Whole genome amplification (WGA) for archiving and genotyping of clinical isolates ofCryptosporidiumspecies

Abstract

SUMMARYClinical and environmental isolates of pathogens are often unique and may be unculturable, yielding a very limited amount of DNA for genetic studies.Cryptosporidiumin particular are difficult to propagate. Whole genome amplification (WGA) is a valuable technique for amplifying genomic material. In this study, we tested 5 WGA commercial kits usingCryptosporidiumclinical isolates. DNA of 5C. hominisand 5C. parvumclinical isolates andC. parvumIOWA reference strain were used. The majority of the samples were amplified by all of the kits tested. The integrity and fidelity of the amplified genomic DNA were assessed by sequence analysis of several PCR products of varying length. We found evidence that one kit in particular may be more error prone while another seemed the more suitable kit forCryptosporidiumclinical samples, generating high molecular weight DNA from all the samples with high fidelity. Thus WGA was found to be a useful technique for producing amplified DNA suitable for downstream genotyping techniques and archiving ofCryptosporidiumclinical isolates.