Marine Rotifer Brachionus Research Articles

Cross-reactivities of mammalian MAPKs antibodies in rotifer and copepod: Application in mechanistic studies in aquatic ecotoxicology

The mitogen-activated protein kinases (MAPKs) family is known to mediate various biological processes in response to diverse environmental pollutants. Although MAPKs are well characterized and studied in vertebrates, in invertebrates the cross-reactivities of MAPKs antibodies were not clearly known in response to environmental pollutants due to limited information of antibody epitopes with material resources for invertebrates. In this paper, we performed phylogenetic analysis of MAPKs genes in the marine rotifer Brachionus koreanus and the copepods Paracyclopina nana and Tigriopus japonicus. Also in rotifer and copepods, several studies of Western blot of MAPK signaling pathways were shown in response to environmental pollutants, including multi-walled carbon nanotubes (MWCNTs), water-accommodated fractions (WAFs) of crude oil, and microplastics. This paper will provide a better understanding of the underlying mechanistic scenario in terms of cross-reactivities of mammalian antibodies in rotifer and copepod.

The prevention of saltwater algal pond contamination using the electron transport chain disruptor, rotenone

Rotenone is a classic inhibitor of Complex I of the electron transport chain which is common to all life. Algae are known to be rotenone-insensitive because in addition to Complex I they have alternative pathways for the oxidation of NADH2. It is this differential sensitivity to rotenone that can be exploited to reduce many types of predators while not affecting biodiesel-producing algae. A universal tool like rotenone is needed to reduce predation in open ponds as predation is well known to limit cultivation to a few weeks in the warmer months. In this study, we tested the effect of rotenone on the mortality and reproduction of two marine algae strains, Nannochloropsis oculata and Tetraselmis suecica, and two marine rotifer strains, Brachionus manjavacas and Brachionus rotundiformis. The LC50s for the two rotifers ranged from 0.18 to 0.35μM while N. oculata was unaffected to at least 7.6μM. Rotifer reproduction was reduced at rotenone concentrations lower than those that caused mortality. Rotenone applications to produce 1 million gallons of biodiesel in one year are estimated to cost $1684. This provides algae farmers an important and inexpensive tool to manage grazing predators in algae mass cultures and avoid pond crashes.

해수산 rotifer Brachionus rotundiformis의 유성생식유도에 관한 염분 및 기사용 배양수의 효과

This study examined the effects of salinity and medium used on the induction of sexual reproduction for the mass production of the resting eggs in the marine rotifer Brachionus rotundiformis for 9 days. After exposure to various salinities, the mixis rate in all treatments began to increase on the third day and decreased after the 4th or 5th day. The resting eggs appeared on the 4th day at 10‰, 15‰, 20‰, 25‰. In particular, the fertilization rates with 33.3% and 31.3% at 20‰ and 25‰ on day 6 were higher than those of the others, respectively. Mean mixis rate for 9 day at 0‰ and 20‰ were the highest with 25.3% and 20.2%, respectively, and the fertilization rate (14.0%) at 25‰ was the highest. In the medium experiment used, the mixis rate in all treatments were similar to the results of the salinity experiment. The resting eggs were only found at 0% and 25% of the used medium treatments. While there were no significant differences in the mean fertilization rate of all treatments, the mean mixis rate (22.3%) in the 25% treatment was higher than that of the control (0% treatment). As a result, salt stress was more effective than using the used medium to induce sexual reproduction in B. rotundiformis. The optimal salinity was considered to be 20-25‰, which caused a high mixis and fertilization rate in the rotifer. These results provide basic data for the mass production of rotifer resting eggs.

Molecular and expression analysis of manganese superoxide dismutase (Mn–SOD) gene under temperature and starvation stress in rotifer Brachionus calyciflorus

Superoxide dismutase (SOD) is an important antioxidant enzyme that protects organs from damage by reactive oxygen species. We cloned cDNA encoding SOD activated with manganese (Mn-SOD) from the rotifer Brachionus calyciflorus Pallas. The full-length cDNA of Mn-SOD was 1,016 bp and had a 669 bp open reading frame encoding 222 amino acids. The deduced amino acid sequence of B. calyciflorus Mn-SOD showed 89.1, 71.3, and 62.1 % similarity with the Mn-SOD of the marine rotifer Brachionus plicatilis, the nematode Caenorhabditis elegans, and the fruit fly Drosophila melanogaster, respectively. The phylogenetic tree constructed based on the amino acid sequences of Mn-SODs from B. calyciflorus and other organisms revealed that this rotifer is closely related to nematodes. Analysis of the mRNA expression of Mn-SOD under different conditions revealed that expression was enhanced 5.6-fold (p < 0.001) at 30 °C after 2 h, however, low temperature (15 °C) promoted Mn SOD temporarily (2.5-fold, p < 0.001) and then decreased to normal level (p > 0.05). Moderate starvation promoted Mn-SOD mRNA expression (p 12 < 0.01, p 36 < 0.05), which reached a maximum value (15.3 times higher than control, p 24 < 0.01) at 24 h. SOD and CAT activities also elevated at the 12 h-starved group. These results indicate that induction of Mn-SOD expression by stressors likely plays an important role in aging of B. calyciflorus.

Synergistic toxicity of Macondo crude oil and dispersant Corexit 9500A® to the Brachionus plicatilis species complex (Rotifera)

Using the marine rotifer Brachionus plicatilis acute toxicity tests, we estimated the toxicity of Corexit 9500A®, propylene glycol, and Macondo oil. Ratios of 1:10, 1:50 and 1:130 for Corexit 9500A®:Macondo oil mixture represent: maximum exposure concentrations, recommended ratios for deploying Corexit (1:10–1:50), 1:130 the actual dispersant:oil ratio used in the Deep Water Horizon spill. Corexit 9500A® and oil are similar in their toxicity. However, when Corexit 9500A® and oil are mixed, toxicity to B. manjavacas increases up to 52-fold. Extrapolating these results to the oil released by the Macondo well, suggests underestimation of increased toxicity from Corexit application. We found small differences in sensitivity among species of the B. plicatilis species complex, likely reflecting phylogenetic similarity. Just 2.6% of the water-accommodated fraction of oil inhibited rotifer cyst hatching by 50%, an ecologically significant result because rotifer cyst in sediments are critical resources for the recolonization of populations each Spring.

Rotifer ( Brachionus “Cayman”) culture performance improvement through l-carnitine addition is not related to fatty acid metabolism

The purpose of this paper was to investigate the nature of the effect of l-carnitine on the cultures of the marine rotifer Brachionus “Cayman” and its microflora. Xenic and gnotobiotic experiments were set up with rotifers, fed axenic wild-type yeast ( Saccharomyces cerevisiae) or xenic microalgae ( Tetraselmis suecica) and incubated with l-carnitine concentrations of 0.1, 1, 10, 60, 100 and 1000 mg L −1. Axenic neonates were obtained by separating amictic eggs from the rotifers by blending, prior to disinfection with glutaraldehyde and hatching. The gnotobiotic cultures had a significantly lower growth rate and egg ratio (%), compared to both xenic trials. The xenic cultures fed algae performed significantly better than those fed baker's yeast, when comparing population density, growth rate and egg ratio. A significant effect of l-carnitine addition was only found in yeast-fed cultures. Microbial assays, conducted with similar doses of autoclaved l-carnitine, revealed that several bacterial species present in the community of conventional Brachionus “Cayman” cultures, could possibly utilize l-carnitine for growth. These results suggest that the improvement of population density, growth rate and egg ratio in xenic rotifer cultures supplemented with l-carnitine, is most likely due to the stimulation by this compound of certain species of (beneficial) microorganisms, which are in turn valorized by the rotifers, yielding improved culture performance. Because of the absence of a positive effect under gnotobiotic conditions, a direct effect of l-carnitine on fatty acid metabolism, as suggested in literature, is unlikely.

Fatty acid as markers to demonstrating trophic relationships among diatoms, rotifers and green-lipped mussels

Green-lipped mussels Perna viridis, collected from Peng Chau, Hong Kong were allotted into two treatment groups, each containing three experimental tanks. The first treatment group comprised of mussels fed with the diatom Thalassiosira pseudonana only, whereas the second treatment group contained mussels fed with the marine rotifer Brachionus plicatilis, which was in turn fed with diatom T. pseudonana. The mussels were fed two times each day over the experimental period of 14 days. On Days 4, 7 and 14, three mussels were collected from each tank of each treatment group and treated as a single replicate. Fatty acid profiles of diatoms, marine rotifers and the three organs (digestive gland, mantle margin and adductor muscle) of the two mussel groups were analyzed. Results showed that monosaturated fatty acid (MUFA) 16:1n7 was conserved along the food chain among diatoms, marine rotifers and green-lipped mussels. This suggested that 16:1n7 or the ratio of 16:1n7 to saturated fatty acid (SFA) 16:0 can be a trophic marker for diatom T. pseudonana and elevated amounts of 16:1n7 in mussels can reflect the dominance of diatoms in its diet. The present results also showed that rotifers could accumulate MUFA 18:1n7 and PUFA 20:4n6 which were transferred up to mussels, especially 18:1n7, as zooplankton have the ability to synthesize or actively accumulate certain fatty acids that they need for growth or reproduction. There was an increase in the amount of 18:1n7 in the digestive gland of mussels fed with rotifers but the level of this fatty acid remained unchanged in those fed with diatoms, further confirming that 18:1n7 can be used as a marker for the presence of rotifers in trophic relationship studies. The relatively faster responses in the digestive gland of mussels to acquire the fatty acid signatures from their food suggested that the fatty acid profiles in the digestive gland can be a good marker to show a short-term fluctuation of food conditions in the marine environment.

Preliminary Trials on the Effect of Lighting for the Population Growth of the Rotifer, Brachionus plicatilis

The effect of lighting to rotifer culture water was investigated preliminarily using freshwater Chlorella as food in terms of vitamin B 12 (VB 12 ) production. The marine rotifer Brachionus plicatilis was batch-cultured for five days at 25°C in 25 psu seawater with or without lighting (L:D = 13:11 and L:D= 0:24). Rotifers were fed VB 12 -enriched or VB 12 -flee Chlorella vulgaris with or without cobalt compound supplementation (cobalt (II) sulfate heptahydrate: CoSO 4 ·7H 2 O). When VB 12 -enriched Chlorella was fed, rotifer with lighting showed better population growth than in complete dark. On the other hand, no difference was observed in the population growth between light and dark groups when VB 12 -free Chlorella was used. Nevertheless if the cobalt compound was supplemented to VB 12 -free Chlorella, the light group showed higher population growth than non-supplemented groups and dark groups, and a much higher amount of VB 12 was detected from the tanks than those from other groups. In addition to that, even in the group fed VB 12 -free Chlorella without Co supplementation a daily increase of a small amount of VB 12 was observed when they were lit up. From these experimental results, we can conclude that lighting plays an important role for the population growth of rotifers and Co compound supplementation to rotifer culture water promotes the reproduction performance of rotifers due to the enhanced production of VB 12 by lighting.

해양생태독성평가를 위한 표준시험생물로서의 식물플랑크톤에 관한 연구

A series of experiments was conducted to identify the potential of five phytoplankton species as standard test species for marine ecotoxicological tests. The candidate phytoplankton species are Skeletonema costatum, Heterosigma akashiwo, Prorocentrum micans, Isochrysis galbana, and Tetraselmis suecica. Salinity tolerance and sensitivity on potassium dichromate as a reference material were identified. Toxicity of eleven ocean dumped sewage sludges and four red tide expellent extracts were estimated by the inhibition of population growth rates (PGR) of marine diatom S. costatum, While most species revealed relatively weak tolerance on salinity, T. suecica demonstrated the highest salinity tolerance ranged from <TEX>$5\~35$</TEX> psu and the others <TEX>$15\~35$</TEX> psu. H. akashiwo revealed the highest sensitivity as 72h <TEX>$IC_{50}$</TEX>=0.76mg/L and T. suecica the lowest as 72h <TEX>$IC_{50}$</TEX>=8.89mg/L on potassium dichromate. Sludge extracts from industrial waste, domestic sewage and livestock farm waste sludge showed high toxicity as 72h <TEX>$IC_{50}$</TEX><<TEX>$2\%$</TEX> and lowest toxicity from filtration bed sludge as 72h <TEX>$IC_{50}$</TEX>=<TEX>$30.50\%$</TEX> NOEC (No Observed Effective Concentration) of sludge extract ranged from <<TEX>$0.4\%$</TEX> to <TEX>$1.6\%$</TEX> and this indicated high phytotoxicity of ocean dumped sewage sludge. The test sensitivity of phytoplankton PGR inhibition was much higher than those of marine rotifer Brachionus plicatilis mortality test and bioluminescent inhibition test by marine bacteria Vibrio fischeri, and comparable with the sea urchin (Strongylocentrotus intermedius) fertilization test. As a result the phytotoxicity test using phytoplankton PGR inhibition (<TEX>$IC_{50}$</TEX>) must be a useful tool for marine phyto-toxicological evaluation of ocean dumped materials.

Low-temperature preservation (at 4 oC) of marine rotifer Brachionus

Low-temperature preservation (at 4 oC) of marine rotifer Brachionus

Relative Growth Rates of the Capitulum and Its Plates in Octolasmis mülleri (Cirripedia: Thoracica: Poecilasmatidae)

ABSTRACT Fifty-two recently metamorphosed, juvenile, and young adult pedunculate barnacles, Octolasmis mulleri, were grown individually under laboratory conditions at 25-27°C, fed the marine rotifer Brachionus sp. and the brine shrimp Artemia salina. Growth of the animals was monitored over 10 weeks by periodic measurements of the capitular length, the area of the capitulum, and the areas of the capitular plates. The mean plate areas, individually and collectively, increase with growth, whereas the average plate coverage of the capitulum does not change.

Effect of disinfectants on the hatching of marine rotifer resting eggs Brachionus plicatilis Muller

Scanning electron microscopy indicated that stored resting eggs of the marine rotifer Brachionus plicatilis Muller were colonized by numerous bacterial cells that affect the hatchability of resting eggs. The effectiveness of chemical disinfectants on the hatching of the resting eggs was studied using selected chemicals and antibacterial drags. The hatching success rate of the fresh and stored eggs was 26 ± 5.8% and 0%, respectively. Sodium hypochlorite (1.0 p.p.m.) increased the hatching rate to 48 ± 1.3% in freshly-collected and 28 ± 1.8% in stored eggs. Physical disintegration of the bacterial mat occurred after 1 h of treatment with sodium hypochlorite. Among antibacterial drugs, sodium nifurstyrenate (Erubaju) enhanced the hatching success rate to 40 ± 1.2% and 20 ± 1.1% for the freshly-collected eggs and stored resting eggs, respectively. However, tetracycline and oxytetracycline did not significantly influence the hatching of the stored eggs. These observations are discussed in relation to the development of an improved method for the processing and hatching of rotifer resting eggs.

Effect of some vertebrate and invertebrate hormones on the population growth, mictic female production, and body size of the marine rotifer Brachionus plicatilis Müller

Eight vertebrate and invertebrate hormones were screened for their effect on population growth, mictic female production, and body size of the marine rotifer Brachionus plicatilis. Growth hormone (GH) or human chorionic gonadotropin (HCG) at 0.0025–25 I.U. ml−1 and estradiol-17s (E2), triiodothyronine (T3), 20-hydroxyecdysone (20-HE), 5-hydroxytryptamine (5-HT), gamma-aminobutyric acid (GABA) or juvenile hormone (JH) at 0.05–50 mg 1−1 were added to 5-ml of Nannochloropsis oculata suspension (7 × 106 cells m1−1). From an initial density of 1 individual m1−1, rotifers were cultured with hormones for 48 hours in 22 ppt seawater at 25 °C, in darkness. Rotifers were counted and classified into female types and transferred to a new algal food suspension without hormone every other day until day 8 when body size was measured.

Surface glycoproteins potentially involved in mate recognition in nine freshwater rotifer species

A surface glycoprotein in the marine rotifer Brachionus plicatilis functions as a contact mate recognition pheromone (MRP). A polyclonal antibody to this glycoprotein (anti-MRP) binds to the body surface of nine freshwater rotifer species at sites important in mating behavior of the species studied. Binding among brachionid species was concentrated in the corona and foot as expected from previous observations of mating behavior. In non-brachionid species other than bdelloids, binding occurred in areas where males contacted females and initiated mating. Although the antibody bound to female Brachionus calycifloris, it did not decrease the number of male mating attempts. These results are discussed in light of other research on anti-MRP blokkage of male mating in B. plicatilis and B. rotundiformis strains.

Characteristics of the mate‐recognition pheromone inbrachionus plicatilis(rotifera)

The chemical recognition systems used by aquatic invertebrates to discriminate mates are poorly understood. We describe the results of analysis of a surface glycoprotein that serves as a mate‐recognition pheromone (MRP) for the marine rotifer Brachionus plicatilis. Binding of a polyclonal antibody to the MRP signal on females is described. The binding of anti‐MRP to purified MRP in nitrocellulose membranes is non‐linear. Anti‐MRP also can be used to detect MRP in crude homogenates of rotifer proteins where MRP comprises 0.06 to 0.14% of total protein. As little as 1–2 ng of MRP can be detected in crude homogenates. We estimated that a single B. plicatilis female has about 0.11 ng of MRP concentrated primarily in her corona. Males can discriminate conspecific females from those of other species, females from different geographic populations, and males from females in their own populations. Monosaccharide analysis of the MRP demonstrated the presence of mannose, glucose, N‐acetylglucosamine, fucose, and N‐a...

Culture technology of marine rotifers and the implications for intensive culture of marine fish in Japan

In order to expand the practice of rearing marine finfish larvae, it is necessary to increase and stabilize the production of live feed. Latest researches on rotifer culture technology in Japan have aimed at these objectives. This paper summarizes the progress on the technology of high-density rotifer mass cultures. The high-density culture of the marine rotifer Brachionus rotundiformis (so-called S-type) enables production that is at least 100 times more efficient than that by the conventional method, with less cost for production. The technology is based on the use of highly concentrated algal biomass as food for rotifers. Rotifers are mass-cultured at 10000-30000 individuals mL-1 by feeding them exclusively on condensed freshwater Chlorella. Cultures are generally supplied with constant O2; pH is adjusted to 7 by addition of HCI to avoid an increase of free ammonia in the culture system. The excess suspended organic matter can be efficiently removed by a nylon filtration mat.

Resting eggs of the marine rotifer Brachionus plicatilis M�ller: development, and effect of irradiation on hatching

Resting eggs of the marine rotifer Brachionus plicatilis M�ller: development, and effect of irradiation on hatching

Identification of a sex pheromone from a rotifer

The first isolation and characterization of a sex pheromone from a zooplankter is reported. A 29 kdalton glycoprotein on the surface of females of the marine rotifer Brachionus plicatilis acts as a contact-mating pheromone. This glycoprotein (gp29) is glycosylated with oligosaccharides containing Nacetylglucosamine, mannose, and fucose residues, and these oligosaccharides are necessary for male recognition of females. Males detect this signal by contact chemoreception with receptors located in their corona. Binding of purified glycoprotein to male receptors reduces mating attempts by 93%. An antibody to the glycoprotein binds to females, reducing male mating attempts by 86%. When purified gp29 is bound to sepharose beads, it is sufficient to elicit male mating behavior. This glycoprotein is likely to play a key role in the evolution and maintenance of reproductive isolation in rotifers.

Studies on the Formation and Hatching of Fertilized Eggs of the Rotifer Brachionus plicatilis-VIII. Dietary Value of Neonates from Rotifer Brachionus plicatilis Resting Eggs for Red Sea Bream Larvae.

The dietary value of hatched neonates from the marine rotifer Brachionus plicatilis resting eggs was investigated through larval rearing trials of red sea bream Pagrus major and chemical analysis. Rotifer resting eggs were mass-produced in 50m3 tanks by feeding Nannochloropsis oculata and baker's yeast. Eggs were preserved at 5°C under total darkness for 1-2 years and hatched at 23°C under continuous light. After nutritional enrichment with the use of N. oculata and squid oil, hatched neonates or commonly mass-cultured rotifers (control) were fed to fish larvae. Newly hatched fish larvae were stocked in 100l (80l in water volume) polycarbonate tanks at 15 or 30/l. No significant differences in larval growth (notochord length and dry weight) were found on Day 20 between groups fed on neonates from resting eggs and those fed on control rotifers. The survival rate of fish larvae fed on neonates (38%) were, however, significantly lower than the group fed on control diets (69%). This may be due to the fact that the uniform lorica size and synchronous growth of neonates during the enrichment culture after simultaneous resting egg hatching did not fit the mouth size of the fish larvae. The amount and composition of fatty acids of neonates were the same as those of the control rotifers.

Isolation and purification of glutathione S-transferases from Brachionus plicatilis and B. calyciflorus (Rotifera)

1. 1. The enzyme glutathione S-transferase (GST), a critical element in xenobiotic metabolism, was isolated from the marine rotifer Brachionus plicatilis and its freshwater congener B. calyciflorus. 2. 2. In B. plicatilis, GST comprised 4.2% of cytosolic protein and was present as three separate isozymes with mol. wts 30,000, 31,400 and 33,700. Specific activity of crude homogenates was 56 nmol min −1 mg −1 protein, while that of affinity chromatography purified GST was 1850. 3. 3. In B. calyciflorus, GST was present as two isozymes with mol. wts of 26,300 and 28,500, representing 1.0% of cytosolic protein. Crude GST specific activity was 1750 nmol min −1 mg −1 protein and purified was 72,400. 4. 4. Rotifer GSTs are unusual because they are monomers whereas all other animals thus far investigated possess dimeric GSTs.