Abstract
Metagenome assembly is an efficient approach to reconstruct microbial genomes from metagenomic sequencing data. Although short-read sequencing has been widely used for metagenome assembly, ...linked- and long-read sequencing have shown their advancements in assembly by providing long-range DNA connectedness. Many metagenome assembly tools were developed to simplify the assembly graphs and resolve the repeats in microbial genomes. However, there remains no comprehensive evaluation of metagenomic sequencing technologies, and there is a lack of practical guidance on selecting the appropriate metagenome assembly tools. This paper presents a comprehensive benchmark of 19 commonly used assembly tools applied to metagenomic sequencing datasets obtained from simulation, mock communities or human gut microbiomes. These datasets were generated using mainstream sequencing platforms, such as Illumina and BGISEQ short-read sequencing, 10x Genomics linked-read sequencing, and PacBio and Oxford Nanopore long-read sequencing. The assembly tools were extensively evaluated against many criteria, which revealed that long-read assemblers generated high contig contiguity but failed to reveal some medium- and high-quality metagenome-assembled genomes (MAGs). Linked-read assemblers obtained the highest number of overall near-complete MAGs from the human gut microbiomes. Hybrid assemblers using both short- and long-read sequencing were promising methods to improve both total assembly length and the number of near-complete MAGs. This paper also discussed the running time and peak memory consumption of these assembly tools and provided practical guidance on selecting them.
Aerobic granulation of nitrifying activated sludge could enhance the removal of 17α-ethinylestradiol (EE2) via abiotic nitration induced by reactive nitrogen species, cometabolism by ...ammonia-oxidizing bacteria and biodegradation by heterotrophic bacteria. Zero-valent iron (ZVI), a promising and low-cost material, has previously been applied to effectively enhance biological wastewater treatment. The impact and the effect mechanism of ZVI on nitrifying granular sludge (NGS) for EE2 removal was investigated in this study. The results showed that the addition of ZVI achieved better EE2 removal, though ZVI was not conducive to the accumulation of nitrite in NGS which reduced the abiotic transformation of EE2. Moreover, ZVI enriched heterotrophic denitrifying bacteria such as Arenimonas, thus changing the EE2 removal pathway and improving the degradation and mineralization of EE2. In addition, ZVI reduced the emission risk of the greenhouse gas N2O and strengthened the stability of the granules. Metagenomic analysis further revealed that the functional genes related to EE2 mineralization, nitrite oxidation, N2O reduction and quorum sensing in NGS were enriched with ZVI addition. This study provides meaningful guidance for ZVI application in the NGS process to achieve efficient and simultaneous removal of ammonia and emerging contaminants.
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•ZVI changed the EE2 removal pathways and improved EE2 degradation and mineralization.•Heterotrophic denitrifying bacteria Arenimonas played a key role in EE2 removal.•ZVI pushed complete nitrification but ammonia removal was not affected.•ZVI reduced the emission risk of N2O and strengthened the stability of the granules.
The Phylum Cressdnaviricota consists of a large number of circular Rep‐encoding single‐stranded (CRESS)‐DNA viruses. Recently, metagenomic analyzes revealed their ubiquitous distribution in a diverse ...range of eukaryotes. Data relating to CRESS‐DNA viruses in humans remains scarce. Our study investigated the presence and genetic diversity of CRESS‐DNA viruses in human vaginal secretions. Vaginal swabs were collected from 28 women between 29 and 43 years old attending a fertility clinic in New York City. An exploratory metagenomic analysis was performed and detection of CRESS‐DNA viruses was confirmed through analysis of near full‐length sequences of the viral isolates. A phylogenetic tree was based on the REP open reading frame sequences of the CRESS‐DNA virus genome. Eleven nearly complete CRESS‐DNA viral genomes were identified in 16 (57.1%) women. There were no associations between the presence of these viruses and any demographic or clinical parameters. Phylogenetic analysis indicated that one of the sequences belonged to the genus Gemycircularvirus within the Genomoviridae family, while ten sequences represented previously unclassified species of CRESS‐DNA viruses.
Novel species of CRESS‐DNA viruses are present in the vaginal tract of adult women. Although they be transient commensal agents, the potential clinical implications for their presence at this site cannot be dismissed.
Forest soils represent important terrestrial carbon (C) pools, where C is primarily fixed in plant biomass and then is incorporated in the biomass of fungi and bacteria. Although classical concepts ...assume that fungi are the main decomposers of the recalcitrant organic matter within plant and microbial biomass, whereas bacteria are considered to mostly utilize simpler compounds, recent studies have shown that fungi and bacteria overlap in substrate utilization. Here, we studied the microbial contribution to the recycling of dead biomass by analyzing the bacterial and fungal communities in soil microcosms supplemented with 13C-labeled biomass of plant, fungal, and bacterial origin using a combination of DNA-stable isotope probing and metagenomics. Both fungi and bacteria contributed actively to the degradation of complex components of plant and microbial biomass. Specific families of carbohydrate-active enzymes (CAZyme) were involved in the degradation of each biomass type. Moreover, the analysis of five bacterial metagenome-assembled genomes indicated the key role of some bacterial genera in the degradation of plant biomass (Cytophaga and Asticcacaulis) and microbial biomass (Herminiimonas). The enzymatic systems utilized by bacteria are highly complex and complementary but also highly diverse among taxa. The results confirm the importance of bacteria, in addition to fungi, as decomposers of complex organic matter in forest soils.
•Both fungi and bacteria actively degrade complex plant and microbial biomass.•The pool of CAZymes was distinct in fungi and bacteria.•Fungal communities encode more specific CAZymes that degrade plant biomass.•Bacterial communities are richer in CAZymes that target microbial biomass.•Bacteria use structurally variable but complementary enzymatic systems.
•mONS can provide rapid, accurate information on pathogenic strains in PJI samples.•The species identified by mONS from fresh samples corresponded with both routine culture and mNGS results.•This ...study showed proof of concept that mONS can function as a rapid, accurate tool in PJI diagnostic microbiology.
Pathogen identification is crucial for the diagnosis and management of periprosthetic joint infection (PJI). Although culturing methods are the foundation of pathogen detection in PJI, false-negative results often occur. Oxford nanopore sequencing (ONS) is a promising alternative for detecting pathogens and providing information on their antimicrobial resistance (AMR) profiles, without culturing.
To evaluate the capability of metagenomic ONS (mONS) in detecting pathogens from PJI samples, both metagenomic next-generation sequencing (mNGS) and mONS were performed in 15 osteoarticular samples from nine consecutive PJI patients according to the modified Musculoskeletal Infection Society (MSIS) criteria. The sequencing data generated from both platforms were then analyzed for pathogen identification and AMR detection using an in-house-developed bioinformatics pipeline.
Our results showed that mONS could be applied to detect the causative pathogen and characterize its AMR features in fresh PJI samples. By real-time sequencing and analysis, pathogen identification and AMR detection from the initiation of sequencing were accelerated.
We showed proof of concept that mONS can function as a rapid, accurate tool in PJI diagnostic microbiology. Despite efforts to reduce host DNA, the high proportion of host DNA was still a limitation of this method that prevented full genome analysis.
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•Partial-denitrification is a feasible way to supply nitrite for anammox.•Nitrite accumulation rate (NAR) increased with pH during denitrification.•The higher NAR at pH 9.0 was ...correlated with an enrichment of Thauera.•Thauera harbored more nitrate reductase than nitrite reductase.•A process for achieving partial denitrification/anammox in WWTP was proposed.
Partial-denitrification (nitrate to nitrite) can supply nitrite for anammox which can reduce organic matter consumption in wastewater treatment plants (WWTPs). In order to achieve stable partial-denitrification, the effect of pH on denitrification were investigated for 420 days in three reactors with influent pH of 5.0, 7.0 and 9.0. The results indicate that the nitrite accumulation rate (NAR) increased with pH, with average effluent NARs being 21%, 38% and 57% in the above reactors, respectively. The sludge cultivated at a high pH of 9.0 was resistant to pH shock, with a high NAR being maintained at 83% when it was exposed to a low pH of 5.0. Metagenomic analysis showed that the higher NAR at pH 9.0 was correlated with an enrichment of Thauera, which harbored more nitrate reductase (8098 hits) than nitrite reductase (2950 hits). Based on these findings, a novel process was proposed for achieving partial-denitrification/anammox in mainstream WWTPs.
In this study, enhanced nitrogen removal through in situ enrichment of anammox bacteria was successfully obtained in a full-scale municipal wastewater treatment plant (WWTP). The WWTP was an ...anaerobic-anoxic-oxic (AAO) process and upgraded by adding moving carriers into the anoxic zone. Enhanced nitrogen removal was obtained during almost two years of operation. The significant nitrogen removal might be associated with the in situ enrichment of anammox bacteria on the adding carriers, as revealed by the comprehensive results of molecular analysis and 15N-stable isotope tracing tests. Quantitative PCR results indicated that anammox bacteria in the anoxic-carrier biofilms presented a higher abundance than flocculent sludge (16S rRNA: P < 0.005; HzsB: P < 0.042). The 16S rRNA amplicon sequencing showed significant differences in the phylum Planctomycetes (P < 0.001) between anoxic-carrier biofilms and flocculent sludge. And metagenomic sequencing analysis further revealed the anammox relative abundance in the anoxic-carrier biofilms was significantly higher than the reported level in the flocculent sludge of conventional WWTPs. In addition, 15N-stable isotope tracing tests showed that anammox could be combined with nitrate reduction by the anoxic-carrier biofilms. Thus, enriched anammox bacteria might contribute to nitrogen loss and lead to improvements in the nitrogen removal, which was also supported by the mass balance analysis of organic carbon, nitrogen, and phosphorus of the WWTP. Overall, this study suggests that anoxic-carrier biofilms might be a candidate to enhance nitrogen removal through partial anammox in municipal WWTPs.
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•The carriers added in the anoxic zone of the municipal WWTP could enhance N-removal.•The anoxic-carrier biofilms enriched with both denitrifies and anammox bacteria.•Anoxic biofilms show comparable AMX activity as denitrification by 15N isotope tracer.•Coupling nitrate reduction with anammox might enhance N-loss via anoxic biofilms.
Background
: We conducted this retrospective study to reveal the accuracy of metagenomic next-generation sequencing (mNGS) for diagnosing osteoarticular infections from fresh abscess specimens ...obtained from patients in an HIV-naive population.
Methods
: We retrospectively analyzed hospital records at three participating TB-specialized hospitals for patients admitted with suggestive diagnoses of osteoarticular tuberculosis between January 2018 and August 2019. Abscess specimens obtained from each patient were tested
via
pathogen culture, GeneXpert
Mycobacterium tuberculosis
(MTB)/rifampicin (RIF), and mNGS assay.
Results
: A total of 82 abscess samples were collected from patients with osteoarticular infections, including 53 cases with (64.6%) bacterial, 21 (25.6%) with mycobacterial, 7 (8.5%) with fungal, and 1 (1.2%) with actinomycetal organisms detected. Analysis of mNGS assay results identified potential pathogens in all cases, with
M. tuberculosis
complex (MTBC) most frequently isolated, followed by
Staphylococcus aureus
and
Brucella melitensis
. Conventional culture testing identified causative pathogens in only 48.4% of samples, a significantly lower rate than the mNGS pathogen identification rate (100%,
p
< 0.01). Culture-positive group specimens yielded significantly greater numbers of sequence reads than did culture-negative group specimens (
p
< 0.01). Of patients receiving surgical interventions and mNGS-guided treatment, 76 (92.7%) experienced favorable outcomes by the time of follow-up assessment at 3 months post-treatment. Notably, MTBC detection in two patients experiencing treatment failure suggests that they had mixed infections with MTBC and other pathogens.
Conclusion
: Results presented here demonstrate that mNGS has a greater pathogen detection rate in osteoarticular infections than conventional culture-based methods.
Tropheryma whipplei is the causative agent of Whipple's disease, which is a rare multiorgan systemic disease. We report two cases of Tropheryma whipplei infection, all routine tests were negative and ...it was finally detected by mNGS. This may help clinicians increase awareness of the diagnosis and treatment of acute severe pneumonia and interstitial pneumonia caused by Tropheryma whipplei.
Stimulating Methanothrix-dominant communities with ethanol is recently considered as a promising strategy of improving the efficiency and stability of anaerobic digestion (AD), while the effects on ...methanogenic pathway and energy metabolism linked to the establishment of direct interspecies electron transfer (DIET) were not investigated yet. The results showed that, Methanothrix species were the dominant and metabolically active methanogens in the methanogenic sludge fed with the ethanol-type fermentation products, and the abundance of genes that encoded the key enzymes involved in the reduction of carbon dioxide was significantly higher than that fed with the other products, such as propionate and butyrate. Conversely, the abundance of genes that encoded the key enzymes involved in acetate decarboxylation among all the methanogenic sludge were nearly same. In the presence of ethanol, the abundance of gene for pilA significantly increased. The gene for pliA was primarily derived from Sphaerochaeta, Sedimentibacter and Pseudomonas species that were specially abundant and metabolically active. Further analysis showed that, the abundance of genes that encoded V/A-type ATPase in the methanogenic digesters fed with the ethanol-type fermentation products was 1.3–1.5 folds higher than that fed with the other products. As a result, the concentration of total ATP in the cells was increased by 1.8–2.3 folds. These results, and the fact that DIET is the only electron donor to support the reduction of carbon dioxide in Methanothrix species for the first time revealed the mechanisms involved in the establishment of DIET-based methanogenic metabolism with ethanol.
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•Ethanol significantly stimulated methanogenic pathway and energy metabolism during AD.•Methanothrix were the dominant and metabolically active methanogens.•The abundance of genes for carbon dioxide reduction in Methanothrix was increased.•The abundance of genes that encoded ATPase in Methanothrix was increased.
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