scholarly journals iSeq 100 for metagenomic pathogen screening in ticks

2021 ◽  
Vol 14 (1) ◽  
Author(s):  
Ju Yeong Kim ◽  
Myung-hee Yi ◽  
Alghurabi Areej Sabri Mahdi ◽  
Tai-Soon Yong
Keyword(s):  
Large Scale ◽  
High Throughput Sequencing ◽  
Sequence Similarity ◽  
Haemaphysalis Longicornis ◽  
Differential Abundance ◽  
Control Programs ◽  
Blood Sucking ◽  
Pathogen Screening ◽  
Vector Borne ◽  
Differential Abundance Analysis

Abstract Background Ticks are blood-sucking ectoparasites that play a pivotal role in the transmission of various pathogens to humans and animals. In Korea, Haemaphysalis longicornis is the predominant tick species and is recognized as the vector of pathogens causing various diseases such as babesiosis, borreliosis, rickettsiosis, and severe fever with thrombocytopenia syndrome. Methods In this study, the targeted high-throughput sequencing of the 16S rRNA V4 region was performed using the state-of-the-art sequencing instrument, iSeq 100, to screen bacterial pathogens in H. longicornis, and the findings were compared with those using conventional PCR with specific primers. Microbiome analyses were performed with EzBioCloud, a commercially available ChunLab bioinformatics cloud platform. ANOVA-Like Differential Expression tool (ALDEx2) was used for differential abundance analysis. Results Rickettsia spp. were detected in 16 out of 37 samples using iSeq 100, and this was confirmed using a PCR assay. In the phylogenetic analysis using gltA and ompA sequences of the detected Rickettsia, the highest sequence similarity was found with ‘Candidatus Rickettsia jingxinensis’ isolate Xian-Hl-79, ‘Ca. R. jingxinensis’ isolate F18, and ‘Ca. R. longicornii‘ isolate ROK-HL727. In the microbiome study, Coxiella AB001519, a known tick symbiont, was detected in all 37 tick samples. Actinomycetospora chiangmaiensis was more abundant in Rickettsia-positive samples than in Rickettsia-negative samples. Conclusions In this study, iSeq 100 was used to investigate the microbiome of H. longicornis, and the potentially pathogenic Rickettsia strain was detected in 16 out of 37 ticks. We believe that this approach will aid in large-scale pathogen screening of arthropods to be used in vector-borne disease control programs. Graphical Abstract

scholarly journals iSeq100 for metagenomic pathogen screening in ticks

2021 ◽  
Author(s):  
Ju Yeong Kim ◽  
Myung-hee Yi ◽  
Alghurabi Areej Sabri Mahdi ◽  
Tai-Soon Yong
Keyword(s):  
Large Scale ◽  
High Throughput Sequencing ◽  
Sequence Similarity ◽  
Haemaphysalis Longicornis ◽  
Conventional Pcr ◽  
Control Programs ◽  
Blood Sucking ◽  
Pathogen Screening ◽  
Vector Borne ◽  
Severe Fever

Abstract Background: Ticks are blood sucking ectoparasites that play a pivotal role in the transmission of various pathogens to humans and animals. In Korea, Haemaphysalis longicornis is the predominant tick species and is recognized as the vector of pathogens causing various diseases such as babesiosis, borreliosis, rickettsiosis, and severe fever with thrombocytopenia syndrome. Methods: In this study, we developed a method to screen bacterial pathogens in H. longicornis using targeted high-throughput sequencing of the 16S rRNA V4 region using the state-of-the-art sequencing instrument, iSeq 100, and compared the findings with those of conventional PCR with specific primers. Results: Rickettsia spp. were detected in 18 out of 39 samples using iSeq 100 but in only 14 samples using conventional PCR. In the phylogenetic analysis using gltA and ompA sequences of the detected Rickettsia, the highest sequence similarity was found with Candidatus Rickettsia jingxinensis isolate Xian-Hl-79, Ca. R. jingxinensis isolate F18, and Ca. R. longicornii isolate ROK-HL727. In the microbiome study, Coxiella AB001519, a known tick symbiont, was detected in all 39 tick samples. The Actinomycetospora chiangmaiensis group was more abundant in Rickettsia spp.-positive samples than in Rickettsia spp.-negative samples. Conclusions: Thus, we demonstrated that iSeq100 can rapidly and economically screen potential pathogens in ticks and can be applied to large-scale pathogen screening in arthropods for vector-borne disease control programs.

scholarly journals Randomized quantile residuals for diagnosing zero-inflated generalized linear mixed models with applications to microbiome count data

2021 ◽  
Vol 22 (1) ◽  
Author(s):  
Wei Bai ◽  
Mei Dong ◽  
Longhai Li ◽  
Cindy Feng ◽  
Wei Xu
Keyword(s):  
Count Data ◽  
Large Scale ◽  
Linear Models ◽  
Type I Error ◽  
Error Rates ◽  
Type I ◽  
Simulation Studies ◽  
Differential Abundance ◽  
Nominal Level ◽  
Differential Abundance Analysis

Abstract Background For differential abundance analysis, zero-inflated generalized linear models, typically zero-inflated NB models, have been increasingly used to model microbiome and other sequencing count data. A common assumption in estimating the false discovery rate is that the p values are uniformly distributed under the null hypothesis, which demands that the postulated model fit the count data adequately. Mis-specification of the distribution of the count data may lead to excess false discoveries. Therefore, model checking is critical to control the FDR at a nominal level in differential abundance analysis. Increasing studies show that the method of randomized quantile residual (RQR) performs well in diagnosing count regression models. However, the performance of RQR in diagnosing zero-inflated GLMMs for sequencing count data has not been extensively investigated in the literature. Results We conduct large-scale simulation studies to investigate the performance of the RQRs for zero-inflated GLMMs. The simulation studies show that the type I error rates of the GOF tests with RQRs are very close to the nominal level; in addition, the scatter-plots and Q–Q plots of RQRs are useful in discerning the good and bad models. We also apply the RQRs to diagnose six GLMMs to a real microbiome dataset. The results show that the OTU counts at the genus level of this dataset (after a truncation treatment) can be modelled well by zero-inflated and zero-modified NB models. Conclusion RQR is an excellent tool for diagnosing GLMMs for zero-inflated count data, particularly the sequencing count data arising in microbiome studies. In the supplementary materials, we provided two generic R functions, called and , for calculating the RQRs given fitting outputs of the R package .

Comparative physical mapping reveals features of microsynteny between Glycine max, Medicago truncatula, and Arabidopsis thaliana

Genome ◽  
2004 ◽  
Vol 47 (1) ◽  
pp. 141-155 ◽  
Author(s):  
H H Yan ◽  
J Mudge ◽  
D-J Kim ◽  
R C Shoemaker ◽  
D R Cook ◽  
...  
Keyword(s):  
Arabidopsis Thaliana ◽  
Glycine Max ◽  
Medicago Truncatula ◽  
Physical Mapping ◽  
Large Scale ◽  
Sequence Similarity ◽  
Artificial Chromosome ◽  
Bac Contig ◽  
Cross Hybridization ◽  
Bac Contigs

To gain insight into genomic relationships between soybean (Glycine max) and Medicago truncatula, eight groups of bacterial artificial chromosome (BAC) contigs, together spanning 2.60 million base pairs (Mb) in G. max and 1.56 Mb in M. truncatula, were compared through high-resolution physical mapping combined with sequence and hybridization analysis of low-copy BAC ends. Cross-hybridization among G. max and M. truncatula contigs uncovered microsynteny in six of the contig groups and extensive microsynteny in three. Between G. max homoeologous (within genome duplicate) contigs, 85% of coding and 75% of noncoding sequences were conserved at the level of cross-hybridization. By contrast, only 29% of sequences were conserved between G. max and M. truncatula, and some kilobase-scale rearrangements were also observed. Detailed restriction maps were constructed for 11 contigs from the three highly microsyntenic groups, and these maps suggested that sequence order was highly conserved between G. max duplicates and generally conserved between G. max and M. truncatula. One instance of homoeologous BAC contigs in M. truncatula was also observed and examined in detail. A sequence similarity search against the Arabidopsis thaliana genome sequence identified up to three microsyntenic regions in A. thaliana for each of two of the legume BAC contig groups. Together, these results confirm previous predictions of one recent genome-wide duplication in G. max and suggest that M. truncatula also experienced ancient large-scale genome duplications.Key words: Glycine max, Medicago truncatula, Arabidopsis thaliana, conserved microsynteny, genome duplication.

scholarly journals Tree Diversity Reduces Fungal Endophyte Richness and Diversity in a Large-Scale Temperate Forest Experiment

Diversity ◽  
2019 ◽  
Vol 11 (12) ◽  
pp. 234 ◽  
Author(s):  
Eric A. Griffin ◽  
Joshua G. Harrison ◽  
Melissa K. McCormick ◽  
Karin T. Burghardt ◽  
John D. Parker
Keyword(s):  
Phylogenetic Diversity ◽  
Large Scale ◽  
High Throughput Sequencing ◽  
Spatial Scales ◽  
Fungal Endophytes ◽  
Fungal Endophyte ◽  
Tree Diversity ◽  
Trophic Levels ◽  
Community Diversity ◽  
And Function

Although decades of research have typically demonstrated a positive correlation between biodiversity of primary producers and associated trophic levels, the ecological drivers of this association are poorly understood. Recent evidence suggests that the plant microbiome, or the fungi and bacteria found on and inside plant hosts, may be cryptic yet important drivers of important processes, including primary production and trophic interactions. Here, using high-throughput sequencing, we characterized foliar fungal community diversity, composition, and function from 15 broadleaved tree species (N = 545) in a recently established, large-scale temperate tree diversity experiment using over 17,000 seedlings. Specifically, we tested whether increases in tree richness and phylogenetic diversity would increase fungal endophyte diversity (the “Diversity Begets Diversity” hypothesis), as well as alter community composition (the “Tree Diversity–Endophyte Community” hypothesis) and function (the “Tree Diversity–Endophyte Function” hypothesis) at different spatial scales. We demonstrated that increasing tree richness and phylogenetic diversity decreased fungal species and functional guild richness and diversity, including pathogens, saprotrophs, and parasites, within the first three years of a forest diversity experiment. These patterns were consistent at the neighborhood and tree plot scale. Our results suggest that fungal endophytes, unlike other trophic levels (e.g., herbivores as well as epiphytic bacteria), respond negatively to increasing plant diversity.

scholarly journals MiDRMpol: A High-Throughput Multiplexed Amplicon Sequencing Workflow to Quantify HIV-1 Drug Resistance Mutations against Protease, Reverse Transcriptase, and Integrase Inhibitors

Viruses ◽  
2019 ◽  
Vol 11 (9) ◽  
pp. 806
Author(s):  
Shambhu G. Aralaguppe ◽  
Anoop T. Ambikan ◽  
Manickam Ashokkumar ◽  
Milner M. Kumar ◽  
Luke Elizabeth Hanna ◽  
...  
Keyword(s):  
Drug Resistance ◽  
High Throughput ◽  
Large Scale ◽  
High Throughput Sequencing ◽  
Polymorphism Analysis ◽  
Resistance Mutations ◽  
Subtype C ◽  
Bioinformatics Pipeline ◽  
Drug Resistance Mutations ◽  
Hiv 1

The detection of drug resistance mutations (DRMs) in minor viral populations is of potential clinical importance. However, sophisticated computational infrastructure and competence for analysis of high-throughput sequencing (HTS) data lack at most diagnostic laboratories. Thus, we have proposed a new pipeline, MiDRMpol, to quantify DRM from the HIV-1 pol region. The gag-vpu region of 87 plasma samples from HIV-infected individuals from three cohorts was amplified and sequenced by Illumina HiSeq2500. The sequence reads were adapter-trimmed, followed by analysis using in-house scripts. Samples from Swedish and Ethiopian cohorts were also sequenced by Sanger sequencing. The pipeline was validated against the online tool PASeq (Polymorphism Analysis by Sequencing). Based on an error rate of <1%, a value of >1% was set as reliable to consider a minor variant. Both pipelines detected the mutations in the dominant viral populations, while discrepancies were observed in minor viral populations. In five HIV-1 subtype C samples, minor mutations were detected at the <5% level by MiDRMpol but not by PASeq. MiDRMpol is a computationally as well as labor efficient bioinformatics pipeline for the detection of DRM from HTS data. It identifies minor viral populations (<20%) of DRMs. Our method can be incorporated into large-scale surveillance of HIV-1 DRM.

scholarly journals MetaDEGalaxy: Galaxy workflow for differential abundance analysis of 16s metagenomic data

F1000Research ◽  
2019 ◽  
Vol 8 ◽  
pp. 726
Author(s):  
Mike W.C. Thang ◽  
Xin-Yi Chua ◽  
Gareth Price ◽  
Dominique Gorse ◽  
Matt A. Field
Keyword(s):  
Microbial Communities ◽  
Sequence Data ◽  
Metagenomic Data ◽  
Marker Genes ◽  
Metagenomic Sequencing ◽  
Differential Analysis ◽  
Biomedical Sciences ◽  
Metagenomic Sequence ◽  
Differential Abundance ◽  
Differential Abundance Analysis

Metagenomic sequencing is an increasingly common tool in environmental and biomedical sciences.  While software for detailing the composition of microbial communities using 16S rRNA marker genes is relatively mature, increasingly researchers are interested in identifying changes exhibited within microbial communities under differing environmental conditions. In order to gain maximum value from metagenomic sequence data we must improve the existing analysis environment by providing accessible and scalable computational workflows able to generate reproducible results. Here we describe a complete end-to-end open-source metagenomics workflow running within Galaxy for 16S differential abundance analysis. The workflow accepts 454 or Illumina sequence data (either overlapping or non-overlapping paired end reads) and outputs lists of the operational taxonomic unit (OTUs) exhibiting the greatest change under differing conditions. A range of analysis steps and graphing options are available giving users a high-level of control over their data and analyses. Additionally, users are able to input complex sample-specific metadata information which can be incorporated into differential analysis and used for grouping / colouring within graphs.  Detailed tutorials containing sample data and existing workflows are available for three different input types: overlapping and non-overlapping read pairs as well as for pre-generated Biological Observation Matrix (BIOM) files. Using the Galaxy platform we developed MetaDEGalaxy, a complete metagenomics differential abundance analysis workflow. MetaDEGalaxy is designed for bench scientists working with 16S data who are interested in comparative metagenomics.  MetaDEGalaxy builds on momentum within the wider Galaxy metagenomics community with the hope that more tools will be added as existing methods mature.

scholarly journals A Field Efficacy Evaluation of In2Care Mosquito Traps in Comparison with Routine Integrated Vector Management at Reducing Aedes aegypti

2021 ◽  
Vol 37 (4) ◽  
pp. 242-249
Author(s):  
Eva A. Buckner ◽  
Katie F. Williams ◽  
Samantha Ramirez ◽  
Constance Darrisaw ◽  
Juliana M. Carrillo ◽  
...  
Keyword(s):  
Aedes Aegypti ◽  
Large Scale ◽  
Pathogen Transmission ◽  
Small Scale ◽  
Integrated Vector Management ◽  
Control Programs ◽  
Vector Management ◽  
Scale Field ◽  
Large Scale Field ◽  
Eggs And Larvae

ABSTRACT Aedes aegypti is the predominant vector of dengue, chikungunya, and Zika viruses. This mosquito is difficult to control with conventional methods due to its container-inhabiting behavior and resistance to insecticides. Autodissemination of pyriproxyfen (PPF), a potent larvicide, has shown promise as an additional tool to control Aedes species in small-scale field trials. However, few large-scale field evaluations have been conducted. We undertook a 6-month-long large-scale field study to compare the effectiveness and operational feasibility of using In2Care Mosquito Traps (In2Care Traps, commercially available Aedes traps with PPF and Beauveria bassiana) compared to an integrated vector management (IVM) strategy consisting of source reduction, larviciding, and adulticiding for controlling Ae. aegypti eggs, larvae, and adults. We found that while the difference between treatments was only statistically significant for eggs and larvae (P &lt; 0.05 for eggs and larvae and P &gt; 0.05 for adults), the use of In2Care Traps alone resulted in 60%, 57%, and 57% fewer eggs, larvae, and adults, respectively, collected from that site compared to the IVM site. However, In2Care Trap deployment and maintenance were more time consuming and labor intensive than the IVM strategy. Thus, using In2Care Traps alone as a control method for large areas (e.g., &gt;20 ha) may be less practical for control programs with the capacity to conduct ground and aerial larviciding and adulticiding. Based on our study results, we conclude that In2Care Traps are effective at suppressing Ae. aegypti and have the most potential for use in areas without sophisticated control programs and within IVM programs to target hotspots with high population levels and/or risk of Aedes-borne pathogen transmission.

scholarly journals A New Paralog Removal Pipeline Resolves Conflict between RAD-seq and Enrichment

2020 ◽  
Author(s):  
Wenbin Zhou ◽  
John Soghigian ◽  
Qiu-yun (Jenny) Xiang
Keyword(s):  
High Throughput Sequencing ◽  
Sequence Similarity ◽  
Phylogenetic Analyses ◽  
Disjunct Distribution ◽  
Divergence Times ◽  
Target Enrichment ◽  
Sequencing Technologies ◽  
Duplication Events ◽  
The Witch ◽  
Phylogenomic Analyses

ABSTRACTTarget enrichment and RAD-seq are well-established high throughput sequencing technologies that have been increasingly used for phylogenomic studies, and the choice between methods is a practical issue for plant systematists studying the evolutionary histories of biodiversity of relatively recent origins. However, few studies have compared the congruence and conflict between results from the two methods within the same group of organisms, especially in plants, where extensive genome duplication events may complicate phylogenomic analyses. Unfortunately, currently widely used pipelines for target enrichment data analysis do not have a vigorous procedure for remove paralogs in Hyb-Seq data. In this study, we employed RAD-seq and Hyb-Seq of Angiosperm 353 genes in phylogenomic and biogeographic studies of Hamamelis (the witch-hazels) and Castanea (chestnuts), two classic examples exhibiting the well-known eastern Asian-eastern North American disjunct distribution. We compared these two methods side by side and developed a new pipeline (PPD) with a more vigorous removal of putative paralogs from Hyb-Seq data. The new pipeline considers both sequence similarity and heterozygous sites at each locus in identification of paralogous. We used our pipeline to construct robust datasets for comparison between methods and downstream analyses on the two genera. Our results demonstrated that the PPD identified many more putative paralogs than the popular method HybPiper. Comparisons of tree topologies and divergence times showed significant differences between data from HybPiper and data from our new PPD pipeline, likely due to the error signals from the paralogous genes undetected by HybPiper, but trimmed by PPD. We found that phylogenies and divergence times estimated from our RAD-seq and Hyb-Seq-PPD were largely congruent. We highlight the importance of removal paralogs in enrichment data, and discuss the merits of RAD-seq and Hyb-Seq. Finally, phylogenetic analyses of RAD-seq and Hyb-Seq resulted in well-resolved species relationships, and revealed ancient introgression in both genera. Biogeographic analyses including fossil data revealed a complicated history of each genus involving multiple intercontinental dispersals and local extinctions in areas outside of the taxa’s modern ranges in both the Paleogene and Neogene. Our study demonstrates the value of additional steps for filtering paralogous gene content from Angiosperm 353 data, such as our new PPD pipeline described in this study. [RAD-seq, Hyb-Seq, paralogs, Castanea, Hamamelis, eastern Asia-eastern North America disjunction, biogeography, ancient introgression]

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