Abstract
This study reports the first complete mitochondrial genomes of three egg parasitoid wasps parasitizing Nilaparvata lugens-Pseudoligosita nephotetticum, Anagrus frequens, and Anagrus nilaparvatae. Genome sizes ranged from 15,429 to 15,889 bp, with all three mitogenomes displaying strong A + T bias, standard gene content, and characteristic strand asymmetries. While A. frequens and A. nilaparvatae exhibited conserved gene orders, extensive gene rearrangements, including multiple inversions in both protein-coding genes (PCGs) and tRNAs, were observed in P. nephotetticum. Codon usage analyses revealed a preference for codons ending in A or U. The non-synonymous (Ka) to synonymous (Ks) substitution ratio analysis identified signs of positive selection in multiple PCGs, particularly in atp8, nad6, and nad3, suggesting possible adaptive evolution related to host-searching behavior. Secondary structure analyses showed the loss of trnL1 in all Anagrus species, while trnS1 and trnR lacked the DHU arm, indicating possible derived traits in Mymaridae. Phylogenetic analysis was the first time to describe the relationship of the genus Anagrus within Mymaridae from the perspective of 13 protein genes. Furthermore, the grouping of (Pseudoligosita + Megaphragma) + Trichogramma was supporting the distinct evolutionary lineage of Pseudoligosita. This work provides new molecular resources and phylogenetic insight for Chalcidoidea, with implications for parasitoid evolution and biological control strategies.