Branches Of Phylogenetic Tree Research Articles

A phylogenomic and molecular markers based analysis of the phylum Chlamydiae: proposal to divide the class Chlamydiia into two orders, Chlamydiales and Parachlamydiales ord. nov., and emended description of the class Chlamydiia.

The phylum Chlamydiae contains nine ecologically and genetically diverse families all placed within a single order. In this work, we have completed a comprehensive comparative analysis of 36 sequenced Chlamydiae genomes in order to identify shared molecular characteristics, namely conserved signature insertions/deletions (CSIs) and conserved signature proteins (CSPs), which can serve as distinguishing characteristics of supra-familial clusters within the phylum Chlamydiae. Our analysis has led to the identification of 32 CSIs which are specific to clusters within the phylum Chlamydiae at various phylogenetic depths. Importantly, 17 CSIs and 98 CSPs were found to be specific for the family Chlamydiaceae while another 3 CSI variants and 15 CSPs were specific for a grouping of the families Criblamydiaceae, Parachlamydiaceae, Simkaniaceae and Waddliaceae. These two clusters were also found to be distinguishable in 16S rRNA based phylogenetic trees, concatenated protein based phylogenetic trees, character compatibility based phylogenetic analyses, and on the basis of 16S rRNA gene sequence identity and average amino acid identity values. On the basis of the identified molecular characteristics, branching in phylogenetic trees, and the genetic distance between the two clusters within the phylum Chlamydiae we propose a division of the class Chlamydiia into two orders: an emended order Chlamydiales, containing the family Chlamydiaceae and the closely related Candidatus family Clavichlamydiaceae, and the novel order Parachlamydiales ord. nov. containing the families Parachlamydiaceae, Simkaniaceae and Waddliaceae and the Candidatus families Criblamydiaceae, Parilichlamydiaceae, Piscichlamydiaceae, and Rhabdochlamydiaceae. We also include a brief discussion of the reunification of the genera Chlamydia and Chlamydophila.

Evidence for the presence of key chlorophyll-biosynthesis-related proteins in the genus Rubrobacter (Phylum Actinobacteria) and its implications for the evolution and origin of photosynthesis.

Homologs showing high degree of sequence similarity to the three subunits of the protochlorophyllide oxidoreductase enzyme complex (viz. BchL, BchN, and BchB), which carries out a central role in chlorophyll-bacteriochlorophyll (Bchl) biosynthesis, are uniquely found in photosynthetic organisms. The results of BLAST searches and homology modeling presented here show that proteins exhibiting a high degree of sequence and structural similarity to the BchB and BchN proteins are also present in organisms from the high G+C Gram-positive phylum of Actinobacteria, specifically in members of the genus Rubrobacter (R. x ylanophilus and R. r adiotolerans). The results presented exclude the possibility that the observed BLAST hits are for subunits of the nitrogenase complex or the chlorin reductase complex. The branching in phylogenetic trees and the sequence characteristics of the Rubrobacter BchB/BchN homologs indicate that these homologs are distinct from those found in other photosynthetic bacteria and that they may represent ancestral forms of the BchB/BchN proteins. Although a homolog showing high degree of sequence similarity to the BchL protein was not detected in Rubrobacter, another protein, belonging to the ParA/Soj/MinD family, present in these bacteria, exhibits high degree of structural similarity to the BchL. In addition to the BchB/BchN homologs, Rubrobacter species also contain homologs showing high degree of sequence similarity to different subunits of magnesium chelatase (BchD, BchH, and BchI) as well as proteins showing significant similarity to the BchP and BchG proteins. Interestingly, no homologs corresponding to the BchX, BchY, and BchZ proteins were detected in the Rubrobacter species. These results provide the first suggestive evidence that some form of photosynthesis either exists or was anciently present within the phylum Actinobacteria (high G+C Gram-positive) in members of the genus Rubrobacter. The significance of these results concerning the origin of the Bchl-based photosynthesis is also discussed.

HIV-1 subtype diversity and transmission clusters among men having sex with men who recently got HIV-l infection, in Zhejiang province

To understand the HIV-1 subtype diversity and transmission characteristics in men having sex with men (MSM) in Zhejiang province. A total of 233 newly diagnosed as HIV-1 positive patients in 2011 were screened out by BED capture enzyme immunoassay (BED-CEIA). Among them, 107 eligible subjects were enrolled for further molecular epidemiological study. Viral RNA was extracted from plasma samples and followed by reverse transcription PCR and nested PCR for amplification of pol gene fragments, sequencing, and bioinformatics analysis. There were no statistically significant differences regarding the social demographic distribution between the subjects under study and those recently infected MSM population. The rate of success for sequence acquisition was 94.4% (101/107). The highest proportion of subtype was CRF01_AE (62.4%), followed by CRF07_BC (31.7%) and with three cases of subtype B, one case of CRF55_01B and two cases of unique recombinant form (CRF01_AE/B and CRF01_AE/CRF07_BC). The phylogenetic trees were mainly divided into CRF01_AE cluster 1, cluster 2 and CRF07_BC cluster 3. The strains located in Hangzhou were diffused in the branches of phylogenetic tree. 10 transmission clusters were found, in which 80% involved two or more regions and 90% was associated with patients residing in Hangzhou. Three surveillance drug resistance mutations (M46I, T215S and G190A) were found in three samples (each sample harbored only one resistance mutation). The overall rate of transmitted drug resistance (TDR) was 2.97%. The increasing complexity of HIV was noticed in MSM in Zhejiang province. However, the prevalence of TDR was low. Cross-regional HIV transmission in MSM was common, which inferred from the study. Hangzhou might play a central regional role in the intra-provincial spread of HIV, to form an interwoven complex network in the MSM population.

Phylogenomic and molecular demarcation of the core members of the polyphyletic pasteurellaceae genera actinobacillus, haemophilus, and pasteurella.

The genera Actinobacillus, Haemophilus, and Pasteurella exhibit extensive polyphyletic branching in phylogenetic trees and do not represent coherent clusters of species. In this study, we have utilized molecular signatures identified through comparative genomic analyses in conjunction with genome based and multilocus sequence based phylogenetic analyses to clarify the phylogenetic and taxonomic boundary of these genera. We have identified large clusters of Actinobacillus, Haemophilus, and Pasteurella species which represent the “sensu stricto” members of these genera. We have identified 3, 7, and 6 conserved signature indels (CSIs), which are specifically shared by sensu stricto members of Actinobacillus, Haemophilus, and Pasteurella, respectively. We have also identified two different sets of CSIs that are unique characteristics of the pathogen containing genera Aggregatibacter and Mannheimia, respectively. It is now possible to demarcate the genera Actinobacillus sensu stricto, Haemophilus sensu stricto, and Pasteurella sensu stricto on the basis of discrete molecular signatures. The other members of the genera Actinobacillus, Haemophilus, and Pasteurella that do not fall within the “sensu stricto” clades and do not contain these molecular signatures should be reclassified as other genera. The CSIs identified here also provide useful diagnostic targets for the identification of current and novel members of the indicated genera.

Time and Knowability in Evolutionary Processes

Historical sciences like evolutionary biology reconstruct past events by using the traces that the past has bequeathed to the present. Markov chain theory entails that the passage of time reduces the amount of information that the present provides about the past. Here we use a Moran process framework to show that some evolutionary processes destroy information faster than others. Our results connect with Darwin’s principle that adaptive similarities provide scant evidence of common ancestry whereas neutral and deleterious similarities do better. We also describe how the branching in phylogenetic trees affects the information that the present supplies about the past.

Rapid Evolution of the Cerebellum in Humans and Other Great Apes

Humans' unique cognitive abilities are usually attributed to a greatly expanded neocortex, which has been described as "the crowning achievement of evolution and the biological substrate of human mental prowess". The human cerebellum, however, contains four times more neurons than the neocortex and is attracting increasing attention for its wide range of cognitive functions. Using a method for detecting evolutionary rate changes along the branches of phylogenetic trees, we show that the cerebellum underwent rapid size increase throughout the evolution of apes, including humans, expanding significantly faster than predicted by the change in neocortex size. As a result, humans and other apes deviated significantly from the general evolutionary trend for neocortex and cerebellum to change in tandem, having significantly larger cerebella relative to neocortex size than other anthropoid primates. These results suggest that cerebellar specialization was a far more important component of human brain evolution than hitherto recognized and that technical intelligence was likely to have been at least as important as social intelligence in human cognitive evolution. Given the role of the cerebellum in sensory-motor control and in learning complex action sequences, cerebellar specialization is likely to have underpinned the evolution of humans' advanced technological capacities, which in turn may have been a preadaptation for language.

Adaptive evolution and environmental durability jointly structure phylodynamic patterns in avian influenza viruses.

Author SummaryHuman populations have experienced several pandemics involving new subtypes of influenza virus over the past century. All of these pandemic strains contained gene segments that originated in wild birds, a host pool that supports a very large and genetically diverse array of influenza viruses. However, once an avian strain establishes itself within the human population, the genetic diversity of the resulting human subtypes is typically quite low compared to that of their avian counterparts. Here we compare the evolutionary dynamics of human and avian influenza viruses in North America and test different hypotheses that might explain these two contrasting evolutionary patterns. Our analysis shows that a combination of characteristics of the host (especially demography) and virus (such as durability in water and mutability) explains the diversity observed. Using a theoretical model, we show that the combination of the short lifespan of wild birds, and greater durability of viruses in aquatic environments, is key to maintaining the high levels of influenza diversity observed in wild birds.

Intrafamilial Transmission of Vaccinia virus during a Bovine Vaccinia Outbreak in Brazil: A New Insight in Viral Transmission Chain

Bovine vaccinia (BV) is an emerging zoonosis caused by the Vaccinia virus (VACV), genus Orthopoxvirus (OPV), Poxviridae family. In general, human cases are related to direct contact with sick cattle but there is a lack of information about human-to-human transmission of VACV during BV outbreaks. In this study, we epidemiologically and molecularly show a case of VACV transmission between humans in São Francisco de Itabapoana County, Rio de Janeiro state. Our group collected samples from the patients, a 49-year-old patient and his son. Our results showed that patients had developed anti-OPV IgG or IgM antibodies and presented neutralizing antibodies against OPV. The VACV isolates displayed high identity (99.9%) and were grouped in the same phylogenetic tree branch. Our data indicate that human-to-human VACV transmission occurred during a BV outbreak, raising new questions about the risk factors of the VACV transmission chain.

Automatic Detection of Key Innovations, Rate Shifts, and Diversity-Dependence on Phylogenetic Trees

A number of methods have been developed to infer differential rates of species diversification through time and among clades using time-calibrated phylogenetic trees. However, we lack a general framework that can delineate and quantify heterogeneous mixtures of dynamic processes within single phylogenies. I developed a method that can identify arbitrary numbers of time-varying diversification processes on phylogenies without specifying their locations in advance. The method uses reversible-jump Markov Chain Monte Carlo to move between model subspaces that vary in the number of distinct diversification regimes. The model assumes that changes in evolutionary regimes occur across the branches of phylogenetic trees under a compound Poisson process and explicitly accounts for rate variation through time and among lineages. Using simulated datasets, I demonstrate that the method can be used to quantify complex mixtures of time-dependent, diversity-dependent, and constant-rate diversification processes. I compared the performance of the method to the MEDUSA model of rate variation among lineages. As an empirical example, I analyzed the history of speciation and extinction during the radiation of modern whales. The method described here will greatly facilitate the exploration of macroevolutionary dynamics across large phylogenetic trees, which may have been shaped by heterogeneous mixtures of distinct evolutionary processes.

Conserved signature indels and signature proteins as novel tools for understanding microbial phylogeny and systematics: identification of molecular signatures that are specific for the phytopathogenic genera Dickeya, Pectobacterium and Brenneria

Genome sequences are enabling applications of different approaches to more clearly understand microbial phylogeny and systematics. Two of these approaches involve identification of conserved signature indels (CSIs) and conserved signature proteins (CSPs) that are specific for different lineages. These molecular markers provide novel and more definitive means for demarcation of prokaryotic taxa and for identification of species from these groups. Genome sequences are also enabling determination of phylogenetic relationships among species based upon sequences for multiple proteins. In this work, we have used all of these approaches for studying the phytopathogenic bacteria belonging to the genera Dickeya, Pectobacterium and Brenneria. Members of these genera, which cause numerous diseases in important food crops and ornamental plants, are presently distinguished mainly on the basis of their branching in phylogenetic trees. No biochemical or molecular characteristic is known that is uniquely shared by species from these genera. Hence, detailed studies using the above approaches were carried out on proteins from the genomes of these bacteria to identify molecular markers that are specific for them. In phylogenetic trees based upon concatenated sequences for 23 conserved proteins, members of the genera Dickeya, Pectobacterium and Brenneria formed a strongly supported clade within the other Enterobacteriales. Comparative analysis of protein sequences from the Dickeya, Pectobacterium and Brenneria genomes has identified 10 CSIs and five CSPs that are either uniquely or largely found in all genome-sequenced species from these genera, but not present in any other bacteria in the database. In addition, our analyses have identified 10 CSIs and 17 CSPs that are specifically present in either all or most sequenced Dickeya species/strains, and six CSIs and 19 CSPs that are uniquely found in the sequenced Pectobacterium genomes. Finally, our analysis also identified three CSIs and one CSP that are specifically shared by members of the genera Pectobacterium and Brenneria, but absent in species of the genus Dickeya, indicating that the former two genera shared a common ancestor exclusive of Dickeya. The identified CSIs and CSPs provide novel tools for identification of members of the genera Dickeya and Pectobacterium and for delimiting these taxa in molecular terms. Descriptions of the genera Dickeya and Pectobacterium have been revised to provide information for these molecular markers. Biochemical studies on these CSIs and CSPs, which are specific for these genera, may lead to discovery of novel properties that are unique to these bacteria and which could be targeted to develop antibacterial agents that are specific for these plant-pathogenic bacteria.

猪瘟病毒编码区基因选择压力及重组分析

We detected the selection pressure and recombination of all 50 strains Classical Swine Fever Virus (CSFV) available complete coding region, based on the maximum likelihood estimation and six algorithms within Recombinant Detect Program 3.44, to provide the first complete overview of the evolution of CSFV. The results showed the overall <italic>ω </italic>(nonsynonymous/synonymous) of 12 proteins under one-radio model were all lower than 1, indicating that the selection pressure of CSFV was subject to strong purifying selection. Among the 12 proteins, NS3, NS4A and NS4B proteins underwent the stronger purifying selection; meanwhile Npro, E^rns, E1, E2 and NS5A proteins were detected the positive selection sites, underwent the stonger positive selection, indicating that the adaptive evolution of CSFV were related to the host immune response. Furthermore, 4 putative recombination virus strains, and the newly found HCLV/99, ALD and SWH, were detected in 50 sample sequences by recombination analysis. The proteins related to the genome recombination were involved in the immunity and virulence of CSFV, and the recombination changed the phylogenetic tree branch of recombination strain. However, the recombination loci were detected in most proteins except NS3, NS4A and NS4B proteins which processed the shorter genetic distance. Our research suggested that the mutation and recombination under the selection pressure of the host immune system promoted the evolution of coding region in CSFV. These studies would provide the foundation for vaccine targets and the more and more complex situation of CSFV epidemiological research.

Analysis of genetic variation of hemagglutinin and three internal genes of influenza B virus isolated in Zhejiang province from 1999 to 2012

To explore the characteristics of the genetic variation of hemagglutinin( HA) and three internal genes coding for the nucleoprotein ( NP) , matrix protein ( M) and nonstructural protein ( NS) of influenza B virus. A total of 31 strains of influenza B virus were isolated in Zhejiang province from 1999 to 2012, and then were amplified and sequenced the genes of HAl , NP, M and NS. The phylogenetic tree was constructed, the nucleotide substitution rate of the above individual gene was estimated and the variation sites of amino acids were analyzed. The 31 isolated strains of influenza B virus were divided into two distinct lineages Victoria and Yamagata in the phylogenetic tree of HAl gene,represented by B/Victoria/2/87 and B/Yamagata/16/88. Phylogenetic analysis of the NP gene showed that the NP gene of Victoria-like influenza B strains which were isolated after 2010 was highly homologous with Yamagata-like isolates, and thereby they were found to be on the same branch of the phylogenetic tree of the NP gene. Nucleotide substitution rates of HAl , NP, M and NS genes were estimated to be 2. 29 x 10 -3 ,1. 39 X 10-3 ,1. 78 X 10-3 ,1. 30 X 10-3 /site per year, respectively. Variations of amino acid of HAl domain of Victoria-like isolates mainly included K48E ,L58P ,N75K,K80R,K129N/S,N165K,S172P ,Sl97N/D and A202V; while those in Yamagata-like isolates were R48K, S1501, N166Y, N203S, G230D and D233N. Determined amino acid sequences of NP of Victoria-like influenza B isolates were similar to Yamagata-like isolates after 2010 and variations happened on four characteristic amino acid sites, naming A60D, I233V, N513S and V5341, compared with previous Victoria-like influenza B isolates. Significant variation was found among influenza B strains isolated in Zhejiang province from 1999 to 2012. The surface HAl gene evolved more rapidly than internal genes. Gene reassortment and gene mutation were the main evolutionary mechanism of influenza B virus.

Sphaerospora sensu stricto: Taxonomy, diversity and evolution of a unique lineage of myxosporeans (Myxozoa)

Myxosporeans (Myxozoa) are eukaryotic parasites, primarily of fish, whose classification is in a state of flux as taxonomists attempt to synthesize the traditional morphology-based system with emerging DNA sequence-based phylogenies. The genus Sphaerospora Thélohan, 1892, which includes pathogenic species that cause significant impacts on fisheries and aquaculture, is one of the most polyphyletic taxa and exemplifies the current challenges facing myxozoan taxonomists. The type species, S. elegans, clusters within the Sphaerospora sensu stricto clade, members of which share similar tissue tropism and long insertions in their variable rRNA gene regions. However, other morphologically similar sphaerosporids lie in different branches of myxozoan phylogenetic trees. Herein, we significantly extend taxonomic sampling of sphaerosporids with SSU+LSU rDNA and EF-2 sequence data for 12 taxa including three representatives of the morphologically similar genus Polysporoplasma Sitjà-Bobadilla et Álvarez-Pellitero, 1995. These taxa were sampled from different vertebrate host groups, biogeographic realms and environments. Our phylogenetic analyses and statistical tests of single and concatenated datasets revealed Sphaerospora s. s. as a strongly supported monophyletic lineage, that clustered sister to the whole myxosporean clade (freshwater+marine lineages). Generally, Sphaerospora s. s. rDNA sequences (up to 3.7kb) are the longest of all myxozoans and indeed metazoans. The sphaerosporid clade has two lineages, which have specific morphological, biological and sequence traits. Lineage A taxa (marine Sphaerospora spp.) have a single binucleate sporoplasm and shorter AT-rich rDNA inserts. Lineage B taxa (freshwater/brackish Sphaerospora spp.+marine/brackish Polysporoplasma spp.) have 2–12 uninucleate sporoplasms and longer GC-rich rDNA inserts. Lineage B has four subclades that correlate with host group and habitat; all Polysporoplasma species, including the type species, cluster together in one of these subclades. We thus suppress the genus Polysporoplasma and the family Polysporoplasmidae and emend the generic diagnosis of the genus Sphaerospora. The combination of morphological, biological and DNA sequence data applied in this study helped to elucidate an important part of the taxonomic puzzle within the phylum Myxozoa.

Phylogenetic relationship among some groups of orthopteran based on complete sequences of the mitochondrial &lt;I&gt;COI&lt;/I&gt; gene

Complete sequences of the COI gene were used to reconstruct the phylogenetic relationship among 56 species from Orthoptera. We also analyzed the reliability of Orthoptera phylogenetic relationship using translated amino acid sequences of the COI genes. The COI sequences were divided into three data sets on the basis of different codon positions to calculate the Partitioned Bremer support (PBS), and to test the phylogenetic signal in different codon positions of protein-coding genes. The result supports the monophyly of Caelifera and Ensifera; but the monophyly of Acrididae, Catantopidae, Oedipodidae, Arcypteridae and Gomphoceridae are not supported. The P-distances among families vary from 0.107 to 0.153, which are smaller than those of other families, being consist with the classification that these 5 families should be merged into one family (Acrididae). Chrotogonidae and Pyrgomorphidae belong to the superfamily Pyrgomorphoidea. Pamphagidae should be a family alone consistent with Otte's taxonomic system. According to the PBS values, the 3rd and 1st codon positions contribution more for the Phylogenetic tree branches than the 2nd, and longer sequences contain more informative sites. We further demonstrated that it is feasible for phylogenetic studies at family level to use the genetic distances among COI sequences from different species of Orthopera.

The Revised Classification of Eukaryotes

This revision of the classification of eukaryotes, which updates that of Adl et al. [J. Eukaryot. Microbiol. 52 (2005) 399], retains an emphasis on the protists and incorporates changes since 2005 that have resolved nodes and branches in phylogenetic trees. Whereas the previous revision was successful in re-introducing name stability to the classification, this revision provides a classification for lineages that were then still unresolved. The supergroups have withstood phylogenetic hypothesis testing with some modifications, but despite some progress, problematic nodes at the base of the eukaryotic tree still remain to be statistically resolved. Looking forward, subsequent transformations to our understanding of the diversity of life will be from the discovery of novel lineages in previously under-sampled areas and from environmental genomic information.

A genome sequence-based approach to taxonomy of the genus Nocardia

The genus Nocardia includes both pathogens and producers of useful secondary metabolites. Although 16S rRNA analysis is required to accurately discriminate among phylogenetic relationships of the Nocardia species, most branches of 16S rRNA-based phylogenetic trees are not reliable. In this study, we performed in silico analyses of the genome sequences of Nocardia species in order to understand their diversity and classification for their identification and applications. Draft genome sequences of 26 Nocardia strains were determined. Phylogenetic trees were prepared on the basis of multilocus sequence analysis of the concatenated sequences of 12 genes (atpD-dnaJ-groL1-groL2-gyrB-recA-rpoA-secA-secY-sodA-trpB-ychF) and a bidirectional best hit. To elucidate the evolutionary relationships of these genes, the genome-to-genome distance was investigated on the basis of the average nucleotide identity, DNA maximal unique matches index, and genome-to-genome distance calculator. The topologies of all phylogenetic trees were found to be essentially similar to each other. Furthermore, whole genome-derived and multiple gene-derived relationships were found to be suitable for extensive intra-genus assessment of the genus Nocardia.

Genetic Variation and Differentiation ofGekko geckofrom Different Populations Based on Mitochondrial CytochromebGene Sequences and Karyotypes

Black-spotted and red-spotted tokay geckos are distributed in different regions and have significant differences in morphological appearance, but have been regarded as the same species, Gekko gecko, in taxonomy. To determine whether black-spotted and red-spotted tokay geckos are genetically differentiated, we sequenced the entire mitochondrial cytochrome b gene (1147 bp) from 110 individuals of Gekko gecko collected in 11 areas including Guangxi China, Yunnan China, Vietnam, and Laos. In addition, we performed karyotypic analyses of black-spotted tokay geckos from Guangxi China and red-spotted tokay geckos from Laos. These phylogenetic analyses showed that black-spotted and red-spotted tokay geckos are divided into two branches in molecular phylogenetic trees. The average genetic distances are as follows: 0.12-0.47% among six haplotypes in the black-spotted tokay gecko group, 0.12-1.66% among five haplotypes in the red-spotted tokay gecko group, and 8.76-9.18% between the black-spotted and red-spotted tokay geckos, respectively. The karyotypic analyses showed that the karyotype formula is 2n = 38 = 8m + 2sm + 2st + 26t in red-spotted tokay geckos from Laos compared with 2n = 38 = 8m + 2sm + 28t in black-spotted tokay geckos from Guangxi China. The differences in these two kinds of karyotypes were detected on the 15th chromosome. The clear differences in genetic levels between black-spotted and red-spotted tokay geckos suggest a significant level of genetic differentiation between the two.

Near full-lenght genome characterization of a newly derived unique recombinant form AG HIV-1 circulating in Siberia

Background Genotyping of HIV-1 variants isolated from patients of Siberian region (Russia) in 2006-2010 revealed a sharp increase of 02_AG recombinant variant proportion among HIV-1 genetic variants, circulating in the Siberia, from 2% in 2006 to 52% in 2010. The majority of HIV 02_AG variants isolated in Siberia form a separate branch of phylogenetic tree. The objective of this analysis was to study structure of the near full-length genome sequence of HIV-1 strains recombinant form AG rapidly spreading in Siberia. Material and methods Sample 10.RU.6637 was collected in 2009 from a 28year-old man, infected in 2008 through heterosexual contact. Three regions of the viral genome were independently amplified using nested PCR from the cDNA to contain a nearly full-length genome of HIV-1. Sequencing of all amplicons was performed by using cycle sequencing and dye termination on an automated sequencer, DNA sequences were assembled using Sequencher software. Jumping profile hidden Markov model (jpHMM) program was used to analyze the subtype assignment of all sequences retrieved. The software Simplot v3.5.1 was initially used to perform bootscanning analyses of a query sequence against a set of other sequences. Phylogenetic trees were constructed with the program PhyML v.3.0 using a maximum likelihood approach. Results Sequence 10.RU.6637 HIV-1 from Siberia was submitted to Genbank under accession number JN230353. Points of recombination were determined and uniqueness of 10. RU.6637 HIV-1 genome structure was shown through indepth genetic analysis. Recombinant breakpoint analysis of 10.RU.6637 HIV-1 genome sequence revealed that it was recombinant form between CRF 02_AG and subsubtype A1. We will designate this variant 02_AG/IDU A HIV-1, to reflect its close relationship to the CRF 02_AG and A1 (IDU A) subtype from Russia. Conclusion The majority of HIV 02_AG variants circulating in Siberia represented of a newly derived unique recombinant form 02_AG/IDU A HIV-1. Wider spread of HIV-1 URF02_AG/IDU A is possible in the territory of Russia in the coming years.

POSITIVE CORRELATION BETWEEN DIVERSIFICATION RATES AND PHENOTYPIC EVOLVABILITY CAN MIMIC PUNCTUATED EQUILIBRIUM ON MOLECULAR PHYLOGENIES

The hypothesis of punctuated equilibrium proposes that most phenotypic evolution occurs in rapid bursts associated with speciation events. Several methods have been developed that can infer punctuated equilibrium from molecular phylogenies in the absence of paleontological data. These methods essentially test whether the variance in phenotypes among extant species is better explained by evolutionary time since common ancestry or by the number of estimated speciation events separating taxa. However, apparent "punctuational" trait change can be recovered on molecular phylogenies if the rate of phenotypic evolution is correlated with the rate of speciation. Strong support for punctuational models can arise even if the underlying mode of trait evolution is strictly gradual, so long as rates of speciation and trait evolution covary across the branches of phylogenetic trees, and provided that lineages vary in their rate of speciation. Species selection for accelerated rates of ecological or phenotypic divergence can potentially lead to the perception that most trait divergence occurs in association with speciation events.

Abstract 964: Intra-tumor heterogeneity and Darwinian selection revealed by multi-region exome sequencing of renal cell carcinomas

Abstract Background: Genetic intra-tumor heterogeneity (ITH) may foster tumor adaptation by providing selectable phenotypes for Darwinian evolution. Extensive genetic heterogeneity may also hinder personalized medicine strategies that rely on the portrayal of the mutational landscape from single tumor biopsies. Methods: To examine ITH, we have subjected multiple biopsies from primary renal cell carcinomas and associated metastatic sites to exome capture sequencing (n=2), SNP-array based chromosomal aberration and ploidy profiling analysis (n=4). Phylogenetic relationships of tumor regions were reconstructed by clonal ordering of non-synonymous somatic mutations. The phenotypic consequences of genetic ITH were characterised by immunohistochemistry, mutation functional analysis and mRNA expression profiling. Results: Phylogenetic reconstruction revealed branched evolutionary tumor growth with 63-69% of somatic mutations identified from single biopsies not detectable across all sequenced tumor regions. Independent tumor suppressor gene loss of function mutations with distinct regional distributions were detected within individual tumors: SETD2 harbored 5 different mutations in 2 tumors and PTEN and KDM5C two different mutations in one tumor, each. Thus, despite genetic divergence during tumor progression, phenotypic convergent evolution occurred, indicating a high degree of early mutational diversity. ITH was observed for a mutation in the kinase domain of mTOR, correlating with S6 and 4EBP phosphorylation specifically in cancer regions carrying the mutation and constitutive activation of mTOR kinase activity in vitro. Expression of a renal cancer-specific prognostic signature differed between tumor regions. Chromosomal aberration analysis revealed extensive ITH with 26/30 tumor biopsies from four tumors harboring divergent allelic imbalance profiles. Ploidy profiling revealed heterogeneity in two out of four tumors and identified an aneuploid tumor cell population in a metastasis that probably evolved from a tetraploid intermediate detectable in the primary tumor. Conclusions: Genetic ITH was present in all tumors and occurs through spatially separated heterogeneous somatic mutations and chromosomal and ploidy aberrations leading to both, phenotypic intra-tumor diversity (mTOR activating mutation) and convergent loss of function(SETD2, PTEN and KDM5C). ITH can lead to underestimation of the tumor genomic landscape portrayed from single biopsies and may present significant challenges to personalized medicine and biomarker development. ITH, associated with heterogeneous protein function, may foster tumor adaptation and therapeutic failure through Darwinian selection. Targeting mutations commonly located on the trunks, rather than the branches of phylogenetic trees, may improve therapeutic outcomes. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 964. doi:1538-7445.AM2012-964