Report Of Brown Rot Research Articles

First Report of Brown Rot Caused by Porogramme epimiltina on Gastrodia elata in China.

Gastrodia elata Blume is a valuable medicinal plant in China with great significance in medicine (Li et al. 2023). From 2022 to 2023, G. elata tuber rot occurred in about 50 households in the main cultivation areas of G. elata (27°39' N, 104°16' E) in Yiliang County, Zhaotong, Yunnan Province, southwest China. The planting area of G. elata was 776 ha, and the incidence rate was 10%. Symptoms present as light brown lesions on the surface of the tuber, sunken, soft and foul-smelling. Infected G. elata tubers were randomly collected from each household, packed into transparent plastic bags, and strains were isolated in the laboratory as follows. The tubers of 15 infected G. elata tubers were surface-sterilized with 0.5% NaOCl for 2 min, rinsed five times with sterile water, and dried. Symptomatic tissues from the margin between necrotic and healthy tissues were cut into 5 × 5 mm pieces, placed onto potato dextrose agar (PDA), and incubated at 28 ºC in the dark for 3 days. Hyphal tips of fungi growing from the samples were transferred onto new PDA plates and incubated until they produced conidia. Two fungal strains (Charliezhao 425 and 433) with the same morphological characteristics were obtained from the samples. Colonies were whitish and grew rapidly, irregularly turning pale orange at the edge or center of the mycelium pad on a two-week-old petri dish, and finally dark red，spore oval to spherical, 2.7 to 5.3 × 2.3 to 3.5 μm (n=50). The morphological characteristics of the isolates resembled Porogramme epimiltina (Mao et al. 2023; Kubayashi et al. 2001). Genomic DNA of two representative isolates (Charliezhao 425 and 433) was extracted using the DN14 cetyltrimethylammonium bromide rapid plant genome extraction kit (Aidlab Biotechnologies Co., Ltd, Beijing). The ITS and TEF1 genes were amplified by polymerase chain reaction using the primers ITS1/ITS4 (White et al. 1990) and EF1-983F/EF1-2218R (Rehner et al, 2005), respectively. All sequences were deposited in GenBank (accession no. OR905803, OR905804 for ITS, OR939812, OR939813 for TEF1). A BLASTN homology search with the ITS nucleotide sequences showed that they had 98.99 to 99.15% identity with P. epimiltina isolate OP997539 (588/594 bp) and isolate OP997539 (584/589 bp), respectively; and the TEF1 sequences had 95.41 to 95.59% % identity to isolates OP556566 (540/565 bp) and isolate OP556566 (542/567 bp), respectively. To complete Koch's hypothesis, the surfaces of 5 mature and healthy G. elata tubers were disinfected with 1% NaClO solution for 1 minute, rinsed with sterile water 5 times, and dried at 25 ℃ for 30 minutes. Conidial suspensions (106 spores/ml) were collected from two isolates (Charliezhao 425 and 433) and sprayed on G. elata tuber, and the control treated with distilled water. All G. elata tubers were incubated at 25℃ with 80% relative humidity. The experiment had three replicates. After 7 days of culture, there were obvious rotten and smelly on the inoculated tubers. No symptoms were observed in the control groups. The pathogen was re-isolated from all inoculated birch tubers and confirmed as P. epimiltina by morphological and molecular analysis, which fulfilled Koch's hypothesis. Gastrodia elata is a valuable and extensively used herbal Traditional Chinese Medicine with a wide range of clinical applications. As far as we know, this is the first report of P. epimiltina causing brown rot of G. elata in China.

First Report of Brown Rot Caused by Monilia yunnanensis on Sweet Cherry in China.

Chinese cherry industry has developed rapidly over the past few years, with the planting acreage continuously expanding, from Shandong province to Liaoning, Shaanxi, Hebei, Sichuan etc. Monilia spp. are the most important causal agents of brown rot of cherry, to date, M. fructicola, M. mumecola, and M. fructigena were reported to cause brown rot of cherry in China (Chen et al. 2013; Yin et al. 2014; Liu et al. 2012). In May 2023, fruit of sweet cherry cultivar 'Hongdeng' (Prunus avium L.) with symptoms resembling brown rot were collected from Tongchuan City, Shaanxi Province. Conidia on diseased tissues were spread on a water agar (WA, 1.5% agar and distilled water) medium and isolated with a glass needle under a professional single spore separation microscope (Wuhan Heipu Science and Technology Ltd., Wuhan, China). If no conidia were present, fruit pieces (5 × 5 mm) at the intersection of healthy and diseased tissues were surface sterilized with a sodium hypochlorite solution (1%) for 30 s and washed three times in sterilized water, followed by 75% ethanol for 30 s, then washed three times in sterilized water. After the tissue pieces were dried, they were placed on potato dextrose agar (PDA; 200 g of potato, 20 g of dextrose, and agar at 20 g/L) and incubated at 22 °C for about twenty days to produce spores and then single spore isolation was carried out. Thirty single-spore isolates were obtained and all were morphologically similar. The isolates produced white-gray colonies with even margins and concentric rings of sporogenous mycelium after 3 days incubation, and abundant black-colored stromata on the PDA medium after 15 days of incubation at 22°C. Conidia were one-celled, hyaline, ellipsoid to lemon shape (14.12 × 10.37 μm), with 1-2 germs which is similar to M. yunnanensis on peach. The genomic DNA of the isolates was extracted as described previously (Chi et al. 2009). The pathogen identity was confirmed by multiplex PCR which resulted in a 237bp amplicon, which is diagnostic of M. yunnanensis (Hu et al. 2011). Further sequencing of the internal transcribed spacer (ITS) region 1 and 2 and 5.8S gene (accession number: OR192774) indicated 100% identity with that of M. yunnanensis isolates (accession numbers: MW355895, ON024742). The average daily growth of mycelium on PDA at 22°C was 11.44 mm. Koch's postulates were fulfilled by inoculating 20 mature sweet cherry fruits of cv. 'Van' with mycelial plugs in a drilled hole. After 3 days of incubation at 22℃ in an airtight plastic tray with wet paper, the inoculated fruit developed typical brown rot symptoms. The developing spores on inoculated fruit were confirmed to be M. yunnanensis based on ITS sequence. All control fruit inoculated with a PDA plug remained healthy. M. yunnanensis was first reported as the causal agent of brown rot of peach in China (Hu et al. 2011). Later studies demonstrated that it is also pathogen on other fruits, e.g. hawthorn (Zhao et al. 2013), plum (Yin et al. 2015), apricot (Yin et al. 2017), apple, and pear in China (Zhu et al. 2016). To our knowledge, this is the first report of cherry brown fruit rot caused by M. yunnanensis, indicating the high risk of this species to cherry production, and effective strategies must be taken to prevent the possible control failure in practice.

First Report of Brown Rot Caused by Monilinia polystroma on Sweet Cherry and Almond in Italy.

Brown rot decay is an important disease of pome and stone fruits. In Italy, the main pathogens on stone fruits are Monilinia laxa, M. fructigena, and M. fructicola (Spitaler et al. 2022a). In addition, Monilinia polystroma (G. Leeuwen) L. M. Kohn (van Leeuwen et al. 2002), was recently found in Italy on peach (Martini et al. 2014), pear (Martini et al. 2015), plum (Abate et al. 2018), apple (Rosati et al. 2021), and quince (Spitaler et al. 2022b). In South Tyrol province, sweet cherry (Prunus avium L.) and almond (Prunus dulcis Mill. D. A. Webb), plants of the Rosaceae family and belonging to stone fruits, were observed to be frequently affected by brown rot. Affected cherries as well as almonds showed brown lesions, covered by yellowish or buff-colored stroma in concentric rings. Symptomatic cherries became shriveled, while symptomatic almonds remained firm. To determine the pathogen, single spore isolates were obtained from five symptomatic fruits, each from a cherry orchard of the cultivar Kordia in July 2021 and almond trees of the variety Dulcis in August 2021. Both sample sites were situated in Vadena/Pfatten. Infestation in the cherry orchard, covered by a rain-protection foil, was determined to be about 1 %. In almond, over 50 % of the fruits of various ripening stages showed brown rot symptoms. On potato dextrose agar (PDA) at 22 °C and a photoperiod of 16 h, isolates from both fruits matched the morphological characteristics of M. polystroma (Vasić et al. 2016) within 14 days. DNA was extracted from mycelium and the rRNA encoding gene region using ITS4 and ITS6 primers as well as a genomic sequence of unknown function using the primers UniMon_Forw and UniMon_Rev (Petróczy et al. 2012), were amplified and sequenced. MegaBLAST analysis revealed 100 % identity with M. polystroma sequences of the NCBI GenBank (rRNA encoding region: NR_154198; genomic region: JN128836). Sequences were deposited in GenBank under the accession numbers OP642545/OP654171 (cherry) and OP642546/OP654172 (almond). Pathogenicity was confirmed with mature cherries cultivar Duroncino or almost mature almond fruit of the variety Dulcis, respectively: 16 samples each for both fruits were surface-sterilized by dipping in 75 % ethanol for 10 s and subsequent rinsing with sterile water for 10 s. Mycelial plugs (1 mm) were dislodged from a 7-day old colony and inserted in a 1 mm hole into the fruits. Incubation was performed in plastic boxes under the conditions described above. PDA-inoculated fruit were used as controls. All cherries and all almonds were completely covered by brown rot lesions 7 days and 15 days post inoculation, respectively. Control fruits remained symptomless. Conidia were produced in branched chains on mycelium-inoculated fruit. Conidia were one-celled, limoniform, hyaline, measuring 13.1 to 22.2 × 9.7 to 14.8 µm (cherry) and 14.1 to 20.8 × 10.7 to 15.3 µm (almond). Additionally, 16 fruits each were inoculated with 20 µL conidial suspension (5 x 10^3 spores/mL) from mycelium-inoculated fruits. All cherries as well as all almonds were completely covered by brown rot lesions 7 days and 15 days post inoculation. Control fruits remained symptomless. To confirm identity, the fungus was isolated from five spore-inoculated fruits each for cherry and almond. The isolates showed identical morphological features and sequence identity as the original isolates. To our knowledge, this is also the first report of M. polystroma on almond, while the pathogen has already been reported on sweet cherry in Poland (Poniatowska et al. 2016). These additional host plants identified in this study suggest a broad impact of M. polystroma on Italian stone fruit production. Due to the economically important cultivation of stone fruit, further knowledge about the pathogens' host range will help to assign brown rot symptoms to M. polystroma and to improve targeted control strategies.

First report of brown rot caused by Monilia mumecola on Chinese plum (Prunus salicina) in Fujian, China.

Brown rot caused by Monilinia spp. (anamorph Monilia) is a common disease in stone fruits worldwide, but the species in different hosts or regions may vary. Monilia mumecola is a recently identified species, only reported in some regions. Although the pathogen has been found on plum in Yunnan, Hubei, and Zhejiang provinces, and Chongqing municipality in China (Yin et al. 2015), it has not been reported in southeast coast of China. In May 2022, brown rot with grey spores on fruit was observed in a plum orchard with 15% disease incidence in Sanming City, Fujian Province, located in southeast coast of China. Four single-spored isolates were obtained from germinating conidia on the water agar for further investigation. The colonies on potato dextrose agar (PDA) were initially white, gradually turned gray to brown, with lobbed margins and rare sporulation. Average mycelial growth rate ranged from 0.74 to 1.08 cm/day at 25 oC. Conidia were lemon-shaped or subglobose, hyaline, with an average size of 17.64 to 19.35×11.14 to 14.44 μm (n=30). Each isolate produced one to three or four germ tubes. Such characteristics are similar to M. mumecola (Yin et al. 2015). To confirm the identity of the isolates, genomic DNA was extracted and species-specific primers of Hu et al. (2011) were used to amplify the 712 bp sequence. In addition, the ITS region, partial genes of glyceraldehyde-3- phosphate dehydrogenase (GAPDH) and beta-tubulin (TUB2) were also amplified using primers sets ITS1/ITS4 (Glass and Donaldson 1995), Mon-G3pdhF/Mon-G3pdhR and Mon-TubF1/ Mon-TubR1, respectively (Hu et al. 2011). Sequences obtained for those three regions were 478, 762 and 1527 bp, respectively. Each region of all four isolates was identical, so one sequence for each region was submitted to GenBank with accession numbers OQ207672, OQ225251 and OQ225252, respectively, which had 100% identity with M. mumecola HQ908786 (ITS), HQ908784 (GAPDH), and HQ908775 (TUB2) using BLAST analysis in NCBI database, respectively. Pathogenicity was conducted with mycelium plugs from the edge of 7-day-old colony on three mature 'Angeleno' plums fruit (Prunus salicina) creating nine inoculations with three wounds per fruit, and each wound was 5 mm in diameter and 2 mm in depth. The same amount fruit and wounds inoculated with PDA plugs without fungi were used as a control. Brown rot symptoms were observed on all inoculated plums 4 days post-inoculation under room temperature with 100% humidity, whereas control plums remained symptomless. Fungal colonies re-isolated from the lesions showed the same morphological features as the original isolate , thus fulfilling Koch's postulates. To our knowledge, this is the first report of M. mumecola on plum in Fujian Province of China. The findings in this studies have important management implications for local plum growers because more than one Monilia species have been reported, where only M. fructicola was present in this region.

First Report of Brown Rot Caused by Phytophthora nicotianae on Navel Orange Fruit in China.

Navel orange (Citrus sinensis Osbeck cv. Newhall) is widely planted in southern China. From September to November 2021, severe outbreaks of Phytophthora brown rot were observed on navel orange fruit in three local orchards in Ganzhou City (28.80N, 115.53E), Jiangxi Province, China, with a disease incidence of 25 to 35%. Symptomatic fruit was mostly observed 1-m from the ground. Initial symptoms on infected fruit were circular, pale-brown to brown, water-soaked, slightly sunken lesions, covered with sparse white mycelia-like growth. As the disease progressed, the lesions turned dark brown and enlarged on the fruit surface. Three to four infected fruits were randomly collected from each orchard, placed in transparent plastic bags and immediately brought back to the laboratory for isolations. Infected fruits were surface-disinfested with 70% ethanol for 60 sec, and rinsed three times with sterile water. Symptomatic tissues from the margin between necrotic and healthy tissues were cut into 5 mm × 5 mm pieces, placed onto potato dextrose agar and incubated at 28°C for 5 days. Nine isolates were obtained. Colonies of three isolates (JFRL 03-16, 03-18, 03-19) in 10-day-old 20% V8 juice agar consisted of abundant, white, cottony aerial mycelia. Hyphal swellings and coenocytic mycelium were observed. Sporangia were ovoid, ellipsoid to spherical, papillate, and ranged in size from 17.2 to 60.1 µm × 15.8 to 48.6 µm (x ̅=46.2 ± 5.5 × 32.4 ± 4.8 µm, n=50). Chlamydospores were spherical, suborbicular, and ranged from 17.8 to 45.9 µm diam (x ̅=30.5 ± 3.5 µm, n=50). Oospores were not observed in pure cultures. These morphological characteristics were consistent with those of P. nicotianae (LaMondia et al. 2014). Genomic DNA was extracted from a representative isolate, JFRL 03-18, using the NuClean Plant Genomic DNA kit (CWBIO, China). The internal transcribed spacer (ITS) region, ras-related protein ypt1 (YPT), β-tubulin (TUB) gene were amplified by Polymerase Chain Reaction using primers ITS1/ITS4 (White et al. 1990), Yph1F/Yph2R (Schena et al. 2008), and TUBUF2/TUBUR1 (Kroon et al. 2004), respectively. All sequences were deposited in GenBank (Accession Nos. ON231777 for ITS, ON246910 for YPT, ON246908 for TUB). BLASTN homology search for these nucleotide sequences showed 100% identical to the ITS (MH341621), YPT (MK058408), TUB (MH760160) sequences of P. nicotianae. Sequences of twelve Phytophthora species and Pythium ostracodes were downloaded from GenBank. The phylogenetic tree of combined ITS, YPT, TUB sequences showed that the isolate JFRL 03-18 clustered with P. nicotianae. To complete Koch's postulates, zoospore suspensions were prepared from the cultures grown on 10-day-old V8 juice agar of isolates (JFRL 03-16, 03-18, 03-19). Pathogenicity tests were performed on healthy and surface-disinfested navel orange fruit. Nine fruits were gently wounded with a needle, inoculated with 10 µl zoospore suspension (104 zoospores/ml) of three isolates separately, and three fruit treated with sterilized water as controls. All fruit were incubated at 25℃ with 80% relative humidity and the test was repeated three times. After 7 days of incubation, the fruit inoculated with P. nicotianae showed similar brown rot symptoms and the control fruit remained symptomless. The pathogen was re-isolated from all inoculated fruits and confirmed as P. nicotianae by morphological and molecular analysis. Phytophthora nicotianae was previously reported on Hamlin sweet orange (Citrus sinensis (L.) Osbeck) fruit causing Phytophthora brown rot in Florida (Graham and Timmer 1995; Hao et al. 2018). To our knowledge, this is the first report of P. nicotianae causing Phytophthora brown rot of navel orange fruit in China. Based on the severity of this disease, local growers should develop and implement integrated disease management strategies for control.

First Report of Brown Rot Caused by Monilinia polystroma on Apple in Italy.

Brown rot is a common apple disease in Italy, caused by Monilinia fructicola, M. laxa and M. fructigena (Martini et al. 2013). In September 2020, in a 'Jeromine' apple orchard under integrated pest management located in Scarnafigi (44°39'N, 7°33'E, north-western of Italy), fruits (8.6%) showing brown to blackish firm lesions (6.0 to 8.0 cm diameter) were observed. In some fruits, rots were covered by yellowish stromata. Two isolates (MPI1; MPI2) were obtained from two symptomatic apples and cultured on potato dextrose agar (PDA) for 7 days at 25°C in 12-h light/12-h dark regime. A white-to-greyish mycelium with slightly undulate margins and irregular, black stromata developed on PDA after 12 days incubation. Conidia, observed in branched monilioid chains, (Suppl. Fig. 1) were one-celled, globose, limoniform, hyaline, 38 to 58 μm (mean: 48) × 20 to 44 μm (mean: 33). Based on morphology, the isolates were tentatively identified as Monilinia polystroma (G.C.M. Leeuwen) Kohn. A polymerase chain reaction with primers ITS1 and ITS4 was performed on internal transcribed spacer (ITS) region 1 and 2 and 5.8S gene. The sequenced amplicons (435 bp - 445 bp; GenBank Accession No. MW600854; MW600855) showed 100% identity to the reference isolate of M. polystroma (HQ846944) and to other isolates from apples (AM937114; JX315717) and plum (GU067539). The ITS region of M. polystroma had five nucleotides to distinguish it from the closest species M. fructigena (Zhu et al. 2016; MH862738) (Suppl. Fig. 2). The pathogenicity of both isolates was tested on mature 'Jeromine' apples (10.1% total soluble solids). Three replicates of six apples per isolate were surface disinfected with 1% NaClO. A mycelial plug (5 mm) from colony grown on PDA was inserted using a cork borer into a hole (6 mm) in each fruit (Vasić et al. 2016). Apples inoculated with sterile PDA plugs were used as control. Fruits were placed at 22 ± 1 °C, 85% relative humidity and 12 h light/12 h dark regime. Lesion size was measured after 3, 6 and 9 days of incubation. All inoculated fruits developed typical brown rot symptoms 6 days after inoculation and yellowish stromata appeared on the surface; control fruit remained healthy (Suppl. Fig. 3). The virulence of both isolates was statistically similar (Suppl. Table 1). M. polystroma was reisolated from all inoculated fruits and confirmed by molecular methods. This is the first report of M. polystroma on apple in Italy. M. polystroma was previously reported on apple in Hungary (Petróczy et al. 2009), on apricot in Switzerland (Hilber-Bodmer et al. 2012), on peach and pear in Italy (Martini et al. 2014; 2015), on plum in China (Zhu et al. 2016), and on apple in Serbia (Vasić et al. 2018). The emergence of this pathogen for pome and stone fruit production in Europe stimulates to study its biology and epidemiology, and its fitness and management, as compared to the other endemic Monilinia species.

First Report of Brown Rot on Peach, Nectarine, Cherry, and Plum Fruits Caused by Monilinia fructicola in Bulgaria

First Report of Brown Rot on Peach, Nectarine, Cherry, and Plum Fruits Caused by <i>Monilinia fructicola</i> in Bulgaria

First Report of Brown Rot Caused by Monilia mumecola on Chaenomeles lagenaria in China

Chaenomeles lagenaria (Loisel) koidz (Rosaceae) is a traditional medicinal plant widespread in China known for its antitumor, heart-strengthening, and antiphlogistic properties (Zhang et al. 2014). Recently, symptoms of brown rot on C. lagenaria have been observed causing 60% yield losses in Changyang county, Hubei province, China. The pathogen mainly infected flowers and fruits. Infected flowers exhibited symptoms of small auburn spots on the tip of bracts and sepals at the beginning, developing along the main vein toward the base of the flowers, resulting in the decay of pedicel. Infected fruits developed round, dark-brown spots with tawny liquid on the surface and progressively rotted and withered. As the disease progressed, a white mycelium occurred on infected tissue in the presence of increased humidity. Symptomatic fruits and flowers were cut into small pieces and disinfected with 1% NaOCl solution for 1 min, rinsed in sterile water, air dried, and placed on potato dextrose agar (PDA). Pure cultures were obtained by using the single-spore isolation method and incubating at 25°C in light. The radial mycelial growth was 7.5 mm/day; compact colony with wavy edges was formed on PDA after 12 days of growth on a 9-cm Petri dish. The mycelium was colorless at first and turned incanus with time. Hyphae were septate and branched. Strings of spherical, colorless conidia with a size of 4 to 22 × 6 to 10 μm developed on short conidiophores. Based on morphological characteristics of mycelia and conidia, isolates were identified as Monilia mumecola (Wei 1979). The internal transcribed spacer (ITS) region of rDNA was amplified using ITS1 and ITS4 primers and sequenced. BLAST analysis of the PCR product showed 99% identity to M. mumecola (AB125620, AB125613). The sequence of isolate was shown in the supplementary file. To verify the pathogenicity, fresh disinfected fruits of C. lagenaria were used for the inoculation test. A group of 10 healthy fruits were inoculated with mycelium plugs; meanwhile, another group of 10 fruits were inoculated with conidial suspension at a concentration of 1 × 10⁶ per milliliter. Ten more noninoculated fruits served as controls. All inoculated and noninoculated fruits were placed in a light cultivation room at 25°C with saturated humidity. Five days later, inoculated fruits in both groups began to brown. By 10 days after inoculation, 100% of the inoculated fruits showed symptoms identical to those observed in the orchard. Control fruits remained symptomless. M. mumecola was reisolated from the inoculated fruits. The experiment was repeated twice. M. mumecola has only been reported to cause brown rot on Prunus mume in Japan (Harada et al. 2004) and peach in China (Hu et al. 2011). To our knowledge, this is the first report of brown rot caused by M. mumecola on C. lagenaria in China.

First report of brown rot caused by Rhizopus arrhizus on tomato in Pakistan

First report of brown rot caused by Rhizopus arrhizus on tomato in Pakistan

First report of brown rot caused by Monilinia fructicola (Winter) Honey on sweet cherry in Turkey

First report of brown rot caused by Monilinia fructicola (Winter) Honey on sweet cherry in Turkey

First Report of Brown Rot Caused by Monilinia fructicola on Apple in Brazil

First Report of Brown Rot Caused by <i>Monilinia fructicola</i> on Apple in Brazil

First Report of Brown Rot Caused by Monilinia fructicola on Nectarine Fruit in Montenegro

First Report of Brown Rot Caused by <i>Monilinia fructicola</i> on Nectarine Fruit in Montenegro

First Report of Brown Rot Caused by Cryptococcus pseudolongus on Fruiting Body of Shiitake (Lentinula edodes) in Korea

First Report of Brown Rot Caused by <i>Cryptococcus pseudolongus</i> on Fruiting Body of Shiitake (<i>Lentinula edodes</i>) in Korea

First Report of Brown Rot Caused by Monilinia fructicola on Chinese Sour Cherry (Cerasus pseudocerasus) in Southwestern China

First Report of Brown Rot Caused by <i>Monilinia fructicola</i> on Chinese Sour Cherry (<i>Cerasus pseudocerasus</i>) in Southwestern China

First Report of Brown Rot on Peach Caused by Monilia polystroma in Slovenia

First Report of Brown Rot on Peach Caused by <i>Monilia polystroma</i> in Slovenia

First Report of Brown Rot on Apple Fruit Caused by Monilinia fructicola in Pennsylvania

First Report of Brown Rot on Apple Fruit Caused by <i>Monilinia fructicola</i> in Pennsylvania

First Report of Brown Rot Caused by Monilia mumecola on Chinese Sour Cherry in Chongqing Municipality, China.

Cherry is widely planted in China, from Liaoning, Beijing, Hebei, Shandong, Zhejiang, Jiangsu, and Anhui provinces (eastern China), to Shaanxi, Sichuan, Chongqing, and Guizhou provinces (western China). The brown rot fungus Monilinia fructigena causes considerable production losses in cherry production in Liaoning Province (3). In May 2013, Chinese sour cherry (Prunus pseudocerasus) cv. Wupi displaying symptoms of brown rot was found in an orchard in Chongqing municipality. Diseased cherry fruit had a brown rot sporulating with grayish, conidial tufts. The fruit later succumbed to the soft rot or shivered and became a mummy. Single-spore isolations on PDA resulted in colonies with concentric rings of pigmented mycelium with lobbed margins. Conidia were broadly ellipsoid to subglobose, occasionally even globose, with an average size of 16 × 12.7 μm. Multiple germ tubes were produced from each conidium, a germination pattern unique to Monilia mumecola (1,2,4). The pathogen identity was confirmed by multiplex PCR as described by Hu et al. (2). The PCR resulted in a 712-bp amplicon, which is diagnostic of M. mumecola. Further sequencing of the internal transcribed spacer (ITS) region 1 and 2 and 5.8S gene further indicated 100% identity with that of M. mumecola isolates from China (Accession No. HQ908786) and from Japan (AB125613, AB125614, and AB125620). Koch's postulates were confirmed by inoculating mature cherry fruit with mycelia plugs. Inoculated fruit were placed in a sterilized moist chamber, and incubated at 22°C with 12 h light/dark cycle. Inoculated fruit developed typical brown rot symptoms only 2 days after inoculation, while the control fruit, inoculated with a sterile PDA plug, remained healthy. The pathogen isolated from inoculated symptomatic fruit was confirmed to be M. mumecola based on morphological characteristics and germination pattern. It should be noted that the conidia on inoculated fruit showed an average size of 20 × 15.3 μm, significantly bigger than that of from PDA, and most produced more than three germ tubes. The inoculation experiments were performed in triplicates. M. mumecola was first reported as the causal agent of brown rot of mume in Japan in 2004 (1). Later studies demonstrated that it is also pathogen on other stone fruits, e.g., peach, nectarine (2), and apricot (4). To our knowledge, this is the first report of cherry brown fruit rot caused by M. mumecola, and the first report of M. mumecola in Chongqing municipality.

First Report of Brown Rot Caused by Monilinia fructicola on Apple in Italy.

Monilinia fructicola (G. Wint.) Honey, the causal agent of brown rot, is one of the most important fungal pathogens of stone fruit but may also affect pome fruits. The pathogen is common in North America, Oceania, South America, and Asia. It is a quarantined pathogen in Europe (3), but was recently detected in apple from the Czech Republic, Germany, and Serbia (1,2,4). In January 2012, during a survey for fungal postharvest pathogens, stored apple (Malus domestica Borkh.) belonging to the cultivars Gala and Pink Lady showing brown rot symptoms were observed in the Emilia Romagna region, Italy. Typical decay spots were circular and brown, tending toward black. Decayed tissues remained firm, and numerous grayish pustules containing spores appeared on rotted areas. The pathogen was isolated on V8 juice agar and culture plates were incubated at 25°C in darkness for 5 days. A conidial suspension was spread on malt extract agar and single spores were selected. The colonies were morphologically identified as M. fructigena. Two colonies developing a gray mass of spores in concentric rings with the reverse side black were further studied by molecular tools. The colony margins were even and the conidia were one-celled, limoniform, hyaline, and 12.1 to 17.4 × 8.4 to 11.2 μm. 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. fructicola sequences (GenBank Accession Nos. HQ893748.1 and FJ515894.1). Pathogenicity was confirmed using surface-sterilized mature 'Gala' apples, wounded with a sterile needle, and inoculated with an isolate conidial suspension (103 spores/ml). A 20 μl droplet was placed in the wound; control fruits received sterile water without conidia. After 5 days of incubation at 20°C in plastic containers with high humidity, typical symptoms of brown rot developed on inoculated fruits, while control fruits remained symptomless. The fungus isolated from inoculated fruit exhibited the same morphological and molecular features shown by the original isolates. To our knowledge, this is the first report of the fungus M. fructicola on apple in Italy. Further studies are necessary to determine geographic distribution, prevalence and economic importance of this quarantine organism in Italy.

First Report of Brown Rot Caused by Monilinia fructicola on Nectarine in Serbia.

In August 2011, nectarine (Prunus persica (L.) Batsch var. nucipersica (Suckow) C. K. Schneid) fruit originated from Oplenac region with symptoms of fruit rot was collected at a green market in Belgrade. Fruit had large, brown, sunken lesions covered with grayish brown tufts. Symptoms resembled those caused by species of Monilinia including M. laxa, M. fructigena, or M. fructicola (2). In order to isolate the causal organism, small superficial fragments of pericarp were superficially disinfected with commercial bleach and placed on potato dextrose agar (PDA). The majority (32 out of 33) isolates formed rosetted non-sporulating colonies with lobed margins resembling those of M. laxa. However, one isolate (Npgm) produced an abundant, grayish-white colony with even margins and concentric rings of sporogenous mycelium, resembling those described for M. fructicola (2). Conidia were one-celled, hyaline, ellipsoid to lemon shaped, 7.38 to 14.76 × 4.92 to 9.84 μm, and borne in branched monilioid chains. The average daily growth on PDA at 24°C was 10.9 mm. A single-spore isolate of Npgm was identified as M. fructicola based on the morphology of colony and conidia, temperature requirements, and growth rate (2). Morphological identification was confirmed by an amplified product of 535 bp using genomic DNA extracted from the mycelium of pure culture and species-specific PCR for the detection of M. fructicola (2). The ribosomal internal transcribed spacer (ITS) region of rDNA of Npgm was amplified and sequenced using primers ITS1/ITS4. Sequence analysis of ITS region revealed 100% nucleotide identity between the isolate Npgm (GenBank Accession No. JX127303) and 17 isolates of M. fructicola from different parts of the world, including four from Europe (FJ411109, FJ411110, GU967379, JN176564). Pathogenicity of the isolate Npgm was confirmed by inoculating five surface-disinfected mature nectarine and five apple fruits by placing a mycelial plug under the wounded skin of the fruit. Nectarine and apple fruits inoculated with sterile PDA plugs served as a negative controls. After a 3-day incubation at 22°C, inoculated sites developed brown lesions and the pathogen was succesfully reisolated. There were no symptoms on the control nectarine or apple fruits. M. fructicola is commonly present in Asia, North and South America, New Zealand, and Australia, while in the EPPO Region the pathogen is listed as an A2 quarantine organism (3). In Europe, the first discovery of M. fructicola was reported in France and since then, it has been found in Hungary, Switzerland, the Czech Republic, Spain, Slovenia, Italy, Austria, Poland, Romania, Germany, and Slovakia (1). Most recently, M. fructicola was found on stored apple fruits in Serbia (4). To our knowledge, this is the first report of M. fructicola decaying peach fruit in Serbia. These findings suggest that the pathogen is spreading on its principal host plants and causing substantial economic losses in the Serbian fruit production.

First Report of Brown Rot Caused by Monilinia fructicola in Sweet Cherry in Maryland.

Monilinia fructicola (G. Wint.) Honey is the most important causal agent of brown rot of stone fruits in North America. In July 2010, 20 sweet cherry fruit (Prunus avium) of unknown variety with symptoms resembling brown rot were collected from one commercial orchard in Maryland. Each cherry fruit came from a different tree. Symptoms included necrotic areas up to 10 mm in diameter with brown conidia and conidiophores developing from the infection center. Spores from nine symptomatic fruit collected each from different trees of a single orchard were suspended in sterile water, spread onto the surface of 1% agar plates, and incubated at 22°C. After 12 h, single, germinated spores were transferred onto 9-cm petri dishes with potato dextrose agar (PDA). Nine fungal colonies, each from a different fruit, were investigated in three replicates for cultural characteristics on separate petri dishes containing PDA. They were very similar in morphology and grew 12.4 mm per day on average at 22°C, forming branched, monilioid chains of grayish colonies with concentric rings and little sporulation. Rich sporulation was observed on tomato sauce medium (250 ml tomato sauce and 20 g agar in 750 ml water). The lemon-shaped spores had an average size of 15 × 10 μm, which is consistent with M. fructicola. Two colonies were randomly selected to identify the pathogen to the species level using a PCR technique based on cytochrome b sequence amplifications (2). Resulting gel electrophoresis patterns were consistent with M. fructicola. Koch's postulates were fulfilled by inoculating 15 mature sweet cherry fruits of cv. Bing with a conidial suspension (105 spores/ml) of one of the single-spore isolates from cherry. Fruit were stab-inoculated at a point to a depth of 2 mm using a sterile needle. A 10-μl droplet was placed on each wound; control fruit received sterile water without conidia. After 3 days of incubation at room temperature in airtight plastic bags, the inoculated fruit developed typical brown rot symptoms with lesions that were 20.6 mm in diameter. The developing spores on inoculated fruit were confirmed to be M. fructicola. All control fruit remained healthy. The entire detached fruit experiment was repeated 1 week later. M. fructicola is assumed to be the main causal agent of brown rot of sweet cherry in the northeastern United States, but recent studies show that M. laxa is also causing the disease on sweet cherry in many northeastern states (1). For this reason, it is important to delineate species for accurate disease assessments. This study confirms assumptions that M. fructicola is a causal agent of sweet cherry in Maryland.