Molecular cloning of a cysteine proteinase cDNA from adult Ancylostoma ceylanicum by the method of rapid amplification of cDNA ends using polymerase chain reaction

Abstract

The hookworm Ancylostoma ceylanicum is a parasite of great importance in human and veterinary medicine. The most promising vaccination trials against hookworm infections are based on antigens belonging to the proteinase family. The aim of the present research was to isolate a cysteine proteinase gene from A. ceylanicum. This was achieved by rapid amplification of cDNA ends using polymerase chain reaction (RACE-PCR). A set of consensus oligonucleotide primers was designed to anneal to the conserved coding regions of cysteine proteinase. The PCR products were cloned and sequenced. The novel sequence displayed a high degree of homology with genes of cysteine proteinases known from other hookworm species. In the coding region the nucleotide identity with accp-1, the cysteine proteinase gene of A. caninum, reaches 84.3%. Analysis of the expression of acey-1, the cysteine proteinase gene of A. ceylanicum, suggests that it is produced exclusively in the gland cells of either adult worms or blood-feeding stages of A. ceylanicum.