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1.
Int Heart J ; 60(5): 1176-1183, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31564708

RESUMO

Recently, the potential role of gut microbiome (GM) in cardiovascular diseases has been revealed. Heart failure (HF) is one of the most prevalent cardiovascular diseases worldwide; however, whether GM dysbiosis participates in the development of HF remains largely unknown. This study aimed to investigate the specific changes in GM composition and function in isoproterenol (ISO)-induced HF in rats.The rats were divided into C (control), 4w-HF (ISO, 2.5 mg/kg/day for 4 weeks, intraperitoneally), and 2w-HF (ISO, 2.5 mg/kg/day for 2 weeks, intraperitoneally) groups. The cardiac structure and function in rats were assessed, and metagenomic analyses were then performed. Compared with the healthy control group, we found that the Shannon diversity index and microbial gene count in the 4w-HF and 2w-HF groups was drastically decreased. High-throughput sequencing showed that the three groups differed in intestinal bacterial community composition. Overgrowth of bacteria, such as Prevotella, was observed in the 4w-HF group, with reduced growth of bacteria, such as Roseburia, Lactobacillus, and Butyrivibrio, associated with healthy status compared with the C group on the genus level. Concomitant with the alteration of GM composition, underrepresentation of health-linked microbial function was observed in both the 4w-HF and 2w-HF groups compared with the C group.Iso-induced HF rats showed a significant decrease in the diversity and richness of the intestinal microbiome, with a downregulation of the key intestinal bacterial groups and overgrowth of bacteria considered to be involved in inflammatory responses as well as a decrease in health-linked microbial function. Our data indicated that altered GM may be a potential player in the pathogenesis and progression of HF.


Assuntos
Bactérias/classificação , Microbioma Gastrointestinal/efeitos dos fármacos , Insuficiência Cardíaca/microbiologia , Isoproterenol/efeitos adversos , Metagenômica/métodos , Animais , Bactérias/genética , Modelos Animais de Doenças , Ecocardiografia , Insuficiência Cardíaca/induzido quimicamente , Insuficiência Cardíaca/diagnóstico por imagem , Sequenciamento de Nucleotídeos em Larga Escala , Injeções Intraperitoneais , Isoproterenol/farmacologia , Masculino , Filogenia , Ratos , Análise de Sequência de DNA
2.
BMC Bioinformatics ; 20(1): 453, 2019 Sep 05.
Artigo em Inglês | MEDLINE | ID: mdl-31488068

RESUMO

BACKGROUND: Metagenomics caused a quantum leap in microbial ecology. However, the inherent size and complexity of metagenomic data limit its interpretation. The quantification of metagenomic traits in metagenomic analysis workflows has the potential to improve the exploitation of metagenomic data. Metagenomic traits are organisms' characteristics linked to their performance. They are measured at the genomic level taking a random sample of individuals in a community. As such, these traits provide valuable information to uncover microorganisms' ecological patterns. The Average Genome Size (AGS) and the 16S rRNA gene Average Copy Number (ACN) are two highly informative metagenomic traits that reflect microorganisms' ecological strategies as well as the environmental conditions they inhabit. RESULTS: Here, we present the ags.sh and acn.sh tools, which analytically derive the AGS and ACN metagenomic traits. These tools represent an advance on previous approaches to compute the AGS and ACN traits. Benchmarking shows that ags.sh is up to 11 times faster than state-of-the-art tools dedicated to the estimation AGS. Both ags.sh and acn.sh show comparable or higher accuracy than existing tools used to estimate these traits. To exemplify the applicability of both tools, we analyzed the 139 prokaryotic metagenomes of TARA Oceans and revealed the ecological strategies associated with different water layers. CONCLUSION: We took advantage of recent advances in gene annotation to develop the ags.sh and acn.sh tools to combine easy tool usage with fast and accurate performance. Our tools compute the AGS and ACN metagenomic traits on unassembled metagenomes and allow researchers to improve their metagenomic data analysis to gain deeper insights into microorganisms' ecology. The ags.sh and acn.sh tools are publicly available using Docker container technology at https://github.com/pereiramemo/AGS-and-ACN-tools .


Assuntos
Dosagem de Genes , Tamanho do Genoma , Metagenoma/genética , Metagenômica/métodos , RNA Ribossômico 16S/genética , Benchmarking , Variações do Número de Cópias de DNA , Bases de Dados Genéticas , Oceanos e Mares , Fatores de Tempo
3.
Genome Biol ; 20(1): 153, 2019 08 02.
Artigo em Inglês | MEDLINE | ID: mdl-31375138

RESUMO

We describe a method that adds long-read sequencing to a mix of technologies used to assemble a highly complex cattle rumen microbial community, and provide a comparison to short read-based methods. Long-read alignments and Hi-C linkage between contigs support the identification of 188 novel virus-host associations and the determination of phage life cycle states in the rumen microbial community. The long-read assembly also identifies 94 antimicrobial resistance genes, compared to only seven alleles in the short-read assembly. We demonstrate novel techniques that work synergistically to improve characterization of biological features in a highly complex rumen microbial community.


Assuntos
Resistência Microbiana a Medicamentos/genética , Metagenômica/métodos , Microbiota/genética , Análise de Sequência de DNA/métodos , Vírus/genética , Animais , Bovinos , Repetições Palindrômicas Curtas Agrupadas e Regularmente Espaçadas , Transferência Genética Horizontal , Genes Microbianos , Fases de Leitura Aberta , Prófagos/genética , Rúmen/microbiologia , Rúmen/virologia , Vírus/isolamento & purificação
4.
Genome Biol ; 20(1): 154, 2019 08 06.
Artigo em Inglês | MEDLINE | ID: mdl-31387630

RESUMO

We develop a metagenomic data analysis pipeline, MicroPro, that takes into account all reads from known and unknown microbial organisms and associates viruses with complex diseases. We utilize MicroPro to analyze four metagenomic datasets relating to colorectal cancer, type 2 diabetes, and liver cirrhosis and show that including reads from unknown organisms significantly increases the prediction accuracy of the disease status for three of the four datasets. We identify new microbial organisms associated with these diseases and show viruses play important prediction roles in colorectal cancer and liver cirrhosis, but not in type 2 diabetes. MicroPro is freely available at https://github.com/zifanzhu/MicroPro .


Assuntos
Doença , Metagenômica/métodos , Microbiota/genética , Software , Fenômenos Fisiológicos Virais , Neoplasias Colorretais/virologia , Diabetes Mellitus Tipo 2/virologia , Sequenciamento de Nucleotídeos em Larga Escala , Humanos , Cirrose Hepática/virologia
5.
Nat Biotechnol ; 37(8): 953-961, 2019 08.
Artigo em Inglês | MEDLINE | ID: mdl-31375809

RESUMO

Ruminants provide essential nutrition for billions of people worldwide. The rumen is a specialized stomach that is adapted to the breakdown of plant-derived complex polysaccharides. The genomes of the rumen microbiota encode thousands of enzymes adapted to digestion of the plant matter that dominates the ruminant diet. We assembled 4,941 rumen microbial metagenome-assembled genomes (MAGs) using approximately 6.5 terabases of short- and long-read sequence data from 283 ruminant cattle. We present a genome-resolved metagenomics workflow that enabled assembly of bacterial and archaeal genomes that were at least 80% complete. Of note, we obtained three single-contig, whole-chromosome assemblies of rumen bacteria, two of which represent previously unknown rumen species, assembled from long-read data. Using our rumen genome collection we predicted and annotated a large set of rumen proteins. Our set of rumen MAGs increases the rate of mapping of rumen metagenomic sequencing reads from 15% to 50-70%. These genomic and protein resources will enable a better understanding of the structure and functions of the rumen microbiota.


Assuntos
Archaea/genética , Bactérias/genética , Metagenoma , Metagenômica/métodos , Rúmen/microbiologia , Animais , Proteínas Arqueais , Proteínas de Bactérias , Bovinos/microbiologia , Bases de Dados de Proteínas , Genoma Arqueal , Genoma Bacteriano , Filogenia , Ovinos/microbiologia
6.
Nat Commun ; 10(1): 3066, 2019 07 11.
Artigo em Inglês | MEDLINE | ID: mdl-31296857

RESUMO

Metagenomic sequence classification should be fast, accurate and information-rich. Emerging long-read sequencing technologies promise to improve the balance between these factors but most existing methods were designed for short reads. MetaMaps is a new method, specifically developed for long reads, capable of mapping a long-read metagenome to a comprehensive RefSeq database with >12,000 genomes in <16 GB or RAM on a laptop computer. Integrating approximate mapping with probabilistic scoring and EM-based estimation of sample composition, MetaMaps achieves >94% accuracy for species-level read assignment and r2 > 0.97 for the estimation of sample composition on both simulated and real data when the sample genomes or close relatives are present in the classification database. To address novel species and genera, which are comparatively harder to predict, MetaMaps outputs mapping locations and qualities for all classified reads, enabling functional studies (e.g. gene presence/absence) and detection of incongruities between sample and reference genomes.


Assuntos
Biologia Computacional/métodos , Análise de Dados , Metagenômica/métodos , Algoritmos , Conjuntos de Dados como Assunto , Sequenciamento de Nucleotídeos em Larga Escala/métodos , Metagenoma/genética , Microbiota/genética , Análise de Sequência de DNA/métodos , Software
7.
BMC Infect Dis ; 19(1): 591, 2019 Jul 08.
Artigo em Inglês | MEDLINE | ID: mdl-31286879

RESUMO

BACKGROUND: We report a rare case of Toscana virus infection imported into Switzerland in a 23-year old man who travelled to Imperia (Italy) 10 days before onset of symptoms. Symptoms included both meningitis and as well epididymitis. This is only the fourth case of Toscana virus reported in Switzerland. CASE PRESENTATION: The patient presented with lymphocytic meningitis and scrotal pain due to epididymitis. Meningitis was initially treated with ceftriaxone. Herpes simplex, tick-borne encephalitis, enterovirus, measles, mumps, rubella and Treponema pallidum were excluded with specific polymerase chain reaction (PCR) or serology. In support of routine diagnostic PCR and serology assays, unbiased viral metagenomic sequencing was performed of cerebrospinal fluid and serum. Toscana virus infection was identified in cerebrospinal fluid and the full coding sequence could be obtained. Specific PCR in cerebrospinal fluid and blood and serology with Immunoglobulin (Ig) M and IgG against Toscana virus confirmed our diagnosis. Neurological symptoms recovered spontaneously after 5 days. CONCLUSIONS: This case of Toscana virus infection highlights the benefits of unbiased metagenomic sequencing to support routine diagnostics in rare or unexpected viral infections. With increasing travel histories of patients, physicians should be aware of imported Toscana virus as the agent for viral meningitis and meningoencephalitis.


Assuntos
Infecções por Bunyaviridae , Epididimite , Meningite Viral , Metagenômica/métodos , Vírus da Febre do Flebótomo Napolitano , Adulto , Anticorpos Antivirais/sangue , Infecções por Bunyaviridae/diagnóstico , Infecções por Bunyaviridae/imunologia , Infecções por Bunyaviridae/virologia , Epididimite/diagnóstico , Epididimite/imunologia , Epididimite/virologia , Humanos , Itália , Masculino , Meningite Viral/diagnóstico , Meningite Viral/imunologia , Meningite Viral/virologia , Técnicas de Diagnóstico Molecular , RNA Viral/genética , Vírus da Febre do Flebótomo Napolitano/genética , Vírus da Febre do Flebótomo Napolitano/imunologia , Análise de Sequência de RNA , Suíça , Adulto Jovem
8.
Nat Biotechnol ; 37(8): 877-883, 2019 08.
Artigo em Inglês | MEDLINE | ID: mdl-31332325

RESUMO

Spatial structuring is important for the maintenance of natural ecological systems1,2. Many microbial communities, including the gut microbiome, display intricate spatial organization3-9. Mapping the biogeography of bacteria can shed light on interactions that underlie community functions10-12, but existing methods cannot accommodate the hundreds of species that are found in natural microbiomes13-17. Here we describe metagenomic plot sampling by sequencing (MaPS-seq), a culture-independent method to characterize the spatial organization of a microbiome at micrometer-scale resolution. Intact microbiome samples are immobilized in a gel matrix and cryofractured into particles. Neighboring microbial taxa in the particles are then identified by droplet-based encapsulation, barcoded 16S rRNA amplification and deep sequencing. Analysis of three regions of the mouse intestine revealed heterogeneous microbial distributions with positive and negative co-associations between specific taxa. We identified robust associations between Bacteroidales taxa in all gut compartments and showed that phylogenetically clustered local regions of bacteria were associated with a dietary perturbation. Spatial metagenomics could be used to study microbial biogeography in complex habitats.


Assuntos
Microbioma Gastrointestinal , Genoma Bacteriano , Metagenômica/métodos , Animais , Código de Barras de DNA Taxonômico , DNA Bacteriano/genética , Metagenoma , Camundongos , Técnicas Analíticas Microfluídicas , RNA Bacteriano/genética , RNA Ribossômico 16S , Análise de Sequência de DNA/métodos
9.
Nat Biotechnol ; 37(8): 937-944, 2019 08.
Artigo em Inglês | MEDLINE | ID: mdl-31359005

RESUMO

Characterization of microbiomes has been enabled by high-throughput metagenomic sequencing. However, existing methods are not designed to combine reads from short- and long-read technologies. We present a hybrid metagenomic assembler named OPERA-MS that integrates assembly-based metagenome clustering with repeat-aware, exact scaffolding to accurately assemble complex communities. Evaluation using defined in vitro and virtual gut microbiomes revealed that OPERA-MS assembles metagenomes with greater base pair accuracy than long-read (>5×; Canu), higher contiguity than short-read (~10× NGA50; MEGAHIT, IDBA-UD, metaSPAdes) and fewer assembly errors than non-metagenomic hybrid assemblers (2×; hybridSPAdes). OPERA-MS provides strain-resolved assembly in the presence of multiple genomes of the same species, high-quality reference genomes for rare species (<1%) with ~9× long-read coverage and near-complete genomes with higher coverage. We used OPERA-MS to assemble 28 gut metagenomes of antibiotic-treated patients, and showed that the inclusion of long nanopore reads produces more contiguous assemblies (200× improvement over short-read assemblies), including more than 80 closed plasmid or phage sequences and a new 263 kbp jumbo phage. High-quality hybrid assemblies enable an exquisitely detailed view of the gut resistome in human patients.


Assuntos
Bactérias/efeitos dos fármacos , Bactérias/genética , Metagenômica/métodos , Microbiota/efeitos dos fármacos , Análise de Sequência de DNA/métodos , Antibacterianos/farmacologia , Farmacorresistência Bacteriana , Fezes/microbiologia , Sequenciamento de Nucleotídeos em Larga Escala/métodos , Humanos , Metagenoma , Nanoporos , Software
10.
Nat Methods ; 16(8): 731-736, 2019 08.
Artigo em Inglês | MEDLINE | ID: mdl-31308552

RESUMO

Metagenomic sequencing has enabled detailed investigation of diverse microbial communities, but understanding their spatiotemporal variability remains an important challenge. Here, we present decomposition of variance using replicate sampling (DIVERS), a method based on replicate sampling and spike-in sequencing. The method quantifies the contributions of temporal dynamics, spatial sampling variability, and technical noise to the variances and covariances of absolute bacterial abundances. We applied DIVERS to investigate a high-resolution time series of the human gut microbiome and a spatial survey of a soil bacterial community in Manhattan's Central Park. Our analysis showed that in the gut, technical noise dominated the abundance variability for nearly half of the detected taxa. DIVERS also revealed substantial spatial heterogeneity of gut microbiota, and high temporal covariances of taxa within the Bacteroidetes phylum. In the soil community, spatial variability primarily contributed to abundance fluctuations at short time scales (weeks), while temporal variability dominated at longer time scales (several months).


Assuntos
Algoritmos , Bactérias/genética , Fezes/microbiologia , Microbioma Gastrointestinal , Metagenômica/métodos , Microbiologia do Solo , Análise Espaço-Temporal , Bactérias/classificação , Humanos , RNA Ribossômico 16S , Análise de Sequência de DNA , Manejo de Espécimes
11.
APMIS ; 127(10): 660-670, 2019 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-31344275

RESUMO

Multiple approaches were employed to detect pathogens from bone margins associated with Diabetic Foot Osteomyelitis (DFO). Intra-operative bone specimens of 14 consecutive subjects with suspected DFO were collected over a six-month study period from Liverpool Hospital. Infected bone and a proximal bone margins presumed to be 'clean/non-infected' were collected. Bone material was subjected to conventional culture, DNA sequencing and microscopy. In total, eight of 14 (57%) proximal bone margins had no growth by conventional culture but were identified in all proximal bone specimens by DNA sequencing. Proximal margins had lower median total microbial counts than infected specimens, but these differences were not statistically significant. Pathogens identified by sequencing in infected specimens were identified in proximal margins and the microbiomes were similar (ANOSIM = 0.02, p = 0.59). Using a combination of SEM and/or PNA-FISH, we visualized the presence of microorganisms in infected bone specimens and their corresponding proximal margins of seven patients (50%) with DFO. We identify that bacteria can still reside in what seems to be proximal 'clean' margins. The significance and implications of clinical outcomes requires further analysis from a larger sample size that incorporates differences in surgical and post-operative approaches, correlating any outcomes back to culture-sequence findings.


Assuntos
Bactérias/isolamento & purificação , Técnicas Bacteriológicas/métodos , Osso e Ossos/microbiologia , Pé Diabético/microbiologia , Histocitoquímica/métodos , Metagenômica/métodos , Osteomielite/microbiologia , Bactérias/classificação , Bactérias/genética , Osso e Ossos/cirurgia , Pé Diabético/patologia , Pé Diabético/cirurgia , Humanos , Hibridização in Situ Fluorescente , Microscopia Eletrônica de Varredura , Osteomielite/patologia , Osteomielite/cirurgia , Análise de Sequência de DNA
13.
BMC Genomics ; 20(Suppl 5): 423, 2019 Jun 06.
Artigo em Inglês | MEDLINE | ID: mdl-31167634

RESUMO

BACKGROUND: High throughput sequencing has spurred the development of metagenomics, which involves the direct analysis of microbial communities in various environments such as soil, ocean water, and the human body. Many existing methods based on marker genes or k-mers have limited sensitivity or are too computationally demanding for many users. Additionally, most work in metagenomics has focused on bacteria and archaea, neglecting to study other key microbes such as viruses and eukaryotes. RESULTS: Here we present a method, MiCoP (Microbiome Community Profiling), that uses fast-mapping of reads to build a comprehensive reference database of full genomes from viruses and eukaryotes to achieve maximum read usage and enable the analysis of the virome and eukaryome in each sample. We demonstrate that mapping of metagenomic reads is feasible for the smaller viral and eukaryotic reference databases. We show that our method is accurate on simulated and mock community data and identifies many more viral and fungal species than previously-reported results on real data from the Human Microbiome Project. CONCLUSIONS: MiCoP is a mapping-based method that proves more effective than existing methods at abundance profiling of viruses and eukaryotes in metagenomic samples. MiCoP can be used to detect the full diversity of these communities. The code, data, and documentation are publicly available on GitHub at: https://github.com/smangul1/MiCoP .


Assuntos
Biologia Computacional/métodos , Fungos/genética , Marcadores Genéticos , Metagenômica/métodos , Microbiota , Análise de Sequência de DNA/métodos , Vírus/genética , Algoritmos , Fungos/classificação , Genoma Fúngico , Genoma Viral , Sequenciamento de Nucleotídeos em Larga Escala/métodos , Humanos , Vírus/classificação
15.
Nat Biotechnol ; 37(7): 783-792, 2019 07.
Artigo em Inglês | MEDLINE | ID: mdl-31235920

RESUMO

The gold standard for clinical diagnosis of bacterial lower respiratory infections (LRIs) is culture, which has poor sensitivity and is too slow to guide early, targeted antimicrobial therapy. Metagenomic sequencing could identify LRI pathogens much faster than culture, but methods are needed to remove the large amount of human DNA present in these samples for this approach to be feasible. We developed a metagenomics method for bacterial LRI diagnosis that features efficient saponin-based host DNA depletion and nanopore sequencing. Our pilot method was tested on 40 samples, then optimized and tested on a further 41 samples. Our optimized method (6 h from sample to result) was 96.6% sensitive and 41.7% specific for pathogen detection compared with culture and we could accurately detect antibiotic resistance genes. After confirmatory quantitative PCR and pathobiont-specific gene analyses, specificity and sensitivity increased to 100%. Nanopore metagenomics can rapidly and accurately characterize bacterial LRIs and might contribute to a reduction in broad-spectrum antibiotic use.


Assuntos
Bactérias/isolamento & purificação , Bronquite/diagnóstico , DNA Bacteriano/genética , Metagenômica/métodos , Nanoporos , Pneumonia Bacteriana/diagnóstico , Antibacterianos/farmacologia , Bactérias/efeitos dos fármacos , Bactérias/genética , Bronquite/microbiologia , Farmacorresistência Bacteriana/genética , Genoma Bacteriano , Sequenciamento de Nucleotídeos em Larga Escala/métodos , Humanos , Projetos Piloto , Pneumonia Bacteriana/microbiologia , Sensibilidade e Especificidade
16.
BMC Bioinformatics ; 20(1): 305, 2019 Jun 04.
Artigo em Inglês | MEDLINE | ID: mdl-31164077

RESUMO

BACKGROUND: Strain-level RNA virus characterization is essential for developing prevention and treatment strategies. Viral metagenomic data, which can contain sequences of both known and novel viruses, provide new opportunities for characterizing RNA viruses. Although there are a number of pipelines for analyzing viruses in metagenomic data, they have different limitations. First, viruses that lack closely related reference genomes cannot be detected with high sensitivity. Second, strain-level analysis is usually missing. RESULTS: In this study, we developed a hybrid pipeline named TAR-VIR that reconstructs viral strains without relying on complete or high-quality reference genomes. It is optimized for identifying RNA viruses from metagenomic data by combining an effective read classification method and our in-house strain-level de novo assembly tool. TAR-VIR was tested on both simulated and real viral metagenomic data sets. The results demonstrated that TAR-VIR competes favorably with other tested tools. CONCLUSION: TAR-VIR can be used standalone for viral strain reconstruction from metagenomic data. Or, its read recruiting stage can be used with other de novo assembly tools for superior viral functional and taxonomic analyses. The source code and the documentation of TAR-VIR are available at https://github.com/chjiao/TAR-VIR .


Assuntos
Vírus de RNA/genética , Software , Humanos , Metagenômica/métodos , Vírus de RNA/classificação , Análise de Sequência de RNA
17.
BMC Infect Dis ; 19(1): 495, 2019 Jun 04.
Artigo em Inglês | MEDLINE | ID: mdl-31164085

RESUMO

BACKGROUND: There is currently no research on the diagnostic value of metagenomic next-generation sequencing (mNGS) for a single pathogens in CSF. The aim of this study was to analyse the value of mNGS for identifying Streptococcus pneumoniae (S. pneumoniae) in paediatric bacterial meningitis. METHODS: Bacterial meningitis (BM) cases from October 23, 2014, to December 31, 2016, and December 1, 2017, to July 31, 2018 at Beijing Children's Hospital were reviewed. Clinical features and pathogens were analysed. RESULTS: We diagnosed 135 patients with BM in this study. A total of 43 S. pneumoniae were identified by combination methods. 26/135 (19.3%) patients had positive results in S. pneumoniae by blood and/or cerebrospinal fluid (CSF) culture. Alere BinaxNow®Streptococcus pneumoniae Antigen test was positive in 35/135(25.9%) cases. 32/135 (23.7%) S. pneumoniae were identified by mNGS. Six CSF samples were identified as S. pneumoniae only by mNGS technology. Taking culture as the gold standard, the sensitivity and specificity of mNGS for diagnosing S. pneumoniae meningitis were 73.1 and 88.1%, respectively. The positive predictive value (PPV) and negative predictive value (NPV) of diagnosing S. pneumoniae meningitis by mNGS were 59.4 and 93.2%, respectively. When comparison between mNGS and combined tests (culture and Alere BinaxNow®Streptococcus pneumoniae Antigen test), the sensitivity and specificity of mNGS for S. pneumoniae identification were 70.3 and 93.9%, the PPV and NPV in the identification of S. pneumoniae by mNGS were 81.4 and 89.3%, respectively. The difference in number of unique reads of S. pneumoniaein from CSF sample (< 14 days onset) and CSF sample (> 14 days from onset) was statistically significant (170.5 VS. 13, P = 0.019). The difference in the collected time of CSF for culture and mNGS was statistically significant (4 days VS. 14 days, P < 0.001). CONCLUSIONS: mNGS has high sensitivity and specificity for S. pneumoniae identification. The pathogen load (number of unique reads) of S. pneumonia is related to the CSF collection time. mNGS was less affected than culture by the use of antibiotics before CSF collection.


Assuntos
Sequenciamento de Nucleotídeos em Larga Escala , Meningites Bacterianas/diagnóstico , Metagenômica/métodos , Streptococcus pneumoniae/genética , Streptococcus pneumoniae/isolamento & purificação , Adolescente , Fatores Etários , Antígenos de Bactérias/análise , Antígenos de Bactérias/sangue , Antígenos de Bactérias/líquido cefalorraquidiano , Antígenos de Bactérias/genética , Criança , Pré-Escolar , Testes Diagnósticos de Rotina , Feminino , Humanos , Lactente , Recém-Nascido , Masculino , Meningites Bacterianas/sangue , Meningites Bacterianas/líquido cefalorraquidiano , Meningites Bacterianas/microbiologia , Pediatria/métodos , Reação em Cadeia da Polimerase/métodos , Valor Preditivo dos Testes , Sensibilidade e Especificidade
18.
BMC Bioinformatics ; 20(Suppl 11): 276, 2019 Jun 06.
Artigo em Inglês | MEDLINE | ID: mdl-31167633

RESUMO

BACKGROUND: A crucial task in metagenomic analysis is to annotate the function and taxonomy of the sequencing reads generated from a microbiome sample. In general, the reads can either be assembled into contigs and searched against reference databases, or individually searched without assembly. The first approach may suffer from fragmentary and incomplete assembly, while the second is hampered by the reduced functional signal contained in the short reads. To tackle these issues, we have previously developed GRASP (Guided Reference-based Assembly of Short Peptides), which accepts a reference protein sequence as input and aims to assemble its homologs from a database containing fragmentary protein sequences. In addition to a gene-centric assembly tool, GRASP also serves as a homolog search tool when using the assembled protein sequences as templates to recruit reads. GRASP has significantly improved recall rate (60-80% vs. 30-40%) compared to other homolog search tools such as BLAST. However, GRASP is both time- and space-consuming. Subsequently, we developed GRASPx, which is 30X faster than GRASP. Here, we present a completely redesigned algorithm, GRASP2, for this computational problem. RESULTS: GRASP2 utilizes Burrows-Wheeler Transformation (BWT) and FM-index to perform assembly graph generation, and reduces the search space by employing a fast ungapped alignment strategy as a filter. GRASP2 also explicitly generates candidate paths prior to alignment, which effectively uncouples the iterative access of the assembly graph and alignment matrix. This strategy makes the execution of the program more efficient under current computer architecture, and contributes to GRASP2's speedup. GRASP2 is 8-fold faster than GRASPx (and 250-fold faster than GRASP) and uses 8-fold less memory while maintaining the original high recall rate of GRASP. GRASP2 reaches ~ 80% recall rate compared to that of ~ 40% generated by BLAST, both at a high precision level (> 95%). With such a high performance, GRASP2 is only ~3X slower than BLASTP. CONCLUSION: GRASP2 is a high-performance gene-centric and homolog search tool with significant speedup compared to its predecessors, which makes GRASP2 a useful tool for metagenomics data analysis, GRASP2 is implemented in C++ and is freely available from http://www.sourceforge.net/projects/grasp2 .


Assuntos
Genes , Metagenômica/métodos , Análise de Sequência de DNA/métodos , Homologia de Sequência do Ácido Nucleico , Software , Algoritmos , Organismos Aquáticos/genética , Microbiota/genética , Curva ROC , Fatores de Tempo
19.
Gene ; 711: 143942, 2019 Aug 30.
Artigo em Inglês | MEDLINE | ID: mdl-31238090

RESUMO

In the work, metagenomic sequencing was conducted to investigate the microbial gene catalogue in two samples of phosphinothricin (PPT)-utilized soils from South China. The gene sets contained an overwhelming majority of prevalent microbial genes, and were largely shared between these two samples. Several genus with high abundance were shared, such as norank_d__Bacteria, Nitrososphaera, Candidatus_Nitrosotalea, Candidatus_Nitrosocosmicus, and Rhodanobacter. Bacitracin resistance genes (61.4%) were the most dominant antibiotic resistance genes in two samples, followed by multidrug resistance efflux pump (12.5%). A lot of common virulence factors with high abundance were found in two samples, such as Alginate, Capsule I, ClpC, FbpABC, and HitABC, many of which were used for the iron uptake system. Total 57 putative PPT acetyltransferase were annotated, and two of them were found to be novel putative acetyltransferases for acetylation and detoxification of PPT. In conclusion, the work revealed microbial gene catalogue of PPT-utilized soils and found two novel putative PPT acetyltransferases using metagenomics. The work facilitates the understanding of impact of PPT on complex microbial community structure and physiology resides in PPT-utilized soils. Moreover, two annotated PPT acetyltransferases show important potential for the development of transgenic herbicide-resistant crops.


Assuntos
Aminobutiratos/metabolismo , Bactérias/classificação , Proteínas de Bactérias/genética , Metagenômica/métodos , Bactérias/genética , Bactérias/metabolismo , Resistência Microbiana a Medicamentos , Filogenia , Análise de Sequência de DNA/métodos , Microbiologia do Solo
20.
BMC Bioinformatics ; 20(1): 310, 2019 Jun 11.
Artigo em Inglês | MEDLINE | ID: mdl-31185897

RESUMO

BACKGROUND: Metagenomics experiments often make inferences about microbial communities by sequencing 16S and 18S rRNA, and taxonomic assignment is a fundamental step in such studies. This paper addresses the weaknesses in two types of metrics commonly used by previous studies for measuring the performance of existing taxonomic assignment methods: Sequence count based metrics and Binary error measurement. These metrics made performance evaluation results biased, less informative and mutually incomparable. RESULTS: We investigated weaknesses in two types of metrics and proposed new performance metrics including Average Taxonomy Distance (ATD) and ATD_by_Taxa, together with the visualized ATD plot. CONCLUSIONS: By comparing the evaluation results from four popular taxonomic assignment methods across three test data sets, we found the new metrics more robust, informative and comparable.


Assuntos
Classificação/métodos , Metagenômica/métodos , Bactérias/genética , Bases de Dados Genéticas , Microbiota , Filogenia , RNA Ribossômico 16S/genética , RNA Ribossômico 18S/genética
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