İBRAHİM CÜNEYT BALCIOĞLU, Naser SEVİL, HANDE DAĞCI, Aslı TETİK, KORAY ERGÜNAY, Kor YERELİ, Ahmet ÖZBİLGİN, NEVİN TURGAY, Seray ÖZENSOY TÖZ, Özgür KURT

Genotyping of Giardia lamblia in a Cohort of Turkish Patients:A Search for a Relationship between Symptoms and Genotypes

Giardia lamblia ile yapılan moleküler düzeydeki çalışmalar iki farklı genotipin farklı klinik belirtilere sahip olduğunu ortaya çıkarmıştır. Bununla birlikte, hangi genotipin ne tür klinik belirtilere neden olduğuna dair henüz kesin bulgular elde edilmiş değildir. Bu araştırmada Türkiye'nin Manisa yöresinden elde edilen G. lamblia izolatları polimeraz zincir reaksiyonu (PZR) ve sonrasında DNA dizi analizi yöntemleriyle araştırılmış ve saptanan genotipler ile hastalardaki klinik semptomlar arasındaki ilişkiler incelenmiştir. Toplam 63 hastadan DNA örnekleri izole edilmiş ve bunların 54'üne Xho I restriksiyon enzimi kullanılarak RFLP analizi uygulanmıştır. Sonuçlar incelendiğinde, 54 örneğin 38'inin (%70.37) A, 16'sının ise (%29.63) B genotipine ait olduğu belirlenmiştir. Kadın hasta sayısının B genotipinde anlamlı düzeyde daha fazla olduğu (p=0.18), ayrıca hastalarda birlikte bulunan karın ağrısı ve ishal yakınması ile G. lamblia B genotipi arasında istatistiksel olarak anlamlı bir ilişki olduğu tespit edilmiştir (ki-kare testi, 10.52; P

Bir Türk Hasta Kohortunda Giardia lamblia Genotiplendirilmesi:Semptomlar ile Genotipler Arası İlişkinin Araştırılması

Recent surveys investigating the molecular biology of Giardia lamblia revealed two distinct assemblages with different clinical outcomes. However, there is not a universal compromise about the clinical effects of each assemblage, warranting further studies. Here, we report the results of the first analyses of the assemblages of G. lamblia in Manisa province located in western Turkey, together with their relationships with the symptoms and DNA sequence analyses of the PCR products. DNA samples were isolated from the stools of 63 patients infected with G. lamblia and 54 DNA samples, amplified successfully with PCR, were digested with the enzyme Xho I for RFLP. Thirty-eight of 54 samples (70.4%) were found to be in Assemblage A, while the remaining 16 samples (29.6%) were found to be in Assemblage B. The number of female patients was found significantly higher in Assemblage B (P=0.18). There was a statistically significant relationship between the occurrence of both abdominal pain and diarrhea and Assemblage B (chi-square, 10.52; P<0.05). No other statistically significant relationship was detected between the assemblages and neither with the symptoms nor with the age groups of the patients. The comparison of the DNA sequences of the PCR products from two assemblage B (one subtype B1 and one B) and one assemblage A samples both with each other and with other DNA sequences in the NCBI website by multialignment analyses, revealed specific regions for assemblages B (B1-B) and A on tpi gene region. Further studies with more patients are required to assess these initial results. Now, our aim is to design a probe for tpi gene region to set up a real-time PCR assay that is easier to conduct and requiring shorter time for the analyses

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Caccio SM, Ryan U: Molecular epidemiology of giardiasis. Molecul Biochem Parasitol, 160, 75-80, 2008.

Volotão AC, Costa-Macedo LM, Haddad FS, Brandão A, Peralta JM, Fernandes O: Genotyping of Giardia duodenalis from human and animal samples from Brazil using beta-giardin gene: A phylogenetic analysis. Acta Trop, 102 (1): 10-19, 2007.

Hill DR: Giardia lamblia. In, Mandell GI, Douglas R, Bennett J (Eds): Principles and practice of infectious diseases. pp. 2888-2894, Churchill Livingstone, New York, 2000.

Sousa MC, Morais JB, Machado JE Poiares Da Silva J: Genotyping of Giardia lamblia human isolates fro Portugal by PZR-RFLP and sequencing. J Eukaryot Microbiol, 53 (S1): 174-176, 2006.

Amar CFL, Dear PH, McLauchlin J: Detection and genotyping by real time PCR/RFLP analyses of Giardia duodenalis from human faeces. J Medical Microbiol, 52, 681-683, 2003.

Berrilli F, Di Cave D, D'Orazi C, Orecchia P, Xhelilaj L, Bejko D, Caça P, Bebeci D, Cenko F, Donia D, Divizia M: Prevalence and genotyping of human isolates of Giardia duodenalis from Albania. Parasitol Int, 55, 295- , 2006.

Yason JA, Rivera WL: Genotyping of Giardia duodenalis isolates among residents of slum area in Manila, Philippines. Parasitol Res, 101 (3): 687, 2007.

Monis PT, Andrews RH, Mayrhofer G, Ey PL: Genetic diversity within the morphological species Giardia intestinalis and its relationship to host origin. Infect Genet Evol, 3, 29-38, 2003.

Thompson RC, Hopkins RM, Homan WL: Nomenclature and genetic groupings of Giardia infecting mammals. Parasitol Today, 16 (5): 210-213, 2000.

Kohli A, Bushen OY, Pinkerton RC, Houpt E Newman RD, Sears CL, Lima AA, Guerrant RL: Giardia duodenalis assemblage, clinical presentation and markers of intestinal inflammation in Brazilian children. Trans R Soc Trop Med Hyg, 102, 718-725, 2008.

Amar CF, Dear PH, Pedraza-Díaz S, Looker N, Linnane E, McLauchlin J: Sensitive PCR-restriction fragment length polymorphism assay for detection and genotyping of Giardia duodenalis in human feces. J Clin Microbiol, 40 (2): 446-452, 2002.

Homan WL, Mank TG: Human giardiasis: Genotype linked differences in clinical symptomatology. Int J Parasitol, 31, 822-826, 2001.

Aydin AF, Besirbellioglu BA, Avci IY, Tanyuksel M, Araz E, Pahsa A: Classification of Giardia duodenalis parasites in Turkey into groups A and B using restriction fragment length polymorphism. Diagn Microbiol Infect Dis, 50 (2): 147-151, 2004.

Paintlia AS, Mahajan RC, Chakraborti A, Sehgal R, Ganguly NK: Characterization of Giardia lamblia groups A and B from North India by isoenzyme and random amplified polymorphic DNA analysis. Parasitol Res, 85 (6): 510-512, 1999.

Read C, Walters J, Robertson ID, Thompson RC: Correlation between genotype of Giardia duodenalis and diarrhoea. Int J Parasitol, 32 (2): 229-231,

Kohli A, Bushen OY, Pinkerton RC, Houpt E Newman RD, Sears CL, Lima AA, Guerrant RL: Giardia duodenalis assemblage, clinical presentation and markers of intestinal inflammation in Brazilian children. Trans R Soc Trop Med Hyg, 102, 718-725, 2008.

Şahin A, Karslı T, Elmacı C, Balcıoğlu MS: Beta-Lactoglobulin Gene Types in Turkish Fat-tailed Sheep Breeds. Kafkas Univ Vet Fak Derg, 17 (6): 1033, 2011.

Cedillo-Rivera R, Darby JM, Enciso-Moreno JA, Ortega-Pierres G, Ey PL: Genetic homogeneity of axenic isolates of Giardia intestinalis derived from acute and chronically infected individuals in Mexico. Parasitol Res, 90, 119-123, 2003.

Giangaspero A, Berilli F, Brandonisio O: Giardia and Cryptosporidium and public health: The epidemiological scenario from the Italian perspective. Parasitol Res, 101, 1169-1182, 2007.

Haque R, Roy S, Kabir M, Stroup SE, Mondal D, Houpt ER: Giardia assemblage A infection and diarrhoea in Bangladesh. J Infect Dis, 192 (12): 2173, 2005.

Lebbad M, Ankarklev J, Tellez A, Leiva B, Andersson JO, Svard S: Dominance of Giardia assemblage B in Le´on, Nicaragua. Acta Trop, 106, 53, 2008.

Özbilgin A, Kilimcioğlu AA, Kurt Ö, Ertan P, Değerli K, Yereli K, Ok ÜZ, Onağ A: Çocuklarda giardiasisin metronidazol ve tek doz ornidazolle sağaltımlarının karşılaştırılması. T Parazitol Derg, 23 (3): 243-246, 1999.

Balcioglu IC, Kurt Ö, Limoncu ME, Dinç G, Gümüş M, Kilimcioglu AA, Kayran E, Özbilgin A: Rural life, lower socioeconomic status and parasitic infections. Parasitology Int, 56, 129-133, 2007.

Marshall MM, Naumovitz D, Ortega Y, Sterling CR: Waterborne protozoan pathogens. Clin Microbiol Rev, 10, 67-85, 1997.

Sprong H, Cacciò SM, van der Giessen JW; ZOOPNET Network And Partners: Identification of zoonotic genotypes of Giardia duodenalis. PLoS Negl Trop Dis, 3 (12): e558, 2009.

Itagaki T, Kinoshita S, Aoki M, Itoh N, Saeki H, Sato N, Uetsuki J, Izumiyama S, Yagita K, Endo T: Genotyping of Giardia intestinalis from domestic and wild animals in Japan using glutamete dehydrogenase gene sequencing. Vet Parasitol, 133 (4): 283-287, 2005.

Minvielle MC, Molina NB, Polverino D, Basualdo JA: First genotyping of Giardia lamblia from human and animal feces in Argentina, South America. Mem Inst Oswaldo Cruz, 103 (1): 98-103, 2008.