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Original Article

Detection and genotyping of Giardia intestinalis isolates using intergenic spacer (IGS)-based PCR

Jong-Ho Lee1, Jongweon Lee1, Soon-Jung Park1, Tai-Soon Yong1, Ui-Wook Hwang2
The Korean Journal of Parasitology 2006;44(4):343-353.
Published online: December 20, 2006

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Corresponding author (tsyong212@yumc.yonsei.ac.kr)
• Received: October 25, 2006   • Accepted: November 17, 2006

Copyright © 2006 by The Korean Society for Parasitology

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Detection and genotyping of Giardia intestinalis isolates using intergenic spacer (IGS)-based PCR
Korean J Parasitol. 2006;44(4):343-353.   Published online December 20, 2006
DOI: https://doi.org/10.3347/kjp.2006.44.4.343
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Detection and genotyping of Giardia intestinalis isolates using intergenic spacer (IGS)-based PCR
Korean J Parasitol. 2006;44(4):343-353.   Published online December 20, 2006
DOI: https://doi.org/10.3347/kjp.2006.44.4.343
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Detection and genotyping of Giardia intestinalis isolates using intergenic spacer (IGS)-based PCR
Image Image Image Image Image Image Image
Fig. 1 Nucleotide sequence alignment of 3' part of the IGS from 13 G. intestinalis isolates. The abbreviations with initial letters CA, K and RD mean Chinese, Korean and Laotian, respectively. GK (GenBank accession no. X52949) nucleotide sequence was retrieved from the EMBL database. WB and GS were purchased from American Type Culture Collection. The dots indicate identical nucleotides and the hyphens indicate base deletions. Underlines show the position of the primer sequences for selective amplification of each assemblage.
Fig. 1 Continued.
Fig. 1 Continued.
Fig. 2 Phylogenetic relationship between the 3 G. intestinalis IGS assemblages determined by neighbor joining method. The results showed that the genotype of 13 G. intestinalis isolates could be classified into the 3 different assemblages, A1, A2, and B.
Fig. 3 Assemblage specificity of IGS-PCR. The 176 bp, 261 bp and 319 bp amplified products were specifically shown in the assemblage A1, A2 and B lane, respectively. Lane M represents 100 bp ladder from MBI Fermentas. The 500 bp and 200 bp bands were shown.
Fig. 4 Specificity of IGS-PCR in recognizing genomic DNAs of other bacteria, protozoa, and non-human Giardia isolates. Lane M, 100 bp ladder from MBI Fermentas. The 500 bp and 200 bp bands were shown. Lane 1-3, G. intestinalis isolates WB, K1, GS; Lane 4, E. coli DH5α; Lane 5, V. vulnificus; Lane 6, E. histolytica; Lane 7, T. vaginalis; Lane 8-9, dog Giardia isolates GD1, GD2; IGS-PCR did not show any amplification products with other bacteria or protozoans including Giardia isolates from other animals.
Fig. 5 Sensitivity of IGS-PCR. PCR product obtained using different amount of genomic DNA of WB (top), K1 (middle) and GS (bottom) isolate. PCR product was detectable with 100 pg of template DNA of K1 isolate, and using the DNA of WB or GS isolates, 10 pg of template DNA was enough to produce PCR products. Lane M represents 100 bp ladder from MBI Fermentas. The 500 bp and 200 bp bands were shown.

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Fig. 1

Fig. 1

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Fig. 5

Detection and genotyping of Giardia intestinalis isolates using intergenic spacer (IGS)-based PCR
Isolates Geographic origin Host Nature of sample Reference
WB Afghanistan Human Trophozoites (Yong et al., 2002)
GK Australia Human Unknowna) (Healey et al., 1990)
GS USA Human Trophozoites (Yong et al., 2002)
K1 Korea, Seoul Human Trophozoites (Yong et al., 2002)
K2 Korea, Seoul Human Cysts This study
K-L Korea, Seoul Human Cysts (Yong et al., 2002)
CA9 China, Anhui Human Cysts (Yong et al., 2002)
CA13 China, Anhui Human Cysts (Yong et al., 2002)
CA14 China, Anhui Human Cysts (Yong et al., 2002)
CA17 China, Anhui Human Cysts (Yong et al., 2002)
CA18 China, Anhui Human Cysts This study
RD25 Laos, Saravane Human Cysts (Yong et al., 2002)
RD37 Laos, Saravane Human Cysts (Yong et al., 2002)
GD1 Korea, Seoul Dog Cysts This study
GD2 Korea, Seoul Dog Cysts This study
GM Korea, Pajoo Field mouse Cysts This study
Function Name Nucleotide sequence
Upstream primer for IGS region GLF 5´-GACCGTCGTGAGACAGGTTAG-3´
Downstream primer for IGS region GSR 5´-CCTGCTGCCGTCCTTGGATG-3´
Upstream primer for group A1 GA1F 5´-GGTTTGCCGTGATATGCA-3´
Upstream primer for group A2 GA2F 5´-CCCTCCAGAGCAGMRTGAGA-3´
Upstream primer for group B GBF 5´-GRCAGGTGTGCTCAGGTG-3´
Downstream primer for group A & B GABR 5´-AGCACCTTGTCCTATAYAGT-3´
Table 1. Giardia isolates examined in this study

rDNA nucleotide sequence was retrieved from the EMBL database.

Table 2. The primers used for the PCR amplification in this study. GLF and GSR primers were used for amplication of the whole IGS region, and other 4 primers were used for selective amplication of each assemblage, respectively

Table 1.

Table 2.

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