The genus Ulva is one of the most commonly commercially cultivated seaweeds both on land in ponds and in the open sea. As a crop, its cultivation is accompanied by three pest groups: (1) diseases caused by pathogenic bacteria, algae, virii, and fungi, which cause perforations, deterioration, and decreased growth of thalli; (2) various grazers, e.g. small crustaceans, gastropods, and non-defined invertebrates, which decrease growth and damage the thalli; and (3) an epiphytic biofilm of microorganisms, which also causes damage to the end-crop decreasing its yield and value. Mitigation categories against Ulva pests may be divided into three categories: (A) pro-active human intervention by washing the infected thalli with acid, fresh water, proprietary chemicals or CO2-saturated seawater, the use of methanolic Ulva extracts, or exudate from the green seaweed, all of which are reported to restrict pest activity; (B) using the beneficial properties of probiotic bacteria thereby eliciting antifouling activity, with selective abilities against pathogens of Ulva spp.; (C) enhancing the effects of environmental changes on pests, i.e. acidification and increased seawater temperature, which mainly decrease the grazer population and thereby their pressure. A summary of these studies is presented as a guide for rapid responses and applied management techniques for the amelioration of cultivation diseases, grazers, and harmful epiphytes affecting the foliose green alga Ulva spp.

This mini review deals with selected aspects of the health and production of the red algal, agarophyte Gracilaria spp. Amongst common macroalgal diseases in the rhodophytes, “ice-ice” presents itself as perhaps the most common first appearing as an apical tip bleaching phenomenon. This phenomenon is widely studied and reported to be caused by defined bacteria. Regrading grazers, the consumers of Gracilaria are presented with resistance against attack. Finally, harmful epiphytes play a dominant role in Gracilaria production. The review provides information and guidance to aspects of various common pests of Gracilaria such as their attachment, damage caused, preferred conditions and various defense practices against their damaging impacts. Understanding and manipulating the red algal microbiome, under specific and varying cultivation conditions, will be a key factor in the holistic control of disease and pest management.

Intensive pond cultivation of Gracilaria may achieve maximal yields by enrichment of seawater with ammonia and CO2. In the present study, we examined the use of organic waste material as a source of ammonia and inorganic carbon. A biofiltration system was constructed, consisting of several fermentation tanks and a fluidized bed reactor for the required bioconversion of organic wastes. Gracilaria conferta was cultured in a 30 m(2) pond from which seawater was circulated through the biofiltration system for a period of nine months. As compared to the filtered seawater used for water supply, the outlets of the fermentation tanks and the fluidized bed reactor showed significant increases in dissolved inorganic carbon and ammonia concentrations and significant decreases in oxygen concentrations and redox potentials. The most efficient organic waste compound tested had the highest C/N ratio. No significant differences were found between the Gracilaria yields of the control and biofiltration pond systems. These results might contribute a fundamental improvement in the economy of Gracilaria pond cultivation by water recycling through such a biofiltration system. Statement of relevance: The system might contribute a fundamental improvement in the economy of Gracilaria pond cultivation.

While large grazers can often be excluded effectively from algal aquaculture operations, smaller herbivores such as small crustaceans and gastropods may be more difficult to control. The susceptibility of three Gracilaria species to herbivores was evaluated in multiple-choice experiments with the amphipod Ampithoe ramondi and the crab Acanthonyx lunulatus. Both mesograzers are common along the Mediterranean coast of Israel. When given a choice, the amphipod preferred to consume Gracilaria lemaneiformis significantly more than either G. conferta or G. cornea. The crab, however, consumed equivalent amounts of G. lemaneiformis and G. conferta, but did not consume G. cornea. Organic content of these algae, an important feeding cue for some mesograzers, could not account for these differences. We further assessed the susceptibility of a candidate species for aquaculture, G. lemaneiformis, against local algae, including common epiphytes. When given a choice of four algae, amphipods preferred the green alga Ulva lactuca over Jania rubens. However, consumption of U. lactuca was equivalent to those of G. lemaneiformis and Padina pavonica. In contrast, the crab showed a marked and significant preference for G. lemaneiformis above any of the other three algae offered. Our results suggest that G. cornea is more resistant to herbivory from common mesograzers and that, contrary to expectations, mixed cultures or epiphyte growth on G. lemaneiformis cannot reduce damage to this commercially appealing alga if small herbivores are capable of recruiting into culture ponds. Mixed cultures may be beneficial when culturing other Gracilaria species.

Intensive cultivation of seaweeds in tanks and ponds has been developed ill Israel during the past twenty years. The achievements and problems presented in this review are based on major studies from other parts of the world. The advantages and disadvantages of on-land free-floating Cultivation are analyzed ill it detailed and comprehensive way, using physical. chemical, and biological variables in tank cultures. The common target variables of this analysis are the growth rate and yield of the cultured seaweeds. The main species Gracilaria. Ulva. and Porphyra, which are presented here, are of economic value for the food and food additive market. This tank culture is compared to the upscale pond culture of bigger facilities. The unique subject of integrated seaweed cultivation with fish or marine invertebrates is discussed separately. Finally. ail economic analysis is presented in order to serve as a guideline for decision-making in seaweed cultivation for commercial products, and for considering future developments.

Absorption and fluorescence spectra. photosynthetic rates. pigment concentrations., and cell size were investigated during photoacclimation of Porphyra leucosticta, it red marine macroalga that thrives during winter in the eastern Mediterranean. Optical properties were measured oil intact thalli using image analysis or from crude extracts after disrupting algal tissue. Absorption spectra front crude extracts showed major peaks at 485, 620. and 664 run for chlorophyll a (chl a) and 500, 564 and 615 nm for phycoerythrin (PE) and phycocyanin (PC). Absorption peaks from intact thalli were less Conspicuous than those from crude extracts, possibly due 10 PE contributing to the bulk absorption of light by chl a in intact cells. Fluorescence emission of intact thalli was about threefold higher than that of crude extracts at 420 and 560 mm. Image analysis revealed differences in maximal fluorescence emissiors as related to cell size, cell age, and thallus functional areas, with female cells having nearly a twofold higher emission than vegetative cells or asexual spores. Changes in the fluorescence emissions observed during acclimation to irradiance could reflect the extent of energy transfer from PE and PC to chl a. variable photosynthetic efficiency or optical properties of the samples. This study suggests that image analysis can provide insights on seaweed photoaccliniation that seem to be lost when disrLlp:ing their tissues to crude extracts. For example, the high PE to chl a ratios observed with image analysis in fresh tissues May confer a unique photoprotective role of PE when P. leucosticta becomes exposed to high irradiances, even during winter time.

A comprehensive study of the major environmental factors affecting growth and chemical composition of Gelidium crinale is presented here for the first time. The purpose of this study was to analyze the significance of the major growth factors and to formulate them into regression models. For this purpose summer and winter experiments were conducted in small aerated seawater tanks under various light and fertilizer conditions. The effects of carbon source, pH control and microelements were also tested. Finally, an annual growth experiment was conducted in order to support the two-season experiment. The major limiting seasonal factor was found to be temperature, which positively affected weekly growth rate, dry weight and gel strength, while negatively affecting the agar percentage. Ammonium was also revealed as a major limiting factor, positively affecting chlorophyll and protein concentrations, and negatively affecting carbohydrate concentration. Other growth factors, such as light intensity, pH, carbon source and microelements, were less- or non-effective, being already almost optimal under ambient conditions.

Currently, Gelidium and Pterocladia (Gelidiales) are collected or harvested only from the sea. Despite several attempts to develop a cultivation technology for Gelidium, no successful methodology has yet been developed. Initial steps towards developmental efforts in Portugal, Spain, South Africa and Israel have been published. More developments have probably been performed but have not been published. Two different technological concepts have been tested for Gelidium cultivation: (1) the attachment of Gelidium fragments to concrete cylinders floating in the sea, and (2) free-floating pond cultivation technology. These vegetative cultivation technologies might be partially optimized by controlling physical, chemical and biological growth factors. The pond cultivation technology is the much more controllable option. The effects of all factors are discussed in detail in this review. It seems that the main difficulty with cultivation of Gelidium is its low growth rate. The claimed yields of the two technologies are far from being economically attractive at this stage of their development. It seems that in order to introduce Gelidium into commercial cultivation, major efforts in genetic improvement through selection or genetic engineering will be required. Only high yield strains will have the potential to compete economically with the present harvesting tradition. However, accumulated experience with genetic improvement of other useful seaweed species suggests that this is possible.

Studies on Caulerpa prolifera rhizoids were carried out to determine the possibility of mass culture, because the rhizoids produce a bio-adhesive. Rhizoids can be induced by cutting the base of a blade and floating it in a media or planting it in sand. Measurement of rhizoid production included determination of number, length, and the weight of attached sand grains. The growth experiments were for 1–2 weeks and fronds growth was compared to rhizoid production. Optimal conditions for rhizoid growth included low levels of nitrogen and phosphate (less than 5 and 2μM, respectively), low irradiance (30μmolphotonm−2s−1), moderate temperature (22–28°), continuous shaking, addition of microelements and auxin (1ppm) and initially detached fronds followed by attachment. Under these optimal conditions maximal weekly growth reached 70–170 rhizoids per blade, 7–11mm length and maximal attachment to sand grains. Blade growth of C. prolifera responded similarly to rhizoid production and reached a weekly growth rate of 30–130%.

The rhizoid section of the green alga Caulerpa prolifera (Cp) is active in attaching the developing plant to the substratum. A model system for the study of the adhesion of Cp rhizoids has been developed and identification of two putative adhesive polypeptides of Caulerpa (Vn-Cp) was revealed by immunodetection. A method for fast induction of new rhizoids was established using blade-base cutting followed by a few days of incubation. The new rhizoids were gently enclosed between two cover glasses and incubated until firm attachment developed. While analyzing protein extracts, two ∼60–70 kDa polypeptides (Vn-Cp I and Vn-Cp II) were identified by immunodetection with monoclonal antibodies to human vitronectin (Vn). The relative concentration values of the Vn-Cp proteins increased significantly in the ‘cell-wall’ fraction of the attached rhizoids during the incubation period. However, Vn-Cp proteins were not detected in non-attached rhizoids. Furthermore, the Vn-Cp proteins were also detectable on glass substratum subsequent to attached rhizoid removal. The induction and accumulation of Vn-Cp proteins on the ‘cell-wall’ of Caulerpa rhizoids and the firm attachment of the rhizoids to the glass substratum during the incubation period suggest that Vn-Cp proteins play a significant role in adhesion, which may be similar to the function of vitronectin in other adhesion systems. Furthermore, the high accumulation of Vn-Cp proteins on the glass substratum during attachment of new rhizoids suggests that the Vn-Cp proteins are secreted to the extracellular matrix and directly connect rhizoids to the glass substratum as an intermediate compound. These unique properties of Cp make it an excellent model system for the establishment of high amounts of adhesive material for future research.

Agar oligosaccharides in the neoagarobiose series were prepared by partial enzyme hydrolysis, separated on Biogel P2 and P4, and analyzed by high‐performance anion exchange chromatography with pulsed amperometric detection, yielding neoagarosaccharide fractions with a disaccharide repetition degree ranging from 1 (neoagarobiose) to more than 8 (neoagarohexadecaose). These fractions were analyzed for their biological activity toward the marine red alga Gracilaria conferta (Schousboe ex Montagne) J. et G. Feldmann in terms of increase of oxygen consumption, release of hydrogen peroxide, elimination of epiphytic bacteria, and induction of thallus tip bleaching. The structure–activity and dose–response relationships of neoagarosaccharides were very similar in the respiratory and oxidative burst responses and in their bactericidal properties, with neoagarosaccharides consisting of 6 to 8 disaccharide repeating units being the most active. All these responses were competitively inhibited by the reduced form of neoagarohexaose, neoagarohexaitol. In contrast, the tip‐bleaching response was light dependent, required much higher concentrations of neoagarosaccharides, and was not inhibited by neoagarohexaitol, suggesting that it is an unspecific oxidative stress reaction. Putative structural effects on the recognition of endogenous agar‐oligosaccharide elicitors by G. conferta are discussed.

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.

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.

Use of light, transmission, and scanning electronmicroscopes revealed that the epidermal cell wall ofthe red algal agarophytes McLachlan and J. Agardh consists of adecklamelle and outer and inner wall layers. The twospecies differed, with having asignificantly thicker outer wall and a more diffusedecklamelle. After induction, the zooids of would attach to glass slides and the twospecies of via an adhesion pad. Within a few days, 3–5 celled germlings penetrated thedecklamelle and outer wall layer of both basiphytes. By the time the epiphyte germlings reached the 15celled stage, they had penetrated the inner walllayer. The differences in epidermal cell wallconstruction between the two basiphytes may play arole in the ability of zooids of toattach in nature where epiphytization of is infrequent.

Elicitation of Gracilaria conferta (Schousboe ex Montagne) J. et G. Feldmann with oligoagars resulted in a defense response that was strong enough to kill epiphytic bacteria associated with the alga. Up to 60% of the resident bacterial flora of healthy plants was eliminated within 60 min after addition of neoagarohexaose to the algal medium. Single isolates of agar‐degrading bacteria that had been isolated previously from healthy or decaying algal tissues proved to be more sensitive. Some of them were generally unable to survive on healthy G. conferta. Others survived on unelicited plants. Approximately 90% of these more resistant agar degraders were eliminated within 15 min after elicitation. The bacterial degradation of dead tissue of G. conferta resulted in a release of elicitors. The elicitors accumulated in the medium and reached high enough concentrations within 24 h to induce a hypersensitive response in healthy algae. The eliciting agent could be destroyed with β‐agarase and was thus probably oligoagar. Application of antibiotics prevented the accumulation of the elicitor, which indicated that bacteria were responsible for its release from the algal biomass. The hypersensitive response of G. conferta after contact with oligoagars is thus a true defense response, because it enables the plant to protect itself efficiently from enzymatic attacks on its cell wall.

Use of light, transmission, and scanning electronmicroscopes revealed that the epidermal cell wall ofthe red algal agarophytes Gracilaria tikvahiaeMcLachlan and G. cornea J. Agardh consists of adecklamelle and outer and inner wall layers. The twospecies differed, with G. cornea having asignificantly thicker outer wall and a more diffusedecklamelle. After induction, the zooids of Ulvalactuca would attach to glass slides and the twospecies of Gracilaria via an adhesion pad. Within a few days, 3–5 celled germlings penetrated thedecklamelle and outer wall layer of both basiphytes. By the time the epiphyte germlings reached the 15celled stage, they had penetrated the inner walllayer. The differences in epidermal cell wallconstruction between the two basiphytes may play arole in the ability of zooids of U. lactuca toattach in nature where epiphytization of G.cornea is infrequent.

The effect of environmental parameters on the growthof Porphyra linearis gametophytes was examinedunder controlled conditions, and related to themultilinear regression growth model recently developedfor this seaweed under coastal conditions in theeastern Mediterranean. Growth chambers, a gradienttable, special culture devices and analytical methodswere combined for this culture study.The major factors significantly controlling thegrowth rate of the P. linearis gametophytein glass dishes were: photoperiod, temperature, agein culture, photosynthetic photon flux (PPF), salinityand water dynamics. Maximal growth occurred underdaylength of 12 h, medium temperature (15–20 °C), low PPF (70–140 μmol photon m-2s-1), ambient salinity (30–40 ppt), 1–3 h ofdaily air exposure, and water velocity of 4 cm s-1.Photosynthesis and respiration rates weredominantly affected by daylength and temperature,while the concentration of pigments was dominantlyaffected by PPF and temperature.These conditions correspond well to the optimalnatural growth environment of this local species andare in agreement with the optimum estimated throughthe recently developed outdoor mathematical growthmodel.

Certain forms of oligocellulose and certainbacterially excreted peptides were identified asendogenous and exogenous elicitors, respectively, ofa tip bleaching response in Gracilaria conferta(Schousboe ex Montagne) J. et G. Feldmann. Thehalf-maximal tip bleaching response was observed when31.1 μM cellobiose or 11.6 μM cellotetraosewere present in the growth medium. In contrast, noresponse was detected after exposure to glucose,cellotriose, cellopentaose or maltooligosaccharides.The response was thus strongly dependent on themolecular size of the oligocellulose and onlysaccharides that consisted of an even number ofglucose residues were elicitor-active. Three bacterialspecies that had earlier been identified as potentialinducers of the tip bleaching symptom excretedelicitor-active compounds into the growth medium.These compounds were protease-sensitive and thuspeptides or proteins. The tip bleaching-inducingcompound that was excreted by one Cytophaga-likeorganism was partially purified. It could be extractedfrom culture supernatants with chloroform and itsmolecular size was between 700 and 1500 Da,corresponding with a structure of 4–20 amino acids.Various endogenous and exogenous elicitors are thusrecognized by G. conferta and allow this alga torespond hypersensitively to the maceration of its cellwall skeleton or just to the presence of certainepiphytic organisms.

Acclimation responses of the red alga Gracilaria tenuistipitata var. liui collected on the northwest coast of Philippines were determined in laboratory setups and outdoor cultivation tanks in Haifa, Israel. Growth under laboratory conditions was influenced by all three variables studied, namely, temperature (20 or 30 °C), salinity (20, 30 or39‰) and seawater pH (6.5, 7.0, 8.0 or ≥ 9.0). In 250 mL flasks lacking pH control growth was influenced by temperature only at 20 ‰, whereas at 39 ‰, growth rates were similar at 20 or 30 °C. In 500 mL cylinders in which pH was controlled, growth rates were significantly different at a pH of 6.5 and 7.0 for all salinities, with maximal rates occurring in 39 ‰. At pH 8.0, and above, growth rates between salinities were similar and reduced to approximately 50% at a pH of 9.0 compared to rates at a pH of 6.5. Photosynthesis responses generally resembled growth responses both, in 250 mL and 500 mL cultures. In 40-L outdoor tanks, weekly growth and agar yields were apparently enhanced by increasing light intensities (up to full sunlight) and nutrient concentrations (up to 0.2 mM PO3 2- and 2.0 mM NH4 +), and rates averaged four times higher than rates determined in the smaller flask cultures. This study shows broad salinity tolerance of G. tenuistipitata var. liui and its ability to sustain growth rates that are among the highest measured for Gracilaria spp. in outdoor cultures.

Photosynthetic (oxygen evolution) and growth (biomass increase) responses to ambient pH and inorganic carbon (Ci) supply were determined for Porphyralinearis grown in 0.5 L glass cylinders in the laboratory, or in 40 L fibreglass outdoor tanks with running seawater. While net photosynthetic rates were uniform at pH 6.0–8.0, dropping only at pH 8.7, growth rates were significantly affected by pH levels other than that of seawater (c. pH 8.3). In glass cylinders, weekly growth rates averaged 76% at external pH 8.0, 13% at pH 8.7 and 26% at pH 7.0. Photosynthetic O2 evolution on a daily basis(i.e. total O2 evolved during day time less total O2 consumed during night time) was similar to the growth responses at all experimental pH levels, apparently due to high dark respiration rates measured at acidic pH. Weekly growth rates averaged 53% in algae grown in fibreglass tanks aerated with regular air (360 mg L-1 CO2) and 28% in algae grown in tanks aerated with CO2-enriched air (750 mg L-1 CO2). The pH of the seawater medium in which P. linear is was grown increased slightly during the day and only rarely reached 9.0. The pH at the boundary layer of algae submerged in seawater increased in response to light reaching, about pH 8.9 within minutes, or remained unchanged for algae submerged in a CO2-free artificial sea water medium. Photosynthesis of P. linearissaturated at Ci concentrations of seawater (K0.5560 μM at pH 8.2) and showed low photosynthetic affinity for CO2(K0.5 61 μM) at pH 6.0. It is therefore concluded that P. linearisuses primarily CO2 with HCO3 - being an alternative source of Ci for photosynthesis. Its fast growth could be related to the enzyme carbonic anhydrase whose activity was detected intra- and extracellularly.