Soong Deok Lee

Gymnophalloides seoi Eggs from the Stool of a 17th Century Female Mummy Found in Hadong, Republic of Korea

It was previously reported that paleoparasitological clues for parasites infecting humans could be found in the feces of mummies of the Joseon Dynasty (1392–1910) in the Republic of Korea. Here, we report the presence of trematode eggs, including Clonorchis sinensis, Metagonimus yokogawai, and Gymnophalloides seoi (a human parasite known in Korea since 1993) in the feces of a recently excavated female mummy in Hadong, Republic of Korea. This is the first report of the discovery of a G. seoi infection in a human mummy. Since Hadong is currently not an endemic area for G. seoi, we speculate that the parasite might have occurred frequently along coastal areas of the Korean peninsula several hundred years ago and that the endemic areas contracted to, more or less, restricted regions since that time.

Paleoparasitological report on Ascaris aDNA from an ancient East Asian sample.

In this study, Ascaris DNA was extracted and sequenced from a medieval archaeological sample in Korea. While Ascaris eggs were confirmed to be of human origin by archaeological evidence, it was not possible to pinpoint the exact species due to close genetic relationships among them. Despite this shortcoming, this is the first Ascaris ancient DNA (aDNA) report from a medieval Asian country and thus will expand the scope of Ascaris aDNA research.

Technical Note: Efficiency of Total Demineralization and Ion-Exchange Column for DNA Extraction from Bone

We investigated whether a combination of recently introduced methods, total demineralization and ion-exchange columns, would increase DNA recovery from old bone. Ten bone samples taken after a burial period of approximately 60 years were used in this study. Bone powder was digested using total or incomplete demineralization. DNA was extracted by the standard organic method. The DNA extract was purified with ion-exchange columns or QIAquick spin columns. The efficiency of different DNA extraction methods was compared in terms of DNA concentration, inhibitors generated by real-time PCR, and conventional STR typing results. The mean DNA concentration using the total demineralization method is approximately 3 times higher than that using the incomplete demineralization method. For DNA purification, the method using QIAquick spin columns appeared to yield approximately double the DNA than the method using ion-exchange columns. Furthermore, 2 out of 10 samples showed higher levels of inhibition with C(T) values of IPC > or =30 cycles when using only ion-exchange columns. In STR results, total demineralization yielded more locus profiles by 4.2 loci than incomplete demineralization, and QIAquick spin columns also yielded more locus profiles by 3.5 loci than ion-exchange columns. Total demineralization of bone powder significantly increased DNA yield and improved STR typing results. However, the use of ion-exchange columns was not efficient when compared with the method using QIAquick spin columns. It is suggested that the combination of total demineralization and QIAquick spin columns lead to greatly improved STR typing results.

Acute traumatic death of a 17th century general based on examination of mummified remains found in Korea

Recently, we examined one of the most perfectly preserved mummies of the Joseon Dynasty (1392-1910) ever found in Korea. The individual was an elderly man and a high-ranking general who had lived sometime during the 16th or 17th century in Korea. When computerized tomography (CT) radiographs were taken, a fracture line was observed on the left side of the mandible. A post-factum dissection also provided crucial clues to the cause of death. First of all, blood clots were still evident at the fracture site, indicating that the mandibular fracture had occurred just before death. Second, we also found feces exclusively in the sigmoid colon or rectum, but not in the stomach, small intestine or colon. This told us that our subject had not eaten anything during his final 2 days (even though there was no indication that he would have had any difficulty eating during that time). Therefore, we presume that this case might not be one of chronic or wasting disease, but rather a case of sudden death. By virtue of the varied specialties of the researchers involved in this study, we were able to piece together a partly very clear and partly very plausible story for our 17th century mummy subject. Considering the high level of preservation of remains and artifacts found in lime soil mixture barrier (LSMB) tombs, not to mention the rich supplementary information available from historical documents, similarly successful studies are promised in forthcoming days and years.

The potential for non-invasive study of mummies: validation of the use of computerized tomography by post factum dissection and histological examination of a 17th century female Korean mummy.

The socio‐cultural antipathies of some descendants with regard to invasive examinations of age‐old human remains make permission for dissection of Korean mummies of the Joseon Dynasty (1392–1910) difficult to obtain. Overcoming this obstacle necessitated the use of non‐invasive techniques, such as multi‐detector computerized tomography (MDCT) and endoscopic examination, enabling determination of the preservation status of internal organs of mummies without significantly damaging the mummies themselves. However, MDCT alone cannot clearly differentiate specific mummified organs. Therefore, in much the same way as diagnostic radiologists make their MDCT readings on living patients more reliable by means of comparison with accumulated post‐factum data from autopsies or histological studies, examinations of mummies by invasive techniques should not be decried as mere destruction of age‐old human remains. Rather, providing that due permission from descendants and/or other relevant authorities can be obtained, dissection and histological examination should be performed whenever opportunities arise. Therefore, in this study, we compared the radiological data acquired from a 17th century mummy with our dissection results for the same subject. As accumulation of this kind of data could be very crucial for correct interpretation of MDCT findings on Korean mummies, we will perform similar trials on other Korean mummies found in forthcoming days if conditions permit.

Independent validation of DNA-based approaches for age prediction in blood

Numerous molecular biomarkers have been proposed as predictors of chronological age. Among them, T-cell specific DNA rearrangement and DNA methylation markers have been introduced as forensic age predictors in blood because of their high prediction accuracy. These markers appear highly promising, but for better application to forensic casework sample analysis the proposed markers and genotyping methods must be tested further. In the current study, signal-joint T-cell receptor excision circles (sjTRECs) and DNA methylation markers located in the ELOVL2, C1orf132, TRIM59, KLF14, and FHL2 genes were reanalyzed in 100 Korean blood samples to test their associations with chronological age, using the same analysis platform used in previous reports. Our study replicated the age association test for sjTREC and DNA methylation markers in the 5 genes in an independent validation set of 100 Koreans, and proved that the age predictive performance of the previous models is relatively consistent across different population groups. However, the extent of age association at certain CpG loci was not identical in the Korean and Polish populations; therefore, several age predictive models were retrained with the data obtained here. All of the 3 models retrained with DNA methylation and/or sjTREC data have a CpG site each from the ELOVL2 and FHL2 genes in common, and produced better prediction accuracy than previously reported models. This is attributable to the fact that the retrained model better fits the existing data and that the calculated prediction accuracy could be higher when the training data and the test data are the same. However, it is notable that the combination of different types of markers, i.e., sjTREC and DNA methylation, improved prediction accuracy in the eldest group. Our study demonstrates the usefulness of the proposed markers and the genotyping method in an independent dataset, and suggests the possibility of combining different types of DNA markers to improve prediction accuracy.

Age estimation via quantification of signal-joint T cell receptor excision circles in Koreans

The estimation of age from biological samples (i.e., remains) at crime scenes could provide useful information about both victims and other persons related to criminal activities. Signal-joint T cell receptor excision circle (sjTREC) levels in peripheral blood decline with age, and negative correlations between sjTREC levels and age have been demonstrated in several ethnic groups. To validate the utility of sjTREC for age estimation in Koreans, Taqman qPCR was used to quantify the sjTREC level in samples obtained from 172 individuals ranging from 16 to 65 years old. We modified the previously reported method by using a shorter amplicon and confirmed the efficiency and utility of this method in this report. Our results showed that the linear negative regression curve between sjTREC levels and age was characterized by r=-0.807 and a standard error of 8.49 years. These results indicate that sjTREC level is an effective age estimation method in Koreans. The value of the standard error of quantification was not different from previous reports for other population groups.

Forensic DNA methylation profiling from evidence material for investigative leads.

DNA methylation is emerging as an attractive marker providing investigative leads to solve crimes in forensic genetics. The identification of body fluids that utilizes tissue-specific DNA methylation can contribute to solving crimes by predicting activity related to the evidence material. The age estimation based on DNA methylation is expected to reduce the number of potential suspects, when the DNA profile from the evidence does not match with any known person, including those stored in the forensic database. Moreover, the variation in DNA implicates environmental exposure, such as cigarette smoking and alcohol consumption, thereby suggesting the possibility to be used as a marker for predicting the lifestyle of potential suspect. In this review, we describe recent advances in our understanding of DNA methylation variations and the utility of DNA methylation as a forensic marker for advanced investigative leads from evidence materials. [BMB Reports 2016; 49(7): 359-369]

Autosomal short tandem repeat analysis of ancient DNA by coupled use of mini- and conventional STR kits.

Abstract:  Multiplex autosomal short tandem repeat (STR) genotyping enables researchers to obtain genetic information from ancient human samples. In this study, we tested newly developed AmpFℓSTR® MiniFiler™ kit for autosomal STR analysis of ancient DNA (aDNA), using human femurs (n = 8) collected from medieval Korean tombs. After extracting aDNA from the bones, autosomal STR analyses were repeated for each sample using the AmpFℓSTR® MiniFiler™ and Identifiler™ kits. Whereas only 21.87% of larger‐sized loci profiles could be obtained with the Identifiler™ kit, 75% of the same loci profiles were determined by MiniFiler™ kit analysis. This very successful amplification of large‐sized STR markers from highly degraded aDNA suggests that the MiniFiler™ kit could be a useful complement to conventional STR kit analysis of ancient samples.

Forensic application of SNP-based resequencing array for individual identification

In forensic field investigations using single nucleotide polymorphism (SNP) have been performed for various purposes. Based on the characteristics of SNP, it is essential to have a multi-amplification technology and a platform to analyze the amplified SNP markers accurately. Here, we have developed a platform based on the resequencing array of Affymetrix analyzing 169 SNP markers amplified via multiplex PCR and verified its forensic application. From the 1000 genomes database, the SNP markers were selected under the condition of less than 0.04 fixation index (Fst) and 0.3 linkage disequilibrium (LD) R(2) value, and 0.4-0.5 minor allele frequency (MAF). It was identified that more than 120 out of 169 SNPs were able to be typed with approximately 10pg of DNA and artificially degraded samples in various tests. The DNA extracted from bones also showed a similar rate of success. The results indicated that our platform has a potential role to assist the current short tandem repeat (STR) method in analyzing harsh samples, such as bone or degraded DNA. With a possibility to expand the platform, it was expected to apply to various uses in different areas in the future.