Leonardo Albarracin

Transcriptomic Analysis of the Innate Antiviral Immune Response in Porcine Intestinal Epithelial Cells: Influence of Immunobiotic Lactobacilli

Lactobacillus rhamnosus CRL1505 and Lactobacillus plantarum CRL1506 are immunobiotic strains able to increase protection against viral intestinal infections as demonstrated in animal models and humans. To gain insight into the host–immunobiotic interaction, the transcriptomic response of porcine intestinal epithelial (PIE) cells to the challenge with viral molecular associated pattern poly(I:C) and the changes in the transcriptomic profile induced by the immunobiotics strains CRL1505 and CRL1506 were investigated in this work. By using microarray technology and reverse transcription PCR, we obtained a global overview of the immune genes involved in the innate antiviral immune response in PIE cells. Stimulation of PIE cells with poly(I:C) significantly increased the expression of IFN-α and IFN-β, several interferon-stimulated genes, cytokines, chemokines, adhesion molecules, and genes involved in prostaglandin biosynthesis. It was also determined that lactobacilli differently modulated immune gene expression in poly(I:C)-challenged PIE cells. Most notable changes were found in antiviral factors (IFN-α, IFN-β, NPLR3, OAS1, OASL, MX2, and RNASEL) and cytokines/chemokines (IL-1β, IL-6, CCL4, CCL5, and CXCL10) that were significantly increased in lactobacilli-treated PIE cells. Immunobiotics reduced the expression of IL-15 and RAE1 genes that mediate poly(I:C) inflammatory damage. In addition, lactobacilli treatments increased the expression PLA2G4A, PTGES, and PTGS2 that are involved in prostaglandin E2 biosynthesis. L. rhamnosus CRL1505 and L. plantarum CRL1506 showed quantitative and qualitative differences in their capacities to modulate the innate antiviral immune response in PIE cells, which would explain the higher capacity of the CRL1505 strain when compared to CRL1506 to protect against viral infection and inflammatory damage in vivo. These results provided valuable information for the deeper understanding of the host–immunobiotic interaction and their effect on antiviral immunity. The comprehensive transcriptomic analyses successfully identified a group of genes (IFN-β, RIG1, RNASEL, MX2, A20, IL27, CXCL5, CCL4, PTGES, and PTGER4), which can be used as prospective biomarkers for the screening of new antiviral immunobiotics in PIE cells and for the development of novel functional food and feeds, which may help to prevent viral infections.

Exopolysaccharides from Lactobacillus delbrueckii OLL1073R-1 modulate innate antiviral immune response in porcine intestinal epithelial cells

HighlightsEPSs from L. delbrueckii OLL1073R‐1 (LDR‐1) modulate TLR3‐mediated immune response in PIE cells.EPS from LDR‐1 improves type I interferon production and interferon‐induced antiviral genes.EPS from LDR‐1 differentially regulates the expression of inflammatory cytokines in response to TLR3 activation.TLR2 and negative regulators of TLR pathway are involved in the immunomodulatory activity of EPS from LDR‐1. Abstract Previous studies demonstrated that the extracellular polysaccharides (EPSs) produced by Lactobacillus delbrueckii OLL1073R‐1 (LDR‐1) improve antiviral immunity, especially in the systemic and respiratory compartments. However, it was not studied before whether those EPSs are able to beneficially modulate intestinal antiviral immunity. In addition, LDR‐1‐host interaction has been evaluated mainly with immune cells while its interaction with intestinal epithelial cells (IECs) was not addressed before. In this work, we investigated the capacity of EPSs from LDR‐1 to modulate the response of porcine IECs (PIE cells) to the stimulation with the Toll‐like receptor (TLR)‐3 agonist poly(I:C) and the role of TLR2, TLR4, and TLR negative regulators in the immunoregulatory effect. We showed that innate immune response triggered by TLR3 activation in porcine IECs was differentially modulated by EPS from LDR‐1. EPSs treatment induced an increment in the expression of interferon (IFN)‐&agr; and IFN‐&bgr; in PIE cells after the stimulation with poly(I:C) as well as the expression of the antiviral factors MxA and RNase L. Those effects were related to the reduced expression of A20 in EPS‐treated PIE cells. EPS from LDR‐1 was also able to reduce the expression of IL‐6 and proinflammatory chemokines. Although further in vivo studies are needed, our results suggest that these EPSs or a yogurt fermented with LDR‐1 have potential to improve intestinal innate antiviral response and protect against intestinal viruses.

Development of an in vitro immunobiotic evaluation system against rotavirus infection in bovine intestinal epitheliocytes

The bovine intestinal epithelial cell line (BIE cells) expresses the Toll-like receptor (TLR)3 and is able to mount an antiviral immune response after the stimulation with poly(I:C). In the present study, we aimed to further characterise the antiviral defence mechanisms in BIE cells by evaluating the innate immune response triggered by rotavirus (RV) infection. In addition, we attempted to determine whether immunobiotic bifidobacteria are able to confer protection of BIE cells against RV infection by beneficially modulating the antiviral immune response. RV OSU (porcine) and UK (bovine) effectively infected BIE cells, while a significant lower capacity to infect BIE cells was observed for human (Wa) and murine (EW) RV. We observed that viral infection in BIE cells triggered TLR3/RIG-I-mediated immune responses with activation of IRF3 and TRAF3, induction of interferon beta (IFN-β) and up-regulation of inflammatory cytokines. Our results also demonstrated that preventive treatments with Bifidobacterium infantis MCC12 or Bifidobacterium breve MCC1274 significantly reduced RV titres in infected BIE cells and differentially modulated the innate immune response. Of note, both strains significantly improved the production of the antiviral factor IFN-β in RV-infected BIE cells. In conclusion, this work provides comprehensive information on the antiviral immune response of BIE cells against RV, that can be further studied for the development of strategies aimed to improve antiviral defences in bovine intestinal epithelial cells. Our results also demonstrate that BIE cells could be used as a newly immunobiotic evaluation system against RV infection for application in the bovine host.

Modulation of porcine intestinal epitheliocytes immunetranscriptome response by Lactobacillus jensenii TL2937.

In order to evaluate probiotic strains applicable for the beneficial immunomodulation of the porcine gut (immunobiotics), we previously developed a porcine intestinal epitheliocyte cell line (PIE cells). Here, transcriptomic studies using PIE cells were performed considering that this information would be valuable for understanding the mechanisms involved in the protective activity of the immunobiotic strain Lactobacillus jensenii TL2937 against intestinal inflammatory damage in pigs. In addition, those studies would provide criteria for selecting biomarkers for the screening of new immunobiotic strains. We performed microarray analysis to investigate the transcriptomic response of PIE cells to the challenge with heat-stable enterotoxigenic Escherichia coli (ETEC) pathogen-associated molecular patterns (PAMPs) and, the changes induced by L. jensenii TL2937 in that response. The approach allowed us to obtain a global overview of the immune genes involved in the response of PIE cells to heat-stable ETEC PAMPs. We observed that L. jensenii TL2937 differently modulated gene expression in ETEC PAMPs-challenged PIE cells. Microarray and RT-PCR analysis indicated that the most remarkable changes in PIE cells transcriptomic profile after heat-stable ETEC PAMPs challenge were observed in chemokines, adhesion molecules, complement and coagulation cascades factors. In addition, an anti-inflammatory effect triggered by TL2937 strain in PIE cells was clearly demonstrated. The decrease in the expression of chemokines (CCL8, CXCL5, CXCL9, CXCL10, and CXCL11), complement (C1R, C1S, C3, and CFB), and coagulation factors (F3) by L. jensenii TL2937 supports our previous reports on the immunoregulatory effect of this strain. These results provided clues for the better understanding of the mechanism underlying host-immunobiotic interaction in the porcine host. The comprehensive transcriptomic profiles of PIE cells provided by our analyses successfully identified a group of genes, which could be used as prospective biomarkers for the screening and evaluation of new anti-inflammatory immunobiotics for the prevention of inflammatory intestinal disorders in pigs.

Draft Genome Sequence of a Probiotic Strain, Lactobacillus fermentum UCO-979C

ABSTRACT This report describes a draft genome sequence of Lactobacillus fermentum strain UCO-979C. The reads generated by a Ion Torrent PGM were assembled into contigs, with a total size of 2.01 Mb. The data were annotated using the NCBI GenBank and RAST servers. Specific features of the genome are highlighted.

Draft Genome Sequence of Lactobacillus plantarum MPL16, a Wakame-Utilizing Immunobiotic Strain Isolated from Swine Feces

ABSTRACT The genome of the immunomodulatory Lactobacillus plantarum MPL16, a strain able to ferment wakame (Undaria pinnatifida), is described here. The reads were assembled into contigs with a total size 3,278,495 bp. The genome information will be useful for further specific genetic studies of this strain that evaluate its immunomodulatory and biotechnological properties.

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 the Immunobiotic Strain Lactobacillus jensenii TL2937

ABSTRACT The genome of the immunomodulatory strain Lactobacillus jensenii TL2937 is described here. The draft genome has a total length of 1,678,416 bp, a G+C content of 34.3%, and 1,470 predicted protein-coding sequences. The genome information will be useful for gaining insight into the immunomodulatory properties of the TL2937 strain in the porcine host.

Draft Genome Sequence of Lactobacillus plantarum CRL1506, an Immunomodulatory Strain Isolated from Goat Milk

ABSTRACT This report describes a draft genome sequence of Lactobacillus plantarum CRL1506, a probiotic strain with immunomodulatory properties isolated from goat milk. The reads generated by a whole-genome shotgun (WGS) strategy on an Illumina MiSeq sequencer were assembled into contigs with a total size of 3,228,096 bp. The draft genome sequence of L. plantarum CRL1506 will be useful for further studies of specific genetic features of this strain and for understanding the mechanisms of its immunobiotic properties.

Draft Genome Sequence of Lactobacillus plantarum TL2766, a Strain with the Ability To Ferment Wakame

ABSTRACT The genome sequence of Lactobacillus plantarum TL2766, a strain with the ability to ferment wakame (Undaria pinnatifida), is described here. The reads were assembled into contigs, with a total size of 3,310,195 bp. The genome information will be useful for further specific genetic studies of this strain and for its biotechnological applications.