DNA fingerprinting of Cryptosporidium parvum isolates using amplified fragment length polymorphism (AFLP)

The genetic variability of 10 Cryptosporidium parvum isolates of human and animal origin was investigated using amplified fragment length polymorphism (AFLP). Analysis of fluorescent dye-labeled amplified products was carried out using an ABI PRISM(TM) 377 DNA sequencer and ASI PRISM(TM) GeneScan so ftware. One-hundred and twelve primer combinations were evaluated using a s ingle C. parvum isolate. The patterns generated were highly reproducible. F or subsequent study, a subset of 9 primer pairs that yielded 30-90 DNA frag ments after the polymerase chain reaction, within the size range of 50-500 bp, was used to screen the 10 C. parvum isolates, including 7 bovine, 1 equ ine, and 2 of human origin. The animal isolates produced identical fingerpr int patterns with every primer combination tested. Of the 2 human isolates tested. 1 of the isolates, passaged in calves, generated the same AFLP DNA banding patterns as the animal isolates. whereas the other isolate, obtaine d directly from human feces, produced unique patterns. Polymorphism, detect ed by comparison of the fingerprint patterns of the latter human isolate wi th the common pattern shared by all other isolates, ranged from 17 to 35% f or the 9 primer pairs. The results show that AFLP is a useful method for di fferentiating C. parvum isolates into 2 distinct genotypes.