Nymph

Bourbon virus (BRBV) was first isolated in 2014 from a resident of Bourbon County, Kansas, USA, who died of the infection. In 2015, an ill Payne County, Oklahoma, resident tested positive for antibodies to BRBV, before fully recovering. We retrospectively tested for BRBV in 39,096 ticks from northwestern Missouri, located 240 km from Bourbon County, Kansas. We detected BRBV in 3 pools of Amblyomma americanum (L.) ticks: 1 pool of male adults and 2 pools of nymphs. Detection of BRBV in A. americanum, a species that is aggressive, feeds on humans, and is abundant in Kansas and Oklahoma, supports the premise that A. americanum is a vector of BRBV to humans. BRBV has not been detected in nonhuman vertebrates, and its natural history remains largely unknown.

Huanglongbing (HLB) causes considerable economic losses to citrus industries worldwide. Its management depends on controlling of the Asian citrus Psyllid (ACP), the vector of the bacterium, Candidatus Liberibacter asiaticus (CLas), the causal agent of HLB. Silencing genes by RNA interference (RNAi) is a promising tool to explore gene functions as well as control pests. In the current study, abnormal wing disc (awd) gene associated with wing development in insects is used to interfere with the flight of psyllids. Our study showed that transcription of awd is development-dependent and the highest level was found in the last instar (5(th)) of the nymphal stage. Micro-application (topical application) of dsRNA to 5(th) instar of nymphs caused significant nymphal mortality and adult wing-malformation. These adverse effects in ACP were positively correlated with the amounts of dsRNA used. A qRT-PCR analysis confirmed the dsRNA-mediated transcriptional down-regulation of the awd gene. Significant down-regulation was required to induce a wing-malformed phenotype. No effect was found when dsRNA-gfp was used, indicating the specific effect of dsRNA-awd. Our findings suggest a role for awd in ACP wing development and metamorphosis. awd could serve as a potential target for insect management either via direct application of dsRNA or by producing transgenic plants expressing dsRNA-awd. These strategies will help to mitigate HLB by controlling ACP.

The Asian citrus psyllid (ACP), Diaphorina citri Kuwayama, is one of the most important citrus pests. ACP is the vector of the phloem-limited bacteria Candidatus Liberibacter americanus and Candidatus Liberibacter asiaticus, the causal agents of the devastating citrus disease huanglongbing (HLB). The management of HLB is based on the use of healthy young plants, eradication of infected plants and chemical control of the vector. RNA interference (RNAi) has proven to be a promising tool to control pests and explore gene functions. Recently, studies have reported that target mRNA knockdown in many insects can be induced through feeding with double-stranded RNA (dsRNA). In the current study, we targeted the cathepsin D, chitin synthase and inhibitor of apoptosis genes of adult and nymph ACP by feeding artificial diets mixed with dsRNAs and Murraya paniculata leaves placed in dsRNAs solutions, respectively. Adult ACP mortality was positively correlated with the amount of dsRNA used. Both nymphs and adult ACP fed dsRNAs exhibited significantly increased mortality over time compared with that of the controls. Moreover, qRT-PCR analysis confirmed the dsRNA-mediated RNAi effects on target mRNAs. These results showed that RNAi can be a powerful tool for gene function studies in ACP and perhaps for HLB control.

IntroductionThe pear psylla Cacopsylla chinensis is a destructive pear pest whose nymphs exhibit seasonal polymorphism between summer-form and winter-form morphs. Whether the two morphs differ in susceptibility to Beauveria bassiana and the underlying mechanisms have not been characterized.MethodsBioassays at three spore concentrations (2.5×106, 2.5×107, and 2.5×108 spores/mL) were conducted to compare fungal virulence against fourth-instar nymphs of both morphs. Hemocyte counts and transcriptome sequencing were performed at 24 h and 36 h post-infection, respectively. qRT-PCR and enzymatic activity assays were used to further validate immune-related gene expression and enzyme activities.ResultsB. bassiana exhibited significantly higher virulence against summer-form nymphs than winter-form nymphs across all concentrations; at the highest concentration, only 8% of summer-form survived to day 5, compared with approximately 40% survival in winter-form, representing an approximately 79-fold difference in LC50 between the two morphs. Transcriptome analysis revealed that winter-form nymphs triggered a much broader transcriptional response with 1,673 differentially expressed genes (1,344 upregulated), accompanied by coordinated induction of immune genes including phenoloxidase, lysozyme and catalase, alongside full activation of energy metabolism pathways such as oxidative phosphorylation and the TCA cycle. In contrast, summer-form nymphs showed weak immune and metabolic activation. qRT-PCR and enzyme activity assays further verified that key immune-related transcripts and enzyme activities were markedly more induced in winter-form nymphs.DiscussionThese findings demonstrate that winter-form nymphs possess a more robust immune-metabolic defense network against B. bassiana, whereas summer-form nymphs are immune-suppressed and highly susceptible, revealing the molecular basis of morph-specific pathogen resistance shaped by seasonal polyphenism and providing a mechanistic basis for seasonally optimized biocontrol strategies.