Quantification of Perkinsus marinus in the eastern oyster Crassostreavirginica using modern stereological techniques

摘要

Dermo disease, caused by the protozoan parasites Perkinsus spp., is responsible for high mortalities of bivalve molluscs worldwide. In order to improve the knowledge of the pathogenesis of Dermo, accurate techniques to estimate the number of parasites in tissue sections are required. This study is aimed to quantify the number and tissue distribution of different stages of P. marinus in a natural population of Crassostrea virginica from Wickford (Rhode Island, US) by the application of modern stereology and immunohistochemistry. Mean total number of trophozoites in eight oysters collected in July 2005, of mean shell length (mean plus or minus SD) of 101.2 plus or minus 5.2 mm, were (mean plus or minus SE) 11.80 plus or minus 3.91 million and 11.55 plus or minus 3.88 million for the optical disector and optical fraction-ator methods respectively. The mean empirical error between both stereological approaches was 3.8 plus or minus 1.0%. Trophozoites were detected intracellularly in the following tissues: intestine (30.1%), Leydig tissue (21.3%), hemocytes (14.9%), digestive gland (11.4%), gills (6.1%), connective tissues (except Leydig and mantle) (5.7%), gonads (4.1%), palps (2.2%), muscle (1.9%), mantle connective (0.8%), pericardium (0.7%), mantle epithelium (0.1%), and heart (0.1%). The remaining 0.6% of trophozoites were found extracellularly throughout different tissues. Percentages of trophozoite stages were (mean plus or minus SE): large, log-phase trophonts (i.e. signet-rings): 97.0 plus or minus 1.2%; meronts: 2.0 plus or minus 0.9%; clusters of small, log-phase trophonts (i.e. merozoites): 1.0 plus or minus 0.5%. These techniques could be useful to follow parasite distribution and progression in experimental infections, and further explore mechanisms of dermo pathogenesis.