Analysis and cytologic characterization of hemocytes from freshwater mussels ( Quadrula sp.).

Background: Freshwater mussels are among the most endangered taxa in North America and minimally invasive techniques to evaluate their health are needed. Objective: The objective of this study was to develop a standardized approach for identifying and enumerating the cellular components of freshwater mussel hemolymph. Methods: Hemocyte clumping, total hemocyte count, and hemocyte morphology were compared in untreated hemolymph or hemolymph treated with formalin, sodium citrate, sodium heparin, EDTA, water, orl-cysteine. Morphology was then used to categorize hemocytes and perform a 100-cell differential. Results: Treatment with formalin or >25 mg/mLl-cysteine reduced hemocyte clumping, although only formalin significantly increased the total hemocyte count. However, formalin also induced crenation that impaired hemocyte identification. Both EDTA and sodium citrate-induced hemocyte degranulation while sodium citrate and >40 mg/mLl-cysteine-induced cell lysis. Hemocytes could be categorized into 2 groups of granulocytes (eosinophilic or basophilic) and 2 groups of agranulocytes (large or small) for performing a cytologic differential. The differential was not significantly altered by anticoagulant treatments providing cell morphology was adequate for obtaining a differential. Eosinophilic granulocytes predominated (59%) with fewer large agranulocytes (27%) and basophilic granulocytes (13%). Small agranulocytes comprised 2% of the total population. Conclusions: No single treatment provided an optimal method to evaluate freshwater mussel hemolymph. Maximal hemocyte counts were obtained following formalin treatment.l-cysteine reduced clumping and maintained hemocyte morphology for performing a cytologic differential. These techniques provide a standardized approach for the hematologic evaluation of freshwater mussels. [ABSTRACT FROM AUTHOR]