Host specificity of plant-associated bacteria is negatively associated with genome size and host abundance along a latitudinal gradient.

Host specialization plays a critical role in the ecology and evolution of plant-microbe symbiosis. Theory predicts that host specialization is associated with microbial genome streamlining and is influenced by the abundance of host species, both of which can vary across latitudes, leading to a latitudinal gradient in host specificity. Here, we quantified the host specificity and composition of plant-bacteria symbioses on leaves across 329 tree species spanning a latitudinal gradient. Our analysis revealed a predominance of host-specialized leaf bacteria. The degree of host specificity was negatively correlated with bacterial genome size and the local abundance of host plants. Additionally, we found an increased host specificity at lower latitudes, aligning with the high prevalence of small bacterial genomes and rare host species in the tropics. These findings underscore the importance of genome streamlining and host abundance in the evolution of host specificity in plant-associated bacteria along the latitudinal gradient.

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.

Metabolism and physiology of pathogenic bacterial obligate intracellular parasites.

Bacterial obligate intracellular parasites (BOIPs) represent an exclusive group of bacterial pathogens that all depend on invasion of a eukaryotic host cell to reproduce. BOIPs are characterized by extensive adaptation to their respective replication niches, regardless of whether they replicate within the host cell cytoplasm or within specialized replication vacuoles. Genome reduction is also a hallmark of BOIPs that likely reflects streamlining of metabolic processes to reduce the need for de novo biosynthesis of energetically costly metabolic intermediates. Despite shared characteristics in lifestyle, BOIPs show considerable diversity in nutrient requirements, metabolic capabilities, and general physiology. In this review, we compare metabolic and physiological processes of prominent pathogenic BOIPs with special emphasis on carbon, energy, and amino acid metabolism. Recent advances are discussed in the context of historical views and opportunities for discovery.

PoMeLo: a systematic computational approach to predicting metabolic loss in pathogen genomes.

BACKGROUND: Genome streamlining, the process by which genomes become smaller and encode fewer genes over time, is a common phenomenon among pathogenic bacteria. This reduction is driven by selection for minimized energy expenditure in a nutrient-rich environment. As pathogens evolve to become more reliant on the host, metabolic genes and resulting capabilities are lost in favor of siphoning metabolites from the host. Characterizing genome streamlining, gene loss, and metabolic pathway degradation can be useful in assessing pathogen dependency on host metabolism and identifying potential targets for host-directed therapeutics. RESULTS: PoMeLo (Predictor of Metabolic Loss) is a novel evolutionary genomics-guided computational approach for identifying metabolic gaps in the genomes of pathogenic bacteria. PoMeLo leverages a centralized public database of high-quality genomes and annotations and allows the user to compare an unlimited number of genomes across individual genes and pathways. PoMeLo runs locally using user-friendly prompts in a matter of minutes and generates tabular and visual outputs for users to compare predicted metabolic capacity between groups of bacteria and individual species. Each pathway is assigned a Predicted Metabolic Loss (PML) score to assess the magnitude of genome streamlining. Optionally, PoMeLo places the results in an evolutionary context by including phylogenetic relationships in visual outputs. It can also initially compute phylogenetically-weighted mean genome sizes to identify genome streamlining events. Here, we describe PoMeLo and demonstrate its use in identifying metabolic gaps in genomes of pathogenic Treponema species. CONCLUSIONS: PoMeLo represents an advance over existing methods for identifying metabolic gaps in genomic data, allowing comparison across large numbers of genomes and placing the resulting data in a phylogenetic context. PoMeLo is freely available for academic and non-academic use at https://github.com/czbiohub-sf/pomelo .

Genomic analysis of the marine yeast Rhodotorula sphaerocarpa ETNP2018 reveals adaptation to the open ocean.

BACKGROUND: Despite a rising interest in the diversity and ecology of fungi in marine environments, there are few published genomes of fungi isolated from the ocean. The basidiomycetous yeast (unicellular fungus) genus Rhodotorula are prevalent and abundant in the open ocean, and they have been isolated from a wide range of other environments. Many of these environments are nutrient poor, such as the Antarctica and the Atacama deserts, raising the question as to how Rhodotorula yeasts may have adapted their metabolic strategies to optimize survival under low nutrient conditions. In order to understand their adaptive strategies in the ocean, the genome of R. sphaerocarpa ETNP2018 was compared to that of fourteen representative Rhodotorula yeasts, isolated from a variety of environments. RESULTS: Rhodotorula sphaerocarpa ETNP2018, a strain isolated from the oligotrophic part of the eastern tropical North Pacific (ETNP) oxygen minimum zone (OMZ), hosts the smallest of the fifteen genomes and yet the number of protein-coding genes it possesses is on par with the other strains. Its genome exhibits a distinct reduction in genes dedicated to Major Facilitator Superfamily transporters as well as biosynthetic enzymes. However, its core metabolic pathways are fully conserved. Our research indicates that the selective pressures of the ETNP OMZ favor a streamlined genome with reduced overall biosynthetic potential balanced by a stable set of core metabolisms and an expansion of mechanisms for nutrient acquisition. CONCLUSIONS: In summary, this study offers insights into the adaptation of fungi to the oligotrophic ocean and provides valuable information for understanding the ecological roles of fungi in the ocean.

Metagenome-assembled genomes reveal greatly expanded taxonomic and functional diversification of the abundant marine Roseobacter RCA cluster.

BACKGROUND: The RCA (Roseobacter clade affiliated) cluster belongs to the family Roseobacteracea and represents a major Roseobacter lineage in temperate to polar oceans. Despite its prevalence and abundance, only a few genomes and one described species, Planktomarina temperata, exist. To gain more insights into our limited understanding of this cluster and its taxonomic and functional diversity and biogeography, we screened metagenomic datasets from the global oceans and reconstructed metagenome-assembled genomes (MAG) affiliated to this cluster. RESULTS: The total of 82 MAGs, plus five genomes of isolates, reveal an unexpected diversity and novel insights into the genomic features, the functional diversity, and greatly refined biogeographic patterns of the RCA cluster. This cluster is subdivided into three genera: Planktomarina, Pseudoplanktomarina, and the most deeply branching Candidatus Paraplanktomarina. Six of the eight Planktomarina species have larger genome sizes (2.44-3.12 Mbp) and higher G + C contents (46.36-53.70%) than the four Pseudoplanktomarina species (2.26-2.72 Mbp, 42.22-43.72 G + C%). Cand. Paraplanktomarina is represented only by one species with a genome size of 2.40 Mbp and a G + C content of 45.85%. Three novel species of the genera Planktomarina and Pseudoplanktomarina are validly described according to the SeqCode nomenclature for prokaryotic genomes. Aerobic anoxygenic photosynthesis (AAP) is encoded in three Planktomarina species. Unexpectedly, proteorhodopsin (PR) is encoded in the other Planktomarina and all Pseudoplanktomarina species, suggesting that this light-driven proton pump is the most important mode of acquiring complementary energy of the RCA cluster. The Pseudoplanktomarina species exhibit differences in functional traits compared to Planktomarina species and adaptations to more resource-limited conditions. An assessment of the global biogeography of the different species greatly expands the range of occurrence and shows that the different species exhibit distinct biogeographic patterns. They partially reflect the genomic features of the species. CONCLUSIONS: Our detailed MAG-based analyses shed new light on the diversification, environmental adaptation, and global biogeography of a major lineage of pelagic bacteria. The taxonomic delineation and validation by the SeqCode nomenclature of prominent genera and species of the RCA cluster may be a promising way for a refined taxonomic identification of major prokaryotic lineages and sublineages in marine and other prokaryotic communities assessed by metagenomics approaches. Video Abstract.

A Large-Scale Genome-Based Survey of Acidophilic Bacteria Suggests That Genome Streamlining Is an Adaption for Life at Low pH.

The genome streamlining theory suggests that reduction of microbial genome size optimizes energy utilization in stressful environments. Although this hypothesis has been explored in several cases of low-nutrient (oligotrophic) and high-temperature environments, little work has been carried out on microorganisms from low-pH environments, and what has been reported is inconclusive. In this study, we performed a large-scale comparative genomics investigation of more than 260 bacterial high-quality genome sequences of acidophiles, together with genomes of their closest phylogenetic relatives that live at circum-neutral pH. A statistically supported correlation is reported between reduction of genome size and decreasing pH that we demonstrate is due to gene loss and reduced gene sizes. This trend is independent from other genome size constraints such as temperature and G + C content. Genome streamlining in the evolution of acidophilic bacteria is thus supported by our results. The analyses of predicted Clusters of Orthologous Genes (COG) categories and subcellular location predictions indicate that acidophiles have a lower representation of genes encoding extracellular proteins, signal transduction mechanisms, and proteins with unknown function but are enriched in inner membrane proteins, chaperones, basic metabolism, and core cellular functions. Contrary to other reports for genome streamlining, there was no significant change in paralog frequencies across pH. However, a detailed analysis of COG categories revealed a higher proportion of genes in acidophiles in the following categories: "replication and repair," "amino acid transport," and "intracellular trafficking". This study brings increasing clarity regarding the genomic adaptations of acidophiles to life at low pH while putting elements, such as the reduction of average gene size, under the spotlight of streamlining theory.

Enhanced production of polyhydroxyalkanoates in Pseudomonas putida KT2440 by a combination of genome streamlining and promoter engineering.

Polyhydroxyalkanoates (PHAs), a class of bioplastics produced by a variety of microorganisms, have become the ideal alternatives for oil-derived plastics due to their superior physicochemical and material characteristics. Pseudomonas putida KT2440 can produce medium-chain-length PHA (mcl-PHA) from various substrates. In this study, a novel strategy of the large-scale deletion of genomic islands (GIs) coupling with promoter engineering was developed in P. putida KT2440 for constructing the minimal genome cell factories (MGF) capable of efficiently producing mcl-PHA. Firstly, P. putida KTU-U13, a 13 GIs- and upp-deleted mutant derived from the parental strain P. putida KT2440, was used as a starting strain for further deletion of GIs to generate a series of genome-reduced strains. Subsequently, the two minimal genome strains KTU-U24 and KTU-U27, which had a 7.19% and 8.35% reduction relative to the genome size of KT2440 and were advantageous over the strain KTU (KT2440∆upp) and KTU-U13 in several physiological traits such as the maximum specific growth rate, plasmid transformation efficiency, heterologous protein expression capacity and PHA production capacity, were selected as the chassis cells for PHA metabolic engineering. To prevent the formation of the by-product gluconic acid, the glucose dehydrogenase gene was deleted in KTU-U24 and KTU-U27, resulting in KTU-U24∆gcd and KTU-U27∆gcd. To enhance the transcriptional level of PHA synthase genes (phaC) and the supply of the precursor acetyl-CoA, a strong endogenous promoter P46 was inserted into upstream of the phaC operon and pyruvate dehydrogenase gene in the genome of KTU-U24∆gcd and KTU-U27∆gcd, to generate KTU-U24∆gcd-P46CA and KTU-U27∆gcd-P46CA, with the PHA yield of 50.5 wt% and 53.8 wt% (weight percent of PHA in cell dry weight). Finally, KTU-U27∆gcd-P46CA, the most minimal KT2440 chassis currently available, was able to accumulate the PHA to 55.82 wt% in a 5-l fermentor, which is the highest PHA yield obtained with P. putida KT2440 so far. This study suggests that genome streamlining in combination with promoter engineering may be a feasible strategy for the development of the MGF for the efficient production of high value products. Moreover, further streamlining of the P. putida KT2440 genome has great potential to create the optimal chassis for synthetic biology applications.

An Extremely Streamlined Macronuclear Genome in the Free-Living Protozoan Fabrea salina.

Ciliated protists are among the oldest unicellular organisms with a heterotrophic lifestyle and share a common ancestor with Plantae. Unlike any other eukaryotes, there are two distinct nuclei in ciliates with separate germline and somatic cell functions. Here, we assembled a near-complete macronuclear genome of Fabrea salina, which belongs to one of the oldest clades of ciliates. Its extremely minimized genome (18.35 Mb) is the smallest among all free-living heterotrophic eukaryotes and exhibits typical streamlined genomic features, including high gene density, tiny introns, and shrinkage of gene paralogs. Gene families involved in hypersaline stress resistance, DNA replication proteins, and mitochondrial biogenesis are expanded, and the accumulation of phosphatidic acid may play an important role in resistance to high osmotic pressure. We further investigated the morphological and transcriptomic changes in the macronucleus during sexual reproduction and highlighted the potential contribution of macronuclear residuals to this process. We believe that the minimized genome generated in this study provides novel insights into the genome streamlining theory and will be an ideal model to study the evolution of eukaryotic heterotrophs.

A myxozoan genome reveals mosaic evolution in a parasitic cnidarian.

BACKGROUND: Parasite evolution has been conceptualized as a process of genetic loss and simplification. Contrary to this model, there is evidence of expansion and conservation of gene families related to essential functions of parasitism in some parasite genomes, reminiscent of widespread mosaic evolution-where subregions of a genome have different rates of evolutionary change. We found evidence of mosaic genome evolution in the cnidarian Myxobolus honghuensis, a myxozoan parasite of fish, with extremely simple morphology. RESULTS: We compared M. honghuensis with other myxozoans and free-living cnidarians, and determined that it has a relatively larger myxozoan genome (206 Mb), which is less reduced and less compact due to gene retention, large introns, transposon insertion, but not polyploidy. Relative to other metazoans, the M. honghuensis genome is depleted of neural genes and has only the simplest animal immune components. Conversely, it has relatively more genes involved in stress resistance, tissue invasion, energy metabolism, and cellular processes compared to other myxozoans and free-living cnidarians. We postulate that the expansion of these gene families is the result of evolutionary adaptations to endoparasitism. M. honghuensis retains genes found in free-living Cnidaria, including a reduced nervous system, myogenic components, ANTP class Homeobox genes, and components of the Wnt and Hedgehog pathways. CONCLUSIONS: Our analyses suggest that the M. honghuensis genome evolved as a mosaic of conservative, divergent, depleted, and enhanced genes and pathways. These findings illustrate that myxozoans are not as genetically simple as previously regarded, and the evolution of some myxozoans is driven by both genomic streamlining and expansion.

Genomic adaptations enabling Acidithiobacillus distribution across wide-ranging hot spring temperatures and pHs.

BACKGROUND: Terrestrial hot spring settings span a broad spectrum of physicochemistries. Physicochemical parameters, such as pH and temperature, are key factors influencing differences in microbial composition across diverse geothermal areas. Nonetheless, analysis of hot spring pools from the Taupo Volcanic Zone (TVZ), New Zealand, revealed that some members of the bacterial genus, Acidithiobacillus, are prevalent across wide ranges of hot spring pHs and temperatures. To determine the genomic attributes of Acidithiobacillus that inhabit such diverse conditions, we assembled the genomes of 19 uncultivated hot spring Acidithiobacillus strains from six geothermal areas and compared these to 37 publicly available Acidithiobacillus genomes from various habitats. RESULTS: Analysis of 16S rRNA gene amplicons from 138 samples revealed that Acidithiobacillus comprised on average 11.4 ± 16.8% of hot spring prokaryotic communities, with three Acidithiobacillus amplicon sequence variants (ASVs) (TVZ_G1, TVZ_G2, TVZ_G3) accounting for > 90% of Acidithiobacillus in terms of relative abundance, and occurring in 126 out of 138 samples across wide ranges of temperature (17.5-92.9 °C) and pH (1.0-7.5). We recovered 19 environmental genomes belonging to each of these three ASVs, as well as a fourth related group (TVZ_G4). Based on genome average nucleotide identities, the four groups (TVZ_G1-TVZ_G4) constitute distinct species (ANI < 96.5%) of which three are novel Acidithiobacillus species (TVZ_G2-TVZ_G4) and one belongs to Acidithiobacillus caldus (TVZ_G1). All four TVZ Acidithiobacillus groups were found in hot springs with temperatures above the previously known limit for the genus (up to 40 °C higher), likely due to significantly higher proline and GC contents than other Acidithiobacillus species, which are known to increase thermostability. Results also indicate hot spring-associated Acidithiobacillus have undergone genome streamlining, likely due to thermal adaptation. Moreover, our data suggest that Acidithiobacillus prevalence across varied hot spring pHs is supported by distinct strategies, whereby TVZ_G2-TVZ_G4 regulate pH homeostasis mostly through Na+/H+ antiporters and proton-efflux ATPases, whereas TVZ_G1 mainly relies on amino acid decarboxylases. CONCLUSIONS: This study provides insights into the distribution of Acidithiobacillus species across diverse hot spring physichochemistries and determines genomic features and adaptations that potentially enable Acidithiobacillus species to colonize a broad range of temperatures and pHs in geothermal environments. Video Abstract.

Genome Size Affects Fitness in the Eukaryotic Alga Dunaliella tertiolecta.

Genome size is tightly coupled to morphology, ecology, and evolution among species [1-5], with one of the best-known patterns being the relationship between cell size and genome size [6, 7]. Classic theories, such as the "selfish DNA hypothesis," posit that accumulating redundant DNA has fitness costs but that larger cells can tolerate larger genomes, leading to a positive relationship between cell size and genome size [8, 9]. Yet the evidence for fitness costs associated with relatively larger genomes remains circumstantial. Here, we estimated the relationships between genome size, cell size, energy fluxes, and fitness across 72 independent lineages in a eukaryotic phytoplankton. Lineages with relatively smaller genomes had higher fitness, in terms of both maximum growth rate and total biovolume reached at carrying capacity, but paradoxically, they also had lower energy fluxes than lineages with relative larger genomes. We then explored the evolutionary trajectories of absolute genome size over 100 generations and across a 10-fold change in cell size. Despite consistent directional selection across all lineages, genome size decreased by 11% in lineages with absolutely larger genomes but showed little evolution in lineages with absolutely smaller genomes, implying a lower absolute limit in genome size. Our results suggest that the positive relationship between cell size and genome size in nature may be the product of conflicting evolutionary pressures, on the one hand, to minimize redundant DNA and maximize performance-as theory predicts-but also to maintain a minimum level of essential function. VIDEO ABSTRACT.

Influence of Substrate Concentration on the Culturability of Heterotrophic Soil Microbes Isolated by High-Throughput Dilution-to-Extinction Cultivation.

The vast majority of microbes inhabiting oligotrophic shallow subsurface soil environments have not been isolated or studied under controlled laboratory conditions. In part, the challenges associated with isolating shallow subsurface microbes may persist because microbes in deeper soils are adapted to low nutrient availability or quality. Here, we use high-throughput dilution-to-extinction culturing to isolate shallow subsurface microbes from a conifer forest in Arizona, USA. We hypothesized that the concentration of heterotrophic substrates in microbiological growth medium would affect which microbial taxa were culturable from these soils. To test this, we diluted cells extracted from soil into one of two custom-designed defined growth media that differed by 100-fold in the concentration of amino acids and organic carbon. Across the two media, we isolated a total of 133 pure cultures, all of which were classified as Actinobacteria or Alphaproteobacteria The substrate availability dictated which actinobacterial phylotypes were culturable but had no significant effect on the culturability of Alphaproteobacteria We isolated cultures that were representative of the most abundant phylotype in the soil microbial community (Bradyrhizobium spp.) and representatives of five of the top 10 most abundant Actinobacteria phylotypes, including Nocardioides spp., Mycobacterium spp., and several other phylogenetically divergent lineages. Flow cytometry of nucleic acid-stained cells showed that cultures isolated on low-substrate medium had significantly lower nucleic acid fluorescence than those isolated on high-substrate medium. These results show that dilution-to-extinction is an effective method to isolate abundant soil microbes and that the concentration of substrates in culture medium influences the culturability of specific microbial lineages.IMPORTANCE Isolating environmental microbes and studying their physiology under controlled conditions are essential aspects of understanding their ecology. Subsurface ecosystems are typically nutrient-poor environments that harbor diverse microbial communities-the majority of which are thus far uncultured. In this study, we use modified high-throughput cultivation methods to isolate subsurface soil microbes. We show that a component of whether a microbe is culturable from subsurface soils is the concentration of growth substrates in the culture medium. Our results offer new insight into technical approaches and growth medium design that can be used to access the uncultured diversity of soil microbes.

Strong Purifying Selection Is Associated with Genome Streamlining in Epipelagic Marinimicrobia.

Marine microorganisms inhabiting nutrient-depleted waters play critical roles in global biogeochemical cycles due to their abundance and broad distribution. Many of these microbes share similar genomic features including small genome size, low % G + C content, short intergenic regions, and low nitrogen content in encoded amino acid residue side chains (N-ARSC), but the evolutionary drivers of these characteristics are unclear. Here, we compared the strength of purifying selection across the Marinimicrobia, a candidate phylum which encompasses a broad range of phylogenetic groups with disparate genomic features, by estimating the ratio of nonsynonymous and synonymous substitutions (dN/dS) in conserved marker genes. Our analysis reveals that epipelagic Marinimicrobia that exhibit features consistent with genome streamlining have significantly lower dN/dS values when compared with their mesopelagic counterparts. We also found a significant positive correlation between median dN/dS values and % G + C content, N-ARSC, and intergenic region length. We did not identify a significant correlation between dN/dS ratios and estimated genome size, suggesting the strength of selection is not a primary factor shaping genome size in this group. Our findings are generally consistent with genome streamlining theory, which postulates that many genomic features of abundant epipelagic bacteria are the result of adaptation to oligotrophic nutrient conditions. Our results are also in agreement with previous findings that genome streamlining is common in epipelagic waters, suggesting that microbes inhabiting this region of the ocean have been shaped by strong selection together with prevalent nutritional constraints characteristic of this environment.

Occurrence and Fate of Ultramicrobacteria in a Full-Scale Drinking Water Treatment Plant.

Ultramicrobacteria (UMB) are omnipresent and numerically dominate in freshwater, as microbes can present in drinking water systems, however, the UMB communities that occur and their removal behaviors remain poorly characterized in drinking water treatment plants (DWTPs). To gain insights into these issues, we profiled bacterial cell density, community structure and functions of UMB and their counterpart large bacteria (LB) using flow cytometry and filtration paired with 16S rRNA gene high-throughput sequencing in a full-scale DWTP. Contrary to the reduction of bacterial density and diversity, the proportion of UMB in the total bacteria community increased as the drinking water treatment process progressed, and biological activated carbon facilitated bacterial growth. Moreover, UMB were less diverse than LB, and their community structure and predicted functions were significantly different. In the DWTP, UMB indicator taxa were mainly affiliated with α/ß/γ-Proteobacteria, Deinococcus-Thermus, Firmicutes, Acidobacteria, and Dependentiae. In particular, the exclusive clustering of UMB at the phylum level, e.g., Parcubacteria, Elusimicrobia, and Saccharibacteria, confirmed the fact that the ultra-small size of UMB is a naturally and evolutionarily conserved trait. Additionally, the streamlined genome could be connected to UMB, such as candidate phyla radiation (CPR) bacteria, following a symbiotic or parasitic lifestyle, which then leads to the observed high connectedness, i.e., non-random intra-taxa co-occurrence patterns within UMB. Functional prediction analysis revealed that environmental information processing and DNA replication and repair likely contribute to the higher resistance of UMB to drinking water treatment processes in comparison to LB. Overall, the study provides valuable insights into the occurrence and fate of UMB regarding community structure, phylogenetic characteristics and potential functions in a full-scale DWTP, and it is a useful reference for beneficial manipulation of the drinking water microbiome.

Parallel Evolution of Genome Streamlining and Cellular Bioenergetics across the Marine Radiation of a Bacterial Phylum.

Diverse bacterial and archaeal lineages drive biogeochemical cycles in the global ocean, but the evolutionary processes that have shaped their genomic properties and physiological capabilities remain obscure. Here we track the genome evolution of the globally abundant marine bacterial phylum Marinimicrobia across its diversification into modern marine environments and demonstrate that extant lineages are partitioned between epipelagic and mesopelagic habitats. Moreover, we show that these habitat preferences are associated with fundamental differences in genomic organization, cellular bioenergetics, and metabolic modalities. Multiple lineages present in epipelagic niches independently acquired genes necessary for phototrophy and environmental stress mitigation, and their genomes convergently evolved key features associated with genome streamlining. In contrast, lineages residing in mesopelagic waters independently acquired nitrate respiratory machinery and a variety of cytochromes, consistent with the use of alternative terminal electron acceptors in oxygen minimum zones (OMZs). Further, while epipelagic clades have retained an ancestral Na+-pumping respiratory complex, mesopelagic lineages have largely replaced this complex with canonical H+-pumping respiratory complex I, potentially due to the increased efficiency of the latter together with the presence of the more energy-limiting environments deep in the ocean's interior. These parallel evolutionary trends indicate that key features of genomic streamlining and cellular bioenergetics have occurred repeatedly and congruently in disparate clades and underscore the importance of environmental conditions and nutrient dynamics in driving the evolution of diverse bacterioplankton lineages in similar ways throughout the global ocean.IMPORTANCE Understanding long-term patterns of microbial evolution is critical to advancing our knowledge of past and present role microbial life in driving global biogeochemical cycles. Historically, it has been challenging to study the evolution of environmental microbes due to difficulties in obtaining genome sequences from lineages that could not be cultivated, but recent advances in metagenomics and single-cell genomics have begun to obviate many of these hurdles. Here we present an evolutionary genomic analysis of the Marinimicrobia, a diverse bacterial group that is abundant in the global ocean. We demonstrate that distantly related Marinimicrobia species that reside in similar habitats have converged to assume similar genome architectures and cellular bioenergetics, suggesting that common factors shape the evolution of a broad array of marine lineages. These findings broaden our understanding of the evolutionary forces that have given rise to microbial life in the contemporary ocean.

Reconstruction of Diverse Verrucomicrobial Genomes from Metagenome Datasets of Freshwater Reservoirs.

The phylum Verrucomicrobia contains freshwater representatives which remain poorly studied at the genomic, taxonomic, and ecological levels. In this work we present eighteen new reconstructed verrucomicrobial genomes from two freshwater reservoirs located close to each other (Tous and Amadorio, Spain). These metagenome-assembled genomes (MAGs) display a remarkable taxonomic diversity inside the phylum and comprise wide ranges of estimated genome sizes (from 1.8 to 6 Mb). Among all Verrucomicrobia studied we found some of the smallest genomes of the Spartobacteria and Opitutae classes described so far. Some of the Opitutae family MAGs were small, cosmopolitan, with a general heterotrophic metabolism with preference for carbohydrates, and capable of xylan, chitin, or cellulose degradation. Besides, we assembled large copiotroph genomes, which contain a higher number of transporters, polysaccharide degrading pathways and in general more strategies for the uptake of nutrients and carbohydrate-based metabolic pathways in comparison with the representatives with the smaller genomes. The diverse genomes revealed interesting features like green-light absorbing rhodopsins and a complete set of genes involved in nitrogen fixation. The large diversity in genome sizes and physiological properties emphasize the diversity of this clade in freshwaters enlarging even further the already broad eco-physiological range of these microbes.

Theory of prokaryotic genome evolution.

Bacteria and archaea typically possess small genomes that are tightly packed with protein-coding genes. The compactness of prokaryotic genomes is commonly perceived as evidence of adaptive genome streamlining caused by strong purifying selection in large microbial populations. In such populations, even the small cost incurred by nonfunctional DNA because of extra energy and time expenditure is thought to be sufficient for this extra genetic material to be eliminated by selection. However, contrary to the predictions of this model, there exists a consistent, positive correlation between the strength of selection at the protein sequence level, measured as the ratio of nonsynonymous to synonymous substitution rates, and microbial genome size. Here, by fitting the genome size distributions in multiple groups of prokaryotes to predictions of mathematical models of population evolution, we show that only models in which acquisition of additional genes is, on average, slightly beneficial yield a good fit to genomic data. These results suggest that the number of genes in prokaryotic genomes reflects the equilibrium between the benefit of additional genes that diminishes as the genome grows and deletion bias (i.e., the rate of deletion of genetic material being slightly greater than the rate of acquisition). Thus, new genes acquired by microbial genomes, on average, appear to be adaptive. The tight spacing of protein-coding genes likely results from a combination of the deletion bias and purifying selection that efficiently eliminates nonfunctional, noncoding sequences.

Evolutionary Dynamics of Chloroplast Genomes in Low Light: A Case Study of the Endolithic Green Alga Ostreobium quekettii.

Some photosynthetic organisms live in extremely low light environments. Light limitation is associated with selective forces as well as reduced exposure to mutagens, and over evolutionary timescales it can leave a footprint on species' genomes. Here, we present the chloroplast genomes of four green algae (Bryopsidales, Ulvophyceae), including the endolithic (limestone-boring) alga Ostreobium quekettii, which is a low light specialist. We use phylogenetic models and comparative genomic tools to investigate whether the chloroplast genome of Ostreobium corresponds to our expectations of how low light would affect genome evolution. Ostreobium has the smallest and most gene-dense chloroplast genome among Ulvophyceae reported to date, matching our expectation that light limitation would impose resource constraints reflected in the chloroplast genome architecture. Rates of molecular evolution are significantly slower along the phylogenetic branch leading to Ostreobium, in agreement with the expected effects of low light and energy levels on molecular evolution. We expected the ability of Ostreobium to perform photosynthesis in very low light to be associated with positive selection in genes related to the photosynthetic machinery, but instead, we observed that these genes may be under stronger purifying selection. Besides shedding light on the genome dynamics associated with a low light lifestyle, this study helps to resolve the role of environmental factors in shaping the diversity of genome architectures observed in nature.

Indispensability of Horizontally Transferred Genes and Its Impact on Bacterial Genome Streamlining.

Why are certain bacterial genomes so small and compact? The adaptive genome streamlining hypothesis posits that selection acts to reduce genome size because of the metabolic burden of replicating DNA. To reveal the impact of genome streamlining on cellular traits, we reduced the Escherichia coli genome by up to 20% by deleting regions which have been repeatedly subjects of horizontal transfer in nature. Unexpectedly, horizontally transferred genes not only confer utilization of specific nutrients and elevate tolerance to stresses, but also allow efficient usage of resources to build new cells, and hence influence fitness in routine and stressful environments alike. Genome reduction affected fitness not only by gene loss, but also by induction of a general stress response. Finally, we failed to find evidence that the advantage of smaller genomes would be due to a reduced metabolic burden of replicating DNA or a link with smaller cell size. We conclude that as the potential energetic benefit gained by deletion of short genomic segments is vanishingly small compared with the deleterious side effects of these deletions, selection for reduced DNA synthesis costs is unlikely to shape the evolution of small genomes.