Molecular identification of the spongy moth parasitoid Phryxe heraclei (Tachinidae) in Kazakhstan.

The spongy moth (Lymantria dispar) is a harmful phytophagous insect, causing significant damage to forest ecosystems. During the collection of Lymantria dispar samples, a parasitoid developing in the pupae of this pest was discovered. A parasitic fly emerged after two days of incubation. Judging from morphological characteristics, the fly was identified as a dipteriae from the family Tachinidae. To determine the species, a molecular genetic analysis was conducted.

gDNA was extracted using the CTAB method. Barcoding was performed using universal primers targeting 16S rRNA (5'-TARTYCAACATCGRGGTC-3' and 5'-CYGTRCDAAGGTAGCATA-3') gene. Then, species identification was performed using ONT sequencing. Reads were mapped against a database of 88274 dipteran rRNA sequences using minimap2 (V.2.28), further processed with SAMtools (V.1.21). Quality metrics visualized in R (V.4.4.2) using Rsamtools and ggplot2 packages.

The sequencing analysis generated 5,066 total reads, of which 4,751 (93.78%) were successfully mapped to reference sequences. A total of 704 reads were mapped to the Phryxe heraclei mitochondrial 16S rRNA gene (GenBank accession no. AB465921.1), with mapping quality scores (MAPQ) ranging from 40 to 60. While other species such as Pexopsis pilosa and Bothria japonica showed mapped reads, these alignments were predominantly of low quality (MAPQ <20) with scattered coverage patterns.

A literature analysis showed that this parasitoid is one of the main natural enemies of the spongy moth in the Far East. However, there are no data on its detection or research in Kazakhstan. Nevertheless, the molecular analysis provided strong evidence supporting the identification of the parasitoid as Phryxe heraclei.

This research was funded by the Ministry of Agriculture of the Republic of Kazakhstan (Program No. BR22887230).