Detection and molecular characterization of the Wolbachia Endobacteria in the Culex pipiens (Diptera: Culicidae) specimens collected from Kayseri province of Turkey

Abstract

This study was performed to investigate Wolbachia endobacteria in Culex pipiens specimens collected from Kayseri province of Turkey. For this aim, totally 10 genomic DNA pools each including 6-15 Cx. pipiens specimens which were collected and identified within the scope of a project (No: 107O533) supported by TUBITAK, were examined by using the amplification of surface protein gene (wsp) region of the Wolbachia. The sequences from this gene were highly variable and could be used to resolve the phylogenetic relationships of different Wolbachia strains. After the genomic DNA extraction from the pools, PCR analyses were carried out with Wolbachia specific primer pair which was amplified a 590-632 bp region of the wsp gene. Out of 6 of the 10 examined genomic DNA pools were found to be positive (60.0%) by PCR analyses. The minimum infection rate of Wolbachia spp. in the totally analyzed 118 Cx. pipens specimens was determined as 5.08. One of the amplicon from the positive isolates was gel purified and sequenced in terms of wsp gene region of Wolbachia by using the same primers. Pair wise analyses of the obtained DNA sequences and multiple alignments with some other Wolbachia strains available in the GenBank were done and phylogenies were investigated. The obtained isolate (WolKys1) was deposited in GenBank International Nucleotide Sequence Database with the accession number JX474753. The phylogenetic analyses revealed that the obtained WolKys1 isolate belongs to Wolbachia Super Group B and wPIP group. According to the phylogenetic comparisons the WolKys1 showed 100.0% identity with some other Wolbachia isolates under the Group B. In conclusion, this study reports the first molecular detection and characterization of Wolbachia endobacteria in Cx. pipiens populations in Turkey

Keywords

• Cx. pipiens
• molecular characterization
• Turkey
• Wolbachia

Kayseri yöresinden toplanmış Culex pipiens örneklerinde Wolbachia Endobakterisinin belirlenmesi ve moleküler karakterizasyonu

Abstract

Bu çalışma, Kayseri yöresinden toplanmış Culex pipiens örneklerinde Wolbachia endobakterisini araştırmak amacıyla yapılmıştır. Bu amaçla TÜBİTAK tarafından desteklenen 107O533 kod no’lu araştırma projesi kapsamında, Kayseri yöresinden toplanmış, Cx. pipiens olarak identifiye edilmiş ve her birinde 6-15 adet C. pipiens türü içeren 10 adet genomik DNA havuzu materyal olarak belirlenmiştir. Genomik DNA havuzları Wolbachia yüzey protein (wsp) gen bölgesinin amplifikasyonu yönünden incelenmiştir. Bu gen bölgesinin sekansı yüksek değişkenlik göstermekte olup farklı Wolbachia suşları arasındaki filogenetik ilişkilerin analizinde kullanılabilmektedir. Havuzlardan genomik DNA ekstraksiyonunu takiben wsp gen bölgesinin 590-632 bp’lik kısmını amplifiye eden Wolbachia spesifik primerler ile PCR analizleri yapılmıştır. İncelenen 10 havuzun 6’sı (%60,0) PCR analizleriyle pozitif bulunmuştur. İncelenen toplam 118 Cx. pipiens türünde Wolbachia spp. ile minimum enfeksiyon oranı 5.08 olarak belirlenmiştir. Pozitif izolatlardan birine ait amplikon jel pürifiye edilmiş ve söz konusu gen bölgesi yönünden aynı primerler ile sekanslanmıştır. Elde edilen DNA dizisinin GenBank’ta mevcut diğer bazı Wolbachia suşları ile pairwise analizleri ve multiple alignmentları yapılarak filogenisi araştırılmıştır. Elde edilen izolat (WolKys1) JX474753 aksesyon numarası ile GenBank International Nucleotide Sequence Database’e kaydedilmiştir. Filogenetik analiz sonucu WolKys1 izolatının Wolbachia B süper grubu ve wPIP grubu içinde yer aldığı belirlenmiştir. Filogenetik kıyaslamalara göre WolKys1 izolatının B grubu altındaki diğer bazı Wolbachia izolatları ile %100 identiklik gösterdiği saptanmıştır. Sonuç olarak bu çalışma ile Türkiye’de ilk kez Cx. pipiens populasyonlarında Wolbachia endobakterisinin moleküler olarak belirlenmesi ve karakterizasyonu yapılmıştır

Keywords

• Cx. pipiens
• moleküler karakterize
• Türkiye
• Wolbachia

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