2.2 Materials and Methods
2.2.1 Zebrafish Care
Laboratory reared wild-type (Tropical 5D) D. rerio were maintained in a recirculating AHAB system (Aquatic Habitats, Inc., Apopka, FL, USA) on a 14:10 h light/dark cycle. Water quality was maintained at 28-29°C, pH 7.0-7.5, and 60 ppm artificial seawater (ASW; Instant Ocean, Foster & Smith, Rhinelander, WI, USA). Adult fish were fed twice daily ad libitum with Artemia nauplii in the morning and Zeigler’s Adult Zebrafish Complete Diet (Zeigler Bros., Inc., Gardners, PA, USA) in the afternoon.
2.2.2 Maternal (F0) Dietary Benzo(a)pyrene Exposure
Sixty healthy reproductively mature female zebrafish (approximately 1 year old) were selected as the F0 generation. F0 females were fed either acetone-treated
…show more content…
(control) or BaP-treated (nominally equivalent to 12.5, 125, or 1250 μg BaP/g diet) decapsulated brine shrimp eggs (for measured concentrations, see 2.3.1 Analytical Chemistry).
The BaP concentrations and exposure regime were adapted with modification from a previous study finding multigenerational effects of benzo(a)pyrene exposure in zebrafish (Corrales et al. 2014b). To prepare the spiked diet, 36 g of decapuslated brine shrimp eggs were spiked with 27 mL of acetone containing BaP (0, 0.01667, 0.1667, or 1.667 μg/L). The spiked diets were air dried and stored in amber jars at room temperature. F0 females were housed at a density of 5 fish per 9.4 L aquarium (Pacific Coast Distributing, Inc., Phoenix, AZ, USA) with hanging filters, with three replicate tanks per treatment group. Water quality and photoperiod were maintained as above. Fish were allowed to acclimate for a week while being transitioned to a diet of untreated decapsulated brine shrimp eggs. The first day of the acclimation period and the day …show more content…
before the start of the exposure, the females were spawned with naïve males in breeder tanks at a 5:3 ratio to purge them of their existing mature oocytes. During the exposure, fish were fed 1% body weight twice/day of the corresponding BaP-treated food for 21 days (nominally equivalent to 0, 250, 2500, 25000 ng BaP/g fish). On day 22, the experimental females were spawned with naïve males in breeder tanks at a 5:3 ratio and the eggs were collected, maintained in embryo medium (30% Danieau), and placed in an incubator at 28°C. On days 23 and 24, the females were euthanized for subsequent analyses. 2.2.3 Analytical Chemistry Concentrations of BaP in fish food were measured on days 0 and 21.
Briefly, 2-10 mg fish food was spiked 100 µL with D12-benzo(a)pyrene internal standard (AccuStandard, New Haven, CT, USA) and extracted by sonicating 10 min in 0.5 mL 1:1 hexane:acetone followed by centrifugation for 1 min at 10,000 x g. The organic layer was transferred to an autosampler vial and the pellet extracted twice more, combining the organic fractions. Extracts were concentrated under N2 to 200 µL, exchanged into hexane and spiked with D12-pyrene (AccuStandard) as a secondary internal standard to calculate recoveries of D12-benzo(a)pyrene, and diluted to 1 mL with hexane. Triplicate blanks containing no fish food were processed along with samples. Samples were analyzed by gas chromatography-mass spectrometry (GC-MS) in electron ionization mode using selected ion monitoring (Agilent 7890A GC, 5975C MS; Agilent, Wilmington, DE, USA). Analytes were separated using an Agilent DB-5ms column (0.25 mm ID 30 m; 0.25 µm film), 2 µL injection, and thermal gradient (80°C for 1 min; 2°0C/min to 300°C, 300°C for 10 min) at 1.2 mL/min constant flow. BaP and the mass-labeled internal standards were detected at 252 and 264 m/z, respectively. Recoveries of D12-benzo(a)pyrene were 95±3%, and the method detection limit was 0.07
ng/mg.
The Artemia franciscana can survive in extreme conditions of salinity, water depth, and temperature (Biology 108 laboratory manual, 2010), but do A. franciscana prefer these conditions or do they simply cope with their surroundings? This experiment explored the extent of the A. franciscanas preference towards three major stimuli: light, temperature, and acidity. A. franciscana are able to endure extreme temperature ranges from 6 ̊ C to 40 ̊ C, however since their optimal temperature for breeding is about room temperature it can be inferred that the A. franciscana will prefer this over other temperatures (Al Dhaheri and Drew, 2003). This is much the same in regards to acidity as Artemia franciscana, in general thrive in saline lakes, can survive pH ranges between 7 and 10 with 8 being ideal for cysts(eggs) to hatch (Al Dhaheri and Drew, 2003). Based on this fact alone the tested A. franciscana should show preference to higher pH levels. In nature A. franciscana feed by scraping food, such as algae, of rocks and can be classified as a bottom feeder; with this said, A. franciscana are usually located in shallow waters. In respect to the preference of light intensity, A. franciscana can be hypothesized to respond to light erratically (Fox, 2001; Al Dhaheri and Drew, 2003). Using these predictions, and the results of the experimentation on the A. franciscana and stimuli, we will be able to determine their preference towards light, temperature, and pH.
The Zebra Mussel (Dreissena polymorpha), is a small freshwater mollusk that is an invasive species. It has slowly been making its way into the United States. It has done this by showing up in lakes and in rivers. The mussels get their name due to the striped pattern on their shells. They are a relatively small species, only growing to the size of a human finger nail but there have been cases of larger (“Zebra Mussels,” 2013, para. 2). Mussels live a short life span of 4-5 years and most do not make into adulthood. They live in freshwater at depths of 6 to 24 feet. The female produces 30,000 to 1,000,000 eggs per year starting at the age of two. Zebra Mussels are free moving and can move with an external organ called the byssus (“Zebra Mussels,” 2013, para. 3). But they can be moved around by currents and other objects too. Zebra Mussels are a threat to ecosystems. They damage the natural habitat of lakes and rivers and cause negative effects to the native aquatic life. They multiply in vast majority casing a lake of food and space for other animals. And also the U.S. has spent millions on the removal of these pests from our water (“Zebra Mussels,” 2014, para. 8). The Zebra Mussel has now made its way into Colorado. Just a few has been found in the waters of Colorado but a few is enough to cause a big stir. Extreme precautions have now been put into effect to make sure that the threat of Zebra Mussels is under control.
To begin the lab, the variable treatment was prepared as the Loggerlite probe, used to later measure oxygen consumption, warmed up for approximately 10 minutes. To prepare the variable treatment, 200ml of Sodium and Ammo-lock water was measured in a container and a pre-prepared “tea bag” of tobacco was steeped in the room temperature treated water until a light yellow color was visible. After preparing the tobacco solution the preparation for the live goldfish began as two beakers were filled with 100 ml of treated water. Each beaker was weighed before addi...
Zebra mussels have now found there way to the United States originating from the Caspian Sea and sought habitation, originally, in Lake St. Claire when ballast water brought them in. From there, zebra mussels have spread and have caused havoc to the environments and its biodiversity, specifically, Presque Isle. Zebra mussels are filter feeders of zooplankton and phytoplanktons, making the water appear clearer, but not cleaner. They have negatively impacted the existence of clams, walleye and several other organisms. In order to preserve the biodiversity of Presque Isle, zebra mussels must be culled and controlled. Several methods that can be utilized to achieve this goal is by chlorination, poison, scraping, etc.
Guillermo Aragon English 121 Leslie McCutchen October 09, 2015 In the Deep End Since the beginning of filmmaking producers have made documentaries to present an argument, an idea, or to portray a kind of reality. In an attempt to introduce an argument of some kind, the filmmaker’s intent is to engage the audiences and to also appeal to their values and emotions. To do this, producers use a number of rhetorical deceives, such as logo, ethos, and pathos, to help get their audience to believe and think in a certain way. Gabriela Cowperthwaite, director of the 2013 documentary Blackfish, uses these rhetorical techniques throughout the film, which tells the story about Tilikum, an orca whale that killed three professional trainers while in captivity.
Lionfish are not currently listed as threatened in their native range. However, the increase in pollution in coral reefs can negatively affect the lionfish's primary food sources (NOAA, 2011). If lionfish are unable to adapt to declines in their prey species, their numbers will decrease. Although between the increasing reef pollution and the Lionfish’s voracious appetite reef populations will more than likely be damaged beyond repair, they may even be close to elimination by the time the Lionfish population actually declines (Albins, 2013).
The experiment measured the survival rate, the growth rate, and the size of the brine shrimp at the time harvested in various environments. To obtain these measurements, three environments were created: sea water, brackish water, and freshwater. For this experiment the scientists used 5 liter plastic buckets. Every two days, half of the water from each bucket was discarded and new water, of each respective salinity, was added into each bucket...
...et al. (2011). Using fluorescent imaging, the researchers found evidence of abnormal vascularization, neuron branching, and neuromast cell development in zebrafish (Danio rerio) exposed to the known endocrine disruptor during early life stages. Aluru et al. (2010) determined that maternal exposure to BPA can cause multiple adverse effects on developing offspring. Unfertilized rainbow trout eggs were treated with three different concentrations, fertilized, and resulting juveniles were observed throughout development. Aluru et al. (2010) concluded that oocyte exposure to BPA leads to modified stress performance, delayed hatching times, and growth suppression in juvenile rainbow trout. The following image is taken from Aluru et al. (2010), showing both the decrease in body size and production of yolk observed in juvenile rainbow trout hatched from BPA-exposed oocytes.
The lionfish not only grow at an alarming rate, but they are able to reproduce at an alarming rate in non-native oceans. An adult lionfish will grow between 12-15 inches in length in their native habitat, but have reported to grow at an alarming rate and have been caught at 18 inches and all the way up to 20 inches off the coast of Florida. This species re-produces so quickly due to the fact that the female lionfish will produce to gelatinous egg packets of 12,000 to 15,000 eggs a piece. These eggs packets will break apart and drift in the current, allowing these eggs to spread apart extremely quickly. These egg masses can float and drift apart for about 25 days before tiny microscopic lionfish larvae are born (Lionfish – Pterois volitans). These larvae will reach full maturity in less than a year, allowing them to reproduce and keep the cycle of the invasion going. In warmer climate waters such as the Atlantic and Caribbean ocean a lionfish is able to spawn every four days (Lionfish – Pterois volitans). Since the nutrient rich waters of these oceans allow the lionfish to n...
White Shark: Mindless Killer or Misunderstood Creature Over the years, many people have been attacked by sharks throughout the world. However, does this warrant the hunting of White sharks? When we stop to think at why this might happen; many people might believe that sharks are terrible creatures that are out for human blood.
An example of an additive is di-butyl phthalate and biphenyl A (BPA), these substances are incorporated while the plastics are being manufactured to provide tensile flexibility and tensile strength. The microplastics tend to accumulate and adsorb toxic materials from the seawater around them, such as organic pollutants and heavy metals. The chemicals are not bound to the microplastic surface, and as a result, when ingested they tend to leach into the animal tissues (Do Sul, J. A. I, 2014). The impacts from such chemical leaches include hepatic stress in fish, mortality, and endocrine
While concluding my data for the three lures, I found data that could be useful and help other fishermen in the sport of fishing. The science behind fishing is environmental, chemistry, and physics. The majority of fishing anglers are knowledgeable to science in the sport of fishing. Due to history and past experimentation the experiment of Gulp, live bait, and frozen bait illustrates the concepts of environmental, chemistry, and physics.
The Nile Perch is an important fish in Africa and a prized catch for a sport fishermen, and could be very expensive to sell. The Nile Perch has black eyes with a silver and blue tinge. The Lates Niloticus, also known as the Nile perch, is one of the most prized fish to catch for fishermen. However, other people in the area do not see the fish as such a prize, the fish’s diet consists of the fish ordinary people are used to consuming in their own diet. The average Nile perch weighs in at five hundred pounds and up to six feet long. The fish tends to cause disarray when introduced to new areas, consuming fish of native populations. The fish has very distinctive features, its body is a slight blue color and it also has black eyes with a yellow ring surrounding them. Surprisingly, the female fish is usually larger
Far below the surface of the ocean, lies a world very different from our own. Among the many inhabitants of this world are fish. The fish of the sea are dazzling, colorful creatures. Their outer beauty is enhanced by the wide array of colors that encompass them and vary to include all shades of the rainbow. Although flat from a distance, fish are anything but. Their insides are made up of many layers including, a vertebrate, skeleton, muscles, fins, gills, and scales. Every fish has a distinctive nose and tail. They are cold blooded animals, who lay eggs to produce young and breathe as water passes over their gills. Fish are incredibly graceful and skillful swimmers who have perfected the sport as a means of escape from predators lurking in the water. Although fish are among the many wonders of the sea, they are also very useful on land. Seafood is one of the healthiest proteins, and, because of the wide variety and types available, fish is often the main course in healthy meals around the world. Finally, fish are peaceful as well as beautiful a...
Nutrition and feeding play an important role in sustainable aquaculture and therefore, feed resources as well as costs continue to dominate aquaculture needs. Fish require adequate nutrition in order to grow and survive and nature offers a great diversity of food to fish including plants and animals. Protein is the major dietary nutrient affecting performance of fish (Lovell, 1989). It provides the essential and non essential amino acids which are necessary for muscle formation and enzymatic function and in part provides energy for maintenance (Yang et al., 2002). Garling and Wilson (1976) reported a level of 25% and 36% crude protein in diets as optimum for warm water fishes. Variation in dietary protein requirement of fish is due to difference