• Title/Summary/Keyword: Environmental DNA

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PCR Diagnosis of Entamoeba histolytica Cysts in Stool Samples

  • Moon, Joung-Ho;Cho, Shin-Hyeong;Yu, Jae-Ran;Lee, Won-Ja;Cheun, Hyeng-Il
    • Parasites, Hosts and Diseases
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    • v.49 no.3
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    • pp.281-284
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    • 2011
  • Amebiasis is a protozoan disease caused by Entamoeba histolytica and a potential health threat in areas where sanitation and hygiene are inappropriate. Highly sensitive PCR methods for detection of E. histolytica in clinical and environmental samples are extremely useful to control amebiasis and to promote public health. The present study compared several primer sets for small subunit (SSU) rDNA and histone genes of E. histolytica cysts. A 246 bp of the SSU rDNA gene of pure cysts contained in phosphate-buffered saline (PBS) and in stool samples was successfully amplified by nested PCR, using the 1,147-246 bp primer set, of the primary PCR products which were pre-amplified using the 1,147 bp primer as the template. The detection limit of the nested PCR using the 1,147-246 primer set was 10 cysts in both groups (PBS and stool samples). The PCR to detect histone gene showed negative results. We propose that the nested PCR technique to detect SSU rDNA can be used as a highly sensitive genetic method to detect E. histolytica cysts in stool samples.

Characterization and Expression of Chironomus riparius Alcohol Dehydrogenase Gene under Heavy Metal Stress (중금속 노출에 따른 리파리 깔다구에서의 ADH 유전자의 발현 및 특성)

  • Park, Ki-Yun;Kwak, Inn-Sil
    • Environmental Analysis Health and Toxicology
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    • v.24 no.2
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    • pp.107-117
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    • 2009
  • Metal pollution of aquatic ecosystems is a problem of economic and health importance. Information regarding molecular responses to metal exposure is sorely needed in order to identify potential biomarkers. To determine the effects of heavy metals on chironomids, the full-length cDNA of alcohol dehydrogenase (ADH3) from Chironomus riparius was determined through molecular cloning and rapid amplification of cDNA ends (RACE). The expression of ADH3 was analyzed under various cadmium and copper concentrations. A comparative and phylogenetic study among different orders of insects and vertebrates was carried out through analysis of sequence databases. The complete cDNA sequence of the ADH3 gene was 1134 bp in length. The sequence of C. riparius ADH3 shows a low degree of amino acid identity (around 70%) with homologous sequences in other insects. After exposure of C. riparius to various concentrations of copper, ADH3 gene expression significantly decreased within 1 hour. The ADH3 gene expression was also suppressed in C. riparius after cadmium exposure for 24 hour. However, the effect of cadmium on ADH3 gene expression was transient in C. riparius. The results show that the suppression of ADH3 gene by copper exposure could be used as a possible biomarker in aquatic environmental monitoring and imply differential toxicity to copper and cadmium in C. riparius larvae.

Analysis of Vitellogenin Gene Expression in Synechogobius hastus (Gobiidae) (풀망둑 난황전구단백질 유전자발현 추적기법)

  • 계명찬
    • Korean Journal of Environmental Biology
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    • v.22 no.1
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    • pp.206-212
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    • 2004
  • In an effort to develop the tools for monitoring the contamination of xenoestrogen in the aquatic environment of Korea, reverse transcription-polymerase chain reaction (RT-PCR) analysis of vitellogenin (VTG) mRNA expression were optimized in Synechogobius hastus. Based on the partial VTG cDNA sequence VTG mRNA level in livers from male fishes was analyzed by RT-PCR. As an internal control beta actin mRNA was amplified. 3 ${\mu}g$ of total RNA was reverse transcribed in 20 $\mu$l reaction using murine leukemia virus 〔MuLV〕 reverse transcriptase. Subsequent PCR using the 1 ${\mu}g$ of cDNA resulted in linear increase in PCR product of VTG in female liver cDNA from 10 to 30 cycles of amplification. On the contrary, in male, PCR product first detected at 28 cycles of amplification and linearly increased during 38 cycles of amplification, suggesting that male S. hastus expresses minute amount of VTG mRNA which is $2^{-18}$ equivalent of female. In conclusion, the optimized protocol of VTG mRNA expression in the liver of male S. hastus will be promising the environmental monitoring the xenoestrogen contamination in the western coast and estuaries in Korea.

Studies on Toxicological Evaluation of Freshwater Sediment using a PLHC-1 Cell Comet Assay (PLHC-1세포주의 Comet assay를 이용한 하천 퇴적토의 생태독성평가)

  • Bak, Jeong-Ah;Hwang, In-Young;Baek, Seung-Hong;Kim, Young-Sug
    • Korean Journal of Environmental Biology
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    • v.29 no.1
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    • pp.23-30
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    • 2011
  • In this study, the Comet assay (evaluation of DNA damage) used the fish hepatocellular carinoma cell, PLHC-1, was tried to the sediment extract obtained from freshwater to understand its applicability as a tool for monitoring sediment toxicity. In parallel, induced EROD (7-ethoxyresorufin- O-deethylase) activity and DNA damage (TEM values) in PLHC-1 cells were measured for establishing the tandem endpoints of the PLHC-1cell test to test the ecotoxicity of sediment. Among several study sites in a small river passed through downtown and industrial park area, one of them, site B, showed a higher level of EROD activity and DNA damage than other sites. It indicates that a tandem endpoints of PLHC-1 cells could be useful tools for assessing the toxicity of sediment. The sensitivity of Comet assay with PLHC-1 cells was a little higher than that with a blood cell of frog tadpoles to the solvent extract of sediment. According to the results, a PLHC-1 cell-Comet assay could be used as a useful tool for evaluating ecotoxicity of the freshwater sediment. In addition, more detailed studies are needed to the contaminated site.

A Reliable "Direct from Field" PCR Method for Identification of Mycorrhizal Fungi from Associated Roots

  • Kuhnann, Christoph;Kim, Seak-Jin;Lee, Sang-Sun;Harms, Carsten
    • Mycobiology
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    • v.31 no.4
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    • pp.196-199
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    • 2003
  • A very reliable and specific method for the identification of fungi in ectotrophic mycorrhizal symbiosis was developed using a specific PCR assay based on the amplification of the ITS1 region. To obtain specific data, an ITS-diagnostic assay was carried out that reveals genera and species specific sequences. Here, an application of one method is presented, which covers the identification of pure mycelia, basidiocarps as well as mixed samples such as ectomycorrhizal roots that were mingled with remains of the host plant. For this purpose a protocol was established that allowed the extraction of DNA from single mycorrhizal roots. In order to perform a specific ITS analysis we generated a new ITS-primer(ITS8) by a multiple alignment of five different genera and species of mycorrhizal fungi. The utilization of ITS1 and ITS8 resulted in specific PCR amplicons, which were characterized by sequencing without purification steps, even when the template DNA was associated with roots.

The Role of Gene-environment Interaction in Environmental Carcinogenesis (환경성 발암 기전에서 유전자-환경 상호작용의 역할)

  • Han, So-Hee;Lee, Kyoung-Mu
    • Journal of Environmental Health Sciences
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    • v.36 no.1
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    • pp.1-13
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    • 2010
  • Evidences supporting gene-environment interaction are accumulating in terms of environmental exposure including lifestyle factors and related genetic variants. One form of defense mechanism against cancer development involves a series of genes whose role is to metabolize (activation/detoxification) and excrete potentially toxic compounds and to repair subtle mistakes in DNA. The purpose of this article is to provide a brief review of the notion of gene-environment interaction, environmental/occupational carcinogens and related cancers, and previous studies of gene-environment interaction on cancers caused by exposure to carcinogenesis. With a number of studies on the interaction between lifestyle factors (e.g., smoking and diet) and genetic polymorphisms in genes involved in xenobiotic metabolism and DNA repair excluded, only several studies have been conducted on the interactive effects between polymorphisms of CYPs, GSTs, ERCCs, XRCCs and environmental/occupational carcinogens such as vinyl chloride, benzo[a]pyrene, and chloroform on carcinogenesis or genotoxicity. Future studies may need to be conducted with sufficient number of subjects and based on occupational cohorts to provide useful information in terms of advanced risk assessment and regulation of exposure level.

Dynamics of Bacterial Communities Analyzed by DGGE during Cyanobacterial Bloom in Daechung Reservoir, Korea (대청호 수화발생시기의 미생물 다양성 및 계통분류학적 분석)

  • Ko, So-Ra;Ahn, Chi-Yong;Lee, Young-Ki;Oh, Hee-Mock
    • Korean Journal of Environmental Biology
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    • v.29 no.3
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    • pp.225-235
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    • 2011
  • The change of microbial communities during cyanobacterial bloom was comparatively analyzed by 16S rDNA PCR-DGGE in Daechung Reservoir during 2003~2005. Morphological analysis showed that Cyanophyceae dominated algal community in the bloom. Dominant cyanobacteria were Microcystis, Planktothrix (Oscillatoria), Phormidium and Anabaena. We used 16S rDNA-denaturing gradient gel electrophoresis (DGGE) profiles and phylogenetic affiliations of the DGGE bands to analyze the community structure and diversity of the predominant microbial community. The DGGE band patterns demonstrated that the most frequent bands were identified as Microcystis during the monitoring periods, Planktothrix also dominated on September 2003 and 2004, whereas Anabaena was showed a peak on September 2005 and Aphanizomenon on August 2003. DGGE and phylogenetic analysis provided us new information that could not be obtained by traditional, morphological analysis. The relationship between cyanobacteria and other aquatic bacteria can be traced and their genetic diversity also identified in detail.

Analysis of Microbial Community During the Anaerobic Dechlorination of Perchloroethylene and Trichloroethylene (Perchloroethylene과 Trichloroethylene의 혐기적 탈염소화 및 미생물 군집 분석)

  • Lee Jae-Won;Kim Byung-Hyuk;Ahn Chi-Yong;Kim Hee-Sik;Yoon Byung-Dae;Oh Hee-Mock
    • Korean Journal of Microbiology
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    • v.41 no.4
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    • pp.281-286
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    • 2005
  • In this study, the anaerobic enrichment cultivation was performed with the sediments and the dredged soils from the cities of Ulsan, Masan, Yeosu, Gwangyang, Ansan and Seongnam. Acetate as an electron donor and PCE (perchloroethylene) or TCE (trichloroethylene) as an electron acceptor were injected into the serum bottle with an anaerobic medium. After the incubation of 12 weeks, the removal efficiency of PCE was highest at $70\%$ in the treatment with the sediment of Ulsan. Also, the bacterial community structure was analyzed by D-DGGE (double denatured gradient gel electrophoresis) through PCR-based 16S rDNA approaches. The dominant species id the anaerobic enrichment were found to belong to the genus of Desulfovibrio.

Bioluminescence capability and intensity in the dinoflagellate Alexandrium species

  • Park, Sang Ah;Jeong, Hae Jin;Ok, Jin Hee;Kang, Hee Chang;You, Ji Hyun;Eom, Se Hee;Yoo, Yeong Du;Lee, Moo Joon
    • ALGAE
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    • v.36 no.4
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    • pp.299-314
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    • 2021
  • Some species in the dinoflagellate genus Alexandrium are bioluminescent. Of the 33 formally described Alexandrium species, the bioluminescence capability of only nine species have been tested, and eight have been reported to be bioluminescent. The present study investigated the bioluminescence capability of seven Alexandrium species that had not been tested. Alexandrium mediterraneum, A. pohangense, and A. tamutum were bioluminescent, but A. andersonii, A. hiranoi, A. insuetum, and A. pseudogonyaulax were not. We also measured the bioluminescent intensity of A. affine, A. fraterculus, A. mediterraneum, A. ostenfeldii, A. pacificum, A. pohangense, A. tamarense, and A. tamutum. The mean 200-second-integrated bioluminescence intensity per cell ranged from 0.02 to 32.2 × 104 relative luminescence unit per cell (RLU cell-1), and the mean maximum bioluminescence intensity per cell per second (BLMax) ranged from 0.01 to 10.3 × 104 RLU cell-1 s-1. BLMax was significantly correlated with the maximum growth rates of Alexandrium species, except for A. tamarense. A phylogenetic tree based on large subunit ribosomal DNA (LSU rDNA) showed that the bioluminescent species A. affine, A. catenella, A. fraterculus, A. mediterraneum, A. pacificum, and A. tamarense formed a large clade. However, the toxicity or mixotrophic capability of these species was split. Thus, their bioluminescence capability in this clade was more consistent than their toxicity or mixotrophic capability. Phylogenetic trees based on LSU rDNA and the luciferase gene of Alexandrium were consistent except for A. pohangense. The results of the present study can provide a basis for understanding the interspecific diversity in bioluminescence of Alexandrium.