Entomopoxvirus infection induces changes in both juvenile hormone and ecdysteroid levels in larval Mythimna separata

Abstract

Insect viruses are among the most important pathogens of lepidopteran insects. Virus-infected larvae often show developmental defects including a prolonged larval period and a failure to pupate, but the mechanisms by which insect viruses regulate host development need further investigation. In this study, the regulation of host endocrinology by a lepidopteran entomopoxvirus (EPV), Mythimna separata EPV (MySEV), was examined. When fourth instar M. separata were inoculated with MySEV occlusion bodies, pupation was prevented and the insects died during the final (sixth) larval instar. Liquid chromatography-MS analysis revealed that juvenile hormone (JH) titres in the haemolymph of MySEV-infected sixth instars were higher than those in mock-infected larvae. JH esterase (JHE) activity was also examined by kinetic assay using a colorimetric substrate. The level of JHE activity in the haemolymph of MySEVinfected larvae was generally lower than that found in mock-infected larvae. In contrast, ecdysteroid titre in the haemolymph of final-instar MySEV-infected larvae was lower than that found in mock-infected larvae when measured by radioimmunoassay. A statistically significant difference in the release of ecdysteroids from prothoracic glands (PGs) that were dissected from MySEV-or mock-infected sixth instar Day 3 larvae was not found following prothoracicotropic hormone (PTTH) exposure. Our results indicate that the release of ecdysteroids was reduced not by infection of the PGs by MySEV, but by reduced PTTH production from the brain. Taken together our study suggests that EPVs retard host development by both reducing ecdysone titre and maintaining status quo levels of JH by preventing its metabolism.

A virus carries a gene encoding juvenile hormone acid methyltransferase, a key regulatory enzyme in insect metamorphosis

“…MySEV was administered to newly moulted 4th instar larvae of the oriental armyworm Mythimna separata , and gene and protein expression were analysed from day 1 of 5th instar to early 6th instar (final instar). In accordance with previous results 10 , MySEV-infected insects developed at almost the same rate as mock-infected insects during 4th and 5th instars, but their development was prolonged substantially during the final (6th) instar compared to mock-infected insects; they eventually died as larvae, whereas all control insects pupated (Table 1 ). Reverse transcription PCR (RT-PCR) followed by melting curve analysis confirmed the expression of the MySEV SAM-dependent MTase gene in MySEV-infected insects throughout the period examined.…”

“…We next analysed gene and protein expression under the same conditions used in a previous study where the physiology of MySEV-infected insects, including JH titres, was examined 10 . MySEV was administered to newly moulted 4th instar larvae of the oriental armyworm Mythimna separata , and gene and protein expression were analysed from day 1 of 5th instar to early 6th instar (final instar).…”

“…JH titres in normal insects are regulated by synthesis with JHAMT and catabolism with esterases, mainly with JH esterase (JHE). Several possible mechanisms have been proposed to explain the higher JH titres in EPV-infected insects than in uninfected insects 10 , 11 . For example, JHE is produced in fat body, which is a major replication tissue for EPV.…”

“…1 ). These phenotypic characteristics of EPV infections may be advantageous for EPV transmission in host insect populations via increased virus production or inhibition of pupation-associated behaviour 10 . Similar pathological phenotypes have been reported for insect viruses in the family Baculoviridae 12 , and for an insect-pathogenic fungus in the family Clavicipitaceae, Metarhizium rileyi 13 .…”

“…However, no genetic evidence for such enzymes targeting 20E has been found in EPV genomes. Previous studies indicated higher JH titres in the haemolymph of larvae infected with EPVs belonging to the genus Betaentomopoxvirus (EPVs infecting lepidopteran insects) than in non-infected larval haemolymph 10 , 11 . A survey using available genome sequence data of five Betaentomopoxvirus species revealed that they possessed a plausible candidate gene responsible for this JH titre elevation.…”