A better understanding of the transmission of HLB between the psyllids and citrus hosts in natural landscape is crucial to formulating effective control strategies. A spatially-explicit agent-based model, which simulates the actions and interactions of autonomous agents within the epidemiological system, has been developed to investigate how ACP and HLB spread in the Central Valley of California, an intermixed landscape of residential and commercial citrus. This study is a practical extension of the mathematical model with purposes to quantify the influence of input epidemiologic parameters on disease progress under sensitivity analysis, and investigate the efficiency of ACP/HLB management strategies by running scenario-based simulations. Although there are numerous hypothetical management strategies for testing, we focus more on survey design , and biological and chemical control strategies . In addition, we incorporated these management strategies into disease modelling with consideration of the social-economic perspective of citrus growers. Growers’ awareness of ACP/HLB and their attitude toward control strategies will change along with disease development, and we evaluate the set-points that yield optimal operation cost and sustainable control using a cost-benefit analysis. Our results indicate that climate change can have a large effect on the performance and spatiotemporal distribution of ACP and biological control agent populations in central California. ACP spread occurs more frequently and faster within commercial citrus clusters, but comparatively slower for low density or well separated residential areas. For different spray strategy scenarios,vertical rack system a comparison between simulation outputs confirms that the synchronize rate for coordinated spray plays an important role in slowing ACP epidemic development.
The Asian citrus psyllid vectors causal pathogens of huanglongbing or citrus greening disease. Its control is critical in all habitats including organic citrus grown in the United States and other regions of the world. Three separate organic programs, organic insecticides applied alone or with horticultural mineral oil and insecticidal soap were compared with one conventional program for impact on ACP and beneficial insects including release of nymphal parasitoid Tamarixia radiata in bearing citrus in southwest Florida. During the dormant winter season, Pyganic applied alone or with 435 oil or M-pede applied in November, December and January and Danitol applied in November and January all provided significant reduction in ACP with residual lasting longer for Danitol.Organic programs 2 and 3 rotated organic insecticides with 435 oil or M-pede resulting in 50% reduction in use of insecticides while providing better ACP control than program 1 with organic insecticides only. However, ACP was reduced more in the conventional program. Tamarixia radiata, ectoparasitoid of ACP nymphs was released in all programs but recovered more from ACP nymphs in the organic program compared to the conventional program. Lacewings, spiders, ants and ladybeetles were observed in all programs that also may have contributed to ACP reduction. Best yields were obtained in programs using organic insecticides with 435 oil or conventional insecticides. Monthly applications of Pyganic with 435 oil during dormant winter period and rotation of organic insecticides with oil during growing season appear to be reasonable options for ACP management in organic citrus. Organic insecticides will also be suitable for conventional citrus growers as selective options to avoid excessive use of non-selective insecticides, to limit pesticide resistance and harm to beneficial insects.
Candidatus Liberibacter asiaticus colonizes the citrus phloem and causes the most destructive citrus disease. Understanding how the bacterium interacts with citrus is important to improve or devise new methods of disease control. The objective of this work was to assess Las movement in citrus plants, from the initial infection site to the root, which is the part of the tree that apparently first suffers after pathogen infection. Experiments were carried out using 208 three-year-old, 60-cm high, potted Valencia/Volkamer lemon. The plants were grafted at the stem 60 cm above the substrate surface with 3- to 4-cm-long buds from Las-positive branches, and assessed over time at different distances from the inoculation site to determine the presence of the bacterium. Two evaluation procedures were used in three experiments carried out in two time periods. In the first, the stem of the inoculated plants were pruned sequentially at 10, 20, 30, and 40 cm below the inoculation site at 14, 21, 28, 35, and 42 DPI. The new leaves that developed at the top of the remaining stems were assessed regularly for HLB symptoms and through qPCR at six months post inoculation. This procedure was used in two experiments carried out from August 2015 to January 2016 and from March to August 2016 . In the second procedure, fibrous roots and one-centimeter long bark rings were removed just below the inoculation site and, at 10-cm intervals from it, up to near the soil level. The evaluations were made at 7, 14, 21, 28, 35, 42, and 49 days post inoculation , with the samples processed and analyzed through qPCR. This procedure was used in just one experiment carried out from March to April 2016. The fastest Las speed were 1.14, 1.43, and 2.02 cm per day for E1, E2, and E3, respectively. The frequencies of the evaluated sites positive for Las, at all evaluation dates, were then subjected to regression analysis. The logistic model better described Las distribution in the plants over time.
The probability of detecting Las at any portion of the plant was a function of time and distance from the inoculum. The longer the time between the inoculation and sampling dates and the shorter the distance from the inoculum, at any given time, the higher the probability of testing positive. Extrapolations made using the regression model allowed estimations of minimum 51 and 131 days for Las to reach the root of 2- and 7-m high citrus trees from a single inoculation site at the top of the tree. This fast movement explains the difficulties of curing an HLB affected tree and reinforces the importance of the implementation of areawide preventive actions to control the disease in citrus orchards. One of the long-term solutions for cultivation of citrus in presence of HLB may be development of disease resistant cultivars. In the genus Citrus, resistance to HLB is not known although some tolerant varieties have been reported. We have identified resistance and significant field tolerance in many citrus relative genera native to Australia. Field and greenhouse studies indicated that many naturally occurring hybrids of citrus and Eremocitrus/Microcitrus are resistant to HLB. Since the HLB resistance in the Australian types appears to be heritable, we have generated hundreds of citrus hybrids by crossing Citrus and Poncirus with disease resistant/tolerant, sexually compatible Australian citrus relatives. The hybrid plants are challenged by exposure to psyllids carrying HLB-associated Liberibacter pathogen. Preliminary evaluations of the hybrids in green house experiments conducted in Fort Pierce, FL indicate that a significant number of the novel hybrids appear to be disease resistant. Further testing to confirm HLB resistance is in progress. The promising hybrids will be useful in understanding the basis of HLB resistance and in developing methods to impart resistance to commercial citrus cultivars. The Asian citrus psyllid is the insect vector responsible for the worldwide spread of ‘Candidatus Liberibacter asiaticus’ , the bacterial pathogen associated with citrus greening disease. Developmental changes in the insect vector impact pathogen transmission,mobile grow rack such that D. citri transmission of CLas is more efficient when bacteria are acquired by nymphs as compared to adults. We hypothesize that expression changes in the D. citri immune system, including the insect’s commensal microbiota, occur during development and regulate vector competency. In support of this hypothesis, more proteins, with greater fold changes, were differentially expressed in response to CLas in adults as compared to nymphs, including insect proteins involved in bacterial adhesion and immunity. Discovery of protein interaction networks has broad applicability in the study of hostmicrobe relationships. Using Protein Interaction Reporter technology, we show how protein interaction networks in D. citri are regulated during development and in response to CLas-infected citrus trees. Notably, a hemocyanin protein highly upregulated in response to CLas was found to physically interact with the CLas coenzyme A biosynthesis enzyme phosphopantothenoylcysteine synthetase/decarboxylase. In addition, hemocyanin was found to physically interact with several other D. citri signaling and stress response proteins.
Co-evolved protein interaction networks at the host-microbe interface are highly specific targets for controlling the insect vector responsible for the spread of citrus greening.Huanglongbing , also known as citrus greening, is one of the most destructive citrus diseases worldwide and is seen as a major threat to the multimillion dollar citrus industry in California. The vector of two of the bacterial species associated with this disease, Candidatus Liberibacter asiaticus and Ca. L. americanus , is the Asian citrus psyllid , Diaphorina citri. In August 2008, the first ACP in California was found in a trap located in San Diego County. The detection triggered an ongoing state wide risk-based survey and HLB testing program for ACP and HLB host plants in California. Since 2008, a total of 135,000 citrus trees and 252,000 ACP samples have been tested so far by the California Department of Food and Agriculture using the United States Department of Agriculture HLB Work Instruction which utilizes a TaqMan real-time PCR assay based on 16S rDNA primers and probe developed by Li et al. . After eight years of challenging Li’s 16S HLB primers with hundreds of thousands of environmental samples, we have found that Li’s primers are highly specific but on rare occasions will cross amplify other closely related bacteria such as Ca. L. crescens . Lcr has never been reported in citrus before, and has only been reported in Babaco papaya during a survey for Papaya Bunchytop Disease in Puerto Rico. Currently there are six recognized species of Candidatus Liberibacter, four of which are pathogenic and two that have not been shown to cause symptoms in plants. Lcr falls in the latter category and is also the only member of this group that has successfully been cultured in artificial media. To date, Lcr has been confirmed in six citrus samples collected in three different counties in California and one from Hawaii. The detections were made during routine HLB testing in which the samples produced Fam Ct values between 33-36. A 1,149 bp fragment of the 16S ribosomal RNA gene was amplified from the DNA extracts using conventional PCR with primers Ol1 and OI2c. The sequence showed 99% identity with the corresponding regions of Ca. L. crescens strain BT-1. All samples had the same two single nucleotide polymorphisms differing with positions characteristic to Lcr. This is the first report of Lcr in citrus. Follow up sampling and testing of the CA trees confirmed all were negative for the HLB associated bacterium. Only one tree was retested for Lcr and was confirmed positive. Plans to retest the other trees for Lcr are underway. We evaluated the percentage of psyllid population with Las and its correlation with insect population captured in sticky cards and the HLB management in the property over a year. Assessment was carried out in four citrus regions in São Paulo state according to the Fundecitrus Phytosanitary Alert System. Cards were installed, read and replaced every two weeks on the period of February 2014 to February 2015 in the regions of Avaré, Santa Cruz do Rio Pardo , Araraquara and Bebedouro . In each reading, up to 50 psyllids per regions were collected and detection of insects Las+ was made by qPCR. Bi-monthly averages of the percentage of psyllids Las+ were compared. Santa Cruz do Rio Pardo region had 71% of Las+ samples and differed from Bebedouro region that had 56%. Avaré and Araraquara regions had 68 and 66%, respectively. The number of psyllids increased gradually between July and August in Avaré and Santa Cruz regions, from September to October in Araraquara region and intermittently from October in Bebedouro region. Potentially infective psyllid was higher in early spring in Avaré and Santa Cruz, with the largest population; Araraquara were higher starting in October, peaked in December ; Bebedouro region reaches the highest values in January and February . Percentage of Las+ psyllids collected in properties without HLB management were higher and statistically different from the averages on the properties that adopted psyllid control. The relationship between the percentage of psyllids Las+ and the number of captured psyllids in the region in a given time denotes the most critical time of year that psyllids Las+ intake in the orchards can occur.