Molecular evolution under increasing transposable element burden in Drosophila: A speed limit on the evolutionary arms race.

Genome architecture is profoundly influenced by transposable elements (TEs), and natural selection against their harmful effects is a critical factor limiting their spread. Genome defense by the piRNA silencing pathway also plays a crucial role in limiting TE proliferation. How these two forces jointly determine TE abundance is not well understood. To shed light on the nature of factors that predict TE success, we test three distinct hypotheses in the Drosophila genus. First, we determine whether TE abundance and relaxed genome-wide purifying selection on protein sequences are positively correlated. This serves to test the hypothesis that variation in TE abundance in the Drosophila genus can be explained by the strength of natural selection, relative to drift, acting in parallel against mildly deleterious non-synonymous mutations. Second, we test whether increasing TE abundance is correlated with an increased rate of amino-acid evolution in genes encoding the piRNA machinery, as might be predicted by an evolutionary arms race model. Third, we test whether increasing TE abundance is correlated with greater codon bias in genes of the piRNA machinery. This is predicted if increasing TE abundance selects for increased efficiency in the machinery of genome defense

Highly Constrained Intergenic Drosophila Ultraconserved Elements Are Candidate ncRNAs

Eukaryotes contain short (∼80–200 bp) regions that have few or no substitutions among species that represent hundreds of millions of years of evolutionary divergence. These ultraconserved elements (UCEs) are candidates for containing essential functions, but their biological roles remain largely unknown. Here, we report the discovery and characterization of UCEs from 12 sequenced Drosophilaspecies. We identified 98 elements ≥80 bp long with very high conservation across the Drosophila phylogeny. Population genetic analyses reveal that these UCEs are not present in mutational cold spots. Instead we infer that they experience a level of selective constraint almost 10-fold higher compared with missense mutations in protein-coding sequences, which is substantially higher than that observed previously for human UCEs. About one-half of these Drosophila UCEs overlap the transcribed portion of genes, with many of those that are within coding sequences likely to correspond to sites of ADAR-dependent RNA editing. For the remaining UCEs that are in nongenic regions, we find that many are potentially capable of forming RNA secondary structures. Among ten chosen for further analysis, we discovered that the majority are transcribed in multiple tissues of Drosophila melanogaster. We conclude that Drosophilaspecies are rich with UCEs and that many of them may correspond to novel noncoding RNAs

Local adaptation and the evolution of genome architecture in threespine stickleback.

Theory predicts that local adaptation should favour the evolution of a concentrated genetic architecture, where the alleles driving adaptive divergence are tightly clustered on chromosomes. Adaptation to marine vs. freshwater environments in threespine stickleback has resulted in an architecture that seems consistent with this prediction: divergence among populations is mainly driven by a few genomic regions harbouring multiple quantitative trait loci (QTL) for environmentally adapted traits, as well as candidate genes with well-established phenotypic effects. One theory for the evolution of these "genomic islands" is that rearrangements remodel the genome to bring causal loci into tight proximity, but this has not been studied explicitly. We tested this theory using synteny analysis to identify micro- and macro-rearrangements in the stickleback genome and assess their potential involvement in the evolution of genomic islands. To identify rearrangements, we conducted a de novo assembly of the closely-related tubesnout (Aulorhyncus flavidus) genome and compared this to the genomes of threespine stickleback and two other closely related species. We found that small rearrangements, within-chromosome duplications, and Lineage-Specific Genes (LSGs) were enriched around genomic islands, and that all three chromosomes harbouring large genomic islands have experienced macro-rearrangements. We also found that duplicates and micro-rearrangements are 9.9x and 2.9x more likely to involve genes differentially expressed between marine and freshwater genotypes. While not conclusive, these results are consistent with the explanation that strong divergent selection on candidate genes drove the recruitment of rearrangements to yield clusters of locally adaptive loci

Antagonistic pleiotropy in the bifunctional surface protein FadL (OmpP1) during adaptation of Haemophilus influenzae to chronic lung infection associated with Chronic Obstructive Pulmonary Disease

Tracking bacterial evolution during chronic infection provides insights into how host selection pressures shape bacterial genomes. The human-restricted opportunistic pathogen nontypeable Haemophilus influenzae (NTHi) infects the lower airways of patients suffering chronic obstructive pulmonary disease (COPD) and contributes to disease progression. To identify bacterial genetic variation associated with bacterial adaptation to the COPD lung, we sequenced the genomes of 92 isolates collected from the sputum of 13 COPD patients over 1 to 9years. Individuals were colonized by distinct clonal types (CTs) over time, but the same CT was often reisolated at a later time or found in different patients. Although genomes from the same CT were nearly identical, intra-CT variation due to mutation and recombination occurred. Recurrent mutations in several genes were likely involved in COPD lung adaptation. Notably, nearly a third of CTs were polymorphic for null alleles of ompP1 (also called fadL), which encodes a bifunctional membrane protein that both binds the human carcinoembryonic antigen-related cell adhesion molecule 1 (hCEACAM1) receptor and imports long-chain fatty acids (LCFAs). Our computational studies provide plausible three-dimensional models for FadL's interaction with hCEACAM1 and LCFA binding. We show that recurrent fadL mutations are likely a case of antagonistic pleiotropy, since loss of FadL reduces NTHi's ability to infect epithelia but also increases its resistance to bactericidal LCFAs enriched within the COPD lung. Supporting this interpretation, truncated fadL alleles are common in publicly available NTHi genomes isolated from the lower airway tract but rare in others. These results shed light on molecular mechanisms of bacterial pathoadaptation and guide future research toward developing novel COPD therapeutics.IMPORTANCE Nontypeable Haemophilus influenzae is an important pathogen in patients with chronic obstructive pulmonary disease (COPD). To elucidate the bacterial pathways undergoing in vivo evolutionary adaptation, we compared bacterial genomes collected over time from 13 COPD patients and identified recurrent genetic changes arising in independent bacterial lineages colonizing different patients. Besides finding changes in phase-variable genes, we found recurrent loss-of-function mutations in the ompP1 (fadL) gene. We show that loss of OmpP1/FadL function reduces this bacterium's ability to infect cells via the hCEACAM1 epithelial receptor but also increases its resistance to bactericidal fatty acids enriched within the COPD lung, suggesting a case of antagonistic pleiotropy that restricts DeltafadL strains' niche. These results show how H. influenzae adapts to host-generated inflammatory mediators in the COPD airways

Phase Variation in HMW1A Controls a Phenotypic Switch in Haemophilus influenzae Associated with Pathoadaptation during Persistent Infection

Genetic variants arising from within-patient evolution shed light on bacterial adaptation during chronic infection. Contingency loci generate high levels of genetic variation in bacterial genomes, enabling adaptation to the stringent selective pressures exerted by the host. A significant gap in our understanding of phase-variable contingency loci is the extent of their contribution to natural infections. The human-adapted pathogen nontypeable Haemophilus influenzae (NTHi) causes persistent infections, which contribute to underlying disease progression. The phase-variable high-molecular-weight (HMW) adhesins located on the NTHi surface mediate adherence to respiratory epithelial cells and, depending on the allelic variant, can also confer high epithelial invasiveness or hyperinvasion. In this study, we characterize the dynamics of HMW-mediated hyperinvasion in living cells and identify a specific HMW binding domain shared by hyperinvasive NTHi isolates of distinct pathological origins. Moreover, we observed that HMW expression decreased over time by using a longitudinal set of persistent NTHi strains collected from chronic obstructive pulmonary disease (COPD) patients, resulting from increased numbers of simple-sequence repeats (SSRs) downstream of the functional P2hmw1A promoter, which is the one primarily driving HMW expression. Notably, the increased SSR numbers at the hmw1 promoter region also control a phenotypic switch toward lower bacterial intracellular invasion and higher biofilm formation, likely conferring adaptive advantages during chronic airway infection by NTHi. Overall, we reveal novel molecular mechanisms of NTHi pathoadaptation based on within-patient lifestyle switching controlled by phase variation. IMPORTANCE Human-adapted bacterial pathogens have evolved specific mechanisms to colonize their host niche. Phase variation is a contingency strategy to allow adaptation to changing conditions, as phase-variable bacterial loci rapidly and reversibly switch their expression. Several NTHi adhesins are phase variable. These adhesins are required for colonization but also immunogenic, in such a way that bacteria with lower adhesin levels are better equipped to survive an immune response, making their contribution to natural infections unclear. We show here that the major NTHi adhesin HMW1A displays allelic variation, which can drive a phase-variable epithelial hyperinvasion phenotype. Over time, hmw1A phase variation lowers adhesin expression, which controls an NTHi lifestyle switch from high epithelial invasiveness to lower invasion and higher biofilm formation. This reversible loss of function aligns with the previously stated notion that epithelial infection is essential for NTHi infection establishment, but once established, persistence favors gene inactivation, in this case facilitating biofilm growth

Lung transcriptional unresponsiveness and loss of early influenza virus control in infected neonates is prevented by intranasal Lactobacillus rhamnosus GG

Respiratory viral infections contribute substantially to global infant losses and disproportionately affect preterm neonates. Using our previously established neonatal murine model of influenza infection, we demonstrate that three-day old mice are exceptionally sensitive to influenza virus infection and exhibit high mortality and viral load. Intranasal pre- and post-treatment of neonatal mice with Lactobacillus rhamnosus GG (LGG), an immune modulator in respiratory viral infection of adult mice and human preterm neonates, considerably improves neonatal mice survival after influenza virus infection. We determine that both live and heat-killed intranasal LGG are equally efficacious in protection of neonates. Early in influenza infection, neonatal transcriptional responses in the lung are delayed compared to adults. These responses increase by 24 hours post-infection, demonstrating a delay in the kinetics of the neonatal anti-viral response. LGG pretreatment improves immune gene transcriptional responses during early infection and specifically upregulates type I IFN pathways. This is critical for protection, as neonatal mice intranasally pre-treated with IFNβ before influenza virus infection are also protected. Using transgenic mice, we demonstrate that the protective effect of LGG is mediated through a MyD88-dependent mechanism, specifically via TLR4. LGG can improve both early control of virus and transcriptional responsiveness and could serve as a simple and safe intervention to protect neonates

Screening of the mutant library obtained by natural transformation for the identification of Haemophilus influenziae genes involved in human epithelial invasion

Trabajo presentado en el XXIV Congreso de la Sociedad Española de Microbiología, celebrado en L’Hospitalet de Llobregat (Barcelona) del 10 al 13 de julio de 2013.Se ha empleado la transformación natural como estrategia de cribado de genes/agrupamientos genéticos implicados en la invasión del epitelio respiratorio humano por el patógeno respiratorio H. Influenzae. Este trabajo constituye la primera búsqueda sistemática de invasinas de H. Influenzae mediante un cribado positivo de ganancia de función.MINECO SAF2012-31168; Departamento de Salud Gobierno Foral de Navarra; CIBERES, (España); Universidad British Columbia-UBC, (Canadá).Peer Reviewe

Transformation of natural genetic variation into haemophilus influenzae genomes

Science, Faculty ofZoology, Department ofReviewedFacult