Diagnosis of drowning using 16S ribosomal RNA gene of phytoplankton

In forensic medicine, the diagnosis of a corpse immersed in water that is differentiation between death from drowning or dead on entering the water, is made mainly using the diatom test by acid digestion. However, the diatom test by acid digesting is technically complicated and requires a larger quantity of sample. Moreover, the procedure is hazardous due to the use of strong acid and is time consuming. Alternative methods for the diagnosis of drowning by detecting plankton genes living in water using molecular biological technique have been reported. By performing one PCR amplification using various organs of a corpse immersed in water, this method is anticipated to permit rapid and accurate diagnosis of death from drowning by identifying the genes of phytoplankton that had entered solid organs via blood circulation. However, the currently used PCR method for the detection of phytoplankton requires DNA purification, necessitating treatment of tissues as well as differentiation between sea water and fresh water phytoplankton. Thus this method has the disadvantages of being labor intensive and having the risk of contamination. To overcome these issues, first we targeted synechococcus sp (Bacteria; Cyanobacteria; Oscillatoriophycideae; Chroococcales), a picoplankton found in both sea water and fresh water, and designed primers specific for its 16S ribosomal RNA genes (16S rDNA). We succeeded to find that phytoplankton DNA used the quantity of the organization from the organ of the drowned body using a direct PCR method without DNA extraction to check the death from being drowned from autopsy findings and a diatom test by acid digestion and an autopsy case to be diagnosed. Furthermore, we quantitatively assayed the 16S rDNA gene of picoplankton from tissues of drowned rabbits and non-drowned rabbits immersed in water after death. The research was to investigate the quantities of picoplankton DNA in lung, liver, kidney tissues and blood in drowned and non-drowned rabbits, and the sensitivity of detection of picoplankton DNA by direct PCR for the diagnosis of death from drowning. The results verified that the detection of phytoplankton DNA in the liver and kidney is the most important evidence for the diagnosis of death from drowning.