Intestinal parasites and genotyping of Giardia duodenalis in children: first report of genotype B in isolates from human clinical samples in Mexico

Abstract

Giardia duodenalis is one of the most prevalent enteroparasites in children. This parasite produces several clinical manifestations. The aim of this study was to determine the prevalence of genotypes of G. duodenalis causing infection in a region of southeastern Mexico. G. duodenalis cysts were isolated (33/429) from stool samples of children and molecular genotyping was performed by polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) analysis, targeting the triosephosphate isomerase ( tpi ) and glutamate dehydrogenase ( gdh ) genes. The tpi gene was amplified in all of the cyst samples, either for assemblage A (27 samples) or assemblage B (6 samples). RFLP analysis classified the 27 tpi -A amplicons in assemblage A, subgenotype I. Samples classified as assemblage B were further analysed using PCR-RFLP of the gdh gene and identified as assemblage B, subgenotype III. To our knowledge, this is the first report of assemblage B of G. duodenalis in human clinical samples from Mexico.

Fig. 1. : identification of subgenotypes A by polymerase chain reaction (PCR) amplification based on the triosephosphate isomerase ( tpi ) gene. Lane 1: molecular size standards; 2, 3: positive control ( Giardia duodenalis subgenotype A-I); 4: negative control; 5-8: Giardia isolates. Silver stained polyacrylamide gel shows in the Lane 2: the amplicon of the PCR (476-bp) based on tpi gene of G. duodenalis for assemblages A; 3, 5-8: DNA pattern of restriction fragment length polymorphism analysis (437-bp) after digestion with Rsa I corresponding to the subgenotype A-I.

Fig. 2. : identification of subgenotypes B by polymerase chain reaction (PCR) amplification based on the triosephosphate isomerase ( tpi ) and glutamate dehydrogenase ( gdh ) genes. Lane 1: molecular size standards; 2, 6: amplicons of the PCR (140-bp) based on tpi gene of Giardia duodenalis for assemblage B; 3, 7: amplicons of the PCR (432-bp) based on gdh gene of G. duodenalis ; 4, 8: DNA pattern of restriction fragment length polymorphism analysis (300-bp and 130-bp) after digestion with Rsa I corresponding to the subgenotype B-III; 5: negative control.