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The effectiveness of vermiculture using earthworms largely depends on the species. In Ukraine, the cultivation of the red California worm, commonly recognised as Eisenia fetida, is becoming increasingly popular. Although this name typically refers to this specific species, Dendrobaena veneta is sometimes included as well. While both species are utilised in vermiculture for industrial production, they are indeed different types of worms. The sale of brood stock, regardless of the name used, relies primarily on physical characteristics assessed through visual inspection. However, trying to identify earthworm species by their physical features alone is often a guessing game, since these creatures lack the intricate and distinctive structures found in other animals. Tracing the origins of products made from such earthworms becomes an even greater challenge. To reduce potential errors and prevent misuse, a genetic method has been developed to identify two specific earthworm species using single-nucleotide polymorphisms (SNPs) in the mitochondrial COI gene. Research was performed on four species of earthworms from the Lumbricidae family at the genetics laboratory of the Institute of Pig Breeding and AIP of the NAAS. Forward primers and reverse primers targeting the cytochrome c oxidase (COI) region were developed for the mitochondrial genome of E. fetida, D. veneta, and other representatives of the Lumbricidae family. In the forward primer sequences of E. fetida and D. veneta, one nucleotide substitution was observed in the reverse three. Despite this, specific PCR products were obtained using these primer pairs for the species E. fetida, E. andrei, D. veneta and Lumbricus terrestris. Nevertheless, specific PCR products were successfully obtained using these primer pairs for the species E. fetida, E. andrei, D. veneta, and L. terrestris. The size of the amplification products, which is 253 base pairs, matches the length predicted from the nucleotide sequence. An analysis of restriction fragments from amplified mitochondrial COI gene DNA samples of four species from the Lumbricidae family revealed three distinct sets of bands. As expected, the restriction fragments for E. fetida and E. andrei were identical. In contrast, the restriction fragments of D. veneta exhibited different characteristics, as predicted by bioinformatic analysis. Furthermore, it was confirmed that DNA from L. terrestris is suitable for use as a negative control. Therefore, the proposed method for genetic identification of the earthworm species E. fetida and D. veneta using PCR-RFLP may be useful for routine analyses.

 

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