Phylogenetic relationships of Korean Sparassis latifolia based on morphological and ITS rDNA characteristics

Recent studies based on morphological characteristics and molecular analyses have revealed that the characteristics of Sparassis crispa from Asia are not concordant with those of collections from Europe and North America. Consequently, the Asian isolate was redefined as Sparassis latifolia. This study is the first report of Sparassis latifolia collected in Korea. The taxonomic relationships and replacement of Sparassis species were inferred from a comparison of the morphological characteristics and by molecular sequence analysis of the internal transcribed spacer (ITS) rDNA regions. In particular, this study focused on the phylogenetic relationships inferred from the biogeographical distribution of isolates within the genus Sparassis.     

Multigene Phylogeography of Bactrocera caudata (Insecta: Tephritidae): Distinct Genetic Lineages in Northern and Southern Hemispheres

Bactrocera caudata is a pest of pumpkin flower. Specimens of B. caudata from the northern hemisphere (mainland Asia) and southern hemisphere (Indonesia) were analysed using the partial DNA sequences of the nuclear 28S rRNA and internal transcribed spacer region 2 (ITS-2) genes, and the mitochondrial cytochrome c oxidase subunit I (COI), cytochrome c oxidase subunit II (COII) and 16S rRNA genes. The COI, COII, 16S rDNA and concatenated COI+COII+16S and COI+COII+16S+28S+ITS-2 nucleotide sequences revealed that B. caudata from the northern hemisphere (Peninsular Malaysia, East Malaysia, Thailand) was distinctly different from the southern hemisphere (Indonesia: Java, Bali and Lombok), without common haplotype between them. Phylogenetic analysis revealed two distinct clades (northern and southern hemispheres), indicating distinct genetic lineage. The uncorrected ‘p’ distance for the concatenated COI+COII+16S nucleotide sequences between the taxa from the northern and southern hemispheres (‘p’ = 4.46-4.94%) was several folds higher than the ‘p’ distance for the taxa in the northern hemisphere (‘p’ = 0.00-0.77%) and the southern hemisphere (‘p’ = 0.00%). This distinct difference was also reflected by concatenated COI+COII+16S+28S+ITS-2 nucleotide sequences with an uncorrected ''p'' distance of 2.34-2.69% between the taxa of northern and southern hemispheres. In accordance with the type locality the Indonesian taxa belong to the nominal species. Thus the taxa from the northern hemisphere, if they were to constitute a cryptic species of the B. caudata species complex based on molecular data, need to be formally described as a new species. The Thailand and Malaysian B. caudata populations in the northern hemisphere showed distinct genetic structure and phylogeographic pattern.     

Genetic structure of three Sorbus latifolia (Lam.) Pers. taxa endemic to northern Bavaria

The Franconian Alb (Bavaria, Germany) is rich in endemic Sorbus taxa, considered as apomictic microspecies and derived by hybridization between Sorbus aria aggregate and Sorbus torminalis (Sorbus latifolia aggregate). Molecular studies using the AFLP technique, neighbour joining, Bayesian clustering, principal coordinate analysis (PCo) and voucher studies were used to investigate genetic structure and origin of adult plants and cultivated offspring of three endemic S. latifolia taxa, namely Sorbus cordigastensis, Sorbus franconica and Sorbus adeana and probable parental species from the S. aria agg. and S. torminalis. The S. latifolia taxa, adults and progeny, showed low genetic variability and a more or less clonal structure, confirming an apomictic mode of reproduction. The investigated S. latifolia taxa were remarkably different among each other, confirming their status as microspecies. The AFLP data confirmed the hybrid origin of the S. latifolia taxa, they were 1.3–1.5 times more closely related to S. aria agg. than to S. torminalis. The S. aria agg. showed a complicate genetic structure and fell into four main groups, two intermediate groups besides Sorbus pannonica and Sorbus aria sensu stricto (S. aria s.str.). Some progeny of S. pannonica was more variable than expected and clustered partly with other groups indicating gene flow within S. aria agg. Different subgroups of the S. aria aggregate may be parental for the S. latifolia taxa, contributing to the remarkable genetic distances between them.     

Two hidden taxa in the Japanese encephalitis vector mosquito,Culex tritaeniorhynchus,and the potential for long-distance migration from overseas to Japan

The Culex vishnui subgroups, particularly Culex tritaeniorhynchus, are considered the primary vectors of the Japanese encephalitis virus (JEV) in Asia. Recent molecular phylogenetic analyses of JEV isolates from Asian countries have shown that JEVs with diverse genetic variants are present in Asia. Furthermore, some JEV strains have been found to have crossed the East China Sea and been introduced into Japan. In this study, the possibility of overseas migration of the JE vector mosquito, Cx. tritaeniorhynchus was examined from the genetic, physical, and meteorological perspectives. Molecular phylogenetic analysis was performed based on both whole coding sequences and on the barcoding region of the mitochondrial cytochrome c oxidase subunit I (COI) gene of Cx. vishnui subgroups collected from Asian countries. Culex tritaeniorhymchus was classified into two genetically independent taxa by COI sequences: the Japanese type (Ct-J), which inhabits Japan except for the Amami Islands of southern Japan, and the continental type (Ct-C), which inhabits the Asian region except for Japan. It was confirmed that approximately 10% of Cx. tritaeniorhynchus trapped during the summer in western Kyushu were Ct-C, and that they could fly for up to 38 h continuously. The meteorological analysis also confirmed that the atmospheric flow occurring over the continent coincided with the date of Ct-C capture. This is the first report showing the existence of two taxa in Cx. tritaeniorhynchus. Their physical and physiological characteristics suggest the possibility of long-distance migration from overseas regions to Japan across the East China Sea. Future efforts are expected to provide evidence to support the occurrence of long-distance migration of Cx. tritaeniorhynchus with JEV.     

Comparison of Visceral Leishmaniasis Diagnostic Antigens in African and Asian Leishmania donovani Reveals Extensive Diversity and Region-specific Polymorphisms

Background

Visceral leishmaniasis (VL), caused by infection with Leishmania donovani complex, remains a major public health problem in endemic regions of South Asia, East Africa, and Brazil. If untreated, symptomatic VL is usually fatal. Rapid field diagnosis relies principally on demonstration of anti-Leishmania antibodies in clinically suspect cases. The rK39 immunochromatographic rapid diagnostic test (RDT) is based on rK39, encoded by a fragment of a kinesin-related gene derived from a Brazilian L. chagasi, now recognised as L. infantum, originating from Europe. Despite its reliability in South Asia, the rK39 test is reported to have lower sensitivity in East Africa. A reason for this differential response may reside in the molecular diversity of the rK39 homologous sequences among East African L. donovani strains.

Methodology/Principal Findings

Coding sequences of rK39 homologues from East African L. donovani strains were amplified from genomic DNA, analysed for diversity from the rK39 sequence, and compared to South Asian sequences. East African sequences were revealed to display significant diversity from rK39. Most coding changes in the 5′ half of repeats were non-conservative, with multiple substitutions involving charge changes, whereas amino acid substitutions in the 3′ half of repeats were conservative. Specific polymorphisms were found between South Asian and East African strains. Diversity of HASPB1 and HASPB2 gene repeat sequences, used to flank sequences of a kinesin homologue in the synthetic antigen rK28 designed to reduce variable RDT performance, was also investigated. Non-canonical combination repeat arrangements were revealed for HASPB1 and HASPB2 gene products in strains producing unpredicted size amplicons.

Conclusions/Significance

We demonstrate that there is extensive kinesin genetic diversity among strains in East Africa and between East Africa and South Asia, with ample scope for influencing performance of rK39 diagnostic assays. We also show the importance of targeted comparative genomics in guiding optimisation of recombinant/synthetic diagnostic antigens.     

Genetic relationship and genetic diversity among Typha taxa from East Asia based on AFLP markers

The genetic relationship and diversity among four Typha taxa in East Asia were evaluated using amplified fragment length polymorphism (AFLP) markers. Three AFLP selective primer combinations generated a total of 707 amplification products, of which 704 (99.6%) were polymorphic. The unweighted pair-group method with arithmetic mean (UPGMA) dendrogram and principal component analysis (PCA) plot confirmed the taxonomic status of four separate species. East Asian Typha taxa separated into two groups: the first with Typha angustifolia and the second with T. orientalis, T. laxmanni, and T. latifolia with a high bootstrap value for UPGMA (93%) and a low first score for PCA (25%). The two clusters corresponded with two sections based on the bracteoles in the female flower: section Bracteolatae and section Ebracteolatae. T. angustifolia showed the highest genetic diversity among the four Typha taxa (percentage of polymorphic loci [PPL] = 71%, H_o = 0.157), whereas T. latifolia had the lowest genetic diversity (PPL = 40%, H_o = 0.117). Genetic diversity was related to the presence of the gap between male and female inflorescences. A positive correlation between genetic distance and geographic distance was clearly found in the two species with continuous inflorescences (T. latifolia and T. orientalis). This positive correlation was not observed in the other species with discontinuous spikes (T. angustifolia and T. laxmanni).     

A new Sparassis species from Spain described using morphological and molecular data

Sparassis miniensis, collected in Pinus pinaster forests in Galicia (northwest Iberian Peninsula) is described as a new species, based on morphological and molecular data. Sparassis miniensis is morphologically distinct from all other species in the genus Sparassis based on scattered flabellae, which are strongly laciniated, azonate, and arise from an orange to rose-purplish base. The sporadic presence of clamp connections is restricted to subhymenial hyphae. Molecular data from LSU-rDNA, ITS and partial gene coding RNA polymerase subunit II (rpb2) suggest a close relationship between the new species S. miniensis and S. brevipes, another European species producing large fruiting bodies but with entire flabellae and no clamp connections.     

Morphological and genetic characteristics of newly crossbred cauliflower mushroom (Sparassis latifolia)

Cauliflower mushroom (Sparassis latifolia or S. crispa) is popular for food and medicine. Importance of new varieties of Sparassis was raised and studied widely by protection system of UPOV. In this study, 10 crossbred strains of Sparassis latifolia that specifically expressed distinctive features during basidiocarp formation and mycelium growth were applied to sawdust medium inoculated with S. latifolia mycelia. The 10 crossbred strains were divided into 3 groups on the basis of morphological (size of marginal wave and basidiocarp color) and genetic characteristics. Each phenotype of the parent and crossbred strains represented 3 marginal wave-sizes (large, medium, and small) and 3 color notations (NN155D, 163C, and 8D). Our result suggests that morphological characteristics of cauliflower mushroom can be affected by various environmental and genetic stimuli under artificial conditions such as crossbreed. Also this research showed genetic differences among breeding isolates and their morphological characteristics were correlated with the molecular data within parent and crossed strain.     

The Methanol Dehydrogenase Gene,mxaF, as a Functional and Phylogenetic Marker for Proteobacterial Methanotrophs in Natural Environments

The mxaF gene, coding for the large (α) subunit of methanol dehydrogenase, is highly conserved among distantly related methylotrophic species in the Alpha-, Beta- and Gammaproteobacteria. It is ubiquitous in methanotrophs, in contrast to other methanotroph-specific genes such as the pmoA and mmoX genes, which are absent in some methanotrophic proteobacterial genera. This study examined the potential for using the mxaF gene as a functional and phylogenetic marker for methanotrophs. mxaF and 16S rRNA gene phylogenies were constructed based on over 100 database sequences of known proteobacterial methanotrophs and other methylotrophs to assess their evolutionary histories. Topology tests revealed that mxaF and 16S rDNA genes of methanotrophs do not show congruent evolutionary histories, with incongruencies in methanotrophic taxa in the Methylococcaceae, Methylocystaceae, and Beijerinckiacea. However, known methanotrophs generally formed coherent clades based on mxaF gene sequences, allowing for phylogenetic discrimination of major taxa. This feature highlights the mxaF gene’s usefulness as a biomarker in studying the molecular diversity of proteobacterial methanotrophs in nature. To verify this, PCR-directed assays targeting this gene were used to detect novel methanotrophs from diverse environments including soil, peatland, hydrothermal vent mussel tissues, and methanotroph isolates. The placement of the majority of environmental mxaF gene sequences in distinct methanotroph-specific clades (Methylocystaceae and Methylococcaceae) detected in this study supports the use of mxaF as a biomarker for methanotrophic proteobacteria.     

Spirometra species from Asia: Genetic diversity and taxonomic challenges

Despite considerable controversy concerning the taxonomy of species within the genus Spirometra, human sparganosis and spirometrosis mainly in Asia and Europe has long been confidently ascribed to Spirometra erinaceieuropaei. Recently, the mitochondrial genomes of purported “S. erinaceieuropaei”, “Spirometra decipiens” and “Spirometra ranarum” from Asia have been determined. However, it has been pointed out that the morphological criteria used for identifying these species are unsuitable and thus these identifications are questionable. In the present study, therefore, Spirometra samples from Asia were re-examined based on mitochondrial cytochrome c oxidase subunit 1 gene sequences and the identification of these species was discussed. Haplotype network and phylogenetic analyses revealed that: i) two distinct Spirometra species, Type I and Type II, are present in Asia and neither of which is close to likely European “S. erinaceieuropaei”; ii) Type I is genetically diverse and widely distributed, however Type II is known so far from Japan and Korea; iii) “S. decipiens” and “S. ranarum” reported from Asia are conspecific with Type I; iv) Type I is probably conspecific with Spirometra mansoni, and Type II may represent an undescribed species.     

Genetic diversity and geographic distribution of Bemisia tabaci species complex in Nepal

Bemisia tabaci species complex comprises at least 44 cryptic species worldwide. Here, we analyze the nucleotide sequences of mitochondrial cytochrome oxidase subunit 1 (COI) gene obtained from 76 samples of B. tabaci collected from 23 districts in Nepal. This is the first genetic and geographic study of B. tabaci species complex in Nepal. Our phylogenetic study identified the presence of three cryptic species—Asia I, Asia II 1, and Asia II 5—with high interspecific but low intraspecific variations. Among the three cryptic species, Asia II 5 was the most prevalent in Nepal, constituting 64.47% of all the sequenced samples. Based on haplotype network analysis of COI sequences, Asia II 1 was more genetically diversified than the other two cryptic species. Our results provided useful information on the genetic diversity and geographic distribution of B. tabaci in Nepal, which help monitor population changes of B. tabaci at cryptic species level and develop sustainable management strategies for its control.     

RPS8—a New Informative DNA Marker for Phylogeny of Babesia and Theileria Parasites in China

Piroplasmosis is a serious debilitating and sometimes fatal disease. Phylogenetic relationships within piroplasmida are complex and remain unclear. We compared the intron–exon structure and DNA sequences of the RPS8 gene from Babesia and Theileria spp. isolates in China. Similar to 18S rDNA, the 40S ribosomal protein S8 gene, RPS8, including both coding and non-coding regions is a useful and novel genetic marker for defining species boundaries and for inferring phylogenies because it tends to have little intra-specific variation but considerable inter-specific difference. However, more samples are needed to verify the usefulness of the RPS8 (coding and non-coding regions) gene as a marker for the phylogenetic position and detection of most Babesia and Theileria species, particularly for some closely related species.     

An ITS-based phylogenetic framework for the genus Vorticella: finding the molecular and morphological gaps in a taxonomically difficult group

Vorticella includes more than 100 currently recognized species and represents one of the most taxonomically challenging genera of ciliates. Molecular phylogenetic analysis of Vorticella has been performed so far with only sequences coding for small subunit ribosomal RNA (SSU rRNA); only a few of its species have been investigated using other genetic markers owing to a lack of similar sequences for comparison. Consequently, phylogenetic relationships within the genus remain unclear, and molecular discrimination between morphospecies is often difficult because most regions of the SSU rRNA gene are too highly conserved to be helpful. In this paper, we move molecular systematics for this group of ciliates to the infrageneric level by sequencing additional molecular markers—fast-evolving internal transcribed spacer (ITS) regions—in a broad sample of 66 individual samples of 28 morphospecies of Vorticella collected from Asia, North America and Europe. Our phylogenies all featured two strongly supported, highly divergent, paraphyletic clades (I, II) comprising the morphologically defined genus Vorticella. Three major lineages made up clade I, with a relatively well-resolved branching order in each one. The marked divergence of clade II from clade I confirms that the former should be recognized as a separate taxonomic unit as indicated by SSU rRNA phylogenies. We made the first attempt to elucidate relationships between species in clade II using both morphological and multi-gene approaches, and our data supported a close relationship between some morphospecies of Vorticella and Opisthonecta, indicating that relationships between species in the clade are far more complex than would be expected from their morphology. Different patterns of helix III of ITS2 secondary structure were clearly specific to clades and subclades of Vorticella and, therefore, may prove useful for resolving phylogenetic relationships in other groups of ciliates.     

The genus Laetiporus (Basidiomycota, Polyporales) in East Asia

Relationships among East Asian, North American and European Laetiporus sulphureus s. lat., a cosmopolitan brown rot species complex, were assessed with phylogenetic analyses and incompatibility tests. Three East Asian taxa, Laetiporus cremeiporus sp. nov., Laetiporus montanus and Laetiporus versisporus, are described and illustrated as well as compared with related taxa from Southeast Asia, North America and Europe. Phylogenetic analyses showed that L. cremeiporus and L. versisporus are clearly distinct species among Laetiporus taxa. The three conifer inhabiting species, Laetiporus conifericola, Laetiporus huroniensis and L. montanus, are closely related to each other. The European population of L. montanus exhibits two sequence variants of the EF1α: one is the same as observed in L. sulphureus in Europe and the other is that observed in East Asian population of L. montanus. A key to the known species of Laetiporus in the northern hemisphere is provided.     

Phylogenetic Distribution of Unusual Triheme to Tetraheme Cytochrome Subunit in the Reaction Center Complex of Purple Photosynthetic Bacteria

To understand the evolutionary relationship between triheme and tetraheme cytochrome subunits in the reaction center complex, genes located downstream of that coding for the M subunit of the reaction center complex (pufM) were amplified by PCR and analyzed in six established and two unidentified species of the genus Rhodovulum and five species of the genus Rhodobacter. All the Rhodovulum species tested had the pufC gene coding for the reaction-center-bound cytochrome subunit, while all the Rhodobacter species were found to have the pufX gene at the corresponding position. Analyses of the amino acid sequences of the pufC gene products showed that the cytochrome subunits of all the Rhodovulum species have three heme-binding-motifs and lack a methionine residue probably working as the sixth axial-ligand to one of the three hemes. Phylogenetic relationships among Rhodovulum species based on the pufC gene products were basically consistent with those based on 16S rRNA sequences, suggesting that the basic characteristics of the triheme cytochrome subunit have been conserved during the evolutionary process of the Rhodovulum species.     

Phylogeography and genetic diversity of the red squirrel (Sciurus vulgaris) in China: Implications for the species' postglacial expansion history

The geographical distribution of Sciurus vulgaris spans much of the Palearctic from western Europe and the UK eastward to the pacific coast of East Asia. S. vulgaris occurs in China only in the far northwest and the northeast. Understanding of the species’ postglacial expansion history in East Asia has been limited by the paucity of molecular data. In this study, we used partial D-loop and cytochrome b gene sequences to assess mitochondrial DNA variation in S. vulgaris in China. Our objectives were to (1) determine phylogeographical patterns of S. vulgaris in China; (2) understand the species’ postglacial expansion history in this region; and (3) quantify genetic diversity levels within S. vulgaris populations in China. We identified a supported phylogenetic group in S. vulgaris from China, and found no tendency for haplotypes to cluster by geographic region. Our analysis of S. vulgaris from China and other regions supports the hypothesis that the Calabria region of southern Italy is a glacial refugium for the species. We tentatively propose a postglacial expansion pattern for the squirrels: migrating from Calabria via Central and Eastern Europe to Russia and from there to China, and firstly to the northwest and then to northeast in China. We found high levels of genetic diversity in S. vulgaris populations across China as a whole, and discussed its influential factors.