Mapping the Burkholderia cenocepacia niche response via high-throughput sequencing

Abstract

Determining how an organism responds to its environment by altering gene expression is key to understanding its ecology. Here, we used RNA-seq to comprehensively and quantitatively assess the transcriptional response of the bacterial opportunistic cystic fibrosis (CF) pathogen and endemic soil dweller, Burkholderia cenocepacia, in conditions mimicking these 2 environments. By sequencing 762 million bases of cDNA from 2 closely related B. cenocepacia strains (one isolated from a CF patient and one from soil), we identified a number of potential virulence factors expressed under CF-like conditions, whereas genes whose protein products are involved in nitrogen scavenging and 2-component sensing were among those induced under soil-like conditions. Interestingly, 13 new putative noncoding RNAs were discovered using this technique, 12 of which are preferentially induced in the soil environment, suggesting that ncRNAs play an important role in survival in the soil. In addition, we detected a surprisingly large number of regulatory differences between the 2 strains, which may represent specific adaptations to the niches from which each strain was isolated, despite their high degree of DNA sequence similarity. Compared with the CF strain, the soil strain shows a stronger global gene expression response to its environment, which is consistent with the need for a more dynamic reaction to the heterogeneous conditions of soil.cystic fibrosis ͉ RNA-seq ͉ soil ͉ transcriptomics

RNA-seq reveals cooperative metabolic interactions between two termite-gut spirochete species in co-culture

“…Gene expression values were determined by normalizing the number of reads mapped to a particular gene (excluding ambiguous regions) divided by the size of the gene (also excluding ambiguous regions). The resulting value is the normalized reads per kilobase, in a manner consistent with the gene expression index calculations of previously published reports (Yoder-Himes et al, 2009). In order to adjust for intensity between samples, the ribosomal signal from each sample was used as a standard, and each sample's intensity was multiplied by a factor that would yield an equal ribosomal RNA (rRNA) signal.…”

“…Breakthroughs in next-generation sequencing not only allow the comparison of expression from highly similar sequences, but also dramatically expedite the pace at which data can be gathered from less-well-studied microbes (Holt et al, 2008;Yoder-Himes et al, 2009) and complex biological environments (Gilbert et al, 2008;Urich et al, 2008). Although transcriptomic calculations are now possible, studies of co-cultures and intact environments are challenging, as they entail inherent difficulties when making comparisons and contrasts.…”

Dual RNA-seq of pathogen and host

“…The first studies in prokaryotes reported the sequencing of rRNA-depleted samples to ~1-7 M reads/library (Oliver et al, 2009;Perkins et al, 2009;Yoder-Himes et al, 2009). This was sufficient to infer differential mRNA expression.…”

“…Many of the pioneering studies were performed using pathogenic bacteria, including S. Typhi (Perkins et al, 2009), Bacillus anthracis (Passalacqua et al, 2009), Burkholderia cenocepacia (Yoder-Himes et al, 2009), Helicobacter pylori or Chlamydia trachomatis (Albrecht et al, 2010), and identified a plethora of novel genes including many new sRNAs (for reviews see (Croucher and Thomson, 2010;Guell et al, 2011;Sorek and Cossart, 2010). The original RNA-seq protocol has since been further modified to increase the informational output: For instance, the usage of a specific exonuclease in a method referred to as differential RNA-seq (dRNA-seq) allowed the selective degradation of processed RNA molecules leaving only primary transcripts, thereby enabling the identification of TSSs on a genome-wide scale .…”

Analysis of genomic characters reveals that four distinct gene clusters are correlated with different functions in Burkholderia cenocepacia AU 1054

“…In a recent study, the gene expression pattern of B. cenocepacia was determined under two conditions, CFlike (cystic fibrosis like) condition patients and soil-like conditions (Yoder-Himes et al 2009). Compared to the strains under soil-like conditions, 124 genes were determined to be expressed more highly under CF-like conditions.…”