Targeted Sequencing Toward Precision Wound Care

The Need for Improved Assessment of Combat Injury Infections: Combat trauma is, by nature, severely invasive. The characteristics of these injuries, particularly in the case of wound infection, require highly specialized treatment approaches. Wound management is primarily guided by subjective observation and is based on history with previous patients, which may not coincide with the personalized treatment needed for a particular individual. A more comprehensive and quantitative measurement of the factors impacting wound infections is urgently needed to more effectively guide treatment, reduce morbidity, shorten hospital stays, and improve rehabilitation for combat-wounded Service members.

The Promise of Modern Genomics: Rapid technical progress in the field of DNA sequencing has enabled tremendous advancements in medicine. In particular, genomics-based methods have been applied to the evaluation and exploration of groups and communities of microbes and their impact on human health and disease. Accordingly, these technologies hold great promise for benefiting combat casualty care through in-depth examination of wound infections, for instance, through detection of bacterial genes that promote antibiotic resistance. The inherent limitation of applying whole genome sequencing to microbes, however, is that the overwhelming background created by human genomic sequence in clinical samples makes confident detection of individual microbial genes nearly impossible. New approaches are therefore needed to bring the power of DNA sequencing to management of wound infections.

A Targeted Solution and Approach: An alternative approach to whole genome DNA sequencing is to target only regions of interest within a genome. By applying a pre-treatment platform that enriches a sample only for specific genes of interest, downstream sequencing becomes much more sensitive for detection of individual genes, and the resultant data focuses only on those genes of interest. We propose to develop and validate a targeted sequencing platform designed to selectively sequence and detect thousands of microbial genes across many microbial species. These genes will be pre-selected based on their relevance to wound infection and healing and on their potential for impacting patient care. This will include genes associated with clinically problematic bacterial strains, antibiotic resistance, and promotion of excessive growth and virulence. This panel of genes and the accompanying targeted sequencing platform will facilitate high-confidence detection of microbial factors from a wound infection that, by other methods, would otherwise be difficult or impossible to detect.

The first step of our approach is to identify and select microbial genes with anticipated roles in progression of wound infections. These genes will be selected from existing data from previous studies, prior experience developing similar detection panels for antibiotic resistance, and existing databases for both resistance and bacterial virulence. Once constructed, the panel will be validated for sensitivity and specificity by testing against control bacteria known to contain genetic regions targeted in the designed panel. To validate utility of the designed targeted panel for informing wound care, it will then be applied to samples from extremity wounds from Service member patients. Detection of targeted genes will be examined to determine whether they predict

healing outcomes and infection or associate with administered antibiotics.

Innovation for the Wounded Warrior: The targeted sequencing approach proposed here has never been applied to the evaluation of combat wound infection, and these data types have never been explored for their potential to inform clinical decisions. Targeted sequencing will enable analysis of microbial genes that impact infection and healing in the immediate clinical environment. This approach has the potential to change the landscape of patient care outcomes, guiding precise interventions applied on an individual patient level. Targeted sequencing technologies have a world of possible applications yet unexplored, and this effort presents an outstanding opportunity for carrying their benefit to military medicine.