Toward a resolution of Campanulid phylogeny, with special reference to the placement of Dipsacales

Broad-scale phylogenetic analyses of the angiosperms and of the Asteridae have failed to confidently resolve relationships among the major lineages of the campanulid Asteridae (i.e., the euasterid II of APG II, 2003). To address this problem we assembled presently available sequences for a core set of 50 taxa, representing the diversity of the four largest lineages (Apiales, Aquifoliales, Asterales, Dipsacales) as well as the smaller ""unplaced"" groups (e.g., Bruniaceae, Paracryphiaceae, Columelliaceae). We constructed four data matrices for phylogenetic analysis: a chloroplast coding matrix (atpB, matK, ndhF, rbcL), a chloroplast non-coding matrix (rps16 intron, trnT-F region, trnV-atpE IGS), a combined chloroplast dataset (all seven chloroplast regions), and a combined genome matrix (seven chloroplast regions plus 18S and 26S rDNA). Bayesian analyses of these datasets using mixed substitution models produced often well-resolved and supported trees. Consistent with more weakly supported results from previous studies, our analyses support the monophyly of the four major clades and the relationships among them. Most importantly, Asterales are inferred to be sister to a clade containing Apiales and Dipsacales. Paracryphiaceae is consistently placed sister to the Dipsacales. However, the exact relationships of Bruniaceae, Columelliaceae, and an Escallonia clade depended upon the dataset. Areas of poor resolution in combined analyses may be partly explained by conflict between the coding and non-coding data partitions. We discuss the implications of these results for our understanding of campanulid phylogeny and evolution, paying special attention to how our findings bear on character evolution and biogeography in Dipsacales.

Coding behavioural data for cladistic analysis: using dynamic homology without parsimony

Many of the controversies around the concept of homology rest on the subjectivity inherent to primary homology propositions. Dynamic homology partially solves this problem, but there has been up to now scant application of it outside of the molecular domain. This is probably because morphological and behavioural characters are rich in properties, connections and qualities, so that there is less space for conflicting character delimitations. Here we present a new method for the direct optimization of behavioural data, a method that relies on the richness of this database to delimit the characters, and on dynamic procedures to establish character state identity. We use between-species congruence in the data matrix and topological stability to choose the best cladogram. We test the methodology using sequences of predatory behaviour in a group of spiders that evolved the highly modified predatory technique of spitting glue onto prey. The cladogram recovered is fully compatible with previous analyses in the literature, and thus the method seems consistent. Besides the advantage of enhanced objectivity in character proposition, the new procedure allows the use of complex, context-dependent behavioural characters in an evolutionary framework, an important step towards the practical integration of the evolutionary and ecological perspectives on diversity. (C) The Willi Hennig Society 2010.

Systematic revision and cladistic analysis of the Brazilian subfamily Sodreaninae (Opiliones: Gonyleptidae)

Sodreaninae is reviewed and all ten species are combined under its type genus, Sodreana Mello-Leitao, 1922, according to a cladistic analysis of morphological characters, which revealed a pectinate pattern of clades. The subfamily is endemic to the Brazilian Atlantic rainforest from Santa Catarina state to Rio de Janeiro state. Sodreana is herein considered a senior synonym of Stygnobates Mello-Leitao, 1927, Zortalia Mello-Leitao, 1936, Gertia B. Soares & H. Soares, 1946 and Annampheres H. Soares, 1979. The following new combinations are proposed: Sodreana barbiellinii (Mello-Leitao, 1927), Sodreana hatschbachi (B. Soares & H. Soares, 1946), Sodreana inscripta (Mello-Leitao, 1939), Sodreana leprevosti (B. Soares & H. Soares, 1947b), Sodreana bicalcarata (Mello-Leitao, 1936). Sodreana granulata (Mello-Leitao, 1937) is revalidated from the synonymy of Sodreana sodreana Mello-Leitao, 1922. Three new species are described: Sodreana glaucoi from Ilhabela and Boraceia, Sao Paulo state; S. curupira from Parque Nacional da Serra dos Orgaos, Rio de Janeiro state, and S. caipora from Ubatuba, Sao Paulo state. Sodreaninae species are restricted to forested areas and most occur in the southern part of the coastal Atlantic rainforest, one species occurs in interior Atlantic rainforest. The biogeographical analysis (Brooks Parsimony Analysis) resulted in a single and fully resolved most parsimonious tree with three main: components: northern (Bahia and Serra do Espinhaco), southern (Santa Catarina, Parana, Serra do Mar of Sao Paulo), and central (Espirito Santo, Serra da Bocaina, southern state of Rio de Janeiro, Serra dos Orgaos, Serra da Mantiqueira, Serra do Mar of Sao Paulo).

Evolutionary history of Dengue virus type 4: Insights into genotype phylodynamics

Dengue virus type 4 (DENV-4) circulates in tropical and subtropical countries from Asia and the Americas. Despite the importance of dengue virus distribution, little is known about the worldwide viral spread. Following a Bayesian phylogenetic approach we inferred the evolutionary history of 310 isolates sampled from 37 countries during the time period 1956-2008 and the spreading dynamics for genotypes I and II. The region (tropical rainforest biome) comprised by Malaysia-Thailand was the most likely ancestral area from which the serotype has originated and spread. Interestingly, cross-correlation analysis on demographic time series with the Asian sequences showed a statistically significant negative correlation that could be suggestive of competition among genotypes within the same serotype. (C) 2011 Elsevier B.V. All rights reserved.

Relation of genetic phylogeny and geographical distance of tick-borne encephalitis virus in central Europe

Tick-borne encephalitis virus (TBEV) is the most important arboviral agent causing disease of the central nervous system in central Europe. In this study, 61 TBEV E gene sequences derived from 48 isolates from the Czech Republic, and four isolates and nine TBEV strains detected in ticks from Germany, covering more than half a century from 1954 to 2009, were sequenced and subjected to phylogenetic and Bayesian phylodynamic analysis to determine the phylogeography of TBEV in central Europe. The general Eurasian continental east-to-west pattern of the spread of TBEV was confirmed at the regional level but is interlaced with spreading that arises because of local geography and anthropogenic influence. This spread is reflected by the disease pattern in the Czech Republic that has been observed since 1991. The overall evolutionary rate was estimated to be approximately 8x10(-4) substitutions per nucleotide per year. The analysis of the TBEV E genes of 11 strains isolated at one natural focus in Zd`ar Kaplice proved for the first time that TBEV is indeed subject to local evolution.

High genetic diversity in field isolates of Trypanosoma theileri assessed by analysis of cathepsin L-like sequences disclosed multiple and new genotypes infecting cattle in Thailand

In this study, we describe the first survey in Thailand of Trypanosoma theileri, a widespread and prevalent parasite of cattle that is transmitted by tabanid flies. Investigation of 210 bovine blood samples of Thai cattle from six farms by hematocrit centrifuge technique (HCT) revealed 14 samples with trypanosomes morphologically compatible to T. theileri. Additional animals were positive for T. theileri by PCR based on the Cathepsin L-like sequence (TthCATL-PCR) despite negative by HCT, indicating cryptic infections. Results revealed a prevalence of 26 +/- 15% (95% CI) of T. theileri infection. Additionally, 12 samples positive for T. theileri were detected in cattle from other 11 farms. From a total of 30 blood samples positive by HCT and/or PCR from 17 farms, seven were characterized to evaluate the genetic polymorphism of T. theileri through sequence analysis of PCR-amplified CATL DNA sequences. All CATL sequences of T. theileri from Thai cattle clustered with sequences of the previously described phylogenetic lineages TthI and TthII, supporting only two major lineages of T. theileri in cattle around the world. However, 11 of the 29 CATL sequences analyzed showed to be different, disclosing an unexpectedly large polymorphic genetic repertoire, with multiple genotypes of T. theileri not previously described in other countries circulating in Thai cattle. (C) 2011 Elsevier B.V. All rights reserved.

Phylogeny of Triatoma sherlocki (Hemiptera: Reduviidae: Triatominae) Inferred from Two Mitochondrial Genes Suggests Its Location Within the Triatoma brasiliensis Complex

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Strain differentiation of Trichophyton rubrum by randomly amplified polymorphic DNA and analysis of rDNA nontranscribed spacer

Trichophyton rubrum is the most common pathogen causing dermatophytosis. Molecular strain-typing methods have recently been developed to tackle epidemiological questions and the problem of relapse following treatment. A total of 67 strains of T rubrum were screened for genetic variation by randomly amplified polymorphic DNA (RAPD) analysis, with two primers, 5'-d[GGTGCGGGAA]-3' and 5'-d[CCCGTCAGCA]-3', as well as by subrepeat element analysis of the nontranscribed spacer of rDNA, using the repetitive subelements TRS-1 and TRS-2. A total of 12 individual patterns were recognized with the first primer and 11 with the second. Phylogenetic analysis of the RAPID products showed a high degree of similarity (> 90 %) among the epidemiologically related clinical isolates, while the other strains possessed 60% similarity. Specific amplification of TRS-1 produced three strain-characteristic banding patterns (PCR types); simple patterns representing one copy of TRS-1 and two copies of TRS-2 accounted for around 85 % of all isolates. It is concluded that molecular analysis has important implications for epidemiological studies, and RAPID analysis is especially suitable for molecular typing in T. rubrum.

Analysis of Bothrops jararacussu venomous gland transcriptome focusing on structural and functional aspects: I - gene expression profile of highly expressed phospholipases A(2)

Snake venom glands are a rich source of bioactive molecules such as peptides, proteins and enzymes that show important pharmacological activity leading to in local and systemic effects as pain, edema, bleeding and muscle necrosis. Most studies on pharmacologically active peptides and proteins from snake venoms have been concerned with isolation and structure elucidation through methods of classical biochemistry. As an attempt to examine the transcripts expressed in the venom gland of Bothrops jararacussu and to unveil the toxicological and pharmacological potential of its products at the molecular level, we generated 549 expressed sequence tags (ESTs) from a directional cDNA library. Sequences obtained from single-pass sequencing of randomly selected cDNA clones could be identified by similarities searches on existing databases, resulting in 197 sequences with significant similarity to phospholipase A(2) (PLA(2)), of which 83.2% were Lys49-PLA(2) homologs (BOJU-1), 0.1% were basic Asp49-PLA(2)s (BOJU-II) and 0.6% were acidic Asp49-PLA(2)s (BOJU-III). Adjoining this very abundant class of proteins we found 88 transcripts codifying for putative sequences of metalloproteases, which after clustering and assembling resulted in three full-length sequences: BOJUMET-I, BOJUMET-II and BOJUMET-III; as well as 25 transcripts related to C-type lectin like protein including a full-length cDNA of a putative galactose binding C-type lectin and a cluster of eight serine-proteases transcripts including a full-length cDNA of a putative serine protease. Among the full-length sequenced clones we identified a nerve growth factor (Bj-NGF) with 92% identity with a human NGF (NGHUBM) and an acidic phospholipase A2 (BthA-I-PLA(2)) displaying 85-93% identity with other snake venom toxins. Genetic distance among PLA(2)s from Bothrops species were evaluated by phylogenetic analysis. Furthermore, analysis of full-length putative Lys49-PLA(2) through molecular modeling showed conserved structural domains, allowing the characterization of those proteins as group II PLA(2)s. The constructed cDNA library provides molecular clones harboring sequences that can be used to probe directly the genetic material from gland venom of other snake species. Expression of complete cDNAs or their modified derivatives will be useful for elucidation of the structure-function relationships of these toxins and peptides of biotechnological interest. (C) 2004 Elsevier SAS. All rights reserved.

Molecular approaches for structural characterization of Bothrops L-amino acid oxidases with antiprotozoal activity: cDNA cloning, comparative sequence analysis, and molecular modeling

Two L-amino acid oxidases (LAAOs) were identified by random sequencing of cDNA libraries from the venom glands of Bothrops moojeni (BmooLAAO) and Bothrops jararacussu (Bjussu LAAO). Phylogenetic analysis involving other SV-LAAOs showed sequence identities within the range 83-87% being closely related to those from Agkistrodon and Trimeresurus. Molecular modeling experiments indicated the FAD-binding, substrate-binding, and helical domains of Bmoo and Bjussu LAAOs. The RMS deviations obtained by the superposition of those domains and that from Calloselasma rhodostoma LAAO crystal structure confirm the high degree of structural similarity between these enzymes. Purified BjussuLAAO-I and BmooLAAO-I exhibited antiprotozoal activities which were demonstrated to be hydrogen-peroxide mediated. This is the first report on the isolation and identification of cDNAs encoding LAAOs from Bothrops venom. The findings here reported contribute to the overall structural elucidation of SV-LAAOs and will advance the understanding on their mode of action. (c) 2006 Elsevier B.V. All rights reserved.

5S rDNA variation and its phylogenetic inference in the genus Leporinus (Characiformes : Anostomidae)

5S rDNA sequences have proven to be valuable as genetic markers to distinguish closely related species and also in the understanding of the dynamic of repetitive sequences in the genomes. In the aim to contribute to the knowledge of the evolutionary history of Leporinus (Anostomidae) and also to contribute to the understanding of the 5S rDNA sequences organization in the fish genome, analyses of 5S rDNA sequences were conducted in seven species of this genus. The 5S rRNA gene sequence was highly conserved among Leporinus species, whereas NTS exhibit high levels of variations related to insertions, deletions, microrepeats, and base substitutions. The phylogenetic analysis of the 5S rDNA sequences clustered the species into two clades that are in agreement with cytogenetic and morphological data.

Molecular systematics of the armored neotropical catfish subfamily Neoplecostominae (Siluriformes: Loricariidae)

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Partial cDNA sequence analysis of myosin Va from rainbow trout (Oncorhynchus mykiss) and its relationship to myosin v isoforms from other vertebrates.

Partial cDNA sequences of myosin V from rainbow trout Oncorhynchus mykiss were analyzed and showed high similarity to MVa from other vertebrates. Phylogenetic analysis has shown that events resulting in the formation of paralogous copies of myosin Va, Vb, and Vc occurred before the divergence of vertebrates into different classes. Expression analysis of myosin Va, Vb, and Vc in different O. mykiss tissues revealed MVa exclusively expressed in hypophysis and brain whereas Vb and Vc were expressed in practically all tissues analyzed. The nucleotide sequence for myosin V was explored in a fish species for the first time and these results represent an important start in understanding the organization, evolution, and expression of myosins in early vertebrates. The data presented here represent contributions to the knowledge of rainbow trout genome. A better understanding of this economically important species could assist in development of improved strains of this fish for aquaculture.

On the larval stages of Macrocoeloma diplacanthum (Decapoda : Brachyura : Majidae), and a review of mithracine phylogenetic aspects

Larval development of Macrocoeloma diplacanthum (Stimpson) consists of two zoeal stages, followed by the megalopa. Each larval stage is described in detail. The duration of the zoeal stages was 2-3 (2.4 +/- 0.5) and 3-4 (3.5 +/- 0.5) days for the first and second zoea, respectively, the megalopa phase appearing 6-8 (7.0 +/- 0.5) days after hatching. Unlike for other majids, zoeal stages of M. diplacanthum can be readily distinguished by their distended forehead with strong underlying muscle bands, undercut dorsal carapace spine, and spine on the terminal endopod segment of the first maxilliped. No other known mithracine or majid zoeae exhibit this combination of features. Our zoeal account of M. diplacanthum from Mexico is remarkably consistent with Floridian specimens previously described. However, we have found some differences between descriptions, which could be attributed to natural variation or inadequate description. Previous attempts to evaluate the relationships within Mithacinae have been based on larval characters widely distributed throughout Majidae and therefore are considered inadequate to infer sister-group relationships. The phylogenetic analysis of majids suggested that the position of Mithracinae is still uncertain, as is its monophyletic status. We recommend that additional characters, particularly of the megalopa phase, be sought for a better resolution of majid evolutionary history.

The phylogenetic relationships of the charismatic poster frogs, Phyllomedusinae (Anura, Hylidae)

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)