Türkiye’de turunçgil ağaçlarında moleküler tekniklerle Spiroplasma citri’nin belirlenmesi

Abstract

Spiroplasma citri, kurak ve yarı kurak iklimde turunçgiller üzerindeki turunçgil palamutlaşma hastalığı (Citrus Stubborn Disease (CSD))’na neden olan bir etmendir. Bu etmenin turunçgil bahçeleri ve fidanlıklarda bulunan genç fidanlardaki enfeksiyonunun erken aşamada tespiti ve karakterizasyonu, hastalığın Spiroplasma citri’den ari bölgelere yayılmasını önlemek için büyük önem taşımaktadır. Bu çalışma, Türkiye'deki farklı Spiroplasma citri suşlarının muhtemel varlığını araştırmak amacıyla yapılmıştır. Eylül ayında Adana ilinin Kozan ilçesinde farklı köylerde bulunan turunçgil bahçelerindeki Stubborn hastalığına benzer çoğunlukla boğum arası kısalma, yapraklarda kaşıklanma ve leke, sezon dışı çiçeklenme ve tipik küçük palamut tipi meyve simptomları gösteren ağaçlardan yaprak ve meyve örneği toplanmıştır. Türk izolatlarının saptanması ve karakterizasyonu için, polimeraz zincir reaksiyonu (PZR) Spiralin-f/r, P89-r/f ve P58-6f/4r primer çiftleri kullanılarak gerçekleştirilmiştir. Spiralin-f/r primerlerinde pozitif sonuç saptanmamıştır. Buna karşın, testlenen tüm örnekler P89-r/f ve P58-6f/4r primer çiftlerine pozitif sonuç vermiştir. P89-r/f primer çiftinden elde edilen PZR ürünleri sekanslanmış ve filogenetik ağaç oluşturulmuştur. Elde edilen filogenetik ağaç, önemli bir turunçgil üretim merkezi olan Adana ilindeki turunçgil ağaçlarında saptanan Spiroplasma citri izolatlarının, Fas, Mısır ve Amerika Birleşik Devletleri (ABD)'nden izole olanlardan daha ziyade Meksika, İspanyol, Cezayir ve İtalyan izolatlarıyla yakından ilişkili olduğunu kanıtlamıştır.

Keywords

• Turunçgil,Filogenetik ilişki,Gen dizileme,Stubborn

Detection of Spiroplasma citri from citrus trees in Turkey by molecular techniques

Abstract

Spiroplasma citri is the causal agent of Citrus Stubborn Disease (CSD) on cultivated citrus in the arid and semi-arid climate. Detection and characterization of this agent at the early stage of infection in citrus orchard and young citrus saplings in the nursery is of utmost importance to prevent the introduction and spread of the disease in Spiroplasma citri free areas. The current study was conducted in order to investigate the possible existence of different Spiroplasma citri strains in Turkey. Leaf and fruit samples were collected from ten years old citrus trees showing mostly short internodes, upright and mottled leaves, off-season blossoming and typical small and acorn-shaped fruits symptoms associated with citrus stubborn diseases in different villages of Kozan town of Adana province in Turkey. Polymerase chain reaction (PCR) was performed using Spiralin-f/r, P89-r/f and P58-6f/4r primer couples for the detection and characterization of the Turkish isolates. No positive sample was detected with the primers Spiralin-f/r. On the contrary, all samples reacted positively to the P89-r/f and P58-6f/4r primer couples. The obtained amplicon from the P89-r/f primers pair was sequenced and the phylogenetic tree was constructed. Phylogenetic tree showed that the Turkish Spiroplasma citri isolates are closely related to the Mexican, Spanish, Algerian, and Italian than those isolated from Morocco, Egypt and USA.

Keywords

• Citrus,Phylogenetic relation,Sequencing,Stubborn

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