scholarly journals Genomic Characterization of Non-Mucus-Adherent Derivatives of Lactobacillus rhamnosus GG Reveals Genes Affecting Pilus Biogenesis

2014 ◽  
Vol 80 (22) ◽  
pp. 7001-7009 ◽  
Author(s):  
Pia Rasinkangas ◽  
Justus Reunanen ◽  
François P. Douillard ◽  
Jarmo Ritari ◽  
Virva Uotinen ◽  
...  
Keyword(s):  
Gene Cluster ◽  
Lactobacillus Rhamnosus ◽  
Illumina Miseq ◽  
Class Iii ◽  
Dot Blot ◽  
Content Type ◽  
Intestinal Mucus ◽  
Pilus Biogenesis ◽  
Dot Blot Analysis ◽  
The Impact

ABSTRACTLactobacillus rhamnosusGG is one of the best-characterized lactic acid bacteria and can be considered a probiotic paradigm. Comparative and functional genome analysis showed thatL. rhamnosusGG harbors a genomic island including thespaCBA-srtC1gene cluster, encoding the cell surface-decorating host-interacting pili. Here, induced mutagenesis was used to study pilus biogenesis inL. rhamnosusGG. A combination of two powerful approaches, mutation selection and next-generation sequencing, was applied toL. rhamnosusGG for the selection of pilus-deficient mutants from an enriched population. The isolated mutants were first screened by immuno-dot blot analysis using antiserum against pilin proteins. Relevant mutants were selected, and the lack of pili was confirmed by immunoelectron microscopy. The pilosotype of 10 mutant strains was further characterized by analyzing pilin expression using Western blot, dot blot, and immunofluorescence methods. A mucus binding assay showed that the mutants did not adhere to porcine intestinal mucus. Comparative genome sequence analysis using the Illumina MiSeq platform allowed us to determine the nature of the mutations in the obtained pilus-deficient derivatives. Three major classes of mutants with unique genotypes were observed: class I, with mutations in thesrtC1gene; class II, with a deletion containing thespaCBA-srtC1gene cluster; and class III, with mutations in thespaAgene. Only a limited number of collateral mutations were observed, and one of the pilus-deficient derivatives with a deficientsrtC1gene contained 24 other mutations. This strain, PB12, can be considered a candidate for human trials addressing the impact of the absence of pili.

scholarly journals Impact of Vitamin B12on Formation of the Tetrachloroethene Reductive Dehalogenase in Desulfitobacterium hafniense Strain Y51

2012 ◽  
Vol 78 (22) ◽  
pp. 8025-8032 ◽  
Author(s):  
Anika Reinhold ◽  
Martin Westermann ◽  
Jana Seifert ◽  
Martin von Bergen ◽  
Torsten Schubert ◽  
...  
Keyword(s):  
Gene Cluster ◽  
De Novo ◽  
Anaerobic Bacteria ◽  
Vitamin B ◽  
Content Type ◽  
Reductive Dehalogenase ◽  
Gene Level ◽  
Desulfitobacterium Hafniense ◽  
Pce Dehalogenase ◽  
The Impact

ABSTRACTCorrinoids are essential cofactors of reductive dehalogenases in anaerobic bacteria. Microorganisms mediating reductive dechlorination as part of their energy metabolism are either capable ofde novocorrinoid biosynthesis (e.g.,Desulfitobacteriumspp.) or dependent on exogenous vitamin B12(e.g.,Dehalococcoidesspp.). In this study, the impact of exogenous vitamin B12(cyanocobalamin) and of tetrachloroethene (PCE) on the synthesis and the subcellular localization of the reductive PCE dehalogenase was investigated in the Gram-positiveDesulfitobacterium hafniensestrain Y51, a bacterium able to synthesize corrinoidsde novo. PCE-depleted cells grown for several subcultivation steps on fumarate as an alternative electron acceptor lost the tetrachloroethene-reductive dehalogenase (PceA) activity by the transposition of thepcegene cluster. In the absence of vitamin B12, a gradual decrease of the PceA activity and protein amount was observed; after 5 subcultivation steps with 10% inoculum, more than 90% of the enzyme activity and of the PceA protein was lost. In the presence of vitamin B12, a significant delay in the decrease of the PceA activity with an ∼90% loss after 20 subcultivation steps was observed. This corresponded to the decrease in thepceAgene level, indicating that exogenous vitamin B12hampered the transposition of thepcegene cluster. In the absence or presence of exogenous vitamin B12, the intracellular corrinoid level decreased in fumarate-grown cells and the PceA precursor formed catalytically inactive, corrinoid-free multiprotein aggregates. The data indicate that exogenous vitamin B12is not incorporated into the PceA precursor, even though it affects the transposition of thepcegene cluster.

scholarly journals Comparative Genomic and Functional Analysis of Lactobacillus casei and Lactobacillus rhamnosus Strains Marketed as Probiotics

2013 ◽  
Vol 79 (6) ◽  
pp. 1923-1933 ◽  
Author(s):  
François P. Douillard ◽  
Angela Ribbera ◽  
Hanna M. Järvinen ◽  
Ravi Kant ◽  
Taija E. Pietilä ◽  
...  
Keyword(s):  
Gene Cluster ◽  
Transcription Initiation ◽  
Lactobacillus Rhamnosus ◽  
Comparative Genomic ◽  
Phenotypic Analysis ◽  
Carbohydrate Utilization ◽  
Content Type ◽  
Host Signaling ◽  
Toll Like Receptor 2 ◽  
Genome Content

ABSTRACTFourLactobacillusstrains were isolated from marketed probiotic products, includingL. rhamnosusstrains from Vifit (Friesland Campina) and Idoform (Ferrosan) andL. caseistrains from Actimel (Danone) and Yakult (Yakult Honsa Co.). Their genomes and phenotypes were characterized and compared in detail withL. caseistrain BL23 andL. rhamnosusstrain GG. Phenotypic analysis of the new isolates indicated differences in carbohydrate utilization betweenL. caseiandL. rhamnosusstrains, which could be linked to their genotypes. The two isolatedL. rhamnosusstrains had genomes that were virtually identical to that ofL. rhamnosusGG, testifying to their genomic stability and integrity in food products. TheL. caseistrains showed much greater genomic heterogeneity. Remarkably, all strains contained an intactspaCBApilus gene cluster. However, only theL. rhamnosusstrains produced mucus-binding SpaCBA pili under the conditions tested. Transcription initiation mapping demonstrated that the insertion of aniso-IS30element upstream of the pilus gene cluster inL. rhamnosusstrains but absent inL. caseistrains had constituted a functional promoter driving pilus gene expression. AllL. rhamnosusstrains triggered an NF-κB response via Toll-like receptor 2 (TLR2) in a reporter cell line, whereas theL. caseistrains did not or did so to a much lesser extent. This study demonstrates that the twoL. rhamnosusstrains isolated from probiotic products are virtually identical toL. rhamnosusGG and further highlights the differences between these andL. caseistrains widely marketed as probiotics, in terms of genome content, mucus-binding and metabolic capacities, and host signaling capabilities.

scholarly journals Lactobacillus rhamnosus GG Genomic and Phenotypic Stability in an Industrial Production Process

2020 ◽  
Vol 86 (6) ◽  
Author(s):  
Marianne Stage ◽  
Anita Wichmann ◽  
Mette Jørgensen ◽  
Natalia Ivonne Vera-Jimenéz ◽  
Malue Wielje ◽  
...  
Keyword(s):  
Production Process ◽  
Gene Cluster ◽  
Industrial Production ◽  
Lactobacillus Rhamnosus ◽  
Probiotic Properties ◽  
Phenotypic Stability ◽  
Content Type ◽  
Freeze Dried ◽  
Probiotic Strains

ABSTRACT Lactobacillus rhamnosus GG is one of the most widely marketed and studied probiotic strains. In L. rhamnosus GG, the spaCBA-srtC1 gene cluster encodes pili, which are important for some of the probiotic properties of the strain. A previous study showed that the DNA sequence of the spaCBA-srtC1 gene cluster was not present in some L. rhamnosus GG variants isolated from liquid dairy products. To examine the stability of the L. rhamnosus GG genome in an industrial production process, we sequenced the genome of samples of L. rhamnosus GG (DSM 33156) collected at specific steps of the industrial production process, including the culture collection stock, intermediate fermentations, and final freeze-dried products. We found that the L. rhamnosus GG genome sequence was unchanged throughout the production process. Consequently, the spaCBA-srtC1 gene locus was intact and fully conserved in all 31 samples examined. In addition, different production batches of L. rhamnosus GG exhibited consistent phenotypes, including the presence of pili in final freeze-dried products, and consistent characteristics in in vitro assays of probiotic properties. Our data show that L. rhamnosus GG is highly stable in this industrial production process. IMPORTANCE Lactobacillus rhamnosus GG is one of the best-studied probiotic strains. One of the well-characterized features of the strain is the pili encoded by the spaCBA-srtC1 gene cluster. These pili are involved in persistence in the gastrointestinal tract and are important for the probiotic properties of L. rhamnosus GG. Previous studies demonstrated that the L. rhamnosus GG genome can be unstable under certain conditions and can lose the spaCBA-srtC1 gene cluster. Since in vitro studies have shown that the loss of the spaCBA-srtC1 gene cluster decreases certain L. rhamnosus GG probiotic properties, we assessed both the genomic stability and phenotypic properties of L. rhamnosus GG throughout an industrial production process. We found that neither genomic nor phenotypic changes occurred in the samples. Therefore, we demonstrate that L. rhamnosus GG retains the spaCBA-srtC1 cluster and exhibits excellent genomic and phenotypic stability in the specific industrial process examined here.

Does combination of Lactobacillus rhamnosus Heriz I and beta glucan improve quality of life in women with breast cancer receiving chemotherapy?

2019 ◽  
Vol 50 (3) ◽  
pp. 569-578 ◽  
Author(s):  
Alireza Monshikarimi ◽  
Alireza Ostadrahimi ◽  
Mohammad Asghari Jafarabadi ◽  
Jamal EivaziZiaei ◽  
Abolfazl Barzeghari ◽  
...  
Keyword(s):  
Breast Cancer ◽  
Quality Of Life ◽  
Placebo Group ◽  
Intervention Group ◽  
Lactobacillus Rhamnosus ◽  
Controlled Clinical Trial ◽  
Beta Glucan ◽  
Content Type ◽  
The Impact

Purpose This paper aims to study the impact of combination Lactobacillus Rhamnosus Heriz I and Beta glucan on quality of life (QOL) in breast cancer (BC) patients receiving chemotherapy. Design/methodology/approach In all, 30 women with BC were enrolled in this randomized double-blind placebo-controlled clinical trial. Intervention group received either daily one capsule of L. rhamnosus strain Heriz I (2 × 107 CFU) and two 10-mg capsules of soluble1–3,1–6,D-beta glucan and the placebo group receiving placebo, interval between two courses of chemotherapy. Health-related QOL was evaluated at baseline and end of the trial using the EORTC QOL Questionnaire version.3.0 (EORTC QLQ-C30). Findings The results showed that combination of L. rhamnosus Heriz I and Beta glucan supplementation in BC patients during chemotherapy improved functional scales score from 37.3 ± 12.1 to 39.8 ± 6.8 and this increase in comparison with placebo was significant after adjusting baseline value (p = 0.015). Changes in symptoms scores were not significant after intervention (p = 0.05) but the decreased score (66.1 ± 12.9 to 60.6 ± 9.4) was considerable clinically. As well as, changes in global health status/QOL score in intervention group in comparison with the placebo group was not significant (p = 0.93). Originality/value The findings suggest that combination of L. rhamnosus Heriz I and Beta glucan may be associated for improving functional scales and symptoms in patients with BC who underwent chemotherapy. To prove positive effect of supplements on other aspects of QOL, further studies are needed.

scholarly journals Cloning and Sequencing of the Genes Downstream of thewbf Gene Cluster of Vibrio cholerae Serogroup O139 and Analysis of the Junction Genes in Other Serogroups

1999 ◽  
Vol 67 (10) ◽  
pp. 5033-5040 ◽  
Author(s):  
Shanmuga Sozhamannan ◽  
Ying Kang Deng ◽  
Manrong Li ◽  
Alexander Sulakvelidze ◽  
James B. Kaper ◽  
...  
Keyword(s):  
Vibrio Cholerae ◽  
Gene Cluster ◽  
Hot Spot ◽  
Dot Blot ◽  
Sequence Element ◽  
O Antigen ◽  
Conserved Genes ◽  
Emergent Pathogen ◽  
Dot Blot Analysis ◽  
Similar Organization

ABSTRACT The DNA sequence of the O-antigen biosynthesis cluster (wbf) of a recently emergent pathogen, Vibrio cholerae serogroup O139, has been determined. Here we report the sequence of the genes downstream of the O139 wbfX gene and analysis of the genes flanking the wbf gene cluster in other serogroups. The gene downstream of wbfX, designatedrjg (right junction gene), is predicted to be not required for O-antigen biosynthesis but appears to be a hot spot for DNA rearrangements. Several variants of the rjg gene (three different insertions and a deletion) have been found in other serogroups. DNA dot blot analysis of 106 V. choleraestrains showed the presence of the left and right junction genes,gmhD and rjg, respectively, in all strains. Further, these genes mapped to a single I-CeuI fragment in all 21 strains analyzed by pulsed-field gel electrophoresis, indicating a close linkage. The insertion sequence element IS1358, found in both O1 and O139 wb* regions, is present in 61% of the strains tested; interestingly, where present, it is predominantly linked to the wb* region. These results indicated a cassette-like organization of the wb* region, with the conserved genes (gmhD and rjg) flanking the divergent, serogroup-specific wb* genes and IS1358. A similar organization of the wb* region in other serogroups raises the possibility of the emergence of new pathogens by homologous recombination via the junction genes.

scholarly journals Piliation of Lactobacillus rhamnosus GG Promotes Adhesion, Phagocytosis, and Cytokine Modulation in Macrophages

2015 ◽  
Vol 81 (6) ◽  
pp. 2050-2062 ◽  
Author(s):  
Cynthia E. Vargas García ◽  
Mariya Petrova ◽  
Ingmar J. J. Claes ◽  
Ilke De Boeck ◽  
Tine L. A. Verhoeven ◽  
...  
Keyword(s):  
Close Contact ◽  
Lactobacillus Rhamnosus ◽  
Interleukin 10 ◽  
Raw 264.7 Cells ◽  
Raw 264.7 ◽  
Phagocytic Cells ◽  
Content Type ◽  
Intestinal Mucus ◽  
Anti Inflammatory

ABSTRACTRecently,spaCBA-encoded pili on the cell surface ofLactobacillus rhamnosusGG were identified to be key molecules for binding to human intestinal mucus and Caco-2 intestinal epithelial cells. Here, we investigated the role of the SpaCBA pilus ofL. rhamnosusGG in the interaction with macrophagesin vitroby comparing the wild type with surface mutants. Our results show that SpaCBA pili play a significant role in the capacity for adhesion to macrophages and also promote bacterial uptake by these phagocytic cells. Interestingly, our data suggest that SpaCBA pili also mediate anti-inflammatory effects by induction of interleukin-10 (IL-10) mRNA and reduction of interleukin-6 (IL-6) mRNA in a murine RAW 264.7 macrophage cell line. These pili appear to mediate these effects indirectly by promoting close contact with the macrophages, facilitating the exertion of anti-inflammatory effects by other surface molecules via yet unknown mechanisms. Blockage of complement receptor 3 (CR3), previously identified to be a receptor for streptococcal pili, significantly decreased the uptake of pilus-expressing strains in RAW 264.7 cells, while the expression of IL-10 and IL-6 mRNA by these macrophages was not affected by this blocking. On the other hand, blockage of Toll-like receptor 2 (TLR2) significantly reduced the expression of IL-6 mRNA irrespective of the presence of pili.

scholarly journals Functional Dynamics of the Gut Microbiome in Elderly People during Probiotic Consumption

mBio ◽  
2015 ◽  
Vol 6 (2) ◽  
Author(s):  
Emiley A. Eloe-Fadrosh ◽  
Arthur Brady ◽  
Jonathan Crabtree ◽  
Elliott F. Drabek ◽  
Bing Ma ◽  
...  
Keyword(s):  
Gut Microbiota ◽  
Lactobacillus Rhamnosus ◽  
Probiotic Bacteria ◽  
Healthy Human ◽  
Content Type ◽  
Treatment Comparison ◽  
Functional Dynamics ◽  
Single Organism ◽  
Probiotic Treatment ◽  
The Impact

ABSTRACT A mechanistic understanding of the purported health benefits conferred by consumption of probiotic bacteria has been limited by our knowledge of the resident gut microbiota and its interaction with the host. Here, we detail the impact of a single-organism probiotic, Lactobacillus rhamnosus GG ATCC 53103 (LGG), on the structure and functional dynamics (gene expression) of the gut microbiota in a study of 12 healthy individuals, 65 to 80 years old. The analysis revealed that while the overall community composition was stable as assessed by 16S rRNA profiling, the transcriptional response of the gut microbiota was modulated by probiotic treatment. Comparison of transcriptional profiles based on taxonomic composition yielded three distinct transcriptome groups that displayed considerable differences in functional dynamics. The transcriptional profile of LGG in vivo was remarkably concordant across study subjects despite the considerable interindividual nature of the gut microbiota. However, we identified genes involved in flagellar motility, chemotaxis, and adhesion from Bifidobacterium and the dominant butyrate producers Roseburia and Eubacterium whose expression was increased during probiotic consumption, suggesting that LGG may promote interactions between key constituents of the microbiota and the host epithelium. These results provide evidence for the discrete functional effects imparted by a specific single-organism probiotic and challenge the prevailing notion that probiotics substantially modify the resident microbiota within nondiseased individuals in an appreciable fashion. IMPORTANCE Probiotic bacteria have been used for over a century to promote digestive health. Many individuals report that probiotics alleviate a number of digestive issues, yet little evidence links how probiotic microbes influence human health. Here, we show how the resident microbes that inhabit the healthy human gut respond to a probiotic. The well-studied probiotic Lactobacillus rhamnosus GG ATCC 53103 (LGG) was administered in a clinical trial, and a suite of measurements of the resident microbes were taken to evaluate potential changes over the course of probiotic consumption. We found that LGG transiently enriches for functions to potentially promote anti-inflammatory pathways in the resident microbes.

scholarly journals Functional Characterization of a Mucus-Specific LPXTG Surface Adhesin from Probiotic Lactobacillus rhamnosus GG

2011 ◽  
Vol 77 (13) ◽  
pp. 4465-4472 ◽  
Author(s):  
Ingemar von Ossowski ◽  
Reetta Satokari ◽  
Justus Reunanen ◽  
Sarah Lebeer ◽  
Sigrid C. J. De Keersmaecker ◽  
...  
Keyword(s):  
Molecular Mechanisms ◽  
Current Knowledge ◽  
Functional Characterization ◽  
Lactobacillus Rhamnosus ◽  
Probiotic Strain ◽  
Surface Proteins ◽  
Adhesive Interaction ◽  
Content Type ◽  
Intestinal Mucus ◽  
Mucosal Surfaces

ABSTRACTIn spite of the wealth of clinical evidence supporting the health benefits ofLactobacillus rhamnosusGG in humans, there is still a lack of understanding of the molecular mechanisms behind its probiosis. Current knowledge suggests that the health-promoting effects of this probiotic strain might be partly dependent on its persistence in the intestine and adhesion to mucosal surfaces. Moreover,L. rhamnosusGG contains mucus-binding pili that might also explain the occupation of its ecological niche as a comparatively less stringent allochthonous intestine-dwelling bacterium. To uncover additional surface proteins involved in mucosal adhesion, we investigated the adherence properties of the only predicted protein (LGG_02337) inL. rhamnosusGG that exhibits homology with a known mucus-binding domain. We cloned a recombinant form of the gene for this putative mucus adhesin and established that the purified protein readily adheres to human intestinal mucus. We also showed that this mucus adhesin is visibly distributed throughout the cell surface and participates in the adhesive interaction betweenL. rhamnosusGG and mucus, although less prominently than the mucus-binding pili in this strain. Based on primary structural comparisons, we concluded that the current annotation of the LGG_02337 protein likely does not accurately reflect its predicted properties, and we propose that this mucus-specific adhesin be called the mucus-binding factor (MBF). Finally, we interpret our results to mean thatL. rhamnosusGG MBF, as an active mucus-specific surface adhesin with a presumed ancillary involvement in pilus-mediated mucosal adhesion, plays a part in the adherent mechanisms during intestinal colonization by this probiotic.

scholarly journals Polymorphisms, Chromosomal Rearrangements, and Mutator Phenotype Development during Experimental Evolution of Lactobacillus rhamnosus GG

2016 ◽  
Vol 82 (13) ◽  
pp. 3783-3792 ◽  
Author(s):  
François P. Douillard ◽  
Angela Ribbera ◽  
Kun Xiao ◽  
Jarmo Ritari ◽  
Pia Rasinkangas ◽  
...  
Keyword(s):  
Gene Cluster ◽  
Chromosomal Rearrangements ◽  
Lactobacillus Rhamnosus ◽  
Probiotic Strain ◽  
Shearing Stress ◽  
Genomic Integrity ◽  
Mutator Phenotype ◽  
Content Type ◽  
Is Elements ◽  
The Stability

ABSTRACTLactobacillus rhamnosusGG is a lactic acid bacterium widely marketed by the food industry. Its genomic analysis led to the identification of a gene cluster encoding mucus-binding SpaCBA pili, which is located in a genomic island enriched in insertion sequence (IS) elements. In the present study, we analyzed by genome-wide resequencing the genomic integrity ofL. rhamnosusGG in four distinct evolutionary experiments conducted for approximately 1,000 generations under conditions of no stress or salt, bile, and repetitive-shearing stress. Under both stress-free and salt-induced stress conditions, the GG population (excluding the mutator lineage in the stress-free series [see below]) accumulated only a few single nucleotide polymorphisms (SNPs) and no frequent chromosomal rearrangements. In contrast, in the presence of bile salts or repetitive shearing stress, some IS elements were found to be activated, resulting in the deletion of large chromosomal segments that include thespaCBA-srtC1pilus gene cluster. Remarkably, a high number of SNPs were found in three strains obtained after 900 generations of stress-free growth. Detailed analysis showed that these three strains derived from a founder mutant with an altered DNA polymerase subunit that resulted in a mutator phenotype. The present work confirms the stability of the pilus production phenotype inL. rhamnosusGG under stress-free conditions, highlights the possible evolutionary scenarios that may occur when this probiotic strain is extensively cultured, and identifies external factors that affect the chromosomal integrity of GG. The results provide mechanistic insights into the stability of GG in regard to its extensive use in probiotic and other functional food products.IMPORTANCELactobacillus rhamnosusGG is a widely marketed probiotic strain that has been used in numerous clinical studies to assess its health-promoting properties. Hence, the stability of the probiotic functions ofL. rhamnosusGG is of importance, and here we studied the impact of external stresses on the genomic integrity ofL. rhamnosusGG. We studied three different stresses that are relevant for understanding its robustness and integrity under bothex vivoconditions, i.e., industrial manufacturing conditions, andin vivoconditions, i.e., intestinal tract-associated stress. Overall, our findings contribute to predicting the genomic stability ofL. rhamnosusGG and its ecological performance.

The More Control, the Better?

2014 ◽  
Vol 26 (2) ◽  
pp. 81-91 ◽  
Author(s):  
Jeeyun Oh ◽  
Mun-Young Chung ◽  
Sangyong Han
Keyword(s):  
Mixed Design ◽  
Negative Effects ◽  
Story Structure ◽  
Content Type ◽  
High User ◽  
User Control ◽  
Rigorous Research ◽  
Control Versus ◽  
The Media ◽  
The Impact

Despite of the popularity of interactive movie trailers, rigorous research on one of the most apparent features of these interfaces – the level of user control – has been scarce. This study explored the effects of user control on users’ immersion and enjoyment of the movie trailers, moderated by the content type. We conducted a 2 (high user control versus low user control) × 2 (drama film trailer versus documentary film trailer) mixed-design factorial experiment. The results showed that the level of user control over movie trailer interfaces decreased users’ immersion when the trailer had an element of traditional story structure, such as a drama film trailer. Participants in the high user control condition answered that they were less fascinated with, absorbed in, focused on, mentally involved with, and emotionally affected by the movie trailer than participants in the low user control condition only with the drama movie trailer. The negative effects of user control on the level of immersion for the drama trailer translated into users’ enjoyment. The impact of user control over interfaces on immersion and enjoyment varies depending on the nature of the media content, which suggests a possible trade-off between the level of user control and entertainment outcomes.

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