Phages drive bacterial diversity, profoundly influencing microbial communities, from microbiomes to the drivers of global biogeochemical cycling. Aiming to broaden our understanding of Escherichia coli (MG1655, K-12) phages, we screened 188 Danish wastewater samples and isolated 136 phages. Ninety-two of these have genomic sequences with less than 95% similarity to known phages, while most map to existing genera several represent novel lineages. The isolated phages are highly diverse, estimated to represent roughly one-third of the true diversity of culturable virulent dsDNA Escherichia phages in Danish wastewater, yet almost half (40%) are not represented in metagenomic databases, emphasising the importance of isolating phages to uncover diversity. Seven viral families, Myoviridae, Siphoviridae, Podoviridae, Drexlerviridae, Chaseviridae, Autographviridae, and Microviridae, are represented in the dataset. Their genomes vary drastically in length from 5.3 kb to 170.8 kb, with a guanine and cytosine (GC) content ranging from 35.3% to 60.0%. Hence, even for a model host bacterium, substantial diversity remains to be uncovered. These results expand and underline the range of coliphage diversity and demonstrate how far we are from fully disclosing phage diversity and ecology.

Over a century of bacteriophage research has uncovered a plethora of fundamental aspects of their biology, ecology, and evolution. Furthermore, the introduction of community-level studies through metagenomics has revealed unprecedented insights on the impact that phages have on a range of ecological and physiological processes.

The phyllosphere microbiome plays an important role in plant fitness. Recently, bacteriophages have been shown to play a role in shaping the bacterial community composition of the phyllosphere. However, no studies on the diversity and abundance of phyllosphere bacteriophage communities have been carried out until now. In this study, we extracted, sequenced, and characterized the dsDNA and ssDNA viral community from a phyllosphere for the first time. We sampled leaves from winter wheat (Triticum aestivum), where we identified a total of 876 virus operational taxonomic units (vOTUs), mostly predicted to be bacteriophages with a lytic lifestyle. Remarkably, 848 of these vOTUs corresponded to new viral species, and we estimated a minimum of 2.0 × 10⁶ viral particles per leaf. These results suggest that the wheat phyllosphere harbors a large and active community of novel bacterial viruses. Phylloviruses have potential applications as biocontrol agents against phytopathogenic bacteria or as microbiome modulators to increase plant growth-promoting bacteria.

Despite the massive developments within culture-independent methods for detection of microorganisms during the last decade, culture-based methods remain a cornerstone in microbiology. Yet, the problem of rapid, accurate and inexpensive identification of bacterial isolates down to species/strain level remains unresolved. We have developed a new method for bacterial DNA enrichment and tagmentation allowing fast (99%) consensus reads. Whereas, the information from high quality reads is used to retrieve species level taxonomy, comparison of LCp allows for strain level differentiation.

A common procedure for studying the microbiome is binning the sequenced contigs into metagenome-assembled genomes. Currently, unsupervised and self-supervised deep learning based methods using co-abundance and sequence based motifs such as tetranucleotide frequencies are state-of-the-art for metagenome binning. Taxonomic labels derived from alignment based classification have not been widely used. Here, we propose TaxVAMB, a metagenome binning tool based on semi-supervised bi-modal variational autoencoders, combining tetranucleotide frequencies and contig co-abundances with contig annotations returned by any taxonomic classifier on any taxonomic rank. TaxVAMB outperforms all other binners on CAMI2 human microbiome datasets, returning on average 40% more near-complete assemblies than the next best binner. On real long-read datasets TaxVAMB recovers on average 13% more near-complete bins and 14% more species. When used in a single-sample setup, TaxVAMB on average returns 83% more high quality bins than VAMB. TaxVAMB bins incomplete genomes drastically better than any other tool, returning 255% more high quality bins of incomplete genomes than the next best binner. Our method has immediate research and industrial applications, as well as methodological novelty which can be translated to other biological problems with semi-supervised multimodal datasets.

Isolating single phages using plaque assays is a laborious and time-consuming process. Whether single isolated phages are the most lyse-effective, the most abundant in viromes, or those with the highest ability to make plaques in solid media is not well known. With the increasing accessibility of high-throughput sequencing, metaviromics is often used to describe viruses in environmental samples. By extracting and sequencing metaviromes from organic waste with and without exposure to a host-of-interest, we show a host-related phage community’s shift, as well as identify the most enriched phages. Moreover, we isolated plaque-forming single phages using the same virome–host matrix to observe how enrichments in liquid media correspond to the metaviromic data. In this study, we observed a significant shift (p = 0.015) of the 47 identified putative Pseudomonas phages with a minimum twofold change above zero in read abundance when adding a Pseudomonas syringae DC3000 host. Surprisingly, it appears that only two out of five plaque-forming phages from the same organic waste sample, targeting the Pseudomonas strain, were highly abundant in the metavirome, while the other three were almost absent despite host exposure. Lastly, our sequencing results highlight how long reads from Oxford Nanopore elevates the assembly quality of metaviromes, compared to short reads alone.