Because of the presence of host-adapted Cryptosporidium species and genotypes, molecular tools can help assess the source and hazardous potential of Cryptosporidium oocysts in water. The development and use of molecular tools in the analysis of environmental samples have gone through several phases. Earlier polymerase chain reaction (PCR) tools were designed for the detection of Cryptosporidium oocysts in clinical samples. Subsequently, a genotyping component was incorporated into many of these assays to differentiate Cryptosporidium oocysts of anthroponotic origins from zoonotic origins. These tools were mostly based on the sequences of bovine C. parvum isolates, and were intended for the detection for C. parvum in clinical samples, thus they do not detect and differentiate many Cryptosporidium spp. and distant C. parvum genotypes. More recently, new molecular tools that are Cryptosporidium genus-specific and have the ability to differentiate Cryptosporidium species and genotype have been introduced, which has resulted in the finding of five major Cryptosporidium parasites in humans: the C. parvum human and bovine genotypes, C. meleagridis, C. felis, and C. canis. Current problems in molecular detection of Cryptosporidium oocysts include: the availability of only a limited number of tools for species differentiation, most of which are based on the small subunit rRNA gene; the nonspecificity of some species differentiation tools; the misinterpretation of data because of lack of information of recent findings; and, the existence of erroneous data in the database and publications. Nevertheless, in conjunction with immunomagnetic separation (IMS), some PCR-based tools have been successfully used in the detection, differentiation, and tracking of Cryptosporidium oocysts in storm water, surface water, and wastewater. Results of these studies have shown that a significant proportion of Cryptosporidium oocysts in water do not have high human-infective potential, which would have been overestimated by the recommended ICR method or method 1622/1623. Despite the recent progress, much more needs to be done before molecular tools can be used in routine analysis of water samples. Specifically, rigorous standardization and testing have yet to be carried out in order to: develop quality assurance and quality control procedures; develop protocols that allow the extraction of PCR-quality nucleic acid without using the expensive and pathogen-specific IMS; reduce turnaround times to allow close to real-time detection; incorporate quantitative and high resolution typing procedures for analysis of samples in special situations (such as outbreaks or bioterrorism); and, take advantage of new techniques such as biosensors and microarrays. The use of molecular tools can potentially generate data that are useful in the risk assessment of various types of water in different environmental settings, and for watershed management and source water protection. Includes references, tables, figures.
