Plagiochila cucullifolia var. anomala var. nov. from Ecuador, with notes on discordant molecular and morphological variation in Plagiochila

Plagiochila cucullifolia Jack & Steph. var. anomala J. Heinrichs & Gradst. var. nov. is described and illustrated. The new variety is known from a single locality in southern Ecuador and differs from P. cucullifolia var. cucullifolia by the flat, not saccate leaves, somewhat smaller plant size and weaker leaf dentation. According to phylogenetic analyses of 35 nrDNA ITS1 and ITS2 sequences of Plagiochila, the two varieties of P. cucullifolia form a monophyletic lineage and are placed in a well supported clade together with five other species of Plagiochila sect. Hylacoetes: P. dimorpha Lindenb. & Gottsche var. ecuadorica (Inoue) J. Heinrichs, P. flabelliflora Steph., P. patriciae J. Heinrichs & H. Anton, P. macrostachya Lindenb. and P. turgida Herzog. Within Plagiochila, nrITS sequence variation is not concordant with morphological diversification. ITS sequences of Plagiochila cucullifolia s.str. and of P. dimorpha var. ecuadorica differ in only 23 aligned positions whereas two sequences of P. subplana Lindenb. differ in 86 aligned positions. Morphologically, P. cucullifolia s.str. and P. dimorpha var. ecuadorica differ in more than 20 characters and previously these two taxa were placed in separate genera. P. subplana phenotypes show considerable variation in leaf shape and dentation but the extremes are linked by numerous intermediates.     

A taxonomic study of KoreanSmilacina

Several taxonomic problems of KoreanSmilacina were examined by statistical methods, based on morphological data. Consequently,S. davurica described in Korean flora differs greatly from originalS. davurica, and closely related toS. bicolor morphologically. Therefore, the present authors treated this new taxon as a variety ofS. bicolor, S. bicolor var.flavovirens N. S. Lee et J. Y. Kim var. nov (“Yondusomdae” in Korean). The distribution in South Korea ofS. bicolor var.bicolor is known as only Mt. Chiri, but this species is collected in Mt. Dukyou, also. The bounds of morphological variations ofS. japonica var.japonica, the short plant size and no hairs on the stem, were revealed. The morphological characteristics ofS. japonica var.japonica andS. japonica var.mandshurica is obscure to distinguish. According to the present study,S. japonica var.mandshurica differs fromS. japonica var.japonica in plant size, size of adaxial leaf epidermal cell, ratio of style/ovary length and shape of stigma. The unrecorded taxon in Korean flora,S. robusta, was given a new Korean name as “Kunsomdae.” AlthoughS. japonica var.mandshurica is the variety ofS. japonica var.japonica, it is more closely related toS. robusta than toS. japonica var.japonica.     

Bacterial Endosymbionts of Pyrodinium bahamense var. compressum

The study presents evidence in support of the bacterial theory associated with the toxicity of Pyrodinium bahamense var. compressum. Bacterial endosymbionts from Philippine P. bahamense var. compressum strain Pbc MZRVA 042595 were isolated and identified via 16S rDNA sequence analysis. Taxonomic diversity of the identified culturable intracellular microbiota associated with Philippine P. bahamense var. compressum was established to be limited to the Phyla Proteobacteria, Actinobacteria, and Firmicutes. Major endosymbionts identified included Moraxella spp., Erythrobacter spp., and Bacillus spp., whereas Pseudomonas putida, Micrococcus spp., and Dietzia maris were identified as minor isolates. All identified strains except D. maris, P. putida, and Micrococcus spp. were shown to contain either saxitoxin or neo saxitoxin or both at levels ≤73 ng/10⁷ bacterial cells based on high-performance liquid chromatography analysis. Paralytic shellfish poisoning-like physiologic reactions in test animals used in the mouse assay were recorded for the endosymbionts except for P. putida. The study is the first to elucidate the possible contribution of bacterial endosymbionts in the toxicity of P. bahamense var. compressum isolated in the Philippines.     

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.     

A new meristematic fungus,Pseudotaeniolina globosa

A new species of Pseudotaeniolina, a genus of anamorphic, melanized fungi with meristematic development, is described. The species is compared to morphologically similar taxa among which are Trimmatostroma and Coniosporium. Its novelty is supported by SSU (small subunit) and ITS (internal transcribed spacer) rDNA sequence data. This revised version was published online in June 2006 with corrections to the Cover Date.     

Genetic relatedness among Filobasidiella species

The three accepted species of Filobasidiella, F. neoformans, F. depauperata, and F. lutea, are compared morphologically and by molecular analysis. Sequences of the internally transcribed spacer (ITS) and the small subunit (SSU) gene of the ribosomal RNA (rRNA) gene cluster were obtained, and analysed by Neighbor-joining and Maximum parsimony methods. The three species of Filobsidiella are shown to form a single monophyletic clade, rooted by Tremella mesenterica. F. lutea was recovered as a distinct, but closely related taxon with the Filobasidiella clade. This is the first report of DNA sequences from herbarium specimens of F. lutea.This revised version was published online in October 2005 with corrections to the Cover Date.     

Taxonomic status of Monotropastrum humile, with special reference to M. humile var. glaberrimum (Ericaceae, Monotropoideae)

Taxonomic treatment of the achlorophyllous monotropoid plant Monotropastrum humile is still unclear and confusing because of the lack of detailed morphological analyses and molecular phylogeny. In particular, the taxonomic status of a glabrous variety, M. humile var. glaberrimum, is under debate. Our detailed examination of the morphological characteristics of living plants revealed that M. humile var. glaberrimum can be easily distinguished from the putative conspecific taxon M. humile var. humile by characteristics not previously recognized, namely the shape and color of the floral disc. Most morphological features characterizing Cheilotheca were also found in M. humile var. glaberrimum. Moreover, there was considerable nucleotide differentiation in the internal transcribed spacer (ITS)2 sequences of M. humile var. humile and var. glaberrimum. Molecular analysis of the phylogenetic relationship of M. humile var. humile, var. glaberrimum, and other monotropoids using ITS2 sequences showed that two varieties of M. humile formed a monophyletic clade with a member of a different genus, Monotropa L., but obvious phylogenetic relationships among these three taxa were not obtained. Thus we conclude that Monotropastrum humile var. glaberrimum should be treated as a distinct species. However, the generic affiliation of M. humile var. glaberrimum could not be determined because of its intermediate character state combination and the insufficient characterization of related species. We strongly suggest that Monotropastrum as a whole needs re-evaluation.     

Genetic variability and taxonomic revision of the genus Auxenochlorella (Shihira et Krauss) Kalina et Puncocharova (Trebouxiophyceae,Chlorophyta)

The monotypic genus Auxenochlorella with its type species A. protothecoides is so far only known from specific habitats such as the sap of several tree species. Several varieties were described according to physiological performances in culture on different organic substrates. However, two strains designated as Auxenochlorella were isolated from other habitats (an endosymbiont of Hydra viridis and an aquatic strain from an acidic volcano stream). We studied those isolates and compared them with six strains of Auxenochlorella belonging to different varieties. The integrative approach used in this study revealed that all strains showed similar morphology but differed in their SSU and ITS rDNA sequences. The Hydra endosymbiont formed a sister taxon to A. protothecoides, which included the varieties protothecoides, galactophila, and communis. The variety acidicola is not closely related to Auxenochlorella and represented its own lineage within the Trebouxiophyceae. In view of these results, we propose a new species of Auxenochlorella, A. symbiontica, for the Hydra symbiont, and a new genus Pumiliosphaera, with its type species, P. acidophila, for acidophilic strain. These results are supported by several compensatory base changes in the conserved region of ITS‐2 and ITS‐2 DNA barcodes.     

Population variation and phylogeny in the endangered <Emphasis Type="BoldItalic">Chamaesyce skottsbergii</Emphasis> (Euphorbiaceae) based on RAPD and ITS analyses

Chamaesyce skottsbergii var. skottsbergii is federally listed as an endangered taxon, and is found in small and isolated populations restricted to calcareous soils in dry shrubland habitats on the Hawaiian islands of Oahu and Molokai. Concern over the genetic relationship among these disjunct populations arose as a result of threats to the habitat of the Oahu population. The populations were examined using random amplified polymorphic DNA (RAPD) markers and sequence analysis of the internal transcribed spacer (ITS) region of the rDNA cistron. Chamaesyce skottsbergii var. vaccinioides, a closely related variety found in several small populations on Molokai, was used for baseline comparison of the genetic divergence among populations. RAPD analysis demonstrated that variation within and among populations is the highest for any Hawaiian species examined. Polymorphism was greater than 95% within populations and was 99.4% at the species level. Similarly, measures of genetic similarity indicate that differentiation among these populations is higher than is known for some species. Both RAPD and ITS sequence analysis indicate that populations of C. skottsbergii var. skottsbergii on Oahu and Molokai are genetically distinct, and the extent of this genetic differentiation supports the recognition of these populations as distinct varieties. The Molokai population is in fact much more closely related to var. vaccinioides than to var. skottsbergii on Oahu, and thus should be recognized by the previously used variety name, C. skottsbergii var. audens. Further conservation measures for each of the varieties are addressed.     

Taxonomic position of a Chinese <Emphasis Type="Italic">Pleurotus</Emphasis> “Bai-Ling-Gu”: it belongs to <Emphasis Type="Italic">Pleurotus eryngii</Emphasis> (DC.: Fr.) Quél. and evolved independently in China

The Chinese Bai-Ling-Gu is a mushroom named Pleurotus eryngii var. tuoliensis C.J. Mou. This species has been identified as P. nebrodensis or P. eryngii var. nebrodensis. We examined its taxonomic position by analysis of mating, cultivation, and rDNA sequences, and concluded as follows. (1) Bai-Ling-Gu mated with P. eryngii var. eryngii, and the F₁ and F₂ formed fruit bodies. (2) Bai-Ling-Gu mated with P. eryngii var. ferulae, and the F₁ formed fruit bodies. (3) In the di-mon mating test, P. eryngii var. nebrodensis from Sicily mated with monokaryons of P. eryngii var. eryngii but mated hardly at all with those of Bai-Ling-Gu and P. eryngii var. ferulae. The di-mon mating pattern of Bai-Ling-Gu resembled those of P. eryngii var. ferulae. (4) The partial sequences of rDNA ITS1 and IGS1 from the epitype of P. nebrodensis were identical with those from P. eryngii var. nebrodensis from Sicily but differed from those from Bai-Ling-Gu. (5) The strains of P. eryngii var. eryngii and P. eryngii var. ferulae were in a group, the strains of P. eryngii var. nebrodensis from Sicily were in another group, and the strains of Bai-Ling-Gu were in the other group in both the phylogenetic trees based on the ITS1 and the IGS1 sequences. These results led to the conclusion that Bai-Ling-Gu is a variety of P. eryngii and evolved independently in China. It is satisfactory to identify Bai-Ling-Gu with P. eryngii var. tuoliensis C.J. Mou.     

Growth pattern and cellular nitrogen and phosphorus contents of the dinoflagellate Peridinium penardii (Lemm.) Lemm. causing a freshwater red tide in a reservoir

The population growth pattern and related changes in both the nitrogen and phosphorus contents in the cell of the dinoflagellate Peridinium penardii (Lemm.) Lemm., which formed a freshwater red tide in a reservoir, were studied in situ. An exponential increase with time in population density was found. A specific growth rate of 0.25 d^–1 was observed. The cellular content of phosphorus per cell decreased from 6.0 × 10^–5 µg to 9.2 × 10^–6 µg along with an increase in population density from 8.0 × 10² cells ml^–1 to 2.5 × 10⁴ cells ml^–1. A prominent change in the cellular nitrogen did not occur. Decreasing cell content and continuous uptake of phosphorus were advantageous for P. penardii to form a freshwater red tide under P-limited conditions.     

Sequence analyses of the ITS regions and the matK gene for determining phylogenetic relationships of Diospyros kaki (persimmon) with other wild Diospyros (Ebenaceae) species

To elucidate the relationships among Diospyros kaki and species closely related in previous studies, the nuclear ribosomal internal transcribed spacer (ITS) DNA sequence and the chloroplast matK gene were sequenced and compared with those of nine Diospyros species from Thailand, four species from temperate regions, and one species of southern Africa, D. lycioides. Maximum parsimony, maximum likelihood, and neighbor joining analyses of the matK and ITS data sets revealed that D. kaki is closely related to two diploid species, D. oleifera and D. glandulosa. D. kaki, D. glandulosa, and D. oleifera were placed differently in the trees obtained from ITS and matK data sets, suggesting that hybridization and/or introgression may have occurred during the development of these species. D. kaki was not found to be closely related to D. ehretioides, a diploid species from Thailand. These results differed from a prior analysis of this genus performed with chloroplast DNA (cpDNA) restriction site mutations in 3.2- and 2.1-kb amplified sequences. The results supported Ng’s hypothesis that D. glandulosa and D. kaki may share a common ancestor. D. oleifera was also closely associated with D. kaki.     

Organelle loss in the endosymbiont ofGymnodinium acidotum (Dinophyceae)

Summary The freshwater dinoflagellateGymnodinium acidotum is known to harbor a cryptomonad endosymbiont whose chloroplasts give the organism its blue-green coloration. Every cell examined from a wild population possessed chloroplasts, mitochondria, and other organelles which are of endosymbiotic origin. Transmission electron microscopy and fluorescence microscopy revealed that only 33% of these cells possessed the nucleus of the endosymbiont. The lack of a cryptomonad nucleus in some cells did not appear to affect the cells' ability to photosynthesize or move in response to varying levels of illumination. This represents the first report of a host/endosymbiont relationship in which a significant number of individuals from a given population lack a major endosymbiont organelle.     

Molecular diversity of oligotrophic and neurotropic members of the black yeast genus Exophiala,with accent on E. dermatitidis

Analysis of ITS rDNA of the black yeast Exophiala dermatitidis revealed a close phylogenetic relationship to the meristematic fungus Sarcinomyces phaeomuriformis. As most strains ofS. phaeomuriformis have a yeast-like phenotype corresponding to the anamorph genus Exophiala, a new combination in Exophiala is proposed. On the basis of ITS sequence, M-13 fingerprint and SSU intron data, two main entities could be distinguished within E. dermatitidis. One of these (B) contained prevalently strains from environmental sources, while the other (A) mainly comprised strains from clinical sources. This may be due to a difference in virulence. All strains from severe brain and disseminated infections in East Asia clustered in group A. However, strains of group A caused a relatively mild fungemia in patients outside East Asia. This revised version was published online in June 2006 with corrections to the Cover Date.     

Cultivation of the sapropelic ciliate Plagiopyla nasuta Stein and isolation of the endosymbiont Methanobacterium formicicum

The sapropelic ciliate Plagiopyla nasuta was isolated and cultured in monoculture. Optimal conditions for growth were: 15–20°C, pH about 7, and about 2% of oxygen in the headspace. Cultures of P. nasuta produced methane. Epifluorescence microscopy revealed the presence of methanogenic bacteria as endosymbionts. An endosymbiont of the ciliate was isolated and identified as Methanobacterium formicicum. In the ciliate cell these methanogens were found to be closely associated with microbody-like organelles. No mitochondria could be detected.     

Amoebae of the genera Vannella Bovee, 1965 and Platyamoeba isolated from fish and their phylogeny inferred from SSU rRNA gene and ITS sequences

Twenty strains of flattened amoebae including 17 isolated from fish were characterised morphologically both at light microscopical and ultrastructural levels and assigned to either the genus Vannella Bovee, 1965 or the genus Platyamoeba Page, 1969. Sequence-based phylogenetic analyses of SSU rRNA genes from a data set representing a total of 29 strains of flattened amoebae strongly indicated that morphological features discriminating between these genera do not reflect phylogenetic relationships of representative strains. Contrary to a previous study, strains of this expanded assemblage formed clusters that did not reflect their environmental origin. Monophyletic groups were of mixed origins and contained freshwater as well as marine strains of both genera isolated in geographically distant localities of various continents. These findings were supported by results of phylogenetic analyses of selected strains based on ITS sequences. However, topologies of acquired ITS trees were not congruent with results inferred from SSU rRNA analyses.     

基于宏基因组测序的扶桑绵粉蚧内共生菌多样性研究

[目的] 扶桑绵粉蚧是一种重要的入侵害虫,也被列为我国的检疫性有害生物。内共生菌的群落结构及与扶桑绵粉蚧关系的研究,将为研究扶桑绵粉蚧的入侵生物学和防治提供新的思路和参考。[方法] 利用宏基因组测序技术对室内饲养扶桑绵粉蚧内共生菌的物种组成、丰度和多样性进行了研究。[结果] 共鉴定获得29个门,47个纲,105个目,178个科,245个属,299个种。变形菌门、γ-变形菌纲、肠杆菌目、肠杆菌科、伯克氏菌属、Candidatus分别为各分类阶元的优势种群。共预测了36505个基因,其中77.27%的基因分布在200~499 nt的范围内。基因集KEGG注释表明,扶桑绵粉蚧内共生菌代谢相关基因最多,达384个,占49.48%;环境信息处理基因最少,仅8个,占1.03%;代谢相关基因最多的为氨基酸代谢基因,为64个,占28.19%。基因集COG注释表明,翻译、核糖体结构和生物发生的基因最多,其次为氨基酸转运和代谢基因,分别占27.64%和12.31%。[结论] 初步确定了扶桑绵粉蚧内共生菌菌群的群落结构和优势菌群,并初步分析了这些内共生菌的功能基因情况,为进一步研究内共生菌与扶桑绵粉蚧之间的关系提供了基础,以期从内共生菌的角度为该虫的防控提供新的策略。     

Three new species of anamorphic yeasts phenotypically and phylogenetically related toCandida mesenterica: The description ofCandida fungicola sp. nov.,Candida sagamina sp. nov., andCandida fukazawae sp. nov. isolated from fruit bodies of mushrooms

Four strains of anamorphic yeasts isolated from fruit bodies of mushrooms collected in Japan were found to represent three new species of the genusCandida. These species resembleCandida mesenterica in characteristics commonly employed in the classification of yeasts. On the basis of DNA-DNA reassociation, however, they were clearly distinguished fromC. mesenterica and from one another. Three new species,Candida fungicola, Candida sagamina, andCandida fukazawae, are proposed for these yeasts. The analysis of SSU rDNA sequences suggested that these three species were closely related to each other and toC. mesenterica andC. suecica.     

秦巴山区三种人参属药用植物叶绿体基因组特征分析

为明确珠子参、羽叶三七和秀丽假人参3种药用植物叶绿体基因组特征与系统发育关系,该文以秦巴山区3种人参属药用植物为研究对象,运用生物信息学技术,分析其叶绿体基因组特征及密码子使用偏好性,并探讨三者之间的亲缘关系。结果表明:(1)3种人参属药用植物的叶绿体基因组为典型的四分体结构,序列全长为 156 071～156 104 bp,总 GC 含量为 38.10%,基因组大小相似度较高。(2)均注释到 133 个基因,包括 88 个蛋白编码基因、37 个tRNA基因和 8 个 rRNA 基因。(3)3种人参属药用植物叶绿体密码子使用偏好性相似,密码子第 3 位碱基以 A/U 结尾为主,密码子使用模式在受到突变影响的同时,主要受到自然选择的影响。(4)系统发育结果显示,3种人参属药用植物的亲缘关系较近,并且秀丽假人参同羽叶三七亲缘关系更近。综上认为,秀丽假人参与珠子参基源植物之间存在近缘关系,这项发现对于珠子参中药材的资源开发利用和分子鉴定,以及进一步研究人参属物种的分类、系统发育和进化机制提供了重要依据。     

Molecular variation of Spiranthes sinensis (Orchidaceae) in Japan, with special reference to systematic treatment of seasonally differentiated groups and a dwarf form, f. gracilis, from Yakushima Island

Molecular variations of Spiranthes sinensis Ames var. australis (R.Br.) H. Hara et Kitam. ex Kitam. in Japan were examined to evaluate the validity of the seasonally differentiated groups and a dwarf form of the species, which is endemic to Yakushima Island, Japan. Sequence differences in the plastid trnL-F locus clearly distinguished Japanese S. sinensis var. australis from S. sinensis var. sinensis collected from Ryukyu. In contrast, the trnL-F sequence of S. sinensis var. australis from Sabah, Malaysia, clearly differed from that of Japanese S. sinensis var. australis, suggesting genetic heterogeneity of Spiranthes sinensis var. australis in Asia. Moreover, a molecular analysis based on the sequences of nuclear ITS1 regions indicated that there are two major groups of S. sinensis var. australis in Japan, with a geographic distribution boundary on Kyushu Island. However, the trnL-F and ITS1 sequences did not support the genetic differentiation of the seasonally differentiated groups or the dwarf form from the other Japanese individuals. Based on these molecular data, the systematic treatment of physiological and morphological variations in the Japanese population of S. sinensis. var. australis is discussed.