Prandial Correlations and Structure of the Ingestive Behavior of Pigs in Precision Feeding Programs.

The feeding behavior of growing-finishing pigs was analyzed to study prandial correlations and the probability of starting a new feeding event. The data were collected in real-time based on 157,632 visits by a group of 70 growing-finishing pigs (from 30.4 to 115.5 kg body weight, BW) to automatic feeders. The data were collected over 84 days, during which period the pigs were kept in conventional (by phase and by group) or precision (with daily and individual adjustments) feeding programs. A criterion to delimit each meal was then defined based on the probability of an animal starting a new feeding event within the next minute since the last visit. Prandial correlations were established between meal size and interval before meal (pre-prandial) or interval after meal (post-prandial) using Pearson correlation analysis. Post-prandial correlations (which can be interpreted as hunger-regulating mechanisms) were slightly stronger than pre-prandial correlations (which can be interpreted as satiety regulation mechanisms). Both correlations decreased as the animals' age increased but were little influenced by the feeding programs. The information generated in this study allows a better understanding of pigs' feeding behavior regulation mechanisms and could be used in the future to improve precision feeding programs.

Nitric oxide-releasing chitosan nanoparticles alleviate the effects of salt stress in maize plants.

Nitric oxide (NO) is a signaling molecule involved in plant response to various abiotic stresses. However, the application of NO donors in agriculture is hampered by the instability of these compounds. Despite the successful uses of NO-releasing nanoparticles for biomedical purposes and the variety of nanomaterials developed as carrier systems of agrochemicals, the potential applications of nanocarriers for NO delivery in plants have not yet been tested. Herein, we report the synthesis and characterization of chitosan nanoparticles (CS NPs) containing the NO donor S-nitroso-mercaptosuccinic acid (S-nitroso-MSA). The efficiency of these NO-releasing NPs in mitigating the deleterious effects of salinity on maize plants was compared to that of the non-encapsulated NO donor. The NPs were synthesized through ionotropic gelation process, and mercaptosuccinic acid (MSA), the NO donor precursor, was encapsulated into CS NPs (91.07% encapsulation efficiency). Free thiol groups of MSA-CS NPs were nitrosated, leading to S-nitroso-MSA-CS NPs (NO-releasing NPs). The incorporation of S-nitroso-MSA into CS NPs allowed a sustained NO release. Treatments of salt-stressed maize plants with S-nitroso-MSA-CS NPs resulted in a higher leaf S-nitrosothiols content compared to that of free S-nitroso-MSA. Moreover, S-nitroso-MSA-CS NPs were more efficient than was the free NO donor in the amelioration of the deleterious effects of salinity in photosystem II activity, chlorophyll content and growth of maize plants because the protective action of the nanoencapsulated S-nitroso-MSA was achieved at lower dosages. Overall, these results demonstrate the positive impact of S-nitroso-MSA nanoencapsulation in increasing NO bioactivity in maize plants under salt stress.

In vitro evaluation of the probiotic potential of bacteriocin producer Lactobacillus sakei 1.

Lactobacillus sakei 1 is a food isolate that produces a heat-stable antimicrobial peptide (sakacin 1, a class IIa bacteriocin) inhibitory to the opportunistic pathogen Listeria monocytogenes. Bacterial isolates with antimicrobial activity may be useful for food biopreservation and also for developing probiotics. To evaluate the probiotic potential of L. sakei 1, it was tested for (i) in vitro gastric resistance (with synthetic gastric juice adjusted to pH 2.0, 2.5, or 3.0); (ii) survival and bacteriocin production in the presence of bile salts and commercial prebiotics (inulin and oligofructose); (iii) adhesion to Caco-2 cells; and (iv) effect on the adhesion of L. monocytogenes to Caco-2 cells and invasion of these cells by the organism. The results showed that L. sakei 1 survival in gastric environment varied according to pH, with the maximum survival achieved at pH 3.0, despite a 4-log reduction of the population after 3 h. Regarding the bile salt tolerance and influence of prebiotics, it was observed that L. sakei 1 survival rates were similar (P > 0.05) for all de Man Rogosa Sharpe (MRS) broth formulations when tests were done after 4 h of incubation. However, after incubation for 24 h, the survival of L. sakei 1 in MRS broth was reduced by 1.8 log (P < 0.001), when glucose was replaced by either inulin or oligofructose (without Oxgall). L. sakei 1 was unable to deconjugate bile salts, and there was a significant decrease (1.4 log) of the L. sakei 1 population in regular MRS broth plus Oxgall (P < 0.05). In spite of this, tolerance levels of L. sakei 1 to bile salts were similar in regular MRS broth and in MRS broth with oligofructose. Lower bacteriocin production was observed in MRS broth when inulin (3,200 AU/ml) or oligofructose (2,400 AU/ml) was used instead of glucose (6,400 AU/ml). L. sakei 1 adhered to Caco-2 cells, and its cell-free pH-neutralized supernatant containing sakacin 1 led to a significant reduction of in vitro listerial invasion of human intestinal Caco-2 cells.

Vaginal lactobacilli as potential probiotics against Candida spp

Urogenital infections affect millions of people every year worldwide. The treatment of these diseases usually requires the use of antimicrobial agents, and more recently, the use of probiotic lactic acid bacteria (LAB) cultures for the management of vaginal infections has been extensively studied. In this work, 11 vaginal lactobacilli isolates, previously obtained from healthy patients, were studied to screen microorganisms with probiotic properties against Candida spp. The LAB were tested for their ability of auto-aggregation, coaggregation with C. albicans, C. glabrata, C. krusei, and C. tropicalis, adhesion to Caco-2 epithelial cells and production of lactic acid and hydrogen peroxide (H2O2). All lactobacilli isolates tested were able to auto-aggregate (ranging from 25.3 percent to 75.4 percent assessed at 4 hours of incubation) and to co-aggregate with the four Candida species into different degrees; among them L. crispatus showed the highest scores of coaggregation. The highest amount of lactic acid was produced by L. salivarius (13.9 g/l), followed by L. johnsonii (6.5 g/l), L. acidophilus (5.5 g/l), and L. jensenii (5.4 g/l). All isolates produced H2O2 , but the highest levels (3 -10 mg/l) were observed for L. acidophilus, L. crispatus, L. gasseri, L. johnsonii, and L. vaginalis. Only L. agilis, L. jensenii, L. johnsonii and L. ruminus were able to adhere to epithelial Caco-2 cells. Among the isolates evaluated, L agilis, L. jensenii, L. johnsonii, and L. ruminus exhibited simultaneously several desirable properties as potential probiotic strains justifying future studies to evaluate their technological properties in different pharmaceutical preparations for human use.

Vaginal lactobacilli as potential probiotics against Candida SPP.

Urogenital infections affect millions of people every year worldwide. The treatment of these diseases usually requires the use of antimicrobial agents, and more recently, the use of probiotic lactic acid bacteria (LAB) cultures for the management of vaginal infections has been extensively studied. In this work, 11 vaginal lactobacilli isolates, previously obtained from healthy patients, were studied to screen microorganisms with probiotic properties against Candida spp. The LAB were tested for their ability of auto-aggregation, co-aggregation with C. albicans, C. glabrata, C. krusei, and C. tropicalis, adhesion to Caco-2 epithelial cells and production of lactic acid and hydrogen peroxide (H2O2). All lactobacilli isolates tested were able to auto-aggregate (ranging from 25.3% to 75.4% assessed at 4 hours of incubation) and to co-aggregate with the four Candida species into different degrees; among them L. crispatus showed the highest scores of co-aggregation. The highest amount of lactic acid was produced by L. salivarius (13.9 g/l), followed by L. johnsonii (6.5 g/l), L. acidophilus (5.5 g/l), and L. jensenii (5.4 g/l). All isolates produced H2O2, but the highest levels (3 - 10 mg/l) were observed for L. acidophilus, L. crispatus, L. gasseri, L. johnsonii, and L. vaginalis. Only L. agilis, L. jensenii, L. johnsonii and L. ruminus were able to adhere to epithelial Caco-2 cells. Among the isolates evaluated, L agilis, L. jensenii, L. johnsonii, and L. ruminus exhibited simultaneously several desirable properties as potential probiotic strains justifying future studies to evaluate their technological properties in different pharmaceutical preparations for human use.

Improved cure of bacterial vaginosis with single dose of tinidazole (2 g), Lactobacillus rhamnosus GR-1, and Lactobacillus reuteri RC-14: a randomized, double-blind, placebo-controlled trial.

Bacterial vaginosis (BV) is the most prevalent vaginal infection worldwide and is characterized by depletion of the indigenous lactobacilli. Antimicrobial therapy is often ineffective. We hypothesized that probiotic Lactobacillus rhamnosus GR-1 and Lactobacillus reuteri RC-14 might provide an adjunct to antimicrobial treatment and improve cure rates. Sixty-four Brazilian women diagnosed with BV were randomly assigned to receive a single dose of tinidazole (2 g) supplemented with either 2 placebo capsules or 2 capsules containing L. rhamnosus GR-1 and L. reuteri RC-14 every morning for the following 4 weeks. At the end of treatment (day 28), the probiotic group had a significantly higher cure rate of BV (87.5%) than the placebo group (50.0%) (p = 0.001). In addition, according to the Gram-stain Nugent score, more women were assessed with "normal" vaginal microbiota in the probiotic group (75.0% vs. 34.4% in the placebo group; p = 0.011). This study shows that probiotic lactobacilli can provide benefits to women being treated with antibiotics for an infectious condition.

Prevalence and characterization of Enterococcus spp. isolated from Brazilian foods.

Enterococci can be used in the food industry as starter or probiotic cultures. However, enterococci are also implicated in severe multi-resistant nosocomial infections. In this study, the prevalence of enterococci in selected Brazilian foodstuffs (raw and pasteurized milk, meat products, cheeses and vegetables) was evaluated. Phenotypic and PCR protocols were used for species identification. Tests for production of gelatinase, haemolysin, bacteriocin and bile salt hydrolysis were done with all enterococci isolates, whereas molecular determination of virulence markers (genes esp, gel, ace, as, efaA, hyl and cylA) and antibiotic resistance was checked only for Enterococcus faecium and Enterococcus faecalis isolates. The antibiotic-resistant isolates were assayed for biofilm formation and adhesion to mammalian cells. From the 120 food samples analyzed, 52.5% were positive for enterococci, meat and cheese being the most contaminated. E. faecium was the predominant species, followed by E. faecalis, E. casseliflavus and Enterococcus gallinarum. Phenotypic tests indicated that 67.7% of isolates hydrolyzed bile salts, 15.2% produced bacteriocin, 12.0% were beta-hemolytic and 18.2% produced gelatinase. Antibiotic resistance (gentamicin, tetracycline and erythromycin) and genes encoding for virulence traits were more frequent in E. faecalis than in E. faecium. Three E. faecium isolates were resistant to vancomycin. Among antibiotic-resistant isolates, 72.4% of E. faecalis were able to form biofilm and 13.8% to adhere to Caco-2 cells. Antibiotic-resistant E. faecalis and E. faecium isolates were grouped by RAPD-PCR and a scattered distribution was noted, indicating that resistance was not related to a particular clone. The spread of virulence/resistance traits in isolates of the two species and different RAPD-types suggest the pathogenic potential of both species. By contrast, the recovery of bacteriocinogenic E. faecium isolates with no virulence traits suggests their potential for biotechnological applications. In conclusion, our results showed that enterococci from Brazilian foods present important dualist aspects for food safety.

Influence of peroxyacetic acid and nisin and coculture with Enterococcus faecium on Listeria monocytogenes biofilm formation.

Biofilm formation is a matter of concern in food industries because biofilms facilitate the survival of pathogenic bacteria such as Listeria monocytogenes, which may contaminate food-processing equipment and products. In this study, nisin and two Enterococcus faecium strains were evaluated for their effect on biofilm formation by L. monocytogenes cultured in brain heart infusion broth and on stainless steel coupons. Elimination of preformed L. monocytogenes biofilms by peroxyacetic acid also was tested. Adhesion control experiments were performed with pure cultures of L. monocytogenes after swab collection of adhered cells, which were then enumerated on PALCAM agar plates and visualized by scanning electron microscopy. Formation of a biofilm was recorded when the number of adhered cells was at least 10(3) CFU/cm2. When L. monocytogenes was cocultured with E. faecium bac-, the number of adhered L. monocytogenes cells was 2.5 log lower (P = 0.002) when initially compared with the control culture, but after 6 h of incubation a biofilm was again detected. However, in coculture on stainless steel coupons, E. faecium bac+ inhibited L. monocytogenes adherence and did not allow biofilm formation for up to 48 h (P < 0.001). In the presence of nisin or after treatment with peroxyacetic acid, bacterial growth was reduced (P < 0.001) up to 4.6 and 5.6 log CFU/cm2, respectively, when compared with L. monocytogenes cultures on untreated coupons. However, after these treatments, cells were still present, and after 24 h of incubation, a renewed biofilm was detected in L. monocytogenes cultures treated with nisin. Although all tested conditions reduced L. monocytogenes growth to some extent, only coculture with E. faecium bac+ efficiently reduced biofilm formation, suggesting a potential control strategy for this pathogen.

Correlation between API 20 STREP and multiplex PCR for identification of Enterococcus spp. isolated from Brazilian foods

We evaluated the suitability of API 20 STREP and multiplex PCR to speciate 52 Enterococcus spp. obtained from Brazilian foods. A high percentage of isolates (78.9 percent) presented discrepant results between evaluated tests. Similar results were obtained for six E. faecalis and five E. faecium. The PCR multiplex was more effective than API 20 STREP for complete identification of the isolates.

A identificação das espécies de 52 Enterococcus spp. isolados de amostras de alimentos foi realizada empregando-se duas metodologias: sistema API 20 STREP e PCR multiplex. Os resultados obtidos revelaram que 78,9 por cento dos isolados apresentaram resultados diferentes nos dois testes utilizados. Apenas seis E. faecalis e cinco E. faecium apresentaram resultados concordantes pelos dois métodos. PCR multiplex permitiu a identificação completa de um número significantemente maior de enterococos do que o sistema API 20 STREP.

Fate of Helicobacter pylori artificially inoculated in lettuce and carrot samples

H. pylori é o agente da maioria dos casos de gastrites e úlceras pépticas. Entretanto, sua forma de transmissão não foi completamente elucidada. Para avaliar a sobrevivência de H. pylori (uma cepa de referência e uma clínica), artificialmente inoculado em amostras de cenoura e alface, porções de 25g das amostras receberam aproximadamente 106 UFC/g de H. pylori, e foram embaladas em atmosfera normal e/ou modificada (3 per center oxigênio, 10 per center dióxido de carbono, e 87 per center nitrogênio). Em seguida, foram armazenadas a 8ºC, com enumeração diária da população de H. pylori em ágar Columbia sangue (CBA) e/ou Helicobacter pylori Special Peptone Agar (HPSPA). Quando usamos CBA com antibióticos, o isolado clínico de H. pylori (HP1) foi detectado por até 72 horas nas amostras de alface e cenoura sanitizadas. Em amostras de cenoura esterilizadas, H. pylori HP1 permaneceu viável por até 96 horas. CBA sem antibióticos permitiu a recuperação de H. pylori ATCC 43629, a partir de amostras de cenoura embaladas em ambas as atmosferas, por até 120 horas. Nossos resultados reforçam que a transmissão de H. pylori, através do consumo de água e alimento contaminados, ainda não pode ser descartado.