First Report of Peach latent mosaic viroid Infecting Peach in Egypt.

Peach latent mosaic viroid (PLMVd) is a widespread pathogen of stone fruit trees in some European and Mediterranean countries and also in North America. To access the presence of the viroid in Egypt, a survey was conducted that covered five commercial peach orchards in the El Khatatba Region in Al Minufiya Governorate. During 2003 and 2004, 73 peach trees (cv. Florida grafted on Nemagard rootstock) were visually inspected and sampled. No symptoms characteristic of PLMVd infection, such as mosaic, delayed growth, or fruit suture cracking, were observed. All samples were tested for the presence of PLMVd using dot-blot hybridization and reverse transcription (RT)-PCR. Aliquots (5 µl) of total nucleic acids extracted from approximately 2 mg of leaf tissue were spotted onto positively charged nylon membranes and hybridized under stringent conditions with a digoxigenin-labeled riboprobe (2). The extracts (1 µl) also were used in RT-PCR as described (1). Only 1 of the 73 peach trees was positive for PLMVd using these detection techniques. The RT-PCR product was of the size expected for PLVMd and was cloned and sequenced. The 339 nucleotide sequence was deposited in GenBank as Accession No. DQ839564. The sequence of this Egyptian PLMVd isolate was 94% identical to the reference PLMVd variant (GenBank Accession No. M83545) and most closely (95%) related to Canadian isolate variant 16 (GenBank Accession No. AJ550911). Such a low incidence compared with other countries may be because the survey was restricted to a limited number of samples, conducted on newly reclaimed lands where no sources of infection existed before, and material with relatively low PLMVd incidence might have been used for planting. Although the incidence of PLMVd was low in this survey, the occurrence represents a threat to the stone fruit tree industry in this country and regular screening of PLMVd in certification programs is suggested. To our knowledge, this is the first report of PLMVd on peach in Egypt. References: (1) S. Loreti et al. EPPO Bull. 29:433, 1999. (2) A. M. Shamloul et al. Acta Hortic. 386:522, 1995.

A Survey of Citrus Viroids in Campania (Southern Italy).

Within a certification program of citrus, a survey covering 14 citrus fields not previously screened were randomly selected in seven areas of the Campania Region of southern Italy to evaluate the presence and distribution of citrus viroids in the region. During the last 3 years, 112 trees were visually inspected and sampled including 14 'Navelina', 8 'Washington navel', 13 'Biondo Commune', 10 'Tarocco', 6 'Valencia late', 1 'Sanguinello', and 1 'Vanilla' sweet oranges (Citrus sinensis), four sour oranges (C. aurantium), 21 'Commune' and 7 'Monreale' clementines (C. clementina), 8 'Common' and 2 'Avana' mandarins (C. reticulata), 8 'Ovale di Sorrento' and 6 'Zagara' lemons (C. lemon), 2 Tangelos (C. reticulata × C. paradisi), and 1 grapefruit (C. paradisi). Nearly all (37 of 38) mandarin and clementine trees showed cachexia-like symptoms. Severe bark scaling that is characteristic of exocortis disease was observed on a single 'Ovale di Sorrento' lemon, the only tree incidentally found to be grafted onto the exocortis-sensitive trifoliate orange (Poncirus trifoliata) rootstock, since sour orange (C. aurantium) is the main rootstock used in Campania. Samples (104 of 112) that had been graft-inoculated into Arizona 861-S1 'Etrog citron' (C. medica L.) on rough lemon (C. jambhiri Lush.) rootstock showed typical viroid symptoms. Viroid infection was confirmed by sequential polyacrylamide gel electrophoresis analysis (2) and molecular hybridization of imprinted membranes (1). Citrus exocortis viroid (CEVd), Hop stunt viroid (HSVd), and Citrus viroid III (CVd-III) were the most frequently detected, accounting respectively, for 67.9, 86.6, and 84.8% of the infected samples. Citrus viroid IV (CVd-IV) and Citrus bent leaf viroid (CBLVd) were found in 24.1 and 13.4% of the sources. CBLVd was detected in sweet orange ('Tarocco' and 'Biondo Commune') and lemon ('Ovale di Sorrento' and 'Zagara'), while CVd-IV was detected in sweet orange ('Tarocco', 'Biondo Commune', and 'Washington navel'), lemon ('Ovale di Sorrento' and 'Zagara'), clementine ('Commune'), and sour orange. Most trees (82.1%) were infected with more than one viroid species. The most frequent viroid combinations were CEVd + HSVd + CVd-III (40.2%), HSVd + CVd-III (16.1%), and CEVd + CBLVd + HSVd + CVd-III + CVd-IV (12.5%). To our knowledge, this is the first report of CEVd, CBLVd, HSVd, CVd-III, and CVd-IV in the Campania Region and the first report of CBLVd and CVd-IV in Italy. References: (1) A. Palacio et al. Eur. J. Plant Pathol.105:897, 2000. (2) R. F. Rivera-Bustamante et al. Anal. Biochem. 156:91, 1986.

Apple dimple fruit viroid: Fulfillment of Koch's Postulates and Symptom Characteristics.

The previously reported properties of a small circular RNA isolated from plants of cultivar Starking Delicious with symptoms similar to those of the dapple apple disease induced by Apple scar skin viroid (ASSVd), strongly suggested that it is a distinct viroid species of genus Apscaviroid to which ASSVd also belongs. Nevertheless, the autonomous replication of this novel RNA, termed Apple dimple fruit viroid (ADFVd), and the nature of the symptoms incited when inoculated free of other pathogens that might be present in the original source, were not determined. We addressed these questions by mechanically inoculating purified ADFVd into young apple seedlings (Golden). Dot blot hybridization of leaf extracts collected ten months after inoculation showed the presence of ADFVd in most of the inoculated seedlings, thus proving the ability of this RNA to replicate independently. Material from these ADFVd-infected seedlings grafted onto the ASSVd indicator Starkrimson incited the appearance on the fruit skin of discolored spots more or less depressed which were particularly frequent around the calyx cavity. Apple cultivars such as Gala, Pink Lady, and Braeburn reacted to ADFVd with similar symptoms, occasionally accompanied by scar skin in Braeburn, whereas ADFVd replicated without eliciting any visible symptoms in other apple cultivars such as Golden, or in the pear indicator Fieud 37. No interference in symptom expression or viroid accumulation were observed in plants inoculated first with ASSVd and then with ADFVd.

Variability of the coat protein gene of Citrus psorosis virus in Campania, southern Italy.

Variability of the Coat protein (CP) gene of Citrus psorosis virus (CPsV) was assessed serologically, and by sequence analyses of two genomic regions located in the 3' (region C) and 5' (region V) halves of the gene. Analysis of 53 psorosis field sources from Campania, Italy, with 23 monoclonal antibodies revealed nine serogroups and at least ten different epitopes. Sequence analysis of 19 of these sources showed limited nucleotide diversity of the CP gene in the population. Diversity was slightly higher in region V than in region C. Phylogenetic analysis of the V and C regions of the CP showed that the Campania sources of CPsV were clearly separated from the CPsV-4 isolate from Florida. For C region, most of the CPsV sources clustered together, whereas two clusters were observed for region V. The ratio between nonsynonymous and synonymous substitutions for regions C (0.083) and V (0.345) indicated negative selective pressure for amino acid changes, more intense in the C region. No correlation was found between serogroups and specific aminoacid sequences, field location or citrus cultivar.