The thanatomicrobiome (thanatos, Greek for death) is a relatively new term and is the study of the microbes colonizing the internal organs and orifices after death. Recent scientific breakthroughs in an initial study of the thanatomicrobiome have revealed that a majority of the microbes within the human body are the obligate anaerobes, Clostridium spp., in the internal postmortem microbial communities. We hypothesized that time-dependent changes in the thanatomicrobiome within internal organs can estimate the time of death as a human body decays. Here we report a cross-sectional study of the sampling of 27 human corpses from criminal cases with postmortem intervals between 3.5–240 hours. The impetus for examining microbial communities in different internal organs is to address the paucity of empirical data on thanatomicrobiomic succession caused by the limited access to these organs prior to death and a dearth of knowledge regarding the movement of microbes within remains. Our sequencing results of 16S rRNA gene amplicons of 27 postmortem samples from cadavers demonstrated statistically significant time-, organ-, and sex-dependent changes. These results suggest that comprehensive knowledge of the number and abundance of each organ’s signature microorganisms could be useful to forensic microbiologists as a new source of data for estimating postmortem interval.

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Death is a universal phenomenon; however, is there “life after death?” This topic has been investigated for centuries but still there are gray areas that have yet to be elucidated. Forensic microbiologists are developing new applications to investigate the dynamic and coordinated changes in microbial activity that occur when a human host dies. There is currently a paucity of explorations of the thanatomicrobiome (thanatos-, Greek for death) and epinecrotic communities (microbial communities residing in and/or moving on the surface of decomposing remains). Ongoing studies can help clarify the structure and function of these postmortem microbiomes. Human microbiome studies have revealed that 75–90% of cells in the body prior to death are microbial. Upon death, putrefaction occurs and is a complicated process encompassing chemical degradation and autolysis of cells. Decomposition also involves the release of contents of the intestines due to enzymes under the effects of abiotic and biotic factors. These factors likely have predictable effects on postmortem microbial communities and can be leveraged for forensic studies. This mini review provides a critical examination of emerging research relating to thanatomicrobiome and epinecrotic communities, how each is studied, and possible strategies of stochastic processes.

Human thanatomicrobiome studies have established that an abundant number of putrefactive bacteria within internal organs of decaying bodies are obligate anaerobes, Clostridium spp. These microorganisms have been implicated as etiological agents in potentially life-threatening infections; notwithstanding, the scale and trajectory of these microbes after death have not been elucidated. We performed phylogenetic surveys of thanatomicrobiome signatures of cadavers’ internal organs to compare the microbial diversity between the 16S rRNA gene V4 hypervariable region and V3-4 conjoined regions from livers and spleens of 45 cadavers undergoing forensic microbiological studies. Phylogenetic analyses of 16S rRNA gene sequences revealed that the V4 region had a significantly higher mean Chao1 richness within the total microbiome data. Permutational multivariate analysis of variance statistical tests, based on unweighted UniFrac distances, demonstrated that taxa compositions were significantly different between V4 and V3-4 hypervariable regions (p < 0.001). Of note, we present the first study, using the largest cohort of criminal cases to date, that two hypervariable regions show discriminatory power for human postmortem microbial diversity. In conclusion, here we propose the impact of hypervariable region selection for the 16S rRNA gene in differentiating thanatomicrobiomic profiles to provide empirical data to explain a unique concept, the Postmortem Clostridium Effect.

Most forensic research that is used to better understand how to estimate the postmortem interval (PMI) entails the study of the physiochemical characteristics of decomposition and the effects that environmental factors have on the decomposition process. Forensic entomology exploits the life cycles of arthropods like Diptera (blow flies or flesh flies) and Coleoptera (beetles) deposited on the decaying carcass to determine PMI. Forensic taphonomy, from the Greek word taphos meaning burial, studies the creation of the fossils of decomposed cadavers to ascertain information as to the nature and time of death. Compared to other areas of taphonomy, there have been relatively few forensic science studies that have investigated the impact of human decomposition on the microbial changes occurring on or in a corpse or in the soil communities underneath a body. Such research may facilitate the critical determination of PMI. Therefore, the scope of this review is to provide a concise summary of the current progress in the newly emerging field of microbial diversity and the next-generation metagenomic sequencing approaches for assessing these communities in humans and in the soil beneath decomposing human.

Genomic studies have estimated there are approximately 10(3)-10(6) bacterial species per gram of soil. The microbial species found in soil associated with decomposing human remains (gravesoil) have been investigated and recognized as potential molecular determinants for estimates of time since death. The nascent era of high-throughput amplicon sequencing of the conserved 16S ribosomal RNA (rRNA) gene region of gravesoil microbes is allowing research to expand beyond more subjective empirical methods used in forensic microbiology. The goal of the present study was to evaluate microbial communities and identify taxonomic signatures associated with the gravesoil human cadavers. Using 16S rRNA gene amplicon-based sequencing, soil microbial communities were surveyed from 18 cadavers placed on the surface or buried that were allowed to decompose over a range of decomposition time periods (3-303 days). Surface soil microbial communities showed a decreasing trend in taxon richness, diversity, and evenness over decomposition, while buried cadaver-soil microbial communities demonstrated increasing taxon richness, consistent diversity, and decreasing evenness. The results show that ubiquitous Proteobacteria was confirmed as the most abundant phylum in all gravesoil samples. Surface cadaver-soil communities demonstrated a decrease in Acidobacteria and an increase in Firmicutes relative abundance over decomposition, while buried soil communities were consistent in their community composition throughout decomposition. Better understanding of microbial community structure and its shifts over time may be important for advancing general knowledge of decomposition soil ecology and its potential use during forensic investigations.

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