Incidence of Giardia lamblia Subspecies by PCR-RFLP in Stool Specimens of Hospitalized Children at Urmia Mutahhari Hospital, West Azerbaijan Province, Iran.
AbstractBackground: Giardia lamblia is one of the most prevalent intestinal flagellate protozoa that infects a wide range of vertebrate hosts causing severe intestinal disorder in children.This study was performed to determine subspecies of G.lamblia by the PCRRFLP method, targeting the glutamate dehydrogenase(gdh)locus, in hospitalized children at Urmia Mutahhari Hospital, West Azerbaijan Province,Iran and determining the infection transformational storages in this area.Methods: Overall,720 stool specimens were collected from the hospitalized children,34 samples were positive and Giardia cysts were detected under the microscope. Cysts were partially purified by the sucrose density gradient method and then washed with sterile distilled water to remove effectively the PCR inhibitors. Genomic DNA of G.lamblia isolates was extracted by freeze-thaw cycles followed by phenol/ chloroform/isoamyl alcohol method. The single step PCR-RFLP assay was used to differentiate the assemblages between A and B, which were found in humans. In this method,432 bp expected size was amplified, and then for detection of subspecies, specificrestriction RsaI and BspLI enzymes were used.Results: Totally 34 samples were positive in terms of Giardia cyst out of 720 examined samples microscopically, so the parasite spread rate is reported 4.72%. Analysis PCR-RFLP on these samples revealed that 28 samples (93.3%) have the genotype BIII and 2 samples (6.7%) belong to the subgroup BIV.Conclusion: PCR-RFLP is a proper analytical method for determining the genotypeamong parasite types, using the glutamate dehydrogenizes zone's genes. Based on the results, an animal origin of infection cycle is suggested.
Savioli L, Smith H, Thompson A. Giardia and Cryptosporidiumjoin the “Neglected Diseases Initiative”. Trends Parasitol. 2006; 22:203-8.
Ali SA, Hill DR. Giardia intestinalis. Curr OpinInfect Dis. 2003; 16: 453–60.
Ajjampur SSR, Sankaran P, Kannan A, Sathyakumar K, Sarkar R, Beryl P,et al. Giardia duodenalis assemblages associated with Diarrhea in Children in South India Identified by PCRRFLP. Am J Trop Med Hyg. 2009; 80(1):16–19.
Fallah E, Nahavandi k, Jamali R, MahdaviPoor B, Asgharzadeh M. Molecular identification of Giardia duodenalis isolates from human and animal reservoirs by PCR-RFLP. J Biol Sci. 2008; 2:172-84.
Read CM, Monis PT, Thompson RC. Discrimination of all genotypes of Giardia duodenalis at the glutamate dehydrogenaselocus using PCR-RFLP. Infect Genet. 2004; 4:125-30.
Moshira M. F. Helmy, Hisham S. Abdel-Fattah, Rashed L. Real-time PCR/RFLP assay to detect Giardia intestinalis genotypes in human isolates with diarrhea in Egypt. J Parasitol. 2009; 95(4):1–5.
Monis PT, Andrews RH, Mayrhofer G, Ey PL. Genetic diversity within the morphological species Giardia intestinalisand its relationship to host origin. Infect Genet. 2003; 3:29-38.
Itagaki T, Kinoshita S, Aoki M, ItohN, Saeki H, Sato N, et al. Genotyping of Giardia intestinal is from domesticated wild animals in Japan using glutamate dehydrogenase gene sequencing. Vet Parasitol. 2005; 133(4):283-87.
O'Handley RM, Olson ME, Fraser D, Adams P, Thompson RC. Prevalence and genotypic characterization of Giardia in dairy calves from Western Australia and Western Canada. Vet Parasitol.2000; 90:193-200.
Eligio-Garcia L, Cortes-Campos A, Jimenez- Cardosoe E. Genotype of Giardia intestinalis isolates from children and dogs and its relationship to host origin. Parasitol Res. 2005; 97(1):1-6.
Leonhard S, Pfister K, Beelitz P, Wielinga C, Thompson RC. The molecular characterization of Giardia from dogs in southern Germany. Vet Parasitol. 2007; 30, (1):33-8.
Traub RJ, Monis PT, Robertson I, Irwin P, Mencke N, Thompson RC. Epidemiological and molecular evidence support the zoonotictransmission of Giardia among humans and dogs . 149 living in the same community. Parasitology. 2004; 128(3):253-62.
Robertson LJ, Forberg T, HermansenL, Hamnes IS, Gjerde B. Giardia duodenalis cysts isolated from wild moose and reindeer in Norway: genetic characterization by PCRRFLP and sequence analysis at two genes. J Wildl Dis. 2007; 43(4):576-85.
Van Keulen H, Macechko PT, Wade S, Schaaf S, Wallis PM, Erlandsen SL. Presence of human Giardia in domestic, farm and wild animals, and environmental samples suggest a zoonotic potential for giardiasis. Vet Parasitol. 2002; 108:97-107.
Hazrati Tappeh Kh, Mohammadzadeh H, Khashaveh Sh, Rezapor B. Prevalence of intestinal parasitic infections among primary school attending students in Nazloo-Chay rural region of Urmia, West Azerbaijan province, Iran in 2008. Urmia Journal of Medical Science. 2011; 4(16):18-21.
Hazrati Tappeh Kh, Mohammadzadeh H, Khashaveh Sh, Rezapor B. Prevalence of intestinal parasitic infections among primary school attending students in Barandooz-Chay rural region of Urmia, West Azerbaijan province, Iran in 2008. Afr J Microbiol Res. 2011; 5(5):28-32.
Saebi E. Protozoal diseases in Iran. Textbook of Clinical Parasitology. 4th ed. Ayexh: Tehran; 2005.
Luchtel D.L, Lawrence W.P and Dewalle F.B. Electron microscopy of Giardia lamblia cycts. Applied Env Microbiol. 1980; 40(4):821-32.
Bertrand I, Albertini L, Schwartzbrod J. Comparison of two target genes lamblia in human feces by PCR and PCR-Restriction Fragment Length Polymorphism.J Clin Microbiol. 2005; 43(12):5940-44.
Fallah E, Nahavandi K.H, Jamali R, Mahdavi B and Asgharzadeh M. Genetic characterization of Giardia intestinalis strains from patients having sporadic giardiasis by using PCR assay. J Med Sci. 2008; 4:1727-48.
Babaei Z, Oormazdi H, Akhlaghi A, Rezaie S, Razmjou1 E, Soltani- Arabshahi SK,et al. Molecular characterization of the Iranian isolates of Giardia lamblia:application of the glutamate dehydrogenase gene. Iran J Public Health. 2008; 37(2):75-82.
Boontanom P, Mungthin M, Tan-ariya, P, NaaglorT, Yoova L. Epidemiology of giardiasis and genotypic characterization of Giardia duodenalisin preschool children of a rural community, central Thailand. Trop Biomed. 2011; 28(1): 32–9.
Sulaiman IM, Fayer R, Bern C, Gilman RH, Trout JM, Schantz PM, et al. Triose phosphate isomerase gene characterization and potential zoonotic transmission of Giardia duodenalis. Emerg Infect Dis. 2003; 9(11):1444-52.
Dalir-Naghadeh B, Tavassoli M, Hatefinia A.Z. Study of epidemiologic measures of association between Giardia sp. infection with occurrence of diarrhea in calves. J Vet Res. 2008; 62(6):363-66.