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RNA-Seq Provides Clues to Bacterial Niche Specialization

NEW YORK (GenomeWeb News) – In a paper scheduled to appear online this week in the Proceedings of the National Academy of Sciences, an international research team used RNA sequencing to uncover gene expression and regulatory changes associated with bacterial adaptation to two distinct niches.

Burkholderia cenocepacia can cause health problems for individuals with cystic fibrosis and other immunocompromised patients. To better understand the niche adaptation of this bug, researchers compared two B. cenocepacia strains isolated from cystic fibrosis patients and soil samples. As it turned out, growth conditions had a dramatic influence on the expression of genes and non-coding RNAs.

But, the team reported, gene expression and other differences were also quite common between strains even when grown under the same conditions. Together, the results may start to explain how B. cenocepacia survives in and exploits different ecological niches.

"We have uncovered a surprising number of strain-specific responses to each environment, which may aid in the identification of therapeutic targets for new vaccines or antibiotics, which is important medically because this organism is resistant to a broad spectrum of antibiotics," senior author Rotem Sorek, a molecular geneticist at Israel's Weizmann Institute of Science, and his co-authors wrote.

B. cenocepacia is found in at least two very different environments: the soil, where it sometimes colonizes the area surrounding plant roots, and the human respiratory tract, where it has been linked to necrotizing pneumonia and even death in cystic fibrosis patients and others.

In an effort to understand how B. cenocepacia adapts to each niche, Sorek and his colleagues used RNA-seq to compare the gene expression and regulatory pathways in AU1054, a strain isolated from the sputum of a cystic fibrosis patient, and HI2424, a strain isolated from agricultural soil in New York State. Each strain was tested several times and grown in both cystic fibrosis and soil extract media.

After growing the strains in media and temperatures mimicking cystic fibrosis and soil conditions, the team isolated RNA, enriched for messenger RNA, and generated cDNA libraries. Researchers at the US Department of Energy's Joint Genome Institute then sequenced the cDNA on an Illumina Genome Analyzer.

After tossing out ribosomal RNA reads and reads that didn't meet the team's established criteria, the researchers were left with 83,940,138 base pairs of cDNA from mRNA transcripts.

In general, the team reported that HI2424 had more highly expressed genes than AU1054, though they found that differential gene expression in both strains depended on the growth conditions used.

For instance, growth under cystic fibrosis-like conditions increased the expression of core housekeeping genes on chromosome 1. In contrast, strains grown in soil-like conditions often induced genes on chromosomes 2 and 3.

The researchers also found that growing B. cenocepacia under the cystic fibrosis-like conditions induced virulence factors, whereas the soil-mimicking conditions upped the expression of genes involved in nitrogen scavenging and environmental sensing.

In addition, the researchers identified what appear to be 13 new non-coding RNAs. A dozen of these are preferentially expressed in the soil conditions, fueling speculation that non-coding RNAs may aid B. cenocepacia's soil survival.

Finally, when the team directly compared the two strains under the same conditions, they found that 179 genes were differentially expressed in sputum-like conditions while 120 genes were differentially expressed in soil-like conditions.

The results provide insights into the differences between the strains as well as the adaptive techniques used by B. cenocepacia in general. And, the researchers noted, by honing in on AU1054 genes that are specifically expressed under cystic fibrosis conditions it may be possible to find key virulence factors.

"Although further investigation is required to tease apart the precise roles of the identified differentially regulated B. cenocepacia genes and [non-coding RNAs] in either niche, this study establishes RNA-seq as a powerful method for the quantitative and qualitative examination of bacterial transcriptomes," the authors concluded.

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