THE EFFECTS OF HYPOXIA AND HYPERCAPNIA ON DISEASE SUSCEPTIBILITY TO VIBRIO PARAHAEMOLYTICUS IN THE PENAEID SHRIMP, LITOPENAEUS VANNAMEI, AND THE GRASS SHRIMP, PALAEMONETES PUGIO

Christina Marie Mikulski

A thesis submitted to the Graduate Faculty of the University of Charleston, South Carolina in partial fulfillment of the requirements for the Degree of Master of Science

ABSTRACT
Estuarine organisms routinely encounter fluctuations in dissolved oxygen, carbon dioxide, and pH, which can vary both seasonally and diurnally. Although hypoxia occurs naturally, eutrophication further exacerbates its severity and duration. Research has shown that environmental stress, such as hypoxia, affects the immune response of invertebrates and vertebrates and has been linked to increased disease incidence. This research investigated the effects of hypoxia, hypercapnia, and low pH on disease susceptibility in both penaeid and palaemonid shrimp. Specific pathogen free juvenile shrimp, Litopenaeus vannamei, were acquired from mariculture farms along the South Carolina coast. Grass shrimp, Palaemonetes pugio, were collected from local tidal creeks in Charleston. Both species were acclimated to test conditions in the laboratory. Animals were injected intramuscularly with a previously determined LD₅₀ dose of a known pathogenic strain of Vibrio parahaemolyticus. Disease susceptibility was measured for shrimp held under normoxia (Po₂ = 150-155 torr, Pco₂ = 0.23, pH = 7.6-8.0 for L. vannamei, Po₂ = 150-155 torr, Pco₂ = 0.23 torr, pH = 8.0-8.2 for P. pugio) and two levels of hypoxia in filtered artificial seawater kept at 30 ppt. The penaeid shrimp were challenged under normocapnic hypoxia (Po₂ = 45 torr, Pco₂ = 0.23 torr, pH = 7.8-8.1) and hypercapnic hypoxia (Po₂ = 30 torr, Pco₂ = 15.2 torr, pH = 6.8-7.0). Grass shrimp were challenged under two levels of hypercapnic hypoxia (Po₂ = 45 torr and 30 torr, Pco₂ = 15.2 torr, pH = 6.7-7.0). Both the juvenile L. vannamei and the P. pugio held under hypercapnic hypoxia at 30 torr oxygen displayed significantly lower 48 hour survival (15.7% and 3.1%, respectively) than animals held in normoxic water (28.7% and 29.4%, respectively). The increase in disease susceptibility was not due to enhanced bacterial growth under these conditions. There was no significant decrease in survival in L. vannamei under normocapnic hypoxia at 45 torr oxygen or in P. pugio under hypercapnic hypoxia at 45 torr oxygen.

Total hemocyte count was measured in adult L. vannamei held under normoxia and hypercapnic hypoxia for 48 hours without bacterial challenge. Total hemocyte count was significantly reduced in L. vannamei held under hypercapnic hypoxia (Po₂ = 30 torr, Pco₂ = 15.2 torr, pH = 6.8-7.0) when compared to shrimp held under normoxia (Po₂= 150-155 torr, Pco₂= 0.23 torr, pH = 7.6-8.0) at 4, 8 16, and 24 hours (39.3%, 65.9%, 65.7%, and 59.6% of normoxic values, respectively). Total hemocyte count was not significantly different between the two treatments at 48 hours. These results show that hypercapnic hypoxia at 30 torr oxygen, 15.2 torr CO₂, and a pH range of 6.8-7.0 decreases disease resistance in both L. vannamei and P. pugio and decreases total hemocyte count in L. vannamei. This may have important implications regarding the health of these organisms when they are exposed to hypercapnic hypoxia in both the natural environment and in aquaculture.

Published:
Mikulski, C. M., L. E. Burnett, and K. G. Burnett.  2000.  The Effects of Hypercapnic Hypoxia on the Survival of Shrimp Challenged with Vibrio parahaemolyticus.  J. Shellfish Res.19:301-311.