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Phylogenetic analysis of the genus Plasmodium based on the gene encoding adenylosuccinate lyase.

Phylogenetic studies of the genus Plasmodium have been performed using sequences of the nuclear, mitochondrial and plastid genes. Here we have analyzed the adenylosuccinate lyase (ASL) gene, which encodes an enzyme involved in the salvage of host purines needed by malaria parasites for DNA synthesis. The ASL gene is present in several eukaryotic as well as prokaryotic organisms and does not have repeat regions, which facilitates the accuracy of the alignment. Furthermore, it has been shown that ASL is not subject to positive natural selection. We have sequenced the ASL gene of several different Plasmodium species infecting humans, rodents, monkeys and birds and used the obtained sequences along with the previously known P. falciparum ASL sequence, for structural and phylogenetic analysis of the genus Plasmodium. The genetic divergence of ASL is comparable with that observed in other nuclear genes such as cysteine proteinase, although ASL cannot be considered conserved when compared to aldolase or superoxide dismutase, which exhibit a slower rate of evolution. Nevertheless, a protein like ASL has a rate of evolution that provides enough information for elucidating evolutionary relationships. We modeled 3D structures of the ASL protein based on sequences used in the phylogenetic analysis and obtained a consistent structure for four different species despite the divergence observed. Such models would facilitate alignment in further studies with a greater number of plasmodial species or other Apicomplexa.

Adenylosuccinate Lyase↗

Soft-tissue anatomy of the extant hominoids: a review and phylogenetic analysis.

This paper reports the results of a literature search for information about the soft-tissue anatomy of the extant non-human hominoid genera, Pan, Gorilla, Pongo and Hylobates, together with the results of a phylogenetic analysis of these data plus comparable data for Homo. Information on the four extant non-human hominoid genera was located for 240 out of the 1783 soft-tissue structures listed in the Nomina Anatomica. Numerically these data are biased so that information about some systems (e.g. muscles) and some regions (e.g. the forelimb) are over-represented, whereas other systems and regions (e.g. the veins and the lymphatics of the vascular system, the head region) are either under-represented or not represented at all. Screening to ensure that the data were suitable for use in a phylogenetic analysis reduced the number of eligible soft-tissue structures to 171. These data, together with comparable data for modern humans, were converted into discontinuous character states suitable for phylogenetic analysis and then used to construct a taxon-by-character matrix. This matrix was used in two tests of the hypothesis that soft-tissue characters can be relied upon to reconstruct hominoid phylogenetic relationships. In the first, parsimony analysis was used to identify cladograms requiring the smallest number of character state changes. In the second, the phylogenetic bootstrap was used to determine the confidence intervals of the most parsimonious clades. The parsimony analysis yielded a single most parsimonious cladogram that matched the molecular cladogram. Similarly the bootstrap analysis yielded clades that were compatible with the molecular cladogram; a (Homo, Pan) clade was supported by 95% of the replicates, and a (Gorilla, Pan, Homo) clade by 96%. These are the first hominoid morphological data to provide statistically significant support for the clades favoured by the molecular evidence.

Anatomy, Comparative↗

Phylogenetic analysis of 5'-noncoding regions from the ABA-responsive rab16/17 gene family of sorghum, maize and rice provides insight into the composition, organization and function of cis-regulatory modules.

Phylogenetic analysis of sequences from gene families and homologous genes from species of varying divergence can be used to identify conserved noncoding regulatory elements. In this study, phylogenetic analysis of 5'-noncoding sequences was optimized using rab17, a well-characterized ABA-responsive gene from maize, and five additional rab16/17 homologs from sorghum and rice. Conserved 5'-noncoding sequences among the maize, sorghum, and rice rab16/17 homologs were identified with the aid of the software program FootPrinter and by screening for known transcription-factor-binding sites. Searches for 7 of 8 (7/8)bp sequence matches within aligned 5'-noncoding segments of the rab genes identified many of the cis-elements previously characterized by biochemical analysis in maize rab17 plus several additional putative regulatory elements. Differences in the composition of conserved noncoding sequences among rab16/17 genes were related to variation in rab gene mRNA levels in different tissues and to response to ABA treatment using qRT-PCR. Absence of a GRA-like element in the promoter of sorghum dhn2 relative to maize rab17 was correlated with an approximately 85-fold reduction of dhn2 RNA in sorghum shoots. Overall, we conclude that phylogenetic analysis of gene families among rice, sorghum, and maize will help identify regulatory sequences in the noncoding regions of genes and contribute to our understanding of grass gene regulatory networks.

5' Untranslated Regions↗

Phylogenetic analysis of the Chlamydia trachomatis major outer membrane protein and examination of potential pathogenic determinants.

Phylogenetic analysis was utilized to investigate biological relationships (tissue tropism, disease presentation, and epidemiologic success), as evidenced by coevolution, among human strains of Chlamydia trachomatis. Nucleotide sequences of omp1, the gene encoding the major outer membrane protein (MOMP) of C. trachomatis, were determined for 40 strains representing 11 serovars. These data were combined with available omp1 sequences from GenBank for an analysis encompassing a total of 69 strains representing 17 serovars infecting humans. Phylogenetic analysis of the nucleotide and inferred amino acid sequences showed no evolutionary relationships among serovars that corresponded to biological or pathological phenotypes (tissue tropism, disease presentation, and epidemiologic success). In addition, no specific residues that may have evolved to play a role in determining biologically relevant characteristics of chlamydia, such as tissue specificity, disease presentation, and epidemiologic success, were apparent in the MOMP. These results suggest that variation in MOMP may have arisen from a need to be diverse in the presence of immune pressure rather than as a function of pathogenicity. Therefore, the role of MOMP in disease pathogenesis and infection may be passive, and it may not be the major ligand responsible for directing infection of various human cell types.

Bacterial Outer Membrane Proteins↗

Sampling properties of DNA sequence data in phylogenetic analysis.

We inferred phylogenetic trees from individual genes and random samples of nucleotides from the mitochondrial genomes of 10 vertebrates and compared the results to those obtained by analyzing the whole genomes. Individual genes are poor samples in that they infrequently lead to the whole-genome tree. A large number of nucleotide sites is needed to exactly determine the whole-genome tree. A relatively small number of sites, however, often results in a tree close to the whole-genome tree. We found that blocks of contiguous sites were less likely to lead to the whole-genome tree than samples composed of sites drawn individually from throughout the genome. Samples of contiguous sites are not representative of the entire genome, a condition that violates a basic assumption of the bootstrap method as it is applied in phylogenetic studies.

Animals↗

Phylogenetic analysis of the Hsp70 sequences reveals the monophyly of Metazoa and specific phylogenetic relationships between animals and fungi.

To understand the early evolution of the Metazoa, it is necessary to determine the correct phylogenetic status of diploblastic animals. Despite cladistic studies of morphological characters and recent molecular phylogenetic studies, it remains uncertain whether diploblasts are monophyletic or paraphyletic, and how the phyla of diploblasts are phylogenetically related. The heat shock protein 70 (Hsp70) sequences, because of their ubiquity and high degree of conservation, could provide a useful model for phylogenetic analysis. We have sequenced almost the entire nucleic acid sequence of cytoplasmic Hsp70 from eight diploblastic species. Our data support the monophyly of diploblastic animals. However, the phylogenetic relationships of the diploblast groups were not significantly resolved. Our phylogenetic trees also support the monophyly of Metazoa with high bootstrap values, indicating that animals form an extremely robust clade.

Amino Acid Sequence↗

Usefulness and limitation of phylogenetic analysis for hepatitis C virus core region: application to isolates from Egyptian and Yemeni patients.

We report here the nucleotide sequences of the core region of HCV isolates from Egyptian and Yemeni patients and the method for classifying these HCV isolates by phylogenetic analysis. Sequence comparison suggested that the genotypes of these isolates were the same. Preliminary phylogenetic analysis of the HCV core region indicated that the genotypes of both isolates were 1c. However, an additional phylogenetic tree of the HCV core region constructed using a greater number of HCV isolates than that used in the preliminary analysis and on the basis of alignment of nucleotide sequences in an appropriate length indicated that the genotypes of these isolates were 4 and not 1c. For a more detailed analysis, the nucleotide sequences of the HCV E1 region as well as the core region for the same Yemeni patient were determined. A phylogenetic tree of the E1 region confirmed that the genotype of the HCV isolate from the Yemeni patient was 4. These data indicate that even when classifying HCV isolates using phylogenetic analysis, the misclassification would occur if care is not taken regarding the number and sequence lengths of the isolates included in the analysis.

Base Sequence↗

Identification of cytochrome P-450 1A (CYP1A) genes from two teleost fish, toadfish (Opsanus tau) and scup (Stenotomus chrysops), and phylogenetic analysis of CYP1A genes.

Cytochrome P-450-mediated responses to environmental challenges are well known in diverse animal taxa, but the evolution of the complex gene superfamily coding for these enzymes is poorly understood. Here we report a phylogenetic analysis of the cytochrome P-450 1A (CYP1A) genes including two new sequences determined from teleost fish, toadfish (Opsanus tau) and scup (Stenotomus chrysops). Degenerate PCR primers were used to amplify a 1.2 kbp fragment from liver cDNA. The toadfish PCR product was used as a probe to identify a full-length CYP1A clone from a toadfish liver cDNA library. The entire coding region of the scup CYP1A was obtained by rapid amplification of cDNA ends (RACE) using specific primers based on the sequence of the partial PCR product. The predicted protein sequences for toadfish and scup CYP1A shared 78% and 83% amino acid identity with rainbow trout CYP1A1 respectively. Amino acid identity with mammalian CYP1A proteins ranged from 51 to 60% for 505 aligned positions. Phylogenetic analysis of four teleost fish CYP1A genes (trout, toadfish, scup and plaice) and 12 mammalian CYP1A genes suggests a monophyletic origin of the teleost genes, with the trout gene being most divergent, and indicates three distinct groupings: mammalian 1A1, mammalian 1A2, and fish 1A. This supports the idea that the gene duplication event which gave rise to CYP1A1 and CYP1A2 occurred after the divergence of the lines leading to mammals and fish. These results establish a molecular phylogeny within the CYP1A subfamily, the first such detailed phylogenetic analysis within a cytochrome P-450 family.

Amino Acid Sequence↗

Phylogenetic analysis of the full-length SARS-CoV sequences: evidence for phylogenetic discordance in three genomic regions.

The origin of the severe acute respiratory syndrome-coronavirus (SARS-CoV) remains unclear. Evidence based on Bayesian scanning plots and phylogenetic analysis using maximum likelihood (ML) and Bayesian methods indicates that SARS-CoV, for the largest part of the genome ( approximately 80%), is more closely related to Group II coronaviruses sequences, whereas in three regions in the ORF1ab gene it shows no apparent similarity to any of the previously characterized groups of coronaviruses. There is discordant phylogenetic clustering of SARS-CoV and coronaviruses sequences, throughout the genome, compatible with either ancient recombination events or altered evolutionary rates in different lineages, or a combination of both.

Evolution, Molecular↗

Typing multidrug-resistant Staphylococcus aureus: conflicting epidemiological data produced by genotypic and phenotypic methods clarified by phylogenetic analysis.

An outbreak of an unusual tetracycline-sensitive, rifampicin- and ciprofloxacin-resistant, methicillin-resistant Staphylococcus aureus (MRSA) strain at a large teaching hospital was investigated. Two typing methods, phage typing and restriction fragment length polymorphism (RFLP) by pulsed-field gel electrophoresis (RFLP-PFGE), gave conflicting results which were clarified by phylogenetic analysis. Phage typing identified all the "epidemic-associated" strains as identical, while RFLP-PFGE further divided these strains into four pulsotypes. Phylogenetic analysis showed these four pulsotypes were related genetically and also recognized a second strain of MRSA causing a continuing cross-infection problem. Variation in the RFLP-PFGE pattern was shown to occur following lysogenization of phage-sensitive MRSA. These results indicate that in analyzing outbreaks caused by subgroups of clonal organisms like MRSA, it is necessary to use at least two typing methods and that conflicts between these could be resolved by phylogenetic analysis.

Bacterial Typing Techniques↗

Whole genome-based phylogenetic analysis of free-living microorganisms.

A phylogenetic 'tree of life' has been constructed based on the observed presence and absence of families of protein-encoding genes observed in 11 complete genomes of free-living microorganisms. Past attempts to reconstruct the evolutionary relation-ships of microorganisms have been limited to sets of genes rather than complete genomes. Despite apparent rampant lateral gene transfer among microorganisms, these results indicate a single robust underlying evolutionary history for these organisms. Broadly, the tree produced is very similar to the small subunit rRNA tree although several additional phylogenetic relationships appear to be resolved, including the relationship of Archaeoglobus to the methanogens studied. This result is in contrast to notions that a robust phylogenetic reconstruction of microorganisms is impossible due to their genomes being composed of an incomprehensible amalgam of genes with complicated histories and suggests that this style of genome-wide phylogenetic analysis could become an important method for studying the ancient diversification of life on Earth. Analyses using informational and operational subsets of the genes showed that this 'tree of life' is not dependent on the phylogenetically more consistent informational genes.

Computational Biology↗

African origin of GB virus C determined by phylogenetic analysis of a complete genotype 5 genome from South Africa.

GB virus C (GBV-C), a positive-strand RNA virus, currently infects approximately one-sixth of the world's population. This virus exists as a collection of genotypes whose global distribution correlates with geographical origin. Genotyping of GBV-C isolates by phylogenetic analysis has relied upon the use of 5'-untranslated region (5'-UTR) sequences, however, complete genome sequences are used to demonstrate definitively their existence and geographical correlation. Initial identification of the fifth genotype from South Africa was based upon phylogenetic analysis of the 5'-UTR. It was sought to confirm this classification by analysis of full-length E2 genes from South African isolates and by analysis of a complete genotype 5 genome. Analysis of full-length E2 genes from 28 GBV-C-infected South African individuals revealed the existence of a unique group of 18 isolates, distinct from the other four genotypes. Bootstrap analysis provided strong support (95 %) for this fifth group. The remaining isolates were either genotype 1 (n=8) or 2 (n=2). Analysis of human E2 gene sequences, with the E2 gene from the chimpanzee variant GBV-Ctro included as the outgroup, produced a tree rooted on the genotype 1 branch. The complete genome nucleotide sequence of South African genotype 5 isolate D50 was determined. Phylogenetic analysis of the 5'-UTR and open reading frame produced congruent trees that grouped the sequences into five major genotypes. Inclusion of the corresponding region of the chimpanzee isolate GBV-Ctro in the analysis produced trees rooted on the branch leading to the genotype 5 isolate D50, suggesting an ancient African origin of GBV-C.

Flaviviridae Infections↗

Gallus gallus aggrecan gene-based phylogenetic analysis of selected avian taxonomic groups.

Mitochondrial DNA (mtDNA) sequences remain the most widely used for phylogenetic analysis in birds. A major limitation of mtDNA sequences, however, is that mitochondria genes are inherited as a single linkage group. Here we describe the use of a 540-bp DNA sequence corresponding to the G3 domain of Gallus gallus nuclear aggrecan gene (AGCI) for phylogenetic analysis of the main groups of Galliformes including Phasianidae, Numididae, and Odontophoridae. We also included species from Cracidae and Megapodiidae which are considered by some as Craciformes and others, including here as Galliformes. The uncorrected sequence divergence of the G3 fragments ranges from 1% among the grouses to 36% between some of the distant groups within Galliformes. These sequences contain 39-48% AT nucleotides and the ratios of transition versus transversion are above 1.5 in majority of the comparisons. Using G3 sequences from an Anseriform, Oxyura jamaicensis, as out-groups, phylogenetic trees were obtained using maximum parsimony and distance algorithms and bootstrap analyses. These trees were consistent with those described using Avian sarcoma and leucosis virus gag genes and those from amino acid sequences of hemoglobin and lysozyme c. Our data also support relationships among Galliformes which were defined using mtDNA sequences. In addition to the general support of the five main families of Galliformes, our data are also consistent with previous work that showed Francolinus africanus and Gallus gallus are in the same clade and that Tetraoninae is a well-supported monophyletic subfamily within Phasianidae. The results presented here suggest that the AGC1 sequences meet the criterion of novel nuclear DNA sequences that can be used to help resolve the relationships among Galliformes.

Aggrecans↗

Identification and phylogenetic analysis of hepatitis C virus in forensic blood samples obtained from injecting drug users.

Injecting drug users (IDUs) are a high-risk group for contracting hepatitis B virus (HBV), hepatitis C virus (HCV), and human immunodeficiency virus (HIV) infections. In Japan, data on the prevalence of those blood-borne viruses among IDUs are very limited. Blood samples were obtained from 12 cadavers of IDUs sent to Nagoya City University for the purpose of judicious autopsy and two alive IDUs with hepatitis C referred to a local hospital at the same period. The viruses were detected by polymerase chain reaction and phylogenetic analysis was performed. Two (16.6%) of the 12 autopsy cases were positive for HCV, but no case was positive for either HBV or HIV. Phylogenetic analysis of the two HCV isolates revealed that one was classified into genotype 1b and another was genotype 2b. Furthermore, nucleotide sequences of two isolates recovered from IDUs with hepatitis C were identical, that indicated the transmission of HCV between them, and those HCV were phylogenetically classified into genotype 2a. The prevalence of HCV infection among IDUs in Japan, despite the case of judicious autopsy, seems to be high, but HIV infection seems to be rare. The transmission of HCV between IDUs was demonstrated, and this indicates that phylogenetic analysis would applicable to also forensic analysis. HCV isolates identified in this study did not phylogenetically segregate, thus multiple transmission route of HCV among IDUs seems be exist in Japan.

Adult↗

A phylogenetic analysis elucidating a case of patient-to-patient transmission of hepatitis C virus during surgery.

A phylogenetic hepatitis C virus (HCV) assay based on the core-Envelope 1 (C-E1) region was developed and used to elucidate a case of a patient-to-patient transmission. The index patient showed clinical symptoms of hepatitis seven weeks after surgery for hallux valgus under general anaesthesia. She progressed to a chronic persistent infection as indicated by positive HCV PCR results two years after surgery. Before her operation, a patient with HCV antibodies and positive HCV PCR had undergone surgery in the same room. There were two possibilities whereby the index patient could have been infected with hepatitis C, either through her work as a nurse or by transmission during surgery. By sequencing the 5' non-coding region PCR product, we found that both patients were infected with genotype 1a. Phylogenetic analysis with the variable C-E1 region suggested that the two patients clustered together with a bootstrap 100% in a tree with 75 sequence references. We further performed a phylogenetic analysis in this region with the genotype 1a reference sequences and an additional 25 genotype 1a sequences consecutively collected from Danish patients with HCV. The two patients still clustered together, supported by a high bootstrap 1000 value of 999. Homology analyses combined with the epidemiological findings indicate that the patient operated on in the same room before the index case was the most likely source of transmission. The mode of transmission could not be conclusively established, but a reusable part of the anaesthetic respiratory circuit is a possibility and a well known risk.

Adult↗

HIV-1 subtyping using phylogenetic analysis of pol gene sequences.

HIV-1 pol gene sequencing is now used routinely in France to identify mutations associated with resistance to reverse transcriptase (RT) or protease (PR) inhibitors. These sequences may also provide other information, such as the HIV-1 subtype. HIV-1 subtyping was compared using the RT and PR gene sequences to heteroduplex mobility assay (HMA) of the envelope gene. The RT and PR genes of 51 samples that had been subtyped earlier by HMA were sequenced. Sequences were aligned and subtypes were determined by phylogenetic analysis with reference HIV sequences. HMA gave the following subtypes: A (20), B (19), C (1), D (3), F (1), G (3) and CRF01-AE (4). Phylogenetic analysis of the RT gene gave: A (5), B (19), C (2), D (3), F (1), G (6), J (2), CRF01_AE (4), CFR02_AG (7) and undetermined (2). PR gene analysis did not infer subtypes with sufficient confidence. HMA and RT subtyping was not in agreement in nine cases. RT subtyping can identify CFR02_AG and CRF01_AE variants from A subtype RT. It was shown that phylogenetic analysis of the RT gene could provide a useful method for HIV-1 subtyping. The length of the amplicon and the relative performance of each primer pair used in this study favoured RT sequences as a subtyping tool. One potential advantage over env subtyping HMA is the ability to identify some circulating recombinant forms (CRFs).

Base Sequence↗

Phylogenetic analysis of general bacterial porins: a phylogenomic case study.

Bacterial porin proteins allow for the selective movement of hydrophilic solutes through the outer membrane of Gram-negative bacteria. The purpose of this study was to clarify the evolutionary relationships among the Type 1 general bacterial porins (GBPs), a porin protein subfamily that includes outer membrane proteins ompC and ompF among others. Specifically, we investigated the potential utility of phylogenetic analysis for refining poorly annotated or mis-annotated protein sequences in databases, and for characterizing new functionally distinct groups of porin proteins. Preliminary phylogenetic analysis of sequences obtained from GenBank indicated that many of these sequences were incompletely or even incorrectly annotated. Using a well-curated set of porins classified via comparative genomics, we applied recently developed bayesian phylogenetic methods for protein sequence analysis to determine the relationships among the Type 1 GBPs. Our analysis found that the major GBP classes (ompC, phoE, nmpC and ompN) formed strongly supported monophyletic groups, with the exception of ompF, which split into two distinct clades. The relationships of the GBP groups to one another had less statistical support, except for the relationships of ompC and ompN sequences, which were strongly supported as sister groups. A phylogenetic analysis comparing the relationships of the GenBank GBP sequences to the correctly annotated set of GBPs identified a large number of previously unclassified and mis-annotated GBPs. Given these promising results, we developed a tree-parsing algorithm for automated phylogenetic annotation and tested it with GenBank sequences. Our algorithm was able to automatically classify 30 unidentified and 15 mis-annotated GBPs out of 78 sequences. Altogether, our results support the potential for phylogenomics to increase the accuracy of sequence annotations.

Algorithms↗

Phylogenetic analysis of the main neutralization and hemagglutination determinants of all human adenovirus prototypes as a basis for molecular classification and taxonomy.

Human adenoviruses (HAdV) are responsible for a wide spectrum of diseases. The neutralization epsilon determinant (loops 1 and 2) and the hemagglutination gamma determinant are relevant for the taxonomy of HAdV. Precise type identification of HAdV prototypes is crucial for detection of infection chains and epidemiology. epsilon and gamma determinant sequences of all 51 HAdV were generated to propose molecular classification criteria. Phylogenetic analysis of epsilon determinant sequences demonstrated sufficient genetic divergence for molecular classification, with the exception of HAdV-15 and HAdV-29, which also cannot be differentiated by classical cross-neutralization. Precise sequence divergence criteria for typing (<2.5% from loop 2 prototype sequence and <2.4% from loop 1 sequence) were deduced from phylogenetic analysis. These criteria may also facilitate identification of new HAdV prototypes. Fiber knob (gamma determinant) phylogeny indicated a two-step model of species evolution and multiple intraspecies recombination events in the origin of HAdV prototypes. HAdV-29 was identified as a recombination variant of HAdV-15 (epsilon determinant) and a speculative, not-yet-isolated HAdV prototype (gamma determinant). Subanalysis of molecular evolution in hypervariable regions 1 to 6 of the epsilon determinant indicated different selective pressures in subclusters of species HAdV-D. Additionally, gamma determinant phylogenetic analysis demonstrated that HAdV-8 did not cluster with -19 and -37 in spite of their having the same tissue tropism. The phylogeny of HAdV-E4 suggested origination by interspecies recombination between HAdV-B (hexon) and HAdV-C (fiber), as in simian adenovirus 25, indicating additional zoonotic transfer. In conclusion, molecular classification by systematic sequence analysis of immunogenic determinants yields new insights into HAdV phylogeny and evolution.

Adenovirus Infections, Human↗