The plant virus Tomato Spotted Wilt Tospovirus activates the immune system of its main insect vector, Frankliniella occidentalis

Abstract

Tospoviruses have the ability to infect plants and their insect vectors. Tomato spotted wilt virus (TSWV), the type species in the Tospovirus genus, infects its most important insect vector, Frankliniella occidentalis, the western flower thrips (WFT). However, no detrimental effects on the life cycle or cytopathological changes have been reported in the WFT after TSWV infection, and relatively few viral particles can be observed even several days after infection. We hypothesized that TSWV infection triggers an immune response in the WFT. Using subtractive cDNA libraries to probe WFT DNA macroarrays, we found that the WFT's immune system is activated by TSWV infection. The activated genes included (i) those encoding antimicrobial peptides, such as defensin and cecropin; (ii) genes involved in pathogen recognition, such as those encoding lectins; (iii) those encoding receptors that activate the innate immune response, such as Toll-3; and (iv) those encoding members of signal transduction pathways activated by Toll-like receptors, such as JNK kinase. Transcriptional upregulation of these genes after TSWV infection was confirmed by Northern analysis, and the kinetics of the immune response was measured over time. Several of the detected genes were activated at the same time that viral replication was first detected by reverse transcription-PCR. To our knowledge, this is the first report of the activation of an insect vector immune response by a plant virus. The results may lead to a better understanding of insects' immune responses against viruses and may help in the future development of novel control strategies against plant viruses, as well as human and animal viruses transmitted by insect vectors.

FIG. 1.

Selected areas of WFT DNA macroarrays after hybridization with riboprobes derived from noninfected versus TSWV-infected subtracted cDNA libraries. Selected regions of nylon membranes show the detection of defensin D (A), hemolin (B), Ikk kinase (C), and complement-like TEP-1 (D) (arrows).

FIG. 2.

(A) Northern analysis of activated immune-related WFT genes after TSWV infection. Riboprobes are indicated at the left. Samples were collected from 0 to 240 hpe, meaning time feeding on TSWV-infected plants. Samples from 0 to 24 hpe represent insects at the first-instar larval stage, those from 96 hpe represent the second-instar larval stage, and those from 240 hpe represent the adult stage. Actin was used as a negative control for activation and as a loading control. (B) Graphic representation of the data showed in panel A. Relative signal intensity is the signal intensity of each gene in the Northern blot normalized to that of the corresponding actin signal, measured with a Storm 860 PhosphoImager.

FIG. 3.

RT-PCR of dissected WFT guts with primers specific to the TSWV complementary M RNA. Samples were collected from 0 to 240 hpe, meaning time feeding on TSWV-infected plants.