Molecular diagnostics of viruses in apple rootstocks using RT-qPCR

Authors

DOI:

https://doi.org/10.31489/2026feb1/12-20

Keywords:

Malus domestica, rootstock, viruses, viroids, RT-qPCR diagnostics

Abstract

The development of intensive horticulture in Kazakhstan requires the use of virus-free planting material, particularly apple rootstocks (Malus domestica), free of viral and viroid infections. Latent forms of pathogens, which do not exhibit visible symptoms but are easily transmitted through vegetative propagation, are consid ered the most hazardous. The objective of this study was to evaluate the phytosanitary status of apple root stocks used in nurseries in southern Kazakhstan by employing reverse transcription quantitative polymerase chain reaction (RT-qPCR). A total of 24 samples from field and laboratory collections were analyzed. RNA extraction was performed using the “PhytoSorb®” kit, and diagnostics were conducted on the Bio-Rad CFX96 platform using multiplex LETGEN test kits. Four pathogens were identified: Apple chlorotic leaf spot virus (ACLSV), Apple stem pitting virus (ASPV), Apple green crinkle associated virus (AGCaV), and Apple hammerhead viroid (AHVd). The viruses ASPV and AGCaV were found to be the most prevalent, occurring in both field and laboratory samples. These findings confirm the widespread circulation of these pathogens within the apple rootstock propagation system. The results emphasize the need for the implementation of regular molecular diagnostics, sanitation programs, and certification measures aimed at preventing the spread of latent infections and ensuring the sustainable development of horticulture in the region.

Author Biographies

M. Askarova, JSC "Almaty Technological University"

PhD student, Department of Biotechnology, JSC "Almaty Technological University"; Senior Researcher, Department of Plant Protection, LLP "Kazakh Research Institute of Fruit and Vegetable Growing", Almaty, Republic of Kazakhstan

Z. Yussupova , LLP "Kazakh Research Institute of Fruit and Vegetable Growing"

Master of Agricultural Sciences,Researcher, "In Vitro Gene Pool" Laboratory, LLP "Kazakh Research Institute of Fruit and Vegetable Growing", Almaty, Republic of Kazakhstan

A. Madenova, LLP "Kazakh Research Institute of Fruit and Vegetable Growing"

PhD, Associate Professor, Leading Researcher, "In Vitro Gene Pool" Laboratory, LLP "Kazakh Research Institute of Fruit and Vegetable Growing", Almaty, Republic of Kazakhstan

R. Abdikarimova, LLP "Kazakh Research Institute of Fruit and Vegetable Growing"

Master of Agricultural Sciences,Junior Researcher, "In Vitro Gene Pool" Laboratory, LLP "Kazakh Research Institute of Fruit and Vegetable Growing", Almaty, Republic of Kazakhstan

B. Kabylbekova, LLP "Kazakh Research Institute of Fruit and Vegetable Growing"

PhD, «In vitro генофонды» зертханасының меңгерушісі, «Қазақ жеміс және көкөніс шаруашылығы ғылыми-зерттеу институты» ЖШС, Алматы қ., Қазақстан Республикасы

References

Ромаданова Н. В., Кушнаренко С. В. Биотехнология получения безвирусных саженцев яблони // Вестник Карагандинского университета. Серия: Биология. – 2021. – № 3(103). – С. 102–118. – Режим доступа: https://dist.buketov.edu.kz/apart/2021-103-3/13.pdf.

Кабылбекова Б. Ж., Нурсейтова Т., Юсупова З., Ковальчук И., Чуканова Н., Уразаева М., Маденова А. К. Определение состава вирусных болезней клоновых подвоев косточковых культур в Казахстане и особенности размножения in vitro здоровых растений // Ізденістер, нәтижелер – Исследования, результаты. – 2024. – № 3(103). – С. 86–97. – Режим доступа: https://journal.kaznaru.edu.kz/index.php/research/article/view/696.

Kerimbek N., Khusnitdinova M., Taskuzhina A., Kapytina A., Pozharsky A., Sagitov A., Gritsenko D. Mapping the distribution of viruses in wild apple populations in the southeast region of Kazakhstan // Forests. – 2025. – Vol. 16(7). – Article 1119. – DOI: https://doi.org/10.3390/f16071119.

Kerimbek N., Kapytina A. I., Khusnitdinova M. A., Taskuzhina A. K. Assessment of occurrence and diversity of apple viruses in South Kazakhstan // Ғылым жәнi білім (Ғылыми журнал). – 2024. – Т. 1, № 2. – С. 79–87. – Режим доступа: https://www.researchgate.net/publication/391587146.

Kairova A. Zh., Baymoldina G. M., Sharipova A. B., Argyngaliyeva Zh. T., Shaimardanova A. S., Akhmetova A. K., Kairov E. A. Assessment of apple (Malus domestica Borkh.) rootstocks for resistance to fire blight (Erwinia amylovora) in Southern Kazakhstan // Applied Sciences. – 2023. – Vol. 13(20). – P. 11530. – DOI: https://doi.org/10.3390/app132011530.

Wunsch A., Hoff B., Miranda Sazo M., van Zoeren J., Lamour K. H., Hurtado Gonzales O. P., Fuchs M. Viruses of apple are seedborne but likely not vertically transmitted // Viruses. – 2024. – Vol. 16(1). – P. 95. – DOI: https://doi.org/10.3390/v16010095.

Ben Mansour K., Koloniuk I., Brožová J., Komínková M., Přibylová J., Sarkisova T., Sedlák J., Špak J., Komínek P. High throughput sequencing reveals apple virome diversity and novel viruses in the Czech Republic // Viruses. – 2025. – Vol. 17(5). – Article 650. – DOI: https://doi.org/10.3390/v17050650.

Omasheva M. Y., Pozharskiy A., Maulenbay A., Ryabushkina N. A. SSR genotyping of Kazakhstani apple varieties: Identification of alleles associated with resistance to highly destructive pathogens // Biotechnology: Theory and Practice. – 2016. – June. – P. 350–356. – Режим доступа: https://biotechlink.org/index.php/journal/article/view/168.

Gritsenko D., Aubakirova K., Voitsekhovskiy I., Galiakparov N. Simultaneous detection of five apple viruses by RT-PCR // International Journal of Biology and Chemistry. – 2020. – Vol. 13(1). – DOI: https://doi.org/10.26577/ijbch.2020.v13.i1.13.

OEPP/EPPO. Certification scheme for pathogen-tested material of Malus, Pyrus and Cydonia (Standard PM 4/27(1)). – Paris: European and Mediterranean Plant Protection Organization, 1999. – Режим доступа: https://gd.eppo.int/standards/PM4/PM4-27-1.

Mekuria G., Rhamesh S. A., Alberts E., Bertozzi T., Wirthensohn M., Collins G., Sedgley M. Comparison of ELISA and RT-PCR for detection of Prunus necrotic ringspot virus and Prune dwarf virus in almond (Prunus dulcis) // Journal of Virological Methods. – 2003. – Vol. 114. – P. 65–69. – Режим доступа: https://pubmed.ncbi.nlm.nih.gov/14599680.

Galymbek K., Madenova A., Bakirov S. Phytosanitary diagnosis of apple fungal diseases in Almaty region // Research on Crops. – 2024. – Vol. 25. – P. 499–508. – DOI: http://dx.doi.org/10.31830/2348-7542.2024.ROC-1112.

Fazio G., Bettoni J. C., Costa L. C., Hurtado-Gonzales O. P., Al Rwahnih M., Steinberger A., Robinson T. Virus studies in the Geneva® apple rootstock breeding program // New York Fruit Quarterly. – 2022. – P. 20–24. – Режим доступа: https://www.researchgate.net/publication/362123931.

James D. Perspective on strategies for controlling the spread of Plum pox virus, causal agent of sharka/plum pox disease // III International Symposium on Plum Pox Virus. – 2016. – Vol. 1163. – P. 129–136. – DOI: https://doi.org/10.17660/ActaHortic.2017.1163.19.

Manzoor S., Nabi S. U., Ayaz A., Rasool B., Sharma S. K., Chesti M. H., Rabbee M. F. Unveiling viral threats to temperate pome fruits: Characterization, transmission, and sustainable management strategies // Frontiers in Microbiology. – 2025. – Vol. 16. – Article 1560720. – DOI: https://doi.org/10.3389/fmicb.2025.1560720.

Wright A. A., Cross A. R., Harper S. J. A bushel of viruses: Identification of seventeen novel putative viruses by RNA seq in six apple trees // PLoS ONE. – 2020. – Vol. 15(1). – e0227669. – DOI: https://doi.org/10.1371/journal.pone.0227669.

Pedrelli A., Nali C., Panattoni A., Pellegrini E., Cotrozzi L. Apple chlorotic leaf spot virus infection affects the physiology and biochemistry of leaves and the quality of fruits of Tuscany (Italy) autochthonous apple varieties // Plant Pathology. – 2025. – Vol. 74(1). – DOI: https://doi.org/10.1111/ppa.14004.

Serra P., Messmer A., Sanderson D., James D., Flores R. Apple hammerhead viroid-like RNA is a bona fide viroid: Autonomous replication and structural features support its inclusion as a new member in the genus Pelamoviroid // Virus Research. – 2018. – Vol. 249. – P. 8–15. – DOI: https://doi.org/10.1016/J.VIRUSRES.2018.03.001.

Sedlák J., Přibylová J., Koloňuk I., Špak J., Lenz O., Semerák M. Elimination of Solanum nigrum ilarvirus 1 and apple hammerhead viroid from apple cultivars using antivirals ribavirin, rimantadine, and zidovudine // Viruses. – 2023. – Vol. 15(8). – P. 1684. – DOI: https://doi.org/10.3390/v15081684.

Маденова А. К., Калдыбаева Д. И., Әбдікәрімова Р. А., Қуан А. А. Идентификация вирусных заболеваний яблони в казахстанских садах методом ОТ-ПЦР-РВ // Ізденістер, нәтижелер – Исследования, результаты. – 2024. – № 3(103). – С. 97–107. – DOI: https://doi.org/10.37884/3-2024/11.

Guo X., Huang S., Wu S., Liu Y., Wang M., Song C., Bai T., Song S., Pang H., Jiao C., Zheng S. Identification and analysis of apple virus diseases based on virome sequencing technology // Acta Horticulturae Sinica. – 2022. – Vol. 49(7). – P. 1415–1428. – DOI: https://doi.org/10.16420/j.issn.0513-353x.2021-0450.

Kabylbekova B., Nurseitova T., Yussupova Z., Turdiyev T., Kovalchuk I., Dolgikh S., Soltanbekov S., Seisenova A., Madenova A. Application of In Vitro Techniques for Elimination of Plum Pox Virus (PPV) and Apple Chlorotic Leaf Spot Virus (ACLSV) in Stone Fruits // Horticulturae. – 2025. – Vol. 11(6). – P. 633. – DOI: https://doi.org/10.3390/horticulturae11060633.

Downloads

Published

2026-03-30

How to Cite

Askarova, M., Yussupova , Z., Madenova, A., Abdikarimova, R., & Kabylbekova, B. (2026). Molecular diagnostics of viruses in apple rootstocks using RT-qPCR. Fundamental and Experimental Biology, 12131(1), 12–20. https://doi.org/10.31489/2026feb1/12-20

Issue

Section

Articles