Age influences the olfactory profiles of the migratory oriental armyworm mythimna separate at the molecular level

Abstract

Background: The oriental armyworm Mythimna separata (Walk) is a serious migratory pest; however, studies on its olfactory response and its underlying molecular mechanism are limited. To gain insights to the olfactory mechanism of migration, olfactory genes were identified using antennal transcriptome analysis. The olfactory response and the expression of olfactory genes for 1-day and 5-day-old moths were respectively investigated by EAG and RT-qPCR analyses.

Results: Putative 126 olfactory genes were identified in M. separata, which included 43 ORs, 13 GRs, 16 IRs, 37 OBPs, 14 CSPs, and 3 SNMPs. RPKM values of IR75d and 10 ORs were larger than co-receptors IR25a and ORco, and the RPKM value of PR2 was larger than that of other ORs. Expression of GR1 (sweet receptor) was higher than that of other GRs. Several sex pheromones activated evident EAG responses where the responses of 5-day-old male moths to the sex pheromones were significantly greater than those of female and 1-day old male moths. In accordance with the EAG response, 11 pheromone genes, including 6 PRs and 5 PBPs were identified in M. separate, and the expression levels of 7 pheromone genes in 5-day-old moths were significantly higher than those of females and 1-day-old moths. PR2 and PBP2 might be used in identifying Z11-16: Ald, which is the main sex pheromone component of M. separata. EAG responses to 16 plant volatiles and the expression levels of 43 olfactory genes in 1-day-old moths were significantly greater than that observed in the 5-day-old moths. Heptanal, Z6-nonenal, and benzaldehyde might be very important floral volatiles for host searching and recognized by several olfactory genes with high expression. Some plant volatiles might be important to male moths because the EAG response to 16 plant volatiles and the expression of 43 olfactory genes were significantly larger in males than in females.

Conclusions: The findings of the present study show the effect of adult age on olfactory responses and expression profile of olfactory genes in the migratory pest M. separate.

Keywords: Co-receptor; Host searching; Migration; Mythimna separata; Olfaction; Pre-mating status; Sex pheromone.

Fig. 1

eggNOG annotation of aligned genes from M. separata antennal transcriptome. The Y-axis shows the number of the unigene, the X-axis the category of annotation

Fig. 2

Analysis of different expressed genes (red) in the male and female antennae of M. separata. If log2 (fold change) > 1, then this gene is expressed at a higher level in the male antennae; if log2 < −1, then the gene shows a higher expression in the female antennae. The Y-axis shows the p-value of the unigene, the X-axis the fold-change of gene expression (FPKM) in male antennae to female. A total of 28 genes showed a higher expression in the male antennae, and 118 genes were expressed at higher levels in the female antennae. FPKM: fragments per kb per million fragments; DEGs: differentially expressed genes

Fig. 3

Aligned putative OR gene sequences of M. separata (black circle). Bootstrap values < 50% were ignored. Msep, M. separata, Bmor, B. mori, Hvir, Heliothis virescens, Ofur, O. furnacalis, Slitu, S. litura

Fig. 4

Aligned putative GR gene sequences of M. separata (black circle), D. melanogaster (red lines) and other moth species (black lines). Bootstrap values < 50% were ignored. Msep, M. separata, Dmel, D. melanogaster, Bmor, B. mori

Fig. 5

Aligned putative IR gene sequences of M. separata (black circle). Bootstrap values < 50% were ignored. Msep, M. separata, Dmel, D. melanogaster, Bmor, B. mori

Fig. 6

Aligned putative OBP gene sequences of M. separata (black circle). Bootstrap values lower than 50% were ignored. Msep, M. separata, Bmor, B. mori, Slitu, S. litura, Hvir, Heliothis virescens, Csup, C. suppressalis

Fig. 7

Aligned putative CSP gene sequences of M. separata (black circle). Bootstrap values < 50% were ignored. Msep, M. separata, Bmor, B. mori, Hvir, Heliothis virescens, Csup, C. suppressalis

Fig. 8

Aligned putative SNMP gene sequences of M. separata (black circle). Bootstrap values < 50% were ignored. Msep, M. separata, Dmel, D. melanogaster, Bmor, B. mori, Msex, M. sexta, Hvir, Heliothis virescens, Harm, Helicoverpa armigera

Fig. 9

Expression levels of olfactory genes in male and female M. separata antennae measured by RNA-Seq. Expression was calculated with log scale of RPKM value

Fig. 10

Expression levels of olfactory genes in male and female antennae with different day time measured by RT-qPCR. Gene expression was calculated relative to the Actin and AK as reference genes and expression in 1-day-old female antennae was arbitrarily defined as control for all genes. Gene expression in other tissue were normalized to 1-day-old female antennae. Log scale of gene expression was used to generate heatmap. F1, 1-day-old female, F5, 5-day-old female, M1, 1-day-old male, M5, 5-day-old male

Fig. 11

EAG response of male and female moth with different day time to sex pheromone with 10⁻⁴ (left) and 10⁻² (right) concentration. F1, 1-day-old female, F5, 5-day-old female, M1, 1-day-old male, M5, 5-day-old male

Fig. 12

EAG response of male and female moth with different day time to plant volatile with 10⁻⁴ (left) and 10⁻² (right) concentration. F1, 1-day-old female, F5, 5-day-old female, M1, 1-day-old male, M5, 5-day-old male