Location: Harry K. Dupree Stuttgart National Aquaculture Research Cntr
2017 Annual Report
Objectives
Objective 1. Characterize the cellular and molecular immune responses governing resistance/susceptibility to infectious disease (Flavobacterium columnare), and develop strategies to enhance immune protection through immunomodulatory compounds, nutritional status, or water chemistry.
Subobjective 1A. Describe the cellular, transcriptional, and pathological responses of mucosal tissues in healthy and Flavobacterium columnare-challenged fish.
Subobjective 1B. Investigate the effects of copper sulfate, short-term feed deprivation, and water chemistry on the mucosal immune system.
Objective 2. Assess microbial community diversity, prevalence, and distribution under homeostatic conditions, after columnaris disease challenge, and following environmental perturbation.
Subobjective 2A. Profile the composition and population dynamics of microbes that reside on the skin and gill of healthy and columnaris-challenged fish.
Subobjective 2B. Investigate the effects of copper sulfate, short-term feed deprivation, and water chemistry on microbial community composition.
Approach
In Objective 1 our research team will utilize next-generation sequencing to establish temporal profiles of transcriptional responses to Flavobacterium (F.) columnare in gill and skin; use flow cytometry to monitor the type and abundance of immune cells trafficking to the gill of challenged fish; and characterize the histopathological changes to an F. columnare challenge and draw parallels to the shifts in immune-related transcripts and cell phenotypes. Studies will also examine the transcriptional and immunological consequences to alterations in water chemistry and to chemical compounds such as copper sulfate. In Objective 2 the core microbiota associated with mucosa in healthy fish and F. columnare infected fish will be characterized using using high-throughput 16S v4 amplicon sequencing. This work will identify the core microbial communities commonly residing in the mucosal barriers of both warmwater bass and channel catfish; vital information towards identifying bacterial species that can be exploited to improve fish health.
Progress Report
ARS scientists characterized the Flavobacterium (F.) columnare virulence mechanism of biofilm formation, which is a critical step of infectivity. Biofilms are an important stage of pathogenesis of the host, which requires a sustained host: pathogen interaction at mucosal sites (gill, skin). Studies to better understand biofilm formation and other virulence mechanisms under different in vitro environmental conditions continue to be conducted. This work will benefit in the development of new vaccines and novel therapies to treat disease.
ARS scientists continue to study the processes by which, mucosal adaptive immune responses (skin) are activated through the use of different immunogens. These works seek to characterize the humoral immune response under various environmental conditions, including those represented in industry, to produce efficient and effective therapies for the treatment of industry relevant diseases.
Copper sulfate toxicity studies were continued on various species of fish in five reconstituted waters ranging from very soft to very hard. These tests determine the LC50 (acute toxicity), as well as the range of useable reconstituted waters, and the highest concentration of copper that does not cause mortality. The work being conducted by ARS scientists is vital for industry farmers to understand the amount copper sulfate they can use to treat their fish. This is the first comprehensive research to demonstrate safety of copper across multiple finfish species under similar culture conditions.
ARS scientists initiated a collaboration with Kentucky State University (Frankfort, Kentucky) on the toxicity of copper sulfate to largemouth bass and the transfer of their innate resistance to copper toxicity to channel catfish. This is the first study to establish that innate resistance of a finfish species to copper toxicity can be transferred.
We completed research studies with USDA/ARS (Leetown, West Virginia) on the effectiveness and safety of copper sulfate to control fungus on rainbow trout eggs. Industry partners have expressed interest in this cheap and effective means to mitigate losses due to egg fungus.
We continued studies to determine the minimum amount of calcium in reconstituted water that allows Flavobacterium columnare, the bacteria responsible for columnaris disease, to attach to gills of channel catfish. We are collaborating with researchers at the USDA/ARS lab (Stoneville, Mississippi). This research will establish an understanding of the physiology of columnaris disease and help us find ways to mitigate it.
We are collaborating with scientists in Denmark, Germany and the U.S., to establish the importance of the potent disinfectant peracetic acid to the global aquaculture industry. We continue our research at Bowling Green State University (Bowling Green, Ohio) and the Freshwater Institute (Shepherdstown, West Virginia) to quantify the amount of aquatic fungus in recirculating aquaculture systems and to treat these systems with peracetic acid to eliminate these pathogens. After evaluating our research, the major manufacturer of peracetic acid in the U.S. has contacted us expressing interest in introducing this product to the aquaculture industry.
A collaboration was initiated with our lab by the Department of Mycology, Real Jardín Botánico (Madrid, Spain) due to our stature in the area of Saprolegnia (aquatic fungus) research. This project is focused on the identification of genotypes and establishing the population structure of Saprolegnia parasitica world-wide. This is the first study to compile a global database of this detrimental aquatic pathogen.
We initiated a collaboration with USDA/ARS (Franklin, Maine) and the Freshwater Institute (Shepherdstown, West Virginia) to evaluate the safety of peracetic acid to Atlantic salmon and salmon eggs. One of the most commercially important food fish in the world, salmon hatcheries are plagued by aquatic fungi, and there is a paucity of available treatment methods. Salmon producers are anxiously searching for a replacement for formalin, which many countries are in the process of restricting due to its carcinogenicity.
Accomplishments
1. Development of a recombinant protein vaccine to protect catfish against columnaris disease. Flavobacterium (F.) columnare is the causative agent of columnaris disease which severely impacts channel catfish production in the United States. ARS researchers in Stuttgart, Arkansas had previously identified F. columnare proteins which activate the adaptive immune response in fish. A new recombinant protein vaccine has been developed resulting in excellent immune protection against columnaris disease, possibly improving resistance to the disease. ARS researchers in Stuttgart, Arkansas have shown that during experimental tank studies that the use of the recombinant protein as an immunogen provided significant protection when subjected to a laboratory disease challenge. The ARS scientists will build on this research and continue to both optimize and develop effective recombinant protein-based vaccines to prevent columnaris disease and help catfish producers reduce overall production losses, potentially increasing profitability, and thereby allowing for industry expansion.
2. Determined treatment of copper sulfate to control fungus on rainbow trout eggs. This research addresses egg mortality resulting from Saprolegnia (fungus) in rainbow trout hatcheries. A range-finding study by ARS researchers in Stuttgart, Arkansas in collaboration with ARS researchers in Leetown, West Virginia determined that treating rainbow trout eggs daily with 10ppm copper sulfate until eyes develop will prevent fungus from growing and destroying the eggs. As the 2nd largest food-fish production species in the U.S., industry partners have communicated the need for this inexpensive and effective means to reduce losses due to egg fungus. This accomplishment will provide direct benefits through increased survival rates in rainbow trout hatcheries which will allow more fry to be produced, and potentially increase profitability.
Review Publications
Liu, D., Behrens, S., Pedersen, L., Straus, D.L., Meinelt, T. 2016. Peracetic acid is an optimal disinfectant for fish-microalgae integrated multi-trophic aquaculture systems. Aquaculture Report. 4:136-142. Available:http://www.sciencedirect.com/science/article/pii/S2352513416300977.
Liu, D., Straus, D.L., Pedersen, L., Meinelt, T. 2017. Pulse versus continuous peracetic acid applications: effects on Rainbow trout performance, biofilm formation and water quality. Aquacultural Engineering. 77:72-79.
Liu, D., Pedersen, L., Straus, D.L., Kloas, W., Meinelt, T. 2017. Alternative prophylaxis/disinfection in aquaculture - Adaptable stress induced by peracetic acid at low concentration and its application strategy in RAS. Aquaculture. 474:82-85.
Lange, M.D., Farmer, B.D., Declercq, A.M., Peatman, E., Decostere, A., Beck, B.H. 2017. Sickeningly sweet: L-rhamnose stimulates Flavobacterium columnare biofilm formation and virulence. Journal of Fish Diseases. doi:10.1111/jfd.12629.
