RESUMO
Monilinia spp. are well-known pathogens causing brown rot of fruit trees in many fruit production areas worldwide. In Italy, three Monilinia species are particularly significant with regard to fruit trees, causing blossom and twig blight and brown rot in fruits: Monilinia laxa (Aderhold and Ruhland) Honey, M. fructicola (Winter) Honey, and M. fructigena (Aderhold and Ruhland). In 2009, a new species, M. polystroma, was distinguished from M. fructigena based on morphological and molecular characteristics in Europe (3). M. polystroma is not known to occur in Italy and to date has been reported from the Czech Republic (1), Hungary (3), Poland (4), Serbia (5), and Switzerland (2). In July 2013, during a survey for fungal postharvest pathogens, stored peaches (Prunus persica (L.) Batsch) belonging to different cultivars showing brown rot symptoms were observed in the Emilia Romagna and Sardinia regions of Italy. Typical decay spots were circular and brown, tending toward black, and 5% of peaches presented a large number of yellowish or buff-colored stromata and firm decayed tissues, the symptoms originated by M. polystroma. The pathogen was isolated on V8 agar (V8A) and culture plates were incubated at 25°C in darkness for 5 days. A conidial suspension was spread on malt extract agar (MEA) and single spores were selected. M. polystroma colonies grown on potato dexstrose agar (PDA) were yellowish in color. Irregular black stromatal crusts occurred on the edges of the colonies after 10 to 12 days of incubation and on the margin was present sporogenous tissue slightly elevated above the colony surface, color buff/pale luteous (1). The conidia were one-celled, ovoid or limoniform, smooth and hyaline, and 12 to 20 × 8 to 12 µm in distilled water when grown on V8A at 22°C. The ribosomal ITS1-5.8S-ITS2 region was PCR-amplified from genomic DNA obtained from mycelium using primers ITS1 and ITS4. A BLAST search in GenBank revealed the highest similarity (99%) to M. polystroma sequences (GenBank Accession No. GU067539). Pathogenicity was confirmed using surface-sterilized mature 'Red Heaven' peaches. The fruits were wounded (2 × 2 × 2 mm) twice with a sterile needle and inoculated with 2-mm plugs of 7-day-old mycelia from fungal colony margins. The sample unit was represented by 10 fruits. Control fruits were inoculated with PDA. After 7 days of incubation at 20°C in plastic containers with high humidity, typical symptoms of brown rot developed on both the wounds of all inoculated fruits, while control fruits remained symptomless. By the 14th day, all fruits had rotted and the yellowish exogenous stromata appeared on the surface of infected peaches. The fungus isolated from inoculated fruit exhibited the same morphological and molecular features of the original isolates; the molecular analysis performed using the primers by Petroczy (3) confirmed the result of the PCR with ITS1 and ITS4 primers. To our knowledge, this is the first report of M. polystroma on peach in Italy. This is relevant because the new pathogen could spread into other European countries that are main peach producers (such as Spain), causing economic losses. Bringing it to the attention of the scientific community allows the arrangement of research studies for assessing potential resistances with a significant impact on disease control management. Further studies are necessary to determine geographic distribution, prevalence, and economic importance of this organism in Italy. References: (1) EPPO Reporting Service. 2011/134: First reports of Monilinia polystroma in Hungary and the Czech Republic. No. 6, 2011. (2) M. Hilber-Bodmer et al. Plant Dis. 96:146, 2012. (3) M. Petroczy and L. Palkovics. Eur. J. Plant Pathol. 125:343, 2009. (4) A. Poniatowska et al. Eur. J. Plant Pathol. 135:855, 2013. (5) M. Vasic et al. Plant Dis. 97:145, 2013.
RESUMO
Italy could be considered the main apple-producing country in the European Union. Italian apple (Malus domestica L. Borkh.) production is estimated at approximately 2.1 million tons and encompasses a wide range of cultivars, harvested from August to November. Colletotrichum acutatum, which causes severe losses to strawberry production, was a regulated organism for all European countries until 2008, when it was removed from the EPPO quarantine pathogen list because of its wide distribution in strawberry-production areas. During the growing season of 2010, fungi were isolated from apple fruits exhibiting bitter rot symptoms after 4 months of storage in several packinghouses in the Emilia Romagna Region. The apples belonged to the Golden Delicious, Granny Smith, Pink Lady and Crisp Pink cultivars. Lesions on the fruit surface were circular and 1 to 3 cm in diameter. When lesions enlarged, they became sunken with relatively firm rotten tissues. The fungal fruiting structures, acervuli, were distributed sparsely or densely on old lesions, and under humid conditions, they discharged an orange conidial mass. Conidia observed with a light microscope appeared hyaline and fusiform, 8 to 16 × 2.5 to 4 µm, with two pointed ends or one rounded end. The fungal isolates were grown on potato dextrose agar (PDA) plates incubated at 25°C. After 7 days, colonies were white, becoming gray to pale orange, and when viewed from the reverse side, the color ranged from pink to reddish orange. Both cultural and morphological characteristics of the pathogen were similar to those described for C. acutatum J.H. Simmonds (3), which is responsible for bitter rot of apple (2). Koch's postulates were performed with one representative isolate from each host by artificial inoculation of 30 healthy apples from the cultivars listed above. Fruit surfaces were disinfected with 70% ethanol, wounded with a sterile needle, and then inoculated with 20 µl of a spore suspension (105 conidia ml-1) prepared from a 15-day-old culture on PDA. Inoculated fruits were sealed in a plastic bag and incubated at 25°C for 10 days. In 92% of fruits, symptoms appeared 10 days later, forming lesions with cream-to-salmon pink fruiting structures. The fungus was reisolated onto PDA from the lesions on the inoculated apples. After 7 days of incubation, the colonies and the morphology of conidia were the same as those of the original isolates. The tests were performed on all four cultivars with similar results. The PCR analysis, carried out using universal primers ITS1 and ITS4 (4) directly from single-spore-derived mycelium (1), resulted in an amplification product with 100% sequence homology with C. acutatum isolate AB626881 from GenBank database. Considering the results obtained, to our knowledge, this is the first report of C. acutatum in Italy causing bitter rot on apple. The disease is common in practically all countries where apples are commercially grown and since the losses could be severe under prolonged warm and wet weather conditions, C. acutatum could represent a serious issue for the Italian apple industry. References: (1) M. Iotti and A. Zambonelli. Mycol. Res. 110:60, 2006. (2) A. L. Jones et al. Plant Dis. 80:1294, 1996. (3) B. C. Sutton. Page 1 in: Colletotrichum: Biology, Pathology and Control. Brit. Soc. Plant Pathol. Oxon. UK 1992. (4) T. J. White et al. Page 315 in: PCR Protocols. A Guide to Methods and Applications. Academic Press, San Diego, CA, 1990.