The Use of Probiotics in Pacific White Shrimp (litopenaeus Vannamei) Aquaculture to Increase Growth and Survival and To Reduce Gene Expression of Heat Shock Protein 70 While Subjected to Acute Thermal Stress

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THE USE OF PROBIOTICS IN PACIFIC WHITE SHRIMP (LITOPENAEUS VANNAMEI) AQUACULTURE TO INCREASE GROWTH AND SURVIVAL AND TO REDUCE GENE EXPRESSION OF HEAT SHOCK PROTEIN 70 WHILE SUBJECTED TO ACUTE THERMAL STRESS Jasmine Smalls Faculty Advisor: Dr. Dennis McIntosh Abstract Shrimp farmers face numerous challenges associated with the vulnerability of shrimp to disease and environmental stress. Pacific white shrimp (Litopenaeus vannamei) is one of the most commonly produced and high yielding cultured shrimp species. Growth and survival are two factors that dictate the overall production; probiotics which are ‘beneficial bacteria,’ can help improve both. Moreover, extreme temperatures are also a concern for shrimp production and can negatively impact growth and survival. Elevated temperatures can trigger the expression of Heat Shock Proteins (HSPs) in shrimp, which aid in repairing denatured proteins. To assess the impact of probiotics on shrimp, two experiments were conducted. The first experiment, which consisted of three trials, analyzed the effect of probiotics on growth and survival of post-larvae (PL) shrimp. Initially, only one trial was intended, however, because there were setbacks, two additional trials were conducted. After multiple weeks, the remaining PL shrimp from each group were counted to ascertain survival, and group weighed to calculate growth. For trial one, the Media Control (MC) appeared to produce the fastest growth followed by ISO 11 and ISO 12. PL in the MC also appeared to have the highest survival at harvest. However, a mortality event was experienced during the first trial due to various confounding factors including excessive temperatures, feeding rates, and stocking density, which prompted the initiation of trial two. For trial two, ISO 5 seemed to have the fastest growth followed by the MC and ISO 11. Survival for the Negative Control (NC), along with ISO 11 and ISO 12 were higher at the time of harvest compared to the MC and ISO 5. However, system design and sample size, in addition to other limiting factors, which influenced shrimp performance undermine the validity of the findings from trial two, so a third trial was planned. During the third trial, probiotics did not affect growth. At the time of harvest, it was found that nearly all containers contained the same number of shrimp, as nearly half of the shrimp from all experimental containers died during the first few days of the study. Due to the errors and issues experienced during all three trials, a definitive conclusion cannot be made regarding the true effectiveness of the applied probiotics on shrimp performance. In the second experiment, the objective was to determine if any of the selected probiotics effect gene expression of HSPs in juvenile shrimp. Shrimp were divided into groups and assigned to one of 12 ((3 probiotics + 1 control) x 3 temperatures)) treatments. Shrimp were pre-exposed to assigned probiotics for six days prior to the planned temperature stress event. Shrimp were sampled in two phases, exposure and recovery, to measure HSP expression with reverse transcriptase PCR. Results were compared across treatments to determine which probiotic had the greatest effect on shrimp. During this study, the intention was to collect the hepatopancreas, because it is this organ which highly expresses and regulates gene function in shrimp. However, due to a mistake in organ identification, the foregut was extracted instead. In addition to the foregut, a section of the abdominal muscle was also collected. Also, due to improper handling, and contamination of tissue samples, there was a statistically significant difference found in the quality of RNA between the NC and ISO 11 treatments at different temperatures across all sampling periods. Similarly, there was a statistically significant difference found in the expression of the HSP 70 between the NC and ISO 11 treatment at different temperatures across all sampling periods, as a possible result of thermal stress and high level of contamination. Due to the high level of contamination that was experienced during this study, a definitive conclusion cannot be made regarding the true effectiveness of the applied probiotics on mitigating shrimp stress.

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