Jillian F. Banfield’s research while affiliated with University of Melbourne and other places

Genomes from metagenomes have revolutionised our understanding of microbial diversity, ecology, and evolution, propelling advances in basic science, biomedicine, and biotechnology. Assembly algorithms that take advantage of increasingly available long-read sequencing technologies bring the recovery of complete genomes directly from metagenomes within reach. However, assessing the accuracy of the assembled long reads, especially from complex environments that often include poorly studied organisms, poses remarkable challenges. Here we show that erroneous reporting is pervasive among long-read assemblers and can take many forms, including multi-domain chimeras, prematurely circularized sequences, haplotyping errors, excessive repeats, and phantom sequences. Our study highlights the need for rigorous evaluation of the algorithms while they are in development, and options for users who may opt for more accurate reads than shorter runtimes.

Genome editing is transforming plant biology by enabling precise DNA modifications. However, delivery of editing systems into plants remains challenging, often requiring slow, genotype-specific methods such as tissue culture or transformation¹. Plant viruses, which naturally infect and spread to most tissues, present a promising delivery system for editing reagents. However, many viruses have limited cargo capacities, restricting their ability to carry large CRISPR-Cas systems. Here we engineered tobacco rattle virus (TRV) to carry the compact RNA-guided TnpB enzyme ISYmu1 and its guide RNA. This innovation allowed transgene-free editing of Arabidopsis thaliana in a single step, with edits inherited in the subsequent generation. By overcoming traditional reagent delivery barriers, this approach offers a novel platform for genome editing, which can greatly accelerate plant biotechnology and basic research.

To date, the only known Lanthanide (Ln)-dependent enzymes are pyrroloquinoline quinone-dependent alcohol dehydrogenases. When compared to their Ca dependent counterparts, there is an emerging picture that Ln-dependent versions of these enzymes are generally more efficient, are preferentially upregulated in the presence of Ln when there is functional redundancy, and may even be evolutionarily older. Ln-utilising microbes have furthermore evolved diverse means of solubilizing and acquiring Ln, enabling them to utilise Ln even at trace concentrations. The ocean is the largest dissolved organic carbon pool on Earth, yet the diversity and prevalence of Ln-dependent carbon metabolisms in the ocean is unknown. Here we show that Ln-utilising methanol-, ethanol- and putative sorbose- and glucose-dehydrogenase genes are ubiquitous in the ocean and are highly transcribed, despite extremely low concentrations of Ln in seawater. These enzymes occur in the genomes of 20% of marine microbes, with several individual organisms hosting dozens of unique Ln-utilising enzymes. We found that active microbial methanol oxidation in the ocean is almost entirely Ln-dependent. The widespread biological utility of Ln may help to explain the nutrient-like vertical concentration profiles of these elements in ocean waters and may exert an influence on rare earth element concentration patterns. Microbial Ln-utilisation is a poorly understood component of marine rare earth element biogeochemistry, with potentially important implications for the carbon cycle. The ocean microbiome will be a rich resource for future research into biologically inspired solutions to lanthanide extraction and purification.

Mediterranean grasslands, vital natural and agricultural ecosystems, experience seasonal variation in water content that likely affect microbial activity. We used metagenomics-informed stable isotope probing to investigate how the activities of microorganisms in Angelo Reserve (2160 mm rainfall) and Hopland (956 mm rainfall) soil change over depth and the seasons. At both sites, we find that the relative abundances of organisms in shallow soil changes relatively little but the most abundant organisms vary greatly with soil depth. Notably the highest levels of isotope incorporation, indicative of growth, occurs in deep soils. The active part of the 0-10 cm soil community varies over time, especially in Hopland soils during the fall rewetting. We defined a large, novel clade of Actinomycetota with notable capacity for thiosulfate oxidation whose representatives are prevalent and active in deep soils (>20 cm) across both ecosystems. Active Saccharibacteria unexpectedly encode nucleotide synthesis genes that enabled isotope incorporation while growing in shallow Angelo soils over all time periods. In contrast to predicted episymbiotic lifestyles of Saccharibacteria, other highly active bacteria are predicted predators. Obligately predatory Pseudobdellovibrio are active in intermediate depth Hopland soils whereas bacteria of the order Haliangiales are active in shallow Angelo soils. Supporting predatory lifestyles of Haliangiales, we used in silico structure prediction to assemble a large protein complex that we identify as a contractile injection system. Overall, the results indicate the potential for active carbon turnover in deep grassland soil and strong seasonal changes in the active members of microbial communities, despite relatively minor shifts in community composition.

Background Underground research laboratories (URLs) provide a window on the deep biosphere and enable investigation of potential microbial impacts on nuclear waste, CO2 and H2 stored in the subsurface. We carried out the first multi-year study of groundwater microbiomes sampled from defined intervals between 140 and 400 m below the surface of the Horonobe and Mizunami URLs, Japan. Results We reconstructed draft genomes for > 90% of all organisms detected over a four year period. The Horonobe and Mizunami microbiomes are dissimilar, likely because the Mizunami URL is hosted in granitic rock and the Horonobe URL in sedimentary rock. Despite this, hydrogen metabolism, rubisco-based CO2 fixation, reduction of nitrogen compounds and sulfate reduction are well represented functions in microbiomes from both URLs, although methane metabolism is more prevalent at the organic- and CO2-rich Horonobe URL. High fluid flow zones and proximity to subsurface tunnels select for candidate phyla radiation bacteria in the Mizunami URL. We detected near-identical genotypes for approximately one third of all genomically defined organisms at multiple depths within the Horonobe URL. This cannot be explained by inactivity, as in situ growth was detected for some bacteria, albeit at slow rates. Given the current low hydraulic conductivity and groundwater compositional heterogeneity, ongoing inter-site strain dispersal seems unlikely. Alternatively, the Horonobe URL microbiome homogeneity may be explained by higher groundwater mobility during the last glacial period. Genotypically-defined species closely related to those detected in the URLs were identified in three other subsurface environments in the USA. Thus, dispersal rates between widely separated underground sites may be fast enough relative to mutation rates to have precluded substantial divergence in species composition. Species overlaps between subsurface locations on different continents constrain expectations regarding the scale of global subsurface biodiversity. Conclusions Our analyses reveal microbiome stability in the sedimentary rocks and surprising microbial community compositional and genotypic overlap over sites separated by hundreds of meters of rock, potentially explained by dispersal via slow groundwater flow or during a prior hydrological regime. Overall, microbiome and geochemical stability over the study period has important implications for underground storage applications. Supplementary Information The online version contains supplementary material available at 10.1186/s40793-024-00649-3.

Snowmelt in high-elevation watersheds triggers a microbial bloom and crash that affects nitrogen (N) export. Predicting watershed N dynamics as snowpack declines is a challenge because the mechanisms that underlie this microbial bloom and crash are uncertain. Using a multi-omic approach, we show that the dynamic molecular properties of dissolved organic N, plus high gene expression for peptidases that recycle microbial biomass, suggested that microbial turnover provided N for biosynthesis during the microbial bloom. Amino acid fermentation by Bradyrhizobia produced organic acids that also fueled denitrification and dissimilatory nitrate reduction to ammonia (DNRA) during snowmelt. Nitrification in spring was driven by Spring-Adapted Nitrososphaerales, which utilized ammonium derived from osmolyte degradation by Winter-Adapted Solirubrobacteraceae. High DNRA gene expression after snowmelt suggested significant nitrate retention, increasing watershed N retention potential. However, declining snowfall may compromise microbial regulation of soil N retention, with implications for watershed N export

In mine wastewaters, three microbial sulfur oxidation pathways have the potential to cause different water quality outcomes. These outcomes can differ from abiotic models of sulfate and acidity predictions currently used to monitor potential sulfur risks. However, studies integrating microbiology and geochemistry in active mine tailings impoundments are very limited. Here, we developed a novel diagnostic approach to detect microbially driven sulfur pathways. Within this 28-day study, eight on-site, 500 L mesocosms were filled with water extracted directly from the water cap of an active Ni/Cu mine tailings impoundment. Diverse combinations of tailings, sulfur compounds, and nitrate amendments were added to the mesocosms simulating common operational variations experienced by active tailings impoundments. Mesocosm results linked complete SOx, S4I, and incomplete SOx + rDSR pathway occurrence (metagenomes, inferred from the identity, i.e. 16S rRNA) and activity (mRNA) to physiochemistry and sulfur geochemistry. By integrating the three lines of evidence, the diagnostic approach was able to identify which sulfur pathways were active under varying physiochemical conditions and how geochemical outcomes were affected. A relationship emerged between acid generation and soxCD expression (soxCD expression indicates the complete SOx pathway activity). However, observed proton yields and sulfate concentrations were less than those predicted by complete SOx pathway activity alone. This indicates other sulfur pathways, e.g. the partial S4I pathway (within Thiomonas and Halothiobacillus), and/or activity of the incomplete SOx pathway (within Thiobacillus and Desulfurivibrio) when either not coupled to rDSR, or paired with use of nitrate, influenced overall sulfur outcomes along with the complete SOx pathway.

The ribosome is the universal translator of the genetic code and is shared across all life. Despite divergence in ribosome structure over the course of evolution, the peptidyl transferase center (PTC), the catalytic site of the ribosome, has been thought to be nearly universally conserved. Here, we identify clades of archaea that have highly divergent ribosomal RNA sequences in the PTC. To understand how these PTC sequences fold, we determined cryo-EM structures of the Pyrobaculum calidifontis ribosome. We find that sequence variation leads to the rearrangement of key PTC base triples and differences between archaeal and bacterial ribosomal proteins also enable sequence variation in archaeal PTCs. Finally, we identify a novel archaeal ribosome hibernation factor that differs from known bacterial and eukaryotic hibernation factors and is found in multiple archaeal phyla. Overall, this work identifies factors that regulate ribosome function in archaea and reveals a larger diversity of the most ancient sequences in the ribosome.

Asgard archaea are of great interest as the progenitors of Eukaryotes, but little is known about the mobile genetic elements (MGEs) that may shape their ongoing evolution. Here, we describe MGEs that replicate in Atabeyarchaeia, a wetland Asgard archaea lineage represented by two complete genomes. We used soil depth–resolved population metagenomic data sets to track 18 MGEs for which genome structures were defined and precise chromosome integration sites could be identified for confident host linkage. Additionally, we identified a complete 20.67 kbp circular plasmid and two family-level groups of viruses linked to Atabeyarchaeia, via CRISPR spacer targeting. Closely related 40 kbp viruses possess a hypervariable genomic region encoding combinations of specific genes for small cysteine-rich proteins structurally similar to restriction-homing endonucleases. One 10.9 kbp integrative conjugative element (ICE) integrates genomically into the Atabeyarchaeum deiterrae-1 chromosome and has a 2.5 kbp circularizable element integrated within it. The 10.9 kbp ICE encodes an expressed Type IIG restriction-modification system with a sequence specificity matching an active methylation motif identified by Pacific Biosciences (PacBio) high-accuracy long-read (HiFi) metagenomic sequencing. Restriction-modification of Atabeyarchaeia differs from that of another coexisting Asgard archaea, Freyarchaeia, which has few identified MGEs but possesses diverse defense mechanisms, including DISARM and Hachiman, not found in Atabeyarchaeia. Overall, defense systems and methylation mechanisms of Asgard archaea likely modulate their interactions with MGEs, and integration/excision and copy number variation of MGEs in turn enable host genetic versatility.

Rice paddies contribute substantially to atmospheric methane (CH 4 ) and these emissions are expected to increase as the need to feed the human population grows. Here, we show that two independent rice genotypes overexpressing genes for PLANT PEPTIDES CONTAINING SULFATED TYROSINE ( PSY ) reduced cumulative CH 4 emissions by 38% (PSY1) and 58% (PSY2) over the growth period compared with controls. Genome-resolved metatranscriptomic data from rhizosphere soils reveal lower ratios of gene activities for CH 4 production versus consumption, decrease in activity of H 2 -producing genes, and increase in bacterial H 2 oxidation pathways in the PSY genotypes. Metabolic modeling using metagenomic and metabolomic data predicts elevated levels of H 2 oxidation and suppressed H 2 production in the PSY rhizosphere. The H 2 -oxidizing bacteria have more genes for utilization of gluconeogenic acids than H 2 -producing counterparts, and their activities were likely stimulated by the observed enrichment of gluconeogenic acids (mostly amino acids) in PSY root exudates. Together these results suggest that decreased CH 4 emission is due to the reduction of H 2 available for hydrogenotrophic methanogenesis. The combination of rice phenotypic characterization, microbiome multi-omic analysis, and metabolic modeling described here provides a powerful strategy to discover the mechanisms by which specific plant genotypes can alter biogeochemical cycles to reduce CH 4 emissions.