Sang Eon Shin

Molecular Identification of Necrophagous Muscidae and Sarcophagidae Fly Species Collected in Korea by Mitochondrial Cytochrome c Oxidase Subunit I Nucleotide Sequences

Identification of insect species is an important task in forensic entomology. For more convenient species identification, the nucleotide sequences of cytochrome c oxidase subunit I (COI) gene have been widely utilized. We analyzed full-length COI nucleotide sequences of 10 Muscidae and 6 Sarcophagidae fly species collected in Korea. After DNA extraction from collected flies, PCR amplification and automatic sequencing of the whole COI sequence were performed. Obtained sequences were analyzed for a phylogenetic tree and a distance matrix. Our data showed very low intraspecific sequence distances and species-level monophylies. However, sequence comparison with previously reported sequences revealed a few inconsistencies or paraphylies requiring further investigation. To the best of our knowledge, this study is the first report of COI nucleotide sequences from Hydrotaea occulta, Muscina angustifrons, Muscina pascuorum, Ophyra leucostoma, Sarcophaga haemorrhoidalis, Sarcophaga harpax, and Phaonia aureola.

The First Survey of Forensically Important Entomofauna Collected from Medicolegal Autopsies in South Korea

Forensic entomology applies insect evidence to legal problems such as the estimation of minimum postmortem interval (mPMI). For this purpose, knowledge of the insect fauna that are attracted to human cadavers in each geographic region is a prerequisite. Despite many studies investigating the insect fauna attracted to meat, there has been no survey of the entomofauna on human cadavers in the East Asian temperate climate zone, particularly in Korea. Therefore, this study reports the entomofauna collected from medicolegal autopsies in northeastern Seoul and its suburbs. Insect samples were collected from 35 medicolegal autopsies in 2010, 2011, and 2013. Molecular and morphological methods were utilized for taxonomic identification. Among 1398 individual samples belonging to 3 orders, 13 families, 18 genera, and 32 species, the dominant family and species were Calliphoridae and Lucilia sericata, respectively. Despite its limited scale, this study provides a snapshot of the general entomofauna that are attracted to human cadavers in this region.

First report of external ophthalmomyiasis caused by Lucilia sericata Meigen in a healthy patient without predisposing risk factors

A 72-year-old man with no medical history initially presented to the emergency room with severe tearing, redness, foreign body sensation, and pain in the left eye. He reported no previous history of any periocular trauma, malignancy, surgery, or systemic illness. On presentation, the patient only showed left periorbital edema and erythema in the left eyelid with no evidence of any skin malignancy. On slit lamp examination, multiple small whitish motile organisms were observed on the left conjunctival fornices. The organisms were removed, preserved, and identified as the third-stage larvae of Lucilia sericata (green bottle fly). The patient was treated with topical antibiotic and steroid eye drops and the inflammation resolved 1 week after treatment initiation. This is the first report of external ophthalmomyiasis caused by facultative parasite, L. sericata maggots in a healthy patient without any predisposing factors.

Molecular identification of forensically important calliphoridae and sarcophagidae species using ITS2 nucleotide sequences

The application of insect evidence to forensic investigations is mainly based on the estimation of postmortem interval and the identification of insect species from samples that are collected from the crime scene. Due to the limited number of expert taxonomists, species identification is one of the major barriers for crime scene investigators to utilize forensic entomology. Therefore, the molecular identification of species, using mitochondrial cytochrome c oxidase subunit I (COI) gene, has been suggested as an alternative strategy. However, in some cases, these maternally inherited markers cause confusion; hence, nuclear DNA markers such as ITS2 are also required as supporting tools. Eleven Calliphoridae and 5 Sarcophagidae fly species collected from Korea were utilized for PCR amplification and nucleotide sequencing of ITS2 locus. Species Identifier software was used for sequence analysis and comparison. The results demonstrated that 11 Korean Calliphoridae and 5 Korean Sarcophagidae fly species could be distinguished using ITS2 nucleotide sequences. In particular, the sister species, Lucilia illustris and Lucilia caesar were also distinguished, despite the very low level of interspecific diversity. However, when compared with previously reported ITS2 nucleotide sequences, several identification failures were noted. This is the first study that widely analyzed nucleotide sequences of the ITS2 locus from Calliphoridae and Sarcophagidae fly species collected in Korea.

Identification of Forensically Important Calliphoridae and Sarcophagidae Species Collected in Korea Using SNaPshot Multiplex System Targeting the Cytochrome c Oxidase Subunit I Gene

Estimation of postmortem interval (PMI) is paramount in modern forensic investigation. After the disappearance of the early postmortem phenomena conventionally used to estimate PMI, entomologic evidence provides important indicators for PMI estimation. The age of the oldest fly larvae or pupae can be estimated to pinpoint the time of oviposition, which is considered the minimum PMI (PMImin). The development rate of insects is usually temperature dependent and species specific. Therefore, species identification is mandatory for PMImin estimation using entomological evidence. The classical morphological identification method cannot be applied when specimens are damaged or have not yet matured. To overcome this limitation, some investigators employ molecular identification using mitochondrial cytochrome c oxidase subunit I (COI) nucleotide sequences. The molecular identification method commonly uses Sangers nucleotide sequencing and molecular phylogeny, which are complex and time consuming and constitute another obstacle for forensic investigators. In this study, instead of using conventional Sangers nucleotide sequencing, single-nucleotide polymorphisms (SNPs) in the COI gene region, which are unique between fly species, were selected and targeted for single-base extension (SBE) technology. These SNPs were genotyped using a SNaPshot® kit. Eleven Calliphoridae and seven Sarcophagidae species were covered. To validate this genotyping, fly DNA samples (103 adults, 84 larvae, and 4 pupae) previously confirmed by DNA barcoding were used. This method worked quickly with minimal DNA, providing a potential alternative to conventional DNA barcoding. Consisting of only a few simple electropherogram peaks, the results were more straightforward compared with those of the conventional DNA barcoding produced by Sangers nucleotide sequencing.

Complete mitochondrial DNA analysis of Hydrotaea ignava (Insecta, Diptera, Muscidae)

Abstract The complete mitochondrial genome sequenced and analyzed from a black garbage fly, Hydrotaea ignava which is critically important species for forensic investigations. The size of mitochondrial genome is 17,026 bp with 40.8% A, 11.2% C, 8.2% G and 39.8% T distribution. This is the longest within complete mitochondrial genome records of the Muscidae species. The mitochondrial genome is composed of 13 protein coding, two ribosomal RNA and 22 tRNA genes and a putative control region. Furthermore, phylogenetic relationships of the superfamily Muscoidea evaluated due to mitochondrial protein coding genes. The results showed that the H. ignava placed in the paraphyletic Muscidae family and early diverged from a clade including Muscidae, Anthomyiidae and Scathophagidae species. This is the first complete mitochondrial genome for the genus Hydrotaea.

Complete mitogenome and phylogenetic analysis of hide beetle Dermestes maculatus (Insecta, Coleoptera, Dermestidae)

Abstract In this study, the complete mitochondrial genome of hide beetle Dermestes maculatus which was collected from Seoul, South Korea was sequenced by next-generation sequencing. The size of mitochondrial genome is 17,026 bp that composed of 13 protein coding, two ribosomal RNA and 22 tRNA genes which has the identical gene orientation with the other Bostrichiformia species. Additionally, the phylogenetic tree of the D. maculatus in the infraorder Bostrichiformia was reconstructed by using 13 protein-coding genes of complete mitochondrial genome. The results showed that the family Dermestidae is positioned in the infraorder Bostrichiformia early branched than family Bostrichidae. This study provides the first complete mitochondrial genome from the genus Dermestes.