Genomic GC Research Articles

The dynamics of genome size and GC contents evolution in genus Nicotiana.

Hybridization and Polyploidization are most common of the phenomenon observed in plants, especially in the genus Nicotiana leading to the duplication of genome. Although genomic changes associated with these events has been studied at various levels but the genome size and GC content variation is less understood because of absence of sufficient genomic data. In this study the flow cytometry technique was used to uncover the genome size and GC contents of 46 Nicotiana species and we compared the genomic changes associated with the hybridization events along evolutionary time scale. The genome size among Nicotiana species varied between 3.28 pg and 11.88 pg whereas GC contents varied between 37.22% and 51.25%. The tetraploid species in genus Nicotiana including section Polydiclae, Repandae, Nicotiana, Rustica and Sauveolentes revealed both up and downsizing in their genome sizes when compared to the sum of genomes of their ancestral species. The genome sizes of three homoploid hybrids were found near their ancestral species. Loss of large genome sequence was observed in the evolutionary more aged species (>10 Myr) as compared to the recently evolved one's (<0.2 Myr). The GC contents were found homogenous with a mean difference of 2.46% among the Nicotiana species. It is concluded that genome size change appeared in either direction whereas the GC contents were found more homogenous in genus Nicotiana.

Evolution of genome size and GC content in the tribe Carduinae (Asteraceae)

Genome size and GC content are basic species-specific attributes often delimiting genera or higher taxa, which enable the identification of polyploidy, hybridization and other modes of genome or karyotype evolution. The evolution of these genomic traits can often occur as a result of the selective pressure of the environment. Here, we reconstruct the evolution of these genomic traits in subtribe Carduinae (Asteraceae) in the context of changes in chromosome numbers. Using flow cytometry, genome size and GC content were estimated for 119 taxa and mapped onto a phylogenetic tree constructed using sequences from seven genetic markers. In addition, the genomic data were compared with the length of stomatal guard cells and achene size (length, weight) to evaluate the extent to which genomic characters could evolve adaptively in this subtribe. We found strong phylogenetic signals for the analysed genomic and phenotypic traits, which delimited most Carduinae genera or clades in agreement with the reconstructed phylogeny. Monoploid genome size was positively correlated with genomic GC content and stomatal guard cell length. In Cirsium, whose species were the focus of the majority of the analyses, the large-genomed subgen. Lophiolepis had smaller guard cells, which might be related to it occurring in more xeric habitats compared to subgen. Cirsium. In contrast, the achenes of the large-genomed subgen. Lophiolepis were larger, possibly in response to the summer drought, whereas achene weight and length were independent of genome size across the subtribe. Thus, genome size in the subtribe Carduinae might evolve under weak environmental control, at least under that mediated by the size of guard cells or achenes. Achene size was related positively to GC content, which could have evolved adaptively in response to summer drought. In Carduus and the North American Cirsium taxa, there is an increase in average chromosome size with reduction in monoploid chromosome number, suggesting descending dysploidy associated with chromosomal fusion. Polyploidy is relatively rare in this subtribe and was confirmed only in five of the species studied, including Cirsium vulgare, an invasive species that likely originated via distant (intersubgeneric) hybridization, as suggested by its intermediate genomic and achene features combined with the conflict between its morphology and phylogenetic position. Phylogenetic reconstruction, differences in genomic parameters, as well as stomatal guard cell and achene sizes support the separation of a monophyletic Lophiolepis from the remainder of Cirsium. Genome and achene size results also indicate that the early diverging Cirsium italicum can be separated from the rest of the monophyletic Lophiolepis+Picnomon+Notobasis clade.

Disentangling the taxonomic structure of the Allium paniculatum species complex in central and eastern Europe using molecular, cytogenetic and morphological tools

Allium section Codonoprasum includes a group of evolutionarily young taxa with unclear taxonomic boundaries, evolutionary relationships and complex synonymy. The most complicated taxon in this section is A. paniculatum, which until recently was considered to be the most widespread and morphologically most variable species, with a distribution extending from the Mediterranean area to central Europe and the Pontic region. A recent taxonomic study has shown that true A. paniculatum is a morphologically and genetically distinct species occurring in southern Ukraine and southern Russia. The clarification of its taxonomic identity questioned the taxonomic identity of populations referred to as A. paniculatum in other parts of Europe, including populations in central (Slovakia, Hungary, northeast Serbia) and the western part of eastern Europe (Romania, Ukraine), from where four other species of this complex (i.e. A. fuscum, A. fussii, A. marginatum and A. podolicum) were described and later frequently merged as synonyms with A. paniculatum. Here, the diversification within this complex in the abovementioned region is explored, using various biosystematic methods (morphometry, leaf anatomy and epidermal micromorphometry, karyology, estimation of genome size and GC content using flow cytometry, and molecular fingerprinting using Amplified Fragment Length Polymorphisms). By this means the existence of three well-separated population groups were revealed in the material studied, taxonomically corresponding to A. fuscum, A. marginatum and the group A. paniculatum + A. podolicum. The genetic patterns indicate that populations of at least some of these taxa might have survived in their current ranges or in neighbouring areas during the last glacial and the early Holocene. Allium fuscum is a species that inhabits shaded rocky outcrops in the wide neighbourhood of the Iron Gate area (south-western Romania, north-western Bulgaria and northeast Serbia). Allium marginatum is a species closely related to A. fuscum, inhabiting dry grasslands and dry oak forests in the Pannonian and Transylvanian Basins. Two eastern-European taxa, A. podolicum and A. paniculatum,which are very similar inmorphology, genome size and genetic markers, are considered to be conspecific and named A. paniculatum. This species inhabits zonal steppe and forest steppe regions in eastern Europe from western Ukraine to southern Russia. In addition, a taxonomic and nomenclatural treatment, as well as a key for identifying the recognized species of the A. paniculatum complex in the region studied, are presented.

Edaphic controls on genome size and GC content of bacteria in soil microbial communities

Nutrient limitation has been shown to reduce bacterial genome size and influence nucleotide composition; however, much of this work has been conducted in marine systems and the factors which shape soil bacterial genomic traits remain largely unknown. Here we determined average genome size, GC content, codon usage, and amino acid content from 398 soil metagenomes across a broad geographic range and used machine-learning to determine the environmental parameters that most strongly explain the distribution of these traits. We found that genomic trait averages were most related to pH, which we suggest is primarily due to the correlation of pH with several environmental parameters, particularly soil carbon content. Low pH soils had higher carbon to nitrogen ratios (C:N) and tended to have communities with lower GC content and larger genomes, potentially a response to increased physiological stress and a requirement for metabolic diversity. Conversely, communities in high pH and low soil C:N had smaller genomes and higher GC content—indicating potential resource driven selection against AT base pairs, which have a higher C:N than GC base pairs. Similarly, we found that nutrient conservation also applied to amino acid stoichiometry, where bacteria in soils with low C:N ratios tended to code for amino acids with lower C:N. Together, these relationships point towards fundamental mechanisms that underpin genome size, and nucleotide and amino acid selection in soil bacteria.

Genomic Legacies of Ancient Adaptation Illuminate GC-Content Evolution in Bacteria.

Bacterial evolution is characterized by strong purifying selection as well as rapid adaptive evolution in changing environments. In this context, the genomic GC content (genomic GC) varies greatly but presents some level of phylogenetic stability, making it challenging to explain based on current hypotheses. To illuminate the evolutionary mechanisms of the genomic GC, we analyzed the base composition and functional inventory of 11,083 representative genomes. A phylogenetically constrained bimodal distribution of the genomic GC, which mainly originated from parallel divergences in the early evolution, was demonstrated. Such variation of the genomic GC can be well explained by DNA replication and repair (DRR), in which multiple pathways correlate with the genomic GC. Furthermore, the biased conservation of various stress-related genes, especially the DRR-related ones, implies distinct adaptive processes in the ancestral lineages of high- or low-GC clades which are likely induced by major environmental changes. Our findings support that the mutational biases resulting from these legacies of ancient adaptation have changed the course of adaptive evolution and generated great variation in the genomic GC. This highlights the importance of indirect effects of natural selection, which indicates a new model for bacterial evolution. IMPORTANCE GC content has been shown to be an important factor in microbial ecology and evolution, and the genomic GC of bacteria can be characterized by great intergenomic heterogeneity, high intragenomic homogeneity, and strong phylogenetic inertia, as well as being associated with the environment. Current hypotheses concerning direct selection or mutational biases cannot well explain these features simultaneously. Our findings of the genomic GC showing that ancient adaptations have transformed the DRR system and that the resulting mutational biases further contributed to a bimodal distribution of it offer a more reasonable scenario for the mechanism. This would imply that, when thinking about the evolution of life, diverse processes of adaptation exist, and combined effects of natural selection should be considered.

Comparative genomics and pangenomics of vancomycin-resistant and susceptible Enterococcus faecium from Irish hospitals.

Introduction. Enterococcus faecium has emerged as an important nosocomial pathogen, which is increasingly difficult to treat due to the genetic acquisition of vancomycin resistance. Ireland has a recalcitrant vancomycin-resistant bloodstream infection rate compared to other developed countries.Hypothesis/Gap statement. Vancomycin resistance rates persist amongst E. faecium isolates from Irish hospitals. The evolutionary genomics governing these trends have not been fully elucidated.Methodology. A set of 28 vancomycin-resistant isolates was sequenced to construct a dataset alongside 61 other publicly available Irish genomes. This dataset was extensively analysed using in silico methodologies (comparative genomics, pangenomics, phylogenetics, genotypics and comparative functional analyses) to uncover distinct evolutionary, coevolutionary and clinically relevant population trends.Results. These results suggest that a stable (in terms of genome size, GC% and number of genes), yet genetically diverse population (in terms of gene content) of E. faecium persists in Ireland with acquired resistance arising via plasmid acquisition (vanA) or, to a lesser extent, chromosomal recombination (vanB). Population analysis revealed five clusters with one cluster partitioned into four clades which transcend isolation dates. Pangenomic and recombination analyses revealed an open (whole genome and chromosomal specific) pangenome illustrating a rampant evolutionary pattern. Comparative resistomics and virulomics uncovered distinct chromosomal and mobilomal propensity for multidrug resistance, widespread chromosomal point-mutation-mediated resistance and chromosomally harboured arsenals of virulence factors. Interestingly, a potential difference in biofilm formation strategies was highlighted by coevolutionary analysis, suggesting differential biofilm genotypes between vanA and vanB isolates.Conclusions. These results highlight the evolutionary history of Irish E. faecium isolates and may provide insight into underlying infection dynamics in a clinical setting. Due to the apparent ease of vancomycin resistance acquisition over time, susceptible E. faecium should be concurrently reduced in Irish hospitals to mitigate potential resistant infections.

A simple stochastic model describing the evolution of genomic GC content in asexually reproducing organisms

A genome’s nucleotide composition can usually be summarized with (G)uanine + (C)ytosine (GC) or (A)denine + (T)hymine (AT) frequencies as GC% = 100% − AT%. Genomic AT/GC content has been linked to environment and selective processes in asexually reproducing organisms. A model is presented relating the evolution of genomic GC content over time to AT rightarrow GC and GC rightarrow AT mutation rates. By employing Itô calculus it is shown that if mutation rates are subject to random perturbations, that can vary over time, several implications follow. In particular, an extra Brownian motion term appears influencing genomic nucleotide variability; the greater the random perturbations the more genomic nucleotide variability. This can have several interpretations depending on the context. For instance, reducing the influence of the random perturbations on the AT/GC mutation rates and thus genomic nucleotide variability, to limit fitness decreasing and deleterious mutations, will likely suggest channeling of resources. On the other hand, increased genomic nucleotide diversity may be beneficial in variable environments. In asexually reproducing organisms, the Brownian motion term can be considered to be inversely reflective of the selective pressures an organism is subjected to at the molecular level. The presented model is a generalization of a previous model, limited to microbial symbionts, to all asexually reproducing, non-recombining organisms. Last, a connection between the presented model and the classical Luria–Delbrück mutation model is presented in an Itô calculus setting.

The complete chloroplast genome of two Firmiana species and comparative analysis with other related species.

Firmiana is a small genus within the subfamily Sterculioideae of the Malvaceae. There are nine Firmiana species distributed in South and South-west China, most of which are endangered. Due to the shortage of plastid genomes data, the phylogenetic relationships and the evolutionary history of this genus remain unclear. Therefore, the complete chloroplast genomes of F. calcarean and F. hainanensis were sequenced using high-throughput sequencing and then compared with the chloroplast genomes of other reported Firmiana species. The genome size of F. calcarean and F. hainanensis is 161,263 and 160,031bp long, respectively, containing a total of 131 genes (including 85 protein coding genes, 37 tRNAs, 8 rRNAs, and one pseudogene). Comparative analysis revealed that the genome structure, GC content, gene content and order, as well as the RNA editing sites within the chloroplast genomes of F. calcarean and F. hainanensis were similar to previously reported Firmiana species. ML phylogenetic analysis revealed that F. danxiaensis, F. hainanensis, F. calcarean, F. simplex, and F. major form a sister group to F. colorata, F. pulcherrima, and F. kwangsiensis. The SSRs, long repeats, and 21 highly divergent regions (Pi > 0.01) identified in this study might provide potential DNA markers for further population genetics and phylogenetic studies of Firmiana. Our findings can help design new species-specific molecular markers and the general framework to further explore the evolutionary history of Firmiana and to address their conservation challenges.

A discussion of RNA virus taxonomy based on the 2020 International Committee on Taxonomy of Viruses report.

RNA viruses have a higher mutation rate than DNA viruses; however, RNA viruses are insufficiently studied outside disease settings. The International Committee on Taxonomy of Viruses (ICTV) is an organization set up by virologists to standardize virus classification. To better understand ICTV taxonomy and the characteristics and rules of different RNA virus families, we analyzed the 3,529 RNA viruses included in the 2020 ICTV report using five widely used metrics: length, host, GC content, number of predicted ORFs, and sequence similarity. The results show that host type has a significant influence on viral genome length and GC content. The genome lengths of virus members within the same genus are quite similar: 98.28% of the genome length differences within any particular genus are less than 20%. The species within those genera containing segmented viruses also have a similar length and number of segments. The number of predicted ORFs in the RNA viral genomes also shows a strong, statistically significant correlation with genome length. We suggest that due to the high mutation rate of RNA virus genomes, current RNA virus classification should mainly rely on protein similarities rather than nucleic acid similarities.

The chloroplast genome of Spiraea thunbergii (Rosaceae)

Spiraea thunbergii (S. thunbergia) is a very common ornamental shrub in the garden, with important horticultural and economic value. In this study, we assembled the complete chloroplast (cp) genome of S. thunbergia into a typical quadripartite structure. The genome size and GC content of the S. thunbergia cp genome are 155,922 bp and 36.76%, respectively. It contains 84 protein-coding genes, 37 tRNA genes, and 8 rRNA genes. The phylogenetic tree supported that S. thunbergii is closely related to Spiraea mongolica in the Rosaceae family. The study will provide significant genomic resource for elucidating the phylogenetic relationship of Spiraea.

Comparative genomic analysis reveals the origin and global distribution of melon necrotic virus isolates

The melon necrotic spot virus (MNSV) is the family member of Tombusviridae and genus member of Gammacarmovirus. It can be widely found from almost everywhere across the globe, infecting host plants like rockmelon, cucumber, muskmelon and watermelon. In this study, we attempted to search for country-specific genome markers of MNSV via the comparison of several genome characteristics across isolates from different countries such as South Korea, China, India, Japan, Netherlands, Canada, Australia, Brazil, USA and Spain. As a result, we discovered that the whole genome length and GC content can be used to preliminarily distinguish South Korea, Japan and Australia MNSV isolates. Besides, single nucleotide polymorphism (SNP) markers and protein profiles (RNA-dependent RNA polymerase (RdRp) and coat protein (CP)) can further differentiate Japan and South Korea MNSV isolates and identify two Japan isolates that are postulated to have South Korea origin. The motif and phylogenetic analysis revealed the previously described divergent group from USA and Australia regions. Interestingly, the RdRp and CP protein models for all isolates are identical and this is a strong indication of the high MNSV functional conservation. In short, the information yielded from this study will in turn aid in future virulence-SNP association studies essential for cost-effective biological control of this virus via genetic engineering in order to reduce harvest loss.

Comparative Analysis of Chloroplast Genomes within Saxifraga (Saxifragaceae) Takes Insights into Their Genomic Evolution and Adaption to the High-Elevation Environment

Saxifraga species are widely distributed in alpine and arctic regions in the Northern hemisphere. Highly morphological diversity within this genus brings great difficulties for species identification, and their typical highland living properties make it interesting how they adapt to the extreme environment. Here, we newly generated the chloroplast (cp) genomes of two Saxifraga species and compared them with another five Saxifraga cp genomes to understand the characteristics of cp genomes and their potential roles in highland adaptation. The genome size, structure, gene content, GC content, and codon usage pattern were found to be highly similar. Cp genomes ranged from 146,549 bp to 151,066 bp in length, most of which comprised 130 predicted genes. Yet, due to the expansion of IR regions, the second copy of rps19 in Saxifraga stolonifera was uniquely kept. Through sequence divergence analysis, we identified seven hypervariable regions and detected some signatures of regularity associated with genetic distance. We also identified 52 to 89 SSRs and some long repeats among seven Saxifraga species. Both ML and BI phylogenetic analyses confirmed that seven Saxifraga species formed a monophyletic clade in the Saxifragaceae family, and their intragenus relationship was also well supported. Additionally, the ndhI and ycf1 genes were considered under positive selection in species inhabiting relatively high altitudes. Given the conditions of intense light and low CO2 concentration in the highland, the products of these two genes might participate in the adaptation to the extreme environment.

Geoalkalibacter halelectricus SAP-1 sp. nov. possessing extracellular electron transfer and mineral-reducing capabilities from a haloalkaline environment.

The extracellular electron transfer (EET)-capable electroactive microorganisms (EAMs) play crucial roles in mineral cycling and interspecies electron transfer in different environments and are used as biocatalysts in microbial electrochemical technologies. Studying EAMs from extreme environments is desired to advance the electromicrobiology discipline, understanding their unique metabolic traits with implications to extreme microbiology, and develop specific bioelectrochemical applications. Here, we present a novel haloalkaliphilic bacterium named Geoalkalibacter halelectricus SAP-1, isolated from a microbial electroactive biofilm enriched from the haloalkaline lake sediments. It is a rod-shaped Gram-negative heterotrophic anaerobe that uses various carbon and energy sources and respires on soluble and insoluble terminal electron acceptors. Besides 16S-rRNA and whole-genome sequence-based phylogeny, the GGDC values of 21.7%, ANI of 78.5%, and 2.77% genomic DNA GC content difference with the closest validly named species Geoalkalibacter ferrihydriticus (DSM 17813T ) confirmed its novelty. When grown with the solid-state electrode as the only electron acceptor, it produced 460 ± 23 μA/cm2 bioelectrocatalytic current, thereby confirming its electroactivity. Further electrochemical analysis revealed the presence of membrane redox components with a high formal potential, putatively involved in the direct mode of EET. These are distinct from EET components reported for any known electroactive microorganisms, including well-studied Geobacter spp., Shewanella spp., and Desulfuromonas acetexigens. The capabilities of G. halelectricus SAP-1 to respire on soluble and insoluble electron acceptors including fumarate, SO4 2- , Fe3+ , and Mn4+ suggests its role in cycling these elements in haloalkaline environments.

Comparative analysis of two genomes of Chlamydia pecorum isolates from an Alpine chamois and a water buffalo

BackgroundTo date, whole genome sequencing has been performed mainly for isolates of Chlamydia trachomatis, C. pneumoniae, C. psittaci and C. abortus, but only a few isolates of C. pecorum have been entirely sequenced and this makes it difficult to understand its diversity and population structure. In this study the genome of two C. pecorum strains isolated from the lung of an Alpine chamois affected with pneumonia (isolate PV7855) and the brain of a water buffalo affected with meningoencephalomyelitis (isolate PV6959), were completely sequenced with MiSeq system (Illumina) and analyzed in their most polymorphic regions.ResultsThe genome length and GC content of the two isolates were found to be consistent with other C. pecorum isolates and the gene content of polymorphic membrane proteins and plasticity zone was found to be very similar. Some differences were observed in the phospholipase genes for both isolates and in the number of genes in the plasticity zone, such as the presence of some hypothetical proteins in PV6959, not present in any other genomes analyzed in this study. Interestingly, PV6959 possesses an extra pmp and has an incomplete tryptophan biosynthesis operon. Plasmids were detected in both isolates.ConclusionsGenome sequencing of the two C. pecorum strains did not reveal differences in length and GC content despite the origin from different animal species with different clinical disease. In the plasticity zone, the differences in the genes pattern might be related to the onset of specific symptoms or infection of specific hosts. The absence of a tryptophan biosynthesis pathway in PV6959 may suggest a strict relationship between C. pecorum and its host.

Whole genome assembly of the armored loricariid catfish Ancistrus triradiatus highlights herbivory signatures

The catfish Ancistrus triradiatus belongs to the species-rich family Loricariidae. Loricariids display remarkable traits such as herbivory, a benthic lifestyle, the absence of scales but the presence of dermal bony plates. They are exported as ornamental fish worldwide, with escaped fishes becoming a threat locally. Although genetic and phylogenetic studies are continuously increasing and developmental genetic investigations are underway, no genome assembly has been formally proposed for Loricariidae yet. We report a high-quality genome assembly of Ancistrus triradiatus using long and short reads, and a newly assembled transcriptome. The genome assembly is composed of 9530 scaffolds, including 85.6% of ray-finned fish BUSCOs, and 26,885 predicted protein-coding genes. The genomic GC content is higher than in other catfishes, reflecting the higher metabolism associated with herbivory. The examination of the SCPP gene family indicates that the genes presumably triggering scale loss when absent, are present in the scaleless A. triradiatus, questioning their explanatory role. The analysis of the opsin gene repertoire revealed that gene losses associated to the nocturnal lifestyle of catfishes were not entirely found in A. triradiatus, as the UV-sensitive opsin 5 is present. Finally, most gene family expansions were related to immunity except the gamma crystallin gene family which controls pupil shape and sub-aquatic vision. Thus, the genome of A. triradiatus reveals that fish herbivory may be related to the photic zone habitat, conditions metabolism, photoreception and visual functions. This genome is the first for the catfish suborder Loricarioidei and will serve as backbone for future genetic, developmental and conservation studies.

Comparative analyses of chloroplast genomes from Six Rhodiola species: variable DNA markers identification and phylogenetic relationships within the genus

BackgroundAs a valuable medicinal plant, Rhodiola has a very long history of folk medicine used as an important adaptogen, tonic, and hemostatic. However, our knowledge of the chloroplast genome level of Rhodiola is limited. This drawback has limited studies on the identification, evolution, genetic diversity and other relevant studies on Rhodiola.ResultsSix Rhodiola complete chloroplast genomes were determined and compared to another Rhodiola cp genome at the genome scale. The results revealed a cp genome with a typical quadripartite and circular structure that ranged in size from 150,771 to 151,891 base pairs. High similarity of genome organization, gene number, gene order, and GC content were found among the chloroplast genomes of Rhodiola. 186 (R. wallichiana) to 200 (R. gelida) SSRs and 144 pairs of repeats were detected in the 6 Rhodiola cp genomes. Thirteen mutational hotspots for genome divergence were determined and could be used as candidate markers for phylogenetic analyses and Rhodiola species identification. The phylogenetic relationships inferred by members of Rhodiola cluster into two clades: dioecious and hermaphrodite. Our findings are helpful for understanding Rhodiola's taxonomic, phylogenetic, and evolutionary relationships.ConclusionsComparative analysis of chloroplast genomes of Rhodiola facilitates medicinal resource conservation, phylogenetic reconstruction and biogeographical research of Rhodiola.

Comprehensive Phylogenomics of Methylobacterium Reveals Four Evolutionary Distinct Groups and Underappreciated Phyllosphere Diversity.

Methylobacterium is a group of methylotrophic microbes associated with soil, fresh water, and particularly the phyllosphere, the aerial part of plants that has been well studied in terms of physiology but whose evolutionary history and taxonomy are unclear. Recent work has suggested that Methylobacterium is much more diverse than thought previously, questioning its status as an ecologically and phylogenetically coherent taxonomic genus. However, taxonomic and evolutionary studies of Methylobacterium have mostly been restricted to model species, often isolated from habitats other than the phyllosphere and have yet to utilize comprehensive phylogenomic methods to examine gene trees, gene content, or synteny. By analyzing 189 Methylobacterium genomes from a wide range of habitats, including the phyllosphere, we inferred a robust phylogenetic tree while explicitly accounting for the impact of horizontal gene transfer (HGT). We showed that Methylobacterium contains four evolutionarily distinct groups of bacteria (namely A, B, C, D), characterized by different genome size, GC content, gene content, and genome architecture, revealing the dynamic nature of Methylobacterium genomes. In addition to recovering 59 described species, we identified 45 candidate species, mostly phyllosphere-associated, stressing the significance of plants as a reservoir of Methylobacterium diversity. We inferred an ancient transition from a free-living lifestyle to association with plant roots in Methylobacteriaceae ancestor, followed by phyllosphere association of three of the major groups (A, B, D), whose early branching in Methylobacterium history has been heavily obscured by HGT. Together, our work lays the foundations for a thorough redefinition of Methylobacterium taxonomy, beginning with the abandonment of Methylorubrum.

RNA-seq Sample Preparation Kits Strongly Affect Transcriptome Profiles of a Gas-Fermenting Bacterium.

ABSTRACTTranscriptome analysis via RNA sequencing (RNA-seq) has become a standard technique employed across various biological fields of study. The rapid adoption of the RNA-seq approach has been mediated, in part, by the development of different commercial RNA-seq library preparation kits compatible with standard next-generation sequencing (NGS) platforms. Generally, the essential steps of library preparation, such as rRNA depletion and first-strand cDNA synthesis, are tailored to a specific group of organisms (e.g., eukaryotes versus prokaryotes) or genomic GC content. Therefore, the selection of appropriate commercial products is of crucial importance to capture the transcriptome of interest as closely to the native state as possible without introduction of technical bias. However, researchers rarely have the resources and time to test various commercial RNA-seq kits for their samples. This work reports a side-by-side comparison of RNA-seq data from Clostridium autoethanogenum obtained using three commercial rRNA removal and strand-specific library construction products of NuGEN Technologies, Qiagen, and Zymo Research and assesses their performance relative to published data. While all three vendors advertise their products as suitable for prokaryotes, we found significant differences in their performance regarding rRNA removal, strand specificity, and most importantly, transcript abundance distribution profiles. Notably, RNA-seq data obtained with Qiagen products were most similar to published data and delivered the best results in terms of library strandedness and transcript abundance distribution range. Our results highlight the importance of finding appropriate organism-specific workflows and library preparation products for RNA-seq studies.IMPORTANCE RNA-seq is a powerful technique for transcriptome profiling while involving elaborate sample processing before library sequencing. We show that RNA-seq library preparation kits can strongly affect the outcome of an RNA-seq experiment. Although library preparation benefits from the availability of various commercial kits, choosing appropriate products for the specific samples can be challenging for new users or for users working with unconventional organisms. Evaluating the performance of different commercial products requires significant financial and time investments infeasible for most researchers. Therefore, users are often guided in their choice of kits by published data involving similar input samples. We conclude that important consideration should be given to selecting sample processing workflows for any given organism.

Diverse Genomic Traits Differentiate Sinking-Particle-Associated versus Free-Living Microbes throughout the Oligotrophic Open Ocean Water Column.

ABSTRACTBacteria and archaea are central to the production, consumption, and remineralization of dissolved and particulate organic matter and contribute critically to carbon delivery, nutrient availability, and energy transformations in the deep ocean. To explore environmentally relevant genomic traits of sinking-particle-associated versus free-living microbes, we compared habitat-specific metagenome-assembled genomes recovered throughout the water column in the North Pacific Subtropical Gyre. The genomic traits of sinking-particle-associated versus free-living prokaryotes were compositionally, functionally, and phylogenetically distinct. Substrate-specific transporters and extracellular peptidases and carbohydrate-active enzymes were more enriched and diverse in particle-associated microbes at all depths than in free-living counterparts. These data indicate specific roles for particle-attached microbes in particle substrate hydrolysis, uptake, and remineralization. Shallow-water particle-associated microbes had elevated genomic GC content and proteome nitrogen content and reduced proteome carbon content in comparison to abyssal particle-associated microbes. An inverse trend was observed for their sympatric free-living counterparts. These different properties of attached microbes are postulated to arise in part due to elevated organic and inorganic nitrogen availability inside sinking particles. Particle-attached microbes also were enriched in genes for environmental sensing via two-component regulatory systems, and cell-cell interactions via extracellular secretion systems, reflecting their surface-adapted lifestyles. Finally, particle-attached bacteria had greater predicted maximal growth efficiencies than free-living bacterioplankton at all depths. All of these particle-associated specific genomic and proteomic features appear to be driven by microhabitat-specific elevated nutrient and energy availability as well as surface-associated competitive and synergistic ecological interactions. Although some of these characteristics have been previously postulated or observed individually, we report them together here in aggregate via direct comparisons of cooccurring free-living and sinking-particle-attached microbial genomes from the open ocean.

Phenotype-genotype analysis of Latilactobacills curvatus from different niches: Carbohydrate metabolism, antibiotic resistance, bacteriocin, phage fragments and linkages with CRISPR-Cas systems

The potential probiotic function of Latilactobacills curvatus has attracted the attention of researchers. To explore the differences in the genomes of L. curvatus, nine strains were isolated from various sources, including feces and fermented vegetables and compared with 25 strains from the NCBI database. The findings indicated that the average genome size, GC content, and CDS of L. curvatus were 1.94 MB, 41.9%, and 1825, respectively. Its core genome is associated with transcription, translation, carbohydrate transport and metabolism, and defense functions. The pan-genome of L. curvatus was in a closed state. The genetic diversity of L. curatus is mainly manifested in its ability to use carbohydrates, antibiotic resistance, bacteriocin operon, and polymeric regularly interspaced short palindromic repeats (CRISPR)-Cas for bacterial immunity. The CRISPR system of 34 strains of L. curvatus was predominantly found to be of the IIA type with a few IIC and IE types. These findings will contribute to a better understanding of this species.