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Sachdeva, R.

Publications and source records attributed to Sachdeva, R..

At least 19 recordsLinked to original sources

Genetic exchange drives relatedness of large linear and circular extrachromosomal elements of archaea

Archaeal extra-chromosomal elements (ECEs) play key roles in archaeal host biology and evolution, yet little is known about their diversity and interactions with each other. Here, we report 42 complete 66 - 165 kbp genomes: 23 "Klingons", 8 "Romulans" and "Romulan-like elements", and 11 "Ferengi", three new families of archaeal ECEs from diverse terrestrial environments. Methylation patterns, phylogenetics and gene sharing analyses suggest that Klingons and Romulans associate with Candidatus Methanoperedens and Ferengi are likely hosted by Candidatus Bathyarchaeaota. Like known archaeal viruses and plasmids, these ECEs both have circular genomes and replicate bi-directionally, unlike the linear Borg and mini-Borg ECEs that also replicate in Methanoperedens. The new ECE genomes all encode genes for methylation, glycosylation, protein degradation, and novel TnpB-like enzymes, which may interact with novel CRISPR-like arrays. All genomes also encode proteins with strong structural similarity to archaeal viral capsid proteins, and some are predicted to multimerize into pentameric or hexameric subunit capsid-like arrays and thus may encapsulate themselves. The three new ECE families share gene content with each other and with Borgs and mini-Borgs, often concentrated in specific genomic regions. Discordance among single gene and genomic phylogenies suggests that apparent relatedness of the five distinct ECE types stems from extensive horizontal gene transfer, rather than evolution from a common ancestor. The three new families of ECEs broaden our understanding of the diversity of archaeal ECEs and highlight the role of horizontal gene transfer in modulating the archaeal and archaeal ECE gene pools.

genomics↗

Rice soil methane emissions are modulated by microbial cohorts that shape redox succession over the growing season

Rice paddies are major anthropogenic methane sources, yet the community interactions regulating emissions remain poorly resolved. This field experiment combined genome-resolved metagenomics, metatranscriptomics, and biogeochemistry to identify microbial cohorts directly and indirectly involved in methane cycling over a rice growing season. Cohorts routed reducing equivalents to different terminal electron-accepting processes and changed in abundance in response to depth and time-dependent shifts in environmental conditions. Methanogenic activity peaked late in the season in the rhizosphere and surface soil, whereas activity in 2-10 cm bulk soil peaked earlier, indicating that methanogenesis was temporally partitioned across soil compartments. Geobacter-like extracellular electron transfer was the most highly expressed alternate sink for reducing equivalents. Methane consumption was vertically partitioned between surface type I methanotrophs and deeper type II methanotrophs and anaerobic methane-oxidizing archaea. We infer that methane emissions reflect seasonal shifts in microbial cohorts that redirect methanogenic substrates through competing redox processes.

microbiology↗

LINE-1 transposon derepression and epigenetic remodeling in Retinoblastoma

Retinoblastoma (RB), the most common pediatric intraocular malignancy, is initiated by biallelic inactivation of the RB1 tumor suppressor gene. Although LINE-1 (L1) retrotransposon activation has been linked to genomic instability and tumor evolution in several adult cancers, its contribution to pediatric malignancies remains poorly understood. Here, we investigated L1 expression in retinoblastoma samples from an Indian pediatric cohort by integrating transcriptomic, locus-specific, and immunohistochemistry. Transcriptomic profiling revealed increased L1 expression in RB tumors compared with control retinal tissues, which was validated by ORF1p immunostaining in tumor sections. Locus-specific analysis identified a subset of transcriptionally active L1 loci, suggesting selective activation of young L1 elements. We also observed altered expression of genes associated with proliferation, chromatin regulation, and oncogenic signaling, including MYCN, MDM2, E2F3, SOX4 and CREBBP. Our findings identify previously underexplored L1 activation as a molecular feature associated with retinoblastoma and suggest that disruption of transposon silencing accompanies the RB transcriptional landscape. Validation in larger patient cohorts will be important; however, this study provides a foundation for exploring L1 activity as a potential molecular marker and future therapeutic vulnerability in retinoblastoma. HighlightsO_LITranscriptomic profiling reveals increased LINE-1 expression in retinoblastoma. C_LIO_LIORF1p protein is robustly expressed in retinoblastoma. C_LIO_LIA subset of LINE-1 loci exhibits transcriptional activation, suggesting locus-specific derepression. C_LI In BriefRetinoblastoma is initiated by RB1 loss, but the contribution of transposable elements to its molecular landscape remains poorly understood. Here, we show that retinoblastoma is associated with LINE-1 promoter hypomethylation, increased LINE-1 transcript expression, ORF1p accumulation, and preferential activation of selected LINE-1 loci. These changes occur together with altered expression of oncogenic signalling pathways and host factors involved in transposable element repression, identifying dysregulated LINE-1 control as a molecular feature of this paediatric cancer. Graphical Abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=115 SRC="FIGDIR/small/740688v1_ufig1.gif" ALT="Figure 1"> View larger version (24K): org.highwire.dtl.DTLVardef@197e89org.highwire.dtl.DTLVardef@1c26c04org.highwire.dtl.DTLVardef@14180a4org.highwire.dtl.DTLVardef@16493bb_HPS_FORMAT_FIGEXP M_FIG C_FIG Retinoblastoma exhibits epigenetic activation of LINE-1 characterized by promoter hypomethylation, increased expression of young LINE-1 elements, and ORF1p accumulation. This is accompanied by coordinated transcriptomic remodeling and disruption of host mechanisms that normally restrain transposable elements, linking LINE-1 activation to the molecular landscape of retinoblastoma.

cancer biology↗

The origin and evolution of archaeal Borg extrachromosomal elements

Borgs are giant linear extrachromosomal elements (ECEs) of methane-oxidizing Methanoperedens archaea whose evolutionary origin and ecosystem distribution remain unknown. Here we detected 240 Borgs in diverse saline, soil, and freshwater ecosystems. 27 encode methyl-coenzyme M reductases central to methane metabolism, eight have full rRNA operons related to those of Methanoperedens, and some contain up to four ribosomal proteins. We also identified 323 mini-Borgs whose proteome content and gene phylogenies classify as a distinct ECE type. Based on 105 complete and near-complete genomes, Borgs and mini-Borgs share core genes in conserved order, indicating common ancestry. Phylogenetic and diversity analyses suggest that Borgs evolved via gene acquisition into a backbone inherited from mini-Borgs. This evolutionary trajectory mirrors those proposed for giant viruses of both eukaryotes and bacteria.

microbiology↗

Metagenomic strain-resolved DNA modification patterns link extrachromosomal genetic elements to host strains

DNA modification is central to microbial defense against extrachromosomal genetic elements (ECEs), consequently ECEs tend to adopt their hosts modification patterns. Shared ECE-host modification patterns enable linking ECEs to their hosts, but modification detection tools are designed for single genomes and are ineffective at metagenome scale. Here, we present MODIFI, software for detecting DNA modifications in metagenomes. MODIFI assumes that each k-mer in a metagenome is mostly unmodified and calculates background signal levels for that k-mer from PacBio HiFi reads, eliminating the need for matched control experiments. MODIFI ECE-host linkages were validated using >1,000 isolate and mock microbiome datasets. Illustrating the approach, we identified 315 strain-resolved, non-redundant ECE-host linkages in environmental and human metagenomes. In infant gut microbiomes, a chromosomal inversion in Enterococcus faecalis alters host and associated plasmid methylation motifs simultaneously. Overall, MODIFI solves a major bottleneck in DNA modification analysis and provides a foundational tool for understanding microbial epigenomics.

microbiology↗

Jumbo circular extrachromosomal elements of methane-oxidizing archaea with variably extensive metabolic and defense gene repertoires

Archaeal extrachromosomal elements (ECEs) are arguably the least well understood of all genetic elements, and few have > 200 kbp ("jumbo") genomes. Here, we report circular, jumbo ECEs with genomes of up to 535 kbp in length that associate with anaerobic methane-oxidizing Methanoperedens archaea. Notably, a 409-kbp genome related to jumbo ECEs is integrated into a subset of the [~]4.2 Mbp Methanoperedens chromosomes at the tRNA-Asp genes. This represents the largest integrative element in Archaea and supports the jumbo ECE - host association. Multiple genome alignment and phylogenetic analyses suggest that the large sizes were developed by extensive DNA acquisition from Methanoperedens. The newly identified ECEs encode, and in some cases express, metabolic genes such as tetrahydromethanopterin S-methyltransferase exclusively involved in methane oxidation, and genes for nitrogen and sulfur compound transformations. Also encoded are defense systems, some of which are absent in hosts, such as hybrid Type I/Type III-A CRISPR-Cas systems. In contrast to viruses and plasmids, they have host-like replication machinery and occur at stable copy ratios of 1.44 {+/-} 0.24 : 1 to the host. Overall, our results reveal a spectrum of jumbo ECEs of Methanoperedens, ranging from plasmid-like to minichromosome-like.

microbiology↗

Revealing the pervasive landscape of MGE-host interactions in situ with single-cell genomics

Mobile genetic elements (MGEs), including plasmids and viruses, drive microbial evolution and ecosystem dynamics, yet their distribution, host range, and functions remain poorly understood, especially among uncultivated lineages. Using [~]60,000 single-cell amplified genomes (SAGs) from host-associated and environmental microbiomes, we identified MGEs internal to or attached to individual cells, directly linking them to their hosts. Between 25% and 75% of cells contained at least one MGE, with gut-derived SAGs showing the highest MGE load. While most MGEs exhibited narrow, species-specific host ranges, a subset spanned multiple species and higher taxonomic ranks. We detected MGE clusters and complete, nearly identical genomes repeatedly associated with hosts across phyla, suggesting both genuinely broad host ranges and recurrent DNA entry events. We also identified microbial species acting as major donors or recipients in antibiotic resistance gene (ARG) exchange, and MGEs, including unclassified types that mediate extensive horizontal transfer of auxiliary functions. Together, these findings illuminate MGE-host associations and underscore that the genetic content of microbial cells is fundamentally linked to their MGEs in nature.

microbiology↗

Megaplasmids associate with Escherichia coli and other Enterobacteriaceae

Humans and animals are ubiquitously colonized by Enterobacteriaceae, a bacterial family that contains both commensals and clinically significant pathogens. Here, we report Enterobacteriaceae megaplasmids of up to 1.58 Mbp in length in infant and adult guts, and other microbiomes. Of 19 complete plasmid genomes, one was reconstructed from an E. coli isolate; others were linked to species of Citrobacter and Enterobacter via analysis of genome modification patterns. The detection of related plasmids in different Enterobacteriaceae, conjugation machinery, and more diverse modified motifs in certain plasmids compared to hosts suggests that these elements are self-transmissible, with a broad host range. The plasmids encode multi-drug efflux systems and potential secreted effectors. Up to 208 tRNAs are encoded and include sequence variants that may counter tRNA-centric defense mechanisms. Overall, the vast megaplasmid coding capacity may broaden host range, increase competitiveness, control invasion by other elements, and counter programmed cell death.

microbiology↗

Phage-based microbiome manipulation elucidates interactions within community context

Testing microbial interaction hypotheses remains a major challenge in microbiome research. Bacteriophages have the potential to probe organism-organism interactions within complex microbiomes, yet are rarely used for this purpose. Here, we isolated nine narrow host range phages that replicate in several ecologically important Variovorax species. As Variovorax CL14 degrades the plant root-stunting hormone auxin, we used phages to eliminate V. CL14 from a rhizosphere consortium and reestablished the stunted root phenotype. We used three of the phages to deplete another Variovorax, SCN45, from complex communities to test correlation network-based hypotheses of thiamine interdependencies. Genome-resolved metagenomics revealed that specific taxa decreased in relative abundance following Variovorax depletion and could be rescued by thiamine supplementation. Thus, we confirmed thiamine production as the mechanistic basis for interdependence. These experiments lay the foundation for research that employs wildtype and engineered phages to test interaction hypotheses and for targeted microbiome manipulation.

microbiology↗

Circular 23S rRNA within archaeal ribosomes

The ribosomal RNAs (rRNAs) that form the core of ribosomes are believed to occur as linear molecules. Here, we investigated rRNAs from diverse and mostly uncultivated archaea and found evidence that, in at least eight phylum-level archaeal lineages, the 23S rRNAs within mature ribosomes are circular. Sequencing of total cellular RNA indicates that the transcriptional abundances of archaeal circular 23S rRNAs vastly exceed those of linear counterparts, and linear versions are often undetectable. As the majority of rRNAs derive from mature ribosomes, the data suggested that the circular molecules are within the ribosomes. Thus, we directly sequenced RNA extracted from isolated ribosomes of a model archaeon, Methanosarcina acetivorans, and confirmed that the assembled 23S rRNAs are circular. Structural modeling places the 5 and 3 ends of the linear precursors of archaeal 23S rRNAs in close proximity to form a GNRA tetraloop (in which N is A, C, G, or U and R is A or G), consistent with their further circularization within ribosomes. We confirm the existence of circular 16S rRNA intermediates in transcriptomes of most archaea, yet a circular form is not evident in some distinct archaeal groups. Overall, the results uncover unexpected variations in the processing required to generate mature rRNAs and the conformation of functional molecules in archaeal ribosomes.

microbiology↗

Assemblies of long-read metagenomes suffer from diverse errors

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.

bioinformatics↗

Overturning circulation structures the microbial functional landscape of the South Pacific

Global overturning circulation partitions the deep ocean into regions with unique physicochemical characteristics, but the extent to which these water masses represent distinct ecosystems remains unknown. Here, we integrate extensive genomic information with hydrography and water mass age to delineate microbial taxonomic and functional boundaries across the South Pacific. Prokaryotic richness steeply increases with depth in the surface ocean, forming a "phylocline", below which richness is consistently high, dipping slightly in highly aged water. Reconstructed genomes self-organize into six spatially-distinct taxonomic cohorts and ten functionally-distinct biomes that are primarily structured by wind-driven circulation at the surface and density-driven circulation at depth. Overall, water physicochemistry, modulated at depth by water age, drives microbial diversity and functional potential in the pelagic ocean.

microbiology↗

The active subset of grassland soil microbiomes changes with soil depth, water availability and prominently features predatory bacteria and episymbionts

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.

bioinformatics↗

Convergent evolution of viral-like Borg archaeal extrachromosomal elements and giant eukaryotic viruses

Borgs are huge extrachromosomal elements of anaerobic methane-oxidizing archaea. They exist in exceedingly complex microbiomes, lack cultivated hosts and have few protein functional annotations, precluding their classification as plasmids, viruses or other. Here, we used in silico structure prediction methods to investigate potential roles for [~]10,000 Borg proteins. Prioritizing analysis of multicopy genes that could signal importance for Borg lifestyles, we uncovered highly represented de-ubiquitination-like Zn-metalloproteases that may counter host targeting of Borg proteins for proteolysis. Also prevalent are clusters of multicopy genes for production of diverse glycoconjugates that could contribute to decoration of the host cell surface, or of putative capsid proteins that we predict multimerize into hexagonal arrays. Features including megabase-scale linear genomes with inverted terminal repeats, genomic repertoires for energy metabolism, central carbon compound transformations and translation, and pervasive direct repeat regions are shared with giant viruses of eukaryotes, although analyses suggest that these parallels arose via convergent evolution. If Borgs are giant archaeal viruses they would fill the gap in the tri(um)virate of giant viruses of all three domains of life. One Sentence SummaryProtein analyses, informed by in silico protein structure prediction, revealed that Borgs share numerous features with giant eukaryotic viruses, suggesting that Borgs have a viral-like lifestyle and evolutionary convergence of large extrachromosomal elements across the Domains of Life.

microbiology↗

Reduced methane emissions in transgenic rice genotypes are associated with altered rhizosphere microbial hydrogen cycling

Rice paddies contribute substantially to atmospheric methane (CH4) 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 CH4 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 CH4 production versus consumption, decrease in activity of H2-producing genes, and increase in bacterial H2 oxidation pathways in the PSY genotypes. Metabolic modeling using metagenomic and metabolomic data predicts elevated levels of H2 oxidation and suppressed H2 production in the PSY rhizosphere. The H2-oxidizing bacteria have more genes for utilization of gluconeogenic acids than H2-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 CH4 emission is due to the reduction of H2 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 CH4 emissions.

microbiology↗

Characterization of sediment and granite hosted deep underground research laboratories reveals diverse microbiome functions, limited temporal variation and substantial genomic conservation

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. 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. Overall, microbiome and geochemical stability over the study period has important implications for underground storage applications.

microbiology↗

Genetic elements and defense systems drive diversification and evolution in Asgard archaea

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, wetland Asgard archaea phylum represented by two complete genomes. We used soil depth-resolved population metagenomic datasets 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 kilobase pair (kbp) circular plasmid (the first reported for Asgard archaea) and two 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 circularizable plasmid-like MGE integrates genomically into an Atabeyarchaeia chromosome and has a 2.5 kbp circularizable element integrated within it. The 10.9 kbp MGE encodes a highly expressed methylase with a sequence specificity matching an active methylation motif identified by PacBio 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.

microbiology↗

Asgard archaea modulate potential methanogenesis substrates in wetland soil

The roles of Asgard archaea in eukaryogenesis and marine biogeochemical cycles are well studied, yet their contributions in soil ecosystems are unknown. Of particular interest are Asgard archaeal contributions to methane cycling in wetland soils. To investigate this, we reconstructed two complete genomes for soil-associated Atabeyarchaeia, a new Asgard lineage, and the first complete genome of Freyarchaeia, and defined their metabolism in situ. Metatranscriptomics highlights high expression of [NiFe]-hydrogenases, pyruvate oxidation and carbon fixation via the Wood-Ljungdahl pathway genes. Also highly expressed are genes encoding enzymes for amino acid metabolism, anaerobic aldehyde oxidation, hydrogen peroxide detoxification and glycerol and carbohydrate breakdown to acetate and formate. Overall, soil-associated Asgard archaea are predicted to be non-methanogenic acetogens, likely impacting reservoirs of substrates for methane production in terrestrial ecosystems. One-Sentence SummaryComplete genomes of Asgard archaea, coupled with metatranscriptomic data, indicate roles in production and consumption of carbon compounds that are known to serve as substrates for methane production in wetlands.

microbiology↗