Effect of dichromate on population and growth of various protozoa isolated from industrial effluents

Three protozoa belonging to genera Euglena, Vorticella and Stylonychia collected from industrial wastes were cultured in a medium containing inorganic salts, basically meant for the growth of algae. Protozoa showed rapid growth in the medium. Hexavalent chromium (K2Cr2O7) at a concentration of 5 micrograms/L in the medium adversely affected the growth of protozoa. At the end of eight days of Cr administration, the population of Euglena, Vorticella and Stylonychia increased 8-, 4.5- and 10-fold, respectively, as against 30-, 6.75- and 50-fold increase in the control cultures. No apparent death phase and no change in activity or morphology of protozoa was observed at this Cr concentration. The protozoa were also exposed to different metal ions, viz. Pb (2.42 mmol/L), Cr (0.48 mmol/L), Cd (0.36 mmol/L), administered in the culture medium for a period of 2 years. The metal tolerance for S. mytilus and V. microstoma was Pb > Cr > Cd. E. proxima could not tolerate any of the long-term metal treatments. Because of the ability of these protozoa to tolerate high concentrations of heavy metals, their potential role in remediation of heavy metals from industrial wastewater is considered.     

Cloning and characterization of endosymbiotic algae isolated fromParamecium bursaria

Summary The green parameciumParamecium bursaria has many endosymbiotic algae in its cytoplasm. Here, we cloned and characterized endosymbiotic algae fromP. bursaria and examined in detail the interaction between the cloned algae and algae-free paramecia. Homogenates ofP. bursaria were cultured on agar plates containing various kinds of media to establish clones of the endosymbiotic algae. Many algal colonies were obtained from poorly nutritious medium (CA medium) after one month in culture. Algae were picked up from these colonies and inoculations were repeated 9 times on agar plates containing CA medium. On enriched media including bacto-peptone, glucose, proteose-peptone and/or yeast extract, however, bacteria and mold grew rapidly and no algal colonies were formed. When the cloned algae were cultured in liquid CA medium, they grew faster than on agar plates and the numbers stayed constant at 1 × 10⁷ algae/ml after 7 days in culture. They revealed high infectivity to algae-free paramecia, and an incubation period of 24 h and at least 1 × 10³ algae/paramecium were required to achieve successful infection (80–90%). The growth and infection rate did not change through 74 repeated inoculations of algae in liquid CA medium. Optical microscopic observations revealed marked morphological similarity between endosymbiotic algae and free-livingChlorella, but the latter showed no infectivity to algae-free paramecia. The cloned endosymbiotic algae presented here will provide an excellent opportunity to examine the mechanism of symbiont-host interaction.     

Antibiotic treatment enhances C2 toxin production by Alexandrium tamarense in batch cultures

The effects of a mixture of penicillin G and streptomycin on the growth and C2 toxin production of a marine dinoflagellate, Alexandrium tamarense CI01, were investigated to determine if antibiotic treatment would increase the toxin yield of the cultured algae in batch cultures. Algal growth and toxin production were both enhanced markedly when the culture was supplemented with the antibiotics, each at an initial concentration of 100 unit ml⁻¹ in medium,² but were severely inhibited when the concentration was 500 unit ml⁻¹ or higher. Short-term pretreatment of algal inocula with the antibiotics at 100, 500, and 1000 unit ml⁻¹ all produced the enhancing effects on the algal cultures in an autoclaved medium. A prolonged antibiotic pretreatment of the algal culture followed by repeated sterile cultivation resulted in an algal culture free of cultivable bacteria. This “drug-treated” culture became more resistant to the toxicity and more responsive to the enhancing effects of the antibiotics. Our results indicated that the antibiotics can enhance growth and C2 toxin productivity not only through their inhibition of the growth of bacteria that compete for nutrients with the coexisting algae, but also through their direct effects on the physiology of the algae. Supplementation of the two antibiotics therefore is an efficient way to increase the yield of C2 toxin in the production cultures of A. tamarense CI01.     

Barley straw as an inhibitor of algal growth II: laboratory studies

The presence of rotting barley straw in water inhibited the growth of several planktonic and filamentous algae in laboratory culture. The inhibitory effect was produced progressively during decomposition of the straw at 20 °C and reached a maximum after six months. When the straw was autoclaved, all inhibitory activity was lost. Algae recovered and continued to grow normally when transferred from cultures containing rotting straw to sterile culture medium. Addition of liquor from rotting straw also inhibited algal growth. The capacity to inhibit growth remained in the liquor after passage through a 0.2 ,m filter but was removed by activated carbon. The inhibitory effect of straw shows promise as a practical means of limiting the growth of a range of algae which can cause problems in aquatic systems.author for correspondence     

The 96-well twin-layer system: a novel approach in the cultivation of microalgae

A novel system for the growth and maintenance of microalgae has been developed that allows the cultivation of a large number of strains with little manual effort. The system is based on a 96-well microtiter plate in which a membrane filter constitutes the bottom of each well. Algal strains are immobilised on the membranes and provided with culture medium through contact with layers of glass fibre located beneath the membranes in a special cultivation chamber. The configuration effectively separates culture medium from algal cells which allows the simultaneous exchange of the culture medium for 96 strains within a few minutes without the need to transfer the algae. If necessary, algal strains can be transferred using multi-channel pipettes. We demonstrate that a large variety of microalgal strains including delicate flagellates can be reliably grown in the system under axenic conditions and without cross-contamination. As an array system, the 96-well twin-layer system using immobilised algae is also amenable to high-throughput and massively parallel applications increasingly sought after in algal bio- and environmental technology.     

CYANOPHYTA-BACTERIA SYSTEMS: EFFECTS OF ADDED CARBON COMPOUNDS OR PHOSPHATE ON ALGAL GROWTH AT LOW NUTRIENT CONCENTRATIONS1

Planktonic blue-green algae are known to be always associated with bacteria. Earlier work has shown that the addition of a bacteria-assimilable carbon source to a normal Zehnder-Gorham culture medium (No. 11) will produce enhanced growth of these algae when atmospheric CO₂ has become the limiting factor. In new work, phosphate-rich culture media were diluted so that they simulated nutrient levels found in the Great Lakes, e.g., Lake Erie. At these low concentrations and when atmospheric CO₂ was not available in a sufficient supply, the addition of sucrose to either a 1/100 or a 1/1000 diluted Z-G, medium (10 mg or 2 mg of sucrose, respectively) also produced enhanced, growth of the tested blue-green algae. The stimulation of algal growth was apparently due to an increased bacterial production of CO₂ and possibly other carbon compounds approaching the composition of the CO₂ molecule. The literature suggests that during vigorous algal growth in lakes, atmospheric CO₂ will be severely limiting. Productive lakes always contain nonliving organic matter. The presence of bacteria-assimilable matter is probably one of the important factors leading to algal bloom.     

Harvest of Scenedesmus sp. with bioflocculant and reuse of culture medium for subsequent high-density cultures

The optimal flocculating conditions for harvesting high-density cultures of Scenedesmus sp. were investigated using inorganic coagulants and the bioflocculant produced by Paenibacillus polymyxa AM49. The flocculated medium as nutrients for subsequent algal cultivation was also tested. Consecutive treatment with 8.5 mM CaCl(2) and 0.2 mM FeCl(3) as coagulants and 1% bioflocculant from the culture broth of P. polymyxa AM49 showed the highest flocculating activity of up to 95% for high density algal cultures. The medium flocculated with the coagulants and bioflocculant showed less than 8% decrease in the growth yield in the subsequent algal cultivation. Furthermore, a 20% or 50% fresh BG11 medium supplement allowed the flocculated medium to maintain a high growth yield in subsequent algal cultivation. These results suggest that the flocculation method presented here is efficient and bio-friendly, and allows the reuse of the flocculated medium, thereby contributing to the economic cultivation and harvest of microalgae.     

Intrinsic autotrophic biomass yield and productivity in algae: experimental methods for strain selection

Using an analogy with fed-batch heterotrophic growth, the algal photoautotrophic yield Φ(DW) (in grams of dry weight biomass synthesized per micromole of absorbed photons) was derived from the algae batch growth behavior in nutrient-replete medium. At known levels of incident light, the yield Φ(DW) enables the estimate of a maximum productivity, and is therefore critical to compare and select algal cultures and growth conditions for large-scale production. The algal culture maximum growth rate was shown to be an unreliable indicator of autotrophic biomass yield. The developed carbonate addition method (carbonate addition, neutralization, and sealing) alleviated carbon limitations otherwise seen in aerated batch cultures, leading to two to five fold higher yield estimates. The fully defined FLX growth medium with variable ionic strengths (FLX1-100) supported excellent growth in most cultures tested. The chosen experimental methods and versatile FLX medium proved well-suited for small sample volumes and a high number of samples.     

Experimental demonstration of the roles of bacteria and bacterivorous protozoa in plankton nutrient cycles

We have used a model food chain composed of a natural bacterial assemblage, a pennate diatom and a bacterivorous microflagellate to investigate the factors controlling the relative importance of bacteria and protozoa as sources for regenerated nitrogen in plankton communities. In bacterized diatom cultures in which diatom growth was nitrogen-limited, the carbon:nitrogen (C:N) ratio of the bacterial substrate greatly affected which population was responsible for the uptake of nitrogen. When nitrogen was added as NH ₄ ⁺ and the cultures were supplemented with glucose, the bacteria competed successfully with the algae for NH ₄ ⁺ and prevented the growth of algae by rapidly assimilating all NH ₄ ⁺ in the cultures. Bacterivorous protozoa inoculated into these cultures grazed the bacterial population and remineralized NH ₄ ⁺ , thus relieving the nitrogen limitation of algal growth and allowing an increase in algal biomass. In contrast, bacteria in cultures supplemented with the amino acid glycine (C:N = 2) were major remineralizers of nitrogen, and the influence of protozoan grazing was minimal. We conclude that the relative importance of bacteria and protozoa as nutrient regenerators in the detrital food loop is dependent largely on the overall carbon:nutrient ratio of the bacterial substrate. The role of bacterivorous protozoa as remineralizers of a growth-limiting nutrient is maximal in situations where the carbon:nutrient ratio of the bacterial substrate is high.     

EFFECT OF GRAZING-ASSOCIATED INFOCHEMICALS ON GROWTH AND MORPHOLOGICAL DEVELOPMENT IN SCENEDESMUS ACUTUS (CHLOROPHYCEAE)

Populations of the phenotypically plastic alga Scenedesmus acutus Meyen were cultured in standard medium or in medium with filtered water from a Daphnia culture to examine the expression of ecomorphs in both water types. A rapid formation of four- and eight-celled coenobia was observed in the presence of Daphnia water, but not in standard medium. Moreover, cell dimensions were increased in the Daphnia water containing medium. Population growth rates were comparable in both water types; however, the carrying capacity appeared significantly lower in Daphnia water. Daphnia needed to feed on digestible food to produce the colony-inducing chemicals; medium from starved animals and Daphnia fed polystyrene beads appeared inactive. Neither suspensions of homogenate of Scenedesmus and Cryptomonas nor auxins affected colony formation or growth rate in S. acutus. The colony-inducing infochemicals are probably not constituents of the algae themselves; however, modification of algal constituents in the digestive system of daphniids may be involved.     

固氮蓝藻“红圈病”的研究——Ⅰ.“红圈病”在大面积培养蓝藻中的危害以及防范措施

对蓝藻“红圈病”的病症、发病条件、传播程度方式、危害进行了研究。结果表明,发病条件与蓝藻最佳生长条件基本一致;病原通过水体和带病藻体传播,对藻的产量和质量均造成严重危害。同时探讨了综合防治措施。     

两株溶藻细菌的分离及初步研究

选择藻华现象严重的巢湖作为采样点,取3个不同位置的水样通过0.22μm的纤维滤膜过滤,培养后加入适应期的藻液中,取黄化藻液作为分离菌种的材料,初筛菌株经反复试验获得有较强抑藻能力的菌株,经生理生化鉴定及16S rDNA分子鉴定其种属。初筛得到45个菌株,有两个菌株WD1和WD2表现出溶藻作用。两株细菌的菌液经离心、高温灭菌、细胞破碎等处理对供试藻鱼腥藻也均有不同程度的抑制作用。分离得到两株溶藻细菌,WD1为约氏不动杆菌,WD2为门多萨假单胞菌。     

Interactions between the diatom Thallasiosira pseudonanna and an associated pseudomonad in a mariculture system.

The marine diatom Thallasiosira pseudonanna (3H) and several bacteria associated with it were isolated from batch cultures at the University of Delaware mariculture facility. The interaction between the algae and each of the bacteria was investigated. One of the isolates, T827/2B (Pseudomonas sp.), was incapable of surviving in f/2 culture medium unless the algae were present. When the algae and T827/2B were grown together in the f/2 medium, the bacterial growth was stimulated and the algal growth was inhibited. Bacterial filtrate had a similar effect on the algae, indicating that the bacterial effect is an indirect one most likely resulting from the excretion of a harmful compound into the medium. Preliminary characterization of the material excreted by the bacteria indicates that it s proteinaceous in nature. The interactions observed does not fit into any single category of interactions but can be explained as a combination of competition and indirect parasitism.     

A BATCH CULTURE METHOD FOR MICROALGAE AND CYANOBACTERIA WITH CO2 SUPPLY THROUGH POLYETHYLENE MEMBRANES1

A new method for CO₂ supply to photoautotrophic organisms was developed, and its applicability for measuring specific growth rates in shaken batch cultures of cyanobacteria and unicellular algae was shown. Small bags containing a concentrated carbonate buffer with a CO₂ partial pressure of 32 mbar were prepared from a thin foil of low density polyethylene (LDPE). These bags were inserted as CO₂ reservoirs (CRs) into polystyrene culture flasks with gas‐permeable screw caps, which were suitable to photometric growth measurement. CO₂ was released directly into the medium with membrane‐controlled kinetics. The CRs were not depleted within 1 week, although the atmosphere in the culture vessel exchanged rapidly with the ambient air. Rates of initial growth and final densities of the cultures of six different unicellular algal species and one cyanobacterium were markedly increased by diffusive CO₂ supply from the CR. In the presence of a CR, growth was exponential during the first 2 d in all cultures studied. The method described allowed a high number of measurements of specific growth rates with relatively simple experimental setup.     

Influence of low doses of mutagens on growth of algae on a solid medium

The effects of low doses of two mutagens (UV-light and ethyl methansulphonate) on the growth of algae on a solid minimal (anorganic) medium were studied. It was supposed that low doses of mutagens will be more suitable for the induction of growth mutations. The UV-light had an inhibitory effect in the whole range of the applied doses on the growth of colonies of algae from the cells inoculated on the solid medium immediately after irradiation. Ethyl methansulphonate produced growth stimulation if applied in the lowest doses. The growth was inhibited in a further range of doses and then there appeared the range of lethal doses. The growth responses to the influence of these mutagens were different in all the three algal species used and to the previous cultivation conditions before their exposure to the mutagens. It is certain that most of these growth responses are only modifications. The influence of ethyl methansulphonate differed according to the method of application. If it affected the algae for a long time (it means if they were inoculated directly on the solid medium containing mutagens), or, if the algae were exposed to its influence immediately before their inoculation on the solid medium, the growth responses of colonies were quite different. Growth responses with the single studied strains differed quantitatively only.     

Interactions between the diatom Thallasiosira pseudonanna and an associated pseudomonad in a mariculture system

The marine diatom Thallasiosira pseudonanna (3H) and several bacteria associated with it were isolated from batch cultures at the University of Delaware mariculture facility. The interaction between the algae and each of the bacteria was investigated. One of the isolates, T827/2B (Pseudomonas sp.), was incapable of surviving in f/2 culture medium unless the algae were present. When the algae and T827/2B were grown together in the f/2 medium, the bacterial growth was stimulated and the algal growth was inhibited. Bacterial filtrate had a similar effect on the algae, indicating that the bacterial effect is an indirect one most likely resulting from the excretion of a harmful compound into the medium. Preliminary characterization of the material excreted by the bacteria indicates that it s proteinaceous in nature. The interactions observed does not fit into any single category of interactions but can be explained as a combination of competition and indirect parasitism.     

柄杆菌对固氮蓝藻生物量及色素的影响

柄杆菌对固氮蓝藻生物量和色素的影响研究结果表明:多态柄杆菌(Caulobacter polymorphus)017-41或新月柄杆菌(Caulobacter creseentus)CB_2的活菌、死菌及破碎细胞悬液分别与鱼腥藻(Anabaena)、念珠藻(Nostoc)不同藻珠混合培养时,试验组生长量均优于对照组;对衰老黄化的藻培养物的生长刺激作用尤为显著;试验组藻培养物的藻蓝或藻红素含量亦明显高于对照组。其作用机理尚待进一步阐明。     

藻类中添加剂的应用研究进展

藻类富含多种营养元素和活性物质,具有重要的经济价值和广阔的应用前景。现阶段藻类培养多采用露天跑道池,成本低,但易受环境影响。不适宜的环境条件(温度、酸性、重金属、紫外、盐度和光强胁迫等)会造成藻类生产成本上升、产品产量及品质下降等后果,严重制约藻类养殖业及相关产业的发展。添加剂不仅能有效促进藻类生长,还能缓解环境胁迫对其带来的逆境伤害。将近年来添加剂在藻类生长及抗逆方面的应用进行系统汇总,并对已阐明的几种添加剂的作用机理进行分析整理。添加剂在藻类中的应用研究大多停留在生长及生理指标的测定,如藻细胞密度、光系统活性、渗透调节物质含量、脂质含量、过氧化氢酶和硝酸还原酶活力等,仅有少部分研究是利用分子技术测定防御基因的表达情况,尝试进一步探究藻类抗逆性分子机制。旨在为研究者进一步明确常用添加剂在藻类中的信号传导机制及改善非生物胁迫造成的产品产量及品质的问题提供理论依据,具有重要现实意义。     

Astaxanthin production by a green alga,Haematococcus pluvialis accompanied with morphological changes in acetate media

Two acetate containing media were developed for astaxanthin production by a green unicellular alga, Haematococcus pluvialis. The basal medium, a vegetative growth medium facilitated the algal cell growth, whereas the modified medium was likely to induce morphological changes with the formation of large cysts and bleached cells which seemed to consequently enhance the carotenoid biosynthesis. In the two-stage culture, the injection of ferrous ion with acetate into the basal medium on the fourth day, was greatly stimulative for both the algal cell growth and the astaxanthin formation at a high light intensity. In addition, carotenoid precursors, mevalonate and pyruvate were effective on the carotenoid formation in the modified medium. Pyruvate was an especially good carbon source both for the algal cell growth and the carotenoid synthesis.     

Cage Culture Turbidostat: a Device for Rapid Determination of Algal Growth Rate

The present cage culture turbidostat consists of a growth chamber and a control unit. The microorganisms (photoautotrophic algae) are kept in the growth chamber by porous membranes (pore size 1 to 3 μm) which retain the algae but allow efficient exchange of the growth medium. Flow rate and composition of the medium can therefore be varied independently of algal population density. A reciprocating pumping mode of the medium is introduced to obtain more gentle clearance of membranes than that provided by rotation or stirring in other membrane fermentors. Pulsed light and a light-emitting diode/light-sensitive transistor couple are used to monitor the turbidity of the culture, independent of external light needed for growth. The control unit keeps the turbidity constant by frequent activation of the dilution pump. Theoretical analysis of growth in the turbidostat shows that integrated activation time of the dilution pump is proportional to the growth rate of the organism. Theoretical analysis was also used to determine minimum flow-rate and nutrient concentration of medium to cover the requirement of the algae. Experiments with three different marine diatoms were carried out, and they demonstrated that the growth rate could be determined every hour and that the cultures could be kept at constant turbidity over 10 to 14 days at least.