scholarly journals Comparative Genome Analysis of Bifidobacterium longum subsp. infantis Strains Reveals Variation in Human Milk Oligosaccharide Utilization Genes among Commercial Probiotics

Nutrients ◽  
2020 ◽  
Vol 12 (11) ◽  
pp. 3247 ◽  
Author(s):  
Rebbeca M. Duar ◽  
Giorgio Casaburi ◽  
Ryan D. Mitchell ◽  
Lindsey N.C. Scofield ◽  
Camila A. Ortega Ramirez ◽  
...  
Keyword(s):  
Human Milk ◽  
Bifidobacterium Longum ◽  
Gene Clusters ◽  
In Vitro Models ◽  
Breastfed Infants ◽  
Fitness Advantage ◽  
Probiotic Strains ◽  
Commercial Probiotics

Dysbiosis is associated with acute and long-term consequences for neonates. Probiotics can be effective in limiting the growth of bacteria associated with dysbiosis and promoting the healthy development of the infant microbiome. Given its adaptation to the infant gut, and promising data from animal and in vitro models, Bifidobacterium longum subsp. infantis is an attractive candidate for use in infant probiotics. However, strain-level differences in the ability of commercialized strains to utilize human milk oligosaccharides (HMOs) may have implications in the performance of strains in the infant gut. In this study, we characterized twelve B. infantis probiotic strains and identified two main variants in one of the HMO utilization gene clusters. Some strains possessed the full repertoire of HMO utilization genes (H5-positive strains), while H5-negative strains lack an ABC-type transporter known to bind core HMO structures. H5-positive strains achieved significantly superior growth on lacto-N-tetraose and lacto-N-neotetraose. In vitro, H5-positive strains had a significant fitness advantage over H5-negative strains, which was also observed in vivo in breastfed infants. This work provides evidence of the functional implications of genetic differences among B. infantis strains and highlights that genotype and HMO utilization phenotype should be considered when selecting a strain for probiotic use in infants.

scholarly journals Targeted high-resolution taxonomic identification of Bifidobacterium longum subsp. infantis using human milk oligosaccharide metabolizing genes

2021 ◽  
Author(s):  
Lauren Tso ◽  
Kevin S Bonham ◽  
Alyssa Fishbein ◽  
Sophie Rowland ◽  
Vanja Klepac-Ceraj ◽  
...  
Keyword(s):  
Human Milk ◽  
Bifidobacterium Longum ◽  
Gene Clusters ◽  
Species Group ◽  
Human Breast Milk ◽  
Metagenomic Sequencing ◽  
Sequencing Data ◽  
Specific Primers ◽  
Breastfed Infants ◽  
Shotgun Metagenomic Sequencing

Bifidobacterium longum subsp. infantis (B. infantis) is one of few microorganisms capable of metabolizing human breast milk and is a pioneer colonizer in the guts of breastfed infants. One current challenge is differentiating B. infantis from its close relatives, B. longum and B. suis, by molecular methods. These two organisms are classified in the same species group as B. infantis but do not share the ability to metabolize human milk oligosaccharides (HMOs). Here, we compared HMO-metabolizing genes across different Bifidobacterium genomes to develop B. infantis specific primers and determine if they alone can be used to quickly characterize B. infantis with shotgun metagenomic sequencing data. We showed that B. infantis is uniquely identified by the presence of five HMO-metabolizing gene clusters, used this characterization to test for its prevalence in infants, and validated the results using the B. infantis-specific primers. By examining stool samples from a cohort of US children and pregnant women using shotgun metagenomic sequencing, we observed that only 18 of 204 (8.8%) of children under 2 years old harbored B. infantis. These results highlight the importance of developing and improving approaches to identify B. infantis. A more accurate characterization may provide insights into regional differences of B. infantis prevalence in infant gut microbiota.

Assessment of the synbiotic properites of human milk oligosaccharides and Bifidobacterium longum subsp. infantis in vitro and in humanised mice

2017 ◽  
Vol 8 (2) ◽  
pp. 281-289 ◽  
Author(s):  
S. Musilova ◽  
N. Modrackova ◽  
P. Hermanova ◽  
T. Hudcovic ◽  
R. Svejstil ◽  
...  
Keyword(s):  
Caesarean Section ◽  
Human Milk ◽  
Pathogenic Bacteria ◽  
Bifidobacterium Longum ◽  
Mode Of Delivery ◽  
Human Milk Oligosaccharides ◽  
Milk Oligosaccharides ◽  
High Concentrations

The mode of delivery plays a crucial role in infant gastrointestinal tract colonisation, which in the case of caesarean section is characterised by the presence of clostridia and low bifidobacterial counts. Gut colonisation can be modified by probiotics, prebiotics or synbiotics. Human milk oligosaccharides (HMOs) are infant prebiotics that show a bifidogenic effect. Moreover, genome sequencing of Bifidobacterium longum subsp. infantis within the infant microbiome revealed adaptations for milk utilisation. This study aimed to evaluate the synbiotic effect of B. longum subsp. infantis, HMOs and human milk (HM) both in vitro and in vivo (in a humanised mouse model) in the presence of faecal microbiota from infants born by caesarean section. The combination of B. longum and HMOs or HM reduced the clostridia and G-bacteria counts both in vitro and in vivo. The bifidobacterial population in vitro significantly increased and produce high concentrations of acetate and lactate. In vitro competition assays confirmed that the tested bifidobacterial strain is a potential probiotic for infants and, together with HMOs or HM, acts as a synbiotic. It is also able to inhibit potentially pathogenic bacteria. The synbiotic effects identified in vitro were not observed in vivo. However, there was a significant reduction in clostridia counts in both experimental animal groups (HMOs + B. longum and HM + B. longum), and a specific immune response via increased interleukin (IL)-10 and IL-6 production. Animal models do not perfectly mimic human conditions; however, they are essential for testing the safety of functional foods.

scholarly journals Combinational effects of prebiotic oligosaccharides on bifidobacterial growth and host gene expression in a simplified mixed culture model and neonatal mice

2016 ◽  
Vol 116 (2) ◽  
pp. 270-278 ◽  
Author(s):  
Tatsuya Ehara ◽  
Hirohisa Izumi ◽  
Muneya Tsuda ◽  
Yuki Nakazato ◽  
Hiroshi Iwamoto ◽  
...  
Keyword(s):  
Gut Microbiota ◽  
Mixed Culture ◽  
Bifidobacterium Longum ◽  
Intestinal Content ◽  
Cell Numbers ◽  
Breastfed Infants ◽  
Neonatal Mice ◽  
Culture Model

AbstractIt is important to provide formula-fed infants with a bifidobacteria-enriched gut microbiota similar to those of breastfed infants to ensure intestinal health. Prebiotics, such as certain oligosaccharides, are a useful solution to this problem, but the combinational benefits of these oligosaccharides have not been evaluated. This study investigated the benefits of oligosaccharide combinations and screened for an optimal combination of oligosaccharides to promote healthy gut microbiota of formula-fed infants. In vitro and in vivo experiments were performed to assess the bifidogenic effects of lactulose (LAC) alone and LAC combined with raffinose (RAF) and/or galacto-oligosaccharide (GOS), using a mixed culture model and neonatal mice orally administered with these oligosaccharides and Bifidobacterium breve. In the in vitro culture model, the combination of the three oligosaccharides (LAC–RAF–GOS) significantly increased cell numbers of B. breve and Bifidobacterium longum (P<0·05) compared with either LAC alone or the combination of two oligosaccharides, and resulted in the production of SCFA under anaerobic conditions. In the in vivo experiment, the LAC–RAF–GOS combination significantly increased cell numbers of B. breve and Bacteroidetes in the large intestinal content (P<0·05) and increased acetate concentrations in the caecal content and serum of neonatal mice. Genes related to metabolism and immune responses were differentially expressed in the liver and large intestine of mice administered with LAC–RAF–GOS. These results indicate a synergistic effect of the LAC–RAF–GOS combination on the growth of bifidobacteria and reveal possible benefits of this combination to the gut microbiota and health of infants.

scholarly journals Multiple Selection Criteria for Probiotic Strains with High Potential for Obesity Management

Nutrients ◽  
2021 ◽  
Vol 13 (3) ◽  
pp. 713
Author(s):  
Jeanne Alard ◽  
Benoit Cudennec ◽  
Denise Boutillier ◽  
Véronique Peucelle ◽  
Amandine Descat ◽  
...  
Keyword(s):  
Gut Microbiota ◽  
Leptin Receptor ◽  
Body Weight Gain ◽  
Bifidobacterium Longum ◽  
Protective Effects ◽  
Metabolic Dysfunction ◽  
Diet Induced Obesity ◽  
Probiotic Strains

Since alterations of the gut microbiota have been shown to play a major role in obesity, probiotics have attracted attention. Our aim was to identify probiotic candidates for the management of obesity using a combination of in vitro and in vivo approaches. We evaluated in vitro the ability of 23 strains to limit lipid accumulation in adipocytes and to enhance the secretion of satiety-promoting gut peptide in enteroendocrine cells. Following the in vitro screening, selected strains were further investigated in vivo, single, or as mixtures, using a murine model of diet-induced obesity. Strain Bifidobacterium longum PI10 administrated alone and the mixture of B. animalis subsp. lactis LA804 and Lactobacillus gasseri LA806 limited body weight gain and reduced obesity-associated metabolic dysfunction and inflammation. These protective effects were associated with changes in the hypothalamic gene expression of leptin and leptin receptor as well as with changes in the composition of gut microbiota and the profile of bile acids. This study provides crucial clues to identify new potential probiotics as effective therapeutic approaches in the management of obesity, while also providing some insights into their mechanisms of action.

scholarly journals Targeted High-Resolution Taxonomic Identification of Bifidobacterium longum subsp. infantis Using Human Milk Oligosaccharide Metabolizing Genes

Nutrients ◽  
2021 ◽  
Vol 13 (8) ◽  
pp. 2833
Author(s):  
Lauren Tso ◽  
Kevin S. Bonham ◽  
Alyssa Fishbein ◽  
Sophie Rowland ◽  
Vanja Klepac-Ceraj
Keyword(s):  
Human Milk ◽  
Regional Differences ◽  
Bifidobacterium Longum ◽  
Gene Clusters ◽  
Species Group ◽  
Human Breast Milk ◽  
Specific Primers ◽  
Breastfed Infants ◽  
Close Relatives ◽  
Us Children

Bifidobacterium longum subsp. infantis (B. infantis) is one of a few microorganisms capable of metabolizing human breast milk and is a pioneer colonizer in the guts of breastfed infants. One current challenge is differentiating B. infantis from its close relatives, B. longum and B. suis. All three organisms are classified in the same species group but only B. infantis can metabolize human milk oligosaccharides (HMOs). We compared HMO-metabolizing genes across different Bifidobacterium genomes and developed B. infantis-specific primers to determine if the genes alone or the primers can be used to quickly characterize B. infantis. We showed that B. infantis is uniquely identified by the presence of five HMO-metabolizing gene clusters, tested for its prevalence in infant gut metagenomes, and validated the results using the B. infantis-specific primers. We observed that only 15 of 203 (7.4%) children under 2 years old from a cohort of US children harbored B. infantis. These results highlight the importance of developing and improving approaches to identify B. infantis. A more accurate characterization may provide insights into regional differences of B. infantis prevalence in infant gut microbiota.

Modulation of Matrix Metalloproteinases by Plant-Derived Products

2020 ◽  
Vol 20 ◽  
Author(s):  
Nur Najmi Mohamad Anuar ◽  
Nurul Iman Natasya Zulkafali ◽  
Azizah Ugusman
Keyword(s):  
Clinical Trials ◽  
Natural Products ◽  
Matrix Metalloproteinases ◽  
Animal Studies ◽  
Current Knowledge ◽  
In Vitro Models ◽  
In Vivo Studies ◽  
Extracellular Matrix Components

: Matrix metalloproteinases (MMPs) are a group of zinc-dependent metallo-endopeptidase that are responsible towards the degradation, repair and remodelling of extracellular matrix components. MMPs play an important role in maintaining a normal physiological function and preventing diseases such as cancer and cardiovascular diseases. Natural products derived from plants have been used as traditional medicine for centuries. Its active compounds, such as catechin, resveratrol and quercetin, are suggested to play an important role as MMPs inhibitors, thereby opening new insights into their applications in many fields, such as pharmaceutical, cosmetic and food industries. This review summarises the current knowledge on plant-derived natural products with MMP-modulating activities. Most of the reviewed plant-derived products exhibit an inhibitory activity on MMPs. Amongst MMPs, MMP-2 and MMP-9 are the most studied. The expression of MMPs is inhibited through respective signalling pathways, such as MAPK, NF-κB and PI3 kinase pathways, which contribute to the reduction in cancer cell behaviours, such as proliferation and migration. Most studies have employed in vitro models, but a limited number of animal studies and clinical trials have been conducted. Even though plant-derived products show promising results in modulating MMPs, more in vivo studies and clinical trials are needed to support their therapeutic applications in the future.

scholarly journals Human Saliva-Mediated Hydrolysis of Eugenyl-β-D-Glucoside and Fluorescein-di-β-D-Glucoside in In Vivo and In Vitro Models

Biomolecules ◽  
2021 ◽  
Vol 11 (2) ◽  
pp. 172
Author(s):  
Mariusz Dziadas ◽  
Adam Junka ◽  
Henryk Jeleń
Keyword(s):  
Oral Cavity ◽  
Control Sample ◽  
Rapid Test ◽  
Human Saliva ◽  
In Vitro Models ◽  
Microbial Hydrolysis ◽  
Amoxicillin Trihydrate ◽  
Potassium Clavulanate

Eugenyl-β-D-glucopyranoside, also referred to as Citrusin C, is a natural glucoside found among others in cloves, basil and cinnamon plants. Eugenol in a form of free aglycone is used in perfumeries, flavourings, essential oils and in medicinal products. Synthetic Citrusin C was incubated with human saliva in several in vitro models together with substrate-specific enzyme and antibiotics (clindamycin, ciprofloxacin, amoxicillin trihydrate and potassium clavulanate). Citrusin C was detected using liquid chromatography with tandem mass spectrometry (LC-MS/MS). Citrusin C was completely degraded only when incubated with substrate-specific A. niger glucosidase E.C 3.2.1.21 (control sample) and when incubated with human saliva (tested sample). The addition of antibiotics to the above-described experimental setting, stopped Citrusin C degradation, indicating microbiologic origin of hydrolysis observed. Our results demonstrate that Citrusin C is subjected to complete degradation by salivary/oral cavity microorganisms. Extrapolation of our results allows to state that in the human oral cavity, virtually all β-D-glucosides would follow this type of hydrolysis. Additionally, a new method was developed for an in vivo rapid test of glucosidase activity in the human mouth on the tongue using fluorescein-di-β-D-glucoside as substrate. The results presented in this study serve as a proof of concept for the hypothesis that microbial hydrolysis path of β-D-glucosides begins immediately in the human mouth and releases the aglycone directly into the gastrointestinal tract.

scholarly journals O23: CHARACTERISING PATIENT-DERIVED COLORECTAL CANCER TISSUE-ORIGINATED ORGANOIDAL SPHEROIDS FOR HIGH-THROUGHPUT MICROFLUIDIC APPLICATIONS

2021 ◽  
Vol 108 (Supplement_1) ◽  
Author(s):  
MI Khot ◽  
M Levenstein ◽  
R Coppo ◽  
J Kondo ◽  
M Inoue ◽  
...  
Keyword(s):  
Colorectal Cancer ◽  
Fluid Flow ◽  
High Throughput ◽  
Microfluidic Devices ◽  
In Vitro Models ◽  
Fluorescent Imaging ◽  
Cancer Tissue ◽  
Cell Models

Abstract Introduction Three-dimensional (3D) cell models have gained reputation as better representations of in vivo cancers as compared to monolayered cultures. Recently, patient tumour tissue-derived organoids have advanced the scope of complex in vitro models, by allowing patient-specific tumour cultures to be generated for developing new medicines and patient-tailored treatments. Integrating 3D cell and organoid culturing into microfluidics, can streamline traditional protocols and allow complex and precise high-throughput experiments to be performed with ease. Method Patient-derived colorectal cancer tissue-originated organoidal spheroids (CTOS) cultures were acquired from Kyoto University, Japan. CTOS were cultured in Matrigel and stem-cell media. CTOS were treated with 5-fluorouracil and cytotoxicity evaluated via fluorescent imaging and ATP assay. CTOS were embedded, sectioned and subjected to H&E staining and immunofluorescence for ABCG2 and Ki67 proteins. HT29 colorectal cancer spheroids were produced on microfluidic devices using cell suspensions and subjected to 5-fluorouracil treatment via fluid flow. Cytotoxicity was evaluated through fluorescent imaging and LDH assay. Result 5-fluorouracil dose-dependent reduction in cell viability was observed in CTOS cultures (p&lt;0.01). Colorectal CTOS cultures retained the histology, tissue architecture and protein expression of the colonic epithelial structure. Uniform 3D HT29 spheroids were generated in the microfluidic devices. 5-fluorouracil treatment of spheroids and cytotoxic analysis was achieved conveniently through fluid flow. Conclusion Patient-derived CTOS are better complex models of in vivo cancers than 3D cell models and can improve the clinical translation of novel treatments. Microfluidics can streamline high-throughput screening and reduce the practical difficulties of conventional organoid and 3D cell culturing. Take-home message Organoids are the most advanced in vitro models of clinical cancers. Microfluidics can streamline and improve traditional laboratory experiments.

scholarly journals Homozygous MTAP deletion in primary human glioblastoma is not associated with elevation of methylthioadenosine

2021 ◽  
Vol 12 (1) ◽  
Author(s):  
Yasaman Barekatain ◽  
Jeffrey J. Ackroyd ◽  
Victoria C. Yan ◽  
Sunada Khadka ◽  
Lin Wang ◽  
...  
Keyword(s):  
Clinical Relevance ◽  
Homozygous Deletion ◽  
In Vitro Models ◽  
Arginine Methyltransferase ◽  
Human Glioblastoma ◽  
Primary Glioblastoma ◽  
Methylthioadenosine Phosphorylase ◽  
Metabolomic Profiling

AbstractHomozygous deletion of methylthioadenosine phosphorylase (MTAP) in cancers such as glioblastoma represents a potentially targetable vulnerability. Homozygous MTAP-deleted cell lines in culture show elevation of MTAP’s substrate metabolite, methylthioadenosine (MTA). High levels of MTA inhibit protein arginine methyltransferase 5 (PRMT5), which sensitizes MTAP-deleted cells to PRMT5 and methionine adenosyltransferase 2A (MAT2A) inhibition. While this concept has been extensively corroborated in vitro, the clinical relevance relies on exhibiting significant MTA accumulation in human glioblastoma. In this work, using comprehensive metabolomic profiling, we show that MTA secreted by MTAP-deleted cells in vitro results in high levels of extracellular MTA. We further demonstrate that homozygous MTAP-deleted primary glioblastoma tumors do not significantly accumulate MTA in vivo due to metabolism of MTA by MTAP-expressing stroma. These findings highlight metabolic discrepancies between in vitro models and primary human tumors that must be considered when developing strategies for precision therapies targeting glioblastoma with homozygous MTAP deletion.

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