First report of Botrytis cinerea causing gray mould on Neomarica longifolia
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Gray mould of Neomarica longifolia (yellow iris) is reported here for the first time in Brazil and worldwide. The identity of the fungus was investigated and both morphology and molecular information demonstrated it to belong to Botrytis cinerea. Koch’s postulates were performed demonstrating that B. cinerea was the etiological agent of the disease.
KeywordsEtiology Ornamental Taxonomy Yellow iris
In July 2017, plants of N. longifolia kept in a greenhouse in the campus of the Universidade Federal de Viçosa (Viçosa, state of Minas Gerais, Brazil) for later use in an experiment, showed gray mould symptoms on flowers and flower buds (Fig. 1b).
Samples of diseased flower parts were taken to the laboratory for analysis. A dried sample was deposited in the herbarium of the Universidade Federal de Viçosa, as VIC 44317. A fungus was found in association with all diseased organs. A pure culture of this fungus was obtained by direct transfer of conidia from colonies on flowers onto PDA plates with the help of a sterile fine pointed needle. One representative culture was deposited in the local culture collection (Coleção Octávio de Almeida Drumond) - Acc. N° COAD 2291. Fungal structures were scraped from the infected flowers with a scalpel and mounted on lactofucsin and lactoglycerol and observed under a light microscope (Olympus BX 53) equipped with a Motic (Moticam 5) digital camera.
The fungus had the following morphology (Fig. 1c–e): Conidiophores subcylindrical, 300˗2000 × 12.5–22.5 μm, branched, smooth, light brown, pale near the apex; Conidiogenous cells ampulliform, 10–20 × 7.5–12.5 μm; Conidia ellipsoid or subespheric, 6–12 × 6–10 μm, with slightly protruding hilum, smooth, light brown or subhyaline. This morphology was readily recognized as typical for Botrytis cinerea as described in Ellis and Waller (1974).
DNA extraction was performed with the Wizard Genomic DNA Purification Kit, by following the manufacturer’s instuctions. Primers ITS4 and ITS5 (White et al. 1990) were used to amplify the ITS region of the 5.8S rRNA gene. Sequencing was performed by Macrogen Korea (www.macrogen.com). Sequences were edited and deposited in GenBank (Accession N° MH175197). A BLASTn search of sequences in GenBank indicated that COAD 2291 had 100% identity with the 199Jb14 isolate of Botrytis cinerea (Accession N° KU516438) as well as to many other B. cinerea isolates.
A conidial suspension of COAD 2291 was prepared by flooding a 7 days-old colony on PDA with 10 mL of sterile distilled water and scraping the surface of the colony with a rubber spatula. The concentration of the suspension was adjusted to 106 conidia/ml with the help of a haemocytometer prior to use. Two healthy N.longifolia individuals grown in 1 L plastic pots had their flower buds brush-inoculated with the conidial suspension. Two healthy plants treated with tap water alone served as controls. After inoculation, the plants were left in a dew chamber for 48 hs. After this period, the plants were transferred to a greenhouse bench and observed daily. Typical symptoms of the disease appeared two days after inoculation only on inoculated individuals.
Botrytis cinerea is a polyphagous pathogen attacking more than 230 different plant species (Horst 1998). Its wide range of hosts, its ability to survive as a saprophyte and the production of resistance structures make management of this pathogen difficult (Jarvis 1977). Little is known in terms of pathogenic fungi occurring on the entire genus Neomarica – even for the far more broadly cultivated N. caerulea. Only three records of fungi on Neomarica are listed in Farr and Rossman (2018) and only one includes an identification of the fungus at the species level. The specie is Cercospora neomaricae on N. caerulea (Macedo and Barreto 2008). The sole previous mention in the literature of a fungal pathogen of N. longifolia is also of a polyphagous pathogen – S. rolfsii (Inácio et al. 2017), but it is likely that further investigations will reveal more specialized pathogenic fungi on this host.
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