First report of 'Candidatus Phytoplasma asteris' associated with yellowing, scorching and decline of almond trees in India.

The almond, a commercially important tree nut crop worldwide, is native to the Mediterranean region. Stone fruit trees are affected by at least 14 'Candidatus Phytoplasma' species globally, among which 'Candidatus Phytoplasma asteris' is one of the most widespread phytoplasma infecting Prunus dulcis, causing aster yellows disease. Recently, almond plantations of Nauni region were consistently affected by phytoplasma, as evidenced by visible symptoms, fluorescent microscopic studies and molecular characterization. During several surveys from May to September 2020-2022, almond aster yellows phytoplasma disease showing symptoms such as chlorosis, inward rolling, reddening, scorching and decline with an incidence as high as 40%. Leaf samples were collected from symptomatic almond trees and the presence of phytoplasma was confirmed through fluorescent microscopic studies by employing DAPI (4, 6-diamino-2-phenylindole) that showed distinctive light blue flourescent phytoplasma bodies in phloem sieve tube elements. The presence of phytoplasma in symptomatic almond trees was further confirmed using nested PCR with specific primer pairs followed by amplification of 16S rDNA and 16S-23S rDNA intergenic spacer (IS) fragments. Sequencing and BLAST analysis of expected amplicon of the 16S rDNA gene confirmed that the almond phytoplasma in Himachal Pradesh was identical to the aster yellows group phytoplasma. Phylogenetic analysis of 16S rDNA almond phytoplasma also grouped 'Prunus dulcis' aster yellows phytoplasma within 16SrI-B subgroup showed 94% nucleotide identity with 'Prunus dulcis' phytoplasma PAEs3 and 'Prunus dulcis' phytoplasma PAE28 from Iran. This research presents the first host report of 'Candidatus Phytoplasma asteris' infecting almonds in India, expanding the knowledge of the diversity and distribution of phytoplasma strains affecting almond trees globally.

Malus pumila 'Spy 227' and Apple stem pitting virus: graft incompatibility and epinasty.

Apple stem pitting foveavirus (ASPV) is one of the most important and widespread virus infecting apples in the world. Of late, the virus has been found to be invariably associated with most of the apple plantations of Shimla district of Himachal Pradesh based on DAS-ELISA results. Bioassay of viruses in vegetatively propagated crops including apple is considered to be an essential component in indexing programmes for the production of virus free propagating material. Woody indicator Malus pumila 'Spy 227' was used for the detection of ASPV through double grafting method. Graft incompatibility and epinasty symptoms were observed on Malus pumila Spy 227 indicator plants. Further, molecular identification of the virus isolate was done by cloning and sequencing of the test isolate. Partial sequence analysis of the coat protein gene showed 89 % nucleotide identity in BLASTN analysis with ASPV isolate from China (Accession No. JF895517). This is the first record of ASPV producing Graft incompatibility on Spy 227 indicator plants.