Previously, we showed that prophylactic addition of glucose to Harsha Lake water samples could inhibit cyanobacteria growth, at least for a short period of time. The current study tested cyanobacterial control with gl...Previously, we showed that prophylactic addition of glucose to Harsha Lake water samples could inhibit cyanobacteria growth, at least for a short period of time. The current study tested cyanobacterial control with glucose for the entire Harsha Lake bloom season. Water samples (1000 ml) were collected weekly from Harsha Lake during the algal-bloom season starting June 9 and lasting until August 24, 2022. To each of two 7-liter polypropylene containers, 500 ml of Harsha Lake water was added, and the containers were placed in a controlled environment chamber. To one container labeled “Treated,” 0.15 g of glucose was added, and nothing was added to the container labeled “Control.” After that, three 25 ml samples from each container were collected and used for 16S rRNA gene sequencing each week. Then 1000 ml of Harsha Lake water was newly collected each week, with 500 ml added to each container, along with the addition of 0.15 g glucose to the “Treated” container. Sequencing data were used to examine differences in the composition of bacterial communities between Treated and Control containers. Treatment with glucose altered the microbial communities by 1) reducing taxonomic diversity, 2) largely eliminating cyanobacterial taxa, and 3) increasing the relative abundance of subsets of non-cyanobacterial taxa (such as Proteobacteria and Actinobacteriota). These effects were observed across time despite weekly inputs derived directly from Lake water. The addition of glucose to a container receiving weekly additions of Lake water suppressed the cyanobacterial populations during the entire summer bloom season. The glucose appears to stimulate the diversity of certain bacterial taxa at the expense of the cyanobacteria.展开更多
The diazotrophic community in biological soil crusts(biocrusts)is the key supplier of nitrogen in dryland.To date,there is still limited information on how biocrust development influences the succession of diazotrophi...The diazotrophic community in biological soil crusts(biocrusts)is the key supplier of nitrogen in dryland.To date,there is still limited information on how biocrust development influences the succession of diazotrophic community,and what are the most important factors mediating diazotrophic communities during biocrust succession.Using the high throughput nifH amplicon sequencing,the diazotrophs in soils at different developmental stages of biocrust were comparatively studied.The results evidenced the decreases of TOC/TN ratio and pH value with biocrust development.Nostoc and Scytonema were the most dominant diazotrophic genera at all biocrust stages,while Azospirillum and Bradyrhizobium were abundant only in bare soil.Diazotrophic co-occurrence networks tended to be less complex and less connected with biocrust succession.The soil TOC/TN ratio was the most dominant factor mediating diazotrophic diversity,community composition and assembly processes,while diazotrophic-diversity and NO3–-N/NH4+-N ratio were positively correlated with the nitrogenase activity during biocrust succession.This study provided novel understandings of nitrogen fixation and succession patterns of diazotrophic community,by showing the effects of biocrust succession on diazotrophic diversity,community composition,community assembly and co-occurrence networks,and recognizing TOC/TN ratio as the most dominant factor mediating diazotrophs during biocrust succession.展开更多
Objective Viral encephalitis is an infectious disease severely affecting human health.It is caused by a wide variety of viral pathogens,including herpes viruses,flaviviruses,enteroviruses,and other viruses.The laborat...Objective Viral encephalitis is an infectious disease severely affecting human health.It is caused by a wide variety of viral pathogens,including herpes viruses,flaviviruses,enteroviruses,and other viruses.The laboratory diagnosis of viral encephalitis is a worldwide challenge.Recently,high-throughput sequencing technology has provided new tools for diagnosing central nervous system infections.Thus,In this study,we established a multipathogen detection platform for viral encephalitis based on amplicon sequencing.Methods We designed nine pairs of specific polymerase chain reaction(PCR)primers for the 12 viruses by reviewing the relevant literature.The detection ability of the primers was verified by software simulation and the detection of known positive samples.Amplicon sequencing was used to validate the samples,and consistency was compared with Sanger sequencing.Results The results showed that the target sequences of various pathogens were obtained at a coverage depth level greater than 20×,and the sequence lengths were consistent with the sizes of the predicted amplicons.The sequences were verified using the National Center for Biotechnology Information BLAST,and all results were consistent with the results of Sanger sequencing.Conclusion Amplicon-based high-throughput sequencing technology is feasible as a supplementary method for the pathogenic detection of viral encephalitis.It is also a useful tool for the high-volume screening of clinical samples.展开更多
With the gradual maturity of sequencing technology,many microbiome studies have published,driving the emergence and advance of related analysis tools.R language is the widely used platform for microbiome data analysis...With the gradual maturity of sequencing technology,many microbiome studies have published,driving the emergence and advance of related analysis tools.R language is the widely used platform for microbiome data analysis for powerful functions.However,tens of thousands of R packages and numerous similar analysis tools have brought major challenges for many researchers to explore microbiome data.How to choose suitable,efficient,convenient,and easy-to-learn tools from the numerous R packages has become a problem for many microbiome researchers.We have organized 324 common R packages for microbiome analysis and classified them according to application categories(diversity,difference,biomarker,correlation and network,functional prediction,and others),which could help researchers quickly find relevant R packages for microbiome analysis.Furthermore,we systematically sorted the integrated R packages(phyloseq,microbiome,MicrobiomeAnalystR,Animalcules,microeco,and amplicon)for microbiome analysis,and summarized the advantages and limitations,which will help researchers choose the appropriate tools.Finally,we thoroughly reviewed the R packages for microbiome analysis,summarized most of the common analysis content in the microbiome,and formed the most suitable pipeline for microbiome analysis.This paper is accompanied by hundreds of examples with 10,000 lines codes in GitHub,which can help beginners to learn,also help analysts compare and test different tools.This paper systematically sorts the application of R in microbiome,providing an important theoretical basis and practical reference for the development of better microbiome tools in the future.All the code is available at GitHub github.com/taowenmicro/EasyMicrobiomeR.展开更多
The monkeypox virus(MPXV)has triggered a current outbreak globally.Genome sequencing of MPXV and rapid tracing of genetic variants will benefit disease diagnosis and control.It is a significant challenge but necessary...The monkeypox virus(MPXV)has triggered a current outbreak globally.Genome sequencing of MPXV and rapid tracing of genetic variants will benefit disease diagnosis and control.It is a significant challenge but necessary to optimize the strategy and application of rapid full-length genome identification and to track variations of MPXV in clinical specimens with low viral loads,as it is one of the DNA viruses with the largest genome and the most AT-biased,and has a significant number of tandem repeats.Here we evaluated the performance of metagenomic and amplicon sequencing techniques,and three sequencing platforms in MPXV genome sequencing based on multiple clinical specimens of five mpox cases in Chinese mainland.We rapidly identified the full-length genome of MPXV with the assembly of accurate tandem repeats in multiple clinical specimens.Amplicon sequencing enables cost-effective and rapid sequencing of clinical specimens to obtain high-quality MPXV genomes.Third-generation sequencing facilitates the assembly of the terminal tandem repeat regions in the monkeypox virus genome and corrects a common misassembly in published sequences.Besides,several intra-host single nucleotide variations were identified in the first imported mpox case.This study offers an evaluation of various strategies aimed at identifying the complete genome of MPXV in clinical specimens.The findings of this study will significantly enhance the surveillance of MPXV.展开更多
Insecticide resistance in Panonychus citri is a major obstacle to mite control in citrus orchards.Pyrethroid insecticides are continually used to control mites in China,although resistance to pyrethroids has evolved i...Insecticide resistance in Panonychus citri is a major obstacle to mite control in citrus orchards.Pyrethroid insecticides are continually used to control mites in China,although resistance to pyrethroids has evolved in some populations.Here,the resistance to the pyrethroid fenpropathrin was investigated and 7 out of 8 field-collected populations of P citri exhibited a high level of resistance,ranging from 171-fold to 15391-fold higher than the susceptible(SS)comparison strain.Three voltage-gated sodium channel(VGSC)mutations were identified in the tested populations:L1031V,F1747L,and F17511.Amplicon sequencing was used to evaluate the frequency of these mutations in the 19 field populations.L1031V and F1751I were present in all populations at frequencies of 11.6%-82.1%and 0.5%-31.8%,respectively,whereas the F1747L mutation was only present in 12 populations from Chongqing,Sichuan,Guangxi,and Yunnan provinces.Introduction of these mutations singly or in combination into transgenic flies significantly increased their resistance to fenpropathrin and these flies also exhibited reduced mortality after exposure to the pyrethroids permethrin andβ-cypermethrin.Panonychus citri VGSC homology modeling and ligand docking indicate that F1747 and F1751 form direct binding contacts with pyrethroids,which are lost with mutation,whereas L1031 mutation may diminish pyrethroid effects through an allosteric mechanism.Overall,the results provide molecular markers for monitoring pest resistance to pyrethroids and offer new insights into the basis of pyrethroid actions on sodium channels.展开更多
Notoginseng Radix et Rhizoma(Sanqi in Chinese)is a precious traditional Chinese herbal medicine.It has the effect of dispersing blood stasis and stopping bleeding,reducing swelling and fixing pain.However,it tends to ...Notoginseng Radix et Rhizoma(Sanqi in Chinese)is a precious traditional Chinese herbal medicine.It has the effect of dispersing blood stasis and stopping bleeding,reducing swelling and fixing pain.However,it tends to contaminate with harmful fungi during storage,which may make it much less effective.In order to understand the fungal contamination of Notoginseng Radix et Rhizoma and master its composition of the exogenous fungi.The surface fungi of Notoginseng Radix et Rhizoma samples collected from six Chinese provinces and districts were investigated by using dilution plate method.Detection of aflatoxins by UPLC-MS/MS.The results showed that Penicillium citrinum was dominantly isolated from Notoginseng Radix et Rhizoma samples from No.1 to No.4.Aspergillus flavus,which produces aflatoxin,was dominantly isolated from Notoginseng Radix et Rhizoma samples from No.5 and No.6.In addition,kinds of mycotoxin were assayed which were produced by three of those identified A.flavus.All three fungi strains produced aflatoxin B1(AFB1)and one strain HBSQ1-5 additionally produced other three kinds of mycotoxin,AFB2,AFG1 and AFG2.It is the results implied that it will be very important to take serious cautions when using Notoginseng Radix et Rhizoma.As well as,understanding the composition of the exogenous fungi of Notoginseng Radix et Rhizoma and the strains of toxin-producing fungi,which can play an important role in guiding the storage of Notoginseng Radix et Rhizoma.展开更多
Studies have shown that gut microbiota metabolites can enter the central nervous system via the blood-spinal cord barrier and cause neuroinflammation, thus constituting secondary injury after spinal cord injury. To in...Studies have shown that gut microbiota metabolites can enter the central nervous system via the blood-spinal cord barrier and cause neuroinflammation, thus constituting secondary injury after spinal cord injury. To investigate the correlation between gut microbiota and metabolites and the possible mechanism underlying the effects of gut microbiota on secondary injury after spinal cord injury, in this study, we established mouse models of T8–T10 traumatic spinal cord injury. We used 16 S rRNA gene amplicon sequencing and metabolomics to reveal the changes in gut microbiota and metabolites in fecal samples from the mouse model. Results showed a severe gut microbiota disturbance after spinal cord injury, which included marked increases in pro-inflammatory bacteria, such as Shigella, Bacteroides, Rikenella, Staphylococcus, and Mucispirillum and decreases in anti-inflammatory bacteria, such as Lactobacillus, Allobaculum, and Sutterella. Meanwhile, we identified 27 metabolites that decreased and 320 metabolites that increased in the injured spinal cord. Combined with pathway enrichment analysis, five markedly differential amino acids(L-leucine, L-methionine, L-phenylalanine, L-isoleucine and L-valine) were screened out, which play a pivotal role in activating oxidative stress and inflammatory responses following spinal cord injury. Integrated correlation analysis indicated that the alteration of gut microbiota was related to the differences in amino acids, which suggests that disturbances in gut microbiota might participate in the secondary injury through the accumulation of partial metabolites that activate oxidative stress and inflammatory responses. Findings from this study provide a new theoretical basis for improving the secondary injury after spinal cord injury through fecal microbial transplantation.展开更多
High-throughput sequencing of amplicons has been widely used to precisely and efficiently identify species compositions and analyze community structures,greatly promoting biological studies involving large amounts of ...High-throughput sequencing of amplicons has been widely used to precisely and efficiently identify species compositions and analyze community structures,greatly promoting biological studies involving large amounts of complex samples,especially those involving environmental and pathogen-monitoring ones.Commercial library preparation kits for amplicon sequencing,which generally require multiple steps,including adapter ligation and indexing,are expensive and time-consuming,especially for applications at a large scale.To overcome these limitations,a“one-step PCR approach”has been previously proposed for constructions of amplicon libraries using long fusion primers.However,efficient amplifications of target genes and accurate demultiplexing of pooled sequencing data remain to be addressed.To tackle these,we present an integrative protocol for one-step PCR amplicon library construction(OSPALC).High-quality reads have been generated by this approach to reliably identify species compositions of mock bacterial communities and environmental samples.With this protocol,the amplicon library is constructed through one regular PCR with long primers,and the total cost per DNA/cDNA sample decreases to just 7%of the typical cost via the multi-step PCR approach.Empirically tested primers and optimized PCR conditions to construct OSPALC libraries for 16S rDNA V4 regions are demonstrated as a case study.Tools to design primers targeting at any genomic regions are also presented.In principle,OSPALC can be readily applied to construct amplicon libraries of any target genes using DNA or RNA samples,and will facilitate research in numerous fields.展开更多
Environmental pollution and the spread of pathogenic microorganisms pose a significant threat to the health of humans and the planet.Thus,understanding and detecting microorganisms is crucial for maintaining a healthy...Environmental pollution and the spread of pathogenic microorganisms pose a significant threat to the health of humans and the planet.Thus,understanding and detecting microorganisms is crucial for maintaining a healthy living environment.Nanopore sequencing is a single-molecule detection method developed in the 1990s that has revolutionized various research fields.It offers several advantages over traditional sequencing methods,including low cost,label-free,time-saving detection speed,long sequencing reading,real-time monitoring,convenient carrying,and other significant advantages.In this review,we summarize the technical principles and characteristics of nanopore sequencing and discuss its applications in amplicon sequencing,metagenome sequencing,and whole-genome sequencing of environmental microorganisms,as well as its in situ application under some special circumstances.We also analyze the advantages and challenges of nanopore sequencing in microbiology research.Overall,nanopore sequencing has the potential to greatly enhance the detection and understanding of microorganisms in environmental research,but further developments are needed to overcome the current challenges.展开更多
Extreme environmental conditions are key factors in the formation of the structure and diversity of microbial communities. In meromictic ecosystems, extreme conditions and a stable stratification of physical, chemical...Extreme environmental conditions are key factors in the formation of the structure and diversity of microbial communities. In meromictic ecosystems, extreme conditions and a stable stratification of physical, chemical and biological parameters lead to diversity and heterogeneity of microenvironments. Lake Doroninskoe is located in an extreme geographical area and diff ers from other known meromictic reservoirs of the world by a low level of illumination in the chemocline and a rare type of alkaline water in sedimentary rocks formed by evaporative concentration. To understand the key factors that shape the composition and structure of the microbial community, the macro-and micro-variations in space and time are of great importance. We investigated the short-term dynamics of the structure and diversity of microbial communities of the meromictic soda lake, Lake Doroninskoe, at day and night using highthroughput sequencing and bioinformatics. Metagenomic analysis of 16 S rRNA gene amplicons showed that the microbial communities had a high taxonomic diversity both at day and night. Sixteen bacterial and three archaeal phyla were identified. Proteobacteria were dominant and comprised 75% during the day, increasing to 90% at night. Metabolically stable denitrifying bacteria that were able to use a variety of alternative electron acceptors and electron donors were prevalent in Lake Doroninskoe. They belonged to the families Enterobacteriaceae(class Gammaproteobacteria) and Alcaligenaceae(class Betaproteobacteria). Statistically significant differences between day and night microbial communities were found. During the day, the microbial community was the most diverse. We discuss the peculiarities of the underexplored shortterm dynamics of the structure and diversity of the microbial communities of the meromictic soda Lake Doroninskoe, and propose topics for prospective studies.展开更多
Amplicon sequencing of functional genes is a powerful technique to explore the diversity and abundance of microbes involved in biogeochemical processes. One such key process, denitrification, is of particular importan...Amplicon sequencing of functional genes is a powerful technique to explore the diversity and abundance of microbes involved in biogeochemical processes. One such key process, denitrification, is of particular importance because it can transform nitrate(NO3-) to N2 gas that is released to the atmosphere. In nitrogen limited alpine wetlands, assessing bacterial denitrification under the stress of wetland desertification is fundamental to understand nutrients, especially nitrogen cycling in alpine wetlands, and thus imperative for the maintenance of healthy alpine wetland ecosystems. We applied amplicon sequencing of the nirS gene to analyze the response of denitrifying bacterial community to alpine wetland desertification in Zoige, China. Raw reads were processed for quality, translated with frameshift correction, and a total of 95,316 nirS gene sequences were used for rarefaction analysis, and 1011 OTUs were detected and used in downstream analysis. Compared to the pristine swamp soil, edaphic parameters including water content, organic carbon, total nitrogen, total phosphorous, available nitrogen, available phosphorous and potential denitrification rate were significantly decreased in the moderately degraded meadow soil and in severely degraded sandy soil. Diversity of the soil nirS-type denitrifying bacteria communities increased along the Zoige wetland desertification, and Proteobacteria and Chloroflexi were the dominant denitrifying bacterial species. Genus Cupriavidus(formerly Wautersia), Azoarcus, Azospira, Thiothrix, and Rhizobiales were significantly(P<0.05) depleted along the wetland desertification succession. Soil available phosphorous was the key determinant of the composition of the nirS gene containing denitrifying bacterial communities. The proportion of depleted taxa increased along the desertification of the Zoige wetland, suggesting that wetland desertification created specific physicochemical conditions that decreased the microhabitats for bacterial denitrifiers and the denitrification related g展开更多
文摘Previously, we showed that prophylactic addition of glucose to Harsha Lake water samples could inhibit cyanobacteria growth, at least for a short period of time. The current study tested cyanobacterial control with glucose for the entire Harsha Lake bloom season. Water samples (1000 ml) were collected weekly from Harsha Lake during the algal-bloom season starting June 9 and lasting until August 24, 2022. To each of two 7-liter polypropylene containers, 500 ml of Harsha Lake water was added, and the containers were placed in a controlled environment chamber. To one container labeled “Treated,” 0.15 g of glucose was added, and nothing was added to the container labeled “Control.” After that, three 25 ml samples from each container were collected and used for 16S rRNA gene sequencing each week. Then 1000 ml of Harsha Lake water was newly collected each week, with 500 ml added to each container, along with the addition of 0.15 g glucose to the “Treated” container. Sequencing data were used to examine differences in the composition of bacterial communities between Treated and Control containers. Treatment with glucose altered the microbial communities by 1) reducing taxonomic diversity, 2) largely eliminating cyanobacterial taxa, and 3) increasing the relative abundance of subsets of non-cyanobacterial taxa (such as Proteobacteria and Actinobacteriota). These effects were observed across time despite weekly inputs derived directly from Lake water. The addition of glucose to a container receiving weekly additions of Lake water suppressed the cyanobacterial populations during the entire summer bloom season. The glucose appears to stimulate the diversity of certain bacterial taxa at the expense of the cyanobacteria.
基金the National Natural Science Foundation of China(32071548,31670503,42077206)the National Key Research and Development Program of China(2018YFE0107000)+2 种基金the 13th Five-year Informatization Plan of Chinese Academy of Sciences(XXH13503-03-106)the National Science Fund for Distinguished Young Scholars(41925028)China Biodiversity Observation Networks(Sino BON).
文摘The diazotrophic community in biological soil crusts(biocrusts)is the key supplier of nitrogen in dryland.To date,there is still limited information on how biocrust development influences the succession of diazotrophic community,and what are the most important factors mediating diazotrophic communities during biocrust succession.Using the high throughput nifH amplicon sequencing,the diazotrophs in soils at different developmental stages of biocrust were comparatively studied.The results evidenced the decreases of TOC/TN ratio and pH value with biocrust development.Nostoc and Scytonema were the most dominant diazotrophic genera at all biocrust stages,while Azospirillum and Bradyrhizobium were abundant only in bare soil.Diazotrophic co-occurrence networks tended to be less complex and less connected with biocrust succession.The soil TOC/TN ratio was the most dominant factor mediating diazotrophic diversity,community composition and assembly processes,while diazotrophic-diversity and NO3–-N/NH4+-N ratio were positively correlated with the nitrogenase activity during biocrust succession.This study provided novel understandings of nitrogen fixation and succession patterns of diazotrophic community,by showing the effects of biocrust succession on diazotrophic diversity,community composition,community assembly and co-occurrence networks,and recognizing TOC/TN ratio as the most dominant factor mediating diazotrophs during biocrust succession.
基金supported by the National Key Research and Development Program(grant number:2022YFC2305304).
文摘Objective Viral encephalitis is an infectious disease severely affecting human health.It is caused by a wide variety of viral pathogens,including herpes viruses,flaviviruses,enteroviruses,and other viruses.The laboratory diagnosis of viral encephalitis is a worldwide challenge.Recently,high-throughput sequencing technology has provided new tools for diagnosing central nervous system infections.Thus,In this study,we established a multipathogen detection platform for viral encephalitis based on amplicon sequencing.Methods We designed nine pairs of specific polymerase chain reaction(PCR)primers for the 12 viruses by reviewing the relevant literature.The detection ability of the primers was verified by software simulation and the detection of known positive samples.Amplicon sequencing was used to validate the samples,and consistency was compared with Sanger sequencing.Results The results showed that the target sequences of various pathogens were obtained at a coverage depth level greater than 20×,and the sequence lengths were consistent with the sizes of the predicted amplicons.The sequences were verified using the National Center for Biotechnology Information BLAST,and all results were consistent with the results of Sanger sequencing.Conclusion Amplicon-based high-throughput sequencing technology is feasible as a supplementary method for the pathogenic detection of viral encephalitis.It is also a useful tool for the high-volume screening of clinical samples.
基金This study was financially supported by the Agricultural Science and Technology Innovation Program(CAAS-ZDRW202308)the Natural Science Foundation of China(42277297,42090060,U21A20182)+2 种基金Jiangsu Funding Program for Excellent Postdoctoral Talent(2022ZB325)Scientific and technology innovation project of China Academy of Chinese Medical Sciences(C12021A04115)the Fundamental Research Funds for the Central public welfare research institutes(ZZ13-YQ-095).
文摘With the gradual maturity of sequencing technology,many microbiome studies have published,driving the emergence and advance of related analysis tools.R language is the widely used platform for microbiome data analysis for powerful functions.However,tens of thousands of R packages and numerous similar analysis tools have brought major challenges for many researchers to explore microbiome data.How to choose suitable,efficient,convenient,and easy-to-learn tools from the numerous R packages has become a problem for many microbiome researchers.We have organized 324 common R packages for microbiome analysis and classified them according to application categories(diversity,difference,biomarker,correlation and network,functional prediction,and others),which could help researchers quickly find relevant R packages for microbiome analysis.Furthermore,we systematically sorted the integrated R packages(phyloseq,microbiome,MicrobiomeAnalystR,Animalcules,microeco,and amplicon)for microbiome analysis,and summarized the advantages and limitations,which will help researchers choose the appropriate tools.Finally,we thoroughly reviewed the R packages for microbiome analysis,summarized most of the common analysis content in the microbiome,and formed the most suitable pipeline for microbiome analysis.This paper is accompanied by hundreds of examples with 10,000 lines codes in GitHub,which can help beginners to learn,also help analysts compare and test different tools.This paper systematically sorts the application of R in microbiome,providing an important theoretical basis and practical reference for the development of better microbiome tools in the future.All the code is available at GitHub github.com/taowenmicro/EasyMicrobiomeR.
基金supported by the National Key Research and Development Program of China(2022YFC2303401,2022YFC2304100,2016YFD0500301,2021YFC0863300)the Beijing Science and Technology Plan(Z211100002521017)the National Natural Science Foundation of China(82241080)。
文摘The monkeypox virus(MPXV)has triggered a current outbreak globally.Genome sequencing of MPXV and rapid tracing of genetic variants will benefit disease diagnosis and control.It is a significant challenge but necessary to optimize the strategy and application of rapid full-length genome identification and to track variations of MPXV in clinical specimens with low viral loads,as it is one of the DNA viruses with the largest genome and the most AT-biased,and has a significant number of tandem repeats.Here we evaluated the performance of metagenomic and amplicon sequencing techniques,and three sequencing platforms in MPXV genome sequencing based on multiple clinical specimens of five mpox cases in Chinese mainland.We rapidly identified the full-length genome of MPXV with the assembly of accurate tandem repeats in multiple clinical specimens.Amplicon sequencing enables cost-effective and rapid sequencing of clinical specimens to obtain high-quality MPXV genomes.Third-generation sequencing facilitates the assembly of the terminal tandem repeat regions in the monkeypox virus genome and corrects a common misassembly in published sequences.Besides,several intra-host single nucleotide variations were identified in the first imported mpox case.This study offers an evaluation of various strategies aimed at identifying the complete genome of MPXV in clinical specimens.The findings of this study will significantly enhance the surveillance of MPXV.
基金supported by the Science and Technology Basic Resources Investigation Program of China(2018FY101105)the National Natural Science Foundation of China(31972272,31871969)+1 种基金the Fundamental Research Funds for the Central Universities(SWU-XDPY22001)of Chinathe China Agricultural Research System of MOA and MARA.
文摘Insecticide resistance in Panonychus citri is a major obstacle to mite control in citrus orchards.Pyrethroid insecticides are continually used to control mites in China,although resistance to pyrethroids has evolved in some populations.Here,the resistance to the pyrethroid fenpropathrin was investigated and 7 out of 8 field-collected populations of P citri exhibited a high level of resistance,ranging from 171-fold to 15391-fold higher than the susceptible(SS)comparison strain.Three voltage-gated sodium channel(VGSC)mutations were identified in the tested populations:L1031V,F1747L,and F17511.Amplicon sequencing was used to evaluate the frequency of these mutations in the 19 field populations.L1031V and F1751I were present in all populations at frequencies of 11.6%-82.1%and 0.5%-31.8%,respectively,whereas the F1747L mutation was only present in 12 populations from Chongqing,Sichuan,Guangxi,and Yunnan provinces.Introduction of these mutations singly or in combination into transgenic flies significantly increased their resistance to fenpropathrin and these flies also exhibited reduced mortality after exposure to the pyrethroids permethrin andβ-cypermethrin.Panonychus citri VGSC homology modeling and ligand docking indicate that F1747 and F1751 form direct binding contacts with pyrethroids,which are lost with mutation,whereas L1031 mutation may diminish pyrethroid effects through an allosteric mechanism.Overall,the results provide molecular markers for monitoring pest resistance to pyrethroids and offer new insights into the basis of pyrethroid actions on sodium channels.
基金the CAMS Innovation Fund for Medical Sciences(CIFMS)(2021-I2M-1-032 and 2017-I2M-1-013)the National Key Research and Development Project(2022YFC3501504).
文摘Notoginseng Radix et Rhizoma(Sanqi in Chinese)is a precious traditional Chinese herbal medicine.It has the effect of dispersing blood stasis and stopping bleeding,reducing swelling and fixing pain.However,it tends to contaminate with harmful fungi during storage,which may make it much less effective.In order to understand the fungal contamination of Notoginseng Radix et Rhizoma and master its composition of the exogenous fungi.The surface fungi of Notoginseng Radix et Rhizoma samples collected from six Chinese provinces and districts were investigated by using dilution plate method.Detection of aflatoxins by UPLC-MS/MS.The results showed that Penicillium citrinum was dominantly isolated from Notoginseng Radix et Rhizoma samples from No.1 to No.4.Aspergillus flavus,which produces aflatoxin,was dominantly isolated from Notoginseng Radix et Rhizoma samples from No.5 and No.6.In addition,kinds of mycotoxin were assayed which were produced by three of those identified A.flavus.All three fungi strains produced aflatoxin B1(AFB1)and one strain HBSQ1-5 additionally produced other three kinds of mycotoxin,AFB2,AFG1 and AFG2.It is the results implied that it will be very important to take serious cautions when using Notoginseng Radix et Rhizoma.As well as,understanding the composition of the exogenous fungi of Notoginseng Radix et Rhizoma and the strains of toxin-producing fungi,which can play an important role in guiding the storage of Notoginseng Radix et Rhizoma.
基金supported by the National Natural Science Foundation of China,Nos. 81771346, 82071383the Natural Science Foundation of Shandong Province (Key Project),No. ZR2020KH007+3 种基金the Taishan Scholar Youth Program of Shandong Province,No. tsqn201812156Academic Promotion Program of Shandong First Medical University,Nos. 2019QL025, 2019RC021Spring Industry Leader Talent Support Plan,No. 201984Rongxiang Regenerative Medicine Fund,No. 2019SDRX-23 (all to BN)。
文摘Studies have shown that gut microbiota metabolites can enter the central nervous system via the blood-spinal cord barrier and cause neuroinflammation, thus constituting secondary injury after spinal cord injury. To investigate the correlation between gut microbiota and metabolites and the possible mechanism underlying the effects of gut microbiota on secondary injury after spinal cord injury, in this study, we established mouse models of T8–T10 traumatic spinal cord injury. We used 16 S rRNA gene amplicon sequencing and metabolomics to reveal the changes in gut microbiota and metabolites in fecal samples from the mouse model. Results showed a severe gut microbiota disturbance after spinal cord injury, which included marked increases in pro-inflammatory bacteria, such as Shigella, Bacteroides, Rikenella, Staphylococcus, and Mucispirillum and decreases in anti-inflammatory bacteria, such as Lactobacillus, Allobaculum, and Sutterella. Meanwhile, we identified 27 metabolites that decreased and 320 metabolites that increased in the injured spinal cord. Combined with pathway enrichment analysis, five markedly differential amino acids(L-leucine, L-methionine, L-phenylalanine, L-isoleucine and L-valine) were screened out, which play a pivotal role in activating oxidative stress and inflammatory responses following spinal cord injury. Integrated correlation analysis indicated that the alteration of gut microbiota was related to the differences in amino acids, which suggests that disturbances in gut microbiota might participate in the secondary injury through the accumulation of partial metabolites that activate oxidative stress and inflammatory responses. Findings from this study provide a new theoretical basis for improving the secondary injury after spinal cord injury through fecal microbial transplantation.
基金supported by the National Natural Science Foundation of China(31961123002,31872228)the Fundamental Research Funds for the Central Universities of China(202041001)+1 种基金the Young Taishan Scholars Program of Shandong Province(tsqn201812024)the National Science Foundation(DEB-1927159).
文摘High-throughput sequencing of amplicons has been widely used to precisely and efficiently identify species compositions and analyze community structures,greatly promoting biological studies involving large amounts of complex samples,especially those involving environmental and pathogen-monitoring ones.Commercial library preparation kits for amplicon sequencing,which generally require multiple steps,including adapter ligation and indexing,are expensive and time-consuming,especially for applications at a large scale.To overcome these limitations,a“one-step PCR approach”has been previously proposed for constructions of amplicon libraries using long fusion primers.However,efficient amplifications of target genes and accurate demultiplexing of pooled sequencing data remain to be addressed.To tackle these,we present an integrative protocol for one-step PCR amplicon library construction(OSPALC).High-quality reads have been generated by this approach to reliably identify species compositions of mock bacterial communities and environmental samples.With this protocol,the amplicon library is constructed through one regular PCR with long primers,and the total cost per DNA/cDNA sample decreases to just 7%of the typical cost via the multi-step PCR approach.Empirically tested primers and optimized PCR conditions to construct OSPALC libraries for 16S rDNA V4 regions are demonstrated as a case study.Tools to design primers targeting at any genomic regions are also presented.In principle,OSPALC can be readily applied to construct amplicon libraries of any target genes using DNA or RNA samples,and will facilitate research in numerous fields.
基金grateful to the financial support from the National Natural Science Foundation of China(Nos.22025407,21974144)Institute of Chemistry,Chinese Academy of Sciences。
文摘Environmental pollution and the spread of pathogenic microorganisms pose a significant threat to the health of humans and the planet.Thus,understanding and detecting microorganisms is crucial for maintaining a healthy living environment.Nanopore sequencing is a single-molecule detection method developed in the 1990s that has revolutionized various research fields.It offers several advantages over traditional sequencing methods,including low cost,label-free,time-saving detection speed,long sequencing reading,real-time monitoring,convenient carrying,and other significant advantages.In this review,we summarize the technical principles and characteristics of nanopore sequencing and discuss its applications in amplicon sequencing,metagenome sequencing,and whole-genome sequencing of environmental microorganisms,as well as its in situ application under some special circumstances.We also analyze the advantages and challenges of nanopore sequencing in microbiology research.Overall,nanopore sequencing has the potential to greatly enhance the detection and understanding of microorganisms in environmental research,but further developments are needed to overcome the current challenges.
基金conducted according to the project IX.137.1.1“Biodiversity of natural and natural-technogenic ecosystems of Transbaikalia(Cenrtal Asia)as indicators of regional climate changes”(АААА-А17-117011210078-9)
文摘Extreme environmental conditions are key factors in the formation of the structure and diversity of microbial communities. In meromictic ecosystems, extreme conditions and a stable stratification of physical, chemical and biological parameters lead to diversity and heterogeneity of microenvironments. Lake Doroninskoe is located in an extreme geographical area and diff ers from other known meromictic reservoirs of the world by a low level of illumination in the chemocline and a rare type of alkaline water in sedimentary rocks formed by evaporative concentration. To understand the key factors that shape the composition and structure of the microbial community, the macro-and micro-variations in space and time are of great importance. We investigated the short-term dynamics of the structure and diversity of microbial communities of the meromictic soda lake, Lake Doroninskoe, at day and night using highthroughput sequencing and bioinformatics. Metagenomic analysis of 16 S rRNA gene amplicons showed that the microbial communities had a high taxonomic diversity both at day and night. Sixteen bacterial and three archaeal phyla were identified. Proteobacteria were dominant and comprised 75% during the day, increasing to 90% at night. Metabolically stable denitrifying bacteria that were able to use a variety of alternative electron acceptors and electron donors were prevalent in Lake Doroninskoe. They belonged to the families Enterobacteriaceae(class Gammaproteobacteria) and Alcaligenaceae(class Betaproteobacteria). Statistically significant differences between day and night microbial communities were found. During the day, the microbial community was the most diverse. We discuss the peculiarities of the underexplored shortterm dynamics of the structure and diversity of the microbial communities of the meromictic soda Lake Doroninskoe, and propose topics for prospective studies.
基金supported by the Natural Science Foundation of China (Grant No. 41201256)
文摘Amplicon sequencing of functional genes is a powerful technique to explore the diversity and abundance of microbes involved in biogeochemical processes. One such key process, denitrification, is of particular importance because it can transform nitrate(NO3-) to N2 gas that is released to the atmosphere. In nitrogen limited alpine wetlands, assessing bacterial denitrification under the stress of wetland desertification is fundamental to understand nutrients, especially nitrogen cycling in alpine wetlands, and thus imperative for the maintenance of healthy alpine wetland ecosystems. We applied amplicon sequencing of the nirS gene to analyze the response of denitrifying bacterial community to alpine wetland desertification in Zoige, China. Raw reads were processed for quality, translated with frameshift correction, and a total of 95,316 nirS gene sequences were used for rarefaction analysis, and 1011 OTUs were detected and used in downstream analysis. Compared to the pristine swamp soil, edaphic parameters including water content, organic carbon, total nitrogen, total phosphorous, available nitrogen, available phosphorous and potential denitrification rate were significantly decreased in the moderately degraded meadow soil and in severely degraded sandy soil. Diversity of the soil nirS-type denitrifying bacteria communities increased along the Zoige wetland desertification, and Proteobacteria and Chloroflexi were the dominant denitrifying bacterial species. Genus Cupriavidus(formerly Wautersia), Azoarcus, Azospira, Thiothrix, and Rhizobiales were significantly(P<0.05) depleted along the wetland desertification succession. Soil available phosphorous was the key determinant of the composition of the nirS gene containing denitrifying bacterial communities. The proportion of depleted taxa increased along the desertification of the Zoige wetland, suggesting that wetland desertification created specific physicochemical conditions that decreased the microhabitats for bacterial denitrifiers and the denitrification related g
