DNA Analysis and the Postmortem Submersion Interval from the Microbiome of Waterlogged Skeletal Remains

The extraction of DNA from waterlogged, and thus compromised bone, has not been studied previously under natural conditions. Forensic laboratories also often struggle to determine what bone type to sample and may be following less than optimal protocols to amplify DNA and generate usable profiles from skeletons recovered from water, which are vital in identification. Another method of identifying remains is using the postmortem submersion interval (PMSI), or time of submersion after death until discovery. The aims of this investigation are to: (1) examine the effects of freshwater on the quality and quantity of DNA retrievable from waterlogged skeletal remains; and (2) to determine which microbes are involved in the aquatic decomposition of skeletal remains and whether their succession is related to the postmortem submersion interval. The study will determine which extraction method, Organic Phenol-Chloroform or solid-phase (ChargeSwitch gDNA Plant Kit), and bone type (rib or scapula) is best for the isolation of DNA from skeletal remains found in water. In addition, the variance of quality and quantity of DNA in different bone types and different bodies of water (e.g. freshwater lake and freshwater river) will also be identified. Mixed effects modelling in R will be used to compare method efficacy and bone type. The study also aims to determine which microbes are involved in the aquatic decomposition of skeletal remains and how microbial succession and indicator species may be used to establish a PMSI estimation method for skeletal remains. Data analysis will follow the Mothur SOP; data will be archived in Forensic Microbiome Database (FMD) and the Sequence Read Archive (SRA). This proposal addresses both (1) best practices for obtaining DNA from waterlogged bone in order to identify missing persons, and (2) the estimation of the postmortem submersion interval from the changing microbiome colonizing waterlogged bone over time, which may indicate specific missing persons who match the time frame. Products expected are interim and final reports, PhD written dissertation and project presentation, at least three conference presentations and three published peer-reviewed papers. The impact will be disseminated to forensic laboratories as suggested protocols for use of preferred bone and method for DNA extraction in waterlogged samples. "Note: This project contains a research and/or development component, as defined in applicable law," and complies with Part 200 Uniform Requirements - 2 CFR 200.210(a)(14). ca/ncf