Mortality of cutworm larvae is not enhanced by Agrotis segetum granulovirus and Agrotis segetum nucleopolyhedrovirus B coinfection relative to single infection by either virus

Abstract

Mixed infections of insect larvae with different baculoviruses are occasionally found. They are of interest from an evolutionary as well as from a practical point of view when baculoviruses are applied as biocontrol agents. Here, we report mixed-infection studies of neonate larvae of the common cutworm, Agrotis segetum, with two baculoviruses, Agrotis segetum nucleopolyhedrovirus B (AgseNPV-B) and Agrotis segetum granulovirus (AgseGV). By applying quantitative PCR (qPCR) analysis, coinfections of individual larvae were demonstrated, and occlusion body (OB) production within singly infected and coinfected larvae was determined in individual larvae. Mixtures of viruses did not lead to changes in mortality rates compared with rates of single-virus treatments, indicating an independent action within host larvae under our experimental conditions. AgseNPV-B-infected larvae showed an increase in OB production during 2 weeks of infection, whereas the number of AgseGV OBs did not change from the first week to the second week. Fewer OBs of both viruses were produced in coinfections than in singly infected larvae, suggesting a competition of the two viruses for larval resources. Hence, no functional or economic advantage could be inferred from larval mortality and OB production from mixed infections of A. segetum larvae with AgseNPV-B and AgseGV.

FIG 1

Mean mortality rates of the first week (days 2 to 7) (A) and the second week (days 8 to 14) (B) of single (GV, NPV_L, and NPV_H)- and mixed (GV-NPV_L and GV-NPV_H)-virus treatments. Different letters above the columns indicate significant differences between treatments (one-way analysis of variance [ANOVA] followed by Tukey's honest significant difference [HSD] test for pairwise comparisons between treatments, significance at P values of <0.05). The vertical lines represent standard deviations. Pie charts above each column represent the ratio of noninfected (white), coinfected (black), AgseGV only-infected (light gray), and AgseNPV-B only-infected (dark gray) larvae from each treatment (n = number of larvae measured in qPCR).

FIG 2

Comparison of the observed and expected mortality rates of GV-NPV_L and GV-NPV_H treatments observed in the first (A) and second (B) weeks postinfection. Expected mortality values were calculated from survival rates of replicates of GV, NPV_L, and NPV_H treatments, assuming independent action (for details, see Results). Statistical analyses were conducted separately for the observed and expected mortality of each treatment. Different letters indicate significant differences (Student's t test, significance at P values of <0.05). Vertical lines represent standard deviations.

FIG 3

Median OB production of AgseNPV-B (A) and AgseGV (B) within singly infected and coinfected larvae, which succumbed during the first (days 2 to 7) and second (days 8 to 14) weeks postinfection. The minimums of the y axes represent the lower limits of detection (LOD) for AgseGV (10⁵ OBs/larva) and AgseNPV-B (10³ OBs/larva). Different letters indicate significant differences (Wilcoxon rank sum test, significance at P values of <0.05) between treatments.

FIG 4

Correlation analysis (Spearman's rank correlation coefficient [r_s]) of larval AgseGV and AgseNPV-B OB production within coinfected A. segetum larvae. Only larvae from the GV-NPV_H treatment that died within the first (A) and second (B) weeks were considered. Vertical lines, median AgseGV OB production; horizontal lines, median AgseNPV-B OB production; solid lines, mixed GV-NPV_H treatment; dashed lines, single GV and NPV_H treatments are drawn as a reference.