Presence Of Bile Salts Research Articles

Albendazole nanosuspension coated granules for the rapid localized release and treatment of colorectal cancer

Albendazole (ABZ), an anthelmintic drug, has been repurposed to treat various types of cancers. However, poor solubility of ABZ, resulting in low bioavailability, limits its application. Nanosuspension is a versatile method for enhancing the dissolution of hydrophobic molecules, but a successful drying has been the biggest challenge in the field. The objective of this research is to formulate and optimize ABZ nanosuspension (NS) coated granules for rapid delivery of ABZ for the treatment of colorectal cancer. ABZ NS was prepared by dual centrifugation method using Kollidon® VA64 and sodium lauryl sulphate (SLS) as stabilizers. The processing method was optimized to obtain a stable nanosuspension with particle size < 300 nm. The optimized ABZ NS was coated on microcrystalline cellulose (MCC) to form the nano-coated granules (NCG) and filled in EUDRACAP® for colon targeted delivery. The ABZ NS and NCG achieved ∼ 60 % and ∼55 % drug release, respectively, in presence of bile salt at colonic pH. Half-maximal inhibitory concentration (IC50) of ABZ NS was found to be 1.18 ± 0.081 µM and 3.59 ± 0.080 µM in two colorectal cancer cell lines: HCT 116 and HT-29, respectively. In addition, In vitro 3D tumor assay revealed that ABZ NS has superior tumor growth inhibition activity compared to the control and pure ABZ. The preparation of ABZ NCG in EUDRACAP® could be a promising approach to achieve colon targeted delivery and to repurpose ABZ for the treatment of colorectal cancer.

As next-generation probiotics: acetic acid bacteria isolated from Kombucha beverages produced with Anatolian hawthorn leaves.

This research examined acetic acid bacteria (AAB) isolated from Kombucha beverages produced with Anatolian hawthorn (Crataegus orientalis) as next-generation probiotics. Eighty-six AAB were isolated from the samples and investigated in terms of biosafety, viability in vitro gastrointestinal conditions, technological and bioactive properties, and also in vitro adhesion abilities. Seventy-six isolates demonstrating γ-hemolysis exhibited resistance to erythromycin and ampicillin. Besides, these isolates survived at low pH and in the presence of bile salts. However, the majority of AAB isolates showed tolerance to phenol, pepsin, and pancreatin. Also, twenty-one isolates showed protease enzyme activity, while eight isolates had amylase enzyme activity. Despite most of the isolates showed viability at 1.5% salt, only 19 isolates survived at 10% salt. Most AAB isolates exhibited inhibition zones ranging from 8 to 26mm against test bacteria, their antioxidant activities were above 80%. Additionally, some isolates exhibited auto-aggregation ability ranging from 0.66 to 23.62% and co-aggregation ability ranging from 1.18 to 71.32%, while hydrophobicity ranged from 1.32 to 69.87% toward xylene. Finally, the indigenous 76 AAB isolates that had remarkable probiotic properties were characterized based on 16S rRNA gene sequencing, and the isolates belonged to Komagateibacter sp. (64.47%), Komagateibacter saccharivorans (15.79%), K. rhaeticus (11.84%), and Gluconobacter sp. (7.90%). As a result, the isolates identified as Gluconobacter sp. A21, Komagataeibacter sp. A139, Gluconobacter sp. A141, and Komagataeibacter sp. A146, which showed high viability under gastrointestinal conditions, safe and acceptable in terms of technological, bioactive, and adhesion properties and could be evaluated as next-generation probiotics.

Efficiency of Encapsulation of Thioflavin T (ThT) into Polar and Nonpolar Environments of Different Bile Salt Aggregates: A Femtosecond Fluorescence Study.

The binding of Thioflavin T (ThT) with various bile salts, a potential host molecule, has been analyzed by steady-state and time-resolved fluorescence spectroscopy. A comparative study has been executed to investigate the influence of confinement of different bile salts, namely, sodium cholate (NaCh), sodium taurocholate (NaTC), and sodium deoxycholate (NaDC) on binding and excited state torsional motion of ThT molecules. The changes in absorption and emission properties of probe molecules were found to be sensitive to increasing bile salt concentration in aqueous 0.2 (M) NaCl solutions. The photophysics of ThT mainly depends on hydrophobicity, morphology, and size of bile salt aggregates in solution. In the presence of bile salts, the emission intensity and emission lifetime of ThT increase significantly, indicating encapsulation of dye. Moreover, we have also investigated the effect of the ionic strength of the medium by sodium chloride (NaCl) on the spectroscopic properties of ThT in the restricted surroundings of aqueous bile salts. It is observed that the fluorescence lifetime of ThT in bile salts increases significantly in the presence of NaCl. The encapsulation efficiency of ThT in bile salt aggregates has been assessed by iodide (I-) as an external ionic quencher. We found that NaDC aggregates are more efficient in the modulation of photophysical properties of ThT and also provide better protection efficiency to decrease the nonradiative deactivation processes.

Probiotic potential of acetic acid bacteria isolated from kombucha in New Zealand in vitro

Kombucha is a fermented tea beverage containing live microorganisms, mostly beneficial strains of yeast and acetic acid bacteria (AAB), but empirical evidence is limited supporting the probiotic potential of kombucha. This study reports the in vitro probiotic potential of 36 AAB strains isolated from three Kombucha samples commercially available in New Zealand. Nine representative AAB strains, belonging to three species (Komagataeibacter rhaeticus, Acetobacter musti and Gluconobacter potus), were examined for their primary probiotic characteristics such as tolerance to bile salts, NaCl, and low pH, and temperature. Three non-cellulose forming strains (A. musti LOAAB1, G. potus LOAAB2 and G. potus GBAAB3) were assayed for their cell surface characteristics such as auto-aggregation, co-aggregation with pathogenic bacteria, and hydrophobicity. Antimicrobial, antioxidant activities and enzymatic activities were also investigated for the strains of interest. Results indicated that nine strains were able to grow under low pH in the presence of bile salts, suggesting their potential to survive in the human gut. Six K. rhaeticus strains produced cellulosic pellicles, a potential source of prebiotics for beneficial bacteria. The three AAB strains LOAAB1, LOAAB2 and GBAAB3 showed promising cell surface characteristics, such as auto-aggregation rates (>80 %), co-aggregation with four pathogenic bacteria (13.24–43.47 %), hydrophobicity (42.12 % to 50.20 %), and antioxidant activities (>90 %). All nine strains tested negative for enzymatic activities (haemolytic, proteolytic, phospholipase, and gelatinase), suggesting that they are safe to consume. Together, these data indicate the potential for the three AAB strains to be further investigated as probiotic sources with more in vivo tests for applications in the food and beverages industry.

Selective depletion of Campylobacter jejuni via T6SS dependent functionality: an approach for improving chickens gut health

The targeted depletion of potential gut pathogens is often challenging because of their intrinsic ability to thrive in harsh gut environments. Earlier, we showed that Campylobacter jejuni (C. jejuni) exclusively uses the Type-VI Secretion System (T6SS) to target its prey such as Escherichia coli (E. coli), and phenotypic differences between T6SS-negative and T6SS-positive C. jejuni isolates toward bile salt sensitivity. However, it remains unclear how the target-driven T6SS functionality prevails in a polymicrobial gut environment. Here, we investigated the fate of microbial competition in an altered gut environment via bacterial T6SS using a T6SS-negative and -positive C. jejuni or its isogenic mutant of the hemolysin-coregulated protein (hcp). We showed that in the presence of bile salt and prey bacteria (E. coli), T6SS-positive C. jejuni experiences enhanced intracellular stress leading to cell death. Intracellular tracking of fluorophore-conjugated bile salts confirmed that T6SS-mediated bile salt influx into C. jejuni can enhance intracellular oxidative stress, affecting C. jejuni viability. We further investigated whether the T6SS activity in the presence of prey (E. coli) perturbs the in vivo colonization of C. jejuni. Using chickens as primary hosts of C. jejuni and non-pathogenic E. coli as prey, we showed a marked reduction of C. jejuni load in chickens cecum when bile salt solution was administered orally. Analysis of local antibody responses and pro-inflammatory gene expression showed a reduced risk of tissue damage, indicating that T6SS activity in the complex gut environment can be exploited as a possible measure to clear the persistent colonization of C. jejuni in chickens.

Fit-for-purpose heterodivalent single-domain antibody for gastrointestinal targeting of toxin B from Clostridium difficile.

Single-domain antibodies (sdAbs), such as VHHs, are increasingly being developed for gastrointestinal (GI) applications against pathogens to strengthen gut health. However, what constitutes a suitable developability profile for applying these proteins in a gastrointestinal setting remains poorly explored. Here, we describe an in vitro methodology for the identification of sdAb derivatives, more specifically divalent VHH constructs, that displayextraordinary developability properties for oral delivery and functionality in the GI environment. We showcase this by developing a heterodivalent VHH construct that cross-inhibits the toxic activity of the glycosyltransferase domains (GTDs) from three different toxinotypes of cytotoxin B (TcdB) from lineages of Clostridium difficile. We show that the VHH construct possesseshigh stability and binding activity under gastric conditions, in the presence of bile salts, and at high temperatures. We suggest that the incorporation of early developability assessment could significantly aid in the efficient discovery of VHHs and related constructs fit for oral delivery and GI applications.

Probiotic Abilities of Lactobacillus spp. Isolated from Algerian Fermented Wheat ‘Hamoum’ intended to Use in Broiler Production

Background: The use of probiotics has become popular in the broiler poultry industry due to its positive results on the health of animals, as well as the safety of consumers. In general, probiotics are isolated and selected In vitro from several sources. Despite this, only a few studies focused on isolating probiotics from cereals, which are the main natural food source for poultry. This study aimed to evaluate and identify an autochthonous Lactobacillus spp. isolated from Algerian fermented wheat “Hamoum” as potential probiotics for use in broiler feed. Methods: This research was conducted over the period between 2019-2022. Five bacterial isolates of the genus Lactobacillus were tested for their hemolysis activity, exopolysaccharides production, the ability to withstand the simulated hostile conditions of the chicken’s gut and their capacity to acidify the concentrate feed beverage. Finally, molecular identification was performed using the REP-PCR technique. Result: These isolates were found to produce exopolysaccharides and have no hemolytic activity. Also, could tolerate simulated gastric acidity. However, only three isolates were able to survive in the presence of bile salts and were found to have a high capacity for acidifying medium in a relatively short time. These isolates were identified as Lactobacillus acidophilus and Lactobacillus plantarum subsp plantarum. Those strains can be considered safe candidate probiotics for use in broiler feeds feed. Further studies are needed to investigate and evaluate the effectiveness of these isolates through an in vivo experiment on broiler chickens.

Comparative Genome Analysis and Characterization of the Probiotic Properties of Lactic Acid Bacteria Isolated from the Gastrointestinal Tract of Wild Boars in the Czech Republic.

Probiotics are crucial components for maintaining a healthy gut microbiota in pigs, especially during the weaning period. Lactic acid bacteria (LAB) derived from the gastrointestinal tract of wild boars can serve as an abundant source of beneficial probiotic strains with suitable properties for use in pig husbandry. In this study, we analyzed and characterized 15strains of Limosilactobacillusmucosae obtained from the gut contents of wild boars to assess their safety and suitability as probiotic candidates. The strains were compared using pan-genomic analysis with 49 L.mucosae strains obtained from the NCBI database. All isolated strains demonstrated their safety by showing an absence of transferrable antimicrobial resistance genes and hemolysin activity. Based on the presence of beneficial genes, five candidates with probiotic properties were selected and subjected to phenotypic profiling. These five selected isolates exhibited the ability to survive conditions mimicking passage through the host's digestive tract, such as low pH and the presence of bile salts. Furthermore, five selected strains demonstrated the presence of corresponding carbohydrate-active enzymes and the ability to utilize various carbohydrate substrates. These strains can enhance the digestibility of oligosaccharide or polysaccharide substrates found in food or feed, specifically resistant starch, α-galactosides, cellobiose, gentiobiose, and arabinoxylans. Based on the results obtained, the L.mucosae isolates tested in this study appear to be promising candidates for use as probiotics in pigs.

Biofilm-producing Escherichia coli O104:H4 overcomes bile salts toxicity by expressing virulence and resistance proteins.

We investigated bile salts' ability to induce phenotypic changes in biofilm production and protein expression of pathogenic Escherichia coli strains. For this purpose, 82 pathogenic E. coli strains isolated from humans (n=70), and animals (n=12), were examined for their ability to form biofilms in the presence or absence of bile salts. We also identified bacterial proteins expressed in response to bile salts using sodium dodecyl-sulfate polyacrylamide gel electrophoresis (SDS-electrophoresis) and liquid chromatography-mass spectrometry (LC-MS/MS). Lastly, we evaluated the ability of these strains to adhere to Caco-2 epithelial cells in the presence of bile salts. Regarding biofilm formation, two strains isolated from an outbreak in Republic of Georgia in 2009 were the only ones that showed a high and moderate capacity to form biofilm in the presence of bile salts. Further, we observed that those isolates, when in the presence of bile salts, expressed different proteins identified as outer membrane proteins (i.e. OmpC), and resistance to adverse growth conditions (i.e. F0F1, HN-S, and L7/L12). We also found that these isolates exhibited high adhesion to epithelial cells in the presence of bile salts. Together, these results contribute to the phenotypic characterization of E. coli O104: H4 strains.

Agro-ecosystem of honeybees as source for native probiotic yeasts

Probiotic microorganisms are used to improve the health and wellness of people and the research on this topic is of current relevance and interest. Fifty-five yeasts, coming from honeybee’s ecosystem and belonging to Candida, Debaryomyces, Hanseniaspora, Lachancea, Metschnikowia, Meyerozyma, Starmerella and Zygosacchromyces genera and related different species, were evaluated for the probiotic traits. The resistance to gastrointestinal conditions, auto-aggregation, cell surface hydrophobicity or biofilm formation abilities as well as antimicrobial activity against common human pathogenic bacteria were evaluated. The safety analysis of strains was also carried out to exclude any possible negative effect on the consumer’s health. The influence of proteinase treatment of living yeasts and their adhesion to Caco-2 cells were also evaluated. The greatest selection occurred in the first step of survival at the acidic pH and in the presence of bile salts, where more than 50% of the strains were unable to survive. Equally discriminating was the protease test which allowed the survival of only 27 strains belonging to the species Hanseniaspora guilliermondii, Hanseniaspora uvarum, Metschnikowia pulcherrima, Metschnikowia ziziphicola, Meyerozyma caribbica, Meyerozyma guilliermondii, Pichia kluyveri, Pichia kudriavzevii and Pichia terricola. An integrated analysis of the results obtained allowed the detection of seven yeast strains with probiotic aptitudes, all belonging to the Meyerozyma genus, of which three belonging to M. guillermondii and four belonging to M. caribbica species.

Genetic and Phenotypic Characterization of Bacillus velezensis Strain BV379 for Human Probiotic Applications.

Bacterial spore-forming Bacillaceae species, including Bacillus subtilis and Heyndrickxia coagulans, are increasingly utilized for probiotic dietary supplementation. Bacillus velezensis is a Bacillus species that is frequently used as a direct-fed microbial in animal feed but less so as a probiotic for humans. The objective of this study was to characterize the suitability of the Bacillus velezensis strain BV379 for probiotic applications by (1) in silico screening for both adverse genetic elements and putatively beneficial traits, (2) in vitro evaluation of interactions with human intestinal epithelial cells, and (3) in vitro characterization of BV379 spore viability at various temperatures, pH, and in the presence of bile salt. In silico screening of the BV379 genome revealed few genes encoding Bacillaceae-associated toxins, virulence factors, and enzymes involved in the production of toxins. While BV379 encodes five antimicrobial resistance genes, minimum inhibitory concentration assays determined that BV379 is susceptible to all eight clinically relevant antibiotics tested. Preliminary cell culture experiments showed that BV379 lysates did not adversely impact human intestinal epithelial cell viability and monolayer permeability. It was also determined that BV379 spores can easily tolerate the harsh pH, bile salt, and microaerobic conditions typical of the GI tract. Altogether, the results presented herein support the safety and potential of Bacillus velezensis strain BV379 for use as an oral probiotic.

Saccharomyces cerevisiae SC-2201 Attenuates AOM/DSS-Induced Colorectal Cancer by Modulating the Gut Microbiome and Blocking Proinflammatory Mediators.

Colorectal cancer is the third most common cancer in the world today, and studies have shown that the ratio of Candida to Saccharomyces cerevisiae increased, and the abundance of S. cerevisiae in the intestines of patients with colorectal cancer decreased, which suggests that there is an imbalance in the proportion of fungi in the intestines of patients with colorectal cancer. The objective of this study was to screen S. cerevisiae isolate from traditional Chinese fermentation starters and assess its ability to ameliorate dysbiosis and to alleviate the carcinogenic process of azoxymethane/dextran sodium sulfate-induced colorectal cancer in mice model. S. cerevisiae strain SC-2201 was isolated and exhibited probiotic properties, including the ability to survive in an acidic pH environment and in the presence of bile salts in the gastrointestinal tract, as well as antioxidant activities. Oral administration of S. cerevisiae SC-2201 not only alleviated weight loss but also reduced colonic shortening and histological damage in azoxymethane/dextran sodium sulfate-induced colorectal cancer in mice. Furthermore, the administration of S. cerevisiae SC-2201 suppressed the expression of proinflammatory mediators, such as interleukin-1β, interleukin-6, cyclooxygenase-2, vascular endothelial growth factor, nucleotide-binding domain, leucine-rich repeat, and pyrin domain-containing protein 3. Specifically, the analysis of gut bacteriome showed a significant decrease in Bacteroidota and Campylobacterota levels, as well as an increase in Proteobacteria level in the colorectal cancer group, which was alleviated by supplementation with S. cerevisiae SC-2201. The analysis of the mycobiome revealed a significant increase in the levels of Basidiomycota, Apiosordaria, Naganishia, and Taphrina genera in the colorectal cancer group, which were alleviated after supplementation with S. cerevisiae SC-2201. However, the levels of Xenoramularia, Entoloma, and Keissleriella were significantly increased after administration with S. cerevisiae SC-2201. Overall, the findings of this study demonstrate that S. cerevisiae SC-2201 possesses potential probiotic properties and can effectively attenuate the development of colorectal cancer, highlighting its cancer-preventive potential. This is the first report of a S. cerevisiae strain isolated from traditional Chinese fermentation starters which showed good probiotic properties, and mitigated azoxymethane/dextran sodium sulfate-induced colorectal cancer by modulating the gut microbiome and blocking proinflammatory mediators in mice.

The reversible activation of norovirus by metal ions.

Murine norovirus (MNV) undergoes extremely large conformational changes in response to the environment. The T = 3 icosahedral capsid is composed of 180 copies of ~58-kDa VP1 comprised of N-terminus (N), shell (S), and C-terminal protruding (P) domains. At neutral pH, the P domains are loosely tethered to the shell and float ~15 Å above the surface. At low pH or in the presence of bile salts, the P domain drops onto the shell and this movement is accompanied by conformational changes within the P domain that enhance receptor interactions while blocking antibody binding. While previous crystallographic studies identified metal binding sites in the isolated P domain, the ~2.7-Å cryo-electron microscopy structures of MNV in the presence of Mg2+ or Ca2+ presented here show that metal ions can recapitulate the contraction observed at low pH or in the presence of bile. Further, we show that these conformational changes are reversed by dialysis against EDTA. As observed in the P domain crystal structures, metal ions bind to and contract the G'H' loop. This movement is correlated with the lifting of the C'D' loop and rotation of the P domain dimers about each other, exposing the bile salt binding pocket. Isothermal titration calorimetry experiments presented here demonstrate that the activation signals (bile salts, low pH, and metal ions) act in a synergistic manner that, individually, all result in the same activated structure. We present a model whereby these reversible conformational changes represent a uniquely dynamic and tissue-specific structural adaptation to the in vivo environment.IMPORTANCEThe highly mobile protruding domains on the calicivirus capsids are recognized by cell receptor(s) and antibodies. At neutral pH, they float ~15 Å above the shell but at low pH or in the presence of bile salts, they contract onto the surface. Concomitantly, changes within the P domain block antibody binding while enhancing receptor binding. While we previously demonstrated that metals also block antibody binding, it was unknown whether they might also cause similar conformational changes in the virion. Here, we present the near atomic cryo-electron microscopy structures of infectious murine norovirus (MNV) in the presence of calcium or magnesium ions. The metal ions reversibly induce the same P domain contraction as low pH and bile salts and act in a synergistic manner with the other stimuli. We propose that, unlike most other viruses, MNV facilely changes conformations as a unique means to escape immune surveillance as it moves through various tissues.

Preparation of probiotic-loaded solid lipid nanoparticles and in vitro survival in gastrointestinal conditions

In the present study, Probiotics (Lactobacillus plantarum) loaded-solid lipid nanoparticles were prepared and characterized. Probiotics have proven to possess significant potential in addressing a range of conditions, encompassing neurodegenerative disorders, cancers, cardiovascular diseases, and inflammatory conditions. Additionally, they show efficacy in sustaining a harmonious gut microbiota ecosystem. SLNs were prepared by using solvent emulsification-diffusion technique. The size and morphology of prepared probiotic-loaded SLNs were examined by transmission electron microscopy. The activity or viability of probiotics might be compromised in the challenging gastric conditions of the stomach or in the presence of bile salts. Furthermore, they may face vulnerability to thermal or oxidative stress during preparation and storage. Hence, there is a need for stable probiotic formulations to surmount diverse physicochemical, biopharmaceutical, and biological barriers, ensuring maximal therapeutic effectiveness and clinical applicability. In vitro experiments were carried out with the aim of examining the viability of bacterial cells under gastrointestinal conditions. Encapsulation of the cells protect the cell numbers (colony forming unit) when compared to free bacteria. Nevertheless, the number of probiotic cells decreased in gastrointestinal acidic condition in contrast to free cells. Overall encapsulation of Lactobacillus plantarum in SLN’s plays an important role in enhancing viability and stability which consequently enhance the survival of bacteria against gastrointestinal environmental conditions.

Evaluation of the probiotic potential of yeast isolated from kombucha in New Zealand

The current study investigated the in vitro probiotic potential of yeast isolated from kombucha, a tea beverage fermented with a symbiotic culture of acetic acid bacteria and yeast. A total of 62 yeast strains were previously isolated from four different commercial kombucha samples sold in New Zealand. Fifteen representative isolates belonging to eight different species were evaluated for their growth under different conditions (temperature, low pH, concentrations of bile salts, and NaCl). Cell surface characteristics, functional and enzymatic activities of the selected strains were also studied in triplicate experiments. Results showed that six strains (Dekkera bruxellensis LBY1, Sachizosaccharomyces pombe LBY5, Hanseniaspora valbyensis DOY1, Brettanomyces anomalus DOY8, Pichia kudraivzevii GBY1, and Saccharomyces cerevisiae GBY2) were able to grow under low-acid conditions (at pH 2 and pH 3) and in the presence of bile salts. This suggests their potential to survive passage through the human gut. All 15 strains exhibited negative enzymatic activity reactions (haemolytic, gelatinase, phospholipase, and protease activities), and thus, they can be considered safe to consume. Notably, two of the fifteen strains (Pichia kudraivzevii GBY1 and Saccharomyces cerevisiae GBY2) exhibited desirable cell surface hydrophobicity (64.60–83.87%), auto-aggregation (>98%), co-aggregation, resistance to eight tested antibiotics (ampicillin, chloramphenicol, colistin sulphate, kanamycin, nalidixic acid, nitrofurantoin, streptomycin, and tetracycline), and high levels of antioxidant activities (>90%). Together, our data reveal the probiotic activities of two yeast strains GBY1 and GBY2 and their potential application in functional food production.

In vitro gastrointestinal digestion of marine oil emulsions and liposomal solutions: fate of LC-PUFAs upon lipolysis.

The bioaccessibility and bioavailability of dietary fatty acids depend on the lipid to which they are esterified, the organisation of theses lipids in water and their recognition by lipolytic enzymes. In this work, we studied the release of marine long-chain polyunsaturated fatty acids (LC-PUFA), depending on their presentation either in the form of phospholipids (PL) or triacylglycerol (TAG). Two formulations based on marine PL or TAG extracted from salmon heads (Salmo salar) were prepared. Lipolysis was first tested in vitro by using individual gastrointestinal lipases and phospholipases to identify the enzymes involved in the digestion. Second, the lipolysis of the prepared formulations by a combination of enzymes was tested under in vitro conditions mimicking the physiological conditions found in the GI tract, both in the stomach and in the upper small intestine, in order to evaluate digestibility of TAG and LC-PUFA-containing liposomes. The in vitro results showed that TAG emulsion was hydrolyzed by porcine pancreatic extracts (PPE) and pure pancreatic lipase (PPL) with its cofactor, colipase, and to a lesser extent by pancreatic-lipase-related protein 2 (PLRP2) and a gastric extract (RGE) containing gastric lipase while no hydrolysis was observed with purified pancreatic phospholipase A2 (PLA2) and carboxyl ester hydrolase (CEH). The PL substrate was found to be hydrolysed by PLA2, PPE and PLRP2. Their phospholipase activities on liposomes formulation was dependent on the presence of bile salts. Using a two-step in vitro digestion model, we measured the kinetics of fatty acid release from TAG and PL during the gastric and intestinal phases of digestion. The highest overall lipolysis level was obtained with liposomes (around 75%) during the intestinal phase while they were preserved during the gastric phase. The overall lipolysis level of TAG emulsion was lower (around 33%), while it started already in the gastric phase. In conclusion, liposomes appear as a better delivery system for intestinal absorption of LC-PUFA than TAG. In addition, their resistance to lipolysis under gastric condition can protect LC-PUFA and provide a gastric stable delivery system for other molecules.

Acinetobacter baumannii Survival under Infection-Associated Stresses Depends on the Expression of Resistance-Nodulation-Division and Major Facilitator Superfamily Efflux Pumps.

Multidrug efflux transporters are major contributors to the antibiotic resistance of Acinetobacter baumannii in clinical settings. Previous studies showed that these transporters are tightly integrated into the physiology of A. baumannii and have diverse functions. However, for many of the efflux pumps, such functions remain poorly defined. In this study, we characterized two putative drug efflux pumps, AmfAB and AmfCD (Acinetobacter Major Facilitator), that are homologous to EmrAB-like transporters from Escherichia coli and other Gram-negative bacteria. These pumps comprise the Major Facilitator Superfamily (MFS) transporters AmfB and AmfD and the periplasmic membrane fusion proteins AmfA and AmfC, respectively. We inactivated and overproduced these pumps in the wild-type ATCC 17978 strain and its derivative strains lacking the major efflux pumps from the Resistance-Nodulation-Division (RND) superfamily and characterized antibiotic susceptibilities and growth of the strains under stresses typical during human infections. We found that neither AmfAB nor AmfCD contribute to the antibiotic non-susceptibility phenotypes of A. baumannii. The two pumps, however, are critical for the adaptation and growth of the bacterium under acidic stress, whereas AmfCD also contributes to growth under conditions of low iron, high temperature, and in the presence of bile salts. These functions are dependent on the presence of the RND pumps, the inactivation of which further diminishes A. baumannii survival and growth. Our results suggest that MFS transporters contribute to stress survival by affecting the permeability properties of the A. baumannii cell envelope.

Genomic and transcriptomic analysis of Ligilactobacillus salivarius IBB3154-in search of new promoters for vaccine construction.

The genome of the strain Ligilactobacillus salivarius IBB3154 was sequenced, and transcriptome analysis was carried out at two different temperatures, allowing the determination of gene expression levels in response to environmental changes (temperature). Genes with higher expression at 42°C were identified. The use of a reporter gene (β- glucuronidase) did not confirm the transcriptomic results; it was found that the promoters of the genes sasA1 and sasA2 were active in the presence of bile salts. This opens up new opportunities for the overexpression of genes of other bacterial species in Ligilactobacillus cells in the intestinal environment.

Probiotic Potential of Bacillus sp. 62A Isolated from a Marine Extreme Environment.

Antimicrobial resistance is an important health concern globally, and probiotics are considered an alternative to minimize it. The present study examined the in vitro probiotic characteristics and in vivo immunomodulatory potential of Bacillus sp. 62A - an extremophile bacterium. Bacillus sp. 62A was evaluated in vitro for its cytotoxicity, hemolytic activity, antibiotic susceptibility, and resistance to gastrointestinal conditions (bile salts, low pH, and intestinal adherence). Additionally, the immunomodulatory effect of Bacillus sp. 62A was studied in mice. The animals were supplemented daily with phosphate-buffered saline (control) and Bacillus sp. 62A at 1 × 108 colony forming units (CFU). Samples were taken on days 5 and 10. Isolated splenocytes were challenged with Escherichia coli for immunological analyses and immune-related gene expression. Serum and feces were collected for IgA and IgG determination. Bacillus sp. 62A did not show cytotoxicity, hemolytic activity, or resistance to antibiotics. Furthermore, the bacterium has autoaggregation and intestinal adhesion capacities and grows in the presence of bile salts and low pH. Bacillus supplementation in mice improved respiratory burst activity, nitric oxide production, and IL-1β and IL-6 gene expressions, mainly at 10days. After E. coli challenge, Bacillus supplementation in mice induced an anti-inflammatory response through a decrease in immunological parameters and an increase in IL-10 gene expression. Moreover, serum IgA and IgG and fecal IgG augmented in supplemented mice. In conclusion, Bacillus sp. 62A has biosafe and immunomodulatory probiotic potential.

Probiotic potential of Bacillus Isolates from Polish Bee Pollen and Bee Bread.

The main goal of this study was the evaluation of the probiotic potential of 10 Bacillus spp. strains isolated from 5 bee bread and 3 bee pollen samples. The antagonistic interaction with Staphylococcus aureus and Escherichia coli was a primary criterion for the preliminary selection of the isolates. Three out of ten strains-PY2.3 (isolated from pollen), BP20.15 and BB10.1 (both isolated from bee bread)-were found to be possible probiotic strains. All these strains are safe for humans (exhibiting [Formula: see text]-hemolytic activity) and meet all essential requirements for probiotics in terms of viability in the presence of bile salts and acid conditions, hydrophobicity, auto-aggregation, and co-aggregation with the cells of important human pathogenic bacteria. They also assimilate more than 30% of cholesterol after 24h of incubation. These three isolates are resistant to penicillin but sensitive (or exhibit moderate resistance) to the other nine antibiotics tested herein. On the basis of whole-genome sequencing, BP20.15 and BB10.1 were classified as B. subtilis and PY2.3 as B. velezensis. Moreover, genomic analyses revealed that all these isolates are potential producers of different antimicrobial compounds, including bacteriocins and secondary metabolites. The outcomes of this study have proven that some of the Bacillus strains isolated from bee pollen or bee bread are potential probiotics.