D. Proulx

Algae and waste water

Microalgal cultures offer an interesting alternative for waste water treatment (urban, industrial or agricultural effluents) because they provide a tertiary biotreatment coupled with the production of potentially valuable biomass, which can be used for several purposes. We review the main abiotic, biotic and operative factors playing a role in the cultivation of microalgae. Various types of bioreactors are scrutinized keeping in view that the main limitation upon the type of usable bioreactors is the enormous volume of water to be treated. The choice of suitable microalgae and cyanobacteria is examined in terms of productivity and easiness of harvesting. The possible alternatives to harvesting are also reviewed with an emphasis on immobilized systems. Finally, the need for more research and development is discussed.

Nitrogen and phosphorus removal by high latitude mat-forming cyanobacteria for potential use in tertiary wastewater treatment

As part of a program to develop biological wastewatertreatment systems for cold climate areas four strainsof filamentous, mat-forming cyanobacteria isolatedfrom Arctic and Antarctic environments were evaluatedfor their nutrient stripping and growth capabilities. A tropical strain, Phormidium bohneri, known forits excellent performance in wastewater treatment, wasused as a comparison. Experiments were done inartificial media under controlled batch cultureconditions to avoid interactions with indigenousmicroorganisms such as bacteria and protozoa. Theculture medium simulated real effluents containinghigh concentrations of nitrate and phosphate.Temperatures (5, 15 and 25°C) and irradiances(80, 210, 350, 640 and 1470 µmol photon m-2s-1) wereselected according to situations encountered in avariety of field conditions. For all irradiancelevels, growth was satisfactory at 15 and 25 °C,but limited at 5 °C. At 25 °C a satisfactory nitrogen removal rate (3.5and 4.0 mg N L-1d-1) was obtained forone polar strain (Phormidium tenue) and thecontrol P. bohneri. At 15 °C, the bestnitrogen removal rate (3.5 mg N L-1d-1)was measured with P. bohneri while the best ratefor the polar strains was around 2.3 mg NL-1d-1. At 15 °C, a phosphorusremoval rate of 0.6 mg P L-1d-1 wasobtained with P. bohneri and polar strains P. tenue and Oscillatoria O-210. Nitrogen(NO3-) and phosphorus (PO43-)uptake rates increased as a function of irradianceover the range 80 to 350 μmolphoton m-2s-1. Our results indicate thattertiary biological wastewater treatment at lowtemperatures (5 °C) cannot be anticipated withthe polar strains tested, because they arepsychrotrophic rather than psychrophilic and thus growtoo slowly under conditions of extreme cold. However, it appears that these cyanobacteria would beuseful for wastewater treatment at moderately cooltemperatures (c. 15 °C), which are commonduring spring and fall in northern climates.

Biological tertiary treatment of urban wastewaters with chitosan-immobilizedPhormidium

SummaryChitosan:Phormidium aggregates (chitosan: algae=1:2, dry weight basis) were used as a biological tertiary treatment to remove the nitrogen (NH4+, NO2-, NO3-) and phosphorus (PO43-) from a secondary effluent. In a batch system, 71 and 92% of P−PO43-were removed after 6 and 24 h, respectively. The orthophosphate removal rate was identical for all three concentrations of algae-chitosan tested (3.3, 4.6, 5.9 g d. wt.·l-1), and was 90 μg±2 μg P−PO43-·l-1·h-1, for a 90% removal. Under control conditions (chitosan flakes only added to the effluent) 73 and 78% of PO43-were removed after 6 and 24 h respectively. A 95% removal of inorganic nitrogen (NH4+, NO2-, NO3-) was attained after 4–6 h withPhormidium immobilized on chitosan flakes, as compared to 30% with chitosan flakes alone (5 g d. wt.·l-1). The system gave a similar performance when operated semi-continuously over 5 days at a daily retention time of 1.0. In the presence of chitosan-immobilized algae, medium P−PO43-levels were reduced by 87.3%±6.4% after 24 h (61.1 μg±7.0 μg P·l-1·h-1). The reduction of inorganic nitrogen in the medium was 98% after 24 h (370 μg±50 μg N·l-1·h-1). In the presence of chitosan alone, some 60% orthophosphate removal was recorded, whereas no reduction of nitrogen was observed. Disappearance of orthophosphate was attributed to its co-precipitation with calcium released from the chitosan by abrasion. The presence of the algae protected the chitosan from abrasion andPhormidium directly assimilated the orthophosphate and inorganic nitrogen, thus reducing their levels in the effluent.

Polar cyanobacteria versus green algae for tertiary waste-water treatment in cool climates

Forty-nine strains of filamentous, mat-forming cyanobacteria isolated from the Arctic, subarctic and Antarctic environments were screened for their potential use in outdoor waste-water treatment systems designed for cold north-temperate climates. The most promising isolate (strain E18, Phormidium sp. from a high Arctic lake) grew well at low temperatures and formed aggregates (flocs) that could be readily harvested by sedimentation. We evaluated the growth and nutrient uptake abilities of E18 relative to a community of green algae (a Chlorococcalean assemblage, denoted Vc) sampled from a tertiary treatment system in Valcartier, Canada. E18 had superior growth rates below 15°C Canada. (µ = 0.20 d-1 at 10°C under continuous irradiance of 225 µmol photon m-2 s-1) and higher phosphate uptake rates below 10°C (k = 0.050 d-1 at 5°C) relative to Vc (µ=0.087 d-1 at 10°C and k = 0.020 d-1 at 5°C, respectively). The green algal assemblage generally performed better than E18 at high temperatures (at 25°C, µ = 0.39 d-1 and k = 0.34 d-1 for Vc; µ = 0.28 d-1 and k = 0.33 d-1 for E18). However, E18 removed nitrate more efficiently than Vc at most temperatures including 25°C. Polar cyanobacteria such as strain E18 are appropriate species for waste-water treatment in cold climates during spring and autumn. Under warmer summer conditions, fast-growing green algae such as the Vc assemblage are likely to colonize and dominate, but warm-water Phormidium isolates could be used at that time.

Effects of a bacterial biofilm on intensive Daphnia culture

Abstract In aquatic systems microorganisms tend to adhere to solid-liquid interfaces. The role played by bacteria in the nutrition of zooplankton organisms has been well recognized. The purpose of this experiment was to know if a larger area of microbial film, resulting from the addition of extra walls (fourfold), would increase the production of Daphnia magna. Daphnid cultures were carried out in plexiglass tanks which received identical initial populations and were fed Scenedesmus grown on secondary urban effluent. The effect of the biofilm was further evaluated by including control tanks which were cleaned daily. Results showed that the presence of the biofilm and additional surfaces increased the density, biomass and biomass harvest of D. magna fourfold, while the biofilm had no observable effect on the biochemical composition of the Daphnia. The increased productivity in the tanks with the extra surface area can be explained by both their more extensive biofilm and a ‘wall effect’ acting on the spatial distribution of the organisms. The exact role of microorganisms is difficult to pinpoint; however their effect appears mostly qualitative and seems to be twofold, that is, in Daphnia nutrition and through the detoxification of the culture medium by nitrification. In view of the significant differences obtained at the bench-scale level (13 litres), there is little doubt that increasing the extent of biofilm, by the addition of surfaces in daphnid-rearing tanks, will result in higher productivities in large-scale installations.

Traitement tertiaere d'un effluent domestique secondaire par culture intensive de la cyanobactérie Phormidium bohneri

Abstract Phormidium bohneri, a self‐flocculating cyanobacterium, was grown outdoors in a 75 1 intensive culture basin (semi‐continuous system) and used for the tertiary treatment of domestic wastewater. The behavior, growth and purification potential of P. bohneri were studied. The nutrient removal efficiency (max.: Ni = 83%, 12.5 mgN 1‐1 d‐1; Pi =81%, 1.3 mg P l‐1 d‐1) of this process allows a quite rapid treatment of the secondary effluent (hydraulic retention time=1d). Stripping account for about 62% of nitrogen (NH3) removal while 38% is assimilated by P. bohneri. Inorganic phosphorus is removed mainly by precipitation (57%) and to a lesser extent is taken‐up by Phormidium (43%). The cyanobacterial biomass (P: 1.1%, N: 8.6%, protein: 53.5%, dry weight basis) can be easily harvested after the treatment by settling.

Establishment of a real-time PCR method for quantification of geosmin-producing Streptomyces spp. in recirculating aquaculture systems.

Geosmin and 2-methylisoborneol (MIB) have been associated with off-flavour problems in fish and seafood products, generating a strong negative impact for aquaculture industries. Although most of the producers of geosmin and MIB have been identified as Streptomyces species or cyanobacteria, Streptomyces spp. are thought to be responsible for the synthesis of these compounds in indoor recirculating aquaculture systems (RAS). The detection of genes involved in the synthesis of geosmin and MIB can be a relevant indicator of the beginning of off-flavour events in RAS. Here, we report a real-time polymerase chain reaction (qPCR) protocol targeting geoA sequences that encode a germacradienol synthase involved in geosmin synthesis. New geoA-related sequences were retrieved from eleven geosmin-producing Actinomycete strains, among them two Streptomyces strains isolated from two RAS. Combined with geoA-related sequences available in gene databases, we designed primers and standards suitable for qPCR assays targeting mainly Streptomyces geoA. Using our qPCR protocol, we succeeded in measuring the level of geoA copies in sand filter and biofilters in two RAS. This study is the first to apply qPCR assays to detect and quantify the geosmin synthesis gene (geoA) in RAS. Quantification of geoA in RAS could permit the monitoring of the level of geosmin producers prior to the occurrence of geosmin production. This information will be most valuable for fish producers to manage further development of off-flavour events.

Impact of water quality on the bacterial populations and off‐flavours in recirculating aquaculture systems

A variety of factors affecting water quality in recirculating aquaculture systems (RAS) are associated with the occurrence of off-flavours. In this study, we report the impact of water quality on the bacterial diversity and the occurrence of the geosmin-synthesis gene (geoA) in two RAS units operated for 252 days. Unit 2 displayed a higher level of turbidity and phosphate, which affected the fresh water quality compared with unit 1. In the biofilter, nitrification is one of the major processes by which high water quality is maintained. The bacterial population observed in the unit 1 biofilter was more stable throughout the experiment, with a higher level of nitrifying bacteria compared with the unit 2 biofilter. Geosmin appeared in fish flesh after 84 days in unit 2, whereas it appeared in unit 1 after 168 days, but at a much lower level. The geoA gene was detected in both units, 28 days prior to the detection of geosmin in fish flesh. In addition, we detected sequences associated with Sorangium and Nannocystis (Myxococcales): members of these genera are known to produce geosmin. These sequences were observed at an earlier time in unit 2 and at a higher level than in unit 1. This study confirms the advantages of new molecular methods to understand the occurrence of geosmin production in RAS.

Comparison of extenders, dilution ratios and theophylline addition on the function of cryopreserved walleye semen

Walleye (Stizostedion vitreum) is a species of interest for the diversification of North American aquaculture production, and semen cryopreservation is of particular value to this effort. To test the hypothesis that adjusting semen extender composition and dilution ratio increases sperm quality after thawing, three extenders (Ext1, Ext2, Ext3; all with DMSO as a cryoprotectant) and three dilution ratios (semen/extender: 1:5, 1:9, 1:15) were screened. The best results were obtained when semen was diluted at a 1:15 ratio with Ext 1, Rathbun extender supplemented with 7% DMSO, 4 mg/ml BSA and 7.5 mg/ml ProFam, a soy-based protein (P = 0.05, n = 6). This method resulted in 46 +/- 3% motility of the thawed spermatozoa and a mortality rate of 39 +/- 4% whereas Ext2 and Ext3 resulted in motility rates of only 10 and 5%. respectively. To test an additional hypothesis that phosphodiesterase inhibition improves sperm function, we assessed the fertility of sperm frozen in optimal conditions and thawed in the presence or absence of 5 mM theophylline (n = 5). The best result was achieved in water without theophylline, with fertilization rates ranging from 28.51 +/- 6.84 to 59.02 +/- 1.06% eyed-up stage, and theophylline reduced fertility (P < 0.05). Our data show that Ext1 at a dilution ratio of one part semen to 15 parts extender should be used for walleye semen cryopreservation and that the fertilizing media does not benefit from theophylline supplementation.

Growth of Daphnia magna on urban wastewaters tertiarily treated with Scenedesmus sp.

Abstract The productivity and composition of Daphnia magna were measured for high density populations grown in 5-litre flow-through tanks supplied with a tertiary effluent. This effluent was urban secondary wastewaters biologically treated with intensive cultures of a microalga Scenedesmus sp. Three daily feeding rates (575, 1150, 1750 mg dry weight algae day −1 tank −1 ), obtained by adjusting the flow of a tertiary effluent containing 115 mg dry weight of algae litre −1 , allowed average productions of 0·9, 1·6 and 2·6 g of Daphnia (wet basis) litre −1 week −1 which were proportional to the rations. When the flow of tertiary effluent was increased to 40 litres day −1 tank −1 ( ad libitum ), the average yield rose to 3·6 g of daphnids (wet basis) litre −1 week −1 . The maximum population reached 48 000 individuals litre −1 . The daphnid/algae evergy conversion ratio was 0·4. Algae were removed by Daphnia at a maximum efficiency of 95%. The composition of Daphnia biomasses was 59·5% protein, 9·5% fat and 5·6% carbohydrate on a dry weight basis. No deficiencies in methionine and linolenic acid were noted. Results indicated the remarkable potential of urban wastewaters for the production of protein and the high productivity of Daphnia magna .