Characterization and Haplotype Analysis of Colletotrichum truncatum in Greenhouse Tomato in Turkey

Abstract

Anthracnose caused by Colletotrichum truncatum (Schwein.) Andrus and W.D. Moore, is an economically important disease of most tropical crops. In recent years, it has been reported that it is also pathogenic on tomatoes. In this study, the infected fruits were obtained from Antalya province in 2019. Isolates were purified by taking single spore. Conidia were measured as 22.5-32.5 × 3.75 µm. DNA isolation was carried out using the CTAB method. After the PCR amplification, the PCR product was run on agarose gel, visualized with a UV transilluminator, and sequenced. Phylogenetic analysis was conducted in MEGA 7. Based on morphological and phylogenetic analysis, the CT isolate was identified as C. truncatum. Pathogenicity tests were carried out using tomato leaves and cherry tomatoes. The inoculated leaves and tomatoes were incubated on a moist filter paper in climate chambers under 27 °C temperature and 12:12 h light-dark conditions. Acervuli were observed on infected tissues on the 7th day of inoculation. Haplotype, the number of haplotypes, and nucleotide diversity were analyzed by DnaSP 6.0 software. 8 haplotypes were determined according to the ITS sequence of 46 C. truncatum isolates from different countries. The median-joining network analysis of haplotypes was drawn using the NETWORK 10 program. It was determined that the CT isolate reported with this study from Turkey and the other reference isolates reported on tomatoes were in the H1 which is the most common haplotype.

Keywords

• Colletotrichum
• Haplotype
• Phylogenetic analysis
• Network analysis
• Tomato

Türkiye'de Sera Domatesinde Colletotrichum truncatum’un Karakterizasyonu ve Haplotip Analizi

Öz

C. truncatum'un (Schwein.) Andrus and W.D. Moore neden olduğu antraknoz çoğu tropikal ürünün ekonomik olarak önemli bir hastalığıdır. Son yıllarda domateste de patojen olduğu rapor edilmiştir. Bu çalışmada, enfekteli meyveler 2019 yılında Antalya ilinden elde edilmiştir. İzolatlar tek spor alınarak saflaştırıldı. Konidiler 22.5-32.5 × 3.75 µm olarak ölçülmüştür. DNA izolasyonu CTAB metodu kullanılarak gerçekleştirildi. PCR amplifikasyonundan sonra, PCR ürünleri agaroz jelde koşturuldu, UV cihazı ile görüntülendi ve sekanslandı. Filogenetik analiz MEGA 7'de yapılmıştır. Morfolojik ve filogenetik analizlere dayanarak CT izolatı C. truncatum olarak tanımlanmıştır. Patojenite testleri domates yaprakları ve çeri domatesleri kullanılarak gerçekleştirildi. İnokule edilen yapraklar ve domatesler, 27 °C sıcaklık ve 12:12 saat aydınlık-karanlık koşulları altındaki iklim odasında nemli bir filtre kağıdı üzerinde inkübe edilmiştir. İnokülasyonun 7. gününde enfekteli dokularda acervuli gözlenmiştir. Haplotip, haplotip sayısı ve nükleotid çeşitliliği DnaSP 6.0 yazılımı ile analiz edilmiştir. Farklı ülkelerden 46 C. truncatum izolatının ITS sekansına göre 8 haplotip belirlenmiştir. Haplotiplerin medyan birleştirme ağı analizi NETWORK 10 programı kullanılarak çizilmiştir. Türkiye'den bu çalışma ile rapor edilen CT izolatının ve domateste rapor edilen diğer referans izolatların en yaygın haplotip olan H1'de olduğu belirlenmiştir.

Anahtar Kelimeler

• Domates,
• Colletotrichum,
• Haplotip,
• Filogenetik analiz,
• Network analizi

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