The oriental armyworm, Mythimna separata, is a serious agricultural pest in China. Seasonal and roundtrip migration has recently led to sudden, localized outbreaks and crop losses. To evaluate genetic differentiation between populations in eastern and western China and elucidate gene flow, the genetic structure of 20 natural populations from nine provinces was examined using seven microsatellite markers. The results indicated high genetic diversity. However, little to moderate (0 < F_ST < 0.15) genetic differentiation was detected, and there was no correlation between genetic distance and geographical distance. Bayesian clustering analysis identified three groups whereas discriminant analysis of principal components identified ten clusters that were considered as two clear‐cut clusters and one admixed group. Gene flow occurred frequently in most population pairs, and an asymmetrical migration rate was detected in several pairwise population comparisons. The bottleneck test showed that few populations had experienced recent bottlenecks. Correspondingly, large‐scale and long‐distance migration of M. separata has caused low genetic differentiation and frequent gene exchange. Our findings are important for studying genetic evolution and help to improve predictions of M. separata outbreaks in China.

The oriental armyworm larvae samples were collected from maize (Z. mays) fields from nine provinces at 20 geographical sites in China. Seven  polymorphic microsatellite loci, A1–91, A3–81, JGTT29, MS1, MS2, MS3, and MS4, developed for M. separata were scored for genotyping. The PCR products of the target band were genotyped by capillary electrophoresis with two‐color fluorescent detection technique using an ABI3730XL  automatic sequencer. (Applied Biosystems). Microsatellite alleles for genotyping were estimated with GeneMarker 2.2 (SoftGenetics LLC) and were confirmed manually.

The microsatellite data file incorporates a total of 542 individuals from 20 geographical populations of the oriental armyworm, Mythimna separata in China. Seven polymorphic microsatellite loci, including A3-91, A3-81, JGTT29, MS1, MS2, MS3, and MS4 are used to genotype. Twenty populations are HBB, HBC, HNH, HNL, HNX, JLC, JLS, LNF, NME, SDJ, SDW, SXB, SXF, SXH, SXL, SXM, SXW, SXX, SYC, and YNZ, respectively. This dataset are associated to the article “Microsatellite-based analysis of genetic structure and gene flow of Mythimna separata (Walker) (Lepidoptera: Noctuidae) in China (ECE-2019-03-00269.R2)”, which has been accepted by journal Ecology and Evolution.

Seven polymorphic microsatellite loci, A1–91, A3–81, JGTT29, MS1, MS2, MS3, and MS4, developed for M. separata were scored for genotyping (Li et al., 2018; Zhang & Zhai, 2008). The missing values were denoted by 0.