Isolation, Characterization, and Comparative Genomics of the Novel Potential Probiotics from Canine Feces.

Lactic acid bacteria (LAB) are commonly used as probiotics; however, not all LAB strains have the same beneficial effects. To successfully use LAB as probiotics in canines, LAB species should originate from the canine intestinal tract as they display host specificity. The objective of this study was to investigate the phenotypic and genomic traits of potential probiotic LAB isolated from canine fecal samples. Twenty LAB samples were evaluated for their potential probiotic characteristics including resistance to low pH, bile salts, hydrophobicity, auto-aggregation, co-aggregation, adhesion to epithelia or mucosa, and production of inhibitory compounds. Additionally, we evaluated their safety and other beneficial effects on canine health, such as DPPH free radical scavenging, and ß-galactosidase. Four strains demonstrated potential probiotic characteristics and were selected: Enterococcus hirae Pom4, Limosilactobacillus fermentum Pom5, Pediococcus pentosaceus Chi8, and Ligilactobacillus animalis FB2. Safety evaluations showed that all strains lacked hemolytic activity, could not produce biogenic amines, and did not carry any pathogenic genes. In addition, L. fermentum Pom5 and P. pentosaceus Chi8 displayed susceptibility to all antibiotics and concordant with the absence of antibiotic resistance genes. Based on their phenotypic and genomic characteristics, L. fermentum Pom5 and P. pentosaceus Chi8 were identified as potential probiotic candidates for canines.

Host-dependent roles of the viral 5' untranslated region (UTR) in RNA stabilization and cap-independent translational enhancement mediated by the 3' UTR of Red clover necrotic mosaic virus RNA1.

The genome of Red clover necrotic mosaic virus (RCNMV) consists of RNA1 and RNA2, both lacking a cap structure and a poly(A)tail. RNA1 has a translational enhancer element (3'TE-DR1) in the 3' untranslated region (UTR). In this study, we analyzed the roles of 5' and 3' UTRs of RNA1 in 3'TE-DR1-mediated cap-independent translation in cowpea and tobacco BY-2 protoplasts using a dual-luciferase (Luc) reporter assay system. Most mutations introduced into RNA1 5' UTR in reporter Luc mRNA abolished or greatly reduced cap-independent translation in BY-2 protoplasts, whereas those mutations had no or much milder effects if any on translational activity in cowpea protoplasts. Our results suggest that a stem-loop structure predicted in the 5' proximal region of RNA1 plays important roles in both translation and RNA stability. We also show that 3'TE-DR1-mediated cap-independent translation relies on a ribosome-scanning mechanism in both protoplasts.

A viral noncoding RNA generated by cis-element-mediated protection against 5'->3' RNA decay represses both cap-independent and cap-dependent translation.

Positive-strand RNA viruses use diverse mechanisms to regulate viral and host gene expression for ensuring their efficient proliferation or persistence in the host. We found that a small viral noncoding RNA (0.4 kb), named SR1f, accumulated in Red clover necrotic mosaic virus (RCNMV)-infected plants and protoplasts and was packaged into virions. The genome of RCNMV consists of two positive-strand RNAs, RNA1 and RNA2. SR1f was generated from the 3' untranslated region (UTR) of RNA1, which contains RNA elements essential for both cap-independent translation and negative-strand RNA synthesis. A 58-nucleotide sequence in the 3' UTR of RNA1 (Seq1f58) was necessary and sufficient for the generation of SR1f. SR1f was neither a subgenomic RNA nor a defective RNA replicon but a stable degradation product generated by Seq1f58-mediated protection against 5'-->3' decay. SR1f efficiently suppressed both cap-independent and cap-dependent translation both in vitro and in vivo. SR1f trans inhibited negative-strand RNA synthesis of RCNMV genomic RNAs via repression of replicase protein production but not via competition of replicase proteins in vitro. RCNMV seems to use cellular enzymes to generate SR1f that might play a regulatory role in RCNMV infection. Our results also suggest that Seq1f58 is an RNA element that protects the 3'-side RNA sequences against 5'-->3' decay in plant cells as reported for the poly(G) tract and stable stem-loop structure in Saccharomyces cerevisiae.