Application of Environmental DNA Metabarcoding in Biodiversity Research and Monitoring

doi:10.13560/j.cnki.biotech.bull.1985.2021-0176

Abstract

Abstract:

Environmental DNA metabarcoding（eDNA）is used to identify species and assess biodiversity by extracting environmental DNA from water,soil and air and using primers for PCR amplification and high-throughput sequencing. As a new monitoring technology,it is faster,more accurate and less destructive to the natural environment while compared with traditional monitoring technology. Therefore,it has changed the way we investigate the biodiversity of the earth to some extent. In this paper,the development and operation process of environmental DNA metabarcoding technology are reviewed. The application status in water,soil and air environment is summarized,and the unique indirect biodiversity monitoring methods are summarized. Concurrently,the advantages and existing problems are discussed in order to provide new ideas and inspirations for the follow-up research and monitoring of biodiversity.

Key words: eDNA metabarcoding, biodiversity, biomonitoring

KANG Zi-qing, ZHANG Yin-long, WU Yong-bo, XIE Dong, XUE Jian-hui, HUA Jian-feng. Application of Environmental DNA Metabarcoding in Biodiversity Research and Monitoring[J]. Biotechnology Bulletin, 2022, 38(1): 299-310.

Figures/Tables 1

References 114

[1]	曲建升, 李延梅, 王雪梅, 等. 生物多样性研究发展态势与挑战[J]. 科学观察, 2009, 4(6):1-8.
	Qu JS, Li YM, Wang XM, et al. International research profiles and challenges of biodiversity science[J]. Sci Focus, 2009, 4(6):1-8.
[2]	Butchart SHM, Walpole M, Collen B, et al. Global biodiversity:indicators of recent declines[J]. Science, 2010, 328(5982):1164-1168. doi: 10.1126/science.1187512 pmid: 20430971
[3]	Gavrilescu M, Demnerová K, Aamand J, et al. Emerging pollutants in the environment:present and future challenges in biomonitoring, ecological risks and bioremediation[J]. N Biotechnol, 2015, 32(1):147-156. doi: 10.1016/j.nbt.2014.01.001 URL
[4]	Siddig AAH, Ellison AM, Ochs A, et al. How do ecologists select and use indicator species to monitor ecological change? Insights from 14 years of publication in Ecological Indicators[J]. Ecol Indic, 2016, 60:223-230. doi: 10.1016/j.ecolind.2015.06.036 URL
[5]	Thomsen PF, Willerslev E. Environmental DNA - An emerging tool in conservation for monitoring past and present biodiversity[J]. Biol Conserv, 2015, 183:4-18. doi: 10.1016/j.biocon.2014.11.019 URL
[6]	Holdaway R, Research L, Wood J, et al. Using DNA meta barcoding to assess New Zealand’s terrestrial biodiversity[J]. NZ J Ecol, 2017, 41(2):251-262.
[7]	Levy-Booth DJ, Campbell RG, Gulden RH, et al. Cycling of extracellular DNA in the soil environment[J]. Soil Biol Biochem, 2007, 39(12):2977-2991. doi: 10.1016/j.soilbio.2007.06.020 URL
[8]	Pietramellara G, Ascher J, Borgogni F, et al. Extracellular DNA in soil and sediment:fate and ecological relevance[J]. Biol Fertil Soils, 2009, 45(3):219-235. doi: 10.1007/s00374-008-0345-8 URL
[9]	Ogram A, Sayler GS, Barkay T. The extraction and purification of microbial DNA from sediments[J]. J Microbiol Methods, 1987, 7(2/3):57-66. doi: 10.1016/0167-7012(87)90025-X URL
[10]	Hebert PD, Cywinska A, Ball SL, et al. Biological identifications through DNA barcodes[J]. Proc Biol Sci, 2003, 270(1512):313-321. doi: 10.1098/rspb.2002.2218 URL
[11]	Erickson DL, Spouge J, Resch A, et al. Dna barcoding in land plants:developing standards to quantify and maximize success[J]. Taxon, 2008, 57(4):1304-1316. pmid: 19779570
[12]	杨耀华. DNA条形码技术的应用进展[J]. 中国医药指南, 2013, 11(17):484-485.
	Yang YH. Advances in the application of DNA barcoding technology[J]. Guide China Med, 2013, 11(17):484-485.
[13]	武宇鹏, 丁亮, 李捷, 等. DNA条形码的应用进展及讨论[J]. 环境昆虫学报, 2011, 33(1):99-106.
	Wu YP, Ding L, Li J, et al. DNA barcoding:current progresses and discussions[J]. J Environ Entomol, 2011, 33(1):99-106.
[14]	Valentini A, Taberlet P, Miaud C, et al. Next-generation monitoring of aquatic biodiversity using environmental DNA metabarcoding[J]. Mol Ecol, 2016, 25(4):929-942. doi: 10.1111/mec.13428 pmid: 26479867
[15]	Ruppert KM, Kline RJ, Rahman MS. Past, present, and future perspectives of environmental DNA(eDNA)metabarcoding:a systematic review in methods, monitoring, and applications of global eDNA[J]. Glob Ecol Conserv, 2019, 17:e00547.
[16]	Dickie IA, Boyer S, Buckley HL, et al. Towards robust and repeatable sampling methods in eDNA-based studies[J]. Mol Ecol Resour, 2018, 18(5):940-952. doi: 10.1111/men.2018.18.issue-5 URL
[17]	SAYER C, SHILLAND E, GREAVES H, et al. Managing Britain’s ponds-conservation lessons from a Norfolk farm[J]. British Wildlife, 2013, 25(October):21-28.
[18]	Zhang S, Lu Q, et al. Assessment of fish communities using environmental DNA:Effect of spatial sampling design in lentic systems of different sizes[J]. Mol Ecol Resour, 2020, 20(1):242-255. doi: 10.1111/1755-0998.13105 pmid: 31625686
[19]	Strickland GJ, Roberts JH. Utility of eDNA and occupancy models for monitoring an endangered fish across diverse riverine habitats[J]. Hydrobiologia, 2019, 826(1):129-144. doi: 10.1007/s10750-018-3723-8
[20]	樊晓燕, 高景峰. 空气微生物群落样品采集、解析方法及研究进展[J]. 安全与环境学报, 2018, 18(1):357-363.
	Fan XY, Gao JF. On the sample collection, community analysis and research advances of the airborne microbial communities[J]. J Saf Environ, 2018, 18(1):357-363.
[21]	Collins RA, Wangensteen OS, O’Gorman EJ, et al. Persistence of environmental DNA in marine systems[J]. Commun Biol, 2018, 1:185. doi: 10.1038/s42003-018-0192-6 pmid: 30417122
[22]	Jo T, Murakami H, Yamamoto S, et al. Effect of water temperature and fish biomass on environmental DNA shedding, degradation, and size distribution[J]. Ecol Evol, 2019, 9(3):1135-1146. doi: 10.1002/ece3.2019.9.issue-3 URL
[23]	Strickler KM, Fremier AK, Goldberg CS. Quantifying effects of UV-B, temperature, and pH on eDNA degradation in aquatic microcosms[J]. Biol Conserv, 2015, 183:85-92. doi: 10.1016/j.biocon.2014.11.038 URL
[24]	Seymour M, Durance I, Cosby BJ, et al. Acidity promotes degradation of multi-species environmental DNA in lotic mesocosms[J]. Commun Biol, 2018, 1:4. doi: 10.1038/s42003-017-0005-3 pmid: 30271891
[25]	Kumar G, Eble JE, Gaither MR. A practical guide to sample preservation and pre-PCR processing of aquatic environmental DNA[J]. Mol Ecol Resour, 2020, 20(1):29-39. doi: 10.1111/men.v20.1 URL
[26]	Robinson CV, Porter TM, Wright MTG, et al. Propylene glycol-based antifreeze is an effective preservative for DNA metabarcoding of benthic arthropods[J]. Freshwater Science, 2021, 40(1):77-87. doi: 10.1086/712232 URL
[27]	Shu L, Ludwig A, Peng ZG. Standards for methods utilizing environmental DNA for detection of fish species[J]. Genes, 2020, 11(3):296. doi: 10.3390/genes11030296 URL
[28]	李雁达, 张永吉. DNA提取试剂盒的综述[J]. 全科口腔医学电子杂志, 2019, 6(28):26.
	Li YD, Zhang YJ. A review of DNA extraction kits[J]. Electron J Gen Stomatol, 2019, 6(28):26.
[29]	张瑞福, 曹慧, 崔中利, 等. 土壤微生物总DNA的提取和纯化[J]. 微生物学报, 2003, 43(2):276-282.
	Zhang RF, Cao H, Cui ZL, et al. Extraction and purification of soil microbial total DNA[J]. Acta Microbiol Sin, 2003, 43(2):276-282.
[30]	Brooks JP, Edwards DJ, Harwich MD, et al. The truth about metagenomics:quantifying and counteracting bias in 16S rRNA studies[J]. BMC Microbiol, 2015, 15(1):1-14. doi: 10.1186/s12866-014-0320-5 URL
[31]	Bell KL, Burgess KS, Okamoto KC, et al. Review and future prospects for DNA barcoding methods in forensic palynology[J]. Forensic Sci Int Genet, 2016, 21:110-116. doi: 10.1016/j.fsigen.2015.12.010 URL
[32]	Aguayo J, Fourrier-Jeandel C, Husson C, et al. Assessment of passive traps combined with high-throughput sequencing to study airborne fungal communities[J]. Appl Environ Microbiol, 2018, 84(11):e02637-17.
[33]	Banchi E, Pallavicini A, Muggia L. Relevance of plant and fungal DNA metabarcoding in aerobiology[J]. Aerobiologia, 2020, 36(1):9-23. doi: 10.1007/s10453-019-09574-2 URL
[34]	Fišer Pečnikar Ž, Buzan EV. 20 years since the introduction of DNA barcoding:from theory to application[J]. J Appl Genet, 2014, 55(1):43-52. doi: 10.1007/s13353-013-0180-y pmid: 24203863
[35]	林森杰, 王路, 郑连明, 等. 海洋生物DNA条形码研究现状与展望[J]. 海洋学报, 2014, 36(12):1-17.
	Lin SJ, Wang L, Zheng LM, et al. Current status and future prospect of DNA barcoding in marine biology[J]. Acta Oceanol Sin, 2014, 36(12):1-17.
[36]	Zhang GK, Chain FJJ, Abbott CL, et al. Metabarcoding using multiplexed markers increases species detection in complex zooplankton communities[J]. Evol Appl, 2018, 11(10):1901-1914. doi: 10.1111/eva.12694 URL
[37]	Olds BP, Jerde CL, Renshaw MA, et al. Estimating species richness using environmental DNA[J]. Ecol Evol, 2016, 6(12):4214-4226. doi: 10.1002/ece3.2016.6.issue-12 URL
[38]	Lim NKM, Tay YC, Srivathsan A, et al. Next-generation freshwater bioassessment:eDNA metabarcoding with a conserved metazoan primer reveals species-rich and reservoir-specific communities[J]. R Soc Open Sci, 2016, 3(11):160635. doi: 10.1098/rsos.160635 URL
[39]	Shaw JLA, Clarke LJ, Wedderburn SD, et al. Comparison of environmental DNA metabarcoding and conventional fish survey methods in a river system[J]. Biol Conserv, 2016, 197:131-138. doi: 10.1016/j.biocon.2016.03.010 URL
[40]	Hollingsworth PM. Refining the DNA barcode for land plants[J]. PNAS, 2011, 108(49):19451-19452. doi: 10.1073/pnas.1116812108 pmid: 22109553
[41]	Poczai P, Hyvönen J. Nuclear ribosomal spacer regions in plant phylogenetics:problems and prospects[J]. Mol Biol Rep, 2010, 37(4):1897-1912. doi: 10.1007/s11033-009-9630-3 pmid: 19626457
[42]	张宇, 郭良栋. 真菌DNA条形码研究进展[J]. 菌物学报, 2012, 31(6):809-820.
	Zhang Y, Guo LD. Progress of fungal DNA barcode[J]. Mycosystema, 2012, 31(6):809-820.
[43]	Shokralla S, Spall JL, Gibson JF, et al. Next-generation sequencing technologies for environmental DNA research[J]. Mol Ecol, 2012, 21(8):1794-1805. doi: 10.1111/j.1365-294X.2012.05538.x pmid: 22486820
[44]	van Dijk EL, Jaszczyszyn Y, et al. The third revolution in sequencing technology[J]. Trends Genet, 2018, 34(9):666-681. doi: 10.1016/j.tig.2018.05.008 URL
[45]	Piper AM, Batovska J, Cogan NOI, et al. Prospects and challenges of implementing DNA metabarcoding for high-throughput insect surveillance[J]. Gigascience, 2019, 8(8):giz092.
[46]	Edgar RC, Flyvbjerg H. Error filtering, pair assembly and error correction for next-generation sequencing reads[J]. Bioinformatics, 2015, 31(21):3476-3482. doi: 10.1093/bioinformatics/btv401 URL
[47]	Scott R, Zhan AB, Brown EA, et al. Optimization and performance testing of a sequence processing pipeline applied to detection of nonindigenous species[J]. Evol Appl, 2018, 11(6):891-905. doi: 10.1111/eva.12604 pmid: 29928298
[48]	Andruszkiewicz EA, Starks HA, Chavez FP, et al. Biomonitoring of marine vertebrates in Monterey Bay using eDNA metabarcoding[J]. PLoS One, 2017, 12(4):e0176343. doi: 10.1371/journal.pone.0176343 URL
[49]	Chain FJJ, Brown EA, MacIsaac HJ, et al. Metabarcoding reveals strong spatial structure and temporal turnover of zooplankton communities among marine and freshwater Ports[J]. Diversity Distrib, 2016, 22(5):493-504. doi: 10.1111/ddi.2016.22.issue-5 URL
[50]	Sohlberg E, Bomberg M, Miettinen H, et al. Revealing the unexplored fungal communities in deep groundwater of crystalline bedrock fracture zones in Olkiluoto, Finland[J]. Front Microbiol, 2015, 6:573. doi: 10.3389/fmicb.2015.00573 pmid: 26106376
[51]	Bienert F, De Danieli S, et al. Tracking earthworm communities from soil DNA[J]. Mol Ecol, 2012, 21(8):2017-2030. doi: 10.1111/j.1365-294X.2011.05407.x URL
[52]	Horton DJ, Kershner MW, Blackwood CB. Suitability of PCR primers for characterizing invertebrate communities from soil and leaf litter targeting metazoan 18S ribosomal or cytochrome oxidase I(COI)genes[J]. Eur J Soil Biol, 2017, 80:43-48. doi: 10.1016/j.ejsobi.2017.04.003 URL
[53]	Calvignac-Spencer S, Merkel K, et al. Carrion fly-derived DNA as a tool for comprehensive and cost-effective assessment of mammalian biodiversity[J]. Mol Ecol, 2013, 22(4):915-924. doi: 10.1111/mec.12183 pmid: 23298293
[54]	Yamamoto N, Bibby K, Qian J, et al. Particle-size distributions and seasonal diversity of allergenic and pathogenic fungi in outdoor air[J]. Isme J, 2012, 6(10):1801-1811. doi: 10.1038/ismej.2012.30 URL
[55]	Nicolaisen M, West JS, Sapkota R, et al. Fungal communities including plant pathogens in near surface air are similar across northwestern Europe[J]. Front Microbiol, 2017, 8:1729. doi: 10.3389/fmicb.2017.01729 pmid: 28943873
[56]	Kraaijeveld K, de Weger LA, Ventayol García M, et al. Efficient and sensitive identification and quantification of airborne pollen using next-generation DNA sequencing[J]. Mol Ecol Resour, 2015, 15(1):8-16. doi: 10.1111/1755-0998.12288 pmid: 24893805
[57]	Leese F, Bouchez A, Abarenkov K, et al. Why we need sustainable networks bridging countries, disciplines, cultures and generations for aquatic biomonitoring 2. 0:A perspective derived from the DNAqua-net COST action[M]// Next Generation Biomonitoring:Part 1. Amsterdam:Elsevier, 2018:63-99.
[58]	Thomsen PF, Kielgast J, Iversen LL, et al. Detection of a diverse marine fish fauna using environmental DNA from seawater samples[J]. PLoS One, 2012, 7(8):e41732. doi: 10.1371/journal.pone.0041732 URL
[59]	Civade R, Dejean T, Valentini A, et al. Spatial representativeness of environmental DNA metabarcoding signal for fish biodiversity assessment in a natural freshwater system[J]. PLoS One, 2016, 11(6):e0157366. doi: 10.1371/journal.pone.0157366 URL
[60]	Ortega A, Geraldi NR, Duarte CM. Environmental DNA identifies marine macrophyte contributions to Blue Carbon sediments[J]. Limnol Oceanogr, 2020, 65(12):3139-3149. doi: 10.1002/lno.v65.12 URL
[61]	Rivera SF, Vasselon V, et al. DNA metabarcoding and microscopic analyses of sea turtles biofilms:Complementary to understand turtle behavior[J]. PLoS One, 2018, 13(4):e0195770. doi: 10.1371/journal.pone.0195770 URL
[62]	Lopes CM, Baêta D, Valentini A, et al. Lost and found:Frogs in a biodiversity hotspot rediscovered with environmental DNA[J]. Mol Ecol, 2020: mec. 15594.
[63]	Takeuchi A, Sado T, Gotoh RO, et al. New PCR primers for metabarcoding environmental DNA from freshwater eels, genus Anguilla[J]. Sci Rep, 2019, 9(1):7977. doi: 10.1038/s41598-019-44402-0 pmid: 31138865
[64]	Song JW, Small MJ, Casman EA. Making sense of the noise:The effect of hydrology on silver carp eDNA detection in the Chicago area waterway system[J]. Sci Total Environ, 2017, 605/606:713-720. doi: 10.1016/j.scitotenv.2017.06.255 URL
[65]	马竹欣. 利用环境DNA技术调查入侵种克氏原螯虾在元阳梯田的分布[D]. 昆明:云南大学, 2016.
	Ma ZX. Distribution of invasive crayfish Procambarus clarkii in Yuanyang terrace revealed by eDNA[D]. Kunming:Yunnan University, 2016.
[66]	Willerslev E, Cappellini E, Boomsma W, et al. Ancient biomolecules from deep ice cores reveal a forested southern Greenland[J]. Science, 2007, 317(5834):111-114. pmid: 17615355
[67]	Willerslev E. Diverse plant and animal genetic records from Holocene and Pleistocene sediments[J]. Science, 2003, 300(5620):791-795. pmid: 12702808
[68]	Pedersen MW, Ruter A, Schweger C, et al. Postglacial viability and colonization in North America’s ice-free corridor[J]. Nature, 2016, 537(7618):45-49. doi: 10.1038/nature19085 URL
[69]	Guardiola M, Uriz MJ, Taberlet P, et al. Deep-sea, deep-sequencing:metabarcoding extracellular DNA from sediments of marine canyons[J]. PLoS One, 2015, 10(10):e0139633. doi: 10.1371/journal.pone.0139633 URL
[70]	Guardiola M, Wangensteen OS, Taberlet P, et al. Spatio-temporal monitoring of deep-sea communities using metabarcoding of sediment DNA and RNA[J]. PeerJ, 2016, 4:e2807. doi: 10.7717/peerj.2807 URL
[71]	Sinniger F, Pawlowski J, Harii S, et al. Worldwide analysis of sedimentary DNA reveals major gaps in taxonomic knowledge of deep-sea benthos[J]. Front Mar Sci, 2016, 3:1-14.
[72]	Clark DE, Pilditch CA, Pearman JK, et al. Environmental DNA metabarcoding reveals estuarine benthic community response to nutrient enrichment - Evidence from an in situ experiment[J]. Environ Pollut, 2020, 267:115472. doi: S0269-7491(20)36160-1 pmid: 32891048
[73]	Xie Y, Zhang X, Yang J, et al. eDNA-based bioassessment of coastal sediments impacted by an oil spill[J]. Environ Pollut, 2018, 238:739-748. doi: 10.1016/j.envpol.2018.02.081 URL
[74]	张宛宛. 基于DNA宏条形码技术的浮游植物群落多样性监测研究[D]. 南京:南京大学, 2017.
	Zhang WW. Study on biodiversity monitoring of phytoplankton community based on DNA barcoding technology[D]. Nanjing:Nanjing University, 2017.
[75]	Buée M, Reich M, Murat C, et al. 454 Pyrosequencing analyses of forest soils reveal an unexpectedly high fungal diversity[J]. New Phytol, 2009, 184(2):449-456. doi: 10.1111/j.1469-8137.2009.03003.x pmid: 19703112
[76]	Andersen K, Bird KL, Rasmussen M, et al. Meta-barcoding of ‘dirt’ DNA from soil reflects vertebrate biodiversity[J]. Mol Ecol, 2012, 21(8):1966-1979. doi: 10.1111/j.1365-294X.2011.05261.x pmid: 21917035
[77]	于水强, 王文娟, B. Larry Li. 环境DNA技术在地下生态学中的应用[J]. 生态学报, 2015, 35(15):4968-4976.
	Yu SQ, Wang WJ, Li B. Applied environmental DNA technology to study underground ecology[J]. Acta Ecol Sin, 2015, 35(15):4968-4976.
[78]	Lamb EG, Winsley T, Piper CL, et al. A high-throughput belowground plant diversity assay using next-generation sequencing of the trnL intron[J]. Plant Soil, 2016, 404(1/2):361-372. doi: 10.1007/s11104-016-2852-y URL
[79]	Khaliq I, Hardy GESJ, White D, et al. eDNA from roots:a robust tool for determining Phytophthora communities in natural ecosystems[J]. PFEMS Microbiol Ecol, 2018, 94(5):fiy048.
[80]	Singer D, Duckert C, et al. High-throughput sequencing of litter and moss eDNA reveals a positive correlation between the diversity of Apicomplexa and their invertebrate hosts across alpine habitats[J]. Soil Biol Biochem, 2020, 147:107837. doi: 10.1016/j.soilbio.2020.107837 URL
[81]	Schnell IB, Thomsen PF, Wilkinson N, et al. Screening mammal biodiversity using DNA from leeches[J]. Curr Biol, 2012, 22(8):R262-R263. doi: 10.1016/j.cub.2012.02.058 URL
[82]	Thomsen PF, Sigsgaard EE. Environmental DNA metabarcoding of wild flowers reveals diverse communities of terrestrial arthropods[J]. Ecol Evol, 2019, 9(4):1665-1679. doi: 10.1002/ece3.2019.9.issue-4 URL
[83]	Yan DF, Mills JG, Gellie NJC, et al. High-throughput eDNA monitoring of fungi to track functional recovery in ecological restoration[J]. Biol Conserv, 2018, 217:113-120. doi: 10.1016/j.biocon.2017.10.035 URL
[84]	Wangensteen OS, Cebrian E, Palacín C, et al. Under the canopy:Community-wide effects of invasive algae in Marine Protected Areas revealed by metabarcoding[J]. Mar Pollut Bull, 2018, 127:54-66. doi: S0025-326X(17)30990-6 pmid: 29475694
[85]	Smith MB, Rocha AM, Smillie CS, et al. Natural bacterial communities serve as quantitative geochemical biosensors[J]. mBio, 2015, 6(3):e00326-e00315.
[86]	Coelho FJ, Cleary DF, Costa R, et al. Multitaxon activity profiling reveals differential microbial response to reduced seawater pH and oil pollution[J]. Mol Ecol, 2016, 25(18):4645-4659. doi: 10.1111/mec.2016.25.issue-18 URL
[87]	Frontalini F, Greco M, Di Bella L, et al. Assessing the effect of mercury pollution on cultured benthic foraminifera community using morphological and eDNA metabarcoding approaches[J]. Mar Pollut Bull, 2018, 129(2):512-524. doi: S0025-326X(17)30840-8 pmid: 29033170
[88]	An C, Woo C, Yamamoto N. Introducing DNA-based methods to compare fungal microbiota and concentrations in indoor, outdoor, and personal air[J]. Aerobiologia, 2018, 34(1):1-12. doi: 10.1007/s10453-017-9490-6 URL
[89]	West JS, Atkins SD, Emberlin J, et al. PCR to predict risk of airborne disease[J]. Trends Microbiol, 2008, 16(8):380-387. doi: 10.1016/j.tim.2008.05.004 pmid: 18595713
[90]	Mahaffee WF, Stoll R. The ebb and flow of airborne pathogens:monitoring and use in disease management decisions[J]. Phytopathology, 2016, 106(5):420-431. doi: 10.1094/PHYTO-02-16-0060-RVW pmid: 27003505
[91]	Degois J, Clerc F, Simon X, et al. First metagenomic survey of the microbial diversity in bioaerosols emitted in waste sorting plants[J]. Ann Work Expo Health, 2017, 61(9):1076-1086.
[92]	Kumari P, Woo C, Yamamoto N, et al. Variations in abundance, diversity and community composition of airborne fungi in swine houses across seasons[J]. Sci Rep, 2016, 6:37929. doi: 10.1038/srep37929 URL
[93]	Yang W, Guo M, et al. Detection and analysis of fine particulate matter and microbial aerosol in chicken houses in Shandong Province, China[J]. Poult Sci, 2018, 97(3):995-1005. doi: 10.3382/ps/pex388 URL
[94]	Tong X, Xu H, Zou L, et al. High diversity of airborne fungi in the hospital environment as revealed by meta-sequencing-based microbiome analysis[J]. Sci Rep, 2017, 7:39606. doi: 10.1038/srep39606 URL
[95]	Korpelainen H, Pietiläinen M. Biodiversity of pollen in indoor air samples as revealed by DNA metabarcoding[J]. Nord J Bot, 2017, 35(5):602-608. doi: 10.1111/njb.01623 URL
[96]	Coombs K, Taft D, et al. Variability of indoor fungal microbiome of green and non-green low-income homes in Cincinnati, Ohio[J]. Sci Total Environ, 2018, 610/611:212-218. doi: 10.1016/j.scitotenv.2017.07.274 URL
[97]	Deiner K, Bik HM, Mächler E, et al. Environmental DNA metabarcoding:Transforming how we survey animal and plant communities[J]. Mol Ecol, 2017, 26(21):5872-5895. doi: 10.1111/mec.2017.26.issue-21 URL
[98]	Glenn TC. Field guide to next-generation DNA sequencers[J]. Mol Ecol Resour, 2011, 11(5):759-769. doi: 10.1111/j.1755-0998.2011.03024.x URL
[99]	Darling JA, Mahon AR. From molecules to management:adopting DNA-based methods for monitoring biological invasions in aquatic environments[J]. Environ Res, 2011, 111(7):978-988. doi: 10.1016/j.envres.2011.02.001 pmid: 21353670
[100]	Miya M, Minamoto T, Yamanaka H, et al. Use of a filter cartridge for filtration of water samples and extraction of environmental DNA[J]. JoVE, 2016(117):54741.
[101]	Jain M, Fiddes IT, Miga KH, et al. Improved data analysis for the MinION nanopore sequencer[J]. Nat Methods, 2015, 12(4):351-356. doi: 10.1038/NMETH.3290
[102]	Abad D, Albaina A, Aguirre M, et al. Is metabarcoding suitable for estuarine plankton monitoring? A comparative study with microscopy[J]. Mar Biol, 2016, 163(7):1-13. doi: 10.1007/s00227-015-2782-x URL
[103]	Yang J, Zhang X, Xie Y, et al. Zooplankton community profiling in a eutrophic freshwater ecosystem-lake Tai basin by DNA metabarcoding[J]. Sci Rep, 2017, 7(1):1773. doi: 10.1038/s41598-017-01808-y URL
[104]	Šigut M, Kostovčík M, Šigutová H, et al. Performance of DNA metabarcoding, standard barcoding, and morphological approach in the identification of host-parasitoid interactions[J]. PLoS One, 2017, 12(12):e0187803. doi: 10.1371/journal.pone.0187803 URL
[105]	Ryan U, Paparini A, Oskam C. New technologies for detection of enteric parasites[J]. Trends Parasitol, 2017, 33(7):532-546. doi: S1471-4922(17)30083-1 pmid: 28385423
[106]	Shi ZY, Yang CQ, Hao MD, et al. FuzzyID2:a software package for large data set species identification via barcoding and metabarcoding using hidden Markov models and fuzzy set methods[J]. Mol Ecol Resour, 2018, 18(3):666-675. doi: 10.1111/men.2018.18.issue-3 URL
[107]	Takahara T, Minamoto T, et al. Estimation of fish biomass using environmental DNA[J]. PLoS One, 2012, 7(4):e35868. doi: 10.1371/journal.pone.0035868 URL
[108]	Ushio M, Masayuki H, Murakami R, et al. Quantitative monitoring of multispecies fish environmental DNA using high-throughput sequencing[J]. Metabarcoding and Metagenomics, 2018, 2:e23297.
[109]	Yamamoto S, Minami K, Fukaya K, et al. Environmental DNA as a ‘snapshot’ of fish distribution:a case study of Japanese jack mackerel in maizuru bay, sea of Japan[J]. PLoS One, 2016, 11(3):e0149786. doi: 10.1371/journal.pone.0149786 URL
[110]	Raemy M, Ursenbacher S. Detection of the European pond turtle(Emys orbicularis)by environmental DNA:is eDNA adequate for reptiles?[J]. Amphibia-Reptilia, 2018, 39(2):135-143. doi: 10.1163/15685381-17000025 URL
[111]	Goldberg CS, Strickler KM, Fremier AK. Degradation and dispersion limit environmental DNA detection of rare amphibians in wetlands:Increasing efficacy of sampling designs[J]. Sci Total Environ, 2018, 633:695-703. doi: 10.1016/j.scitotenv.2018.02.295 URL
[112]	Elbrecht V, Leese F. Can DNA-based ecosystem assessments quantify species abundance? testing primer bias and biomass——sequence relationships with an innovative metabarcoding protocol[J]. PLoS One, 2015, 10(7):e0130324. doi: 10.1371/journal.pone.0130324 URL
[113]	Coble AA, Flinders CA, Homyack JA, et al. eDNA as a tool for identifying freshwater species in sustainable forestry:a critical review and potential future applications[J]. Sci Total Environ, 2019, 649:1157-1170. doi: 10.1016/j.scitotenv.2018.08.370 URL
[114]	李飞龙, 杨江华, 等. 环境DNA宏条形码监测水生态系统变化与健康状态[J]. 中国环境监测, 2018, 34(6):37-46.
	Li FL, Yang JH, et al. Using environmental DNA metabarcoding to monitor the changes and health status of aquatic ecosystems[J]. Environ Monit China, 2018, 34(6):37-46.