The sequencing of the complete genome of a Tomato black ring virus (TBRV) and of the RNA2 of three Grapevine chrome mosaic virus (GCMV) isolates from grapevine reveals the possible recombinant origin of GCMV.

The complete genome of a Tomato black ring virus isolate (TBRV-Mirs) (RNA1, 7,366 nt and RNA2, 4,640 nt) and the RNA2 sequences (4,437; 4,445; and 4,442 nts) of three Grapevine chrome mosaic virus isolates (GCMV-H6, -H15, and -H27) were determined. All RNAs contained a single open reading frame encoding polyproteins of 254 kDa (p1) and 149 kDa (p2) for TBRV-Mirs RNA1 and RNA2, respectively, and 146 kDa for GCMV RNA2. p1 of TBRV-Mirs showed the highest identity with TBRV-MJ (94 %), Beet ringspot virus (BRSV, 82 %), and Grapevine Anatolian ringspot virus (GARSV, 66 %), while p2 showed the highest identity with TBRV isolates MJ (89 %) and ED (85 %), followed by BRSV (65 %), GCMV (58 %), and GARSV (57 %). The amino acid identity of RNA2 sequences of four GCMV isolates (three from this study and one from GenBank) ranged from 91 to 98 %, the homing protein being the most variable. The RDP3 program predicted putative intra-species recombination events for GCMV-H6 and recognized GCMV as a putative inter-species recombinant between GARSV and TBRV. In both cases, the recombination events were at the movement protein level.

First Report of Cylindrocarpon pauciseptatum Associated with Root Rot and Decline of Peach in Southern Italy (Apulia Region).

During a survey for the sanitary status of stone fruits in southern Italy (Apulia region), symptoms of low vigor, sparse foliage, and chlorosis of leaves, frequently leading to decline or death of the plants, were observed on 3- to 5-year-old peach trees (Prunus persica) cvs. Tardi Belle, Zee Lady, and O'Henry grafted on GF677. Brown-to-black discolorations of the wood were observed in cross-sections of the trunks just below the graft union. Samples were collected from May to June 2010 from two symptomatic orchards in Brindisi and Foggia provinces. Small pieces of brownish, vascular wood and necrotic root tissues were surface disinfested, placed onto potato dextrose agar (PDA), and incubated for 7 days at 25°C in the dark. Single-conidial isolates were subsequently grown on PDA at 25°C for 10 days. Fungal colonies were presumptively identified as members of the genus Cylindrocarpon on the basis of their morphological and conidial characteristics. On PDA, the isolates developed abundant mycelium, which gradually became yellowish or partially brownish. Macroconidia were predominantly three septate, straight and cylindrical with both ends broadly rounded. Chlamydospores and ovoidal microconidia were observed on synthetic nutrient-poor agar (1). Sequence of the ribosomal internal transcribed spacer (ITS) region was obtained using universal primers (ITS6-ITS4) and deposited in GenBank (Accession No. HE577846). This sequence revealed 100% genetic identity with a sequence from Cylindrocarpon pauciseptatum Schroers & Crous (Accession No. EF607090), a recently described species (3). In nature, several species of the genus Cylindrocarpon affect a large number of woody plants, mainly grapevine, olive, and stone fruits, in which they attack the root surface (2). To verify Koch's postulates, the roots of 20 3-month-old peach seedlings (GF305) were dipped for 30 min in a spore suspension of the fungus (1 × 108 conidia ml-1). Seedlings were then transplanted in an artificial soil mix and held under controlled conditions in a greenhouse at 24°C. Typical black-foot symptoms developed on 92% of the inoculated plants within 3 months, whereas the control plants, whose roots had been dipped in distilled water, remained healthy. C. pauciseptatum was reisolated from infected tissues and internal vascular lesions of 45% of the inoculated plants, but none of the plants used as controls, fulfilling Koch's postulates. To our knowledge, this is the first report of this pathogen on peach in the Apulia Region of Italy. Currently, C. pauciseptatum is limited to a few orchards where presumably it was introduced with infected propagating material from extra-regional nurseries. C. pauciseptatum has the potential to negatively affect the stone fruit industry in Italy including reducing nursery production and productivity and vigor of trees in orchards, or even rapid death of young trees. References: (1) W. Gams et al. CBS Course of Mycology. 4th ed. Centraalbureau voor Schimmelcultures, Baarn, the Netherlands, 1998. (2) M. E. S. Hernandez et al. Eur. J. Plant Pathol. 104:347, 1998. (3) H. J. Schroers et al. Mycol. Res. 112:82, 2008.

Transmission of Grapevine Leafroll Viruses by Planococcus ficus (Hemiptera: Pseudococcidae) and Ceroplastes rusci (Hemiptera: Coccidae).

Grapevine leafroll associated virus-3 (GLRaV-3) and Grapevine leafroll associated virus-5 (GLRaV-5), two members of the genus Ampelovirus associated with grapevine leafroll disease, were transmitted by the mealybug Planococcus ficus and the soft scale insect Ceroplastes rusci from infected to healthy vines under experimental conditions. The efficiencies of transmission of GLRaV-3 and GLRaV-5 by P. ficus were 23.3 and 8.3%, respectively, and by C. rusci were 3.3 and 1.7%, respectively. Juvenile instars of P. ficus were more efficient in transmission of the viruses than adult females. This is the first report of the ability of C. rusci to transmit these viruses to grapevines.

Grapevine fleck virus-like viruses in Vitis.

Two sets of degenerate primers for the specific amplification of 572-575 nt and 386 nt segments of the methyltransferase and RNA- dependent RNA polymerase cistrons of members of the genera Tymovirus and Marafivirus and of the unassigned virus Grapevine fleck virus (GFkV) were designed on the basis of available sequences. These primers were used for amplifying and subsequent cloning and sequencing part of the open reading frame 1 of the genome of GFkV, Grapevine asteroid mosaic-associated virus (GAMaV) and of another previously unreported virus, for which the name Grapevine red globe virus (GRGV) is proposed. Computer-assisted analysis of the amplified genome portions showed that the three grapevine viruses are phylogenetically related with one another and with sequenced tymoviruses and marafiviruses. The relationships with tymoviruses was confirmed by the type of ultrastructural modifications induced in the host cells. RdRp-specific degenerate primers were successfully used for the aspecific detection of the three viruses in crude grapevine sap extracts. Specific virus identification was obtained with RT-PCR using antisense virus-specific primers.