A moorable,automated plankton sampler

A moorable, automated plankton sampler is described, designed to obtain series of plankton samples over extended time periods in the absence of a surface vessel. The sampler consists of one or more net-containing boxes, a unit for generating as well as measuring water flow through the net boxes, a programmeable control unit, and a frame. Each net box contains several nets on a vinyl strip, used to move the nets from the storage chamber into the fishing position and then into the preservative chamber. Preservation of samples is in a formalin-brine mixture.Net boxes can be added and the number of nets per box changed; the prototype is described with 1 net box with 10 nets. Volume of water to be filtered, time between sample collections, number of net boxes and the number of nets in each box is programmed into the control unit before sampler deployment. Collections made with the sampler are compared with those made with a SCOR net and a pump.     

A computerized plankton counter

A portable computerized plankton counting system is presented, which has the potential to enumerate 255 different species per sample. The software is used to count phytoplankton samples and produce summary data. The hardware allows the results to be reproduced in the form of a hardcopy, stored on a cassette tape, or transferred to a larger computer for analysis.     

Phage-mediated bioluminescent detection of Bacillus anthracis

Aims:  Bacillus anthracis , the causative agent of anthrax, is a serious human pathogen. The aim of this study was to provide the proof of principle results for the development of a 'bioluminescent' reporter bacteriophage that was capable of specifically detecting B. anthracis .
Methods and Results:  The reporter phage was engineered by integrating the bacterial luxA and luxB reporter genes into a nonessential region of the lysogenic Wβ phage genome. This resulted in a phage that was capable of specifically infecting and conferring a bioluminescent phenotype to B. anthracis viable cells. No processing or cell preparation was required; the phage and cells were simply mixed, and the samples were analysed for bioluminescence. A bioluminescent signal was evident after 16 min postinfection of vegetative cells. The strength and time required to generate the signal was dependent on the number of cells present. Nevertheless, 10³ CFU ml⁻¹ was detectable within 60 min. The utility of the bioluminescent phage was analysed using spores as the starting material. The Wβ:: luxAB phage was able to transduce a bioluminescent signal to germinating spores within 60 min.
Conclusions:  This proof of principle study demonstrates that the reporter phage displays promise as a tool for the rapid detection of B. anthracis .
Significance and Impact of the Study:  The new methodology offers the potential for the detection of viable cells from either environmental or clinical samples.     

Characteristics of plankton communities in Chinese integrated fish ponds: effects of excessive grazing by planktivorous carps on plankton communities

Biomass and production of plankton communities were investigated in two Chinese integrated fish culture ponds in August, Dianshanhu Pond (with high density of planktivorous carp) and Pingwang Pond (with low density of planktivorous carp). The plankton communities were composed of rotifers, protozoans, phytoplankton (<40 µm) and bacteria. The large phytoplankton (>40 µm), cladocerans and copepods were rare because of grazing pressure by the carp. The density or biomass of bacteria (1.93 × 10⁷ and 2.20 × 10⁷ cells ml^–1 on average in Dianshanhu and Pingwang Ponds, respectively), picophytoplankton (24.6 and 18.5 mg m^–3 Chla on average) and rotifers (5372 and 20733 ind. 1^–1 on average) exceeded the maximum values reported for natural waters.The average [³H]thymidine uptake rates were 694 and 904 pmoles 1^–1 h^–1 (13.4 and 20.6 µgC 1^–1) and the bacterial production by the >2 µm fraction amounted 21–28% of total [³H] thymidine uptake rate in both ponds. The mean chlorophylla concentrations were 59.1 and 183 mg m^–3 in Dianshanhu and Pingwang Ponds, respectively. 82.4% and 65.3% of the total Chla was contributed by the <10 µm nano- and picophytoplankton in each pond, respectively. In particular, the picophytoplankton contribution amounted 41.2% of thtal Chla in Dianshanhu Pond. Primary production was 2.5 and 3.4 gC m^–2 d^–1 in each pond, respectively, and >50% of production was contributed by picophytoplankton. The mean biomasses of protozoa were 168 µg 1^–1 and 445 µg 1^–1 and those of rotifers were 763 µg 1^–1 and 1186 µg 1^–1 in Dianshanhu and Pingwang Ponds, respectively. The ecological efficiencies expressed in terms of the ratios of primary production to zooplankton production were 0.22 and 0.31, for the two ponds.     

The effect of fluidic conditions on the continuous-flow bioluminescent detection of ATP in a microfluidic device

The effect of fluidic conditions on the bioluminescent detection of ATP in a microfluidic device was surveyed using homemade detector system. The bioluminescent reaction of ATP was directly affected by the retention time and flow rates of the solutions in this diffusion-based mixing and reaction system due to the laminar flow in the microchannel. ATP and enzyme solutions were separately injected into the microfluidic device at different flow rates through 3 inlet ports. The ATP solution was injected through the middle port, while the enzyme solution was injected in the two remaining ports. When the ratio of ATP to enzyme solution was fixed, the optimum flow rates of enzyme, ATP, and enzyme solution was 3.5, 8.0, and 3.5 μL/min, respectively. The optimal total flow rate was 15 μL/min. The detection limit for the concentration of ATP at optimal conditions was less than 10⁻⁷ M.     

Microtiter plate tests for segregation of bioluminescent bacteria

It has been recently shown that bioluminescence imaging can be usefully applied to provide new insights into bacterial self‐organization. In this work we employ bioluminescence imaging to record images of nutrient rich liquid cultures of the lux‐gene reporter Escherichia coli in microtiter plate wells. The images show that patterns of inhomogenous bioluminescence form along the three‐phase contact lines. The paper analyzes the dependencies of the average number of luminous aggregates (clouds) on various environmental factors. In particular, our results show that optimal (neutral) pH and high aeration rates determine the highest mean number of clouds, and that spatiotemporal patterns do not form in the pH buffered suspensions. In addition, a sigmoidal (switch‐like) dependence of the number of aggregates on the rate of aeration was observed. The obtained bioluminescence imaging data was interpreted by employing the Keller–Segel–Fisher (KSF) model of chemotaxis and logistic growth, adapted to systems of metabolically flexible (two‐state) bacteria. The modified KSF model successfully simulated the observed switch‐like responses. The results of the microtiter plate tests and their simulations indicate that the segregation of bacteria with different activities proceeds in the three‐phase contact line region. Copyright © 2015 John Wiley & Sons, Ltd.     

密云水库的浮游生物群落

2002年4～10月对密云水库浮游生物群落的调查结果显示：浮游植物有6门,58属,122种,细胞密度为565．30～10^4cells／L,4月份硅藻(Bacillariophyta)占优势,6～10月蓝藻(Cyanophyta)占绝对优势．浮游植物优势种群有水华微囊藻(Microcystis flas-aquae)、颗粒直链藻(Melosira granulata)、梅尼小环藻(Cyclotella meneghiniana)、角甲藻(Ceratium hirundinella)等。浮游动物有36种,密度为4761ind/L;浮游动物具有原生动物在数量上占绝对优势．轮虫次之,枝角类与桡足类数量较少的特点。浮游动物的优势种群有弹跳虫(Halteria grandinella)、急游虫(Stromlridium uiHde)、针簇多肢轮虫(Polyarthra trigla)、螺形龟甲轮虫(Keratella cochlearis)、广布中剑水蚤(Mesocyclops leuckarti)、长额象鼻蚤(Bosmlna longirostris)等．与1980年监测结果相比,密云水库浮游生物总密度上升较快,群落结构和优势种群也发生了明显变化．浮游生物优势种群的指示作用显示,在植物生长季节,库区水体已进入富营养状态。     

Protein structure and bioluminescent spectra for firefly bioluminescence

Modern theory on general and specific effects of microenvironment on emission spectra was used for explanation of spectral differences for both natural and mutant forms of beetle luciferases, as well as for bioluminescence emitter oxyluciferin in model systems. For the analysis, both authors' and other published data were used. It was shown that active site mutations that resulted in spectral shifts of bioluminescence as a rule caused substantial decrease in the catalytic activity of the enzyme. At the same time, mutations in the conservative regions of the protein amino acid sequence that were in the periphery of the protein globe resulted in red shift of the bioluminescence spectra without affecting catalytic activity. Correlation was observed between the value of spectral shift and polarizability of the introduced amino acid residue: the higher the polarizability, the larger was the red shift of bioluminescence. Copyright © 2002 John Wiley & Sons, Ltd.     

Comparison of the spectral emission of lux recombinant and bioluminescent marine bacteria

The purpose of the present paper was to study the influence of bacteria harbouring the luciferase‐encoding Vibrio harveyi luxAB genes upon the spectral emission during growth in batch‐culture conditions. In vivo bioluminescence spectra were compared from several bioluminescent strains, either naturally luminescent (Vibrio fischeri and Vibrio harveyi) or in recombinant strains (two Gram‐negative Escherichia coli::luxAB strains and a Gram‐positive Bacillus subtilis::luxAB strain). Spectral emission was recorded from 400 nm to 750 nm using a highly sensitive spectrometer initially devoted to Raman scattering. Two peaks were clearly identified, one at 491–500 nm (± 5 nm) and a second peak at 585–595 (± 5 nm) with the Raman CCD. The former peak was the only one detected with traditional spectrometers with a photomultiplier detector commonly used for spectral emission measurement, due to their lack of sensitivity and low resolution in the 550–650 nm window. When spectra were compared between all the studied bacteria, no difference was observed between natural or recombinant cells, between Gram‐positive and Gram‐negative strains, and growth conditions and growth medium were not found to modify the spectrum of light emission. Copyright © 2003 John Wiley & Sons, Ltd.     

A review of bioluminescent ATP techniques in rapid microbiology

Use of firefly luciferase to assay adenosine triphosphate (ATP) extracted from microorganisms provides an easy means to enumerate microbes within minutes. The small amount of light produced is proportional to ATP and thus microbial number. The average bacterium contains around 10^?15 g ATP per cell. Present reagents permit detection of 10³ cells per tube. Luminometers currently on the market detect about 10^?12 g ATP. Proper extraction of ATP from the microbes is an essential part of any protocol, as is the removal of non-microbial ATP from, for example, somatic cells also present in samples. The technique may be applied to a wide range of samples, for example food and beverages and clinical samples such as urine. The ATP assay gives a global measure of microbial numbers, i.e. it is not species specific unless a species separation step is included in the protocol.     

A moorable,automated plankton sampler: comments on Lewis & Heckl (1991)

Comments on a recently described moorable, automated plankton sampler are given, mainly because it was designed to capture large zooplankton. However, the need for automatic devices for sampling phyto- and zooplankton is stressed. A new design for such a device is presented. A preliminary test was made using standard continuous-flow (auto-analyser) equipment, a cultured flagellate and formalin as a fixative.     

Attributes of the plankton flora at Bushehr,Iran

The plankton flora on the northeastern coast of the Gulf of Persia consists of many diatom species, the coccolithophores Gephyrocapsa oceanica and Coccolithus huxleyi, and the blue-green alga, Trichodesmium thiebautii. These are prevalent throughout the year and always at low concentrations, with an average maximum in January of 14463 cells/liter and minimum in June of 802/liter. Such comparative constancy suggests that the flora has the attribute of stability. The individual species fluctuate in a patternless, uncorrelated manner, so that the flora is characterized by the attribute of unpredictability. The turbidity of the shallow water reduces the light so that light is usually neither limiting nor inhibitory. There is a small amount of nitrate always available and ample phosphate and silicate. Pure culture studies of several species show growth from about 12° to 34°. The water was 34° in August of 1977. The flora's responsiveness to these light, nutrient, and temperature quantities makes possible its recovery to normal after advective disturbance in June 1977.Contribution number 4574 from the Woods Hole Oceanographic Institution.Contribution number 4574 from the Woods Hole Oceanographic Institution.     

The use of dinoflagellate bioluminescence to characterize cell stimulation in bioreactors

Bioluminescent dinoflagellates are flow-sensitive marine organisms that produce light emission almost instantaneously upon stimulation by fluid shear in a shear stress dose-dependent manner. In the present study we tested the hypothesis that monitoring bioluminescence by suspended dinoflagellates can be used as a tool to characterize cellular response to hydrodynamic forces in agitated bioreactors. Specific studies were performed to determine: (1) impeller configurations with minimum cell activation, (2) correlations of cellular response and an integrated shear factor, and (3) the effect of rapid acceleration in agitation. Results indicated that (1) at a volumetric mass transfer coefficient of 3 x 10(-4) s(-1), marine impeller configurations were less stimulatory than Rushton configurations, (2) bioluminescence response and a modified volumetric integrated shear factor had an excellent correlation, and (3) rapid acceleration in agitation was highly stimulatory, suggesting a profound effect of temporal gradients in shear in increasing cell stimulation. By using bioluminescence stimulation as an indicator of agitation-induced cell stimulation and/or damage in microcarrier cultures, the present study allows for the verification of hypotheses and development of novel mechanisms of cell damage in bioreactors.     

混合营养型浮游生物生态学研究进展

目前已知的混合营养型浮游生物集中在甲藻门、黄藻门、裸藻门、隐藻门、金藻门、绿藻门和原生动物界的纤毛虫门,根据营养特性可分为3个生理类群。混合营养型浮游生物分布广泛,从淡水湖泊到开阔海洋,从赤道到两极,无论是寡营养还是富营养环境都发现有它们的存在。混合营养型浮游生物可进行光合营养和吞噬营养,营养策略的采用受控于环境因子。在低光照、寡营养等特殊环境中,混养生物具有重要生态地位。它可以成为主要摄食细菌者,占总生物量和生产力的大部分,比光合营养的浮游植物或吞噬营养的浮游动物更具有竞争优势。20世纪90年代以来对有害赤潮的研究,加深了对混合营养型浮游生物的认识。相信通过实验和模型的结合以及新技术的应用,将进一步推动混合营养型浮游生物生态学研究的深入。     

Heavy metal uptake by plankton and other seston particles

Abstract

The distribution of heavy metals Cd, Cu and Pb between the dissolved phase and the suspended matter has been studied in a stagnant fresh-water lake Zoommeer with the aim of finding a link between heavy metals and seston particles. Phytoplankton and Zooplankton were identified to species level and their density was determined. The average surface area and average volume, respectively, of each plankton species was calculated from the measured dimensions of 20–200 specimens of each species. Heavy metal concentrations in the dissolved phase and the particulate matter were determined by differential pulse anodic stripping voltammetry.

The seston particles were divided into 10 subdivisions and the total surface area and volume, respectively, of each subdivision was taken as an independent variable for the subsequent multiple regression analysis to find the possible correlations with the heavy metal concentrations. The obtained models can explain a very large part (up to 98% for Pb, 99% for Cd and 87% for Cu) of the variation in heavy metal concentration. An adsorption process appears to govern Cd and Pb uptake by Chlorophyceae and Dinophyceae. In addition, both Cd and Pb can penetrate into Chlorophyceae. In the case of Cu, a specific interaction with the Cyanophyceae has been found. In general, the uptake of heavy metals is highly specific for both the respective metal and the organism.     

Physical control of plankton population abundance and dynamics in intertidal rock pools

Little is known about the population structure and dynamics of plankton of intertidal rock pools. A numerical model was developed for rock pool plankton with growth limited by both tidal washout and the stress associated with adverse conditions in high-shore pools. This model predicts that a stress tolerant species will tend to have maximum population densities in high-shore pools and that populations will fluctuate in opposite phase to the spring-neap tidal cycle. Conversely, where a species is susceptible to stress in high-shore pools, the maximum population density is likely to occur lower on the shore, and numbers in upper shore pools will cycle in phase with the spring-neap cycle. These two alternative predictions were sufficient to classify the dynamics of the most abundant species in time series taken from rock pools in the Isle of Man. The dinoflagellate Oxyrrhis marina followed the predictions of the stress tolerant model. In comparison, the spatiotemporal patterns of other taxa, including a ciliate, a dinoflagellate and cryptophytes, suggested stress-susceptible life histories.     

BIOLUMBASE--the database of natural and transgenic bioluminescent organisms.

The Institute of Biophysics SB RAS hosts and maintains a specialized collection of luminous bacteria (CCIBSO 836) containing over 700 strains isolated in various regions of the world's oceans. The culture collection is a source of lux genes and biologically active substances. The wide application of bioluminescence in medicine and ecology has given importance to analysing information on the structure and functioning of bioluminescence systems in natural and transgenic microorganisms, as well as on their features that are closely interrelated with bioluminescence. The aims of our BIOLUMBASE database are: gathering information on microorganisms with lux genes, their analysis and free access, and distribution of this data throughout the global network. The database includes two sections, natural and transgenic luminous microorganisms, and is updated by our own experimental results, the published literature and internet resources. For the future, a publicly available internet site for BIOLUMBASE is planned. This will list the strains and provide comprehensive information on the properties and functions of luminous bacteria, the mechanisms of regulation of bioluminescence systems, constructs with lux genes, and applications of bioluminescence in microbiology, ecology, medicine and biotechnology. It is noteworthy that this database will also be useful for evaluation of biological hazards of transgenic strains. Users will be able to carry out bibliographic and strain searches starting from any feature of interest.     

Observations on the plankton of some Costa Rican lakes

We sampled 30 lakes in Costa Rica in the wet season (July–August) of 1991 for phytoplankton (with integrated and whole water samples), and 17 for zooplankton (with net tows). Taxa of plankton and community richness were poorly related to geography, morphology, chemistry, and other biota. Neither the zooplankton nor the phytoplankton appeared to influence the composition of the other, and neither were apparently influenced by the presence of fish.Phytoplankton richness reflected primarily sampling method, but also tended to decrease with elevation and with Secchi disk depth, and tended to increase with pH and alkalinity. Chlorophytes were the most abundant division in 14 lakes; these lakes tended to be unstratified, turbid, and located at higher elevation. Diatoms were common in 4 of the 7 lakes with elevated silica (over 30 ppm). Each lake showed at least a 3 : 1 dominance by copepods, cladocera, or insect larvae. Copepods dominated 7 of the 17 lakes, most of which were shallow, turbid, and had low alkalinity. Cladocera dominated 7 lakes that were typically deeper and located at low-to mid-elevations. Insect larvae dominated two small, turbid lakes.     

A bioluminescent signal system: detection of chemical toxicants in water.

Prototype technologies of a bioluminescent signal system (BSS) based on the luminous bacterium Photobacterium phosphoreum and three enzymatic bioluminescence systems have been proposed for detecting and signalling the presence of toxicants in water systems. A number of pesticides, mostly known as poisonous substances, similar in their structures and physicochemical properties, have been taken as model compounds of chemical agents. The effect of toxicants (organophosphates, derivatives of dithiocarbamide acid, and pyrethroid preparations) on the bioluminescence of the four systems has been analysed. EC(50) and EC(80) have been determined and compared to the maximum permissible concentration for each of the analysed substances. The triple-enzyme systems with ADH and trypsin have been shown to be more sensitive to organophosphorous compounds (0.13-11 mg/L), while the triple-enzyme system with trypsin is highly sensitive to lipotropic poison, a derivative of dithiocarbamine acid (0.03 mg/L). Sensitivities of the triple-enzyme systems to pyrethroid preparations are similar to those of luminous bacteria (0.9-5 mg/L). The results can be used to construct an alarm-test bioluminescence system for detecting chemical toxicants, based on intact bacteria or enzyme systems.     

A luxCDABE-based bioluminescent bioreporter for the detection of phenol

A bioluminescent reporter strain, Acinetobacter sp. DF4-8, was constructed for the detection of phenol by inserting a mopR-like promoter upstream of the Vibrio fischeri bioluminescent luxCDABE gene cassette in a modified mini-Tn5 construct. When introduced into the chromosome of Acinetobacter sp. DF4, the bioreporter produced a sensitive bioluminescent response to phenol at concentrations ranging from 2.5 to 100 ppm. This response was linear (R ²=0.986) in the range from 20 to 90 ppm. A significant bioluminescent response was also recorded when strain DF4-8 was incubated with slurries from aged, phenol-contaminated soil. Received 13 February 2002/ Accepted in revised form 11 July 2002