Novel strains of a pandemic plant virus, tomato spotted wilt orthotospovirus, increase vector fitness and modulate virus transmission in a resistant host

Abstract

Tomato spotted wilt orthotospovirus (TSWV) is one of the most successful pandemic agricultural pathogens transmitted by several species of thrips in a persistent propagative manner. Current management strategies for TSWV heavily rely on growing single-gene resistant cultivars of tomato ("Sw-5b" gene) and pepper ("Tsw" gene) deployed worldwide. However, the emergence of resistance-breaking strains (RB) in recent years has compounded the threat of TSWV to agricultural production worldwide. Despite this, an extensive study on the thrips transmission biology of RB strains is currently lacking. It is also unclear whether mutualistic TSWV-thrips interactions vary across different novel strains with disparate geographical origins. To address both critical questions, we studied whether and how four novel RB strains of TSWV (two sympatric and two allopatric), along with a non-RB strain, impact western flower thrips (WFT) fitness and whether this leads to differences in TSWV incidence, symptom severity (virulence), and virus accumulation in two differentially resistant tomato cultivars. Our findings show that all RB strains increased WFT fitness by prolonging the adult period and increasing fecundity compared to non-RB and non-viruliferous controls, regardless of the geographical origin of strains or the TSWV titers in individual thrips, which were substantially low in allopatric strains. TSWV accumulation in thrips varied at different developmental stages and was unrelated to the infected tissues from which thrips acquired the virus. However, it was significantly positively correlated to that in WFT-inoculated susceptible plants, but not the resistant ones. The TSW incidences were high in tomato plants infected with all RB strains, ranging from 80% to 90% and 100% in resistant and susceptible plants, respectively. However, TSW incidence in the non-RB-infected susceptible tomato plants was 80%. Our findings provide new insights into how novel strains of TSWV, by selectively offering substantial fitness benefits to vectors, modulate transmission and gain a potential epidemiological advantage over non-RB strains. This study presents the first direct evidence of how vector-imposed selection pressure, besides the one imposed by resistant cultivars, may contribute to the worldwide emergence of RB strains.

Keywords: Orthotospovirus tomatomaculae; insect vector biology; resistance-breaking strains; vector-virus interactions; western flower thrips.

Figure 1

An overview of methods followed to study the transmission biology of resistance-breaking strains of TSWV. For each strain, ~800 neonate larvae were allowed a 72-h acquisition access period on non-infected tomato leaves or those infected with various TSWV strains to obtain non-viruliferous and viruliferous thrips, respectively. Different strains of TWSV used in the study were sympatric (from Bushland, TX): Tom-BL1, Tom-BL2; allopatric (from California) Tom-CA and (from Mexico) Tom-MX; and non-resistance breaking (non-RB). After virus acquisition, a cohort of 10 second instar thrips were transferred to a 9-cm sterile Petri dish (affixed with an insect mesh in the center) containing bean pods to record egg-to-adult developmental time every 12 h. Another large cohort of larvae was transferred to a different set of 14-cm Petri dishes through various developmental stages. At each stage, TSWV accumulation was quantified in 10 individual thrips per strain (and non-viruliferous control) using RT-qPCR. On the first day of adulthood, a subset of adult thrips was reared individually on 9-cm Petri dishes (one adult/Petri dish, eight Petri dishes/strain including control) to record the adult period and fecundity. To record fecundity, individual bean pods were replaced every 2 days with fresh ones. The collected old pods were then individually incubated in a new Petri dish for 3 days to record the number of emerging larvae. To study TSWV transmission, a large subset of larvae (30/plant) were allowed a 72-h inoculation access period on 3-week-old non-infect tomato plants from two different cultivars (one resistant and susceptible each, 10 plants/cultivar/strain) including non-viruliferous thrips as a control.

Figure 2

(A) Total and (B) stage-wise egg-to-adult developmental time of thrips individually infected with different strains of TWSV: Tom-BL1, Tom-BL2, Tom-CA, Tom-MX, and non-RB, with non-viruliferous control. Different letters indicate significant differences at P < 0.05.

Figure 3

(A) Adult period and (B) fecundity of thrips individually infected with different strains of TWSV: Tom-BL1, Tom-BL2, Tom-CA, Tom-MX, and non-RB, with non-viruliferous control. Different letters indicate significant differences at P < 0.05.

Figure 4

TWSV accumulation across different developmental stages of individual thrips infected with different strains of TWSV: Tom-BL1, Tom-BL2, Tom-CA, Tom-MX, and Non-RB, quantified using RT-qPCR. Different letters indicate significant differences at P < 0.05.

Figure 5

TSWV accumulation in thrips-inoculated tomato plants infected with different strains of TWSV: Tom-BL1, Tom-BL2, Tom-CA, Tom-MX, and Non-RB. TSWV was quantified 4 weeks post-inoculation from the topmost leaf using RT-qPCR. Different letters indicate significant differences at P < 0.05.

Figure 6

Regression analysis to assess functional relationships between TSWV copy numbers in thrips with those in (A) infected tissues from which they acquired the virus and thrips-inoculated, (B) susceptible, and (C) resistant plants used in transmission studies.