Acid-Fast Stain Identifies Schistosoma Eggs

Schistosomiasis, also known as snail fever, is a disease caused by parasitic flatworms called schistosomes. The urinary tract or the intestines may be infected and signs and symptoms may include abdominal pain, diarrhea, bloody stool, or blood in the urine.

Microscopic identification of eggs in stool or urine is the most practical method for diagnosis. Stool examination is performed when infection with Schistosoma mansoni or S. japonicum is suspected, and urine examination should be performed if S. haematobium is suspected. Eggs can be present in the stool in infections with all Schistosoma species.

Scientists at the University of Lisbon examined whether the Ziehl–Neelsen (ZN) stain, also known as the acid- fast stain, would be helpful in detection and identification of Schistosoma eggs. In histological sections, S. mansoni eggshells appear as ZN positive and S. haematobium shells as ZN negative. The staining target of the responsible ZN component (carbolfuchsin) in the shell is unknown and because carbolfuchsin is supposed to stain mycolic acids in the mycobacterial cell wall, unidentified substances in the eggshell were proposed as target. Fuchsin is a known nucleic acid stain, and it was already shown that mycobacteria with insufficiently retained carbolfuchsin may be invisible in bright-field microscopy; yet, they can be easily detected because of a strong red fluorescence when excited with green light.

The team prestained a smear of S. mansoni eggs with the nucleic acid stain 4ʹ,6-diamidine-2ʹ-phenylindole dihydrochloride (DAPI) and then stained with the classical ZN procedure. The smear was observed using bright-field and fluorescent microscopy where carbolfuchsin fluoresces red (ZN-fluo) and DAPI fluoresces blue (DAPI-fluo). In bright-field microscopy, the shell appeared to stain very little, whereas the miracidium within the intact egg and outside the egg appeared acid-fast negative, apparently only retaining the counterstain methylene blue. Contrary to this, fluorescent microscopy showed strong staining of the miracidium with carbolfuchsin (ZN-fluo) and DAPI (DAPI-fluo).

The authors concluded that because acid-fast stains and low-cost light- emitting diode fluorescent microscopy are now commonly used in many regions where schistosomiasis is endemic, it may be the time to revisit the staining mechanisms of acid-fast stains and investigate the use of these stains for their capacity to improve the detection of Schistosoma eggs. The study was published in the April 2016 issue of The American Journal of Tropical Medicine and Hygiene.

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