Identifation And Pathogenicity Test Of One-celled Spore Isolated From Diseased Leaves Of Marssonina Coronaria

Marssonina leaf blotch of apple caused by Marssonina coronaria (Ell.& Davis) Davis leads to severe premature leaf fall since early summer, and occurres in all apple-growing region in China. In recent years, in the process of disease survey, specimen collection and pathogen isolation, we found that most of the fructification formed on the lesion of the diseased leaves in autumn were full filled with one-celled spore, two-celled conidia of this fungus were rare. Other scholars also found this phenomenon. However, the relationship between one-celled spore and two-celled condia, the status of one-celled spore in the life history of this fungus and its pathogenicity are not clear. In this study, we got the cultures of one-celled spore isolated from diseased apple leaves caused by M.coronaria by using single spore isolation; identified the one-celled spore through morphology observation and molecular genetic analysis; did its germination and pathogenicity test; and monitored the occurrence of one-celled spore stage in the orchard. The main results are as follows::1. One-celled spore were successfully isolated from disease apple leaves caused by M. coronaria. The means of singlespore isolation and media of PDA, PCDA, 10%V8 were recommend for isolation of one-celled spore among three means of isolation and four kinds of media used in this study.2. New propagules produced by the culture got from one-celled spore were different depending on the condtion of incubation. Cultures incubated on PCDA at 25℃in the dark for 14 days, or incubated at 10℃in the dark and 24℃under light (14 h dark/ 10 h light) for 14 days, then put the culture at 25℃in the dark for another 14 days, it produced two-celled condia; while cultures incubated on PCDA at 25℃in the dark for 14 days, then put the culture at 10℃in the dark and 24℃under light (14 h dark/ 10 h light) for another 14 days, or incubated at 10℃in the dark and 24℃under light (14 h dark/ 10 h light) for 14 days, both one-celled spore and two-celled conidia were observed. Additionally, cultures got from two-celled condia could product the same propagules when incubated under the same condition.3. Although one-celled spore and two-celled conidia shows significant difference on size and morphology, the symptoms of diseased leaves with either one-celled spore or conidia, the morphology of their acervuli and colonies, and their ITS sequence were very similar, which indicated that both one-celled spore and two-celled conidium belongs to M. coronaria.4. The germination test for one-celled spore showed that the spore could germinate on PDA, PCDA and 10%V8 at 25℃in the dark for 24-96h, but the germination rate was less than 1%; light was not necessary for the spore germination; the germinated spores was observed on PCDA which incubated at 25℃and 20℃in the dark for 24-96h, but not at 30℃. The germinated one-celled spore was not observed on both side of excised apple leaves after inoculation with or without injury within 5 days, and the symptom was not observed either.5. The survey of Marssonina leaf blotch of apple and monitoring for the occurrence of one-celled spore stage in the orchard indicated that the disease occurred in all apple producting regions in shannxi province; Fuji was more susceptible than Qinguan and Gala; apple trees aged 15-20 years were more susceptible than which below 15 years old; the The fructification formed on the diseased apple leaves caused by M. coronaria in July were all acervuli full filled with two-celled conidia, while in September, most fructification produced one-cell spore, and two-celled conidia were rarely observed.