J Indian Acad Forensic Med. Jan- March 2012, Vol. 34, No. 1 ISSN 0971-0973

Review Research Paper Application of Genetics and Molecular Biology In Forensic Odontology

* Rohit Malik, ** Deepankar Misra, ***PC Srivastava, **** Akansha Misra

Abstract Various methods of Forensic Science have been applied to solve many mysteries of world including criminal investigation. Forensic odontology researches are usually associated to the dentist‘s field of action by using bite marks, dental records and radiographs plays an important role in solving many crimes. These methods help in age determination and sex identification of the people who have lost their identity after death. Nevertheless, since the development of genetics and molecular biology there were an increase in number and quality of solved cases. The present article emphasizes the importance to associate certain forensic biology areas to traditional investigation methods in human identification, especially with forensic odontology. It also shows that in some situations, teeth are an important source to genetic analysis and molecular studies. Finally it was concluded that knowledge of forensic genetics applied to traditional forensic investigations would produce an increase of information to Justice.

Key Words: Forensic Odontology, Genetics, Human Identification

Introduction: This human identification methodology Forensic odontology researches are relies upon three important steps: DNA isolation usually consists of analysis, expertise and or extraction, amplification of specific DNA evaluation of dentistry related episodes, but it regions using PCR technique (Polymerase can also be extended to other areas if it is Chain Reaction) and molecule profiles analysis. necessary for justice and administration [2] Human individuality identification tests using interests. The most common studied area where DNA are based upon polymorphism of several a dentist acts as a forensic expert is related to mini and micro satellite loci. This polymorphism oral maxillo-facial trauma caused by different in autosomal chromosomes is considered a sources such as mechanical, physical and great individuality indicator. First, in order to do chemical and dental analysis used for human the amplification phase highly specific probes identification. [1] were developed which allow simultaneous Development of genetics in the 80's scanning in several loci of the sample. After the allowed innovations to medical field as well to material amplification using PCR technique, they forensic sciences. This was due to discovery of are analysed in agarose or polyacrylamide gel or specific mini satellite regions of human genome in automatic sequencer. Depending on the which had the so called DNA (deoxyribonucleic quality and quantity of the obtained sample, acid) "digital impressions". Its analysis led to analysis can be done from nuclear DNA or human individuality information. From that point, mitochondrial (mt DNA), both show advantages genetics research dealing with DNA and limitations. [3] polymorphism has had a great development Computer aid in several laboratory steps associated to biotechnology and forensic mathematics increase the reliability of Corresponding Author: examinations to determine sex, age estimation, **Senior Lecturer parenthood and human identification. Therefore Department of Oral Medicine and Radiology Institute of Dental Studies and Technology it became mandatory that forensic experts in Modinagar, UP, India several areas of criminal investigation, forensic E-mail: [email protected] medicine and dentistry would associate classical *Professor investigation techniques to molecular biology ***Assoc. Prof., Dept. of Forensic Medicine, analysis and DNA examination in order to Rohilkhand Medical College & Hospital, Bareilly, UP, achieve more reliable, objective and specific ****PG Student, results facing complex cases. Dept. of Oral & Maxillofacial Pathology Expertise examination must show a multi disciplinary approach; therefore the

55

J Indian Acad Forensic Med. Jan- March 2012, Vol. 34, No. 1 ISSN 0971-0973 present paper presents a scientific literature choice of materials, but as time goes by those review aiming to point out the importance of sources become inappropriate, leading towards forensic genetics used along with traditional the mineralized tissue sources like bones and investigation methods in human identification, teeth. Bones are an important source to obtain mainly with forensic odontology. It also intends genetic material since they are located inside to show the tooth forensic relevance as a source the body and are mineralized; also the cortical of material to genetic and molecular studies. part protects the medullary part from external Molecular Biology Studies: factors and micro-organism that may degrade Nowadays, Forensic Anthropology has the DNA. [7] molecular biology as a powerful allied, mainly in Teeth as Genetic Material Source: species, family background, sex and age Teeth also are a good source to obtain investigation. Since that, determination of genetic material. This is true mainly because of species using bone, tooth, hair or organic fluid their great tissue resistance (enamel, dentin, samples was done searching for exclusive cementum and pulp) against external injuries. [8] human molecular indicators. Those indicators Pulp tissue is a loose connective tissue and it are polymorphic and easily detectable in the degrades easily when compared to others dental population. They can refer to a gene, a restricted tissues. Dental pulp is protected by tooth site or any other DNA sequence which presents structures and therefore can present better different allelic version to that locus. The condition than others soft tissue for DNA mechanisms that explain polymorphism can be extraction. Lessing et al [9] showed that pulp single changes in nucleotides (substitution) such can be a source of DNA in teeth that had been as SNP (Single Nucleotide Polymorphism), kept or obtained in different conditions. VNTR (variable number of tandem repeats) and Amelogenin can also be studied from STR (short tandem repeats) and the insertion or dental material that had been through adverse exclusion of DNA sequences (Indels), such as situations. There are several techniques to Alu insertions. It is important to add that some obtain dental material to extract DNA such as indicators are specific to a determined tooth grinding or crushing, tooth horizontal population, this happens because the difference sectioning, pulp extirpation by tooth irrigation of frequency between two big populations is and sectioning and nitrogen liquid cryogenic more than 50% (such as Europeans and pulverization. [10] Africans). Those indicators are named ancestor Some legal precautions must be indicators (AIMs). [4] followed when dealing with dental material as In order to increase age estimative source to obtain DNA since the process precision of decomposed bodies such as bones destroys the material used in the examination. several researches have been developed trying Those precautions involve proper teeth to relate mainly the racemic mixture of aspartic identification, describing all characteristics and if acid (D/L) from tooth tissues with one‘s it is possible, taking pictures and radiographs in chronological age. It is well known that aspartic the original positions when they are removed acid, particularly in its dextrogen form (D), from dental arch. Those precautions are done to linearly increases along aging. [5] Other preserve dental characteristics of evidences in biochemical indicators such as gelatinase A and order to prove their genuine value when doubts glutamic acid from dentine are also being are raised concerning their origin or the results researched to be used in age estimation. [6] of the person identity. Human Identification Using DNA: After the dental recording and filing Genetic material can be obtained from phase and before the handling of samples, it is several biological sources such as body fluids, necessary to decontaminate dental surface, soft tissues and mineralized tissues. When Sodium Hypochlorite is substance most used, dealing with living suspects in parenthood test but it is necessary to have an optimization (father, mother and son), genetic material is among concentration, time and applying method preferably obtained from either blood (leukocyte) of this substance. [11] or buccal mucosa. Multi Disciplinary Approach in On the other hand, if it is necessary to Human Identification: verify genetic relation involving pot-mortem In Forensic dentistry, DNA examination material, time from death and corpse condition can be used together with traditional techniques are relevant factors to choose which method of showing great results to identify destroyed or DNA extraction is to be used. In a recent death advanced decomposed stage corpses. [12] case blood, viscera and soft tissue are the first Besides that, DNA analysis obtained from oral

56

J Indian Acad Forensic Med. Jan- March 2012, Vol. 34, No. 1 ISSN 0971-0973 mucosa cells has great importance in dental charred with fuel. DNA was obtained from dental impressions, also known as bite marks. pulp extracted from intra osseous third molar. Normally, the primary investigation approach of Facing an overwhelming result this type of evidence is related to the analysis of presented in the forensic literature and from the dental characteristics left in the victim or in the even more reliable techniques, DNA object. Nevertheless, when those marks do not examinations and Molecular Biology analysis give a conclusive result, biological material became an essential tool to help or solve collection from the place where bite was applied investigation matters that had been considered is extremely important to find the identity of the irresolvable in crime investigation and Forensic person who was responsible for the mark. medicine. Therefore it is mandatory that those Among the techniques to obtain DNA from who are in forensic investigations must acquire human skin there is the double swab technique knowledge about forensic genetics to provide which consists of applying a swab with distilled greater justice and benefit the society. water followed by another dry swab on the bite References: mark. This technique showed good results to 1. Smith BC. Introduction to DNA analysis. Dent Clin North Am. 2001; obtain biological material to be researched. [13] 45: 229-35. In the Borgula et al [14] experimental 2. Edwards A, Civitello A, Hammond HA, Caskey CT. DNA Typing and Genetic Mapping with Trimeric and Tetrameric Tandem Repeats. research was demonstrated that is possible to Am J Hum Genet. 1991; 49: 746-56. analyse the genotype of specific bacteria found 3. Shriver MD, Mei R, Parra EJ, Sonpar V, Halder J, Tishkoff AS, et al. in the oral cavity (Streptococcus) of individuals Large-scale SNP analysis reveals clustered and continuous as an alternative when it is not possible to obtain patterns of human genetic variation. Hum Genomics. 2005; 2: 81-9. 4. Slavkin HC. Sex, enamel and forensic dentistry: a search for the DNA of the one who caused the bite mark. identity. J Am Dent Assoc. 1997; 128: 1021-5. In more complex cases such as 5. Othani S. Estimation of age from dentin by utilizing the racemization presented by Bilge et al [15] a multi field of aspartic acid: influence of pH. Forensic Sci Int. 1995; 75: 181-7. approach investigation was necessary to identify 6. Sajdok J, Pilin A, Pudil F, Zídková J, Kás J. A new method of age estimation based on the changes in human non-collagenous a corpse. Anthropologic techniques, forensic proteins from dentin. Forensic Sci Int. 2006; 156: 245-9. dentistry, computer superposition (face/cranium) 7. Iwamura ESM, Soares-Vieira JA, Muñoz DR. Human identification and DNA investigation were used. Sex was and analysis of DNA in bones. Rev Hosp Clin Fac Med Sao Paulo. determined by cranium characteristics and as 2004; 59: 383-8. 8. Pretty IA, Sweet D. A look at forensic dentistry. Part 1: the role of well by amelogenin analysis. Age was estimated teeth in the determination of human identity. Br Dent J. 2001; 190: using longitudinal divided crown measurements 359-66. and computer superposition showed a positive 9. Lessig R, Edelmann J. Individualisation of dental tissue - an aid for identification between victim facial structures odontological identification? J Forensic Odontostomatol. 1995; 13: 1-3. and the found head. 10. Sweet D, Hildebrand D. Recovery of DNA from human teeth by DNA was extracted from dental pulp, cryogenic grinding. J Forensic Sci. 1998; 43: 1199-1202. bones, muscle tissues and compared to genetic 11. Kemp BM, Smith DG. Use of bleach to eliminate contaminating profile of the victim's presumed daughter and DNA from the surface of bones and teeths. Forensic Sci Int. 2005; 154: 53-61. wife. Fatherhood indication was verified in 11 12. Sweet D, Lorente M, Lorente JA, Valenzuela A, Villanueva E. An examined loci. In another case, Sweet et al [16] improved method to recover saliva from human skin: the double presented an identification of human parts from swab technique. J Forensic Sci. 1997; 42: 320-2. a woman that had been disappeared for 3 years. 13. Atsü SS, Gökdemir K, Kedici PS, Ikyaz YY. Bite marks in forensic odontology. J Forensic Odontostomatol. 1998; 16: 30-4. Investigations showed that the presumed victim 14. Borgula LM, Robinson FG, Rahimi M, Chew KE, Birchmeier KR, had 3 smear cell laminas in the laboratory files. Owens SG, et al. Isolation and genotypic comparison of oral DNA was extracted and compared to genetic streptococci from experimental bite marks. J Forensic Odontostomatol. 2003; 21: 23-30. profile obtained from the dental sample of the 15. Bilge Y, Kedici PS, Alakoç YD, Ûlküer KÛ, Ilkyaz YY. The found corpse. The result was positive, showing identification of a dismembered human body: a multidisciplinary coincidence in 8 of the 8 examined loci, approach. Forensic Sci Int. 2003; 137: 141-6. including amelogenin. 16. Sweet D, Hidel brand D, Phillips D. Identification of a skeleton using DNA from teeth and PAP smear. J Forensic Sci. 1999; 44: 630-3. In the identification of charred bodies, 17. Soares-Vieira JA, Billerbeck AEC, Iwamura ESM, Cardoso LA, the great resistance of mineralized tissues Muñoz DR. Post-mortem forensic identity testing: application of allows the victims to be identified not only by PCR to the identification of fire victim. Sao Paulo Med J. 2000; 118: DNA extracted from bone material [17], but also 75-7. from dental material. Sweet et al [18] showed an 18. Sweet D, Sweet CHW. DNA analysis of dental pulp to link incinerated remains of homicide victim to crime scene. J Forensic identification of a homicide victim that had been Sci. 1995; 40: 310-4.

57