General Insect Management 757
via suppression of resistance genes as has been reported in Sitobion avenae (Xu et al. 2014) and Aphis
gossypii (Gong et al. 2014).
RNAi studies with stink bug species have not been reported in the literature, but some insights can be
gained from published work on the plant bug Lygus lineolaris. Suppression of the Inhibitor of Apoptosis
(IAP) gene via dsRNA injection resulted in increased Lygus mortality whereas feeding of the IAP dsRNA
at a dietary concentration of 1,000 ppm had no phenotypic effect (Walker and Allen 2011, Allen and Walker
2012). The digestive physiology of hemipteran species accounts for one apparent barrier to oral delivery of
dsRNAs. As with stink bugs, Lygus bugs engage in extra-oral digestion, a process that includes the injec-
tion of salivary secretions comprising lytic enzymes into plant feeding sites, creating a slurry of plant mate-
rial that then is ingested. Salivary secretions from hemipteran species are known to contain digestive
enzymes such as lipases, phosphatases, pectinases, and proteases (Kaloshian and Walling 2005). The
report that Lygus salivary extracts also exhibit potent dsRNAse activity suggests that dsRNAs are likely
to suffer degradation prior to ingestion (Allen and Walker 2012). Similarly, salivary- and hemolymph-
nucleases have been identified as likely barriers to RNAi in the pea aphid (Christiaens et al. 2014).
An alternative strategy for management of hemipteran species relies on topical delivery of dsRNAs
through the insect cuticle, a route that bypasses the harsh environment of the insect gut. This approach
has been used with some success to deliver dsRNAs and achieve target gene silencing in lepidopteran
(Wang et al. 2011), dipteran (Pridgeon et al. 2008), and hemipteran species (El-Shesheny et al. 2013,
Killiny et al. 2014). In the case of the Asian citrus psyllid, Diaphorina citri (Kuwayama), deposition of
dsRNA on the ventral side of the thorax resulted in suppression of five CYP4 (cytochrome P450 mono-
oxygenase) genes and an increase in sensitivity to the neonicotinoid insecticide imidacloprid (Killiny et al.
2014). In the aforementioned studies, the concentrations of dsRNA required for a topical effect ranged from
50–200 ppm, a prohibitively high rate for actual field application on row crops impacted by stink bugs.
The identification of formulations that can package dsRNA, confer environmental stability, and improve
delivery to insect cells may result in the lower use rates required for topical applications. Delivery agents
including cationic lipids and dendrimers, cyclodextrin polymers, mesoporous silica, and various forms
of polethyleneimine are being evaluated for RNAi human therapeutics (for review, see Zhou et al. 2013).
Preliminary studies with dipteran and lepidopteran species suggest that some of these delivery agents
can enable dsRNA delivery to otherwise recalcitrant insect species (Whyard et al. 2009, Zhang et al.
2010b, He et al. 2013). The requirements for effective delivery presumably would differ depending on
whether the route of delivery is via topical exposure or ingestion. In the latter case, formulations would
need to protect dsRNAs from stink bug salivary- and gut- nucleases and enable uptake into gut cells. In
addition, because of the nature of stink bug feeding, these formulations must presumably penetrate plant
tissue in order to be accessible for ingestion. More research is needed to evaluate the feasibility of this
approach and the impact of dsRNA stabilization on both bioavailability and efficacy.
16.5 Acknowledgments
We thank Ted R. Schultz (National Museum of Natural History, Washington, DC) for his help concern-
ing the identification of the ant species used by the Chinese in biological control during the Ancient
History Period. We also thank Marlin E. Rice (Ames, IA) for his input on this chapter.
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