Naming the unnamed: over 65,000 Candidatus names for unnamed Archaea and Bacteria in the Genome Taxonomy Database.

Thousands of new bacterial and archaeal species and higher-level taxa are discovered each year through the analysis of genomes and metagenomes. The Genome Taxonomy Database (GTDB) provides hierarchical sequence-based descriptions and classifications for new and as-yet-unnamed taxa. However, bacterial nomenclature, as currently configured, cannot keep up with the need for new well-formed names. Instead, microbiologists have been forced to use hard-to-remember alphanumeric placeholder labels. Here, we exploit an approach to the generation of well-formed arbitrary Latinate names at a scale sufficient to name tens of thousands of unnamed taxa within GTDB. These newly created names represent an important resource for the microbiology community, facilitating communication between bioinformaticians, microbiologists and taxonomists, while populating the emerging landscape of microbial taxonomic and functional discovery with accessible and memorable linguistic labels.

Streptomyces odonnellii sp. nov., a proteolytic streptomycete isolated from soil under cerrado (savanna) vegetation cover.

A novel streptomycete, strain 594T, isolated from Brazilian soil collected under cerrado (savanna) vegetation cover is described. Strain 594T produced thermophilic chitinolytic proteases in assays containing feather meal and corn steep liquor as sole sources of carbon and nitrogen. The strain produced white to grey aerial mycelium and spiral chains of spiny-surfaced spores on the aerial mycelium and did not produce diffusible pigments. The ll-isomer of diaminopimelic acid was present in the cell wall and menaquinones were predominantly MK-9(H6) (52 %) and MK-9(H8) (30 %) with 6 % MK-9(H4) and slightly less than 1 % MK-9(H2). Polar lipids present were phosphatidylethanolamine, diphosphatidylglycerol, phosphatidylglycerol, phosphatidylinositol and an unknown phospholipid. The major fatty acids were anteiso-C15 : 0, anteiso-C16 : 0, anteiso-C14 : 0 and anteiso-C17 : 0. The G+C content of the genomic DNA was 70.4 mol%. Phylogenetic analysis of the nearly complete 16S rRNA gene sequence indicated that it differed from described Streptomyces species. Multilocus sequence analysis (MLSA) using five housekeeping genes (atpD, gyrB, rpoB, recA and trpB) comparing Streptomyces type strains showed that the MLSA distance of strain 594T to the most closely related species was greater than the 0.007 threshold. The in silico DNA-DNA relatedness between the genome sequence of strain 594T and that of the phylogenetically nearest species was well below the species level recommendation. There was thus multiple evidence justifying the description of this strain as representing a novel species, for which the name Streptomyces odonnellii sp. nov. is proposed. The type strain is 594T (=IMPPG 594T=DSM 41949T=NRRL B-24891T).

An in-depth evaluation of metagenomic classifiers for soil microbiomes.

BACKGROUND: Recent endeavours in metagenomics, exemplified by projects such as the human microbiome project and TARA Oceans, have illuminated the complexities of microbial biomes. A robust bioinformatic pipeline and meticulous evaluation of their methodology have contributed to the success of these projects. The soil environment, however, with its unique challenges, requires a specialized methodological exploration to maximize microbial insights. A notable limitation in soil microbiome studies is the dearth of soil-specific reference databases available to classifiers that emulate the complexity of soil communities. There is also a lack of in-vitro mock communities derived from soil strains that can be assessed for taxonomic classification accuracy. RESULTS: In this study, we generated a custom in-silico mock community containing microbial genomes commonly observed in the soil microbiome. Using this mock community, we simulated shotgun sequencing data to evaluate the performance of three leading metagenomic classifiers: Kraken2 (supplemented with Bracken, using a custom database derived from GTDB-TK genomes along with its own default database), Kaiju, and MetaPhlAn, utilizing their respective default databases for a robust analysis. Our results highlight the importance of optimizing taxonomic classification parameters, database selection, as well as analysing trimmed reads and contigs. Our study showed that classifiers tailored to the specific taxa present in our samples led to fewer errors compared to broader databases including microbial eukaryotes, protozoa, or human genomes, highlighting the effectiveness of targeted taxonomic classification. Notably, an optimal classifier performance was achieved when applying a relative abundance threshold of 0.001% or 0.005%. The Kraken2 supplemented with bracken, with a custom database demonstrated superior precision, sensitivity, F1 score, and overall sequence classification. Using a custom database, this classifier classified 99% of in-silico reads and 58% of real-world soil shotgun reads, with the latter identifying previously overlooked phyla using a custom database. CONCLUSION: This study underscores the potential advantages of in-silico methodological optimization in metagenomic analyses, especially when deciphering the complexities of soil microbiomes. We demonstrate that the choice of classifier and database significantly impacts microbial taxonomic profiling. Our findings suggest that employing Kraken2 with Bracken, coupled with a custom database of GTDB-TK genomes and fungal genomes at a relative abundance threshold of 0.001% provides optimal accuracy in soil shotgun metagenome analysis.

Proposal of names for 329 higher rank taxa defined in the Genome Taxonomy Database under two prokaryotic codes.

The Genome Taxonomy Database (GTDB) is a taxonomic framework that defines prokaryotic taxa as monophyletic groups in concatenated protein reference trees according to systematic criteria. This has resulted in a substantial number of changes to existing classifications (https://gtdb.ecogenomic.org). In the case of union of taxa, GTDB names were applied based on the priority of publication. The division of taxa or change in rank led to the formation of new Latin names above the rank of genus that were only made publicly available via the GTDB website without associated published taxonomic descriptions. This has sometimes led to confusion in the literature and databases. A number of the provisional GTDB names were later published in other studies, while many still lack authorships. To reduce further confusion, here we propose names and descriptions for 329 GTDB-defined prokaryotic taxa, 223 of which are suitable for validation under the International Code of Nomenclature of Prokaryotes (ICNP) and 49 under the Code of Nomenclature of Prokaryotes described from Sequence Data (SeqCode). For the latter, we designated 23 genomes as type material. An additional 57 taxa that do not currently satisfy the validation criteria of either code are proposed as Candidatus.

Metagenomics Databases for Bacteria.

The booming sequencing technologies have turned metagenomics into a widely used tool for microbe-related studies, especially in the areas of clinical medicine and ecology. Accordingly, the toolkit of metagenomics data analysis is growing stronger to provide multiple approaches for solving various biological questions and understanding the component and function of microbiome. As part of the toolkit, metagenomics databases play a central role in the creation and maintenance of processed data such as definition of taxonomic classifications, annotation of gene functions, sequence alignment, and phylogenetic tree inference. The availability of a large quantity of high-quality bacterial genomic sequences contributes significantly to the construction and update of metagenomics databases, which constitute the core resource for metagenomics data analysis at various scales. This chapter presents the key concepts, technical options, and challenges for metagenomics projects as well as the curation processes and versatile functions for the four representative bacterial metagenomics databases, including Greengenes, SILVA, Ribosomal Database Project (RDP), and Genome Taxonomy Database (GTDB).

Genome analysis of the candidate phylum MBNT15 bacterium from a boreal peatland predicted its respiratory versatility and dissimilatory iron metabolism.

Uncultured bacteria of the candidate phylum MBNT15, distantly related to Desulfobacterota, have been identified in a broad range of mostly organic-rich aquatic environments. We assembled a near-complete genome of a member of MBNT15 from a boreal peatland metagenome and used genomic data to analyze the metabolic pathways of this bacterium and its ecological role. This bacterium, designated SHF-111, was predicted to be rod shaped, it lacks flagellar machinery but twitching motility is encoded. Genome-based phylogenetic analysis supported the phylum-level classification of the MBNT15 lineage. Genome annotation and metabolic reconstruction revealed the presence of the Embden-Meyerhof, Entner-Doudoroff and pentose phosphate pathways, as well as the complete tricarboxylic acid (TCA) cycle, and suggested a facultatively anaerobic chemoheterotrophic lifestyle with the ability to ferment peptides, amino acids, fatty acids and simple sugars, and completely oxidize these substrates through aerobic and anaerobic respiration. The SHF-111 genome encodes multiple multiheme c-type cytochromes that probably enable dissimilatory iron reduction. Consistently, the relative abundance of MBNT15 in peatlands positively correlated with iron concentration. Apparently, in the wetland ecosystem, MBNT15 representatives play the role of scavengers, carrying out the complete mineralization of low molecular weight organic substances formed as a result of microbial degradation of complex polymeric substrates. Comparative genome analysis of the MBNT15 phylum revealed that vast majority of its members are capable of aerobic respiration and dissimilatory iron reduction and some species also can reduce sulfur and nitrogen compounds, but not sulfate. Based on phylogenetic and genomic analyses, the novel bacterium is proposed to be classified as Candidatus Deferrimicrobium borealis, within a candidate phylum Deferrimicrobiota.

Phylogenomics reveals the basis of adaptation of Pseudorhizobium species to extreme environments and supports a taxonomic revision of the genus.

The family Rhizobiaceae includes many genera of soil bacteria, often isolated for their association with plants. Herein, we investigate the genomic diversity of a group of Rhizobium species and unclassified strains isolated from atypical environments, including seawater, rock matrix or polluted soil. Based on whole-genome similarity and core genome phylogeny, we show that this group corresponds to the genus Pseudorhizobium. We thus reclassify Rhizobium halotolerans, R. marinum, R. flavum and R. endolithicum as P. halotolerans sp. nov., P. marinum comb. nov., P. flavum comb. nov. and P. endolithicum comb. nov., respectively, and show that P. pelagicum is a synonym of P. marinum. We also delineate a new chemolithoautotroph species, P. banfieldiae sp. nov., whose type strain is NT-26T (=DSM 106348T=CFBP 8663T). This genome-based classification was supported by a chemotaxonomic comparison, with increasing taxonomic resolution provided by fatty acid, protein and metabolic profiles. In addition, we used a phylogenetic approach to infer scenarios of duplication, horizontal transfer and loss for all genes in the Pseudorhizobium pangenome. We thus identify the key functions associated with the diversification of each species and higher clades, shedding light on the mechanisms of adaptation to their respective ecological niches. Respiratory proteins acquired at the origin of Pseudorhizobium were combined with clade-specific genes to enable different strategies for detoxification and nutrition in harsh, nutrient-poor environments.

A review of computational tools for generating metagenome-assembled genomes from metagenomic sequencing data.

Metagenomic sequencing provides a culture-independent avenue to investigate the complex microbial communities by constructing metagenome-assembled genomes (MAGs). A MAG represents a microbial genome by a group of sequences from genome assembly with similar characteristics. It enables us to identify novel species and understand their potential functions in a dynamic ecosystem. Many computational tools have been developed to construct and annotate MAGs from metagenomic sequencing, however, there is a prominent gap to comprehensively introduce their background and practical performance. In this paper, we have thoroughly investigated the computational tools designed for both upstream and downstream analyses, including metagenome assembly, metagenome binning, gene prediction, functional annotation, taxonomic classification, and profiling. We have categorized the commonly used tools into unique groups based on their functional background and introduced the underlying core algorithms and associated information to demonstrate a comparative outlook. Furthermore, we have emphasized the computational requisition and offered guidance to the users to select the most efficient tools. Finally, we have indicated current limitations, potential solutions, and future perspectives for further improving the tools of MAG construction and annotation. We believe that our work provides a consolidated resource for the current stage of MAG studies and shed light on the future development of more effective MAG analysis tools on metagenomic sequencing.

Out From the Shadows - Resolution of the Taxonomy of the Family Cryomorphaceae.

The family Cryomorphaceae for many years has been a poorly defined taxonomic group within the order Flavobacteriales, phylum Bacteroidetes. Members of the Cryomorphaceae, apparently consisting of multiple-family level clades, have been mostly but not exclusively detected in saline ecosystems. The problems with the taxonomy of this group have stemmed from inadequate resolution of taxonomic groups using 16S rRNA gene sequences, sparse numbers of cultivated taxa, and limited phenotypic distinctiveness. The Genome Tiaxonomc Database (GTDB), which is based on normalized taxonomic ranks includes Cryomorphaceae as containing the genera Owenweeksia and Schleiferia. This is at odds with the official taxonomy that places these genera in the family Schleiferiaceae. The other Cryomorphaceae affiliated species have even more uncertain taxonomic positions including Cryomorpha ignava. To clarify the taxonomy of Cryomorphaceae, genomes were generated for all type strains of the family Cryomorphaceae lacking such data. The GTDB-toolkit (GTDB-tk) was used to place taxa in the GTDB, which revealed novelty at the family level for some of these type strains. 16S rRNA gene sequences and concatenated protein sequences were used to further evaluate the taxonomy of the order Flavobacteriales. From the data, the GTDB enabled successful clarification of the taxonomy of the family Cryomorphaceae. A number of placeholder families were given Latinized names. It is proposed that the family Cryomorphaceae is emended to include only the species Cryomorpha ignava. The family Schleiferiaceae is emended to account for the expansion of its membership. Luteibaculum oceani represents a new family designated Luteibaculaceae fam. nov. Vicingus serpentipes is the representative of Vicingaceae fam. nov. while Salibacter halophilus represents Salibacteraceae fam. nov.

CAMITAX: Taxon labels for microbial genomes.

BACKGROUND: The number of microbial genome sequences is increasing exponentially, especially thanks to recent advances in recovering complete or near-complete genomes from metagenomes and single cells. Assigning reliable taxon labels to genomes is key and often a prerequisite for downstream analyses. FINDINGS: We introduce CAMITAX, a scalable and reproducible workflow for the taxonomic labelling of microbial genomes recovered from isolates, single cells, and metagenomes. CAMITAX combines genome distance-, 16S ribosomal RNA gene-, and gene homology-based taxonomic assignments with phylogenetic placement. It uses Nextflow to orchestrate reference databases and software containers and thus combines ease of installation and use with computational reproducibility. We evaluated the method on several hundred metagenome-assembled genomes with high-quality taxonomic annotations from the TARA Oceans project, and we show that the ensemble classification method in CAMITAX improved on all individual methods across tested ranks. CONCLUSIONS: While we initially developed CAMITAX to aid the Critical Assessment of Metagenome Interpretation (CAMI) initiative, it evolved into a comprehensive software package to reliably assign taxon labels to microbial genomes. CAMITAX is available under Apache License 2.0 at https://github.com/CAMI-challenge/CAMITAX.

Availability of Nanopore sequences in the genome taxonomy for Vibrionaceae systematics: Rumoiensis clade species as a test case.

Whole genome sequence comparisons have become essential for establishing a robust scheme in bacterial taxonomy. To generalize this genome-based taxonomy, fast, reliable, and cost-effective genome sequencing methodologies are required. MinION, the palm-sized sequencer from Oxford Nanopore Technologies, enables rapid sequencing of bacterial genomes using minimal laboratory resources. Here we tested the ability of Nanopore sequences for the genome-based taxonomy of Vibrionaceae and compared Nanopore-only assemblies to complete genomes of five Rumoiensis clade species: Vibrio aphrogenes, V. algivorus, V. casei, V. litoralis, and V. rumoiensis. Comparison of overall genome relatedness indices (OGRI) and multilocus sequence analysis (MLSA) based on Nanopore-only assembly and Illumina or hybrid assemblies revealed that errors in Nanopore-only assembly do not influence average nucleotide identity (ANI), in silico DNA-DNA hybridization (DDH), G+C content, or MLSA tree topology in Vibrionaceae. Our results show that the genome sequences from Nanopore-based approach can be used for rapid species identification based on the OGRI and MLSA.

Vibrio ishigakensis sp. nov., in Halioticoli clade isolated from seawater in Okinawa coral reef area, Japan.

Five novel strains showing non-motile, alginolytic, halophilic and fermentative features were isolated from seawater samples off Okinawa in coral reef areas. These strains were characterized by an advanced polyphasic taxonomy including genome based taxonomy using multilocus sequence analysis (MLSA) and in silico DNA-DNA similarity (in silico DDH). Phylogenetic analyses on the basis of 16S rRNA gene sequences revealed that the isolates could be assigned to the genus Vibrio, however they were not allocated into any distinct cluster with known Vibrionaceae species. MLSA based on eight protein-coding genes (gapA, gyrB, ftsZ, mreB, pyrH, recA, rpoA, and topA) showed the vibrios formed an outskirt branch of Halioticoli clade. The experimental DNA-DNA hybridization data revealed that the five strains were in the range of being defined as conspecific but separate from nine Halioticoli clade species. The G+C contents of the Vibrio ishigakensis strains were 47.3-49.1mol%. Both Amino Acid Identity and Average Nucleotide Identity of the strain C1(T) against Vibrio ezurae HDS1-1(T), Vibrio gallicus HT2-1(T), Vibrio halioticoli IAM 14596(T), Vibrio neonatus HDD3-1(T) and Vibrio superstes G3-29(T) showed less than 95% similarity. The genome-based taxonomic approach by means of in silico DDH values also supports the V. ishigakensis strains being distinct from the other known Halioticoli clade species. Sixteen traits (growth temperature range, DNase and lipase production, indole production, and assimilation of 10 carbon compounds) distinguished these strains from Halioticoli clade species. The names V. ishigakensis sp. nov. is proposed for the species of Halioticoli clade, with C1(T) as the type strain (JCM 19231(T)=LMG 28703(T)).