Cyanobacterial distributions are shaped by abiotic factors including temperature, light and nutrient availability as well as biotic factors such as grazing and viral infection. In this study, we investigated the abundances of T4-like and T7-like cyanophages and the extent of picocyanobacterial infection in the cold, high-nutrient-low-chlorophyll, sub-Antarctic waters of the southwest Pacific Ocean during austral spring. Synechococcus was the dominant picocyanobacterium, ranging from 4.7 × 10 to 1.2 × 10 cells∙mL, while Prochlorococcus abundances were relatively low overall, ranging from 1.0 × 10 to 3.9 × 10 cells∙mL. Using taxon-specific, single-virus and single-cell polony methods, we found that cyanophages were on average 15-fold, and up to 50-fold, more abundant than cyanobacteria in these waters. T4-like cyanophages (ranging from 1.7 × 10 to 6.5 × 10 phage·mL) were 2.7-fold more abundant than T7-like cyanophages (ranging from 3.1 × 10 to 2.8 × 10 phage·mL). Picocyanobacteria were primarily infected by T4-like cyanophages with more Synechococcus (4.8%-12.1%) infected than Prochlorococcus (2.5%-6.2%), whereas T7-like cyanophages infected less than 1% of both genera. These infection levels translated to daily mortality in the range of 5.7%-26.2% and 2.9%-14.3% of the standing stock of Synechococcus and Prochlorococcus, respectively. Our findings suggest that T4-like cyanophages are significant agents of cyanobacterial mortality in the cold, low-iron, sub-Antarctic waters of the South Pacific Ocean.

The leaf surface, known as the phylloplane, presents an oligotrophic and heterogeneous environment due to its topography and uneven distribution of resources. Although it is a challenging environment, leaves support abundant bacterial communities that are spatially structured. However, the factors influencing these spatial distribution patterns are not well understood. To study the changes in population density and spatial distribution of bacteria in synthetic communities, the behaviour of two common bacterial groups in the Arabidopsis thaliana leaf microbiota-Methylobacterium (methylobacteria) and Sphingomonas (sphingomonads)-was examined. Using synthetic communities consisting of two or three species, the hypothesis was tested that the presence of a third species affects the density and spatial interaction of the other two species. Results indicated that methylobacteria exhibit greater sensitivity to changes in population densities and spatial patterns, with higher intra-genus competition and lower densities and aggregation compared to sphingomonads. Pairwise comparisons were insufficient to explain the shifts observed in three-species communities, suggesting that higher-order interactions influence the structuring of complex communities. This emphasises the role of multispecies interactions in determining spatial patterns and community dynamics on the phylloplane.

The Canadian province of Alberta contains substantial oilsands reservoirs, consisting of bitumen, clay and sand. Extracting oil involves separating bitumen from inorganic particles using hot water and chemical diluents, resulting in liquid tailings waste with ecotoxicologically significant compounds. Ongoing efforts aim to reclaim tailings-affected areas, with protist colonisation serving as one assessment method of reclamation progress. Oilsands-associated protist communities have mainly been evaluated using amplicon sequencing of the 18S rRNA V4 region; however, this barcode may overlook important protist groups. This study examined how community assessment methods between the V4 and V9 regions differ in representing protist diversity across four oilsands-associated environments. The V9 barcode identified more operational taxonomical units (OTUs) for Discoba, Metamonada and Amoebozoa compared with the V4. A comparative shotgun metagenomics approach revealed few eukaryotic contigs but did recover a complete Paramicrosporidia mitochondrial genome, only the second publicly available from microsporidians. Both V4 and V9 markers were informative for assessing community diversity in oilsands-associated environments and are most effective when combined for a comprehensive taxonomic estimate, particularly in anoxic environments.

Shotgun and proximity-ligation metagenomic sequencing were used to generate thousands of metagenome assembled genomes (MAGs) from the untreated wastewater, activated sludge bioreactors, and anaerobic digesters from two full-scale municipal wastewater treatment facilities. Analysis of the antibiotic resistance genes (ARGs) in the pool of contigs from the shotgun metagenomic sequences revealed significantly different relative abundances and types of ARGs in the untreated wastewaster compared to the activated sludge bioreactors or the anaerobic digesters (p < 0.05). In contrast, these results were statistically similar when comparing the ARGs in the pool of MAGs, suggesting that proximity-ligation metagenomic sequencing is particularly useful for pairing ARGs with their hosts but less adept at discerning quantitative differences in ARG types and relative abundances. For example, numerous MAGs of the genera Acinetobacter, Enterococcus, Klebsiella and Pseudomonas were identified in the untreated wastewater, many of which harboured plasmid-borne and/or chromosomal-borne ARGs; none of these MAGs, however, were detected in the activated sludge bioreactors or anaerobic digesters. In conclusion, this research demonstrates that the antibiotic resistome undergoes significant transitions in both the relative abundance and the host organisms during the municipal wastewater treatment process.

Actinobacteria have frequently been reported in the Andean Puna, including strains of the genus Micrococcus. These strains demonstrate resistance to high levels of UV radiation, arsenic, and multiple antibiotics, and possess large linear plasmids. A comparative analysis of the sequences and putative functions of these plasmids was conducted. The presence of large regions with high sequence identity (exceeding 30 kb in total) in all three studied Micrococcus megaplasmids indicates a clear evolutionary link among these elements. Genes related to essential plasmid functions were primarily found within these conserved regions, while genes associated with resistance to metals and antibiotics resided in accessory regions. Moreover, the abundance of open reading frames related to transposition and recombination, along with local deviations from the average GC content, provides evidence for the mosaic nature and considerable genetic plasticity of these plasmids. This study presents evidence of a common ancestor for linear plasmids in Micrococcus and suggests that horizontal gene transfer likely occurs frequently within Andean lakes, providing the native microbial community with a beneficial gene pool to withstand extreme conditions. Additionally, the successful transfer of the linear plasmid pLMA1 by a DNase-insensitive, conjugation-type mechanism and its potential use as a genetic vector is demonstrated.

Anaerobic digestion (AD) of organic wastes relies on the interaction and cooperation of various microorganisms. Phages are crucial components of the microbial community in AD systems, but their diversity and interactions with the prokaryotic populations are still inadequately comprehended. In this study, 2121 viral operational taxonomic units (vOTUs) were recovered from 12 anaerobic fatty acid-fed reactors. Notably, 63.1% of these vOTUs could not be assigned to any known family, revealing a substantial presence of uncharted phages specifically associated with AD environments. Over half of the vOTUs associated with hosts had the capability to infect multiple hosts, ranging from 2 to 49, with a prevalent tendency to infect 2-5 hosts. In silico predictions of phage-host linkages uncovered that only a small fraction of vOTUs were shared across different functional groups, including fermentative bacteria, syntrophic fatty acid-oxidising bacteria (SFOB) and methanogens. Phages linked to hosts in all three groups primarily consisted of generalists and temperate species, especially those linked to SFOB. Additionally, metabolic reconstruction identified auxiliary metabolic genes participating in fatty acid degradation, methanogenesis and energy conservation. The present study provides insights into phage characteristics and their in situ interactions with prokaryotic hosts, highlighting their ecological role in AD systems.

Plant detritus is abundant in grasslands but decomposes slowly and is relatively nutrient-poor, whereas animal carcasses are labile and nutrient-rich. Recent studies have demonstrated that labile nutrients from carcasses can significantly alter the long-term soil microbial function at an ecosystem scale. However, there is a paucity of knowledge on the functional and structural response and temporal scale of soil microbiomes beneath large herbivore carcasses. This study compared microbiome functions and structures of soil beneath Connochaetes taurinus (hereafter 'wildebeest') carcasses at various postmortem intervals of decomposition to matched control samples over 18 months. Microbial functions were compared by their community-level physiological profiles determined by sole-carbon substrate utilisation and structures by metagenomic sequences using 16S rRNA gene markers. Overall metabolism and metabolic diversity remained increased and functionally dissimilar to control soils throughout the experimental period, with successive sole-carbon substrate utilisation observed. Conversely, diversity was initially reduced and structurally dissimilar from the control soil but recovered within the experimental period. The study contributes to the knowledge of carcass decomposition by investigating the long-term soil microbiome dynamics resulting from large herbivore carcasses decomposing in a mesic grassland. Microbial functional succession and ecologically relevant bacterial biomarkers of soil beneath the decomposing carcasses were identified for various postmortem intervals.

The unintended microbiological production of hydrogen sulphide (HS) poses a significant challenge in engineered systems, including sewage treatment plants, landfills and aquaculture systems. Although sulphur-rich amino acids and other substrates conducive to non-sulphate-based HS production are frequently present, the capacity and potential of various microorganisms to perform sulphate-free HS production remain unclear. In this study, we identify the identity, activity and genomic characteristics of bacteria that degrade cysteine to produce HS in anaerobic enrichment bioreactors seeded with material from aquaculture systems. Our comparison with canonical sulphate-reducing bacteria reveals that both sulphur sources contribute to microbial HS production, with cysteine facilitating a more rapid process compared to sulphate. 16S rRNA amplicon sequencing and metagenomic analysis identified four bacterial families-Dethiosulfatibacteraceae, Fusobacteriaceae, Vibrionaceae and Desulfovibrionaceae-as central to non-sulphate HS production. Metagenome- and metatranscriptome-assembled genomes elucidated the primary cysteine degradation pathway mediated by cysteine desulphidase cyuA and indicated that some bacteria may also utilise cysteine as a carbon source in sulphate-based HS production.

Rhizobia and legumes form a symbiotic relationship resulting in the formation of root structures known as nodules, where bacteria fix nitrogen. Legumes release flavonoids that are detected by the rhizobial nodulation (Nod) protein NodD, initiating the transcriptional activation of nod genes and subsequent synthesis of Nod Factors (NFs). NFs then induce various legume responses essential for this symbiosis. Although evidence suggests differential regulation of nodD expression and NF biosynthesis during symbiosis, the necessity of this regulation for the formation of nitrogen-fixing nodules remains uncertain. Here, we demonstrate that deletion of the Rlv3841 NodD regulatory domain results in a constitutively active protein (NodD) capable of activating NF biosynthesis gene expression without the presence of flavonoids. Optimised constitutive expression of nodD or nodD3 in nodD null mutants led to wild-type levels of nodulation and nitrogen fixation in pea and M. truncatula, respectively, indicating that flavonoid-regulated nodD expression is not essential for supporting symbiosis. These findings illustrate that transcriptional control of flavonoid-independent NodD regulators can be employed to drive NF biosynthesis, which holds potential for engineering symbiosis between rhizobia and cereals equipped with reconstituted NF receptors.

Symbiotic cnidarians, such as sea anemones and corals, rely on their mutualistic microalgal partners (Symbiodiniaceae) for survival. Marine heatwaves can disrupt this partnership, and it has been proposed that introducing experimentally evolved, heat-tolerant algal symbionts could enhance host thermotolerance. To test this hypothesis, the sea anemone Exaiptasia diaphana (a coral model) was inoculated with either the heterologous wild type or heat-evolved algal symbiont, Cladocopium proliferum, and homologous wild-type Breviolum minutum. The novel symbioses persisted for 1.5 years and determined holobiont thermotolerance during a simulated summer heatwave. Anemones hosting SS8, one of the six heat-evolved strains tested, exhibited the highest thermotolerance. Notably, anemones hosting the wild-type C. proliferum (WT10) were the second most thermally tolerant group, whereas anemones hosting the heat-evolved SS5 or SS9 strains were among the most thermosensitive. Elevated temperatures led to an increase in the levels of many amino acids and a decrease in tricarboxylic acid (TCA) metabolites in all anemone hosts, potentially indicating an increase in autophagy and a reduction in energy and storage production. Some consistent differences were observed in changes in metabolite levels between anemone groups in response to elevated temperature, suggesting that the algal symbiont influenced host metabolome and nutritional budget.

Evolutionary novelty has been one of the central themes in the field of evolutionary biology for many years. Structural and functional innovations such as scales in the reptiles, fins in the fishes and mammary glands in the mammals have been the focus of the studies. Insights obtained from these studies have shaped the criterion for the identification of novelty as well as provide the framework for studying novelty. In this article, I argue that unicellular organisms present an excellent opportunity for the investigation of evolutionary novelty. Even though bacteria share some fundamental aspects of novelty with higher organisms, there are definite departures. Here, I outline these departures in four different contexts: criterion for the identification of novelty, types of evolutionary novelties, level of biological complexity that bacteria embody and, most importantly, the role of the environment. Identifying the role of the environment allows the categorisation of novelty as probable or improbable and adaptive or latent. This categorisation of novel traits, based on the role of the environment, can facilitate the study of novelty in bacteria. Insights obtained from such studies are crucial for understanding the fundamental aspects of evolutionary novelty.

Bacterial degradation of ubiquitous and persistent steroids such as steroid hormones is important for their removal from the environment. Initial studies of steroid degradation in anaerobic bacteria suggested that ring-cleaving hydrolases are involved in oxygen-independent sterane skeleton degradation. However, the enzymes involved in ring A cleavage of the common intermediate androsta-1,4-diene-3,17-dione have remained unknown. Here, we enriched a ring A hydrolase from cholesterol/nitrate grown Sterolibacterium denitrificans and from Escherichia coli after heterologous expression of its gene. This enzyme specifically cleaves the cyclic 1,3-diketone of the central degradation intermediate, androsta-1,3,17-trione to 1,17-dioxo-2,3-seco-androstan-3-oate (DSAO), a hallmark reaction of anaerobic steroid degradation. The highly conserved ring A hydrolase was identified in all known and many previously unknown steroid-degrading proteobacteria. Using enriched enzymes, we enzymatically produced DSAO from the chemically synthesised androsta-1-en-3,17-dione precursor, allowing the identification of subsequent metabolites involved in ring A degradation. The results obtained suggest the involvement of an additional hydrolase, an aldolase, and a β-oxidation-like cascade for complete ring A degradation to form the three-ring 5,10-seco-1,2,3,4-tetranorandrosta-5,17-dione. The results identified a key enzyme of anaerobic steroid degradation that may serve as a functional marker for monitoring steroid contaminant degradation at anoxic environmental sites.

Human activities such as agriculture and urban development are linked to water quality degradation. Canada represents a large and heterogeneous landscape of freshwater lakes, where variations in climate, geography and geology interact with land cover alteration to influence water quality differently across regions. In this study, we investigated the influence of water quality and land use on bacterial communities across 12 ecozones. At the pan-Canadian scale, total phosphorus (TP) was the most significant water quality variable influencing community structure, and the most pronounced shift was observed at 110 μg/L of TP, corresponding to the transition from eutrophic to hypereutrophic conditions. At the regional scale, water quality significantly explained bacterial community structure in all ecozones. In terms of land use effect, at the pan-Canadian scale, agriculture and, to a lesser extent, urbanisation were significant land use variables influencing community structure. Regionally, in ecozones characterised by extensive agriculture, this land cover variable was consistently significant in explaining community structure. Likewise, in extensively urbanised ecozones, urbanisation was consistently significant in explaining community structure. Overall, these results demonstrate that bacterial richness and community structure are influenced by water quality and shaped by agriculture and urban development in different ways.

Streptofilum capillatum was recently described and immediately caught scientific attention, because it forms a phylogenetically deep branch in the streptophytes and is characterised by a unique cell coverage composed of piliform scales. Its phylogenetic position and taxonomic rank are still controversial discussed. In the present study, we isolated further strains of Streptofilum from biocrusts in sand dunes and Arctic tundra soil. Molecular and morphological characterisation including transmission electron microscopy confirmed that both new strains belong to Streptofilum. The Arctic strain is described as a new species, Streptofilum arcticum sp. nov., based on molecular differences, a specific sarcinoid morphology and unique ultrastructure with massive cell coverage composed of pili-shaped scales. A comprehensive characterisation of the ecophysiological traits of both new Streptofilum isolates and the original one revealed a broad temperature tolerance, a rapid recovery of photosynthetic performance after desiccation, an efficient photosynthesis at low light and a tolerance to high-light conditions. In addition, Streptofilum could cope with UV irradiation, but only S. capillatum grew under UV exposure. All Streptofilum strains are well-adapted to water-deprived terrestrial habitats such as biocrusts. From this study it can be concluded that already early-branching streptophytes were able to tolerate terrestrial conditions.

Legionella pneumophila, the causative agent of Legionnaires' disease, employs the Icm/Dot Type IV secretion system (T4SS) to replicate in amoebae and macrophages. The opportunistic pathogen responds to stress by forming 'viable but non-culturable' (VBNC) cells, which cannot be detected by standard cultivation-based techniques. In this study, we document that L. pneumophila enters the VBNC state after exposure to heat stress at 50°C for 30 h, at 55°C for 5 h or at 60°C for 30 min, while still retaining metabolic activity and intact cell membranes. Resuscitation of heat-induced VBNC L. pneumophila neither occurred in amoebae nor in macrophages. VBNC L. pneumophila showed impaired uptake by phagocytes, formation of Legionella-containing vacuoles (LCVs), and Icm/Dot-dependent secretion of effector proteins. The T4SS was rapidly inactivated already upon exposure to 50°C for 3-5 h, while the bacteria were still culturable. The Legionella quorum sensing (Lqs)-LvbR network is implicated in VBNC induction, since the ∆lvbR and ∆lqsR mutant strains showed a more pronounced heat sensitivity than the parental strain, and the ∆lqsA mutant was less heat sensitive. Taken together, our results reveal that heat exposure of L. pneumophila rapidly inactivates the Icm/Dot T4SS before the VBNC state is induced, thus impairing resuscitation by amoebae.

The microbiome influences critical aspects of mosquito biology and variations in microbial composition can impact the outcomes of laboratory studies. To investigate how biotic and abiotic conditions in an insectary affect the composition of the mosquito microbiome, a single cohort of Aedes aegypti eggs was divided into three batches and transferred to three different climate-controlled insectaries within the Liverpool School of Tropical Medicine. The bacterial microbiome composition was compared as mosquitoes developed, the microbiome of the mosquitoes' food sources was characterised, environmental conditions over time in each insectary were measured, and mosquito development and survival were recorded. While developmental success was similar across all three insectaries, differences in microbiome composition were observed between mosquitoes from each insectary. Environmental conditions and bacterial input via food sources varied between insectaries, potentially contributing to the observed differences in microbiome composition. At both adult and larval stages, specific members of the mosquito microbiome were associated with particular insectaries; the insectary with less stable and cooler conditions resulted in a slower pupation rate and higher diversity of the larval microbiome. These findings underscore that even minor inconsistencies in rearing conditions can affect the composition of the mosquito microbiome, which may influence experimental outcomes.

Ecological assembly-the process of ecological community formation through species introductions-has recently seen exciting theoretical advancements across dynamical, informational, and probabilistic approaches. However, these theories often remain inaccessible to non-theoreticians, and they lack a unifying lens. Here, I introduce the assembly graph as an integrative tool to connect these emerging theories. The assembly graph visually represents assembly dynamics, where nodes symbolise species combinations and edges represent transitions driven by species introductions. Through the lens of assembly graphs, I review how ecological processes reduce uncertainty in random species arrivals (informational approach), identify graphical properties that guarantee species coexistence and examine how the class of dynamical models constrain the topology of assembly graphs (dynamical approach), and quantify transition probabilities with incomplete information (probabilistic approach). To facilitate empirical testing, I also review methods to decompose complex assembly graphs into smaller, measurable components, as well as computational tools for deriving empirical assembly graphs. In sum, this math-light review of theoretical progress aims to catalyse empirical research towards a predictive understanding of ecological assembly.

Cellulolytic flagellates are essential for the symbiotic digestion of lignocellulose in the gut of lower termites. Most species are associated with host-specific consortia of bacterial symbionts from various phyla. 16S rRNA-based diversity studies and taxon-specific fluorescence in situ hybridization revealed a termite-specific clade of Actinomycetales that colonise the cytoplasm of Trichonympha spp. and other gut flagellates, representing the only known case of intracellular Actinomycetota in protists. Comparative analysis of eleven metagenome-assembled genomes from lower termites allowed us to describe them as new genera of Bifidobacteriaceae. Like the previously investigated Candidatus Ancillula trichonymphae, they ferment sugars via the bifidobacterium shunt but, unlike their free-living relatives, experienced significant genome erosion. Additionally, they acquired new functions by horizontal gene transfer from other gut bacteria, including the capacity to produce hydrogen. Members of the genus Ancillula (average genome size 1.56 ± 0.2 Mbp) retained most pathways for the synthesis of amino acids, including a threonine/serine exporter, providing concrete evidence for the basis of the mutualistic relationship with their host. By contrast, Opitulatrix species (1.23 ± 0.1 Mbp) lost most of their biosynthetic capacities, indicating that an originally mutualistic symbiosis is on the decline.

The Hepatincolaceae (Alphaproteobacteria) are a group of bacteria that inhabit the gut of arthropods and other ecdysozoans, associating extracellularly with microvilli. Previous phylogenetic studies, primarily single-gene analyses, suggested their relationship to the Holosporales, which includes intracellular bacteria in protist hosts. However, the genomics of Hepatincolaceae is still in its early stages. In this study, the number of available Hepatincolaceae genomes was increased to examine their evolutionary and functional characteristics. It was found that the previous phylogenetic grouping with Holosporales was incorrect due to sequence compositional biases and that Hepatincolaceae form an independent branch within the Hepatincolaceae. This led to a reinterpretation of their features, proposing a new evolutionary scenario that involves an independent adaptation to host association compared to the Holosporales, with distinct specificities. The Hepatincolaceae exhibit greater nutritional flexibility, utilising various molecules available in the host gut and thriving in anaerobic conditions. However, they have a less complex mechanism for modulating host interactions, which are likely less direct than those of intracellular bacteria. In addition, representatives of Hepatincolaceae show several lineage-specific traits related to differences in host species and life conditions.

Deep-sea sediments contain a large number of Thaumarchaeota that are phylogenetically distinct from their pelagic counterparts. However, their ecology and evolutionary adaptations are not well understood. Metagenomic analyses were conducted on samples from various depths of a 750-cm sediment core collected from the Mariana Trench Challenger Deep. The abundance of Thaumarchaeota and archaeal amoA generally decreased with depth, except for an unexpected peak midway through the core. The thaumarchaeotal metagenome-assembled genomes were classified into diverse phylogenetic clusters associated with amoA-NP-γ, amoA-NP-θ, and amoA-NP-δ of ammonia-oxidising Thaumarchaeota and non-ammonia-oxidising lineages. The most abundant group was within amoA-NP-γ, which is usually found in coastal and shallow habitats, indicating potential niche expansion from marine shallow to hadal environments. This benthic group showed within-species genomic variations compared to the previously identified Hadal water group, suggesting microdiversification of hadal Thaumarchaeota along with niche separation between benthic and pelagic environments. Evolutionary adaptations associated with the benthic-to-pelagic transition included reduced genome size, loss of motility/cell adhesion, altered energy metabolism, and different mechanisms for substrate acquisition and regulation (e.g., ammonium). These findings offer new insights into the evolution of hadal Thaumarchaeota and demonstrate, for the first time, intraspecies-level genomic variation in Thaumarchaeota related to the benthic-versus-pelagic niche partitioning in the deep ocean.

Active geothermal systems are relatively rare in Antarctica and represent metaphorical islands ideal to study microbial dispersal. In this study, we tested the macro-ecological concept that high dispersal rates result in communities being dominated by either habitat generalists or specialists by investigating the microbial communities on four geographically separated geothermal sites on three Antarctic volcanoes (Mts. Erebus, Melbourne, and Rittman). We found that the microbial communities at higher temperature (max 65°C) sites (Tramway Ridge on Erebus and Rittmann) were unique from each other and were dominated by a variety of novel Archaea from class Nitrososphaeria, while lower temperature (max 50°C) sites (Western Crater on Erebus and Melbourne) had characteristically mesophilic communities (Planctomycetes, Acidobacteriota, etc.) that were highly similar. We found that 97% of the detected microbial taxa were regional habitat specialists, with no generalists, with community assembly driven by high dispersal rates and drift (25% and 30% of community assembly, respectively), not environmental selection. Our results indicate that for microbial communities experiencing high dispersal rates between isolated communities, habitat specialists may tend to out-compete habitat generalists.