Journal of Citrus Pathology

Due to their small size, locating pathogenic RNAs, such as viroids, in plant tissue and cell organelles has been difficult. Viroids are small circular single-stranded RNA plant pathogens that reduce plant growth, vigor, and yield in economically important crops such as potato, tomato, hops and citrus. Viroid infections in plants are largely diagnosed by dot blot hybridization, polyacrylamide gel electrophoresis (PAGE) or gels, or real-time quantitative polymerase chain reaction (qPCR). Because traditional plant in situ hybridization studies for viroids are often limited by the lack of signal amplification and binding specificity due to the small target sequence, we examined the use of RNAscope™ (Advanced Cell Diagnostics Inc., Newark, CA). This in situ hybridization method increases the detection by amplifying the signal of a single target, to detect the cellular distribution of citrus exocortis viroid (CEVd) with higher sensitivity and specificity. We found that after optimization, CEVd was localized in nuclei of infected cells as clearly distinguishable punctate red dots visible with light microscopy at low magnification, suggesting that the RNAscope™ can be used to study viroids in situ.

High-throughput sequencing (HTS) of citrus indicator hosts, originally inoculated from field samples and showing transient chlorotic flecking or oak leaf patterns, revealed the presence of the first South African variant of citrus virus A (CiVA). This virus was first identified in citrus in Italy and was classified as a member of the second species (Coguvirus eburi) of the genus Coguvirus within the order Bunyavirales. The South African sequence variants of CiVA share 95.26-95.55% (RNA1) and 94.82-97.85% (RNA2) nucleotide sequence identity with the CiVA sequences from the Italian isolate. The discovery of CiVA in South African citrus orchards indicates a wide distribution of CiVA and further investigations are required to determine the association of CiVA with citrus disease symptoms.

Citrus tristeza virus (CTV) is the most important viral pathogen of citrus. CTV-induced stem pitting negatively impacts grapefruit and sweet orange production. The mechanisms of stem pitting development in CTV-infected citrus remain unclear. This study evaluated the utility of high-resolution CT scanning as a tool to study stem pitting in live citrus material. CT scans were used to easily identify pits based on differences in tissue density. Stem pits were also mapped and modelled three-dimensionally along the length of the stem. Nano-CT scanning proved to be a potentially valuable, non-destructive method for stem pitting characterization in citrus.

Citrus is the highest-value fruit crop in terms of international trade. However, citrus species are susceptible to several diseases caused by different pathogens which directly cause a decrease in production leading to economic losses. In the last half-century, the citrus industry in Uruguay has had a strong socio-economic impact and is also constantly evolving to stay competitive in world markets, by introduction of new varieties and improvement of production practices to obtain high yielding orchards. Nevertheless, despite the existence since 2014 of the Uruguayan National Citrus Sanitary and Certification Program, scarce information is available regarding the virus and viroid status of commercial citrus in Uruguay. The incidence of citrus tristeza virus (CTV), citrus psorosis virus (CPsV), satsuma dwarf virus (SDV), citrus exocortis viroid (CEVd), hop stunt viroid (HSVd), citrus dwarfing viroid (CDVd), citrus bark cracking viroid (CBCVd) and citrus bent leaf viroid (CBLVd) was investigated in this study, as well as CTV genotypes prevalent in the country. Molecular diagnostic assays were used to test 1175 samples including Valencia and Navel sweet oranges, Mandarin hybrids, Clementines and lemons, which were randomly collected from seven citrus-producing provinces. Only 6% of the samples were negative for the pathogens screened, while 93% of them were CTV positive. SDV, CBLVd and CBCVd were not detected. Co-infections were frequently detected, finding plants with up to four simultaneous pathogens, including CTV in all co-infected plants. This is the first comprehensive survey of several citrus-infecting viruses and viroids in Uruguay, as well as a determination of the CTV genotypes prevalent in the country.

Interaction between viruses is one of the major factors that determines viral population structure or equilibrium, which is a determinant of virus pathogenesis. If we could manipulate virus interactions, we could potentially limit the effects of disease. Using citrus tristeza virus (CTV) as a model, we examined if we could alter the equilibrium of a population by adding different CTV genotypes or other citrus pathogens. We found that population structure could be altered through the addition of specific CTV genotypes, disrupting existing interactions and selectively changing the titer of specific genotypes, while the addition of other citrus viruses or viroids did not have an effect.

IOCV is pleased to present “Citrus Rootstocks: Their Characters and Reactions”, an unpublished manuscript by the late Prof. W.P. Bitters, University of California, Riverside. Based upon Prof. Bitters research and many other sources, this work was compiled between the late 1960’s and 1986 (additional information in preface). It represents a comprehensive (for that era) treatment of many facets of citrus rootstock physiology, horticulture, and pathology. Rootstocks serve as both conduits for citrus disease development and potential disease management tools, as well as tools in dealing with abiotic stresses. The history of the use of rootstocks for citrus has largely been determined by these factors, and they continue to greatly influence citrus rootstock research and usage. Although many advances in knowledge regarding citrus rootstocks have been made since 1986, the basic information presented remains important for all citriculturists. We hope that this Special Topic in IOCV’s Journal of Citrus Pathology makes this valuable resource more widely available.