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Abstract: One of the key steps of the molecular identification of bark beetles is obtaining a sufficient quantity of high-quality DNA extract. In this study, we investigated the influence of different storage procedures for Ips typographus (L.) specimens and various DNA extraction protocols on the quantity and quality of DNA intended for use in molecular diagnostics. Adult beetles were frozen at -20 °C, either dry or in ethanol. We tested four different protocols for DNA extraction. We compared the quantity of extracted DNA and assessed its quality with PCR and Sanger sequencing. Different storage protocols had no significant effect on the quantity of DNA extracted. However, freezing specimens in ethanol provided higher-quality DNA for molecular applications. Only two of the extraction protocols produced sequenceable amplicons, and the difference in the amount of extracted DNA between them was not significant. We propose the optimal combination of storing specimens in ethanol at -20°C and using the Nucleospin Insect DNA extraction kit from Macherey Nagel, enabling a timeefficient identification process.
Keywords: early detection, specimen storage, total DNA extraction, PCR, polymerase chain reaction, Sanger sequencing, molecular diagnostics
Published in DiRROS: 02.02.2024; Views: 958; Downloads: 369
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Abstract: The northern spruce bark beetle, Ips duplicatus (Sahlberg), has invaded Central Europe in the past century and continues to spread southwards and westwards. It has caused damage in some parts of Europe in recent decades and poses a serious threat to spruce forests across the continent. From 2018 to 2021, we conducted intensive monitoring to determine the status of I. duplicatus in Slovenia. We used different types of traps and pheromone lures that attract I. duplicatus beetles and took bark samples from bark beetle-infested trees. Of the 26 traps used during the four years of monitoring, I. duplicatus was found in 16 traps in the central part of Slovenia in 2020. Adult beetles were caught in traps containing both I. typographus and I. duplicatus lures. The identity of this species was confirmed using morphological keys and molecular methods. One to five I. duplicatus beetles were found per trap, totaling 25 specimens. However, we did not confirm any I. duplicatus in samples taken from trees in 2021. We conclude that I. duplicatus is present in Slovenia, but our results suggest that the species is only present locally and that the population is small.
Keywords: biological invasions, forest pests, temperate forests, conifers, damage, forest pest monitoring, confirmation
Published in DiRROS: 05.09.2023; Views: 655; Downloads: 331
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Keywords: varstvo gozdov, morfološke analize, Pinus nigra, žuželke, ličinke, Monochamus sp., črni bor
Published in DiRROS: 16.12.2022; Views: 615; Downloads: 217
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Keywords: varstvo gozdov, morfološke analize, Xylosandrus crassisculus, azijski ambrozijski podlubnik
Published in DiRROS: 16.11.2022; Views: 749; Downloads: 0

Keywords: varstvo gozdov, morfološke analize, Xylosandrus crassisculus, azijski ambrozijski podlubnik
Published in DiRROS: 16.11.2022; Views: 685; Downloads: 0