Obligate cross-feeding expands the metabolic niche of bacteria.

Bacteria frequently engage in obligate metabolic mutualisms with other microorganisms. However, it remains generally unclear how the resulting metabolic dependencies affect the ecological niche space accessible to the whole consortium relative to the niche space available to its constituent individuals. Here we address this issue by systematically cultivating metabolically dependent strains of different bacterial species either individually or as pairwise cocultures in a wide range of carbon sources. Our results show that obligate cross-feeding is significantly more likely to expand the metabolic niche space of interacting bacterial populations than to contract it. Moreover, niche expansion occurred predominantly between two specialist taxa and correlated positively with the phylogenetic distance between interaction partners. Together, our results demonstrate that obligate cross-feeding can significantly expand the ecological niche space of interacting bacterial genotypes, thus explaining the widespread occurrence of this type of ecological interaction in natural microbiomes.

Isolation and Characterization of Shewanella Phage Thanatos Infecting and Lysing Shewanella oneidensis and Promoting Nascent Biofilm Formation.

Species of the genus Shewanella are widespread in nature in various habitats, however, little is known about phages affecting Shewanella sp. Here, we report the isolation of phages from diverse freshwater environments that infect and lyse strains of Shewanella oneidensis and other Shewanella sp. Sequence analysis and microscopic imaging strongly indicate that these phages form a so far unclassified genus, now named Shewanella phage Thanatos, which can be positioned within the subfamily of Tevenvirinae (Duplodnaviria; Heunggongvirae; Uroviricota; Caudoviricetes; Caudovirales; Myoviridae; Tevenvirinae). We characterized one member of this group in more detail using S. oneidensis MR-1 as a host. Shewanella phage Thanatos-1 possesses a prolate icosahedral capsule of about 110 nm in height and 70 nm in width and a tail of about 95 nm in length. The dsDNA genome exhibits a GC content of about 34.5%, has a size of 160.6 kbp and encodes about 206 proteins (92 with an annotated putative function) and two tRNAs. Out of those 206, MS analyses identified about 155 phage proteins in PEG-precipitated samples of infected cells. Phage attachment likely requires the outer lipopolysaccharide of S. oneidensis, narrowing the phage's host range. Under the applied conditions, about 20 novel phage particles per cell were produced after a latent period of approximately 40 min, which are stable at a pH range from 4 to 12 and resist temperatures up to 55°C for at least 24 h. Addition of Thanatos to S. oneidensis results in partial dissolution of established biofilms, however, early exposure of planktonic cells to Thanatos significantly enhances biofilm formation. Taken together, we identified a novel genus of Myophages affecting S. oneidensis communities in different ways.

Characterization of ExeM, an Extracellular Nuclease of Shewanella oneidensis MR-1.

Bacterial extracellular nucleases have multiple functions in processes as diverse as nutrient acquisition, natural transformation, biofilm formation, or defense against neutrophil extracellular traps (NETs). Here we explored the properties of ExeM in Shewanella oneidensis MR-1, an extracellular nuclease, which is widely conserved among species of Shewanella, Vibrio, Aeromonas, and others. In S. oneidensis, ExeM is crucial for normal biofilm formation. In vitro activity measurements on heterologously produced ExeM revealed that this enzyme is a sugar-unspecific endonuclease, which requires Ca2+ and Mg2+/Mn2+ as co-factors for full activity. ExeM was almost exclusively localized to the cytoplasmic membrane fraction, even when a putative C-terminal membrane anchor was deleted. In contrast, ExeM was not detected in medium supernatants. Based on the results we hypothesize that ExeM predominantly interacts with DNA in close proximity to the cell, e.g., to promote biofilm formation and defense against NETs, or to control uptake of DNA.

Functional specificity of extracellular nucleases of Shewanella oneidensis MR-1.

Bacterial species such as Shewanella oneidensis MR-1 require extracellular nucleolytic activity for the utilization of extracellular DNA (eDNA) as a source of nutrients and for the turnover of eDNA as a structural matrix component during biofilm formation. We have previously characterized two extracellular nucleases of S. oneidensis MR-1, ExeM and ExeS. Although both are involved in biofilm formation, they are not specifically required for the utilization of eDNA as a nutrient. Here we identified and characterized EndA, a third extracellular nuclease of Shewanella. The heterologously overproduced and purified protein was highly active and rapidly degraded linear and supercoiled DNAs of various origins. Divalent metal ions (Mg(2+) or Mn(2+)) were required for function. endA is cotranscribed with phoA, an extracellular phosphatase, and is not upregulated upon phosphostarvation. Deletion of endA abolished both extracellular degradation of DNA by S. oneidensis MR-1 and the ability to use eDNA as a sole source of phosphorus. PhoA is not strictly required for the exploitation of eDNA as a nutrient. The activity of EndA prevents the formation of large cell aggregates during planktonic growth. However, in contrast to the findings for ExeM, endA deletion had only minor effects on biofilm formation. The findings strongly suggest that the extracellular nucleases of S. oneidensis exert specific functions required under different conditions.