An improved microscopic method for the diagnosis of Enterocytozoon

hepatopenaei in shrimp farms

Hepatopancreatic microsporidiosis caused by Enterocytozoon hepatopenaei (abbreviated as

EHP), is an emerging parasite of penaeid shrimp in several shrimp farming countries in Asia.

Light microscopic examination of the stained clinical smear is an inexpensive method of

diagnosing microsporidian infections even though it does not allow species level identification.

An improved microscopic method for the diagnosis of EHP in shrimp was standardized by

employing conventional concentration techniques for parasitic stages by sedimentation

(formalin-diethyl ether) or flotation (Sheather’s sugar solution) followed by a microsporidia-

specific staining technique by modified trichrome stain (Ryan-Blue formulation). The method

was found ideal for biological samples such as faeces, hepatopancreas and pond sediments.

Ryan-Blue stains the microsporidian spore pinkish-red due to dye content chromotrope 2R and

provides a good differentiation from the lightly stained bluish background debris materials by

aniline blue, so that the ovoid and refractile spores stand out for easy visualization by light

microscopy. The method showed varying sensitivity depending upon the level of infection in

shrimp.

Figure 1. Spores of Enterocytozoon hepatopenaei from shrimp faeces after sedimentation, stained with Chromotrope 2R stain

K. P. JithendranPrincipal scientist,

Central Institute of Brackishwater Aquaculture, Chennai-600 028

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