Production of Antibodies to Citrus Tristeza Virus in Transgenic Citrus

Citrus tristeza virus (CTV) is the most important virus disease of citrus in the world. CTV causes death of trees on sour orange rootstock and/or stem pitting of scions regardless of rootstock which results in trees of low vigor, reduced yield with reduction in size and quality of fruit. The purpose of this project was to produce monoclonal antibodies (MABs) to CTV coat protein (CP), develop single domain antibodies (dAbs) or Fab fragments which neutralize the infection by binding to the virus, and to produce transformed plants which express the dAbs. The objectives of this research have been met and putative transgenic tobacco and citrus plants have been developed. These putative transgenic plants are presently undergoing evaluation to determine the level of dAbs expression and to determine their resistance to CTV. Additionally, the CTV genome has been sequenced and the CP gene of several biologically characterized CTV strains molecular characterized. This has indicated a correlation between CP sequence homology and biological activity, and the finding of DI RNAs associated with some CTV strains. Several MABs have been produced which enable broad spectrum identification of CTV strains while other MABs enable differentiation between mild and severe strains. The use of selected MAbs and determination of the CP gene sequence has enabled predictions of biological activities of unknown CTV isolates. The epitopes of two MABs, one reacting selectively with severe CTV strains and the other reacting with all strains, have been characterized at the molecular level.

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Occurrence of Stem-Pitting Strains of Citrus tristeza virus in Croatia

Plant Disease ◽

10.1094/pd-89-0342b ◽

2005 ◽

Vol 89 (3) ◽

pp. 342-342 ◽

Cited By ~ 2

Author(s):

S. Černi ◽

D. Škorić ◽

M. Krajačić ◽

Ž Gatin ◽

C. Santos ◽

...

Keyword(s):

Mediterranean Basin ◽

Citrus Tristeza Virus ◽

Enzyme Linked Immunosorbent Assay ◽

Cp Gene ◽

Plant Pathol ◽

The Mediterranean ◽

Tristeza Virus ◽

Stem Pitting ◽

The Mediterranean Basin ◽

Citrus Tristeza

Citrus is grown in Croatia (approximately 1,500 ha of citrus groves) on the Dalmatian Coast and Islands between 42 and 43°30′N. The major species, Citrus unshiu Marc. (Satsuma mandarin), is grafted on trifoliate rootstock. The presence of Citrus tristeza virus (CTV) in Satsumas in the Neretva Valley Region was previously reported (3). During the course of a biomolecular characterization of isolates from Croatia, 15 budsticks were collected from field-infected, enzyme-linked immunosorbent assay (ELISA)-positive sources during the autumn of 2003 near Kaštela, Split, Metković (Neretva Valley), and on the island of Vis. Isolates were propagated by graft transmission to Madam Vinous sweet orange (SwO) and maintained in an insect-proof greenhouse at 21 to 33°C. Eight months later, the bark of terminal twigs was peeled off, and the wood was examined for the occurrence of pits. Typical tristeza stem-pitting (SP) was observed in four isolates originating from cvs. Fukumoto navel, Washington navel, and Ichimaru Satsuma and C. wilsonii. The bark from the infected sources was analyzed using immunocapture reverse transcription-polymerase chain reaction (RT-PCR) with primers CTV1 and CTV10 (1), targeting the whole coat protein (CP) gene. The PCR products of the expected size (669 nucleotides) were obtained and TA cloned (pGEM-T Easy Vector; Promega, Madison, WI) in E. coli cells. Thirty-two clones harboring the CTV CP gene were sequenced. Two of the SP isolates contained four genomic variants that differed an average of 2.0% from the severe SwO SP strains SY568 and Nuaga (4) from California and Japan, respectively. The other two SP isolates contained four variants that differed as little as 1.6% from the severe SwO SP from India, CTV-Puna, and CTV-Bangalore (2). The net average distance between these two clusters of sequences is 5.2%. One sequence from each of the four SP isolates was deposited in GenBank (Accession Nos. AY791841 to AY791844). These findings were confirmed by direct observation of SP symptoms in a Satsuma orchard in the Neretva Valley during the spring of 2004. No other conspicuous symptoms that could be attributed to CTV were observed in the field. Most Satsumas were introduced to the Neretva Region from Japan between 1964 and 1984. Together with the fact that the related Nuaga strain was also isolated from Satsumas in Japan (4), our results suggest that SwO SP strains were introduced into Croatia at the same time and have been spreading for several decades. It has been generally believed that this kind of CTV strains either do not exist in the Mediterranean basin or, when found (e.g., Spain), are immediately eradicated. The findings reported here suggest that the epidemiological scenario for the Mediterranean Basin requires revision. References: (1) G. Nolasco et al. Eur. J. Plant Pathol. 108:293, 2002. (2) A. Roy et al. Arch. Virol. 148:707, 2003. (3) A. Šarić and I. Dulić. Agric. Conspectus Sci. 55:171, 1990. (4) G. Suastika et al. J. Gen. Plant Pathol. 67:73, 2001.

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A model for the spread and control of citrus tristeza virus disease

Journal of Applied Statistics ◽

10.1080/02664768900000039 ◽

1989 ◽

Vol 16 (3) ◽

pp. 315-320

Author(s):

Ruth Marcus ◽

Hovav Talpaz ◽

Moshe Bar-Joseph

Keyword(s):

Citrus Tristeza Virus ◽

Virus Disease ◽

And Control ◽

Tristeza Virus ◽

Citrus Tristeza

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Genetic Diversity and Phylogenetic Analysis of Citrus tristeza virus Isolates from Turkey

Advances in Virology ◽

10.1155/2019/7163747 ◽

2019 ◽

Vol 2019 ◽

pp. 1-11 ◽

Cited By ~ 1

Author(s):

Gözde Erkiş-Güngör ◽

Bayram Çevik

Keyword(s):

Phylogenetic Analysis ◽

Citrus Tristeza Virus ◽

Molecular Characteristics ◽

Cloning And Sequencing ◽

Phylogenetic Groups ◽

The 1960S ◽

Tristeza Virus ◽

Virus Isolates ◽

Stem Pitting ◽

Citrus Tristeza

The presence of Citrus tristeza virus (CTV) in Turkey has been known since the 1960s and the virus was detected in all citrus growing regions of the country. Even though serological and biological characteristics of CTV have been studied since the 1980s, molecular characteristics of CTV isolates have not been studied to date in Turkey. In this study, molecular characteristics of 15 CTV isolates collected from different citrus growing regions of Turkey were determined by amplification, cloning, and sequencing of their major coat protein (CP) genes. The sequence analysis showed that the CP genes were highly conserved among Turkish isolates. However, isolates from different regions showed more genetic variation than isolates from the same region. Turkish isolates were clustered into three phylogenetic groups showing no association with geographical origins, host, or symptoms induced in indicator plants. Phylogenetic analysis of Turkish isolates with isolates from different citrus growing regions of the world including well-characterized type isolates of previously established strain specific groups revealed that some Turkish isolates were closely related to severe quick decline or stem pitting isolates. The results demonstrated that although CTV isolates from Turkey are considered biologically mild, majority of them contain severe components potentially causing quick decline or stem pitting.

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Stem Pitting Strains of Citrus Tristeza Virus in Indonesia

International Organization of Citrus Virologists Conference Proceedings (1957-2010) ◽

10.5070/c5160069w8 ◽

1991 ◽

Vol 11 (11) ◽

Author(s):

A. Muharam ◽

A. M. Whittle

Keyword(s):

Citrus Tristeza Virus ◽

Tristeza Virus ◽

Stem Pitting ◽

Citrus Tristeza

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Stem-Pitting Citrus tristeza virus Predominantly Transmitted by the Brown Citrus Aphid from Mixed Infections Containing Non-Stem-Pitting and Stem-Pitting Isolates

Plant Disease ◽

10.1094/pdis-10-10-0772 ◽

2011 ◽

Vol 95 (8) ◽

pp. 913-920 ◽

Cited By ~ 5

Author(s):

R. H. Brlansky ◽

Avijit Roy ◽

V. D. Damsteegt

Keyword(s):

Citrus Tristeza Virus ◽

Fruit Size ◽

Haplotype Diversity ◽

Aphid Transmission ◽

Single Strand Conformation Polymorphism ◽

Mixed Infections ◽

Tristeza Virus ◽

Stem Pitting ◽

Citrus Spp ◽

Citrus Tristeza

Citrus tristeza virus (CTV) is a phloem-limited Closterovirus that produces a variety of symptoms in various Citrus spp. One of these symptoms is stem pitting (SP). SP does not occur in all Citrus spp. but when it does it may cause low tree vigor, decline, and an economic reduction in fruit size and yield. Historically, the first appearance of CTV-SP in a citrus area often occurs after the introduction of the most efficient CTV vector, the brown citrus aphid (BCA), Toxoptera citricida. Hypotheses for this association range from the introduction of these strains in new planting materials to the increased ability of BCA to transmit SP strains from existing CTV sources. It is known that CTV often exists as a complex of isolates or subisolates. Single and multiple BCA transmissions have been used to separate different genotypes or strains of CTV from mixed CTV infected plants. This study was initiated to determine what the BCA transmits when an exotic severe SP CTV isolate B12 from Brazil or B408 from Dominican Republic are mixed with a non-SP (NSP) isolate, FS627 from Florida. Biological and molecular data was generated from grafted mixtures of these isolates and their aphid-transmitted subisolates. Single-strand conformation polymorphism patterns of the 5′ terminal region of open reading frame (ORF) 1a, the overlapping region of ORF1b and ORF2, and the major coat protein gene region of NSP and SP CTV-grafted plants remained unchanged but the patterns of doubly inoculated plants varied. The haplotype diversity within SP isolates B12, B408, and mixtures of NSP and SP isolates (FS627/B12 and FS627/B408) and aphid-transmitted subisolates from doubly inoculated plants was determined by analysis of the haplotype nucleotide sequences. Aphid transmission experiments, symptoms, and molecular analyses showed that SP-CTV was more frequently transmitted with or without NSP-CTV from mixed infections.

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Citrus Tristeza Virus Isolates of the Same Genotype Differ in Stem Pitting Severity in Grapefruit

Plant Disease ◽

10.1094/pdis-12-19-2586-re ◽

2020 ◽

Vol 104 (9) ◽

pp. 2362-2368

Author(s):

Glynnis Cook ◽

Beatrix Coetzee ◽

Rachelle Bester ◽

Johannes H. J. Breytenbach ◽

Chanel Steyn ◽

...

Keyword(s):

Citrus Tristeza Virus ◽

Aphid Transmission ◽

Common Source ◽

Citrus Paradisi ◽

Full Genome ◽

Full Genome Sequencing ◽

Reading Frame ◽

Tristeza Virus ◽

Stem Pitting ◽

Citrus Tristeza

Two isolates of the T68 genotype of citrus tristeza virus (CTV) were derived from a common source, GFMS12, by single aphid transmission. These isolates, named GFMS12-8 and GFMS12-1.3, induced stem pitting with differing severity in ‘Duncan’ grapefruit (Citrus × paradisi [Macfad.]). Full-genome sequencing of these isolates showed only minor nucleotide sequence differences totaling 45 polymorphisms. Numerous nucleotide changes, in relatively close proximity, were detected in the p33 open reading frame (ORF) and the leader protease domains of ORF1a. This is the first report of full-genome characterization of CTV isolates of a single genotype, derived from the same source, but showing differences in pathogenicity. The results demonstrate the development of intragenotype heterogeneity known to occur with single-stranded RNA viruses. Identification of genetic variability between isolates showing different pathogenicity will enable interrogation of specific genome regions for potential stem pitting determinants.

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INCIDENCE OF CITRUS TRISTEZA VIRUS, THE STEM PITTING ISOLATES IN THAILAND

Acta Horticulturae ◽

10.17660/actahortic.2008.773.6 ◽

2008 ◽

pp. 51-56

Author(s):

R. Sdoodee ◽

P. Tothaum ◽

J. Worapattamasri ◽

P. Jantaradsamee ◽

S. Patnantawech ◽

...

Keyword(s):

Citrus Tristeza Virus ◽

Tristeza Virus ◽

Stem Pitting ◽

Citrus Tristeza

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Citrus Diseases Exotic to Florida: Citrus Tristeza Virus–Stem Pitting (CTV-SP)

EDIS ◽

10.32473/edis-pp149-2022 ◽

2022 ◽

Vol 2022 (1) ◽

Author(s):

Amit Levy ◽

Ozgur Batuman ◽

Peggy Sieburth ◽

Ajia Paolillo ◽

Kuang-Ren Chung ◽

...

Keyword(s):

Citrus Tristeza Virus ◽

Viral Disease ◽

Original Version ◽

Educational Tool ◽

Citrus Industry ◽

Citrus Diseases ◽

Florida Citrus ◽

Tristeza Virus ◽

Stem Pitting ◽

Citrus Tristeza

This document is one in a series designed to provide important information on the causal agent, symptoms, and transmission of exotic citrus diseases. This information can be used as an educational tool to raise awareness about these diseases and for scouting and identification efforts. Disseminating information about the diseases to the citrus industry may prevent their introduction and spread in Florida. This document will focus on the exotic viral disease caused by isolates of citrus tristeza virus–stem pitting (CTV-SP). Original version: Chung, Kuang-Ren, and Ronald Brlansky. 2006. “Citrus Diseases Exotic to Florida: Citrus Tristeza Virus– Stem Pitting (CTV-SP)”. EDIS 2006 (7). https://doi.org/10.32473/edis-pp149-2006.

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