Brachionus Plicatilis Research Articles

Genome size evolution at the speciation level: The cryptic species complex Brachionus plicatilis(Rotifera)

BackgroundStudies on genome size variation in animals are rarely done at lower taxonomic levels, e.g., slightly above/below the species level. Yet, such variation might provide important clues on the tempo and mode of genome size evolution. In this study we used the flow-cytometry method to study the evolution of genome size in the rotifer Brachionus plicatilis, a cryptic species complex consisting of at least 14 closely related species.ResultsWe found an unexpectedly high variation in this species complex, with genome sizes ranging approximately seven-fold (haploid '1C' genome sizes: 0.056-0.416 pg). Most of this variation (67%) could be ascribed to the major clades of the species complex, i.e. clades that are well separated according to most species definitions. However, we also found substantial variation (32%) at lower taxonomic levels - within and among genealogical species - and, interestingly, among species pairs that are not completely reproductively isolated. In one genealogical species, called B. 'Austria', we found greatly enlarged genome sizes that could roughly be approximated as multiples of the genomes of its closest relatives, which suggests that whole-genome duplications have occurred early during separation of this lineage. Overall, genome size was significantly correlated to egg size and body size, even though the latter became non-significant after controlling for phylogenetic non-independence.ConclusionsOur study suggests that substantial genome size variation can build up early during speciation, potentially even among isolated populations. An alternative, but not mutually exclusive interpretation might be that reproductive isolation tends to build up unusually slow in this species complex.

Pengaruh Penambahan Vitamin B pada Media Blotong 12 Kering terhadap Pertumbuhan Populasi Dunaliella salina <br><i>[Effect of Adding Vitamin B on Blotong Dry Media 12 Againts Population Growth Dunaliella salina]<i>

Abstract Dunaliella salina is one natural food that is good enough for the larvae of sea urchins and also can be used as food Brachionus plicatilis and Artemia in aquaculture feed Artemia. Blotong is precipitated in the process of purification juice before cooking and crystallized into sugar. Purification is a process to separate the juice from dirt or fibers. Sugarcane is used as fertilizer by using oven dried first. Vitamin B is needed some algae to build 12 proteins and essential as a coenzyme necessary for DNA synthesis in the formation of new cells. Vitamin B is a 12 coenzyme B -dependent methionine synthase and vitamin B -dependent ribonucleotide reductase. Vitamin B 12 12 12 dependent methionine synthase function for synthesis methionin (essential amino acids), whereas vitamin B 12 dependent ribonucleotide reductase functions to DNA synthesis. This study aims to determine the effect of vitamin B in dry blotong media on the growth of D. salina 12 population and to determine the optimal dosage of vitamin B in dry blotong media which can produce the highest 12 population growth of D. salina. This research used Completely Randomized Design (CRD) with seven treatments and three replications. The study used namely: Fertilizer blotong dried with the addition of vitamin B 10 g / L (A), dose of 12 12 ug / L (B), the dose of 14 ug / L (C), a dose of 16 ug / L (D), dose 18 ug / L (E), a dose of 20 ug / L (F) and control treatment of dry sugarcane fertilizer without adding vitamin B12 or dose 0 g / L (G). The main parameter used is the population of D. salina, while supporting the water quality parameters (salinity, pH and water temperature). Data were analyzed with ANOVA (analysis of variants) and to know the differences among the treatments performed Duncan's Multiple Range Test. The results showed that the addition of vitamin B in dry blotong media had no effect on the growth of population D. salina (p> 0.05).

A novel growth-promoting protein in the conditioned media from the rotifer Brachionus plicatilis at an early exponential growth phase

We confirmed the existence of growth-promoting substances in the conditioned media (CM) from the rotifer Brachionus plicatilis at an early exponential growth phase and isolated a novel protein with a growth-promoting activity from the crude extract (CE) of rotifer cells. CM was prepared from the culture media where rotifers had been cultured at an early exponential growth phase and filtered through a 0.22-μm filter membrane. The growth-promoting activity was determined using rotifers in CM for 5 days. As a result, the increase of rotifers added with CM was significantly higher than that of the control in artificial seawater (P < 0.001). Moreover, the growth-promoting activity of CM was dose-dependent and inactivated by heat treatment at 80°C for 60 min. Meanwhile, CM filtered through a <10 kDa ultrafiltration membrane showed a low activity, whereas proteinase K treatment resulted in a complete inactivation. These results suggest that the rotifer secrets growth-promoting proteins into CM. CE also contained a protein with the activity and properties similar to those found in CM. Then, CE was subjected to purification of a growth-promoting protein for convenience using various types of chromatography after fractionation with 30–80% saturated ammonium sulfate. Subsequently, a protein with an approximate molecular weight of 25000 was isolated, and its N-terminal amino acid sequence was determined to be PAVVDFTAVWFGPLQMIKP. An orthologue was found in the EST database of B. plicatilis, the full sequence of which showed about 50% identity to the corresponding regions of thioredoxins from other organisms.

Effects of calorie restriction on the expression of manganese superoxide dismutase and catalase under oxidative stress conditions in the rotifer Brachionus plicatilis

Calorie restriction (CR) in the rotifer Brachionus plicatilis extends its lifespan, as it enhances the expression of antioxidant enzymes such as manganese superoxide dismutase (Mn SOD) and catalase. Here we show that CR also increased the mRNA levels of these antioxidant enzymes upon exposure to oxidative stress. Rotifers cultured under CR showed a higher survival rate than those fed ad libitum (AL) upon exposure to 0.05–0.2 μM juglone, an oxidative stress inducer. The relative mRNA levels of Mn SOD and catalase before exposure to juglone were slightly higher in the CR rotifers than in their AL counterparts, although these differences were not statistically significant. AL rotifers showed no apparent upregulation of the mRNA levels of these antioxidant enzymes upon exposure to 0.025 and 0.05 μM juglone. In contrast, the CR rotifers increased the mRNA levels of Mn SOD and catalase by up to 5.4-fold and 4.2-fold, respectively, resulting in significant differences between their levels in AL and CR rotifers under oxidative stress conditions. Furthermore, the protein level of catalase was clearly higher in CR than in AL rotifers 6 h after exposure to oxidative stress. These results suggest that the upregulation of Mn SOD and catalase genes is involved in CR-induced resistance to oxidative stress in the rotifer.

Patterns and processes in the genetic differentiation of the Brachionus calyciflorus complex, a passively dispersing freshwater zooplankton

Elucidating the evolutionary patterns and processes of extant species is an important objective of any research program that seeks to understand population divergence and, ultimately, speciation. The island-like nature and temporal fluctuation of limnetic habitats create opportunities for genetic differentiation in rotifers through space and time. To gain further understanding of spatio-temporal patterns of genetic differentiation in rotifers other than the well-studied Brachionus plicatilis complex in brackish water, a total of 318 nrDNA ITS sequences from the B. calyciflorus complex in freshwater were analysed using phylogenetic and phylogeographic methods. DNA taxonomy conducted by both the sequence divergence and the GMYC model suggested the occurrence of six potential cryptic species, supported also by reproductive isolation among the tested lineages. The significant genetic differentiation and non-significant correlation between geographic and genetic distances existed in the most abundant cryptic species, BcI-W and Bc-SW. The large proportion of genetic variability for cryptic species Bc-SW was due to differences between sampling localities within seasons, rather than between different seasons. Nested Clade Analysis suggested allopatric or past fragmentation, contiguous range expansion and long-distance colonization possibly coupled with subsequent fragmentation as the probable main forces shaping the present-day phylogeographic structure of the B. calyciflorus species complex.

Effects of Salinity on Population Growth, Nutritive Value, and Feeding Cost of the Enriched Rotifer, Brachionus plicatilis

Effects of Salinity on Population Growth, Nutritive Value, and Feeding Cost of the Enriched Rotifer, Brachionus plicatilis

Automatic measurement of Acartia tonsa nauplii density, and estimation of stage distribution

Copepods are increasingly seen as an important feed in the culture of marine fish. Calanoid species such as Acartia tonsa have been studied for mass culture purposes. In this work, a plankton counter developed for automatic measurement of density in rotifer ( Brachionus plicatilis) cultures has been tested for Acartia nauplii. The plankton counter, after adaptation, can estimate both the size and density of the nauplii within reasonable margins of error. Size estimates were validated against measurements made by microscope, and density estimates against manual counts. The plankton counter makes it possible to automatically monitor metrics like hatching success, mortality and development rate in Acartia cultures, as well as monitoring of densities in larval first feeding tanks. The latter opens the opportunity for appetite based feeding of larvae. The area measurement can be used to estimate the biomass and the development stage of the nauplii.

Effect of enriched Brachionus plicatilis and Artemia salina nauplii by microalga Tetraselmis chuii (Bütcher) grown on four different culture media on the growth and survival of Sparus aurata larvae

constituted 22.3% of the total fatty acids, due to the presence of DHA which formed 8.0% of the total fatty acids, while ARA and EPA were absent. The results show that B. plicatilis and newly hatched Artemia enriched with T. chuii grown on was effective for good growth and survival rate of S. aurata larvae.

Seasonal changes in zooplankton community structure in Lake Küçükçekmece, İstanbul, Turkey

The zooplankton community of Lake Küçükçekmece includes individuals from freshwater groups Rotifera and Cladocera, and also marine species Acartia clausi of the subclass Copepoda; Tintinnopsis nana, Favella sp. Jorgensen, 1924, and Strobilidium spiralis from the group Ciliata; Micronereis sp. Linnaeus, 1758 and Spio sp. Fabricius, 1785 from the class Polychaeta; Pleurobrachia pileus from the phylum Ctenophora; and Gammarus sp. Fabricius, 1775 from the order Amphipoda. Rotifera was the dominant group for each season studied. In autumn, dominant specimens included Bosmina longirostris, Spio sp. Fabricius, 1785, Pleurobrachia pileus, the cypris and nauplius larva of Balanus sp., and veliger larva. In winter, dominant species included Synchaeta pectinata, Filinia longirostris, and Epiphanes sp. Ehrenberg, 1832. In spring, dominate species included Asplanchna priodonta and Strobilidium spiralis. In summer, the dominant species was Brachionus plicatilis. Polyarthra major, Filinia longiseta, Keratella cochlearis, and Brachionus, which were found in the lake, are indicator species of eutrophication. Marine species Tintinnopsis nana peaked during spring at the first and second sampling stations, while Acartia clausi peaked during summer at the second station.

Bacillus cereus and Bacillus subtilis used as probiotics in rotifer (Brachionus plicatilis) cultures

Since live food is a key factor in fish and invertebrate larvae rearing, the objective of this study was to evaluate the probiotic potential of two Gram positive Bacillus species: Bacillus cereus (CCBM-2) and Bacillus subtilis (CCBM- 64), originally isolated from marine sediments in the Colombian Caribbean, in rotifer Brachionus plicatilis cultures. The enzymatic activity and antibacterial activity against pathogens of B. cereus and B. subtilis were measured in vitro. Direct supplementation of the rotifer culture water with both bacteria was also carried out and the effect was measured in vivo by estimating the density of rotifers and bacterial counts in the culture. The results indicated that extracellular products of both bacteria strongly inhibited the growth of AeromonA. hydrophila and Vibrio alginolyticus isolated from diseased fish. Enzyme API ZYM assays showed that both bacteria have esterase lipase, leucine arylamidase, acid phosphatase, lipase, and Naphthol-AS-BI- phosphohydrolase activities. Furthermore, it was observed that the addition of B. subtilis to the rotifer culture water resulted in a significant increase in rotifer numbers and a reduction of Vibrio levels. B. subtilis appears to be a promising probiotic for rotifer cultures, but further research is required to determine the capacity in controlling A. hydrophila and V. alginolyticus when administered via rotifers to fish larvae.

Pastas de Rhodomonas salina (Cryptophyta) como alimento para Brachionus plicatilis (Rotifera)

Rotifers are an important live feed for first feeding larvae of many fish species. The use of concentrated algae cells in the mass culture of the rotifer Brachionus plicatilis (Brachionidae) has opened new horizons for research on this organism. Pastes of Rhodomonas salina (Pyrenomonadaceae) obtained either by centrifugation or flocculation with chitosan were preserved, with or without vitamin C, at -20 degrees C for four weeks and were evaluated biochemically (proteins, lipids, pigments and fatty acids contents) and subsequently, were used to feed the rotifer Brachionus plicatilis at a ratio of 25 mg/L/day. Four different microalgae pastes were prepared: (1) centrifuged and preserved with vitamin C (CV), (2) centrifuged and preserved without vitamin C (C), (3) flocculated and with vitamin C (FV) and (4) flocculated without vitamin C (F). All treatments showed similar contents of proteins and total lipids with respect to control culture (a fresh culture of R. salina), with mean values of 40.0 +/- 2.32% and 12.0 +/- 1.45%, respectively. The pheophytin a/chlorophyll a ratio, a general indicator of the chemical status of microalgal concentrates, was similar (0.09-0.11) between centrifuged pastes and control culture, but was found to be higher in flocculated pastes (1.28-1.48). The fatty acid profile varied with respect to the control culture, mainly in the proportion of the essential polyunsaturated fatty acids (PUFAs): eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA). Total PUFAs, EPA and DHA contents were statistically similar between centrifuged pastes and control culture (PUFAs: 47%, EPA: 4% and DHA: 4.7%), whereas values obtained for flocculated pastes were significantly lower. The rotifers grew equally well when fed with centrifuged pastes or control culture (maximum density: 320 rotifers/mL; instantaneous growth rate: 0.23 rotifers/day, fecundity: 1.49 eggs/female and productivity: 43 x 10(3) rotifers/L/day. No significant effect of vitamin C was found when used as a paste preservative. We concluded that centrifugation is an effective harvesting method, and that freezing to -20 degrees C for four weeks (no vitamin added), may help maintain the nutritional quality of R. salina paste, similar to fresh microalgae and can be offered to Brachionus plicatilis.

Population Growth of Six Iranian Brachionus Rotifer Strains in Response to Salinity and Food Type

AbstractIntrinsic rates of population increase (r) were evaluated as a measure of population dynamics of four strains of Brachionus plicatilis and two strains of B. urceolaris from Iran in response to different salinities and feeding algae. Each rotifer strain was cultured at four salinities: 5, 20, 25 and 30‰ and fed with two microalgal species: Chlorella vulgaris and Nannochloropsis oculata. Salinity of 5‰ was critical for all the examined strains, at which r was at minimum and was different from the other salinities (P &lt; 0.05). For B. plicatilis strains, the maximum r was observed in those fed on Chlorella at salinities of 10 and 30‰ (64 ± 0.01 day−1). While, in B. urceolaris, maximum r was for Nannochloropsis fed rotifers at salinity of 20‰ (0.69 ± 0.01 day−1). Maximum final population density (FD) was obtained for a strain of B. urceolaris fed on Nannochloropsis at 20‰ (329.3 ± 10.9 ind.mL−1). FD was relatively lower in B. plicatilis strains among all examined salinities. ANOVA showed the significant effect of salinity and rotifer strain, and algae × rotifer strain on both r and FD, and salinity × rotifer × algae on FD (P &lt; 0.05). (© 2010 WILEY‐VCH Verlag GmbH &amp; Co. KGaA, Weinheim)

The relationship between enrichment, fatty acid profiles and bacterial load in cultured rotifers (Brachionus plicatilis L-strain) and Artemia (Artemia salina strain Franciscana)

Rotifers and Artemia are generally used as first foods in marine finfish aquaculture. Because of their poor nutritional value and the incapacity of marine fish to elongate or desaturate 18-carbon of longer polyunsaturated fatty acids (PUFA), it is a common practice to enrich live foods with commercial products. Since live foods represent a significant vector for transmitting bacterial contaminants, this study describes the impact of using different enrichment strategies for rotifers and Artemia on bacterial load, in addition to fatty acid profiles. Rotifers were reared in continuous culture while Artemia were obtained from cysts; both were enriched for 24h. Total bacterial counts were obtained after a 7-d incubation on marine agar. Docosahexaenoic acid (DHA) levels varied from 9.8 to 34.4% and from 8.3 to 23.2% respectively, for rotifers and Artemia. Eicosapentaenoic acid (EPA) levels ranged from 3.2 to 7.3% for rotifers and from 5.1 to 9.0% for Artemia, while arachidonic acid (ARA) levels varied, respectively, from 0.7 to 2.9 and from 1.4 to 3.7. Total bacterial counts varied from 0.9×108 to 56.6×108 for rotifers and from 0.2×109 to 11.7×109 for Artemia. These results demonstrate the importance of the enrichment strategy on the fatty acid composition and the bacterial contamination of live food.

Local adaptation in rotifer populations

The adaptation of organisms to their environment has been a subject of study for a long time. One method to study adaptations in populations involves comparing contemporary populations of the same species under different selective regimes, in what is known as a ‘local adaptation’ study. A previous study of the cyclically parthenogenetic rotifer Brachionus plicatilis found high heritabilities for some life-history traits. Some of these life-history traits significantly differed among six populations from Eastern Spain and data suggested some traits to have higher evolutionary rates than neutral genetic markers. Here, by studying the same B. plicatilis populations, we examine the variation and possible local adaptation of their main life-history traits, closely related to fitness, in relation to habitat salinity and temperature. These environmental factors have been shown to play a key role in the ecological differentiation among co-generic species of B. plicatilis. The results obtained in this study show that: (1) the seasonality of rotifer populations from Eastern Spain has profoundly influenced sexual reproduction strategies; (2) salinity is probably a key factor in the ecological specialization of some populations; and (3) rotifer populations harbour high variability in their fitness components.

Evaluating bioencapsulation of florfenicol in rotifers (Brachionus plicatilis)

The rotifer Brachionus plicatilis is one of the most important live-feed organisms used in the larval culture of marine fish. Bioencapsulation of florfenicol in rotifers was investigated by delivering it directly to the organisms as particles and the doses ranged from 50 to 1000 mg L⁻¹. Analysis of the florfenicol concentrations in rotifers was performed using high-performance liquid chromatography. The uptake increased with the dose and the enrichment time, and the highest concentration achieved in this study, 1.25 ng rotifer⁻¹, was obtained using a dose of 1000 mg L⁻¹ and 150 min of enrichment. The optimal time for satisfactory enrichment of rotifers with florfenicol was found to be dose dependent and ranged from 30 to 120 min for doses between 400 and 800 mg L⁻¹. The parameters affecting the concentrations achieved in rotifers were the size of the florfenicol particles, enrichment time and dose.

Developmental stages in diapausing eggs: an investigation across monogonont rotifer species

Monogononts reproduce by parthenogenesis punctuated by events of mixis that generate ‘resting eggs’, which actually are embryos whose development is arrested. We document the nuclei number and position of the resting eggs of nine rotifer species (Brachionus plicatilis, B. calyciflorus, B. manjavacas, Plationus patulus, Epiphanes senta, E. chihuahuaensis, Rhinoglena frontalis, Lecane bulla and Sinantherina socialis) in the attempt to assess the stage at which dormant embryos are arrested. The morphology of the entire embryos was reconstructed visualising nuclear DNA using confocal microscopy, and their developmental stage identified. Two groups of species were identified: dormant embryos of one group possessed on average less than 30 nuclei, with low variation within and between species, and the stage may correspond to early gastrula; embryos in the second group contain relatively more nuclei, averaging around 40–60, with higher variation within species. The two groups of species seem not to reflect any phylogenetic relationship. Consequences of the dormant embryo stages are discussed.

Effect of experimental methodology on estimation of density at sex initiation in cyclically parthenogenetic rotifers

Sexual reproduction is density-dependent in several cyclically parthenogenetic rotifer species and induced by a chemical signal produced by the rotifers. As population density increases, the chemical accumulates in the medium until reaching a threshold that triggers sex. The density threshold for sex initiation is of ecological and evolutionary importance due to the large impact that timing of sex has on fitness in cyclical parthenogenetic rotifers. Here we study the effects of experimental methodology in combination with intrinsic features of the biological system on the estimation of the density required to produce enough of this chemical to initiate sex (i.e., density threshold). We have developed a theoretical model describing the dynamics of both population density and the concentration of the chemical signal. Model analysis showed that the time at which sex is recorded in an experiment, the initial density (i.e., experimental volume) and the population growth rate can affect the estimation of density thresholds. Results from an experiment in which density at sex initiation was tested for 29 clones of Brachionus plicatilis at two culture volumes, revealed that density threshold is negatively dependent on the experimental volume. We conclude that in order to estimate density thresholds so that they are comparable across studies, experimental methodology should be standardized.

Effect of female aging on the morphology and hatchability of resting eggs in the rotifer Brachionus plicatilis Müller

This study examined the morphology and hatchability of Brachionus plicatilis resting eggs as a function of the aging of maternal fertilized mictic females. One-hundred twenty fertilized B. plicatilis (Australian strain) were individually cultured and monitored daily until death. All cultures were maintained at 25°C, 11 ppt, and fed the micro-algae Tetraselmis tetrathele. Resting eggs produced by the females were investigated using two parameters: egg morphology, and hatching rate. Under these culture conditions, females normally produce 1–6 (mean ± SD = 2.7 ± 1.2) resting eggs during their lifetime. However, the number of resting eggs with abnormal morphology increased as a function of maternal age. Among resting eggs with normal morphology (n = 225), 82.2% were produced during the first and second spawning, and had hatching rates of more than 60%, while the hatching rates were below 30% in resting eggs with a spawning order of >2. Thus, the quality of B. plicatilis resting eggs was negatively correlated with maternal age.

Axenic culture of Brachionus plicatilis using antibiotics

The rotifer Brachionus plicatilis culture is composed of complex microcosms including bacteria, protozoans, algae, and fungi. Previous studies reported methods to establish axenic rotifer cultures, but further refinement of these techniques is needed, for molecular biological research which requires pure culture to isolate nucleic acids from rotifers only. In order to render rotifer culture axenic, we tested five antibiotics: ampicillin (Amp), chloramphenicol (Cp), kanamycin (Km), nalidixic acid (Na), and streptomycin (Sm) at 30–100 μg/ml. Except for Cp, which reduces rotifer reproduction, all other antibiotics at the tested concentrations did not affect rotifer reproduction or show any toxic effects. A rotifer disinfection method was finally established by treating the resting eggs with 0.25% (w/v) sodium hypochlorite (NaOCl) for 3 min, washing with sterilized sea water, and then exposing the neonates to an Amp, Km, Na, and Sm mixture. Using four nutrient media, we confirmed that this protocol renders the rotifer culture bacterial and fungus free. The axenic rotifer culture generated here is useful not only for genetic analysis of Brachionus plicatilis, but for studying the rotifer life cycle without bacterial influence.

Molecular evolution of the membrane associated progesterone receptor in the Brachionus plicatilis (Rotifera, Monogononta) species complex

Many studies have investigated physiological roles of the membrane associated progesterone receptor (MAPR), but little is known of its evolution. Marked variations in response to exogenous progesterone have been reported for four brachionid rotifer species, suggesting differences in progesterone signaling and reception. Here we report sequence variation for the MAPR gene in the Brachionus plicatilis species complex. Phylogenetic analysis of this receptor is compared with relatedness based on cytochrome c oxidase subunit 1 sequences. Nonsynonymous to synonymous site substitution rate ratios, amino acid divergence, and variations in predicted phosphorylation sites are examined to assess evolution of the MAPR among brachionid clades.