Impact Of Horizontal Gene Transfer Research Articles

Unraveling the genetic diversity and evolution of Arbuscular Mycorrhizal Fungi (AMF)

The analysis of genetic differences among arbuscular mycorrhizal fungi (AMF) species has shed light on the genetic diversity and evolution of these fascinating organisms. AMF are essential constituents of land ecosystems because of their mutualistic symbiotic associations with plant roots. Their capability to amplify nutrient absorption, enhance water assimilation, and stimulate overall plant development plays a pivotal function in sustaining thriving ecosystems. By extensively examining the genomes of various AMF classifications, researchers have uncovered their evolutionary connections and gained understanding of the impact of horizontal gene transfer (HGT) on their genetic makeup. This knowledge has not only upgraded our comprehension of the evolutionary past of AMF but has also furnished valuable insights into the mechanisms that govern the symbiotic interactions between fungi and plants. In future studies on AMF genome variance, it is crucial to prioritize functional characterization, understand co-evolutionary dynamics with host plants, uncover mechanisms and implications of HGT, and integrate metagenomic approaches. By identifying the specific genes and genetic variations that contribute to AMF's ability to form mutualistic associations with plants, scientists can gain insights into the underlying mechanisms of this important ecological relationship. Furthermore, through the analysis of genetic divergence in both AMF and their host plants, researchers can elucidate the co-evolutionary dynamics that have shaped their mutualistic symbiosis. Equally, understanding the contribution of horizontal gene transfer (HGT) to AMF genetic diversity can provide insights into the evolutionary history and adaptive abilities of these fungi. Ultimately, the incorporation of metagenomic techniques into future studies on AMF genetic diversity will facilitate a broader understanding of the microorganism populations associated with these fungi. By focusing on these domains, it is possible to advance our understanding of AMF biology and unlock novel opportunities for innovative applications in agriculture and ecosystem management.

Genome mining reveals abiotic stress resistance genes in plant genomes acquired from microbes via HGT.

Colonization by beneficial microbes can enhance plant tolerance to abiotic stresses. However, there are still many unknown fields regarding the beneficial plant-microbe interactions. In this study, we have assessed the amount or impact of horizontal gene transfer (HGT)-derived genes in plants that have potentials to confer abiotic stress resistance. We have identified a total of 235 gene entries in fourteen high-quality plant genomes belonging to phyla Chlorophyta and Streptophyta that confer resistance against a wide range of abiotic pressures acquired from microbes through independent HGTs. These genes encode proteins contributed to toxic metal resistance (e.g., ChrA, CopA, CorA), osmotic and drought stress resistance (e.g., Na+/proline symporter, potassium/proton antiporter), acid resistance (e.g., PcxA, ArcA, YhdG), heat and cold stress resistance (e.g., DnaJ, Hsp20, CspA), oxidative stress resistance (e.g., GST, PoxA, glutaredoxin), DNA damage resistance (e.g., Rad25, Rad51, UvrD), and organic pollutant resistance (e.g., CytP450, laccase, CbbY). Phylogenetic analyses have supported the HGT inferences as the plant lineages are all clustering closely with distant microbial lineages. Deep-learning-based protein structure prediction and analyses, in combination with expression assessment based on codon adaption index (CAI) further corroborated the functionality and expressivity of the HGT genes in plant genomes. A case-study applying fold comparison and molecular dynamics (MD) of the HGT-driven CytP450 gave a more detailed illustration on the resemblance and evolutionary linkage between the plant recipient and microbial donor sequences. Together, the microbe-originated HGT genes identified in plant genomes and their participation in abiotic pressures resistance indicate a more profound impact of HGT on the adaptive evolution of plants.

Modulating Gene Expression within a Microbiome Based on Computational Models.

Simple SummaryDevelopment of computational biology methodologies has provided comprehensive understanding of the complexity of microbiomes, and the extensive ways in which they influence their environment. This has awakened a new research goal, aiming to not only understand the mechanisms in which microbiomes function, but to actively modulate and engineer them for various purposes. However, current microbiome engineering techniques are usually manually tailored for a specific system and neglect the different interactions between the new genetic information and the bacterial population, turning a blind eye to processes such as horizontal gene transfer, mutations, and other genetic alterations. In this work, we developed a generic computational method to automatically tune the expression of heterologous genes within a microbiome according to given preferences, to allow the functionality of the engineering process to propagate in longer periods of time. This goal was achieved by treating each part of the gene individually and considering long term fitness effects on the environment, providing computational and experimental evidence for this approach.Recent research in the field of bioinformatics and molecular biology has revealed the immense complexity and uniqueness of microbiomes, while also showcasing the impact of the symbiosis between a microbiome and its host or environment. A core property influencing this process is horizontal gene transfer between members of the bacterial community used to maintain genetic variation. The essential effect of this mechanism is the exposure of genetic information to a wide array of members of the community, creating an additional “layer” of information in the microbiome named the “plasmidome”. From an engineering perspective, introduction of genetic information to an environment must be facilitated into chosen species which will be able to carry out the desired effect instead of competing and inhibiting it. Moreover, this process of information transfer imposes concerns for the biosafety of genetic engineering of microbiomes as exposure of genetic information into unwanted hosts can have unprecedented ecological impacts. Current technologies are usually experimentally developed for a specific host/environment, and only deal with the transformation process itself at best, ignoring the impact of horizontal gene transfer and gene-microbiome interactions that occur over larger periods of time in uncontrolled environments. The goal of this research was to design new microbiome-specific versions of engineered genetic information, providing an additional layer of compatibility to existing engineering techniques. The engineering framework is entirely computational and is agnostic to the selected microbiome or gene by reducing the problem into the following set up: microbiome species can be defined as wanted or unwanted hosts of the modification. Then, every element related to gene expression (e.g., promoters, coding regions, etc.) and regulation is individually examined and engineered by novel algorithms to provide the defined expression preferences. Additionally, the synergistic effect of the combination of engineered gene blocks facilitates robustness to random mutations that might occur over time. This method has been validated using both computational and experimental tools, stemming from the research done in the iGEM 2021 competition, by the TAU group.

Evaluation of existing guidelines for their adequacy for the food and feed risk assessment of microorganisms obtained through synthetic biology.

EFSA was asked by the European Commission to evaluate synthetic biology (SynBio) developments for agri‐food use in the near future and to determine whether or not they are expected to constitute potential new hazards/risks. Moreover, EFSA was requested to evaluate the adequacy of existing guidelines for risk assessment of SynBio and if updated guidance is needed. The scope of this Opinion covers food and feed risk assessment, the variety of microorganisms that can be used in the food/feed chain and the whole spectrum of techniques used in SynBio. This Opinion complements a previously adopted Opinion with the evaluation of existing guidelines for the microbial characterisation and environmental risk assessment of microorganisms obtained through SynBio. The present Opinion confirms that microbial SynBio applications for food and feed use, with the exception of xenobionts, could be ready in the European Union in the next decade. New hazards were identified related to the use or production of unusual and/or new‐to‐nature components. Fifteen cases were selected for evaluating the adequacy of existing guidelines. These were generally adequate for assessing the product, the production process, nutritional and toxicological safety, allergenicity, exposure and post‐market monitoring. The comparative approach and a safety assessment per se could be applied depending on the degree of familiarity of the SynBio organism/product with the non‐genetically modified counterparts. Updated guidance is recommended for: (i) bacteriophages, protists/microalgae, (ii) exposure to plant protection products and biostimulants, (iii) xenobionts and (iv) feed additives for insects as target species. Development of risk assessment tools is recommended for assessing nutritional value of biomasses, influence of microorganisms on the gut microbiome and the gut function, allergenic potential of new‐to‐nature proteins, impact of horizontal gene transfer and potential risks of living cell intake. A further development towards a strain‐driven risk assessment approach is recommended.

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.

Impact of horizontal gene transfer on emergence and stability of cooperative virulence in Salmonella Typhimurium

Intestinal inflammation fuels the transmission of Salmonella Typhimurium (S.Tm). However, a substantial fitness cost is associated with virulence expression. Mutations inactivating transcriptional virulence regulators generate attenuated variants profiting from inflammation without enduring virulence cost. Such variants interfere with the transmission of fully virulent clones. Horizontal transfer of functional regulatory genes (HGT) into attenuated variants could nevertheless favor virulence evolution. To address this hypothesis, we cloned hilD, coding for the master regulator of virulence, into a conjugative plasmid that is highly transferrable during intestinal colonization. The resulting mobile hilD allele allows virulence to emerge from avirulent populations, and to be restored in attenuated mutants competing against virulent clones within-host. However, mutations inactivating the mobile hilD allele quickly arise. The stability of virulence mediated by HGT is strongly limited by its cost, which depends on the hilD expression level, and by the timing of transmission. We conclude that robust evolution of costly virulence expression requires additional selective forces such as narrow population bottlenecks during transmission.

Population Biology and Comparative Genomics of Campylobacter Species.

The zoonotic pathogen Campylobacter is the leading cause for bacterial foodborne infections in humans. Campylobacters are most commonly transmitted via the consumption of undercooked poultry meat or raw milk products. The decreasing costs of whole genome sequencing enabled large genome-based analyses of the evolution and population structure of this pathogen, as well as the development of novel high-throughput molecular typing methods. Here, we review the evolutionary development and the population diversity of the two most clinically relevant Campylobacter species; C. jejuni and C. coli. The state-of-the-art phylogenetic studies showed clustering of C. jejuni lineages into host specialists and generalists with coexisting lifestyles in chicken and livestock-associated hosts, as well as the separation of C. coli isolates of riparian origin (waterfowl, water) from C. coli isolated from clinical and farm-related samples. We will give an overview of recombination between both species and the potential impact of horizontal gene transfer on host adaptation in Campylobacter. Additionally, this review briefly places the current knowledge of the population structure of other Campylobacter species such as C. lari, C. concisus and C. upsaliensis into perspective. We also provide an overview of how molecular typing methods such as multilocus sequence typing (MLST) and whole genome MLST have been used to detect and trace Campylobacter outbreaks along the food chain.

A Comprehensive Coexpression Network Analysis in Vibrio cholerae

Research into the evolution and pathogenesis of Vibrio cholerae has benefited greatly from the generation of high-throughput sequencing data to drive molecular analyses. The steady accumulation of these data sets now provides a unique opportunity for in silico hypothesis generation via coexpression analysis. Here, we leverage all published V. cholerae RNA sequencing data, in combination with select data from other platforms, to generate a gene coexpression network that validates known gene interactions and identifies novel genetic partners across the entire V. cholerae genome. This network provides direct insights into genes influencing pathogenicity, metabolism, and transcriptional regulation, further clarifies results from previous sequencing experiments in V. cholerae (e.g., transposon insertion sequencing [Tn-seq] and chromatin immunoprecipitation sequencing [ChIP-seq]), and expands upon microarray-based findings in related Gram-negative bacteria.IMPORTANCE Cholera is a devastating illness that kills tens of thousands of people annually. Vibrio cholerae, the causative agent of cholera, is an important model organism to investigate both bacterial pathogenesis and the impact of horizontal gene transfer on the emergence and dissemination of new virulent strains. Despite the importance of this pathogen, roughly one-third of V. cholerae genes are functionally unannotated, leaving large gaps in our understanding of this microbe. Through coexpression network analysis of existing RNA sequencing data, this work develops an approach to uncover novel gene-gene relationships and contextualize genes with no known function, which will advance our understanding of V. cholerae virulence and evolution.

Horizontal Gene Transfer and Endophytes: An Implication for the Acquisition of Novel Traits.

Horizontal gene transfer (HGT), an important evolutionary mechanism observed in prokaryotes, is the transmission of genetic material across phylogenetically distant species. In recent years, the availability of complete genomes has facilitated the comprehensive analysis of HGT and highlighted its emerging role in the adaptation and evolution of eukaryotes. Endophytes represent an ecologically favored association, which highlights its beneficial attributes to the environment, in agriculture and in healthcare. The HGT phenomenon in endophytes, which features an important biological mechanism for their evolutionary adaptation within the host plant and simultaneously confers “novel traits” to the associated microbes, is not yet completely understood. With a focus on the emerging implications of HGT events in the evolution of biological species, the present review discusses the occurrence of HGT in endophytes and its socio-economic importance in the current perspective. To our knowledge, this review is the first report that provides a comprehensive insight into the impact of HGT in the adaptation and evolution of endophytes.

Transcriptome analysis of Xanthomonas oryzae pv. oryzicola exposed to H2O2 reveals horizontal gene transfer contributes to its oxidative stress response.

Xanthomonas oryzae pv. oryzicola (Xoc), the causal agent of bacterial leaf streak, is one of the most severe seed-borne bacterial diseases of rice. However, the molecular mechanisms underlying Xoc in response to oxidative stress are still unknown. In this study, we performed a time-course RNA-seq analysis on the Xoc in response to H2O2, aiming to reveal its oxidative response network. Overall, our RNA sequence analysis of Xoc revealed a significant global gene expression profile when it was exposed to H2O2. There were 7, 177, and 246 genes that were differentially regulated at the early, middle, and late stages after exposure, respectively. Three genes (xoc_1643, xoc_1946, xoc_3249) showing significantly different expression levels had proven relationships with oxidative stress response and pathogenesis. Moreover, a hypothetical protein (XOC_2868) showed significantly differential expression, and the xoc_2868 mutants clearly displayed a greater H2O2 sensitivity and decreased pathogenicity than those of the wild-type. Gene localization and phylogeny analysis strongly suggests that this gene may have been horizontally transferred from a Burkholderiaceae ancestor. Our study not only provides a first glance of Xoc's global response against oxidative stress, but also reveals the impact of horizontal gene transfer in the evolutionary history of Xoc.

A putative antimicrobial peptide from Hymenoptera in the megaplasmid pSCL4 of Streptomyces clavuligerus ATCC 27064 reveals a singular case of horizontal gene transfer with potential applications.

Streptomyces clavuligerus is a Gram‐positive bacterium that is a high producer of secondary metabolites with industrial applications. The production of antibiotics such as clavulanic acid or cephamycin has been extensively studied in this species; nevertheless, other aspects, such as evolution or ecology, have received less attention. Furthermore, genes that arise from ancient events of lateral transfer have been demonstrated to be implicated in important functions of host species. This approximation discovered relevant genes that genomic analyses overlooked. Thus, we studied the impact of horizontal gene transfer in the S. clavuligerus genome. To perform this task, we applied whole‐genome analysis to identify a laterally transferred sequence from different domains. The most relevant result was a putative antimicrobial peptide (AMP) with a clear origin in the Hymenoptera order of insects. Next, we determined that two copies of these genes were present in the megaplasmid pSCL4 but absent in the S. clavuligerus ATCC 27064 chromosome. Additionally, we found that these sequences were exclusive to the ATCC 27064 strain (and so were not present in any other bacteria) and we also verified the expression of the genes using RNAseq data. Next, we used several AMP predictors to validate the original annotation extracted from Hymenoptera sequences and explored the possibility that these proteins had post‐translational modifications using peptidase cleavage prediction. We suggest that Hymenoptera AMP‐like proteins of S. clavuligerus ATCC 27064 may be useful for both species adaptation and as an antimicrobial molecule with industrial applications.

In vivo competition and horizontal gene transfer among distinct Staphylococcus aureus lineages as major drivers for adaptational changes during long-term persistence in humans

BackgroundThe airways of the majority of adolescent cystic fibrosis (CF) patients are persistently colonized or infected by Staphylococcus aureus. Using whole genome sequencing, we studied the evolutionary traits within a S. aureus population in the airways of a CF patient hypothesizing that horizontal gene transfer (HGT) and inter-bacterial interaction play a major role in adaptation during long-term persistence.ResultsWhole genome sequencing of 21 S. aureus isolates spanning 13 years resulted in seven lineages defined by the spa types t012, t021, t331, t338, t364, t056, and t2351. Of these, the successfully persisting lineages t012 and t021 were closely related suggesting the evolution of t021 from t012, which was further corroborated by a nearly identical, syntenic set of mobile genetic elements. During transformation from t012 to t021, an increase of genomic changes including HGT from other S. aureus lineages was detected.ConclusionsIn summary, our in vivo data enabled us to conceptualize an evolutionary model showing the impact of HGT and inter-bacterial interaction on bacterial long-term adaptation to the human host during CF.

Genome-wide analysis of horizontally acquired genes in the genus Mycobacterium

Horizontal gene transfer (HGT) was attributed as a major driving force for the innovation and evolution of prokaryotic genomes. Previously, multiple research endeavors were undertaken to decipher HGT in different bacterial lineages. The genus Mycobacterium houses some of the most deadly human pathogens; however, the impact of HGT in Mycobacterium has never been addressed in a systematic way. Previous initiatives to explore the genomic imprints of HGTs in Mycobacterium were focused on few selected species, specifically among the members of Mycobacterium tuberculosis complex. Considering the recent availability of a large number of genomes, the current study was initiated to decipher the probable events of HGTs among 109 completely sequenced Mycobacterium species. Our comprehensive phylogenetic analysis with more than 9,000 families of Mycobacterium proteins allowed us to list several instances of gene transfers spread across the Mycobacterium phylogeny. Moreover, by examining the topology of gene phylogenies here, we identified the species most likely to donate and receive these genes and provided a detailed overview of the putative functions these genes may be involved in. Our study suggested that horizontally acquired foreign genes had played an enduring role in the evolution of Mycobacterium genomes and have contributed to their metabolic versatility and pathogenicity.

Evolutionary transitions between beneficial and phytopathogenic Rhodococcus challenge disease management.

Understanding how bacteria affect plant health is crucial for developing sustainable crop production systems. We coupled ecological sampling and genome sequencing to characterize the population genetic history of Rhodococcus and the distribution patterns of virulence plasmids in isolates from nurseries. Analysis of chromosome sequences shows that plants host multiple lineages of Rhodococcus, and suggested that these bacteria are transmitted due to independent introductions, reservoir populations, and point source outbreaks. We demonstrate that isolates lacking virulence genes promote beneficial plant growth, and that the acquisition of a virulence plasmid is sufficient to transition beneficial symbionts to phytopathogens. This evolutionary transition, along with the distribution patterns of plasmids, reveals the impact of horizontal gene transfer in rapidly generating new pathogenic lineages and provides an alternative explanation for pathogen transmission patterns. Results also uncovered a misdiagnosed epidemic that implicated beneficial Rhodococcus bacteria as pathogens of pistachio. The misdiagnosis perpetuated the unnecessary removal of trees and exacerbated economic losses.

Protist Evolution: Stealing Genes to Gut It Out

The amount and evolutionary impact of horizontal gene transfer in eukaryotes remain contentious issues. Anew phylogenomic study suggests that gene transfer from prokaryotes has contributed significantly to the adaptation and metabolic evolution of Blastocystis, the most widespread human gut eukaryotic parasite.

Unparalleled replacement of native mitochondrial genes by foreign homologs in a holoparasitic plant.

Horizontal gene transfer (HGT) among flowering plant mitochondria occurs frequently and, in most cases, leads to nonfunctional transgenes in the recipient genome. Parasitic plants are particularly prone to this phenomenon, but their mitochondrial genomes (mtDNA) have been largely unexplored. We undertook a large-scale mitochondrial genomic study of the holoparasitic plant Lophophytum mirabile (Balanophoraceae). Comprehensive phylogenetic analyses were performed to address the frequency, origin, and impact of HGT. The sequencing of the complete mtDNA of L.mirabile revealed the unprecedented acquisition of host-derived mitochondrial genes, representing 80% of the protein-coding gene content. All but two of these foreign genes replaced the native homologs and are probably functional in energy metabolism. The genome consists of 54 circular-mapping chromosomes, 25 of which carry no intact genes. The likely functional replacement of up to 26 genes in L.mirabile represents a stunning example of the potential effect of rampant HGT on plant mitochondria. The use of host-derived genes may have a positive effect on the host-parasite relationship, but could also be the result of nonadaptive forces.

Key experimental evidence of chromosomal DNA transfer among selected tuberculosis-causing mycobacteria

Horizontal gene transfer (HGT) is a major driving force of bacterial diversification and evolution. For tuberculosis-causing mycobacteria, the impact of HGT in the emergence and distribution of dominant lineages remains a matter of debate. Here, by using fluorescence-assisted mating assays and whole genome sequencing, we present unique experimental evidence of chromosomal DNA transfer between tubercle bacilli of the early-branching Mycobacterium canettii clade. We found that the obtained recombinants had received multiple donor-derived DNA fragments in the size range of 100 bp to 118 kbp, fragments large enough to contain whole operons. Although the transfer frequency between M. canettii strains was low and no transfer could be observed among classical Mycobacterium tuberculosis complex (MTBC) strains, our study provides the proof of concept for genetic exchange in tubercle bacilli. This outstanding, now experimentally validated phenomenon presumably played a key role in the early evolution of the MTBC toward pathogenicity. Moreover, our findings also provide important information for the risk evaluation of potential transfer of drug resistance and fitness mutations among clinically relevant mycobacterial strains.

Intraspecies Genomic Diversity and Long-Term Persistence of Bifidobacterium longum.

Members of genus Bifidobacterium are Gram-positive bacteria, representing a large part of the human infant microbiota and moderately common in adults. However, our knowledge about their diversity, intraspecific phylogeny and long-term persistence in humans is still limited. Bifidobacterium longum is generally considered to be the most common and prevalent species in the intestinal microbiota. In this work we studied whole genome sequences of 28 strains of B. longum, including 8 sequences described in this paper. Part of these strains were isolated from healthy children during a long observation period (up to 10 years between isolation from the same patient). The three known subspecies (longum, infantis and suis) could be clearly divided using sequence-based phylogenetic methods, gene content and the average nucleotide identity. The profiles of glycoside hydrolase genes reflected the different ecological specializations of these three subspecies. The high impact of horizontal gene transfer on genomic diversity was observed, which is possibly due to a large number of prophages and rapidly spreading plasmids. The pan-genome characteristics of the subspecies longum corresponded to the open pan-genome model. While the major part of the strain-specific genetic loci represented transposons and phage-derived regions, a large number of cell envelope synthesis genes were also observed within this category, representing high variability of cell surface molecules. We observed the cases of isolation of high genetically similar strains of B. longum from the same patients after long periods of time, however, we didn’t succeed in the isolation of genetically identical bacteria: a fact, reflecting the high plasticity of microbiota in children.

Transduction-mediated horizontal gene transfer in the oral microbiome.

The recent article, “The impact of horizontal gene transfer on the adaptive ability of the human oral microbiome” by Roberts and Kreth (2014) provides a valuable review of the mechanisms and consequences of horizontal gene transfer (HGT) in the oral microbiome. It discusses the three well-known mechanisms of HGT (i.e., transformation, conjugation and transduction), as well as the recently described mechanism of vesicle-mediated gene transfer, although the bulk of the attention is given to transformation occurring in oral biofilms. The authors emphasize the potential importance of HGT in the oral microbiome by pointing to the demonstrated transfer of genes (in vitro) from transient oral bacteria, unable to colonize oral biofilms, to members of the established oral microbiome community. Thus, the genes (and their consequences) may remain in the oral cavity long after the (bacterial) sources of the genes are gone. In this regard, it should be noted that there have been several recent reports of transient oral infections following the ingestion of certain foods, such as cheeses (Razavi et al., 2007; Kamodyova et al., 2014). Whilst the magnitude of the bacteriophage load in the oral cavity is acknowledged by the authors, the potential of bacteriophages for HGT via transduction in the oral cavity is, however, not as clearly or accurately described. For example, the authors state that “Studies on saliva have isolated bacteriophages able to lysogenize specific bacterial pathogens such as Enterococcus faecalis (Bachrach et al., 2003; Stevens et al., 2009) and Aggregatibacter actinomycetemcomitans (Sandmeier et al., 1995; Willi et al., 1997).” While the Bachrach et al paper did report on E. faecalis phages isolated from saliva, the Stevens et al article reported on bacteriophages induced from E. faecalis isolated from infected root canals (NOT saliva), and the source of the A. actinomycetemcomitans from which phages were induced in the works by Sandmeier et al, and Willi et al, was subgingival plaque (again, NOT saliva). This is of some significance since it demonstrates that there are multiple niches (not just saliva) harboring bacterial viruses in the oral cavity, and phages in any of these niches could be a vehicle for HGT. Secondly, although HGT may occur by temperate phages via specialized transduction, it is also possible for HGT to occur by virulent phages through generalized transduction. Consequently, “…bacteriophages able to lysogenize…” may not be the only explanation for transduction-mediated HGT. This was not considered in the discussion of bacteriophages and HGT. Further, it is quite curious that the authors state that “…there have been no reported observations of transduction in the oral cavity, or relevant in vitro models…” since the title of the Willi et al. article cited in this review is “Transduction of antibiotic resistance markers among Actinobacillus actinomycetemecomitans strains by temperate bacteriophage Aa phi 23” (emphasis added). This study clearly demonstrated the capacity of bacteriophages (through transduction) to horizontally transfer genes between strains of oral bacteria, at least in vitro. Finally, it is customary that the references cited should be the original source of the information in question (Annesley, 2011). Reports of E. faecalis bacteriophages in the oral cavity (e.g., Natkin, 1967) were published more than 30 years prior to the two articles cited above by the authors. Similarly, isolation of A. actinomycetemcomitans bacteriophages was originally reported (Stevens et al., 1982) more than 10 years prior to the articles (Sandmeier et al., 1995; Willi et al., 1997) cited by the authors.

The evolution of fungal metabolic pathways.

Fungi contain a remarkable range of metabolic pathways, sometimes encoded by gene clusters, enabling them to digest most organic matter and synthesize an array of potent small molecules. Although metabolism is fundamental to the fungal lifestyle, we still know little about how major evolutionary processes, such as gene duplication (GD) and horizontal gene transfer (HGT), have interacted with clustered and non-clustered fungal metabolic pathways to give rise to this metabolic versatility. We examined the synteny and evolutionary history of 247,202 fungal genes encoding enzymes that catalyze 875 distinct metabolic reactions from 130 pathways in 208 diverse genomes. We found that gene clustering varied greatly with respect to metabolic category and lineage; for example, clustered genes in Saccharomycotina yeasts were overrepresented in nucleotide metabolism, whereas clustered genes in Pezizomycotina were more common in lipid and amino acid metabolism. The effects of both GD and HGT were more pronounced in clustered genes than in their non-clustered counterparts and were differentially distributed across fungal lineages; specifically, GD, which was an order of magnitude more abundant than HGT, was most frequently observed in Agaricomycetes, whereas HGT was much more prevalent in Pezizomycotina. The effect of HGT in some Pezizomycotina was particularly strong; for example, we identified 111 HGT events associated with the 15 Aspergillus genomes, which sharply contrasts with the 60 HGT events detected for the 48 genomes from the entire Saccharomycotina subphylum. Finally, the impact of GD within a metabolic category was typically consistent across all fungal lineages, whereas the impact of HGT was variable. These results indicate that GD is the dominant process underlying fungal metabolic diversity, whereas HGT is episodic and acts in a category- or lineage-specific manner. Both processes have a greater impact on clustered genes, suggesting that metabolic gene clusters represent hotspots for the generation of fungal metabolic diversity.