Genomic Characterization of Lactobacillus delbrueckii TUA4408L and Evaluation of the Antiviral Activities of its Extracellular Polysaccharides in Porcine Intestinal Epithelial Cells

In lactic acid bacteria, the synthesis of exopolysaccharides (EPS) has been associated with some favorable technological properties as well as health-promoting benefits. Research works have shown the potential of EPS produced by lactobacilli to differentially modulate immune responses. However, most studies were performed in immune cells and few works have concentrated in the immunomodulatory activities of EPS in non-immune cells such as intestinal epithelial cells. In addition, the cellular and molecular mechanisms involved in the immunoregulatory effects of EPS have not been studied in detail. In this work, we have performed a genomic characterization of Lactobacillus delbrueckii subsp. delbrueckii TUA4408L and evaluated the immunomodulatory and antiviral properties of its acidic (APS) and neutral (NPS) EPS in porcine intestinal epithelial (PIE) cells. Whole genome sequencing allowed the analysis of the general features of L. delbrueckii TUA4408L genome as well as the characterization of its EPS genes. A typical EPS gene cluster was found in the TUA4408L genome consisting in five highly conserved genes epsA-E, and a variable region, which includes the genes for the polymerase wzy, the flippase wzx, and seven glycosyltransferases. In addition, we demonstrated here for the first time that L. delbrueckii TUA4408L and its EPS are able to improve the resistance of PIE cells against rotavirus infection by reducing viral replication and regulating inflammatory response. Moreover, studies in PIE cells demonstrated that the TUA4408L strain and its EPS differentially modulate the antiviral innate immune response triggered by the activation of Toll-like receptor 3 (TLR3). L. delbrueckii TUA4408L and its EPS are capable of increasing the activation of interferon regulatory factor (IRF)-3 and nuclear factor κB (NF-κB) signaling pathways leading to an improved expression of the antiviral factors interferon (IFN)-β, Myxovirus resistance gene A (MxA) and RNaseL

Draft genome sequence of probiotic lactobacillus brevis TUCO-5E, isolated from porcine milk

This report describes the draft genome sequence of Lactobacillus brevisTUCO-5E, a probiotic strain isolated from porcine maternal milk. The reads were generated by a whole-genome sequencing (WGS) strategy on an Illumina MiSeq sequencer and were assembled into contigs with a total estimated size of 2,461,089 bp. A total of 2,455 open reading frames (ORFs) were predicted, including 2,301 protein-coding sequences. The draft genome sequence of L. brevis TUCO-5E will be useful for further studies of specific genetic features and for understanding the mechanisms of its probiotic properties in the porcine host.Fil: Quilodrán-Vega, Sandra Rayén. Universidad de Concepción. Facultad de Medicina Veterinaria. Departamento de Medicina Preventiva y Patología; ChileFil: Albarracín, Leonardo Miguel. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Tucumán. Centro de Referencia para Lactobacilos; ArgentinaFil: Hebert, Elvira Maria. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Tucumán. Centro de Referencia para Lactobacilos; ArgentinaFil: Saavedra, Maria Lucila. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Tucumán. Centro de Referencia para Lactobacilos; ArgentinaFil: Fonseca, Alexis. Universidad de Concepción. Departmento de Farmacología; ChileFil: Salas-Burgos, Alexis. Universidad de Concepción. Departmento de Farmacología; ChileFil: Kitazawa, Haruki. Tohoku University. Graduate School of Agricultural Science. Food and Feed Immunology Group; JapónFil: Villena, Julio Cesar. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Tucumán. Centro de Referencia para Lactobacilos; Argentina. Universidad Nacional de Tucumán; Argentin

Phylogenetic tree based on 16S rRNA gene sequences.

<p>Blue rectangle depicts the clade for “<i>Ca</i>. V. ishoeyi”. In bold is shown the position for “<i>Ca</i>. V. ishoeyi” into the clade, and the numbers next to the nodes indicates the <i>a posteriori</i> probability in percentage of 0 to 1. The tree was calculated using Bayesian approach with MrBayes tool.</p

Main genomic features among some representative sulfur-oxidizing bacteria and the draft genome of “<i>Ca</i>. V. ishoeyi”.

<p>Main genomic features among some representative sulfur-oxidizing bacteria and the draft genome of “<i>Ca</i>. V. ishoeyi”.</p

Biosynthetic gene clusters, identified as terpene and undetermined type in “<i>Ca</i>. V. ishoeyi”.

<p><b>A. Terpene gene cluster.</b> Sphere indicates the Phytoene synthase (phythoen_synt) domain. <b>B</b>. <b>Undetermined biosynthetic gene cluster.</b> Spheres show the Acyltransferase (AT), involved in biosynthesis of Polyketide synthase (PKS) compounds, Aminotransferase (Amino), involved in biosynthesis of Polyketide synthase/nonribosomal peptide synthetases (PKS/NRPS) compounds, Adenylation (A) and Carrier Protein (CP; Thiolation) domains, from undetermined compounds (Other). Asterisks indicate domains used as a rule for gene cluster classification. Identified genes are indicated in the backbone of the gene clusters and the product list is available in the <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0188371#pone.0188371.s007" target="_blank">S5A and S5B Table</a>.</p

Circular representation of the draft genome of “<i>Ca</i>. V. ishoeyi”.

<p>Moving out from the innermost circle: the black circle describes the GC% content in a window = 1000 bp; the green circle represents the CDSs of the antisense DNA strand, and the red circle the CDSs of the sense DNA strand; the outermost circle shows all scaffolds, each one alternating between blue and black.</p

Genomic features of <i>“Candidatus</i> Venteria ishoeyi”, a new sulfur-oxidizing macrobacterium from the Humboldt Sulfuretum off Chile

<div><p>The Humboldt Sulfuretum (HS), in the productive Humboldt Eastern Boundary Current Upwelling Ecosystem, extends under the hypoxic waters of the Peru-Chile Undercurrent (<i>ca</i>. 6°S and <i>ca</i>. 36°S). Studies show that primeval sulfuretums held diverse prokaryotic life, and, while rare today, still sustain species-rich giant sulfur-oxidizing bacterial communities. We here present the genomic features of a new bacteria of the HS, “<i>Candidatus</i> Venteria ishoeyi” (“<i>Ca</i>. V. ishoeyi”) in the family <i>Thiotrichaceae</i>.Three identical filaments were micro-manipulated from reduced sediments collected off central Chile; their DNA was extracted, amplified, and sequenced by a Roche 454 GS FLX platform. Using three sequenced libraries and through <i>de novo</i> genome assembly, a draft genome of 5.7 Mbp, 495 scaffolds, and a N50 of 70 kbp, was obtained. The 16S rRNA gene phylogenetic analysis showed that “<i>Ca</i>. V. ishoeyi” is related to non-vacuolate forms presently known as <i>Beggiatoa</i> or <i>Beggiatoa</i>-like forms. The complete set of genes involved in respiratory nitrate-reduction to dinitrogen was identified in “<i>Ca</i>. V. ishoeyi”; including genes likely leading to ammonification. As expected, the sulfur-oxidation pathway reported for other sulfur-oxidizing bacteria were deduced and also, key inorganic and organic carbon acquisition related genes were identified. Unexpectedly, the genome of “<i>Ca</i>. V. ishoeyi” contained numerous CRISPR repeats and an I-F CRISPR-Cas type system gene coding array. Findings further show that, as a member of an eons-old marine ecosystem, “<i>Ca</i>. V. ishoeyi” contains the needed metabolic plasticity for life in an increasingly oxygenated and variable ocean.</p></div

Schematic representation of the dissimilatory sulfite reductase (DSR) system, identified in “<i>Ca</i> V. ishoeyi”.

<p>In yellow: <i>dsrAB</i> genes coding for sulfite reductases: in pink: <i>dsrEFH</i> genes, coding for heterohexameric protein; in black: the <i>dsrC</i> gene, coding for sulfurtransferase; in red: <i>dsrMKJOP</i> genes, coding for transmembrane electron-transporting complex; and, in blue: the <i>dsrL</i> gene, coding for intracellular sulfur oxidation protein, are shown. F1, F2, and F3 represent the frame arrangement in the sense strand DNA.</p