Location: Aquatic Animal Health Research
2020 Annual Report
Objectives
The additional funds will enhance research on current Objective 2 and two new objectives that are presently going through OSQR review at this time which are as follows:
Current Objective 2: Develop vaccines and probiotics that provide protection against bacterial and parasitic pathogens and identify mechanisms of immunity by identifying and characterizing host mechanisms responding to infection and host-pathogen interactions that can be used to develop approaches that reduce losses to disease.
New Objective 1: Identify virulence factors critical for pathogenesis of major catfish pathogens to guide the development of novel and cost-effective disease interventions.
New Objective 2: Improve prevention and control strategies for bacterial and parasitic diseases of catfish and shrimp.
Objective 1: Identify and characterize genetic diversity, mechanisms of pathogenesis and virulence factors in Flavobacterium (F.) columnare.
Subobjective 1.A.: Comparative bacterial genome analysis of Flavobacterium columnare isolates of different genetic types and virulence.
Subobjective 1.B.: Molecular basis of lipopolysaccharide (LPS) and capsular polysaccharide (CPS) antigenic diversity in Flavobacterium columnare.
Subobjective 1.C.: Characterize the growth and survival of F. columnare in fish mucus.
Objective 2: Develop vaccines and probiotics that provide protection against bacterial and parasitic pathogens and identify mechanisms of immunity by identifying and characterizing host mechanisms responding to infection and host-pathogen interactions that can be used to develop approaches that reduce losses to disease.
Subobjective 2.A.: Chemical mutagenesis of Flavobacterium columnare to modify the capsular polysaccharide (CPS) to develop attenuated vaccines.
Subobjective 2.B.: Evaluate the protective effect of a DNA vaccine encoding Ich immobilization antigens to protect catfish against Ich.
Subobjective 2.C.: Develop a waterborne challenge model and recombinant protein vaccine to protect channel catfish against virulent Aeromonas hydrophila.
Approach
Catfish and tilapia farmers continue to identify disease as a major problem in their industries. For instance, columnaris disease, caused by Flavobacterium (F.) columnare, is one of the top two diseases diagnosed in the industry. Since 2009, a virulent strain of Aeromonas (A.) hydrophila has greatly impacted the catfish industry and resulted in losses of greater than 12 million U.S. dollars. Ichthyophthirius (I.) multifiliis, the parasite that causes Ich, is responsible for annual losses of 1.2 million U.S. dollars to the catfish industry alone. An increased understanding of the pathogen, host responses to the pathogen, and host-pathogen interactions is necessary for disease prevention and control. This in-house project will expand our knowledge of these and will use new and existing knowledge to develop approaches to reduce disease losses in catfish and tilapia aquaculture. Development of disease prevention strategies will increase the profitability and sustainability of these important aquaculture industries.
Objective 1 recognizes that although columnaris disease has been intensely studied in the past, important questions concerning genetic diversity of isolates impacting aquaculture and mechanisms of pathogenesis have newly emerged. A greater understanding of these factors will enhance our ability to improve existing and develop new prevention strategies practical for use in the catfish and tilapia industries. Research conducted in this objective will utilize comparative genome analyses to identify genetic similarities and differences among F. columnare isolates of different genomovars (genetic types) and will correlate the genetic differences with variation in virulence. The genome sequences will be exploited to determine the molecular basis of lipopolysaccharide and capsular polysaccharide antigenic diversity in F. columnare. The growth of F. columnare in catfish mucus will be used as a model to determine the proteomic changes that occur in F. columnare during the colonization of catfish and how these changes are involved in virulence.
Objective 2 acknowledges that even though there is a commercially available vaccine for F. columnare and experimental vaccines exist for other bacterial and parasite pathogens, there is a need to develop improved disease prevention methods and identify the mechanisms responsible for protective immunity. Research conducted will utilize chemical mutagenesis to modify the capsular polysaccharide of F. columnare to develop more effective attenuated vaccines. A DNA vaccine for I. multifiliis will be developed based on proteins of the parasite that have been previously demonstrated to be protective. A reproducible waterborne challenge model for virulent A. hydrophila will be developed and will allow for more effective testing of treatment or prevention strategies. A recombinant protein vaccine for A. hydrophila will be developed based on secreted proteins of the bacterium that are identified as protective. This research will increase our understanding of the host immune responses against pathogens and will develop improved and new vaccines for prevention of disease in catfish and tilapia aquaculture.
Progress Report
This progress report represents the final report for this project as it termed during this annual report cycle. Replaced by new project #6010-32000-027-00D, "Integrated Research to Improve Aquatic Animal Health in Warmwater Aquaculture." Please refer to new project for additional information.
Columnaris disease, caused by Flavobacterium (F.) columnare, is an important bacterial pathogen impacting the catfish industry. Research completed under this project advanced our understanding of the genetic diversity, mechanism of pathogenesis and virulence factors in F. columnare. Research established the existence of four phylogenetically distinct genetic groups in the species F. columnare with biological relevance. A multiplex polymerase chain reaction (PCR) assay was developed to assign an unknown isolate to genetic group. The PCR assay was used to determine the genetic group of F. columnare involved in columnaris disease cases throughout the catfish industry. The results demonstrated that isolates belonging to genetic groups 1, 2, and 3 are involved in catfish disease cases, but genetic group 2 isolates appear to be the most predominant type impacting the catfish industry. Research was conducted to determine the influence of catfish mucus on the virulence of F. columnare. Research demonstrated that the bacterium can replicate and survive for up to 3 months in water containing catfish mucus and while growing in this environment the bacterium increased the production of proteases likely responsible for the greater capacity to cause disease in catfish.
Outbreaks of virulent Aeromonas (A.) hydrophila have cost U.S. catfish aquaculture an estimated $60-70 million. A reproducible immersion-based challenge model was developed which allowed for conducting laboratory experiments to determine the pathogenic mechanisms of A. hydrophila and identify potential methods for treatment and prevention. The role of iron and organic matter, such as chitin, in virulence was elucidated, and research demonstrated that catfish feeding practices impact the susceptibility of fish to virulent A. hydrophila. Research revealed that kaolin clay significantly blocked the movement and binding ability of the bacterium to catfish mucus, and laboratory studies demonstrated that addition of the clay to water protects catfish against infection. A killed A. hydrophila vaccine, delivered via immersion in the water, was developed and provided a high level of protective immunity.
Ichthyophthirius multifiliis (Ich) is a parasite of fish and causes severe losses to aquaculture industries worldwide. Research was conducted to characterize the immune responses of fish to the pathogen and results demonstrated the importance of specific antibodies for protective immunity. A deoxyribonucleic acid (DNA) vaccine encoding Ich antigens was developed and tested for the ability to provide protection from parasite challenge. The results demonstrated that vaccination of catfish with the DNA vaccine triggered the immune response and provided partial protective immunity, providing proof of concept for this vaccine as a preventative measure for Ich.
Inland, low salinity shrimp farmers in west Alabama have recently reported abnormally low survivals at harvest. Reduced survivals have also been reported by farms in Florida and Texas. Research was conducted to compare the performance of shrimp from different sources at three different farms. The results demonstrated that while the source of shrimp did influence survival on one farm, overall performance was acceptable from all three sources.
Research demonstrated that resistance of Nile tilapia to commercially important pathogens, such as Streptococcus (S.) iniae and S. agalactiae, is a heritable trait and can be markedly improved through selective breeding.
Accomplishments
1. The severity of motile aeromonas septicemia caused by virulent aeromonas hydrophila in channel catfish is influenced by nutrients and microbes in water. Outbreaks of motile Aeromonas septicemia (MAS) disease, caused by virulent Aeromonas hydrophila (vAh), has severely impacted catfish farming in the southeastern United States since 2009. The development of control and prevention strategies has been hindered by a lack of understanding for the conditions that trigger disease in the field. ARS researchers in Auburn, Alabama, assessed the effect of nutrients and selected microbes on the growth of A. hydrophila and severity of disease in channel catfish. Laboratory experiments demonstrated that fish feed can serve as a source of nutrients for vAh as indicated by vigorous growth of the bacterium in water containing fish feed; and these bacteria were highly virulent to catfish. The research also found that a potential probiotic bacterium could protect fish against vAh infection, resulting in approximately 54% lower fish mortality. This research sheds light on the importance of optimizing feeding practices in intensive catfish farming.
Review Publications
Mabrok, M., Putita, C., Lafrentz, B.R., Kayansamruaj, P., Dong, H., Rodkhum, C. 2020. Development of a species-specific polymerase chain reaction for highly sensitive detection of Flavobacterium columnare targeting chondroitin AC lyase gene. Aquaculture. 521(2020):734597. https://doi.org/10.1016/j.aquaculture.2019.734597.
Shoemaker, C.A., Lafrentz, B.R., Garcia, J.C., Xu, D., Beck, B.H. 2020. Streptococcosis in fish. In: AFS-FHS (American Fisheries Society-Fish Health Section). FHS Blue Book: Suggested Procedures for the Detection and Identification of Certain Finfish and Shellfish Pathogens. AFS-FHS. Bethesda, Maryland. Chapter 1.3.3.
Xu, D., Zhang, D., Shoemaker, C.A., Beck, B.H. 2019. Immune response of channel catfish (Ictalurus punctatus) against Ichthyophthirius multifiliis post vaccination using DNA vaccines encoding immobilization antigens. Fish and Shellfish Immunology. https://doi.org/10.1016/j.fsi.2019.08.071.
Zhang, D., Xu, D., Shoemaker, C.A., Beck, B.H. 2020. The severity of Motile Aeromonas septicemia caused by virulent Aeromonas hydrophila in channel catfish is influenced by nutrients and microbes in water. Aquaculture. https://doi.org/10.1016/j.aquaculture.2019.734898.
Qiao, G., Xu, C., Sun, Q., Xu, D., Zhang, M., Chen, P., Li, Q. 2019. Effects of dietary poly-beta-hyroxybutyrate supplementation on the growth, immune response and intestinal microbiota of soiny mullet (Liza haematocheila). Fish and Shellfish Immunology. 91:251-263.
