Variation In Gene Content Research Articles

Evolution of the pheV-tRNA integrated genomic island in Escherichia coli.

Escherichia coli exhibit extensive genetic diversity at the genome level, particularly within their accessory genome. The tRNA integrated genomic islands (GIs), a part of the E. coli accessory genome, play an important role in pathogenicity. However, studies examining the evolution of GIs have been challenging due to their large size, considerable gene content variation and fragmented assembly in draft genomes. Here we examined the evolution of the GI integrated at pheV-tRNA (GI-pheV), with a primary focus on uropathogenic E. coli (UPEC) and the globally disseminated multidrug resistant ST131 clone. We show the gene content of GI-pheV is highly diverse and arranged in a modular configuration, with the P4 integrase encoding gene intP4 the only conserved gene. Despite this diversity, the GI-pheV gene content displayed conserved features among strains from the same pathotype. In ST131, GI-pheV corresponding to the reference strain EC958 (EC958_GI-pheV) was found in ~90% of strains. Phylogenetic analyses suggested that GI-pheV in ST131 has evolved together with the core genome, with the loss/gain of specific modules (or the entire GI) linked to strain specific events. Overall, we show GI-pheV exhibits a dynamic evolutionary pathway, in which modules and genes have evolved through multiple events including insertions, deletions and recombination.

Giant transposons promote strain heterogeneity in a major fungal pathogen.

No "one size fits all" option exists for treating fungal infections in large part due to genetic and phenotypic variation among strains. Accounting for strain heterogeneity is thus fundamental for developing efficacious treatments and strategies for safeguarding human health. Here, we report significant progress towards achieving this goal by uncovering a previously hidden mechanism generating heterogeneity in the major human fungal pathogen Aspergillus fumigatus : giant transposons called Starships that span dozens of kilobases and mobilize fungal genes as cargo. By conducting the first systematic investigation of these unusual transposons in a single fungal species, we demonstrate their contributions to population-level variation at the genome, pangenome and transcriptome levels. The Starship atlas we developed will not only help account for variation introduced by these elements in laboratory experiments but will serve as a foundational resource for determining how Starships shape clinically-relevant phenotypes, such as antifungal resistance and pathogenicity.

Koliella bifissiva sp. nov (Chlorellaceae, Chlorophyta) and Analysis of Its Organelle Genomes.

Chlorellacean members are common in aquatic or subaerial habitats, and many of them have significant economic value. Taxonomic reports and organelle genome data for the Nannochloris clade, an important subgroup within this family, are limited, hindering the understanding and exploitation of this clade. In this study, a fusiform-celled strain, FACHB-3607, was isolated from a pond in China. Through examination of morphological characteristics and phylogenetic analyses of rbcL, 18S rDNA, and ITS, it was identified as a new species within the Nannochloris clade, named Koliella bifissiva sp. nov. In addition, this study provided a first insight into the organellar genomes of the genus Koliella. The K. bifissiva chloroplast had a 99.8 kb genome, and the mitochondrion had a 40.8 kb genome, which are moderate sizes within the Nannochloris clade. Phylogenomic analysis showed that K. bifissiva is most closely related to Nannochloris sp. "desiccata", followed by Marvania. In contrast, Picochlorum was the most distantly related species. The organelle genomes of the Nannochloris clade display dynamic evolution, reflected in variations in genome size, gene content and order, and selection pressure. This research enhances our knowledge of species diversity and evolutionary history in the Nannochloris clade.

Host specificity and cophylogeny in the “animal-gut bacteria-phage” tripartite system

Cophylogeny has been identified between gut bacteria and their animal host and is highly relevant to host health, but little research has extended to gut bacteriophages. Here we use bee model to investigate host specificity and cophylogeny in the “animal-gut bacteria-phage” tripartite system. Through metagenomic sequencing upon different bee species, the gut phageome revealed a more variable composition than the gut bacteriome. Nevertheless, the bacteriome and the phageome showed a significant association of their dissimilarity matrices, indicating a reciprocal interaction between the two kinds of communities. Most of the gut phages were host generalist at the viral cluster level but host specialist at the viral OTU level. While the dominant gut bacteria Gilliamella and Snodgrassella exhibited matched phylogeny with bee hosts, most of their phages showed a diminished level of cophylogeny. The evolutionary rates of the bee, the gut bacteria and the gut phages showed a remarkably increasing trend, including synonymous and non-synonymous substitution and gene content variation. For all of the three codiversified tripartite members, however, their genes under positive selection and genes involving gain/loss during evolution simultaneously enriched the functions into metabolism of nutrients, therefore highlighting the tripartite coevolution that results in an enhanced ecological fitness for the whole holobiont.

Pangenome evaluation of gene essentiality in Streptococcus pyogenes.

Bacterial species often consist of strains with variable gene content, collectively referred to as the pangenome. Variations in the genetic makeup of strains can alter bacterial physiology and fitness. To define biologically relevant genes of a genome, genome-wide transposon mutant libraries have been used to identify genes essential for survival or virulence in a given strain. Such phenotypic studies have been conducted in four different genotypes of the human pathogen Streptococcus pyogenes, yet challenges exist in comparing results across studies conducted in different genetic backgrounds and conditions. To advance genotype to phenotype inferences across different S. pyogenes strains, we built a pangenome database of 249 S. pyogenes reference genomes. We systematically re-analyzed publicly available transposon sequencing datasets from S. pyogenes using a transposon sequencing-specific analysis pipeline, Transit. Across four genetic backgrounds and nine phenotypic conditions, 355 genes were essential for survival, corresponding to ~24% of the core genome. Clusters of Orthologous Genes (COG) categories related to coenzyme and lipid transport and growth functions were overrepresented as essential. Finally, essential operons across S. pyogenes genotypes were defined, with an increased number of essential operons detected under in vivo conditions. This study provides an extendible database to which new studies can be added, and a searchable html-based resource to direct future investigations into S. pyogenes biology.IMPORTANCEStreptococcus pyogenes is a human-adapted pathogen occupying restricted ecological niches. Understanding the essentiality of genes across different strains and experimental conditions is important to direct research questions and efforts to prevent the large burden of disease caused by S. pyogenes. To this end we systematically reanalyzed transposon sequencing studies in S. pyogenes using transposon sequencing-specific methods, integrating them into an extendible meta-analysis framework. This provides a repository of gene essentiality in S. pyogenes which was used to highlight specific genes of interest and for the community to guide future phenotypic studies.

Quality assurance for next-generation sequencing diagnostics of rare neurological diseases in the European Reference Network.

In the past decade, next-generation sequencing (NGS) has revolutionised genetic diagnostics for rare neurological disorders (RND). However, the lack of standardised technical, interpretative, and reporting standards poses a challenge for ensuring consistent and high-quality diagnostics globally. To address this, the European Reference Network for Rare Neurological Diseases (ERN-RND) collaborated with the European Molecular Genetics Quality Network (EMQN) to establish an external quality assessment scheme for NGS-based diagnostics in RNDs. The scheme, initiated in 2021 with a pilot involving 29 labs and followed by a second round in 2022 with 42 labs, aimed to evaluate the performance of laboratories in genetic testing for RNDs. Each participating lab analysed genetic data from three hypothetical cases, assessing genotyping, interpretation, and clerical accuracy. Despite a majority of labs using exome or genome sequencing, there was considerable variability in gene content, sequencing quality, adherence to standards, and clinical guidance provision. Results showed that while most labs provided correct molecular diagnoses, there was significant variability in reporting technical quality, adherence to interpretation standards, reporting strategies, and clinical commentary. Notably, some labs returned results with the potential for adverse medical outcomes. This underscores the need for further harmonisation, guideline development, and external quality assessment in the evolving landscape of genomic diagnostics for RNDs. Overall, the experience with the scheme highlighted the generally good quality of participating labs but emphasised the imperative for ongoing improvement in data analysis, interpretation, and reporting to enhance patient safety.

Morphological, Metabolomic and Genomic Evidences on Drought Stress Protective Functioning of the Endophyte Bacillus safensis Ni7.

The metabolomic and genomic characterization of an endophytic Bacillus safensis Ni7 was carried out in this study. This strain has previously been isolated from the xerophytic plant Nerium indicum L. and reported to enhance the drought tolerance in Capsicum annuum L. seedlings. The effects of drought stress on the morphology, biofilm production, and metabolite production of B. safensis Ni7 are analyzed in the current study. From the results obtained, the organism was found to have multiple strategies such as aggregation and clumping, robust biofilm production, and increased production of surfactin homologues under the drought induced condition when compared to non-stressed condition. Further the whole genome sequencing (WGS) based analysis has demonstrated B. safensis Ni7 to have a genome size of 3,671,999bp, N50 value of 3,527,239, and a mean G+C content of 41.58%. Interestingly the organism was observed to have the presence of various stress-responsive genes (13, 20U, 16U,160, 39, 17M, 18, 26, and ctc) and genes responsible for surfactin production (srfAA, srfAB, srfAC, and srfAD), biofilm production (epsD, epsE, epsF, epsG, epsH, epsI, epsK, epsL, epsM, epsN, and pel), chemotaxis (cheB_1, cheB_2, cheB_3, cheW_1, cheW_2 cheR, cheD, cheC, cheA, cheY, cheV, and cheB_4), flagella synthesis (flgG_1, flgG_2, flgG_3, flgC, and flgB) as supportive to the drought tolerance. Besides these, the genes responsible for plant growth promotion (PGP), including the genes for nitrogen (nasA, nasB, nasC, nasD, and nasE) and sulfur assimilation (cysL_1&L_2, cysI) and genes for phosphate solubilization (phoA, phoP_1& phoP_2, and phoR) could also be predicted. Along with the same, the genes for catalase, superoxide dismutase, protein homeostasis, cellular fitness, osmoprotectants production, and protein folding could also be predicted from its WGS data. Further pan-genome analysis with plant associated B. safensis strains available in the public databases revealed B. safensis Ni7 to have the presence of a total of 5391 gene clusters. Among these, 3207 genes were identified as core genes, 954 as shell genes and 1230 as cloud genes. This variation in gene content could be taken as an indication of evolution of strains of Bacillus safensis as per specific conditions and hence in the case of B. safensis Ni7 its role in habitat adaptation of plant is well expected. This diversity in endophytic bacterial genes may attribute its role to support the plant system to cope up with stress conditions. Overall, the study provides genomic evidence on Bacillus safensis Ni7 as a stress alleviating microbial partner in plants.

Presence of phage-plasmids in multiple serovars of Salmonella enterica.

Evidence is accumulating in the literature that the horizontal spread of antimicrobial resistance (AMR) genes mediated by bacteriophages and bacteriophage-like plasmid (phage-plasmid) elements is much more common than previously envisioned. For instance, we recently identified and characterized a circular P1-like phage-plasmid harbouring a bla CTX-M-15 gene conferring extended-spectrum beta-lactamase (ESBL) resistance in Salmonella enterica serovar Typhi. As the prevalence and epidemiological relevance of such mechanisms has never been systematically assessed in Enterobacterales, in this study we carried out a follow-up retrospective analysis of UK Salmonella isolates previously sequenced as part of routine surveillance protocols between 2016 and 2021. Using a high-throughput bioinformatics pipeline we screened 47 784 isolates for the presence of the P1 lytic replication gene repL, identifying 226 positive isolates from 25 serovars and demonstrating that phage-plasmid elements are more frequent than previously thought. The affinity for phage-plasmids appears highly serovar-dependent, with several serovars being more likely hosts than others; most of the positive isolates (170/226) belonged to S. Typhimurium ST34 and ST19. The phage-plasmids ranged between 85.8 and 98.2 kb in size, with an average length of 92.1 kb; detailed analysis indicated a high amount of diversity in gene content and genomic architecture. In total, 132 phage-plasmids had the p0111 plasmid replication type, and 94 the IncY type; phylogenetic analysis indicated that both horizontal and vertical gene transmission mechanisms are likely to be involved in phage-plasmid propagation. Finally, phage-plasmids were present in isolates that were resistant and non-resistant to antimicrobials. In addition to providing a first comprehensive view of the presence of phage-plasmids in Salmonella, our work highlights the need for a better surveillance and understanding of phage-plasmids as AMR carriers, especially through their characterization with long-read sequencing.

A review on recent taxonomic updates of gut bacteria associated with social bees, with a curated genomic reference database.

Honeybees and bumblebees play a crucial role as essential pollinators. The special gut microbiome of social bees is a key factor in determining the overall fitness and health of the host. Although bees harbor relatively simple microbial communities at the genus level, recent studies have unveiled significant genetic divergence and variations in gene content within each bacterial genus. However, a comprehensive and refined genomics-based taxonomic database specific to social bee gut microbiomes remains lacking. Here, we first provided an overview of the current knowledge on the distribution and function of social bee gut bacteria, as well as the factors that influence the gut population dynamics. We then consolidated all available genomes of the gut bacteria of social bees and refined the species-level taxonomy, by constructing a maximum-likelihood core genome phylogeny and calculating genome-wide pairwise average nucleotide identity. On the basis of the refined species taxonomy, we constructed a curated genomic reference database, named the bee gut microbe genome sequence database (BGM-GDb). To evaluate the species-profiling performance of the curated BGM-GDb, we retrieved a series of bee gut metagenomic data and inferred the species-level composition using metagenomic intra-species diversity analysis system (MIDAS), and then compared the results with those obtained from a prebuilt MIDAS database. We found that compared with the default database, the BGM-GDb excelled in aligned read counts and bacterial richness. Overall, this high-resolution and precise genomic reference database will facilitate research in understanding the gut community structure of social bees.

Analysis of twelve genomes of the bacterium Kerstersia gyiorum from brown-throated sloths (Bradypus variegatus), the first from a non-human host.

Kerstersia gyiorum is a Gram-negative bacterium found in various animals, including humans, where it has been associated with various infections. Knowledge of the basic biology of K. gyiorum is essential to understand the evolutionary strategies of niche adaptation and how this organism contributes to infectious diseases; however, genomic data about K. gyiorum is very limited, especially from non-human hosts. In this work, we sequenced 12 K. gyiorum genomes isolated from healthy free-living brown-throated sloths (Bradypus variegatus) in the Parque Estadual das Fontes do Ipiranga (São Paulo, Brazil), and compared them with genomes from isolates of human origin, in order to gain insights into genomic diversity, phylogeny, and host specialization of this species. Phylogenetic analysis revealed that these K. gyiorum strains are structured according to host. Despite the fact that sloth isolates were sampled from a single geographic location, the intra-sloth K. gyiorum diversity was divided into three clusters, with differences of more than 1,000 single nucleotide polymorphisms between them, suggesting the circulation of various K. gyiorum lineages in sloths. Genes involved in mobilome and defense mechanisms against mobile genetic elements were the main source of gene content variation between isolates from different hosts. Sloth-specific K.gyiorum genome features include an IncN2 plasmid, a phage sequence, and a CRISPR-Cas system. The broad diversity of defense elements in K. gyiorum (14 systems) may prevent further mobile element flow and explain the low amount of mobile genetic elements in K. gyiorum genomes. Gene content variation may be important for the adaptation of K. gyiorum to different host niches. This study furthers our understanding of diversity, host adaptation, and evolution of K. gyiorum, by presenting and analyzing the first genomes of non-human isolates.

Whole Genome Sequencing and Comparative Genomics of Six Staphylococcus schleiferi and Staphylococcus coagulans Isolates.

Staphylococcus schleiferi and Staphylococcus coagulans, closely related bacterial species within the Staphylococcus genus, present a challenge in classification and diagnosis due to their close genetic proximity and overlapping phenotypic features. Moreover, our understanding of the virulence mechanisms in staphylococcal species, beyond the extensively studied Staphylococcus aureus, remains limited, underscoring the importance of using comparative data to enhance our insights into virulence within these bacterial species. This study employed a comprehensive approach, utilizing comparative genomics, to identify genomic distinctions between S. schleiferi and S. coagulans, aiming to address the challenges in the accurate classification and diagnosis of these organisms and identify unique features. Whole genome sequencing was performed on six clinical isolates, and their genomes were compared to identify variations in gene content and virulence factors. De novo assembly and annotation revealed two samples as S. coagulans and four samples as S. schleiferi. Analysis of the core genomes revealed conserved regions crucial for defining species identity, while accessory genomic elements contained unique genes, possibly impacting the pathogenicity of the species.

Genome assembly of Medicago truncatula accession SA27063 provides insight into spring black stem and leaf spot disease resistance.

Medicago truncatula, model legume and alfalfa relative, has served as an essential resource for advancing our understanding of legume physiology, functional genetics, and crop improvement traits. Necrotrophic fungus, Ascochyta medicaginicola, the causal agent of spring black stem (SBS) and leaf spot is a devasting foliar disease of alfalfa affecting stand survival, yield, and forage quality. Host resistance to SBS disease is poorly understood, and control methods rely on cultural practices. Resistance has been observed in M. truncatula accession SA27063 (HM078) with two recessively inherited quantitative-trait loci (QTL), rnpm1 and rnpm2, previously reported. To shed light on host resistance, we carried out a de novo genome assembly of HM078. The genome, referred to as MtHM078 v1.0, is comprised of 23 contigs totaling 481.19 Mbp. Notably, this assembly contains a substantial amount of novel centromere-related repeat sequences due to deep long-read sequencing. Genome annotation resulted in 98.4% of BUSCO fabales proteins being complete. The assembly enabled sequence-level analysis of rnpm1 and rnpm2 for gene content, synteny, and structural variation between SBS-resistant accession SA27063 (HM078) and SBS-susceptible accession A17 (HM101). Fourteen candidate genes were identified, and some have been implicated in resistance to necrotrophic fungi. Especially interesting candidates include loss-of-function events in HM078 because they fit the inverse gene-for-gene model, where resistance is recessively inherited. In rnpm1, these include a loss-of-function in a disease resistance gene due to a premature stop codon, and a 10.85 kbp retrotransposon-like insertion disrupting a ubiquitin conjugating E2. In rnpm2, we identified a frameshift mutation causing a loss-of-function in a glycosidase, as well as a missense and frameshift mutation altering an F-box family protein. This study generated a high-quality genome of HM078 and has identified promising candidates, that once validated, could be further studied in alfalfa to enhance disease resistance.

Genome evolution and transcriptome plasticity is associated with adaptation to monocot and dicot plants in Colletotrichum fungi.

Colletotrichum fungi infect a wide diversity of monocot and dicot hosts, causing diseases on almost all economically important plants worldwide. Colletotrichum is also a suitable model for studying gene family evolution on a fine scale to uncover events in the genome associated with biological changes. Here we present the genome sequences of 30 Colletotrichum species covering the diversity within the genus. Evolutionary analyses revealed that the Colletotrichum ancestor diverged in the late Cretaceous in parallel with the diversification of flowering plants. We provide evidence of independent host jumps from dicots to monocots during the evolution of Colletotrichum, coinciding with a progressive shrinking of the plant cell wall degradative arsenal and expansions in lineage-specific gene families. Comparative transcriptomics of 4 species adapted to different hosts revealed similarity in gene content but high diversity in the modulation of their transcription profiles on different plant substrates. Combining genomics and transcriptomics, we identified a set of core genes such as specific transcription factors, putatively involved in plant cell wall degradation. These results indicate that the ancestral Colletotrichum were associated with dicot plants and certain branches progressively adapted to different monocot hosts, reshaping the gene content and its regulation.

The gut microbial composition in polycystic ovary syndrome with hyperandrogenemia and its association with steroid hormones.

Background: Polycystic ovary syndrome (PCOS) is characterized by excess androgens, ovulatory dysfunction, and polycystic ovaries. The mechanisms underlying ovulatory and metabolic disorders in PCOS remain elusive, hampering therapeutic development. Enhanced metabolic health correlates with increased microbiota gene content and microbial diversity. We aimed to explore the impact of gut microbiota and serum steroids on PCOS regulation associated with androgen excess. Methods: The fecal samples of patients with hyperandrogenic PCOS (n = 14) and control group with PCOS (n = 14) were analyzed by 16S rRNA gene sequencing. The peripheral venous blood of all subjects was collected to detect serum hormones. The association between gut microbiota and serum hormones was analyzed with the R language. Results: Our findings reveal that the hyperandrogenic PCOS group exhibits lower richness and diversity of gut microbiota compared to the control group. Characteristic genera in PCOS patients with hyperandrogenism include Bifidobacterium, Enterobacteriaceae_unclassified, Streptococcus, Saccharimonadaceae, Enterococcus, and Eubacterium_nodatum_group. Five hormones, including 5β-androsterone, deoxycorticosterone, corticosterone, 11-dehydrocorticosterone, and cortexolone, emerge as potential serum biomarkers for identifying patients with hyperandrogenic-PCOS (HA-PCOS). Furthermore, a lower vitamin D3 level may act as a susceptibility factor, suggesting that vitamin D3 supplementation could serve as a potential intervention for PCOS with hyperandrogenism. Conclusion: Specific fecal microbiota and serum steroids may be used as characteristic markers for clinical diagnosis of hyperandrogenic-PCOS. This research enhances our understanding of the intricate interplay among hormones, gut microbiota, and hyperandrogenemia in patients with PCOS.

Comparative genomics of clinical Stenotrophomonas maltophilia isolates reveals genetic diversity which correlates with colonization and persistence in vivo.

Stenotrophomonas maltophilia is a Gram-negative emerging opportunistic pathogen often present in people with respiratory diseases such as cystic fibrosis (CF). People with CF (pwCF) experience lifelong polymicrobial infections of the respiratory mucosa. Our prior work showed that Pseudomonas aeruginosa promotes persistence of S. maltophilia in mouse respiratory infections. As is typical for environmental opportunistic pathogens, S. maltophilia has a large genome and a high degree of genetic diversity. In this study, we evaluated the genomic content of S. maltophilia, combining short and long read sequencing to construct nearly complete genomes of 10 clinical isolates. The genomes of these isolates were then compared with all publicly available S. maltophilia genome assemblies, and each isolate was then evaluated for colonization/persistence in vivo, both alone and in coinfection with P. aeruginosa. We found that while the overall genome size and GC content were fairly consistent between strains, there was considerable variability in both genome structure and gene content. Similarly, there was significant variability in S. maltophilia colonization and persistence in experimental mouse respiratory infections in the presence or absence of P. aeruginosa. Ultimately, this study gives us a greater understanding of the genomic diversity of clinical S. maltophilia isolates, and how this genomic diversity relates to both interactions with other pulmonary pathogens and to host disease progression. Identifying the molecular determinants of infection with S. maltophilia can facilitate development of novel antimicrobial strategies for a highly drug-resistant pathogen.

Lateral gene transfer generates accessory genes that accumulate at different rates within a grass lineage.

Lateral gene transfer (LGT) is the movement of DNA between organisms without sexual reproduction. The acquired genes represent genetic novelties that have independently evolved in the donor's genome. Phylogenetic methods have shown that LGT is widespread across the entire grass family, although we know little about the underlying dynamics. We identify laterally acquired genes in five de novo reference genomes from the same grass genus (four Alloteropsis semialata and one Alloteropsis angusta). Using additional resequencing data for a further 40 Alloteropsis individuals, we place the acquisition of each gene onto a phylogeny using stochastic character mapping, and then infer rates of gains and losses. We detect 168 laterally acquired genes in the five reference genomes (32-100 per genome). Exponential decay models indicate that the rate of LGT acquisitions (6-28 per Ma) and subsequent losses (11-24% per Ma) varied significantly among lineages. Laterally acquired genes were lost at a higher rate than vertically inherited loci (0.02-0.8% per Ma). This high turnover creates intraspecific gene content variation, with a preponderance of them occurring as accessory genes in the Alloteropsis pangenome. This rapid turnover generates standing variation that can ultimately fuel local adaptation.

Genome assembly and analysis of Lactuca virosa: implications for lettuce breeding.

Lettuce (Lactuca sativa L.) is a leafy vegetable crop with ongoing breeding efforts related to quality, resilience, and innovative production systems. To breed resilient and resistant lettuce in the future, valuable genetic variation found in close relatives could be further exploited. Lactuca virosa (2x = 2n = 18), a wild relative assigned to the tertiary lettuce gene pool, has a much larger genome (3.7 Gbp) than Lactuca sativa (2.5 Gbp). It has been used in interspecific crosses and is a donor to modern crisphead lettuce cultivars. Here, we present a de novo reference assembly of L. virosa with high continuity and complete gene space. This assembly facilitated comparisons to the genome of L. sativa and to that of the wild species L. saligna, a representative of the secondary lettuce gene pool. To assess the diversity in gene content, we classified the genes of the 3 Lactuca species as core, accessory, and unique. In addition, we identified 3 interspecific chromosomal inversions compared to L. sativa, which each may cause recombination suppression and thus hamper future introgression breeding. Using 3-way comparisons in both reference-based and reference-free manners, we show that the proliferation of long-terminal repeat elements has driven the genome expansion of L. virosa. Further, we performed a genome-wide comparison of immune genes, nucleotide-binding leucine-rich repeat, and receptor-like kinases among Lactuca spp. and indicated the evolutionary patterns and mechanisms behind their expansions. These genome analyses greatly facilitate the understanding of genetic variation in L. virosa, which is beneficial for the breeding of improved lettuce varieties.

Characterization of novel recombinant mycobacteriophages derived from homologous recombination between two temperate phages.

Comparative analyses of mycobacteriophage genomes reveals extensive genetic diversity in genome organization and gene content, contributing to widespread mosaicism. We previously reported that the prophage of mycobacteriophage Butters (cluster N) provides defense against infection by Island3 (subcluster I1). To explore the anti-Island3 defense mechanism, we attempted to isolate Island3 defense escape mutants on a Butters lysogen, but only uncovered phages with recombinant genomes comprised of regions of Butters and Island3 arranged from left arm to right arm as Butters-Island3-Butters (BIBs). Recombination occurs within two distinct homologous regions that encompass lysin A, lysin B, and holin genes in one segment, and RecE and RecT genes in the other. Structural genes of mosaic BIB genomes are contributed by Butters while the immunity cassette is derived from Island3. Consequently, BIBs are morphologically identical to Butters (as shown by transmission electron microscopy) but are homoimmune with Island3. Recombinant phages overcome antiphage defense and silencing of the lytic cycle. We leverage this observation to propose a stratagem to generate novel phages for potential therapeutic use.

The Effect of Methodological Considerations on the Construction of Gene-Based Plant Pan-genomes.

Pan-genomics is an emerging approach for studying the genetic diversity within plant populations. In contrast to common resequencing studies that compare whole genome sequencing data with a single reference genome, the construction of a pan-genome (PG) involves the direct comparison of multiple genomes to one another, thereby enabling the detection of genomic sequences and genes not present in the reference, as well as the analysis of gene content diversity. Although multiple studies describing PGs of various plant species have been published in recent years, a better understanding regarding the effect of the computational procedures used for PG construction could guide researchers in making more informed methodological decisions. Here, we examine the effect of several key methodological factors on the obtained gene pool and on gene presence-absence detections by constructing and comparing multiple PGs of Arabidopsis thaliana and cultivated soybean, as well as conducting a meta-analysis on published PGs. These factors include the construction method, the sequencing depth, and the extent of input data used for gene annotation. We observe substantial differences between PGs constructed using three common procedures (de novo assembly and annotation, map-to-pan, and iterative assembly) and that results are dependent on the extent of the input data. Specifically, we report low agreement between the gene content inferred using different procedures and input data. Our results should increase the awareness of the community to the consequences of methodological decisions made during the process of PG construction and emphasize the need for further investigation of commonly applied methodologies.

Exploring environmental intra-species diversity through non-redundant pangenome assemblies.

At the genome level, microorganisms are highly adaptable both in terms of allele and gene composition. Such heritable traits emerge in response to different environmental niches and can have a profound influence on microbial community dynamics. As a consequence, any individual genome or population will contain merely a fraction of the total genetic diversity of any operationally defined "species", whose ecological potential can thus be only fully understood by studying all of their genomes and the genes therein. This concept, known as the pangenome, is valuable for studying microbial ecology and evolution, as it partitions genomes into core (present in all the genomes from a species, and responsible for housekeeping and species-level niche adaptation among others) and accessory regions (present only in some, and responsible for intra-species differentiation). Here we present SuperPang, an algorithm producing pangenome assemblies from a set of input genomes of varying quality, including metagenome-assembled genomes (MAGs). SuperPang runs in linear time and its results are complete, non-redundant, preserve gene ordering and contain both coding and non-coding regions. Our approach provides a modular view of the pangenome, identifying operons and genomic islands, and allowing to track their prevalence in different populations. We illustrate this by analysing intra-species diversity in Polynucleobacter, a bacterial genus ubiquitous in freshwater ecosystems, characterized by their streamlined genomes and their ecological versatility. We show how SuperPang facilitates the simultaneous analysis of allelic and gene content variation under different environmental pressures, allowing us to study the drivers of microbial diversification at unprecedented resolution.