An ultra-deep sequencing strategy to detect sub-clonal TP53 mutations in presentation chronic lymphocytic leukemia cases using multiple polymerases

Chronic lymphocytic leukaemia (CLL) is the most common clonal B-cell disorder characterized by clonal diversity, a relapsing and remitting course, and in its aggressive forms remains largely incurable. Current front-line regimes include agents such as fludarabine, which act primarily via the DNA damage response pathway. Key to this is the transcription factor p53. Mutations in the TP53 gene, altering p53 functionality, are associated with genetic instability, and are present in aggressive CLL. Furthermore, the emergence of clonal TP53 mutations in relapsed CLL, refractory to DNA-damaging therapy, suggests that accurate detection of sub-clonal TP53 mutations prior to and during treatment may be indicative of early relapse. In this study, we describe a novel deep sequencing workflow using multiple polymerases to generate sequencing libraries (MuPol-Seq), facilitating accurate detection of TP53 mutations at a frequency as low as 0.3%, in presentation CLL cases tested. As these mutations were mostly clustered within the regions of TP53 encoding DNA-binding domains, essential for DNA contact and structural architecture, they are likely to be of prognostic relevance in disease progression. The workflow described here has the potential to be implemented routinely to identify rare mutations across a range of diseases

Volatile Organic Compounds Emitted by Fungal Associates of Conifer Bark Beetles and their Potential in Bark Beetle Control

Conifer bark beetles attack and kill mature spruce and pine trees, especially during hot and dry conditions. These beetles are closely associated with ophiostomatoid fungi of the Ascomycetes, including the genera Ophiostoma, Grosmannia, and Endoconidiophora, which enhance beetle success by improving nutrition and modifying their substrate, but also have negative impacts on beetles by attracting predators and parasites. A survey of the literature and our own data revealed that ophiostomatoid fungi emit a variety of volatile organic compounds under laboratory conditions including fusel alcohols, terpenoids, aromatic compounds, and aliphatic alcohols. Many of these compounds already have been shown to elicit behavioral responses from bark beetles, functioning as attractants or repellents, often as synergists to compounds currently used in bark beetle control. Thus, these compounds could serve as valuable new agents for bark beetle management. However, bark beetle associations with fungi are very complex. Beetle behavior varies with the species of fungus, the stage of the beetle life cycle, the host tree quality, and probably with changes in the emission rate of fungal volatiles. Additional research on bark beetles and their symbiotic associates is necessary before the basic significance of ophiostomatoid fungal volatiles can be understood and their applied potential realized