Cyclic lipopeptide (CLP) synthesis gene clusters in pseudomonads display a high degree of synteny, and the structures of the peptides synthesized are very similar. Accordingly, the genomic island encoding the synthesis of syringomycin and syringopeptin in P. syringae pv. syringae closely resembles that of P. fluorescens In5, which contains genes coding for synthesis of the antifungal and anti-oomycete peptides nunamycin and nunapeptin, respectively. However, the regulation of syringomycin and syringopeptin synthesis is different from that of nunamycin and nunapeptin synthesis. While CLP synthesis in the plant pathogen P. syringae pv. syringae is induced by plant signal molecules, such compounds do not significantly influence synthesis of nunamycin and nunapeptin in P. fluorescens In5. Instead, fungal-associated molecules positively regulate antifungal peptide synthesis in P. fluorescens In5, while the synthesis of the global regulator GacA in P. fluorescens In5 is positively regulated by plant signal molecules but not fungal-associated molecules.

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.

Mobile genetic elements (MGEs) are instrumental in natural prokaryotic genome editing, permitting genome plasticity and allowing microbes to accumulate immense genetic diversity. MGEs include DNA elements such as plasmids, transposons and Insertion Sequences (IS-elements), as well as bacteriophages (phages), and they serve as a vast communal gene pool. These mobile DNA elements represent a human health risk as they can add new traits, such as antibiotic resistance or virulence, to a bacterial strain. Sequencing libraries targeting circular MGEs, referred to as mobilomes, allows the expansion of our current understanding of the mechanisms behind the mobility, prevalence and content of these elements. However, metamobilomes from bacterial communities are not studied to the same extent as metagenomics, partly because of methodological biases arising from multiple displacement amplification (MDA), often used in previous metamobilome publications. In this study, we show that MDA is detrimental to the detection of larger-sized plasmids if small plasmids are present by comparing the abundances of reads mapping to plasmids in a wastewater sample spiked with a mock community of selected plasmids with and without MDA. Furthermore, we show that it is possible to produce samples consisting almost exclusively of circular MGEs and obtain a catalog of larger, complete, circular MGEs from complex samples without the use of MDA. Importance Mobile genetic elements (MGEs) can transport genetic information between genomes in different bacterial species, adding new traits, potentially generating dangerous multidrug-resistant pathogens. In fact, plasmids and circular MGEs can encode bacterial genetic specializations such as virulence, resistance to metals, antimicrobial compounds, and bacteriophages, as well as the degradation of xenobiotics. For this reason, circular MGEs are crucial to investigate, but they are often missed in metagenomics and ecological studies. In this study, we present, for the first time, an improved method, which reduces the bias towards small MGEs and we demonstrate that this method can unveil larger, complete circular MGEs from complex samples without the use of multiple displacement amplification. This method may result in the detection of larger-sized plasmids that have hitherto remained unnoticed and therefore has the potential to reveal novel accessory genes, acting as possible targets in the development of preventive strategies directed at pathogens.

Despite the ecological significance of viral communities, phages remain understudied. Current genomic databases lack high-quality phage genomes linked to specific bacteria. Bacteria of the Erwinia genus are known to colonize the phyllosphere of plants, both as commensals and pathogens. We isolated three Erwinia billingiae phages; Zoomie, Pecta, and Snitter from organic household waste. Based on sequence similarity to their closest relatives, we propose that they represent three new genera “Pectavirus” within the Zobellviridae family, “Snittervirus” in the subfamily Tempevirinae, family Drexlerviridae, and “Zoomievirus” within the Autographviridae family, which may constitute a yet undefined sub-family together with the genus Limelightvirus.