Dhana Raj Boina

Quantifying Dispersal of Diaphorina citri (Hemiptera: Psyllidae) by Immunomarking and Potential Impact of Unmanaged Groves on Commercial Citrus Management

ABSTRACT Diaphorina citri Kuwayama (Hemiptera: Psyllidae) is an important pest of citrus. It is an efficient vector of three bacterial pathogens that are the presumptive causal agents of huanglongbing (HLB) or citrus greening disease. The movement patterns and dispersal capabilities of D. citri require study to better understand the spread of HLB and to improve management strategies for D. citri. A recently developed immunomarking technique that uses crude food proteins (chicken egg albumin, bovine casein, and soy protein) was evaluated for marking and tracking movement of D. citri in Florida citrus groves. In general, both egg and milk protein markers exhibited longer residual activity (35 d) than the soy protein marker (20 d) when applied to citrus leaves with a residual activity order of egg > milk > soy protein. However, residues of all three protein markers decreased with a simulated rain; this was more pronounced for soy protein than for egg and milk proteins. Temperature did not significantly affect acquisition of markers by adult D. citri. Egg, milk, and soy protein markers were detected on >90% of adult D. citri for up to 10, 10, and 5 d, respectively, after field application. Addition of tetrasodium ethylenediamine tetraacetic acid (water softener) and/or Silwet L-77 (wetting agent) to marker solutions did not affect longevity of detection. Each of the protein markers was detected on ≥80% of exposed D. citri for up to 30 d after direct application to adults. A field study was conducted to measure movement of D. citri between replicated pairs of 0.4 ha managed and unmanaged citrus plots separated by 60–100 m. Approximately 70% of captured D. citri were found marked 3 d after application of proteins in the field. Using two marker proteins, it was determined that D. citri moved bi-directionally between managed and unmanaged (abandoned) groves within 3 d with a greater number of D. citri adults moving from unmanaged into managed plots than from managed into unmanaged plots (net movement). These data indicate frequent movement by adult D. citri between groves and suggest that unmanaged groves may act as refuge sites for D citri, leading to reinfestation of nearby managed groves.

Antifeedant and sublethal effects of imidacloprid on Asian citrus psyllid, Diaphorina citri

BACKGROUND Asian citrus psyllid (ACP), Diaphorina citri Kuwayama, transmits the causal bacteria of the devastating citrus disease huanglongbing (HLB). Because of the variation in spatial and temporal uptake and systemic distribution of imidacloprid applied to citrus trees and its degradation over time in citrus trees, ACP adults and nymphs are exposed to concentrations that may not cause immediate mortality but rather sublethal effects. The objective of this laboratory study was to determine the effects of sublethal concentrations of imidacloprid on ACP life stages. RESULTS Feeding by ACP adults and nymphs on plants treated daily with a sublethal concentration (0.1 microg mL(-1)) of imidacloprid significantly decreased adult longevity (8 days), fecundity (33%) and fertility (6%), as well as nymph survival (12%) and developmental rate compared with untreated controls. The magnitude of these negative effects was directly related to exposure duration and concentration. Furthermore, ACP adults that fed on citrus leaves treated systemically with lethal and sublethal concentrations of imidacloprid excreted significantly less honeydew (7-94%) compared with controls in a concentration-dependent manner suggesting antifeedant activity of imidacloprid. CONCLUSIONS Sublethal concentrations of imidacloprid negatively affect development, reproduction, survival and longevity of ACP, which likely contributes to population reductions over time. Also, reduced feeding by ACP adults on plants treated with sublethal concentrations of imidacloprid may potentially decrease the capacity of ACP to successfully acquire and transmit the HLB causal pathogen.

Morphological characterization of the antennal sensilla of the Asian citrus psyllid, Diaphorina citri Kuwayama (Hemiptera: Psyllidae), with reference to their probable functions

The Asian citrus psyllid, Diaphorina citri Kuwayama (Hemiptera: Psyllidae), is one of the most significant economic pests of citrus worldwide. This insect vectors three phloem-restricted bacteria in the genus Candidatus Liberibacter that cause huanglongbing (citrus greening), the most severe disease limiting citrus production worldwide. We examined the external morphology of the antennal sensilla of male and female D. citri using scanning electron microscopy (SEM) and determined the putative functions of the identified sensilla using transmission electron microscopic (TEM) techniques. The filiform antennae of D. citri were of the conventional type comprised of a basal scape, pedicel and a long, thread-like flagellum, which is composed of eight flagellomeres. Eleven morphologically unique sensillar types were found and described on the antennae of male and female D. citri. Of those identified, the two apical setae, multiporous types I and II sensilla trichoidea, and the antennal rhinaria were porous and may be involved in perception of host- and mate-related volatile chemicals. However, the aporous types I, II and III sensilla trichoidea may have mechanosensory functions and the chaetica sensilla, cavity sensilla and unidentified uniporous sensilla may be involved in proprioception, thermo-hygroreception and cold detection, respectively. The shape, external morphology and array of sensilla on the antennae of male and female D. citri were similar. The only major difference observed was in the morphology of the short apical setae, whose tips were recessed inward in females but not so in males. The results are discussed in relation to plausible roles of the identified sensilla in mate and host location by this species.

Influence of Posttreatment Temperature on the Toxicity of Insecticides Against Diaphorina citri (Hemiptera: Psyllidae)

ABSTRACT The psyllid Diaphorina citri Kuwayama (Hemiptera: Psyllidae) is one of the most important pests of citrus worldwide because it efficiently vectors three bacteria in the genus Candidatus Liberibacter that cause the devastating citrus greening disease (huanglongbing). Current management practices for this insect pest rely on multiple sprays of foliar insecticides and one or two applications of soil systemic insecticides per season. Effective psyllid and disease management in Florida requires insecticide applications throughout the entire season over wide ranging temperature and environmental conditions. Using a petri dish bioassay technique, the effect of posttreatment temperature (range, 17–37°C) on the toxicity of selected organophosphate (chlorpyrifos and dimethoate), carbamate (carbaryl), avermectin (abamectin), pyrethroid (bifenthrin, zeta-cypermethrin, fenpropathrin, and lambda-cyhalothrin), and neonicotinoid (acetamiprid, imidacloprid, and thiamethoxam) insecticides was evaluated against adult D. citri. The toxicity of both organophosphates showed a positive temperature correlation within the 17–37°C range. Similarly, carbaryl (carbamate) and abamectin (avermectin) exhibited increased toxicity with increasing temperature from 17 to 37°C, with abamectin showing higher overall temperature-dependent toxicity against D. citri adults than carbaryl. With the exception of bifenthrin, which showed a positive temperature-dependent toxicity correlation between 27 and 37°C, all other pyrethroids tested exhibited a negative correlation over the temperature range examined. The toxicity of fenpropathrin and lambda-cyhalothrin dramatically decreased with increasing temperature from 17 to 37°C. The neonicotinoids imidacloprid and thiamethoxam exhibited a mixed response to increasing temperature, whereas acetamiprid showed a positive temperature correlation. However, all three neonicotinoids showed positive temperature-dependent toxicity correlations against D. citri adults over the temperature range tested. These data will enable citrus growers to choose the most effective insecticides for D. citri control from the various classes currently available depending on the prevailing temperature conditions.

Chemical control of the Asian citrus psyllid and of huanglongbing disease in citrus

By 2014, huanglongbing (HLB), the most destructive disease of citrus, and its insect vector, the Asian citrus psyllid (ACP), Diaphorina citri (Kuwayama), became established in all major citrus-growing regions of the world, including the United States, with the exception of California. At present, application of insecticides is the most widely followed option for reducing ACP populations, while application of antibiotics for suppressing HLB disease/symptoms is being practiced in some citrus-growing regions. Application of insecticides during the dormant winter season, along with cultivation of HLB-free seedlings and early detection and removal of symptomatic and asymptomatic trees, has been very effective in managing ACP. Area-wide management of ACP by application of insecticides at low volume in large areas of citrus cultivation has been shown to be effective in managing HLB and reducing management costs. As insecticide resistance is a major problem in sustainable management of ACP, rotation/alternation of insecticides with different chemistries and modes of action needs to be followed. Besides control of the insect vector, use of antibiotics has temporarily suppressed the symptoms of HLB in diseased trees. Recent efforts to discover and screen existing as well as new compounds for their antibiotic and antimicrobial activities have identified some promising molecules for HLB control. There is an urgent need to find a sustainable solution to the HLB menace through chemical control of ACP populations and within HLB-infected trees through the judicious use of labeled insecticides (existing and novel chemistries) and antibiotics in area-wide management programs with due consideration to the insecticide resistance problem.

Effect of pyriproxyfen, a juvenile hormone mimic, on egg hatch, nymph development, adult emergence and reproduction of the Asian citrus psyllid, Diaphorina citri Kuwayama

BACKGROUND The Asian citrus psyllid (ACP), Diaphorina citri Kuwayama, is a vector of bacteria presumably responsible for huanglongbing (HLB) disease in citrus. In this laboratory study, an investigation was made of the activity of pyriproxyfen, a juvenile hormone mimic, on ACP eggs, nymphs and adults to evaluate its potential as a biorational insecticide for inclusion in an integrated pest management (IPM) program for ACP. RESULTS Irrespective of egg age, timing or method of treatment, a significantly lower percentage of eggs (5-29%) hatched after exposure to 64 and 128 microg mL(-1) of pyriproxyfen. Only 0-36% of early instars (first, second and third) and 25-74% of late instars (fourth and fifth) survived to adults following exposure to 16, 32 and 64 microg mL(-1) of pyriproxyfen. However, 15-20% of adults that emerged following treatment as late instars exhibited morphological abnormalities. Furthermore, pyriproxyfen adversely affected reproduction (fecundity and fertility) of adults that emerged from treated fifth instars or that were treated topically with 0.04 microg as adults. CONCLUSIONS Application of pyriproxyfen at 64 microg mL(-1) resulted in greater inhibition of egg hatch and suppression of adult emergence compared with lower rates. Pyriproxyfen also markedly reduced female fecundity and egg viability for adults that were exposed either as fifth instars or as newly emerged adults. The ovicidal, larvicidal and reproductive effects against ACP suggest that pyriproxyfen is suitable for integration into an IPM program for ACP.

Antennal Sensilla of Tamarixia radiata (Hymenoptera: Eulophidae), a Parasitoid of Diaphorina citri (Hemiptera: Psyllidae)

ABSTRACT Tamarixia radiata (Waterston) (Hymenoptera: Eulophidae) is an effective idiobiont ectoparasitoid of the psyllid Diaphorina citri Kuwayama (Hemiptera: Psyllidae), vector of the huanglongbing (citrus greening disease) pathogen. We examined the external and functional morphology of the antennal sensilla of adult male and female T. radiata by using scanning and transmission electron microscopy, respectively, to gain insights into the behavioral ecology of this parasitoid. The geniculate antennae of male and female T. radiata were composed of a long scapula-shaped scape with a basal radicula, a barrel-shaped pedicel, and a long flagellum with a basal ring-like annulus. Five morphologically distinct sensilla, including two types of aporous trichoid sensilla (AST-1 and AST-2), one multiporous trichoid sensillum (MST), one multiporous placoid sensillum (MPS), and one aporous basiconic capitate peg sensillum were identified on the antennae of both sexes. The antennal structures of T. radiata were sexually dimorphic. Male antennae consisted of four funicular flagellomeres and possessed a greater number of olfactory MST than female antennae, suggesting their possible function in perception of mate-related volatile cues. Female antennae were characterized by three funicular flagellomeres and a greater number of MPS than male antennae, suggesting their possible function in the perception of host-related volatile cues. The results are discussed in relation to plausible roles of the identified sensilla in mate and host location by this important parasitoid species.

Blockage of chloride channels and anion transporters with pesticidal natural products and their synthetic analogs

Ligand-gated chloride channels mediate a variety of functions in excitable membranes of nerve and muscle in insects, and have a long history as targets for neurotoxic insecticides. Recent findings from our laboratory confirm that the natural product silphinenes and their semi-synthetic analogs share a mode of action with the established ligand-gated chloride channel antagonist, picrotoxinin. The silphinenes are non-selective, being roughly equipotent on insect and mammalian receptors, but also possess lethal and neurotoxic effects on a dieldrin-resistant strain of Drosophila melanogaster. These findings suggest that silphinenes act on insect GABA receptors in a way that is different from picrotoxinin, and it is possible that resistant insect populations in the field could be controlled with insecticidal compounds derived from the silphinenes. Voltage-gated chloride channels and anion transporters provide additional classes of validated targets for insecticidal/nematicidal action. Anion transporter blockers are toxic to insects via an action on the gut, and RNAi studies implicate voltage-gated chloride channels in nematode muscle as another possible target. There was no cross resistance to DIDS in a dieldrin-resistant strain of Drosophilamelanogaster, and no evidence for neurotoxicity. The potent paralytic actions of anion transporter blockers against nematodes, and stomach poisoning activity against lepidopteran larvae suggests they are worthy of further investigation as commercial insecticidal/nematicidal agents.

NEUROTOXICOLOGY OF bis(n)-TACRINES ON Blattella germanica AND Drosophila melanogaster ACETYLCHOLINESTERASE

A series of bis(n)-tacrines were used as pharmacological probes of the acetylcholinesterase (AChE) catalytic and peripheral sites of Blattella germanica and Drosophila melanogaster, which express AChE-1 and AChE-2 isoforms, respectively. In general, the potency of bis(n)-tacrines was greater in D. melanogaster AChE (DmAChE) than in B. germanica AChE (BgAChE). The change in potency with tether length was high in DmAChE and low in BgAChE, associated with 90-fold and 5.2-fold maximal potency gain, respectively, compared to the tacrine monomer. The optimal tether length for Blattella was 8 carbons and for Drosophila was 10 carbons. The two species differed by only about twofold in their sensitivity to tacrine monomer, indicating that differential potency occurred among dimeric bis(n)-tacrines due to structural differences in the peripheral site. Multiple sequence alignment and in silico homology modeling suggest that aromatic residues of DmAChE confer higher affinity binding, and the lack of same at the BgAChE peripheral site may account, at least in part, to the greater overall sensitivity of DmAChE to bis(n)-tacrines, as reflected by in vitro assay data. Topical and injection assays in cockroaches found minimal toxicity of bis(n)-tacrines. Electrophysiological studies on D. melanogaster central nervous system showed that dimeric tacrines do not readily cross the blood brain barrier, explaining the observed nonlethality to insects. Although the bis(n)-tacrines were not good insecticide candidates, the information obtained in this study should aid in the design of selective bivalent ligands targeting insect, pests, and disease vectors.