Browsing by Subject "Karenia brevis"
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Item Brevetoxin: How Is It Made and Why(2012-10-19) Thompson, NatalieKarenia brevis is the major harmful algal bloom-forming species in the Gulf of Mexico, and produces neurotoxins, known as brevetoxins, that cause large fish kills, neurotoxic shellfish poisoning, and human respiratory distress. Brevetoxins are polyethers that bind voltage-sensitive sodium channels, opening them for prolonged periods of time. Clonal cultures of K. brevis exhibit unique brevetoxin profiles, which not only differ from one another, but also change when subjected to different environmental conditions. The brevetoxin structures were elucidated 30 years ago without any breakthroughs for the biosynthetic pathway. These unique ladder-like polyethers have 10 (PbTx-1) or 11 (PbTx-2) rings, indicating that they are synthesized as secondary metabolites by polyketide synthases. The extensive size of the genome and the lack of histones and nucleosomes combined with the additional regulatory step of a trans-splicing spliced leader sequence make normal molecular techniques ineffective in determining the genes involved in toxin synthesis. The goal of this project is to identify a potential link between toxin, gene, and function. One objective is to take the next step towards identifying the genes associated with the synthesis and regulation of brevetoxins and to help elucidate the hypothesized gene clusters of multi-protein enzymatic complexes involved in brevetoxin production, one for each backbone. The second objective is to make an effort to determine the in vivo function of the costly brevetoxins by identifying possible ion channels, which could be osmotically regulated by the toxins. Genes for polyketide synthases (PKS) were identified in K. brevis, obtained from Expressed Sequence Tag (EST) libraries. In this work, reverse transcription polymerase chain reactions (RT-PCR) were used to generate pools of complementary DNA (cDNA), which was used in real-time quantitative polymerase chain reactions (qPCR) to give relative amounts of PKS transcripts. K. brevis clones have shown a significant increase in toxin production after a rapid shift from high salinity to low salinity, indicating a regulation of brevetoxin synthesis. To gain a better understanding of regulation of toxin production during algal blooms, we compared the toxin levels under different conditions to the transcript levels of PKS genes, as determined by quantitative RT-PCR. In a separate line of investigation, an in silico analysis of the EST library was performed to identify ion channel genes expressed by K. brevis, which may be the in vivo binding site of brevetoxin. The information generated from this project will help to elucidate the effects of environmental variations on toxin production and the biological function of toxin production -- valuable information for the shellfish industries and public health.Item Differences in growth and toxicity of Karenia(Texas A&M University, 2006-08-16) Neely, Tatum ElizabethHarmful algal blooms (HABs) in the Gulf of Mexico are primarily caused by dense aggregations of the dinoflagellate species, Karenia brevis. Karenia brevis produces a highly toxic neurotoxin, brevetoxin which has been shown to cause Neurotoxic Shellfish Poisoning (NSP) and respiratory distress in humans in addition to a wide range of negative impacts upon natural ecosystems. Karenia mikimotoi is a co-existing species present during K. brevis blooms. K. mikimotoi has caused major HAB events in other parts of the ocean, but has not been recognized as a major contributor to toxicity of blooms in the Gulf of Mexico. K. brevis and K. mikimotoi have both been associated with the presence of unidentified hemolytic toxins. Production of hemolysins has not previously been investigated for either species to date in the Gulf of Mexico. Presence of hemolysins may affect toxicity and the overall impact of HABs. Therefore, detection of hemolysins is imperative for accurate identification of potential harmful impacts of such blooms. The primary goal of this research is to define whether either species is capable of producing hemolytic activity independent of brevetoxin activity; and to identify if there is significant differentiation between a variety of clonal isolates regarding toxicity and growth rate when subjected to variable experimental conditions.Item Investigation of the Population Genetic Structure of the Toxic Dinoflagellate Karenia brevis in the Gulf of Mexico(2012-08-15) Henrichs, Darren 1983-Karenia brevis is the major harmful bloom forming dinoflagellate in the Gulf of Mexico. The toxin produced by this dinoflagellate can cause large fish kills, marine mammal mortality, respiratory irritation, and neurotoxic shellfish poisoning in humans. Blooms can occur anywhere in the Gulf of Mexico (hereafter Gulf) but are predominantly observed off the west coast of Florida and the coast of Texas. The west coast of Florida has been hypothesized to be the origin for blooms of K. brevis in other regions within the Gulf based upon the frequent formation of blooms in this region. To investigate this possibility, microsatellite markers were used to determine the population-genetic structure of K. brevis in the Gulf of Mexico. The difficulties of culturing K. brevis required development and use of a single-cell PCR amplification protocol for preserved cells. Lugol's iodine-preserved bloom samples of K. brevis were destained with sodium thiosulfate and subjected to two rounds of PCR amplification. The destaining protocol resulted in the successful, simultaneous amplification of five microsatellite markers from single cells of K. brevis. A total of 18, highly polymorphic microsatellite markers are available for K. brevis. Each marker was amplified from 40 cultures of K. brevis isolated from water samples from Florida and Texas. Observed genetic diversity was high but similar to the genetic diversity observed in other phytoplankton species. No genetic divergence was detected between isolates from Florida and isolates from Texas. Single cells from a total of 38 field samples were analyzed at five microsatellite markers to determine if population-genetic structure was present in K. brevis in the Gulf. Significant genetic divergence between several individual samples was detected, reflecting the high genetic diversity present within the species. Observed genetic divergence was low between blooms from the west coast of Florida and the coast of Texas and supports the hypothesis of a common origin for blooms of K. brevis in the Gulf of Mexico.Item LISST Instruments as a Tool in Phytoplankton Ecology(2012-10-12) Railey, Lauren 1987-Laser in situ scattering and transmissometry (LISST) instruments are used to measure the particle size distributions (PSDs) and volume concentration of individual and groups of phytoplankton in water. The objective of this research was to test the LISST?s ability in detecting monospecific blooms in-situ and the ability to detect aggregation after diatoms were subjected to different temperatures and bacteria concentrations. The PSDs of ten harmful algal bloom (HAB) species were measured with the LISST characterizing the peak location, peak height, peak width, and peak range resulting in a scattering signature for each species. Each species had specific characteristics that would allow for their detection with the LISST, though microscope observations would be needed for complete accuracy. The LISST was able to detect HABs placed in natural seawater collected off the Texas coast. Blooms of four HAB species before they reached full? bloom concentrations were detected making the LISST a possible low cost, effective tool in the early detection and monitoring of HABs. The diatom, Odontella aurita, was used to test how well the LISST could monitor aggregation, an important process in the termination of many phytoplankton blooms. Increasing temperature causes an increase in transparent exopolymer particle (TEP) production in diatoms, which is a critical sticky particle that increases the probability of aggregation. An increase in temperature can also cause an increase in bacteria concentration that can positively effect TEP production and thus aggregation. O. aurita was grown at 20 ?C and 28 ?C and showed a significant increase in TEP abundance with temperature (p = 0.002), though no relationship between TEP production and bacteria concentration existed. Coomassie stained particles (CSP) are proteinaceous gel-like particles, which are currently understudied. CSP was consistently produced though it did not appear to be dependent upon any single factor. The increase in ocean temperatures has implications for an increase in phytoplankton blooms making the monitoring and understanding of these blooms even more important as they can affect the carbon cycle and potentially the microbial loop.Item Physical Mechanisms Driving Harmful Algal Blooms Along the Texas Coast(2012-12-12) Ogle, Marcus 1982-Commonly referred to as ?red tide?, harmful algal blooms (HABs) formed by Karenia brevis occur frequently in the Gulf of Mexico (GOM). A bloom is defined as cell abundances >105 cells L-1. This thesis will focus primarily on Karenia brevis, formerly known as Gymnodinium breve, in the Gulf of Mexico. K. brevis is harmful because it produces brevetoxin, a ladder-frame polyether that acts as a potent neurotoxin in vertebrates. K. brevis commonly causes fish kills, respiratory irritation in humans, and Neurotoxic Shellfish Poisoning (NSP) if ingested. Blooms of K. brevis occur almost annually along the West Florida Shelf (WFS) in the late summer and early fall, when the coastal current is favorable for bloom initiation. Along the Texas-Louisiana shelf (TLS) however, blooms of K. brevis are infrequent and sporadic. While much is known of the blooms along the WFS due to their frequent presence, little is known of the mechanisms driving the blooms along the TLS due to their inconsistent presence. To understand the stochastic nature of HABs along the TLS, historical data of bloom occurrences from 1996 to present were compared with NOAA station PTAT2 wind, sea-level pressure, air and water temperature data and NCEP NARR-A sea-level pressure data. The difference in the monthly-mean along-shore component of the wind was statistically significant between bloom and non-bloom years in September (p<<0.001) and April (p=0.0015), with bloom years having a strong downcoast current. Monthly mean water temperature values yielded similar results between bloom and non-bloom years. Both March and September monthly-mean water temperature values were lower during non-bloom years with p-values of 0.01 and 0.048, respectively. These results suggest the possibly of forecasting for HABs along the TLS with currently measured, publicly available data.