Program Overview: Texas A&M, Fish Health and Disease Program

Haitham Mohammed

Department of Rangeland, Wildlife and Fisheries Management, College of Agriculture and Life Sciences, Texas A&M University, College Station, TX, 77843

From a Catfish Pond to the Ocean: Adaptive Responses of Aeromonas hydrophila to High Salinity

¹Ozgur Erdogan, ¹Noha I. Elbanna, ¹Ty J. Werdel, ¹Haitham H. Mohammed

¹Department of Rangeland, Wildlife and Fisheries Management, College of Agriculture and Life Sciences, Texas A&M University, College Station, TX, 77843

Aeromonas hydrophila, a Gram-negative, facultative anaerobic pathogen, is responsible for motile Aeromonas septicemia (MAS) in freshwater fish. The disease causes massive mortality and economic losses in both farmed and wild fish worldwide. The virulent pathotype of A. hydrophila (vAh) has severely impacted the U.S. catfish aquaculture, the largest aquaculture sector in the U.S., with annual losses of tens of millions of pounds of market-size fish. Despite the severity of the problem, the current preventive and control measures are either limited or ineffective.  According to an analysis of farm-level risk factors associated with MAS outbreaks in farmed catfish, salt (sodium chloride, NaCl) significantly reduces vAh infection rates. Therefore, in this study, we investigated the physiological and morphological responses of vAh under increasing osmotic stress, simulating a shift from freshwater pond environments to a more saline aquatic environment. We evaluated vAh’s growth, morphology, biofilm formation, survival, motility, and virulence in varying concentrations of sodium chloride in the culture medium. Our results showed that vAh has high salt tolerance. However, high salinity significantly decreased motility, biofilm formation, and bacterial growth. Moreover, high salinity has induced structural cell changes, suggesting that vAh may activate stress-response mechanisms to tolerate the higher osmotic stress levels. These findings suggest that the bacterium exhibits physiological alterations under elevated salinity, which potentially limits vAh virulence. At high salinity, vAh cells produce an elongated morphotype, which permits them to cope with adverse conditions, persist longer in the environment, and repopulate after stress when the conditions become favorable. Understanding both the susceptibility and adaptive capabilities of vAh under varying salinity levels provides valuable insights into pathogen ecology and supports the development of novel approaches to control MAS in aquaculture systems.

Silent Waters, Active Pathogens: Molecular Mapping of Columnaris Disease in Texas Fish

Md. Inja-Mamun Haque, Noha I. ElBanna, and Haitham H. Mohammed

Fish Health and Disease Laboratory, Department of Rangeland, Wildlife and Fisheries Management, Texas A&M University, College Station, TX-77843

Columnaris disease is a bacterial infection affecting freshwater fish, including numerous commercially and recreationally important species worldwide. In the southeastern United States, it is the most frequently reported disease in catfish farms, as documented by case summary reports from the Aquatic Research and Diagnostic Laboratory at Mississippi State University. The disease presents clinically as an external infection, characterized by gill necrosis, mouth rot, skin ulcerations, and fin erosion. The etiological agents are Gram-negative, yellow-pigmented bacteria collectively known as columnaris-causing bacteria (CCB), including four species, Flavobacterium columnare, F. covae, F. davisii, and F. Oreochromis, with different host associations. Current columnaris treatment strategies predominantly depend on using a combination of systemic antibiotics in fish feed and external disinfectants to manage morbidity and mortality. However, the continued use of these treatments, especially antibiotics, has led to the emergence of antibiotic-resistant strains, challenging sustainable disease management in aquaculture. Recent columnaris outbreaks in South Texas have significantly affected several key fish species, including hybrid catfish (Ictalurus furcatus × I. punctatus), hybrid tilapia (Oreochromis mossambicus × O. niloticus), and red drum (Sciaenops ocellatus), as well as sport fish such as largemouth bass (Micropterus salmoides), bluegill (Lepomis macrochirus), and crappie (Pomoxis spp.). The aim of this study was to identify the predominant bacterial species associated with columnaris disease outbreaks in Texas, assess their antibiotic resistance, characterize their biofilm-forming potential, evaluate their responses to disinfectants in both biofilm and planktonic forms, and analyze their phylogenetic relationships to uncover strain-level diversity. Putative CCB isolates (n = 40) were recovered from diseased fish submitted to the Fish Health and Disease Laboratory at Texas A&M University during outbreaks in 2023-2024. Following established microbiological protocols and subsequent DNA extraction procedures, a multiplex polymerase chain reaction (PCR) assay was utilized, employing four reference CCB strains provided by USDA-ARS in Auburn, Alabama, to ascertain species-level identity. Among the isolates examined, F. covae was the most prevalent, accounting for 60% of the total (n = 24), followed by F. davisii, which constituted 40% (n = 16). No isolates were identified as F. columnare or F. oreochromis. Antibiotic susceptibility testing of 13 isolates was performed to FDA-approved antibiotics for the treatment of food fish following the standard Kirby-Bauer disc diffusion method. All isolates were sensitive to oxytetracycline and florfenicol, with F. covae showing an average inhibition zone of 36.09 ± 2.67 mm and 45.03 ± 0.18 mm, respectively. F. davisii exhibited smaller zones at 32.83 ± 0.73 mm and 42.72 ± 0.09 mm. In contrast, Romet-30 resistance was widespread, as 77% (10 out of 13) of isolates were resistant. F. covae had minimal inhibition (5.75 ± 0.58 mm), while F. davisii showed complete resistance to Romet-30. No significant differences were observed between F. covae and F. davisii regarding their ability to form biofilms or grow as planktonic cells. The impact of typical disinfectants on both biofilm and planktonic forms was explored and will be discussed. Phylogenetic analysis confirmed the evolutionary positioning of the collected isolates to reference CCB strains, offering insights into diversity at the strain level. The findings highlight the predominance of F. covae and F. davisii in the recent columnaris outbreaks in Texas, underscoring the necessity for alternative therapeutics aimed at these particular species to enhance disease control in the future.

Formalin-Inactivated Vaccine Candidates Against Pathogenic Vibrios in Gilthead Seabream (Sparus aurata)

1Noha I. ElBanna, 2Alaa E. Eissa, 3Salah M. Aly

1Aquaculture Diseases Control Department, Fish Farming & Technology Institute, Suez Canal University, Ismailia, Egypt, 41522; 2Aquatic Animal Medicine and Management Department, College of Veterinary Medicine, Cairo University, Giza, Egypt, 12613; 3Pathology Department, College of Veterinary Medicine, Suez Canal University, Ismailia, Egypt,41522.

Vibriosis is one of the most serious and economically devastating bacterial diseases impacting global mariculture. With the rising challenge of antimicrobial resistance and the demand for sustainable aquaculture practices, vaccination offers a promising biosecurity approach. This study investigated the protective efficiencies and the potential immune mechanisms of vibrio monovalent and polyvalent autogenous formalin-inactivated whole-cell bacterins (FKC) in cultured Gilthead seabream (Sparus aurata) in Egypt. Autogenous Vibrio FKC vaccines were intraperitoneally administered at 0.1 ml with a final concentration of 1×10⁸ CFU/mL.  One hundred and forty Gilthead seabream were randomly divided into five equal groups and allocated into twenty glass tanks (4 tanks/ group, 7 fish /tank). The groups were assigned as follows: the 1st group received a monovalent Vibrio alginolyticus vaccine, while the 2nd group was vaccinated with a monovalent Vibrio parahaemolyticus O11: K40 vaccine. The 3rd group was immunized with a polyvalent FKC bacterin containing V. parahaemolyticus O11: K40 and V. alginolyticus, whereas the 4th group received a polyvalent vaccine comprising V. parahaemolyticus O3: K6 and V. alginolyticus. The 5th group, the control, was injected with sterile phosphate-buffered saline at the same dose and via the same route. The fish were raised under a 12:12 light/dark cycle, with seawater maintained at 20–22°C and 35 ppt salinity.  Two months after immunization, the challenge infection with homologous virulent strains showed protection rates of 100% and 83.3% for monovalent vaccines (V. alginolyticus and V. parahaemolyticus O11:K40), and 91.75% and 75% for polyvalent vaccines (V. parahaemolyticus O11:K40 & V. alginolyticus and V. parahaemolyticus O3:K6 & V. alginolyticus).  The tested vaccine preparations significantly increased (P < 0.05) the agglutination antibody titer, phagocytic activity, and respiratory burst activity when compared to the non-vaccinated control group. Vaccinated fish histopathological examination revealed significantly increasing size and numbers of splenic and renal Melano-macrophage centers. These responses were significantly higher (P < 0.05) than non-immunized fish throughout the 4 months.  Our findings support the potential of inactivated autogenous Vibrio vaccines to provoke protection against vibriosis in Gilthead seabream cultured in Egypt, they were superior in the monovalent FKC V. alginolyticus vaccine and polyvalent FKC of V. parahaemolyticus O11: K40, with V. alginolyticus vaccine that could be a viable method for the prevention and control of vibriosis in marine aquaculture.