Sexual cycle properties and changes in carotenoid levels in ovary of Japanese mitten crab <Emphasis Type="Italic">Eriocheir japonica</Emphasis> from rivers of Primorye

The sexual cycle of Japanese mitten crab Eriocheir japonica (de Haan, 1835) in its northern range in water bodies of the Primorye proved to have specific properties including changed seasonal dynamics of gonad maturity stages, delayed spawning period, and reduced breeding season. The ovary color regularly changed during the gonad growth in females, which was accompanied by an increase in total carotenoid content from 1.4 to 22.2 mg/100 g tissue and by changes in the proportion of major pigments. The relative content of the carotene fraction notably decreased and that of astaxanthin slightly decreased during gonad maturation. The revealed changes in carotenoid levels in the ovary point to the activity and significance of pigment metabolism in the reproductive cycle of E. japonica.     

Gonadogenesis and color characteristics of ovaries in Japanese mitten crab Eriocheir japonicus

The gonadogenesis was studied in adult and juvenile females of Japanese mitten crab Eriocheir japonicus (Crustacea: Decapoda, Grapsida) inhabiting the rivers of the Maritime Territory. The morphometric parameters of oocytes at different stages of maturity were determined using the methods of computer morphometry and color characteristics were evaluated using the Munsell Book of Color. As a result, a color table was compiled for the ovaries from the beginning of development to gonad maturity, which included light yellow (sandy), yellow, beige, light purple, light brown, dark purple, brown (chocolate), and dark brown (brown). The regular changes in the ovary color of E. japonicus proved to closely correlate with the gonadogenesis, namely, with the composition of cells at each stage of gonad maturity.     

Characterization and expression analysis of <Emphasis Type="Italic">prohibitin</Emphasis> in the testis of Chinese mitten crab <Emphasis Type="Italic">Eriocheir sinensis</Emphasis>

Prohibitin plays a key role in maintaining mitochondrial membrane integrity and retaining its normal function. We have initially cloned and sequenced the cDNA of prohibitin from testis of the crab Eriocheir sinensis. The 1,357 bp Prohibitin cDNA comprises a 105 bp 5′ untranslated region, a 427 bp 3′ untranslated region and a 825 bp open reading frame. Protein alignment substantiates that the Prohibitin has 70.2, 69.8, 70.5, 70.9, 72.4, 70.6 and 74.9% identity with its homologues in Mus musculus, Homo sapiens, Gallus gallus, Danio rerio, Xenopus tropicalis, Drosophila mojavensis and Aedes aegypti, respectively. In situ hybridization revealed that the Prohibitin mRNA was mainly localized around the proacrosomal vesicle and nucleus membrane in early-stage spermatid. In the following middle stage, Prohibitin mRNA was situated inside the invaginated region of half-moon-like nucleus and surrounded the proacrosomal vesicle. In late-stage spermatid, the mRNA was aggregated in the acrosomal tubule, the band between the acrosome and cup-like nucleus, remanent cytoplasm as well. In the mature sperm, mRNA was only found in the acrosomal tubule and the limited space between the nucleus and acrosome. Therefore, we presume that Prohibitin may fulfill critical functions in the spermiogenesis of Eriocheir sinensis.     

Identification and characteristics of a novel gene, <Emphasis Type="Italic">EJO1</Emphasis>, in the Chinese mitten crab (<Emphasis Type="Italic">Eriocheir japonica sinensis</Emphasis>) ovary

EJO1, a novel gene presumably involved in the ovary development of the Chinese mitten crab (Eriocheir japonica sinensis), was identified and characterized by suppression subtractive hybridization and cDNA macroarray analysis. EJO1 expression was 2.6-fold higher at stage III than at stage II during the ovary development of the mitten crab. EJO1 is 876 bp in length containing a 759 bp open reading frame which encodes a 252-amino-acid protein. Homology analysis showed that no sequence significantly matching EJO1 was found in SwissProt, so it was deduced as a novel gene (GenBank accession number: AY185917). The EJO1 protein is probably a secretion protein with a signal peptide of 17 amino acids. The pI/Mw deduced from the amino acid sequence was 6.18/28.18 kDa. Expression profile showed that EJO1 mRNA is highly expressed in the heart, intestine, and ovary of the crab, while there is little or no expression in muscle and hepatopancreas. The differential expression of EJO1 at the different developmental stages of the ovary was further confirmed by Northern blot analysis. In conclusion, EJO1 is a novel gene differentially expressed in the ovary of the Chinese mitten crab, which may play an important role in the ovary development.     

Aquacultural characteristics of <Emphasis Type="Italic">Rhizoclonium</Emphasis><Emphasis Type="Italic">riparium</Emphasis> and an evaluation of its biomass growth potential

A study was made of environmental factors affecting the growth of Rhizoclonium riparium in order to evaluate its suitability for large-scale culturing. The results indicate that under the natural conditions prevailing at Taishi, Taiwan, this species can grow year-round, with a monthly biomass production (oven-dried) of 945–1540 kg ha⁻¹ pond surface (assuming a pond depth of 1 m). The specific growth rate ranged from –2.1 to 10.4% per day. Salinity and temperature, both influenced the rate significantly, with optimal values at 20% and 25 °C, respectively. Short (2-mm) lengths of filaments had a higher specific growth rate than longer (20 mm) filaments. Under rotational culturing conditions, the specific growth rate was reduced when flow was increased.     

<Emphasis Type="Italic">Ty</Emphasis>1<Emphasis Type="Italic">/copia</Emphasis>- and <Emphasis Type="Italic">Ty</Emphasis>3<Emphasis Type="Italic">/gypsy</Emphasis>-like DNA sequences in <Emphasis Type="Italic">Helianthus </Emphasis>species

Two repeated DNA sequences isolated from a partial genomic DNA library of Helianthus annuus, p HaS13 and p HaS211, were shown to represent portions of the int gene of a Ty3 /gypsy retroelement and of the RNase-Hgene of a Ty1 /copia retroelement, respectively. Southern blotting patterns obtained by hybridizing the two probes to BglII- or DraI-digested genomic DNA from different Helianthus species showed p HaS13 and p HaS211 were parts of dispersed repeats at least 8 and 7 kb in length, respectively, that were conserved in all species studied. Comparable hybridization patterns were obtained in all species with p HaS13. By contrast, the patterns obtained by hybridizing p HaS211 clearly differentiated annual species from perennials. The frequencies of p HaS13- and p HaS211-related sequences in different species were 4.3x10(4)-1.3x10(5) copies and 9.9x10(2)-8.1x10(3) copies per picogram of DNA, respectively. The frequency of p HaS13-related sequences varied widely within annual species, while no significant difference was observed among perennial species. Conversely, the frequency variation of p HaS211-related sequences was as large within annual species as within perennials. Sequences of both families were found to be dispersed along the length of all chromosomes in all species studied. However, Ty3 /gypsy-like sequences were localized preferentially at the centromeric regions, whereas Ty1/ copia-like sequences were less represented or absent around the centromeres and plentiful at the chromosome ends. These findings suggest that the two sequence families played a role in Helianthusgenome evolution and species divergence, evolved independently in the same genomic backgrounds and in annual or perennial species, and acquired different possible functions in the host genomes.     

Flower color alteration in <Emphasis Type="Italic">Lotus japonicus</Emphasis> by modification of the carotenoid biosynthetic pathway

To establish a model system for alteration of flower color by carotenoid pigments, we modified the carotenoid biosynthesis pathway of Lotus japonicus using overexpression of the crtW gene isolated from marine bacteria Agrobacterium aurantiacum and encoding β-carotene ketolase (4,4′-β-oxygenase) for the production of pink to red color ketocarotenoids. The crtW gene with the transit peptide sequence of the pea Rubisco small subunit under the regulation of the CaMV35S promoter was introduced to L. japonicus. In most of the resulting transgenic plants, the color of flower petals changed from original light yellow to deep yellow or orange while otherwise exhibiting normal phenotype. HPLC and TLC analyses revealed that leaves and flower petals of these plants accumulated novel carotenoids, believed to be ketocarotenoids consisting of including astaxanthin, adonixanthin, canthaxanthin and echinenone. Results indicated that modification of the carotenoid biosynthesis pathway is a means of altering flower color in ornamental crops.     

<Emphasis Type="Italic">Humulus japonicus</Emphasis> Accelerates the Decomposition of <Emphasis Type="Italic">Miscanthus sacchariflorus</Emphasis> and <Emphasis Type="Italic">Phragmites australis</Emphasis> in a Floodplain

Humulus japonicus in communities of Miscanthus sacchariflorus and Phragmites australis can grow large enough to overtop other species in the Amsa-dong floodplain. Because of strong winds and the weight of Humulus, plants of M. sacchariflorus and P. australis fell in mid-August and were subject to decomposition under its dense shading. To assess the effects of H. japonicus on nutrient cycling in these communities, we collected fresh samples of M. sacchariflorus and P. australis in litterbags and decomposed them under H. japonicus for 9 months, beginning in August. Biomass and organic contents from M. sacchariflorus during this incubation period were 49–51% and 44–48%, whereas those of P. australis were 49–61% and 32–52%, respectively. Their annual k values were 1.61–1.74 and 1.46–3.54, respectively. Initial N concentrations in M. sacchariflorus and P. australis were 13 and 20 mg g⁻¹, while C:N ratios were 31 and 21, respectively. These results indicate that H. japonicus is responsible for the collapse of M. sacchariflorus and P. australis in August and also accelerates their nutrient cycling through rapid decomposition, thereby increasing nutrient circulation in floodplains.     

Morphology transition genes in the dimorphic fission yeast <Emphasis Type="Italic">Schizosaccharomyces</Emphasis><Emphasis Type="Italic">japonicus</Emphasis>

The ability of the saprophytic fungus Schizosaccharomyces japonicus to alternate between unicellular yeast form and multicellular true mycelium makes this organism an attractive non-pathogenic model for the investigation of di- and polymorphism critical for pathogenicity in many pathogenic fungi. In a previous work we described three mutations that made the cells unable to form hyphae. Here we report on the isolation of additional mycelium-minus mutants and show that the mutations represent seven genes. Apart from the inhibition of the yeast-to-mycelium transition, the mutations also cause drastic changes in the yeast phase. Cells of myc3-34 and myc4-35 grow isotropically with apolar distribution of actin and randomised localisation of division planes. The mutants myc1-4, myc1-10, myc1-36, myc5-39 and myc6-43 form sep-like, branching microhyphae containing bipolarly growing cells. All mutants are defective in the polarisation of vacuole distribution, and myc3-34, myc4-35 and myc7-56 are also defective in vacuole fusion. The diversity of the mutant phenotypes indicates that several cellular processes must take place simultaneously for the transition from the yeast phase into the mycelial phase.     

<Emphasis Type="Italic">Agrobacterium</Emphasis><Emphasis Type="Italic">tumefaciens</Emphasis>-mediated transformation of callus cells of <Emphasis Type="Italic">Crataegus</Emphasis><Emphasis Type="Italic">aronia</Emphasis>

A genetic transformation system has been developed for callus cells of Crataegus aronia using Agrobacterium tumefaciens. Callus culture was established from internodal stem segments incubated on Murashige and Skoog (MS) medium supplemented with 5 mg l⁻¹ Indole-3-butyric acid (IBA) and 0.5 mg l⁻¹ 6-benzyladenine (BA). In order to optimize the callus culture system with respect to callus growth and coloration, different types and concentrations of plant growth regulators were tested. Results indicated that the best average fresh weight of red colored callus was obtained on MS medium supplemented with 2 mg l⁻¹ 2,4-dichlorophenoxyacetic acid (2,4-D) and 1.5 mg l⁻¹ kinetin (Kin) (callus maintenance medium). Callus cells were co-cultivated with Agrobacterium harboring the binary plasmid pCAMBIA1302 carrying the mgfp5 and hygromycin phosphotransferase (hptII) genes conferring green fluorescent protein (GFP) activity and hygromycin resistance, respectively. Putative transgenic calli were obtained 4 weeks after incubation of the co-cultivated explants onto maintenance medium supplemented with 50 mg l⁻¹ hygromycin. Molecular analysis confirmed the integration of the transgenes in transformed callus. To our knowledge, this is the first time to report an Agrobacterium-mediated transformation system in Crataegus aronia.     

<Emphasis Type="Italic">In vitro</Emphasis> multiplication of heavy metals hyperaccumulator <Emphasis Type="Italic">Thlaspi caerulescens</Emphasis>

A micropropagation protocol through multiple shoot formation was developed for Thlaspi caerulescens L., one of the most important heavy metals hyperaccumulator plants. In vitro seed-derived young seedlings were used for the initiation of multiple shoots on Murashige and Skoog (MS) medium with combinations of benzylaminopurine (BA; 0.5–1.0 mg dm⁻³), naphthaleneacetic acid (NAA; 0–0.2 mg dm⁻³), gibberellic acid (GA₃; 0–1.0 mg dm⁻³) and riboflavin (0–3.0 mg dm⁻³). The maximum number of shoots was developed on medium containing 1.0 mg dm⁻³ BA and 0.2 mg dm⁻³ NAA. GA₃ (0.5 mg dm⁻³) in combination with BA significantly increased shoot length. In view of shoot numbers, shoot length and further rooting rate, the best combination was 1.0 mg dm⁻³ BA + 0.5 mg dm⁻³ GA₃ + 1.0 mg dm⁻³ riboflavin. Well-developed shoots (35–50 mm) were successfully rooted at approximately 95 % on MS medium containing 20 g dm⁻³ sucrose, 8 g dm⁻³ agar and 1.0 mg dm⁻³ indolebutyric acid. Almost all in vitro plantlets survived when transferred to pots.     

Revision of the <Emphasis Type="Italic">Serrulati</Emphasis> species of <Emphasis Type="Italic">Penstemon</Emphasis> section <Emphasis Type="Italic">Saccanthera</Emphasis> subsection <Emphasis Type="Italic">Serrulati</Emphasis>

A revision of Penstemon sect. Saccanthera subsect. Serrulati includes a new species (P. salmonensis), a new variety (P. triphyllus var. infernalis), and the elevation of a subspecies to species (P. curtiflorus), bringing the total number of species to eight, which are keyed and described, complete with nomenclature and type citations.     

Comparative photosynthesis characteristics of <Emphasis Type="Italic">Calycanthus chinensis</Emphasis> and <Emphasis Type="Italic">Chimonanthus praecox</Emphasis>

Calycanthus chinensis is an endangered plant of the national second-grade protection of China restricted in a small area in Zhejiang Province. We studied parameters of photosynthesis, chlorophyll (Chl) contents, and Chl fluorescence (minimum fluorescence, F₀, maximum fluorescence, F_m, variable fluorescence, F_v, and F_v/F_m) of C. chinensis and Chimonanthus praecox. C. chinensis had lower compensation irradiance but higher saturation irradiance than C. praecox. Hence C. chinensis has more advantage in obtaining and utilizing photon energy and higher Chl content, and is more adaptive to higher temperature and propitious to thermal dissipation than C. praecox. In addition, C. chinensis produces abundant, well-preserved seed with a higher germination rate and a wider adaptability to temperature than C. praecox. Thus C. chinensis is prone to survival and viability, and gets rid of the endangered plant species of the national second-grade protection of China.     

Developmental and ultrastructual characteristics of mouse oocytes grown <Emphasis Type="Italic">in Vitro</Emphasis> from primordial germ cells

The aim of this study was to clarify the developmental and ultrastructual characteristics of oocytes grown in vitro from primordial germ cells. The female genital ridges at 12.5 days post coitus were cultured for 18 days on an insert membrane in Waymouth's MB752/1 medium, supplemented with 15% fetal bovine serum and 1 mM sodium pyruvate; subsequently, the follicles isolated from the tissue were cultured for eight days in Waymouth's medium supplemented with 5 microg/ml insulin, 5 microg/ml transferrin, 5 ng/ml selenium, 10 mIU/ml follicle stimulating hormone, and 100 ng/ml stem cell factor. The primordial germ cells developed in vitro into oocytes of more than 60 microm in diameter. The transmission electron microscopic analysis indicated that the oocytes, which developed in vitro, showed no obvious abnormality in their ultrastructure and had organelles appropriate for the oocyte size. However, a delay in the progressive changes of morphology in some of the organelles during oocyte growth was often found when comparing them to oocytes grown in vivo.     

The <Emphasis Type="Italic">Caenorhabditis</Emphasis><Emphasis Type="Italic">briggsae</Emphasis> genome contains active <Emphasis Type="Italic">CbmaT1</Emphasis> and <Emphasis Type="Italic">Tcb1</Emphasis> transposons

The maT clade of transposons is a group of transposable elements intermediate in sequence and predicted protein structure to mariner and Tc transposons, with a distribution thus far limited to a few invertebrate species. We present evidence, based on searches of publicly available databases, that the nematode Caenorhabditis briggsae has several maT-like transposons, which we have designated as CbmaT elements, dispersed throughout its genome. We also describe two additional transposon sequences that probably share their evolutionary history with the CbmaT transposons. One resembles a fold back variant of a CbmaT element, with long (380-bp) inverted terminal repeats (ITRs) that show a high degree (71%) of identity to CbmaT1. The other, which shares only the 26-bp ITR sequences with one of the CbmaT variants, is present in eight nearly identical copies, but does not have a transposase gene and may therefore be cross mobilised by a CbmaT transposase. Using PCR-based mobility assays, we show that CbmaT1 transposons are capable of excising from the C. briggsae genome. CbmaT1 excised approximately 500 times less frequently than Tcb1 in the reference strain AF16, but both CbmaT1 and Tcb1 excised at extremely high frequencies in the HK105 strain. The HK105 strain also exhibited a high frequency of spontaneous induction of unc-22 mutants, suggesting that it may be a mutator strain of C. briggsae.     

<Emphasis Type="Italic">Galleria</Emphasis><Emphasis Type="Italic">mellonella</Emphasis> are Resistant to <Emphasis Type="Italic">Pneumocystis</Emphasis><Emphasis Type="Italic">murina</Emphasis> Infection

Studying Pneumocystis has proven to be a challenge from the perspective of propagating a significant amount of the pathogen in a facile manner. The study of several fungal pathogens has been aided by the use of invertebrate model hosts. Our efforts to infect the invertebrate larvae Galleria mellonella with Pneumocystis proved futile since P. murina neither caused disease nor was able to proliferate within G. mellonella. It did, however, show that the pathogen could be rapidly cleared from the host.     

New combinations and typifications in <Emphasis Type="Italic">Bistorta</Emphasis>, <Emphasis Type="Italic">Persicaria</Emphasis>, <Emphasis Type="Italic">Polygonum,</Emphasis> and <Emphasis Type="Italic">Rumex</Emphasis> (Polygonaceae)

New combinations are proposed in anticipation of the Polygonaceae treatment in the forthcoming volume of Intermountain Flora: Polygonum kelloggii var. esotericum, P. kelloggii var. watsonii , Rumex densiflorus var. pycnanthus , R. salicifolius var. utahensis, and R. occidentalis var. tomentellus. Typifications are proposed to facilitate ongoing studies in Polygonaceae and to maintain current usage.     

Post-pollination hybridization barriers in <Emphasis Type="Italic">Nicotiana</Emphasis> section <Emphasis Type="Italic">Alatae</Emphasis>

Nicotiana section Alatae contains eight species with variable flower sizes and morphologies. Section members readily hybridize in the glasshouse, but no hybrids have been observed in natural sympatric and parapatric populations. To investigate interspecific crossing relationships with respect to mechanisms preventing hybridization, all members of section Alatae were intercrossed in a complete diallel. We found positive correlation between the pistil length of the pollen donor and interspecific seed set relative to the conspecific cross. Pollen tube growth rate and pollen donor pistil length were positively correlated as well. Furthermore, pollen from short-pistil members of section Alatae could only grow a maximum distance proportional to, but greater than, their own pistil lengths. Our results show that pollen tube growth capacity (i.e., rate and distance), provides a hybridization barrier in long-pistil species × short-pistil species crosses. We also found another hybridization barrier not specifically related to pollen tube growth capacity in short-pistil species × long-pistil species. Taken together, these barriers can generally be described by a ‘pistil-length mismatch’ rule; in section Alatae, pollen has the most success fertilizing ovules from species with pistil lengths similar to their own. This rule could contribute to hybridization barriers in Section Alatae because the species display dramatically different pistil lengths. Electronic supplementary material  The online version of this article (doi:) contains supplementary material, which is available to authorized users.