Investigation of geographical differences among Giardia duodenalis assemblage B populations using diverse haplotypes identified by cloning of PCR amplicons.

Recently, several studies using cloned PCR amplicons have revealed greater haplotype diversity in Giardia duodenalis assemblage B than can be detected using conventional methods. This study aimed to estimate the geographical distribution of G. duodenalis assemblage B using these highly diverse haplotype sets. Samples were collected from two Indonesian sites (Waitabula and Wainyapu). Partial sequences of the glutamate dehydrogenase (gdh), triosephosphate isomerase (tpi), and beta-giardin (bg) loci were used as the PCR targets. Sequence analysis of the cloned PCR amplicons confirmed the presence of unique haplotypes in each region. For further analysis, we employed not only the Indonesian (Waitabula and Wainyapu) haplotypes acquired in this study but also the Kenyan haplotypes detected in our previous study. The phylogenetic relationships among haplotypes were calculated and plotted using multidimensional scaling (MDS). The plot integrating information from the three loci indicated geographically dependent clustering not only between Waitabula and Wainyapu but also between Indonesia and Kenya. This study demonstrates that employing greater haplotype diversity detected using the PCR-cloning method is an effective way to detect geographical differences in G. duodenalis assemblage B.