Detection of Cryptosporidium oocysts in fresh and frozen cattle faeces: comparison of three methods

Abstract

Aims: The aim of this study was to compare the performance of three commonly used screening tests for Cryptosporidium oocysts in fresh and frozen cattle faeces. Methods and Results: Twenty‐nine freshly voided faecal samples were collected from calves from three farms in the northwest of England. Three diagnostic tests for Cryptosporidium were carried out on each sample both before and after freezing – the modified Ziehl‐Neelsen (MZN) and auramine phenol (APh) stains and a commercial enzyme immunoassay (EIA) kit, the ProSpecT Cryptosporidium Microplate assay (Remel, Lenexa, KS). Twelve samples were deemed positive by the reference standard (polymerase chain reaction, PCR). There were some discrepancies between the results of the screening tests and the levels of agreement were quantified. The sensitivity and specificity of each method was determined, with PCR as the gold standard. Sensitivity and specificity of the MZN stain was optimized when samples with fewer than two oocyst‐like bodies were classified as negative. Conclusions: All three screening methods used were effective in detecting Cryptosporidium infection in both fresh and frozen calf faeces. Significance and Impact of the Study: This study has highlighted the value of determining characteristics of tests used for diagnosis and epidemiological studies.

Analytical sensitivity of staining and molecular techniques for the detection of Cryptosporidium spp. oocysts isolated from bovines in water samples: a preliminary study

“…However, the analytical results of this study are consistent with studies that analyzed stool samples from diverse domestic species by both staining techniques, finding no significant differences in sensitivity (De Quadros et al 2006, Brook et al 2008. In the case of water, one study analyzed samples of wastewater treatment plants for the detection of Cryptosporidium spp.…”

“…The main advantages of ZN staining are that it is inexpensive and the slides can be kept as permanent records, whereas slides with AU stain cannot (Brook et al 2008). Moreover, AU requires an expensive fluorescence microscope and background fluorescence could be a problem in AU-stained smears if there is a lot of fecal debris (Weber et al 1991).…”

Unexpected results from large-scale cryptosporidiosis screening study in calves in Tanzania

“…Indeed, Arrowood and Sterling (1989) also reported problems with false-positive results using acid-fast staining techniques, and some fungal spores have been found to be acid-fast (Casemore, 1991;Sunnotel et al, 2006). A study that compared three methods of detecting Cryptosporidium oocysts in cattle feces (mZN, auramine phenol staining and a commercial enzyme immunoassay kit), found that although mZN had a comparable or better sensitivity than the other methods, the specificity was lower (Brook et al, 2008), and the main advantages of this method, according to these authors, were its price and the ease of keeping the slides as a permanent record.…”

Negative Staining Technique of Heine for the Detection of Cryptosporidium spp.: A Fast and Simple Screening Technique~!2009-11-13~!2009-01-06~!2010-02-11~!

“…For instance a sample could be considered positive if five oocysts or more were observed, causing low-level shedding of oocysts to become interpreted as negative samples [27]. This extra loss of sensitivity in turn could be resolved by using repeated stool sample examinations on consecutive days [28]. The modified Ziehl-Neelsen staining is a low cost technique (about 0,15 US$ per sample), while the cost for PCR-based diagnosis is considerably higher (about 7,60 US$ per sample) [29].…”