Lactic acid bacteria reduce bacterial diarrhea in rabbits via enhancing immune function and restoring intestinal microbiota homeostasis.

BACKGROUND: Numerous previous reports have demonstrated the efficacy of Lactic acid bacteria (LAB) in promoting growth and preventing disease in animals. In this study, Enterococcus faecium ZJUIDS-R1 and Ligilactobaciiius animalis ZJUIDS-R2 were isolated from the feces of healthy rabbits, and both strains showed good probiotic properties in vitro. Two strains (108CFU/ml/kg/day) were fed to weaned rabbits for 21 days, after which specific bacterial infection was induced to investigate the effects of the strains on bacterial diarrhea in the rabbits. RESULTS: Our data showed that Enterococcus faecium ZJUIDS-R1 and Ligilactobaciiius animalis ZJUIDS-R2 interventions reduced the incidence of diarrhea and systemic inflammatory response, alleviated intestinal damage and increased antibody levels in animals. In addition, Enterococcus faecium ZJUIDS-R1 restored the flora abundance of Ruminococcaceae1. Ligilactobaciiius animalis ZJUIDS-R2 up-regulated the flora abundance of Adlercreutzia and Candidatus Saccharimonas. Both down-regulated the flora abundance of Shuttleworthia and Barnesiella to restore intestinal flora balance, thereby increasing intestinal short-chain fatty acid content. CONCLUSIONS: These findings suggest that Enterococcus faecium ZJUIDS-R1 and Ligilactobaciiius animalis ZJUIDS-R2 were able to improve intestinal immunity, produce organic acids and regulate the balance of intestinal flora to enhance disease resistance and alleviate diarrhea-related diseases in weanling rabbits.

An update on the taxonomy and functional properties of the probiotic Enterococcus faecium SF68.

Enterococcus faecium SF68 (SF68) is a well-known probiotic with a long history of safe use. Recent changes in the taxonomy of enterococci have shown that a novel species, Enterococcus lactis, is closely related with E. faecium and occurs together with other enterococci in a phylogenetically well-defined E. faecium species group. The close phylogenetic relationship between the species E. faecium and E. lactis prompted a closer investigation into the taxonomic status of E. faecium SF68. Using phylogenomics and ANI, the taxonomic analysis in this study showed that probiotic E. faecium SF68, when compared to other E. faecium and E. lactis type and reference strains, could be re-classified as belonging to the species E. lactis. Further investigations into the functional properties of SF68 showed that it is potentially capable of bacteriocin production, as a bacteriocin gene cluster encoding the leaderless bacteriocin EntK1 together with putative Lactococcus lactis bacteriocins LsbA, and LsbB-like putative immunity peptide (LmrB) were found located in an operon on plasmid pF9. However, bacteriocin expression was not studied. Competitive exclusion experiments in co-culture over 7 days at 37 °C showed that the probiotic SF68 could inhibit the growth of specific E. faecium and Listeria monocytogenes strains, while showing little or no inhibitory activity towards an entero-invasive Escherichia coli and a Salmonella Typhimurium strain, respectively. In cell culture experiments with colon carcinoma HT29 cells, the probiotic SF68 was also able to strain-specifically inhibit adhesion and/or invasion of enterococcal and L. monocytogenes strains, while such adhesion and invasion inhibition effects were less pronounced for E. coli and Salmonella strains. This study therefore provides novel data on the taxonomy and functional properties of SF68, which can be reclassified as Enterococcus lactis SF68, thereby enhancing the understanding of its probiotic nature.

Impacts of water activity on survival of Listeria innocua and Enterococcus faecium NRRL B-2354 in almonds during steam treatments.

Raw almonds have been associated with Salmonella outbreaks and multiple recalls related to Listeria monocytogenes contamination. While steam treatment has been approved for pasteurizing both conventional and organic whole almonds, there is limited understanding of how water activity (aw) influences the effectiveness of steam treatments in decontaminating almonds. Hence, this study aimed to assess and compare the efficacy of steam treatments against Listeria innocua and Enterococcus faecium NRRL B-2354, the known non-pathogenic surrogates, on almonds. It also sought to investigate the impact of almond's aw on bacterial resistance during steam treatments. Almond kernels were inoculated with ~8 log10 CFU/g of either E. faecium or L. innocua and equilibrated to aw 0.25 or 0.45 before being subjected to steam treatments at temperatures of 100-135 °C. Our results revealed that L. innocua exhibited lower resistance to steam compared to E. faecium, with 1.2-2.6 log10 CFU/g reductions for L. innocua and 1.0-2.0 log10 CFU/g reductions for E. faecium when the surface temperature of almonds reached 100-130 °C, depending on the aw of the almonds. The obtained DL. innocua, 100-130°C-values were 2.0-16.6 s, and DE. faecium, 100-130°C-values were 4.0-21.8 s, depending on the aw of almonds. In general, elevating steam temperatures and almond aw decreased the tolerance of L. innocua and E. faecium during steam inactivation. In addition, the z-values indicated that E. faecium on almonds was less sensitive to change in steam temperature compared to L. innocua, especially at lower aw. The zL. innocua-values were 36.6 °C and 35.7 °C, while zE. faecium-values were 48.9 °C and 42.7 °C in almonds with aw 0.25 and 0.45, respectively. Results from this study suggest that steam treatments serve as effective interventions for controlling pathogen contaminations in raw almonds.

Metabolic response of broiler chickens to different doses of Enterococcus faecium ICIS 96 in the diet.

The competitiveness and profitability of the poultry industry in market conditions are hampered by growing problems with the safety and quality of poultry meat. The use of targeted microbial preparations can assist in resolving these problems. Numerous studies of the properties of bacteria of the genus Enterococcus have shown their effectiveness in the practice of poultry farming. The objectives of this study were to identify the effect of various doses of Enterococcus faecium ICIS 96 added to the diet of broiler chickens on their metabolism and to evaluate the productive qualities of the chickens when different doses are used. The experiment was carried out on 72 Cobb-500 cross broiler chickens, divided into 3 groups. Chickens in the first group received a suspension of E. faecium ICIS 96 containing 1 × 108 cells in 1 mL of sterile saline, at a dose of 0.1 mL per 1 kg of live weight. The second group of chickens received a double dose of enterococcus, that is 0.2 mL of the suspension per 1 kg of live weight per day. The results demonstrated that 0.1 mL of a 1 × 108 per mL enterococcus culture suspension in the poultry diet per 1 kg of live weight per day intensified metabolism and increased the live weight of chickens after 40 d of treatment.

La compétitivité et la rentabilité de l'industrie de la volaille dans les conditions du marché sont désavantagées par les problèmes croissants de sécurité et de qualité de la viande de volaille. La solution à ce problème peut être trouvée grâce à l'utilisation de préparations microbiennes ciblées. De nombreuses études sur les propriétés des bactéries du genre Enterococcus suggèrent leur efficacité dans la pratique de l'aviculture. Le but de l'étude est d'évaluer l'effet de différentes doses d'E. faecium ICIS 96 sur le métabolisme et les qualités productives des poulets à griller. L'expérience a été réalisée sur 72 poulets de chair croisés Cobb-500, répartis en 3 groupes. Les poulets du premier groupe expérimental ont reçu une suspension de culture E. faecium ICIS 96, préparée à raison de 1 milliard de cellules microbiennes dans 1 mL de solution saline. Les poulets de chair du deuxième groupe expérimental ont reçu une dose doublée d'entérocoques, soit 0,2 mL d'une suspension de 1 milliard de cellules microbiennes pour 1 kg de poids vif par jour. Les résultats ont démontré que 0,1 mL d'une suspension de culture d'entérocoques dans l'alimentation des volailles pour 1 kg de poids vif par jour intensifiait le métabolisme et, par conséquent, augmentait le poids vif des poulets dans les 40 jours suivant l'ontogenèse post-incubation.(Traduit par Docteur Serge Messier).

Anti-tumor effect of infant-derived Enterococcus via the inhibition of proliferation and inflammation as well as the promotion of apoptosis.

Probiotic Enterococcus hirae WEHI01 and Enterococcus faecium WEFA23 from infants were previously found to effectively inhibit the development of melanoma. In this study, their immunomodulatory and antitumor mechanisms were systemically studied. In vitro assay showed that E. hirae WEHI01 and E. faecium WEFA23 achieved biphasic immune regulation, which was revealed by the activation of resting spleen lymphocytes and RAW264.7 macrophages, as well as the anti-inflammation effect when immune cells were treated with LPS. The antitumor effects of E. hirae WEHI01 and E. faecium WEFA23 in vitro and vivo were then investigated. CCK8 and the cell scratch assay showed that the conditioned media, which were co-incubated with Enterococcus and spleen lymphocytes, significantly inhibited the proliferation and migration of B16F10, HepG-2 and HT-29 cells. The results of the tumor-bearing mice model experiment showed that E. faecium WEFA23 inhibition of the growth of tumors in mice, and the anti-tumor mechanism involved three aspects, namely tumor proliferation (decreasing expressions of LDHA, VEGF, MMP2, MMP9 and HIF-1α), inhibition of the pro-inflammation state (decreasing expressions of IL-6, TGF-ß and IL-17) and the promotion of apoptosis (increasing expression of Bax/Bcl-2, caspase-3 and p53). The results suggest that the two strains of Enterococcus could be promising candidates for treating melanoma with a highly inhibitory effect.

Thermal inactivation of Salmonella Typhimurium and surrogate Enterococcus faecium in mash broiler feed in a laboratory scale circulated thermal bath.

This study compares kinetic parameters of Salmonella and surrogate Enterococcus faecium in mash broiler feed during thermal inactivation. Two-gram samples of mash broiler feed were added into a filtered sample bag and inoculated with nalidixic acid (NaL, 200 ppm) resistant S. Typhimurium or Enterococcus faecium, followed by vacuum-packaging and heating in a circulated thermal water bath at 75°, 85°, and 95°C for 0 to 180 s. Counts of bacterial survival were analyzed on tryptic soy agar and bile esculin agar plus 200 ppm of NaL. Microbial data and thermal kinetic parameters (n = 8, Global-Fit and United States Department of Agriculture [USDA]-Integrated-Predictive-Modeling-Program software) were analyzed by JMP software. Heating mash broiler feed at 75°, 85°, and 95°C decreased (P < 0.05) Salmonella cell counts by >6 log10CFU/g after 180, 60, and 50 s, respectively. Heating E. faecium in feed at 75°, 85°, and 95°C for 180, 120, and 70 s achieved reductions of 3, 6, and >6.5 log10CFU/g, respectively. D-values of linear, Weibull models, and z-value of Salmonella at 75°, 85°, and 95°C were 1.8 to 11.2, 4.2 to 21.8, and 28.6 s, respectively, which were lower (P < 0.05) than those of E. faecium (3.7-18.1, 8.5-34.4, and 34.1 s). Linear with Tail, Linear with Tail and Shoulder, and Weibull with tail equations revealed that E. faecium were more resistant (P < 0.05) to heat than Salmonella as shown by longer "Shoulder-time" (26.5 vs. 16.2 s) and greater "Tail" effect (4.4-4.5 vs. 2.5-2.6 log10CFU/g). Results clearly suggested that E. faecium can be used as a surrogate for Salmonella to validate thermal inactivation during feed manufacture.

Thermal inactivation kinetics of Salmonella enterica and Enterococcus faecium NRRL B-2354 as a function of temperature and water activity in fine ground black pepper.

Fine ground black pepper generally consumed as a seasoning without any further processing has been associated with Salmonella enterica outbreaks. Thermal inactivation kinetics data is necessary to develop a pasteurization process for fine ground black pepper. This study investigates the influence of temperature and water activity on thermal inactivation kinetics of Salmonella in fine ground black pepper. It also assesses the suitability of Enterococcus faecium as a surrogate for Salmonella. Fine ground black pepper of varying water activities, aw (0.40, 0.55, 0.70) was subjected to isothermal treatments at different temperatures (65-80 °C) for five equidistant time points with intervals ranging from 18 s to 250 min. The survival data were used to fit two primary models (log-linear and Weibull) and two secondary models (response surface and Modified Bigelow). Results indicated that among the two primary models, the Weibull model explained the thermal inactivation kinetics better with lower RMSE (0.24 - 0.56 log CFU/g) and AICc values at all aw and temperatures. Water activity and treatment temperature significantly enhanced the thermal inactivation of Salmonella. E. faecium NRRL B-2354 was found to be a suitable surrogate for Salmonella in fine ground black pepper at all tested treatment conditions. The developed modified Bigelow model based on the Weibull model could be applied to predict the inactivation kinetics of Salmonella in black pepper and would benefit the spice industry in identifying process parameters for thermal pasteurization of fine ground black pepper.

Heat resistance comparison of Salmonella and Enterococcus faecium in cornmeal at different moisture levels.

Adequate surrogate identification is critical for validating in-plant thermal process controls for Salmonella inactivation in different food matrices. This study compared the thermal inactivation parameters (D- and z-values) and evaluated the heat resistance of Enterococcus faecium (8459) as a surrogate for a 5-serovar Salmonella cocktail in cornmeal. The cornmeal was spray inoculated with the respective bacteria to achieve ~9 log CFU/g population and set to the desired moisture contents (16, 22, and 28% w.b.). The inoculated cornmeal was then heat-treated at pre-determined temperatures (60, 64, and 68 °C) in sealed aluminum thermal-death-time disks in hot water baths for pre-determined time intervals. Injury-recovery media [brain heart infusion (BHI) agar overlaid with xylose lysine deoxycholate (XLD) agar for Salmonella or BHI agar overlaid m-enterococcus agar for E. faecium] were used for microbial enumeration to account for thermally injured bacterial cells. The D-values of Salmonella in cornmeal at 16, 22, and 28% moisture content were 37.5, 8.4, and 2.4 min at 60 °C, 19.9, 3.5, and 1.1 min at 64 °C, and 10.1, 1.4, and 0.5 min at 68 °C, respectively. The D-values of E. faecium in cornmeal at 16, 22, and 28% moisture content were 140.4, 18.9, and 3.3 min at 60 °C, 78.4, 7.1, and 1.6 min at 64 °C, and 37.3, 2.8, and 0.8 min at 68 °C, respectively. The z-values of E. faecium and Salmonella in cornmeal at 16, 22, and 28% moisture content were 13.9, 9.7, and 12.5 °C, and 14.0, 10.4, and 11.7 °C, respectively. These results indicated similar or higher thermal resistance (D-values) and equivalent thermal sensitivity (z-values) of E. faecium compared to Salmonella at different moisture contents and respective temperatures (P ≤ 0.05). Therefore, E. faecium could be used as a surrogate for Salmonella during thermal process validation of cornmeal processing.

Control of Salmonella in low-moisture foods: Enterococcus faecium NRRL B-2354 as a surrogate for thermal and non-thermal validation.

Salmonella has been implicated in multiple foodborne outbreaks and recalls associated with low water activity foods (LawF). To verify the effectiveness of a process against Salmonella in LawF, validation using a nonpathogenic surrogate strain is essential. Enterococcus faecium NRRL B-2354 strain has been used as a potential surrogate of Salmonella in different processing of LawF. However, the survival of Salmonella and E. faecium in LawF during food processing is a dynamic function of aw, food composition and structure, processing techniques, and other factors. This review assessed pertinent literature on the thermal and non-thermal inactivation of Salmonella and its presumable surrogate E. faecium in various LawF and provided an overview of its suitibility in different LawF. Overall, based on the D-values, survival/reduction, temperature/time to obtain 4 or 5-log reductions, most studies concluded that E. faecium is a suitable surrogate of Salmonella during LawF processing as its magnitude of resistance was slightly greater or equal (i.e., statistical similar) as compared to Salmonella. Studies also showed its unsuitability which either does not provide a proper margin of safety or being overly resistant and may compromise the quality and organoleptic properties of food. This review provides useful information and guidance for future validation studies of LawF.

Pharmacological Effects of Marine-Derived Enterococcus faecium EA9 against Acute Lung Injury and Inflammation in Cecal Ligated and Punctured Septic Rats.

Microorganisms obtained from the marine environment may represent a potential therapeutic value for multiple diseases. This study explored the possible protective role of marine-derived potential probiotic Enterococcus faecium EA9 (E. faecium) against pulmonary inflammation and oxidative stress using the cecal ligation and puncture (CLP) model of sepsis in Wistar rats. Animals were pretreated with E. faecium for 10 days before either sham or CLP surgeries. Animals were sacrificed 72 hours following the surgical intervention. The histological architecture of lung tissues was evaluated as indicated by the lung injury score. In addition, the extend of pulmonary edema was determined as wet/dry weight ratio. The inflammatory cytokines were estimated in lung tissues, including tumor necrosis factor-alpha (TNF-α), interleukin-6 (IL-6), and interleukin-1 beta (IL-1ß) using the enzyme-linked-immunosorbent-assay (ELISA) technique. Moreover, markers for lipid peroxidation such as thiobarbituric acid reaction substances (TBARs), and endogenous antioxidants, including reduced glutathione (GSH) were determined in lung tissues. Finally, the enzymatic activities of antioxidant enzymes such as catalase (CAT), superoxide dismutase (SOD), glutathione peroxidase (GPx), and glutathione reductase (GR) were assayed in the lungs. Pretreatment with E. faecium markedly attenuated CLP-induced lung injury and pulmonary edema. Markers for inflammation, including TNF-α, IL-6, and IL-1ß were augmented in the lung tissues of CLP animals, while E. faecium ameliorated their augmented levels. E. faecium pretreatment also restored the elevated TBARS levels and the prohibited CAT, SOD, and GPx enzymatic activities in CLP animals. GSH levels were corrected by E. faecium in CLP animals. The inflammatory and lipid peroxidation mediators were positively correlated, while antioxidant enzymatic activities were negatively correlated with CLP-induced lung injury and pulmonary edema. Collectively, marine-derived Enterococcus faecium EA9 might be considered as a prospective therapeutic tool for the management of pulmonary dysfunction associated with sepsis.

Enterococcus faecium PNC01 isolated from the intestinal mucosa of chicken as an alternative for antibiotics to reduce feed conversion rate in broiler chickens.

BACKGROUND: The development and utilization of probiotics had many environmental benefits for replacing antibiotics in animal production. Bacteria in the intestinal mucosa have better adhesion to the host intestinal epithelial cells compared to bacteria in the intestinal contents. In this study, lactic acid bacteria were isolated from the intestinal mucosa of broiler chickens and investigated as the substitution to antibiotic in broiler production. RESULTS: In addition to acid resistance, high temperature resistance, antimicrobial sensitivity tests, and intestinal epithelial cell adhesion, Enterococcus faecium PNC01 (E. faecium PNC01) was showed to be non-cytotoxic to epithelial cells. Draft genome sequence of E. faecium PNC01 predicted that it synthesized bacteriocin to perform probiotic functions and bacteriocin activity assay showed it inhibited Salmonella typhimurium from invading intestinal epithelial cells. Diet supplemented with E. faecium PNC01 increased the ileal villus height and crypt depth in broiler chickens, reduced the relative length of the cecum at day 21, and reduced the relative length of jejunum and ileum at day 42. Diet supplemented with E. faecium PNC01 increased the relative abundance of Firmicutes and Lactobacillus, decreased the relative abundance of Bacteroides in the cecal microbiota. CONCLUSION: E. faecium PNC01 replaced antibiotics to reduce the feed conversion rate. Furthermore, E. faecium PNC01 improved intestinal morphology and altered the composition of microbiota in the cecum to reduce feed conversion rate. Thus, it can be used as an alternative for antibiotics in broiler production to avoid the adverse impact of antibiotics by altering the gut microbiota.

Comparative study of multidrug-resistant Enterococcus faecium obtained from different hosts.

Introduction. The possible transfer of antimicrobial resistance genes between Enterococcus faecium isolates from humans and different animal species, including those not covered by monitoring programs (e.g. pet and wildlife), poses a serious threat to public health.Hypothesis/Gap Statement. Little is known about occurrence and mechanisms of phenomenon of multidrug resistance of E. faecium isolated from various host species in Poland.Aim. The aim of the study was to characterize multidrug-resistant E. faecium isolated from humans and animals (livestock, pets and wildlife) in terms of the occurrence of genetic markers determining resistance.Methodology. Bacterial isolates were tested for phenotypic resistance and the presence of genes encoding resistance to macrolides, tetracycline, aminoglycosides, aminocyclitols and phenicols as well as efflux pump (emeA), resolvase (tndX) and integrase (Int-Tn) genes. The quinolone resistance-determining regions of gyrA and parC were sequenced.Results. Human isolates of E. faecium were characterized by high-level resistance to: ciprofloxacin, enrofloxacin, erythromycin (100 %), as well, as aminoglycosides resistance (kanamycin - 100%, streptomycin - 78 %, gentamicin - 78%). Regardless of the animal species, high level of resistance of E. faecium to tetracycline (from 88-100 %), erythromycin (from 82-94 %) and kanamycin (from 36-100 %) was observed. All E. faecium isolates from wildlife were resistant to fluoroquinolones. However, full susceptibility to vancomycin was observed in all isolates tested. Phenotypic antimicrobial resistance of E. faecium was identified in the presence of the following resistance genes: erm(B) (70%), msr(A) (50 %), tet(L) (35 %), tet(K) (34 %), tet(M) (76 %), aac(6')-Ie-aph(2″)-Ia (25%), ant(6)-Ia (31%), aph(3)-IIIa (68 %), (tndX) (23 %), and integrase gene (Int-Tn) (34 %). A correlation between an amino acid substitution at positions 83 and 87 of gyrA and position 80 of parC and the high-level fluoroquinolone resistance in E. faecium has been observed as well.Conclusion. The level and range of antimicrobial resistance and the panel of resistance determinants is comparable between E. faecium isolates, despite host species.

Evaluation of Enterococcus faecium NRRL B-2354 as a surrogate for Salmonella in ground black pepper at different water activities.

Thermal inactivation kinetics of Salmonella in low moisture foods are necessary for developing proper thermal processing parameters for pasteurization. The effect of water activity on thermal inactivation kinetics of Salmonella and Enterococcus faecium NRRL B-2354 in ground black pepper has not been studied previously. Identification of a suitable surrogate assists in conducting in-plant process validations. Ground black pepper was inoculated with a 5-serotype Salmonella cocktail or E. faecium NRRL B-2354, equilibrated to water activities of 0.25, 0.45 or 0.65 in a humidity-controlled chamber, and isothermally treated at different temperatures. The survivor data were used for fitting the log-linear models to obtain the D and z-values of Salmonella and E. faecium in ground black pepper. Modified Bigelow models were developed to evaluate the effects of temperature and water activity on the thermal inactivation kinetics of Salmonella and E. faecium. Water activity and temperature showed significant negative effects on the thermal resistance of Salmonella and E. faecium in ground black pepper. For example, significantly higher D values of Salmonella were observed at water activity of 0.45 (D70°C = 20.5 min and D75°C = 7.8 min) compared to water activity of 0.65 (D70°C = 3.9 min and D75°C = 2.0 min). D-values of E. faecium were significantly higher than those of Salmonella at all three water activities, indicating that E. faecium is a suitable surrogate for Salmonella in thermal processing validation.

The global dissemination of hospital clones of Enterococcus faecium.

BACKGROUND: The hospital-adapted A1 group of Enterococcus faecium remains an organism of significant concern in the context of drug-resistant hospital-associated infections. How this pathogen evolves and disseminates remains poorly understood. METHODS: A large, globally representative collection of short-read genomic data from the hospital-associated A1 group of Enterococcus faecium was assembled (n = 973). We analysed, using a novel analysis approach, global diversity in terms of both the dynamics of the accessory genome and homologous recombination among conserved genes. RESULTS: Two main modes of genomic evolution continue to shape E. faecium: the acquisition and loss of genes, including antimicrobial resistance genes, through mobile genetic elements including plasmids, and homologous recombination of the core genome. These events lead to new clones emerging at the local level, followed by the erosion of signals of clonality through recombination, and in some identifiable cases producing new clonal clusters. These patterns lead to new, emerging lineages which are able to spread globally over relatively short timeframes. CONCLUSIONS: The ability of A1 E. faecium to continually present new combinations of genes for potential selection suggests that controlling this pathogen will remain challenging but establishing a framework for understanding genomic evolution is likely to aid in tracking the threats posed by newly emerging lineages.

Differential analysis of gut microbiota and the effect of dietary Enterococcus faecium supplementation in broiler breeders with high or low laying performance.

The difference in microbiota was examined for breeders with different egg-laying rates, and the impact of dietary Enterococcus faecium (EF) was also determined in the present study. A total of 256 Arbor Acres broiler breeders (48-wk-old) were used in a 2 × 2 factorial design, which encompassed 2 egg-laying rate levels [average (average egg laying: AP, 80.45 ± 0.91%) and low (lower egg laying: LP, 70.61 ± 1.16%)] and 2 different dietary groups [control (no additive), 6 × 108 cfu/kg EF]. The results showed that the AP breeders presented a lower egg weight, feed conversion ratio, abdominal fat rate, and serum leptin level (P(laying) ≤ 0.05) as well as a higher egg-laying rate (P(laying) < 0.01) than the LP breeders. Dietary supplementation with EF improved the egg weight (P(EF) = 0.03) and had a higher concentration of follicle-stimulating hormone (FSH) in the serum (P(EF) = 0.04). The relative expression of Caspase 9, Bax, AMHR, BMP15, and GATA4 in the ovary of AP breeders was lower, whereas the FSHR and BMPR1B expression was higher than that measured in LP breeders (P(laying) ≤ 0.05). LP increased the abundance of Bacteroidetes (phylum), Firmicutes (phylum), Bacteroidia (class), Clostridia (class), Bacteroidales (order), Clostridiales (order), and Lachnospiraceae (family), whereas the AP promoted the enrichment of Proteobacteria (phylum) and Gammaproteobacteria (class) (P(laying) < 0.05). The genera Bacillus, Rhodanobacter, and Streptomyces were positively correlated with the egg-laying rate and BMPR1B expression (P < 0.05) but negatively correlated with the abdominal fat rate (P < 0.05) and Caspase 9 (P < 0.05). These findings indicate that the low reproductive performance breeders had lower microbiota diversity and higher Firmicutes, which triggers the energy storage that led to higher fat deposition. Besides, increases in the abdominal fat rate, leptin level, and apoptosis (Caspase 9, Bax) and reproduction-related gene (BMP15, AMHR, BMPR1B, and GATA4) expression would possibly be the potential mechanisms under which breeders have different reproductive performance. Dietary EF increased the egg weight and serum FSH level and decreased the Bacteroidetes (phylum) in low reproductive breeders.

Competitiveness of three biocontrol candidates against ochratoxigenic Penicillium nordicum under dry-cured meat environmental and nutritional conditions.

The environmental conditions during the ripening of dry-cured meats and their nutritional composition promote the colonisation of their surface by Penicillium spp., including P. nordicum producer of ochratoxin A (OTA). The objective of this work was to study the competitiveness of three potential biocontrol candidates (Debaryomyces hansenii FHSCC 253H, Enterococcus faecium SE920 and Penicillium chrysogenum CECT, 20922) against the ochratoxigenic P. nordicum FHSCC4 under environmental and nutritional conditions simulating the ripening of dry-cured meat products. For this, the nutritional utilisation pattern, niche overlap index (NOI), interactions by dual-culture assays and OTA production were determined. The number of carbon sources (CSs) metabolised depended on the microorganism and the interacting water activity (aw) x temperature conditions. The number of CSs utilised by both filamentous fungi was quite similar and higher than those utilised by D. hansenii and E. faecium. The yeast isolate metabolised a number of CSs much larger than the bacterium. The NOI values showed that, in general, P. nordicum nutritionally dominated E. faecium and D. hansenii regardless of the environmental conditions evaluated. The relationship between the toxigenic and non-toxigenic fungal isolates depended on the aw x temperature combinations, although in none of the conditions a dominance of P. nordicum was observed. According to the interaction assays, both D. hansenii and P. chrysogenum decreased the growth of P. nordicum. The effect of D. hansenii could be attributed to the production of some extra-cellular compounds, while the action of P. chrysogenum is likely related to nutritional competition. In addition, both P. chrysogenum and D. hansenii reduced the OTA levels produced by P. nordicum. The effect of the yeast was more pronounced decreasing the concentration of OTA at quantities lower than the limit established by the Italian legislation. Therefore, P. chrysogenum and D. hansenii can be suggested as biocontrol candidates in the manufacture of dry-cured meat products.

Mode and dynamics of vanA-type vancomycin resistance dissemination in Dutch hospitals.

BACKGROUND: Enterococcus faecium is a commensal of the gastrointestinal tract of animals and humans but also a causative agent of hospital-acquired infections. Resistance against glycopeptides and to vancomycin has motivated the inclusion of E. faecium in the WHO global priority list. Vancomycin resistance can be conferred by the vanA gene cluster on the transposon Tn1546, which is frequently present in plasmids. The vanA gene cluster can be disseminated clonally but also horizontally either by plasmid dissemination or by Tn1546 transposition between different genomic locations. METHODS: We performed a retrospective study of the genomic epidemiology of 309 vancomycin-resistant E. faecium (VRE) isolates across 32 Dutch hospitals (2012-2015). Genomic information regarding clonality and Tn1546 characterization was extracted using hierBAPS sequence clusters (SC) and TETyper, respectively. Plasmids were predicted using gplas in combination with a network approach based on shared k-mer content. Next, we conducted a pairwise comparison between isolates sharing a potential epidemiological link to elucidate whether clonal, plasmid, or Tn1546 spread accounted for vanA-type resistance dissemination. RESULTS: On average, we estimated that 59% of VRE cases with a potential epidemiological link were unrelated which was defined as VRE pairs with a distinct Tn1546 variant. Clonal dissemination accounted for 32% cases in which the same SC and Tn1546 variants were identified. Horizontal plasmid dissemination accounted for 7% of VRE cases, in which we observed VRE pairs belonging to a distinct SC but carrying an identical plasmid and Tn1546 variant. In 2% of cases, we observed the same Tn1546 variant in distinct SC and plasmid types which could be explained by mixed and consecutive events of clonal and plasmid dissemination. CONCLUSIONS: In related VRE cases, the dissemination of the vanA gene cluster in Dutch hospitals between 2012 and 2015 was dominated by clonal spread. However, we also identified outbreak settings with high frequencies of plasmid dissemination in which the spread of resistance was mainly driven by horizontal gene transfer (HGT). This study demonstrates the feasibility of distinguishing between modes of dissemination with short-read data and provides a novel assessment to estimate the relative contribution of nested genomic elements in the dissemination of vanA-type resistance.

Lactiplantibacillus plantarum dfa1 Outperforms Enterococcus faecium dfa1 on Anti-Obesity in High Fat-Induced Obesity Mice Possibly through the Differences in Gut Dysbiosis Attenuation, despite the Similar Anti-Inflammatory Properties.

Fat reduction and anti-inflammation are commonly claimed properties of probiotics. Lactiplantibacillus plantarum and Enterococcus faecium were tested in high fat-induced obesity mice and in vitro experiments. After 16 weeks of probiotics, L. plantarum dfa1 outperforms E. faecium dfa1 on the anti-obesity property as indicated by body weight, regional fat accumulation, serum cholesterol, inflammatory cytokines (in blood and colon tissue), and gut barrier defect (FITC-dextran assay). With fecal microbiome analysis, L. plantarum dfa1 but not E. faecium dfa1 reduced fecal abundance of pathogenic Proteobacteria without an alteration in total Gram-negative bacteria when compared with non-probiotics obese mice. With palmitic acid induction, the condition media from both probiotics similarly attenuated supernatant IL-8, improved enterocyte integrity and down-regulated cholesterol absorption-associated genes in Caco-2 cell (an enterocyte cell line) and reduced supernatant cytokines (TNF-α and IL-6) with normalization of cell energy status (extracellular flux analysis) in bone-marrow-derived macrophages. Due to the anti-inflammatory effect of the condition media of both probiotics on palmitic acid-activated enterocytes was neutralized by amylase, the active anti-inflammatory molecules might, partly, be exopolysaccharides. As L. plantarum dfa1 out-performed E. faecium dfa1 in anti-obesity property, possibly through the reduced fecal Proteobacteria, with a similar anti-inflammatory exopolysaccharide; L. plantarum is a potentially better option for anti-obesity than E. faecium.

Influence of low water activity on the thermal resistance of Salmonella Enteritidis PT30 and Enterococcus faecium as its surrogate in egg powders.

Egg powders are increasingly popular ingredients, due to their functionality and compactness, in industrial food production and preparation at homes. However, there is a lack of studies that evaluate the thermal resistance of Salmonella Enteritidis PT30 and its potential surrogate Enterococcus faecium NRRL B-2354 in egg powders. This study examined the log-linear relationship between the thermal resistance of Salmonella Enteritidis (D-value) and the water activity (aw) of egg powders. The changes of aw in the egg powders with temperature were measured using a Vapor Sorption Analyzer and a high-temperature cell. The D80 ℃-value of S. Enteritidis PT30 and E. faecium inoculated in the egg powders preconditioned to three aw levels (0.3, 0.45, and 0.6) at 20 ℃ were determined using aluminum thermal death test cells. The aw values increased (P < 0.05) in all three egg powders when the temperature of the samples was raised from room temperature to 80 ℃. The D80 ℃-values ranged from 5.3 ± 0.1 to 25.9 ± 0.2 min for S. Enteritidis while 10.4 ± 0.4 to 43.8 ± 0.4 for E. faecium in samples of the three different aw levels. S. Enteritidis PT30 showed a log-linear relationship between D80 ℃-values and aw80 ℃ for the egg powders. This study contributes to our understanding of the impact of aw on the development of thermal treatments for low-moisture foods.

Enterococcus faecium R0026 Combined with Bacillus subtilis R0179 Prevent Obesity-Associated Hyperlipidemia and Modulate Gut Microbiota in C57BL/6 Mice.

Bacillus subtilis and Enterococcus faecium are commonly used probiotics. This study aimed to identify the effect of live combined Bacillus subtilis R0179 and Enterococcus faecium R0026 (LCBE) on obesityassociated hyperlipidemia and gut microbiota in C57BL/6 mice. Forty male C57BL/6 mice were divided into four groups: normal group (N group), model group (M group), low-dose group (L group), and high-dose group (H group). Mice were gavaged with LCBE at 0.023 g/mice/day (L group) or 0.23 g/mice/day (H group) and fed with a high-fat diet for 8 weeks. In vitro E. faecium R0026 showed an ability to lower the low-concentration of cholesterol by 46%, and the ability to lower the highconcentration of cholesterol by 58%. LCBE significantly reduced the body weight gain, Lee index, brown fat index and body mass index of mice on a high-fat diet. Moreover, LCBE markedly improved serum lipids (including serum triglyceride, total cholesterol, low-density lipoprotein and highdensity lipoprotein) while also significantly reducing liver total cholesterol. Serum lipopolysaccharide and total bile acid in L and H groups decreased significantly compared with M group. PCR-DGGE analysis showed that the composition of gut microbiota in the treatment groups was improved. Akkermansia muciniphila was found in H group. The PCA result indicated a similar gut microbiota structure between LCBE treatment groups and normal group while the number of bands and Shannon diversity index increased significantly in the LCBE treatment groups. Finally, qPCR showed Bifidobacterium spp. increased significantly in H group compared with M group, LCBE alleviated liver steatosis and improved brown adipose tissue index.