Abstract
Lateral genetic transfer (LGT) is a major source of phenotypic innovation among bacteria. Determinants for antibiotic resistance and other adaptive traits can spread rapidly, particularly by ...conjugative plasmids, but also phages and natural transformation. Each successive step from the uptake of foreign DNA, its genetic recombination and regulatory integration, to its establishment in the host population presents differential barriers and opportunities. The emergence of successive multidrug-resistant strains of Staphylococcus aureus illustrates the ongoing role of LGT in the combinatorial assembly of pathogens. The dynamic interplay among hosts, vectors, DNA elements, combinations of genetic determinants and environments constructs communities of genetic exchange. These relations can be abstracted as a graph, within which an exchange community might correspond to a path, transitively closed set, clique or near-clique. We provide a set-based definition, and review the features of actual genetic exchange communities (GECs), adopting first a knowledge-driven approach based on literature, and then a synoptic data-centric bioinformatic approach. GECs are diverse, but share some common features.
Differential opportunity and barriers to lateral genetic transfer create bacterial communities of exchange.
Despite the enormous diversity among parasitic angiosperms in form and structure, life-history strategies, and plastid genomes, little is known about the diversity of their mitogenomes. We report the ...sequence of the wonderfully bizarre mitogenome of the hemiparasitic aerial mistletoe Viscum scurruloideum. This genome is only 66 kb in size, making it the smallest known angiosperm mitogenome by a factor of more than three and the smallest land plant mitogenome. Accompanying this size reduction is exceptional reduction of gene content. Much of this reduction arises from the unexpected loss of respiratory complex I (NADH dehydrogenase), universally present in all 300+ other angiosperms examined, where it is encoded by nine mitochondrial and many nuclear nad genes. Loss of complex I in a multicellular organism is unprecedented. We explore the potential relationship between this loss in Viscum and its parasitic lifestyle. Despite its small size, the Viscum mitogenome is unusually rich in recombinationally active repeats, possessing unparalleled levels of predicted sublimons resulting from recombination across short repeats. Many mitochondrial gene products exhibit extraordinary levels of divergence in Viscum, indicative of highly relaxed if not positive selection. In addition, all Viscum mitochondrial protein genes have experienced a dramatic acceleration in synonymous substitution rates, consistent with the hypothesis of genomic streamlining in response to a high mutation rate but completely opposite to the pattern seen for the high-rate but enormous mitogenomes of Silene. In sum, the Viscum mitogenome possesses a unique constellation of extremely unusual features, a subset of which may be related to its parasitic lifestyle.
Multidrug-resistant bacteria are spreading at alarming rates, and despite extensive efforts no new class of antibiotic with activity against Gram-negative bacteria has been approved in over fifty ...years. Natural products and their derivatives have a key role in combating Gram-negative pathogens. Here we report chemical optimization of the arylomycins-a class of natural products with weak activity and limited spectrum-to obtain G0775, a molecule with potent, broad-spectrum activity against Gram-negative bacteria. G0775 inhibits the essential bacterial type I signal peptidase, a new antibiotic target, through an unprecedented molecular mechanism. It circumvents existing antibiotic resistance mechanisms and retains activity against contemporary multidrug-resistant Gram-negative clinical isolates in vitro and in several in vivo infection models. These findings demonstrate that optimized arylomycin analogues such as G0775 could translate into new therapies to address the growing threat of multidrug-resistant Gram-negative infections.
Escherichia coli sequence type 131 (ST131) is a globally disseminated, multidrug resistant (MDR) clone responsible for a high proportion of urinary tract and bloodstream infections. The rapid ...emergence and successful spread of E. coli ST131 is strongly associated with several factors, including resistance to fluoroquinolones, high virulence gene content, the possession of the type 1 fimbriae FimH30 allele, and the production of the CTX-M-15 extended spectrum β-lactamase (ESBL). Here, we used genome sequencing to examine the molecular epidemiology of a collection of E. coli ST131 strains isolated from six distinct geographical locations across the world spanning 2000–2011. The global phylogeny of E. coli ST131, determined from whole-genome sequence data, revealed a single lineage of E. coli ST131 distinct from other extraintestinal E. coli strains within the B2 phylogroup. Three closely related E. coli ST131 sublineages were identified, with little association to geographic origin. The majority of single-nucleotide variants associated with each of the sublineages were due to recombination in regions adjacent to mobile genetic elements (MGEs). The most prevalent sublineage of ST131 strains was characterized by fluoroquinolone resistance, and a distinct virulence factor and MGE profile. Four different variants of the CTX-M ESBL–resistance gene were identified in our ST131 strains, with acquisition of CTX-M-15 representing a defining feature of a discrete but geographically dispersed ST131 sublineage. This study confirms the global dispersal of a single E. coli ST131 clone and demonstrates the role of MGEs and recombination in the evolution of this important MDR pathogen.
Lipopolysaccharide (LPS) resides in the outer membrane of Gram-negative bacteria where it is responsible for barrier function
. LPS can cause death as a result of septic shock, and its lipid A core ...is the target of polymyxin antibiotics
. Despite the clinical importance of polymyxins and the emergence of multidrug resistant strains
, our understanding of the bacterial factors that regulate LPS biogenesis is incomplete. Here we characterize the inner membrane protein PbgA and report that its depletion attenuates the virulence of Escherichia coli by reducing levels of LPS and outer membrane integrity. In contrast to previous claims that PbgA functions as a cardiolipin transporter
, our structural analyses and physiological studies identify a lipid A-binding motif along the periplasmic leaflet of the inner membrane. Synthetic PbgA-derived peptides selectively bind to LPS in vitro and inhibit the growth of diverse Gram-negative bacteria, including polymyxin-resistant strains. Proteomic, genetic and pharmacological experiments uncover a model in which direct periplasmic sensing of LPS by PbgA coordinates the biosynthesis of lipid A by regulating the stability of LpxC, a key cytoplasmic biosynthetic enzyme
. In summary, we find that PbgA has an unexpected but essential role in the regulation of LPS biogenesis, presents a new structural basis for the selective recognition of lipids, and provides opportunities for future antibiotic discovery.
The outer membrane is an essential structural component of Gram-negative bacteria that is composed of lipoproteins, lipopolysaccharides, phospholipids, and integral β-barrel membrane proteins. A ...dedicated machinery, called the Lol system, ensures proper trafficking of lipoproteins from the inner to the outer membrane. The LolCDE ABC transporter is the inner membrane component, which is essential for bacterial viability. Here, we report a novel pyrrolopyrimidinedione compound, G0507, which was identified in a phenotypic screen for inhibitors of
growth followed by selection of compounds that induced the extracytoplasmic σ
stress response. Mutations in
,
, and
conferred resistance to G0507, suggesting LolCDE as its molecular target. Treatment of
cells with G0507 resulted in accumulation of fully processed Lpp, an outer membrane lipoprotein, in the inner membrane. Using purified protein complexes, we found that G0507 binds to LolCDE and stimulates its ATPase activity. G0507 still binds to LolCDE harboring a Q258K substitution in LolC (LolC
), which confers high-level resistance to G0507
but no longer stimulates ATPase activity. Our work demonstrates that G0507 has significant promise as a chemical probe to dissect lipoprotein trafficking in Gram-negative bacteria.
Tandem gene amplification is a frequent and dynamic source of antibiotic resistance in bacteria. Ongoing expansions and contractions of repeat arrays during population growth are expected to manifest ...as cell-to-cell differences in copy number (CN). As a result, a clonal bacterial culture could comprise subpopulations of cells with different levels of antibiotic sensitivity that result from variable gene dosage. Despite the high potential for misclassification of heterogenous cell populations as either antibiotic-susceptible or fully resistant in clinical settings, and the concomitant risk of inappropriate treatment, CN distribution among cells has defied analysis. Here, we use the MinION single-molecule nanopore sequencer to uncover CN heterogeneity in clonal populations of
and
grown from single cells isolated while selecting for resistance to an optimized arylomycin, a member of a recently discovered class of Gram-negative antibiotic. We found that gene amplification of the arylomycin target, bacterial type I signal peptidase LepB, is a mechanism of unstable arylomycin resistance and demonstrate in
that amplification instability is independent of RecA. This instability drives the emergence of a nonuniform distribution of
CN among cells with a range of 1 to at least 50 copies of
identified in a single clonal population. In sum, this remarkable heterogeneity, and the evolutionary plasticity it fuels, illustrates how gene amplification can enable bacterial populations to respond rapidly to novel antibiotics. This study establishes a rationale for further nanopore-sequencing studies of heterogeneous cell populations to uncover CN variability at single-molecule resolution.
Influenza B virus (IBV) causes annual influenza epidemics around the world. Here we use an in vivo plasmablast enrichment technique to isolate a human monoclonal antibody, 46B8 that neutralizes all ...IBVs tested in vitro and protects mice against lethal challenge of all IBVs tested when administered 72 h post infection. 46B8 demonstrates a superior therapeutic benefit over Tamiflu and has an additive antiviral effect in combination with Tamiflu. 46B8 binds to a conserved epitope in the vestigial esterase domain of hemagglutinin (HA) and blocks HA-mediated membrane fusion. After passage of the B/Brisbane/60/2008 virus in the presence of 46B8, we isolated three resistant clones, all harbouring the same mutation (Ser301Phe) in HA that abolishes 46B8 binding to HA at low pH. Interestingly, 46B8 is still able to protect mice against lethal challenge of the mutant viruses, possibly owing to its ability to mediate antibody-dependent cellular cytotoxicity (ADCC).
IL-22 is induced by aryl hydrocarbon receptor (AhR) signaling and plays a critical role in gastrointestinal barrier function through effects on antimicrobial protein production, mucus secretion, and ...epithelial cell differentiation and proliferation, giving it the potential to modulate the microbiome through these direct and indirect effects. Furthermore, the microbiome can in turn influence IL-22 production through the synthesis of L-tryptophan (L-Trp)-derived AhR ligands, creating the prospect of a host-microbiome feedback loop. We evaluated the impact IL-22 may have on the gut microbiome and its ability to activate host AhR signaling by observing changes in gut microbiome composition, function, and AhR ligand production following exogenous IL-22 treatment in both mice and humans.
Microbiome alterations were observed across the gastrointestinal tract of IL-22-treated mice, accompanied by an increased microbial functional capacity for L-Trp metabolism. Bacterially derived indole derivatives were increased in stool from IL-22-treated mice and correlated with increased fecal AhR activity. In humans, reduced fecal concentrations of indole derivatives in ulcerative colitis (UC) patients compared to healthy volunteers were accompanied by a trend towards reduced fecal AhR activity. Following exogenous IL-22 treatment in UC patients, both fecal AhR activity and concentrations of indole derivatives increased over time compared to placebo-treated UC patients.
Overall, our findings indicate IL-22 shapes gut microbiome composition and function, which leads to increased AhR signaling and suggests exogenous IL-22 modulation of the microbiome may have functional significance in a disease setting. Video Abstract.
Changes in transcriptional regulation underlie many of the phenotypic differences observed within and between species of bacteria. Lateral genetic transfer (LGT) can significantly impact the ...transcription factor (TF) genes which drive these transcriptional changes. Although much emphasis has been placed on LGT of intact genes, the units of transfer and recombination do not necessarily correspond to regions delineated by exact gene boundaries. Here we apply phylogenetic and network-based methods to investigate the relationship between units of lateral transfer and recombination within the Escherichia coli - Shigella clade and the topological properties of genes in the E. coli transcriptional regulatory network (TRN).
We carried out a systematic phylogenetic study of genetic transfer among 5282 sets of putatively orthologous genes from 27 strains belonging to the E. coli - Shigella clade. We then used these results to examine the evolutionary histories of TF genes, as well as the transcriptional regulation of lateral genes. We found evidence of LGT in 2655 (50.3%) gene sets: 678 (12.8%) show evidence of recombination breakpoints within the gene boundaries. Thus, within- and whole- gene lateral transfer is widespread among strains of E. coli and Shigella. We found that unlike global regulators, which have mostly evolved vertically, neighbour regulators (genes which regulate adjacent genes on the chromosome) have frequently been subject to transfer within the E. coli - Shigella clade. At least 56 (62%) of the 90 neighbour regulator gene sets examined show evidence of LGT, 19 (34%) of which have internal recombination breakpoints. Neighbour regulators show no evidence of co-transfer with their nearby target genes. Rather, the frequency of recombination breakpoints, and conflicting evolutionary histories among neighbour regulators and their target genes, suggest that the genomic regions encoding these genes have been constructed through successive layering of LGT events within the clade. We find no difference in the relative complexity of regulation (i.e. the number of regulators) of lateral versus vertical genes.
Neighbour regulators show higher frequencies of transfer than other types of regulatory genes. This implicates the topological properties of regulatory genes in the TRN, and their physical proximity to targets on the chromosome, as contributing to successful LGT. The prevalence of recombination breakpoints within regulatory and target gene sets indicates that within-gene transfer has had a significant cumulative effect on the evolution of regulatory interactions in E. coli and Shigella.
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