David A Fitzpatrick

Publications of David A Fitzpatrick

• Horizontal gene transfer in fungi.

  Authors: David A Fitzpatrick

  FEMS microbiology letters. 11/2011; 329(1):1-8.

  Horizontal gene transfer (HGT) is frequently observed in prokaryotes and until recently was assumed to be of limited importance to eukaryotes. However, there is an increasing body of evidence toHorizontal gene transfer (HGT) is frequently observed in prokaryotes and until recently was assumed to be of limited importance to eukaryotes. However, there is an increasing body of evidence to suggest that HGT is an important mechanism in eukaryotic genome evolution, particularly in unicellular organisms. The transfer of individual genes, gene clusters or entire chromosomes can have significant impacts on niche specification, disease emergence or shift in metabolic capabilities. In terms of genomic sequencing, the fungal kingdom is one of the most densely sampled eukaryotic lineages and is at the forefront of eukaryote comparative genomics and enables us to use fungi to study eukaryotic evolutionary mechanisms including HGT. This review describes the bioinformatics-based methodologies commonly used to locate HGT in fungal genomes and investigates the possible mechanisms involved in transferring genetic material laterally into fungal species. I will highlight a number of fungal HGT events and discuss the impact they have played on fungal evolution and discuss the implications HGT may have on the fungal tree of life.

• Reconstructing the fungal tree of life using phylogenomics and a preliminary investigation of the distribution of yeast prion-like proteins in the fungal kingdom.

  Authors: Edgar M Medina, Gary W Jones, David A Fitzpatrick

  Journal of molecular evolution. 09/2011; 73(3-4):116-33.

  We have used three independent phylogenomic approaches (concatenated alignments, single-, and multi-gene supertrees) to reconstruct the fungal tree of life (FTOL) using publicly available fungalWe have used three independent phylogenomic approaches (concatenated alignments, single-, and multi-gene supertrees) to reconstruct the fungal tree of life (FTOL) using publicly available fungal genomes. This is the first time multi-gene families have been used in fungal supertree reconstruction and permits us to use up to 66% of the 1,001,217 genes in our fungal database. Our analyses show that different phylogenomic datasets derived from varying clustering criteria and alignment orientation do not have a major effect on phylogenomic supertree reconstruction. Overall the resultant phylogenomic trees are relatively congruent with one another and successfully recover the major fungal phyla, subphyla and classes. We find that where incongruences do occur, the inferences are usually poorly supported. Within the Ascomycota phylum, our phylogenies reconstruct monophyletic Saccharomycotina and Pezizomycotina subphyla clades and infer a sister group relationship between these to the exclusion of the Taphrinomycotina. Within the Pezizomycotina subphylum, all three phylogenies infer a sister group relationship between the Leotiomycetes and Sordariomycetes classes. However, there is conflict regarding the relationships with the Dothideomycetes and Eurotiomycetes classes. Within the Basidiomycota phylum, supertrees derived from single- and multi-gene families infer a sister group relationship between the Pucciniomycotina and Agaricomycotina subphyla while the concatenated phylogeny infers a poorly supported relationship between the Agaricomycotina and Ustilagomycotina. The reconstruction of a robust FTOL is important for future fungal comparative analyses. We illustrate this point by performing a preliminary investigation into the phyletic distribution of yeast prion-like proteins in the fungal kingdom.

• The copper regulon of the human fungal pathogen Cryptococcus neoformans H99.

  Authors: Chen Ding, Jun Yin, Edgar Mauricio Medina Tovar, David A Fitzpatrick, Desmond G Higgins, Dennis J Thiele

  Molecular microbiology. 08/2011; 81(6):1560-76.

  Cryptococcus neoformans is a human fungal pathogen that is the causative agent of cryptococcosis and fatal meningitis in immuno-compromised hosts. Recent studies suggest that copper (Cu) acquisitionCryptococcus neoformans is a human fungal pathogen that is the causative agent of cryptococcosis and fatal meningitis in immuno-compromised hosts. Recent studies suggest that copper (Cu) acquisition plays an important role in C. neoformans virulence, as mutants that lack Cuf1, which activates the Ctr4 high affinity Cu importer, are hypo-virulent in mouse models. To understand the constellation of Cu-responsive genes in C. neoformans and how their expression might contribute to virulence, we determined the transcript profile of C. neoformans in response to elevated Cu or Cu deficiency. We identified two metallothionein genes (CMT1 and CMT2), encoding cysteine-rich Cu binding and detoxifying proteins, whose expression is dramatically elevated in response to excess Cu. We identified a new C. neoformans Cu transporter, CnCtr1, that is induced by Cu deficiency and is distinct from CnCtr4 and which shows significant phylogenetic relationship to Ctr1 from other fungi. Surprisingly, in contrast to other fungi, we found that induction of both CnCTR1 and CnCTR4 expression under Cu limitation, and CMT1 and CMT2 in response to Cu excess, are dependent on the CnCuf1 Cu metalloregulatory transcription factor. These studies set the stage for the evaluation of the specific Cuf1 target genes required for virulence in C. neoformans.

• Universally distributed single-copy genes indicate a constant rate of horizontal transfer.

  Authors: Christopher J Creevey, Tobias Doerks, David A Fitzpatrick, Jeroen Raes, Peer Bork

  PloS one. 01/2011; 6(8):e22099.

  Single copy genes, universally distributed across the three domains of life and encoding mostly ancient parts of the translation machinery, are thought to be only rarely subjected to horizontal geneSingle copy genes, universally distributed across the three domains of life and encoding mostly ancient parts of the translation machinery, are thought to be only rarely subjected to horizontal gene transfer (HGT). Indeed it has been proposed to have occurred in only a few genes and implies a rare, probably not advantageous event in which an ortholog displaces the original gene and has to function in a foreign context (orthologous gene displacement, OGD). Here, we have utilised an automatic method to identify HGT based on a conservative statistical approach capable of robustly assigning both donors and acceptors. Applied to 40 universally single copy genes we found that as many as 68 HGTs (implying OGDs) have occurred in these genes with a rate of 1.7 per family since the last universal common ancestor (LUCA). We examined a number of factors that have been claimed to be fundamental to HGT in general and tested their validity in the subset of universally distributed single copy genes. We found that differing functional constraints impact rates of OGD and the more evolutionarily distant the donor and acceptor, the less likely an OGD is to occur. Furthermore, species with larger genomes are more likely to be subjected to OGD. Most importantly, regardless of the trends above, the number of OGDs increases linearly with time, indicating a neutral, constant rate. This suggests that levels of HGT above this rate may be indicative of positively selected transfers that may allow niche adaptation or bestow other benefits to the recipient organism.

• Assessment of inactivating stop codon mutations in forty Saccharomyces cerevisiae strains: implications for [PSI] prion- mediated phenotypes.

  Authors: David A Fitzpatrick, Jennifer O'Brien, Ciara Moran, Naushaba Hasin, Elaine Kenny, Paul Cormican, Amy Gates, Derek W Morris, Gary W Jones

  PloS one. 01/2011; 6(12):e28684.

  The yeast prion [PSI(+)] has been implicated in the generation of novel phenotypes by a mechanism involving a reduction in translation fidelity causing readthrough of naturally occurring stop codons.The yeast prion [PSI(+)] has been implicated in the generation of novel phenotypes by a mechanism involving a reduction in translation fidelity causing readthrough of naturally occurring stop codons. Some [PSI(+)] associated phenotypes may also be generated due to readthrough of inactivating stop codon mutations (ISCMs). Using next generation sequencing we have sequenced the genomes of two Saccharomyces cerevisiae strains that are commonly used for the study of the yeast [PSI(+)] prion. We have identified approximately 26,000 and 6,500 single nucleotide polymorphisms (SNPs) in strains 74-D694 and G600 respectively, compared to reference strain S288C. In addition to SNPs that produce non-synonymous amino acid changes we have also identified a number of SNPs that cause potential ISCMs in these strains, one of which we show is associated with a [PSI(+)]-dependent stress resistance phenotype in strain G600. We identified twenty-two potential ISCMs in strain 74-D694, present in genes involved in a variety of cellular processes including nitrogen metabolism, signal transduction and oxidative stress response. The presence of ISCMs in a subset of these genes provides possible explanations for previously identified [PSI(+)]-associated phenotypes in this strain. A comparison of ISCMs in strains G600 and 74-D694 with S. cerevisiae strains sequenced as part of the Saccharomyces Genome Resequencing Project (SGRP) shows much variation in the generation of strain-specific ISCMs and suggests this process is possible under complex genetic control. Additionally we have identified a major difference in the abilities of strains G600 and 74-D694 to grow at elevated temperatures. However, this difference appears unrelated to novel SNPs identified in strain 74-D694 present in proteins involved in the heat shock response, but may be attributed to other SNP differences in genes previously identified as playing a role in high temperature growth.

• Analysis of gene evolution and metabolic pathways using the Candida Gene Order Browser.

  Authors: David A Fitzpatrick, Peadar O'Gaora, Kevin P Byrne, Geraldine Butler

  BMC genomics. 01/2010; 11:290.

  Candida species are the most common cause of opportunistic fungal infection worldwide. Recent sequencing efforts have provided a wealth of Candida genomic data. We have developed the Candida GeneCandida species are the most common cause of opportunistic fungal infection worldwide. Recent sequencing efforts have provided a wealth of Candida genomic data. We have developed the Candida Gene Order Browser (CGOB), an online tool that aids comparative syntenic analyses of Candida species. CGOB incorporates all available Candida clade genome sequences including two Candida albicans isolates (SC5314 and WO-1) and 8 closely related species (Candida dubliniensis, Candida tropicalis, Candida parapsilosis, Lodderomyces elongisporus, Debaryomyces hansenii, Pichia stipitis, Candida guilliermondii and Candida lusitaniae). Saccharomyces cerevisiae is also included as a reference genome. CGOB assignments of homology were manually curated based on sequence similarity and synteny. In total CGOB includes 65617 genes arranged into 13625 homology columns. We have also generated improved Candida gene sets by merging/removing partial genes in each genome. Interrogation of CGOB revealed that the majority of tandemly duplicated genes are under strong purifying selection in all Candida species. We identified clusters of adjacent genes involved in the same metabolic pathways (such as catabolism of biotin, galactose and N-acetyl glucosamine) and we showed that some clusters are species or lineage-specific. We also identified one example of intron gain in C. albicans. Our analysis provides an important resource that is now available for the Candida community. CGOB is available at http://cgob.ucd.ie.

• Gene and genome trees conflict at many levels.

  Authors: Leanne S Haggerty, Fergal J Martin, David A Fitzpatrick, James O McInerney

  Philosophical transactions of the Royal Society of London. Series B, Biological sciences. 09/2009; 364(1527):2209-19.

  Horizontal gene transfer (HGT) plays a significant role in microbial evolution. It can accelerate the adaptation of an organism, it can generate new metabolic pathways and it can completely remodelHorizontal gene transfer (HGT) plays a significant role in microbial evolution. It can accelerate the adaptation of an organism, it can generate new metabolic pathways and it can completely remodel an organism's genome. We examine 27 closely related genomes from the YESS group of gamma proteobacteria and a variety of four-taxon datasets from a diverse range of prokaryotes in order to explore the kinds of effects HGT has had on these organisms.

• Evolution of pathogenicity and sexual reproduction in eight Candida genomes.

  Authors: Geraldine Butler, Matthew D Rasmussen, Michael F Lin, Manuel A S Santos, Sharadha Sakthikumar, Carol A Munro, Esther Rheinbay, Manfred Grabherr, Anja Forche, Jennifer L Reedy [......] Thyagarajan Srikantha, Qiandong Zeng, Judith Berman, Matthew Berriman, Joseph Heitman, Neil A R Gow, Michael C. Lorenz, Bruce W Birren, Manolis Kellis, Christina A Cuomo

  Nature. 07/2009; 459(7247):657-62.

  Candida species are the most common cause of opportunistic fungal infection worldwide. Here we report the genome sequences of six Candida species and compare these and related pathogens andCandida species are the most common cause of opportunistic fungal infection worldwide. Here we report the genome sequences of six Candida species and compare these and related pathogens and non-pathogens. There are significant expansions of cell wall, secreted and transporter gene families in pathogenic species, suggesting adaptations associated with virulence. Large genomic tracts are homozygous in three diploid species, possibly resulting from recent recombination events. Surprisingly, key components of the mating and meiosis pathways are missing from several species. These include major differences at the mating-type loci (MTL); Lodderomyces elongisporus lacks MTL, and components of the a1/2 cell identity determinant were lost in other species, raising questions about how mating and cell types are controlled. Analysis of the CUG leucine-to-serine genetic-code change reveals that 99% of ancestral CUG codons were erased and new ones arose elsewhere. Lastly, we revise the Candida albicans gene catalogue, identifying many new genes.

• Evidence of recent interkingdom horizontal gene transfer between bacteria and Candida parapsilosis.

  Authors: David A Fitzpatrick, Mary E Logue, Geraldine Butler

  BMC evolutionary biology. 02/2008; 8:181.

  BACKGROUND: To date very few incidences of interdomain gene transfer into fungi have been identified. Here, we used the emerging genome sequences of Candida albicans WO-1, Candida tropicalis, CandidaBACKGROUND: To date very few incidences of interdomain gene transfer into fungi have been identified. Here, we used the emerging genome sequences of Candida albicans WO-1, Candida tropicalis, Candida parapsilosis, Clavispora lusitaniae, Pichia guilliermondii, and Lodderomyces elongisporus to identify recent interdomain HGT events. We refer to these as CTG species because they translate the CTG codon as serine rather than leucine, and share a recent common ancestor. RESULTS: Phylogenetic and syntenic information infer that two C. parapsilosis genes originate from bacterial sources. One encodes a putative proline racemase (PR). Phylogenetic analysis also infers that there were independent transfers of bacterial PR enzymes into members of the Pezizomycotina, and protists. The second HGT gene in C. parapsilosis belongs to the phenazine F (PhzF) superfamily. Most CTG species also contain a fungal PhzF homolog. Our phylogeny suggests that the CTG homolog originated from an ancient HGT event, from a member of the proteobacteria. An analysis of synteny suggests that C. parapsilosis has lost the endogenous fungal form of PhzF, and subsequently reacquired it from a proteobacterial source. There is evidence that Schizosaccharomyces pombe and Basidiomycotina also obtained a PhzF homolog through HGT. CONCLUSION: Our search revealed two instances of well-supported HGT from bacteria into the CTG clade, both specific to C. parapsilosis. Therefore, while recent interkingdom gene transfer has taken place in the CTG lineage, its occurrence is rare. However, our analysis will not detect ancient gene transfers, and we may have underestimated the global extent of HGT into CTG species.

• Evaluation of whether accelerated protein evolution in chordates has occurred before, after, or simultaneously with gene duplication.

  Authors: Catrióna R Johnston, Colm O'Dushlaine, David A Fitzpatrick, Richard J Edwards, Denis C Shields

  Molecular biology and evolution. 02/2007; 24(1):315-23.

  Gene duplication and loss are predicted to be at least of the order of the substitution rate and are key contributors to the development of novel gene function and overall genome evolution. AlthoughGene duplication and loss are predicted to be at least of the order of the substitution rate and are key contributors to the development of novel gene function and overall genome evolution. Although it has been established that proteins evolve more rapidly after gene duplication, we were interested in testing to what extent this reflects causation or association. Therefore, we investigated the rate of evolution prior to gene duplication in chordates. Two patterns emerged; firstly, branches, which are both preceded by a duplication and followed by a duplication, display an elevated rate of amino acid replacement. This is reflected in the ratio of nonsynonymous to synonymous substitution (mean nonsynonymous to synonymous nucleotide substitution rate ratio [Ka:Ks]) of 0.44 compared with branches preceded by and followed by a speciation (mean Ka:Ks of 0.23). The observed patterns suggest that there can be simultaneous alteration in the selection pressures on both gene duplication and amino acid replacement, which may be consistent with co-occurring increases in positive selection, or alternatively with concurrent relaxation of purifying selection. The pattern is largely, but perhaps not completely, explained by the existence of certain families that have elevated rates of both gene duplication and amino acid replacement. Secondly, we observed accelerated amino acid replacement prior to duplication (mean Ka:Ks for postspeciation preduplication branches was 0.27). In some cases, this could reflect adaptive changes in protein function precipitating a gene duplication event. In conclusion, the circumstances surrounding the birth of new proteins may frequently involve a simultaneous change in selection pressures on both gene-copy number and amino acid replacement. More precise modeling of the relative importance of preduplication, postduplication, and simultaneous amino acid replacement will require larger and denser genomic data sets from multiple species, allowing simultaneous estimation of lineage-specific fluctuations in mutation rates and adaptive constraints.

• The molecular phylogeny of a nematode-specific clade of heterotrimeric G-protein alpha-subunit genes.

  Authors: Damien M O'Halloran, David A Fitzpatrick, Grace P McCormack, James O McInerney, Ann M Burnell

  Journal of molecular evolution. 08/2006; 63(1):87-94.

  In animal olfactory systems, odorant molecules are detected by olfactory receptors (ORs). ORs are part of the G-protein-coupled receptor (GPCR) superfamily. Heterotrimeric guanine nucleotide bindingIn animal olfactory systems, odorant molecules are detected by olfactory receptors (ORs). ORs are part of the G-protein-coupled receptor (GPCR) superfamily. Heterotrimeric guanine nucleotide binding G-proteins (G-proteins) relay signals from GPCRs to intracellular effectors. G-proteins are comprised of three peptides. The G-protein alpha subunit confers functional specificity to G-proteins. Vertebrate and insect Galpha-subunit genes are divided into four subfamilies based on functional and sequence attributes. The nematode Caenorhabditis elegans contains 21 Galpha genes, 14 of which are exclusively expressed in sensory neurons. Most individual mammalian cells express multiple distinct GPCR gene products, however, individual mammalian and insect olfactory neurons express only one functional odorant OR. By contrast C. elegans expresses multiple ORs and multiple Galpha subunits within each olfactory neuron. Here we show that, in addition to having at least one member of each of the four mammalian Galpha gene classes, C. elegans and other nematodes also possess two lineage-specific Galpha gene expansions, homologues of which are not found in any other organisms examined. We hypothesize that these novel nematode-specific Galpha genes increase the functional complexity of individual chemosensory neurons, enabling them to integrate odor signals from the multiple distinct ORs expressed on their membranes. This neuronal gene expansion most likely occurred in nematodes to enable them to compensate for the small number of chemosensory cells and the limited emphasis on cephalization during nematode evolution.

• Genome phylogenies indicate a meaningful alpha-proteobacterial phylogeny and support a grouping of the mitochondria with the Rickettsiales.

  Authors: David A Fitzpatrick, Christopher J Creevey, James O McInerney

  Molecular biology and evolution. 02/2006; 23(1):74-85.

  Placement of the mitochondrial branch on the tree of life has been problematic. Sparse sampling, the uncertainty of how lateral gene transfer might overwrite phylogenetic signals, and the uncertaintyPlacement of the mitochondrial branch on the tree of life has been problematic. Sparse sampling, the uncertainty of how lateral gene transfer might overwrite phylogenetic signals, and the uncertainty of phylogenetic inference have all contributed to the issue. Here we address this issue using a supertree approach and completed genomic sequences. We first determine that a sensible alpha-proteobacterial phylogenetic tree exists and that it can confidently be inferred using orthologous genes. We show that congruence across these orthologous gene trees is significantly better than might be expected by random chance. There is some evidence of horizontal gene transfer within the alpha-proteobacteria, but it appears to be restricted to a minority of genes ( approximately 23%) most of whom ( approximately 74%) can be categorized as operational. This means that placement of the mitochondrion should not be excessively hampered by interspecies gene transfer. We then show that there is a consistently strong signal for placement of the mitochondrion on this tree and that this placement is relatively insensitive to methodological approach or data set. A concatenated alignment was created consisting of 15 mitochondrion-encoded proteins that are unlikely to have undergone any lateral gene transfer in the timeline under consideration. This alignment infers that the sister group of the mitochondria, for the taxa that have been sampled, is the order Rickettsiales.

• Multiple lineage specific expansions within the guanylyl cyclase gene family.

  Authors: David A Fitzpatrick, Damien M O'Halloran, Ann M Burnell

  BMC evolutionary biology. 02/2006; 6:26.

  BACKGROUND: Guanylyl cyclases (GCs) are responsible for the production of the secondary messenger cyclic guanosine monophosphate, which plays important roles in a variety of physiological responsesBACKGROUND: Guanylyl cyclases (GCs) are responsible for the production of the secondary messenger cyclic guanosine monophosphate, which plays important roles in a variety of physiological responses such as vision, olfaction, muscle contraction, homeostatic regulation, cardiovascular and nervous function. There are two types of GCs in animals, soluble (sGCs) which are found ubiquitously in cell cytoplasm, and receptor (rGC) forms which span cell membranes. The complete genomes of several vertebrate and invertebrate species are now available. These data provide a platform to investigate the evolution of GCs across a diverse range of animal phyla. RESULTS: In this analysis we located GC genes from a broad spectrum of vertebrate and invertebrate animals and reconstructed molecular phylogenies for both sGC and rGC proteins. The most notable features of the resulting phylogenies are the number of lineage specific rGC and sGC expansions that have occurred during metazoan evolution. Among these expansions is a large nematode specific rGC clade comprising 21 genes in C. elegans alone; a vertebrate specific expansion in the natriuretic receptors GC-A and GC-B; a vertebrate specific expansion in the guanylyl GC-C receptors, an echinoderm specific expansion in the sperm rGC genes and a nematode specific sGC clade. Our phylogenetic reconstruction also shows the existence of a basal group of nitric oxide (NO) insensitive insect and nematode sGCs which are regulated by O2. This suggests that the primordial eukaryotes probably utilized sGC as an O2 sensor, with the ligand specificity of sGC later switching to NO which provides a very effective local cell-to-cell signalling system. Phylogenetic analysis of the sGC and bacterial heme nitric oxide/oxygen binding protein domain supports the hypothesis that this domain originated from a cyanobacterial source. CONCLUSION: The most salient feature of our phylogenies is the number of lineage specific expansions, which have occurred within the GC gene family during metazoan evolution. Our phylogenetic analyses reveal that the rGC and sGC multi-domain proteins evolved early in eumetazoan evolution. Subsequent gene duplications, tissue specific expression patterns and lineage specific expansions resulted in the evolution of new networks of interaction and new biological functions associated with the maintenance of organismal complexity and homeostasis.

• A fungal phylogeny based on 42 complete genomes derived from supertree and combined gene analysis.

  Authors: David A Fitzpatrick, Mary E Logue, Jason E Stajich, Geraldine Butler

  BMC evolutionary biology. 02/2006; 6:99.

  BACKGROUND: To date, most fungal phylogenies have been derived from single gene comparisons, or from concatenated alignments of a small number of genes. The increase in fungal genome sequencingBACKGROUND: To date, most fungal phylogenies have been derived from single gene comparisons, or from concatenated alignments of a small number of genes. The increase in fungal genome sequencing presents an opportunity to reconstruct evolutionary events using entire genomes. As a tool for future comparative, phylogenomic and phylogenetic studies, we used both supertrees and concatenated alignments to infer relationships between 42 species of fungi for which complete genome sequences are available. RESULTS: A dataset of 345,829 genes was extracted from 42 publicly available fungal genomes. Supertree methods were employed to derive phylogenies from 4,805 single gene families. We found that the average consensus supertree method may suffer from long-branch attraction artifacts, while matrix representation with parsimony (MRP) appears to be immune from these. A genome phylogeny was also reconstructed from a concatenated alignment of 153 universally distributed orthologs. Our MRP supertree and concatenated phylogeny are highly congruent. Within the Ascomycota, the sub-phyla Pezizomycotina and Saccharomycotina were resolved. Both phylogenies infer that the Leotiomycetes are the closest sister group to the Sordariomycetes. There is some ambiguity regarding the placement of Stagonospora nodurum, the sole member of the class Dothideomycetes present in the dataset.Within the Saccharomycotina, a monophyletic clade containing organisms that translate CTG as serine instead of leucine is evident. There is also strong support for two groups within the CTG clade, one containing the fully sexual species Candida lusitaniae, Candida guilliermondii and Debaryomyces hansenii, and the second group containing Candida albicans, Candida dubliniensis, Candida tropicalis, Candida parapsilosis and Lodderomyces elongisporus. The second major clade within the Saccharomycotina contains species whose genomes have undergone a whole genome duplication (WGD), and their close relatives. We could not confidently resolve whether Candida glabrata or Saccharomyces castellii lies at the base of the WGD clade. CONCLUSION: We have constructed robust phylogenies for fungi based on whole genome analysis. Overall, our phylogenies provide strong support for the classification of phyla, sub-phyla, classes and orders. We have resolved the relationship of the classes Leotiomyctes and Sordariomycetes, and have identified two classes within the CTG clade of the Saccharomycotina that may correlate with sexual status.

• Evidence of positive Darwinian selection in putative meningococcal vaccine antigens.

  Authors: David A Fitzpatrick, Christopher J Creevey, James O McInerney

  Journal of molecular evolution. 08/2005; 61(1):90-8.

  Meningococcal meningitidis is a life-threatening disease. In Europe and the United States the majority of cases are caused by virulent meningococcal strains belonging to serogroup B. Presently thereMeningococcal meningitidis is a life-threatening disease. In Europe and the United States the majority of cases are caused by virulent meningococcal strains belonging to serogroup B. Presently there is no effective vaccine against serogroup B strains, as traditional vaccine antigens such as polysaccharide capsules are unusable as they lead to autoimmunity. The year 2000 saw the publication of the complete genome of Neisseria meningitidis MC58, a virulent serogroup B bacterium. Working in conjunction with the sequencing project, researchers endeavored to locate highly conserved membrane-associated proteins that elicit an immune response. It is hoped that these proteins will provide a basis for novel vaccines against serogroup B strains. A number of potential vaccine antigens have been located and are presently in phase I clinical trials. Recently many reports pertaining to the evidence of positive Darwinian selection in membrane proteins of pathogens have been reported. This study utilized in silico methods to test for evidence of historical positive Darwinian selection in seven such vaccine candidates. We found that two of these proteins show signatures of adaptive evolution, while the remaining proteins show evidence of strong purifying selection. This has significant implications for the design of a vaccine against serogroup B strains, as it has been shown that vaccines that target epitopes that are under strong purifying selection are better than those that target variable epitopes.

• The anhydrobiotic potential and molecular phylogenetics of species and strains of Panagrolaimus (Nematoda, Panagrolaimidae).

  Authors: Adam J Shannon, John A Browne, Jacqueline Boyd, David A Fitzpatrick, Ann M Burnell

  The Journal of experimental biology. 07/2005; 208(Pt 12):2433-45.

  Members of the genus Panagrolaimus are bacterial-feeding nematodes that occupy a diversity of niches ranging from Antarctic and temperate soils to terrestrial mosses. Some members of this genus areMembers of the genus Panagrolaimus are bacterial-feeding nematodes that occupy a diversity of niches ranging from Antarctic and temperate soils to terrestrial mosses. Some members of this genus are able to survive extreme desiccation by entering into a state of suspended animation known as anhydrobiosis. We have assembled a collection of Panagrolaimus species and strains and have investigated their anhydrobiotic phenotypes. Our data show that within the genus Panagrolaimus there is a continuum of strains ranging from those unable to survive exposure to low relative humidity (RH) without prior preconditioning at high RH (slow desiccation strategists), through strains that have limited ability to survive rapid desiccation but whose anhydrobiotic ability improves upon preconditioning, to strains such as P. superbus that can readily survive immediate exposure to severe desiccation (fast desiccation strategists). Using this panel of nematodes we investigated the effect of preincubation at high RH on the accumulation of trehalose and on the nematodes' anhydrobiotic potential. We found that there is a strong correlation between trehalose induction and anhydrobiotic survival in Panagrolaimus. Furthermore, the high trehalose levels observed in fully hydrated P. superbus (10% dry mass) suggest that constitutive expression of trehalose pre-adapts this fast dehydration strategist to combat desiccation. All the strains observed, regardless of survival rates, undertook both coiling and clumping, which has the effect of reducing surface area and slowing the rate of water loss during desiccation. Phylogenetic analyses were carried out to investigate whether the observed anhydrobiotic phenotypes were the result of convergent evolution or represented a single phylogenetic lineage. These analyses, derived from alignments of the rDNA ITS and D3 sequences, indicate that the strongly anhydrobiotic strains of Panagrolaimus form a single phylogenetic lineage, which is separate from the weakly anhydrobiotic strains. The weakly anhydrobiotic strains are also phylogenetically divergent from each other. Our data indicate that Panagrolaimus has the potential to be an excellent model system for the investigation of molecular aspects of nematode anhydrobiosis.

• Evidence of positive Darwinian selection in Omp85, a highly conserved bacterial outer membrane protein essential for cell viability.

  Authors: David A Fitzpatrick, James O McInerney

  Journal of molecular evolution. 03/2005; 60(2):268-73.

  Omp85 is a highly conserved outer membrane protein found in all gram-negative bacteria. It is essential for bacterial cell viability and plays an integral function in the positioning and folding ofOmp85 is a highly conserved outer membrane protein found in all gram-negative bacteria. It is essential for bacterial cell viability and plays an integral function in the positioning and folding of other outer membrane proteins into the bacterial outer membrane. We have employed a maximum likelihood and a maximum parsimony approach to detect evidence of positive Darwinian selection in Omp85 homologues from 10 delta-proteobacteria and have identified 14 amino acid sites that show evidence of being under the influence of adaptive evolution. Interestingly all sites bar one are concentrated within surface loops of the protein that most likely interact with host immune response or the surrounding environment. Alternatively amino acids within membrane-spanning regions of the protein are found to be under purifying selection most likely as a result of structural constraints.

• Does a tree-like phylogeny only exist at the tips in the prokaryotes?

  Authors: Christopher J Creevey, David A Fitzpatrick, Gayle K Philip, Rhoda J Kinsella, Mary J O'Connell, Melissa M Pentony, Simon A Travers, Mark Wilkinson, James O McInerney

  Proceedings. Biological sciences / The Royal Society. 01/2005; 271(1557):2551-8.

  The extent to which prokaryotic evolution has been influenced by horizontal gene transfer (HGT) and therefore might be more of a network than a tree is unclear. Here we use supertree methods to askThe extent to which prokaryotic evolution has been influenced by horizontal gene transfer (HGT) and therefore might be more of a network than a tree is unclear. Here we use supertree methods to ask whether a definitive prokaryotic phylogenetic tree exists and whether it can be confidently inferred using orthologous genes. We analysed an 11-taxon dataset spanning the deepest divisions of prokaryotic relationships, a 10-taxon dataset spanning the relatively recent gamma-proteobacteria and a 61-taxon dataset spanning both, using species for which complete genomes are available. Congruence among gene trees spanning deep relationships is not better than random. By contrast, a strong, almost perfect phylogenetic signal exists in gamma-proteobacterial genes. Deep-level prokaryotic relationships are difficult to infer because of signal erosion, systematic bias, hidden paralogy and/or HGT. Our results do not preclude levels of HGT that would be inconsistent with the notion of a prokaryotic phylogeny. This approach will help decide the extent to which we can say that there is a prokaryotic phylogeny and where in the phylogeny a cohesive genomic signal exists.

• Fatty acid biosynthesis in Mycobacterium tuberculosis: lateral gene transfer, adaptive evolution, and gene duplication.

  Authors: Rhoda J Kinsella, David A Fitzpatrick, Christopher J Creevey, James O McInerney

  Proceedings of the National Academy of Sciences of the United States of America. 10/2003; 100(18):10320-5.

  Mycobacterium tuberculosis is a high GC Gram-positive member of the actinobacteria. The mycobacterial cell wall is composed of a complex assortment of lipids and is the interface between theMycobacterium tuberculosis is a high GC Gram-positive member of the actinobacteria. The mycobacterial cell wall is composed of a complex assortment of lipids and is the interface between the bacterium and its environment. The biosynthesis of fatty acids plays an essential role in the formation of cell wall components, in particular mycolic acids, which have been targeted by many of the drugs used to treat M. tuberculosis infection. M. tuberculosis has approximately 250 genes involved in fatty acid metabolism, a much higher proportion than in any other organism. In silico methods have been used to compare the genome of M. tuberculosis CDC1551 to a database of 58 complete bacterial genomes. The resulting alignments were scanned for genes specifically involved in fatty acid biosynthetic pathway I. Phylogenetic analysis of these alignments was used to investigate horizontal gene transfer, gene duplication, and adaptive evolution. It was found that of the eight gene families examined, five of the phylogenies reconstructed suggest that the actinobacteria have a closer relationship with the alpha-proteobacteria than expected. This is either due to either an ancient transfer of genes or deep paralogy and subsequent retention of the genes in unrelated lineages. Additionally, adaptive evolution and gene duplication have been an influence in the evolution of the pathway. This study provides a key insight into how M. tuberculosis has developed its unique fatty acid synthetic abilities.