Application of <Emphasis Type="Italic">Gracilaria lichenoides</Emphasis> (Rhodophyta) for alleviating excess nutrients in aquaculture

A study was conducted in Xiangshan Bay, Ningbo, China, using red alga Gracilaria lichenoides to alleviate nutrient pollution in shrimp (Litopenaeus vannamei) and fish (Epinephelus awoara) culture ponds. Our results showed that G. lichenoides was efficient at absorbing inorganic nitrogen (IN) and inorganic phosphate (IP), and maintained a more stable dissolved oxygen (DO) level. A total of 506.5 kg (1,013 kg ha⁻¹) of shrimp and 210.5 kg (421 kg ha⁻¹) of fish were harvested from the shrimp/algae (SA) and fish/algae (FA) ponds, respectively. Only 53.5 kg shrimp were harvested from the shrimp pond without Gracilaria (S) due to anoxic asphyxia, and 163 kg fish were harvested from the fish culture pond without Gracilaria (F). Compared with using microalgae, bioremediation by macroalgae has no risk of harmful algal blooms (HABs), and it is easy to control seaweed biomass. During the experiment, there was a better environmental condition (lower chemical oxygen demand, IN, IP and chlorophyll a concentrations) in the ponds with Gracilaria. Furthermore, Gracilaria spp. can be used as food for abalone or other aquacultured animals and thus enhance economic return.     

World-wide use and importance of Gracilaria

The world's first source of agar, from the middle of the seventeenth century, was Gelidium from Japan, but by the beginning of the twentieth century demand for the phycocolloid exceeded of the supply of this alga. Since then Gracilaria has played an important role in the production of agar. Currently agars are obtained from five genera in three orders of red algae and marketed as ‘natural agar’ in squares or strips or as ‘industrial agar’ in powder form. The development of production processes through alkaline hydrolysis of sulphates has allowed a good quality food agar to be obtained from Gracilaria. This does not show the synergistic reaction with locust bean gum apparent with Gelidium agar. The term ‘agaroids’ is applied to Gracilaria agars produced without alkaline hydrolysis of sulphates, with greater sulphate content and much less gel strength. Unlike Gelidium, Gracilaria has to be processed in a short period of time and cannot be allowed to remain in storage for use during years of lower availability. Statistics of imports of agarophytes to Japan during the last 10 years give an indication of the state of the market. During this period there was a marked reduction in Gracilaria imports, mainly from Chile, but also the Philippines, Indonesia and South Africa, mainly due to the overall increase in the capacity of agar production in Gracilaria-producing countries.     

Opportunities and challenges for the development of an integrated seaweed-based aquaculture activity in Chile: determining the physiological capabilities of <Emphasis Type="Italic">Macrocystis</Emphasis> and <Emphasis Type="Italic">Gracilaria</Emphasis> as biofilters

Seaweed production is a reality in Chile. More than ten species are commercially used to produce phycocolloids, fertilizers, plant growth control products, human food or animal fodder and feed additives. These multiple uses of algae offer a number of possibilities for coupling this activity to salmon, abalone and filter-feeder farming. In this context, different experiments carried out in Chile have demonstrated that Gracilaria chilensis and Macrocystis pyrifera have great potential in the development of an integrated aquaculture strategy. The present Integrated Multi-Trophic Aquaculture (IMTA) approach study showed that Gracilaria can be cultured best at 1 m depth whereas Macrocystis has an especially good growth response at 3 m depth. Both species use available nitrogen efficiently. On the other hand, high intensities of solar radiation (UV and PAR) can be critical at low depths of cultivation, and our results indicate that both species show photosynthetic susceptibility mainly at noon during the summer. The demand of Macrocystis for abalone feeding is increasing, thus improving the opportunity for developing an integrated nutrient waste recycling activity in Chile. Although Gracilaria shows a higher nitrogen uptake capacity than Macrocystis, its market value does not yet allow a massive commercial scaling.     

Agar yield and quality of Gracilaria chilensis (Gigartinales,Rhodophyta) in tank culture using fish effluents

Tank cultivation of Gracilaria using fish effluents has permitted a production of 48 kg m^–2 yr^–1 and can reduce the dissolved nitrogen loads in the seawater. We report the yield, gel strength, gelling and melting point of agar from Gracilaria cultivated in tanks with seawater previously utilized in intensive, land-based salmon cultures and compared to a control using directly pumped seawater, over a study period of 22 months. The results show that the highest agar yield (20 to 22%) was obtained when Gracilaria was cultivated with pure seawater as compared to the fish effluents. The gel strength, gelling and melting point were higher in the agar obtained from algae cultured with fish effluents. During the spring, the gel strength, gelling and melting point increased in tanks with fish effluents and decreased in tanks with a supply of pure seawater.     

Experimental tank cultivation of Gracilaria sp. (Gracilariales,Rhodophyta) in Ecuador

This paper describes experiments to grow a local and still unidentified species of Gracilaria in shrimp hatcheries in Ecuador. The experiments used outdoor tanks of 1 and 18 m³ capacity, with continuous aeration and water renewal every two and five days, respectively. The sea water (salinity 34 ppt) was enriched with Guillard's f/2 medium; light and temperature were monitored but not controlled. One kg of fresh seaweed, inoculated into each tank, produced a biomass of ca. 3 kg in a period of 35 days in the 1 m3 tank and 18 kg in 43 days in the 18 m³ tank. We therefore believe that it is technically feasible to use the large infrastructure of existing shrimp hatcheries in Ecuador to produce Gracilaria.     

Saldanha Bay,South Africa: recovery of Gracilaria verrucosa (Gracilariales,Rhodophyta)

Since World War II the greater Saldanha Bay lagoon system, South Africa, has been an important Gracilaria producer. Two agar factories, built in the 1960's, used Gracilaria from Saldanha Bay as their raw material. In the early 1970's the industry was destroyed as a result of dredging and marine construction operations to establish a harbor in the bay for loading ore. These environmental changes destroyed stocks and prevented the previously significant beachings of the seaweed from occurring. After a few years of no or very low commercial production, the resource slowly started to recover. The size of Gracilaria drifts increased over the following eight years to approximately one-third of the original output. This trend seems to continue. Although the stocks and resultant drifts are unlikely to recover fully to their original quantity, current production is already sufficient to ensure re-establishment of a seaweed industry in Saldanha Bay. This could have considerable socio-economic impact on the area.     

Chemical structure and quality of agars from Gracilaria

Agar polymers synthesized by species of the genus Gracilaria constitute a complex mixture of molecules, containing several extremes in structure. Sulphate hemi-esters, methyl ethers and pyruvic ketals can alter in a number of ways the structural regularity of agar based on strictly 3-O-linked β-l-galactopyranose and 4-O-linked α-l-galactopyranose residues. In comparison with agars from Gelidium and Pterocladia, agars from Gracilaria can have higher degrees of sulphation, methoxylation and pyruvylation. The gelling ability of agars from most of Gracilaria species is considerably improved by adopting, before extraction, an alkali pretreatment which converts α-l-galactose 6 sulphate into 3,6-anhydro-α-l-galactose. Native agars obtained from Gracilaria cannot be classified, with few exceptions, as bacteriological grade agar as they have a high content of methoxyls and consequently high gelling temperatures. On the contrary, the genus Gracilaria is considered the most important source of food and sugar-reactive grade agars. Among techniques which can be used to study algal polysaccharides, combined ¹H and ¹³C nuclear magnetic resonance spectroscopy represent the most effective and powerful method for the investigation of the chemical structure of agarocolloids.     

Gracilaria conferta and its epiphytes: 3. Allelopathic inhibition of the red seaweed byUlva cf.lactuca

In a previous study (Svirski et al., 1993), it was found that growth inhibition ofGracilaria spp., when cultured in the presence ofUlva cf.lactuca, was not due to shading or nutrient depletion, but seemed to be caused by competition for inorganic carbon or some type of allelopathy. In the present study, we attempted to differentiate between these two possible influences by (1) growing the two algae in biculture under various conditions, but keeping inorganic carbon levels constant and measuring net photosynthesis, respiration and growth rates, and by (2) measuring growth rates ofGracilaria spp. in the presence of extracts derived from media previously used to growUlva cf.lactuca.Both net photosynthesis and growth rates ofGracilaria spp. in biculture were inhibited, despite CO₂ (and also HCO₃ ^–) levels being kept constantly high in the culture media. It is likely that these responses were due to markedly enhanced rates of dark respiration inGracilaria spp. when grown together withUlva cf.lactuca. Growth ofGracilaria spp. was also inhibited by extracts derived from seawater in whichUlva cf.lactuca had previously been grown. The strong inhibition by ethyl acetate and chloroform extracts indicate an allelopathic effect onGracilaria spp.     

Comparison of growth rate in culture,dry matter content,agar content and agar quality of two New Zealand red seaweeds,Gracilaria chilensis Bird,McLachlanet Oliveira andGracilaria truncata Kraft

Plant growth rates and agar characteristics were compared for two agarophytes,Gracilaria chilensis (Gracilariales, Rhodophyta) andG. truncata, so that the suitability ofG. truncata for mariculture could be assessed.G. chilensis plants grew steadily in the laboratory culture system at rates of 5–8% day^-1 (mean Relative Growth Rate) throughout the 6 week experimental period, with no decline in health.G. truncata plants grew at rates of 2–4% day^-1 for 5 weeks, but during the sixth week their apical tips became necrotic and growth rates fell to zero. There was no significant difference in the dry matter content (as a percentage of their wet weight) between the two species ofGracilaria, with values falling in the range of 16–19%. Slightly higher agar yields were obtained from alkali-treatedG. chilensis (17–20% dry matter) than from untreatedG. truncata (16–18%) although the agar fromG. truncata formed softer gels from which it was more difficult to recover. The gel strength of untreated agar extracted fromG. chilensis was very low (ca. 100 g cm^-2 for a 1% gel) but when an alkali treatment step was included in the extraction process, it increased to 520 g cm^-2 for a 1% gel. Contrary to an earlier report, untreatedG. truncata agar also had a very low gel strength (ca. 100 g cm^-2 for a 1.5% gel), but it rose to only 167 g cm^-2 after alkaline treatment.     

Associational plant refuges: convergent patterns in marine and terrestrial communities result from differing mechanisms

Summary An associational plant refuge occurs when a plant that is susceptible to herbivory gains protection from herbivory when it is associated with another plant. In coastal North Carolina, the abundance of the palatable red alga Gracilaria tikvahiae is positively correlated with the abundance of the unpalatable brown alga Sargassum filipendula during times of increased herbivore activity. To see if grazing by the sea urchin Arbacia punctulata could generate this pattern, controlled experiments were conducted in out-door microcosms and in the laboratory. Gracilaria beneath a canopy of Sargassum was eaten significantly less than Gracilaria alone. When Arbacia were excluded, Gracilaria alone grew significantly more than Gracilaria beneath Sargassum, demonstrating that Sargassum is a competitor of Gracilaria. Experiments investigating Sargassum's deterrent role indicated that Sargassum decreased the foraging range of Arbacia and the rate at which it fed on Gracilaria. Additional experiments with plastic Sargassum mimics indicated that the decreased grazing on Gracilaria was not a result of Sargassum morphology, but was probably attributable to some chemical characteristic of Sargassum. The pattern of increased grazing in monocultures (only Gracilaria present) versus polycultures (both Gracilaria and Sargassum present) demonstrated in this study also has been demonstrated for plant-insect interactions in terrestrial communities. In these communities, insect density is higher in monocultures than in polycultures because insects find and immigrate to monocultures more rapidly, and once in a monoculture, they emigrate from them less often than from polycultures. In this study, urchins did not find and immigrate to monocultures more rapidly, nor did they tend to stay in them once they were found; in fact, they emigrated from monocultures of Gracilaria more rapidly than from Gracilaria and Sargassum polycultures. Increased grazing in Gracilaria monocultures resulted from increased rates of movement and feeding of individual herbivores, not from increased herbivore density as has been reported for terrestrial systems.     

Studies on the yield and gel strength of agar from Gracilaria domingensis Sonder ex Kuetzing (Gracilariales,Rhodophyta) following the addition of calcium

Studies were carried out on the seasonal variation in yield and gel strength of agar from Gacilaria domingensis with and without the addition of calcium chloride. Extraction was done with and without treatment with 1% hydrochloric acid. The results showed an increase in yield and gel strength when an alkaline solution of calcium was used, but the gel strength was low. For commercial use, Gracilaria domingensis should be mixed with better quality Gracilaria species because of its low gel strength.     

Integrated seaweed cultivation on an abalone farm in South Africa

Land-based abalone aquaculture in South Africa, based on the local species Haliotis midae, started in the early 1990s and has grown rapidly in the last decade, with 13 commercial farms now producing over 850 t per annum. Over 6,000 t per annum of kelp Ecklonia maxima are now harvested for this purpose, and some kelp beds are reaching maximum sustainable limits. Research into seaweed aquaculture as feed (Ulva and some Gracilaria) for abalone started in the late 1990s on the southeast coast (where there are no kelp beds) using abalone waste water. A growing body of evidence suggests that a mixed diet of kelp plus other seaweeds can give growth rates at least as good as compound feed, and can improve abalone quality and reduce parasite loads. A pilot scale Ulva lactuca and abalone integrated recirculation unit using 25% recirculation was designed and built on the south west coast of South Africa using one 12,000-L abalone tank containing 13,200 15 ± 2.5 g abalone, connected to two 3,000-L seaweed tanks containing an initial starting biomass of 10 kg of seaweed, replicated 3 times. In an 18-month period, there were no significant differences in abalone health or growth rates, sediment build up and composition, mobile macro fauna densities and species between the recirculation or the flow-through units. Transfer of oxygen generated by the seaweeds to the abalone tanks was poor, resulting in the recirculated abalone tanks having lower (33%) dissolved oxygen concentrations than a comparable flow-through abalone unit. Seaweed nutrient content and specific growth rates in the units were comparable to seaweeds cultivated in fertilized effluent (SGR = 3.2 ± 3.4%.day⁻¹; Yield = 0.2 ± 0.19 kg.m².day⁻¹). Indications were that at this low recirculation ratio the seaweeds in the units were nutrient limited and that there were no negative effects to the abalone being cultivated in such a recirculation unit at this recirculation ratio.     

Recent trends of seaweed production in Chile

In the last fourteen years the production of seaweeds in Chile has ranged from 74 000 to 229 000 wet metric tons per year and has included about twenty species belonging to Phaeophyta and Rhodophyta. The only source of this production has been the exploitation of natural beds, except for Gracilaria, which is the only case of commercial cultivation and contributes significant quantities to total production. Initially most of the raw material was exported but currently important quantities of Gracilaria and several carrageenophytes are being processed by local industry. Changes in production of the main resources are analyzed with consideration of potential demand, level of knowledge about natural beds, and the situation of total Gracilaria farming, in order to attempt predictions for the supply. Current possibilities of applying new technologies to cultivate other economically important Chilean seaweeds are also analyzed and discussed.     

The yield,physical and chemical properties of agar gel from Gracilaria species (Gracilariales,Rhodophyta) of the Kenya coast

Six species of Gracilaria, G. corticata J. Agardh, G. crassa Harvey, G. millardetii J. Agardh, G. salicornia (J. Ag.) Dawson, G. verrucosa (Huds.) Papenfuss and Gracilaria sp, collected from different stations along the Kenya coast were studied. The yield of hot water native agar extract ranges from 8.1–30% of dry weight, with G. verrucosa and G. salicornia having the greatest and the least yield, respectively. The gel-strength of 1.5% agar solution was highest in G. verrucosa (220 g cm^–2) and lowest in G. corticata (< 60 g cm – 2) whereas the highest gelling temperature was recorded for Gracilaria sp. (40.4 °C) and the lowest in G. verrucosa (28.9 °C). 3,6 anhydrogalactose content was the highest in G. verrucosa and the lowest in G. corticata while sulphate content was higher in the latter.     

Inorganic nutrition in pond cultivated Gracilaria conferta (Rhodophyta): nitrogen, phosphate and sulfate

The experience gained in the outdoor cultivation system of Gracilariaconferta in Israel suggests three main inorganic variables: inorganic carbon,macronutrients and micronutrients. The effects of the macronutrientsnitrogen, phosphate and sulfate on growth rate, dry weight yield, and agaryield and quality were assessed in this research. The starvation periodbetween ammonium pulse feedings affected the agar yield and its quality asa function of the water temperature. Phosphate was the secondmacronutrient in the usual 1:10 proportion. Dark starvation as well assulfate starvation decreased agar yield. The results of nitrogen or sulfatestarvation suggest a biphasic process of agar biosynthesis. Starting with anoverall growth stage richer with sulfated agarose, and turning into an agaraccumulation stage with more neutral agarose. The elucidation of theintegrated effects of starvation and seasonality on agar production of Gracilaria is the major contribution of this research.     

Interaction mechanisms between Gracilaria chilensis (Rhodophyta) and epiphytes

Epiphytes can have a considerable effect on Gracilaria production, and Ulva is one of the commonest algal species identified as an epiphyte, reaching loads of 60% (g of epiphytes per g of Gracilaria) in the intertidal cultures of southern Chile. This study evaluates the relative importance of light reduction, addition of weight to the host thalli and nutrient depletion, as mechanisms determining the interaction effects of Ulva epiphytes on Gracilaria cultivation. Using field experiments undertaken during the main Gracilaria growth season (spring), we evaluate the mechanisms of epiphyte-host algae interaction by manipulating artificial epiphytes. The results indicate that Ulva can significantly depress Gracilaria biomass production and that the addition of weight to the host algae and the consequent dislodgement increase, appear to be the main mechanisms involved in the Ulva-Gracilaria interaction. However, the light reduction caused by the epiphytes can also partially explain the reduction in Gracilaria production. Nutrients depletion would not appear to fully account for the detrimental effects of Ulva over Gracilaria in intertidal farming areas of southern Chile.     

Phycological research in the development of the Chinese seaweed industry

The term seaweed industry is employed in a broad sense and includes production both of commercial seaweeds such as Laminaria and Porphyra by phycoculture and of processed seaweed products, such as algin, agar and carrageenan.Before the founding of the People's Republic, China had a very insignificant seaweed industry, producing small quantities of the purple laver Porphyra and the glueweed Gloiopeltis by the primitive rock-cleaning method and the kelps Laminaria and Undaria by the primitive stone-throwing method, both aiming at enhancing the growth of the wild seaweeds. Also, a small quantity of agar was manufactured by the traditional Japanese method of gelling, freezing, thawing and drying the product. The small production was not sufficient to meet the demand of the Chinese people who for ages have appreciated seaweeds and their products for food. Therefore, large quantities of seaweeds and seaweed products had to be imported from various countries, for instance, Eucheuma and Gracilaria from Indonesia and other southeastern Asian countries, Laminaria and agar from Japan, even Porphyra from the USA. Annual Laminaria import from Japan generally amounted to over 10 000 tons and in some years approached 20 000–30 000 tons. Some quantities of the glueweed Gloiopeltis and the vermifuge weed Digenea simplex were exported, mainly to Japan.Since the founding of the People's Republic of China in October, 1949, China has exerted efforts to build up a self-supporting seaweed industry. Now after a lapse of 30-some years, a sizable seaweed industry has been developed. China is now able to produce by phycoculture more than one million tons of fresh seaweeds, including Laminaria, Undaria, Porphyra, Eucheuma, Gracilaria etc. and several thousand tons of seaweed extracts, including algin, agar, carrageenan, mannitol and iodine. At present, China still imports some quantities of seaweeds and seaweed products from various countries but is able to produce sufficient quantities to meet the people's need and even to export some quantities of the seaweeds Laminaria, Undaria and Porphyra and the seaweed products algin and mannitol.At the Tenth International Seaweed Symposium, I presented a paper on the Marine Phycoculture of China, in which I emphasized on the methods of cultivation (Tseng 1981b). Therefore I would like to take this opportunity to supplement the last lecture by presenting a paper on the role of phycological research in the development of China's seaweed industry.     

Eutrophication assessment and bioremediation strategy in a marine fish cage culture area in Nansha Bay,China

On the basis of field data measured during four cruises from January to November 2007, variations in the characteristics of dissolved inorganic nitrogen and phosphate were analyzed in Nansha marine fish cage culture area, Ningbo City, China. Dissolved inorganic nitrogen (DIN) was selected as the parameter to balance seaweed absorption and fish DIN production. The contents of DIN and phosphate varied with different seasons, and eutrophication index (E) value ranged from 2.41 to 15.99, indicating serious eutrophication conditions; the annual average value of N/P of 32.95 indicates a nitrogen surplus in this system. The eutrophication condition in Nansha Bay was mainly caused by the fish cage culture activities. Based on their biological characteristics, Laminaria and Gracilaria were selected as the bioremediation species in winter and spring and summer and autumn, respectively. The optimal co-cultivation proportion of fish cage to Laminaria and Gracilaria in this bay was 1 cage, 450 m² and one cage, 690 m², respectively.     

Characterization of an agar fraction extracted from Gracilaria dura (Gracilariales,Rhodophyta)

The evaluation of biopolymers of biotechnological interest has stressed the importance of renewable sources from the northern Adriatic Sea. In the frame of this research program, the red alga Gracilaria cf. dura, has been studied. Agar was obtained from Gracilaria cf. dura by means of alkali treatment and hot water extraction. In order to purify the extracts, amylase and isopropyl alcohol were used. Analysis of sugars was carried out by means of gas chromatography on alditol acetate derivatives; sulfate content was evaluated by means of infrared spectroscopy. An analysis of molecular weight distribution was carried out by means of high performance gel permeation chromatography coupled with a low angle laser light scattering device as detector. Analytical data were examined in terms of specificity of the algal source utilized, and the agar fractions extracted by means of this procedure were compared with a commercial sample.     

Gracilaria and its epiphytes: 4. The response of two Gracilaria species to Ulva lactuca in a bacteria-limited environment

The responses of Gracilaria lemaneiformis, an easily epiphytized host,and the relatively resistant G. cornea mutant, to the green alga Ulva lactuca were studied using biculture experiments with and withoutantibiotics. Both Gracilaria species grown with and without U.lactuca showed different levels of growth rate, release of hydrogenperoxide and of halogenated hydrocarbons. These quantitative differencesled to a successful response against Ulva lactuca in the case of G.cornea mutant and to a failure in response in the case of G.lemaneiformis. The response of each Gracilaria species to U.lactuca was qualitatively similar to its response to bacteria. This suggeststhe involvement of oligosaccharide elicitors produced in the presence ofepiphytes and bacteria. A clear Gracilaria inhibition was demonstratedwith extracts of the culture medium. It appears that hydrogen peroxide,halogenated hydrocarbons and oligosaccharides may be components of theinhibitory activity of the extracts. The responses of Gracilaria speciesto the presence of U. lactuca suggest the characterization of adefence response.