Patricia Burns

Ninety-eight Lactobacillus plantarum strains isolated from Italian and Argentinean cheeses were evaluated for probiotic potential. After a preliminary subtractive screening based on the presence of msa and bsh genes, 27 strains were characterized. In general, the selected strains showed high resistance to lysozyme, good adaptation to simulated gastric juice, and a moderate to low bile tolerance. The capacity to agglutinate yeast cells in a mannose-specific manner, as well as the cell surface hydrophobicity was found to be variable among strains. Very high β-galactosidase activity was shown by a considerable number of the tested strains, whereas variable prebiotic utilization ability was observed. Only tetracycline resistance was observed in two highly resistant strains which harbored the tetM gene, whereas none of the strains showed β-glucuronidase activity or was capable of inhibiting pathogens. Three strains (Lp790, Lp813, and Lp998) were tested by in vivo trials. A considerable heterogeneity was found among a number of L. plantarum strains screened in this study, leading to the design of multiple cultures to cooperatively link strains showing the widest range of useful traits. Among the selected strains, Lp790, Lp813, and Lp998 showed the best probiotic potential and would be promising candidates for inclusion as starter cultures for the manufacture of probiotic fermented foods.

High-pressure homogenization (HPH) is one of the most promising alternatives to traditional thermal treatment of food preservation and diversification. Its effectiveness on the deactivation of pathogenic and spoilage microorganisms in model systems and real food is well documented. To evaluate the potential of milk treated by HPH for the production of Crescenza cheese with commercial probiotic lactobacilli added, 4 types of cheeses were made: HPH (from HPH-treated milk), P (from pasteurized milk), HPH-P (HPH-treated milk plus probiotics), and P-P (pasteurized milk plus probiotics) cheeses. A strain of Streptococcus thermophilus was used as starter culture for cheese production. Compositional, microbiological, physicochemical, and organoleptic analyses were carried out at 1, 5, 8, and 12 d of refrigerated storage (4°C). According to results obtained, no significant differences among the 4 cheese types were observed for gross composition (protein, fat, moisture) and pH. Differently, the HPH treatment of milk increased the cheese yield about 1% and positively affected the viability during the refrigerated storage of the probiotic bacteria. In fact, after 12 d of storage, the Lactobacillus paracasei A13 cell loads were 8 log cfu/ g, whereas Lactobacillus acidophilus H5 exhibited, in P-P cheese, a cell load decrease of about 1 log cfu/g with respect to the HPH-P cheese. The hyperbaric treatment had a significant positive effect on free fatty acids release and cheese proteolysis. Also, probiotic cultures affected proteolytic and lipolytic cheese patterns. No significant differences were found for the sensory descriptors salty and creamy among HPH and P cheeses as well as for acid, piquant, sweet, milky, salty, creamy, and overall acceptance among HPH, HPH-P, and P-P Crescenza cheeses.

Probiotic bacteria, according to the definition adopted by the World Health Organization in 2002, are live microorganisms, which when administered in adequate amounts confer a health benefit to the host. Recent studies show that the same probiotic strain produced and/or preserved under different storage conditions, may present different responses regarding their susceptibility to the adverse conditions of the gastrointestinal tract, its capacity to adhere to the intestinal epithelium, or its immunomodulating capacity, being the functionality affected without changes in cell viability. This could imply that the control of cell viability is not always enough to guarantee the functionality (probiotic capacity) of a strain. Therefore, a new challenge arises for food technologists and microbiologists when it comes to designing and monitoring probiotic food: to be able to monitor the cell functionality a probiotic microorganism along all the stages the strain goes through from the moment it is produced and included into the food vehicle until to the moment of consumption. Conventional methodological tools or others still to be developed must be used. The application of cell membrane functionality markers, the use of tests of resistence to intestinal barriers, the study of surface properties and the application of in vivo models comes together as complementary tools to assess the actual capacity of a probiotic into a specific food to exert functional effects regardless the number of viable cells present at the moment of consumption.

To determine the protective capacity against Salmonella infection in mice of the cell-free fraction (postbiotic) of fermented milk, produced at laboratory and industrial level.The proteolytic activity (PA) of 5 commercial cultures and 11 autochthonous Lactobacillus strains was evaluated. The DSM-100H culture displayed the highest PA and it was selected for further studies. The capacity of the postbiotics produced by pH-controlled fermentation to stimulate the production of secretory IgA in faeces and to protect mice against Salmonella infection was evaluated. A significant increase in secretory IgA in faeces of mice fed 14 days the postbiotic obtained at the laboratory (F36) was detected compared to control animals. A significantly higher survival was observed in mice fed the F36 and the FiSD (industrial product) compared to controls.The postbiotics obtained showed immunomodulatory and protective capacity against Salmonella infection in mice.The pH-controlled milk fermentation by the proteolytic DSM-100H culture could be a suitable strategy to obtain a food ingredient to be added to a given food matrix, not adequate to host viable cells of probiotics, to confer it enhanced functionality and thus expand the functional food market.

Abstract Domesticated horses live under different conditions compared with their extinct wild ancestors. While housed, medicated and kept on a restricted source of feed, the microbiota of domesticated horses is hypothesized to be altered. We assessed the fecal microbiome of 57 domestic and feral horses from different locations on three continents, observing geographical differences. A higher abundance of eukaryota ( p < 0.05) and viruses ( p < 0.05) and lower of archaea ( p < 0.05) were found in feral animals when compared with domestic ones. The abundance of genes coding for microbe-produced enzymes involved in the metabolism of carbohydrates was significantly higher ( p < 0.05) in feral animals regardless of the geographic origin. Differences in the fecal resistomes between both groups of animals were also noted. The domestic/captive horse microbiomes were enriched in genes conferring resistance to tetracycline, likely reflecting the use of this antibiotic in the management of these animals. Our data showed an impoverishment of the fecal microbiome in domestic horses with diet, antibiotic exposure and hygiene being likely drivers. The results offer a view of the intestinal microbiome of horses and the impact of domestication or captivity, which may uncover novel targets for modulating the microbiome of horses to enhance animal health and well-being.

Progressive adaptation to bile might render some lactobacilli able to withstand physiological bile salt concentrations. In this work, the adaptation to bile was evaluated on previously isolated dairy strains of Lactobacillus delbrueckii subsp. lactis 200 and L. delbrueckii subsp. lactis 200+, a strain derived thereof with stable bile-resistant phenotype. The adaptation to bile was obtained by comparing cytosolic proteomes of both strains grown in the presence or absence of bile. Proteomics were complemented with physiological studies on both strains focusing on glycolytic end-products, the ability to adhere to the human intestinal epithelial cell line HT29-MTX and survival to simulated gastrointestinal conditions. Protein pattern comparison of strains grown with and without bile allowed us to identify 9 different proteins whose production was regulated by bile in both strains, and 17 proteins that showed differences in their levels between the parental and the bile-resistant derivative. These included general stress response chaperones, proteins involved in transcription and translation, in peptidoglycan/exopolysaccharide biosynthesis, in the lipid and nucleotide metabolism and several glycolytic and pyruvate catabolism enzymes. Differences in the level of metabolic end-products of the sugar catabolism were found between the strains 200 and 200+. A decrease in the adhesion of both strains to the intestinal cell line was detected in the presence of bile. In simulated gastric and intestinal juices, a protective effect was exerted by milk improving the survival of both microorganisms. These results indicate that bile tolerance in L. delbrueckii subsp. lactis involves several mechanisms responding to the deleterious impact of bile salts on bacterial physiology.

The influence of two cheese-isolated Lactobacillus strains on cheese composition, acceptability and probiotic capacity was assessed. Soft cheeses with and without the addition of Lactobacillus plantarum I91 or Lactobacillus paracasei I90 were prepared. Gross composition was assessed and secondary proteolysis was described by soluble fractions and free amino acids profiles. Acceptability was determined by a panel of 98 non-trained consumers. Cheeses harboring added Lactobacillus strains were also studied in vivo to evaluate their probiotic capacity. Gross composition of the cheeses was similar for control and treated (Lactobacillus-added) cheeses. Peptidolysis increased in cheeses with added lactobacilli, which was evidenced by a higher free amino acid content. Overall, the acceptability of the cheeses was good: 65%–80% of the consumers said that they “liked very much” or “liked” the cheeses. Cheeses with L. plantarum I91 showed the highest changes in composition and proteolysis and were the most accepted ones. On the contrary, composition of cheeses with L. paracasei I90 was similar to that of the controls, but these samples were less accepted than cheeses without lactobacilli. The oral administration of cheese containing L. plantarum I91 or L. paracasei I90 proved to be safe and able to enhance the number of IgA + cells in the small intestine lamina propria of mice. The use of selected strains of NSLAB exerted a technological and probiotic role: it contributed to the standardization of cheese quality and induced benefic health effects at the gut mucosa in vivo.

The probiotic Lactobacillus paracasei A13, treated at 50 MPa sub-lethal high pressure homogenisation (HPH), was used as adjunct for producing Caciotta cheese. The cell HPH treatment was used because it has been proven to increase the in vitro strain functionality. The starters and Lb. paracasei A13 viability, the cheese hydrolytic patterns and organoleptic profiles were monitored. After cheesemaking and during ripening, the Lb. paracasei A13 gastric acid resistance in cheese and the ability of the cheese, containing HPH-treated or untreated cells, to modulate the gut mucosal immune system in mice were evaluated. Traditional Caciotta was used as controls. The HPH-treated probiotic strain maintained high viability for 14 days whilst the physico-chemical analyses on Caciotta cheese containing HPH-treated cells showed a faster ripening, compared to other cheeses. For functional properties, the 50 MPa treatment increased the Lb. paracasei gastric resistance in Caciotta, maintaining high strain viability, but IL-10 producing capacity was lost by HPH-treatment whilst IgA production was not modified.

The growth of six probiotic commercial strains of lactobacilli was assessed in reconstituted dried whey and buttermilk supplemented with yeast extract, meat peptone, soy peptone, tryptone or casein acid hydrolysate at 0.3%, 0.6% or 1%. The addition of 1% glucose was also tested. Growth and acidification kinetics were determined at 37°C using MRS broth and a commercial culture medium as references. The suitability of whey and buttermilk as cryoprotectants at –20°C and –70°C was also assessed. Whey and buttermilk with 0.3% yeast extract were chosen for the growth of probiotic lactobacilli, since no satisfactory growth was observed without an external nitrogen source, whereas glucose did not improve the growth of any of the strains assayed. In general, buttermilk performed as satisfactorily as the reference media. The effectiveness of these media as cryoprotectants was strain dependent: skimmed milk and whey were the most suitable ones, especially for long‐term storage at –20°C. However, at –70°C, no significant differences were observed between the culture media assessed. The use of whey or buttermilk as culture media for the production of probiotic lactic acid bacteria and for their cryopreservation implies a novel use of these low‐cost products, offering an alternative way of utilizing the by‐products of the dairy industry, helping to minimize their negative impact on the environment.

High pressure homogenization (HPH) is one of the most promising alternatives to traditional thermal treatment for food preservation and diversification. In order to evaluate its potential for the production of fermented milks carrying probiotic bacteria, four types of fermented milks were manufactured from HPH treated and heat treated (HT) milk with and without added probiotics. Microbiological, physicochemical and organoleptic analyses were carried out during the refrigerated period (35 d at 4°C). HPH application to milk did not modify the viability of the probiotic cultures but did increase the cell loads of the starter cultures (ca. 1 log order) compared with traditional products. The coagula from HPH-milk was significantly more compacted ( P <0·05) (higher firmness) than that obtained with HT-milk, and it had the highest values of consistency, cohesiveness and viscosity indexes compared with fermented milks produced without HPH treatment. All the samples received high sensory analysis scores for each descriptor considered. HPH treatment of milk can potentially diversify the market for probiotic fermented milks, especially in terms of texture parameters.

Lactobacillus fermentum Lf2 is a strain which is able to produce high levels (approximately 1 g/l) of crude exopolysaccharide (EPS) when it is grown in optimised conditions. The aim of this work was to characterize the functional aspects of this EPS extract, focusing on its application as a dairy food additive. Our findings are consistent with an EPS extract that acts as moderate immunomodulator, modifying s-IgA and IL-6 levels in the small intestine when added to yogurt and milk, respectively. Furthermore, this EPS extract, in a dose feasible to use as a food additive, provides protection against Salmonella infection in a murine model, thus representing a mode of action to elicit positive health benefits. Besides, it contributes to the rheological characteristics of yogurt, and could function as a food additive with both technological and functional roles, making possible the production of a new functional yogurt with improved texture.

Gut microbiota dysbiosis plays a central role in the development and perpetuation of chronic inflammation in inflammatory bowel disease (IBD) and therefore is key target for interventions with high quality and functional probiotics. The local production of stable probiotic formulations at limited cost is considered an advantage as it reduces transportation cost and time, thereby increasing the effective period at the consumer side. In the present study, we compared the anti-inflammatory capacities of the Bifidobacterium animalis subsp. lactis (B. lactis) INL1, a probiotic strain isolated in Argentina from human breast milk, with the commercial strain B. animalis subsp. lactis BB12. The impact of spray-drying, a low-cost alternative of bacterial dehydration, on the functionality of both bifidobacteria was also investigated. We showed for both bacteria that the spray-drying process did not impact on bacterial survival nor on their protective capacities against acute and chronic colitis in mice, opening future perspectives for the use of strain INL1 in populations with IBD.

The aim of this study was to adjust technological parameters: acidification of the curd (pH 5.25) and time (2, 5, 10 and 20 min) and stretching temperature (58, 62.5 and 68 °C) in order to make a pasta filata cheese carrying a probiotic bacterium at levels higher than 107 CFU/g. A control and a probiotic cheese were produced. Lactobacillus rhamnosus GG was used and its resistance to simulated gastrointestinal digestion (SGD) was evaluated. Gross composition and pH, microbiological analysis, proteolysis, physicochemical and sensory characteristics, volatile compounds, organic acids and sugar profiles were also determined. The probiotic remained above 3 × 107 CFU/g during its shelf life and exhibited high resistance to SGD (matrix protection of about 60%). The addition of the probiotic increased secondary proteolysis (about 30% for SN fraction in trichloracetic and phosphotungstic acids) and the production of diacetyl, acetoin, lactic and acetic acids. Sensory characteristics (smell, astringency, acid taste and residual flavor) were also modified. The development of a probiotic Fior di Latte cheese that might contribute to disease prevention and generate improvements in sensory characteristics compared to traditional products would allow expanding the market of functional foods.

Low levels of High Pressure of Homogenization (HPH) can be applied directly to lactic acid bacteria cells in order to enhance some functional properties. In a previous work we observed that a 50 MPa HPH treatment increased Lactobacillus paracasei A13 hydrophobicity and resistance to simulated gastric digestion. The aim of this work was to assess the in vivo effects of HPH treatment applied to probiotic lactobacilli on their interaction capacity with the gut and on their ability to induce IgA cell proliferation in mice intestine. BALB/c mice received FITC-labelled cultures of strains, previously treated or not (control) at 50 MPa. Fluorescently labelled cells were studied in the intestine of animals sacrificed 10 and 30 min after intragastric intubation. HPH-treated and control cultures of each strain were orally administered to mice for 2, 5 or 7 consecutive days. The number of IgA-producing cells in the gut was studied by immunohistochemistry. HPH treated probiotic lactobacilli modified their interaction with the small intestine. HPH-treated cells induced a higher IgA response compared to untreated ones, in a strain- and feeding period-dependent way. HPH treatment could increase some in vivo functional characteristics of probiotic strains, highlighting the potential of this technique for the development of probiotic cultures.

This study aimed to evaluate the resistance, viability, and functionality of two strains of probiotics immobilized in solid lipid microparticles covered by a complex of gelatin and gum Arabic. Microcapsules were evaluated regarding their size, morphology, and resistance under stress conditions. Encapsulated microorganisms were evaluated concerning during storage for 120 days. Additionally, the effect of encapsulation on the functionality of the microorganisms was investigated using an in vivo assay. Microcapsules had sizes around 80 μm and extreme pH and temperature of 50 °C destabilized them. Encapsulation improved the stability of these microorganisms in the presence of salt and in gastrointestinal conditions. Encapsulated microorganisms maintained their viability during storage and the dosage of S-IgA and cytokines (IL-2, IL-6, IL-10, and TNF-α) in mice indicated that encapsulated microorganisms maintained their functionality. Therefore, the microencapsulation technique may be promising for the improvement of the viability of probiotics under adverse conditions without compromising their immunomodulating capacity.

The aim of this study was to isolate, identify and characterize lactic acid bacteria (LAB) from spontaneously fermented maize silage, and evaluate their performance as spray-dried (SD) cultures to enhance the fermentation and the aerobic stability of maize micro-silos. Eleven strains of LAB were characterized for growth kinetics, the capability to grow in vegetable-based medium (VBM), production of organic acids and the ability to tolerate heat–stress. Three strains (Lactobacillus plantarum Ls71, Pediococcus acidilactici Ls72 and Lactobacillus buchneri Ls141) were selected and further characterized for the ability to grow as single strain or in co-culture in MRS and VMB medium, to survive at freeze and spray-drying process, for their performance as SD bacteria in micro-silos and for the aerobic stability in bucket silos. L. buchneri Ls141 showed the highest growth capability in VBM and produced the highest amount of acetic acid, while L. plantarum Ls71 produced the highest amounts of lactic acid. P. acidilactici Ls72 was the most heat-resistant strain, with a reduction of 0.2 log10 CFU/mL (15 min at 55 °C). The three strains satisfactorily tolerated both spray and freeze-drying. After 4 days of fermentation, all the samples reached a pH value of about 3.7-3.8. A significantly lower cell load of filamentous fungi and yeasts ( 8.7 log10 CFU/g) was observed after 30 d of fermentation. A greater amount of acetic acid, crude protein, ash and ammonia nitrogen/total nitrogen was detected in inoculated silages. A significant reduction of filamentous fungi and yeasts was also observed in inoculated bucket silos after 50 d of fermentation. The aerobic stability was significantly improved in inoculated silage since the temperature remained stable after 16 days (384 h). On the contrary, an increase of 5 °C was observed in control samples after 1 day. The selected strains have the potential to be produced as SD silage inoculant as they were able to accelerate the fermentation process, to control filamentous fungi and yeasts, to improve some nutritional and chemical parameters of silage and to improve aerobic stability.

The roles of an exopolysaccharide (EPS) extract from Lactobacillus fermentum Lf2 were studied individually or combined with a probiotic strain, Bifidobacterium animalis subsp. lactis INL1. EPS in its purified form caused an increase in the levels of cytokine TNF-α; both purified and crude EPS produced an increase in the regulatory cytokine IL-10. BALB/c mice received yoghurt with no additives (Y), with EPS (YE), with bifidobacteria (YB), or both (YEB) for 25 days. Only the YE group presented significantly increased concentrations of total short chain fatty acids (p < 0.05) including acetic and butyric acids; the levels of the Clostridium coccoides cluster also rose over time (p < 0.05) for this group. A possible bifidogenic role was observed for the YEB group, reflected in the increasing levels of the genus Bifidobacterium along time (p < 0.05); this was not observed when the probiotic was administered solely (YB group).

Nowadays, there is a great interest of food industries on the development of innovative dairy products containing probiotic bacteria. Beyond yoghurt, cheeses could be a suitable matrix for the incorporation of probiotics. This work aimed to study the impact of the addition of two probiotic lactobacilli (L. rhamnosus GG and L. acidophilus LA5), either individually or combined, on the physicochemical and sensory characteristics of pasta filata soft cheeses and to evaluate the immunomodulating capacity of the product using an in vivo model. Four cheese types were produced: CC (control cheese); L-C (added with L. acidophilus LA5); GG-C (added with L. rhamnosus GG) and GGL-C (added with both probiotics). No differences in gross composition were found. An increase in secondary proteolysis was observed in GGL-C cheeses which correlated with a greater bitter and aftertaste. On the contrary, the L-C cheese received the highest overall quality score. The translocation assay was negative. Moreover, the three experimental cheeses were able to modulate the immune system of mice by reducing the secretion of pro-inflammatory cytokines in the intestine, downregulating the levels of interleukin (IL)-10 and increasing the secretion of Secretory Immunoglobulin A (S-IgA). No synergistic effect was detected when both probiotics were added.

Our aim was to obtain derivatives of non-intestinal lactobacilli—commonly used in the dairy industry—able to grow under physiological concentrations of bile salts. Six resistant derivatives (five from Lactobacillus delbrueckii subsp. lactis and one from L. acidophilus) growing in the presence of 0.5% bile salts were obtained from 24 strains of L. delbrueckii subsp. bulgaricus, L. delbrueckii subsp. lactis and L. helveticus, and also one strain of L. acidophilus with low resistance to bile salts. Carbohydrate fermentation profiles remained unchanged but the rate of fermentation of several sugars was generally slower in bile-resistant derivatives. No significant differences with respect to the parent strains were observed for the cell-wall hydrophobicity, capacity to grow in milk and survival during frozen storage. Some bile-resistant derivatives were also resistant to low pH, but in other strains this capacity was acquired following progressive adaptation to low pH. Adaptation to bile salts and low pH might be valuable tools for increasing the survival under gastrointestinal conditions of non-intestinal lactobacilli used for the manufacture of dairy products.

In a previous work, bile-salt-resistant derivatives were obtained from non-intestinal lactobacilli. The aim of this work was to investigate the impact of bile adaptation of Lactobacillus delbrueckii subsp. lactis 200 on morphology, surface properties, in vivo interaction capacity with the gut and ability to activate the gut immune response. Electron microscopy studies, growth kinetics in the presence of bovine and porcine bile, the capacity to deconjugate bile acids, hydrophobicity, autoaggregation and co-aggregation capacities were studied for the parental strain and its bile-resistant derivative in vitro. Additionally, survival in intestinal fluid, the interaction with the gut and the immunomodulating capacities were studied in mice. Bile salt adaptation conferred upon the adapted strain a higher capacity to withstand physiological concentrations of bile salts and greater survival capacity in intestinal fluid. However, bile salt exposure reduced cell hydrophobicity, autoaggregation and adhesion capacities, resulting in reduced persistence in the intestinal lumen and delayed capacity to activate the gut immune response. Insight into the effects of bile salts upon the interaction and immunomodulating capacity of lactobacilli with the gut is provided, relating in vitro and in vivo results.

Limosilactobacillus fermentum Lf2 (Lf2) is an autochthonous strain that produces high levels of exopolysaccharides (EPS). The objective of this work was to evaluate the probiotic potential of Lf2 and its relationship with these metabolites in a mouse model of TNBS (trinitrobenzene sulfonic acid)-induced chronic colitis. Mice were treated intrarectally with increasing doses of TNBS resuspended in 50% ethanol for 14 days. In parallel, they received different treatments by gavage (lactose 10% as the matrix): freeze-dried Lf2 (L); purified EPS (E); and lactose 10% (T). A healthy control group (H) was treated with 50% alcohol without TNBS (intrarectally) and 10% lactose (by gavage). In the small intestine, there was a significant increase in IgA levels for the group that received EPS and a decrease in IFN-γ for mice treated with the strain compared to the other groups. In the large intestine, IL-2 and IFN-γ presented the lowest levels in the groups treated with EPS and the strain. The concentrations of acetic and propionic acids in mice that received Lf2 were the highest, while the levels of butyric acid were comparable to the healthy control group. An increase in the abundance of SCFA-producing bacteria was observed for mice treated with EPS and the strain in comparison with the colitis control group. The enzyme activity of catalase was higher in all the treatments compared to the TNBS-induced colitis control mice. To summarize the results obtained, a principal component analysis (PCA) was performed, clearly grouping the treatments in different clusters according to the variables studied. This is one of the first studies to address the role of a potential probiotic strain in a chronic colitis mouse model, trying to elucidate the relationship between its properties and the EPS synthesized.

To ferment buttermilk, a low-cost by-product of the manufacture of butter, with a proteolytic strain of Lactobacillus helveticus, to enhance its value by the production of a functional peptide-enriched powder.Buttermilk was fermented with Lact. helveticus 209, a strain chosen for its high proteolytic activity. To enhance the release of peptidic fractions, during fermentation pH was kept at 6 by using NaOH, Ca(CO)(3) or Ca(OH)(2). Cell-free supernatant was recovered by centrifugation, supplemented or not with maltodextrin and spray-dried. The profile of peptidic fractions released was studied by RP-HPLC. The lactose, Na and Ca content was also determined. The powder obtained was administered to BALB/c mice for 5 or 7 consecutive days, resulting in the proliferation of IgA-producing cells in the small intestine mucosa of the animals.Buttermilk is a suitable substrate for the fermentation with Lact. helveticus 209 and the release of peptide fractions able to be spray-dried and to modulate the gut mucosa in vivo.A powder enriched with peptides released from buttermilk proteins, with potential applications as a functional food additive, was obtained by spray-drying. A novel use of buttermilk as substrate for lactic fermentation is reported.

The capacity of lactic acid bacteria to produce exopolysaccharides (EPS) conferring microorganisms a ropy phenotype could be an interesting feature from a technological point of view. Progressive adaptation to bile salts might render some lactobacilli able to overcome physiological gut barriers but could also modify functional properties of the strain, including the production of EPS. In this work some technological properties and the survival ability in simulated gastrointestinal conditions of Lactobacillus delbrueckii subsp. lactis 193, and Lb. delbrueckii subsp. lactis 193+, a strain with stable bile-resistant phenotype derived thereof, were characterized in milk in order to know whether the acquisition of resistance to bile could modify some characteristics of the microorganism. Both strains were able to grow and acidify milk similarly; however the production of ethanol increased at the expense of the aroma compound acetaldehyde in milk fermented by the strain 193+, with respect to milk fermented by the strain 193. Both microorganisms produced a heteropolysaccharide composed of glucose and galactose, and were able to increase the viscosity of fermented milks. In spite of the higher production yield of EPS by the bile-resistant strain 193+, it displayed a lower ability to increase viscosity than Lb. delbrueckii subsp. lactis 193. Milk increased survival in simulated gastric juice; the presence of bile improved adhesion to the intestinal cell line HT29-MTX in both strains. However, the acquisition of a stable resistance phenotype did not improve survival in simulated gastric and intestinal conditions or the adhesion to the intestinal cell line HT29-MTX. Thus, Lb. delbrueckii subsp. lactis 193 presents suitable technological properties for the manufacture of fermented dairy products; the acquisition of a stable bile-resistant phenotype modified some properties of the microorganism. This suggests that the possible use of bile-resistant derivative strains should be carefully evaluated in each specific application considering the influence that the acquisition of a stable bile-resistant phenotype could have in survival ability in gastric and intestinal conditions and in technological properties.

The aim of this study was to evaluate the influence of microbiological and technological factors on the viability and functionality of probiotic Lactobacillus strains. In particular, the influence of harvesting time, food matrix, refrigerated storage, time of inoculation and refrigerated storage in fermented milk on the resistance to simulated gastric digestion ( RSGD ) was evaluated. Moreover, strain resistance to simulated gastric digestion was compared to human gastric fluid. Results showed that the variables studied affected, in a different way and in a strain‐dependent manner, the RSGD . No direct relation was observed between cell viability and RSGD .

Probiotic microorganisms are mainly bacteria from the genera Lactobacillus and Bifidobacterium that exert a health benefit on the host when consumed on a regular basis as dietary supplements or in foods. The fermented dairy industry was the first in successfully marketing specific strains of probiotic bacteria in products such as yogurts and fresh cheeses. Factors such as growing vegetarianism including veganism among consumers, lactose intolerance, allergy to milk proteins, cholesterol content of dairy products, and religious or dietary habits have limited the growth of the market of probiotics in dairy products and impaired their consumption by people interested in a healthier diet. Plant material such as cereal, legumes, fruits, and vegetables, and their combinations, offer a plethora of possibilities for the development of probiotic products, fermented or not. Much of the knowledge learned in the last 30 years about probiotics in dairy products is paving the way for the development of new nondairy foods and drinks.

Shiga toxin (Stx)-producing Escherichia coli (STEC) is an emergent pathogen associated with foodborne diseases, especially foodstuffs of animal origin. A total of 250 beef samples (ground beef and hamburgers) obtained from retail outlets in Santa Fe and Santo Tomé cities, and 150 milk samples from bulk tank milk from dairy barns of the region were analyzed by selective enrichment and immunomagnetic separation. Escherichia coli O157:H7 stx2, eae and ehxA positive strains were isolated from three (1.2%) beef samples. The strains could be differentiated by pulsed-field gel electrophoresis, phagetyping and genotyping of stx. The milk samples were negative for STEC O157. These findings confirm the role of food of animal origin in the epidemiology of E. coli O157:H7 - associated diseases.

Caseinomacropeptide (CMP) is the 64 C-terminal amino acid residue of κ-casein, formed by chymosin cleavage during cheese manufacture. This study examined the effects of oral administration of a CMP-enriched extract (CMPEE), obtained from a local dairy plant, on the Ca content of mouse femurs. Animals received low (0.1%, w/v), normal (0.5%, w/v) or high (1.2%, w/v) Ca diet for 3 or 8 weeks and CMPEE diluted (1:10) in their drinking water. No significant differences in Ca content were observed in faeces, kidney, urine or blood serum compared with control animals. The oral administration of CMP to mice significantly enhanced the Ca content in femur under a low-Ca diet model, especially during the period of full body development (3 weeks), in which case a significant 12% Ca increase was observed. These findings pave the way for further studies aimed at supplementing infant food with industrially-obtained CMP-enriched extract for enhanced bone health.

To evaluate the effects of spray drying of Lactobacillus rhamnosus 64 on its capacity to modulate the gut immune response and on the attenuation of TNBS‐induced colitis in mice. Lactobacillus rhamnosus 64 was spray dried in cheese whey‐starch solution and administered to mice for 3, 6 or 10 consecutive days. Peritoneal macrophage phagocytic activity, secretory IgA levels in the small intestinal fluid and TNFα, IFNγ, IL‐10, IL‐6 and IL‐2 levels in homogenates of the small and large intestine were determined. The effects of spray drying were also evaluated in an acute model of Trinitrobenzenesulfonic acid (TNBS)‐induced colitis. A shift in the regulation of immune parameters, particularly the cytokine profile, was observed for mice treated with the spray‐dried culture, compared to the profile observed in animals that received the strain as fresh culture (FC). The spray‐dried culture of L. rhamnosus 64 showed anti‐inflammatory properties in murine model of TNBS‐induced colitis. The spray‐drying process of L. rhamnosus 64 in whey‐starch modified its immunomodulating capacity in healthy animals and conferred enhanced protection in an in vivo model of inflammation. Probiotic capacity can be affected by spray drying in relation to the properties observed for the strain as an overnight FC. This fact should be taken into account when producing the culture for its application in the industry.

To assess the variability of secretory immunoglobulin A (S-IgA) in the lumen and feces of mice along a working day. Mice were maintained under a 12 h light–dark cycle, light period starting at 8 AM. S-IgA was determined in feces and intestinal content (after one or three washes) at three points along the day: at the beginning, in the middle and at the end of the light period (ELP). Significant reduction in the content of S-IgA in the small intestine fluid and in feces was observed at the end of the light cycle, which coincides with the end of a regular working day (8 PM) in any given animal facility. It was also observed that three washes of the small intestine were more effective than one flush to recover a significant higher amount of S-IgA, with the smallest coefficient of variation observed by the ELP. A smaller CV would imply a reduced number of animals needed to achieve the same meaningful results. The results may be useful when designing animal trials for the selection of probiotic candidates based on their capacity of activating S-IgA, since it would imply a more rational use of experimental animals.

Breast milk can be a source of potential probiotic bacteria, but the technological capacity of isolates obtained from this source is not always guaranteed. We aimed at isolating lactobacilli from breast milk samples collected in Argentina, focusing on isolates with functional and technological potential as probiotics. Fourteen Lactobacillus and one Bifidobacterium isolates were obtained from 164 samples donated by 104 mothers. The isolates preliminarily identified by MALDI-TOF, and then the identity was confirmed by partial 16S rRNA gene sequencing. Hydrophobicity was determined (hexadecane and xylene partition). The strains were also co-cultured with murine RAW 264.7 macrophages for screening the capacity to induce the anti-inflammatory cytokine interleukin (IL)-10. Hydrophobicity ranged from 7.4 and 95.9%. The strains Lactobacillus gasseri (70a and 70c) and Lactobacillus plantarum (73a and 73b) were the strains with a higher capacity to induce IL-10 production by macrophages. The technological application was evaluated by freezing dried in 10% lactose or 10% polydextrose. The survival was assessed after accelerated (37 °C, 4 weeks) or long-term (5 and 25 °C, 12 months) storage. Except for Lactobacillus gallinarum 94d, strains lost less than 1 Log10 order cfu/g after long-term (12 months) storage at 5 °C in lactose and polydextrose as protectants. A low correlation between survival to accelerated and long-term storage tests was observed. L. gasseri (70a and 70c) and L. plantarum (73a and 73b) deserve further studies as potential probiotics due to their capacity to induce IL-10 from murine macrophages and their hydrophobicity. In special, L. plantarum 73a was able to confer enhanced protection against Salmonella infection by promoting the immunity of the small intestine.

Four commercial starter cultures containing Streptococcus thermophilus and Lactobacillus delbrueckii subsp. bulgaricus (S1, S2, S3 and S4; S3 also contained Limosilactobacillus fermentum) were compared for fermentation, volatile flavor compounds, physicochemical parameters and microbiology, in yogurt prepared from three milk base formulations with increased protein (B1, B2 and B3).The fermentation patterns differed among starters, with Yoflex Mild 1.0 (S4) and SLB95 (S2) showing the longest fermentation time, depending on the formulation. At 21 days, S. thermophilus counts were similar among starters and higher than 8.52 log CFU mL-1 , for all yogurts. The highest counts (6.86 log CFU mL-1 ) for L. delbrueckii subsp. bulgaricus was found for S2 yogurts made from whey protein hydrolysate (B3). Minor water-holding capacity was detected for YF-L811 (S1) yogurts. Yoflex Harmony 1.0 (S3) starter containing Lim. fermentum produced a distinctive volatile profile characterized by aldehydes with respect to yogurts prepared with S1, S2 and S4, which were characterized by ketones.Results indicate the usefulness of carrying out studies similar to the present one to select the most appropriate process conditions depending on the desired product. © 2022 Society of Chemical Industry.

Fil: Sanchez, Borja. Consejo Superior de Investigaciones Cientificas. Instituto Superior de Productos Lacteos de Asturias; Espana;

Buttermilk is a suitable substrate for fermentation with proteolytic strains of L actobacillus in order to release peptide fractions able to enhance the gut mucosal immune system. We aimed to determine the influence of the degree of proteolysis of buttermilk proteins on their functionality. Animals received for seven consecutive days the cell‐free fraction of 10 or 20% (w/v) buttermilk fermented with L actobacillus delbrueckii subsp. lactis 210 at p H 6. The p H was controlled either with N a OH or C a( OH ) 2 . No significant differences in the number of I g A ‐producing cells in the small intestine of mice were found. The functional capacity of the product under study was not affected by the technological variables considered.

The aim of this study was to manufacture pasta filata cheeses added with two probiotic lactobacilli: Lactobacillus rhamnosus GG and Lactobacillus acidophilus LA5, either individually or combined, and to evaluate the effect of the storage temperature (4 and 12 °C) on their chemical, microbiological, and sensorial characteristics. Three cheese types were made: (i) G: containing L. rhamnosus GG, (ii) L: containing L. acidophilus LA5, and (iii) GL: containing both probiotic strains. Gross composition, pH, microbiological, and sensory characteristics were determined. No differences in gross composition were found among them. pH values remained above 5.2 in cheeses stored at 4 °C. However, a postacidification was observed in cheeses ripened at 12 °C. L. acidophilus LA5 was not able to grow, while L. rhamnosus GG grew 1.5 log 10 CFU/g in G and GL cheeses stored at 12 °C, reducing the pH from day 8 onwards. These results emphasize the importance of the storage temperature since the good characteristics of probiotic cheeses are kept if the cold-chain is respected. Thus, the selection of probiotics, together with the food matrix and the starter, should be carefully evaluated.

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Chrysin is a natural flavonoid that despite having numerous biological properties, its therapeutic value is limited due to its very low solubility in aqueous media. In this work, chrysin was conjugated with methoxypolyethylene glycols (mPEGs) of different molecular weights (350, 500, 750, and 2000 g/mol), affording PEGylated chrysins with high yields and excellent purities. In all cases, an increase in the water solubility of the conjugates was observed, which was highest when 500 g/mol of mPEG was used in the PEGylation reaction. Furthermore, in aqueous solution, PEGylated chrysins formed aggregates of ellipsoid shape. Electrochemical studies showed that the redox properties were conserved after PEGylation. While in vitro antibacterial and antifungal studies probed that the intrinsic activity was conserved, in vitro antitumor activities against HepG2 (liver carcinoma cells) and PC3 (prostate cancer cell) showed that PEGylated chrysins retained the cytotoxic activity and the ability of induction of apoptosis for the evaluated human cancer cells.

Using a mouse model of diet-induced obesity in males and females C57BL/6 mice, effects of a yoghurt enriched with proteins on pro-inflammatory cytokine and lipid profiles were assessed. A clear sex-dependent behaviour of the mice was shown; females gained less weight than males independent of the diet. When a high fat diet was implemented with yoghurt, a reduction of cholesterol and triglycerides in liver and blood serum of males, and triglycerides in the liver of females was observed. For male mice fed with yoghurt, a significant reduction of the levels of pro-inflammatory cytokines was detected; in female mice, the anti-inflammatory effect due to yoghurt consumption was observed only to a minor extent. Principal component analysis confirmed that gender or diet was able to group individual animals. A higher variability of data was observed in females compared with males, probably the reason why less significant differences were observed in the former.

Emulsion aggregation toner (E/A Toner) has been developed for digital color copiers &amp; printers.Emulsion aggregation toner has been prepared by using the following process steps: pigment/wax dispersion preparation, emulsion polymerization, homogenization, aggregation, and coalescence. Shape optimized toner particles are obtained by the control of temperature, pH and time.The emulsion aggregation process with stepwise aggregation in particle formation provides a capsule structure and it also exhibits narrow particle size distribution.Narrow size distribution can be obtained by optimization of the latex size and Zeta potential, controlled in the emulsion polymerization step.This study shows the surface controlled toner can provide 2 important advantages for high quality color printing compared to conventional pulverized toner. The first is a method to control the charging of different colors by uniform surface composition. The same level of charge is obtained for each color as well as stable print quality under various conditions. Charging properties can also be controlled by metal ion treatment in the toner-washing step. The second advantage is the prevention of surface wax, which enables high toner flow, even for small particle sizes about (5-micron particle). This process also enables compatibility with an oil-less fusing system and excellent image quality with lower toner mass per unit area even for rough paper surfaces.

What is called “the biotics family” refers to a set of terms that have emerged lately in the fields of health, nutrition, and food technology. The first three members of this family, and for which there are consensus definitions, are probiotic, prebiotic, and synbiotic. In the last decade, this family has grown significantly with the incorporation of many related words containing the “biotic” suffix such as psychobiotic, immunobiotic, oncobiotic, enterobiotic, oralbiotic, postbiotic, paraprobiotic, and proteobiotic. The main problem of the uncontrolled expansion of this family is the proliferation of overlapping, misleading, or poorly defined terms, often lacking consensus on how to define them. This chapter proposes to take a look at these terms and provides a discussion to avoid the misuse of them.

Universidad Nacional del Litoral (UNL), Consejo Nacional de Investigaciones Cientificas y Tecnicas (CONICET)

Ponencia presentada en el XII Congreso Argentino de Ciencia y Tecnologia de Alimentos (CYTAL), celebrado del 7 al 9 de otubre de 2009 en Concordia, Entre Rios (Argentina).

To characterize four novel autochthonous bifidobacteria isolated from monkey faeces and a Bifidobacterium lactis strain isolated from chicken faeces by evaluating their technological and biological/functional potential to be used as probiotics. Different stressors, including food process parameters and storage, can affect their viability and functionality.The resistance to frozen storage, tolerance to lyophilization and viability during storage, thermal, acidic and simulated gastric resistance, surface hydrophobicity and antimicrobial activity against pathogens were studied. Bifidobacterium lactis Bb12 and INL1 were used as reference strains. The results obtained demonstrated that the new isolates presented strain-dependent behaviour. Good results were obtained for thermal resistance, frozen storage at -80°C and lyophilized powders maintained at 5°C. Cell viability during refrigerated storage was higher when the strains were resuspended in milk at pH 5·0 than at 4·5. The surface hydrophobicity ranged between 7 and 98% depending on the strain. The simulated gastric resistance was improved for the strains incorporated in cheese. Regarding antimicrobial activity, bifidobacteria isolated from monkey presented higher inhibitory capacity than the reference strains.This research provides a deeper insight into new strains of bifidobacteria isolated from primates and chicken that have not been previously characterized for their potential use in dairy products and confirm the most robust stress tolerance of B. lactis.The possibility of expanding the available bifidobacteria with the potential to be added to a probiotic food necessarily implies characterizing them from different points of view, especially when considering unknown species. For monkey isolates (which showed higher antimicrobial activity against pathogens), more in-depth knowledge is needed before applying strategies to improve their performance. On the contrary, the chicken isolate B. lactis P32/1 showed similar behaviour to the references B. lactis strains; therefore, it could be considered as a potential probiotic candidate.

Fil: Binetti, Ana Griselda. Consejo Nacional de Investigaciones Cientificas y Tecnicas. Centro Cientifico Tecnologico Conicet - Santa Fe. Instituto de Lactologia Industrial. Universidad Nacional del Litoral. Facultad de Ingenieria Quimica. Instituto de Lactologia Industrial; Argentina

Fil: Burns, Patricia Graciela. Consejo Nacional de Investigaciones Cientificas y Tecnicas. Centro Cientifico Tecnologico Conicet - Santa Fe. Instituto de Lactologia Industrial. Universidad Nacional del Litoral. Facultad de Ingenieria Quimica. Instituto de Lactologia Industrial; Argentina

Fil: Cuffia, Facundo. Consejo Nacional de Investigaciones Cientificas y Tecnicas. Centro Cientifico Tecnologico Conicet - Santa Fe. Instituto de Lactologia Industrial. Universidad Nacional del Litoral. Facultad de Ingenieria Quimica. Instituto de Lactologia Industrial; Argentina

Fil: Guglielmotti, Daniela Marta. Consejo Nacional de Investigaciones Cientificas y Tecnicas. Centro Cientifico Tecnologico Conicet - Santa Fe. Instituto de Lactologia Industrial. Universidad Nacional del Litoral. Facultad de Ingenieria Quimica. Instituto de Lactologia Industrial; Argentina