Potentiation of the humoral immune response elicited by a commercial vaccine against bovine respiratory disease by Enterococcus faecalis CECT7121.

Vaccination against pathogens involved in bovine respiratory disease (BRD) is a useful tool to reduce the risk of this disease however, it has been observed that the commercially available vaccines only partially prevent the infections caused by Pasteurella multocida and Mannheimia haemolytica. Therefore, it is recommended to search for new adjuvant strategies to minimise the economic impact of this respiratory syndrome. A possibility to improve the conventional vaccine response is to modulate the immune system with probiotics, since there is accumulating evidence that certain immunomodulatory strains administered around the time of vaccination can potentiate the immune response. Considering veterinary vaccines are frequently tested in murine models, we have developed an immunisation schedule in BALB/c mice that allows us to study the immune response elicited by BRD vaccine. In order to evaluate a potential strategy to enhance vaccine efficacy, the adjuvant effect of Enterococcus faecalis CECT7121 on the murine specific humoral immune response elicited by a commercial vaccine against BRD was studied. Results indicate that the intragastric administration of E. faecalis CECT7121 was able to induce an increase in the specific antibody titres against the bacterial components of the BRD vaccines (P. multocida and M. haemolytica). The quality of the humoral immune response, in terms of antibody avidity, was also improved. Regarding the cellular immune response, although the BRD vaccination induced a low specific secretion of cytokines in the spleen cell culture supernatants, E. faecalis CECT7121-treated mice showed higher interferon-γ production than immunised control mice. Our results allowed us to conclude that the administration of E. faecalis CECT7121 could be employed as an adjuvant strategy to potentiate humoral immune responses.

Probiotic activity of Enterococcus faecalis CECT7121: effects on mucosal immunity and intestinal epithelial cells.

AIMS: To analyse the effect of Enterococcus faecalis CECT7121 on intestinal epithelial cells (IECs) and its effects on the mucosal immune response. METHODS AND RESULTS: Enterococcus faecalis CECT7121 showed a high adhesion capacity to completely and heterogeneously differentiated human intestinal epithelial cell line (Caco-2 cells). In addition, the contact of this bacterium with Caco-2 cells did not induce inflammatory chemokines (IL-8 and CCL-20). The presence of IgA(+) and IL-6(+) cells in the small intestine, as well as the production of inflammatory cytokines (TNFα, IL-6 and IL-12) in the gut, was determined after intragastric inoculation of Ent. faecalis CECT7121 in BALB/c mice. The administration of Ent. faecalis CECT7121 increased the number of IgA(+) cells in the intestinal lamina propria without modifying the percentage of IL-6(+) cells. No differences were observed in the cytokines measured in the intestinal extracts between probiotic-treated and control mice. CONCLUSIONS: Enterococcus faecalis CECT7121 stimulates local mucosal immunity and adheres to IECs without inducing inflammatory signals. SIGNIFICANCE AND IMPACT OF THE STUDY: Our results indicate that, apart from its already reported systemic immune activity, Ent. faecalis CECT7121 has a modulatory effect at a local level.

Bio-preservation of ground beef meat by Enterococcus faecalis CECT7121.

Meat and particularly ground beef is frequently associated with Food Poisoning episodes and breeches in Food Safety. The main goal of this research was to evaluate the bactericide effect of the probiotic Enterococcus faecalis CECT7121, against different pathogens as: Escherichia coli O157:H7, Staphylococcus aureus, Clostridium perfringens and Listeria monocytogenes, inoculated in ground beef meat. Three studies were performed to evaluate the inhibition of E. faecalis CECT7121 on ground beef meat samples inoculated with pathogens: Study I: Samples (100 g meat) were inoculated with pathogens (10(3) CFU/g)) and E. faecalis CECT7121 (10(4) CFU/g) simultaneously. Study II: Samples were inoculated with E. faecalis CECT7121 24 h before the pathogens. Study III: E. faecalis CECT7121were inoculated 24 h after pathogens. The viable counts were performed at 0, 24, 48 and 72 h post-inoculation. The simultaneous inoculation of E. faecalis CECT7121 with E. coli O157:H7 strains resulted in the absence of viable counts of bacteria at 72 h post-treatment. However, when the probiotic was added 24 h before and 24 h after the pathogen E. coli O157:H7, viable cells were not detected at 24 h and 48 h post-treatment, respectively. Consistently, neither S. aureus nor Cl. perfringens viable bacteria were detected at 48 h in whole assays when inoculated with E. faecalis CECT7121. The same trend than described before was obtained after applying the 3 models assayed for L. monocytogenes. The current assays demonstrated the bactericide activity of E. faecalis CECT7121 strain on bacterial pathogens in ground beef meat.

Bio-preservation of ground beef meat by Enterococcus faecalis CECT7121

Meat and particularly ground beef is frequently associated with Food Poisoning episodes and breeches in Food Safety. The main goal of this research was to evaluate the bactericide effect of the probiotic Enterococcus faecalis CECT7121, against different pathogens as: Escherichia coli O157:H7, Staphylococcus aureus, Clostridium perfringens and Listeria monocytogenes, inoculated in ground beef meat. Three studies were performed to evaluate the inhibition of E. faecalis CECT7121 on ground beef meat samples inoculated with pathogens: Study I: Samples (100 g meat) were inoculated with pathogens (10³ CFU/g)) and E. faecalis CECT7121 (10(4) CFU/g) simultaneously. Study II: Samples were inoculated with E. faecalis CECT7121 24 h before the pathogens. Study III: E. faecalis CECT7121were inoculated 24 h after pathogens. The viable counts were performed at 0, 24, 48 and 72 h post-inoculation. The simultaneous inoculation of E. faecalis CECT7121 with E. coli O157:H7 strains resulted in the absence of viable counts of bacteria at 72 h post-treatment. However, when the probiotic was added 24 h before and 24 h after the pathogen E. coli O157:H7, viable cells were not detected at 24 h and 48 h post-treatment, respectively. Consistently, neither S. aureus nor Cl. perfringens viable bacteria were detected at 48 h in whole assays when inoculated with E. faecalis CECT7121. The same trend than described before was obtained after applying the 3 models assayed for L. monocytogenes. The current assays demonstrated the bactericide activity of E. faecalis CECT7121 strain on bacterial pathogens in ground beef meat.

Assessment of the in vitro efficacy of the novel antimicrobial peptide CECT7121 against human Gram-positive bacteria from serious infections refractory to treatment.

BACKGROUND: Resistant Gram-positive bacteria are causing increasing concern in clinical practice. This work investigated the efficacy of AP-CECT7121 (an antimicrobial peptide isolated from an environmental strain of Enterococcus faecalis CECT7121) against various pathogenic Gram-positive bacteria. METHODS: Strains were isolated from intensive care unit patients unresponsive to standard antibiotic treatments. Inhibitory activity of AP-CECT7121 was assessed using the agar-well diffusion method. The most resistant isolates from each species screened (Enterococcus faecium, Enterococcus faecalis,Staphylococcus aureus, Streptococcus pneumoniae, Streptococcus pyogenes, Clostridium perfringens and Clostridium difficile) were further examined in time-killing curve studies. RESULTS: These bactericidal kinetic experiments demonstrated a rapid killing effect with no viable bacteria being detected within 30 and 90 min for enterococcal and streptococcal strains and 180 min for community-acquired methicillin-resistant S. aureus and C. perfringens: viable counts for C. difficile were threefold decreased after 90 min. CONCLUSIONS: AP-CECT7121 may provide a novel strategy for treating potentially fatal clinical infections in hospitalized patients.

In vitro efficacy of the novel peptide CECT7121 against bacteria isolated from mastitic dairy cattle.

AIMS: To evaluate the in vitro bactericidal activity of the novel antimicrobial peptide (AP) CECT7121 against Gram-positive bacteria from mastitic dairy cattle. METHODS AND RESULTS: A total of 15 Staphylococcus aureus, 10 Streptococcus dysgalactiae, 7 Strep. uberis, 1 Strep. agalactiae strains were isolated from 33 different mastitic dairy cattle, sourced from two dairies in Tandil-Argentina. Isolates from each of the bacterial species screened which developed the lowest inhibition zones in response to the peptide, were further evaluated in a series of time-killing curve studies. No survivors were detected in whole strains (from the three Streptococcal species isolated) within 120 min of incubation in presence of the peptide. The Staph. aureus isolates were less sensitive but, nevertheless, a drop in viable counts to below the detection limit was achieved for all the test strains by the final postincubation sampling point at 180 min. CONCLUSIONS: The study demonstrated the in vitro efficacy of the AP-CECT7121 against a variety strains of Gram positives isolated from mastitic dairy cattle. SIGNIFICANCE AND IMPACT OF THE STUDY: There is urgent global interest in the development of natural alternatives for the control and prevention of mastitis. Confirmation of the in vitro activity of the novel AP-CECT7121 against Gram-positive isolates encourages further research.

Partial characterization of enterocin MR99 from a corn silage isolate of Enterococcus faecalis.

AIMS: To assess the inhibitory activity on Gram-positive and Gram-negative bacteria of several species of enterococci recovered from a natural corn silage. METHODS AND RESULTS: The inhibitory activity of strains of Enterococcus faecalis (58), Enterococcus faecium (35), Enterococcus gallinarum (3) and Enterococcus casseliflavus (4) were studied employing indicator strains from various sources (clinical, food and ATCC). Enterococcus faecalis MR99, the only strain with inhibitory activity, inhibited other enterococci, Listeria spp., Staphylococcus aureus, Clostridium spp., Bacillus spp., Escherichia coli, Shigella sonnei and Shigella flexneri. The bacterium contained only one conjugative pheromone-responsive plasmid. The partially chromatography-purified MR99 enterocin (PPE) had a molecular weight of approx. 5000 Da and a pI of 6.2, was sensitive to proteolytic enzymes and could be extracted in benzene and butanol. It appeared stable to adjustment of pH 4.0, 5.0, 6.0, 7.0 and 8.0 and was resistant to heat. Inactivation was at 15 min at 121 degrees C. Enterocin MR99 was bactericidal on strains of Listeria monocytogenes, Staph. aureus, and bovine mastitis agents, it was bacteriostatic on E. coli. Although enterocins MR99 and AS48 have inhibitory activity on Gram-negative bacilli, PCR studies demonstrated a lack of relationship between them. CONCLUSIONS: The active component had a protein nature, was resistant to heat and presented a wide inhibitory spectrum. SIGNIFICANCE AND IMPACT OF THE STUDY: The biological properties of Ent. faecalis MR99 suggest that this strain merits further investigations so it can be applied in human and veterinary health programmes.

[Evaluation of the bacterial flora in natural corn silage]. / Evaluación de la flora bacteriana en un ensilado natural de maíz.

Silage is the best method for forage conservation, keeping it in a fresh condition and with high nutrient content. In order to study the natural evolution of maize silage without additives during 50 days after sealed, pH, temperature changes, number and type of the indigenous bacteria were studied every 5 days. The initial pH decreased from 6.40 to 4.10 and temperature stabilized at 26 degrees C. The mesophilic aerobic bacteria were quickly reduced across time. Enterococci were descended gradually. Clostridia remained in a low quantity. The microorganisms most frequently isolated were Escherichia coli, Clostridium butyricum, Enterococcus faecalis and Lactobacillus plantarum; this acid lactic bacteria was the principal cause of the abundant production of anaerobic fermentation of sugars in the forage reaching the pH which maintained the silage in adequate conditions.

Evaluación de la flora bacteriana en un ensilado natural de maíz / Evaluation of the bacterial flora in natural corn silage

Silage is the best method for forage conservation, keeping it in a fresh condition and with high nutrient content. In order to study the natural evolution of maize silage without additives during 50 days after sealed, pH, temperature changes, number and type of the indigenous bacteria were studied every 5 days. The initial pH decreased from 6.40 to 4.10 and temperature stabilized at 26 degrees C. The mesophilic aerobic bacteria were quickly reduced across time. Enterococci were descended gradually. Clostridia remained in a low quantity. The microorganisms most frequently isolated were Escherichia coli, Clostridium butyricum, Enterococcus faecalis and Lactobacillus plantarum; this acid lactic bacteria was the principal cause of the abundant production of anaerobic fermentation of sugars in the forage reaching the pH which maintained the silage in adequate conditions.

Evaluación de la flora bacteriana en un ensilado natural de maíz / Evaluation of the bacterial flora in natural corn silage

Silage is the best method for forage conservation, keeping it in a fresh condition and with high nutrient content. In order to study the natural evolution of maize silage without additives during 50 days after sealed, pH, temperature changes, number and type of the indigenous bacteria were studied every 5 days. The initial pH decreased from 6.40 to 4.10 and temperature stabilized at 26 degrees C. The mesophilic aerobic bacteria were quickly reduced across time. Enterococci were descended gradually. Clostridia remained in a low quantity. The microorganisms most frequently isolated were Escherichia coli, Clostridium butyricum, Enterococcus faecalis and Lactobacillus plantarum; this acid lactic bacteria was the principal cause of the abundant production of anaerobic fermentation of sugars in the forage reaching the pH which maintained the silage in adequate conditions.(AU)

Evaluación de la flora bacteriana en un ensilado natural de maíz. / [Evaluation of the bacterial flora in natural corn silage]

Silage is the best method for forage conservation, keeping it in a fresh condition and with high nutrient content. In order to study the natural evolution of maize silage without additives during 50 days after sealed, pH, temperature changes, number and type of the indigenous bacteria were studied every 5 days. The initial pH decreased from 6.40 to 4.10 and temperature stabilized at 26 degrees C. The mesophilic aerobic bacteria were quickly reduced across time. Enterococci were descended gradually. Clostridia remained in a low quantity. The microorganisms most frequently isolated were Escherichia coli, Clostridium butyricum, Enterococcus faecalis and Lactobacillus plantarum; this acid lactic bacteria was the principal cause of the abundant production of anaerobic fermentation of sugars in the forage reaching the pH which maintained the silage in adequate conditions.

[Meningitis due to Enterococcus faecalis]. / Meningitis a Enterococcus faecalis.

Enterococcus faecalis meningitis is an infrequent entity that accounts for less than 1% of all suppurative meningitis in the adult. Usually, this infection affects patients with compromised host defenses or those who have congenital or acquired CNS lesions mainly as intrahospitalary infections. An 85 year old woman from our community (Tandil county) without any predisposed condition, was admitted in the hospital in an unconscious state (grade 3 Glasgow's index), meningeal signs and purulent CSF, from which E. faecalis was isolated. The patient was treated with IV Ampicillin and Gentamycin (17 days), intrathecal Gentamycin (4 days) and IV dexametasona (6 days). The clinical and bacteriological remission was achieved, without any sequel or relapse during 2 years follow up.

Meningitis a Enterococcus faecalis / [Meningitis due to Enterococcus faecalis]

Enterococcus faecalis meningitis is an infrequent entity that accounts for less than 1

of all suppurative meningitis in the adult. Usually, this infection affects patients with compromised host defenses or those who have congenital or acquired CNS lesions mainly as intrahospitalary infections. An 85 year old woman from our community (Tandil county) without any predisposed condition, was admitted in the hospital in an unconscious state (grade 3 Glasgow’s index), meningeal signs and purulent CSF, from which E. faecalis was isolated. The patient was treated with IV Ampicillin and Gentamycin (17 days), intrathecal Gentamycin (4 days) and IV dexametasona (6 days). The clinical and bacteriological remission was achieved, without any sequel or relapse during 2 years follow up.

Meningitis a Enterococcus faecalis. / [Meningitis due to Enterococcus faecalis]

Enterococcus faecalis meningitis is an infrequent entity that accounts for less than 1

of all suppurative meningitis in the adult. Usually, this infection affects patients with compromised host defenses or those who have congenital or acquired CNS lesions mainly as intrahospitalary infections. An 85 year old woman from our community (Tandil county) without any predisposed condition, was admitted in the hospital in an unconscious state (grade 3 Glasgows index), meningeal signs and purulent CSF, from which E. faecalis was isolated. The patient was treated with IV Ampicillin and Gentamycin (17 days), intrathecal Gentamycin (4 days) and IV dexametasona (6 days). The clinical and bacteriological remission was achieved, without any sequel or relapse during 2 years follow up.