Citrus leprosis identification and care

Citrus leprosis (CL) is one of the most serious threats to São Paulo's citrus industry, being a disease of viral and endemic etiology present on the American continent, from Argentina to Mexico. To assess its economic importance, it is estimated that producers in São Paulo spend approximately 80 million dollars/year only on specific acaricides to control the mite vector, preventing the spread of CL. It was first reported in Florida at the beginning of the 20th century, where it caused significant losses and was referred to as “scaly bark" or "nailhead rust”, but it stopped being detected from the 1960s onwards. From the 1920s onwards, a similar disease was described in Argentina, Paraguay and Brazil, considered similar to that in the USA, and came to be called leprosis. The disease is characterized by lesions on leaves, branches and fruits of orange trees; can result in defoliation and intense fruit drop. Branch lesions, if very numerous, can coalesce, resulting in branch death, and result in a downward death of the plant within a few years (Figure 1 AE). Orange trees are the most susceptible; tangerines and pomelos to a lesser extent and lemons are considered immune.

At first, the etiology of CL was controversial, but in 1940, Frezzi, in Argentina, demonstrated the association of the mite, identified as Brevipalpus obovatus, with LC. Musumecci and Rossetti demonstrated in 1963 that LC in Brazil was transmitted by B. phoenicis. Knorr, in Florida, noticed that lesions in LC branches, grafted onto healthy branches, spread, suggesting a viral etiology, a hypothesis that was subsequently confirmed by a series of evidence, such as the presence of cellular changes and particles similar to those caused by Kitajima viruses. et al in the 1970s and mechanical transmission of the virus by Colariccio et al in 1995. CL's presence is confirmed in almost all Latin American countries (Argentina, Paraguay, Brazil, Bolivia, Colombia, Venezuela, Panama, Nicaragua, Honduras, El Salvador, Belize and Mexico).

In 1995, the signatory joined Esalq as a visiting professor, after retiring from the University of Brasília, where he was a professor, at the invitation of professor Raul Machado, to manage the newly created Electron Microscopy Laboratory on the campus. To maintain his research, he decided to begin investigations into plant viruses transmitted by mites. Brevipalpus (VTB), focusing on LC, an activity maintained to this day and which had the support of a multidisciplinary, multi-institutional and multinational team, and financial support from Fapesp. Since 2006, when he compulsorily retired, he has maintained studies on VTB as a volunteer employee/collaborating researcher at Esalq, with the approval of the Department of Phytopathology and Nematology and the institution's management.

In the 24 years of investigations since then, there has been an exponential increase in information not only about CL, but about VTB in general, with the identification and characterization of several of them and advances in knowledge about the biology and systematics of mites Brevipalpus, and the relationships between those who act as vectors and the viruses transmitted.

A significant advance in understanding the CL pathosystem was made when partial sequences of the causative virus were obtained from dsRNA in 2003, with which a tool for its molecular detection (RT-PCR) was developed quickly and accurately, in a large number of samples, having contributed to confirming the presence of LC in the countries already mentioned. Soon afterwards, in 2006, it was possible to obtain the total genome sequence of the CL agent, which was called leprosis virus C (CiLV-C), distinct from other known plant viruses, for which it was proposed to create of a new genre Cilevirus, accepted by International Committee for Virus Taxonomy (ICTV).

Morphological studies had already indicated, in 2003, that there are two distinct types of VTB based on the effects they caused in infected cells. The cytoplasmic type, with short bacilliform particles and the presence of a dense inclusion (viroplasm) in the cytoplasm (Figure 2A). Another, referred to as nuclear, induces a clear viroplasm in the nucleus, with short rod-shaped particles present in the nucleus and cytoplasm (Figure 2B). Molecular studies confirmed this dichotomy, indicating that the cytoplasmic type would correspond to the Cilevirus, such as CiLV-C, while nuclear would correspond to Dichorhavirus. This genus, whose type member is orchid spot virus (OFV) (Figure 3D), is close to the genera Nucleorhabdovirus e Cytorhabdovirus, both plant viruses and are part of the Rhabdoviridae family.

Among the VTB known in Brazil, CoRSV (a dichorhavirus) (Figure 3E) and passion fruit green spot (PfGSV) (Figure 3B), a possible Cilevirus, can cause certain losses, and in addition to them, successive surveys have identified several cases of VTB, such as tawny leprosis (LigLV) (Figure 3A), violet solano annular spot (SvRSV) (Figure 3C), lady's annular spot. da-noite (CeRSV) (Figure 3G), chlorotic spot of the Clerodendrum (ClCSV) (Figure 3F) and others still unidentified, in several species of plants, generally ornamental. In all these cases of VTB, the common characteristic is that the symptoms resulting from the infection are invariably localized (chlorotic, ring, necrotic spots), which do not become systemic, possibly because these VTB are unable to infect phloem cells and, thus, invade the sieved vessels, to be transported remotely within the infected plant.

It is worth highlighting the fact that CL, in reality, is not a single disease, but a syndrome with very similar symptoms, which can be caused by several different VTB, in addition to CiLV-C, which is by far the most economically important and the more widespread (Figure 1 AE). In Colombia, CiLV-C2 was described, another Cilevirus (Figure 1F). In Mexico and Colombia, an OFV isolate was identified as the causal agent of CL (Figure 1J). Other Dichorhaviruses such as leprosis N (CiLV-N) (Figure 1G) detected in a region with a mild climate and higher altitudes in Southeastern Brazil, chlorotic spot (CiCSV) (Figure 1H) identified in Piauí, bright spot (CiBSV) (Figure 1I) in the states of Santa Catarina and Rio Grande do Sul they are considered part of the CL syndrome. The first CL outbreak in orange trees (Figure 1K) outside the American continent, caused by an OFV isolate, was recently reported in South Africa. Although Florida CL disappeared about 50 years ago, analyzes carried out on herbal samples showed that it was caused by another possible OFV isolate. In Colombia, CiLV-C was detected in 2006, and CiLV-C2 in 2013, with indications that this becomes the predominant virus. In Brazil, CiLV-C2 has not yet been registered, nor have OFV isolates affecting citrus plants as occurs in Colombia, Mexico and South Africa. Dichoravirus described among us (CiLV-N, CiCSV and CiBSV) are marginal and rare in occurrence. Studies on CiLV-C diversity show that the isolate originally studied and referred to as Cordeirópolis (CiLV-C CRD) occurs throughout the American continent. But in the state of São Paulo, another isolate detected approximately ten years ago, called São José do Rio Preto (CiLV-C SJP), is rapidly dispersing throughout all orchards. Possibly this isolate would be more efficiently transmitted by populations of B. yothersi present in orchards.

Investigations into this VTB pathosystem include mites Brevipalpus (Figure 4 AB), belonging to the Tenuipalpidae family. Currently, 291 recognized species of Brevipalpus, which are distributed globally from equatorial to subtropical regions. Few of them, however, are involved in the transmission of VTB. Until recently, species were recognized B. obovatus, B.californicus e B. phoenicis as vectors. Mites Brevipalpus They are small (adults measure ca. 0,3mm), flat and red-brown in color (Figure 4A), and most of them reproduce by thelytocal parthenogenesis, females generating females. They feed by sucking cells, which are punctured by the stylet (Figure 4 C, D). There is evidence indicating that VTB particles present in infected cells are ingested together with the cellular content circulating in the mite's body from the midgut, passing through the anterior prosomal gland (=salivary) and introduced into the host plant with saliva at the beginning of feeding. , resulting in infection. You Dichorhavirus, such as OFV, CoRSV and ClCSV would multiply in the vector's tissues, while Cilevirus (CiLV-C, CiLV-C2, PfGSV, SvRSV) would only circulate in the body of the mite vector, without infecting it. There have recently been changes in the morphological criteria for identifying species of Brevipalpus with the introduction of additional characters and also with the use of molecular markers (nuclear or mitochondrial DNA) resulting in important changes in systematics. Thus, the species considered B. phoenicis it was divided into at least eight species, some new and others “resurrected” and this led to the reconfiguration of the species considered as vectors. For example, the main vector species of CiLV-C was identified as B. yothersi, while the CoRSV vector would be B. papayensis. Table 1 lists the VTB and species of Brevipalpus currently recognized as vectors.

Today, the epidemiology of CiLV-C is well known and its management strategies in the São Paulo citrus industry are essentially based on chemical control and other cultural practices to prevent the spread of its vector, B. yothersi. The recent arrival of Greening/HLB caused significant changes in phytosanitary management (intensified use of insecticides, reduction in spacing between plants) in São Paulo's citrus industry, combined with a smaller number of specific acaricides for B. yothersi available. To face this new challenge, several researches are underway to find solutions to keep CL under control.

Regarding the creation of the Kitaviridae family, it was a courtesy provided by the working group responsible for the systematics of Cilevirus, within ICTV. The ICTV recently encouraged the creation of new families for numerous “orphan” genera, and this group proposed including the genera Cilevirus, Higrevirus e Blunervirus, which have a similar genomic organization, in a new family Kitaviridae, honoring the signatory, due to his contributions to the advancement of knowledge about VTB. However, it must be made very clear that such advances would not have been achieved by the work of a single person, and resulted from the participation of a large group, with more than 50 people (teachers, researchers and students), from several Brazilian institutions and the abroad, in addition to research funding agencies such as Fapesp, with which this effort must be shared.

Elliot W. Kitajima, Collaborating Researcher, Department of Phytopathology and Nematology, Escola Superior de Agricultura Luiz de Queiroz, USP, Piracicaba campus

Growing Vegetables and Fruits November 2019

With each new edition, Cultivar Hortaliças e Frutas publishes a series of technical content produced by renowned researchers from all over Brazil, which address the main difficulties and challenges encountered in the field by rural producers. Through research focused on controlling the main pests and diseases in vegetable and fruit cultivation, the Magazine helps farmers in the search for management solutions that increase their profitability. In the November 2019 edition you can also see: 

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