Çanakkale’de yeşil kurt Helicoverpa armigera (Lepidoptera: Noctuidae) popülasyonun genetik çeşitliliği

Öz

Yeşil Kurt, Helicoverpa armigera (Hübner) (Lepidoptera: Noctuidae) dünyada tarımsal alanlarda yaygın olarak bilinen polifag bir zararlıdır. Domates üretiminde kalite ve ürünlerde önemli zararlara sebep olup, Türkiye’ de 1913 yılından beri bilinmektedir. Çalışmanın amacı, yeşil kurdun mitekondrial COI genindeki genetik çeşitliliği ve filogenetik benzerliğinin belirlenmesidir. 2017 ve 2018 yıllarında yeşil kurt ile bulaşık domatesler Çanakkale (14 alandan), Balıkesir (1 alan) ve Bursa (1 alandan) toplandı. Her örnekleme alanın temsil eden larvalar (n=3) kullanılarak genomik DNA izolasyonu yapıldı. COI geninin bir kısmı (536 bp) çoğaltılarak PCR ürünleri direkt olarak dizilimleri belirlendi. DnaSP ile haploit sayıları ve mutasyobları belirlendi. Populasyonlar arası genetik akrabalık düzeyleri UPGMA metodu ile MEGA X programında belirlendi. Sonuç olarak, test edilen 45 örnekte13 haplotip ve 13 mutasyon belirlendi. Buna ilave olarak yeşil kurtta 9 özel haplotip bulundu. Analizler sonucunda farklı cografik alanlardan elde edilen yeşil kurt popülasyonları 2 ayrı grupta yer aldı. İlk grup 12 haplotip (Çanakkale ve Balıkesir popülasyonu) ve diğer grupta ise sadec 1 haplotip (Bursa popülasyonu) yer aldı. Bu çalışma, Türkiye’de yeşil kurdun moleküler karekterizasyonu ve genetik çeşitliliğinin belirlenmesi için ilk kez yapılmıştır.

Anahtar Kelimeler

• Helicoverpa armigera,
• Cotton Bollworm,
• Genetic diveristy,
• mtDNA,
• COI gene

Proje Numarası

FYL-2018/2466

Genetic Diversity of Cotton Bollworm, Helicoverpa armigera (Lepidoptera: Noctuidae) Population in Çanakkale

Öz

The cotton bollworm, Helicoverpa armigera (Hübner) (Lepidoptera: Noctuidae) is a commonly known polyphagous pest in agricultural areas all over the world. It has been a major pest causing significant damages by reducing tomato fruit production and quality since 1913 in Turkey. The objective of the study was to evaluate genetic diversity and phylogenetic relationship of H. armigera based on mitochondrial COI gene. Infested tomatoes with H. armigera larvae were collected from Çanakkale province (14 location), Balıkesir (1 location) and Bursa (1 location) in August and September 2017 and 2018. Larvae representing for each location (n=3) were used for genomic DNA isolation. The partial COI gene (536 bp) was amplified and PCR products were sequenced directly. The numbers of haplotypes and mutations were estimated using the DnaSP. Genetic relationships between the population were evaluated using the UPGMA method in MEGA X program. As a result, we identified 13 haplotypes and 13 mutations in all tested 45 samples. Additionally, 9 private haplotypes were detected among H. armigera population. Analyses revealed that population of H. armigera sampled from different geographic location were established as two distinct groups. The first group comprised 12 haplotypes (Çanakkale and Balıkesir population) while the distinct second group consisted of only 1 haplotype (Bursa population). The presented study was the first attempt to detect molecular characterization and genetic characterization of cotton bollworm in Turkey.

Anahtar Kelimeler

• Helicoverpa armigera
• Cotton bollworm
• Genetic diversity
• mtDNA
• COI gene

Destekleyen Kurum

Çanakkale Onsekiz Mart Üniversitesi BAP

Proje Numarası

FYL-2018/2466

Teşekkür

This research was financially supported by Canakkale Onsekiz Mart University, Scientific Research Projects Commission (FYL-2018/2466). The authors thank Emeritus Prof. Dr. James L. Nation for reviewing and his useful suggestions. The part of this study was published as an abstract in the 1st International Molecular Plant Protection Congress, April 10-13, 2019 in Adana, Turkey.

Kaynakça

1. Albernaz, K.C., Silva-Brandao, K.L., Fresia, P., Consoli, F.L., Omoto, C., 2012. Genetic variability and demographic history of Heliothis virescens (Lepidoptera: Noctuidae) populations from Brazil inferred by mtDNA sequences. Bulletin of Entomological Research. 102: 333-343.
2. Alkan, B., 1948. Orta Anadolu Hububat Zararlıları (Zararlı Hayvan ve Böcekler). A.Ü. Ziraat Fakültesi Yayınları. Sayı:1, Ankara Üniversitesi Basımevi, Ankara, 71 s.
3. Asokan, R., Rebijith, K.B., Krishna Kumar, N.K., Manamohan, M., 2012. Genetic diversity of tomato fruit borer, Helicoverpa armigera Hübner (Lepidoptera: Noctuidae) inferred from mitochondrial cytochrome oxidase-I (mtCO-I). Pest Management in Horticultural Ecosystems. 18 (1): 29-34.
4. Behere, G.T., Tay, W.T., Russell, D.A., Heckel, D.G., Appleton, B.R., Kranthi, K.R., Batterham, P., 2007. Mitochondrial DNA analysis of field populations of Helicoverpa armigera (Lepidoptera: Noctuidae) and of its relationship to H. zea. BMC Evolutionary Biology. 7: 117-127.
5. Behere, G.T., Tay, W.T., Russell, D.A., Batterham, P., 2007. Molecular markers to discriminate among four pest species of Helicoverpa (Lepidoptera: Noctuidae). Bulletin of Entomological Research. 98: 599-603.
6. Behere, G.T., Tay, W.T., Russell, D.A., Kranthi, K.R., Batterham, P., 2013. Population genetic structure of the cotton bollworm Helicoverpa armigera (Hübner) (Lepidoptera: Noctuidae) in India as Inferred from EPIC-PCR DNA Markers. PLoS ONE. 8: e53448.
7. Boore, J.L. 1999. Animal mitochondrial genome. Nucleic Acids Research. 27: 1767-1780.
8. Excoffier, L., Foll, M., Petit, R.J., 2009. Genetic consequences of range expansions. Annual Review of Ecology Evolution and Systematics. 40: 481-501.

9. Excoffier, L., Lischer, H.E.L., 2010. Arlequin suite ver 3.5: A new series of programs to perform population genetics analyses under Linux and Windows. Molecular Ecology Resources. 10: 564-567.
10. Felsenstein, J., 1985. Confidence limits on phylogenies: an approach using the bootstrap. Evolution 39: 783-791.
11. Fitt, G.P., 1989. The ecology of Heliothis species in relation to agroecosystems. Annual Review of Entomology. 34: 17-52.
12. Folmer, O., Black, M., Hoeh, W., Lutz, R., Vrijenhoek, R., 1994. DNA primers for amplification of mitochondrial cytochrome c oxidase subunit I from diverse metazoan invertebrates. Molecular Marine Biology and Biotechnology. 3: 294-299.
13. Fu, Y.X., 1997. Statistical tests of neutrality of mutations against population growth, hitchhiking and background selection. Genetics. 147:915-925.
14. Kimura, M., 1980. A simple method for estimating evolutionary rates of base substitutions through comparative studies of nucleotide sequences. Journal of Molecular Evolution. 16: 111-120.
15. Kumar, S., Stecher, G., Li, M., Knyaz, C., Tamura, K., 2018. MEGA X: Molecular Evolutionary Genetics Analysis across computing platforms. Molecular Biology and Evolution. 35: 1547-1549.
16. Leigh, J.W., Bryant, D., 2015. PopART: Full-feature software for haplotype network construction. Methods in Ecology and Evolution. 6: 1110-1116.
17. Leite, N.A., Alves-Pereira, A., Corrêa, A.S., Zucchi, M.I., Omoto, C., 2014. Demographics and genetic variability of the New World Bollworm (Helicoverpa zea) and the Old World Bollworm (Helicoverpa armigera) in Brazil. PLoS One .9:e11328.
18. Librado, P., Rozas, J., 2009. DnaSP v5: A software for comprehensive analysis of DNA polymorphism data. Bioinformatics. 25: 1451-1452.
19. Mironidis, G.K., Savopoulou-Soultani, M., 2008. Development, survivorship and reproduction of Helicoverpa armigera (Lepidoptera: Noctuidae) under constant and alternating temperatures. Environmental Entomology. 37: 16-28.
20. Nei, M., 1987. Molecular evolutionary genetics. Columbia University Press, New York, NY. 512 pp.
21. Perera, O.P., Allen, K.C., Jain, D., Purcell, M., Little, N.S., Luttrell, R.G., 2015. Rapid Identification of Helicoverpa armigera and Helicoverpa zea (Lepidoptera: Noctuidae) Using Ribosomal RNA Internal Transcribed Spacer 1. Journal of Insect Science. 15 (1): 1-10.
22. Pogue, M.G., 2004. A new synonym of Helicoverpa zea (Boddie) and differentiation of adult males of H. zea and H. armigera (Hübner) (Lepidoptera: Noctuidae: Heliothinae). Annals of the Entomological Society of America. 97: 1222-1226.
23. Rogers, A.R., Harpending, H., 1992. Population growth makes waves in the distribution of pairwise genetic differences. Molecular Biology and Evolution. 9: 552-569.
24. Rozas, J., Ferrer-Mata, A., Sánchez-DelBarrio, J.C., Guirao-Rico, S., Librado, P., Ramos-Onsins, S.E., Sánchez-Gracia, A., 2017. DNA sequence polymorphism analysis of large data sets. Molecular Biology and Evolution 34. (12): 3299-3302.
25. Saitou, N., Nei, M., 1987. The neighbor-joining method: a new method for reconstructing phylogenetic trees. Molecular Biology and Evolution. 4: 406-425.
26. Simon, C., Frati, F., Beckenbach, A., Crespi, B., Liu, H., Flook, P., 1994. Evolution, weighting, and a phylogenetic utility of mitochondrial gene sequences and a compilation of conserved polymerase chain reaction. Annals of the Entomological Society of America. 87: 651-701.
27. Tajima, F., 1989. The effect of change in population size on DNA polymorphism. Genetics. 123: 585-595.
28. Tamura, K., Nei, M., Kumar, S., 2004. Prospects for inferring very large phylogenesis by using the neighbor-joining method. Proceedings of the National Academy of Sciences of the USA. 101: 11030-11035.
29. Thompson, J.D., Higgins, D.G., Gibson, T.J., 1994. CLUSTAL W: Improving the sensitivity of progressive multiple sequence alignment through sequence weighting, position-specific gap penalties and weight matrix choice. Nucleic Acids Research. 22 (22): 4673-4680.
30. Uğurlu, S., Gürkan, M.O., 2007. Insecticide resistance in Helicoverpa armigera from cotton-growing areas in Turkey. Phytoparasitica. 35 (4):376-279.
31. Zhang, D.X., Hewitt, G.M., 1996. Nuclear integrations: challenges for mitochondrial DNA markers. Trends in Ecology & Evolution. 11: 247-251.