Antibiotics involved in Clostridium difficile-associated disease increase colonization factor gene expression

Clostridium difficile is the most common cause of antibiotic-associated diarrhoea. Antibiotics are presumed to disturb the normal intestinal microbiota, leading to depletion of the barrier effect and colonization by pathogenic bacteria. This first step of infection includes adherence to epithelial cells. We investigated the impact of various environmental conditions in vitro on the expression of genes encoding known, or putative, colonization factors: three adhesins, P47 (one of the two S-layer proteins), Cwp66 and Fbp68, and a protease, Cwp84. The conditions studied included hyperosmolarity, iron depletion and exposure to several antibiotics (ampicillin, clindamycin, ofloxacin, moxifloxacin and kanamycin). The analysis was performed on three toxigenic and three non-toxigenic C. difficile isolates using real-time PCR. To complete this work, the impact of ampicillin and clindamycin on the adherence of C. difficile to Caco-2/TC7 cells was analysed. Overall, for the six strains of C. difficile studied, exposure to subinhibitory concentrations (1/2 MIC) of clindamycin and ampicillin led to the increased expression of genes encoding colonization factors. This was correlated with the increased adherence of C. difficile to cultured cells under the same conditions. The levels of gene regulation observed among the six strains studied were highly variable, cwp84 being the most upregulated. In contrast, the expression of these genes was weakly, or not significantly, modified in the presence of ofloxacin, moxifloxacin or kanamycin. These results suggest that, in addition to the disruption of the normal intestinal microbiota and its barrier effect, the high propensity of antibiotics such as ampicillin and clindamycin to induce C. difficile infection could also be explained by their direct role in enhancing colonization by C. difficile.

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Effects of Subinhibitory Concentrations of Antibiotics on Colonization Factor Expression by Moxifloxacin-Susceptible and Moxifloxacin-Resistant Clostridium difficile Strains

Antimicrobial Agents and Chemotherapy ◽

10.1128/aac.00532-09 ◽

2009 ◽

Vol 53 (12) ◽

pp. 5155-5162 ◽

Cited By ~ 24

Author(s):

Cécile Denève ◽

Sylvie Bouttier ◽

Bruno Dupuy ◽

Frédéric Barbut ◽

Anne Collignon ◽

...

Keyword(s):

Clostridium Difficile ◽

Fluoroquinolone Resistance ◽

Reverse Transcription Pcr ◽

Colonization Factor ◽

Expression Of Genes ◽

Genes Encoding ◽

Resistant Strains ◽

Subinhibitory Concentrations ◽

The Impact

ABSTRACT Recent outbreaks of Clostridium difficile infection have been related to the emergence of the NAP1/027 epidemic strain. This strain demonstrates increased virulence and resistance to the C-8-methoxyfluoroquinolones gatifloxacin and moxifloxacin. These antibiotics have been implicated as major C. difficile infection-inducing agents. We investigated by real-time reverse transcription-PCR the impact of subinhibitory concentrations of ampicillin, clindamycin, ofloxacin, and moxifloxacin on the expression of genes encoding three colonization factors, the protease Cwp84, the high-molecular-weight S-layer protein, and the fibronectin-binding protein Fbp68. We have previously shown in six non-NAP1/027 moxifloxacin-susceptible strains that the presence of ampicillin or clindamycin induced an upregulation of these genes, whereas the presence of fluoroquinolones did not. The objective of this study was to analyze the expression of these genes under the same conditions in four NAP1/027 strains, one moxifloxacin susceptible and three moxifloxacin resistant. Two in vitro-selected moxifloxacin-resistant mutants were also analyzed. Moxifloxacin resistance was associated with the Thr82→Ile substitution in GyrA in all but one of the moxifloxacin-resistant strains. The expression of cwp84 and slpA was strongly increased after culture with ampicillin or clindamycin in NAP1/027 strains. Interestingly, after culture with fluoroquinolones, the expression of cwp84 and slpA was only increased in four moxifloxacin-resistant strains, including the NAP1/027 strains and one of the in vitro-selected mutants. The overexpression of cwp84 was correlated with increased production of the protease Cwp84. The historical NAP1/027 moxifloxacin-susceptible strain and its mutant appear to be differently regulated by fluoroquinolones. Overall, fluoroquinolones appear to favor the expression of some colonization factor-encoding genes in resistant C. difficile strains. The fluoroquinolone resistance of the NAP1/027 epidemic strains could be considered an ecological advantage. This could also increase their colonization fitness and promote the infection.

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Role of hyaluronan in regulating self-renewal and osteogenic differentiation of mesenchymal stromal cells and pre-osteoblasts

Clinical Oral Investigations ◽

10.1007/s00784-020-03259-8 ◽

2020 ◽

Vol 24 (11) ◽

pp. 3923-3937

Author(s):

Maria B. Asparuhova ◽

Vivianne Chappuis ◽

Alexandra Stähli ◽

Daniel Buser ◽

Anton Sculean

Keyword(s):

Growth Factor ◽

Osteogenic Differentiation ◽

Bone Matrix ◽

Osteogenic Potential ◽

Differentiation Markers ◽

Self Renewal ◽

Downstream Signaling ◽

Expression Of Genes ◽

Genes Encoding ◽

The Impact

Abstract Objectives The aim of the study was to investigate the impact of two hyaluronan (HA) formulations on the osteogenic potential of osteoblast precursors. Materials and methods Proliferation rates of HA-treated mesenchymal stromal ST2 and pre-osteoblastic MC3T3-E1 cells were determined by 5-bromo-20-deoxyuridine (BrdU) assay. Expression of genes encoding osteogenic differentiation markers, critical growth, and stemness factors as well as activation of downstream signaling pathways in the HA-treated cells were analyzed by quantitative reverse transcription-polymerase chain reaction (qRT-PCR) and immunoblot techniques. Results The investigated HAs strongly stimulated the growth of the osteoprogenitor lines and enhanced the expression of genes encoding bone matrix proteins. However, expression of late osteogenic differentiation markers was significantly inhibited, accompanied by decreased bone morphogenetic protein (BMP) signaling. The expression of genes encoding transforming growth factor-β1 (TGF-β1) and fibroblast growth factor-1 (FGF-1) as well as the phosphorylation of the downstream signaling molecules Smad2 and Erk1/2 were enhanced upon HA treatment. We observed significant upregulation of the transcription factor Sox2 and its direct transcription targets and critical stemness genes, Yap1 and Bmi1, in HA-treated cells. Moreover, prominent targets of the canonical Wnt signaling pathway showed reduced expression, whereas inhibitors of the pathway were considerably upregulated. We detected decrease of active β-catenin levels in HA-treated cells due to β-catenin being phosphorylated and, thus, targeted for degradation. Conclusions HA strongly induces the growth of osteoprogenitors and maintains their stemness, thus potentially regulating the balance between self-renewal and differentiation during bone regeneration following reconstructive oral surgeries. Clinical relevance Addition of HA to deficient bone or bony defects during implant or reconstructive periodontal surgeries may be a viable approach for expanding adult stem cells without losing their replicative and differentiation capabilities.

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Using muscle gene expression to estimate triacylglyceride deposition, and relative contributions of fatty acid synthesis and fatty acid import in intramuscular fat in cattle

Animal Production Science ◽

10.1071/an14247 ◽

2014 ◽

Vol 54 (9) ◽

pp. 1436 ◽

Cited By ~ 4

Author(s):

B. P. Dalrymple ◽

B. Guo ◽

G. H. Zhou ◽

W. Zhang

Keyword(s):

Gene Expression ◽

Fatty Acid ◽

De Novo ◽

Acid Synthesis ◽

Intramuscular Fat ◽

Muscle Gene Expression ◽

Expression Of Genes ◽

Genes Encoding ◽

Muscle Gene ◽

The Impact

Intramuscular fat content (IMF%) in cattle influences the value of individual animals, especially for higher marbling markets. IMF is triacylglyceride (TAG) in lipid droplets in the intramuscular adipocytes. However, there are many different pathways from feed intake to the final common process of TAG synthesis and storage as IMF. To evaluate the relative importance of different pathways we compared changes in the expression of genes encoding proteins involved in the TAG and fatty acid (FA) synthesis pathways in the longissimus muscle of Piedmontese × Hereford (P×H) and Wagyu × Hereford (W×H) crosses. Based on these changes we have estimated the relative contributions of FA synthesised de novo in the intramuscular adipocyte and the uptake of circulating FA (both free and from TAG), from the diet or synthesised de novo in other tissues, to TAG deposition as IMF. We have analysed the impact of different developmental times and different diets on these processes. Increased de novo FA synthesis in intramuscular adipocytes appeared to contribute more than increased FA uptake from circulation to the additional TAG deposition in W×H compared with P×H cattle between 12 and 25 months (forage diet). Changing diet from forage to concentrate appeared to increase the importance of FA uptake from circulation relative to de novo FA synthesis for TAG synthesis in intramuscular adipocytes. These results are consistent with the literature based on analysis of lipid composition. Gene expression appears to provide a simple assay for identification of the source of FA for the deposition of IMF.

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ANALYSIS OF THE EXPRESSION OF BIRCH STRESS TOLERANCE GENES UNDER THE INFLUENCE OF DROUGHT IN THE CENTRAL BLACK EARTH REGION

Forestry Engineering Journal ◽

10.34220/issn.2222-7962/2020.2/3 ◽

2020 ◽

Vol 10 (2) ◽

pp. 23-34

Author(s):

Tat'yana Grodeckaya ◽

Petr Evlakov ◽

Igor Isakov

Keyword(s):

Abiotic Stress ◽

Deciduous Forest ◽

Rna Isolation ◽

Phenylpropanoid Pathway ◽

Resistance Mechanisms ◽

Drought Period ◽

Cell Defense ◽

Expression Of Genes ◽

Genes Encoding ◽

The Impact

Birch is one of the main deciduous forest-forming species in the European part of Russia, but the impact of stress (in particular, drought) greatly limits its distribution. In this regard, it seems relevant to identify resistance mechanisms in order to select promising genotypes for their further reproduction. The aim of this work is to identify drought-tolerant genotypes of European birch (B. pendula Roth.), pubescent birch (B. pubescens Ehrh.) аnd their hybrids. For the study, we have taken samples of birch at the age of 26 years, which remained viable after the droughts of 2010 and 2013. Birch leaves have been selected in the third decade of June 2019 to analyze drought resistance. Leaves of birch, selected during the period with optimal conditions of air temperature and precipitation, have been used as control ones. A modified CTAB method has been proposed for RNA isolation. We studied the expression of genes encoding proteins involved in the activation of cell defense pathways under the influence of abiotic stress (pal, PR-1, PR-10, lea8, DREB2) during the drought period (June 2019). A study of the expression of genes encoding proteins of metabolic pathways that are activated in response to abiotic stress (phenylpropanoid pathway) associated with the pathogenesis of proteins (PR1 and PR10), transcription factors (DREB2), and late embryogenesis proteins (LEA) has been made. As a result of the effects of drought, a significant increase in the expression of pal, PR-1, PR-10 and DREB2 genes has been detected in the analyzed samples. At the same time, changes in lea8 gene expression were detected for two out of ten genotypes. The largest increase in expression for all five genes is shown for birch samples 29-58 and 233. It indicates the development of adaptive mechanisms in these genotypes and can characterize them as the most stable. The studied genes can be recommended as markers for the analysis of stress resistance in various species of woody plants

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Relation between Biofilm and Virulence in Vibrio tapetis: A Transcriptomic Study

Pathogens ◽

10.3390/pathogens7040092 ◽

2018 ◽

Vol 7 (4) ◽

pp. 92 ◽

Cited By ~ 4

Author(s):

Sophie Rodrigues ◽

Christine Paillard ◽

Sabine Van Dillen ◽

Ali Tahrioui ◽

Jean-Marc Berjeaud ◽

...

Keyword(s):

Pathogenic Bacteria ◽

Cell System ◽

Ruditapes Philippinarum ◽

Differentially Expressed ◽

Type Vi Secretion System ◽

Altered Expression ◽

Expression Of Genes ◽

Polysaccharide Synthesis ◽

Genes Encoding ◽

Genetic Level

Marine pathogenic bacteria are able to form biofilms on many surfaces, such as mollusc shells, and they can wait for the appropriate opportunity to induce their virulence. Vibrio tapetis can develop such biofilms on the inner surface of shells of the Ruditapes philippinarum clam, leading to the formation of a brown conchiolin deposit in the form of a ring, hence the name of the disease: Brown Ring Disease. The virulence of V. tapetis is presumed to be related to its capacity to form biofilms, but the link has never been clearly established at the physiological or genetic level. In the present study, we used RNA-seq analysis to identify biofilm- and virulence-related genes displaying altered expression in biofilms compared to the planktonic condition. A flow cell system was employed to grow biofilms to obtain both structural and transcriptomic views of the biofilms. We found that 3615 genes were differentially expressed, confirming that biofilm and planktonic lifestyles are very different. As expected, the differentially expressed genes included those involved in biofilm formation, such as motility- and polysaccharide synthesis-related genes. The data show that quorum sensing is probably mediated by the AI-2/LuxO system in V. tapetis biofilms. The expression of genes encoding the Type VI Secretion System and associated exported proteins are strongly induced, suggesting that V. tapetis activates this virulence factor when living in biofilm.

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Diverse Mechanisms Underlie Enhancement of Enteric Viruses by the Mammalian Intestinal Microbiota

Viruses ◽

10.3390/v11080760 ◽

2019 ◽

Vol 11 (8) ◽

pp. 760 ◽

Cited By ~ 5

Author(s):

Alexa N. Roth ◽

Katrina R. Grau ◽

Stephanie M. Karst

Keyword(s):

Intestinal Microbiota ◽

Pathogenic Bacteria ◽

Enteric Viruses ◽

Enteric Virus ◽

Commensal Bacteria ◽

Virus Infections ◽

Virus Binding ◽

Host Immune Responses ◽

And Function ◽

The Impact

Over the past two decades, there has been tremendous progress in understanding the impact of the intestinal microbiota on mammalian metabolism, physiology, and immune development and function. There has also been substantial advancement in elucidating the interplay between commensal and pathogenic bacteria. Relatively more recently, researchers have begun to investigate the effect of the intestinal microbiota on viral pathogenesis. Indeed, a growing body of literature has reported that commensal bacteria within the mammalian intestinal tract enhance enteric virus infections through a variety of mechanisms. Commensal bacteria or bacterial glycans can increase the stability of enteric viruses, enhance virus binding to host receptors, modulate host immune responses in a proviral manner, expand the numbers of host cell targets, and facilitate viral recombination. In this review, we will summarize the current literature exploring these effects of the intestinal microbiota on enteric virus infections.

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Cellular Mn/Zn ratio influences phosphoglucomutase activity and capsule production in Streptococcus pneumoniae D39

Journal of Bacteriology ◽

10.1128/jb.00602-20 ◽

2021 ◽

Author(s):

Averi L. McFarland ◽

Nabin Bhattarai ◽

Merrin Joseph ◽

Malcolm E. Winkler ◽

Julia E. Martin

Keyword(s):

Streptococcus Pneumoniae ◽

Transition Metal ◽

Branch Point ◽

Pathogenic Bacteria ◽

Biosynthetic Pathway ◽

Capsular Polysaccharide ◽

Direct Role ◽

Expression Levels ◽

Virulence Determinant ◽

The Impact

Capsular polysaccharide (CPS) is a major virulence determinant for human pathogenic bacteria. Although the essential functional roles for CPS in bacterial virulence have been established, knowledge of how CPS production is regulated remains limited. Streptococcus pneumoniae (pneumococcus) CPS expression levels and overall thickness change in response to available oxygen and carbohydrate. These nutrients in addition to transition metal ions can vary significantly between host environmental niches and infection stage. Since pneumococcus must modulate CPS expression among various host niches during disease progression, we examined the impact of nutritional transition metal availability of manganese (Mn) and zinc (Zn) on CPS production. We demonstrate that increased Mn/Zn ratio increase CPS production via Mn-dependent activation of the phosphoglucomutase Pgm, an enzyme that functions at the branch point between glycolysis and the CPS biosynthetic pathway in a transcription-independent manner. Further, we find that the downstream CPS protein CpsB, a Mn-dependent phosphatase, does not promote aberrant dephosporylation of its target capsule-tyrosine kinase CpsD during Mn-stress. Together, these data reveal a direct role for cellular Mn/Zn ratios in the regulation of CPS biosynthesis via direct activation of Pgm. We propose a multilayer mechanism used by the pneumococcus in regulating CPS levels across various host niches. IMPORTANCE Evolving evidence strongly indicates that maintenance of metal homeostasis is essential for establishing colonization and continued growth of bacterial pathogens in the vertebrate host. In this study, we demonstrate the impact of cellular manganese/zinc (Mn/Zn) ratios on bacterial capsular polysaccharide (CPS) production, an important virulence determinant of many human pathogenic bacteria, including Streptococcus pneumoniae. We show that higher Mn/Zn ratios increase CPS production via Mn-dependent activation of the phosphoglucomutase Pgm, an enzyme that functions at the branch point between glycolysis and the CPS biosynthetic pathway. The findings provide a direct role for Mn/Zn homeostasis in the regulation of CPS expression levels and further support the ability for metal cations to act as important cellular signaling mediators in bacteria.

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Exogenous Trehalose Induces Defenses in Wheat Before and During a Biotic Stress Caused by Powdery Mildew

Phytopathology ◽

10.1094/phyto-07-13-0191-r ◽

2014 ◽

Vol 104 (3) ◽

pp. 293-305 ◽

Cited By ~ 26

Author(s):

Christine Tayeh ◽

Béatrice Randoux ◽

Dorothée Vincent ◽

Natacha Bourdon ◽

Philippe Reignault

Keyword(s):

Gene Expression ◽

Powdery Mildew ◽

Lipid Transfer Protein ◽

Control Strategies ◽

Lipid Transfer ◽

Wheat Diseases ◽

Expression Of Genes ◽

Pathogenesis Related Protein ◽

Genes Encoding ◽

The Impact

Powdery mildew would be one of the most damaging wheat diseases without the extensive use of conventional fungicides. Some of the alternative control strategies currently emerging are based on the use of resistance inducers. The disacharride trehalose (TR) is classically described as an inducer of defenses in plants to abiotic stress. In this work, the elicitor or priming effect of TR was investigated in wheat both before and during a compatible wheat–powdery mildew interaction through molecular, biochemical, and cytological approaches. In noninoculated conditions, TR elicited the expression of genes encoding chitinase (chi, chi1, and chi4 precursor), pathogenesis-related protein 1, as well as oxalate oxidase (oxo). Moreover, lipid metabolism was shown to be altered by TR spraying via the upregulation of lipoxygenase (lox) and lipid-transfer protein (ltp)-encoding gene expression. On the other hand, the protection conferred by TR to wheat against powdery mildew is associated with the induction of two specific defense markers. Indeed, in infectious conditions following TR spraying, upregulations of chi4 precursor and lox gene expression as well as an induction of the LOX activity were observed. These results are also discussed with regard to the impact of TR on the fungal infectious process, which was shown to be stopped at the appressorial germ tube stage. Our findings strongly suggest that TR is a true inducer of wheat defense and resistance, at least toward powdery mildew.

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ARID5B regulates metabolic programming in human adaptive NK cells

Journal of Experimental Medicine ◽

10.1084/jem.20172168 ◽

2018 ◽

Vol 215 (9) ◽

pp. 2379-2395 ◽

Cited By ~ 33

Author(s):

Frank Cichocki ◽

Cheng-Ying Wu ◽

Bin Zhang ◽

Martin Felices ◽

Bianca Tesi ◽

...

Keyword(s):

Membrane Potential ◽

Nk Cells ◽

Mitochondrial Membrane Potential ◽

Mitochondrial Membrane ◽

Dna Hypomethylation ◽

Interaction Domain ◽

Direct Role ◽

Expression Of Genes ◽

Transport Chain ◽

Genes Encoding

Natural killer (NK) cells with adaptive immunological properties expand and persist in response to human cytomegalovirus. Here, we explored the metabolic processes unique to these cells. Adaptive CD3−CD56dimCD57+NKG2C+ NK cells exhibited metabolic hallmarks of lymphocyte memory, including increased oxidative mitochondrial respiration, mitochondrial membrane potential, and spare respiratory capacity. Mechanistically, we found that a short isoform of the chromatin-modifying transcriptional regulator, AT-rich interaction domain 5B (ARID5B), was selectively induced through DNA hypomethylation in adaptive NK cells. Knockdown and overexpression studies demonstrated that ARID5B played a direct role in promoting mitochondrial membrane potential, expression of genes encoding electron transport chain components, oxidative metabolism, survival, and IFN-γ production. Collectively, our data demonstrate that ARID5B is a key regulator of metabolism in human adaptive NK cells, which, if targeted, may be of therapeutic value.

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The Two TpsB-like Proteins in Anabaena sp. PCC 7120 Are Involved in Secretion of Selected Substrates.

Journal of Bacteriology ◽

10.1128/jb.00568-20 ◽

2020 ◽

Author(s):

Giang Ngo ◽

Melis Girbas ◽

Hannah Schätzle ◽

Andreas Hammer ◽

Schara Safarian ◽

...

Keyword(s):

Outer Membrane ◽

Pathogenic Bacteria ◽

Carbon Fixation ◽

Membrane Integrity ◽

Genomic Region ◽

Secretion System ◽

Secretion Systems ◽

Expression Of Genes ◽

Genes Encoding ◽

Pcc 7120

The outer membrane of Gram-negative bacteria acts as an initial diffusion barrier that shields the cell from the environment. It contains many membrane-embedded proteins required for functionality of this system. These proteins serve as solute and lipid transporters or as machines for membrane insertion or secretion of proteins. The genome of Anabaena sp. PCC 7120 codes for two outer membrane transporters termed TpsB1 and TpsB2. Those belong to the family of the two-partner secretion system proteins which are characteristic for pathogenic bacteria. Because pathogenicity of Anabaena sp. PCC 7120 has not been reported, the function of these two cyanobacterial TpsB proteins was analyzed. TpsB1 is encoded by alr1659, while TpsB2 is encoded by all5116. The latter is part of a genomic region containing 11 genes encoding TpsA-like proteins. However, tpsB2 is transcribed independently of a tpsA gene-cluster. Bioinformatics analysis revealed the presence of at least 22 genes in Anabaena sp. PCC 7120 putatively coding for substrates of the TpsB-system suggesting a rather global function of the two TpsB proteins. Insertion of a plasmid into each of the two genes, respectively, resulted in phenotypes of altered outer membrane integrity and antibiotic resistance. In addition, the expression of genes coding for the Clp and Deg proteases is dysregulated in these mutants. Moreover, for two of the putative substrates a dependence of the secretion on functional TpsB proteins could be confirmed. We confirm the existence of a two-partner secretion system in Anabaena sp. PCC 7120 and predict a large pool of putative substrates. IMPORTANCE Cyanobacteria are important organisms for the ecosystem considering their contribution to carbon fixation and oxygen production, while at the same time some species produce compounds that are toxic to their environment. As a consequence, cyanobacteria overpopulation might negatively impact the diversity of natural communities. Thus, a detailed understanding of cyanobacterial interaction with the environment including other organisms is required to define their impact on ecosystems. While two-partner secretion systems are well known from pathogenic bacteria, we provide a first description of the cyanobacterial two-partner secretion system.

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