Redescription of Balantidium polyvacuolum Li 1963 (Class: Litostomatea) inhabiting the intestines of Xenocyprinae fishes in Hubei, China

Redescription of Balantidium polyvacuolum Li 1963, collected from the hindgut of Xenocypris davidi and Xenocypris argentea, from Niushan Lake Fishery (30A degrees 19' N, 114A degrees 31' E) in Wuhan City, Hubei Province, China in April and June 2007 is presented in this paper to complete Li's description at both light and scanning electronic microscopic levels. The unique body shape of B. polyvacuolum-highly arched dorsal side and flattened ventral surface-as well as its remarkable concave platelet present in the centroventral were well described and compared with other close Balantidium species. Besides, two types of vestibulum shape are observed in our present work, which may suggest the existence of two subspecies or genotype species of these balantidia.

Hydrolysis and coagulation behavior of polyferric sulfate and ferric sulfate

The hydrolysis behaviors of polyferric sulfate (PFS) and ferric sulfate (FS) under conditions similar to raw wastewater were investigated and the coagulation of biologically pretreated molasses wastewater using PFS and FS was evaluated by studying coagulation efficiency, zeta potential and microscopic surface morphology of flocs. Experimental results show that the hydrolysis behavior of PFS is different from that of FS on the basis of ferron assay. In the case of FS, fast-reacting Fe(III) polymers were the dominant polynuclear species while large fraction of slow-reacting iron polymers is present in PFS. Despite slightly fewer dosages of PFS required as compared to FS, there is no marked difference in the coagulation of molasses effluent between PFS and FS, especially at the optimum dosages. Both coagulants destabilize organic compounds predominantly through charge neutralization-precipitation mechanism. Hydrolysis rate of PFS in synthetic solution is appreciably different from that in raw wastewater. This may due to the effect of sulfate anion introduced as counter-ion as well as depolymerization of larger polymeric Fe(III) species by the organic ligands present in molasses effluent.

The role of cell-surface-bound phosphatases in species competition within natural phytoplankton assemblage: an in situ experiment

Despite it is widely acknowledged that the ability to hydrolyze dissolved organic matter using extracellular phosphatases is diverse in fresh water phytoplankton, the competition within single species related to presence and quantity of cell-surface-bound phosphatases has not been examined in natural conditions yet. Here, we studied phytoplankton species competition in a freshwater reservoir during an in situ experiment. A natural plankton community, with the exclusion of large zooplankton, was enclosed in permeable dialysis bags inside two large containers of different bioavailable phosphate concentrations. Phytoplankton species biomass and the abundance of bacteria were determined in purpose to compare the development of enclosed microbial communities. Total and cell-surface-bound phosphatase activities in the phytoplankton were investigated using the Fluorescently Labelled Enzyme Activity (FLEA) technique that allows for direct microscopic detection of phosphatase-positive cells and, with image cytometry, enables quantification of phosphatase hydrolytic capacity. Production of extracellular phosphatases was not completely inhibited or stopped in the phosphate-enriched environment, phytoplankton cells only showed the activity less often. Under the phosphate-nonenriched conditions, the production of phosphatases was enhanced, but active species did not proliferate amongst phytoplankton assemblage. Further, specific growth rates of the phosphatase-positive species in the non-enriched environment were lower than the same phosphatase-positive species in phosphate-enriched environment. Interestingly, the phosphatase-positive cells of Ankyra ancora increased their size in both treatments equally, although the population in phosphate-enriched environment grew much faster and the cell-specific phosphatase activity was lower. We hypothesize that brand new daughter cells had sufficient phosphorus reserves and therefore did not employ extracellular phosphatases until they matured and needed extra bioavailable phosphorus to support their metabolism before cell division. Based on presented in situ experiment, we propose that the ability to hydrolyze organic polymers and particles with cell-surface-hound phosphatases is advantageous for longer persistence of given population in a phosphate-scarce environment; although phosphatase-positive species cannot dominate the reservoir phytoplankton solely because of specific phosphorus-scavenging strategy.

Constitutive expression of thymidylate synthase from LCDV-C induces a transformed phenotype in fish cells

Thymidylate synthase (TS), an essential enzyme in DNA synthesis and repair, plays a key role in the events of cell cycle regulation and tumor formation. Here, an investigation was presented about subcellular location and biological function of viral TS from lymphocystis disease virus from China (LCDV-C) in fish cells. Fluorescence microscopy revealed that LCDV-C TS was predominantly localized in the cytoplasm in fish cells. Cell cycle analysis demonstrated that LCDV-C TS promoted cell cycle progression into S and G2/M phase in the constitutive expressed cells. As a result, the cells have a faster growth rate compared with the control cells as revealed by cell growth curves. For foci assay, the TS-expressed cells gave rise to foci 4-5 weeks after incubation. Microscopic examination of the TS-induced foci revealed multilayered growth and crisscross morphology characteristic of transformed cells. Moreover, LCDV-C TS predisposed the transfected cells to acquire an anchorage-independent phenotype and could grow in 0.3% soft agar. So the data reveal LCDV-C TS is sufficient to induce a transformed phenotype in fish cells in vitro and exhibits its potential ability in cell transformation. To our knowledge, it is the first report on viral TS sequences associated with transforming activity. (C) 2007 Elsevier Inc. All rights reserved.

High-frequency programmable acoustic wave device realized through ferroelectric domain engineering

Surface acoustic wave devices are extensively used in contemporary wireless communication devices. We used atomic force microscopy to form periodic macroscopic ferroelectric domains in sol-gel deposited lead zirconate titanate, where each ferroelectric domain is composed of many crystallites, each of which contains many microscopic ferroelastic domains. We examined the electro-acoustic characteristics of the apparatus and found a resonator behavior similar to that of an equivalent surface or bulk acoustic wave device. We show that the operational frequency of the device can be tailored by altering the periodicity of the engineered domains and demonstrate high-frequency filter behavior (>8GHz), allowing low-cost programmable high-frequency resonators. © 2014 AIP Publishing LLC.

Microcalorimetric investigation of the toxic action of Cr(VI) on the metabolism of Tetrahymena thermophila BF5 during growth

Tetrahymena thermophila BF5 produce heat by metabolism and movement. Using a TAM air isothermal microcalorimeter, the power-time curves of the metabolism of T thermophila BF5 during growth were obtained and the action on them by the addition of Cr(VI) were studied. The morphological change with Cr(VI) coexisted and biomass change during the process of T thermophila BF5 growth were studied by light microscope. Chromium has been regarded as an essential trace element for life. However, hexavalent chromium is a known carcinogen, mutagen, cytotoxicant and strong oxidizing agent. Cr(VI) of different concentration have different effects on T thermophila BF5 growth with the phenomenon of low dose stimulation (0-3 x 10(-5) mol L-1) and high dose inhibition (3 x 10(-5) to 2.4 x 10(-4) mol L-1). The relationship between the growth rate constant (k) and c is a typical U-shaped curve, which is a characteristic of hormesis. T thermophila BF5 cannot grow at all when the concentration of Cr(VI) is up to 2.4 x 10(-4) mol L-1. The microscopic observations agree well with the results obtained by means of microcalorimetry. And T thermophila BF5 had obviously morphological changes by the addition of Cr(VI). (c) 2006 Elsevier B.V. All rights reserved.

Functional analysis of FP25K of Helicoverpa armigera single nucleocapsid nucleopolyhedrovirus

The fp25k gene of Helicoverpa armigera single nucleocapsid nucleopolyhedrovirus (HearNPV) was studied. HearNPV fp25k gene transcription was found starting from about 18 h post-infection, and protein could be detected from the same time with antiserum against FP25K. To study the function of HearNPV fp25k, a recombinant HearNPV (HaBacWD11) with an enhanced green fluorescent protein (GFP) gene replacing the fp25k was constructed using HaBacHZ8, a bacmid of HearNPV that lacks the polyhedrin gene. Growth curve analysis showed that HaBacWD11 produced higher titres of budded viruses (BVs) than its wild-type counterpart HaBacHZ8-GFP. Electron microscopic analysis indicated that at the late stage of infection, the number of intranuclear enveloped nucleocapsids in HaBacWD11-infected cells was much less than that of HaBacHZ8-GFP. A rescue recombinant virus HaBacWD14 was constructed by reintroducing fp25k gene into HaBacWD11. The growth curve and electron microscopic analysis of the rescued recombinant confirmed that the increase of BV yield and the decrease of the virion production in infected cells were the result of fp25k deletion. The expression of membrane fusion protein (Ha133) and ODV-E66 were studied using the FP25K mutants HaBacWD11 and HaBacHZ8-GFP. Unlike FP25K mutants in Autographa californica multicapsid NPV (AcMNPV), which caused an increase in the expression of membrane fusion protein GP64 and a decrease of ODV-E66, no obvious changes at the expression level of Ha133 and ODV-E66 were observed in HearNPV FP25K mutant.

Effects of longer heavy vehicles on traffic congestion

Two-lane, "microscopic" (vehicle-by-vehicle) simulations of motorway traffic are developed using existing models and validated using measured data from the M25 motorway. An energy consumption model is also built in, which takes the logged trajectories of simulated vehicles as drive-cycles. The simulations are used to investigate the effects on motorway congestion and fuel consumption if "longer and/or heavier vehicles" (LHVs) were to be permitted in the UK. Baseline scenarios are simulated with traffic composed of cars, light goods vehicles and standard heavy goods vehicles (HGVs). A proportion of conventional articulated HGVs is then replaced by a smaller number of LHVs carrying the same total payload mass and volume. Four LHV configurations are investigated: an 18.75 m, 46 t longer semi-trailer (LST); 25.25 m, 50 t and 60 t B-doubles and a 34 m, 82 t A-double. Metrics for congestion, freight fleet energy consumption and car energy consumption are defined for comparing the scenarios. Finally, variation of take-up level and LHV engine power for the LST and A-double are investigated. It is concluded that: (a) LHVs should reduce congestion particularly in dense traffic, however, a low mean proportion of freight traffic on UK roads and low take-up levels will limit this effect to be almost negligible; (b) LHVs can significantly improve the energy efficiency of freight fleets, giving up to a 23% reduction in fleet energy consumption at high take-up levels; (c) the small reduction in congestion caused by LHVs could improve the fuel consumption of other road users by up to 3% in dense traffic, however in free-flowing traffic an opposite effect occurs due to higher vehicle speeds and aerodynamic losses; and (d) underpowered LHVs have potential to generate severe congestion, however current manufacturers' recommendations appear suitable. © 2013 IMechE.

Morphology, biometry and distribution of Difflugia biwae Kawamura, 1918 (Protozoa : Rhizopoda)

The freshwater testate amoeba Difflugia biwae Kawamura, 1918, isolated from Mulan Lake, Hubei Province, China is investigated using light microscopy and scanning electron microscopy. The morphology, biometry and distribution of this little known species are supplied. After careful comparison with three other similar species, including D. delicatula Gauthier-Livre et Thomas, D. elegans Penard and D. oblonga caudata Stepanek, we believe that the characteristics of smooth fusiform shell, conspicuous great collar flare (always larger than body-width) around the aperture, constriction behind the aperture and a somewhat curved aboral horn set D. biwae apart from all other Difflugia species. This species shows a great diversity in total length, collar height and aboral horn length which have high variability (CV between 11.76 and 24.52). However, body width, collar diameter, neck width, body length and aperture diameter are fairly constant with low variability (CV between 5.34 and 8.79) which shows a remarkable uniformity of D. biwae. Also, the size frequency distributions of both body width and body length yield bell-shaped (normally distributed) curves and indicate that D. biwae is a size-monomorphic species, characterized by a main-size class and a small size range. D. biwae is probably endemic to East Asia (China and Japan) because it has such a large size (165-306 mu m) that it would have been easily found in Europe and North America, if it were there. Consequently, D. biwae must have a restricted geographical distribution, disproving the old hypothesis that microscopic organisms are cosmopolitan.

Studies on the food web structure of Lake Donghu using stable carbon and nitrogen isotope ratios

Food web structure was studied by using carbon and nitrogen isotope ratios in a hypereutrophic subtropical Chinese lake, Lake Donghu. High external nutrient loading and the presence of abundant detritus from submersed macrophytes were responsible for the high sediment delta(15)N and delta(13)C, respectively. C-13 was significantly higher in submersed macrophytes than in other macrophytes. The similar delta(13)C values in phytoplankton, zooplankton, zoobenthos, and planktivorous fish indicate that phytoplankton was the major food source for the consumers. By using a delta(15)N mass balance model, we estimate that the contributions of zooplankton to the diet of silver carp and bighead carp were 54% and 74%, respectively, which is in agreement with previous microscopic observations on intestinal contents of these fishes.

Rickettsia-like organism infection in a freshwater cultured fish Ophiocephalus argus C. in China

From 2001 to 2002, a new and emergent infectious disease of Ophiocephalus argus occurred in a fishery in Hubei Province, China, with an incidence of 60% similar to 70% and a mortality as high as 100 %. The diseased fish showed an enlarged abdomen, the millet-like nodules in internal organs, and the swollen kidney which was composed of 5 similar to 10 sarcoma-like bodies in cream or gray-white colour or ulcerated into beandregs-like substance. Light microscopic observation revealed the basophilic or acidphilic inclusions in cytoplasm of the cells and the granulomas, a diffusive chronic inflammation in internal organs. Further analysis under an electron microscope indicated that the intracytoplasmic inclusions were rickettsia-like organisms (RLOs) that are either spherical or coccoid, with variable size, ranging from 0.5 similar to 1.5 mum in diameter, and enclosed within membrane-bound cytoplasmic vacuoles. RLO had a central nucleoid region with some fine filamentous structures and an electron-dense granule. Its cytoplasm contained abundant ribosomal bodies. Occasionally, RLO appeared to be divided by binary fission. RLOs were also observed in the homogenized tissue of infected fish. The results suggested that the death of cultured O. argus was caused by RLO infection.

Induction of apoptosis in a carp leucocyte cell line infected with turbot (Scophthalmus maximus L.) rhabdovirus

A rhabdovirus was observed from the diseased turbot (Scophthalmus maximus L.) with lethal syndrome. In this study, a carp leucocyte (CLC) cell line was used to investigate the infection process and cell death mechanism occurring during the virus infection. Strong cytopathogenic effect (CPE) and the morphological changes, such as extreme chromatin condensation, nucleus fragmentation, and apoptotic body formation, were observed under fluorescence microscopy after DAPI staining in the infected CLC cells. Transmission electron microscopy analysis showed cell shrinkage, plasma membrane blebbing, cytoplasm vacuolization, chromatin condensation, nuclear breakdown and formation of discrete apoptotic bodies. The bullet-shaped nucleocapsids were measured and ranged in size from 110 to 150 nm in length and 40 to 60 nm in diameter. And therefore the virus is called Scophthalmus maximus rhabdovirus (SMRV). Agarose gel electrophoresis analysis of the DNA extracted from infected cells showed typical DNA ladder in the course of SMRV infection. Flow cytometry analysis of SMRV infected CLC cells detected apoptotic peak in the virus infected CLC cells. Virus titre analysis and electron microscopic observation revealed that the virus replication fastigium was earlier than that of the apoptosis occurrence. No apoptosis was observed in the CLC infected with UV-inactivated SMRV. All these supported that SMRV infected CLC cells undergo apoptosis and the virus replication is necessary for apoptosis induction of CLC cells. (C) 2004 Published by Elsevier B.V.

Infection and propagation of lymphocystis virus isolated from the cultured flounder Paralichthys olivaceus in grass carp cell lines

The causative agent of lymphocystis disease that frequently occurs in cultured flounder Paralichthys olivaceus in China is lymphocystis virus (LV). In this study, 13 fish cell lines were tested for their susceptibility to LV. Of these, 2 cell lines derived from the freshwater grass carp Ctenopharyngodon idellus proved susceptible to the LV, and 1 cell line, GCO (grass carp ovary), was therefore used to replicate and propagate the virus. An obvious cytopathic effect (CPE) was first observed in cell monolayers at 1 d post-inoculation, and at 3 d this had extended to about 75% of the cell monolayer. However, no further CPE extension was observed after 4 d. Cytopathic characteristics induced by the LV were detected by Giemsa staining and fluorescence microscopic observation with Hoechst 33258 staining. The propagated virus particles were also observed by electron microscopy. Ultrastructure analysis revealed several distinct cellular changes, such as chromatin compaction and margination, vesicle formation, cell-surface convolution, nuclear fragmentation and the occurrence of characteristic 'blebs' and cell fusion. This study provides a detailed report of LV infection and propagation in a freshwater fish cell line, and presents direct electron microscopy evidence for propagation of the virus in infected cells. A possible process by which the CPEs are controlled is suggested.

New data on Myxobolus longisporus (Myxozoa : Myxobolidae), a gill infecting parasite of carp, Cyprinus carpio haematopterus, from Chinese lakes

The original description of Myxobolus longisporus Nie et Li, 1992, the species infecting gills of Cyprinus carpio haematopterus L., is supplemented with new data on the spore morphology and pathogenicity. Spores are elongate pyriform with pointed anterior end, 15.7 (15.5-16.5) mum long, 6.7 (6-8) mum wide and 5.5 mum thick. Sutural ridge is straight and narrow. Mucus envelope is lacking. Two equal-sized elongate pyriform polar capsules are 8.5 mum long and 2.5 mum wide with convergent long axes. Polar filament coiled perpendicularly to the long axis of the capsule makes 9 (8-10) turns. Posterior end of polar capsules exceeds mid-spore by 15-20%. Cyst-like plasmodia are localised in the gill secondary lamellae. The infection is described in adult big host specimens. Gross lesions manifested as dark red colouration of gill tissues were restricted to the ventral part of the first gill arches. Remarkable site specificity (apical part of secondary lamellae) was observed in the course of development of microscopic lesions. M. longisporus is characterised also on the molecular level using sequences of SSU rRNA gene. Phylogenetic analysis based on these sequences has allowed clearer phylogenetic relationships to be established with other species of the genus Myxobolus sequenced to date.

Cloning of biologically active genomes from a Helicoverpa armigera single-nucleocapsid nucleopolyhedrovirus isolate by using a bacterial artificial chromosome

Purification of genotypes from baculovirus isolates provides understanding of the diversity of baculoviruses and may lead to the development of better pesticides. Here, we report the cloning of different genotypes from an isolate of Helicoverpa armigera single-nucleocapsid nucleopolyhedrovirus (HaSNPV) by using a bacterial artificial chromosome (BAC). A transfer vector (pHZB10) was constructed which contained an Escherichia coli mini-F replicon cassette within the upstream and downstream arms of HaSNPV polyhedrin gene. Hz2e5 cells were co-transfected with wild-type HaSNPV DNA and pHZB10 to generate recombinant viruses by homologous recombination. The DNA of budded viruses (BVs) was used to transform E. coli. One of the bacmid colonies, HaBacHZ8, has restriction enzyme digestion profiles similar to an in vivo cloned strain HaSNPV-G4, the genome of which has been completely sequenced. For testing the oral infectivity, the polyhedrin gene of HaSNPV was reintroduced into HaBacHZ8 to generate the recombinant bacmid HaBacDF6. The results of one-step growth curves, electron microscopic examination, protein expression analysis and bioassays indicated that HaBacDF6 replicated as well as HaSNPV-G4 in vitro and in vivo. The biologically functional HaSNPV bacmids obtained in this research will facilitate future studies on the function genomics and genetic modification of HaSNPV. (C) 2003 Elsevier B.V. All rights reserved.