Phytopathologia Mediterranea

From 2016, witches’ broom and stunting symptoms were observed in Lolium rigidum grown in some fruit tree nurseries in Faragheh (Abarkouh, Yazd province, Iran). Total DNAs were extracted from symptomatic and asymptomatic plants and assessed for phytoplasma presence using direct and nested PCR to detect the 16S ribosomal RNA gene. From all symptomatic L. rigidum plant samples, expected length PCR amplicons were obtained. RFLP analysis with informative restriction enzymes showed identical profiles in all the samples resulted positive, that were also consistent with those of one of the subgroups of the aster yellows phytoplasmas (16SrI). The 16S rRNA gene sequence of Faragheh L. rigidum bushy stunt strain was 100% identical to some ‘Candidatus Phytoplasma asteris’ related strains, and 99.12% similar to the reference ‘Ca. P. asteris’ strain. The virtual RFLP pattern was identical (similarity coefficient 1.00) to the pattern of phytoplasmas in subgroup 16SrI-F. This is the first report of occurrence and molecular identification of this phytoplasma strain in L. rigidum and indicates a potential phytoplasma reservoir for trees in fruit tree nurseries where insect vectors may be present. This phytoplasma strain has been reported in symptomatic stone fruits in Spain and in potato in Ecuador. Further research on the epidemiology of witches’ broom and stunting in L. rigidum is required to develop elimination the phytoplasma from areas surrounding agricultural crops and avoid the risks of epidemics.

A new disease syndrome of zucchini squash was observed in Southern Italy, in 2018 and again in 2020. Affected plants were severely stunted and leaves were bent downwards, small, stiff, thick, leathery, and had interveinal chloroses. In addition, flowers were virescent and fruits were deformed and often cracked. Disease incidence was 20 and 30% in two different zucchini cultivations in Campania region (Southern Italy). Tomato yellow leaf curl New Delhi virus (ToLCNDV) was detected in eight samples, by loop-mediated isothermal amplification–based (LAMP) kit and by PCR and Sanger sequencing of the AV1 gene. Phytoplasmas were detected in the same samples using nested PCR assays with primer pairs P1/P7 and R16F2n/R16R2. Phytoplasma associations in plant samples were confirmed using specific primers for the multilocus genes SecY, tuf and rp. Sequence comparison of multilocus genes and phylogenetic analyses of the 16S rDNA gene confirmed the association of a phytoplasma strain closely related to ‘Candidiatus Phytoplamsa asteris’. This is the first report of mixed infections of ToLCNDV and a putative ‘Ca. Phytoplamsa asteris’ strain in zucchini, associated with a new Squash-Phytoplasma-Begomovirus (SqPB) disease syndrome.

In Europe and the Mediterranean region, ‘Candidatus Liberibacter solanacearum’ (Lso) is associated with emerging diseases of Apiaceae crops, mainly carrot. Emergency measures for import of carrot seed were set, requiring seed to be heat-treated at 50°C or tested as Lso-negative by PCR. The germination response to heat treatment was assessed for 24 carrot cultivar and hybrid seed lots. Ten parsley, five fennel, and two celery seed lots were also analysed. Of these 41 seed lots, 21 were Lso-infected. Water heat treatment significantly decreased germinability compared to dry heat treatment, indicating that dry heat treatment is a cheaper and less detrimental procedure. However, the dry heat treatment significantly decreased seed germination compared to untreated controls in four of 24 seed lots of carrot, four of ten parsley seed lots, three of five fennel seed lots, and one of two celery seed lots. For parsley, the heat treatment reduced germinability to a lesser extent in Lso-infected than Lso-free seed lots. These data show that heat treatment can affect the germination of Apiaceae seeds to varying degrees, depending on species or variety, the type of heat treatment, and the sanitary status of the seeds.

Pumpkin plants showing symptoms of witches’ broom (PuWB) were observed in Xinjiang Uyghur autonomous region, China, in September 2018. A phytoplasma was detected in symptomatic plants by PCR amplifying portions of the 16S ribosomal and tuf genes. In addition, the phylogeny based on these genes sequencing indicated that the PuWB strain clusters with ‘Candidatus Phytoplasma solani’ (subgroup 16SrXII-A). Furthermore, based on in silico and in vitro restriction fragment length polymorphism analyses, the PuWB phytoplasma was confirmed as a ‘Ca. P. solani’-related strain. This was the first record of the occurrence of phytoplasma presence in pumpkins in China, and the first record of 16SrXII phytoplasma infecting pumpkins in the world.