A phylogenetic study of the Anopheles punctulatus group of malaria vectors comparing rDNA sequence alignments derived from the mitochondrial and nuclear small ribosomal subunits

A phylogenetic study of the members of the Anopheles punctulatus group was performed using structural and similarity-based DNA sequence alignments of the small ribosomal subunit (SSU) from both the nuclear and the mitochondrial genomes. The mitochondrial SSU gene (12S, approximately 650 bp) proved to be highly restricted by its secondary structure and displayed little informative sequence variation. Consequently, it was considered unsuitable for a phylogenetic study of these closely related mosquito species. A structural alignment of the nuclear ribosomal DNA SSU (18S, approximately 2000 bp) proved to be more informative than similarity-based alignments. Analyses showed the A. punctulatus group to be monophyletic with two major clades; a Farauti clade containing members displaying an all-black-scaled proboscis (A. farauti 1-3 and 5-7) and the Punctulatus clade containing members displaying extensive white scaling on the apical half of the proboscis (A. farauti 4, A. punctulatus, and An. sp. near punctulatus). Anopheles koliensis was positioned basal to the Farauti clade.     

Differential ecology of Anopheles punctulatus and three members of the Anopheles farauti complex of mosquitoes on Guadalcanal, Solomon Islands, identified by PCR-RFLP analysis

From a series of larval collections made across northern Guadalcanal during the dry season, October–November 1997, four members of the Anopheles punctulatus group of mosquitoes (Diptera: Culicidae) were identified using PCR‐RFLP analysis. Anopheline larvae were found in 54/57 (95%) of the sites sampled, comprising An. farauti Laveran sensu stricto (32 sites), An. farauti species no. 2 (39 sites), An. farauti no. 7 (36 sites) and An. punctulatus Dönitz (10 sites). Anopheles punctulatus occurred only on the coastal plain, where it was associated with the more transient sites. Anopheles farauti sensu lato was more widespread throughout the survey region, with similar proportions of all three sibling species in both transient and permanent sites. Two members of the An. farauti complex, An. farauti s.s. and species no. 2, were found in brackish water. All breeding sites of An. punctulatus were cohabited by An. farauti s.l., sometimes by all three sibling species. Anopheles farauti s.s. was the only species collected on human bait, with a much higher biting rate early in the evening (57 bites/human/hour at 18.30–20.00 hours) than later (0.8 bites/human/hour at 21.00–24.00 hours).     

The mosquito Anopheles (Cellia) oreios sp. n., formerly species 6 of the Australasian Anopheles farauti complex,and a critical review of its biology and relation to disease

Species 6 of the Australasian Anopheles farauti sibling species complex (Diptera: Culicidae) is described and formally named Anopheles oreios Bangs & Harbach, sp. n. Adult, pupal and fourth‐instar larval specimens collected in the Baliem Valley, Papua Province, Indonesia, are characterized and compared with those of Anopheles farauti, Anopheles hinesorum, Anopheles irenicus and Anopheles torresiensis (formerly informally denoted as species 1, 2, 7 and 3, respectively). The variable wings of adult females, the male genitalia, the pupa and the fourth‐instar larva of An. oreios are illustrated and DNA sequence data are included for regions coding for sections of the mitochondrial COI and COII genes. The biology of An. oreios and its relation to malaria transmission are discussed in detail and contrasted with the biology and disease relations of some members of the An. farauti and Anopheles punctulatus sibling species complexes.     

Morphological studies on the Anopheles punctulatus Dönitz complex

SYNOPSIS An account is given of the member species of the Anopheles punctulatus Dönitz complex viz. A. punctulatus Dönitz, A. koliensis Owen, and 2 species previously recognised as A. farauti Laveran and which are now designated A. farauti No. 1 and A. farauti No. 2. Many of the eggs of A. farauti No. 1 were distinctive but the eggs of the other 3 species were similar. Keys for the identification of larve, pupae and adults are given.     

Population structure, mitochondrial polyphyly and the repeated loss of human biting ability in anopheline mosquitoes from the southwest Pacific

Australia and New Guinea contain high levels of endemism and biodiversity, yet there have been few evaluations of population‐level genetic diversity in fauna occurring throughout the Australo‐Papuan region. Using extensive geographical sampling, we examined and compared the phylogenetic relationships, phylogeography and population structure of Anopheles farauti, An. hinesorum and An. irenicus throughout their ranges in the southwest Pacific using mitochondrial (mtDNA COI) and nuclear (ribosomal protein S9 and ribosomal DNA ITS2) loci. Phylogenetic analyses suggest that the ability to utilize humans as hosts has been lost repeatedly, coincident with independent colonizations of the Solomon Islands. As some of the species under investigation transmit malaria in the region, this is a medically important finding. Maximum likelihood and Bayesian phylogenetic analyses of nuclear loci also showed that the three species are monophyletic. However, putative introgression of An. hinesorum mtDNA onto a nuclear background of An. farauti was evident in populations from Queensland, Torres Strait and southern New Guinea. Haplotype networks and pairwise F_ST values show that there is significant genetic structure within New Guinea and Australia in both An. farauti and An. hinesorum, consistent with a long‐term history of low gene flow among populations.     

Allozyme analysis reveals six species within the Anopheles punctulatus complex of mosquitoes in Papua New Guinea

Abstract. Among samples collected from nineteen localities in Papua New Guinea, we have identified six species within the Anopheles punctulatus complex of mosquitoes, by means of cellulose acetate allozyme electrophoresis. An.punctulatus Dönitz sensu stricto was collected from seven villages in the Madang area and from Buksak, Sausi Mission and an area 18 km SW of Tari; An.koliensis Owen from eight villages in the Madang area, from Popondetta and Brown River near Karema; and An.farauti No. 1 from ten coastal areas including Madang, Lorengau, Popondetta, Port Moresby, Rabaul and Wewak. Three newly recognized species, reported here for the first time, are designated as An.farauti No. 4 from Gonoa and Hudini, Madang area; An.farauti No. 5 from Ketarabo near Goroka; and An.farauti No. 6 from Hiwanda near Tari. Three other known members of the complex, An.clowi Rozeboom & Knight, An.farauti No. 2 (Bryan, 1973) and An.farauti No. 3 (Mahon & Meithke, 1982) were not detected in Papua New Guinea. Problems arising with morphological characters for the identification of species in this group are discussed.     

Multiple Independent Losses of Photosynthesis and Differing EvolutionaryRates in the Genus Cryptomonas (Cryptophyceae): Combined Phylogenetic Analyses of DNA Sequences of the Nuclear and the Nucleomorph Ribosomal Operons

In this study, evidence for at least three independent losses of photosynthesis in the freshwater cryptophyte genus Cryptomonas is presented. The phylogeny of the genus was inferred by molecular phylogenetic analyses of the nuclear internal transcribed spacer 2 (nuclear ITS2), partial nuclear large subunit ribosomal DNA (LSU rDNA), and nucleomorph small subunit ribosomal DNA (SSU rDNA, NM). Both concatenated and single data sets were used. In all data sets, the colorless Cryptomonas strains formed three different lineages, always supported by high bootstrap values (maximum parsimony, neighbor joining and maximum likelihood) and posterior probabilities (Bayesian analyses). The three leukoplast-bearing lineages displayed differing degrees of accelerated evolutionary rates in nuclear and nucleomorph rDNA. Also an increase in A+T-content in highly variable regions of the nucleomorph SSU rDNA was observed in one of the leukoplast-bearing lineages.This article contains three online-only supplementary tables.Reviewing Editor: Dr. Yves Van de Peer     

小麦抗白粉病相关基因GST克隆与表达

以从小麦抗白粉病相关基因差异表达分析中获得的EST-3 (Genbank序列号EX567360)为标签,采用电子克隆的方法对其进行延伸,并对电子克隆结果进行半定量RT-PCR验证,最后对白粉菌不同侵染时间进行了表达分析.经RT-PCR扩增,EST-3表达的带型变化趋势与其在抑制性消减杂交SSH-cDNA的差异显示情况一致,且RT-PCR获得的序列与电子克隆的序列一致性达98％.生物信息学分析表明,该序列是由875 bp核苷酸组成的,具有完整的开放阅读框架,编码蛋白为229个氨基酸,GenBank序列号JK841279,含有一个N端和C端谷胱甘肽硫转移酶结构域,该序列与小麦谷胱甘肽硫转移酶基因(GST)一致性较高,达97％.表达分析结果显示,白粉菌侵染24 h表达受到抑制,48 h开始表达,侵染72 h表达最强,96 h又开始下降,表明GST基因属于白粉菌诱导型相关基因,参与小麦对白粉病的应答反应.     

Electrophoretic keys to identify members of the Anopheles punctulatus complex of vector mosquitoes in Papua New Guinea

Abstract. Electrophoretic keys are given for the six species of the Anopheles punctulatus complex (Diptera: Culicidae) known from Papua New Guinea plus An.farauti No. 2 and No. 3 from Australia. The categories ‘faster’, ‘standard’ and ‘slower’ are used in keys to relate allozyme band migration following cellulose acetate electrophoresis to the standard pattern. Alternative keys are given depending on the availability of different species for use as standards.     

基于cpDNA和rDNA ITS片段分析西南地区青杨派杨树的系统发育与进化关系

西南地区青杨派杨树种质资源丰富,可为杨树遗传改良提供珍贵的基因资源,但树种之间形态学差异细微,该研究以山杨作为外类群,测定了西南地区及其他地区杨属青杨派17个种(杂种)共36份样本的3个叶绿体片段(atpF-atpH、trnL-F和matK)和核糖体ITS片段,并对其进行系统发育分析,以探讨西南地区青杨派树种的系统进化关系。结果表明:(1)在所有样本中,3个叶绿体片段atpF-atpH、trnL-F、matK的长度分别为605~634bp、957~1 010bp、819bp,3个片段拼接后的联合序列包含29个变异位点和15个信息位点;ITS片段对齐后的长度为646bp,变异位点19个,信息位点17个。(2)基于叶绿体联合序列和ITS片段的杨属青杨派树种的平均遗传距离分别为0.001 3和0.003 6。叶绿体联合片段的MP和Bayes系统树树型基本一致,青杨派树种可以划分为2组,第1组由青杨、三脉青杨、大青杨和辽杨构成;第2组中的小叶杨、小青杨、川杨、德钦杨、昌都杨、乡城杨、康定杨、西南杨、滇杨和藏川杨不能有效区分,且均与缘毛杨的遗传关系较近。(3)基于ITS的系统树与叶绿体联合片段构建的系统树差异不大,仅在于第2组中的小青杨与小叶杨、川杨等差异明显,与第1组中的树种紧密聚拢。     

Genetic diversity in two sibling species of the <Emphasis Type="Italic">Anopheles punctulatus</Emphasis> group of mosquitoes on Guadalcanal in the Solomon Islands

Background  

The mosquito Anopheles irenicus, a member of the Anopheles punctulatus group, is geographically restricted to Guadalcanal in the Solomon Islands. It shows remarkable morphological similarities to one of its sibling species, An. farauti sensu stricto (An. farauti s.s.), but is dissimilar in host and habitat preferences. To infer the genetic variations between these two species, we have analyzed mitochondrial cytochrome oxidase subunit II (COII) and nuclear ribosomal internal transcribed spacer 2 (ITS2) sequences from Guadalcanal and from one of its nearest neighbours, Malaita, in the Solomon Islands.     

Phylogeny of Rhigonematomorpha based on the complete mitochondrial genome of Rhigonema thysanophora (Nematoda: Chromadorea)

We determined the complete mitochondrial genome sequence of Rhigonema thysanophora, the first representative of Rhigonematomorpha, and used this sequence along with 57 other nematode species for phylogenetic analyses. The R. thysanophora mtDNA is 15 015 bp and identical to all other chromadorean nematode mtDNAs published to date in that it contains 36 genes (lacking atp8) encoded in the same direction. Phylogenetic analyses of nucleotide and amino acid sequence data for the 12 protein‐coding genes recovered Rhigonematomorpha as the sister group to the heterakoid species, Ascaridia columbae (Ascaridomorpha). The organization of R. thysanophora mtDNA resembles the most common pattern for the Rhabditomorpha+Ascaridomorpha+Diplogasteromorpha clade in gene order, but with some substantial gene rearrangements. This similarity in gene order is in agreement with the sequence‐based analyses that indicate a close relationship between Rhigonematomorpha and Rhabditomorpha+Ascaridomorpha+Diplogasteromorpha. These results are consistent with certain analyses of nuclear SSU rDNA for R. thysanophora and some earlier classification systems that asserted phylogenetic affinity between Rhigonematomorpha and Ascaridomorpha, but inconsistent with morphology‐based phylogenetic hypotheses that suggested a close (taxonomic) relationship between rhigonematomorphs and oxyuridomorphs (pinworms). These observations must be tempered by noting that few rhigonematomorph species have been sequenced and included in phylogenetic analyses, and preliminary studies based on SSU rDNA suggest the group is not monophyletic. Additional mitochondrial genome sequences of rhigonematids are needed to characterize their phylogenetic relationships within Chromadorea, and to increase understanding of mitochondrial genome evolution.     

Independent terrestrial origins of the Halosphaeriales (marine Ascomycota)

A phylogenetic study of marine ascomycetes was initiated to test and refine evolutionary hypotheses of marine-terrestrial transitions among ascomycetes. Taxon sampling focused on the Halosphaeriales, the largest order of marine ascomycetes. Approximately 1050 base pairs (bp) of the gene that codes for the nuclear small subunit (SSU) and 600 bp of the gene that codes for the nuclear large subunit (LSU) ribosomal RNAs (rDNA) were sequenced for 15 halosphaerialean taxa and integrated into a data set of homologous sequences from terrestrial ascomycetes. An initial set of phylogenetic analyses of the SSU rDNA from 38 taxa representing 15 major orders of the phylum Ascomycota confirmed a close phylogenetic relationship of the halosphaerialean species with several other orders of perithecial ascomycetes. A second set of analyses, which involved more intensive taxon sampling of perithecial ascomycetes, was performed using the SSU and LSU rDNA data in combined analyses. These second analyses included 15 halosphaerialean taxa, 26 terrestrial perithecial fungi from eight orders, and five outgroup taxa from the Pezizales. In these analyses the Halosphaeriales were polyphyletic and comprised two distinct lineages. One clade of Halosphaeriales comprised 12 taxa from 11 genera and was most closely related to terrestrial fungi of the Microascales. The second clade of halosphaerialean fungi comprised taxa from the genera Lulworthia and Lindra and was an isolated lineage among the perithecial fungi. Both the main clade of Halosphaeriales and the Lulworthia/Lindra clade are supported by the data as being independently derived from terrestrial ancestors.     

Symbiont diversity in scleractinian corals from tropical reefs and subtropical non-reef communities in Taiwan

We examined zooxanthellae diversity in scleractinian corals from southern Taiwan and the Penghu Archipelago, a tropical coral reef and a subtropical non-reefal community, respectively. Zooxanthellae diversity was investigated in 52 species of scleractinian corals from 26 genera and 13 families, using restriction fragment length polymorphism (RFLP), and phylogenetic analyses of the nuclear small-subunit ribosomal DNA (nssrDNA) and large-subunit ribosomal DNA (nlsrDNA). RFLP and phylogenetic analyses of nuclear-encoded ribosomal RNA genes showed that Symbiodinium clade C was the dominant zooxanthellae in scleractinian corals in the seas around Taiwan; Symbiodinium clade D was also found in some species. Both Symbiodinium clade C and D were found in colonies of seven species of scleractinian corals. Symbiodinium clade D was associated with corals that inhabit either shallow water or the reef edge in deep water, supporting the hypothesis that Symbiodinium clade D is a relatively stress-tolerant zooxanthellae found in marginal habitats.Communicated by Biological Editor H.R. Lasker     

Complete mitochondrial genome of Tubulipora flabellaris (Bryozoa: Stenolaemata): the first representative from the class Stenolaemata with unique gene order

Mitochondrial genomes play a significant role in the reconstruction of phylogenetic relationships within metazoans. There are still many controversies concerning the phylogenetic position of the phylum Bryozoa. In this research, we have finished the complete mitochondrial genome of one bryozoan (Tubulipora flabellaris), which is the first representative from the class Stenolaemata. The complete mitochondrial genome of T. flabellaris is 13,763 bp in length and contains 36 genes, which lacks the atp8 gene in contrast to the typical metazoan mitochondrial genomes. Gene arrangement comparisons indicate that the mitochondrial genome of T. flabellaris has unique gene order when compared with other metazoans. The four known bryozoans complete mitochondrial genomes also have very different gene arrangements, indicates that bryozoan mitochondrial genomes have experienced drastic rearrangements. To investigate the phylogenetic relationship of Bryozoa, phylogenetic analyses based on amino acid sequences of 11 protein coding genes (excluding atp6 and atp8) from 26 metazoan complete mitochondrial genomes were made utilizing Maximum Likelihood (ML) and Bayesian methods, respectively. The results indicate the monopoly of Lophotrochozoa and a close relationship between Chaetognatha and Bryozoa. However, more evidences are needed to clarify the relationship between two groups. Lophophorate appeared to be polyphyletic according to our analyses. Meanwhile, neither analysis supports close relationship between Branchiopod and Phoronida. Four bryozoans form a clade and the relationship among them is T. flabellaris + (F. hispida + (B. neritina + W. subtorquata)), which is in coincidence with traditional classification system.     

Phylogenetic relationships within the genus Tetrahymena inferred from the cytochrome c oxidase subunit 1 and the small subunit ribosomal RNA genes

Details of the phylogenetic relationships among tetrahymenine ciliates remain unresolved despite a rich history of investigation with nuclear gene sequences and other characters. We examined all available species of Tetrahymena and three other tetrahymenine ciliates, and inferred their phylogenetic relationships using nearly complete mitochondrial cytochrome c oxidase subunit 1 (cox1) and small subunit (SSU) rRNA gene sequences. The inferred phylogenies showed the genus Tetrahymena to be monophyletic. The three “classical” morphology-and-ecology-based groupings are paraphyletic. The SSUrRNA phylogeny confirmed the previously established australis and borealis groupings, and nine ribosets. However, these nine ribosets were not well supported. Using cox1 gene, the deduced phylogenies based on this gene revealed 12 well supported groupings, called coxisets, which mostly corresponded to the nine ribosets. This study demonstrated the utility of cox1 for resolving the recent phylogeny of Tetrahymena, whereas the SSU rRNA gene provided resolution of deeper phylogenetic relationships within the genus.     

Serial replacement of a diatom endosymbiont in the marine dinoflagellate Peridinium quinquecorne (Peridiniales, Dinophyceae)

To infer the phylogeny of both the host and the endosymbiont of Peridinium quinquecorne Abé, the small subunit (SSU) ribosomal DNA (rDNA) from the host and two genes of endosymbiont origin (plastid‐encoded rbcL and nuclear‐encoded SSU rDNA) were determined. The phylogenetic analysis of the host revealed that the marine dinoflagellate P. quinquecorne formed a clade with other diatom‐harbouring dinoflagellates, including Kryptoperidinium foliaceum (Stein) Lindeman, Durinskia baltica (Levander) Carty et Cox and Galeidinium rugatum Tamura et Horiguchi, indicating a single endosymbiotic event for this lineage. Phylogenetic analyses of the endosymbiont in these organisms revealed that the endosymbiont of P. quinquecorne formed a clade with a centric diatom (SSU data indicated it to be closely related to Chaetoceros), whereas the endosymbionts of other three dinoflagellates formed a clade with a pennate diatom. The discrepancy between the host and the endosymbiont phylogenies suggests a secondary replacement of the endosymbiont from a pennate to a centric diatom in P. quinquecorne.     

Multilocus population genetic analysis of the Southwest Pacific malaria vector Anopheles punctulatus

The population structure and history of the cryptic malaria vector species, Anopheles punctulatus (Doenitz), was investigated throughout Papua New Guinea and the Solomon Islands with the aim of detailing genetic subdivisions and the potential for movement through this biogeographically complex region. We obtained larval collections from over 80 sites and utilised a diverse array of molecular markers that evolve through different processes. Individuals were initially identified to species and genotyped using the ribosomal DNA second internal transcribed spacer. DNA sequencing of a single copy nuclear ribosomal protein S9 and the mitochondrial cytochrome oxidase I loci were then investigated and 12 nuclear microsatellite markers were developed and analysed. Our data revealed three genetically distinct populations – one in Papua New Guinea, the second on Buka Island (Bougainville Province, Papua New Guinea), and the third on Guadalcanal Island (Solomon Islands). Genetic differentiation within Papua New Guinea was much lower than that found in studies of other closely related species in the region. The data does suggest that A. punctulatus has undergone a population bottleneck followed by a recent population and range expansion in Papua New Guinea. Humans and regional economic growth may be facilitating this population expansion, as A. punctulatus is able to rapidly occupy human modified landscapes and traverse unsealed roads. We therefore anticipate extensive movement of this species through New Guinea – particularly into the highlands, with a potential increase in malaria frequency in a warming climate – as well as relatively unrestricted gene flow of advantageous alleles that may confound vector control efforts.     

Taxonomy, molecular phylogeny, and ultrastructural morphology of Olpidiopsis porphyrae sp. nov. (Oomycetes, straminipiles), a unicellular obligate endoparasite of Bangia and Porphyra spp. (Bangiales, Rhodophyta)

Olpidiopsis porphyrae sp. nov., a marine oomycete endoparasite that infects the commercially cultivated red alga Porphyra yezoensis, is described and its phylogenetic position based on molecular data and ultrastructural morphology is discussed. O. porphyrae infects the host Porphyra by means of encysted zoospores. Spherical-shaped holocarpic thalli develop within the cytoplasm of its algal host, which produce monoplanetic, subapically biflagellate zoospores. The characteristic features of this isolate are the ellipsoidal, unicellular thallus and simple holocarpic zoosporangial development, which show morphological similarity with the genus Olpidiopsis. Laboratory infection experiments with a wide range of green, brown, and red algae revealed that O. porphyrae infects several stages of the bangialean red algae (the genera Bangia and Porphyra). Molecular phylogenetic analyses inferred from both SSU rRNA and cox2 genes showed O. porphyrae branched before the main saprolegnian and peronosporalean lineages within the monophyletic oomycete clade, indicating its phylogenetic separation from them. A single or double K-body-like organelle, which contains tubular inclusions, is found located to one side of the zoospore nucleus and shows similarities to homologous organelles previously described in O. saprolegniae. The ultrastructural morphology of O. porphyrae with zoospore initials containing K-bodies and tubular mitochondrial cristae is characteristic of oomycetes. Group I intron-like multiple insertions were found in the SSU rRNA gene of O. porphyrae. This is the first report of SSU group I introns in the class Oomycetes.     

Generic concept in Chlorella‐related coccoid green algae (Chlorophyta,Trebouxiophyceae)

Using a combined set of sequences of SSU and ITS regions of nuclear‐encoded ribosomal DNA, the concept of the experimental algal genus Chlorella was evaluated. Conventionally in the genus Chlorella, only coccoid, solitary algae with spherical morphology that do not possess any mucilaginous envelope were included. All Chlorella species reproduce asexually by autospores. However, phylogenetic analyses showed that within the clade of ‘true’Chlorella species (Chlorella vulgaris, C. lobophora, and C. sorokiniana), taxa with a mucilaginous envelope and colonial lifeform have also evolved. These algae, formerly designated as Dictyosphaerium, are considered as members of the genus Chlorella. In close relationship to Chlorella, five different genera were supported by the phylogenetic analyses: Micractinium (spherical cells, colonial, with bristles), Didymogenes (ellipsoidal cells, two‐celled coenobia, with or without two spines per cell), Actinastrum (ellipsoidal cells within star‐shaped coenobia), Meyerella (spherical cells, solitary, without pyrenoids), and Hegewaldia (spherical cells, colonial, with or without bristles, oogamous propagation). Based on the secondary structures of SSU and ITS rDNA sequences, molecular signatures are provided for each genus of the Chlorella clade.