AVANOS YÖRESİ SOKAK KÖPEKLERİNDE DİROFİLARİA İMMİTİS’İN PREVALANSININ BELİRLENMESİ

Year 2017, Volume: 26 Issue: 2, 140 - 146, 01.07.2017

Feray Yabaneri Abdullah İnci Alparslan Yıldırım Zuhal Önder Arif Çiloğlu Önder Düzlü

Abstract

Bu çalışma, Avanos ve çevresindeki sokak köpeklerinde
filarial nematod enfeksiyonlarının prevalansını tespit
etmek amacıyla yürütülmüştür. Bu amaçla, Mayıs-Eylül
2013 tarihleri arasında farklı yaş, cinsiyet ve ırktan
olmak üzere toplam 138 sokak köpeğinden kan örnekleri alınmıştır. Alınan kan örnekleri öncelikle natif,
modifiye knott ve membran filtrasyon-asit fosfataz
histokimyasal boyama yöntemleri ile perifer kandaki
mikrofilerler yönünden incelenmiştir. Sonraki basamakta toplanan kanlardan genomik DNA ekstraksiyonu
yapılmış ve elde edilen genomik DNA’lar Dirofilaria
immitis’in 5.8S-ITS2-28S ve cytochrome oxidase subunit
1 (COI) spesifik gen bölgelerini parsiyel olarak
amplifiye eden primerler ile PCR analizine tabii
tutulmuşlardır. PCR analizi sonucunda, 138 köpeğin 3’ü
(%2.17) D. immitis pozitif olarak belirlenmiş, diğer
filarial nematod enfeksiyonlarına ise rastlanmamıştır.
Moleküler olarak pozitif belirlenen 3 örneğin 2’sinde
perifer kan muayene yöntemleri ile mikrofilerler saptanmıştır. Pozitif belirlenen izolatlardan birine ait PCR
ürünü jelden pürifiye edilerek her iki gen bölgesi için
çift yönlü sekans analizlerine tabii tutulmuştur. Sekans
analizi sonucu Avanos yöresinden elde edilen D. immitis
izolatının Dünya’da GenBank veri tabanına kayıtlı homolog izolatlarla 5.8S-ITS2-28S ve parsiyel COI gen
bölgelerinin çoklu hizalama analizlerine göre sırasıyla
%99.5-%100 ve %94.3-%100 identiklik gösterdiği belirlenmiştir. Bu çalışma ile Avanos yöresi sokak köpeklerinde D. immitis enfeksiyonlarının hem mikroskobik
hem de moleküler yöntemlerle prevalansı belirlenmiş
ve pozitif belirlenen bir izolatın 5.8S-ITS2-28S ve COI
gen bölgelerinin moleküler karakterizasyonu ve
filogenetik analizi yapılmıştır.

Keywords

Avanos, Dirofilaria immitis, köpek, moleküler karakterizasyon, prevalans

The Prevalence of Dirofilaria Immitis in Stray Dogs in Avanos Province

Abstract

This study was carried out to determine the prevalence
of filarial nematode infections in stray dogs in Avanos
and vicinity. For this aim, a total of 138 blood samples
were collected from stray dogs in different ages, sexes
and breeds between May-September 2013. The blood
samples were firstly examined by wet mount (direct
microscopic examination), modified knott’s and membrane filtration-acid phosphates histochemical staining
techniques in order to detect the circulating microfilaria
in peripheral blood. In the next step, genomic DNA extraction was obtained from collected blood samples, and
the obtained genomic DNAs were analyzed by PCR with
primers amplifying specific gene regions of 5.8S-ITS2-
28S and cytochrome oxidase subunit 1 (COI) of Dirofilaria immitis. Three (2.17%) out of 138 blood samples
were found to be D. immitis positive whereas no other
filarial nematode species were detected. Two out of
three positive samples determined as molecularly positive were also found positive by peripheral blood examination techniques. The PCR product belonging to one of
positive isolates was gel purified and sequenced in both
directions for two gene regions. The identity rates between the obtained D. immitis isolate obtained from
Avanos region and the other homologous isolates from
the world GenBank were determined as 99.5%-100%
and 94.3%-100% according to the phylogenetic analyses of 5.8S-ITS2-28S and COI gene regions, respectively.
Both microscopic and molecular methods the prevalence of D. immitis from stray dogs of Avanos region was
revealed and the molecular characterization and phylogenetic analyses of isolate for 5.8S-ITS2-28S and COI
gene regions were investigated in this study.

Keywords

Avanos, Dirofilaria immitis, dog, molecular characterization, prevalence

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