Phoma tracheiphila is the causal agent of a tracheomycotic disease of citrus called mal secco causing the dieback of twigs and branches. This pathogen is of quarantine concern; therefore, fast and reliable protocols are required to detect it promptly. A specific primer pair and a dual-labeled fluorogenic probe were used in a real-time polymerase chain reaction (PCR) with the Cepheid Smart Cycler II System (Transportable Device TD configuration) to detect this fungus in citrus samples. Real-time PCR assay was compared to modified conventional PCR assay. The sensitivity of the former was evaluated by testing P. tracheiphila DNA dilutions, and the minimum amount detectable was about 500 fg, whereas the linear quantification range was within 100 ng to 1 pg. Conventional PCR sensitivity was 10 pg. Conventional and real-time PCR successfully detected the fungus in woody samples of naturally infected lemon and artificially inoculated sour orange seedlings. Nevertheless, real-time PCR was about 10- to 20-fold more sensitive than conventional PCR, and preliminary results indicate that the former technique achieves quantitative monitoring of the fungus in tissues. Simple and rapid procedures to obtain suitable DNA samples from fungal cultures and citrus woody samples for PCR assays enable diagnosis to be completed in a short time.

Summary Cankers and dieback on London plane caused by Diaporthe scabra have been observed in urban plantings in Catania, Italy. Symptoms were delayed spring flush, trunk cankers, small chlorotic leaves associated with dead terminal or lateral branches and an early defoliation. Isolations from infected wood on PDA led to the isolation of the anamorph Phomopsis scabra with α‐conidia production. The infected wood placed in a moist chamber developed pycnidia with α and β conidia and perithecia of the teleomorph D. scabra. Sequence analysis of the internal transcribed spacers ITS1 and ITS2, the ribosomal intragenic spacer beta‐tubulin 1 and beta‐tubulin 2 genes, congruently indicated that the D. scabra isolate is phylogenetically related to two strains of D. helianthi isolated in Italy from sunflower and to strains of D. eres. Inoculations on young plants of London plane growing in pots confirmed the pathogenicity of the isolated fungus. To our knowledge, this is the first report of the disease in Italy.

In April 2006, a new leaf disease occurred in a private garden in eastern Sicily (Italy) on young, 2-year-old seedlings of Mexican blue palm, Brahea armata S. Watson, in the Arecaceae. Symptoms were detected on 80% of seedlings. The leaves had minute, brown spots that enlarged into dark brown, circular or elliptical lesions, 3 to 6 mm in diameter, and with a necrotic, gray center. The lesions sometimes were surrounded by a chlorotic halo, and older leaves had larger chlorotic areas between spots. Conidia, conidiophores, and terminal vesicles were examined from diseased leaves. A Cylindrocladium sp. was consistently isolated from leaf lesions on Oxoid (Basingstoke, Hampshire, England) potato-dextrose agar after surface disinfestations with 0.8% NaOCl. Cylindrocladium isolates were cultured on carnation leaf agar (CLA) using single hyphal tips. Five isolates were established and identified as Calonectria pauciramosa C.L. Schoch & Crous based on obpyriform to broadly ellipsoidal terminal vesicles, conidiophore branching pattern, conidia size (52 × 4.6 μm), perithecium morphology, and ascopores size (36 × 6.8 μm). Perithecia were obtained with C. pauciramosa tester strains from Italy (G87 and G128) and South Africa (U 971 and U 1670) (2,3) that confirmed both mating types to be present. Further confirmation was obtained by internal transcribed spacer (ITS) analysis. The sequence of rDNA ITS1-5.8 S-ITS2 regions, obtained after amplification with primer ITS1 and ITS4, revealed that the Brahea isolates showed total homology with the sequence of the C. pauciramosa (STE-U 971 from soil) (= Cylindrocladium pauciramosum) available in GenBank. Isolate CBS 120619 from Mexican blue palm was deposited at Centraalbureau voor Schimmelcultures. Spray inoculations of 10 2-year-old Mexican blue palm seedlings were performed with a spore suspension of the fungus adjusted to 105 conidia per ml obtained from 14-day-old single-spore colonies on CLA at 24°C under cool white fluorescent irradiation on a 12-h light/dark regimen. In addition, the following species were similarly inoculated using 10 1-year-old plants: Arecastrum romanzoffianum (Cham.) Becc., B. edulis H. Wendl. ex S. Watson, Chamaerops humilis L., Howea forsteriana Becc., Phoenix canariensis Hort. ex Chabaud., Trachycarpus fortunei (Hook.) H. Wendl., and Ravenea rivularis Jumelle & Perrier. Inoculated, and 10 control plants were placed in separate plastic bags in a growth chamber at 25 ± 1°C. After 7 to 10 days, foliar symptoms including flecks and spots developed on both species of Brahea and on Chamaerops humilis, and on these hosts, pathogenicity tests were repeated. Other palm species and control plants remained healthy. C. pauciramosa was consistently reisolated from inoculated plants on the basis of vesicle shape and conidia sizes of the anamorph. Cylindrocladium candelabrum, Cylindrocladium colhounii, Cylindrocladium floridanum, Cylindrocladium parasiticum, Cylindrocladium pteridis, Cylindrocladium scoparium, and Cylindrocladium theae have been reported as leaf spots pathogens of Arecaceae (1). To our knowledge, this is the first occurrence of C. pauciramosa on Mexican blue palm and the first report of the pathogen on Arecaceae. References: (1) P. W. Crous. Taxonomy and Pathology of Cylindrocladium (Calonectria) and Allied Genera. The American Phytopathological Society, St. Paul MN, 2002. (2) P. W. Crous et al. Stud. Mycol. 50:415, 2004. (3) G. Polizzi and P. W. Crous Eur. J. Plant Pathol. 105:407, 1999.

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