Veterinária
URI permanente desta comunidadehttps://locus.ufv.br/handle/123456789/11842
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10 resultados
Resultados da Pesquisa
Item Potential Control of Listeria monocytogenes by Bacteriocinogenic Enterococcus hirae ST57ACC and Pediococcus pentosaceus ST65ACC Strains Isolated From Artisanal Cheese(Probiotics and Antimicrobial Proteins, 2019-03) Cavicchioli, Valéria Quintana; Camargo, Anderson Carlos; Todorov, Svetoslav Dimitrov; Nero, Luís AugustoBacteriocinogenic Enterococcus hirae ST57ACC and Pediococcus pentosaceus ST65ACC strains, previously isolated from artisanal cheese, were evaluated for their safety with the aim to determine whether they could be used as beneficial strains, especially in the control of Listeria monocytogenes. Both isolates survived simulated gastrointestinal conditions and showed high levels of auto- and co-aggregation with L. monocytogenes, although the hydrophobicity of cells varied. Using the agar-spot test with 33 commercial drugs from different groups, only anti-inflammatory drugs and drugs containing loratadine and propranolol hydrochloride were able to affect the growth of the tested strains. Both strains were resistant to 3 out of 11 antibiotics tested by the disc diffusion method, and low frequencies of antibiotic resistance-encoding genes were observed by PCR analysis. Tested strains neither presented biogenic amine-related genes nor produced these substances. Aside from some antibiotic resistance characteristics, the tested strains were considered safe as they lack other virulence-related genes. E. hirae ST57ACC and P. pentosaceus ST65ACC both presented beneficial properties, particularly their ability to survive gastrointestinal conditions and to aggregate with L. monocytogenes, which can facilitate the elimination of this pathogen. Further studies should be conducted to better understand these interactions.Item Lactobacillus curvatus UFV-NPAC1 and other lactic acid bacteria isolated from calabresa, a fermented meat product, present high bacteriocinogenic activity against Listeria monocytogenes(BMC Microbiology, 2019) Castilho, Nathália Parma Augusto; Colombo, Monique; Oliveira, Leandro Licursi de; Todorov, Svetoslav Dimitrov; Nero, Luís AugustoBacteriocins produced by lactic acid bacteria (LAB) can be considered as viable alternatives for food safety and quality, once these peptides present antimicrobial activity against foodborne pathogens and spoilage bacteria. Fermented foods, such as artisanal sausages and cured meats, are relevant sources of LAB strains capable of producing novel bacteriocins, with particular interest by the food industry.Three LAB strains (firstly named as Lactobacillus curvatus 12, L. curvatus 36 and Weissella viridescens 23) were obtained from calabresa by presenting promising bacteriocinogenic activity, distinct genetic profiles (rep-PCR, RAPD, bacteriocin-related genes) and wide inhibitory spectrum. Among these strains, L. curvatus 12 presented higher bacteriocin production, reaching 25,000 AU/mL after incubation at 25, 30 and 37 °C and 6, 9 and 12 h. Partially purified bacteriocins from L. curvatus 12 kept their inhibitory activity after elution with isopropanol at 60% (v/v). Bacteriocins produced by this strain were purified by HPLC and sequenced, resulting in four peptides with 3102.79, 2631.40, 1967.06 and 2588.31 Da, without homology to known bacteriocins.LAB isolates obtained from calabresa presented high inhibitory activity. Among these isolates, bacteriocins produced by L. curvatus 12, now named as L. curvatus UFV-NPAC1, presented the highest inhibitory performance and the purification procedures revealed four peptides with sequences not described for bacteriocins to date.Item Functional properties of Lactobacillus mucosae strains isolated from brazilian goat milk(Probiotics and Antimicrobial Proteins, 2016-12-10) Nero, Luís Augusto; Moraes, Georgia Maciel Dias de; Abreu, Louricélia Rodrigues de; Egito, Antônio Silvio do; Salles, Hévila Oliveira; Silva, Liana Maria Ferreira da; Todorov, Svetoslav Dimitrov; Santos, Karina Maria Olbrich dosThe search for probiotic candidates among lactic acid bacteria (LAB) isolated from food may uncover new strains with promising health and technological properties. Lactobacillus mucosae strains attracted recent research attention due to their ability to adhere to intestinal mucus and to inhibit pathogens in the gastrointestinal tract, both related to a probiotic potential. Properties of interest and safety aspects of three Lb. mucosae strains (CNPC006, CNPC007, and CNPC009) isolated from goat milk were investigated employing in vitro tests. The presence of genetic factors related to bile salt hydrolase production (bsh), intestinal adhesion properties (msa, map, mub, and ef-tu), virulence, and biogenic amine production were also verified. All strains exhibited the target map, mub, and ef-tu sequences; the msa gene was detected in CNPC006 and CNPC007 strains. Some of the searched sequences for virulence factors were detected, especially in the CNPC009 strain; all strains carried the hyl gene, related to the production of hyaluronidase. Lb. mucosae CNPC007 exhibited a high survival rate in simulated gastric and enteric conditions. Besides, all strains exhibited the bsh sequence, and CNPC006 and CNPC007 were able to deconjugate salts of glycodeoxycholic acid (GDC). Regarding technological properties for dairy product applications, a relatively higher milk acidification and clotting capacity, diacetyl production, and proteolytic activity were registered for CNPC007 in comparison to the other strains. Collectively, the results aim at Lb. mucosae CNPC007 as a promising probiotic candidate for application in dairy products, deserving further studies to confirm and explore its potential.Item Virulence, antibiotic resistance and biogenic amines of bacteriocinogenic lactococci and enterococci isolated from goat milk(International Journal of Food Microbiology, 2014-06-12) Perin, Luana Martins; Miranda, Rodrigo Otávio; Todorov, Svetoslav Dimitrov; Franco, Bernadette Dora Gombossy de Melo; Nero, Luís AugustoThe present study aimed to investigate the virulence, antibiotic resistance and biogenic amine production in bacteriocinogenic lactococci and enterococci isolated from goat milk in order to evaluate their safety. Twenty-nine bacteriocinogenic lactic acid bacteria (LAB: 11 Lactococcus spp., and 18 Enterococcus spp.) isolated from raw goat milk were selected and subjected to PCR to identify gelE, cylA, hyl, asa1, esp, efaA, ace, vanA, vanB, hdc1, hdc2, tdc and odc genes. The expression of virulence factors (gelatinase, hemolysis, lipase, DNAse, tyramine, histamine, putrescine) in different incubation temperatures was assessed by phenotypic methods, as well as the resistance to vancomycin, gentamicin, chloramphenicol, ampicillin and rifampicin (using Etest®). The tested isolates presented distinct combinations of virulence related genes, but not necessarily the expression of such factors. The relevance of identifying virulence-related genes in bacteriocinogenic LAB was highlighted, demanding for care in their usage as starter cultures or biopreservatives due to the possibility of horizontal gene transfer to other bacteria in food systems.Item Investigation of genes involved in nisin production in Enterococcus spp. strains isolated from raw goat milk(Antonie van Leeuwenhoek, 2016-06-02) Perin, Luana Martins; Todorov, Svetoslav Dimitrov; Nero, Luís AugustoDifferent strains of Lactococcus lactis are capable of producing the bacteriocin nisin. However, genetic transfer mechanisms allow the natural occurrence of genes involved in nisin production in members of other bacterial genera, such as Enterococcus spp. In a previous study, nisA was identified in eight enterococci capable of producing antimicrobial substances. The aim of this study was to verify the presence of genes involved in nisin production in Enterococcus spp. strains, as well as nisin expression. The nisA genes from eight Enterococcus spp. strains were sequenced and the translated amino acid sequences were compared to nisin amino-acid sequences previously described in databases. Although containing nisin structural and maturation related genes, the enterococci strains tested in the present study did not present the immunity related genes (nisFEG and nisI). The translated sequences of nisA showed some point mutations, identical to those presented by Lactococcus strains isolated from goat milk. All enterococci were inhibited by nisin, indicating the absence of immunity and thus that nisin cannot be expressed. This study demonstrated for the first time the natural occurrence of nisin structural genes in Enterococcus strains and highlights the importance of providing evidence of a link between the presence of bacteriocin genes and their expression.Item Functional properties of Lactobacillus mucosa e strains isolated from brazilian goat milk(Probiotics and Antimicrobial Proteins, 2016-12-10) Nero, Luís Augusto; Todorov, Svetoslav Dimitrov; Moraes, Georgia Maciel Dias de; Abreu, Louricélia Rodrigues de; Egito, Antônio Silvio do; Salle, Hévila Oliveira; Silva, Liana Maria Ferreira da; Santos, Karina Maria Olbrich dosThe search for probiotic candidates among lactic acid bacteria (LAB) isolated from food may uncover new strains with promising health and technological properties. Lactobacillus mucosae strains attracted recent research attention due to their ability to adhere to intestinal mucus and to inhibit pathogens in the gastrointestinal tract, both related to a probiotic potential. Properties of interest and safety aspects of three Lb. mucosae strains (CNPC006, CNPC007, and CNPC009) isolated from goat milk were investigated employing in vitro tests. The presence of genetic factors related to bile salt hydrolase production (bsh), intestinal adhesion properties (msa, map, mub, and ef-tu), virulence, and biogenic amine production were also verified. All strains exhibited the target map, mub, and ef-tu sequences; the msa gene was detected in CNPC006 and CNPC007 strains. Some of the searched sequences for virulence factors were detected, especially in the CNPC009 strain; all strains carried the hyl gene, related to the production of hyaluronidase. Lb. mucosae CNPC007 exhibited a high survival rate in simulated gastric and enteric conditions. Besides, all strains exhibited the bsh sequence, and CNPC006 and CNPC007 were able to deconjugate salts of glycodeoxycholic acid (GDC). Regarding technological properties for dairy product applications, a relatively higher milk acidification and clotting capacity, diacetyl production, and proteolytic activity were registered for CNPC007 in comparison to the other strains. Collectively, the results aim at Lb. mucosae CNPC007 as a promising probiotic candidate for application in dairy products, deserving further studies to confirm and explore its potential.Item Genetic diversity and some aspects of antimicrobial activity of lactic acid bacteria isolated from goat milk(Applied Biochemistry and Biotechnology, 2015-01-31) Cavicchioli, Valéria Quintana; Dornellas, Wesley dos Santos; Perin, Luana Martins; Pieri, Fábio Alessandro; Franco, Bernadette Dora Gombossy de Melo; Todorov, Svetoslav Dimitrov; Nero, Luís AugustoLactic acid bacteria (LAB, n = 57) were previously obtained from raw goat milk, identified as Lactococcus spp. (n = 24) and Enterococcus spp. (n = 33), and characterized as bacteriocinogenic. Fingerprinting by pulsed field gel electrophoresis (PFGE) demonstrated high genetic diversity, and 30 strains were selected and exhibited strong antimicrobial activity against 46 target strains (LAB, spoilage, and foodborne pathogens). Six strains (Lactococcus lactis: GLc03 and GLc05; and Enterococcus durans: GEn09, GEn12, GEn14, and GEn17) were selected to characterize their bacteriocinogenic features, using Listeria monocytogenes ATCC 7644 as the target. The six strains produced bacteriocins at higher titer when incubated in MRS at 37 °C up to 12 h, when compared to growth at 25 and 30 °C. The produced bacteriocins kept their antimicrobial activity after exposure to 100 °C for 2 h and 121 °C for 20 min; the antimicrobial activity was also observed after treatment at pH 2.0 to 10.0, except for GLc03. L. monocytogenes populations were reduced approximately two logs after treatment with cell-free supernatants from the selected strains. These data show that goat milk can contain a diverse microbiota able to inhibit L. monocytogenes, a common pathogen found in dairy products, and can be potentially employed in biopreservation of food produced under different processing conditions.Item In Vitro evaluation of bacteriocins activity against Listeria monocytogenes biofilm formation(Applied Biochemistry and Biotechnology, 2015-12-10) Camargo, Anderson Carlos; Paula, Otávio Almeida Lino de; Todorov, Svetoslav Dimitrov; Nero, Luís AugustoThe present study aimed to assess the activity of cell-free supernatant (CFS) containing bacteriocins on the formation and maintenance of biofilms developed by Listeria monocytogenes, and the associated effect of bacteriocins and ethylene-diamine-tetra-acetic acid (EDTA) on the formed biofilm. CFS from 9 lactic acid bacteria (LAB) strains was tested for inhibitory activity against 85 L. monocytogenes isolates and 21 LAB strains. Then, 12 L. monocytogenes strains were selected based on genetic profiles and sensitivity to CFS and were subjected to an in vitro assay to assess biofilm formation in microtiter plates, considering different culture media and incubation conditions. Based on these results, 6 L. monocytogenes strains were subjected to the same in vitro procedure to assess biofilm formation, being co-inoculated with CFS. In addition, these strains were subjected to the same in vitro procedure, modified by adding the CFS after biofilm formation. Relevant decrease in biofilm formation was observed in the first experiment, but CFS added after biofilm formation did not eliminate them. CFS from Lactobacillus curvatus ET31 were selected due to its anti-biofilm activity, being associated to EDTA at different concentrations and tested for biofilm control of three strains of L. monocytogenes, using the same in vitro procedure described previously. Concentrated bacteriocin presented poor performance in eliminating formed biofilms, and EDTA concentration presented no evident interference on biofilm elimination. Twelve selected L. monocytogenes strains were positive for investigated virulence makers and negative for luxS gene, recognized as being involved in biofilm formation. Selected L. monocytogenes strains were able to produce biofilms under different conditions. CFSs have the potential to prevent biofilm formation, but they were not able to destroy already formed biofilms. Nevertheless, low concentrations of CFS combined with EDTA caused a relevant reduction in already formed biofilms, but this association was not able to eliminate them. The activity of selected CFS was demonstrated against L. monocytogenes-formed biofilms, being more effective when associated to EDTA at different concentrations.Item Inhibition of herpes simplex virus 1 (HSV-1) and poliovirus (PV-1) by bacteriocins from lactococcus lactis subsp. lactis and enterococcus durans strains isolated from goat milk(International Journal of Antimicrobial Agents, 2017-04-05) Cavicchioli, Valéria Quintana; Carvalho, Otávio Valério de; Paiva, Janine Cerqueira de; Todorov, Svetoslav Dimitrov; Silva Júnior, Abelardo; Nero, Luís AugustoBacteriocins have unusual inhibitory activity, including antiviral properties, and this can be exploited to give alternative applications. Semi–purified bacteriocins of six lactic acid bacteria (LAB) strains isolated from goat milk (two Lactococcus lactis: GLc03 and GLc05, and four Enterococcus durans: GEn09, GEn12, GEn14 and GEn17) were tested for cytotoxicity in Vero cells (50% Cytotoxicity Concentration: CC50), and for their antiviral activities against herpes simplex virus 1 (HVS-1) and poliovirus (PV-1). Semi-purified bacteriocins presented low cytotoxicity, with CC50 varying from 256.2 µg/mL (GLc05) to 1084.5 µg/mL (GEn14). CC10 was determined for all isolates (GLc03: 36.9 µg/mL; GLc05: 51.2 µg/mL; GEn09: 88.1 µg/mL; GEn12: 99.9 µg/mL; GEn14: 275 µg/mL; and GEn17: 62.2 µg/mL) and considered for antiviral activity assays. Antiviral activity before virus adsorption was recorded against PV-1 for GLc05 (4.9%), GEn09 (3.4%), GEn12 (24.7%) and GEn17 (23.5%), and against HSV-1 for GEn12 (27.9%), GEn14 (58.7%) and GEn17 (39.2%). Antiviral activity after virus adsorption was identified against PV-1 for GLc05 (32.7%), GEn09 (91.0%), GEn12 (93.7%) and GEn17 (57.2%), and against HSV-1 for GEn17 (71.6%). The results obtained indicate the potential of some bacteriocins, particularly those produced by E. durans strains investigated in the present study, in viral inhibition and their application as new antiviral agents.Item Safety of Lactobacillus plantarum ST8Sh and its bacteriocin(Probiotics and Antimicrobial Proteins, 2017-02-23) Todorov, Svetoslav Dimitrov; Perin, Luana M.; Nero, Luís Augusto; Carneiro, Bruno M.http://dx.doi.org/10.1007/s12602-017-9260-3; Rahal, Paula; Holzapfel, WilhelmTotal DNA extracted from Lb. plantarum ST8Sh was screened for the presence of more than 50 genes related to production of biogenic amines (histidine decarboxylase, tyrosine decarboxylase, and ornithine decarboxylase), virulence factors (sex pheromones, gelatinase, cytolysin, hyaluronidase, aggregation substance, enterococcal surface protein, endocarditis antigen, adhesion of collagen, integration factors), and antibiotic resistance (vancomycin, tetracycline, erythromycin, gentamicin, chloramphenicol, bacitracin). Lb. plantarum ST8Sh showed a low presence of virulence genes. Only 13 genes were detected (related to sex pheromones, aggregation substance, adhesion of collagen, tetracycline, gentamicin, chloramphenicol, erythromycin, but not to vancomycin, and bacitracin) and may be considered as indication of safety for application in fermented food products. In addition, interaction between Lb. plantarum ST8Sh and drugs from different groups were determined in order to establish possible application of the strain in combination with commercial drugs. Cytotoxicity of the semi-purified bacteriocins produced by Lb. plantarum ST8Sh was depended on applied concentration—highly cytotoxic when applied at 25 μg/mL and no cytotoxicity at 5 μg/mL.