Cryptosporidium and giardia in water in Alexandria: detection and evaluation of viability by flow cytometry and different stains

ABSTRACT

Cryptosporidium oocysts and Giardia cysts have become ubiquitous in surface waters worldwide. The number and extent of outbreaks of waterborne diseases indicate a significant risk for their possible transmission by drinking-water. Since many Egyptian cities depend on surface water as their main source of drinking water, knowledge of the prevalence of waterborne protozoa in water resources is important. The present study was designed to use flow cytometry to detect Cryptosporidium oocysts and Giardia cysts in water samples in Alexandria city in comparison with the standard staining techniques. Testing the viability of the encountered parasites was also carried out comparing flow cytometry and trypan blue vital stain. Thirty water samples were collected from water tanks from different districts of Alexandria city. Samples were subjected to staining techniques and flow cytometry. Stains used were modified Zeihl-Neelsen [MZN], safranin methylene blue [SMeB], modified trichrome, fluorescent stains [phenol auramine and acridine orange]. Viability was evaluated comparing trypan blue stain and flow cytometry using 4'-6-Diamidino-2-phenylindole immunostain [DAPI]. Flow cytometry proved to be much more sensitive than staining techniques with a sensitivity of 100% for both Cryptosporidium oocysts [30 samples] and Giardia cysts [11 samples]. Following flow cytometry, the fluorescent phenol auramine stain had the greatest sensitivity of 94.74% and 80% [18 and 4 samples, respectively]. The percent of live parasites present in each sample was always significantly higher by DAPI than trypan blue stain. The results of the present study clearly demonstrate that incorporation of flow cytometry can improve sensitivity of detection of Giardia cysts and Cryptosporidium oocysts in water samples. Although it is more expensive than the other staining methods, it is rapid, simple and accurate in estimating the quantity and viability of the parasites in each sample. Thus, flow cytometry can be recommended for detection of protozoa in water