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Horizontal gene transfer (HGT) is a major source of phenotypic innovation and a mechanism of niche adaptation in prokaryotes. Quantification of HGT is critical to decipher its myriad roles in microbial evolution and adaptation. Advances in genome sequencing and bioinformatics have augmented our ability to understand the microbial world, particularly the direct or indirect influence of HGT on diverse life forms. Methods for detecting HGT can be classified into phylogenetic-based and parametric or composition-based approaches. Here, we exploited the complementary strengths of both the approaches to construct a high confidence horizontal gene flow network. Our network is unique in its ability to detect the transfer of native genes of a genome to genomes from other taxa, thus establishing donor and recipient organisms (taxa), rather than through a post hoc analysis as is the practice with several other approaches. The scale-free horizontal gene flow network presented here provides new insights into modes of transfer for the exchange of genetic information and also illuminates differential gene flow across phyla.

Horizontal gene transfer (HGT) is well described in prokaryotes: it plays a crucial role in evolution, and has functional consequences in insects and plants. However, less is known about HGT in humans. Studies have reported bacterial integrations in cancer patients, and microbial sequences have been detected in data from well-known human sequencing projects. Few of the existing tools for investigating HGT are highly automated. Thanks to the adoption of Nextflow for life sciences workflows, and to the standards and best practices curated by communities such as nf-core, fully automated, portable, and scalable pipelines can now be developed. Here we present nf-core/hgtseq to facilitate the analysis of HGT from sequencing data in different organisms. We showcase its performance by analysing six exome datasets from five mammals. Hgtseq can be run seamlessly in any computing environment and accepts data generated by existing exome and whole-genome sequencing projects; this will enable researchers to expand their analyses into this area. Fundamental questions are still open about the mechanisms and the extent or role of horizontal gene transfer: by releasing hgtseq we provide a standardised tool which will enable a systematic investigation of this phenomenon, thus paving the way for a better understanding of HGT.

In this issue of Cell Hostamp; Microbe, Millman et al. use bioinformatic and genetic approaches to discover 21 novel antiviral immune systems in prokaryotes. Remarkably, many of these systems bear homology to components of the human innate immune system, suggesting an evolutionary tie between prokaryotic and eukaryotic antiviral defenses.

The genomes of bacteria, archaea, and phage contain small amounts of C

Background Protein histidine phosphorylation (pHis) plays critical roles in prokaryotic signal transduction pathways and various eukaryotic cellular processes. It is estimated to account for 6–10% of the phosphoproteome, however only hundreds of pHis sites have been discovered to date. Due to the inherent disadvantages of experimental methods, it is an urgent task for developing efficient computational approaches to identify pHis sites. Results Here, we present a novel tool, pHisPred, for accurately identifying pHis sites from protein sequences. We manually collected the largest number of experimental validated pHis sites to build benchmark datasets. Using randomized tenfold CV, the weighted SVM-RBF model shows the best performance than other four commonly used classification models (LR, KNN, RF, and MLP). From ten thousands of features, 140 and 150 most informative features were individually selected out for eukaryotic and prokaryotic models. The average AUC and F1-score values of pHisPred were (0.81, 0.40) and (0.78, 0.46) for tenfold CV on the eukaryotic and prokaryotic training datasets, respectively. In addition, pHisPred significantly outperforms other tools on testing datasets, in particular on the eukaryotic one. Conclusion We implemented a python program of pHisPred, which is freely available for non-commercial use at https://github.com/xiaofengsong/pHisPred. Moreover, users can use it to train new models with their own data.

ATP-binding cassette (ABC) transporters are one of the largest families of membrane proteins in prokaryotic organisms. Much is now understood about the structure of these transporters and many reviews have been written on that subject. In contrast, less has been written on the assembly of ABC transporter complexes and this will be a major focus of this book chapter. The complexes are formed from two cytoplasmic subunits that are highly conserved (in terms of their primary and three-dimensional structures) across the whole family. These ATP-binding subunits give rise to the name of the family. They must assemble with two transmembrane subunits that will typically form the permease component of the transporter. The transmembrane subunits have been found to be surprisingly diverse in structure when the whole family is examined, with seven distinct folds identified so far. Hence nucleotide-binding subunits appear to have been bolted on to a variety of transmembrane platforms during evolution, leading to a greater variety in function. Furthermore, many importers within the family utilise a further external substrate-binding component to trap scarce substrates and deliver them to the correct permease components. In this chapter, we will discuss whether assembly of the various ABC transporter subunits occurs with high fidelity within the crowded cellular environment and whether promiscuity in assembly of transmembrane and cytoplasmic components can occur. We also discuss the new AlphaFold protein structure prediction tool which predicts a new type of transmembrane domain fold within the ABC transporters that is associated with cation exporters of bacteria and plants.

Interpreting phylogenetic trees requires a root, which provides the direction of evolution and polarizes ancestor-descendant relationships. But inferring the root using genetic data is difficult, particularly in cases where the closest available outgroup is only distantly related, which are common for microbes. In this chapter, we present a workflow for estimating rooted species trees and the evolutionary history of the gene families that evolve within them using probabilistic gene tree-species tree reconciliation. We illustrate the pipeline using a small dataset of prokaryotic genomes, for which the example scripts can be run using modest computer resources. We describe the rooting method used in this work in the context or other rooting strategies and discuss some of the limitations and opportunities presented by probabilistic gene tree-species tree reconciliation methods.

All living cells sense and respond to changes in external or internal conditions. Without that cognitive capacity, they could not obtain nutrition essential for growth, survive inevitable ecological changes, or correct accidents in the complex processes of reproduction. Wherever examined, even the smallest living cells (prokaryotes) display sophisticated regulatory networks establishing appropriate adaptations to stress conditions that maximize the probability of survival. Supposedly "simple" prokaryotic organisms also display remarkable capabilities for intercellular signalling and multicellular coordination. These observations indicate that all living cells are cognitive.

Indole and its derivatives are widespread across different life forms, functioning as signalling molecules in prokaryotes and with more diverse roles in eukaryotes. A majority of indoles found in the environment are attributed to bacterial enzymes converting tryptophan into indole and its derivatives. The involvement of indoles among lower organisms as an interspecies and intraspecies signal is well known, with many reports showing that inter-kingdom interactions involving microbial indole compounds are equally important as they influence defence systems and even the behaviour of higher organisms. This review summarizes recent advances in our understanding of the functional properties of indole and indole derivatives in diverse eukaryotes. Furthermore, we discuss current perspectives on the role of microbial indoles in human diseases such as diabetes, obesity, atherosclerosis, and cancers. Deciphering the function of indoles as biomarkers of metabolic state will facilitate the formulation of diet-based treatments and open unique therapeutic opportunities.

The transition from prokaryotic lateral gene transfer to eukaryotic meiotic sex is poorly understood. Phylogenetic evidence suggests that it was tightly linked to eukaryogenesis, which involved an unprecedented rise in both genome size and the density of genetic repeats. Expansion of genome size raised the severity of Muller’s ratchet, while limiting the effectiveness of lateral gene transfer (LGT) at purging deleterious mutations. In principle, an increase in recombination length combined with higher rates of LGT could solve this problem. Here, we show using a computational model that this solution fails in the presence of genetic repeats prevalent in early eukaryotes. The model demonstrates that dispersed repeat sequences allow ectopic recombination, which leads to the loss of genetic information and curtails the capacity of LGT to prevent mutation accumulation. Increasing recombination length in the presence of repeat sequences exacerbates the problem. Mutational decay can only be resisted with homology along extended sequences of DNA. We conclude that the transition to homologous pairing along linear chromosomes was a key innovation in meiotic sex, which was instrumental in the expansion of eukaryotic genomes and morphological complexity.

Argonaute proteins are programmable nucleases that have defense and regulatory functions in both eukaryotes and prokaryotes. All known prokaryotic Argonautes (pAgos) characterized so far act on DNA targets. Here, we describe a new class of pAgos that uniquely use DNA guides to process RNA targets. The biochemical and structural analysis of Pseudooceanicola lipolyticus pAgo (PliAgo) reveals an unusual organization of the guide binding pocket that does not rely on divalent cations and the canonical set of contacts for 5’-end interactions. Unconventional interactions of PliAgo with the 5’-phosphate of guide DNA define its new position within pAgo and shift the site of target RNA cleavage in comparison with known Argonautes. The specificity for RNA over DNA is defined by ribonucleotide residues at the cleavage site. The analysed pAgos sense mismatches and modifications in the RNA target. The results broaden our understanding of prokaryotic defense systems and extend the spectrum of programmable nucleases with potential use in RNA technology.

The origin of eukaryotic cell size and complexity is often thought to have required an energy excess supplied by mitochondria. Recent observations show energy demands to scale continuously with cell volume suggesting that eukaryotes do not have higher energetic capacity. However, respiratory membrane area scales super-linearly with cell surface area. Furthermore, the consequences of the contrasting genomic architectures between prokaryotes and eukaryotes have not been precisely quantified. Here, we investigated (1) factors that affect the volumes at which prokaryotes become surface area-constrained, (2) the amount of energy divested to DNA due to contrasting genomic architectures, and (3) the costs and benefits of respiring symbionts. Our analyses suggest that prokaryotes are not surface area-constrained at volumes of 10(0)–10(3) μm(3), the genomic architecture of extant eukaryotes is only slightly advantageous at genomes sizes of 10(6)–10(7) bp, and a larger host cell may have derived a greater advantage (lower cost) from harboring ATP-producing symbionts. This suggests that eukaryotes first evolved without the need for mitochondria as these ranges hypothetically encompass the Last Eukaryote Common Ancestor and its relatives. Our analyses also show that larger and faster-dividing prokaryotes would have a shortage of respiratory membrane area and divest more energy into DNA. We thus argue that, though mitochondria may not have been required by the first eukaryotes, eukaryote diversification was ultimately dependent upon mitochondria.

Biological processes depend on the differential expression of genes over time, but methods to make physical recordings of these processes are limited. Here we report a molecular system for making time-ordered recordings of transcriptional events into living genomes. We do this via engineered RNA barcodes, based on prokaryotic retrons1, which are reverse-transcribed into DNA and integrated into the genome using the CRISPR-Cas system2. The unidirectional integration of barcodes by CRISPR integrases enables reconstruction of transcriptional event timing based on a physical record via simple, logical rules rather than relying on pre-trained classifiers or post-hoc inferential methods. For disambiguation in the field, we will refer to this system as a Retro-Cascorder.

Background Evolutionary rate is a key characteristic of gene families that is linked to the functional importance of the respective genes as well as specific biological functions of the proteins they encode. Accurate estimation of evolutionary rates is a challenging task that requires precise phylogenetic analysis. Here we present an easy to estimate protein family level measure of sequence variability based on alignment column homogeneity in multiple alignments of protein sequences from Clade-Specific Clusters of Orthologous Genes (csCOGs). Results We report genome-wide estimates of variability for 8 diverse groups of bacteria and archaea and investigate the connection between variability and various genomic and biological features. The variability estimates are based on homogeneity distributions across amino acid sequence alignments and can be obtained for multiple groups of genomes at minimal computational expense. About half of the variance in variability values can be explained by the analyzed features, with the greatest contribution coming from the extent of gene paralogy in the given csCOG. The correlation between variability and paralogy appears to originate, primarily, not from gene duplication, but from acquisition of distant paralogs and xenologs, introducing sequence variants that are more divergent than those that could have evolved in situ during the lifetime of the given group of organisms. Both high-variability and low-variability csCOGs were identified in all functional categories, but as expected, proteins encoded by integrated mobile elements as well as proteins involved in defense functions and cell motility are, on average, more variable than proteins with housekeeping functions. Additionally, using linear discriminant analysis, we found that variability and fraction of genomes carrying a given gene are the two variables that provide the best prediction of gene essentiality as compared to the results of transposon mutagenesis in Sulfolobus islandicus. Conclusions Variability, a measure of sequence diversity within an alignment relative to the overall diversity within a group of organisms, offers a convenient proxy for evolutionary rate estimates and is informative with respect to prediction of functional properties of proteins. In particular, variability is a strong predictor of gene essentiality for the respective organisms and indicative of sub- or neofunctionalization of paralogs.

Prokaryotic genomes are generally gene dense and encode relatively few pseudogenes, or nonfunctional/inactivated remnants of genes. However, in certain contexts, such as recent ecological shifts or extreme population bottlenecks (such as those experienced by symbionts and pathogens), pseudogenes can quickly accumulate and form a substantial fraction of the genome. Identification of pseudogenes is, thus, a critical step for understanding the evolutionary forces acting upon, and the functional potential encoded within, prokaryotic genomes. Here, we present Pseudofinder, an open-source software dedicated to pseudogene identification and analysis. With Pseudofinder’s multi-pronged, reference-based approach, we demonstrate its capacity to detect a wide variety of pseudogenes, including those that are highly degraded and typically missed by gene-calling pipelines, as well newly formed pseudogenes, which can have only one or a few inactivating mutations. Additionally, Pseudofinder can detect intact genes undergoing relaxed selection, which may indicate incipient pseudogene formation. Implementation of Pseudofinder in annotation pipelines will not only clarify the functional potential of sequenced microbes, but will also generate novel insights and hypotheses regarding the evolutionary dynamics of bacterial and archaeal genomes.

The article is devoted to the author's results of the algebraic analysis of molecular genetic systems, including a set of structured DNA alphabets and long nucleotide sequences in single-stranded DNA of eukaryotic and prokaryotic genomes. A connection of the system of DNA n-plets alphabets with principles of algebraic holography is shown, which concerns a popular theme of holography principles in genetically inherited physiology. In addition, a relation between DNA n-plets alphabets and the Poincaré disk model of Lobachevski hyperbolic geometry is revealed. This relation can explain known facts of the relationship of physiological phenomena with hyperbolic geometry. Considering long DNA sequences as a bunch of many parallel texts written in different n-plets alphabets led to the discovery of some universal rules of the stochastic organization of genomic DNAs. These rules are discussed concerning the general problem of the biological dualism "probability-vs-determinism". In general, the presented results give pieces of evidence in favor of the efficiency of a model approach to living organisms as quantum-informational algebraic-harmonic essences.

OBJECTIVE: To express and purify the antigenic peptide of adeno-associated virus (AAV) capsid conserved regions in prokaryotic cells and prepare its rabbit polyclonal antibody. METHODS: The DNA sequence encoding the conserved regions of AAV capsid protein was synthesized and cloned into the vector pET30a to obtain the plasmid pET30a-AAV-CR for prokaryotic expression and purification of the conserved peptides. Coomassie blue staining and Western blotting were used to identify the AAV conserved peptides. Japanese big ear white rabbits were immunized with AAV conserved region protein to prepare polyclonal antibody, with the rabbits injected with PBS as the control group. The antibody titer was determined with ELISA, and the performance of the antibody for recognizing capsid protein sequences of AAV1-AAV10 was assessed with Western blotting and immunofluorescence assay. RESULTS: The plasmid pET30a-AAV-CR was successfully constructed, and a recombinant protein with a relative molecular mass of 17000 was obtained. The purified protein induced the production of antibodies against the conserved regions of AAV capsid in rabbits, and the titer of the purified antibodies reached 1:320 000. The antibodies were capable of recognizing a wide range of capsid protein sequences of AAV1-AAV10. CONCLUSION: We successfully obtained the polyclonal antibodies against AAV capsid conserved region protein from rabbits, which facilitate future studies of AAV vector development and the biological functions of AAV.

Background Prokaryote-virus interactions play key roles in driving biogeochemical cycles. However, little is known about the drivers shaping their interaction network structures, especially from the host features. Here, we compiled 7656 species-level genomes in 39 prokaryotic phyla across environments globally and explored how their interaction specialization is constrained by host life history traits, such as growth rate. Results We first reported that host growth rate indicated by the reverse of minimal doubling time was negatively related to interaction specialization for host in host-provirus network across various ecosystems and taxonomy groups. Such a negative linear growth rate-specialization relationship (GrSR) was dependent on host optimal growth temperature (OGT), and stronger toward the two gradient ends of OGT. For instance, prokaryotic species with an OGT ≥ 40 °C showed a stronger GrSR (Pearson’s r = −0.525, P < 0.001). Significant GrSRs were observed with the presences of host genes in promoting the infection cycle at stages of adsorption, establishment, and viral release, but nonsignificant with the presence of immune systems, such as restriction-modification systems and CRISPR-Cas systems. Moreover, GrSR strength was increased with the presence of temperature-dependent lytic switches, which was also confirmed by mathematical modeling. Conclusions Together, our results advance our understanding of the interactions between prokaryotes and proviruses and highlight the importance of host growth rate in interaction specialization during lysogenization.

Vertical inheritance is foundational to Darwinian evolution, but fails to explain major innovations such as the rapid spread of antibiotic resistance among bacteria and the origin of photosynthesis in eukaryotes. While lateral gene transfer (LGT) is recognized as an evolutionary force in prokaryotes, the role of LGT in eukaryotic evolution is less clear. With the exception of the transfer of genes from organelles to the nucleus, a process termed endosymbiotic gene transfer (EGT), the extent of interdomain transfer from prokaryotes to eukaryotes is highly debated. A common critique of studies of interdomain LGT is the reliance on the topology of single-gene trees that attempt to estimate more than one billion years of evolution. We take a more conservative approach by identifying cases in which a single clade of eukaryotes is found in an otherwise prokaryotic gene tree (i.e. exclusive presence). Starting with a taxon-rich dataset of over 13,600 gene families and passing data through several rounds of curation, we identify and categorize the function of 306 interdomain LGT events into diverse eukaryotes, including 189 putative EGTs, 52 LGTs into Opisthokonta (i.e. animals, fungi and their microbial relatives), and 42 LGTs nearly exclusive to anaerobic eukaryotes. To assess differential gene loss as an explanation for exclusive presence, we compare branch lengths within each LGT tree to a set of vertically-inherited genes subsampled to mimic gene loss (i.e. with the same taxonomic sampling) and consistently find shorter relative distance between eukaryotes and prokaryotes in LGT trees, a pattern inconsistent with gene loss. Our methods provide a framework for future studies of interdomain LGT and move the field closer to an understanding of how best to model the evolutionary history of eukaryotes.

Conjugation drives the horizontal transfer of adaptive traits across prokaryotes. One-fourth of the plasmids encode the functions necessary to conjugate autonomously, the others being eventually mobilizable by conjugation. To understand the evolution of plasmid mobility, we studied plasmid size, gene repertoires, and conjugation-related genes. Plasmid gene repertoires were found to vary rapidly in relation to the evolutionary rate of relaxases, for example, most pairs of plasmids with 95% identical relaxases have fewer than 50% of homologs. Among 249 recent transitions of mobility type, we observed a clear excess of plasmids losing the capacity to conjugate. These transitions are associated with even greater changes in gene repertoires, possibly mediated by transposable elements, including pseudogenization of the conjugation locus, exchange of replicases reducing the problem of incompatibility, and extensive loss of other genes. At the microevolutionary scale of plasmid taxonomy, transitions of mobility type sometimes result in the creation of novel taxonomic units. Interestingly, most transitions from conjugative to mobilizable plasmids seem to be lost in the long term. This suggests a source-sink dynamic, where conjugative plasmids generate nonconjugative plasmids that tend to be poorly adapted and are frequently lost. Still, in some cases, these relaxases seem to have evolved to become efficient at plasmid mobilization in trans, possibly by hijacking multiple conjugative systems. This resulted in specialized relaxases of mobilizable plasmids. In conclusion, the evolution of plasmid mobility is frequent, shapes the patterns of gene flow in bacteria, the dynamics of gene repertoires, and the ecology of plasmids., INCEPTION project (PIA/ANR-16-CONV-0005), Equipe FRM (EQU201903007835), Laboratoire d’Excellence IBEID (ANR-10-LABX-62-IBEID) to E.P.C.R. and C.C. PID2020-117923GB-I00 project funded by Spanish Ministry of Science and Innovation to FdlC and MPG-B.

All living cells depend on the intricate organization of molecular components in space and time. Although this notion was historically based on eukaryotic cells, with their structured intracellular architecture and cellular morphologies, it is now recognized that prokaryotes (that is, bacteria and archaea) also possess complex structures. A cytoskeleton is a network of intracellular protein filaments that play a structural or mechanical role (such as scaffolding, pushing, or pulling) in the spatiotemporal organization of cellular processes. Polymerization of protein monomers in a roughly linear fashion into filaments represents an effective means to establish long-range spatial order by bridging the gap between nanometer-sized molecules and micron-sized cells. It is now evident that bacteria and archaea possess numerous kinds of cytoskeletal proteins, including prokaryotic homologues of the eukaryotic actins, tubulins, and intermediate filaments, as well as other types that have been found primarily or exclusively in prokaryotes (Table 1). Understanding the diverse functions and mechanisms of the rapidly growing universe of prokaryotic cytoskeletal proteins will not only advance prokaryotic cell biology and reveal evolutionary principles, but also open up new avenues for the development of anti-microbial agents, de novo protein design, and the construction of minimal and synthetic cells.

A pangenome is the complete set of genes (core and accessory) present in a phylogenetic clade. We hypothesize that a pangenome's accessory gene content is structured and maintained by selection. To test this hypothesis, we interrogated the genomes of 40 Pseudomonas species for statistically significant coincident (i.e. co-occurring/avoiding) gene patterns. We found that 86.7% of common accessory genes are involved in ≥1 coincident relationship. Further, genes that co-occur and/or avoid each other - but are not vertically inherited - are more likely to share functional categories, are more likely to be simultaneously transcribed, and are more likely to produce interacting proteins, than would be expected by chance. These results are not due to coincident genes being adjacent to one another on the chromosome. Together, these findings suggest that the accessory genome is structured into sets of genes that function together within a given strain. Given the similarity of the Pseudomonas pangenome with open pangenomes of other prokaryotic species, we speculate that these results are generalizable. Keyword: pangenome, microbial genomics, selection.

Summary Prokaryotic laccases are emergent biocatalysts. However, they have not been broadly found and characterized in bacterial organisms, especially in lactic acid bacteria. Recently, a prokaryotic laccase from the lactic acid bacterium Pediococcus acidilactici 5930, which can degrade biogenic amines, was discovered. Thus, our study aimed to shed light on laccases from lactic acid bacteria focusing on two Pediococcus laccases, P. acidilactici 5930 and Pediococcus pentosaceus 4816, which have provided valuable information on their biochemical activities on redox mediators and biogenic amines. Both laccases are able to oxidize canonical substrates as ABTS, ferrocyanide and 2,6‐DMP, and non‐conventional substrates as biogenic amines. With ABTS as a substrate, they prefer an acidic environment and show sigmoidal kinetic activity, and are rather thermostable. Moreover, this study has provided the first structural view of two lactic acid bacteria laccases, revealing new structural features not seen before in other well‐studied laccases, but which seem characteristic for this group of bacteria. We believe that understanding the role of laccases in lactic acid bacteria will have an impact on their biotechnological applications and provide a framework for the development of engineered lactic acid bacteria with enhanced properties., Prokaryotic laccases are emergent biocatalysts. However, they have not been broadly found and characterized in bacterial organisms, especially in lactic acid bacteria. Recently, a prokaryotic laccase from the lactic acid bacterium Pediococcus acidilactici 5930, which can degrade biogenic amines, was discovered. Thus, our study aimed to shed light on laccases from lactic acid bacteria focusing on two Pediococcus laccases, P . acidilactici 5930 and Pediococcus pentosaceus 4816, which have provided valuable information on their biochemical activities on redox mediators and biogenic amines. Both laccases are able to oxidize canonical substrates as ABTS, ferrocyanide and 2,6‐DMP, and non‐conventional substrates as biogenic amines. With ABTS as a substrate, they prefer an acidic environment and show sigmoidal kinetic activity, and are rather thermostable. Moreover, this study has provided the first structural view of two lactic acid bacteria laccases, revealing new structural features not seen before in other well‐studied laccases, but which seem characteristic for this group of bacteria. We believe that understanding the role of laccases in lactic acid bacteria will have an impact on their biotechnological applications and provide a framework for the development of engineered lactic acid bacteria with enhanced properties.

Made available in DSpace on 2021-06-25T15:06:38Z (GMT). No. of bitstreams: 0 Previous issue date: 2021-05-05 Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP) Prokaryotes contribute to the health of marine sponges. However, there is lack of data on the assembly rules of sponge-associated prokaryotic communities, especially for those inhabiting biodiversity hotspots, such as ecoregions between tropical and warm temperate southwestern Atlantic waters. The sympatric species Aplysina caissara, Axinella corrugata, and Dragmacidon reticulatum were collected along with environmental samples from the north coast of SAo Paulo (Brazil). Overall, 64 prokaryotic phyla were detected; 51 were associated with sponge species, and the dominant were Proteobacteria, Bacteria (unclassified), Cyanobacteria, Crenarchaeota, and Chloroflexi. Around 64% and 89% of the unclassified operational taxonomical units (OTUs) associated with Brazilian sponge species showed a sequence similarity below 97%, with sequences in the Silva and NCBI Type Strain databases, respectively, indicating the presence of a large number of unidentified taxa. The prokaryotic communities were species-specific, ranging 56%-80% of the OTUs and distinct from the environmental samples. Fifty-four lineages were responsible for the differences detected among the categories. Functional prediction demonstrated that Ap. caissara was enriched for energy metabolism and biosynthesis of secondary metabolites, whereas D. reticulatum was enhanced for metabolism of terpenoids and polyketides, as well as xenobiotics' biodegradation and metabolism. This survey revealed a high level of novelty associated with Brazilian sponge species and that distinct members responsible from the differences among Brazilian sponge species could be correlated to the predicted functions. Sao Paulo State Univ, Inst Biosci, Coastal Campus Sao Vicente, Sao Paulo, Brazil Univ Estado Rio De Janeiro, Biol Inst Roberto Alcantara Gomes, Dept Genet, Rio De Janeiro, Brazil Univ Sao Paulo, Biosci Inst & NP Biomar, Dept Physiol, Ctr Marine Biol, Sao Paulo, Brazil Sao Paulo State Univ, Inst Biosci, Coastal Campus Sao Vicente, Sao Paulo, Brazil FAPESP: 2016/17189-7 FAPESP: 2017/10157-5 FAPESP: 2018/12937-0 FAPESP: 2017/11380-0

Most prokaryotic cells are encased in a surface layer (S-layer) consisting of a paracrystalline array of repeating lattice-forming proteins. S-layer proteins populate a vast and diverse sequence space, performing disparate functions in prokaryotic cells, including cellular defense, cell-shape maintenance, and regulation of import and export of materials. This article highlights recent advances in the understanding of S-layer structure and assembly, made possible by rapidly evolving structural and cell biology methods. We underscore shared assembly principles revealed by recent work and discuss a common molecular framework that may be used to understand the structural organization of S-layer proteins across bacteria and archaea., Highlights Despite enormous sequence diversity in surface (S)-layer proteins, structural diversity is much lower than previously thought. S-layer proteins have a bipartite arrangement with a lattice-forming and an anchoring segment. Novel structural biology methods are revealing the architectures of S-layers in situ. S-layer assembly across prokaryotes is tightly coupled to the cell cycle, including the cell division machinery.

The average amino acid identity (AAI) is an index of pairwise genomic relatedness, and multiple studies have proposed its application in prokaryotic taxonomy and related disciplines. AAI demonstrates better resolution in elucidating taxonomic structure beyond the species rank when compared with average nucleotide identity (ANI), which is a standard criterion in species delineation. However, an efficient and easy-to-use computational tool for AAI calculation in large-scale taxonomic studies is not yet available. Here, we introduce a bioinformatic pipeline, named EzAAI, which allows for rapid and accurate AAI calculation in prokaryote sequences. The EzAAI tool is based on the MMSeqs2 program and computes AAI values almost identical to those generated by the standard BLAST algorithm with significant improvements in the speed of these evaluations. Our pipeline also provides a function for hierarchical clustering to create dendrograms, which is an essential part of any taxonomic study. EzAAI is available for download as a standalone JAVA program at http://leb.snu.ac.kr/ezaai .

Lateral gene transfer (LGT) occurs in a broad range of prokaryotes and eukaryotes, occasionally promoting adaptation. LGT of functional nuclear genes has been reported among some plants, but systematic studies are needed to assess the frequency and facilitators of LGT. We scanned the genomes of a diverse set of 17 grass species that span more than 50 Ma of divergence and include major crops to identify grass-to-grass protein-coding LGT. We identified LGTs in 13 species, with significant variation in the amount each received. Rhizomatous species acquired statistically more genes, probably because this growth habit boosts opportunities for transfer into the germline. In addition, the amount of LGT increases with phylogenetic relatedness, which might reflect genomic compatibility among close relatives facilitating successful transfers. However, genetic exchanges among highly divergent species indicates that transfers can occur across almost the entire family. Overall, we showed that LGT is a widespread phenomenon in grasses that has moved functional genes across the grass family into domesticated and wild species alike. Successful LGTs appear to increase with both opportunity and compatibility.

The classification of life forms into a hierarchical system (taxonomy) and the application of names to this hierarchy (nomenclature) is at a turning point in microbiology. The unprecedented availability of genome sequences means that a taxonomy can be built upon a comprehensive evolutionary framework, a longstanding goal of taxonomists. However, there is resistance to adopting a single framework to preserve taxonomic freedom, and ever increasing numbers of genomes derived from uncultured prokaryotes threaten to overwhelm current nomenclatural practices, which are based on characterised isolates. The challenge ahead then is to reach a consensus on the taxonomic framework and to adapt and scale the existing nomenclatural code, or create a new code, to systematically incorporate uncultured taxa into the chosen framework.

The advent of comparative genomics in the late 1990s led to the discovery of extensive lateral gene transfer in prokaryotes. The resulting debate over whether life as a whole is best represented as a tree or a network has since given way to a general consensus in which trees and networks co-exist rather than stand in opposition. Embracing this consensus allows us to move beyond the question of which is true or false. The future of the tree of life debate lies in asking what trees and networks can, and should, do for science.

A major question remaining in the field of evolutionary biology is how prokaryotic organisms made the leap to complex eukaryotic life. The prevailing theory depicts the origin of eukaryotic cell complexity as emerging from the symbiosis between an α-proteobacterium, the ancestor of present-day mitochondria, and an archaeal host (endosymbiont theory). A primary contribution of mitochondria to eukaryogenesis has been attributed to the mitochondrial genome, which enabled the successful internalisation of bioenergetic membranes and facilitated remarkable genome expansion. It has also been postulated that a key contribution of the archaeal host during eukaryogenesis was in providing 'archaeal histones' that would enable compaction and regulation of an expanded genome. Yet, how the communication between the host and the symbiont evolved is unclear. Here, we propose an evolutionary concept in which mitochondrial TCA cycle signalling was also a crucial player during eukaryogenesis enabling the dynamic control of an expanded genome via regulation of DNA and histone modifications. Furthermore, we discuss how TCA cycle remodelling is a common evolutionary strategy invoked by eukaryotic organisms to coordinate stress responses and gene expression programmes, with a particular focus on the TCA cycle-derived metabolite itaconate.

Advances in transcriptome sequencing allow for simultaneous interrogation of differentially expressed genes from multiple species originating from a single RNA sample, termed dual or multi-species transcriptomics. Compared to single-species differential expression analysis, the design of multi-species differential expression experiments must account for the relative abundances of each organism of interest within the sample, often requiring enrichment methods and yielding differences in total read counts across samples. The analysis of multi-species transcriptomics datasets requires modifications to the alignment, quantification, and downstream analysis steps compared to the single-species analysis pipelines. We describe best practices for multi-species transcriptomics and differential gene expression.

The vertical flux of marine snow particles significantly reduces atmospheric carbon dioxide concentration. In the mesopelagic zone, a large proportion of the organic carbon carried by sinking particles dissipates thereby escaping long term sequestration. Particle associated prokaryotes are largely responsible for such organic carbon loss. However, links between this important ecosystem flux and ecological processes such as community development of prokaryotes on different particle fractions (sinking vs. non-sinking) are yet virtually unknown. This prevents accurate predictions of mesopelagic organic carbon loss in response to changing ocean dynamics. Using combined measurements of prokaryotic heterotrophic production rates and species richness in the North Atlantic, we reveal that carbon loss rates and associated microbial richness are drastically different with particle fractions. Our results demonstrate a strong negative correlation between prokaryotic carbon losses and species richness. Such a trend may be related to prokaryotes detaching from fast-sinking particles constantly enriching non-sinking associated communities in the mesopelagic zone. Existing global scale data suggest this negative correlation is a widespread feature of mesopelagic microbes.

Genetic studies including the quest, cloning and expression of genes encoding proteins responsible for various vital physiological processes and beneficial characteristics of economic perspective have made the biotechnology research progressively auspicious. Due to its great zootechnical and industrial importance somatotropin gene have been cloned from various animal species. Current study was designed to clone mature ovine growth hormone complementary DNA (oGH cDNA) of a sheep breed, Kajli and carry out over expression studies of cloned GH cDNA in a suitable prokaryotic expression system. Sheep GH cDNA was cloned in T/A (thymine / adenine) vector with signal peptide and confirmed by nested polymerase chain reaction (PCR) and restriction digestion. The gene was then ligated in pLEX expression vector and restricted plasmids showed a fragment insert of ~ 600 bps. Restriction analysis confirmed positive clones, were induced for protein expression analysis. The pET vectors (plasmid for expression by T7 RNA polymerase) have an isopropylthio-β-galactoside (IPTG) inducible strong T7 promoter and Escherichia coli expression strain of BL21 (DE3) pLysS contains DNA fragment from T7 phage which harbors RNA polymerase. Therefore, for expressing recombinant proteins, cells were induced with various IPTG concentrations to optimize expression levels. Cells were induced with different IPTG concentrations (0.1 to 0.8 mM) followed by sodium dodecyl sulphate-polyacrylamide gel electrophoresis (SDS-PAGE). Results indicated maximum expression level of oGH at 5 hrs after induction of cells with 0.3 mM IPTG concentration with a molecular weight of 22 kDa. As for as cellular localization of protein is concerned accumulation of expressed oGH is observed in inclusion bodies. The successful expression of the cloned GH cDNA of sheep confirmed the functional viability of the clone. The above mentioned technique of genetic engineering has provided to boost the dairy industry by the production of large quantities of recombinant bovine somatotropin (rbST).