To Compare the Effects of a Standard Versus Hydrophilic Polymer Coated Airway Stent in a Porcine Model: A Randomized, Single-Blinded Study.

BACKGROUND: Stent encrustation with debris and mucostasis is a significant cause of airway injury and comorbidity, leading to ~25% of stent exchanges (1-3). Previous work from our group has shown that the experimental coating can reduce mucous adhesion in bench testing and demonstrated a signal for reducing airway injury and mucostasis in a feasibility study. OBJECTIVES: The aim of this study is to continue our inquiry in a randomized, single-blinded multi-animal trial to investigate the degree of airway injury and mucostasis using silicone stents with and without this specialized coating. METHODS: We modified commercially available silicone stents with a hydrophilic polymer from Toray Industries. We conducted an in vivo survival study in 6 mainstem airways (3 coated and 3 uncoated) of 3 pigs to compare the degree of airway injury and mucostasis between coated versus noncoated stented airways. Both stents were randomized to either left or right mainstem bronchus. The pathologist was blinded to the stent type. RESULTS: We implanted a total of six 14×15 mm silicone stents (1 per mainstem bronchi) into 3 pigs. All animals survived to termination at 4 weeks. All stents were intact; however, 1 uncoated stent migrated out. On average, all the coated stents demonstrated reduced pathology and tissue injury scores (75 vs. 68.3, respectively). The average total dried mucous weight was slightly higher in the coated stents (0.07 g vs. 0.05 g; respectively). CONCLUSION: Coated stents had lower airway injury compared with uncoated stents in this study. Of all the stents, 1 uncoated stent migrated out and was not included in the dried mucous weight totals. This could explain the slightly higher mucous weight in the coated stents. Nevertheless, this current study demonstrates promising results in lowering airway injury in stents incorporated with the hydrophilic coating, and future studies, including a larger number of subjects, would be needed to corroborate our findings.

An IS-mediated, RecA-dependent, bet-hedging strategy in Burkholderia thailandensis.

Adaptation to fluctuating environmental conditions is difficult to achieve. Phase variation mechanisms can overcome this difficulty by altering genomic architecture in a subset of individuals, creating a phenotypically heterogeneous population with subpopulations optimized to persist when conditions change, or are encountered, suddenly. We have identified a phase variation system in Burkholderia thailandensis that generates a genotypically and phenotypically heterogeneous population. Genetic analyses revealed that RecA-mediated homologous recombination between a pair of insertion sequence (IS) 2-like elements duplicates a 208.6 kb region of DNA that contains 157 coding sequences. RecA-mediated homologous recombination also resolves merodiploids, and hence copy number of the region is varied and dynamic within populations. We showed that the presence of two or more copies of the region is advantageous for growth in a biofilm, and a single copy is advantageous during planktonic growth. While IS elements are well known to contribute to evolution through gene inactivation, polar effects on downstream genes, and altering genomic architecture, we believe that this system represents a rare example of IS element-mediated evolution in which the IS elements provide homologous sequences for amplification of a chromosomal region that provides a selective advantage under specific growth conditions, thereby expanding the lifestyle repertoire of the species.

Bacterial populations are often diverse, even when originating from a single cell. This diversity helps microbes survive in fluctuating environmental conditions by increasing the odds of population survival. For example, if environmental conditions change such that only a subpopulation with unique abilities survives, the entire population will be saved. Genomes are naturally dynamic. For example, mobile sections of DNA, called transposable elements, can change their position within a genome. If a transposable element jumps into a gene, it can harm the cell. But if it moves into a different site, it may provide an organism with new features that can help it survive. Most organisms contain multiple copies of transposable elements in their DNA. For example, a subtype of the soil bacterium Burkholderia thailandensis, strain E264, has two identical transposable elements that book-end a region of DNA that contains 157 genes. Lowrey et al. studied this bacterial strain in different environmental conditions to find out more. The experiments revealed that in growing populations of E264, some bacteria had one copy of the region, while others had two or three. In a rich environment, most bacteria had just one copy of the region. However, when grown in challenging conditions, most bacteria contained two or three copies of the region. Moreover, bacteria required at least two copies to form dense communities known as biofilms, which are advantageous for bacterial survival in challenging conditions. Bacteria with only one copy, however, were better adapted to a free-swimming lifestyle. Lowrey et al. further showed that the DNA repair system was required for duplicating the region. Usually, this system finds and recombines identical DNA sequences to repair broken DNA. However, if two identical DNA sequences (a pair of transposable elements) are present, the repair system can recombine them during DNA replication, resulting in the duplication of the DNA between the identical sequences. The same system also reduces the copy number of the region from three or two to just one. Since the repair system is constantly working and DNA recombination is always occurring at a low level, B. thailandensis E264 maintains a genetically diverse population with bacteria containing different copy numbers of the region. This diversity ensures that the strain survives in fluctuating environmental conditions. Transposable elements are hotspots of evolution. They are known to interrupt genes and shrink genomes. Lowrey et al. showed that transposable elements also influence evolution by providing DNA sequences that the DNA repair system can use to duplicate DNA. This process of duplicating genes is more frequent than random genetic mutations, expediting adaptation.

A putative enoyl-CoA hydratase contributes to biofilm formation and the antibiotic tolerance of Achromobacter xylosoxidans.

Achromobacter xylosoxidans has attracted increasing attention as an emerging pathogen in patients with cystic fibrosis. Intrinsic resistance to several classes of antimicrobials and the ability to form robust biofilms in vivo contribute to the clinical manifestations of persistent A. xylosoxidans infection. Still, much of A. xylosoxidans biofilm formation remains uncharacterized due to the scarcity of existing genetic tools. Here we demonstrate a promising genetic system for use in A. xylosoxidans; generating a transposon mutant library which was then used to identify genes involved in biofilm development in vitro. We further described the effects of one of the genes found in the mutagenesis screen, encoding a putative enoyl-CoA hydratase, on biofilm structure and tolerance to antimicrobials. Through additional analysis, we find that a fatty acid signaling compound is essential to A. xylosoxidans biofilm ultrastructure and maintenance. This work describes methods for the genetic manipulation of A. xylosoxidans and demonstrated their use to improve our understanding of A. xylosoxidans pathophysiology.