Chironomid communities are indicators of water pollution because of their ability to thrive under freshwater conditions. However, it is difficult to distinguish between chironomid larvae based on morphology. DNA barcoding, based on nucleotide sequences of marker genes, can be used to identify chironomid larvae. Samples of chironomid larvae were collected from Gwangju Stream and Pungyeongjeong Stream, tributaries of the Yeongsan River in South Korea. We identified 3 subfamilies, 13 genera, 16 species, and 1 cryptic species. There were 7 genera and 10 species from the subfamily Chironominae, 5 genera and 5 species from subfamily Orthocladiinae, 1 genus and 1 species from subfamily Tanipodinae, and the cryptic chironomid species of the family Chironomidae. There were 21 individuals from, 7 species and 1 cryptic species from the Gwangju Stream and 24 individuals, belonging to 10 species from the Pungyeongjeong Stream. The only species detected in both streams was Cricotopus bicinctus. The relationship between water quality and the species detected was difficult to explain, but the number of species showed a tendency to increase at sites where water quality was poor. Additional investigations and studies are needed to understand the relationship between water quality and the chironomid species occurring in these two streams.

본 연구는 국내에 유통되는 두족류 제품에 대한 모니터링을 수행하였다. 문어와 오징어로 표기되어 판매되는 28개 제품을 대상으로 DNA 바코드를 분석하여 원재료의 종을 동정하였다. DNA 바코드 증폭을 위하여 미토콘드리아의 16S ribosomal RNA 및 cytochrome c oxidase subunit I 유전자 부위를 증폭하는 두 종류의 프라이머 세트를 선정하였으며, 이를 이용하여 증폭된 PCR 산물의 염기서열을 분석하였다. 확보한 염기서열은 ‘BLAST Search’를 이용하여 미국국립보건원 GenBank에 등록되어있는 생물 종의 염기서열과 비교하여 유사도와 매칭 점수를 고려하여 최종 종을 동정하였다. 동정결과, 원재료를 오징어로 표기 한 12개 제품은 아메리카대왕오징어(Dosidicus gigas, n=3), 살오징어(Todarodes pacificus, n=9) 종으로 확인되었다. 반면 문어를 원재료로 표기한 16개 제품의 경우, 6개 제품에서 대만주머니낙지(Cistopus taiwanicus, n=1), 하이야주 꾸미(Amphioctopus marginatus, n=1), Scaeurgus unicirrhus (n=1), 아메리카대왕오징어(Dosidicus gigas, n=3)로 동정되어 표기된 원재료와 불일치하였으며, 이 중 3개의 제품은 알레르기 유발 원재료인 오징어가 사용되었음을 확인하였다.

This study was investigated to test whether the zygote recognized the topoisomerase II beta (TOP2B) mediated DNA fragmentation in epididymal spermatozoa or the nuclease degradation in vas deferens spermatozoa by testing for the presence of gammaH2AX (γH2AX). The γH2AX is phosphorylation of histone protein H2AX on serine 139 occurs at sites flanking DNA double-stranded breaks (DSBs). The presence of γH2AX in the pronuclei of mouse zygotes which were injected with DNA broke epididymal spermatozoa was tested by immunohistochemistry at 5 and 9 h post fertilization, respectively. Paternal pronuclei that arose from epididymal spermatozoa treated with divalent cations did not stain for γH2AX at 5 h. On the other hand, in embryos injected with vas deferences spermatozoa that had been treated with divalent cations, γH2AX was only present in paternal pronuclei, and not the maternal pronuclei at 5 h. Interestingly, both pronuclei stained positively for γH2AX for all treatments and controls at 9 h after sperm injection. In conclusion, the embryos recognize DNA that is damaged by nuclease, but not by TOP2B because H2AX in phosphorylated in paternal pronuclei resulting from spermatozoa treated with fragmented DNA from vas deferens spermatozoa treated with divalent cations, but not from epididymal spermatozoa treated the same way.

해외 수입농산물의 양이 점차적으로 증가하여 다양한 병해충의 유입 가능성이 증가함에 따라, 이들을 신속하고 정확하게 판별하기 위하여 DNA 염기서열을 이용하여 형태학적 동정을 보완하고 있다. 현재 바구미과의 Sternochetus속 에 속하는 관리해충은 S. mangiferae와 S. frigidus, 그리고 S. olivieri가 있으며, 태국에서 수입된 망고에서 2018년 9월부터 2019년 1월까지 7차례에 걸쳐 바구미류가 검출되었고, 형태학적 형질 및 DNA 바코드를 이용하여 S. olivieri로 동정하였다. 추가로, Sternochetus속에 속하는 종들의 분류학적 형질을 이용하여 동정의 수단을 제공하였다. 본 연구결 과는 수입 및 반입되는 망고에서 검출되는 바구미류의 신속하고 정확한 진단에 활용될 수 있을 것으로 판단된다.

Due to rapid increase of international trade, many invasive and exotic pests have been introduced in Korea. One of typical example is Solenopsis invicta found in harbor and nearby areas unexpectedly triggering alerts of invasive and exotic pests. Practically, critical limitation to identify these species based on morphology exists because of lack of experts, so that it is very important to develop fast and accurate methods to identify these species. Molecular marker is one of candidates for satisfying these requirements of invasive and exotic pests: usually COI gene has been used for identifying insect species efficiently. Here, we developed web-based integrated platform for identifying invasive and exotic pests. As a first step, we collected 71,146 COI sequences from 529 species which are potentially invasive and exotic pests in Korea. In addition, we are collecting their complete mitochondrial genome sequences for evaluating additional marker regions which can be more effective for identifying species. Web-based interfaces are under development to access these raw data as well as bioinformatic analysis function to identify species based on mitochondrial sequences. Our platform will be a fundamental resources not only to identify invasive and exotic pests effectively but also to understand ecology of these species to find anticipative policies to prevent invasion of these species.

In this study, we developed 11 microsatellite markers specific to A. crataegi using NGS to investigate the genetic relationships of A. crataegi populations from South Korea to circumferential Asian countries (China, Russia, Mongolia, and Japan). Further, two mitochondrial DNA (mtDNA) gene segments (COI and CytB) were sequenced from the samples. The population- and individual-based Principal Coordinates and STRUCTURE analyses collectively suggested that the South Korean population of A. crataegi is most differentiated from the Japanese population, whereas it was closer to Mongolian and Chinese populations. These results collectively suggest that northern populations, in particular, Mongolian populations can be considered as the most genetically compatible one as donee population, when the reintroduction program is launched. †These authors contributed equally to this paper.

During the last couple of decades, molecular techniques are widespread tools for the species diversity. However, their application to taxonomy provoked intense debates between traditional and molecular taxonomists. To prevent every kind of disagreement, it should be required to a threshold for standarizing the species delimitation. Here, we tested three species delimitation methods (ABGD, PTP and bPTP) and compared their results with morphospecies on the widest DNA barcoding dataset. Moreover, a possible threshold for species determination within the superfamily has been suggested. It is believed that it could be an important criterion in detection for hidden or cryptic species, tracing of variation, and can help in identification on immature stages of pests.

한국산 나문재속 식물에 대한 계통학적 유연관계를 밝히고, 분자계통학적 연구를 통해 나문재속 종간 유연관계를 확인할 수 있는 분자마커를 찾아내기 위해 연구를 수행하였다. 핵 리보솜 DNA ITS와 엽록체 DNA matK, psbA-trnH 그리고 trnL-trnF를 분자마커로 사용하였다. ITS 영역은 칠면초와 해홍나물 그리고 해홍나물과 방석나물을 구분하지 못하였다. psbA-trnH와 trnL-trnF 영역의 염기서열은 칠면초와 방석나물을 구분하지 못하였다. 그러나 4종의 분자마커 영역을 조합하여 분석한 결과 나문재속 식물 5종이 각각 독립적인 계통을 형성하는 것을 확인하였다. 따라서 나문재속 계통관계 분석을 위해서 여러 개의 분자마커 조합이 유용할 것으로 판단된다. 나문재속 내 분류군 간의 계통관계를 명확히 밝히기 위해 차후에 좀 더 많은 생태학적, 형태학적 자료를 조사해야 할 것으로 보인다.

유전자 친자확인시험법은 동일 종 내에서도 개체를 식별 할 수 있는 강력한 분자진단체계이다. 반달가슴곰 우수리아 종(Ursus thibetanus ussuricus)은 한반도를 비롯한 중국, 러시아 등지에 분포하나, 우리나라에서는 지역적으로 절멸된 것으로 추정되고 있으며, 현재는 멸종위기야생생물Ⅰ급으 로 지정되어 있고, 북한, 러시아, 중국 등에서 재도입되어 종복원이 진행 중에 있다. 본 연구는 우리나라에 재도입된 반달가슴곰집단의 친자확인, 가계도 작성, 미확인개체 추적 등 방사개체의 관리에 요구되는 분자진단체계의 구축을 위하여 차세대염기서열분석법을 통해 결정한 반달가슴곰 전 장유전체 서열(whole genome sequence, WGS)에서 미세 부수체(microsatellite, MS) 마커 개발을 목적으로 하였다. 전장유전체 서열은 총 12개체의 혈액 DNA를 이용하여 결정하였고, 결정된 서열에서 총 1,730,507개의 단순반복서열(simple tandem repeat, STR)의 정보를 발굴하였다. 이 중 반복단위(repeat unit)의 길이가 2-6 nt로 구성된 STR은 총 5,954개였다. WGS 상에서 검출된 STR의 관찰이형접합율(observed heterozygosity, Ho)을 고려하여 총 97종을 1차 선정하였다. 부-모-자 유전자형의 멘델유전(mendelian inheritance)을 만족하는 48종의 마커를 반달가슴곰 전체 집단에 적용하여 시험하였고, 중합효소연쇄반응(polymerase chain reaction, PCR)을 통해 유전자형을 결정하였다. 유전 자형의 다양성지수와다형정보량(polymorphic information content, PIC), 동일개체출현율을 고려하여 최종 15종의 MS 마커 세트를 구성하였다. 분석결과, 전체 집단에서 출현한 평균 대립유전자의 수는 6.267개였으며, 기대이형접합율(expected heterozygosity)는 평균 0.7242을 나타내었다. 평균 PIC는 0.6718, 동일개체 출현율은 1.867×10-14 수준으로 높았으나, 부권부정율은 0.00337로 다소 낮은 수준을 보였다. 재도입된 1세대집단 전체에서는 평균 대립유전자 수 5.867개, 평균 기대이형접합율 0.7217, 평균 PIC 0.6573을 나타내었고, 후손 전체에서는 평균 대립유전자 수 5.933개, 평균 기대이형접합율 0.7140, 평균 PIC 0.6554를 나타내었다. 하지만, 후손집단을 2세대와 3세대로 구분했을 때, 2세대는 평균 대립유전자 수 5.867, 평균 기대이형접합율 0.722, 평균 PIC 0.657, 3세대는 평균 대립유전자 수 4.467, 평균 기대이형접합율 0.694, 평균 PIC 0.601을 나타내어 세대가 진행될수록 대립유전자의 수와 기대이형접합율, PIC 모두 감소하는 경향을 나타내었다. 최종 선정된 15개의 MS 마커체계를 이용하여 반달가슴곰 집단에 대한 미확인 개체 추적, 친자확인 등을 시험하였다. 최근 포획된 개체들 중에서 개체명이 확인되지 않았던 4개체는 자연출생 2세대, 3세대, 전파발신기가 탈락된 방사 1세대로 확인 되었다. 또한 동일성검사 결과 올해 5월 교통사고를 당한 반달가슴곰은 KM-53이며, 올해 출생한 새끼 반달가슴곰들에 대한 검사를 통해 2개체가 인공수정에 의해 출생한 것으로 확인되었다. 이상의 결과들은 본 연구에서 확립한 MS 마커의 조합이 자연방사와 자연출생 개체들로 구성된 반달 가슴곰 집단의 개체관리를 위한 분자마커체계로 유용하게 이용될 수 있음을 보여주고 있다. 또한 본 연구를 통해서 확립된 MS 마커들은 현재 지리산과 수도산에 서식하고 있는 반달가슴곰 집단의 개체관리, 미확인 개체 추적, 친자확인 등 유전자분석에 활용될 수 있을 것으로 기대되며, 아울러 향후 유전적 다양성 증진, 집단간 교류개체 선정 등에도 필요한 유전 정보를 제공할 수 있을 것이다. 결론적으로 반달가슴곰 전장유전체 서열을 토대로 마련된 분자진단체계는 향후 반달가슴곰의 보호와 생태복원을 위한 종복원에 기여할 것으로 기대된다.

Promalactis established by Meyrick in 1908, is one of the highest species richness genera of the family Oecophoridae. Over 240 described species are mainly distributed from Oriental and Palaearctic regions. They are usually a tiny-sized and exhibit subtle morphological differences which make problems on morphological-based identification. In this study, we performed DNA barcoding study for the Promalactis by using mitochondrial cytochrome c oxidase subunit I (COI). From analyzing 155 COI sequences, we observed the usefulness of the COI in species identification with intraspecific genetic variation (range 0.0-2.1%) and interspecific genetic divergence (range 2.8-15.4%).

Incursions of red imported fire ant (RIFA), Solenopsis invicta into Korea have been increasing. After a first interception of a colony of S. invicta on Gamman pier, Pusan port while intensive surveillance by Animal & Plant Quarantine Agency (APQA) in September 2017, three more RIFA colonies have been found in sea port piers of Pyeongtek, Incheon and Pusan cities. The social forms and mitochondrial DNA haplotypes of the intercepted RIFA colonies were analysed by allelic discrimination assay (peptide nucleic acid probe based RT-PCR) of Gp-9 gene and mt-DNA fragment of 768 bp, which is part of the Cytochrome oxidaseⅠ gene. The colony on Gamman pier, intercepted in Sep. 2017 was previously reported as a haplotype 5 (H5) of mitochondrial DNA and a social form of polygyne. The colony on Hutchison pier of Pusan port, intercepted in June 2018 were confirmed as a H22 haplotype and a monogyne. Those different social forms show different origins of each colonies. Those on piers of Pyeongtek and Incheon ports, also found in 2018 were confirmed samely as H22 and monogyne. However, it could be putatively assumed that those two colonies were differently introduced via different container cargoes, considering those colonies were found in container yards of distantly located different sea ports. More genetic variation analyses using diverse sets of molecular markers such as microsatellites, genome-wide single nucleotide polymorphisms, etc. in nuclear gens are being proceeded for more exact introduction routes (origins).

We conducted quarantine insect species diversity monitoring using DNA barcoding with 517 lepidopteran samples that were obtained from quarantine inspections of foreign vessels entering Korea. The DNA barcode of each sample was treated as a molecular operational taxonomic unit (MOTU). For species delimitation and species identification of the analyzed samples, we applied a 2% cutoff rule and then identified with using BLAST of NCBI GenBank and BOLD System ver. 3.0. Consequently, 517 analyzed samples were delimited as 214 putative species across 20 families. Of these 214 putative species 145 (368 samples) were considered taxonomically identified if the closest BLAST match was no more than 2% different. Therefore the number of samples that were identified to the species level was relatively low, at approximately 71%. 115 of the 145 species were known in Korea. Of the 30 species that were not known in Korea, three, i.e., Noctua pronuba (Noctuidae), Orthosia hibisci (Noctuidae), and Pieris brassicae (Pieridae), were checked as ‘Regulated pests’ in Korea. We suggest that the three regulated pest species could be prevented from being introduced to Korea if monitoring of the vessels that pass the navigation route that contains these three species is performed consistently. Therefore, we suggest that the monitoring of quarantined insect pest enables the prevention of the introduction of alien species.

The black-veined white, Aporia crataegi (Lepidoptera: Pieridae), which is distributed mainly in Eastern Asia is presumed to be extinct in South Korea, only with some numbers of dried specimens left, whereas the species is found casually in circumferential countries. One of the common conservation practices for such species is to launch introduction program, but prior population genetic analysis between donor and donee populations might be essential for long-term conservation. In this study, we developed 11 microsatellite markers specific to A. crataegi using Illumina paired-end sequencing to investigate the genetic relationships of A. crataegi populations from South Korea and circumferential Asian countries (China, Russia, Mongolia, and Japan). Further, two mitochondrial DNA (mtDNA) gene segments (COI and CytB) were sequenced from the samples. The population- and individual-based Principal Coordinates and STRUCTURE analyses collectively suggested that the South Korean population of A. crataegi is most differentiated from the Japanese population, whereas it was closer to Mongolian and Chinese populations. The STRUCTURE analysis based on two concatenated mtDNA gene sequences also supported different genetic composition of Japanese population from the remaining populations including that of South Korea and rather similar genetic composition between the populations of South Korea and Mongolia. These results collectively suggest that northern populations, in particular, Mongolian populations can be considered as the most genetically compatible one as doner population, when reintroduction program is launched.