2. Ozgur Batuman, associate professor, Plant Pathology Department, UF/IFAS Southwest Florida REC; Amit Levy, assistant professor, Plant Pathology Department, UF/IFAS Citrus REC; Mark E. Hilf, research plant pathologist (retired), USDA-ARS-USHRL; Peggy J. Sieburth, biological scientist III (retired), Bureau of Citrus Budwood Registration, FDACS DPI; Ronald H. Brlansky, professor emeritus; and William O. Dawson, professor emeritus, Plant Pathology Department, UF/IFAS Citrus REC; UF/IFAS Extension, Gainesville, FL 32611.

The citrus industry of Florida faces insurmountable challenges against the destructive diseases citrus tristeza and Huanglongbing (HLB, or citrus greening). Though the tristeza causal agent, citrus tristeza virus (CTV), has been in Florida decades longer than HLB, growers have concentrated most of their efforts on combating the more detrimental HLB. The Asian citrus psyllid (Diaphorina citri; ACP) is the insect vector of the bacterial pathogen Candidatus Liberibacter asiaticus and transmits the incurable HLB to all commercial citrus. During our searches for biological and viral controls against the ACP, we consistently detected sequences of CTV in Florida field populations of ACP. This unexpected finding led us to investigate whether ACPs collected from young shoots could be used as a tool to survey CTV in Florida citrus groves. We first surveyed for the most common CTV strains in Florida (T30, T36, and VT/T68) in citrus trees on mostly sour orange (Citrus aurantium) rootstock, the rootstock susceptible to CTV decline. Out of 968 trees sampled across five years (2018-2022), approximately 8.2% were positive for CTV, with more than half of the CTV-positive trees infected with strain T30. Simultaneously, we looked at CTV strains in ACPs during this time and found that approximately 88% of pooled adult and nymph ACPs also had CTV, with over half the positive samples having the T36 strain. As a result of the much higher CTV incidences in the ACPs, we conducted a second investigation into whether we could more easily detect the same CTV strains in ACP nymphs as in CTV-infected citrus tissue. After individually sampling 43 trees and pooling the nymphs from each tree, we detected CTV at about the same incidence in the citrus tissue and the nymphs, but with much less ACP tissue, time, and resources required for detection compared to citrus tissue. Results from this study illustrate the sustained threat of CTV to Florida citrus and demonstrate the ACP as a potential bioindicator for CTV.

The destructive citrus disease, Huanglongbing (HLB) or citrus greening, continues to devastate Florida's citrus industry. A hemipteran insect, the Asian citrus psyllid (ACP), disperses Candidatus Liberibacter asiaticus, one of the putative bacterial pathogens of HLB. This study builds upon ongoing research utilizing high-throughput sequencing to analyze the virome of ACP populations collected from citrus groves throughout Florida. Following the widespread detection of sequences aligning to the genome of citrus tristeza virus (CTV) across consecutive years in the Florida ACP virome, we continued to detect a pervasive amount of CTV in Florida ACPs during subsequent years. Simultaneously, we also detected mixed infections of CTV strains in pooled ACPs from different Florida regions. Predating the HLB epidemic, CTV has been present in Florida for many years and our results confirm its widespread and diverse persistence in Florida citrus groves through a unique lens, the ACP. CTV presence in the ACP likely results from feeding on CTV-infected citrus trees in Florida citrus groves, which may help to understand an overlapping presence of CTV and HLB, both endemic citrus pathosystems in the state, and their role in future integrated pest management strategies.

Citrus tristeza virus (CTV) is a major cause of the decline and eventual death of trees on sour orange rootstocks. This 2-page fact sheet is part of the 2018–2019 Florida Citrus Production Guide. Written by O. Batuman, A. Levy, M. E. Hilf, P. J. Sieburth, W. O. Dawson, and R. H. Brlansky, and published by the Plant Pathology Department, January 2019. PP-181/CG039: 2022–2023 Florida Citrus Production Guide: Tristeza Decline (ufl.edu)

Citrus tristeza virus (CTV) is a major cause of the decline and eventual death of trees on sour orange rootstocks. This 2-page fact sheet is part of the 2019–2020 Florida Citrus Production Guide. Written by O. Batuman, A. Levy, M. E. Hilf, P. J. Sieburth, W. O. Dawson, and R. H. Brlansky, and published by the Plant Pathology Department, March 2019. PP-181/CG039: 2022–2023 Florida Citrus Production Guide: Tristeza Decline (ufl.edu)

All certified citrus nursery trees can trace their origins to fully tested scion and foundation source trees that remain clean and disease-free in screenhouses. Implementation of real-time polymerase chain reaction (PCR) for pathogen testing has lead to an increased requirement for both the testing efficiency and sensitivity in the Florida Bureau of Citrus Budwood Registration testing program. Budwood source citrus trees are tested annually for the vectored diseases Citrus tristeza virus (CTV) and citrus greening (HLB) and once every 6 years for other graft-transmissible diseases. To test the current number (more than 6000) of source trees annually, streamlining of collection and extraction of samples was necessary. Advance preparation allowed greater efficiency during the late summer to early fall months that samples are collected, extracted, and tested for HLB. Specialized equipment, including a new type of grinding apparatus, reduced costs and the time needed for processing. The second yearly collection from the same trees had been carried out during Spring 2008 for CTV ELISA testing. Two real-time CTV PCR tests, used in conjunction from a single extraction, have replaced the second collection and CTV enzyme-linked immunosorbent assay (ELISA) test. After extraction, testing for each pathogen can be completed in one month, previously an impossible task by conventional PCR. Citrus leaf blotch virus was the first pathogen tested using the same extraction as for HLB. One collection, one sample, and one extraction save money and allow for thorough, sensitive budwood testing for multiple citrus pathogens.

Citrus tristeza virus (CTV) has affected how citrus is grown in Florida since the 1950s. The brown citrus aphid, first detected in Florida in 1995, is an efficient vector of CTV and is capable of spreading severe forms of CTV throughout the state. The use of molecular markers for CTV led to the discovery of aphid transmitted stem-pitting forms of CTV (SP-CTV) in Polk County, central Florida. A survey to determine if SP-CTV was present was undertaken for the eleven counties representing 80% of commercial citrus production in Florida. Five sweet orange and two grapefruit sites per county were surveyed using a hierarchical bulk sampling procedure. Immuno-capture reverse transcriptase polymerase chain reaction (ICRT-PCR) with Type 11 primers was used for initial screening followed by other SP-CTV markers for positive samples. Fifty-five percent (42 of 77) of the sites surveyed tested negative in all samples. The majority of sites testing positive, 61%, had a less than a 5% chance of any single tree in that block testing positive. Only six sites had a greater than a 10% chance of any one tree testing positive for the SP-CTV markers. There were two main pattern profiles of markers present. Not all of these isolates have been evaluated in biological indicators so whether they can cause significant damage and what their threat is to the citrus industry has yet to be determined. Currently, a strong Citrus Budwood Registration Program and increasing the number of budwood source trees under protective screen will prevent the spread of severe forms of CTV throughout the nursery industry.