration. Oxygen availability caused a drastic change in the pattern of intermediates and end-products, reinforcing the key-role of the fulfilment of the redox balance. The flux control coefficients for the step catalysed by mannitol-1-phosphate dehydrogenase were calculated and the implications in the design of metabolic engineering strategies are discussed. PMID:10849005

Neves, A R; Ramos, A; Shearman, C; Gasson, M J; Almeida, J S; Santos, H

2000-06-01

285

Pilus Biogenesis in Lactococcus lactis: Molecular Characterization and Role in Aggregation and Biofilm Formation  

PubMed Central

The genome of Lactococcus lactis strain IL1403 harbors a putative pilus biogenesis cluster consisting of a sortase C gene flanked by 3 LPxTG protein encoding genes (yhgD, yhgE, and yhhB), called here pil. However, pili were not detected under standard growth conditions. Over-expression of the pil operon resulted in production and display of pili on the surface of lactococci. Functional analysis of the pilus biogenesis machinery indicated that the pilus shaft is formed by oligomers of the YhgE pilin, that the pilus cap is formed by the YhgD pilin and that YhhB is the basal pilin allowing the tethering of the pilus fibers to the cell wall. Oligomerization of pilin subunits was catalyzed by sortase C while anchoring of pili to the cell wall was mediated by sortase A. Piliated L. lactis cells exhibited an auto-aggregation phenotype in liquid cultures, which was attributed to the polymerization of major pilin, YhgE. The piliated lactococci formed thicker, more aerial biofilms compared to those produced by non-piliated bacteria. This phenotype was attributed to oligomers of YhgE. This study provides the first dissection of the pilus biogenesis machinery in a non-pathogenic Gram-positive bacterium. Analysis of natural lactococci isolates from clinical and vegetal environments showed pili production under standard growth conditions. The identification of functional pili in lactococci suggests that the changes they promote in aggregation and biofilm formation may be important for the natural lifestyle as well as for applications in which these bacteria are used. PMID:23236417

Oxaran, Virginie; Ledue-Clier, Florence; Dieye, Yakhya; Herry, Jean-Marie; Pechoux, Christine; Meylheuc, Thierry; Briandet, Romain; Juillard, Vincent; Piard, Jean-Christophe

2012-01-01

286

Sulfur Amino Acid Metabolism and Its Control in Lactococcus lactis IL1403  

PubMed Central

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

Sperandio, Brice; Polard, Patrice; Ehrlich, Dusko S.; Renault, Pierre; Guédon, Eric

2005-01-01

287

1 JUNE 2006 Bact to basics  

E-print Network

... Alternative Medicine: Some Stuff Works! Natural Gas Shortage Alternative Medicine Extra: 2005: Hottest year grinds to a halt unless the electrons can be dumped. In aerobic organisms like people and plants

Lovley, Derek

288

Malonate-bound structure of the glycerophosphodiesterase from Enterobacter aerogenes (GpdQ) and characterization of the native Fe[supscript 2+] metal-ion preference  

SciTech Connect

The structure of a malonate-bound form of the glycerophosphodiesterase from Enterobacter aerogenes, GpdQ, has been refined at a resolution of 2.2 {angstrom} to a final R factor of 17.1%. The structure was originally solved to 2.9 {angstrom} resolution using SAD phases from Zn{sup 2+} metal ions introduced into the active site of the apoenzyme [Jackson et al. (2007), J. Mol. Biol. 367, 1047-1062]. However, the 2.9 {angstrom} resolution was insufficient to discern significant details of the architecture of the binuclear metal centre that constitutes the active site. Furthermore, kinetic analysis revealed that the enzyme lost a significant amount of activity in the presence of Zn{sup 2+}, suggesting that it is unlikely to be a catalytically relevant metal ion. In this communication, a higher resolution structure of GpdQ is presented in which malonate is visibly coordinated in the active site and analysis of the native metal-ion preference is presented using atomic absorption spectroscopy and anomalous scattering. Catalytic implications of the structure and its Fe{sup 2+} metal-ion preference are discussed.

Jackson, Colin J.; Hadler, Kieran S.; Carr, Paul D.; Oakley, Aaron J.; Yip, Sylvia; Schenk, Gerhard; Ollis, David L. (Queensland); (ANU)

2010-09-20

289

Malonate-bound structure of the glycerophosphodiesterase from Enterobacter aerogenes (GpdQ) and characterization of the native Fe[superscript 2+] metal-ion preference  

SciTech Connect

The structure of a malonate-bound form of the glycerophosphodiesterase from Enterobacter aerogenes, GpdQ, has been refined at a resolution of 2.2 {angstrom} to a final R factor of 17.1%. The structure was originally solved to 2.9 {angstrom} resolution using SAD phases from Zn{sup 2+} metal ions introduced into the active site of the apoenzyme [Jackson et al. (2007), J. Mol. Biol. 367, 1047-1062]. However, the 2.9 {angstrom} resolution was insufficient to discern significant details of the architecture of the binuclear metal centre that constitutes the active site. Furthermore, kinetic analysis revealed that the enzyme lost a significant amount of activity in the presence of Zn2+, suggesting that it is unlikely to be a catalytically relevant metal ion. In this communication, a higher resolution structure of GpdQ is presented in which malonate is visibly coordinated in the active site and analysis of the native metal-ion preference is presented using atomic absorption spectroscopy and anomalous scattering. Catalytic implications of the structure and its Fe{sup 2+} metal-ion preference are discussed.

Jackson, Colin J.; Hadler, Kieran S.; Carr, Paul D.; Oakley, Aaron J.; Yip, Sylvia; Schenk, Gerhard; Ollis, David L. (Queensland); (ANU)

2011-09-28

290

Analysis of a soluble (UreD:UreF:UreG)2 accessory protein complex and its interactions with Klebsiella aerogenes urease by mass spectrometry.  

PubMed

Maturation of the nickel-containing urease of Klebsiella aerogenes is facilitated by the UreD, UreF, and UreG accessory proteins along with the UreE metallo-chaperone. A fusion of the maltose binding protein and UreD (MBP-UreD) was co-isolated with UreF and UreG in a soluble complex possessing a (MBPUreD: UreF:UreG)2 quaternary structure. Within this complex a UreF:UreF interaction was identified by chemical cross-linking of the amino termini of its two UreF protomers, as shown by mass spectrometry of tryptic peptides. A preactivation complex was formed by the interaction of (MBP-UreD:UreF:UreG)2 and urease. Mass spectrometry of intact protein species revealed a pathway for synthesis of the urease pre-activation complex in which individual hetero-trimer units of the (MBP-UreD:UreF:UreG)2 complex bind to urease. Together, these data provide important new insights into the structures of protein complexes associated with urease activation. PMID:23797863

Farrugia, Mark A; Han, Linjie; Zhong, Yueyang; Boer, Jodi L; Ruotolo, Brandon T; Hausinger, Robert P

2013-09-01

291

A monoamine-regulated Klebsiella aerogenes operon containing the monoamine oxidase structural gene (maoA) and the maoC gene.  

PubMed Central

The Klebsiella aerogenes gene maoA, which is involved in the synthesis of monoamine oxidase, was induced by tyramine and the related compounds, subjected to catabolite and ammonium ion repression, and cloned. The nucleotide sequence of the region involved in monoamine oxidase synthesis was determined. Two open reading frames, the maoA gene and a hitherto unknown gene (maoC), were found. These are located between a potential promoter sequence and a transcriptional terminator sequence. A region of the Escherichia coli chromosome that was highly homologous to the Klebsiella maoA gene was found. The potential maoA gene is located at 30.9 min on the E. coli chromosome. Analysis of the amino acid sequences of the first 11 amino acids from the N terminus of the purified monoamine oxidase agrees with those deduced from the nucleotide sequence of the maoA gene. The leader peptide extends over 30 amino acids and has the characteristics of a signal sequence. Primer extension and S1 nuclease mapping of transcripts generated in vivo suggests that the tyramine-induced mRNA starts at a site 62 bases upstream from the ATG initiation codon of the maoC gene. In the putative promoter region, a high degree of similarity to the consensus sequence for the binding site of cyclic AMP receptor protein was found. Thus, the mao region is composed of two cistrons, and the mao operon is regulated by monoamine compounds, glucose, and ammonium ions. Images PMID:1556068

Sugino, H; Sasaki, M; Azakami, H; Yamashita, M; Murooka, Y

1992-01-01

292

Heterologous Production of Methionine Lyase from Brevibacterium linens in Lactococcus lactis and Formation of Volatile Sulfur Compounds  

Microsoft Academic Search

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--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.

Sean B. Hanniffy; Mark Philo; Carmen Pelaez; Michael J. Gasson; Teresa Requena; M. C. Martinez-Cuesta

2009-01-01

293

A General Method for Selection of ?-Acetolactate Decarboxylase-Deficient Lactococcus lactis Mutants To Improve Diacetyl Formation  

PubMed Central

The enzyme acetolactate decarboxylase (Ald) plays a key role in the regulation of the ?-acetolactate pool in both pyruvate catabolism and the biosynthesis of the branched-chain amino acids, isoleucine, leucine, and valine (ILV). This dual role of Ald, due to allosteric activation by leucine, was used as a strategy for the isolation of Ald-deficient mutants of Lactococcus lactis subsp. lactis biovar diacetylactis. Such mutants can be selected as leucine-resistant mutants in ILV- or IV-prototrophic strains. Most dairy lactococcus strains are auxotrophic for the three amino acids. Therefore, the plasmid pMC004 containing the ilv genes (encoding the enzymes involved in the biosynthesis of IV) of L. lactis NCDO2118 was constructed. Introduction of pMC004 into ILV-auxotrophic dairy strains resulted in an isoleucine-prototrophic phenotype. By plating the strains on a chemically defined medium supplemented with leucine but not valine and isoleucine, spontaneous leucine-resistant mutants were obtained. These mutants were screened by Western blotting with Ald-specific antibodies for the presence of Ald. Selected mutants lacking Ald were subsequently cured of pMC004. Except for a defect in the expression of Ald, the resulting strain, MC010, was identical to the wild-type strain, as shown by Southern blotting and DNA fingerprinting. The mutation resulting in the lack of Ald in MC010 occurred spontaneously, and the strain does not contain foreign DNA; thus, it can be regarded as food grade. Nevertheless, its application in dairy products depends on the regulation of genetically modified organisms. These results establish a strategy to select spontaneous Ald-deficient mutants from transformable L. lactis strains. PMID:10049884

Curic, Mirjana; Stuer-Lauridsen, Birgitte; Renault, Pierre; Nilsson, Dan

1999-01-01

294

A general method for selection of alpha-acetolactate decarboxylase-deficient Lactococcus lactis mutants to improve diacetyl formation.  

PubMed

The enzyme acetolactate decarboxylase (Ald) plays a key role in the regulation of the alpha-acetolactate pool in both pyruvate catabolism and the biosynthesis of the branched-chain amino acids, isoleucine, leucine, and valine (ILV). This dual role of Ald, due to allosteric activation by leucine, was used as a strategy for the isolation of Ald-deficient mutants of Lactococcus lactis subsp. lactis biovar diacetylactis. Such mutants can be selected as leucine-resistant mutants in ILV- or IV-prototrophic strains. Most dairy lactococcus strains are auxotrophic for the three amino acids. Therefore, the plasmid pMC004 containing the ilv genes (encoding the enzymes involved in the biosynthesis of IV) of L. lactis NCDO2118 was constructed. Introduction of pMC004 into ILV-auxotrophic dairy strains resulted in an isoleucine-prototrophic phenotype. By plating the strains on a chemically defined medium supplemented with leucine but not valine and isoleucine, spontaneous leucine-resistant mutants were obtained. These mutants were screened by Western blotting with Ald-specific antibodies for the presence of Ald. Selected mutants lacking Ald were subsequently cured of pMC004. Except for a defect in the expression of Ald, the resulting strain, MC010, was identical to the wild-type strain, as shown by Southern blotting and DNA fingerprinting. The mutation resulting in the lack of Ald in MC010 occurred spontaneously, and the strain does not contain foreign DNA; thus, it can be regarded as food grade. Nevertheless, its application in dairy products depends on the regulation of genetically modified organisms. These results establish a strategy to select spontaneous Ald-deficient mutants from transformable L. lactis strains. PMID:10049884

Curic, M; Stuer-Lauridsen, B; Renault, P; Nilsson, D

1999-03-01

295

Identification of Candidate Carrier Proteins for Surface Display on Lactococcus lactis by Theoretical and Experimental Analyses of the Surface Proteome? †  

PubMed Central

Lactococcus lactis is a lactic acid bacterium of proven safety for use in human oral applications. For this purpose, surface display of recombinant proteins is important, and new approaches for it are being sought. Analysis of the bacterial surface proteome is essential in identifying new candidate carrier proteins for surface display. We have made two different predictions of surface-associated proteins of L. lactis MG1363 by using Augur and LocateP software, which yielded 666 and 648 proteins, respectively. Surface proteins of L. lactis NZ9000, a derivative of MG1363, were identified by using a proteomics approach. The surface proteins were cleaved from intact bacteria, and the resulting peptides were identified by mass spectrometry. The latter approach yielded 80 proteins, 34 of which were not predicted by either software. Of the 80 proteins, 7 were selected for further study. These were cloned in frame with a C-terminal hexahistidine tag and overexpressed in L. lactis NZ9000 using nisin-controlled expression. Proteins of correct molecular weight carrying a hexahistidine tag were detected. Their surface localization was confirmed with flow cytometry. Basic membrane protein A (BmpA) was exposed at the highest level. To test BmpA as a candidate carrier protein, the hexahistidine tag was replaced by the B domain of staphylococcal protein A in the genetic construct. The B domain was displayed on the surface with BmpA as a carrier. The advantage of covalent BmpA binding was demonstrated. BmpA was thus shown to be a suitable candidate for a carrier protein in lactococcal surface display. PMID:21183632

Berlec, Ales; Zadravec, Petra; Jevnikar, Zala; Strukelj, Borut

2011-01-01

296

Influence of Additives on High Pressure Stability of ?-Galactosidase from Kluyveromyces Lactis and Invertase from Saccharomyces Cerevisiae  

Microsoft Academic Search

High pressure denaturation of two yeast enzymes, Kluyveromyces lactis ?-galactosidase and Saccharomyces cerevisiae invertase in aqueous solutions of salts and polyols was investigated. Polyols (more than salts) were shown to act as very effective agents against pressure denaturation for both enzymes, leading to an increase in half-lives by factors between 1–10,000. NaBr and KBr were essentially destabilizing compounds, leading to

Violaine Athès; Didier Combes

1998-01-01

297

A Plasmodium falciparum GLURP–MSP3 chimeric protein; expression in Lactococcus lactis, immunogenicity and induction of biologically active antibodies  

Microsoft Academic Search

Plasmodium falciparum malaria is a major cause of morbidity and mortality worldwide. To evaluate the efficacy of a possible vaccine antigen against P. falciparum infection, a fusion protein, derived from P. falciparum Glutamate-rich protein (GLURP) genetically coupled to P. falciparum Merozoite surface protein 3 (MSP3) was produced in Lactococcus lactis as a secreted recombinant GLURP–MSP3 fusion protein. The hybrid protein

Michael Theisen; Soe Soe; Katja Brunstedt; Frank Follmann; Lars Bredmose; Hans Israelsen; Søren M Madsen; Pierre Druilhe

2004-01-01

298

Characterization of the d-Xylulose 5-Phosphate/d-Fructose 6-Phosphate Phosphoketolase Gene (xfp) from Bifidobacterium lactis  

PubMed Central

A d-xylulose 5-phosphate/d-fructose 6-phosphate phosphoketolase (Xfp) from the probiotic Bifidobacterium lactis was purified to homogeneity. The specific activity of the purified enzyme with d-fructose 6-phosphate as a substrate is 4.28 Units per mg of enzyme. Km values for d-xylulose 5-phosphate and d-fructose 6-phosphate are 45 and 10 mM, respectively. The native enzyme has a molecular mass of 550,000 Da. The subunit size upon sodium dodecyl sulfate-polyacrylamide gel electrophoresis (90,000 Da) corresponds with the size (92,529 Da) calculated from the amino acid sequence of the isolated gene (named xfp) encoding 825 amino acids. The xfp gene was identified on the chromosome of B. lactis with the help of degenerated nucleotide probes deduced from the common N-terminal amino acid sequence of both the native and denatured enzyme. Comparison of the deduced amino acid sequence of the cloned gene with sequences in public databases revealed high homologies with hypothetical proteins (26 to 55% identity) in 20 microbial genomes. The amino acid sequence derived from the xfp gene contains typical thiamine diphosphate (ThDP) binding sites reported for other ThDP-dependent enzymes. Two truncated putative genes, pta and guaA, were localized adjacent to xfp on the B. lactis chromosome coding for a phosphotransacetylase and a guanosine monophosphate synthetase homologous to products of genes in Mycobacterium tuberculosis. However, xfp is transcribed in B. lactis as a monocistronic operon. It is the first reported and sequenced gene of a phosphoketolase. PMID:11292814

Meile, Leo; Rohr, Lukas M.; Geissmann, Thomas A.; Herensperger, Monique; Teuber, Michael

2001-01-01

299

Kinetics and Consequences of Binding of Nona and Dodecapeptides to the Oligopeptide Binding Protein (OppA) of Lactococcus lactis  

Microsoft Academic Search

The oligopeptide transport system (Opp) of Lactococcus lactis belongs to the class of binding protein-dependent ABC-transporters. This system has the unique capacity to mediate the uptake of peptides from 4 up to at least 18 residues. Kinetic analysis of peptide binding to the binding protein, OppA*, revealed a relationship between the peptide dissociation constants and the length of the ligand.

Bert Poolman; Wil N. Konings; Antonia Picon; Frank C. Lanfermeijer

1999-01-01

300

Use of PCR-Based Methods for Rapid Differentiation of Lactobacillus delbrueckii subsp. bulgaricus and L. delbrueckii subsp. lactis  

Microsoft Academic Search

Two PCR-based methods, specific PCR and randomly amplified polymorphic DNA PCR (RAPD-PCR), were used for rapid and reliable differentiation of Lactobacillus delbrueckii subsp. bulgaricus and L. delbrueckii subsp. lactis. PCR with a single combination of primers which targeted the proline iminopeptidase (pepIP) gene of L. delbrueckii subsp. bulgaricus allowed amplification of genomic fragments specific for the two subspecies when either

SANDRA TORRIANI; GIACOMO ZAPPAROLI; FRANCO DELLAGLIO

1999-01-01

301

Genetic Manipulation of Lactococcus lactis by Using Targeted Group II Introns: Generation of Stable Insertions without Selection  

Microsoft Academic Search

Despite their commercial importance, there are relatively few facile methods for genomic manipulation of the lactic acid bacteria. Here, the lactococcal group II intron, Ll.ltrB, was targeted to insert efficiently into genes encoding malate decarboxylase (mleS) and tetracycline resistance (tetM) within the Lactococcus lactis genome. Integrants were readily identified and maintained in the absence of a selectable marker. Since splicing

Courtney L. Frazier; Joseph San Filippo; Alan M. Lambowitz; David A. Mills

2003-01-01

302

Unraveling the Role of Surface Mucus-Binding Protein and Pili in Muco-Adhesion of Lactococcus lactis  

PubMed Central

Adhesion of bacteria to mucus may favor their persistence within the gut and their beneficial effects to the host. Interactions between pig gastric mucin (PGM) and a natural isolate of Lactococcus lactis (TIL448) were measured at the single-cell scale and under static conditions, using atomic force microscopy (AFM). In parallel, these interactions were monitored at the bacterial population level and under shear flow. AFM experiments with a L. lactis cell-probe and a PGM-coated surface revealed a high proportion of specific adhesive events (60%) and a low level of non-adhesive ones (2%). The strain muco-adhesive properties were confirmed by the weak detachment of bacteria from the PGM-coated surface under shear flow. In AFM, rupture events were detected at short (100?200 nm) and long distances (up to 600?800 nm). AFM measurements on pili and mucus-binding protein defective mutants demonstrated the comparable role played by these two surface proteinaceous components in adhesion to PGM under static conditions. Under shear flow, a more important contribution of the mucus-binding protein than the pili one was observed. Both methods differ by the way of probing the adhesion force, i.e. negative force contact vs. sedimentation and normal-to-substratum retraction vs. tangential detachment conditions, using AFM and flow chamber, respectively. AFM blocking assays with free PGM or O-glycan fractions purified from PGM demonstrated that neutral oligosaccharides played a major role in adhesion of L. lactis TIL448 to PGM. This study dissects L. lactis muco-adhesive phenotype, in relation with the nature of the bacterial surface determinants. PMID:24260308

Duviau, Marie-Pierre; Meyrand, Mickael; Guerardel, Yann; Castelain, Mickael; Loubiere, Pascal; Chapot-Chartier, Marie-Pierre; Dague, Etienne; Mercier-Bonin, Muriel

2013-01-01

303

Ability of Lactococcus lactis to export viral capsid antigens: a crucial step for development of live vaccines  

Microsoft Academic Search

Thefood grade bacterium Lactococcus lactis is a potential vehicle for protein delivery in the gastrointestinal tract. As a model, we constructed lactococcal strains producing antigens of infectious bursal disease virus (IBDV). IBDV infects chickens and causes depletion of B-lymphoid cells in the bursa of Fabricius and subsequent immunosuppression, morbidity, or acute mortality. The two major IBDV antigens, i.e., VP2 and

Yakhya Dieye; Arjan J. W. Hoekman; Florence Clier; Vincent Juillard; Hein J. Boot; Jean-Christophe Piard

2003-01-01

304

A distinct physiological state of Lactococcus lactis cells that confers survival against a direct and prolonged exposure to severe stresses  

Microsoft Academic Search

When exponential phase cultures of Lactococcus lactis were directly exposed to severe stresses (acid, bile salt, heat, and hydrogen peroxide) for a prolonged period, most of the cells were quickly killed, however, a small number of the cells, approximately 0.01% of the population, was found to survive. How these ‘survivor’ cells might have survived the stresses, when other supposedly-the-same cells

Woojin S Kim; Ji Hyeon Park; Jade E Tandianus; Jun Ren; Ping Su; Noel W Dunn

2002-01-01

305

Glutamate-induced metabolic changes in Lactococcus lactis NCDO 2118 during GABA production: combined transcriptomic and proteomic analysis  

Microsoft Academic Search

GABA is a molecule of increasing nutraceutical interest due to its modulatory activity on the central nervous system and smooth\\u000a muscle relaxation. Potentially probiotic bacteria can produce it by glutamate decarboxylation, but nothing is known about\\u000a the physiological modifications occurring at the microbial level during GABA production. In the present investigation, a GABA-producing\\u000a Lactococcus lactis strain grown in a medium

Roberto Mazzoli; Enrica Pessione; Magali Dufour; Valérie Laroute; Maria Gabriella Giuffrida; Carlo Giunta; Muriel Cocaign-Bousquet; Pascal Loubière

2010-01-01

306

Comparative analysis of Lactococcus lactis bacteriocins and preliminary characterisation of a new proteinase K resistant lactococcin member  

Microsoft Academic Search

Detection of lactic acid bacteria (LAB) bacteriocins producers is of great significance for food industry to establish starter\\u000a bacterial association and to improve food safety. Eighty oneLactococcus lactis strains, isolated from traditional Tunisian dairy products, were screened for their antibacterial activity. Bacteriocin production\\u000a in the supernatant was demonstrated for twelve strains by the well diffusion assay, protease susceptibility and by

Hadda Ouzari; Afef Najjari; Houda Amairi; Maher Gtari; Abdenaceur Hassen; Abdellatif Boudabous

2008-01-01

307

iOD907, the first genome-scale metabolic model for the milk yeast Kluyveromyces lactis.  

PubMed

We describe here the first genome-scale metabolic model of Kluyveromyces lactis, iOD907. It is partially compartmentalized (four compartments), composed of 1867 reactions and 1476 metabolites. The iOD907 model performed well when comparing the positive growth of K. lactis to Biolog experiments and to an online catalogue of strains that provides information on carbon sources in which K. lactis is able to grow. Chemostat experiments were used to adjust non-growth-associated energy requirements, and the model proved accurate when predicting the biomass, oxygen and carbon dioxide yields. When compared to published experiments, in silico knockouts accurately predicted in vivo phenotypes. The iOD907 genome-scale metabolic model complies with the MIRIAM (minimum information required for the annotation of biochemical models) standards for the annotation of enzymes, transporters, metabolites and reactions. Moreover, it contains direct links to Kyoto encyclopedia of genes and genomes (KEGG; for enzymes, metabolites and reactions) and to the Transporters Classification Database (TCDB) for transporters, allowing easy comparisons to other models. Furthermore, this model is provided in the well-established systems biology markup language (SBML) format, which means that it can be used in most metabolic engineering platforms, such as OptFlux or Cobra. The model is able to predict the behavior of K. lactis under different environmental conditions and genetic perturbations. Furthermore, by performing simulations and optimizations, it can be important in the design of minimal media and will allow insights on the milk yeast's metabolism, as well as identifying metabolic engineering targets for improving the production of products of interest. PMID:24777859

Dias, Oscar; Pereira, Rui; Gombert, Andreas K; Ferreira, Eugénio C; Rocha, Isabel

2014-06-01

308

A tool kit for molecular genetics of Kluyveromyces lactis comprising a congenic strain series and a set of versatile vectors.  

PubMed

A set of different marker deletions starting with a ura3 derivative of the Kluyveromyces lactis type strain CBS2359 was constructed. After a first cross to obtain a strain with the opposite mating type that also carried a leu2 allele, continuous back-crosses were used to obtain a congenic strain series with different marker combinations, including deletions in KlHIS3, KlADE2 and KlLAC4. Enzymes involved in carbohydrate metabolism were shown to behave very similarly to the original type strain and other K. lactis strains investigated previously. Moreover, a vector series of Saccharomyces cerevisiae genes flanked by loxP sites was constructed to be used as heterologous deletion cassettes in K. lactis, together with two plasmids for expression of Cre-recombinase for marker regeneration. To increase the frequency of homologous recombination, the Klku80 deletion was also introduced into the congenic strain series. A PCR-based method for determination of mating type is provided. PMID:20522115

Heinisch, Jürgen J; Buchwald, Ulf; Gottschlich, André; Heppeler, Nele; Rodicio, Rosaura

2010-05-01

309

Efficient production of secreted staphylococcal antigens in a non-lysing and proteolytically reduced Lactococcus lactis strain.  

PubMed

Cell surface-exposed and secreted proteins are attractive targets for vaccination against pathogenic gram-positive bacteria. To obtain sufficient amounts of such antigens, efficient protein production platforms are needed. In this study, a pipeline for the production and purification of surface-exposed and secreted antigens of the gram-positive bacterial pathogen Staphylococcus aureus is presented. Cytoplasmic or extracellular production of S. aureus antigens was achieved using the Lactococcus lactis strain PA1001, which lacks the major extracellular protease HtrA and the autolysin AcmA to minimize proteolysis and cell lysis, respectively. For most tested S. aureus antigens, secretory production directed by the signal peptide of the major secreted protein Usp45 of L. lactis resulted in higher yields than intracellular production without a signal peptide. Additionally, secretory production of His-tagged antigens allowed their facile one-step purification from the growth medium by metal affinity chromatography. For three of the purified antigens, biological activity was confirmed through enzyme activity assays. We, furthermore, show that the present pipeline can be used to produce staphylococcal antigens with an N-terminal AVI-tag for site-specific labeling with biotin or a C-terminal cell wall-binding domain for cell surface display. We conclude that our L. lactis-based pipeline allows the efficient production of S. aureus antigens and their subsequent purification in one step. PMID:25176446

Neef, Jolanda; Koedijk, Danny G A M; Bosma, Tjibbe; van Dijl, Jan Maarten; Buist, Girbe

2014-12-01

310

Genome-scale genotype-phenotype matching of two Lactococcus lactis isolates from plants identifies mechanisms of adaptation to the plant niche.  

PubMed

Lactococcus lactis is a primary constituent of many starter cultures used for the manufacturing of fermented dairy products, but the species also occurs in various nondairy niches such as (fermented) plant material. Three genome sequences of L. lactis dairy strains (IL-1403, SK11, and MG1363) are publicly available. An extensive molecular and phenotypic diversity analysis was now performed on two L. lactis plant isolates. Diagnostic sequencing of their genomes resulted in over 2.5 Mb of sequence for each strain. A high synteny was found with the genome of L. lactis IL-1403, which was used as a template for contig mapping and locating deletions and insertions in the plant L. lactis genomes. Numerous genes were identified that do not have homologs in the published genome sequences of dairy L. lactis strains. Adaptation to growth on substrates derived from plant cell walls is evident from the presence of gene sets for the degradation of complex plant polymers such as xylan, arabinan, glucans, and fructans but also for the uptake and conversion of typical plant cell wall degradation products such as alpha-galactosides, beta-glucosides, arabinose, xylose, galacturonate, glucuronate, and gluconate. Further niche-specific differences are found in genes for defense (nisin biosynthesis), stress response (nonribosomal peptide synthesis and various transporters), and exopolysaccharide biosynthesis, as well as the expected differences in various mobile elements such as prophages, plasmids, restriction-modification systems, and insertion sequence elements. Many of these genes were identified for the first time in Lactococcus lactis. In most cases good correspondence was found with the phenotypic characteristics of these two strains. PMID:18039825

Siezen, Roland J; Starrenburg, Marjo J C; Boekhorst, Jos; Renckens, Bernadet; Molenaar, Douwe; van Hylckama Vlieg, Johan E T

2008-01-01

311

Oral immunization with recombinant Lactococcus lactis delivering a multi-epitope antigen CTB-UE attenuates Helicobacter pylori infection in mice.  

PubMed

Urease is an essential virulence factor and colonization factor for Helicobacter pylori (H. pylori) and is considered as an excellent vaccine candidate antigen. However, conventional technologies for preparing an injectable vaccine require purification of the antigenic protein and preparation of an adjuvant. Lactococcus lactis NZ9000 (L. lactis) could serve as an antigen-delivering vehicle for the development of edible vaccine. In previous study, we constructed a multi-epitope vaccine, designated CTB-UE, which is composed of the mucosal adjuvant cholera toxin B subunit (CTB), three Th cell epitopes and two B-cell epitopes from urease subunits. To develop a novel type of oral vaccine against H. pylori, genetically modified L. lactis strains were established to secrete this epitope vaccine extracellularly in this study. Oral prophylactic immunization with recombinant L. lactis significantly elicited humoral anti-urease antibody responses (P < 0.001) and reduced the gastric colonization of H. pylori from 7.14 ± 0.95 to 4.68 ± 0.98 log10 CFU g(-1) stomach. This L. lactis oral vaccine offers a promising vaccine candidate for the control of H. pylori infection. PMID:24687988

Li, Xinyang; Xing, Yingying; Guo, Le; Lv, Xiaobo; Song, Hui; Xi, Tao

2014-10-01

312

Bacteriophage Resistance of a ?thyA Mutant of Lactococcus lactis Blocked in DNA Replication  

PubMed Central

The thyA gene, which encodes thymidylate synthase (TS), of Lactococcus lactis CHCC373 was sequenced, including the upstream and downstream regions. We then deleted part of thyA by gene replacement. The resulting strain, MBP71 ?thyA, was devoid of TS activity, and in media without thymidine, such as milk, there was no detectable dTTP pool in the cells. Hence, DNA replication was abolished, and acidification by MBP71 was completely unaffected by the presence of nine different phages tested at a multiplicity of infection (MOI) of 0.1. Nonreplicating MBP71 must be inoculated at a higher level than CHCC373 to achieve a certain pH within a specified time. For a pH of 5.2 to be reached in 6 h, the inoculation level of MBP71 must be 17-fold higher than for CHCC373. However, by adding a limiting amount of thymidine this could be lowered to just 5-fold the normal amount, while acidification was unaffected with MBP71 up to an MOI of 0.01. It was found that nonreplicating MBP71 produced largely the same products as CHCC373, though the acetaldehyde production of the former was higher. PMID:12039762

Pedersen, Martin B.; Jensen, Peter R.; Janzen, Thomas; Nilsson, Dan

2002-01-01

313

Production of xylitol from D-xylose by recombinant Lactococcus lactis.  

PubMed

The D-xylose reductase from Pichia stipitis CBS 5773 and the xylose transporter from Lactobacillus brevis ATCC 8287 were expressed in active form in Lactococcus lactis NZ9800. Xylitol production was investigated using non-growing recombinant cells in high cell-density under microaerobic conditions in the presence of xylose and glucose. Besides xylose, the recombinant strain with xylose reductase activity reduced l-arabinose and D-ribose in significant extent to the corresponding pentitols. The ratio of xylitol produced per glucose consumed was almost 10-fold higher under glucose limitation than the ratio in the presence of excess initial glucose. The co-expression of the xylose transporter with the xylose reductase did not increase the efficiency of xylitol production appreciably when compared to the strain in which only the xylose reductase gene was expressed. A fed-batch experiment with high initial xylose concentration (160 gl(-1)) under glucose limitation was carried out using the strain co-expressing xylose reductase and xylose transporter genes. The xylitol yield from xylose was 1.0 mol mol(-1) and the ratio of xylitol produced per glucose consumed was 2.5 mol mol(-1). The volumetric productivity was 2.72 gl(-1)h(-1) at 20 h. Of the xylose initially present, 34% was consumed. Analysis of the fermentation metabolites revealed a shift from homolactic to mixed acid fermentation at early stages of the experiment. PMID:15916828

Nyyssölä, Antti; Pihlajaniemi, Anne; Palva, Airi; von Weymarn, Niklas; Leisola, Matti

2005-07-21

314

Bifidobacterium longum L-arabinose isomerase--overexpression in Lactococcus lactis, purification, and characterization.  

PubMed

Bifidobacterium longum NRRL B-41409 L-arabinose isomerase (L-AI) was cloned and overexpressed in Lactococcus lactis using a phosphate-depletion-inducible expression system. The purified B. longum L-AI was characterized using D-galactose and L-arabinose as the substrates. The enzyme was active and stable at acidic pH with an optimum at pH 6.0-6.5. The enzyme showed the highest activity at 55 °C during a 20-min incubation at pH 6.5. The K(m) value was 120 mM for L-arabinose and 590 mM for D-galactose. The V(max) was 42 U mg(-1) with L-arabinose and 7.7 U mg(-1) with D-galactose as the substrates. The enzyme had very low requirement for metal ions for catalytic activity, but it was stabilized by divalent metal ions (Mg(2+), Mn(2+)). The enzyme bound the metal ions so tightly that they could not be fully removed from the active site by EDTA treatment. Using purified B. longum L-AI as the catalyst at 35 °C, equilibrium yields of 36 % D-tagatose and 11 % L-ribulose with 1.67 M D-galactose and L-arabinose, respectively, as the substrates were reached. PMID:22763951

Salonen, Noora; Nyyssölä, Antti; Salonen, Kalle; Turunen, Ossi

2012-09-01

315

Expression of heparan sulfate sulfotransferases in Kluyveromyces lactis and preparation of 3?-phosphoadenosine-5?-phosphosulfate  

PubMed Central

Heparan sulfate (HS) belongs to a major class of glycans that perform central physiological functions. Heparin is a specialized form of HS and is a clinically used anticoagulant drug. Heparin is a natural product isolated from pig intestine. There is a strong demand to replace natural heparin with a synthetic counterpart. Although a chemoenzymatic approach has been employed to prepare synthetic heparin, the scale of the synthesis is limited by the availability of sulfotransferases and the cofactor, 3?-phosphoadenosine-5?-phosphosulfate (PAPS). Here, we present a novel method to produce secreted forms of sulfotransferases in the yeast cells, Kluyveromyces lactis. Five sulfotransferases including N-sulfotransferase, 2-O-sulfotransferase, 3-O-sulfotransferase 1 and 6-O-sulfotransferases 1 and 3 were expressed using this method. Unlike bacterial-expressed sulfotransferases, the yeast proteins can be directly used to modify polysaccharides without laborious purification. The yeast-expressed sulfotransferases also tend to have higher specific activity and thermostability. Furthermore, we demonstrated the possibility for the gram-scale synthesis of PAPS from adenosine 5'-triphosphate at only 1/5000th of the price purchased from a commercial source. Our results pave the way to conduct the enzymatic synthesis of heparin in large quantities. PMID:21224284

Zhou, Xianxuan; Chandarajoti, Kasemsiri; Pham, Truong Quang; Liu, Renpeng; Liu, Jian

2011-01-01

316

Transcriptome and Proteome Exploration to Model Translation Efficiency and Protein Stability in Lactococcus lactis  

PubMed Central

This genome-scale study analysed the various parameters influencing protein levels in cells. To achieve this goal, the model bacterium Lactococcus lactis was grown at steady state in continuous cultures at different growth rates, and proteomic and transcriptomic data were thoroughly compared. Ratios of mRNA to protein were highly variable among proteins but also, for a given gene, between the different growth conditions. The modeling of cellular processes combined with a data fitting modeling approach allowed both translation efficiencies and degradation rates to be estimated for each protein in each growth condition. Estimated translational efficiencies and degradation rates strongly differed between proteins and were tested for their biological significance through statistical correlations with relevant parameters such as codon or amino acid bias. These efficiencies and degradation rates were not constant in all growth conditions and were inversely proportional to the growth rate, indicating a more efficient translation at low growth rate but an antagonistic higher rate of protein degradation. Estimated protein median half-lives ranged from 23 to 224 min, underlying the importance of protein degradation notably at low growth rates. The regulation of intracellular protein level was analysed through regulatory coefficient calculations, revealing a complex control depending on protein and growth conditions. The modeling approach enabled translational efficiencies and protein degradation rates to be estimated, two biological parameters extremely difficult to determine experimentally and generally lacking in bacteria. This method is generic and can now be extended to other environments and/or other micro-organisms. PMID:20019804

Dressaire, Clementine; Gitton, Christophe; Loubiere, Pascal; Monnet, Veronique; Queinnec, Isabelle; Cocaign-Bousquet, Muriel

2009-01-01

317

Catabolism of mannitol in Lactococcus lactis MG1363 and a mutant defective in lactate dehydrogenase.  

PubMed

Mannitol metabolism in Lactococcus lactis MG1363 and in a derivative strain deficient in lactate dehydrogenase (LDH(d)) was characterized. Both strains had the ability to grow on mannitol as an energy source, although this polyol was a poorer substrate for growth than glucose. When compared to glucose, the metabolism of mannitol caused an NADH burden due to formation of an additional NADH molecule at the reaction catalysed by mannitol-1-phosphate dehydrogenase (Mtl1PDH). This resulted in a prominent accumulation of mannitol 1-phosphate (Mtl1P) both in growing and resting cells, suggesting the existence of a severe bottleneck at Mtl1PDH. Growth on mannitol induced the activity of Mtl1PDH in both the LDH(d) and MG1363 strains. The lower accumulation of Mtl1P in mannitol-grown cells when compared to glucose-grown LDH(d) cells, as monitored by in vivo (13)C-NMR, reflects this induction. A clear shift towards the production of ethanol was observed on mannitol, indicating pressure to regenerate NAD(+) when this substrate was used. A strategy to obtain a mannitol-overproducing strain is proposed. PMID:12427938

Neves, Ana Rute; Ramos, Ana; Shearman, Claire; Gasson, Michael J; Santos, Helena

2002-11-01

318

Structure–activity relationships in Kluyveromyces lactis ?-toxin, a eukaryal tRNA anticodon nuclease  

PubMed Central

tRNA anticodon damage inflicted by secreted ribotoxins such as Kluyveromyces lactis ?-toxin and bacterial colicins underlies a rudimentary innate immune system that distinguishes self from nonself species. The intracellular expression of ?-toxin (a 232-amino acid polypeptide) arrests the growth of Saccharomyces cerevisiae by incising a single RNA phosphodiester 3? of the modified wobble base of tRNAGlu. Fungal ?-toxin bears no primary structure similarity to any known nuclease and has no plausible homologs in the protein database. To gain insight to ?-toxin's mechanism, we tested the effects of alanine mutations at 62 basic, acidic, and polar amino acids on ribotoxin activity in vivo. We thereby identified 22 essential residues, including 10 lysines, seven arginines, three glutamates, one cysteine, and one histidine (His209, the only histidine present in ?-toxin). Structure–activity relations were gleaned from the effects of 44 conservative substitutions. Recombinant tag-free ?-toxin, a monomeric protein, incised an oligonucleotide corresponding to the anticodon stem–loop of tRNAGlu at a single phosphodiester 3? of the wobble uridine. The anticodon nuclease was metal independent. RNA cleavage was abolished by ribose 2?-H and 2?-F modifications of the wobble uridine. Mutating His209 to alanine, glutamine, or asparagine abolished nuclease activity. We propose that ?-toxin catalyzes an RNase A-like transesterification reaction that relies on His209 and a second nonhistidine side chain as general acid–base catalysts. PMID:19383764

Keppetipola, Niroshika; Jain, Ruchi; Meineke, Birthe; Diver, Melinda; Shuman, Stewart

2009-01-01

319

A Genome-Scale Integration and Analysis of Lactococcus lactis Translation Data  

PubMed Central

Protein synthesis is a template polymerization process composed by three main steps: initiation, elongation, and termination. During translation, ribosomes are engaged into polysomes whose size is used for the quantitative characterization of translatome. However, simultaneous transcription and translation in the bacterial cytosol complicates the analysis of translatome data. We established a procedure for robust estimation of the ribosomal density in hundreds of genes from Lactococcus lactis polysome size measurements. We used a mechanistic model of translation to integrate the information about the ribosomal density and for the first time we estimated the protein synthesis rate for each gene and identified the rate limiting steps. Contrary to conventional considerations, we find significant number of genes to be elongation limited. This number increases during stress conditions compared to optimal growth and proteins synthesized at maximum rate are predominantly elongation limited. Consistent with bacterial physiology, we found proteins with similar rate and control characteristics belonging to the same functional categories. Under stress conditions, we found that synthesis rate of regulatory proteins is becoming comparable to proteins favored under optimal growth. These findings suggest that the coupling of metabolic states and protein synthesis is more important than previously thought. PMID:24130467

Racle, Julien; Picard, Flora; Girbal, Laurence; Cocaign-Bousquet, Muriel; Hatzimanikatis, Vassily

2013-01-01

320

Metabolic Behavior of Lactococcus lactis MG1363 in Microaerobic Continuous Cultivation at a Low Dilution Rate  

PubMed Central

Minute amounts of oxygen were supplied to a continuous cultivation of Lactococcus lactis subsp. cremoris MG1363 grown on a defined glucose-limited medium at a dilution rate of 0.1 h?1. More than 80% of the carbon supplied with glucose ended up in fermentation products other than lactate. Addition of even minute amounts of oxygen increased the yield of biomass on glucose by more than 10% compared to that obtained under anaerobic conditions and had a dramatic impact on catabolic enzyme activities and hence on the distribution of carbon at the pyruvate branch point. Increasing aeration caused carbon dioxide and acetate to replace formate and ethanol as catabolic end products while hardly affecting the production of either acetoin or lactate. The negative impact of oxygen on the synthesis of pyruvate formate lyase was confirmed. Moreover, oxygen was shown to down regulate the protein level of alcohol dehydrogenase while increasing the enzyme activity levels of the pyruvate dehydrogenase complex, ?-acetolactate synthase, and the NADH oxidases. Lactate dehydrogenase and glyceraldehyde dehydrogenase enzyme activity levels were unaffected by aeration. PMID:11375180

Jensen, Niels Bang Siemsen; Melchiorsen, Claus Rix; Jokumsen, Kirsten Vaever; Villadsen, John

2001-01-01

321

Molecular and Functional Analyses of the metC Gene of Lactococcus lactis, Encoding Cystathionine ?-Lyase  

PubMed Central

The enzymatic degradation of amino acids in cheese is believed to generate aroma compounds and therefore to be essential for flavor development. Cystathionine ?-lyase (CBL) can convert cystathionine to homocysteine but is also able to catalyze an ?,? elimination. With methionine as a substrate, it produces volatile sulfur compounds which are important for flavor formation in Gouda cheese. The metC gene, which encodes CBL, was cloned from the Lactococcus lactis model strain MG1363 and from strain B78, isolated from a cheese starter culture and known to have a high capacity to produce volatile compounds. The metC gene was found to be cotranscribed with a downstream cysK gene, which encodes a putative cysteine synthase. The MetC proteins of both strains were overproduced in strain MG1363 with the NICE (nisin-controlled expression) system, resulting in a >25-fold increase in cystathionine lyase activity. A disruption of the metC gene was achieved in strain MG1363. Determination of enzymatic activities in the overproducing and knockout strains revealed that MetC is essential for the degradation of cystathionine but that at least one lyase other than CBL contributes to methionine degradation via ?,? elimination to form volatile aroma compounds. PMID:10618201

Fernandez, Maria; van Doesburg, Wim; Rutten, Ger A. M.; Marugg, Joey D.; Alting, Arno C.; van Kranenburg, Richard; Kuipers, Oscar P.

2000-01-01

322

Multi-omics approach to study the growth efficiency and amino acid metabolism in Lactococcus lactis at various specific growth rates  

PubMed Central

Background Lactococcus lactis is recognised as a safe (GRAS) microorganism and has hence gained interest in numerous biotechnological approaches. As it is fastidious for several amino acids, optimization of processes which involve this organism requires a thorough understanding of its metabolic regulations during multisubstrate growth. Results Using glucose limited continuous cultivations, specific growth rate dependent metabolism of L. lactis including utilization of amino acids was studied based on extracellular metabolome, global transcriptome and proteome analysis. A new growth medium was designed with reduced amino acid concentrations to increase precision of measurements of consumption of amino acids. Consumption patterns were calculated for all 20 amino acids and measured carbon balance showed good fit of the data at all growth rates studied. It was observed that metabolism of L. lactis became more efficient with rising specific growth rate in the range 0.10 - 0.60 h-1, indicated by 30% increase in biomass yield based on glucose consumption, 50% increase in efficiency of nitrogen use for biomass synthesis, and 40% reduction in energy spilling. The latter was realized by decrease in the overall product formation and higher efficiency of incorporation of amino acids into biomass. L. lactis global transcriptome and proteome profiles showed good correlation supporting the general idea of transcription level control of bacterial metabolism, but the data indicated that substrate transport systems together with lower part of glycolysis in L. lactis were presumably under allosteric control. Conclusions The current study demonstrates advantages of the usage of strictly controlled continuous cultivation methods combined with multi-omics approach for quantitative understanding of amino acid and energy metabolism of L. lactis which is a valuable new knowledge for development of balanced growth media, gene manipulations for desired product formation etc. Moreover, collected dataset is an excellent input for developing metabolic models. PMID:21349178

2011-01-01

323

Genome-scale metabolic model for Lactococcus lactis MG1363 and its application to the analysis of flavor formation.  

PubMed

Lactococcus lactis subsp. cremoris MG1363 is a paradigm strain for lactococci used in industrial dairy fermentations. However, despite of its importance for process development, no genome-scale metabolic model has been reported thus far. Moreover, current models for other lactococci only focus on growth and sugar degradation. A metabolic model that includes nitrogen metabolism and flavor-forming pathways is instrumental for the understanding and designing new industrial applications of these lactic acid bacteria. A genome-scale, constraint-based model of the metabolism and transport in L. lactis MG1363, accounting for 518 genes, 754 reactions, and 650 metabolites, was developed and experimentally validated. Fifty-nine reactions are directly or indirectly involved in flavor formation. Flux Balance Analysis and Flux Variability Analysis were used to investigate flux distributions within the whole metabolic network. Anaerobic carbon-limited continuous cultures were used for estimating the energetic parameters. A thorough model-driven analysis showing a highly flexible nitrogen metabolism, e.g., branched-chain amino acid catabolism which coupled with the redox balance, is pivotal for the prediction of the formation of different flavor compounds. Furthermore, the model predicted the formation of volatile sulfur compounds as a result of the fermentation. These products were subsequently identified in the experimental fermentations carried out. Thus, the genome-scale metabolic model couples the carbon and nitrogen metabolism in L. lactis MG1363 with complete known catabolic pathways leading to flavor formation. The model provided valuable insights into the metabolic networks underlying flavor formation and has the potential to contribute to new developments in dairy industries and cheese-flavor research. PMID:23974365

Flahaut, Nicolas A L; Wiersma, Anne; van de Bunt, Bert; Martens, Dirk E; Schaap, Peter J; Sijtsma, Lolke; Dos Santos, Vitor A Martins; de Vos, Willem M

2013-10-01

324

Kinetics of Lactococcus lactis growth and metabolite formation under aerobic and anaerobic conditions in the presence or absence of hemin.  

PubMed

The study of batch kinetics of Lactococcus lactis cell growth and product formation reveals three distinct metabolic behaviors depending upon the availability of oxygen to the culture and the presence of hemin in the medium. These three cultivation modes, anerobic homolactic fermentation, aerobic heterolactic fermentation, and hemin-stimulated respiration have been studied at pH 6.0 and 30 degrees C with a medium containing a high concentration of glucose (60 g/L). A maximum cell density of 5.78 g/L was obtained in the batch culture under hemin-stimulated respiration conditions, about three times as much as that achieved with anerobic homolactic fermentation (1.87 g/L) and aerobic heterolactic fermentation (1.80 g/L). The maximum specific growth rate was 0.60/h in hemin-stimulated respiration, slightly higher than that achieved in homolactic fermentation (0.56/h) and substantially higher than that in heterolactic fermentation (0.40/h). Alteration of metabolism caused by the supplementation of oxygen and hemin is evidenced by changes in both cell growth kinetics and metabolite formation kinetics, which are characterized by a unique pseudo-diauxic growth of L. lactis. We hypothesise that Lactococcus lactis generates bioenergy (ATP) through simultaneous lactate formation and hemin-stimulated respiration in the primary exponential phase, when glucose is abundant, and utilizes lactate for cell growth and cell maintenance in the stationary phase, after glucose is exhausted. We also examined the applicability of a modified logistic model and the Luedeking-Piret model for cell growth kinetics and metabolite formation kinetics, respectively. PMID:16807924

Lan, Christopher Q; Oddone, Gian; Mills, David A; Block, David E

2006-12-20

325

NAD-dependent lactate dehydrogenase catalyses the first step in respiratory utilization of lactate by Lactococcus lactis?  

PubMed Central

Lactococcus lactis can undergo respiration when hemin is added to an aerobic culture. The most distinctive feature of lactococcal respiration is that lactate could be consumed in the stationary phase concomitantly with the rapid accumulation of diacetyl and acetoin. However, the enzyme responsible for lactate utilization in this process has not yet been identified. As genes for fermentative NAD-dependent l-lactate dehydrogenase (l-nLDH) and potential electron transport chain (ETC)-related NAD-independent l-LDH (l-iLDH) exist in L. lactis, the activities of these enzymes were measured in this study using crude cell extracts prepared from respiratory and fermentation cultures. Further studies were conducted with purified preparations of recombinant LDH homologous proteins. The results showed that l-iLDH activity was hardly detected in both crude cell extracts and purified l-iLDH homologous protein while l-nLDH activity was very significant. This suggested that l-iLDHs were inactive in lactate utilization. The results of kinetic analyses and the effects of activator, inhibitor, substrate and product concentrations on the reaction equilibrium showed that l-nLDH was much more prone to catalyze the pyruvate reduction reaction but could reverse its role provided that the concentrations of NADH and pyruvate were extremely low while NAD and lactate were abundant. Metabolite analysis in respiratory culture revealed that the cellular status in the stationary phase was beneficial for l-nLDH to catalyze lactate oxidation. The factors accounting for the respiration- and stationary phase-dependent lactate utilization in L. lactis are discussed here. PMID:24251099

Zhao, Rui; Zheng, Sui; Duan, Cuicui; Liu, Fei; Yang, Lijie; Huo, Guicheng

2013-01-01

326

Production of human lysozyme in biofilm reactor and optimization of growth parameters of Kluyveromyces lactis K7.  

PubMed

Lysozyme (1,4-?-N-acetylmuramidase) is a lytic enzyme, which degrades the bacterial cell wall. Lysozyme has been of interest in medicine, cosmetics, and food industries because of its anti-bactericidal effect. Kluyveromyces lactis K7 is a genetically modified organism that expresses human lysozyme. There is a need to improve the human lysozyme production by K. lactis K7 to make the human lysozyme more affordable. Biofilm reactor provides high biomass by including a solid support, which microorganisms grow around and within. Therefore, the aim of this study was to produce the human lysozyme in biofilm reactor and optimize the growth conditions of K. lactis K7 for the human lysozyme production in biofilm reactor with plastic composite support (PCS). The PCS, which includes polypropylene, soybean hull, soybean flour, bovine albumin, and salts, was selected based on biofilm formation on PCS (CFU/g), human lysozyme production (U/ml), and absorption of lysozyme inside the support. To find the optimum combination of growth parameters, a three-factor Box-Behnken design of response surface method was used. The results suggested that the optimum conditions for biomass and lysozyme productions were different (27 °C, pH 6, 1.33 vvm for biomass production; 25 °C, pH 4, no aeration for lysozyme production). Then, different pH and aeration shift strategies were tested to increase the biomass at the first step and then secrete the lysozyme after the shift. As a result, the lysozyme production amount (141 U/ml) at 25 °C without pH and aeration control was significantly higher than the lysozyme amount at evaluated pH and aeration shift conditions (p?

Ercan, Duygu; Demirci, Ali

2013-07-01

327

Dose-response effect of Bifidobacterium lactis HN019 on whole gut transit time and functional gastrointestinal symptoms in adults  

PubMed Central

Objective. To assess the impact of Bifidobacterium lactis HN019 supplementation on whole gut transit time (WGTT) and frequency of functional gastrointestinal (GI) symptoms in adults. Material and methods. We randomized 100 subjects (mean age: 44 years; 64% female) with functional GI symptoms to consume a proprietary probiotic strain, B. lactis HN019 (Fonterra Research Centre, Palmerston North, New Zealand), at daily doses of 17.2 billion colony forming units (CFU) (high dose; n = 33), 1.8 billion CFU (low dose; n = 33), or placebo (n = 34) for 14 days. The primary endpoint of WGTT was assessed by X-ray on days 0 and 14 and was preceded by consumption of radiopaque markers once a day for 6 days. The secondary endpoint of functional GI symptom frequency was recorded with a subject-reported numeric (1–100) scale before and after supplementation. Results. Decreases in mean WGTT over the 14-day study period were statistically significant in the high dose group (49 ± 30 to 21 ± 32 h, p < 0.001) and the low dose group (60 ± 33 to 41 ± 39 h, p = 0.01), but not in the placebo group (43 ± 31 to 44 ± 33 h). Time to excretion of all ingested markers was significantly shorter in the treatment groups versus placebo. Of the nine functional GI symptoms investigated, eight significantly decreased in frequency in the high dose group and seven decreased with low dose, while two decreased in the placebo group. No adverse events were reported in any group. Conclusions. Daily B. lactis HN019 supplementation is well tolerated, decreases WGTT in a dose-dependent manner, and reduces the frequency of functional GI symptoms in adults. PMID:21663486

Waller, Philip A; Gopal, Pramod K; Leyer, Gregory J; Ouwehand, Arthur C; Reifer, Cheryl; Stewart, Morgan E; Miller, Larry E

2011-01-01

328

Lack of the host membrane protease FtsH hinders release of the Lactococcus lactis bacteriophage TP712.  

PubMed

The temperate bacteriophage TP712 was unable to plaque on Lactococcus lactis ?ftsH lacking the membrane protease FtsH and complementation in trans restored the WT phenotype. Absence of ftsH did not hinder phage adsorption, phage DNA delivery or activation of the lytic cycle. Thin sections revealed that TP712 virions appeared to be correctly assembled inside the ?ftsH host, but were not released. These virions were infective, demonstrating that a functional host FtsH is required by TP712 to proceed effectively with lysis of the host. PMID:24018314

Roces, Clara; Wegmann, Udo; Campelo, Ana B; García, Pilar; Rodríguez, Ana; Martínez, Beatriz

2013-12-01

329

A System To Generate Chromosomal Mutations inLactococcus lactisWhich Allows Fast Analysis of Targeted Genes  

Microsoft Academic Search

pWV01 derivative pVE6007. Transformation of L. lactis MG1363(pVE6007) with the pORI19 bank of lacto- coccal chromosomal fragments at the permissive temperature allowed replication of several copies of a recombinant plasmid from the bank within a cell because of the provision intransof RepA-Ts from pVE6007. A temperature shift to 37&C resulted in loss of pVE6007 and integration of the pORI19 derivatives

JEAN LAW; GIRBE BUIST; ALFRED HAANDRIKMAN; JAN KOK; GERARD VENEMA; ANDKEES LEENHOUTS

1995-01-01

330

Substrate specificity and mutational analysis of Kluyveromyces lactis ?-toxin, a eukaryal tRNA anticodon nuclease  

PubMed Central

tRNA anticodon damage inflicted by the Kluyveromyces lactis ?-toxin underlies an RNA-based innate immune system that distinguishes self from nonself species. ?-toxin arrests the growth of Saccharomyces cerevisiae by incising a single phosphodiester 3? of the wobble base of tRNAGlu(UUC) to generate a break with 2?,3?-cyclic phosphate and 5?-OH ends. Recombinant ?-toxin cleaves oligonucleotide substrates in vitro that mimic the anticodon stem–loop of tRNAGlu. A single 2?-deoxy sugar substitution at the wobble nucleoside abolishes anticodon nuclease activity. To gain further insights to ?-toxin's substrate specificity, we tested deoxynucleoside effects at positions other than the site of transesterification. The results attest to a stringent requirement for a ribonucleoside at the uridine 5? of the wobble base. In contrast, every other nonwobble ribonucleoside in the anticodon loop can be replaced by a deoxy without significantly affecting ?-toxin's cleavage activity. Whereas either the 5? half or the 3? half of the anticodon stem can be replaced en bloc with DNA without a major effect, simultaneously replacing both strands with DNA interfered strongly, signifying that ?-toxin requires an A-form helical conformation of the anticodon stem. We purified ?-toxin mutants identified previously as nontoxic in vivo and gauged their anticodon nuclease activities in vitro. The results highlight Glu9 and Arg151 as candidate catalytic residues, along with His209 implicated previously. By analogy to other endoribonucleases, we speculate that ?-toxin drives transesterification by general acid-base catalysis (via His209 and Glu9) and transition-state stabilization (via Arg151). PMID:21610213

Jain, Ruchi; Poulos, Michael G.; Gros, Julien; Chakravarty, Anupam K.; Shuman, Stewart

2011-01-01

331

Monte-Carlo Modeling of the Central Carbon Metabolism of Lactococcus lactis: Insights into Metabolic Regulation  

PubMed Central

Metabolic pathways are complex dynamic systems whose response to perturbations and environmental challenges are governed by multiple interdependencies between enzyme properties, reactions rates, and substrate levels. Understanding the dynamics arising from such a network can be greatly enhanced by the construction of a computational model that embodies the properties of the respective system. Such models aim to incorporate mechanistic details of cellular interactions to mimic the temporal behavior of the biochemical reaction system and usually require substantial knowledge of kinetic parameters to allow meaningful conclusions. Several approaches have been suggested to overcome the severe data requirements of kinetic modeling, including the use of approximative kinetics and Monte-Carlo sampling of reaction parameters. In this work, we employ a probabilistic approach to study the response of a complex metabolic system, the central metabolism of the lactic acid bacterium Lactococcus lactis, subject to perturbations and brief periods of starvation. Supplementing existing methodologies, we show that it is possible to acquire a detailed understanding of the control properties of a corresponding metabolic pathway model that is directly based on experimental observations. In particular, we delineate the role of enzymatic regulation to maintain metabolic stability and metabolic recovery after periods of starvation. It is shown that the feedforward activation of the pyruvate kinase by fructose-1,6-bisphosphate qualitatively alters the bifurcation structure of the corresponding pathway model, indicating a crucial role of enzymatic regulation to prevent metabolic collapse for low external concentrations of glucose. We argue that similar probabilistic methodologies will help our understanding of dynamic properties of small-, medium- and large-scale metabolic networks models. PMID:25268481

Murabito, Ettore; Verma, Malkhey; Bekker, Martijn; Bellomo, Domenico; Westerhoff, Hans V.; Teusink, Bas; Steuer, Ralf

2014-01-01

332

Monte-Carlo modeling of the central carbon metabolism of Lactococcus lactis: insights into metabolic regulation.  

PubMed

Metabolic pathways are complex dynamic systems whose response to perturbations and environmental challenges are governed by multiple interdependencies between enzyme properties, reactions rates, and substrate levels. Understanding the dynamics arising from such a network can be greatly enhanced by the construction of a computational model that embodies the properties of the respective system. Such models aim to incorporate mechanistic details of cellular interactions to mimic the temporal behavior of the biochemical reaction system and usually require substantial knowledge of kinetic parameters to allow meaningful conclusions. Several approaches have been suggested to overcome the severe data requirements of kinetic modeling, including the use of approximative kinetics and Monte-Carlo sampling of reaction parameters. In this work, we employ a probabilistic approach to study the response of a complex metabolic system, the central metabolism of the lactic acid bacterium Lactococcus lactis, subject to perturbations and brief periods of starvation. Supplementing existing methodologies, we show that it is possible to acquire a detailed understanding of the control properties of a corresponding metabolic pathway model that is directly based on experimental observations. In particular, we delineate the role of enzymatic regulation to maintain metabolic stability and metabolic recovery after periods of starvation. It is shown that the feedforward activation of the pyruvate kinase by fructose-1,6-bisphosphate qualitatively alters the bifurcation structure of the corresponding pathway model, indicating a crucial role of enzymatic regulation to prevent metabolic collapse for low external concentrations of glucose. We argue that similar probabilistic methodologies will help our understanding of dynamic properties of small-, medium- and large-scale metabolic networks models. PMID:25268481

Murabito, Ettore; Verma, Malkhey; Bekker, Martijn; Bellomo, Domenico; Westerhoff, Hans V; Teusink, Bas; Steuer, Ralf

2014-01-01

333

Dual mutations reveal interactions between components of oxidative phosphorylation in Kluyveromyces lactis.  

PubMed Central

Loss of mtDNA or mitochondrial protein synthesis cannot be tolerated by wild-type Kluyveromyces lactis. The mitochondrial function responsible for rho(0)-lethality has been identified by disruption of nuclear genes encoding electron transport and F(0)-ATP synthase components of oxidative phosphorylation. Sporulation of diploid strains heterozygous for disruptions in genes for the two components of oxidative phosphorylation results in the formation of nonviable spores inferred to contain both disruptions. Lethality of spores is thought to result from absence of a transmembrane potential, Delta Psi, across the mitochondrial inner membrane due to lack of proton pumping by the electron transport chain or reversal of F(1)F(0)-ATP synthase. Synergistic lethality, caused by disruption of nuclear genes, or rho(0)-lethality can be suppressed by the atp2.1 mutation in the beta-subunit of F(1)-ATPase. Suppression is viewed as occurring by an increased hydrolysis of ATP by mutant F(1), allowing sufficient electrogenic exchange by the translocase of ADP in the matrix for ATP in the cytosol to maintain Delta Psi. In addition, lethality of haploid strains with a disruption of AAC encoding the ADP/ATP translocase can be suppressed by atp2.1. In this case suppression is considered to occur by mutant F(1) acting in the forward direction to partially uncouple ATP production, thereby stimulating respiration and relieving detrimental hyperpolarization of the inner membrane. Participation of the ADP/ATP translocase in suppression of rho(0)-lethality is supported by the observation that disruption of AAC abolishes suppressor activity of atp2.1. PMID:11729142

Clark-Walker, G D; Chen, X J

2001-01-01

334

Glucose Metabolism in Lactococcus lactis MG1363 under Different Aeration Conditions: Requirement of Acetate To Sustain Growth under Microaerobic Conditions  

PubMed Central

Lactococcus lactis subsp. lactis MG1363 was grown in batch cultures on a defined medium with glucose as the energy source under different aeration conditions, namely, anaerobic conditions, aerobic conditions, and microaerobic conditions with a dissolved oxygen tension of 5% (when saturation with air was used as the reference). The maximum specific growth rate was high (0.78 to 0.91 h?1) under all aeration conditions but decreased with increasing aeration, and more than 90% of the glucose was converted to lactate. However, a shift in by-product formation was observed. Increasing aeration resulted in acetate, CO2, and acetoin replacing formate and ethanol as end products. Under microaerobic conditions, growth came to a gradual halt, although more than 60% of the glucose was still left. A decline in growth was not observed during microaerobic cultivation when acetate was added to the medium. We hypothesize that the decline in growth was due to a lack of acetyl coenzyme A (acetyl-CoA) needed for fatty acid synthesis since acetyl-CoA can be synthesized from acetate by means of acetate kinase and phosphotransacetylase activities. PMID:12788751

Nordkvist, Mikkel; Jensen, Niels Bang Siemsen; Villadsen, John

2003-01-01

335

The acid tolerant and cold-active ?-galactosidase from Lactococcus lactis strain is an attractive biocatalyst for lactose hydrolysis.  

PubMed

The gene encoding the ?-galactosidase from the dairy Lactococcus lactis IL1403 strain was cloned, sequenced and overexpressed in Escherichia coli. The purified enzyme has a tetrameric arrangement composed of four identical 120 kDa subunits. Biochemical characterization showed that it is optimally active within a wide range of temperatures from 15 to 55 °C and of pH from 6.0 to 7.5. For its maximal activity this enzyme requires only 0.8 mM Fe(2+) and 1.6 mM Mg(2+). Purified protein displayed a high catalytic efficiency of 102 s(-1) mM(-1) for lactose. The enzyme stability was increased by immobilization mainly at low pH (from 4.0 to 5.5) and high temperatures (55 and 60 °C). The bioconversion of lactose using the L. lactis ?-galactosidase allows the production of lactose with a high bioconversion rate (98 %) within a wide range of pH and temperature. PMID:23180374

Vincent, Violette; Aghajari, Nushin; Pollet, Noémie; Boisson, Anaïs; Boudebbouze, Samira; Haser, Richard; Maguin, Emmanuelle; Rhimi, Moez

2013-04-01

336

Identification and Molecular Characterization of the Chromosomal Exopolysaccharide Biosynthesis Gene Cluster from Lactococcus lactis subsp. cremoris SMQ-461  

PubMed Central

The exopolysaccharide (EPS) capsule-forming strain SMQ-461 of Lactococcus lactis subsp. cremoris, isolated from raw milk, produces EPS with an apparent molecular mass of >1.6 × 106 Da. The EPS biosynthetic genes are located on the chromosome in a 13.2-kb region consisting of 15 open reading frames. This region is flanked by three IS1077-related tnp genes (L. lactis) at the 5? end and orfY, along with an IS981-related tnp gene, at the 3? end. The eps genes are organized in specific regions involved in regulation, chain length determination, biosynthesis of the repeat unit, polymerization, and export. Three (epsGIK) of the six predicted glycosyltransferase gene products showed low amino acid similarity with known glycosyltransferases. The structure of the repeat unit could thus be different from those known to date for Lactococcus. Reverse transcription-PCR analysis revealed that the eps locus is transcribed as a single mRNA. The function of the eps gene cluster was confirmed by disrupting the priming glycosyltransferase gene (epsD) in Lactococcus cremoris SMQ-461, generating non-EPS-producing reversible mutants. This is the first report of a chromosomal location for EPS genetic elements in Lactococcus cremoris, with novel glycosyltransferases not encountered before in lactic acid bacteria. PMID:16269783

Dabour, N.; LaPointe, G.

2005-01-01

337

Production of polyomavirus-like particles in a Klgal80 knockout strain of the yeast Kluyveromyces lactis.  

PubMed

VP1, the major coat protein of polyomavirus, assembles intracellularly to virus-like particles if expressed in eukaryotes. Here, the nonconventional yeast Kluyveromyces lactis was used for production of virus-like particles of murine polyomavirus. The heterologous gene of VP1 was integrated in the LAC4 locus of the GAL/LAC genes. Consequently the expression of VP1 is regulated by the interplay of the activator KlGal4p and inhibitor KlGal80p. This cloning strategy couples the production of VP1 to that of the enzyme ? -galactosidase, allowing a fast alternative for monitoring the course of recombinant protein production by measuring the ? -galactosidase activity. A Klgal80 knockout strain was generated for a constitutive expression of VP1 and a continuous VLP production. High-cell-density fermentation showed that (1) Kluyveromyces lactis is generally suitable for VLP production and (2) the Klgal80 knockout strain produces higher amounts of recombinant VP1. Furthermore, VLPs could be purified chromatographically to 87% (w/w) of total protein, and showed a homogeneous species of 45-nm particles and a high resistance against proteolysis compared to conventional in vitro assembled VLPs. This demonstrates the superior stability of virus-like particles produced in yeast. PMID:23302109

Simon, Claudia; Schaepe, Sebastian; Breunig, Karin; Lilie, Hauke

2013-01-01

338

Isolation of nisin-producing Lactococcus lactis WNC 20 strain from nham, a traditional Thai fermented sausage.  

PubMed

A total of 14,020 lactic acid bacteria (LAB) were isolated from nham and screened for bacteriocin production. One Lactococcus lactis strain WNC 20 produced a bacteriocin that not only inhibited closely related LAB, but also some food-borne pathogens including Listeria monocytogenes, Clostridium perfringens, Bacillus cereus and Staphylococcus aureus. Biochemical studies revealed that the bacteriocin was heat-stable even at autoclaving temperature (121 degrees C for 15 min) and was active over a wide pH range (2-10). The bacteriocin was inactivated by alpha-chymotrypsin and proteinase K but not other proteases. The antimicrobial spectrum and some characteristics of this bacteriocin were nearly identical to that of nisin. The gene encoding this bacteriocin was amplified by polymerase chain reaction (PCR) with nisin gene-specific primer. Sequencing of this gene showed identical sequences to nisin Z as indicated by the substitution of asparagine residue instead of histidine at position 27. The ability of the bacteriocin produced by Lc. lactis WNC 20 may be useful in improving the food safety of the fermented product. PMID:12457588

Noonpakdee, W; Santivarangkna, C; Jumriangrit, P; Sonomoto, K; Panyim, S

2003-03-15

339

Milk acidification by Lactococcus lactis is improved by decreasing the level of dissolved oxygen rather than decreasing redox potential in the milk prior to inoculation  

Microsoft Academic Search

Although redox potential is very rarely taken into account in food fermentation it could be as influential as pH on bacterial activities. Lactococcus lactis is already known to exhibit a powerful reducing activity in milk but its reduction activity was shown to occur prior to its acidification activity with a potential interaction between these two lactococcal activities. Therefore, acidification lag-type

Sophie Jeanson; Nadine Hilgert; Marie-Odile Coquillard; Céline Seukpanya; Marc Faiveley; Pascal Neveu; Christophe Abraham; Véra Georgescu; Pascal Fourcassié; Eric Beuvier

2009-01-01

340

Growth Phase-Dependent Proteomes of the Malaysian Isolated Lactococcus lactis Dairy Strain M4 Using Label-Free Qualitative Shotgun Proteomics Analysis  

PubMed Central

Lactococcus lactis is the most studied mesophilic fermentative lactic acid bacterium. It is used extensively in the food industry and plays a pivotal role as a cell factory and also as vaccine delivery platforms. The proteome of the Malaysian isolated L. lactis M4 dairy strain, obtained from the milk of locally bred cows, was studied to elucidate the physiological changes occurring between the growth phases of this bacterium. In this study, ultraperformance liquid chromatography nanoflow electrospray ionization tandem mass spectrometry (UPLC- nano-ESI-MSE) approach was used for qualitative proteomic analysis. A total of 100 and 121 proteins were identified from the midexponential and early stationary growth phases, respectively, of the L. lactis strain M4. During the exponential phase, the most important reaction was the generation of sufficient energy, whereas, in the early stationary phase, the metabolic energy pathways decreased and the biosynthesis of proteins became more important. Thus, the metabolism of the cells shifted from energy production in the exponential phase to the synthesis of macromolecules in the stationary phase. The resultant proteomes are essential in providing an improved view of the cellular machinery of L. lactis during the transition of growth phases and hence provide insight into various biotechnological applications. PMID:24982972

Yap, Theresa Wan Chen; Rabu, Amir; Abu Bakar, Farah Diba; Abdul Rahim, Raha; Mahadi, Nor Muhammad; Illias, Rosli Md.

2014-01-01

341

Growth phase-dependent proteomes of the Malaysian isolated Lactococcus lactis dairy strain M4 using label-free qualitative shotgun proteomics analysis.  

PubMed

Lactococcus lactis is the most studied mesophilic fermentative lactic acid bacterium. It is used extensively in the food industry and plays a pivotal role as a cell factory and also as vaccine delivery platforms. The proteome of the Malaysian isolated L. lactis M4 dairy strain, obtained from the milk of locally bred cows, was studied to elucidate the physiological changes occurring between the growth phases of this bacterium. In this study, ultraperformance liquid chromatography nanoflow electrospray ionization tandem mass spectrometry (UPLC- nano-ESI-MS(E)) approach was used for qualitative proteomic analysis. A total of 100 and 121 proteins were identified from the midexponential and early stationary growth phases, respectively, of the L. lactis strain M4. During the exponential phase, the most important reaction was the generation of sufficient energy, whereas, in the early stationary phase, the metabolic energy pathways decreased and the biosynthesis of proteins became more important. Thus, the metabolism of the cells shifted from energy production in the exponential phase to the synthesis of macromolecules in the stationary phase. The resultant proteomes are essential in providing an improved view of the cellular machinery of L. lactis during the transition of growth phases and hence provide insight into various biotechnological applications. PMID:24982972

Yap, Theresa Wan Chen; Rabu, Amir; Abu Bakar, Farah Diba; Rahim, Raha Abdul; Mahadi, Nor Muhammad; Illias, Rosli Md; Murad, Abdul Munir Abdul

2014-01-01

342

Effect on Components of the Intestinal Microflora and Plasma Neuropeptide Levels of Feeding Lactobacillus delbrueckii, Bifidobacterium lactis, and Inulin to Adult and Elderly Rats  

Microsoft Academic Search

The aim of this study was to compare the effects of the mixture of Lactobacillus delbrueckii subsp. rhamnosus strain GG, Bifidobacterium lactis Bb12, and inulin on intestinal populations of lactobacilli, bifidobacteria, and enterobacteria in adult and elderly rats fed the same (in quality and quantity) diet. The portal plasma levels of two neuropeptides, neuropeptide Y (NPY) and peptide YY (PYY),

Violetta Lesniewska; Ian Rowland; Patrice D. Cani; Audrey M. Neyrinck; Nathalie M. Delzenne; Patrick J. Naughton

2006-01-01

343

Characteristics of the bacteriocin produced by Lactococcus lactis subsp. cremoris CTC 204 and the effect of this compound on the mesophilic bacteria associated with raw beef  

Microsoft Academic Search

Summary >Screening for the bacteriocin production of strains of lactic acid bacteria from various meat and meat products resulted in the detection of a bacteriocin-producing Lactococcus lactis subsp. cremoris CTC 204, isolated from chicken. The bacteriocin inhibited not only closely related lactic acid bacteria (Lactobacillus helveticus), but also pathogenic microorganisms (Staphylococcus aureus, Listeria monocytogenes, Bacillus cereus, and Clostridium perfringens). It

R. Bromberg; I. Moreno; R. R. Delboni; H. C. Cintra; P. T. V Oliveira

2005-01-01

344

Shewanella oneidensis in a lactate-fed pure-culture and a glucose-fed co-culture with Lactococcus lactis with an electrode as electron acceptor  

E-print Network

terminal electron acceptors, such as mineral oxides or electrodes, has positioned Shewanella oneidensis MR lactis with an electrode as electron acceptor Miriam A. Rosenbaum a , Haim Y. Bar b , Qasim K. Beg c electron acceptor). Such transfer of electrons is highly promising for novel applications in wastewater

Segrè, Daniel

345

Genome Sequence Analysis of the Biogenic Amine-Producing Strain Lactococcus lactis subsp. cremoris CECT 8666 (Formerly GE2-14)  

PubMed Central

We here report a 2,801,031-bp annotated draft assembly for the Lactococcus lactis subsp. cremoris GE2-14 genome. This dairy strain produces the biogenic amine putrescine. This sequence may help identify the mechanisms regulating putrescine biosynthesis and throw light on ways to reduce its presence in fermented foods. PMID:25342694

del Rio, Beatriz; Linares, Daniel M.; Fernandez, Maria; Mayo, Baltasar; Martin, M. Cruz; Alvarez, Miguel A.

2014-01-01

346

Cloning, Expression, and Chromosomal Stabilization of the Propionibacterium shermanii Proline Iminopeptidase Gene (pip) for Food-Grade Application in Lactococcus lactis  

PubMed Central

Proline iminopeptidase produced by Propionibacterium shermanii plays an essential role in the flavor development of Swiss-type cheeses. The enzyme (Pip) was purified and characterized, and the gene (pip) was cloned and expressed in Escherichia coli and Lactococcus lactis, the latter species being an extensively studied, primary cheese starter culture that is less fastidious in its growth condition requirements than P. shermanii. The levels of expression of the pip gene could be enhanced with a factor 3 to 5 by using a strong constitutive promoter in L. lactis or the inducible tac promoter in E. coli. Stable replication of the rolling-circle replicating (rcr) plasmid, used to express pip in L. lactis, could only be obtained by providing the repA gene in trans. Upon the integration of pip, clear gene dosage effects were observed and stable multicopy integrants could be maintained upon growth under the selective pressure of sucrose. The multicopy integrants demonstrated a high degree of stability in the presence of glucose. This study examines the possibilities to overexpress genes that play an important role in food fermentation processes and shows a variety of options to obtain stable food-grade expression of such genes in L. lactis. PMID:9835556

Leenhouts, Kees; Bolhuis, Albert; Boot, Johan; Deutz, Inge; Toonen, Marjolein; Venema, Gerard; Kok, Jan; Ledeboer, Aat

1998-01-01

347

Use of synthetic genes for cloning, production and functional expression of the bacteriocins enterocin A and bacteriocin E 50-52 by Pichia pastoris and Kluyveromyces lactis.  

PubMed

The use of synthetic genes may constitute a successful approach for the heterologous production and functional expression of bacterial antimicrobial peptides (bacteriocins) by recombinant yeasts. In this work, synthetic genes with adapted codon usage designed from the mature amino acid sequence of the bacteriocin enterocin A (EntA), produced by Enterococcus faecium T136, and the mature bacteriocin E 50-52 (BacE50-52), produced by E. faecium NRRL B-32746, were synthesized. The synthetic entA and bacE50-52 were cloned into the protein expression vectors pPICZ?A and pKLAC2 for transformation of derived vectors into Pichia pastoris X-33 and Kluyveromyces lactis GG799, respectively. The recombinant vectors were linearized and transformed into competent cells selecting for P. pastoris X-33EAS (entA), P. pastoris X-33BE50-52S (bacE50-52), K. lactis GG799EAS (entA), and K. lactis GG799BE50-52S (bacE50-52). P. pastoris X-33EAS and K. lactis GG799EAS, but not P. pastoris X-33BE50-52S and K. lactis GG799BE50-52S, showed antimicrobial activity in their supernatants. However, purification of the supernatants of the producer yeasts permitted recovery of the bacteriocins EntA and BacE50-52. Both purified bacteriocins were active against Gram-positive bacteria such as Listeria monocytogenes but not against Gram-negative bacteria, including Campylobacter jejuni. PMID:24510220

Jiménez, Juan J; Borrero, Juan; Gútiez, Loreto; Arbulu, Sara; Herranz, Carmen; Cintas, Luis M; Hernández, Pablo E

2014-06-01

348

Increase of stress resistance in Lactococcus lactis via a novel food-grade vector expressing a shsp gene from Streptococcus thermophilus  

PubMed Central

The effects of the expression of a small heat shock protein (shsp) gene from Streptococcus thermophilus on stress resistance in Lactococcus lactis under different environmental stresses were investigated in this study. pMG36e-shsp, an expression vector, was first constructed by inserting a shsp open reading frame (ORF) cloned from S. thermophilus strain St-QC into pMG36e. Then, a food-grade expression vector, pMG-shsp, was generated by deleting the erythromycin resistance gene from pMG36e-shsp. The transformation rate of pMG-shsp was comparable to that of pMG36e-shsp when each of these two vectors was introduced into L. lactis. These results demonstrated that the shsp ORF could successfully used as a food-grade selection marker in both pMG-shsp and pMG36e-shsp. Furthermore, the growth characteristics were almost the same between L. lactis ML23 transformants harboring pMG36e or pMG-shsp. The survival rate of L. lactis ML23 expressing the shsp ORF were increased to 0.032%, 0.006%, 0.0027%, 0.03%, and 0.16% under the following environmental stresses: heat, acid, ethanol, bile salt and H2O2, respectively. These results indicated that the expression of the shsp gene in the food-grade vector pMG-shsp conferred resistance to environmental stresses without affecting the growth characteristics of L. lactis ML23. PMID:24031940

Tian, Hongtao; Tan, Jianxin; Zhang, Lifang; Gu, Xinxi; Xu, Wentao; Guo, Xinghua; Luo, Yunbo

2012-01-01

349

Effects in the use of a genetically engineered strain of Lactococcus lactis delivering in situ IL-10 as a therapy to treat low-grade colon inflammation.  

PubMed

Irritable bowel syndrome (IBS) is a gastrointestinal disorder characterized by chronic abdominal pain, discomfort, and bloating. Interestingly, there is now evidence of the presence of a low-grade inflammatory status in many IBS patients, including histopathological and mucosal cytokine levels in the colon, as well as the presence of IBS-like symptoms in quiescent inflammatory bowel disease (IBD). The use of a genetically engineered food-grade bacterium, such as Lactococcus lactis, secreting the anti-inflammatory cytokine IL-10 has been proven by many pre-clinical studies to be a successful therapy to treat colon inflammation. In this study, we first reproduced the recovery-recurrence periods observed in IBS-patients in a new chronic model characterized by 2 episodes of DiNitro-BenzeneSulfonic-acid (DNBS)-challenge and we tested the effects of a recombinant strain of L. lactis secreting IL-10 under a Stress-Inducible Controlled Expression (SICE) system. In vivo gut permeability, colonic serotonin levels, cytokine profiles, and spleen cell populations were then measured as readouts of a low-grade inflammation. In addition, since there is increasing evidence that gut microbiota tightly regulates gut barrier function, tight junction proteins were also measured by qRT-PCR after administration of recombinant L. lactis in DNBS-treated mice. Strikingly, oral administration of L. lactis secreting active IL-10 in mice resulted in significant protective effects in terms of permeability, immune activation, and gut-function parameters. Although genetically engineered bacteria are, for now, used only as a "proof-of-concept," our study validates the interest in the use of the novel SICE system in L. lactis to express therapeutic molecules, such as IL-10, locally at mucosal surfaces. PMID:24732667

Martín, Rebeca; Martín, Rebeca; Chain, Florian; Chain, Florian; Miquel, Sylvie; Miquel, Sylvie; Natividad, Jane M; Natividad, Jane M; Sokol, Harry; Sokol, Harry; Verdu, Elena F; Verdu, Elena F; Langella, Philippe; Langella, Philippe; Bermúdez-Humarán, Luis G; Bermúdez-Humarán, Luis G

2014-06-01

350

Growth of infants fed formula supplemented with Bifidobacterium lactis Bb12 or Lactobacillus GG: a systematic review of randomized controlled trials  

PubMed Central

Background Growth is an essential outcome measure for evaluating the safety of any new ingredients, including probiotics, added to infant formulae. The aim of this systematic review was to determine the effects of supplementation of infant formulae with Bifidobacterium lactis Bb12 (B lactis) and/or Lactobacillus rhamnosus GG (LGG) compared with unsupplemented formula on the growth of healthy infants. Methods The MEDLINE, EMBASE, and Cochrane Library databases were searched in June 2013 for relevant randomized controlled trials (RCTs) conducted in healthy term infants. Unpublished data were obtained from the manufacturer of B lactis-supplemented formula. The primary outcome measures were weight, length, and head circumference. Results Nine eligible trials were identified. Compared with unsupplemented controls, supplementation of infant formula with B lactis had no effect on weight gain [4 RCTs, n?=?266, mean difference (MD) 0.96 g/day, 95% confidence interval (CI) -0.70 to 2.63)], length gain (4 RCTs, n?=?261, MD ?0.39 mm/month, 95% CI ?1.32 to 0.53), or head circumference gain (3 RCTs, n?=?207, MD 0.56 mm/month, 95% CI ?0.17 to 1.30). Data limited to one small (n?=?105) trial suggest that infants who received standard infant formula supplemented with LGG grew significantly better. No such effect was observed in infants fed hydrolyzed formula supplemented with LGG. Conclusions Supplementation of infant formula with B lactis results in growth similar to what is found in infants fed unsupplemented formula. Limited data do not allow one to reach a conclusion regarding the effect of LGG supplementation on infant growth. PMID:24215626

2013-01-01

351

Oral immunization with Lactococcus lactis-expressing EspB induces protective immune responses against Escherichia coli O157:H7 in a murine model of colonization.  

PubMed

Enterohemorrhagic Escherichia coli (EHEC) have been responsible for several outbreaks of hemolytic-uremic syndrome (HUS) worldwide. HUS is the most common cause of acute renal failure in children and results in fatalities as high as 50% in the elderly. Currently, neither a specific treatment nor a vaccine is available for EHEC. Lactococcus lactis is a generally regarded as safe "GRAS" bacterium that offers a valuable platform for oral vaccine delivery. Toward the development of an oral vaccine against EHEC, we have previously constructed a recombinant L. lactis strain expressing the EHEC antigen, EspB in the cytoplasmic compartment. However, oral immunization of mice with this strain induced weak priming of the immune system. This outcome was attributed to the rather low levels of EspB expressed by this recombinant strain. Therefore, in the present study we optimized the expression of EspB in L. lactis by secreting the antigen either under constitutive or nisin-inducible control. Indeed, oral immunization of mice with the EspB-secreting strains successfully induced specific mucosal and systemic antibody responses. These responses were associated with mixed Th1/Th2 cell responses in Peyer's Patches and mesenteric lymph nodes. Moreover, immunized mice exhibited significant protection against E. coli O157:H7 colonization, as indicated by the reduced amount and/or duration of the bacterial fecal shedding. Our results demonstrate the protective potential of EspB as an oral vaccine against EHEC infection. Additionally, the study demonstrates the efficient delivery of recombinant EspB by the "GRAS" bacterium, L. lactis. The safety profile of L. lactis as a vaccine vehicle can particularly be beneficial to children and elderly as high-risk groups for HUS incidence. PMID:24877767

Ahmed, B; Loos, M; Vanrompay, D; Cox, E

2014-06-30

352

Immune Response Elicited by DNA Vaccination Using Lactococcus lactis Is Modified by the Production of Surface Exposed Pathogenic Protein  

PubMed Central

In this study, we compared immune responses elicited by DNA immunization using Lactococcus lactis or L. lactis expressing the Staphylococcus aureus invasin Fibronectin Binding Protein A (FnBPA) at its surface. Both strains carried pValac:BLG, a plasmid containing the cDNA of Beta-Lactoglobulin (BLG), and were designated LL-BLG and LL-FnBPA+ BLG respectively. A TH2 immune response characterized by the secretion of IL-4 and IL-5 in medium of BLG reactivated splenocytes was detected after either oral or intranasal administration of LL-FnBPA+ BLG. In contrast, intranasal administration of LL-BLG elicited a TH1 immune response. After BLG sensitization, mice previously intranasally administered with LL-BLG showed a significantly lower concentration of BLG-specific IgE than the mice non-administered. Altenatively administration of LL-FnBPA+ BLG didn't modify the BLG-specific IgE concentration obtained after sensitization, thus confirming the TH2 orientation of the immune response. To determine if the TH2-skewed immune response obtained with LL-FnBpA+ BLG was FnBPA-specific or not, mice received another L. lactis strain producing a mutated form of the Listeria monocytogenes invasin Internalin A intranasally, allowing thus the binding to murine E-cadherin, and containing pValac:BLG (LL-mInlA+ BLG). As with LL-FnBPA+ BLG, LL-mInlA+ BLG was not able to elicit a TH1 immune response. Furthermore, we observed that these difference were not due to the peptidoglycan composition of the cell wall as LL-FnBPA+ BLG, LL-mInlA+ BLG and LL-BLG strains shared a similar composition. DNA vaccination using LL-BLG elicited a pro-inflammatory TH1 immune response while using LL-FnBPA+ BLG or LL-mInlA+ BLG elicited an anti-inflammatory TH2 immune response. PMID:24465412

Pontes, Daniela; Azevedo, Marcela; Innocentin, Silvia; Blugeon, Sebastien; Lefevre, Francois; Azevedo, Vasco; Miyoshi, Anderson; Courtin, Pascal; Chapot-Chartier, Marie-Pierre; Langella, Philippe; Chatel, Jean-Marc

2014-01-01

353

Benchmarking Various Green Fluorescent Protein Variants in Bacillus subtilis, Streptococcus pneumoniae, and Lactococcus lactis for Live Cell Imaging  

PubMed Central

Green fluorescent protein (GFP) offers efficient ways of visualizing promoter activity and protein localization in vivo, and many different variants are currently available to study bacterial cell biology. Which of these variants is best suited for a certain bacterial strain, goal, or experimental condition is not clear. Here, we have designed and constructed two “superfolder” GFPs with codon adaptation specifically for Bacillus subtilis and Streptococcus pneumoniae and have benchmarked them against five other previously available variants of GFP in B. subtilis, S. pneumoniae, and Lactococcus lactis, using promoter-gfp fusions. Surprisingly, the best-performing GFP under our experimental conditions in B. subtilis was the one codon optimized for S. pneumoniae and vice versa. The data and tools described in this study will be useful for cell biology studies in low-GC-rich Gram-positive bacteria. PMID:23956387

Overkamp, Wout; Beilharz, Katrin; Detert Oude Weme, Ruud; Solopova, Ana; Karsens, Harma; Kovacs, Akos T.; Kok, Jan

2013-01-01

354

Characterization of the divergent sacBK and sacAR operons, involved in sucrose utilization by Lactococcus lactis.  

PubMed

The divergently transcribed sacBK and sacAR operons, which are involved in the utilization of sucrose by Lactococcus lactis NZ9800, were examined by transcriptional and gene inactivation studies. Northern analyses of RNA isolated from cells grown at the expense of different carbon sources revealed three sucrose-inducible transcripts: one of 3.2 kb containing sacB and sacK, a second of 3.4 kb containing sacA and sacR, and a third of 1.8 kb containing only sacR. The inactivation of the sacR gene by replacement recombination resulted in the constitutive transcription of the sacBK and sacAR operons in the presence of different carbon sources, indicating that SacR acts as a repressor of transcription. PMID:10074089

Luesink, E J; Marugg, J D; Kuipers, O P; de Vos, W M

1999-03-01

355

Recombinant Lactococcus lactis can make the difference in antigen-specific immune tolerance induction, the Type 1 Diabetes case.  

PubMed

Especially in western civilizations, immune diseases that are driven by innocuous (auto- or allo-) antigens are gradually evolving to become pandemic threats. A particularly poignant example is type 1 diabetes, where young children are confronted with the perspective and consequences of total pancreatic ?-cell destruction. Along these disquieting observations we find ourselves equipped with impressively accumulating molecular immunological knowledge on the ins and outs of these pathologies. Often, however, it is difficult to translate this wealth into efficacious medicines. The molecular understanding, the concept of oral tolerance induction, the benefit of using recombinant Lactococcus lactis therein and recent openings towards their clinical use may well enable turning all colors to their appropriate fields on this Rubik's cube. PMID:25185797

Robert, Sofie; Steidler, Lothar

2014-08-29

356

Oral intake of heat-killed cells of Lactococcus lactis strain H61 promotes skin health in women.  

PubMed

We conducted a double-blind, placebo-controlled trial to evaluate the effect of heat-killed cells of Lactococcus lactis strain H61 on various skin properties of Japanese women. Volunteers (age 31-62 years) were randomly assigned to receive test food with or without 60 mg of heat-killed strain H61 (fifteen women in each group; H61 and control groups, respectively) daily for 8 weeks. Results were analysed for three age categories (30s, 40s and 50-60s). Compared with that at week 0, skin hydration at the inner forearm at weeks 4 and 8 decreased in all volunteers (except those in their 50-60s) because of the environmental change from autumn to winter. The oldest H61 group maintained skin hydration at the inner forearm throughout the study. Skin elasticity and melanin content in the cheek decreased and sebum content increased throughout the test period due to seasonal environmental change, regardless of age or H61 treatment. Self-evaluation scores for apparent hair follicles and dryness of the throat at week 8 were higher in the overall H61 group than in the combined placebo group. The 30s H61 group noted marked improvements in self-surveyed skin elasticity at week 8 compared with at week 0 and with the placebo group at week 8. The results of the present study indicate that oral intake of heat-killed cells of L. lactis strain H61 can improve some skin properties and body characteristics in women. This strain would probably be useful in increasing the quality of life in an ageing population. PMID:25191547

Kimoto-Nira, Hiromi; Aoki, Reiji; Sasaki, Keisuke; Suzuki, Chise; Mizumachi, Koko

2012-01-01

357

Proton motive force-driven and ATP-dependent drug extrusion systems in multidrug-resistant Lactococcus lactis.  

PubMed

Three mutants of Lactococcus lactis subsp. lactis MG1363, termed EthR, DauR, and RhoR, were selected for resistance to high concentrations of ethidium bromide, daunomycin, and rhodamine 6G, respectively. These mutants were found to be cross resistant to a number of structurally and functionally unrelated drugs, among which were typical substrates of the mammalian multidrug transporter (P-glycoprotein) such as daunomycin, quinine, actinomycin D, gramicidin D, and rhodamine 6G. The three multidrug-resistant strains showed an increased rate of energy-dependent ethidium and daunomycin efflux compared with that of the wild-type strain. This suggests that resistance to these toxic compounds is at least partly due to active efflux. Efflux of ethidium from the EthR strain could occur against a 37-fold inwardly directed concentration gradient. In all strains, ethidium efflux was inhibited by reserpine, a well-known inhibitor of P-glycoprotein. Ionophores which selectively dissipate the membrane potential or the pH gradient across the membrane inhibited ethidium and daunomycin efflux in the wild-type strain, corresponding with a proton motive force-driven efflux system. The ethidium efflux system in the EthR strain, on the other hand, was inhibited by ortho-vanadate and not upon dissipation of the proton motive force, which suggests the involvement of ATP in the energization of transport. The partial inhibition of ethidium efflux by ortho-vanadate and nigericin in the DauR and RhoR strains suggest that a proton motive force-dependent and an ATP-dependent system are expressed simultaneously. This is the first report of an ATP-dependent transport system in prokaryotes which confers multidrug resistance to the organism. PMID:7961458

Bolhuis, H; Molenaar, D; Poelarends, G; van Veen, H W; Poolman, B; Driessen, A J; Konings, W N

1994-11-01

358

Surface expression of Helicobacter pylori urease subunit B gene E fragment on Lactococcus lactis by means of the cell wall anchor of Staphylococcus aureus protein A  

Microsoft Academic Search

Fragment E of ureB (ureBE) was cloned from a clinical isolate of Helicobacter pylori. A prokaryotic expression vector, pAMJ399, with the ureB fragment E and the Staphylococcus aureus protein A anchor fragment (spaX), was constructed. The fusion protein was expressed under the control of the P170 promoter in Lactococcus lactis. Western blot assay of lactococcal cell wall extracts with a

Dafeng Song; Qing Gu

2009-01-01

359

GAL4 of Saccharomyces cerevisiae activates the lactose-galactose regulon of Kluyveromyces lactis and creates a new phenotype: glucose repression of the regulon.  

PubMed Central

A Kluyveromyces lactis mutant defective in lac9 cannot induce beta-galactosidase or galactokinase activity and is unable to grow on lactose or galactose. When this strain was transformed with the GAL4 positive regulatory gene of Saccharomyces cerevisiae it was able to grow on lactose or galactose as the sole carbon source. Transformants bearing GAL4 exhibited a 4.5-h generation time on galactose or lactose, versus 24 h for the nontransformed lac9 strain. A K. lactis lac9 strain bearing two integrated copies of GAL4 showed 3.5-fold induction of beta-galactosidase activity and 1.8-fold induction of galactokinase activity compared with 15.6-fold and 4.4-fold induction, respectively, for the LAC9 wild-type strain. In transformants bearing 10 integrated copies of GAL4, the induced level of beta-galactosidase was nearly as high as in the LAC9 wild-type strain. In addition to restoring lactose and galactose gene expression, GAL4 in K. lactis lac9 mutant cells conferred a new phenotype, severe glucose repression of lactose and galactose-inducible enzymes. Glucose repressed beta-galactosidase activity 35- to 74-fold and galactokinase activity 14- to 31-fold in GAL4 transformants, compared with the 2-fold glucose repression exhibited in the LAC9 wild-type strain. The S. cerevisiae MEL1 gene was repressed fourfold by glucose in LAC9 cells. In contrast, the MEL1 gene in a GAL4 lac9 strain was repressed 20-fold by glucose. These results indicate that the GAL4 and LAC9 proteins activate transcription in a similar manner. However, either the LAC9 or GAL4 gene or a product of these genes responds differently to glucose in K. lactis. Images PMID:3102945

Riley, M I; Hopper, J E; Johnston, S A; Dickson, R C

1987-01-01

360

Characterization of KlGRR1 and SMS1 Genes, Two New Elements of the Glucose Signaling Pathway of Kluyveromyces lactis?  

PubMed Central

The expression of the major glucose transporter gene, RAG1, is induced by glucose in Kluyveromyces lactis. This regulation involves several pathways, including one that is similar to Snf3/Rgt2-ScRgt1 in Saccharomyces cerevisiae. We have identified missing key components of the K. lactis glucose signaling pathway by comparison to the same pathway of S. cerevisiae. We characterized a new mutation, rag19, which impairs RAG1 regulation. The Rag19 protein is 43% identical to the F-box protein ScGrr1 of S. cerevisiae and is able to complement an Scgrr1 mutation. In the K. lactis genome, we identified a single gene, SMS1 (for similar to Mth1 and Std1), that encodes a protein showing an average of 50% identity with Mth1 and Std1, regulators of the ScRgt1 repressor. The suppression of the rag4 (glucose sensor), rag8 (casein kinase I), and rag19 mutations by the ?sms1 deletion, together with the restoration of RAG1 transcription in the double mutants, demonstrates that Sms1 is a negative regulator of RAG1 expression and is acting downstream of Rag4, Rag8, and Rag19 in the cascade. We report that Sms1 regulates KlRgt1 repressor activity by preventing its phosphorylation in the absence of glucose, and that SMS1 is regulated by glucose, both at the transcriptional and the posttranslational level. Two-hybrid interactions of Sms1 with the glucose sensor and KlRgt1 repressor suggest that Sms1 mediates the glucose signal from the plasma membrane to the nucleus. All of these data demonstrated that Sms1 was the K. lactis homolog of MTH1 and STD1 of S. cerevisiae. Interestingly, MTH1 and STD1 were unable to complement a ?sms1 mutation. PMID:18552281

Hnatova, Martina; Wesolowski-Louvel, Micheline; Dieppois, Guenaelle; Deffaud, Julien; Lemaire, Marc

2008-01-01

361

Production and purification of staphylococcal nuclease in Lactococcus lactis using a new expression-secretion system and a pH-regulated mini-reactor  

PubMed Central

Background Staphylococcal (or micrococcal) nuclease or thermonuclease (SNase or Nuc) is a naturally-secreted nucleic acid degrading enzyme that participates in Staphylococcus aureus spread in the infected host. Purified Nuc protein can be used as an exogenous reagent to clear cellular extracts and improve protein purification. Here, a recombinant form of Nuc was produced and secreted in a Gram-positive host, Lactococcus lactis, and purified from the culture medium. Results The gene segment corresponding to the S. aureus nuclease without its signal peptide was cloned in an expression-secretion vector. It was then fused to a lactococcal sequence encoding a signal peptide, and expressed under the control of a lactococcal promoter that is inducible by zinc starvation. An L. lactis subsp cremoris model strain (MG1363) transformed with the resulting plasmid was grown in either of two media (GM17v and CDM) that are free of animal compounds, allowing GMP (Good Manufacturing Practice) production. Induction conditions (concentration of the metal chelator EDTA and timing of addition) in small-scale pH-regulated fermentors were optimized using LacMF (Lactis Multi-Fermentor), a home-made parallel fermentation control system able to monitor 12 reactors simultaneously. Large amounts of recombinant Nuc (rNuc) were produced and secreted in both media, and rNuc was purified from GM17v medium in a single-step procedure. Conclusions In L. lactis, rNuc production and secretion were optimal after induction by 0.5 mM EDTA in small scale (200 mL) GM17v exponential phase cultures (at an OD600 of 2), leading to a maximal protein yield of 210 mg per L of culture medium. Purified rNuc was highly active, displaying a specific activity of 2000 U/mg. PMID:20492646

2010-01-01

362

The nuclear Kluyveromyces lactis MRF1 gene encodes a mitochondrial class I peptide chain release factor that is important for cell viability  

Microsoft Academic Search

We report the isolation and characterization of the Kluyveromyces lactis MRF1 gene encoding mitochondrial peptide chain release factor mRF-1. Over-expression of the KlMRF1 gene has a strong antisuppressive effect in a Saccharomyces cerevisiae mitochondrial nonsense suppressor strain. Inactivation of KlMRF1 results in a dual phenotype: most cells die after about 10–13 generations, while a small number of cells exceed this

Herman J. Pel; Sophie Rozenfeld; Monique Bolotin-Fukuhara

1996-01-01

363

Nucleotide sequence and analysis of pWC1, a pC194-type rolling circle replicon in Lactococcus lactis.  

PubMed

A 2.8-kb cryptic plasmid showing no homology to either pFX3 (rolling circle, pE194-type) or pCI305 (theta-type) lactococcal replicons was identified in Lactococcus lactis subsp. cremoris 2204. The plasmid, pWC1, was compatible with both pCI3340 (a pCI305 derivative) and pFX3 in L. lactis subsp. cremoris 2204. Sequence analysis of pWC1 showed one major ORF encoding a protein with a deduced size of 316 amino acids (aa). Database comparisons showed that the protein was distinct from the pFX- and pCI-type replication proteins (less than 21% aa identity), but shared significant homology (up to 57% aa identity) with the replication proteins from a different group of rolling circle plasmids (pC194-type) commonly found in gram-positive bacteria. A pC194-type rolling circle plasmid has not been previously described in L. lactis. Further sequence analysis showed a conserved double-stranded origin of replication in pWC1 preceded by a large (118-bp) direct repeat. The chloramphenicol-resistance gene from pC194 was inserted into a nonessential region of pWC1 to give pCP12. The host range of pCP12 included Streptococcus thermophilus, Enterococcus faecalis, and Staphylococcus aureus, but not Escherichia coli. Both pCP12 and to a lesser extent pWC1 generated single-stranded DNA (ssDNA) in L. lactis. A possible single-stranded origin of replication was identified by sequence analysis of pWC1 and by comparing levels of ssDNA produced by pCP12 deletion derivatives. The pWC1 replicon may be a useful addition to other replicons currently available for vector construction. PMID:8700966

Pillidge, C J; Cambourn, W M; Pearce, L E

1996-03-01

364

RNA processing is involved in the post-transcriptional control of the citQRP operon from Lactococcus lactis biovar diacetylactis  

Microsoft Academic Search

The importance of Lactococcus lactis biovar diacetylactis (L. diacetylactis) in the dairy industry is due to its ability to produce aroma compounds, such as acetoin and diacetyl, from citrate. The\\u000a first step in citrate utilization is its uptake by the cells. In L. diacetylactis, the citrate transport system is encoded by the citQRP operon. We have previously proposed that expression

D. Drider; J. M. Santos; C. M. Arraiano; P. López

1998-01-01

365

In vitro inhibition of Citrobacter freundii, a red-leg syndrome associated pathogen in raniculture, by indigenous Lactococcus lactis CRL 1584.  

PubMed

Red-leg syndrome (RLS) is one of the main infectious diseases that cause economic losses in Lithobates catesbeianus hatcheries, Citrobacter freundii being an etiological agent. Treatment or prevention with therapeutics or chemicals results in modifications of the indigenous microbiota, development of antibiotic resistance, presence of their residues in food and enhancement of production costs. Thus, probiotics could be used as an alternative therapy. Lactic acid bacteria are part of the indigenous microbiota of healthy frogs and can prevent pathogen colonization by different mechanisms, including the production of antagonistic substances. In this work, the evaluation and characterization of the inhibition of C. freundii CFb by Lactococcus lactis subsp. lactis CRL 1584, a potentially probiotic candidate, were carried out. This strain produced lactic acid, H(2)O(2) and bacteriocin in static and shaken conditions and inhibited pathogen growth in associative cultures, with an earlier inhibition under agitated conditions. The elimination of each of the antimicrobial metabolites partially abolished the inhibition of the pathogen, suggesting that the inhibitory effect could be attributed to a combined action of the three antagonistic molecules. Electron microphotographs revealed the damage caused by L. lactis CRL 1584 supernatants to C. freundii cells. The addition of pure lactic acid, H(2)O(2) and bacteriocin to the culture media showed that each metabolite caused different morphological modifications in C. freundii, in agreement with the effect on viable cell counts. The results support the possibility that L. lactis CRL 1584 might be considered as a probiotic to be used in the prevention of RLS in raniculture. PMID:21531092

Pasteris, Sergio E; Guidoli, Marcos G; Otero, María C; Bühler, Marta I; Nader-Macías, María E

2011-08-01

366

Characterisation of the Poly(Vinylpyrrolidone)Poly(Vinylacetate-Co-Crotonic Acid) (PVP:PVAc-CA) Interpolymer Complex Matrix Microparticles Encapsulating a Bifidobacterium lactis Bb12 Probiotic Strain  

Microsoft Academic Search

The method of producing poly-(vinylpyrrolidone)-poly-(vinylacetate-co-crotonic acid) (PVP:PVAc-CA) interpolymer complex matrix\\u000a microparticles in supercritical carbon dioxide (scCO2), encapsulating bacteria, has recently been developed. This study was aimed at probing the external and internal structure\\u000a of these microparticles, which can be used in food. The encapsulation efficiency and distribution of encapsulated Bifidobacterium lactis Bb12 within these microparticles were also investigated. Scanning electron

C. I. Mamvura; F. S. Moolman; L. Kalombo; A. N. Hall; M. S. Thantsha

2011-01-01

367

In vitro adherence properties of Lactobacillus rhamnosus DR20 and Bifidobacterium lactis DR10 strains and their antagonistic activity against an enterotoxigenic Escherichia coli  

Microsoft Academic Search

Adhesion and colonisation properties of three probiotic strains namely, Lactobacillus rhamnosus DR20, L. acidophilus HN017, and Bifidobacterium lactis DR10, were determined in vitro using the differentiated human intestinal cell-lines including HT-29, Caco-2, and HT29-MTX, and compared with properties of L. acidophilus LA-1 and L. rhamnosus GG (two commercial probiotic strains). Two independent methods were employed to quantitate the “adhesiveness” of

Pramod K Gopal; Jaya Prasad; John Smart; Harsharanjit S Gill

2001-01-01

368

Enhancing immunity by dietary consumption of a probiotic lactic acid bacterium (Bifidobacterium lactis HN019): optimization and definition of cellular immune responses  

Microsoft Academic Search

Objective: To define the cellular basis for immune enhancement by a probiotic lactic acid bacteria strain (Bifidobacterium lactis HN019); and to determine whether immune enhancement can be optimized by delivery in oligosaccharide-enriched low-fat milk.Design: A double-blind, three-stage before-and-after intervention trial.Setting: Taipei Medical College Hospital, Taipei, Taiwan.Subjects: Fifty healthy Taiwanese citizens (age range 41–81; median 60) randomly allocated to two groups.Interventions:

BL Chiang; YH Sheih; LH Wang; CK Liao; HS Gill

2000-01-01

369

The SWI/SNF KlSnf2 Subunit Controls the Glucose Signaling Pathway To Coordinate Glycolysis and Glucose Transport in Kluyveromyces lactis  

PubMed Central

In Kluyveromyces lactis, the expression of the major glucose permease gene RAG1 is controlled by extracellular glucose through a signaling cascade similar to the Saccharomyces cerevisiae Snf3/Rgt2/Rgt1 pathway. We have identified a key component of the K. lactis glucose signaling pathway by characterizing a new mutation, rag20-1, which impairs the regulation of RAG1 and hexokinase RAG5 genes by glucose. Functional complementation of the rag20-1 mutation identified the KlSNF2 gene, which encodes a protein 59% identical to S. cerevisiae Snf2, the major subunit of the SWI/SNF chromatin remodeling complex. Reverse transcription-quantitative PCR and chromatin immunoprecipitation analyses confirmed that the KlSnf2 protein binds to RAG1 and RAG5 promoters and promotes the recruitment of the basic helix-loop-helix Sck1 activator. Besides this transcriptional effect, KlSnf2 is also implicated in the glucose signaling pathway by controlling Sms1 and KlRgt1 posttranscriptional modifications. When KlSnf2 is absent, Sms1 is not degraded in the presence of glucose, leading to constitutive RAG1 gene repression by KlRgt1. Our work points out the crucial role played by KlSnf2 in the regulation of glucose transport and metabolism in K. lactis, notably, by suggesting a link between chromatin remodeling and the glucose signaling pathway. PMID:23002104

Soulard, Alexandre; Wesolowski-Louvel, Micheline; Lemaire, Marc

2012-01-01

370

Effect of yogurt containing polydextrose, Lactobacillus acidophilus NCFM and Bifidobacterium lactis HN019: a randomized, double-blind, controlled study in chronic constipation  

PubMed Central

Background Constipation is a frequent complaint and the combination of a prebiotic and probiotics could have a potentially synergic effect on the intestinal transit. The present study therefore aims to investigate the combination of polydextrose (Litesse®), L. acidophilus NCFM® and B. lactis HN019 in a yogurt on intestinal transit in subjects who suffer from constipation. Methods Patients with constipation were randomly divided into two groups, Control Group (CG) and Treatment Group (TG), and had to eat 180 ml of unflavored yogurt every morning for 14 days. Those in the CG received only yogurt, while the TG received yogurt containing polydextrose, L. acidophilus NCFM® (ATCC 700396) and B. lactis HN019 (AGAL NM97/09513). Results Favourable clinical response was assessed since Agachan score had a significant reduction at the end of the study in both groups and tended to be better in the TG. The subjects in the treatment group also had a shorter transit time at the end of the intervention compared to the control group (p?=?0.01). Conclusion The product containing yogurt with polydextrose, B. lactis HN019 and L. acidophilus NCFM® significantly shortened colonic transit time after two weeks in the TG compared to CG and may be an option for treatment of constipation. PMID:25056655

2014-01-01

371

Co-evolution of segregation guide DNA motifs and the FtsK translocase in bacteria: identification of the atypical Lactococcus lactis KOPS motif.  

PubMed

Bacteria use the global bipolarization of their chromosomes into replichores to control the dynamics and segregation of their genome during the cell cycle. This involves the control of protein activities by recognition of specific short DNA motifs whose orientation along the chromosome is highly skewed. The KOPS motifs act in chromosome segregation by orienting the activity of the FtsK DNA translocase towards the terminal replichore junction. KOPS motifs have been identified in ?-Proteobacteria and in Bacillus subtilis as closely related G-rich octamers. We have identified the KOPS motif of Lactococcus lactis, a model bacteria of the Streptococcaceae family harbouring a compact and low GC% genome. This motif, 5'-GAAGAAG-3, was predicted in silico using the occurrence and skew characteristics of known KOPS motifs. We show that it is specifically recognized by L. lactis FtsK in vitro and controls its activity in vivo. L. lactis KOPS is thus an A-rich heptamer motif. Our results show that KOPS-controlled chromosome segregation is conserved in Streptococcaceae but that KOPS may show important variation in sequence and length between bacterial families. This suggests that FtsK adapts to its host genome by selecting motifs with convenient occurrence frequencies and orientation skews to orient its activity. PMID:22373923

Nolivos, Sophie; Touzain, Fabrice; Pages, Carine; Coddeville, Michele; Rousseau, Philippe; El Karoui, Meriem; Le Bourgeois, Pascal; Cornet, François

2012-07-01

372

Identification of Restriction-Modification Systems of Bifidobacterium animalis subsp. lactis CNCM I-2494 by SMRT Sequencing and Associated Methylome Analysis  

PubMed Central

Bifidobacterium animalis subsp. lactis CNCM I-2494 is a component of a commercialized fermented dairy product for which beneficial effects on health has been studied by clinical and preclinical trials. To date little is known about the molecular mechanisms that could explain the beneficial effects that bifidobacteria impart to the host. Restriction-modification (R-M) systems have been identified as key obstacles in the genetic accessibility of bifidobacteria, and circumventing these is a prerequisite to attaining a fundamental understanding of bifidobacterial attributes, including the genes that are responsible for health-promoting properties of this clinically and industrially important group of bacteria. The complete genome sequence of B. animalis subsp. lactis CNCM I-2494 is predicted to harbour the genetic determinants for two type II R-M systems, designated BanLI and BanLII. In order to investigate the functionality and specificity of these two putative R-M systems in B. animalis subsp. lactis CNCM I-2494, we employed PacBio SMRT sequencing with associated methylome analysis. In addition, the contribution of the identified R-M systems to the genetic accessibility of this strain was assessed. PMID:24743599

O?Connell Motherway, Mary; Watson, Debbie; Bottacini, Francesca; Clark, Tyson A.; Roberts, Richard J.; Korlach, Jonas; Garault, Peggy; Chervaux, Christian; van Hylckama Vlieg, Johan E. T.; Smokvina, Tamara; van Sinderen, Douwe

2014-01-01

373

Regulation of glycolysis and sugar phosphotransferase activities in Streptococcus lactis: growth in the presence of 2-deoxy-D-glucose.  

PubMed

Streptococcus lactis K1 has the capacity to grow on many sugars, including sucrose and lactose, in the presence of high levels (greater than 500 mM) of 2-deoxy-D-glucose. Initially, growth of the organism was transiently halted by the addition of comparatively low concentrations (less than 0.5 mM) of the glucose analog to the culture. Inhibition was coincident with (i) rapid accumulation of 2-deoxy-D-glucose 6-phosphate (ca. 120 mM) and preferential utilization of phosphoenolpyruvate via the mannose:phosphotransferase system, (ii) depletion of phosphorylated glycolytic intermediates, and (iii) a 60% reduction in intracellular ATP concentration. During the 5- to 10-min period of bacteriostasis the intracellular concentration of 2-deoxy-D-glucose 6-phosphate rapidly declined, and the concentrations of glycolytic intermediates were restored to near-normal levels. When growth resumed, the cell doubling time (Td) and the steady-state levels of 2-deoxy-D-glucose 6-phosphate maintained by the cells were dependent upon the medium concentration of 2-deoxy-D-glucose. Resistance of S. lactis K1 to the potentially toxic analog was a consequence of negative regulation of the mannose:phosphotransferase system by two independent mechanisms. The first, short-term response occurred immediately after the initial "overshoot" accumulation of 2-deoxy-D-glucose 6-phosphate, and this mechanism reduced the activity (fine control) of the mannose:phosphotransferase system. The second, long-term mechanism resulted in repression of synthesis (coarse control) of enzyme IImannose. The two regulatory mechanisms reduced the rate of 2-deoxy-D-glucose translocation via the mannose:phosphotransferase system and minimized the activity of the phosphoenolpyruvate-dependent futile cycle of the glucose analog (J. Thompson and B. M. Chassy, J. Bacteriol. 151:1454-1465, 1982). Phosphoenolpyruvate was thus conserved for transport of the growth sugar and for generation of ATP required for biosynthetic and work functions of the growing cell. PMID:6404888

Thompson, J; Chassy, B M

1983-05-01

374

Catabolism of Glucose and Lactose in Bifidobacterium animalis subsp. lactis, Studied by 13C Nuclear Magnetic Resonance  

PubMed Central

Bifidobacteria are widely used as probiotics in several commercial products; however, to date there is little knowledge about their carbohydrate metabolic pathways. In this work, we studied the metabolism of glucose and lactose in the widely used probiotic strain Bifidobacterium animalis subsp. lactis BB-12 by in vivo 13C nuclear magnetic resonance (NMR) spectroscopy. The metabolism of [1-13C]glucose was characterized in cells grown in glucose as the sole carbon source. Moreover, the metabolism of lactose specifically labeled with 13C on carbon 1 of the glucose or the galactose moiety was determined in suspensions of cells grown in lactose. These experiments allowed the quantification of some intermediate and end products of the metabolic pathways, as well as determination of the consumption rate of carbon sources. Additionally, the labeling patterns in metabolites derived from the metabolism of glucose specifically labeled with 13C on carbon 1, 2, or 3 in cells grown in glucose or lactose specifically labeled in carbon 1 of the glucose moiety ([1-13Cglucose]lactose), lactose specifically labeled in carbon 1 of the galactose moiety ([1-13Cgalactose]lactose), and [1-13C]glucose in lactose-grown cells were determined in cell extracts by 13C NMR. The NMR analysis showed that the recovery of carbon was fully compatible with the fructose 6-phosphate, or bifid, shunt. The activity of lactate dehydrogenase, acetate kinase, fructose 6-phosphate phosphoketolase, and pyruvate formate lyase differed significantly between glucose and lactose cultures. The transcriptional analysis of several putative glucose and lactose transporters showed a significant induction of Balat_0475 in the presence of lactose, suggesting a role for this protein as a lactose permease. This report provides the first in vivo experimental evidence of the metabolic flux distribution in the catabolic pathway of glucose and lactose in bifidobacteria and shows that the bifid shunt is the only pathway involved in energy recruitment from these two sugars. On the basis of our experimental results, a model of sugar metabolism in B. animalis subsp. lactis is proposed. PMID:24077711

Gonzalez-Rodriguez, Irene; Gaspar, Paula; Sanchez, Borja; Gueimonde, Miguel; Neves, Ana Rute

2013-01-01

375

The killer toxin of Kluyveromyces lactis: characterization of the toxin subunits and identification of the genes which encode them.  

PubMed Central

The killer character of the yeast Kluyveromyces lactis is associated with the presence of the linear DNA plasmids k1 and k2 and results from the secretion of a protein toxin into the growth medium. We find that toxin activity co-purifies with three polypeptides which we have termed the alpha- (mol. wt 99,000), beta- (mol. wt 30,000) and gamma- (mol. wt 27,500) subunits. The alpha-subunit appears to contain a single asparagine-linked oligosaccharide chain but neither of the smaller subunits is glycosylated. The N-terminal amino acid sequence of each subunit has been determined. Comparison of these data with the DNA sequence of plasmid k1 indicates that it encodes all three subunits. The alpha- and beta-subunits must be processed from the primary translation product of a single gene by an enzyme related to the KEX2 endopeptidase of Saccharomyces cerevisiae. Images Fig. 1. Fig. 2. Fig. 3. Fig. 4. PMID:3758030

Stark, M J; Boyd, A

1986-01-01

376

Characterization of ?-galacto-oligosaccharides formed via heterologous expression of ?-galactosidases from Lactobacillus reuteri in Lactococcus lactis.  

PubMed

?-Galacto-oligosaccharides (?-GOS) are produced by transgalactosylation reactions of ?-galactosidase (?-Gal) or by conversion of raffinose family oligosaccharides by levansucrase. Similarly to ?-GOS, ?-GOS have the potential to mimic glycan receptors on eukaryotic cells and act as molecular decoys to prevent bacterial infection; however, data on transgalactosylation reactions of ?-Gal remain scarce. The ?-Gal gene sequence from Lactobacillus reuteri was cloned into an ?-Gal negative strain of Lactococcus lactis. Transgalactosylation reactions were achieved using crude cell extracts with melibiose or raffinose as galactosyl donor and fucose, N-acetylglucosamine or lactose as galactosyl acceptor. The composition, sequence and most linkage types of ?-GOS formed with acceptors saccharides were determined by liquid chromatography-tandem mass spectrometry. ?-Gal of Lactobacillus reuteri formed (1???3)-, (1???4)- or (1???6)-linked ?-GOS but exhibited a preference for formation of (1???6)-linkages. Fucose, N-acetylglucosamine and lactose were suitable galactosyl acceptors for ?-Gal of L. reuteri, resulting in formation of (1???3)-, (1???4)- or (1???6)-linked hetero-oligosaccharides. By determining the structural specificity of ?-Gal and increasing the variation of oligosaccharides produced by introducing alternative acceptor sugars, this work supports further studies to assess ?-GOS pathogen adhesion prevention in mammalian hosts. PMID:23942880

Wang, Yvonne; Black, Brenna A; Curtis, Jonathan M; Gänzle, Michael G

2014-03-01

377

Immobilization of ?-d-galactosidase from Kluyveromyces lactis on functionalized silicon dioxide nanoparticles: characterization and lactose hydrolysis.  

PubMed

?-D-Galactosidase (BGAL) from Kluyveromyces lactis was covalently immobilized to functionalized silicon dioxide nanoparticles (10-20 nm). The binding of the enzyme to the nanoparticles was confirmed by Fourier transform-infrared spectroscopy (FTIR) and scanning electron microscopy (SEM). Functionalized nanoparticles showed 87% immobilization yield. Soluble and immobilized enzyme preparation exhibited pH-optima at pH 6.5 and 7.0, respectively, with temperature optima at 35 and 40°C, respectively. Michaelis constant (K(m)) was 4.77 and 8.4mM for free and immobilized BGAL, respectively. V(max) for the soluble and immobilized enzyme was 12.25 and 13.51 U/ml, respectively. Nanoparticle immobilized BGAL demonstrated improved stability after favoring multipoint covalent attachment. Thermal stability of the immobilized enzyme was enhanced at 40, 50 and 65°C. Immobilized nanoparticle-enzyme conjugate retained more than 50% enzyme activity up to the eleventh cycle. Maximum lactose hydrolysis by immobilized BGAL was achieved at 8h. PMID:22230612

Verma, Madan Lal; Barrow, Colin James; Kennedy, J F; Puri, Munish

2012-03-01

378

An alkali-thermostable xylanase from Bacillus pumilus functionally expressed in Kluyveromyces lactis and evaluation of its deinking efficiency.  

PubMed

This work aimed at studying the recombinant expression of an alkali- and thermo-stable xylanase from Bacillus pumilus in Kluyveromyces lactis and its use in deinking of civic paper waste. Efficient expression with a 3-fold increase in the activity than the native organism was achieved. An inducer concentration of 2.5% and medium pH of 9.0 was the best for enzyme expression. Purified enzyme showed an optimum activity at temperatures 50 and 60°C and pH 9.0 and 10.0, respectively. At pH 12.0, enzyme retained 74% and 26% activity after 2 and 3h of incubation, respectively. After incubation at 50 and 60°C for 1h, the enzyme showed 100% retention of activity, and remained active for 4h at 60°C retaining 23% residual activity. Partially purified recombinant enzyme showed higher deinking efficiency (273%) of laser print waste paper than crude xylanase from Bacillus and commercial acidic enzyme. This xylanase with superior stability characteristics could be a suitable candidate in paper and pulp industries. PMID:24709528

Thomas, Leya; Ushasree, Mrudula V; Pandey, Ashok

2014-08-01

379

Structure-Guided Engineering of Lactococcus lactis Alcohol Dehydrogenase LlAdhA for Improved Conversion of Isobutyraldehyde to Isobutanol  

PubMed Central

We have determined the X-ray crystal structures of the NADH-dependent alcohol dehydrogenase LlAdhA from Lactococcus lactis and its laboratory-evolved variant LlAdhARE1 at 1.9 Å and 2.5 Å resolution, respectively. LlAdhARE1, which contains three amino acid mutations (Y50F, I212T, and L264V), was engineered to increase the microbial production of isobutanol (2-methylpropan-1-ol) from isobutyraldehyde (2-methylpropanal). Structural comparison of LlAdhA and LlAdhARE1 indicates that the enhanced activity on isobutyraldehyde stems from increases in the protein’s active site size, hydrophobicity, and substrate access. Further structure-guided mutagenesis generated a quadruple mutant (Y50F/N110S/I212T/L264V), whose KM for isobutyraldehyde is ~17-fold lower and catalytic efficiency (kcat/KM) is ~160-fold higher than wild-type LlAdhA. Combining detailed structural information and directed evolution, we have achieved significant improvements in non-native alcohol dehydrogenase activity that will facilitate the production of next-generation fuels such as isobutanol from renewable resources. PMID:22974724

Liu, Xiang; Bastian, Sabine; Snow, Christopher D.; Brustad, Eric M.; Saleski, Tatyana E.; Xu, Jian-He; Meinhold, Peter; Arnold, Frances H.

2012-01-01

380

The Maltose ABC Transporter in Lactococcus lactis Facilitates High-Level Sensitivity to the Circular Bacteriocin Garvicin ML  

PubMed Central

We generated and characterized a series of spontaneous mutants of Lactococcus lactis IL1403 with average 6- to 11-fold-lowered sensitivities to the circular bacteriocin garvicin ML (GarML). Carbohydrate fermentation assays highlighted changes in carbohydrate metabolism, specifically loss of the ability to metabolize starch and maltose, in these mutants. PCR and sequencing showed that a 13.5-kb chromosomal deletion encompassing 12 open reading frames, mainly involved in starch and maltose utilization, had spontaneously occurred in the GarML-resistant mutants. Growth experiments revealed a correlation between sensitivity to GarML and carbon catabolite repression (CCR); i.e., sensitivity to GarML increased significantly when wild-type cells were grown on maltose and galactose as sole carbohydrates, an effect which was alleviated by the presence of glucose. Among the genes deleted in the mutants were malEFG, which encode a CCR-regulated membrane-bound maltose ABC transporter. The complementation of mutants with these three genes recovered normal sensitivity to the bacteriocin, suggesting an essential role of the maltose ABC transporter in the antimicrobial activity of GarML. This notion was supported by the fact that the level of sensitivity to GarML was dose dependent, increasing with higher expression levels of malEFG over a 50-fold range. To our knowledge, this is the first time a specific protein complex has been demonstrated to be involved in sensitivity to a circular bacteriocin. PMID:22411612

Gabrielsen, Christina; Brede, Dag A.; Hernandez, Pablo E.; Nes, Ingolf F.

2012-01-01

381

The Mitochondrial Genome Integrity Gene, Mgi1, of Kluyveromyces Lactis Encodes the ?-Subunit of F(1)-Atpase  

PubMed Central

In a previous report, we found that mutations at the mitochondrial genome integrity locus, MGI1, can convert Kluyveromyces lactis into a petite-positive yeast. In this report, we describe the isolation of the MGI1 gene and show that it encodes the ?-subunit of the mitochondrial F(1)-ATPase. The site of mutation in four independently isolated mgi1 alleles is at Arg435, which has changed to Gly in three cases and Ile in the fourth isolate. Disruption of MGI1 does not lead to the production of mitochondrial genome deletion mutants, indicating that an assembled F(1) complex is needed for the ``gain-of-function'' phenotype found in mgi1 point mutants. The location of Arg435 in the ?-subunit, as deduced from the three-dimensional structure of the bovine F(1)-ATPase, together with mutational sites in the previously identified mgi2 and mgi5 alleles, suggests that interaction of the ?- and ?- (MGI2) subunits with the ?-subunit (MGI5) is likely to be affected by the mutations. PMID:8978033

Chen, X. J.; Clark-Walker, G. D.

1996-01-01

382

Effects of Roundup(®) and glyphosate on three food microorganisms: Geotrichum candidum, Lactococcus lactis subsp. cremoris and Lactobacillus delbrueckii subsp. bulgaricus.  

PubMed

Use of many pesticide products poses the problem of their effects on environment and health. Amongst them, the effects of glyphosate with its adjuvants and its by-products are regularly discussed. The aim of the present study was to shed light on the real impact on biodiversity and ecosystems of Roundup(®), a major herbicide used worldwide, and the glyphosate it contains, by the study of their effects on growth and viability of microbial models, namely, on three food microorganisms (Geotrichum candidum, Lactococcus lactis subsp. cremoris and Lactobacillus delbrueckii subsp. bulgaricus) widely used as starters in traditional and industrial dairy technologies. The presented results evidence that Roundup(®) has an inhibitory effect on microbial growth and a microbicide effect at lower concentrations than those recommended in agriculture. Interestingly, glyphosate at these levels has no significant effect on the three studied microorganisms. Our work is consistent with previous studies which demonstrated that the toxic effect of glyphosate was amplified by its formulation adjuvants on different human cells and other eukaryotic models. Moreover, these results should be considered in the understanding of the loss of microbiodiversity and microbial concentration observed in raw milk for many years. PMID:22362186

Clair, Emilie; Linn, Laura; Travert, Carine; Amiel, Caroline; Séralini, Gilles-Eric; Panoff, Jean-Michel

2012-05-01

383

Engineering Lactococcus lactis for production of mannitol: high yields from food-grade strains deficient in lactate dehydrogenase and the mannitol transport system.  

PubMed

Mannitol is a sugar polyol claimed to have health-promoting properties. A mannitol-producing strain of Lactococcus lactis was obtained by disruption of two genes of the phosphoenolpyruvate (PEP)-mannitol phosphotransferase system (PTS(Mtl)). Genes mtlA and mtlF were independently deleted by double-crossover recombination in strain L. lactis FI9630 (a food-grade lactate dehydrogenase-deficient strain derived from MG1363), yielding two mutant (Delta ldh Delta mtlA and Delta ldh Delta mtlF) strains. The new strains, FI10091 and FI10089, respectively, do not possess any selection marker and are suitable for use in the food industry. The metabolism of glucose in nongrowing cell suspensions of the mutant strains was characterized by in vivo (13)C-nuclear magnetic resonance. The intermediate metabolite, mannitol-1-phosphate, accumulated intracellularly to high levels (up to 76 mM). Mannitol was a major end product, one-third of glucose being converted to this hexitol. The double mutants, in contrast to the parent strain, were unable to utilize mannitol even after glucose depletion, showing that mannitol was taken up exclusively by PEP-PTS(Mtl). Disruption of this system completely blocked mannitol transport in L. lactis, as intended. In addition to mannitol, approximately equimolar amounts of ethanol, 2,3-butanediol, and lactate were produced. A mixed-acid fermentation (formate, ethanol, and acetate) was also observed during growth under controlled conditions of pH and temperature, but mannitol production was low. The reasons for the alteration in the pattern of end products under nongrowing and growing conditions are discussed, and strategies to improve mannitol production during growth are proposed. PMID:15006767

Gaspar, Paula; Neves, Ana Rute; Ramos, Ana; Gasson, Michael J; Shearman, Claire A; Santos, Helena

2004-03-01

384

Altering renneting pH changes microstructure, cell distribution, and lysis of Lactococcus lactis AM2 in cheese made from ultrafiltered milk.  

PubMed

The objective of this study was to investigate the lysis of a highly autolytic strain of Lactococcus lactis ssp. cremoris AM2 in a model cheese made from concentrated ultrafiltered milk. From the same initial ultrafiltered retentate inoculated with L. lactis AM2, 5 cheeses were made by the addition of rennet at different pH values (6.6, 6.2, 5.8, 5.4, and 5.2). Lysis was monitored by measurement of the release of lactate dehydrogenase, an intracellular marker enzyme, and by immunodetection of intracellular proteins with species-specific antibodies. Confocal scanning laser microscopy (CSLM) was used to investigate the cheese microstructure by staining for protein and fat. Dual staining with a bacterial viability kit with CSLM was performed to reveal the integrity and localization of the bacterial cells. Levels of soluble calcium significantly increased when the pH at which the rennet was added decreased. In cheese renneted at pH 6.6, CSLM revealed an open porous structure containing a dense protein network with fat globules of different sizes distributed in the aqueous phase. In cheese renneted at pH 5.2, the protein network was homogeneous, with a less dense protein network, and an even distribution of fat globules. On d 1, bacterial cells were organized into colonies in cheese renneted at pH 6.6, whereas in cheeses renneted at pH 5.2, bacteria were evenly dispersed as single cells throughout the protein network. Lysis was detected on d 1 in cheeses renneted at high pH values and continued to increase throughout ripening, whereas induction of lysis was delayed in cheeses renneted at lower pH values until the end of ripening. This study demonstrates that alterations in the microstructure of the cheese and the distribution of cells play a role in lysis induction of L. lactis AM2. PMID:16507673

Hannon, J A; Lopez, C; Madec, M-N; Lortal, S

2006-03-01

385

Fructose Utilization in Lactococcus lactis as a Model for Low-GC Gram-Positive Bacteria: Its Regulator, Signal, and DNA-Binding Site  

PubMed Central

In addition to its role as carbon and energy source, fructose metabolism was reported to affect other cellular processes, such as biofilm formation by streptococci and bacterial pathogenicity in plants. Fructose genes encoding a 1-phosphofructokinase and a phosphotransferase system (PTS) fructose-specific enzyme IIABC component reside commonly in a gene cluster with a DeoR family regulator in various gram-positive bacteria. We present a comprehensive study of fructose metabolism in Lactococcus lactis, including a systematic study of fru mutants, global messenger analysis, and a molecular characterization of its regulation. The fru operon is regulated at the transcriptional level by both FruR and CcpA and at the metabolic level by inducer exclusion. The FruR effector is fructose-1-phosphate (F1P), as shown by combined analysis of transcription and measurements of the intracellular F1P pools in mutants either unable to produce this metabolite or accumulating it. The regulation of the fru operon by FruR requires four adjacent 10-bp direct repeats. The well-conserved organization of the fru promoter region in various low-GC gram-positive bacteria, including CRE boxes as well as the newly defined FruR motif, suggests that the regulation scheme defined in L. lactis could be applied to these bacteria. Transcriptome profiling of fruR and fruC mutants revealed that the effect of F1P and FruR regulation is limited to the fru operon in L. lactis. This result is enforced by the fact that no other targets for FruR were found in the available low-GC gram-positive bacteria genomes, suggesting that additional phenotypical effects due to fructose metabolism do not rely directly on FruR control, but rather on metabolism. PMID:15901699

Barriere, Charlotte; Veiga-da-Cunha, Maria; Pons, Nicolas; Guedon, Eric; van Hijum, Sacha A. F. T.; Kok, Jan; Kuipers, Oscar P.; Ehrlich, Dusko S.; Renault, Pierre

2005-01-01

386

?-Casein hydrolysate generated by the cell envelope-associated proteinase of Lactobacillus delbrueckii ssp. lactis CRL 581 protects against trinitrobenzene sulfonic acid-induced colitis in mice.  

PubMed

Lactobacillus delbrueckii ssp. lactis CRL 581, a thermophilic lactic acid bacterium used as a starter culture for the manufacture of several fermented dairy products, possesses an efficient proteolytic system that is able to release a series of potentially bioactive peptides (i.e., antihypertensive and phosphopeptides) from ?- and ?-caseins. Considering the potential beneficial health effects of the peptides released by L. delbrueckii ssp. lactis CRL 581 from milk proteins, the aim of this work was to analyze the anti-mutagenic and anti-inflammatory properties of the casein hydrolysates generated by the cell envelope-associated proteinase of this bacterium. The ability of ?- and ?-casein hydrolysates to suppress the mutagenesis of a direct-acting mutagen 4-nitroquinoline-N-oxide on Salmonella typhimurium TA 98 and TA 100 increased concomitantly with the time of casein hydrolysis. The anti-inflammatory effect of the ?-casein hydrolysate was evaluated using a trinitrobenzene sulfonic acid (TNBS)-induced Crohn's disease murine model. The hydrolysate was administered to mice 10 d before the intrarectal inoculation of TNBS. The mice that received ?-casein hydrolysate previously to TNBS showed decreased mortality rates, faster recovery of initial body weight loss, less microbial translocation to the liver, decreased ?-glucuronidase and myeloperoxidase activities in the gut, and decreased colonic macroscopic and microscopic damage compared with the animals that did not receive this hydrolysate. In addition, ?-casein hydrolysate exerted a beneficial effect on acute intestinal inflammation by increased interleukin 10 and decreased IFN-? production in the gut. Our findings are consistent with the health-promoting attributes of the milk products fermented by L. delbrueckii ssp. lactis CRL 581 and open up new opportunities for developing novel functional foods. PMID:22365194

Espeche Turbay, M B; de Moreno de LeBlanc, A; Perdigón, G; Savoy de Giori, G; Hebert, E M

2012-03-01

387

Identification and Functional Characterization of the Lactococcus lactis CodY-Regulated Branched-Chain Amino Acid Permease BcaP (CtrA)  

PubMed Central

Transcriptome analyses have previously revealed that a gene encoding the putative amino acid transporter CtrA (YhdG) is one of the major targets of the pleiotropic regulator CodY in Lactococcus lactis and Bacillus subtilis. The role of ctrA in L. lactis was further investigated with respect to both transport activity as well as CodY-mediated regulation. CtrA is required for optimal growth in media containing free amino acids as the only amino acid source. Amino acid transport studies showed that ctrA encodes a secondary amino acid transport system that is specific for branched-chain amino acids (BCAAs) (isoleucine, leucine, and valine) and methionine, which is in disagreement with its previously proposed function (a cationic amino acid transporter), which was assigned based on homology. We propose to rename CtrA BcaP, for branched-chain amino acid permease. BcaP is a member of a group of conserved transport systems, as homologs are widely distributed among gram-positive bacteria. Deletion of bcaP resulted in the loss of most of the BCAA uptake activity of L. lactis, indicating that BcaP is the major BCAA carrier of this organism. Deletion of bcaP together with a second (putative) BCAA permease, encoded by brnQ, further reduced the viability of the strain. DNA microarray analysis showed that deletion of bcaP predominantly affects genes belonging to the regulons of the transcriptional regulator CodY, which is involved in global nitrogen metabolism and needs BCAAs for its activation, and of CmbR, which is involved in sulfur amino acid metabolism. PMID:16621821

den Hengst, Chris D.; Groeneveld, Maarten; Kuipers, Oscar P.; Kok, Jan

2006-01-01

388

Reassessing the Role of Staphylococcus aureus Clumping Factor and Fibronectin-Binding Protein by Expression in Lactococcus lactis  

PubMed Central

Since Staphylococcus aureus expresses multiple pathogenic factors, studying their individual roles in single-gene-knockout mutants is difficult. To circumvent this problem, S. aureus clumping factor A (clfA) and fibronectin-binding protein A (fnbA) genes were constitutively expressed in poorly pathogenic Lactococcus lactis using the recently described pOri23 vector. The recombinant organisms were tested in vitro for their adherence to immobilized fibrinogen and fibronectin and in vivo for their ability to infect rats with catheter-induced aortic vegetations. In vitro, both clfA and fnbA increased the adherence of lactococci to their specific ligands to a similar extent as the S. aureus gene donor. In vivo, the minimum inoculum size producing endocarditis in ?80% of the rats (80% infective dose [ID80]) with the parent lactococcus was ?107 CFU. In contrast, clfA-expressing and fnbA-expressing lactococci required only 105 CFU to infect the majority of the animals (P < 0.00005). This was comparable to the infectivities of classical endocarditis pathogens such as S. aureus and streptococci (ID80 = 104 to 105 CFU) in this model. The results confirmed the role of clfA in endovascular infection, but with a much higher degree of confidence than with single-gene-inactivated staphylococci. Moreover, they identified fnbA as a critical virulence factor of equivalent importance. This was in contrast to previous studies that produced controversial results regarding this very determinant. Taken together, the present observations suggest that if antiadhesin therapy were to be developed, at least both of the clfA and fnbA products should be blocked for the therapy to be effective. PMID:11553573

Que, Yok-Ai; Francois, Patrice; Haefliger, Jacques-Antoine; Entenza, Jose-Manuel; Vaudaux, Pierre; Moreillon, Philippe

2001-01-01

389

Enhancement of Ad-CRT/E7-Mediated Antitumor Effect by Preimmunization with L. lactis Expressing HPV-16 E7.  

PubMed

Abstract Although current polyvalent vaccines can prevent development of cervical cancer, they cannot be used to treat patients who already have the disease. Adenovirus expressing calreticulin-E7 (Ad-CRT-E7) has shown promising results in the cervical cancer murine model. We also demonstrated that immunization with Lactococcus lactis encoding HPV-16 E7 (Ll-E7) anchored to its surface induces significant HPV-16 E7-specific immune response. Here, we assessed the combination of both approaches in the treatment of a cervical cancer animal model. Intranasal preimmunization of Ll-E7, followed by a single Ad-CRT/E7 application, induced ?80% of tumor suppression in comparison with controls. Mice treated with a combination of Ll-E7 and Ad-CRT/E7 resulted in a 70% survival rate 300 days post-treatment, whereas 100% of the mice in the control groups died by 50 days. Significant CD8+ cytotoxic T-lymphocytes infiltration was detected in the tumors of mice treated with Ll-E7+Ad-CRT/E7. Tumors with regression showed a greater number of positive cells for in situ TUNEL staining than controls. Our results suggest that preimmunization with Ll-E7 enhances the Ad-CRT/E7-mediated antitumor effect. This treatment provides an enormous advantage over repeated applications of Ad-CRT/E7 by maintaining the effectiveness of the three-dose application of Ad-CRT/E7, but avoiding the high systemic toxicities associated with such repeat treatments. PMID:25216057

Rangel-Colmenero, Blanca R; Gomez-Gutierrez, Jorge G; Villatoro-Hernández, Julio; Zavala-Flores, Laura M; Quistián-Martínez, Deyanira; Rojas-Martínez, Augusto; Arce-Mendoza, Alma Y; Guzmán-López, Santos; Montes-de-Oca-Luna, Roberto; Saucedo-Cárdenas, Odila

2014-11-01

390

Increased Biomass Yield of Lactococcus lactis by Reduced Overconsumption of Amino Acids and Increased Catalytic Activities of Enzymes  

PubMed Central

Steady state cultivation and multidimensional data analysis (metabolic fluxes, absolute proteome, and transcriptome) are used to identify parameters that control the increase in biomass yield of Lactococcus lactis from 0.10 to 0.12 C-mol C-mol?1 with an increase in specific growth rate by 5 times from 0.1 to 0.5 h?1. Reorganization of amino acid consumption was expressed by the inactivation of the arginine deiminase pathway at a specific growth rate of 0.35 h?1 followed by reduced over-consumption of pyruvate directed amino acids (asparagine, serine, threonine, alanine and cysteine) until almost all consumed amino acids were used only for protein synthesis at maximal specific growth rate. This balanced growth was characterized by a high glycolytic flux carrying up to 87% of the carbon flow and only amino acids that relate to nucleotide synthesis (glutamine, serine and asparagine) were consumed in higher amounts than required for cellular protein synthesis. Changes in the proteome were minor (mainly increase in the translation apparatus). Instead, the apparent catalytic activities of enzymes and ribosomes increased by 3.5 times (0.1 vs 0.5 h?1). The apparent catalytic activities of glycolytic enzymes and ribosomal proteins were seen to follow this regulation pattern while those of enzymes involved in nucleotide metabolism increased more than the specific growth rate (over 5.5 times). Nucleotide synthesis formed the most abundant biomonomer synthetic pathway in the cells with an expenditure of 6% from the total ATP required for biosynthesis. Due to the increase in apparent catalytic activity, ribosome translation was more efficient at higher growth rates as evidenced by a decrease of protein to mRNA ratios. All these effects resulted in a 30% decrease of calculated ATP spilling (0.1 vs 0.5 h?1). Our results show that bioprocesses can be made more efficient (using a balanced metabolism) by varying the growth conditions. PMID:23133574

Adamberg, Kaarel; Seiman, Andrus; Vilu, Raivo

2012-01-01

391

Influence of Cofermentation by Amylolytic Lactobacillus plantarum and Lactococcus lactis Strains on the Fermentation Process and Rheology of Sorghum Porridge  

PubMed Central

Amylolytic lactic acid bacteria (ALAB) can potentially replace malt in reducing the viscosity of starchy porridges. However, the drawback of using ALAB is their low and delayed amylolytic activity. This necessitates searching for efficient ALAB and strategies to improve their amylolytic activity. Two ALAB, Lactobacillus plantarum MNC 21 and Lactococcus lactis MNC 24, isolated from Obushera, were used to ferment starches in MRS broth: sorghum, millet, sweet potato, and commercial soluble starch. The amylolytic activity of MNC 21 was comparable to that of the ALAB collection strain Lb. plantarum A6, while that of MNC 24 was extremely low. MNC 21, MNC 24, and their coculture were compared to A6 and sorghum malt for ability to ferment and reduce the viscosity of sorghum porridge (11.6% dry matter). ALAB and the coculture lowered the pH from 6.2 to <4.5 within 12 h, while malt as a carrier of wild starter took about 20 h. Coculturing increased lactic acid yield by 46% and 76.8% compared to the yields of MNC 21 and MNC 24 monocultures, respectively. The coculture accumulated significantly larger (P < 0.05) amounts of maltose and diacetyl than the monocultures. Sorghum malt control and the coculture hydrolyzed more starch in sorghum porridge than the monocultures. The coculture initiated changes in the rheological parameters storage modulus (G?), loss modulus (G?), phase angle (?), and complex viscosity (?*) earlier than its constituent monocultures. The shear viscosity of sorghum porridge was reduced significantly (P < 0.05) from 1950 cP to 110 cP (malt), 281 cP (coculture), 382 cP (MNC 21), 713 cP (MNC 24), and 722 cP (A6). Coculturing strong ALAB with weak ALAB or non-ALAB can be exploited for preparation of nutrient-dense weaning foods and increasing lactic acid yield from starchy materials. PMID:22610432

Byaruhanga, Yusuf B.; Muyanja, Charles M. B. K.; Aijuka, Matthew; Schuller, Reidar B.; Sahlstr?m, Stefan; Langsrud, Thor; Narvhus, Judith A.

2012-01-01

392

Expression of avian influenza haemagglutinin (H5) and chicken interleukin 2 (chIL-2) under control of the ptcB promoter in Lactococcus lactis.  

PubMed

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. PMID:25273565

Szatraj, Katarzyna; Szczepankowska, Agnieszka K; S?czy?ska, Violetta; Florys, Katarzyna; Gromadzka, Beata; Lepek, Krzysztof; P?ucienniczak, Gra?yna; Szewczyk, Bogus?aw; Zagórski-Ostoja, W?odzimierz; Bardowski, Jacek

2014-01-01

393

D-Tagatose production in the presence of borate by resting Lactococcus lactis cells harboring Bifidobacterium longum L-arabinose isomerase.  

PubMed

Bifidobacterium longum NRRL B-41409 L-arabinose isomerase (L-AI) was overexpressed in Lactococcus lactis using a phosphate depletion inducible expression system. The resting L. lactis cells harboring the B. longum L-AI were used for production of D-tagatose from D-galactose in the presence of borate buffer. Multivariable analysis suggested that high pH, temperature and borate concentration favoured the conversion of D-galactose to D-tagatose. Almost quantitative conversion (92 %) was achieved at 20 g L?¹ substrate and at 37.5 °C after 5 days. The D-tagatose production rate of 185 g L?¹ day ?¹ was obtained at 300 g L?¹ galactose, at 1.15 M borate, and at 41 °C during 10 days when the production medium was changed every 24 h. There was no significant loss in productivity during ten sequential 24 h batches. The initial D-tagatose production rate was 290 g L?¹ day?¹ under these conditions. PMID:22903573

Salonen, Noora; Salonen, Kalle; Leisola, Matti; Nyyssölä, Antti

2013-04-01

394

Effect of yoghurt containing Bifidobacterium lactis Bb12® on faecal excretion of secretory immunoglobulin A and human beta-defensin 2 in healthy adult volunteers  

PubMed Central

Background Probiotics are used to provide health benefits. The present study tested the effect of a probiotic yoghurt on faecal output of beta-defensin and immunoglobulin A in a group of young healthy women eating a defined diet. Findings 26 women aged 18-21 (median 19) years residing in a hostel were given 200 ml normal yoghurt every day for a week, followed by probiotic yoghurt containing Bifidobacterium lactis Bb12® (109 in 200 ml) for three weeks, followed again by normal yoghurt for four weeks. Stool samples were collected at 0, 4 and 8 weeks and assayed for immunoglobulin A and human beta-defensin-2 by ELISA. All participants tolerated both normal and probiotic yoghurt well. Human beta-defensin-2 levels in faeces were not altered during the course of the study. On the other hand, compared to the basal sample, faecal IgA increased during probiotic feeding (P = 0.0184) and returned to normal after cessation of probiotic yoghurt intake. Conclusions Bifidobacterium lactis Bb12® increased secretory IgA output in faeces. This property may explain the ability of probiotics to prevent gastrointestinal and lower respiratory tract infections. PMID:22196482

2011-01-01

395

The crystal structures of Lactococcus lactis MG1363 Dps proteins reveal the presence of an N-terminal helix that is required for DNA binding.  

PubMed

Dps proteins play a major role in the protection of bacterial DNA from damage by reactive oxygen species. Previous studies have implicated the extended lysine-containing N-terminal regions of Dps subunits in DNA binding, but this part of the structure has not previously been observed crystallographically. Here the structures of two Dps proteins (DpsA and DpsB) from Lactococcus lactis MG1363 reveal for the first time the presence of an N-terminal alpha helix that extends from the core of the Dps subunit. Consequently, the N-terminal helices are displayed in parallel pairs on the exterior of the dodecameric Dps assemblies. Both DpsA and DpsB bind DNA. Deletion of the DpsA N-terminal helix impaired DNA binding. The N-terminal Lys residues of Escherichia coli Dps have been implicated in DNA binding. Replacement of the lactococcal DpsA Lys residues 9, 15 and 16 by Glu did not inhibit DNA binding. However, DNA binding was inhibited by EDTA, suggesting a role for cations in DNA binding. In contrast to E. coli, Bacillus brevis and Mycobacterium smegmatis Dps:DNA complexes, in which DNA interacts with crystalline Dps phases, L. lactis DNA:Dps complexes appeared as non-crystalline aggregates of protein and DNA in electron micrographs. PMID:16091047

Stillman, Timothy J; Upadhyay, Manisha; Norte, Valia A; Sedelnikova, Svetlana E; Carradus, Maria; Tzokov, Svetomir; Bullough, Per A; Shearman, Claire A; Gasson, Michael J; Williams, Colin H; Artymiuk, Peter J; Green, Jeffrey

2005-08-01

396

Microbial production of palatinose through extracellular expression of a sucrose isomerase from Enterobacter sp. FMB-1 in Lactococcus lactis MG1363.  

PubMed

Sucrose isomerase (SIase) has been used to produce palatinose, a structural isomer of sucrose, which has many beneficial health properties, such as low-glycemic and low-insulinemic indices. A gene corresponding to SIase from Enterobacter sp. FMB-1 was expressed in Lactococcus lactis MG1363 using the P170 expression system. The autoinducible promoter (P170) and an optimized signal peptide (SP310mut2) were used to induce and secrete SIase in L. lactis. One-step Ni-NTA affinity chromatography and Western blot analysis demonstrated that SIase was successfully secreted to the culture supernatant, although 60% of the recombinant enzymes were retained inside the cells. The production of the recombinant SIase was highly correlated with pH (pH 6) and glucose concentration (30g/L) of the medium. The extracellularly produced recombinant SIase was functionally active, effectively transforming 50g/L sucrose to 36g/L palatinose, with a conversion rate of 72% in the culture supernatant. PMID:20620050

Park, Jong-Yul; Jung, Jong-Hyun; Seo, Dong-Ho; Ha, Suk-Jin; Yoon, Jong-Won; Kim, Young-Cheul; Shim, Jae-Hoon; Park, Cheon-Seok

2010-11-01

397

Identification of a novel operon in Lactococcus lactis encoding three enzymes for lactic acid synthesis: phosphofructokinase, pyruvate kinase, and lactate dehydrogenase.  

PubMed Central

The discovery of a novel multicistronic operon that encodes phosphofructokinase, pyruvate kinase, and lactate dehydrogenase in the lactic acid bacterium Lactococcus lactis is reported. The three genes in the operon, designated pfk, pyk, and ldh, contain 340, 502, and 325 codons, respectively. The intergenic distances are 87 bp between pfk and pyk and 117 bp between pyk and ldh. Plasmids containing pfk and pyk conferred phosphofructokinase and pyruvate kinase activity, respectively, on their host. The identity of ldh was established previously by the same approach (R. M. Llanos, A. J. Hillier, and B. E. Davidson, J. Bacteriol. 174:6956-6964, 1992). Each of the genes is preceded by a potential ribosome binding site. The operon is expressed in a 4.1-kb transcript. The 5' end of the transcript was determined to be a G nucleotide positioned 81 bp upstream from the pfk start codon. The pattern of codon usage within the operon is highly biased, with 11 unused amino acid codons. This degree of bias suggests that the operon is highly expressed. The three proteins encoded on the operon are key enzymes in the Embden-Meyerhoff pathway, the central pathway of energy production and lactic acid synthesis in L. lactis. For this reason, we have called the operon the las (lactic acid synthesis) operon. Images PMID:8478320

Llanos, R M; Harris, C J; Hillier, A J; Davidson, B E

1993-01-01

398

Characterisation of thermotolerant cocci from indigenous flora of 'leben' in algerian arid area and DNA identification of atypical Lactococcus lactis strains.  

PubMed

Lactic acid bacteria (LAB) are widely used in food industry and their growth performance is important for the quality of the fermented product. By combining results from conventional isolation methods and molecular investigation of 16S rRNA gene and lactococcal/enterococcal specific genes, we identify at species level catalase negative gram positive thermoresistant cocci isolated from traditional 'leben', a 24-h fermented milk in arid area of west Algeria. Forty strains phenotypically related to cocci LAB were identified as belonging to the species Lactococcus lactis ssp. lactis, Enterococcus faecalis, Enterococcus faecium, and other Enterococcus species. No Streptococcus thermophilus strain was isolated. Ten different phenotype groups were recognized, and the species content of these groups were in some cases different from the expected features usually given in genus and species descriptions. In particular, atypical lactococci, able to grow in 6.5% NaCl, at pH 9.5 and showing high resistance to thermal stresses were isolated. Lactococci, but also enterococci isolated from traditional 'leben' produced in the desert area, may be therefore of interest in milk fermentation. Further studies to assess this source of diversity within the wild microbial population should provide starter new strains for product innovation. PMID:19484304

Bensalah, Farid; Delorme, Christine; Renault, P

2009-08-01

399

Effect of the Abortive Infection Mechanism and Type III Toxin/Antitoxin System AbiQ on the Lytic Cycle of Lactococcus lactis Phages  

PubMed Central

To survive in phage-containing environments, bacteria have evolved an array of antiphage systems. Similarly, phages have overcome these hurdles through various means. Here, we investigated how phages are able to circumvent the Lactococcus lactis AbiQ system, a type III toxin-antitoxin with antiviral activities. Lactococcal phage escape mutants were obtained in the laboratory, and their genomes were sequenced. Three unrelated genes of unknown function were mutated in derivatives of three distinct lactococcal siphophages: orf38 of phage P008, m1 of phage bIL170, and e19 of phage c2. One-step growth curve experiments revealed that the phage mutations had a fitness cost while transcriptional analyses showed that AbiQ modified the early-expressed phage mRNA profiles. The L. lactis AbiQ system was also transferred into Escherichia coli MG1655 and tested against several coliphages. While AbiQ was efficient against phages T4 (Myoviridae) and T5 (Siphoviridae), escape mutants of only phage 2 (Myoviridae) could be isolated. Genome sequencing revealed a mutation in gene orf210, a putative DNA polymerase. Taking these observations together, different phage genes or gene products are targeted or involved in the AbiQ phenotype. Moreover, this antiviral system is active against various phage families infecting Gram-positive and Gram-negative bacteria. A model for the mode of action of AbiQ is proposed. PMID:23813728

Samson, Julie E.; Belanger, Maxime

2013-01-01

400

Lignes directrices canadiennes sur la rhinosinusite bact?rienne aigu?  

PubMed Central

Résumé Objectif Faire un résumé clinique des lignes directrices canadiennes sur la rhinosinusite bactérienne aiguë (RSBA) qui présente des éléments d’intérêt pour les médecins de famille. Source des données Les auteurs des lignes directrices ont effectué une recherche documentaire systématique et ont rédigé des recommandations. Une cote a été donnée à la fois en fonction de la fiabilité des données probantes et de la solidité des recommandations. On a sollicité les commentaires d’experts en la matière venant de l’extérieur, ainsi que l’aval de sociétés médicales canadiennes (Association pour la microbiologie médicale et l’infectiologie Canada, Société canadienne d’allergie et d’immunologie clinique, Société canadienne d’otorhinolaryngologie et de chirurgie cervicofaciale, Association canadienne des médecins d’urgence et Regroupement canadien des médecins de famille en santé respiratoire). Message principal Le diagnostic de la RSBA repose sur la présence de symptômes particuliers et leur durée; l’imagerie ou une culture n’est pas nécessaire dans les cas peu compliqués. Le traitement dépend de la gravité des symptômes, notamment avec des corticostéroïdes intranasaux (CSIN) recommandés comme monothérapie pour les cas de légers à modérés, quoique leurs bienfaits soient modestes. Le recours à des CSIN accompagnés d’antibiotiques est réservé aux patients qui ne répondent pas aux CSIN après 72 heures et comme traitement initial des patients dont les symptômes sont graves. Le choix de l’antibiotique doit tenir compte du pathogène soupçonné, du risque de résistance, des problèmes concomitants et des tendances locales de la résistance aux antimicrobiens. Des thérapies d’appoint comme l’irrigation nasale avec une solution saline sont recommandées. En présence de cas réfractaires au traitement, d’épisodes récurrents et de signes de complications, on devrait demander une consultation en otorhinolaryngologie. Les lignes directrices portent sur les situations particulières à l’environnement canadien des soins de santé, y compris les actions à prendre durant les périodes d’attente prolongées pour avoir une consultation avec un spécialiste ou une imagerie. Conclusion Les lignes directrices canadiennes offrent des recommandations à jour pour le diagnostic et le traitement de la RSBA qui tiennent compte de la compréhension en évolution de la maladie. De plus, les lignes directrices présentent des outils utiles pour aider les cliniciens à cerner les épisodes viraux par opposition à ceux d’origine bactérienne, ainsi qu’à prendre en charge de manière optimale leurs patients atteints de RSBA.

Kaplan, Alan

2014-01-01

401

Effect of the consumption of a fermented dairy product containing Bifidobacterium lactis DN-173 010 on constipation in childhood: a multicentre randomised controlled trial (NTRTC: 1571)  

PubMed Central

Background Constipation is a frustrating symptom affecting 3% of children worldwide. Randomised controlled trials show that both polyethylene glycol and lactulose are effective in increasing defecation frequency in children with constipation. However, in 30–50%, these children reported abdominal pain, bloating, flatulence, diarrhoea, nausea and bad taste of the medication. Two recent studies have shown that the fermented dairy product containing Bifidobacterium lactis strain DN-173 010 is effective in increasing stool frequency in constipation-predominant irritable bowel syndrome patients with a defecation frequency < 3/week and in constipated women with a defecation frequency < 3/week. Goal of this study is to determine whether this fermented dairy product is effective in the treatment of constipated children with a defecation frequency < 3/week. Methods/design It is a two nation (The Netherlands and Poland) double-blind, placebo-controlled randomised multicentre trial in which 160 constipated children (age 3–16 years) with a defecation frequency <3/week will be randomly allocated to consume a fermented dairy product containing Bifidobacterium lactis DN-173 010 or a control product, twice a day, for 3 weeks. During the study all children are instructed to try to defecate on the toilet for 5–10 minutes after each meal (3 times a day) and daily complete a standardized bowel diary. Primary endpoint is stool frequency. Secondary endpoints are stool consistency, faecal incontinence frequency, pain during defecation, digestive symptoms (abdominal pain, flatulence), adverse effects (nausea, diarrhoea, bad taste) and intake of rescue medication (Bisacodyl). Rate of success and rate of responders are also evaluated, with success defined as ? 3 bowel movements per week and ?1 faecal incontinence episode over the last 2 weeks of product consumption and responder defined as a subject reporting a stool frequency ? 3 on the last week of product consumption. To demonstrate that the success percentage in the intervention group will be 35% and the success percentage in the control group (acidified milk without ferments, toilet training, bowel diary) will be 15%, with alpha 0.05 and power 80%, a total sample size of 160 patients was calculated. Conclusion This study is aimed to show that the fermented dairy product containing Bifidobacterium lactis strain DN-173 010 is effective in increasing stool frequency after 3 weeks of product consumption in children with functional constipation and a defecation frequency < 3/week. PMID:19296845

Tabbers, Merit M; Chmielewska, Ania; Roseboom, Maaike G; Boudet, Claire; Perrin, Catherine; Szajewska, Hania; Benninga, Marc A

2009-01-01

402

Cloning and characterization of the KlDIM1 gene from Kluyveromyces lactis encoding the m2(6)A dimethylase of the 18S rRNA.  

PubMed

The KlDIM1 gene encoding the m2(6)A rRNA dimethylase was cloned from a Kluyveromyces lactis genomic library using a PCR amplicon from the Saccharomyces cerevisiae ScDIM1 gene as probe. The KlDIM1 gene encodes a 320-amino acid protein which shows 81% identity to ScDim1p from S. cerevisiae and 25% identity to ksgAp from Escherichia coli. Complementation of the kasugamycin-resistant ksgA-mutant of E. coli lacking dimethylase activity demonstrates that KlDim1p is the functional homologue of the bacterial enzyme. Multiple alignment of dimethylases from prokaryotes and yeasts shows that the two yeast enzymes display distinctive structural motives including a putative nuclear localization signal. PMID:9219342

Housen, I; Demonté, D; Lafontaine, D; Vandenhaute, J

1997-06-30

403

Degradation and debittering of a tryptic digest from beta-casein by aminopeptidase N from Lactococcus lactis subsp. cremoris Wg2.  

PubMed Central

The mode of action of purified aminopeptidase N from Lactococcus lactis subsp. cremoris Wg2 on a complex peptide mixture of a tryptic digest from bovine beta-casein was analyzed. The oligopeptides produced in the tryptic digest before and after aminopeptidase N treatment were identified by analysis of the N- and C-terminal amino acid sequences and amino acid compositions of the isolated peptides and by on-line liquid chromatography-mass spectrometry. Incubation of purified peptides with aminopeptidase N resulted in complete hydrolysis of many peptides, while others were only partially hydrolyzed or not hydrolyzed. The tryptic digest of beta-casein exhibits a strong bitter taste, which corresponds to the strong hydrophobicity of several peptides in the tryptic digest of beta-casein. The degradation of the "bitter" tryptic digest by aminopeptidase N resulted in a decrease of hydrophobic peptides and a drastic decrease of bitterness of the reaction mixture. PMID:8100130

Tan, P S; van Kessel, T A; van de Veerdonk, F L; Zuurendonk, P F; Bruins, A P; Konings, W N

1993-01-01

404

Inhibition kinetics of catabolic dehydrogenases by elevated moieties of ATP and ADP--implication for a new regulation mechanism in Lactococcus lactis.  

PubMed

ATP and ADP inhibit, in varying degrees, several dehydrogenases of the central carbon metabolism of Lactococcus lactis ATCC 19435 in vitro, i.e. glyceraldehyde-3-phosphate dehydrogenase (GAPDH), lactate dehydrogenase (LDH) and alcohol dehydrogenase (ADH). Here we demonstrate mixed inhibition for GAPDH and competitive inhibition for LDH and ADH by adenine nucleotides in single inhibition studies. The nonlinear negative co-operativity was best modelled with Hill-type kinetics, showing greater flexibility than the usual parabolic inhibition equation. Because these natural inhibitors are present simultaneously in the cytoplasm, multiple inhibition kinetics was determined for each dehydrogenase. For ADH and LDH, the inhibitor combinations ATP plus NAD and ADP plus NAD are indifferent to each other. Model discrimination suggested that the weak allosteric inhibition of GAPDH had no relevance when multiple inhibitors are present. Interestingly, with ADH and GAPDH the combination of ATP and ADP exhibits lower dissociation constants than with either inhibitor alone. Moreover, the concerted inhibition of ADH and GAPDH, but not of LDH, shows synergy between the two nucleotides. Similar kinetics, but without synergies, were found for horse liver and yeast ADHs, indicating that dehydrogenases can be modulated by these nucleotides in a nonlinear manner in many organisms. The action of an elevated pool of ATP and ADP may effectively inactivate lactococcal ADH, but not GAPDH and LDH, providing leverage for the observed metabolic shift to homolactic acid formation in lactococcal resting cells on maltose. Therefore, we interpret these results as a regulation mechanism contributing to readjusting the flux of ATP production in L. lactis. PMID:20193044

Cao, Rong; Zeidan, Ahmad A; Rådström, Peter; van Niel, Ed W J

2010-04-01

405

Isolation of a bacteriocin-producing lactococcus lactis and application of its bacteriocin to manage spoilage bacteria in high-value marine fish under different storage temperatures.  

PubMed

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?lactis PSY2 gave increased protection against spoilage bacteria and offers an alternative for the preservation of high-value sea foods. PMID:22555500

Sarika, A R; Lipton, A P; Aishwarya, M S; Dhivya, R S

2012-07-01

406

Phages of non-dairy lactococci: isolation and characterization of ?L47, a phage infecting the grass isolate Lactococcus lactis ssp. cremoris DPC6860.  

PubMed

Lactococci isolated from non-dairy sources have been found to possess enhanced metabolic activity when compared to dairy strains. These capabilities may be harnessed through the use of these strains as starter or adjunct cultures to produce more diverse flavor profiles in cheese and other dairy products. To understand the interactions between these organisms and the phages that infect them, a number of phages were isolated against lactococcal strains of non-dairy origin. One such phage, ?L47, was isolated from a sewage sample using the grass isolate L. lactis ssp. cremoris DPC6860 as a host. Visualization of phage virions by transmission electron microscopy established that this phage belongs to the family Siphoviridae and possesses a long tail fiber, previously unseen in dairy lactococcal phages. Determination of the lytic spectrum revealed a broader than expected host range, with ?L47 capable of infecting 4 industrial dairy strains, including ML8, HP and 310, and 3 additional non-dairy isolates. Whole genome sequencing of ?L47 revealed a dsDNA genome of 128, 546 bp, making it the largest sequenced lactococcal phage to date. In total, 190 open reading frames (ORFs) were identified, and comparative analysis revealed that the predicted products of 117 of these ORFs shared greater than 50% amino acid identity with those of L. lactis phage ?949, a phage isolated from cheese whey. Despite their different ecological niches, the genomic content and organization of ?L47 and ?949 are quite similar, with both containing 4 gene clusters oriented in different transcriptional directions. Other features that distinguish ?L47 from ?949 and other lactococcal phages, in addition to the presence of the tail fiber and the genome length, include a low GC content (32.5%) and a high number of predicted tRNA genes (8). Comparative genome analysis supports the conclusion that ?L47 is a new member of the 949 lactococcal phage group which currently includes the dairy ?949. PMID:24454309

Cavanagh, Daniel; Guinane, Caitriona M; Neve, Horst; Coffey, Aidan; Ross, R Paul; Fitzgerald, Gerald F; McAuliffe, Olivia

2014-01-13

407

Phages of non-dairy lactococci: isolation and characterization of ?L47, a phage infecting the grass isolate Lactococcus lactis ssp. cremoris DPC6860  

PubMed Central

Lactococci isolated from non-dairy sources have been found to possess enhanced metabolic activity when compared to dairy strains. These capabilities may be harnessed through the use of these strains as starter or adjunct cultures to produce more diverse flavor profiles in cheese and other dairy products. To understand the interactions between these organisms and the phages that infect them, a number of phages were isolated against lactococcal strains of non-dairy origin. One such phage, ?L47, was isolated from a sewage sample using the grass isolate L. lactis ssp. cremoris DPC6860 as a host. Visualization of phage virions by transmission electron microscopy established that this phage belongs to the family Siphoviridae and possesses a long tail fiber, previously unseen in dairy lactococcal phages. Determination of the lytic spectrum revealed a broader than expected host range, with ?L47 capable of infecting 4 industrial dairy strains, including ML8, HP and 310, and 3 additional non-dairy isolates. Whole genome sequencing of ?L47 revealed a dsDNA genome of 128, 546 bp, making it the largest sequenced lactococcal phage to date. In total, 190 open reading frames (ORFs) were identified, and comparative analysis revealed that the predicted products of 117 of these ORFs shared greater than 50% amino acid identity with those of L. lactis phage ?949, a phage isolated from cheese whey. Despite their different ecological niches, the genomic content and organization of ?L47 and ?949 are quite similar, with both containing 4 gene clusters oriented in different transcriptional directions. Other features that distinguish ?L47 from ?949 and other lactococcal phages, in addition to the presence of the tail fiber and the genome length, include a low GC content (32.5%) and a high number of predicted tRNA genes (8). Comparative genome analysis supports the conclusion that ?L47 is a new member of the 949 lactococcal phage group which currently includes the dairy ?949. PMID:24454309

Cavanagh, Daniel; Guinane, Caitriona M.; Neve, Horst; Coffey, Aidan; Ross, R. Paul; Fitzgerald, Gerald F.; McAuliffe, Olivia

2014-01-01

408

AcmD, a Homolog of the Major Autolysin AcmA of Lactococcus lactis, Binds to the Cell Wall and Contributes to Cell Separation and Autolysis  

PubMed Central

Lactococcus lactis expresses the homologous glucosaminidases AcmB, AcmC, AcmA and AcmD. The latter two have three C-terminal LysM repeats for peptidoglycan binding. AcmD has much shorter intervening sequences separating the LysM repeats and a lower iso-electric point (4.3) than AcmA (10.3). Under standard laboratory conditions AcmD was mainly secreted into the culture supernatant. An L. lactis acmAacmD double mutant formed longer chains than the acmA single mutant, indicating that AcmD contributes to cell separation. This phenotype could be complemented by plasmid-encoded expression of AcmD in the double mutant. No clear difference in cellular lysis and protein secretion was observed between both mutants. Nevertheless, overexpression of AcmD resulted in increased autolysis when AcmA was present (as in the wild type strain) or when AcmA was added to the culture medium of an AcmA-minus strain. Possibly, AcmD is mainly active within the cell wall, at places where proper conditions are present for its binding and catalytic activity. Various fusion proteins carrying either the three LysM repeats of AcmA or AcmD were used to study and compare their cell wall binding characteristics. Whereas binding of the LysM domain of AcmA took place at pHs ranging from 4 to 8, LysM domain of AcmD seems to bind strongest at pH 4. PMID:23951292

Visweswaran, Ganesh Ram R.; Steen, Anton; Leenhouts, Kees; Szeliga, Monika; Ruban, Beata; Hesseling-Meinders, Anne; Dijkstra, Bauke W.; Kuipers, Oscar P.; Kok, Jan; Buist, Girbe

2013-01-01

409

Epidermal growth factor-expressing Lactococcus lactis enhances growth performance of early-weaned pigs fed diets devoid of blood plasma.  

PubMed

The effect of supplementing Lactococcus lactis (L. lactis) that was engineered to express epidermal growth factor (EGF-LL) to early-weaned pigs fed diets with typical levels of blood plasma (5%) or diets without blood plasma [blood plasma was substituted with soybean (Glycine max) meal and fish meal, based on amino acid supply] was examined. A total of 108 weaned piglets (19-26 d of age; mean initial BW 6.58 kg; 9 pigs per pen) were fed ad libitum according to a 2-phase feeding program without growth promoters. Three pens were assigned to each of 4 treatments: i) blood plasma-containing diet with blank bacterial growth medium (BP-Con), ii) blood plasma-containing diet with fermented EGF-LL (BP-EGF), iii) blood plasma-free diet with blank bacterial growth medium (BPF-Con), and iv) blood plasma-free diet with fermented EGF-LL (BPF-EGF). The amount of epidermal growth factor (EGF) was determined in the fermentation product and pigs were allotted 60 ?g EGF/kg BW/d for 3 wk postweaning. There were no differences in overall growth performance between BP-Con and BP-EGF pigs and no differences in overall growth performance between LoCon and BPF-EGF pigs. Pigs fed BPF-EGF showed increased daily BW gain (410 vs. 260 g/d; P < 0.01) and gain:feed (0.67 vs. 0.58; P < 0.05) compared to BPF-Con pigs in wk 3 postweaning; this was comparable to values for the BP-Con group (400 g/d and 0.64). These results indicate that supplementation with EGF-LL can be effective in enhancing the performance of early-weaned piglets fed a low complexity diet and reduces the need for feeding high-quality animal proteins and antibiotics. PMID:23365266

Bedford, A; Li, Z; Li, M; Ji, S; Liu, W; Huai, Y; de Lange, C F M; Li, J

2012-12-01

410

Induction of antigen-specific tolerance by oral administration of Lactococcus lactis delivered immunodominant DQ8-restricted Gliadin peptide in sensitized NOD AB? DQ8 transgenic mice  

PubMed Central

Active delivery of recombinant autoantigens or allergens at the intestinal mucosa by genetically modified Lactococcus lactis (LL) provides a novel therapeutic approach for the induction of tolerance. Celiac disease is associated with either HLA-DQ2 or HLA-DQ8 restricted responses to specific antigenic epitopes of gliadin, and may be treated by induction of antigen-specific tolerance. We investigated whether oral administration of LL-delivered DQ8-specific gliadin epitope induces antigen-specific tolerance. L. lactis was engineered to secrete a deamidated DQ8 gliadin epitope (LL-eDQ8d) and the induction of antigen-specific tolerance was studied in NOD AB° DQ8 transgenic mice. Tolerance was assessed by delayed-type hypersensitivity reaction, cytokine measurements, eDQ8d-specific proliferation and regulatory T cell analysis. Oral administration of LL-eDQ8d induced suppression of local and systemic DQ8 restricted T-cell responses in NOD AB° DQ8 transgenic mice. Treatment resulted in an antigen-specific decrease of the proliferative capacity of inguinal lymph node cells and lamina propria cells. Production of IL-10 and TGF-? and a significant induction of Foxp3+ regulatory T-cells were associated with the eDQ8d-specific suppression induced by LL-eDQ8d. These data provide support for the development of effective therapeutic approaches for gluten-sensitive disorders using orally administered antigen-secreting LL. Such treatments may be effective even in the setting of established hypersensitivity. PMID:19635921

Huibregtse, Inge L.; Marietta, Eric V.; Rashtak, Shadi; Koning, Frits; Rottiers, Pieter; David, Chella S.; van Deventer, Sander J.H.; Murray, Joseph A.

2012-01-01

411

Lait (1998) 78,165-171 Inra/Elsevier, Paris  

E-print Network

lactis stress genes already known with a vegetative promoter and a CIRCE sequence. ln O. oeni-cristalline #12;166 M.P. Jobin et al. 1. INTRODUCTION Les bactéries lactiques sont capables de fermenter les dans le vin, O. oeni réalise la fermentation malolactique (FML) qui est la décarboxylation du L

Paris-Sud XI, Université de

412

Oral delivery of glutamic acid decarboxylase (GAD)-65 and IL10 by Lactococcus lactis reverses diabetes in recent-onset NOD mice.  

PubMed

Growing insight into the pathogenesis of type 1 diabetes (T1D) and numerous studies in preclinical models highlight the potential of antigen-specific approaches to restore tolerance efficiently and safely. Oral administration of protein antigens is a preferred method for tolerance induction, but degradation during gastrointestinal passage can impede such protein-based therapies, reducing their efficacy and making them cost-ineffective. To overcome these limitations, we generated a tolerogenic bacterial delivery technology based on live Lactococcus lactis (LL) bacteria for controlled secretion of the T1D autoantigen GAD65370-575 and the anti-inflammatory cytokine interleukin-10 in the gut. In combination with short-course low-dose anti-CD3, this treatment stabilized insulitis, preserved functional ?-cell mass, and restored normoglycemia in recent-onset NOD mice, even when hyperglycemia was severe at diagnosis. Combination therapy did not eliminate pathogenic effector T cells, but increased the presence of functional CD4(+)Foxp3(+)CD25(+) regulatory T cells. These preclinical data indicate a great therapeutic potential of orally administered autoantigen-secreting LL for tolerance induction in T1D. PMID:24677716

Robert, Sofie; Gysemans, Conny; Takiishi, Tatiana; Korf, Hannelie; Spagnuolo, Isabella; Sebastiani, Guido; Van Huynegem, Karolien; Steidler, Lothar; Caluwaerts, Silvia; Demetter, Pieter; Wasserfall, Clive H; Atkinson, Mark A; Dotta, Francesco; Rottiers, Pieter; Van Belle, Tom L; Mathieu, Chantal

2014-08-01

413

Effects of ingesting milk fermented by Lactococcus lactis H61 on skin health in young women: A randomized double-blind study.  

PubMed

We conducted a randomized double-blind trial to evaluate the effects of fermented milk produced using only Lactococcus lactis strain H61 as a starter bacterium (H61-fermented milk) on the general health and various skin properties of young women. Healthy female volunteers (n=23; age=19-21r) received H61-fermented milk (10(10) cfu of strain H61/d) or conventional yogurt (10(10) cfu of both Lactobacillus delbrueckii ssp. bulgaricus and Streptococcus thermophilus per day), as a reference food, daily for 4 wk. Before and at the end of 4 wk, blood samples were taken, and skin hydration (inner forearms and cheek) and melanin content, elasticity, and sebum content (cheek only) were measured. Skin hydration at the inner forearm was higher at wk 4 than at wk 0 in both groups. Sebum content in cheek rose significantly after intervention in the H61-fermented milk group, but not the conventional yogurt group. Other skin parameters did not differ in either group. Serum analysis showed that total protein concentration and platelet count were elevated and reactive oxygen species decreased in both groups after the intervention. Although H61-fermented milk and conventional yogurt had similar effects on skin status and some blood characteristics of participants, an increase of sebum content in cheek is preferable to H61-fermented milk. As skin lipids contribute to maintaining the skin barrier, H61-fermented milk would provide beneficial effects on skin for young women. PMID:25022690

Kimoto-Nira, H; Nagakura, Y; Kodama, C; Shimizu, T; Okuta, M; Sasaki, K; Koikawa, N; Sakuraba, K; Suzuki, C; Suzuki, Y

2014-09-01

414

Charged residues on a flap-loop structure of Lactococcus lactis prolidase play critical roles in allosteric behavior and substrate inhibition.  

PubMed

Allosteric behavior and substrate inhibition are unique characteristics of Lactococcus lactis prolidase. We hypothesized that charged residues (Asp36, His38, Glu39, and Arg40), present on one loop essential for catalysis, interact with residues in or near the active site to impart these unique characteristics. Asp36 has a predominant role in the allosteric behavior, as demonstrated through the non-allosteric behavior of the D36S mutant enzyme. In contrast, a double mutant (D36E/R293K) maintained the allostery, indicating that this aspartic acid residue interacts with Arg293, previously shown to be critical in the allostery. Substitution of His38 drastically reduced the substrate inhibition, and substrate specificity of the mutant at Asp36 or His38 showed the influence of these residues to the substrate specificity. These findings confirm the importance of the loop in the enzymatic reaction mechanism and suggest the existence of conformational changes of the loop structure between open and closed states. A variety of mutations at Glu39 and Arg40 showed that these residues influence roles of the loop in the enzyme reaction. On the basis of these results and combined with observations of molecular models of this prolidase, we concluded that Asp36 and His38 interact with the residues in the active site to generate an allosteric subsite and a pseudo-S(1)' site, which are responsible for the allosteric behavior and substrate inhibition. PMID:21875695

Chen, Jian An; Tanaka, Takuji

2011-12-01