Larval settlement of the tropical abalone, Haliotis asinina Linnaeus, using natural and artificial chemical inducers

Many kinds of chemical and biological materials have been used as inducers of settlement of abalone larvae, as well as other species of marine gastropods, with responses being highly variable, even to the same chemical cue. The present study tested chemical inducers, γ-aminobutyric acid (GABA), δ-aminovaleric acid (5-AVA) and l -glutamic acid (GA) and the effects they have on larval settlement of Haliotis asinina . Additionally, a relatively inexpensive commercial substance, monosodium glutamate (MSG), was trialed. The datum provided shows all chemicals to be active inducers of settlement in this study, in order of effectiveness of 5-AVA, GABA, MSG to GA. Induction as adjudged from larval numbers settled was best at 6 h 62%, with 10⁻¹ mM 5-AVA. At 24 h, induction was the highest at 78% when exposed to 10⁻² mM 5-AVA. Larvae that were allowed to settle up to 72 h showed the highest numbers of settled larvae, and declined back to 60% when exposed to 10⁻² 5-AVA and 10⁻¹ mM GABA respectively. Monosodium glutamate, although third in settlement standings would bypass the other chemicals, with regard to cost versus yield. The assessment of settlement surface, rough or smooth proved to be irrelevant, which had no significant impact on larval settlement.     

Ingestion and digestion of 10 species of microalgae by winged pearl oyster Pteria sterna (Gould, 1851) larvae

Ten species of microalgae were tested for ingestion and digestion in Pteria sterna larvae using epifluorescence microscopy to choose an appropriate diet. An experiment was conducted using 2, 4, 5 (straight-hinge) 10 and 22 (umbo stage) day old larvae. Larvae were stocked in 150 ml flasks at 30 ml⁻¹ and fed 100,000 algal cells ml⁻¹ of each species individually. Larvae were fed for 1 h and then were observed under the microscope to detect ingestion; larvae were then sieved and placed in flasks containing filtered seawater and were observed after 1 and 2 h to analyse digestion for the microalgae ingested. Out of the 10 species administered, only Nannochloris sp., Pavlova lutheri and Isochrysis aff. galbana (T-ISO) were ingested, and only the last two species were digested. No ingestion of Phaeodactylum tricornutum, Chaetoceros muelleri, Ch. calcitrans, Thalassiosira weisflogii, Dunaliella salina, Tetraselmis tetrathele and T. suecica was evident at any stage of larval development tested. Only T-ISO and P. lutheri should be used for larval rearing of P. sterna until other species are identified as suitable (ingested and digested) for this species.     

Hemagglutinating activity and electrophoretic pattern of hemolymph serum proteins of Penaeus monodon and Macrobrachium rosenbergii to white spot syndrome virus injections

The giant fresh water prawn Macrobrachium rosenbergii is known to be highly tolerant to white spot syndrome virus (WSSV) infections when compared to the widely cultured marine tiger shrimp Penaeus monodon. At present, the exact mechanism of tolerance by M. rosenbergii to WSSV is not known. In this study, we attempt to study the effect of WSSV injections on the hemagglutination activity of the hemolymph serum of both P. monodon and M. rosenbergii and look for changes if any, on their hemolymph serum protein electrophoretic patterns. Our results show that M. rosenbergii had significantly (p < 0.05) higher hemagglutinating activity against mouse erythrocytes when compared to P. monodon. As the infection progressed to 48 h there was a further increase (p < 0.05) in the hemagglutination activity in M. rosenbergii, while it decreased in P. monodon. 12% SDS-PAGE analysis of the hemolymph serum of M. rosenbergii infected with WSSV did not show any new protein bands, whereas few bands with decreased intensity was observed in moribund P. monodon where the hemagglutinating activity was also observed to be decreased. The results indicate that hemolymph hemagglutinin levels are modulated in crustaceans as a response to viral infections.     

Effect of variation in the protein value of the red macroalga Palmaria palmata on the feeding, growth and gonad composition of the sea urchins Psammechinus miliaris and Paracentrotus lividus (Echinodermata)

The sea urchins Psammechinus miliaris and Paracentrotus lividus were fed three macroalgal diets with varying protein content over a 90-day period. These diets were the red alga Palmaria palmata, which had been grown in seawater enriched with two levels of ammonia-N and contained a protein content of either 41% (High-N) or 32% (Low-N) and the brown alga Laminaria saccharina (23% protein). A significantly different response to the macroalgal diets by the two sea urchin species was found in terms of food consumption rate, food conversion efficiency, gonadal growth and gonad nutritional composition. P. miliaris was able to efficiently utilise the High-N P. palmata, whereas P. lividus exhibited a higher food conversion efficiency (FCE) and specific growth rate (SGR) when fed the Low-N compared with the High-N P. palmata. P. miliaris had a significantly higher food consumption rate and higher gonad index (GI) compared with P. lividus, irrespective of diet type. The lowest FCE, SGR and GI were shown by both species fed L. saccharina. No interspecific difference in gonadal nitrogen content was observed, although a positive relationship was shown between dietary and gonadal nitrogen content for both species. All three macroalgal diets produced bright orange or yellow orange gonad colour in P. lividus. High-N and Low-N P. palmata improved gonad colouration in P. miliaris compared with L. saccharina. The results suggest that P. miliaris can efficiently assimilate high protein plant-based diet, whereas P. lividus is less capable of benefiting from high dietary protein levels and this should be taken into consideration when culturing this species either in land-based or sea-based polyculture systems.     

Dual disease resistance in a selectively bred eastern oyster, Crassostrea virginica, strain tested in Chesapeake Bay

Selective breeding efforts have yielded oyster strains, Crassostrea virginica, with improved survival and resistance against Haplosporidium nelsoni (MSX); however, because of susceptibility to the oyster pathogen Perkinsus marinus (Dermo), their utility has been limited in areas where the two parasites co-occur. Dual resistance to H. nelsoni and P. marinus was achieved through four generations of artificial selection of wild Delaware Bay oyster progeny at a site in the lower York River, Virginia, USA where both diseases are enzootic. During 1993–1995, survival, growth, and disease susceptibility of third generation Delaware Bay (F₃-DEBY) oysters were evaluated at the York River site in comparison to that of similarly selected third generation James River, Virginia oysters (F₃-JR), and first generation Louisiana oysters (F₁-LA), whose parents were naturally selected in the wild for resistance to P. marinus. During 1997–1999, the performance of F₄-DEBY was evaluated at three sites in Virginia in comparison to two groups of first generation oysters whose parents originated from Mobjack Bay and Tangier Sound, Virginia where both P. marinus and H. nelsoni are enzootic. In the presence of high infection pressure from both H. nelsoni and P. marinus, the F₃-DEBY stock showed significantly higher survival and growth than either the F₃-JR or F₁-LA strain. After 15 months of deployment, 79% of F₃-DEBY, 11% of F₃-JR and 17% of F₁-LA oysters were market size (≥76.2 mm) and cumulative mortality was only 16% in F₃-DEBY as compared to 42% in F₃-JR and F₁-LA. At the termination of the study, F₃-DEBY oysters exhibited 22% lower mortality than the F₁-LA stock, which outperformed the F₃-JR stock. Relative performance in respect to disease varied considerably with sample date; however, average H. nelsoni weighted prevalence varied such that F₃-DEBY1-LA=F₃-JR and average P. marinus weighted prevalence varied such that F₁-LA3-DEBY3-JR. In the 1997–1999 trial, F₄-DEBY oysters experienced 34–61% lower mortality, greater growth rate, and consistently lower prevalence and intensity of P. marinus than either Mobjack Bay or Tangier Sound oysters. H. nelsoni prevalences were very low (<12%) in all three stocks. This is the first study to demonstrate that reduced susceptibility to both P. marinus and H. nelsoni can be achieved through selective breeding.     

The effect of diet type and quantity on the development of common sea urchin larvae Echinus esculentus

Several species of sea urchins are now being cultivated for commercial purposes and with the continued increased demand for sea urchin gonads as a food product, new species are being assessed for their aquaculture and market potential. This study focussed on establishing protocols for the production of common sea urchin Echinus esculentus larvae and juveniles to assess its potential as an echinoculture species. Two trials were carried out, the first trial evaluated the influence of three microalgal diets (D=Dunaliella tertiolecta only, mixed D/P=D. tertiolecta plus Phaeodactylum tricornutum and P=P. tricornutum only) on larval morphology. Larval length, width, post-oral arm length and rudiment length were significantly effected by diet. Diets D and D/P prompted more rapid metamorphosis. In the second trial, the effects of different rations of D. tertiolecta were tested. The food ration, standard ration (SR; 1000, 3000, and 5000 cells ml⁻¹) and high ration (HR; 3000, 9000, and 15,000 cells ml⁻¹) were increased as the larvae acquired the 3rd and 4th pair of larval arms. Larvae fed the SR were significantly larger (longer and wider) and had significantly longer rudiments than those in the HR treatment. The number of larvae metamorphosing and settling onto substrates was significantly higher in treatment SR compared to HR. Optimising the larval diet shortened the larval stage from 21–23 days in the first trial to 16 days in the second trial. The maximum percentage of metamorphosing individuals which survived to post-larvae or juveniles (10 days after they were first judged competent to settle) was 46.6%, suggesting E. esculentus is a viable aquaculture candidate.     

Biofilm development within a larval rearing tank of the tropical rock lobster, Panulirus ornatus

The role of bacterial biofilms in disease processes is becoming increasingly recognised in both clinical and environmental settings. Biofilm development within a rearing tank of the tropical rock lobster Panulirus ornatus was studied to evaluate if the biofilm is a reservoir for potentially pathogenic bacteria that cause mass larval mortalities. Within a 5000 L larval rearing tank, fiberglass microscope slides were systematically distributed during a standard rearing attempt to assess biofilm development. Culture-based counts for two media types, TCBS and Marine Agar (MA), demonstrated increased bacterial densities until days 11 and 13 respectively. For both media types, a drop in the plate counts was followed by a subsequent increase towards the end of the experiment. Scanning electron microscopy (SEM) confirmed that cell densities decreased between days 13 and 17, most likely due to sloughing of the biofilm into the water column. SEM images revealed distinct changes in dominant morphologies reflecting a succession of bacterial populations. A dynamic succession of microbial species during biofilm development was also demonstrated using denaturing gradient gel electrophoresis (DGGE) profiling of bacterial 16S rRNA genes in combination with statistical ordination analysis. Prominent changes in the DGGE profiles coincided with the decrease in bacterial numbers observed by SEM and plating on MA between days 13 and 17. Fluorescence in situ hybridization (FISH) identified -Proteobacteria as being numerically abundant in the biofilm. This was supported by results from DGGE analysis, which retrieved only sequences affiliated with - and γ-Proteobacteria. DGGE bands affiliated with Vibrio became dominant towards the end of the larval run (days 21 to 24). A Vibrio harveyi strain isolated from the biofilm late in the larval rearing trial (day 24) demonstrated increased larval mortality in small scale phyllosoma survival studies. The detection of Vibrionaceae at the end of the larval trial coincided with mass phyllosoma mortality and show that the biofilm is a reservoir for potentially pathogenic bacteria.     

Evidences of the intertidal red alga Grateloupia turuturu in turning Vibrio parahaemolyticus into non-culturable state in the presence of light

Grateloupia turuturu, previously known as Grateloupia doryphora, has been widely reported to be an invasive algal species. There are no studies to relate the impact of its existence on its surrounding environment. In this paper, we present our results to show that about 70% of individuals collected from the field could turn Vibrio parahaemolyticus into non-culturable state on both selective (TCBS) and non-selective (2216E) culture medium in 24 h in the presence of light in live algal culture. Total bacteria counts on TCBS and 2216E plates dropped from the initial 565 (± 174) and 1192 (± 60) cfu ml^− 1 respectively to zero in 24 h. This effect disappeared when the alga was grown in darkness. The same effect was not found in two other intertidal macroalgae Laminaria japonica and Palmaria palmata. Further tests showed that the settlement ability of bacteria in seawater was impaired significantly in the presence of this alga in comparison with three other algal species.     

In vitro antimicrobial susceptibility of Photobacterium damselae subsp. piscicida to 15 different antimicrobial agents

The in vitro susceptibilities of 145 Photobacterium damselae subsp. piscicida strains, isolated from different geographical regions, against 15 major fish farming antimicrobial agents were assessed using a disc diffusion method. Ninety-nine percent of all isolates were sensitive to florfenicol and trimethoprime-sulfamethoxazole; 93% of the strains were resistant to erythromycin. Based on the antimicrobial susceptibility, the P. damselae subp. piscicida isolates could be distinguished from each other. While all American and European isolates were sensitive to kanamycin, 49% of the Japanese isolates were resistant to it. Moreover the results clearly indicated increasing resistance to a variety of antibiotics, particular in Japan, but also in some Mediterranean countries.     

Postlarvae and juveniles of a selected line of Penaeus stylirostris are resistant to infectious hypodermal and hematopoietic necrosis virus infection

A susceptibility study of postlarvae (PL) and juvenile Super Shrimp®, a selected line of Penaeus stylirostris, was conducted to compare their resistance to infectious hypodermal and hematopoietic necrosis virus (IHHNV) infection to that of a specific pathogen free (SPF) population of P. vannamei. Super Shrimp® PLs were fed with IHHNV-infected shrimp tissue for 2 days and then maintained on a pelletized ration for an additional 28 days. PLs were sampled at days 0, 1, 2, 3, 4, 6, 10, 15, 20, 25 and 30. There was no apparent mortality during the experimental period. Tissue DNA extracted from the PLs was analyzed for the presence of IHHNV by PCR. Low levels of IHHNV were detected only in DNA extracts from samples at days 1, 2, and 3. No IHHNV DNA was detected from days 4 to 30. The days that the PLs were weakly IHHNV-PCR positive were during the period that they were being fed with IHHNV-tissue, and thus, the IHHNV DNA signal was suspected to be from the infected tissue used as a feed. Through both histology and in situ hybridization, we confirmed that tissues of Super Shrimp® PLs were not infected with IHHNV. PCR results of another IHHNV challenge study with juveniles of Super Shrimp® were similar to those with PLs. These results indicate that IHHNV did not replicate in the PL and juvenile Super Shrimp®. In contrast, P. vannamei juveniles, which were used as a positive control, showed a more intense IHHNV infection, as determined by PCR detection, beginning at 6 days postchallenge and increasing throughout the remainder of the study. In addition, the IHHNV-infected P. vannamei at 30 days postchallenge showed histological changes characteristic of IHHNV infection and had a positive reaction for IHHNV with in situ hybridization. Our studies show that Super Shrimp® are resistant to IHHNV infection. This is the first unequivocal demonstration of resistance to viral infection in shrimp.     

Settlement behavior of American lobster (Homarus americanus): effect of female origin and developmental temperature

The benthic settlement success of American lobster (Homarus americanus) postlarvae is a key factor in population dynamics, with the temperature being a key factor. Given the spatial distribution of the species, its pelagic larvae probably encounter various thermal zones during development. Little information is available on the ability of lobster larvae to acclimate to temperature or if this ability varies among geographic areas. The main objectives of this study were to characterize (i) the settlement behavior of the postlarvae in relation to (a) temperature during larval development and (b) the geographic origin of the maternal parent and (ii) the behavioral response of postlarvae with different thermal histories in the presence/absence of a thermocline. Results did not support the hypothesis of thermal adaptation among local populations, as settlement behavior of postlarvae did not differ among females from different geographic regions. In contrast, larval survival and settlement behavior and postlarval energetic conditions varied with development temperature. Behavioral plasticity in the benthic settlement was observed in relation to temperature structure of the experimental water column. This study suggests that there may be links between development temperature and the natural variability in survival and settlement of American lobsters. Findings emphasize the need for an integrated approach in lobster recruitment studies with the inclusion of behavior in dispersion models.     

Two microalgae Crypthecodinium cohnii and Phaeodactylum tricornutum as alternative source of essential fatty acids in starter feeds for seabream (Sparus aurata)

Despite oils extracted from algae and other microorganisms that may constitute excellent sources of HUFAs, few studies have determined the nutritional value of different microalgal species for young marine fish. Six thousand gilthead seabream (Sparus aurata) postlarvae (73 mg body weight) were fed for 57 days diets containing either fish oil as a single lipid source or 2 and 4% of Cryptecodinum cohnii or 5% Phaeodactylum tricornutum. Fish oil substitution by C. cohnii resulted in improved fish survival and a very good growth performance, in agreement with a higher proportion of DHA in diets and in total lipids of fish. Incorporation of DHA and other fatty acids was proportional to their contents in diet suggesting the good nutritional utilization of homogenized C. cohnii. Lower survival rates were found in fish fed P. tricornutum and could be related to an epithelial degeneration observed in the anterior intestine. This degeneration could be related to a higher lipid content in these fish or to the strong hornlike cornutate processes found in the valves of the diatom P. tricornutum.     

Effects of chloramphenicol, erythromycin, and furazolidone on growth of Isochrysis galbana and Chaetoceros gracilis

This study focused on determining the effects of antibiotics on microalgae used as food for scallop larvae. Six different dose levels of chloramphenicol, erythromycin, and furazolidone were added to cultures of Isochrysis galbana and Chaetoceros gracilis. An in vivo experiment was subsequently conducted to determine the effect of chloramphenicol and erythromycin on larval survival of the Pacific calico scallop Argopecten ventricosus in tanks and on the population of its associated bacteria. Results showed that growth of I. galbana was not significantly affected by chloramphenicol or erythromycin at the test doses of 0.5, 1.0, 3.0, 6.0, 9.0, and 12.0 mg/l. C. gracilis was significantly sensitive to erythromycin and chloramphenicol at doses higher than 0.5 and 3.0 mg/l, respectively. Furazolidone inhibited the growth of both I. galbana and C. gracilis at all test doses. Results showed that exposure of scallop larvae to a dose of 6 mg/l chloramphenicol or erythromycin did not significantly affect growth of I. galbana, significantly enhanced survival of the scallop larvae, and inhibited the growth of Vibrio spp. in tanks. This study demonstrated the adverse effect of chloramphenicol, erythromycin and furazolidone on I. galbana and C. gracilis microalgae but the positive effect on survival of the scallop larvae, decreasing associated bacterial population.     

A biophysical model to assess the trade‐off between larval recruitment and catch in southern Australia's largest prawn fishery

Data from stock assessment surveys, published research and climate sensors were linked to model the interaction between fishing, physical‐oceanographic processes and spatial patterns of larval settlement for western king prawn [Penaeus (Melicertus) latisulcatus]. This information was used to evaluate the trade‐off between larval recruitment and catch during fishing periods that demand high prices but coincide with spawning. Total rates of larval settlement were maximized when tidal currents and atmospheric physical‐forcing components were coupled with simulations of larval swimming behaviour under average gulf temperatures. Average gulf temperatures sustained longer larval durations and increased larval settlement rates by over 12% compared with warmer gulf conditions simulated under a scenario of global warming. Reproductive data coupled with outputs from the biophysical model identified consistent inter‐annual patterns in the areas contributing to larval settlement success. Areas located in the north‐east, and central‐west of the fishery, consistently contributed to over 40% of all larvae reaching a settlement in each year. Harvest sensitivity analyses indicated that changes in the spatial patterns of pre‐Christmas fishing could lead to improvements in overall rates of the larval settlement while maintaining or improving the levels of catch. Future studies to refine the model inputs relating to physical processes, larval behaviour and mortality rates for P. latisulcatus coupled with surveys of juvenile prawn abundance to ground truth the modelled predictions, would allow stock recruitment relationships to be more closely examined and inform adaptive management of the fishery in the future.     

Vibrio harveyi and other bacterial pathogens in cultured summer flounder, Paralichthys dentatus

A monitoring program for Vibrio harveyi and other potential bacterial pathogens of summer flounder was conducted at two facilities in the Northeast United States. Bacterial samples were collected from larval and juvenile fish and live-feed, and identified using API 20E biochemical profiles and 16S rDNA sequencing. Histopathological examinations were conducted in order to relate histological changes with the presence of potential bacterial pathogens. V. harveyi, Vibrio ichthyoenteri, and Photobacterium damselae subsp. damselae, three known pathogens of flatfish, were isolated from diseased summer flounder. Although a high proportion of juvenile summer flounder showed microscopic signs of disease, the presence of these potential bacterial pathogens in fish was not associated with large-scale mortalities. An outbreak of flounder infectious necrotizing enteritis with 30% cumulative mortality occurred when juveniles were transported to a new facility. Isolates of V. harveyi from the disease outbreak were genetically different from the isolates from the commercial hatchery. V. harveyi isolates from both facilities were pathogenic to summer flounder by intraperitoneal injection. P. damselae subsp. damselae, V. ichthyoenteri, and Vibrio scophthalmi, also found in fish with gross lesions, were not pathogenic to juvenile summer flounder by intraperitoneal injection. Our research shows that several potential bacterial pathogens are associated with morbidity and mortality in summer flounder larvae and juveniles, especially in situations of stress. Increased knowledge about the environmental conditions that lead to disease, as well as the interactions of V. harveyi with other microbial species, could lead to the development of management strategies in summer flounder farms.     

单糖组合对藤壶金星幼虫附着的影响研究

藤壶Amphibalanus(=Balanus)amphitrite是典型的海洋污损生物,藤壶金星幼虫受外源信号的指引,寻找到合适的附着地点才能进行附着和变态。针对前期发现的有一定诱导活性的4种单糖Man、Xyl、Glc和Fuc,该研究利用正交试验研究其对藤壶金星幼虫附着的影响。结果发现,单糖组合对藤壶金星幼虫附着具有显著性影响,Man(5.00×10^-8 mol/L)、Glc(1.25×10^-8 mol/L)、Xyl(1.25×10^-8 mol/L)和Fuc(1.90×10^-7 mol/L)为最优组合。组合中4种单糖的影响力由高到低依次是Man>Fuc>Glc>Xyl,且Man、Fuc和Glc的影响具有显著性,Xyl对幼虫附着影响不大。添加单糖除了影响金星幼虫的附着率以外,更重要的是能显著缩短幼虫附着的时间进程。     

脂多糖对细菌生物被膜形成及厚壳贻贝幼虫变态的影响

为探究脂多糖（LPS）对海洋细菌生物被膜形成、海洋贝类幼虫变态所产生的作用,深入了解影响厚壳贻贝附着变态的因素,本实验使用不同浓度的LPS直接刺激厚壳贻贝幼虫,观察其对幼虫附着变态的直接作用;同时在海假交替单胞菌形成生物被膜的过程中添加不同浓度的LPS,分析生物被膜生物学特性的变化,及变化后的生物被膜对厚壳贻贝幼虫附着变态的影响。结果显示,3种浓度的LPS均可直接诱导厚壳贻贝幼虫的变态;10.0mg/L浓度LPS处理后的生物被膜其细菌密度、膜厚度明显降低,且膜厚度降低了12.1%,胞外产物中多糖、脂类显著增加,其中代表性多糖可拉酸的含量增加了35.4%,同时其对幼虫变态的诱导作用也提高了53.3%。因而,在厚壳贻贝幼虫的附着变态过程中,LPS具有直接诱导作用,同时还可以通过调控生物被膜胞外物质,特别是可拉酸的生成,间接影响厚壳贻贝幼虫的附着变态。本研究成果对海洋细菌互作关系研究、厚壳贻贝养殖产业的改善以及海洋防污技术研发具有重要推动价值。     

Spawning and Larval Development of the Four‐Sided Sea Cucumber,Isostichopus badionotus (Selenka 1867), under Controlled Conditions

The objective of this study was to generate information for the development of Isostichopus badionotus culture protocols by determining optimal spawning methods, egg production, length and duration of larval stages, and larval habitat settlement preferences. Studies performed during the spawning seasons of 2010 and 2011 showed that this organism spawns between July and November without mechanical or chemical induction. The larval development has five well‐defined stages: early, mid and late auricularia, doliolaria, and pentactula. Juveniles (654.3 µm length) were obtained on average 25 days after fertilization (DAF), when larvae were incubated at 25 ± 1 C. No preference was observed for any of the materials used for settlement of the larvae. The easy adaptation of I. badionotus to the culture conditions and the competence of the larvae to complete metamorphosis while feeding solely on microalgae concentrates make this species an excellent candidate for farming. However, it is necessary to continue designing techniques for larval culture and to establish appropriate captivity conditions that allow multiannual reproduction.     

Preliminary analysis of recombinant β-tubulin of Pseudocohnilembus persalinus (Ciliophora: Scuticociliatida) as a vaccine antigen candidate against scuticociliatosis

The recombinant β-tubulin of Pseudocohnilembus persalinus was investigated to test whether tubulin could be a suitable antigenic target against scuticociliatosis. The cDNA of P. persalinus β-tubulin has an open reading frame of 1332 bp and had 15 TAA triplets code for glutamine (Q). All 15 TAA/Q triplets of the β-tubulin cDNA were mutated to universal CAA/Q triplets by site-directed mutagenesis and the mutated β-tubulin was expressed in Escherichia coli as a fusion with glutathione S-transferase (GST-BTU). In Western blot analysis, rat sera immunized with recombinant GST-BTU reacted with reduced β-tubulin of P. persalinus. The immunized rat sera showed higher parasiticidal activity than those of control. Ciliates preincubated with heat-inactivated immune sera showed significantly lower proliferation than ciliates preincubated with control sera when exposed to naïve rat sera as a complement source. The present results suggest that the recombinant β-tubulin of scuticociliates may be used as a target antigen for development of vaccine against scuticociliatosis.     

Effects of secondary crops on bacterial growth and nitrogen removal in shrimp farm effluent treatment systems

Secondary crops provide a means of assimilating some effluent nitrogen from eutrophic shrimp farm settlement ponds. However, a more important role may be their stimulation of beneficial bacterial nitrogen removal processes. In this study, bacterial biomass, growth and nitrogen removal capacity were quantified in shrimp farm effluent treatment systems containing vertical artificial substrates and either the banana shrimp Penaeus merguiensis (de Man) or the grey mullet, Mugil cephalus L. Banana shrimp were found to actively graze biofilm on the artificial substrates and significantly reduced bacterial biomass relative to a control (24.5±5.6 mg C m⁻² and 39.2±8.7 mg C m⁻², respectively). Bacterial volumetric growth rates, however, were significantly increased in the presence of the shrimp relative to the control (45.2±11.3 mg C m⁻² per day and 22.0±4.3 mg C m⁻² per day, respectively). Specific growth rate, or growth rate per cell, of bacteria was therefore appreciably stimulated by the banana shrimp. Nitrate assimilation was found to be significantly higher on grazed substrate biofilm relative to the control (223±54 mg N m⁻² per day and 126±36 mg N m⁻² per day, respectively), suggesting that increased bacterial growth rate does relate to enhanced nitrogen uptake. Regulated banana shrimp feeding activity therefore can increase the rate of new bacterial biomass production and also the capacity for bacterial effluent nitrogen assimilation. Mullet had a negligible influence on the biofilm associated with the artificial substrate but reduced sediment bacterial biomass (224±92 mg C m⁻²) relative to undisturbed sediment (650±254 mg C m⁻²). Net, or volumetric bacterial growth in the sediment was similar in treatments with and without mullet, suggesting that the growth rate per cell of bacteria in grazed sediments was enhanced. Similar rates of dissolved nitrogen mineralisation were found in sediments with and without mullet but nitrification was reduced. Presence of mullet increased water column suspended solids concentrations, water column bacterial growth and dissolved nutrient uptake. This study has shown that secondary crops, particularly banana shrimp, can play a stimulatory role in the bacterial processing of effluent nitrogen in eutrophic shrimp effluent treatment systems.