Luis M Cintas

Complutense University of Madrid, Madrid, Madrid, Spain

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Publications (64)171.42 Total impact

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    ABSTRACT: Analysis of 1245 lactic acid bacteria (LAB) isolates obtained from fish, seafood and fish products showed that 197 exerted direct antimicrobial activity against 20 spoilage and food-borne pathogenic microorganisms. Further evaluation of 64 LAB isolates selected on the basis of their direct antimicrobial activity revealed 25 secreted bacteriocins. Biochemical characterization, PCR analysis and/or DNA sequencing of the superoxide dismutase gene (sodA) and/or 16S rRNA gene (16S rDNA), and/or SDS-PAGE analyses of the 64 selected isolates allowed the identification of 24 of them as Enterococcus faecium, 22 as Enterococcus faecalis, seven as Pediococcus pentosaceus, five as Weissella cibaria, three as Lactobacillus sakei subsp. carnosus, one as L. sakei subsp. sakei, one as Lactobacillus curvatus subsp. curvatus and one as Leuconostoc mesenteroides subsp. cremoris. PCR analyses for the detection of genes encoding previously described bacteriocins performed on the 25 bacteriocinogenic strains showed that 19 strains (18 enterococci and 1 lactobacilli) amplified, at least, one of the tested genes, and up to four and two target genes were simultaneously detected in a single enterococcal and lactobacilli strain, respectively. Moreover, in vitro safety evaluation of E. faecium strains was carried out by detection of potential virulence factors, analysis of hemolysin, gelatinase and protease production, and susceptibility testing to antibiotics of relevance for human and veterinary industry. The results reported herein demonstrate the suitability of fish, seafood and fish products for the isolation of LAB (mainly enterococci), including (multi)bacteriocinogenic strains, encoding bacteriocins active against Listeria monocytogenes and other food-borne pathogens of interest for the food industry.
    European Food Research and Technology 05/2015; DOI:10.1007/s00217-015-2465-3 · 1.39 Impact Factor
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    ABSTRACT: In aquaculture, several criteria should be considered to select an appropriate probiotic, including the aquatic origin and safety of the strain and its ability to modulate the host immune response. The properties and effects of probiotics are strain-specific and some factors such as viability, dose and duration of diet supplementation may regulate their immunomodulatory activities. In this study, we assessed the in vitro effect of eight heat-inactivated and viable lactic acid bacteria (LAB) of aquatic origin belonging to the genera Enterococcus, Lactobacillus, Lactococcus, Leuconostoc, Pediococcus and Weissella on the viability and innate immune response of turbot (Scophthalmus maximus L.) leucocytes. Head-kidney leucocytes were incubated with viable and heat-inactivated LAB at different concentrations. After incubation, the viability of leucocytes was evaluated using colorimetric assays (MTT and LDH) and flow cytometry (annexin V/propidium iodide). Heat-inactivated LAB showed no cytotoxic effect while viable LAB exerted variable influence on apoptosis of turbot phagocytes and lymphocytes. Leucocyte respiratory burst activity and phagocytosis were also differentially activated, as viable LAB stimulated leucocytes more efficiently than the heat-inactivated LAB. Our results suggest diverse strain-specific mechanisms of interaction between the evaluated LAB and turbot leucocytes. Furthermore, our work sets up in vitro systems to evaluate the effect of LAB as potential probiotics, which will be useful to develop efficient screening. Copyright © 2015. Published by Elsevier Ltd.
    Fish &amp Shellfish Immunology 02/2015; 44(1). DOI:10.1016/j.fsi.2015.02.021 · 3.03 Impact Factor
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    ABSTRACT: The use of lactic acid bacteria of aquatic origin as probiotics constitutes an alternative strategy to the antibiotic treatment for disease control in aquaculture. Enterococci are currently used as probiotics in human and animal health. In this study, we evaluated the safety of 64 enterococci isolated from rainbow trout (Oncorhynchus mykiss, Walbaum), feed and rearing environment, and their antimicrobial activity against 9 fish pathogens. The 64 enterococcal isolates were identified to the species level by polymerase chain reaction (PCR), using specific primers for the different enterococcal species, and confirmed by superoxide dismutase gene sequencing. Enterococcus faecium and E. hirae were the most common species (42.2 and 35.9%, respectively). A total of 48 isolates (75%) showed phenotypic resistance to at least 1 antibiotic determined by a disk-diffusion method, and 25 isolates (39.1%) harbored at least 1 antibiotic resistance gene [erm(B), tet(M), tet(S), tet(K), tet(L), tet(T), vanC2, and aad(E)], detected by PCR. One (1.6%) isolate produced gelatinase and none produced hemolysin, using a plate assay. The virulence genes gelE (46.9%), efaAfs (17.2%), agg (1.6%), and hyl (1.6%) were detected by PCR. A total of 48 isolates (75%) exerted antimicrobial activity against 1 or more of the tested fish pathogens, using a stab-on-agar test. From these isolates, 21 (43.8%) harbored at least 1 bacteriocin-encoding gene (entP, entL50A and entL50B, hirJM79, entSE-K4, entQ and entA), detected by PCR. None of the enterococci showed bile deconjugation and mucin degradation abilities. A total of 17 enterococcal isolates (26.6%) that did not harbor any antibiotic resistance or virulence factor were considered safe for application as probiotics, including 6 isolates (35.3%) that showed antimicrobial activity against at least 1 fish pathogen and harbored at least 1 bacteriocin-encoding gene. Rainbow trout, feed, and rearing environment constitute an appropriate source for the isolation of enterococci as potential probiotic for aquaculture.
    Foodborne Pathogens and Disease 02/2015; 12(4). DOI:10.1089/fpd.2014.1906 · 2.09 Impact Factor
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    ABSTRACT: We have evaluated the cloning and functional expression of previously described broad antimicrobial spectrum bacteriocins SRCAM 602, OR-7, E-760, and L-1077, by recombinant Pichia pastoris. Synthetic genes, matching the codon usage of P. pastoris, were designed from the known mature amino acid sequence of these bacteriocins and cloned into the protein expression vector pPICZαA. The recombinant derived plasmids were linearized and transformed into competent P. pastoris X-33, and the presence of integrated plasmids into the transformed cells was confirmed by PCR and sequencing of the inserts. The antimicrobial activity, expected in supernatants of the recombinant P. pastoris producers, was purified using a multistep chromatographic procedure including ammonium sulfate precipitation, desalting by gel filtration, cation exchange-, hydrophobic interaction-, and reverse phase-chromatography (RP-FPLC). However, a measurable antimicrobial activity was only detected after the hydrophobic interaction and RP-FPLC steps of the purified supernatants. MALDI-TOF MS analysis of the antimicrobial fractions eluted from RP-FPLC revealed the existence of peptide fragments of lower and higher molecular mass than expected. MALDI-TOF/TOF MS analysis of selected peptides from eluted RP-FPLC samples with antimicrobial activity indicated the presence of peptide fragments not related to the amino acid sequence of the cloned bacteriocins.
    01/2015; 2015:767183. DOI:10.1155/2015/767183
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    ABSTRACT: Turbot (Scophthalmus maximus L.) is an important commercial marine flatfish. Its production may be affected by bacterial diseases that cause severe economical losses, mainly tenacibaculosis and vibriosis, provoked by Tenacibaculum maritimum and Vibrio splendidus, respectively. An alternative or complementary strategy to chemotherapy and vaccination for the control of these diseases is the use of probiotics. In this work, we report the invitro and invivo potential of eight lactic acid bacteria (LAB), previously isolated from fish, seafood and fish products intended for human consumption, as turbot probiotics. Seven out of the eight LAB exerted direct antimicrobial activity against, at least, four strains of T.maritimum and V.splendidus. All LAB survived in seawater at 18°C for 7 days, and withstood exposure to pH 3.0 and 10% (v/v) turbot bile; however, they differed in cell surface hydrophobicity (8.2-21.7%) and in their ability to adhere to turbot skin (1.2-21.7%) and intestinal (0.7-2.1%) mucus. Most of the tested strains inhibited the binding of turbot pathogens to the mucus. Leuconostoc mesenteroides subsp. cremoris SMM69 and Weissella cibaria P71 were selected based on their strong antimicrobial activity against T.maritimum and V.splendidus, good probiotic properties, and different adhesion ability to skin mucus and capacity to inhibit the adhesion of turbot pathogens to mucus. These two LAB strains were harmless when administered by bath to turbot larvae and juveniles; moreover, real-time PCR on the transcription levels of the immunity-related genes encoding IL-1β, TNF-α, lysozyme, C3, MHC-Iα and MHC-IIα in five organs (head-kidney, spleen, liver, intestine and skin) revealed the ability of these LAB to stimulate their expression in turbot juveniles, especially the non-specific immunity associated genes in mucosal tissues. Based on our results, Lc. cremoris SMM69 and W.cibaria P71 may be considered as suitable probiotic candidates for turbot farming.
    Fish &amp Shellfish Immunology 12/2014; 41(2):570-80. DOI:10.1016/j.fsi.2014.10.007 · 3.03 Impact Factor
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    ABSTRACT: This work reports the isolation and taxonomic identification of the cultivable total microbiota (TM) and Lactic Acid Bacteria (LAB) from rainbow trout (Oncorhynchus mykiss, Walbaum) and rearing environment from selected stages of the life-cycle, and the evaluation of the LAB antimicrobial activity against the main fish pathogens. TM and LAB isolates were randomly selected and identified by 16S rRNA and/or superoxide dismutase gene sequencing. Although a great diversity in the TM was observed, Enterobacteriaceae and Aeromonadaceae were clearly prevalent, while the genus Lactococcus was the predominant LAB. From a total of 1620 randomly selected LAB, 1159 isolates (71.5%) showed antimicrobial activity. From these, 248 isolates (21.4%) selected for their activity against, at least, four fish pathogens, were taxonomically identified, being Lactococcus lactis the most common species (164 isolates, 66.1%). Interestingly, 88 isolates (35.5%), including 55 L. lactis isolates, exerted activity against four strains of the rainbow trout pathogen Lactococcus garvieae. Our results demonstrate that rainbow trout and rearing environment are potential sources for the isolation of LAB, mainly lactococci, active against L. garvieae and other fish pathogens. Moreover, this is the first study describing the cultivable TM and LAB from rainbow trout intestine and rearing environment along the fish life-cycle. The host-derived LAB active against fish pathogens comprise potential candidates as probiotics in rainbow trout farming as an alternative or complementary strategy to antibiotics and vaccines for disease prevention. Copyright © 2014 Elsevier Ltd. All rights reserved.
    Anaerobe 11/2014; 32C:7-14. DOI:10.1016/j.anaerobe.2014.11.001 · 2.36 Impact Factor
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    ABSTRACT: In this work, genes encoding gelatinase (gelE) and serine proteinase (sprE), two extracellular proteases produced by Enterococcus faecalis DBH18 were cloned in the protein expression vector pMG36c, containing the constitutive P32 promoter, generating the recombinant plasmids pCG, pSP and pCGSP encoding gelE, sprE and gelE-sprE, respectively. Transformation of non-caseinolytic E. faecalis P36, E. faecalis JH2-2, E. faecium AR24, and E. hirae AR14 strains with these plasmids permitted detection of caseinolytic activity only in the strains transformed with pCG or pCGSP. Complementation of a deletion (knockout) mutant of E. faecalis V583 for production of gelatinase (GelE) with pCG unequivocally supported that gelE is responsible of the caseinolytic activity of the transformed strain, grown in bovine skim milk (BSM). RP-HPLC-MS/MS analysis of hydrolysates of transformed Enterococcus spp. strains grown in BSM permitted the identification of 38 major peptide fragments including peptides with previously reported ACE-inhibitory activity (ACE-IA), antihypertensive and antioxidant activity.
    Journal of Agricultural and Food Chemistry 05/2014; 62(24). DOI:10.1021/jf5006269 · 3.11 Impact Factor
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    ABSTRACT: 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 pPICZαA 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.
    Molecular Biotechnology 02/2014; 56(6). DOI:10.1007/s12033-014-9731-7 · 2.28 Impact Factor
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    ABSTRACT: A fingertip-to-fingertip intraindividual transmission experiment was carried out in 30 healthy volunteers, using four MLST-typed Enterococcus faecium clones. Overall results showed an adequate fit goodness to a theoretical exponential model, whereas four volunteers (13%) exhibited a significantly higher finger-to-finger bacterial transmission efficiency. This observation might have deep consequences in nosocomial epidemiology.
    MicrobiologyOpen 02/2014; DOI:10.1002/mbo3.156 · 2.21 Impact Factor
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    ABSTRACT: a b s t r a c t Cloning of the enterolysin A (EnlA) gene (enlA) from Enterococcus faecalis DAC9 into the pMSP3545-derived pMLG2-protein expression vector encoding EnlA under control of the inducible P nisA promoter permitted the controlled release and heterologous expression of mature EnlA by Lactococcus lactis subsp. cremoris NZ9000 and Lactococcus lactis subsp. lactis IL1403. The nisin-induced expression of enlA by L. lactis NZ9000 (pMLG2) and L. lactis IL1403 (pMLG2), grown in GM17 or bovine skim milk (BSM), caused a noticeable reduction of the optical density (OD 600) of the cultures and death of the growing cells. However, a high angiotensin converting enzyme (ACE)-inhibitory activity (ACE-IA) was only observed in the BSM-derived hydrolysates of L. lactis IL1403 (pMLG2) after 48 h-induction with nisin. Analysis of these hydrolysates by reversed phase high performance liquid chromatography-tandem mass spec-trometry permitted the identification of major peptide fragments with known ACE-IA or sharing at least three C-terminal residues with those displaying ACE-IA.
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    ABSTRACT: Enterococcus faecalis isolates from food and environmental origin were evaluated for their angiotensin-converting enzyme (ACE)-inhibitory activity (ACE-IA) after growth in bovine skim milk (BSM). Most (90% active) but not all (10% inactive) E. faecalis strains produced BSM-derived hydrolysates with high ACE-IA. Known ACE-inhibitory peptides (ACE-IP) and an antioxidant peptide were identified in the E. faecalis hydrolysates by reversed-phase high-performance liquid chromatography-tandem mass spectrometry (RP-HPLC-MS/MS). Antimicrobial activity against Pediococcus damnosus CECT4797 and Listeria ivanovii CECT913 was also observed in the E. faecalis hydrolysates. The incidence of virulence factors in the E. faecalis strains with ACE-IA and producers of ACE-IP was variable but less virulence factors were observed in the food and environmental strains than in the clinical reference strains. Pulsed-field gel electrophoresis (PFGE) and multilocus sequence typing (MLST) based analysis demonstrated that food and environmental E. faecalis strains were genetically different from those of clinical origin. When evaluated, most E. faecalis strains of clinical origin also originated BSM-derived hydrolysates with high ACE-IA due to the production of ACE-IP. Accordingly, the results of this work suggest that most E. faecalis strains of food, environmental and clinical origin produce BSM-derived bioactive peptides with human health connotations and potential biotechnological applications.
    International journal of food microbiology 06/2013; 166(1):93-101. DOI:10.1016/j.ijfoodmicro.2013.06.019 · 3.16 Impact Factor
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    ABSTRACT: Mature sakacin A (SakA, encoded by sapA) and its cognate immunity protein (SakI, encoded by sapiA), and two SakA-derived chimeras mimicking the N-terminal end of mature enterocin P (EntP/SakA) and mature enterocin A (EntA/SakA) together with SakI, were fused to different signal peptides (SP) and cloned into the protein expression vectors pNZ8048 and pMG36c for evaluation of their production and functional expression by different lactic acid bacteria. The amount, antimicrobial activity, and specific antimicrobial activity of SakA and its chimeras produced by Lactococcus lactis subsp. cremoris NZ9000 depended on the SP and the expression vector. Only L. lactis NZ9000 (pNUPS), producing EntP/SakA, showed higher bacteriocin production and antimicrobial activity than the natural SakA-producer Lactobacillus sakei Lb706. The lower antimicrobial activity of the SakA-producer L. lactis NZ9000 (pNUS) and that of the EntA/SakA-producer L. lactis NZ9000 (pNUAS) could be ascribed to secretion of truncated bacteriocins. On the other hand, of the Lb. sakei Lb706 cultures transformed with the pMG36c-derived vectors only Lb. sakei Lb706 (pGUS) overproducing SakA showed a higher antimicrobial activity than Lb. sakei Lb706. Finally, cloning of SakA and EntP/SakA into pPICZαA and pKLAC2 permitted the production of SakA and EntP/SakA by recombinant Pichia pastoris X-33 and Kluyveromyces lactis GG799 derivatives although their antimicrobial activity was lower than expected from their production.
    Journal of Industrial Microbiology 06/2013; 40(9). DOI:10.1007/s10295-013-1302-6 · 2.51 Impact Factor
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    Fish &amp Shellfish Immunology 06/2013; 34(6):1726-1727. DOI:10.1016/j.fsi.2013.03.277 · 3.03 Impact Factor
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    ABSTRACT: Background The microorganisms intended for use as probiotics in aquaculture should exert antimicrobial activity and be regarded as safe not only for the aquatic hosts but also for their surrounding environments and humans. The objective of this work was to investigate the antimicrobial/bacteriocin activity against fish pathogens, the antibiotic susceptibility, and the prevalence of virulence factors and detrimental enzymatic activities in 99 Lactic Acid Bacteria (LAB) (59 enterococci and 40 non-enterococci) isolated from aquatic animals regarded as human food. Results These LAB displayed a broad antimicrobial/bacteriocin activity against the main Gram-positive and Gram-negative fish pathogens. However, particular safety concerns based on antibiotic resistance and virulence factors were identified in the genus Enterococcus (86%) (Enterococcus faecalis, 100%; E. faecium, 79%). Antibiotic resistance was also found in the genera Weissella (60%), Pediococcus (44%), Lactobacillus (33%), but not in leuconostocs and lactococci. Antibiotic resistance genes were found in 7.5% of the non-enterococci, including the genera Pediococcus (12.5%) and Weissella (6.7%). One strain of both Pediococcus pentosaceus and Weissella cibaria carried the erythromycin resistance gene mef(A/E), and another two P. pentosaceus strains harboured lnu(A) conferring resistance to lincosamides. Gelatinase activity was found in E. faecalis and E. faecium (71 and 11%, respectively), while a low number of E. faecalis (5%) and none E. faecium exerted hemolytic activity. None enterococci and non-enterococci showed bile deconjugation and mucin degradation abilities, or other detrimental enzymatic activities. Conclusions To our knowledge, this is the first description of mef(A/E) in the genera Pediococcus and Weissella, and lnu(A) in the genus Pediococcus. The in vitro subtractive screening presented in this work constitutes a valuable strategy for the large-scale preliminary selection of putatively safe LAB intended for use as probiotics in aquaculture.
    BMC Microbiology 01/2013; 13(1):15. DOI:10.1186/1471-2180-13-15 · 2.98 Impact Factor
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    ABSTRACT: The bacteriocin enterocin A (EntA) produced by Enterococcus faecium T136 has been successfully cloned and produced by the yeasts Pichia pastoris X-33EA, Kluyveromyces lactis GG799EA, Hansenula polymorpha KL8-1EA, and Arxula adeninivorans G1212EA. Moreover, P. pastoris X-33EA and K. lactis GG799EA produced EntA in larger amounts and with higher antimicrobial and specific antimicrobial activities than the EntA produced by E. faecium T136.
    Applied and Environmental Microbiology 06/2012; 78(16):5956-61. DOI:10.1128/AEM.00530-12 · 3.95 Impact Factor
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    ABSTRACT: Due to their very potent antimicrobial activity against diverse food-spoiling bacteria and pathogens and their favourable biochemical properties, peptide bacteriocins from Gram-positive bacteria have long been considered promising for applications in food preservation or medical treatment. To take advantage of bacteriocins in different applications, it is crucial to have detailed knowledge on the molecular mechanisms by which these peptides recognize and kill target cells, how producer cells protect themselves from their own bacteriocin (self-immunity) and how target cells may develop resistance. In this review we discuss some important recent progress in these areas for the non-lantibiotic (class II) bacteriocins. We also discuss some examples of how the current wealth of genome sequences provides an invaluable source in the search for novel class II bacteriocins.
    Microbiology 12/2011; 157(Pt 12):3256-67. DOI:10.1099/mic.0.052571-0 · 2.84 Impact Factor
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    ABSTRACT: Replacement of the leader sequence (LS) of the bacteriocin enterocin A (LS(entA)) by the signal peptides (SP) of the protein Usp45 (SP(usp45)), and the bacteriocins enterocin P (SP(entP)), and hiracin JM79 (SP(hirJM79)) permits the production, secretion, and functional expression of EntA by different lactic acid bacteria (LAB). Chimeric genes encoding the SP(usp45), the SP(entP), and the SP(hirJM79) fused to mature EntA plus the EntA immunity genes (entA+entiA) were cloned into the expression vectors pNZ8048 and pMSP3545, under control of the inducible P(nisA) promoter, and in pMG36c, under control of the constitutive P(32) promoter. The amount, antimicrobial activity, and specific antimicrobial activity of the EntA produced by the recombinant Lactococcus lactis, Enterococcus faecium, E. faecalis, Lactobacillus sakei and Pediococcus acidilactici hosts varied depending on the signal peptide, the expression vector, and the host strain. However, the antimicrobial activity and the specific antimicrobial activity of the EntA produced by most of the LAB transformants was lower than expected from their production. The supernatants of the recombinant L. lactis NZ9000 (pNZUAI) and L. lactis NZ9000 (pNZHAI), overproducers of EntA, showed a 1.2- to 5.1-fold higher antimicrobial activity than that of the natural producer E. faecium T136 against different Listeria spp.
    Journal of Biotechnology 08/2011; 156(1):76-86. DOI:10.1016/j.jbiotec.2011.07.038 · 2.88 Impact Factor
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    ABSTRACT: Bacteriocins produced by enterococci, referred to as enterocins, possess great interest for their potential use as biopreservatives in food and feed, as well as alternative antimicrobials in humans and animals. In this context, the aim of the present study was to determine the antimicrobial activity and the presence of bacteriocin structural genes in fecal enterococcal isolates from animal origins. Evaluation of the direct antimicrobial activity of 253 isolates from wild boars (Sus scrofa, n = 69), mullets (Liza ramada, n = 117), and partridges (Perdix perdix, n = 67) against eight indicator bacterial strains (including Listeria monocytogenes, Pediococcus pentosaceus, and Enterococcus spp.) showed that 177 (70%) exerted antimicrobial activity against at least one indicator microorganism. From these isolates, 123 were further selected on the basis of their inhibition group, and 81 were found to be producers of bacteriocins active against Listeria monocytogenes. Analysis of the presence of enterocin structural genes in a subset of 36 isolates showed that 70% harbored one or more of the evaluated genes, those of enterocin P and hiracin JM79 being the most prevalent. These results show that wild animals constitute an appropriate source for the isolation of bacteriocinogenic enterococci.
    Journal of food protection 08/2011; 74(8):1252-60. DOI:10.4315/0362-028X.JFP-11-016 · 1.80 Impact Factor
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    ABSTRACT: In this work, biogenic amine production (histamine, tyramine and putrescine) by a collection of 74 lactic acid bacteria of aquatic origin has been investigated by means of amino acid decarboxylation by growth on decarboxylase differential medium, biogenic amine detection by thin-layer chromatography (TLC) and decarboxylase gene detection by PCR. None of the evaluated strains showed neither production of histamine and putrescine, nor presence of the genetic determinants encoding the corresponding decarboxylase activities. However, the tyrosine decarboxylase gene (tdc) was present in all the enterococcal strains, and tyramine production was detected by TLC in all of them but Enterococcus faecium BCS59 and MV5. Analysis of the tyrosine decarboxylase operon of these strains revealed the presence of an insertion sequence upstream tdc that could be responsible for their lack of tyrosine decarboxylase activity.
    International journal of food microbiology 02/2011; 146(2):212-6. DOI:10.1016/j.ijfoodmicro.2011.02.024 · 3.16 Impact Factor
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    ABSTRACT: Replacement of the signal peptide (SP) of the bacteriocins enterocin P (EntP) and hiracin JM79 (HirJM79), produced by Enterococcus faecium P13 and Enterococcus hirae DCH5, respectively, by the signal peptide of Usp45 (SP(usp45)), the major Sec-dependent protein secreted by Lactococcus lactis, permits the production, secretion, and functional expression of EntP and HirJM79 by L. lactis. Chimeric genes encoding the SP(usp45) fused to either mature EntP (entP), with or without the immunity gene (entiP) or to mature HirJM79 (hirJM79), with or without the immunity gene (hiriJM79), were cloned into the expression vector pMG36c, carrying the P(32) constitutive promoter, and into pNZ8048 under control of the inducible PnisA promoter. The production of EntP and HirJM79 by most of the L. lactis recombinant strains was 1.5- to 3.7-fold higher and up to 3.6-fold higher than by the E. faecium P13 and E. hirae DCH5 control strains, respectively. However, the specific antimicrobial activity of the recombinant EntP was 1.1- to 6.2-fold higher than that produced by E. faecium P13, while that of the HirJM79 was a 40% to an 89% of that produced by E. hirae DCH5. Chimeras of SP(usp45) fused to mature EntP or HirJM79 drive the production and secretion of these bacteriocins in L. lactis in the absence of specific immunity and secretion proteins. The supernatants of the recombinant L. lactis NZ9000 strains, producers of EntP, showed a much higher antimicrobial activity against Listeria spp. than that of the recombinant L. lactis NZ9000 derivatives, producers of HirJM79.
    Applied Microbiology and Biotechnology 01/2011; 89(1):131-43. DOI:10.1007/s00253-010-2849-z · 3.81 Impact Factor

Publication Stats

2k Citations
171.42 Total Impact Points

Institutions

  • 1994–2015
    • Complutense University of Madrid
      • • Department of Nutrition, Bromatology and Food Technology
      • • Department of Nutrition and Bromatology II (Bromatology)
      • • Facultad de Veterinaria
      Madrid, Madrid, Spain
  • 2012
    • Leibniz Institute of Plant Genetics and Crop Plant Research
      Gatersleben, Saxony-Anhalt, Germany
  • 2001
    • University of Groningen
      • Groningen Biomolecular Sciences and Biotechnology Institute (GBB)
      Groningen, Groningen, Netherlands