First report of Alternaria alternata causing fruit rot on Fig (Ficus carica) in Pakistan

Fig (Ficus carica L.) is among the earliest and widely cultivated fruit trees in the world due to its easy adaptation to diverse climates (Solomons et al. 2006). In July 2020, a rot disease was observed on multiple orchards located in Faisalabad- a region of Punjab Province. The symptoms appeared as light brown, circular to oval, and water-soaked lesions (4-8 mm in diameter). In more advanced stages of the disease, the lesions enlarged in size and leading to rot of the entire fruit. Disease incidence on fruit across the fields ranged from 23 to 29%. To isolate the causal agent, segments (5 mm2) were excised from 15 symptomatic fruit, surface disinfested with 70% ethanol for 1 min, washed in three changes of sterilized water, air dried, transferred aseptically to plates containing potato dextrose agar (PDA), and incubated at 25°C for 7 days with a 12-h photoperiod. Nine single spore isolates with similar morphology were isolated from the infected tissues. The cultured isolates consistently yielded dark brown to black colonies on PDA. Conidia were in chains (average conidial dimension 20 to 28 × 8 to 10 μm), olivaceous to dark brown, with a short conical beak with both transversal (two to five) and longitudinal (one to three) septa. Conidiophores were short, septate, hyaline to olivaceous brown, either branched or unbranched, 20 to 52 μm long, and 1 to 3 μm wide. These cultural and morphological characteristics were consistent with the descriptions of Alternaria alternata (Simmons 2007). The genomic DNA from three isolates was extracted using a PrepMan Ultra kit according to the manufacturer’s protocol and amplified using universal primers ITS1/4 (White et al. 1990) and the endopolygalacturonase gene using primers PG3/PG2b (Andrew et al. 2009), and sequenced. The amplified PCR products were deposited in GenBank (accession nos. MW261786, MW433689, MW439319 for ITS and MW249057, MW463344, MW463345 for PG3/PG2b). Blast searches against GenBank showed 99%-100% nucleotide identity with the reference sequences of various A. alternata isolates. The pathogenicity of the representative isolate (PDL 2021) was tested on Fig fruit cv. “Black Mission”. For that, 20 asymptomatic and mature fruit were surface-disinfected with 75% ethanol solution for 30 s. The fruit were inoculated by spraying a spore suspension (106 spores/ml) of A. alternata and stored at 25°C and 80% relative humidity. An equal number of fruit inoculated with sterile water were used as negative controls. Symptoms similar to those on the naturally infected fruits began after 4-5 days of inoculation. The negative controls remained healthy. Koch’s postulates were fulfilled by reisolating (100%) A. alternata from only the inoculated fruit. Previously, the pathogen has been reported to cause fruit rot of Lychee, Peach and Pomegranate in Pakistan (Alam et al 2017a; 2019b; 2019c). The pathogen has been reported to cause fig fruit rot in California (Michailides et al. 1994). Keeping in view the extent of disease on many fruits, further studies are needed on management options to combat the disease in Punjab Province of Pakistan.

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First Report of Phytophthora palmivora Causing Fruit Rot of Fig (Ficus carica L.) in China

Plant Disease ◽

10.1094/pdis-02-13-0158-pdn ◽

2013 ◽

Vol 97 (9) ◽

pp. 1252-1252 ◽

Cited By ~ 4

Author(s):

C. Zhang ◽

W. Zhang ◽

H. Q. Ma ◽

G. Z. Zhang

Keyword(s):

Large Scale ◽

Morphological Characteristics ◽

Pcr Amplification ◽

Daily Rainfall ◽

Ficus Carica ◽

Fruit Rot ◽

Phytophthora Palmivora ◽

Universal Primers ◽

First Report ◽

Immature Fruit

Fresh fig (Ficus carica L.) has been grown on a large scale in Beijing, China, since 2011. In late July 2012, a rot disease occurred on immature fruit of fig after a heavy rain (average daily rainfall 170 mm) in Fangshan District, Beijing, which caused about 30% incidence of green fruit on trees. The symptom first appeared as a water-soaked lesion that was covered with a white, fluffy mass of mycelia, followed by a soft, mushy rot of infected area on the fruit. To isolate the causal agent, mycelia and sporangia from 10 symptomatic fruits were suspended in sterile water, spread on potato dextrose agar (PDA) plates, and incubated at 25°C for 18 h. The isolates from each diseased fruit showed the same colonial characteristics. A single sporangium was isolated under a dissecting microscope and transferred onto PDA to obtain a pure culture. On carrot agar, the colony was white and homogeneous with tidy edge, with a few aerial hyphae. Sporangia were obpyriform with obvious papillae and measured 54.7 to 63.8 (59.3) × 26.5 to 36.3 (30.7) μm. The chlamydospores produced in culture were spherical. The pathogen was identified as Phytophthora palmivora based on the morphological characteristics (3) and confirmed with ITS sequences by PCR amplification using rDNA universal primers ITS1 and ITS4. The resulting sequence (Accession No. KC131229) had a 99% identity to that of P. palmivora (JQ354937) isolated from Pachira aquatica. Koch's postulates were conducted by inoculating six surface-sterilized figs with a PDA plug from a 7-day-old culture, with six noninoculated (PDA plugs only) fruits serving as controls. The inoculated fruits were incubated at room temperature in a plastic box covered with film. Symptoms similar to those on the naturally infected fruits began on wounded fruits 48 h after inoculation and on non-wounded fruits 60 h after inoculation, while the six control fruits remained healthy. P. palmivora was reisolated from the symptomatic fruit tissue. P. palmivora is one of the most severe pathogens on edible figs, being reported by Japanese in 1941 (2). Fruit rot of fig caused by the pathogen was reported in Florida in 1984 (1). To our knowledge, this is the first report of P. palmivora leading to fruit rot on fig in China. References: (1) N. E. El-Gholl and S. A. Alfieri, Jr. Proc. Fla. State Hort. Soc. 97:327, 1984. (2) Y. Nisikado et al. Ber. Ohara Inst. 8:427, 1941. (3) Y. N. Yu. Flora Fungorum Sinicorum: Peronosporales (in Chinese) Vol. 6. Science Press, Beijing, 1998.

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First Report of Fusarium proliferatum Causing Fruit Rot on Muskmelon (Cucumis melo L.) in China

Plant Disease ◽

10.1094/pdis-09-21-2015-pdn ◽

2021 ◽

Author(s):

Xiaoyan Yu ◽

Jing Zhang ◽

Lifeng Guo ◽

Aoran Yu ◽

Xiangjing Wang ◽

...

Keyword(s):

Rna Polymerase Ii ◽

Phylogenetic Trees ◽

Salvia Miltiorrhiza ◽

Disease Incidence ◽

Morphological Characteristics ◽

Fusarium Proliferatum ◽

Fruit Rot ◽

Single Spore ◽

Effective Prevention ◽

First Report

Muskmelon is an economically important crop in the world, especially in China, the largest producer of muskmelon with an annual output up to 12.7 million tonnes (Gómez-García et al. 2020). Since 2018, fruit rot was observed on muskmelon in Malianzhuang Base, the main muskmelon producing area in Shandong Province, whose disease incidence was about 25-30%. Water-soaked dark brown spots were initially appeared on the side of the fruit near the ground, then gradually expanded and covered with white mold with time. To isolate the pathogens, ten muskmelon fruits with typical symptoms were collected from different greenhouses in the base. Small tissues taken from the edge of the diseased and healthy tissues were immersed in 1% NaClO for 2 min, then soaked in 75% ethanol for 30 s, and rinsed 3 times with sterile distilled water (SDW). The sterilized tissues were naturally dried and placed on potato dextrose agar (PDA) amended with streptomycin sulfate (50 mg/L) for 7 days at 28℃. The emerging fungal mycelia were transferred to fresh PDA using the hyphal tip technology. Ten colonies were purified by single spore method and cultured on PDA for 7 days at 28℃ in the dark for morphological and molecular analyses. All colonies were flocculent with abundant white to light purple aerial hyphae, and the undersides of the colonies were observed to be from white to purple over time. Microconidia produced on PDA were hyaline, fusiform, ovoid, single cell without septum, and 4.5 to 12.7 × 2.0 to 3.6 μm in size (n=50). Macroconidia produced on carboxymethylcellulose agar (CMC) were slightly curved at both ends with three to five septa, and 17.6 to 35.7 × 2.8 to 4.0 μm in size (n=30). According to the morphological characteristics, these isolates were preliminarily identified as Fusarium sp. (Leslie and Summerell 2006). To further identify these isolates, genomic DNA of five isolates was extracted by CTAB method (Wu et al. 2001). The internal transcribed spacer (ITS) region of ribosomal DNA, translation elongation factor 1-α (TEF1) region, and the RNA polymerase II second largest subunit (RPB2) were amplified by PCR amplification with primers ITS1/ITS4, EF-1/EF-2, and RPB2-5F2/fRPB2-7cR, respectively (White et al. 1990; O’Donnell et al. 2008; Liu et al. 1999). Sequences of the five isolates were identical. The ITS, EF1-α, and RPB2 gene sequences of isolate NEAU-Mf-10-2 were submitted to NCBI GenBank with accession numbers of MZ950914, MZ960928, and MZ960929, respectively, having 100% similarity to those of Fusarium proliferatum (MK372368, MK952799 and MN245721). Phylogenetic trees were constructed based on the concatenated sequences of EF1-α and RPB2 genes using neighbour-joining and maximum-likelihood algorithms with MEGA 7.0. Two similar tree topologies both showed isolate NEAU-Mf-10-2 clustered with F. proliferatum NRRL 43665. Therefore, isolate NEAU-Mf-10-2 was identified as F. proliferatum based on morphological characteristics and phylogenetic analysis. To fulfill Koch’s postulates, ten muskmelon fruits (var. Tianbao) were soaked in 2% NaClO for 2 min, and then washed three times with SDW. Muskmelon fruits were inoculated by injecting conidia suspension (200 μL, 1×106 spores/mL) with a sterile injector. Ten other surface sterilized muskmelon fruits inoculated with sterile water were used as control. The fruits were placed in a light incubator at 28℃ with 12h light cycles for 7 days. All inoculated fruits showed symptoms highly similar to those of infected muskmelon fruits observed in the field. No symptoms were observed on fruits used as control. The Fusarium isolates were successfully re-isolated from the symptomatic fruits, and identified based on above morphological and molecular biological methods. Previous studies have reported that F. proliferatum can infect Polygonatum cyrtonema, Salvia miltiorrhiza, Allium cepa, A. sativum, and so on. To our knowledge, this is the first report of F. proliferatum causing fruit rot on muskmelon in China, which will provide basic information for designing effective prevention and control strategies on this disease.

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First Report of Alternaria alternata Causing Fruit Rot on Fig (Ficus carica) in Montenegro

Plant Disease ◽

10.1094/pdis-07-13-0770-pdn ◽

2014 ◽

Vol 98 (3) ◽

pp. 424-424 ◽

Cited By ~ 2

Author(s):

N. Latinović ◽

S. Radišek ◽

J. Latinović

Keyword(s):

Alternaria Alternata ◽

Direct Sequencing ◽

Disease Incidence ◽

Morphological Characteristics ◽

Fruit Rot ◽

Conidial Suspension ◽

Culture Collections ◽

First Report ◽

Control Fruit ◽

Rot Disease

In July 2012, a fruit rot disease was observed in several commercial fig tree orchards located in the Podgorica region in Montenegro. Symptoms on fruits initially appeared as small circular to oval, light brown, necrotic, sunken spots located mostly on the areas surrounding the ostiolar canal with an average diameter of 5 to 10 mm, which gradually enlarged in size leading to total fruit rot. Disease incidence on fruit across the fields ranged from 15 to 20% but the disease did not increase further due to hot and dry conditions thereafter. No foliar symptoms were observed. Small pieces (5 mm2) of symptomatic fruits were excised from the junction of diseased and healthy tissue, surface sterilized in 70% ethanol solution for 1 min, washed in three changes of sterile distilled water, air dried, and transferred to potato dextrose agar (PDA). After 2 to 3 days of incubation at 25°C, a fungus was consistently isolated. The isolates had radial growth and produced sooty black colonies. Microscopic observations of the colonies revealed brown septate hyphae and simple or branched conidiophores 30 to 65 μm long and 3 to 4.5 μm wide. Dark brown conidia were in chains (3 to 7), sized 10 to 35 × 5 to 9 μm, ellipsoid to ovoid, with 2 to 5 transverse and a few (1 to 3) to no longitudinal septa. Based on morphological characteristics, the fungus was identified as Alternaria alternata (3). For molecular identification, DNA was extracted from mycelia and conidia of two representative single spore isolates designated as ALT1-fCG and ALT2-fCG. PCR was carried out using internal transcribed spacer (ITS) region primers ITS4/ITS5 and A. alternata species-specific primers AAF2/AAR3 (1). Both primer pairs gave PCR products that were subjected to direct sequencing. BLAST analysis of the 546-bp ITS4/ITS5 (KF438091) and 294-bp AAF2/AAR3 (KF438092) sequences revealed 100% identity with several A. alternata isolates. Pathogenicity tests were conducted on 30 detached almost ripe and healthy fig fruit (cv. Primorka) by spraying them with a conidial suspension of the isolated fungus (106 conidia/ml) with a handheld sprayer. Thirty fruit inoculated with sterile water served as the non-inoculated control. Inoculated and control fruit were kept in a moist chamber at 25°C. Symptoms appeared on inoculated fruit 2 to 3 days after inoculation and all fruit were completely rotted 5 to 6 days after inoculation. Control fruit did not display any symptoms. A. alternata was consistently re-isolated from inoculated fruit, fulfilling Koch's postulates. The fig fruit rot caused by A. alternata has been reported before in California (2) and elsewhere mainly as postharvest pathogen. To our knowledge, this is the first report of fruit rot caused by A. alternata on fig in Montenegro. Considering Podgorica as the largest fig-producing area and the importance of fig as a traditionally grown crop, it could pose a threat to fig production in Montenegro. Voucher specimens are available at the culture collections of the University of Montenegro, Biotechnical Faculty. References: (1) P. Konstantinova et al. Mycol. Res. 106:23, 2002. (2) T. J. Michailides et al. Plant Dis. 78:44-50, 1994. (3) E. G. Simmons. Page 775 in: Alternaria and Identification Manual. CBS Fungal Biodiversity Centre, 2007.

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First report of banana (Musa acuminate L. AAA Cavendish, cv. Formosana) leaf spot disease caused by Curvularia geniculata in China

Plant Disease ◽

10.1094/pdis-09-21-2001-pdn ◽

2021 ◽

Author(s):

Yanxiang Qi ◽

Yanping Fu ◽

Jun Peng ◽

Fanyun Zeng ◽

Yanwei Wang ◽

...

Keyword(s):

Leaf Spot ◽

Disease Incidence ◽

Morphological Characteristics ◽

Universal Primers ◽

Leaf Spot Disease ◽

Leaf Margin ◽

Conidial Suspension ◽

Tropical Fruit ◽

First Report ◽

Spot Disease

Banana (Musa acuminate L.) is an important tropical fruit in China. During 2019-2020, a new leaf spot disease was observed on banana (M. acuminate L. AAA Cavendish, cv. Formosana) at two orchards of Chengmai county (19°48ʹ41.79″ N, 109°58ʹ44.95″ E), Hainan province, China. In total, the disease incidence was about 5% of banana trees (6 000 trees). The leaf spots occurred sporadically and were mostly confined to the leaf margin, and the percentage of the leaf area covered by lesions was less than 1%. Symptoms on the leaves were initially reddish brown spots that gradually expanded to ovoid-shaped lesions and eventually become necrotic, dry, and gray with a yellow halo. The conidia obtained from leaf lesions were brown, erect or curved, fusiform or elliptical, 3 to 4 septa with dimensions of 13.75 to 31.39 µm × 5.91 to 13.35 µm (avg. 22.39 × 8.83 µm). The cells of both ends were small and hyaline while the middle cells were larger and darker (Zhang et al. 2010). Morphological characteristics of the conidia matched the description of Curvularia geniculata (Tracy & Earle) Boedijn. To acquire the pathogen, tissue pieces (15 mm2) of symptomatic leaves were surface disinfected in 70% ethanol (10 s) and 0.8% NaClO (2 min), rinsed in sterile water three times, and transferred to potato dextrose agar (PDA) for three days at 28°C. Grayish green fungal colonies appeared, and then turned fluffy with grey and white aerial mycelium with age. Two representative isolates (CATAS-CG01 and CATAS-CG92) of single-spore cultures were selected for molecular identification. Genomic DNA was extracted from the two isolates, the internal transcribed spacer (ITS), large subunit ribosomal DNA (LSU rDNA), glyceraldehyde-3-phosphate dehydrogenase (GAPDH), translation elongation factor 1-alpha (TEF1-α) and RNA polymerase II second largest subunit (RPB2) were amplified and sequenced with universal primers ITS1/ITS4, LROR/LR5, GPD1/GPD2, EF1-983F/EF1-2218R and 5F2/7cR, respectively (Huang et al. 2017; Raza et al. 2019). The sequences were deposited in GenBank (MW186196, MW186197, OK091651, OK721009 and OK491081 for CATAS-CG01; MZ734453, MZ734465, OK091652, OK721100 and OK642748 for CATAS-CG92, respectively). For phylogenetic analysis, MEGA7.0 (Kumar et al. 2016) was used to construct a Maximum Likelihood (ML) tree with 1 000 bootstrap replicates, based on a concatenation alignment of five gene sequences of the two isolates in this study as well as sequences of other Curvularia species obtained from GenBank. The cluster analysis revealed that isolates CATAS-CG01 and CATAS-CG92 were C. geniculata. Pathogenicity assays were conducted on 7-leaf-old banana seedlings. Two leaves from potted plants were stab inoculated by puncturing into 1-mm using a sterilized needle and placing 10 μl conidial suspension (2×106 conidia/ml) on the surface of wounded leaves and equal number of leaves were inoculated with sterile distilled water serving as control (three replicates). Inoculated plants were grown in the greenhouse (12 h/12 h light/dark, 28°C, 90% relative humidity). Necrotic lesions on inoculated leaves appeared seven days after inoculation, whereas control leaves remained healthy. The fungus was recovered from inoculated leaves, and its taxonomy was confirmed morphologically and molecularly, fulfilling Koch’s postulates. C. geniculata has been reported to cause leaf spot on banana in Jamaica (Meredith, 1963). To our knowledge, this is the first report of C. geniculata on banana in China.

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First Report of Alternaria Leaf Spot of Banana Caused by Alternaria alternata in the United States

Plant Disease ◽

10.1094/pdis-01-13-0007-pdn ◽

2013 ◽

Vol 97 (8) ◽

pp. 1116-1116 ◽

Cited By ~ 3

Author(s):

V. Parkunan ◽

S. Li ◽

E. G. Fonsah ◽

P. Ji

Keyword(s):

United States ◽

Alternaria Alternata ◽

Leaf Spot ◽

Morphological Characteristics ◽

The United States ◽

Its Sequencing ◽

Single Spore ◽

First Report ◽

Alternaria Leaf Spot ◽

Leaf Spots

Research efforts were initiated in 2003 to identify and introduce banana (Musa spp.) cultivars suitable for production in Georgia (1). Selected cultivars have been evaluated since 2009 in Tifton Banana Garden, Tifton, GA, comprising of cold hardy, short cycle, and ornamental types. In spring and summer of 2012, 7 out of 13 cultivars (African Red, Blue Torres Island, Cacambou, Chinese Cavendish, Novaria, Raja Puri, and Veinte Cohol) showed tiny, oval (0.5 to 1.0 mm long and 0.3 to 0.9 mm wide), light to dark brown spots on the adaxial surface of the leaves. Spots were more concentrated along the midrib than the rest of the leaf and occurred on all except the newly emerged leaves. Leaf spots did not expand much in size, but the numbers approximately doubled during the season. Disease incidences on the seven cultivars ranged from 10 to 63% (10% on Blue Torres Island and 63% on Novaria), with an average of 35% when a total of 52 plants were evaluated. Six cultivars including Belle, Ice Cream, Dwarf Namwah, Kandarian, Praying Hands, and Saba did not show any spots. Tissue from infected leaves of the seven cultivars were surface sterilized with 0.5% NaOCl, plated onto potato dextrose agar (PDA) media and incubated at 25°C in the dark for 5 days. The plates were then incubated at room temperature (23 ± 2°C) under a 12-hour photoperiod for 3 days. Grayish black colonies developed from all the samples, which were further identified as Alternaria spp. based on the dark, brown, obclavate to obpyriform catenulate conidia with longitudinal and transverse septa tapering to a prominent beak attached in chains on a simple and short conidiophore (2). Conidia were 23 to 73 μm long and 15 to 35 μm wide, with a beak length of 5 to 10 μm, and had 3 to 6 transverse and 0 to 5 longitudinal septa. Single spore cultures of four isolates from four different cultivars were obtained and genomic DNA was extracted and the internal transcribed spacer (ITS1-5.8S-ITS2) regions of rDNA (562 bp) were amplified and sequenced with primers ITS1 and ITS4. MegaBLAST analysis of the four sequences showed that they were 100% identical to two Alternaria alternata isolates (GQ916545 and GQ169766). ITS sequence of a representative isolate VCT1FT1 from cv. Veinte Cohol was submitted to GenBank (JX985742). Pathogenicity assay was conducted using 1-month-old banana plants (cv. Veinte Cohol) grown in pots under greenhouse conditions (25 to 27°C). Three plants were spray inoculated with the isolate VCT1FT1 (100 ml suspension per plant containing 105 spores per ml) and incubated under 100% humidity for 2 days and then kept in the greenhouse. Three plants sprayed with water were used as a control. Leaf spots identical to those observed in the field were developed in a week on the inoculated plants but not on the non-inoculated control. The fungus was reisolated from the inoculated plants and the identity was confirmed by morphological characteristics and ITS sequencing. To our knowledge, this is the first report of Alternaria leaf spot caused by A. alternata on banana in the United States. Occurrence of the disease on some banana cultivars in Georgia provides useful information to potential producers, and the cultivars that were observed to be resistant to the disease may be more suitable for production. References: (1) E. G. Fonsah et al. J. Food Distrib. Res. 37:2, 2006. (2) E. G. Simmons. Alternaria: An identification manual. CBS Fungal Biodiversity Center, Utrecht, Netherlands, 2007.

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High Susceptibility of Olive Cultivar FS-17 to Alternaria alternata in Southern Spain

Plant Disease ◽

10.1094/pdis-92-8-1252a ◽

2008 ◽

Vol 92 (8) ◽

pp. 1252-1252 ◽

Cited By ~ 11

Author(s):

J. Moral ◽

R. De la Rosa ◽

L. León ◽

D. Barranco ◽

T. J. Michailides ◽

...

Keyword(s):

Alternaria Alternata ◽

Disease Incidence ◽

Southern Spain ◽

Fruit Rot ◽

Planting Density ◽

High Susceptibility ◽

Olive Cultivar ◽

White Skin ◽

Olive Cultivars ◽

Olive Trees

Traditional olive orchards in Spain have been planted at a density of 70 to 80 trees per ha with three trunks per tree. During the last decade, the hedgerow orchard, in which planting density is approximately 2,000 trees per ha, was developed. In 2006 and 2007, we noted a severe outbreak of fruit rot in FS-17, a new cultivar from Italy, in an experimental hedgerow planting in Córdoba, southern Spain. The incidence of fruit rot in ‘FS-17’ was 80% in January of 2006 and 24% in January of 2007. Cvs. Arbosana, IRTA-i18 (a selected clone from ‘Arbequina’), and Koroneiki had no symptoms in either year of the study. Disease incidence in ‘Arbequina’ was <0.1% only in 2006. Affected fruits were soft with gray-white skin and they eventually mummified. Black-green sporodochia were observed on the surface of diseased fruits. A fungus was isolated from diseased fruits on potato dextrose agar (PDA) and incubated at 22 to 26°C with a 12-h photoperiod. After 8 days of growing on PDA, fungal colonies formed conidial chains having a main axis with up to 10 conidia and secondary and tertiary short branches with two to four conidia. Conidia were obpyriform, ovoid, or ellipsoidal, without a beak or with a short beak, had up to four transverse septa, and measured 11.7 to 24.7 (mean 19.6) μm long and 7.7 to 13.0 (mean 9.6) μm wide at the broadest part of the conidium. The length of the beak of conidia was variable, ranging from 0 to 28.6 (mean 5.5) μm. The fungus was identified as Alternaria alternata (1). Pathogenicity tests were performed by spraying 40 mature fruits of ‘FS-17’ with a spore suspension (1 × 106 spores per ml). The same number of control fruits was treated with water. After 21 days, inoculated fruit developed symptoms that had earlier been observed in the field. A. alternata was reisolated from lesions on all infected fruits. The fungus was not isolated from any of the control fruits. The experiment was performed twice. The new growing system and the high susceptibility of some olive cultivars, such as FS-17, may result in a high incidence of disease caused by a pathogen that is generally characterized as weakly virulent. To our knowledge, this is the first report of A. alternata causing a severe outbreak of fruit rot on olive trees in the field. References: (1) B. M. Pryor and T. J. Michailides. Phytopathology 92:406, 2002.

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First Report of Fruit Rot of Melon Caused by Fusarium asiaticum in China

Plant Disease ◽

10.1094/pdis-08-20-1857-pdn ◽

2020 ◽

Author(s):

Fangmin Hao ◽

Quanyu Zang ◽

Weihong Ding ◽

Erlei Ma ◽

Yunping Huang ◽

...

Keyword(s):

Disease Incidence ◽

Elongation Factor ◽

Morphological Characteristics ◽

Its Region ◽

Fruit Rot ◽

Post Inoculation ◽

Basal Cells ◽

First Report ◽

Fusarium Asiaticum ◽

Sterile Needle

Melon (Cucumis melo L.) is a member of the Cucurbitaceae family, an important economical and horticultural crop, which is widely grown in China. In May 2020, fruit rot disease with water-soaked lesions and pink molds on cantaloupe melons was observed in several greenhouses with 50% disease incidence in Ningbo, Zhejiang Province in China. In order to know the causal agent, diseased fruits were cut into pieces, surface sterilized for 1 min with 1% sodium hypochlorite (NaClO), 2 min with 75% ethyl alcohol, rinsed in sterile distilled water three times (Zhou et al. 2018), and then placed on potato dextrose agar (PDA) medium amended with streptomycin sulfate (100 μg/ml) plates at 25°C for 4 days. The growing hyphae were transferred to new PDA plates using the hyphal tip method, putative Fusarium colonies were purified by single-sporing. Twenty-five fungal isolates were obtained and formed red colonies with white aerial mycelia at 25°C for 7 days, which were identified as Fusarium isolates based on the morphological characteristics and microscopic examination. The average radial mycelial growth rate of Fusarium isolate Fa-25 was 11.44 mm/day at 25°C in the dark on PDA. Macroconidia were stout with curved apical and basal cells, usually with 4 to 6 septa, and 29.5 to 44.2 × 3.7 to 5.2 μm on Spezieller Nährstoffarmer agar (SNA) medium at 25°C for 10 days (Leslie and Summerell 2006). To identify the species, the internal transcribed spacer (ITS) region and translational elongation factor 1-alpha (TEF1-α) gene of the isolates were amplified and cloned. ITS and TEF1-α was amplified using primers ITS1/ITS4 and EF1/EF2 (O’Donnell et al. 1998), respectively. Sequences of ITS (545 bp, GenBank Accession No. MT811812) and TEF1-α (707 bp, GenBank Acc. No. MT856659) for isolate Fa-25 were 100% and 99.72% identical to those of F. asiaticum strains MSBL-4 (ITS, GenBank Acc. MT322117.1) and Daya350-3 (TEF1-α, GenBank Acc. KT380124.1) in GenBank, respectively. A phylogenetic tree was established based on the TEF1-α sequences of Fa-25 and other Fusarium spp., and Fa-25 was clustered with F. asiaticum. Thus, both morphological and molecular characterizations supported the isolate as F. asiaticum. To confirm the pathogenicity, mycelium agar plugs (6 mm in diameter) removed from the colony margin of a 2-day-old culture of strain Fa-25 were used to inoculate melon fruits. Before inoculation, healthy melon fruits were selected, soaked in 2% NaClO solution for 2 min, and washed in sterile water. After wounding the melon fruits with a sterile needle, the fruits were inoculated by placing mycelium agar plugs on the wounds, and mock inoculation with mycelium-free PDA plugs was used as control. Five fruits were used in each treatment. The inoculated and mock-inoculated fruits were incubated at 25°C with high relative humidity. Symptoms were observed on all inoculated melon fruits 10 days post inoculation, which were similar to those naturally infected fruits, whereas the mock-inoculated fruits remained symptomless. The fungus re-isolated from the diseased fruits resembled colony morphology of the original isolate. The experiment was conducted three times and produced the same results. To our knowledge, this is the first report of fruit rot of melon caused by F. asiaticum in China.

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First Report of Colletotrichum siamense Causing Anthracnose of Monstera deliciosa in Zhanjiang, China

Plant Disease ◽

10.1094/pdis-08-20-1839-pdn ◽

2020 ◽

Author(s):

Yue Lian Liu ◽

Jian Rong Tang ◽

Yu Han Zhou

Keyword(s):

Disease Incidence ◽

Morphological Characteristics ◽

Pathogenicity Test ◽

Sterile Water ◽

Single Spore ◽

First Report ◽

Rdna Its ◽

Colletotrichum Siamense ◽

Monstera Deliciosa ◽

Pure Cultures

Monstera deliciosa Liebm is an ornamental foliage plant (Zhen et al. 2020De Lojo and De Benedetto 2014). In July of 2019, anthracnose lesions were observed on leaves of M. deliciosa cv. Duokong with 20% disease incidence of 100 plants at Guangdong Ocean University campus (21.17N,110.18E), Guangdong Province, China. Initially affected leaves showed chlorotic spots, which coalesced into larger irregular or circular lesions. The centers of spots were gray with a brown border surrounded by a yellow halo (Supplementary figure 1). Twenty diseased leaves were collected for pathogen isolation. Margins of diseased tissue was cut into 2 × 2 mm pieces, surface-disinfected with 75% ethanol for 30 s and 2% sodium hypochlorite (NaOCl) for 60 s, rinsed three times with sterile water before isolation. Potato dextrose agar (PDA) was used to culture pathogens at 28℃ in dark. Successively, pure cultures were obtained by transferring hyphal tips to new PDA plates. Fourteen isolates were obtained from 20 leaves. Three single-spore isolates (PSC-1, PSC-2, and PSC-3) were obtained ,obtained, which were identical in morphology and molecular analysis (ITS). Therefore, the representative isolate PSC-1 was used for further study. The culture of isolate PSC-1 on PDA was initially white and later became cottony, light gray in 4 days, at 28 °C. Conidia were single celled, hyaline, cylindrical, clavate, and measured 13.2 to 18.3 µm × 3.3 to 6.5 µm (n = 30). Appressoria were elliptical or subglobose, dark brown, and ranged from 6.3 to 9.5 µm × 5.7 to 6.5 µm (n = 30). Morphological characteristics of isolate PSC-1 were consistent with the description of Colletotrichum siamense (Prihastuti et al. 2009; Sharma et al. 2013). DNA of the isolate PSC-1 was extracted for PCR sequencing using primers for the rDNA ITS (ITS1/ITS4), GAPDH (GDF1/GDR1), ACT (ACT-512F/ACT-783R), CAL (CL1C/CL2C), and TUB2 (βT2a/βT2b) (Weir et al. 2012). Analysis of the ITS (accession no. MN243535), GAPDH (MN243538), ACT (MN512640), CAL (MT163731), and TUB2 (MN512643) sequences revealed a 97-100% identity with the corresponding ITS (JX010161), GAPDH (JX010002), ACT (FJ907423), CAL (JX009714) and TUB2 (KP703502) sequences of C. siamense in GenBank. A phylogenetic tree was generated based on the concatenated sequences of ITS, GAPDH, ACT, CAL, and TUB2 which clustered the isolate PSC-1 with C. siamense the type strain ICMP 18578 (Supplementary figure 2). Based on morphological characteristics and phylogenetic analysis, the isolate PSC-1 associated with anthracnose of M. deliciosa was identified as C. siamense. Pathogenicity test was performed in a greenhouse at 24 to 30oC with 80% relative humidity. Ten healthy plants of cv. Duokong (3-month-old) were grown in pots with one plant in each pot. Five plants were inoculated by spraying a spore suspension (105 spores ml-1) of the isolate PSC-1 onto leaves until runoff, and five plants were sprayed with sterile water as controls. The test was conducted three times. Anthracnose lesions as earlier were observed on the leaves after two weeks, whereas control plants remained symptomless. The pathogen re-isolated from all inoculated leaves was identical to the isolate PSC-1 by morphology and ITS analysis, but not from control plants. C. gloeosporioides has been reported to cause anthracnose of M. deliciosa (Katakam, et al. 2017). To the best of our knowledge, this is the first report of C. siamense causing anthracnose on M. deliciosa in ChinaC. siamense causes anthracnose on a variety of plant hosts, but not including M. deliciosa (Yanan, et al. 2019). To the best of our knowledge, this is the first report of C. siamense causing anthracnose on M. deliciosa, which provides a basis for focusing on the management of the disease in future.

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Fusarium acuminatum Associated with Root Rot of Ophiopogon japonicus (Linn. f.) in China

Plant Disease ◽

10.1094/pdis-11-20-2344-pdn ◽

2020 ◽

Author(s):

Tao Tang ◽

Fanfan Wang ◽

Jie Guo ◽

XiaoLiang Guo ◽

Yuanyuan Duan ◽

...

Keyword(s):

Root Rot ◽

Disease Incidence ◽

Morphological Characteristics ◽

Vaccinium Corymbosum ◽

Fruit Rot ◽

Its Sequence ◽

Pathogenicity Test ◽

Direct Pcr ◽

Fusarium Acuminatum ◽

Ophiopogon Japonicus

Ophiopogon japonicus (Linn. f.) is a perennial evergreen in the Liliaceae family that is cultivated in many provinces of China due to its high medicinal and economic value . In April 2019, an unknown root rot disease was observed on the rhizomes of O. japonicus in a commercial production field in Xiangyang City (30.83° N, 112.53° E), Hubei Province. Disease incidence was approximately 10-20%. Symptoms included chlorosis, drooping and rolling of the leaves followed by rapid death of entire plant. Infected roots appeared to be softened, necrotic, and shriveled with reddish fungal growth. Infected tissues were disinfested on surface with 75% ethanol for 30 s and 0.1% HgCl2 for 1 min, rinsed with sterile distilled water, and dried. Small pieces (2 mm × 2 mm) were then excised from disinfested tissue and incubated on potato dextrose agar (PDA) medium at 25 ℃ in the dark. After 3 days of incubation, six isolates with 75% of isolation rate and same colony morphology were sub-cultured and purified by hyphal tip isolation. Purified cultures grew rapidly and media plates (70×70 mm ) were covered with hyphae after 3 to 4 days. Cultures were initially white and became pink or red over 5 days. Microconidia were not observed. Macroconidia were produced from monophialides on branched conidiophores, which were slender, equilaterally curved, and measured 32.5 to 53.5 μm in length and 3.5 to 5.1 μm in width, with three to five septa. All strains were preliminarily identified as Fusarium acuminatum (Eslie and Summerell 2006) on the basis of morphology. To confirm the identity of the pathogen, molecular identification was performed with strain MD1. Following DNA extraction, PCR was performed using the TSINGKE 2×T5 Direct PCR Mix kit. Target areas of amplification were internal transcribed spacer (ITS), RNA polymerase second largest subunit (RPB2) and beta-tubulin gene (TUB2) regions of rDNA, using ITS1,4 (Yin et al. 1990) , RPB2-5f2/7cr (O’Donnell et al. 2010)and Btu-F-F01, Btu-F-R01 primers(Wang et al. 2014), respectively. Nucleotide sequences were deposited in NCBI (GenBank MT525360.1; MW164629; MT588110.1). BLAST analysis of the ITS sequence had 100% similarity to a 517 bp portion of F. acuminatum sequence in GenBank (MK764994.1) ；RPB2 sequence had 100% similarity to a 687 bp portion of F. acuminatum sequence in GenBank (HM068330.1) and TUB2 sequence had 99% similarity to a 964 bp portion of F. acuminatum sequence in GenBank (KT965741.1). A pathogenicity test was performed in laboratory on O. japonicus roots with isolate MD1. Mycelial plugs (5 mm) were excised from the margin of colony cultured for 5 days, and placed on three-years-old tuberous roots covered with wet sterile cotton and kept at 25℃, under 80% relative humidity. Controls were inoculated with non-colonized PDA plugs (5 mm). All treatments had three replicate plants. On incolated plants, white hyphae covered on O. japonicus roots 3 DPI became pink and by 5 DPI, roots had rot symptoms. By comparision, the control plants had no symptoms. The pathogen was reisolated from the inoculated roots and exhibited same morphological characteristics and ITS sequence as those of F. acuminatum. F. acuminatum was reported to cause fruit rot on postharvest pumpkin and Vaccinium corymbosum in China (Li et al. 2019; Wang et al. 2016).To our knowledge, this is the first report of root rot caused by F. acuminatum on O. japonicus in China.

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First Report of Passalora Leaf Spot Caused by Passalora bougainvilleae on Bougainvillea in Mexico

Plant Disease ◽

10.1094/pdis-11-10-0787 ◽

2011 ◽

Vol 95 (6) ◽

pp. 775-775 ◽

Cited By ~ 1

Author(s):

V. Ayala-Escobar ◽

V. Santiago-Santiago ◽

A. Madariaga-Navarrete ◽

A. Castañeda-Vildozola ◽

C. Nava-Diaz

Keyword(s):

Uv Light ◽

Disease Incidence ◽

Morphological Characteristics ◽

The United States ◽

Pathogenicity Test ◽

Commonwealth Mycological Institute ◽

Conidial Suspension ◽

Single Spore ◽

First Report ◽

Bougainvillea Spectabilis

Bougainvillea (Bougainvillea spectabilis Willd) growing in 28 gardens during 2009 showed 100% disease incidence and 3 to 7% disease severity. Bougainvilleas with white flowers were the most affected. Symptoms consisted of light brown spots with dark brown margins visible on adaxial and abaxial sides of the leaves. Spots were circular, 2 to 7 mm in diameter, often surrounded by a chlorotic halo, and delimited by major leaf veins. Single-spore cultures were incubated at 24°C under near UV light for 7 days to obtain conidia. Pathogenicity was confirmed by spraying a conidial suspension (1 × 104 spores/ml) on leaves of potted bougainvillea plants (white, red, yellow, and purple flowers), incubating the plants in a dew chamber for 48 h and maintaining them in a greenhouse (20 to 24°C). Identical symptoms to those observed at the residential gardens appeared on inoculated plants after 45 to 60 days. The fungus was reisolated from inoculated plants that showed typical symptoms. No symptoms developed on control plants treated with sterile distilled water. The fungus produced distinct stromata that were dark brown, spherical to irregular, and 20 to 24 μm in diameter. Conidiophores were simple, born from the stromata, loose to dense fascicles, brown, straight to curved, not branched, zero to two septate, 14 × 2 μm, with two to four conspicuous and darkened scars. The conidia formed singly, were brown, broad, ellipsoid, obclavate, straight to curved with three to four septa, 40 × 4 μm, and finely verrucous with thick hilum at the end. Fungal DNA from the single-spore cultures was obtained using a commercial DNA Extraction Kit (Qiagen, Valencia, CA); ribosomal DNA was amplified with ITS5 and ITS4 primers and sequenced. The sequence was deposited at the National Center for Biotechnology Information Database (GenBank Accession Nos. HQ231216 and HQ231217). The symptoms (4), morphological characteristics (1,2,4), and pathogenicity test confirm the identity of the fungus as Passalora bougainvilleae (Muntañola) Castañeda & Braun (= Cercosporidium bougainvilleae Muntañola). This pathogen has been reported from Argentina, Brazil, Brunei, China, Cuba, El Salvador, India, Indonesia, Jamaica, Japan, Thailand, the United States, and Venezuela (3). To our knowledge, this is the first report of this disease on B. spectabilis Willd in Mexico. P. bougainvilleae may become an important disease of bougainvillea plants in tropical and subtropical areas of Mexico. References: (1) U. Braun and R. R. Castañeda. Cryptogam. Bot. 2/3:289, 1991. (2) M. B. Ellis. More Dematiaceous Hypomycetes. Commonwealth Mycological Institute, Kew, Surrey, UK, 1976. (3) C. Nakashima et al. Fungal Divers. 26:257, 2007. (4) K. L. Nechet and B. A. Halfeld-Vieira. Acta Amazonica 38:585, 2008.

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