Postmortem Submersion Interval (PMSI) Estimation from the Microbiome of Sus scrofa Bone in a Freshwater River

Due to inherent differences between terrestrial and aquatic systems, methods for estimating the postmortem interval (PMI) are not directly applicable to remains recovered from water. Recent studies have explored the use of microbial succession for estimating the postmortem submersion interval (PMSI); however, a non-disturbed, highly replicated and long-term aquatic decomposition study in a freshwater river has not been performed. In this study, porcine skeletal remains (N=200) were submerged in a freshwater river from November 2017-2018 (6322 accumulated degree days (ADD)/353 days) to identify changes and successional patterns in bacterial communities. One cage (e.g., 5 ribs and 5 scapulae) was collected approximately every 250 ADD for twenty-four collections; baseline samples never exposed to water acted as controls. Variable region 4 (V4) of 16S rDNA, was amplified and sequenced via the Illumina MiSeq FGx sequencing platform. Resulting sequences were analyzed using mothur (v1.39.5) and R (v3.6.0). The abundances of bacterial communities differed significantly between sample types. These differences in relative abundance were attributed to Clostridia, Holophagae and Gammaproteobacteria. Phylogenetic diversity increased with ADD for each bone type; comparably, ß-diversity bacterial community structure ordinated chronologically, which was explained with environmental parameters and inferred functional pathways. Models fit using rib samples provided a tighter prediction interval than scapulae, with a prediction of PMSI with root mean square error of within 472.31 (~27 days) and 498.47 (~29 days), respectively.