Does Submersion in a Seawater Environment Affect the Retrieval of Useful DNA from Teeth?
Drake, Sarah Michelle
Definitive post-mortem identification of human remains currently involves identification by visual means, fingerprinting, dental identification or DNA analysis. It is generally agreed that teeth provide the most protected environment for retrieval of DNA from decomposed and/or damaged bodies. Sometimes single teeth are the only body part that has been found. Forensic research has focused on the effects of intense heat and extended post-mortem time on the ability to retrieve DNA from teeth. However, little is known about the degradation of DNA in teeth from bodies decomposing in sea water. Given that the use of dental DNA (as opposed to DNA sourced from a long bone) is an issue which is beginning to be explored for the identification of victims of mass disasters, and the high risk of marine disasters in New Zealand, I decided to study the effect of seawater on the quality of DNA extracted from teeth. Materials and Methods Thirty seven caries-free and periodontal infection-free recently extracted human molars were separated into two groups. Group I were placed into Otago harbour as individual teeth, Group II were surgically embedded into pig jawbones and subjected to the same conditions. The samples were placed into two separate locations (intertidal and fully submerged) and removed at periods from 1-26 weeks. Three human teeth were also processed as a week 0 control. Group III comprised 20 pig teeth still encased in bone the same mandibles (as used for Group II). These pig teeth were included to assess the effect of the presence of an intact periodontal ligament and the teeth were extracted at the various time points when the jaws were recovered. Once removed, the teeth were cleaned then pulverised using the SPEX 6770 freezer mill. The genomic DNA (gDNA) was then extracted via a silica-based extraction technique and used in a qPCR Taqman assay (human or pig as appropriate) for the amplification of the constitutively expressed GAPDH gene. Results The retrieval of adequate amounts of good quality DNA from control teeth confirmed the validity of the protocol utilised. Fragmentation of the human gDNA in the Group I and II teeth had occurred to such an extent that within one week of exposure to marine conditions only negligible amounts of DNA remained to provide a template for qPCR amplification. Following two weeks in seawater the DNA from Groups I and II had degraded to a level where it was no longer able to be detected with GAPDH following the qPCR amplification assay. There was no significant difference in the recovery of DNA between Groups I and II or between the submerged and intertidal locations for either Group. The pig incisor teeth (Group III) also showed a significant decrease in gDNA following one week of exposure to seawater. However, the levels of DNA available for amplification in Group III teeth were significantly higher than Groups I and II for week one and two. Negligible DNA was amplified following eight weeks in a marine environment for Group III teeth. Conclusions The time period over which useful DNA was able to be retrieved from the human teeth analysed, both isolated teeth and teeth encased in pig jaws, was considerably shorter than time periods reported in studies analysing DNA degradation in “seawater” in a laboratory-based setting. The DNA in the Group III teeth (in situ teeth with an intact periodontal ligament) also showed significant degradation following one week in a sea water setting, however within this group sufficient DNA remained after eight weeks submersion to provide a template for amplification. Based on the findings from this study, the use of DNA identification from teeth is likely to be successful if the body had been submerged for 8 weeks or less. The same results are unlikely to be achieved from a tooth found on the beach or near a body.
Advisor: Rich, Alison M; Kieser, Jules; Milne, Trudy; Duncan, Warwick; Firth, Norman
Degree Name: Doctor of Clinical Dentistry
Degree Discipline: Sir John Walsh Research Institute
Publisher: University of Otago
Keywords: Forensic; Odontology; Teeth; DNA; Seawater; Identification
Research Type: Thesis