Data_Sheet_1_Postmortem submersion interval estimation of cadavers recovered from freshwater based on gut microbial community succession.docx

Microbial community succession during decomposition has been proven to be a useful tool for postmortem interval (PMI) estimation. Numerous studies have shown that the intestinal microbial community presented chronological changes after death and was stable in terrestrial corpses with different causes of death. However, the postmortem pattern of intestinal microbial community succession in cadavers retrieved from water remains unclear. For immersed corpses, the postmortem submersion interval (PMSI) is a useful indicator of PMI. To provide reliable estimates of PMSI in forensic investigations, we investigated the gut microbial community succession of corpses submersed in freshwater and explored its potential application in forensic investigation. In this study, the intestinal microbial community of mouse submersed in freshwater that died of drowning or CO2 asphyxia (i.e., postmortem submersion) were characterized by 16S rDNA amplification and high-throughput sequencing, followed by bioinformatic analyses. The results demonstrated that the chronological changes in intestinal bacterial communities were not different between the drowning and postmortem submersion groups. α-diversity decreased significantly within 14 days of decomposition in both groups, and the β-diversity bacterial community structure ordinated chronologically, inferring the functional pathway and phenotype. To estimate PMSI, a regression model was established by random forest (RF) algorithm based on the succession of postmortem microbiota. Furthermore, 15 genera, including Proteus, Enterococcus, and others, were selected as candidate biomarkers to set up a concise predicted model, which provided a prediction of PMSI [MAE (± SE) = 0.818 (± 0.165) d]. Overall, our present study provides evidence that intestinal microbial community succession would be a valuable marker to estimate the PMSI of corpses submerged in an aquatic habitat.</p

Age-dependent seroprevalence of EV71 and CA16 antibody.

<p>Age-related seroprevalence of neutralizing antibodies to EV71 (A) and CA16 (B) in individuals in Guangdong, China, before and after the 2010 HFMD epidemic. Before: before the 2010 HFMD epidemic; after: after the 2010 HFMD epidemic.</p

Overall geometric mean titer (GMT) of EV71 and CA16 antibody.

<p>Before: before the 2010 HFMD epidemic; after: after the 2010 HFMD epidemic. The lines indicate 95% confidence interval.</p

Overall seroprevalence of EV71 and CA16 antibody.

<p>Overall seroprevalence of neutralizing antibodies to EV71 (A) and CA16 (B) in individuals in Guangdong, China, before and after the 2010 HFMD epidemic. Before: before the 2010 HFMD epidemic; after: after the 2010 HFMD epidemic. The lines indicate 95% confidence interval.</p

Phylogenetic relationship among the 562 EV71 C4 strains isolated in mainland China from 1998 to 2010.

<p>257 China strains are from the present study, and 305 China strains are from GenBank. The bootstrap values of 1000 replicates for major lineages are displayed as numbers at the nodes. Strains isolated in Shenzhen from 1998 to 2004 were marked by the sombol •.</p

EV71 reference strains used in this study.

*<p>The GenBank numbers of the 203 reference strains are AY895129-AY895145, EU753363 - EU753418, FJ765416 - FJ765435, GQ121417 - GQ121441, GQ253391 - GQ253423, GQ487666 - GQ487689 and GU353079 - GU353106. All of these strains are part of the 305 GenBank EV71 C4 reference strains collected in Mainland China from 1998 to 2010.</p

Demographic profile of the subjects.

<p>No: number; before: before the 2010 HFMD epidemic; after: after the 2010 HFMD epidemic.</p

Maximum Clade Credibility tree and Bayesian Skyline of the China EV71 C4 strains.

<p>The upper portion of the figure was maximum clade credibility tree with 95% highest posterior density (HPD) intervals for the node times (in years) and posterior probabilities for branching events. Tree was generated by the MCMC method in BEAST on the basis of a multiple alignment of VP1 nucleotide sequences of EV71 C4 strains collected in Guangdong Province from 2008 to 2010 and Genbank EV71 C4 reference strains isolated in mainland China from 1998 to 2010. Below the tree, the Bayesian Skyline with 95% HPD intervals shows the relative measure for genetic diversity through time (values plotted on y-axis).</p

Alignment of deduced amino acid sequences of the VP1 fragment of representative EV71 strains collected in mainland China from 1998 to 2010.

<p>The amino acid substitutions are marked by using small panes.</p