Molecular and biological evidence for a severe seedling yellows strain of Citrus tristeza virus spreading in southern Italy

Citrus tristeza virus (CTV) outbreaks have been reported in the main citrus-growing regions of Italy in the past 10 years. In some areas where eradication efforts failed to suppress spread, high CTV incidence is now observed. Recently, potentially severe CTV strains were detected in Calabria (southern Italy), one of the major citrus-growing area. As a result, investigations of the virulence and molecular features of CTV populations spreading in this region were undertaken. Virus was detected by enzyme-linked immunosorbent assay (ELISA) using a broad spectrum polyclonal antiserum, and was differentiated into potential virulent categories with the severe-strain discriminating monoclonal antibody MCA13. Isolate genotyping was conducted using reverse-transcription polymerase chain reaction (RT-PCR) with multiple molecular markers (MMM), single-strand conformation polymorphism (SSCP) analysis of the amplicons from the genes coding for the coat protein (CP) p25 and the non-structural p20 protein as well as sequence analysis. Based on the serological reactivity, the isolates were differentiated in two distinct serogroups: MCA13-reactive and MCA13 non-reactive. Similarly, based on the molecular profile, the isolates were grouped in two genetically distinct phylogenetic clusters, and associated either with a T30-like or with a T3-like genotype. These data were related to the results of biological indexing on standard indicator plants, which distinguished isolates causing mild or severe seedling yellow reactions. The study has demonstrated the presence of MCA13-reactive isolates associated with a T3-like genotype and causing severe seedling yellows in sour orange, grapefruit and lemon seedlings, and stem pitting in Mexican lime

Assessment of <I>Citrus tristeza virus</I> (CTV) incidence in Calabria, southern Italy: results of a three-year survey

Since 2006 a survey on Citrus tristeza virus (CTV) has been carried out in Calabria, southern Italy, to determine the occurrence of the virus, to evaluate its incidence, to identify and characterize the virus strains, and to monitor the aphid vector populations. Citrus samples were collected from nurseries and orchards located in the five provinces of the region. The virus was not detected in the citrus-growing areas of Catanzaro (CZ) or Crotone (KR), whereas it was found in three orchards in Cosenza (CS), three in Vibo Valentia (VV) and twelve citrus plantings in Reggio Calabria (RC). The highest infection percentages occurred in citrus orchards close to fields already infected with CTV. Infections were detected not only in foreign cultivars, but also in local cultivars such as ‘Comune’ clementine, ‘Moro’, ‘Ovale’ and ‘Tarocco’ sweet orange, suggesting that CTV was transmitted by aphids. CTV occurred in only a few plantlets sampled in two citrus nurseries located near the main areas already infected. Serological differentiation of several CTV field isolates revealed that the mild strains were prevalent. Aphis gossypii (Glover) and A. spiraecola Patch (= A. citricola Van der Goot) were the most frequent aphids in the orchards, whereas Toxoptera aurantii (Boyer de Foscoulombe) and Myzus persicae (Sulzer) occurred with low incidence. The absence of T. citricidus (Kirkaldy) was confirmed

Molecular and biological evidence for a severe seedling yellows strain of Citrus tristeza virus spreading in Calabria (Southern Italy)

Citrus tristeza virus (CTV) outbreaks have been reported in the main citrus-growing regions of Italy in the past 10 years. In some areas where eradication efforts failed to suppress spread, high CTV incidence is now observed. Recently, potentially severe CTV strains were detected in Calabria (southern Italy), one of the major citrusgrowing area. As a result, investigations of the virulence and molecular features of CTV populations spreading in this region were undertaken. Virus was detected by enzyme-linked immunosorbent assay (ELISA) using a broad spectrum polyclonal antiserum, and was differentiated into potential virulent categories with the severe-strain discriminating monoclonal antibody MCA13. Isolate genotyping was conducted using reverse-transcription polymerase chain reaction (RT-PCR) with multiple molecular markers (MMM), single-strand conformation polymorphism (SSCP) analysis of the amplicons from the genes coding for the coat protein (CP) p25 and the non-structural p20 protein as well as sequence analysis. Based on the serological reactivity, the isolates were differentiated in two distinct serogroups: MCA13-reactive and MCA13 non-reactive. Similarly, based on the molecular profile, the isolates were grouped in two genetically distinct phylogenetic clusters, and associated either with a T30-like or with a T3-like genotype. These data were related to the results of biological indexing on standard indicator plants, which distinguished isolates causing mild or severe seedling yellow reactions. The study has demonstrated the presence of MCA13-reactive isolates associated with a T3-like genotype and causing severe seedling yellows in sour orange, grapefruit and lemon seedlings, and stem pitting in Mexican lime