The investigation of Cryptosporidium spp. in water samples by PCR

Amaç: Sivas ve çevresinde insanlarda, insanlara kaynak oluşturan insan ve hayvan çıkartılarıylakirlenme olasılığı yüksek içme ve kullanma sularında bu parazitin prevalansını PCR yöntemiylesaptamaktır.Yöntem: Bu amaçla; Sivas ilçe ve köylerinden toplam 92 örnekten en az 10 litrelik su örneğiplastik bidonlarla toplanmış ve filtre edilmiştir. Modifiye Asit Fast (MAF) boyama yöntemi içinyayma preparatları hazırlanmış ve 100xlik objektifle Cryptosporidium spp. yönündenincelenmiştir. Tüm örneklerden DNA izolasyonu ticari bir kit (QIAGEN -QIAamp DNA StoolMini Kit) kullanılarak yapıldı. Örneklerde Cryptosporidium 18S SSU rRNA lokusunun PCR ileçoğaltılması sonucu tanıya gidilmiştir.Bulgular: İncelenen 92 su örneğinin asit-fast ile hiç birinde PCR ile 2sinde (%2,2)Cryptosporidium spp. saptanmıştır. Pozitif örnekler musluk sularından saptanmış olup kaynaksularında parazit bulunmamıştır. Pozitif örnekler Kangal ve Suşehri ilçelerine bağlı iki köyden eldeedilen musluk sularından saptanmıştır.Sonuç: Çevresel su örneklerinde Cryptosporidium varlığı birkaç yöntemle araştırılmalıdır. Direktve Asit-fast boyama yöntemiyle yapılan incelemeler başarılı bulunmamıştır. Çünkü suörneklerinde ookistlerin yapısal özellikleri bozulmakta ve tanınmaları zorlaşmaktadır.Cryptosporidium spp.nin saptanmasında PCR daha duyarlı bir yöntemdir.

Su örneklerinde PCR ile Cryptosporidium spp. varlığının araştırılması

Objective: Cryptosporidium spp. are intestinal pathogens of humans and animals. Environmentaland tap water samples obtained from different regions in Sivas were examined for the prevalenceof Cryptosporidium infection.Methods: Approximately 10 lt of water collected and filtered from spring and tap waters fromvillages and towns in Sivas province were examined. All samples were examined both with theModified Acid Fast (MAF) staining method and PCR for the presence of Cryptosporidium spp. Agenus specific PCR assay was designed for the specific amplification of the 18S SSU rRNA gene.Results: None of the 92 water samples analyzed by MAF were positive, whoever 2 water samples(2.2%) from the Kangal and Suşehri counties were positive by PCR.Conclusions: The PCR technique is more reliable in diagnosing Cryptosporidium in watersamples. It is possible that the oocysts are decomposed in water and it is difficult to identify themby MAF.

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