Erratum to: ‘Consensus pan-genome assembly of the specialised wine bacterium Oenococcus oeni’

<b>Background</b>\ud \ud <i>Oenococcus oeni</i> is a lactic acid bacterium that is specialised for growth in the ecological niche of wine, where it is noted for its ability to perform the secondary, malolactic fermentation that is often required for many types of wine. Expanding the understanding of strain-dependent genetic variations in its small and streamlined genome is important for realising its full potential in industrial fermentation processes.\ud \ud <b>Results</b>\ud \ud Whole genome comparison was performed on 191 strains of <i>O. oeni</i>; from this rich source of genomic information consensus pan-genome assemblies of the invariant (core) and variable (flexible) regions of this organism were established. Genetic variation in amino acid biosynthesis and sugar transport and utilisation was found to be common between strains. Furthermore, we characterised previously-unreported intra-specific genetic variations in the natural competence of this microbe.\ud \ud <b>Conclusion</b>\ud \ud By assembling a consensus pan-genome from a large number of strains, this study provides a tool for researchers to readily compare protein-coding genes across strains and infer functional relationships between genes in conserved syntenic regions. This establishes a foundation for further genetic, and thus phenotypic, research of this industrially-important species

Deep sequencing reveals divergent expression patterns within the small RNA transcriptomes of cultured and vegetative tissues of sugarcane

Deep sequencing has advanced the discovery and analysis of the small RNA component of transcriptomes and has revealed developmentally-regulated populations of small RNAs consistent with key roles in plant development. To study small RNA transcriptome complexity and explore their roles in sugarcane development, we obtained almost 50 million small RNA reads from suspension cells, embryogenic calli, leaf, apex and a developmental series of stem internodes. The complexity of the small RNA component of the transcriptome varied between tissues. The undifferentiated and young tissue type libraries had lower redundancy levels than libraries generated from maturing and mature tissues. The ratio of 21:24 nt small RNAs also varied widely between different tissue types, as did the proportion of abundant small RNAs derived from each putative origin of small RNA biogenesis. Cluster analysis indicates many abundant small RNAs display developmental expression patterns. There was substantial variation in isomiR composition, abundance and expression patterns within sugarcane microRNA (miRNA) families. Two hundred and fifty-six isomiRs from 36 miRNA families were identified by homology to known miRNA families from a range of plant species. Many isomiRs and miRNA families appear to be developmentally regulated, including a subset of miRNAs that are progressively upregulated during stem internode maturation. Transcribed sequences putatively targeted by abundant sugarcane small RNAs were predicted and miRNA directed cleavage of 18 predicted sugarcane targets were validated by 5′ RACE

Genome Sequences of Three Species of Hanseniaspora Isolated from Spontaneous Wine Fermentations

Members of the genus Hanseniaspora represent a significant proportion of the normal flora of grape berries and play a significant role in wine fermentation. Here, we present genome sequences for three species of Hanseniaspora, H. opuntiae, H. osmophila, and H. uvarum, which were isolated from spontaneous Chardonnay wine fermentation

Incorporating target sequences of developmentally regulated small RNAs into transgenes to enhance tissue specificity of expression in plants

Many applications of plant genetic engineering require tight control of transgene expression to a particular organ, tissue or developmental stage, particularly when off-target expression causes deleterious effects. Most studies have relied on tissue-specific and developmentally regulated promoters to confer the desired transgene expression pattern via transcriptional control. However, the process to identify promoters conferring required expression patterns is slow and expensive, with no certainty of success. Therefore, we investigated the practicality of post-transcriptional control of transgene expression patterns in plants, triggered by binding of endogenous small RNA sequences. Target sequences of several small RNAs, inserted 3′ of a luciferase reporter, altered transgene expression in patterns consistent with developmental abundance of the corresponding small RNAs in sugarcane. Incorporating the target sequences of miR167 or a leaf-specific small RNA selectively reduced reporter expression in leaves by more than 90 % on average, whereas incorporating the miR160-3p target sequence selectively reduced reporter expression in roots by 75 % on average across immature transgenic lines. We anticipate that the strategy should be broadly applicable in plants

Understanding and avoiding transgene silencing

This chapter contains sections titled:\ud \ud - Incidence and Practical Significance of Transgene Silencing\ud \ud - Factors Influencing Transgene Silencing\ud \ud - Box 12.1. An RNAi primer\ud \ud - Mechanisms of Transgene Silencing\ud \ud - Strategies to Avoid Transgene Silencing\ud \ud - Conclusions and Future Prospects\ud \ud - Box 12.2. Avoiding transgene silencing

Design rules for efficient transgene expression in plants

Sustained expression of transgenes in specified developmental patterns is commonly needed in plant biotechnology, but obstructed by transgene silencing. Here, we present a set of gene design rules, tested on the silencing-susceptible beetle luc and bacterial ims genes, expressed in sugarcane. Designs tested independently or in combination included removal of rare codons, removal of RNA instability sequences, blocking of likely endogenous sRNA binding sites and randomization of non-rare codons. Stable transgene expression analyses, on multiple independent lines per construct, showed greatest improvement from the removal of RNA instability sequences, accompanied by greatly reduced transcript degradation evident in northern blot analysis. We provide a set of motifs that readily can be eliminated concurrently with rare codons and undesired structural features such as repeat sequences, using Gene Designer 2.0 software. These design rules yielded 935- and 5-fold increased expression in transgenic callus, relative to the native luc and ims sequences; and gave sustained expression under the control of sugarcane and heterologous promoters over several years in greenhouse and field trials. The rules can be applied easily with codon usage tables from any plant species, providing a simple and effective means to achieve sustained expression of otherwise silencing-prone transgenes in plants

HLA-A alleles including HLA-A29 affect the composition of the gut microbiome:a potential clue to the pathogenesis of birdshot retinochoroidopathy

Birdshot retinochoroidopathy occurs exclusively in individuals who are HLA-A29 positive. The mechanism to account for this association is unknown. The gut microbiome has been causally implicated in many immune-mediated diseases. We hypothesized that HLA-A29 would affect the composition of the gut microbiome, leading to a dysbiosis and immune-mediated eye disease. Fecal and intestinal biopsy samples were obtained from 107 healthy individuals from Portland, Oregon environs, 10 of whom were HLA-A29 positive, undergoing routine colonoscopy. Bacterial profiling was achieved via 16S rRNA metabarcoding. Publicly available whole meta-genome sequencing data from the Human Microbiome Project (HMP), consisting of 298 healthy controls mostly of US origin, were also interrogated. PERMANOVA and sparse partial least squares discriminant analysis (sPLSDA) demonstrated that subjects who were HLA-A29 positive differed in bacterial species composition (beta diversity) compared to HLA-A29 negative subjects in both the Portland (p = 0.019) and HMP cohorts (p = 0.0002). The Portland and HMP cohorts evidenced different subsets of bacterial species associated with HLA-A29 status, likely due to differences in the metagenomic techniques employed. The functional composition of the HMP cohort did not differ overall (p = 0.14) between HLA-A29 positive and negative subjects, although some distinct pathways such as heparan sulfate biosynthesis showed differences. As we and others have shown for various HLA alleles, the HLA allotype impacts the composition of the microbiome. We hypothesize that HLA-A29 may predispose chorioretinitis via an altered gut microbiome.</p

HLA Alleles Associated With Risk of Ankylosing Spondylitis and Rheumatoid Arthritis Influence the Gut Microbiome

Objective HLA alleles affect susceptibility to more than 100 diseases, but the mechanisms that account for these genotype–disease associations are largely unknown. HLA alleles strongly influence predisposition to ankylosing spondylitis (AS) and rheumatoid arthritis (RA). Both AS and RA patients have discrete intestinal and fecal microbiome signatures. Whether these changes are the cause or consequence of the diseases themselves is unclear. To distinguish these possibilities, we examined the effect of HLA–B27 and HLA–DRB1 RA risk alleles on the composition of the intestinal microbiome in healthy individuals. Methods Five hundred sixty‐eight stool and biopsy samples from 6 intestinal sites were collected from 107 healthy unrelated subjects, and stool samples were collected from 696 twin pairs from the TwinsUK cohort. Microbiome profiling was performed using sequencing of the 16S ribosomal RNA bacterial marker gene. All subjects were genotyped using the Illumina CoreExome SNP microarray, and HLA genotypes were imputed from these data. Results Associations were observed between the overall microbial composition and both the HLA–B27 genotype and the HLA–DRB1 RA risk allele (P = 0.0002 and P = 0.00001, respectively). These associations were replicated using the stool samples from the TwinsUK cohort (P = 0.023 and P = 0.033, respectively). Conclusion This study shows that the changes in intestinal microbiome composition seen in AS and RA are at least partially due to effects of HLA‐B27 and HLA–DRB1 on the gut microbiome. These findings support the hypothesis that HLA alleles operate to cause or increase the risk of these diseases through interaction with the intestinal microbiome and suggest that therapies targeting the microbiome may be effective in preventing or treating these diseases