Comparison of fish communities using environmental DNA metabarcoding and capture methods in a plateau Erhai Lake,China

Environmental DNA(eDNA)has been used as an important tool for fish diversity analysis,which can greatly solve the problems in traditional survey methodology.However,little work has been done on the actual monitoring accuracy of eDNA.In this study,we analyzed the current status of fish resources in Erhai Lake in Yunnan,SW China,by dividing the lake into three sectors according to habitat differences,and compared the results of eDNA and traditional capture methods to investigate the shortcomings of the current analysis of eDNA results.A total of 27 fish species were detected by eDNA and traditional capture methods,including 20 and 19 fish species,respectively,and additional differences in fish composition between the two methods.The alpha diversity showed higher fish abundance and lower fish diversity by eDNA method compared to the traditional capture method,demonstrating that eDNA was not superior for use in fish diversity analysis.Fish community similarity analysis showed that community differences were generally significant for eDNA(P<0.05).RDA analysis indicated that environmental factors did not significantly affect fish communities monitored by the eDNA method.However,water temperature,aquatic plants,and water depth had significant(P<0.05)effects on fish communities in the traditional capture method,suggesting that eDNA results are insensitive to the effects of environmental factors.Our results illustrate the effectiveness of eDNA in fish identification and the issues in quantification compared to traditional capture methods.Therefore,combining eDNA with traditional methods is a more effective method for analyzing eDNA metabarcoding,following which the protocols of both quantitative methods can be designed to explore the regularity of eDNA quantification.

Epiphytic zooplankton community profiles in a typical urban wetland as revealed by DNA metabarcoding

Zooplankton,a crucial component of urban wetland,are one of the effective bioindicators for monitoring the feeding stocks of organisms at higher trophic levels and assessing the ecological quality of ecosystems.However,information about the characteristics of epiphytic zooplankton community structure resulted from traditional methods is limited and hindered by the large amount of detritus and sludge attached to the macrophytes.We investigated the epiphytic zooplankton communities associated with macrophytes(Vallisneria,Nymphaea,and Thalia dealbata)in a subtropical wetland using as DNA markers of the 18 S rRNA gene and the mitochondrial cytochrome c oxidase subunit I(COI)gene.A total of 241 OTUs of zooplankton were obtained from COI amplicons,including 194 OTUs of Rotifera,22 of Cladocera,and 25 of Copepoda,while only 62 OTUs of zooplankton were obtained from 18 S rDNA amplicons including 34 OTUs of Rotifera and 28 of Copepoda.The zooplankton communities associated with the three macrophytes were similar,but they differed significantly from those in the open waters.However,there were no significant temporal differences among the zooplankton communities.Epiphytic zooplankton communities were dominated by littoral zooplankton such as Testudinella,Lecane,and Philodina.Microzooplankton,especially littoral species,utilize macrophytes as food sources and as refuges against predation.This further led to an increase inαandβdiversity of zooplankton communities in urban wetlands.Our result suggests that the joint use of multiple molecular markers could improve the taxonomic resolution and generate a comprehensive biodiversity profile of zooplankton.

Methodology for fish biodiversity monitoring with environmental DNA metabarcoding:The primers,databases and bioinformatic pipelines

Environmental DNA(eDNA)originates from cellular material shed by organisms into aquatic or terrestrial environments and can be sampled and monitored using metabarcoding technology,which is revolutionizing fish biodiversity monitoring.Several reviews concerning fish eDNA have focused on standard sampling methods and its applications,though a systematic review focused on marker genes,databases,and bioinformatic pipelines has not yet been published.Here,we present a comprehensive literature review of studies applying metabarcoding technology to fish eDNA for the purpose of fish biodiversity monitoring.We systematically provide the available universal primers used to amplify barcoding sequences from fish eDNA,and then discuss reference barcoding databases,relevant bioinformatic analyses,as well as developed pipelines.The performances of universal primers and their relevant reference databases are summarized.Combined use of multiple primer pairs targeted for more than one gene marker(e.g.,12S,16S,Cytb,COI),and use of both local and public databases are recommended as approaches to improve the sensitivity and reliability of fish eDNA analyses.We also compare the effectiveness of eDNA metabarcoding to traditional approaches for monitoring fish biodiversity and highlight challenges and future perspectives associated with this new tool.Ultimately,we advocate for greater incorporation of eDNA analysis into fish biodiversity assessments to assist environmental managers.

Environmental barcoding of the ectomycorrhizal fungal genus Cortinarius

Understanding the role of ectomycorrhizal fungi in plant communities is hampered by a lack of knowledge about fungal diversity.DNA barcoding of the ectomycorrhizal fungal genus Cortinarius was used to compare fungal diversity in soil from four plant communities:(i)Nothofagus forest(where Cortinarius is common and diverse),(ii)Kunzea forest(where Cortinarius is present but with low diversity),(iii)a Pinus radiata plantation(Cortinarius is not thought to be present)and(iv)a sub-Antarctic island(where known ectomycorrhizal hosts are absent).PCR primers specific for the ITS region of Cortinarius species were developed.Specificity was tested in vitro and in silico against DNA from basidiocarps of Cortinarius and non-Cortinarius species.The primers were tested for their ability to amplify Cortinarius DNA in soil from forests of the three ectomycorrhizal forest communities and a range of soils from the ectomycorrhiza-free subantarctic Campbell Island.High diversity of Cortinarius was associated with soil of all three ectomycorrhizal communities,despite Cortinarius being previously unrecorded from Pinus.Soil from all three communities share some ectomycorrhizal fungi(including fungi shared between native and exotic hosts),having implications for community succession,introduction of exotic fungi and biodiversity assessment.No Cortinarius was detected from Campbell Island samples.The validated molecular protocol assessed species diversity in a rapid and cost effective way.Baseline biodiversity assessment based on DNA barcoding is more effective at detecting diversity than traditional methods,but requires careful consideration of the difference between ectomycorrhizal fungal diversity in soil versus root-tips.

Stable oligonucleotide-functionalized gold nanosensors for environmental biocontaminant monitoring

The global propagation of environmental biocontaminants such as antibiotic resistant pathogens and their antibiotic resistance genes（ARGs） is a public health concern that highlights the need for improved monitoring strategies. Here, we demonstrate the environmental stability and applicability of an oligonucleotide-functionalized gold nanosensor. The mec A ARG was targeted as model biocontaminant due to its presence in clinically-relevant pathogens and to its emergence as an environmental contaminant.mec A-specific nanosensors were tested for antibiotic resistance gene（ARG） detection in ARG-spiked effluent from four wastewater treatment plants（WWTPs）. The mec A-specific nanosensors showed stability in environmental conditions and in high ionic strength（[MgCl_2] 〈 50 m M）, and high selectivity against mismatched targets. Spectrophotometric detection was reproducible with an LOD of 70 pM（≈ 4 × 10~7 genes/μL）, even in the presence of interferences associated with non-target genomic DNA and complex WWTP effluent. This contribution supports the environmental applicability of a new line of cost-effective, field-deployable tools needed for wide-scale biocontaminant monitoring.

Biodiversity of freshwater ciliates(Protista,Ciliophora)in the Lake Weishan Wetland,China:the state of the

Ciliates are core components of the structure of and function of aquatic microbial food webs.They play an essential role in the energy flow and material circulation within aquatic ecosystems.However,studies on the taxonomy and biodiversity of freshwater ciliates,especially those in wetlands in China are limited.To address this issue,a project to investigate the freshwater ciliates of the Lake Weishan Wetland,Shandong Province,commenced in 2019.Here,we summarize our findings to date on the diversity of ciliates.A total of 187 ciliate species have been found,94 of which are identified to species-level,87 to genus-level,and six to family-level.These species show a high morphological diversity and represent five classes,i.e.,Heterotrichea,Litostomatea,Prostomatea,Oligohymenophorea,and Spirotrichea.The largest number of species documented are oligohymenophoreans.A comprehensive database of these ciliates,including morphological data,gene sequences,microscope slide specimens and a DNA bank,has been established.In the present study,we provide an annotated checklist of retrieved ciliates as well as information on the sequences of published species.Most of these species are recorded in China for the first time and more than 20%are tentatively identified as new to science.Additionally,an investigation of environmental DNA revealed that the ciliate species diversity in Lake Weishan Wetland is higher than previously supposed.

Impact of anthropogenic activities on changes of ichthyofauna in the middle and lower Xiang River

With the population increasing,fish communities,as important components in the river system,were inescapably impacted by human activities.To investigate these impacts,we characterized the fish community’s composition by environmental DNA metabarcoding in the urban and suburban sites along Xiang River in central China.The results showed that 32 species were identified,and most species were from Cypriniform.The analysis indicated no significant differences in fish communities among the three urban areas.However,significant differences were observed between the bank solidified and non-solidified areas,which indicated that the channelization significantly influenced the biodiversity and abundances.An alien species,Clarias gariepinus,was discovered in all examined sites,and it was negatively associated with ten native species.Furthermore,human populations and acidification were also negatively related to diversity.Our findings clearly showed that the consequences of anthropogenic activities shaped ichthyofauna and caused biodiversity loss in the urban river.