The greening-causing agent alters the behavioral and electrophysiological responses of the Asian citrus psyllid to a putative sex pheromone.

The Asian Citrus Psyllid (ACP), Diaphorina citri, is a vector of the pathological bacterium Candidatus Liberibacter asiaticus (CLas), which causes the most devastating disease to the citrus industry worldwide, known as greening or huanglongbing (HLB). Earlier field tests with an acetic acid-based lure in greening-free, 'Valencia' citrus orange groves in California showed promising results. The same type of lures tested in São Paulo, Brazil, showed unsettling results. During the unsuccessful trials, we noticed a relatively large proportion of females in the field, ultimately leading us to test field-collected males and females for Wolbachia and CLas. The results showed high rates of Wolbachia and CLas infection in field populations. We then compared the olfactory responses of laboratory-raised, CLas-free, and CLas-infected males to acetic acid. As previously reported, CLas-uninfected males responded to acetic acid at 1 µg. Surprisingly, CLas-infected males required 50 × higher doses of the putative sex pheromone, thus explaining the failure to capture CLas-infected males in the field. CLas infection was also manifested in electrophysiological responses. Electroantennogram responses from CLas-infected ACP males were significantly higher than those obtained with uninfected males. To the best of our knowledge, this is the first report of a pathogen infection affecting a vector's response to a sex attractant.

Yellow Leaf Disease Resistance and Melanaphis sacchari Preference in Commercial Sugarcane Cultivars.

Sugarcane yellow leaf disease (YLD) caused by sugarcane yellow leaf virus (ScYLV) is a major threat for the sugarcane industry worldwide, and the aphid Melanaphis sacchari is its main vector. Breeding programs in Brazil have provided cultivars with intermediate resistance to ScYLV, whereas the incidence of ScYLV has been underestimated partly due to the complexity of YLD symptom expression and identification. Here, we evaluated YLD symptoms in a field assay using eight sugarcane genotypes comprising six well-established commercial high-sucrose cultivars, one biomass yield cultivar, and a susceptible reference under greenhouse conditions, along with estimation of virus titer through RT-qPCR from leaf samples. Additionally, a free-choice bioassay was used to determine the number of aphids feeding on the SCYLV-infected cultivars. Most of the cultivars showed some degree of resistance to YLD, while also revealing positive RT-qPCR results for ScYLV and virus titers with non-significant correlation with YLD severity. The cultivars IACSP01-5503 and IACBIO-266 were similar in terms of aphid preference and ScYLV resistance traits, whereas the least preferred cultivar by M. sacchari, IACSP96-7569, showed intermediate symptoms but similar virus titer to the susceptible reference, SP71-6163. We conclude that current genetic resistance incorporated into sugarcane commercial cultivars does not effectively prevent the spread of ScYLV by its main aphid vector.

Effect of Sugarcane Cultivars Infected with Sugarcane Yellow Leaf Virus (ScYLV) on Feeding Behavior and Biological Performance of Melanaphis sacchari (Hemiptera: Aphididae).

Sugarcane yellow leaf virus (ScYLV), Polerovirus, Luteoviridae, is one of the main viruses that infect sugarcane worldwide. The virus is transmitted by the aphid Melanaphis sacchari in a persistent, circulative manner. To better understand the interactions between ScYLV, sugarcane genotypes and M. sacchari, we explored the effect of sugarcane cultivars on the feeding behavior and biological performance of the vector. The number of nymphs, adults, winged, total number of aphids and dead aphids was assayed, and an electrical penetration graph (EPG) was used to monitor the stylet activities. Multivariate analysis showed changes in the vector's behavior and biology on cultivars, identifying specific groups of resistance. In the cultivar 7569, only 5.5% of the insects were able to stay longer on sustained phloem ingestion, while in the other seven cultivars these values varied from 20% to 60%. M. sacchari showed low phloem activities in cultivars 7569 and Bio266. Overall, cultivar 7569 showed the worst biological performance of aphids, with the insects presenting mechanical difficulties for feeding and a shorter duration of the phloem period, and thus being considered the most resistant. We conclude that ScYLV virus infection in different sugarcane cultivars induced specific changes in the host plant, modifying the behavior of its main vector, which may favor or impair virus transmission.

Aphids Are Unable to Ingest Phloem Sap from the Peduncles of Lime Fruits.

Citrus exports to Europe are regulated enforcing that fruits shall be free from peduncles and leaves, as they represent an important pathway for the entrance of non-European (non-EU) Citrus tristeza virus (CTV) isolates into the European Community. Aphids, are the vectors of CTV and could potentially feed on peduncles of imported fruits and thus spread non-EU isolates of CTV across Europe. We studied the probing behaviour of the main vectors of CTV (Aphis (Toxoptera) citricidus and Aphis gossypii) on lime leaves and peduncles to assess whether they could potentially transmit the virus. Aphids placed on peduncles rejected probing and feeding, tried to escape and spent most of their time on non-probing activities. Our work demonstrated that both A. citricidus and A. gossypii could not ingest sap from the phloem of lime peduncles, as phloem ingestion was never observed. This implies that aphids would not be able to acquire CTV from an infected fruit peduncle and transmit it to a susceptible plant. Our study supports that citrus exports with fruit peduncles to Europe may not be a real risk for the introduction of non-EU isolates of CTV to the European Community.

Drench Application of Systemic Insecticides Disrupts Probing Behavior of Diaphorina citri (Hemiptera: Liviidae) and Inoculation of Candidatus Liberibacter asiaticus.

Candidatus Liberibacter asiaticus (CLas) is a phloem-limited bacterium that is associated with the Huanglongbing (HLB) disease of citrus and transmitted by the psyllid, Diaphorina citri. There are no curative methods to control HLB and the prevention of new infections is essential for HLB management. Therefore, the objective of our study was to determine the effects of systemic insecticides, such as the neonicotinoids imidacloprid, thiamethoxam, and a mixture of thiamethoxam and chlorantraniliprole (diamide) on the probing behavior of CLas-infected D. citri and their effect on CLas transmission. The electrical penetration graph (EPG-DC) technique was used to monitor the stylet penetration activities of CLas-infected D. citri on sweet orange [Citrus sinensis (L.) Osbeck] 'Valencia' treated with systemic insecticides. Systemic insecticides disrupted the probing behavior of CLas-infected D. citri, in a way that affected CLas transmission efficiency, particularly by negatively affecting the stylet activities related to the phloem phase. All insecticides reduced (by 57-73%) the proportion of psyllids that exhibited sustainable phloem ingestion (waveform E2 > 10 min), with significant differences observed on plants treated with thiamethoxam and thiamethoxam + chlorantraniliprole. The transmission rate of CLas with high inoculum pressure (five CLas-infected D. citri per plant and a seven-day inoculation access period) to untreated control plants was 93%. In contrast, CLas transmission was reduced to 38.8% when test plants were protected by systemic insecticides. Our results indicated that all insecticides tested presented a potential to reduce CLas inoculation by an average of 59%; therefore, these insecticides can be used to reduce the spread of HLB.

Control of insect vectors and plant viruses in protected crops by novel pyrethroid-treated nets.

BACKGROUND: Long-lasting insecticide-treated nets (LLITNs) constitute a novel alternative that combines physical and chemical tactics to prevent insect access and the spread of insect-transmitted plant viruses in protected enclosures. This approach is based on a slow-release insecticide-treated net with large hole sizes that allow improved ventilation of greenhouses. The efficacy of a wide range of LLITNs was tested under laboratory conditions against Myzus persicae, Aphis gossypii and Bemisia tabaci. Two nets were selected for field tests under a high insect infestation pressure in the presence of plants infected with Cucumber mosaic virus and Cucurbit aphid-borne yellows virus. The efficacy of Aphidius colemani, a parasitoid commonly used for biological control of aphids, was studied in parallel field experiments. RESULTS: LLITNs produced high mortality of aphids, although their efficacy decreased over time with sun exposure. Certain nets excluded whiteflies under laboratory conditions; however, they failed in the field. Nets effectively blocked the invasion of aphids and reduced the incidence of viruses in the field. The parasitoid A. colemani was compatible with LLITNs. CONCLUSION: LLITNs of appropriate mesh size can become a very valuable tool in combination with biocontrol agents for additional protection against insect vectors of plant viruses under IPM programmes.

A non-persistently transmitted-virus induces a pull-push strategy in its aphid vector to optimize transmission and spread.

Plant viruses are known to modify the behaviour of their insect vectors, both directly and indirectly, generally adapting to each type of virus-vector relationship in a way that enhances transmission efficiency. Here, we report results of three different studies showing how a virus transmitted in a non-persistent (NP) manner (Cucumber mosaic virus; CMV, Cucumovirus) can induce changes in its host plant, cucumber (Cucumis sativus cv. Marumba) that modifies the behaviour of its aphid vector (Aphis gossypii Glover; Hemiptera: Aphididae) in a way that enhances virus transmission and spread non-viruliferous aphids changed their alighting, settling and probing behaviour activities over time when exposed to CMV-infected and mock-inoculated cucumber plants. Aphids exhibited no preference to migrate from CMV-infected to mock-inoculated plants at short time intervals (1, 10 and 30 min after release), but showed a clear shift in preference to migrate from CMV-infected to mock-inoculated plants 60 min after release. Our free-choice preference assays showed that A. gossypii alates preferred CMV-infected over mock-inoculated plants at an early stage (30 min), but this behaviour was reverted at a later stage and aphids preferred to settle and reproduce on mock-inoculated plants. The electrical penetration graph (EPG) technique revealed a sharp change in aphid probing behaviour over time when exposed to CMV-infected plants. At the beginning (first 15 min) aphid vectors dramatically increased the number of short superficial probes and intracellular punctures when exposed to CMV-infected plants. At a later stage (second hour of recording) aphids diminished their feeding on CMV-infected plants as indicated by much less time spent in phloem salivation and ingestion (E1 and E2). This particular probing behaviour including an early increase in the number of short superficial probes and intracellular punctures followed by a phloem feeding deterrence is known to enhance the transmission efficiency of viruses transmitted in a NP manner. We conclude that CMV induces specific changes in a plant host that modify the alighting, settling and probing behaviour of its main vector A. gossypii, leading to optimum transmission and spread of the virus. Our findings should be considered when modelling the spread of viruses transmitted in a NP manner.