Abstract
The aim of this study was to determine the genetic diversity of Giardia duodenalis present in a human population living in a northern Ecuadorian rain forest. All Giardia positive samples (based on an ELISA assay) were analysed using a semi-nested polymerase chain reaction-restriction fragment length polymorphism assay that targets the glutamate dehydrogenase (gdh) gene; those amplified were subsequently genotyped using NlaIV and RsaI enzymes. The gdh gene was successfully amplified in 74 of 154 ELISA positive samples; 69 of the 74 samples were subsequently genotyped. Of these 69 samples, 42 (61%) were classified as assemblage B (26 as BIII and 16 as BIV), 22 (32%) as assemblage A (3 as AI and 19 as AII) and five (7%) as mixed AII and BIII types. In this study site we observe similar diversity in genotypes to other regions in Latin America, though in contrast to some previous studies, we found similar levels of diarrheal symptoms in those individuals infected with assemblage B compared with those infected with assemblage A.
Highlights
2008, Sahagún et al 2008); we, found no evidence of differential pathogenicity between assemblage A and B, which was in agreement with findings by García et al (2002) and Ceu Souza and Poiares da Silva (2004)
Of the 154 ELISA-positive samples analysed for the presence of the gdh gene, only 74 (48%) were positive, likely due to low numbers of cysts in the faeces of some of our samples
This lack of PCR sensitivity for the gdh gene has been noted by other groups, such as Amar et al (2002), Cordón et al (2008) and Tungtrongchitr et al (2010), who suggested that absence of this band was
Summary
Read et al (2004) and Babaei et al (2008). One positive control identified as assemblage AII and another as assemblage BIII were used as references for the RFLP. All ELISA-positive samples were analysed by semi-nested PCR and 74 (48%) exhibited a band at 432 bp specific for the gdh gene (Fig. 1). Of these 74 samples, 69 were genotyped using RFLP (Table II), with the NlaIV restriction enzyme used to distinguish between assemblage A and assemblage B genotypes (Fig. 2A). Some prior studies have suggested that assemblage A infections are more likely to cause diarrhoea than assemblage B (Read et al 2002, Haque et al 2005, Cordón et al. Fig. 1: amplification of the 432 base pairs (bp) band specific for the glutamate dehydrogenase gene of Giardia duodenalis. Lane 1: molecular weight marker 1,000 bp; 2: Giardia positive control 432 bp; 3: Giardia positive 432 bp
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