scholarly journals Analysis procedures for assessing recovery of high quality, complete, closed genomes from Nanopore long read metagenome sequencing

2020 ◽  
Author(s):  
Krithika Arumugam ◽  
Irina Bessarab ◽  
Mindia A. S. Haryono ◽  
Xianghui Liu ◽  
Rogelio E. Zuniga-Montanez ◽  
...  
Keyword(s):  
Activated Sludge ◽  
Filamentous Bacteria ◽  
Community Members ◽  
Functional Roles ◽  
Sludge Flocs ◽  
New Methods ◽  
Sequencing Technologies ◽  
Long Read ◽  
Metagenome Sequencing ◽  
Parallel Sampling

AbstractNew long read sequencing technologies offer huge potential for effective recovery of complete, closed genomes from complex microbial communities. Using long read (MinION) obtained from an ensemble of activated sludge enrichment bioreactors, we 1) describe new methods for validating long read assembled genomes using their counterpart short read metagenome assembled genomes; 2) assess the influence of different correction procedures on genome quality and predicted gene quality and 3) contribute 21 new closed or complete genomes of community members, including several species known to play key functional roles in wastewater bioprocesses: specifically microbes known to exhibit the polyphosphate– and glycogen–accumulating organism phenotypes (namely Accumulibacter and Dechloromonas, and Micropruina and Defluviicoccus, respectively), and filamentous bacteria (Thiothrix) associated with the formation and stability of activated sludge flocs. Our findings further establish the feasibility of long read metagenome–assembled genome recovery, and demonstrate the utility of parallel sampling of moderately complex enrichments communities for recovery of genomes of key functional species relevant for the study of complex wastewater treatment bioprocesses.

scholarly journals Recovery of complete genomes and non-chromosomal replicons from activated sludge enrichment microbial communities with long read metagenome sequencing

2021 ◽  
Vol 7 (1) ◽  
Author(s):  
Krithika Arumugam ◽  
Irina Bessarab ◽  
Mindia A. S. Haryono ◽  
Xianghui Liu ◽  
Rogelio E. Zuniga–Montanez ◽  
...  
Keyword(s):  
Activated Sludge ◽  
Microbial Communities ◽  
Community Members ◽  
Functional Roles ◽  
Sludge Flocs ◽  
Sequencing Technologies ◽  
Complete Genomes ◽  
Long Read ◽  
Metagenome Sequencing ◽  
Parallel Sampling

AbstractNew long read sequencing technologies offer huge potential for effective recovery of complete, closed genomes from complex microbial communities. Using long read data (ONT MinION) obtained from an ensemble of activated sludge enrichment bioreactors we recover 22 closed or complete genomes of community members, including several species known to play key functional roles in wastewater bioprocesses, specifically microbes known to exhibit the polyphosphate- and glycogen-accumulating organism phenotypes (namely Candidatus Accumulibacter and Dechloromonas, and Micropruina, Defluviicoccus and Candidatus Contendobacter, respectively), and filamentous bacteria (Thiothrix) associated with the formation and stability of activated sludge flocs. Additionally we demonstrate the recovery of close to 100 circularised plasmids, phages and small microbial genomes from these microbial communities using long read assembled sequence. We describe methods for validating long read assembled genomes using their counterpart short read metagenome-assembled genomes, and assess the influence of different correction procedures on genome quality and predicted gene quality. Our findings establish the feasibility of performing long read metagenome-assembled genome recovery for both chromosomal and non-chromosomal replicons, and demonstrate the value of parallel sampling of moderately complex enrichment communities to obtaining high quality reference genomes of key functional species relevant for wastewater bioprocesses.

scholarly journals Deep learning-based morphology classification of activated sludge flocs in wastewater treatment plants

2021 ◽  
Author(s):  
Hisashi Satoh ◽  
Yukari Kashimoto ◽  
Naoki Takahashi ◽  
Takashi Tsujimura
Keyword(s):  
Wastewater Treatment ◽  
Deep Learning ◽  
Activated Sludge ◽  
Morphological Change ◽  
Wastewater Treatment Plants ◽  
Filamentous Bacteria ◽  
Sludge Flocs

A deep learning-based two-label classifier 1 recognized a 20% morphological change in the activated flocs. Classifier-2 quantitatively recognized an abundance of filamentous bacteria in activated flocs.

scholarly journals Efficiency of Biological Phosphorus Removal by Filamentous Bacteria

2016 ◽  
Vol 21 (1-2) ◽  
pp. 117-123 ◽  
Author(s):  
Alicja Machnicka ◽  
Klaudiusz Grübel
Keyword(s):  
Activated Sludge ◽  
Phosphorus Removal ◽  
Water Environment ◽  
Treatment Plant ◽  
Chemical Precipitation ◽  
Heterogeneous Structure ◽  
Filamentous Bacteria ◽  
Biological Phosphorus Removal ◽  
High Concentration ◽  
Sludge Flocs

AbstractPhosphorus removal in wastewater treatment plant is carried out by chemical precipitation, advanced biological treatment or a combination of both. One of the biggest problems with high concentration of phosphorus in water environment is eutrophication. Activated sludge flocs have a heterogeneous structure, which consist of a variety of microorganisms. Filamentous bacteria are normally present in the activated sludge and have ability to assimilation of phosphorus. In this study phosphorus accumulation by isolated filamentous bacteria from activated sludge foam was present.

scholarly journals proovframe: frameshift-correction for long-read (meta)genomics

2021 ◽  
Author(s):  
Thomas Hackl ◽  
Florian Trigodet ◽  
A Murat Eren ◽  
Steven J Biller ◽  
John M Eppley ◽  
...  
Keyword(s):  
Gene Prediction ◽  
Genomic Analysis ◽  
Community Members ◽  
Reading Frame ◽  
Complex Samples ◽  
Small Indels ◽  
Sequencing Technologies ◽  
Long Reads ◽  
Long Read ◽  
Improving Accuracy

Long-read sequencing technologies hold big promises for the genomic analysis of complex samples such as microbial communities. Yet, despite improving accuracy, basic gene prediction on long-read data is still often impaired by frameshifts resulting from small indels. Consensus polishing using either complementary short reads or to a lesser extent the long reads themselves can mitigate this effect but requires universally high sequencing depth, which is difficult to achieve in complex samples where the majority of community members are rare. Here we present proovframe, a software implementing an alternative approach to overcome frameshift errors in long-read assemblies and raw long reads. We utilize protein-to-nucleotide alignments against reference databases to pinpoint indels in contigs or reads and correct them by deleting or inserting 1-2 bases, thereby conservatively restoring reading-frame fidelity in aligned regions. Using simulated and real-world benchmark data we show that proovframe performs comparably to short-read-based polishing on assembled data, works well with remote protein homologs, and can even be applied to raw reads directly. Together, our results demonstrate that protein-guided frameshift correction significantly improves the analyzability of long-read data both in combination with and as an alternative to common polishing strategies. Proovframe is available from https://github.com/thackl/proovframe.

scholarly journals MEGAN-LR: New algorithms allow accurate binning and easy interactive exploration of metagenomic long reads and contigs

2017 ◽  
Author(s):  
Daniel H. Huson ◽  
Benjamin Albrecht ◽  
Caner Bagci ◽  
Irina Bessarab ◽  
Anna Gorska ◽  
...  
Keyword(s):  
Reference Database ◽  
Practical Application ◽  
Computational Tools ◽  
New Methods ◽  
Sequencing Technologies ◽  
Long Reads ◽  
Interval Tree ◽  
Long Read ◽  
Taxonomic Binning ◽  
New Algorithms

AbstractBackgroundThere are numerous computational tools for taxonomic or functional analysis of microbiome samples, optimized to run on hundreds of millions of short, high quality sequencing reads. Programs such as MEGAN allow the user to interactively navigate these large datasets. Long read sequencing technologies continue to improve and produce increasing numbers of longer reads (of varying lengths in the range of 10k-1M bps, say), but of low quality. There is an increasing interest in using long reads in microbiome sequencing and there is a need to adapt short read tools to long read datasets.MethodsWe describe a new LCA-based algorithm for taxonomic binning, and an interval-tree based algorithm for functional binning, that are explicitly designed for long reads and assembled contigs. We provide a new interactive tool for investigating the alignment of long reads against reference sequences. For taxonomic and functional binning, we propose to use LAST to compare long reads against the NCBI-nr protein reference database so as to obtain frame-shift aware alignments, and then to process the results using our new methods.ResultsAll presented methods are implemented in the open source edition of MEGAN and we refer to this new extension as MEGAN-LR (MEGAN long read). We evaluate the LAST+MEGAN-LR approach in a simulation study, and on a number of mock community datasets consisting of Nanopore reads, PacBio reads and assembled PacBio reads. We also illustrate the practical application on a Nanopore dataset that we sequenced from an anammox bio-rector community.

scholarly journals Molecular characterization of filamentous bacteria isolated from full-scale activated sludge processes

2007 ◽  
Author(s):  
◽  
Zinhle Marrengane
Keyword(s):  
Activated Sludge ◽  
Morphological Characteristics ◽  
Molecular Techniques ◽  
Support Network ◽  
Bulk Liquid ◽  
Bacterial Cells ◽  
Filamentous Bacteria ◽  
Vast Number ◽  
Sludge Flocs ◽  
Activated Sludge Processes

Activated sludge flocs are responsible for flocculation, settling and dewaterability. It is important to maintain the growth off loc-forming bacteria for efficient sludge settleability and compaction for good quality effluent. Filamentous bacteria on the other hand are believed to provide rigid support network or backbone upon which floc-forming bacteria adhere to form stable activated sludge flocs (Wilderer et al., 2002; Ramothokang et al., 2003). Filamentous bacteria can also be detrimental to the process when they outgrow floc-forming bacteria. Morphologically filamentous bacteria are at an advantage as they have higher outward growth velocity and can extend freely to bulk liquid substrate. Proliferation of filamentous bacteria causes foaming and bulking (Martins et al., 2004). Although chemical alleviation measures to circumvent bulking are present, they are symptomatic (Chang et al., 2004). Eikelboom (1975) developed the first identification keys for the classification of filamentous bacteria that is primarily based on morphological characteristics and microscopic examination. Although very useful, this type of identification has its limitations. For instance some filamentous bacteria can change morphology in response to changes in the environment and although some of them can be morphologically similar they may vary considerably in their physiology and taxonomy (Martins et al., 2004). A vast number of filamentous bacteria are still very poorly understood which could be due to the problems of cultivation due to their slow growing nature and maintenance of cultures (Rossetti et al., 2006). This limitation necessitates a molecular approach to resolve the taxonomy of filamentous bacteria as it is a culture-independent technique which is highly accurate. This project was undertaken to verify the identity of pure cultures of filamentous bacteria isolated previously through the application of molecular techniques. The 16S rDNA are conserved regions in bacterial cells and they can be extracted and specific nucleic acid fragments amplified. Denaturation gradient gel electrophoresis enabled the separation of fragments of identical length but different size and served as an indication of purity (Muyzer et al., 1993).

Application of Image Analysis System for Study of Activated Sludge Flocs

1986 ◽  
Vol 21 (1) ◽  
pp. 130-140 ◽  
Author(s):  
Da-hong Li ◽  
J. J. Ganczarczyk
Keyword(s):  
Image Analysis ◽  
Activated Sludge ◽  
Statistical Processing ◽  
Equivalent Diameter ◽  
Image Analysis System ◽  
Sectional Area ◽  
Cross Sectional ◽  
Computerized Image Analysis ◽  
Sludge Flocs ◽  
Analysis System

Abstract The computerized image analysis system has been successfully used for determination and statistical processing of the following geometric characteristics of activated sludge flocs: longest dimension, breadth, equivalent diameter, cross-sectional area, perimeter, elongation, and circularity. These parameters could be effectively and precisely determined by the system applied. In addition, the studied method, as compared to direct microscope observation and photography floc-sizing methods, was found to be more accurate, less time-consuming, and less dependent on the investigators.

Role of Glycogen as an Intracellular Carbon Reserve of Activated Sludge in the Competitive Growth of Filamentous and Non-Filamentous Bacteria

1991 ◽  
Vol 23 (4-6) ◽  
pp. 899-905 ◽  
Author(s):  
Y. Matsuzawa ◽  
T. Mino
Keyword(s):  
Activated Sludge ◽  
Feeding Pattern ◽  
Filamentous Bacteria ◽  
Competitive Growth ◽  
Maximum Capacity ◽  
Intermittent Feeding ◽  
Feeding Type ◽  
Continuous Feeding ◽  
The Relationship

Activated sludge mixed cultures were cultivated with a glucose containing substrate in order to investigate the relationship between the feeding pattern (continuous or intermittent feeding) and the glycogen reservation capacity of activated sludge. An experimental method to measure the maximum capacity of glycogen reservation in the sludge was developed. Sludge with higher glycogen reservation capacity has an ability to synthesize glycogen faster, which ensures the higher glucose uptake. Therefore, sludge which has high glycogen reservation capacity becomes predominant in intermittently fed reactors. When the feeding pattern was changed from continuous feeding to intermittent feeding, a filamentous bacterium, Type 1701, started to decrease and a gram positive tetrad coccus became predominant. When the feeding pattern was returned to continuous feeding, Type 1701 re-appeared. Type 1701 has lower glycogen reservation capacity than the tetrad coccus. Therefore, the former cannot dominate over the latter in intermittently fed reactors.

scholarly journals Investigation of long non-coding RNAs as regulatory players of grapevine response to powdery and downy mildew infection

2021 ◽  
Vol 21 (1) ◽  
Author(s):  
Garima Bhatia ◽  
Santosh K. Upadhyay ◽  
Anuradha Upadhyay ◽  
Kashmir Singh
Keyword(s):  
Downy Mildew ◽  
Plasmopara Viticola ◽  
Defense Responses ◽  
Protein Coding ◽  
Functional Roles ◽  
Real Time Quantitative Pcr ◽  
Transcriptional Reprogramming ◽  
Sequencing Technologies ◽  
Non Coding Rnas ◽  
Fungal Phytopathogens

Abstract Background Long non-coding RNAs (lncRNAs) are regulatory transcripts of length > 200 nt. Owing to the rapidly progressing RNA-sequencing technologies, lncRNAs are emerging as considerable nodes in the plant antifungal defense networks. Therefore, we investigated their role in Vitis vinifera (grapevine) in response to obligate biotrophic fungal phytopathogens, Erysiphe necator (powdery mildew, PM) and Plasmopara viticola (downy mildew, DM), which impose huge agro-economic burden on grape-growers worldwide. Results Using computational approach based on RNA-seq data, 71 PM- and 83 DM-responsive V. vinifera lncRNAs were identified and comprehensively examined for their putative functional roles in plant defense response. V. vinifera protein coding sequences (CDS) were also profiled based on expression levels, and 1037 PM-responsive and 670 DM-responsive CDS were identified. Next, co-expression analysis-based functional annotation revealed their association with gene ontology (GO) terms for ‘response to stress’, ‘response to biotic stimulus’, ‘immune system process’, etc. Further investigation based on analysis of domains, enzyme classification, pathways enrichment, transcription factors (TFs), interactions with microRNAs (miRNAs), and real-time quantitative PCR of lncRNAs and co-expressing CDS pairs suggested their involvement in modulation of basal and specific defense responses such as: Ca2+-dependent signaling, cell wall reinforcement, reactive oxygen species metabolism, pathogenesis related proteins accumulation, phytohormonal signal transduction, and secondary metabolism. Conclusions Overall, the identified lncRNAs provide insights into the underlying intricacy of grapevine transcriptional reprogramming/post-transcriptional regulation to delay or seize the living cell-dependent pathogen growth. Therefore, in addition to defense-responsive genes such as TFs, the identified lncRNAs can be further examined and leveraged to candidates for biotechnological improvement/breeding to enhance fungal stress resistance in this susceptible fruit crop of economic and nutritional importance.

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