Abstract
The American horseshoe crab (Limulus polyphemus) (HSC) is commonly managed in public aquaria. Despite many institutions housing them, there is a dearth of information in the literature regarding clinical health assessment. In recent HSC hemolymph studies at the National Aquarium, hemocyte and biochemical analyses were performed for two separate populations including free-ranging HSC in Delaware Bay during the spawning season, and for a longitudinal study of animals collected from the bay and followed through managed care. Morbidity and mortality of animals over the first 6 months of animals arriving at the institution were associated with initial threshold values of three analytes. Thresholds where 100% of the animals above the threshold survived and 75% of those below died were: total protein (TP) ≤3.5 g/dL, glucose (GLU) ≤25 mg/dL, and copper (Cu) ≤59 ppm (unpublished data). With this information, a prospective study was initiated to evaluate serial physical examinations and hemolymph assays monthly for seven months. HSC with low TP, GLU, and Cu values were expected to have increased morbidity and mortality.
Monthly assessments included physical exams utilizing a scoring system for behavior and external abnormalities, heart rate (HR), weight, morphometrics (prosomal width and interocular distance), and hemolymph analysis. Hemolymph analysis included total hemocyte count, plasma alanine aminotransferase (ALT), aspartate aminotransferase (AST), albumin, alkaline phosphatase (ALP), amylase, calcium, chloride, creatine kinase (CK), Cu, creatinine, GLU, lipase, magnesium, osmolality, inorganic phosphorus, potassium, sodium, TP, urea nitrogen, and uric acid. Two tank-side point of care devices were utilized to compare glucose (HemoCue 201, HemoCue America, Brea, CA, USA) and total dissolved solids by refractometer and to compare with spectrophotometer analysis for glucose and total protein. Pantone® color strips were compared to hemolymph color in order to determine if certain color codes correlated with specific Cu ranges.
Over the duration of the study, clinical health assessments and clinicopathologic data for the population studied remained largely stable on an individual basis. Adverse effects from monthly handling for examination and hemolymph draw were not appreciable in the majority of the population. A single individual was removed from the study due to a friable arthrodial membrane and spontaneous hemorrhage upon ventroflexion. The only physical factor that varied over time points in all examined HSC was heart rate. Over the course of the study, approximately one third of the studied population developed minor increases in physical exam scores relative to their starting scores, reflective of a worsened external appearance or behavior. All HSC showed an increase in Cu values at month 2. The other analytes of initial interest showed minimal variation over time. These findings support the intra-individual stability of these HSC hemolymph analytes in managed care situations and lay the groundwork for further exploration of hemolymph dyscrasias seen in various illnesses. This study also provides a framework for assessing clinical appearance that may enable greater objectivity in clinicians and husbandry staff who are managing HSC populations in research, zoological, or other scenarios where tracking changes in HSC health over time may be necessary.
Acknowledgements
The authors would like to thank the aquarists at National Aquarium that have been involved with the care of these animals and the time dedicated to this project.
*Presenting author
+Student presenter