Fumie Kasai

Complete Genomic Structure of the Bloom-forming Toxic Cyanobacterium Microcystis aeruginosa NIES-843

Abstract The nucleotide sequence of the complete genome of a cyanobacterium, Microcystis aeruginosa NIES-843, was determined. The genome of M. aeruginosa is a single, circular chromosome of 5 842 795 base pairs (bp) in length, with an average GC content of 42.3%. The chromosome comprises 6312 putative protein-encoding genes, two sets of rRNA genes, 42 tRNA genes representing 41 tRNA species, and genes for tmRNA, the B subunit of RNase P, SRP RNA, and 6Sa RNA. Forty-five percent of the putative protein-encoding sequences showed sequence similarity to genes of known function, 32% were similar to hypothetical genes, and the remaining 23% had no apparent similarity to reported genes. A total of 688 kb of the genome, equivalent to 11.8% of the entire genome, were composed of both insertion sequences and miniature inverted-repeat transposable elements. This is indicative of a plasticity of the M. aeruginosa genome, through a mechanism that involves homologous recombination mediated by repetitive DNA elements. In addition to known gene clusters related to the synthesis of microcystin and cyanopeptolin, novel gene clusters that may be involved in the synthesis and modification of toxic small polypeptides were identified. Compared with other cyanobacteria, a relatively small number of genes for two component systems and a large number of genes for restriction-modification systems were notable characteristics of the M. aeruginosa genome.

Morphology, genetic diversity, temperature tolerance and toxicity of Cylindrospermopsis raciborskii (Nostocales, Cyanobacteria) strains from Thailand and Japan

Cylindrospermopsis raciborskii is a planktonic, nostocalean cyanobacterium, which produces an alkaloid heptatoxin, cylindrospermopsin. We performed morphological observations, 16S rDNA sequence analysis, PCR fingerprint analysis of short tandemly repeated repetitive (STRR) sequences, temperature tolerances and toxin analysis to characterize 24 strains of this toxic cyanobacterium isolated from Thailand and Japan. All strains shared common morphological traits characteristic of C. raciborskii and showed high 16S rDNA sequence similarity, forming a defined cluster together with the reference strains from Australia. In particular, some of the Thai strains shared 99.9% to 100% similarity with the Australian strains. Various combinations of STRR primers revealed different and unique DNA band patterns among strains of C. raciborskii. The phylogenetic tree revealed two main clusters of C. raciborskii strains, the Thai/Japan-Shinobazugaike cluster (cluster I) and the Japan-Gonoike cluster (cluster II). Cluster I was further divided into two subclusters, A (only Thai strains) and B (one Thai strain and the Japan-Shinobazugaike strains). Thus, the results from 16S rDNA and STRR analyses showed no clear geographical distinction between Japanese and Thai strains and between Thai and Australian strains. Thai strains were separated into adaptive and non-adaptive groups to low temperature (15 and 17.5 degrees C) and Japanese strains were composed of only low-temperature-adaptive ones. The toxin cylindrospermopsin was detected in some strains of cluster I-A and in one strain of cluster II. We conclude that C. raciborskii is a species that has recently begun to invade, and a species with different physiological strains or ecotypes in temperature tolerance; the toxin is synthesized without any relation to phylogenetic or genetic clusters and to geography.

Genetic combinations of symbionts in a vegetatively reproducing lichen, Parmotrema tinctorum, based on ITS rDNA sequences

Abstract The genetic combinations between mycobionts and photobionts in Parmotrema tinctorum collected from ca. 60 km2 of the Shimizu district, of Shizuoka City in Japan was investigated based on ITS rDNA sequences. This lichen produces apothecia quite rarely, and in principle propagates vegetatively by isidia. The genetic diversity of the mycobiont comprised four types, while that of the photobiont comprised 21 types. There were 28 different combinations between mycobiont and photobiont. All the photobionts were identified as Trebouxia corticola (s. lat.), based on both molecular phylogenetic results and morphological observation of culture strains obtained in this study. Therefore, P. tinctorum is considered to be highly selective toward the photobiont. The 28 combinations from the small area represent an unexpectedly high diversity, because P. tinctorum is thought to propagate vegetatively. Four possible mechanisms to account the high genetic combinations are suggested: i.e., photobiont exchange, fusion of thalli, and long-distance dispersal of isidia or ascospores. The genetic diversity of photobionts was poor in the urban area, but rich in suburbs and mountainsides. This might be caused by a bottleneck or founder effect in the population recovering from former damage by heavy air pollution, or variable selectivity of P. tinctorum depending on the environments.

Effects of simetryne on growth of various freshwater algal taxa.

The sensitivity of 56 algal strains, representing 7 taxonomic groups to the triazine herbicide, simetryne, was examined using EC50 values for growth. There was a wide range of values from 6.5 to 1500 microg litre(-1). The Volvocales (Chlorophyceae, Chlorophyta) and Cyanophyceae (Cyanophyta) as a whole were the most sensitive, whereas the Desmidiales (Charophyceae, Chlorophyta) and Bacillariophyceae (Chromophyta) were the most tolerant, although sensitivity differed among strains of a single species. Sensitive and tolerant species were both isolated from samples collected at the same site. The results suggest that changes in species composition and relative abundance will occur when herbicides are applied in natural habitats.

Herbicide susceptibility in two green algae, Chlorella vulgaris and Selenastrum capricornutum

Abstract Responses of four strains belonging to two green algae, Chlorella vulgaris and Selenastrum capricornutum to three herbicides, simetryn, pretilachlor and thiobencarb, were compared, using single species toxicity tests for population growth. There was obvious difference in the susceptibility beween the two species. The two strains of C. vulgaris were more tolerant than the two strains of S. capricornutum to all the three herbicides. The difference in the susceptibility between the two species might be explained in part by the characteristics such as a thick cell wall and higher activities of several enzymes in C. vulgaris.

NBRP databases: databases of biological resources in Japan.

The National BioResource Project (NBRP) is a Japanese project that aims to establish a system for collecting, preserving and providing bioresources for use as experimental materials for life science research. It is promoted by 27 core resource facilities, each concerned with a particular group of organisms, and by one information center. The NBRP database is a product of this project. Thirty databases and an integrated database-retrieval system (BioResource World: BRW) have been created and made available through the NBRP home page (http://www.nbrp.jp). The 30 independent databases have individual features which directly reflect the data maintained by each resource facility. The BRW is designed for users who need to search across several resources without moving from one database to another. BRW provides access to a collection of 4.5-million records on bioresources including wild species, inbred lines, mutants, genetically engineered lines, DNA clones and so on. BRW supports summary browsing, keyword searching, and searching by DNA sequences or gene ontology. The results of searches provide links to online requests for distribution of research materials. A circulation system allows users to submit details of papers published on research conducted using NBRP resources.

Assessment of herbicide effects on algal production in the Kokai River (Japan) using a model stream and Selenastrum bioassay

The effects of herbicides on algal production in the Kokai River were assessed using a Selenastrum growth inhibition test and by an investigation of the periphyton formed in a channel carrying river water. Herbicides in the water were analysed at the same intervals as the algal growth test (three times per week) from April to August 1992. Growth of Selenastrum in the river water samples was inhibited severely in mid-May, mostly by the joint effects of butachlor and pretilachlor and from early to late June mostly by simetryn alone. The influence of six other kinds of herbicide was low or not detectable. Selenastrum growth ratios (SGR) in the river water samples were correlated with fluctuations in SG-IP (Selenastrum growth inhibition potential), the sum of the growth inhibition ratios by the respective herbicides at the concentrations detected in the river. However, the expected growth inhibition (SG-IP) was usually larger than the observed value (SGR); the reasons for this are discussed.The rate of accumulation of periphyton, which formed on a substrate in the channel, decreased from early May and stayed at relatively low levels until early June. The low accumulation rate from mid-June to mid-July, when growth of Selenastrum had almost recovered, can be attributed to grazing by invertebrates. These results suggested that algal production in the Kokai River is affected by the single or joint action of herbicides, at least in the period from early May to early June.

Taxonomic revision of Chattonella antiqua, C. marina and C. ovata (Raphidophyceae) based on their morphological characteristics and genetic diversity

Demura M., Noël M.-H., Kasai F., Watanabe M.M. and Kawachi M. 2009. Taxonomic revision of Chattonella antiqua, C. marina and C. ovata (Raphidophyceae) based on their morphological characteristics and genetic diversity. Phycologia 48: 518–535. DOI: 10.2216/08-98.1. Three raphidophyte species, Chattonella antiqua, C. marina and C. ovata, are red tide–forming harmful phytoplankton species, causing mass mortality at fish farms. These species were originally distinguished from each other solely based on their morphological characters. Recent genetic diversity analyses showed that the three species were extremely similar. It was therefore necessary to re-examine these three species to determine if they were independent single species. We compared 104 strains of the three species using morphological characters, nuclear ITS rDNA regions, the chloroplast rbcL gene, the mitochondrion COI gene and selected microsatellite regions. The morphological characters for the three species formed a continuum of variation instead of clearly defined limits. Compared with other selected heterokontophytes, their genetic divergence in the analyzed three regions was found to be at the intraspecies level. Microsatellite markers distinguished each of the 104 strains, but the strains apparently formed a single geographical population. Based on the morphological characters and the phylogenetic analyses, three groups, corresponding to each of the three Chattonella species, were roughly recognized. Nevertheless, this three-group distribution was insufficient to justify their distinction at the species rank. We therefore proposed a taxonomic revision with C. antiqua and C. ovata reduced to varieties of C. marina, this latter having name priority. We proposed a new status: C. marina var. antiqua (Hada) Demura & Kawachi and C. marina var. ovata (Y. Hara & Chihara) Demura & Kawachi.

Recombination, cryptic clades and neutral molecular divergence of the microcystin synthetase (mcy) genes of toxic cyanobacterium Microcystis aeruginosa.

BackgroundThe water-bloom-forming cyanobacterium Microcystis aeruginosa is a known producer of various kinds of toxic and bioactive chemicals. Of these, hepatotoxic cyclic heptapeptides microcystins have been studied most intensively due to increasing concerns for human health risks and environmental damage. More than 70 variants of microcystins are known, and a single microcystin synthetase (mcy) gene cluster consisting of 10 genes (mcyA to mcyJ) has been identified to be responsible for the production of all known variants of microcystins. Our previous multilocus sequence typing (MLST) analysis of the seven housekeeping genes indicated that microcystin-producing strains of M. aeruginosa are classified into two phylogenetic groups.ResultsTo investigate whether the mcy genes are genetically structured similarly as in MLST analysis of the housekeeping genes and to identify the evolutionary forces responsible for the genetic divergence of these genes, we used 118 mcy-positive isolates to perform phylogenetic and population genetic analyses of mcy genes based on three mcy loci within the mcy gene cluster (mcyD, mcyG, and mcyJ), none of which is involved in the production of different microcystin variants. Both individual phylogenetic analysis and multilocus genealogical analysis of the mcy genes divided our isolates into two clades, consistent with the MLST phylogeny based on seven housekeeping loci. No shared characteristics within each clade are known, and microcystin analyses did not identify any compositional trend specific to each clade. Statistical analyses for recombination indicated that recombination among the mcy genes is much more frequent within clades than between, suggesting that recombination has been an important force maintaining the cryptic divergence of mcy genes. On the other hand, a series of statistical tests provided no strong evidence for selection to explain the deep divergence of the mcy genes. Furthermore, analysis of molecular variance (AMOVA) indicated a low level of geographic structuring in the genetic diversity of mcy.ConclusionOur phylogenetic analyses suggest that the mcy genes of M. aeruginosa are subdivided into two cryptic clades, consistent with the phylogeny determined by MLST. Population genetic analyses suggest that these two clades have primarily been maintained as a result of homology-dependent recombination and neutral genetic drift.

Effects of the triazine herbicide, simetryn, on freshwater plankton communities in experimental ponds.

The effects of a triazine herbicide, simetryn, on freshwater phyto- and zooplankton communities and water chemistry were investigated using outdoor experimental ponds. Simetryn was applied at a concentration of 0.1 or 1.0 mg l(-1) 18 days after the start of the experiment. Simetryn treatments reduced pH, dissolved oxygen concentration, chlorophyll a concentration and photosynthetic rate in the pond water, whereas they increased the dissolved nutrient concentration. In the phytoplankton community Volvocales and Cryptophyceae were clearly reduced by the herbicide application. In the 0.1 mg l(-1) simetryn-treated ponds, photosynthetic rate of the phytoplankton community recovered slightly about two weeks after the treatment, which coincided with recovery of centric diatoms and Cryptophyceae, whereas it was completely suppressed until the end of the experiment in the 1.0 mg l(-1) simetryn-treated ponds. The decrease in zooplankton density in the treated ponds was not due to the direct toxic effects of simetryn but to indirect effects resulting from reduced algal productivity which in turn reduced the food supply.