Genome streamlining in Parcubacteria transitioning from soil to groundwater.

BACKGROUND: To better understand the influence of habitat on the genetic content of bacteria, with a focus on members of Candidate Phyla Radiation (CPR) bacteria, we studied the effects of transitioning from soil via seepage waters to groundwater on genomic composition of ultra-small Parcubacteria, the dominating CPR class in seepage waters, using genome resolved metagenomics. RESULTS: Bacterial metagenome-assembled genomes (MAGs), (318 total, 32 of Parcubacteria) were generated from seepage waters and compared directly to groundwater counterparts. The estimated average genome sizes of members of major phyla Proteobacteria, Bacteroidota and Cand. Patescibacteria (Candidate Phyla Radiation - CPR bacteria) were significantly higher in soil-seepage water as compared to their groundwater counterparts. Seepage water Parcubacteria (Paceibacteria) exhibited 1.18-fold greater mean genome size and 2-fold lower mean proportion of pseudogenes than those in groundwater. Bacteroidota and Proteobacteria also showed a similar trend of reduced genomes in groundwater compared to seepage. While exploring gene loss and adaptive gains in closely related CPR lineages in groundwater, we identified a membrane protein, and a lipoglycopeptide resistance gene unique to a seepage Parcubacterium genome. A nitrite reductase gene was also identified and was unique to the groundwater Parcubacteria genomes, likely acquired from other planktonic microbes via horizontal gene transfer. CONCLUSIONS: Overall, our data suggest that bacteria in seepage waters, including ultra-small Parcubacteria, have significantly larger genomes and higher metabolic enrichment than their groundwater counterparts, highlighting possible genome streamlining of the latter in response to habitat selection in an oligotrophic environment.

Iron coatings on carbonate rocks shape the attached bacterial aquifer community.

Most studies of groundwater ecosystems target planktonic microbes, which are easily obtained via water samples. In contrast, little is known about the diversity and function of microbes adhering to rock surfaces, particularly to consolidated rocks. To investigate microbial attachment to rock surfaces, we incubated rock chips from fractured aquifers in limestone-mudstone alternations in bioreactors fed with groundwater from two wells representing oxic and anoxic conditions. Half of the chips were coated with iron oxides, representing common secondary mineralization in fractured rock. Our time-series analysis showed bacteria colonizing the chips within two days, reaching cell numbers up to 4.16 × 105 cells/mm2 after 44 days. Scanning electron microscopy analyses revealed extensive colonization but no multi-layered biofilms, with chips from oxic bioreactors more densely colonized than from anoxic ones. Estimated attached-to-planktonic cell ratios yielded values of up to 106: 1 and 103: 1, for oxic and anoxic aquifers, respectively. We identified distinct attached and planktonic communities with an overlap between 17 % and 42 %. Oxic bioreactors were dominated by proteobacterial genera Aquabacterium and Rhodoferax, while Rheinheimera and Simplicispira were the key players of anoxic bioreactors. Motility, attachment, and biofilm formation traits were predicted in major genera based on groundwater metagenome-assembled genomes and reference genomes. Early rock colonizers appeared to be facultative autotrophs, capable of fixing CO2 to synthesize biomass and a biofilm matrix. Late colonizers were predicted to possess biofilm degrading enzymes such as beta-glucosidase, beta-galactosidase, amylases. Fe-coated chips of both bioreactors featured more potential iron reducers and oxidizers than bare rock chips. As secondary minerals can also serve as energy source, they might favor primary production and thus contribute to subsurface ecosystem services like carbon fixation. Since most subsurface microbes seem to be attached, their contribution to ecosystem services should be considered in future studies.

Host Preference of Beneficial Commensals in a Microbially-Diverse Environment.

Gut bacteria are often described by the neutral term commensals. However, the more we learn about their interactions with hosts, the more apparent it becomes that gut commensals often contribute positively to host physiology and fitness. Whether hosts can prefer beneficial bacteria, and how they do so, is not clear. This is of particular interest in the case of the bacterivore C. elegans, which depends on bacteria as food source, but also as gut colonizers that contribute to its physiology, from development to immunity. It is further unclear to what extent worms living in their microbially-diverse habitats can sense and distinguish between beneficial bacteria, food, and pathogens. Focusing on Enterobacteriaceae and members of closely related families, we isolated gut bacteria from worms raised in compost microcosms, as well as bacteria from the respective environments and evaluated their contributions to host development. Most isolates, from worms or from the surrounding environment, promoted faster development compared to the non-colonizing E. coli food strain. Pantoea strains further showed differential contributions of gut isolates versus an environmental isolate. Characterizing bacterial ability to hinder pathogenic colonization with Pseudomonas aeruginosa, supported the trend of Pantoea gut commensals being beneficial, in contrast to the environmental strain. Interestingly, worms were attracted to the beneficial Pantoea strains, preferring them over non-beneficial bacteria, including the environmental Pantoea strain. While our understanding of the mechanisms underlying these host-microbe interactions are still rudimentary, the results suggest that hosts can sense and prefer beneficial commensals.

Selection of Anabaena sp. PCC 7938 as a Cyanobacterium Model for Biological ISRU on Mars.

Crewed missions to Mars are expected to take place in the coming decades. After short-term stays, a permanent presence will be desirable to enable a wealth of scientific discoveries. This will require providing crews with life-support consumables in amounts that are too large to be imported from Earth. Part of these consumables could be produced on site with bioprocesses, but the feedstock should not have to be imported. A solution under consideration lies in using diazotrophic, rock-weathering cyanobacteria as primary producers: fed with materials naturally available on site, they would provide the nutrients required by other organisms. This concept has recently gained momentum but progress is slowed by a lack of consistency across contributing teams, and notably of a shared model organism. With the hope to address this issue, we present the work performed to select our current model. We started with preselected strains from the Nostocaceae family. After sequencing the genome of Anabaena sp. PCC 7938-the only one not yet available-we compared the strains' genomic data to determine their relatedness and provide insights into their physiology. We then assessed and compared relevant features: chiefly, their abilities to utilize nutrients from Martian regolith, their resistance to perchlorates (toxic compounds present in the regolith), and their suitability as feedstock for secondary producers (here a heterotrophic bacterium and a higher plant). This led to the selection of Anabaena sp. PCC 7938, which we propose as a model cyanobacterium for the development of bioprocesses based on Mars's natural resources. IMPORTANCE The sustainability of crewed missions to Mars could be increased by biotechnologies which are connected to resources available on site via primary producers: diazotrophic, rock-leaching cyanobacteria. Indeed, this could greatly reduce the mass of payloads to be imported from Earth. The concept is gaining momentum but progress is hindered by a lack of consistency across research teams. We consequently describe the selection process that led to the choice of our model strain, demonstrate its relevance to the field, and propose it as a shared model organism. We expect this contribution to support the development of cyanobacterium-based biotechnologies on Mars.

Single-colony sequencing reveals microbe-by-microbiome phylosymbiosis between the cyanobacterium Microcystis and its associated bacteria.

BACKGROUND: Cyanobacteria from the genus Microcystis can form large mucilaginous colonies with attached heterotrophic bacteria-their microbiome. However, the nature of the relationship between Microcystis and its microbiome remains unclear. Is it a long-term, evolutionarily stable association? Which partners benefit? Here we report the genomic diversity of 109 individual Microcystis colonies-including cyanobacteria and associated bacterial genomes-isolated in situ and without culture from Lake Champlain, Canada and Pampulha Reservoir, Brazil. RESULTS: We identified 14 distinct Microcystis genotypes from Canada, of which only two have been previously reported, and four genotypes specific to Brazil. Microcystis genetic diversity was much greater between than within colonies, consistent with colony growth by clonal expansion rather than aggregation of Microcystis cells. We also identified 72 bacterial species in the microbiome. Each Microcystis genotype had a distinct microbiome composition, and more closely related genotypes had more similar microbiomes. This pattern of phylosymbiosis could be explained by co-phylogeny in only two out of the nine most prevalent associated bacterial genera, Roseomonas and Rhodobacter. These phylogenetically associated genera could enrich the metabolic repertoire of Microcystis, for example by encoding the biosynthesis of complementary carotenoid molecules. In contrast, other colony-associated bacteria showed weaker signals of co-phylogeny, but stronger evidence of horizontal gene transfer with Microcystis. These observations suggest that acquired genes are more likely to be retained in both partners (Microcystis and members of its microbiome) when they are loosely associated, whereas one gene copy is sufficient when the association is physically tight and evolutionarily long-lasting. CONCLUSIONS: We have introduced a method for culture-free isolation of single colonies from nature followed by metagenomic sequencing, which could be applied to other types of microbes. Together, our results expand the known genetic diversity of both Microcystis and its microbiome in natural settings, and support their long-term, specific, and potentially beneficial associations. Video Abstract.

Coherence of Microcystis species revealed through population genomics.

Microcystis is a genus of freshwater cyanobacteria, which causes harmful blooms in ecosystems worldwide. Some Microcystis strains produce harmful toxins such as microcystin, impacting drinking water quality. Microcystis colony morphology, rather than genetic similarity, is often used to classify Microcystis into morphospecies. Yet colony morphology is a plastic trait, which can change depending on environmental and laboratory culture conditions, and is thus an inadequate criterion for species delineation. Furthermore, Microcystis populations are thought to disperse globally and constitute a homogeneous gene pool. However, this assertion is based on relatively incomplete characterization of Microcystis genomic diversity. To better understand these issues, we performed a population genomic analysis of 33 newly sequenced genomes mainly from Canada and Brazil. We identified 17 Microcystis clusters of genomic similarity, five of which correspond to monophyletic clades containing at least three newly sequenced genomes. Four out of these five clades match to named morphospecies. Notably, M. aeruginosa is paraphyletic, distributed across 12 genomic clusters, suggesting it is not a coherent species. A few clades of closely related isolates are specific to a unique geographic location, suggesting biogeographic structure over relatively short evolutionary time scales. Higher homologous recombination rates within than between clades further suggest that monophyletic groups might adhere to a Biological Species-like concept, in which barriers to gene flow maintain species distinctness. However, certain genes-including some involved in microcystin and micropeptin biosynthesis-are recombined between monophyletic groups in the same geographic location, suggesting local adaptation.

Population genomics of the symbiotic plasmids of sympatric nitrogen-fixing Rhizobium species associated with Phaseolus vulgaris.

Cultivated common beans are the primary protein source for millions of people around the world who subsist on low-input agriculture, enabled by the symbiotic N2 -fixation these legumes perform in association with rhizobia. Within a single agricultural plot, multiple Rhizobium species can nodulate bean roots, but it is unclear how genetically isolated these species remain in sympatry. To better understand this issue, we sequenced and compared the genomes of 33 strains isolated from the rhizosphere and root nodules of a particular bean variety grown in the same agricultural plot. We found that the Rhizobium species we observed coexist with low genetic recombination across their core genomes. Accessory plasmids thought to be necessary for the saprophytic lifestyle in soil show similar levels of genetic isolation, but with higher rates of recombination than the chromosomes. However, the symbiotic plasmids are extremely similar, with high rates of recombination and do not appear to have co-evolved with the chromosome or accessory plasmids. Therefore, while Rhizobium species are genetically isolated units within the microbial community, a common symbiotic plasmid allows all Rhizobium species to engage in symbiosis with the same host in a single agricultural plot.