scholarly journals First report of Leptosphaeria biglobosa ‘canadensis’ causing blackleg on oilseed rape (Brassica napus) in China

Plant Disease ◽  
2021 ◽  
Author(s):  
Tao Luo ◽  
Guoqing Li ◽  
Long Yang
Keyword(s):  
Brassica Napus ◽  
Oilseed Rape ◽  
Southern China ◽  
Disease Incidence ◽  
Morphological Characteristics ◽  
Gansu Province ◽  
Bootstrap Support ◽  
First Report ◽  
Fungal Isolates ◽  
Leptosphaeria Biglobosa

Oilseed rape (Brassica napus L.) is one of the most important oilseed crops in China. It is widely cultivated in China, with winter oilseed rape in Yangtze River basin and in southern China, and spring oilseed rape in northern China. In August 2017, a survey for Leptosphaeria spp. on spring oilseed rape was conducted in Minle county, Zhangye city, Gansu Province, China. The symptoms typical of blackleg on basal stems of oilseed rape were observed in the field. A large number of black fruiting bodies (pycnidia) were present on the lesions (Fig. 1A). The disease incidence of basal stem infection in the surveyed field was 19%. A total of 19 diseased stems were collected to isolate the pathogen. After surface sterilizing (75% ethanol for 30 s, 5% NaOCl for 60 s, followed by rinsing in sterilized water three times), diseased tissues were cultured on acidified potato dextrose agar (PDA) plates at 20°C for 7 days. Twelve fungal isolates were obtained. All fungal isolates produced typical tan pigment on PDA medium, and produced pycnidia after two weeks (Fig. 1B). Colony morphological characteristics indicated that these isolates might belong to Leptosphaeria biglobosa. To confirm identification, multiple PCR was conducted using the species-specific primers LmacF, LbigF, LmacR (Liu et al. 2006). Genomic DNA of each isolate was extracted using the cetyltrimethylammonium bromide (CTAB) method. DNA samples of L. maculans isolate UK-1 and L. biglobosa isolate W10 (Cai et al. 2015) were used as references. Only a 444-bp DNA band was detected in all 12 isolates and W10, whereas a 333-bp DNA band was detected only in the UK-1 isolate (Fig. 1C). PCR results suggested that these 12 isolates all belong to L. biglobosa. In addition, the internal transcribed spacer (ITS) region of these 12 isolates was analyzed for subspecies identification (Vincenot et al. 2008). Phylogenetic analysis based on ITS sequence showed that five isolates (Lb1134, Lb1136, Lb1138, Lb1139 and Lb1143) belonged to L. biglobosa ‘brassicae’ (Lbb) with 78% bootstrap support, and the other seven isolates (Lb1135, Lb1137, Lb1140, Lb1141, Lb1142, Lb1144 and Lb1145) belonged to L. biglobosa ‘canadensis’ (Lbc) with 95% bootstrap support (Fig. 1D). Two Lbb isolates (Lb1134 and Lb1136) and two Lbc isolates (Lb1142 and Lb1144) were randomly selected for pathogenicity testing on B. napus cultivar Zhongshuang No. 9 (Wang et al. 2002). Conidial suspensions (10 μL, 1 × 107 conidia mL-1) of these four isolates were inoculated on needle-wounded cotyledons (14-day-old seedling), with 10 cotyledons (20 wounded sites) per isolate. A further 10 wounded cotyledons were inoculated with water and served as controls. Seedlings were maintained in a growth chamber at 20°C with 100% relative humidity and a 12-h photoperiod. After 7 days, cotyledons inoculated with the four isolates showed necrotic lesions in the inoculated wounds. Control cotyledons had no symptoms (Fig. 2). Fungi re-isolated from the infected cotyledons showed similar colony morphology as the original isolates. Therefore, L. biglobosa ‘brassicae’ and L. biglobosa ‘canadensis’ appear to be the pathogens causing the observed blackleg symptoms on spring oilseed rape in Gansu, China. In previous studies, L. biglobosa ‘brassicae’ has been found in many crops in China, including oilseed rape (Liu et al. 2014; Cai et al. 2015), Chinese radish (Raphanus sativus) (Cai et al. 2014a), B. campestris ssp. chinensis var. purpurea (Cai et al. 2014b), broccoli (B. oleracea var. italica) (Luo et al. 2018), ornamental kale (B. oleracea var. acephala) (Zhou et al. 2019a), B. juncea var. multiceps (Zhou et al. 2019b), B. juncea var. tumida (Deng et al. 2020) and Chinese cabbage (B. rapa subsp. pekinensis) (Yu et al. 2021 accepted). To the best of our knowledge, this is the first report of L. biglobosa ‘canadensis’ causing blackleg on B. napus in China.

scholarly journals First Report of Flyspeck Caused by Zygophiala wisconsinensis on Sweet Persimmon Fruit in Korea

Plant Disease ◽  
2011 ◽  
Vol 95 (5) ◽  
pp. 616-616 ◽  
Author(s):  
J. Kim ◽  
O. Choi ◽  
J.-H. Kwon
Keyword(s):  
Disease Incidence ◽  
Morphological Characteristics ◽  
Its Rdna ◽  
Control Treatment ◽  
Diospyros Kaki ◽  
Distilled Water ◽  
First Report ◽  
Persimmon Fruit ◽  
Fungal Isolates ◽  
Agar Disc

Sweet persimmon (Diospyros kaki L.), a fruit tree in the Ebenaceae, is cultivated widely in Korea and Japan, the leading producers worldwide (2). Sweet persimmon fruit with flyspeck symptoms were collected from orchards in the Jinju area of Korea in November 2010. The fruit had fungal clusters of black, round to ovoid, sclerotium-like fungal bodies with no visible evidence of a mycelial mat. Orchard inspections revealed that disease incidence ranged from 10 to 20% in the surveyed area (approximately 10 ha) in 2010. Flyspeck symptoms were observed on immature and mature fruit. Sweet persimmon fruit peels with flyspeck symptoms were removed, dried, and individual speck lesions transferred to potato dextrose agar (PDA) and cultured at 22°C in the dark. Fungal isolates were obtained from flyspeck colonies on 10 sweet persimmon fruit harvested from each of three orchards. Fungal isolates that grew from the lesions were identified based on a previous description (1). To confirm identity of the causal fungus, the complete internal transcribed spacer (ITS) rDNA sequence of a representative isolate was amplified and sequenced using primers ITS1 and ITS4 (4). The resulting 552-bp sequence was deposited in GenBank (Accession No. HQ698923). Comparison with ITS rDNA sequences showed 100% similarity with a sequence of Zygophiala wisconsinensis Batzer & Crous (GenBank Accession No. AY598855), which infects apple. To fulfill Koch's postulates, mature, intact sweet persimmon fruit were surface sterilized with 70% ethanol and dried. Three fungal isolates from this study were grown on PDA for 1 month. A colonized agar disc (5 mm in diameter) of each isolate was cut from the advancing margin of a colony with a sterilized cork borer, transferred to a 1.5-ml Eppendorf tube, and ground into a suspension of mycelial fragments and conidia in a blender with 1 ml of sterile, distilled water. The inoculum of each isolate was applied by swabbing a sweet persimmon fruit with the suspension. Three sweet persimmon fruit were inoculated per isolate. Three fruit were inoculated similarly with sterile, distilled water as the control treatment. After 1 month of incubation in a moist chamber at 22°C, the same fungal fruiting symptoms were reproduced as observed in the orchards, and the fungus was reisolated from these symptoms, but not from the control fruit, which were asymptomatic. On the basis of morphological characteristics of the fungal colonies, ITS sequence, and pathogenicity to persimmon fruit, the fungus was identified as Z. wisconsinensis (1). Flyspeck is readily isolated from sweet persimmon fruit in Korea and other sweet persimmon growing regions (3). The exposure of fruit to unusual weather conditions in Korea in recent years, including drought, and low-temperature and low-light situations in late spring, which are favorable for flyspeck, might be associated with an increase in occurrence of flyspeck on sweet persimmon fruit in Korea. To our knowledge, this is the first report of Z. wisconsinensis causing flyspeck on sweet persimmon in Korea. References: (1) J. C. Batzer et al. Mycologia 100:246, 2008. (2) FAOSTAT Database. Retrieved from http://faostat.fao.org/ , 2008. (3) H. Nasu and H. Kunoh. Plant Dis. 71:361, 1987. (4) T. J. White et al. Page 315 in: PCR Protocols: A Guide to Methods and Applications. M. A. Innis et al., eds. Academic Press, Inc., New York, 1990.

scholarly journals First Report of a Chaetomella sp. Causing a Leaf Spot on Sansevieria trifasciata in China

Plant Disease ◽  
2013 ◽  
Vol 97 (7) ◽  
pp. 992-992 ◽  
Author(s):  
Y. L. Li ◽  
Z. Zhou ◽  
W. Lu ◽  
J. R. Ye
Keyword(s):  
Leaf Spot ◽  
Disease Incidence ◽  
Morphological Characteristics ◽  
Leaf Tissue ◽  
Aerial Mycelium ◽  
Control Treatment ◽  
Distilled Water ◽  
First Report ◽  
Potted Plants ◽  
Fungal Isolates

Sansevieria trifasciata originates from tropical West Africa. It is widely planted as a potted ornamental in China for improving indoor air quality (1). In February 2011, leaves of S. trifasciata plants in an ornamental market of Anle, Luoyang City, China, were observed with sunken brown lesions up to 20 mm in diameter, and with black pycnidia present in the lesions. One hundred potted plants were examined, with disease incidence at 20%. The symptomatic leaves affected the ornamental value of the plants. A section of leaf tissue from the periphery of two lesions from a plant was cut into 1 cm2 pieces, soaked in 70% ethanol for 30 s, sterilized with 0.1% HgCl2 for 2 min, then washed five times in sterilized distilled water. The pieces were incubated at 28°C on potato dextrose agar (PDA). Colonies of two isolates were brown with submerged hyphae, and aerial mycelium was rare. Abundant and scattered pycnidia were reniform, dark brown, and 200 to 350 × 100 to 250 μm. There were two types of setae on the pycnidia: 1) dark brown setae with inward curved tops, and 2) straight, brown setae. Conidia were hyaline, unicellular, cylindrical, and 3.75 to 6.25 × 1.25 to 2.50 μm. Morphological characteristics suggested the two fungal isolates were a Chaetomella sp. To confirm pathogenicity, six mature leaves of a potted S. trifasciata plant were wounded with a sterile pin after wiping each leaf surface with 70% ethanol and washing each leaf with sterilized distilled water three times. A 0.5 cm mycelial disk cut from the margin of a 5-day-old colony on a PDA plate was placed on each pin-wounded leaf, ensuring that the mycelium was in contact with the wound. Non-colonized PDA discs were placed on pin-wounded leaves as the control treatment. Each of two fungal isolates was inoculated on two leaves, and the control treatment was done similarly on two leaves. The inoculated plant was placed in a growth chamber at 28°C with 80% relative humidity. After 7 days, inoculated leaves produced brown lesions with black pycnidia, but no symptoms developed on the control leaves. A Chaetomella sp. was reisolated from the lesions of inoculated leaves, but not from the control leaves. An additional two potted plants were inoculated using the same methods as replications of the experiment, with identical results. To confirm the fungal identification, the internal transcribed spacer (ITS) region of rDNA of the two isolates was amplified using primers ITS1 and ITS4 (2) and sequenced. The sequences were identical (GenBank Accession No. KC515097) and exhibited 99% nucleotide identity to the ITS sequence of an isolate of Chaetomella sp. in GenBank (AJ301961). To our knowledge, this is the first report of a leaf spot of S. trifasciata caused by Chaetomella sp. in China as well as anywhere in the world. References: (1) X. Z. Guo et al. Subtropical Crops Commun. Zhejiang 27:9, 2005. (2) T. J. White et al. PCR Protocols: A Guide to Methods and Applications. M. A. Innis et al., eds. Academic Press, San Diego, CA, 1990.

scholarly journals First report of Corynespora cassicola causing black spot on Sarcandra glabra in China

Plant Disease ◽  
2021 ◽  
Author(s):  
Jiang Ni ◽  
B. R. Lin ◽  
Lisha Song ◽  
Guiyu Tan ◽  
Jiang zhan Zhang ◽  
...  
Keyword(s):  
Salvia Miltiorrhiza ◽  
Pathogenic Fungus ◽  
Morphological Characteristics ◽  
Bootstrap Support ◽  
Control Group ◽  
Broad Host Range ◽  
Pathogenicity Test ◽  
Guangxi Province ◽  
First Report ◽  
Fungal Isolates

Sarcandra glabra is an important Chinese medicinal plant, which was widely cultivated under forest in south China. Guangxi province is the main producing areas of this herb. In June 2019, a serious leaf disease was found causing severe defoliation in the S. glabra plantation under bamboo forest in Rongan country, Guangxi province (109°13′N′′E). About 70% of the plants in the plantation (300 ha) showed the similar symptoms. Initially, circular lesions appeared on young leaves as black spots (about 1 to 2 mm). Then, the spots gradually enlarged usually with an obvious yellowish margin (6 to 8 mm). Finally, the lesions coalesced and formed irregular, black, and large necrotic areas, resulting in the leaf abscission. For pathogen isolation, small pieces of tissue (5×5 mm) taken from 25 diseased leaves were sterilized with 75% ethanol for 30 s, subsequently, soaked in 0.1% HgCl2 for 2 min, rinsed three times in sterile distilled water, dried, and then placed aseptically onto the potato dextrose agar (PDA) plates, and incubated at 28 °C (12 h/12 h light/dark). Three days later, the isolates were placed on a new PDA plate for subsequent purification and sporulation. 20 pure fungal isolates were obtained from single spores. Of which, 15 isolates showed similar morphological characteristics.The colonies on PDA were round, dense, gray edge and dark gray in center area. Conidia in culture were appeared light brown, cylindrical in shape, with 0 to 8 septa, and 55 to 165 μm × 5.2 to 13.5 μm in size (mean = 106.2 μm × 8.6 μm, n = 30). These morphological characteristics resemble those of Corynespora sp. (Berk. & M.A. Curtis) C.T. Wei (Ellis et al. 1971). A single-spore isolate (ZD5) was selected from the 15 fungal isolates for a subsequent molecular identification. The genes of internal transcribed spacer (ITS) of ribosomal DNA, β-tublin, and actin were amplified with the primer pairs ITS-1/ITS-4 (White et al. 1990), β-tubulin 2-Bt2a/Bt2b (Glass and Donaldson 1995), ACT-512F/ACT-783R (Carbone and Kohn 1999), respectively. And the ITS, β-tublin, and actin sequences were deposited in the GenBank database with the accession numbers MW362446, MW367029, and MW533122. Blast analysis and neighbor-joining analysis based on ITS, β-tublin, and actin sequences using MEGA 6 revealed that the isolate was placed in the same clade as C. cassicola with 100% bootstrap support. Pathogenicity test was performed on the two-year-old potted S. glabra. Six-mm-diameter mycelial plugs were attached to the healthy leaves of S. glabra for co-culture, while the control group was attached with PDA. All plants were covered with plastic bags for 2 days in order to maintain high humidity and cultured in a greenhouse at 28 °C with a 12-h/12-h light/dark cycle. The symptoms appeared 2 days after co-culture were identical to those observed in the field. The same fungus was re-isolated from the lesions, and further morphological characterization and molecular assays, as described above.The control leaves remained symptomless during the pathogenicity tests. According to the previous literatures, C. cassicola is a plant pathogenic fungus with a broad host range, which can damage diverse tropical plants including Salvia miltiorrhiza (Lu et al. 2019), Solanum americanum (Wagner and Louise 2019), Vitex rotundifolia (Yeh and Kirschner 2017), Cucumis sativus, Lycopersicon esculentum (Hsu et al. 2002), Carica papaya (Tsai et al. 2015),and so on. To our knowledge, this is the first report of C. cassicola causing leaf spot on S. glabra in China.

scholarly journals First Report of Anthracnose Caused by Colletotrichum gloeosporioides on Pistachio (Pistacia vera) in China

Plant Disease ◽  
2011 ◽  
Vol 95 (10) ◽  
pp. 1314-1314 ◽  
Author(s):  
S. Y. Yang ◽  
S. C. Su ◽  
T. Liu ◽  
G. Fan ◽  
J. Wang ◽  
...  
Keyword(s):  
Dna Sequence ◽  
Colletotrichum Gloeosporioides ◽  
Disease Incidence ◽  
Morphological Characteristics ◽  
Northern China ◽  
Gansu Province ◽  
Internal Transcribed Spacers ◽  
Pistacia Vera ◽  
First Report ◽  
Stem Cankers

In the 1990s, pistachio (Pistacia vera L. ‘Kerman’ and ‘Peters’) was introduced in China. They are found in many orchards in Beijing and Gansu and Hebei provinces, northern China. In 2009, a new disease was observed on leaves, stems, and fruits in pistachio orchards in Gansu Province. Disease incidence in 8- to 12-year-old orchards was 30%. Yield losses reached 25%. Symptoms began as discrete, sunken, black spots, approximately 10 mm in diameter, followed by circular lesions that eventually coalesced with tissue death recorded and orange fructifications developed on lesions. Pieces of diseased leaves, stems, and fruits were surfaced sterilized and placed on 2% potato dextrose agar (PDA) at 25°C. A fungus was consistently isolated. After 10 days, cultures on PDA showed aerial, white mycelium that turned gray to grayish black with a salmon-to-orange conidial mass at 25°C and a 12-h photoperiod. Brown, 80 to 120 μm long setae were observed in the acervulus. Conidia were hyaline, fusiform to nearly straight, and averaged 12 to 18 × 3 to 5 μm. On the basis of morphological characteristics, the fungus was identified as Colletotrichum gloeosporioides (Penz.) Sacc. (2). On PDA, 0.5 μg/ml of benomyl was applied for the sensitivity test (3). Benomyl completely inhibited the growth of the fungus. Mycelial DNA was extracted, PCR amplified using ITS1 and ITS4 primers for the ribosomal DNA internal transcribed spacers 1 and 2, and sequenced. The DNA sequence was recorded in GenBank as No. HQ631378. The DNA sequence was blasted showing 99% identity with Accession Nos. GQ144454 and GU004376, for C. gloeosporioides. Pathogenicity tests were conducted under greenhouse conditions at 25°C. Three replicates of 2-year-old ‘Kerman’ plants were inoculated with mycelial PDA plugs placed on 0.5-cm2 stem wounds and then wrapped with Parafilm. Controls were inoculated with PDA plugs without the fungus. After 3 weeks, stem cankers were observed on inoculated plants. Control plants remained healthy. Pathogenicity was also tested on injured leaves and fruits. A 10-μl drop of a spore suspension of 104 conidia/ml was applied on ‘Kerman’ and ‘Peters’ leaves and ‘Kerman’ fruits and placed on plates with a wet filter paper at 25°C. Small, black lesions were observed at 2 days after inoculation. At 7 days, necrotic lesions covered the entire surface. C. gloeosporioides was reisolated from necrotic lesions. Controls did not develop symptoms. C. acutatum has been reported on pistachio in Australia (1), but to our knowledge, this is the first report of anthracnose caused by C. gloeosporioides on pistachio. References: (1) G. J. Ash and V. M. Lanoiselet. Australas. Plant Pathol. 30:365, 2001. (2) J. Y. Lu. Plant Pathogenic Mycology. China Agricultural Press, Beijing, 2001. (3) N. A. R. Peres et al. Plant Dis. 86:620, 2002.

scholarly journals First Report of Anthracnose of Gossypium indicum Lam. caused by Colletotrichum theobromicola in Korea

Plant Disease ◽  
2021 ◽  
Author(s):  
Donghun Kang ◽  
Jungyeon Kim ◽  
Youn Mi Lee ◽  
Balaraju Kotnala ◽  
Yongho Jeon
Keyword(s):  
Disease Incidence ◽  
Morphological Characteristics ◽  
Bootstrap Support ◽  
Pathogenicity Test ◽  
Distilled Water ◽  
First Report ◽  
Spray Method ◽  
Cotton Plants ◽  
Leaf Surface Area ◽  
And Control

In September 2020, typical anthracnose symptoms were observed on cotton (Gossypium indicum Lam.) leaves growing in Hahoe village, Andong, Gyeongbuk Province, Korea. The leaves of the infected plants initially showed spots with halo-lesions which became enlarged and spread to the entire leaf surface area. The infected leaves later became yellowish and chlorotic (Fig. 1A). The disease incidence was at least 90% in the field. For pathogen isolation, fresh samples collected from symptomatic leaves were cut into small pieces (4 to 5 mm2), surface-sterilized in 1% sodium hypochlorite for 1 min, rinsed three times, and macerated in sterile distilled water (SDW). They were spread onto potato dextrose agar (PDA) plates and incubated at 25 °C for 5 days under a 12-h photoperiod. Five isolates were recovered from the infected leaves. Purified fungal colonies were initially white, later turned yellow on PDA medium. Conidia were yellow-colored, smooth-walled, aseptate, straight or slightly distorted, and cylindrical with one end slightly acute or with broadly rounded ends, and with size ranges from 15.3 to 17.5 µm (length) × 4.5 to 5.2 µm (width) (Fig. 1B). The morphological characteristics of the present isolates were consistent with those of Colletotrichum gloeosporioides (Weir et al. 2012). A single isolate, ANUK97, was selected for identification. The multilocus sequence analysis (MLSA) of the actin (ACT), calmodulin (CAL), glyceraldehyde-3-phosphate dehydrogenase (GAPDH), internal transcribed spacer (ITS) rDNA, and β-tubulin (Tub2) were amplified by PCR with the primer pairs of ACT-521F/ACT-783R, CL1C/CL2C, GDF/GDR, ITS1/ITS4, and T1/T2, respectively (White et al. 1990). The resulting sequences were deposited in GenBank under accession numbers MW580367 (ACT), MW580368 (CAL), MW580369 (GAPDH), MW580370 (ITS), and MW580371 (TUB2). A nucleotide BLAST search revealed that ACT, CAL, GAPDH, ITS, and TUB2 sequences be 99% similar to accession numbers MN307380.1, MH155176.1, MK796226.1, MW580370.1, and JX010377.1, respectively of C. theobromicola. Maximum likelihood (ML) phylogenetic analysis was conducted based on a combined dataset of ACT, CAL, GAPDH, ITS, and TUB2 sequences using MEGA-X 10.1.8. The isolate ANUK97 was clustered with a representative strain C. theobromicola CBS124945 100% bootstrap support (Fig. 2). For the pathogenicity test, two-month-old cotton seedlings (n = 10) were inoculated with conidial suspensions (10⁶ spore/mL) of C. theobromicola obtained from 7-day-old PDA cultures at 25 °C by spray method. Seedlings treated with sterile distilled water served as controls. Inoculated and control cotton plants were incubated in the greenhouse at 25 °C under a 12-h photoperiod. After 7 days, necrotic lesions were observed on the artificially inoculated cotton plants, while control plants did not develop any disease symptoms. The pathogen was re-isolated from infected cotton leaves, but not from control plants to fulfill Koch’s postulates. To our knowledge, this is the first report of anthracnose of cotton caused by Colletotrichum theobromicola in Korea.

scholarly journals First Report of Powdery Mildew (Pseudoidium neolycopersici) on Croton (Codiaeum variegatum var. pictum) in China

Plant Disease ◽  
2015 ◽  
Vol 99 (2) ◽  
pp. 288-288 ◽  
Author(s):  
X.-M. Liu ◽  
Y.-X. Wei ◽  
H. Zhang ◽  
F.-X. Zhou ◽  
J.-J. Pu
Keyword(s):  
Powdery Mildew ◽  
Southern China ◽  
Disease Incidence ◽  
Morphological Characteristics ◽  
Oidium Neolycopersici ◽  
Tomato Plants ◽  
First Report ◽  
Pseudoidium Neolycopersici ◽  
Codiaeum Variegatum ◽  
Pathogenicity Tests

Croton (Codiaeum variegatum (Linn.) var. pictum (Lodd.)) is an ornamental plant commonly grown in southern China. In March 2014, severe powdery mildew infections were observed on crotons in gardens of Hainan University (20.1°N and 110.3°E), Haikou, Hainan province. Disease incidence was estimated in a random batch of 100 plants in three replicates, with the average value approaching 80%. Symptoms first appeared as white circular patches on the adaxial surface and expanded to the abaxial surface, petioles, and stems. The top leaves were the most affected. Upper surfaces of the infected leaves were covered by white, dense mycelia. As the disease progressed, infected leaves turned purple on the lower surfaces and curly before becoming necrotic and abscising from the plant. Powdery mildew was more severe in shaded environments, especially during rainy or foggy weather in early spring. Two hundred conidiophores and conidia were observed microscopically. The conidiophores were straight or occasionally flexuous, 62.3 to 127.6 × 6.2 to 10.2 μm, consisting of two to three straight cells. Conidia were born in solitary on the top of conidiophores. Conidia were hyaline, ellipsoidal, 26.4 to 42.2 × 11.7 to 23.4 μm (average 32.5 × 16.5 μm), contained no distinct fibrosin bodies, and produced a subterminal germ tube. The wrinkling pattern of the outer walls of older conidia was angular or reticulated. Appressoria were single and multilobed. Cleistothecia were not observed. Based on morphological characteristics, the fungus was identified as Oidium neolycopersici (2), which was recently renamed Pseudoidium neolycopersici (L. Kiss) (3). The identity was confirmed by sequence analysis. Genomic DNA was extracted from the foliar powdery mildew colonies using Chelex-100 (Bio-Rad, Shanghai, China). The rDNA internal transcribed spacer (ITS) region was amplified with primers ITS1 and ITS4 (5). The ITS sequence of the representative isolates C01 (GenBank Accession No. KJ890378.1) and four other powdery mildew samples collected from crotons in Hainan University was 100% identical to that of P. neolycopersici isolates from tomato plants such as JQ972700 and AB163927. Inoculations were made by gently pressing diseased leaves onto leaves of five healthy plants of croton and tomato (‘Money maker’). Five non-inoculated croton and tomato plants served as controls. Inoculated and non-inoculated plants were maintained in an incubator at 25°C with a 12-h photoperiod. After eight days, typical powdery mildew symptoms developed on 93% of the inoculated plants, while no symptom developed on the non-inoculated plants. The pathogenicity tests were repeated three times. The same fungus was always re-isolated from the diseased tissue according to Koch's postulates. The pathogenicity tests further confirmed that the pathogen from crotons is P. neolycopersici (Basionym. Oidium neolycopersici (KJ890378.1)), which is commonly known as the tomato powdery mildew. P. neolycopersici is also a pathogen of Normania triphylla (1) and papaya (4). To our knowledge, this is the first report of P. neolycopersici infecting croton. The avenue of this pathogen entering gardens of Hainan University remains unknown. The gardens are located far away from tomato farms. Also no symptom of powdery mildew on croton was observed during surveys in other locations in Haikou. The origin of the pathogen warrants additional research. References: (1) D. Delmail et al. Mycotaxon 113:269, 2010. (2) L. Kiss et al. Mycol. Res. 105:684, 2001. (3) L. Kiss et al. Mycol. Res. 115:612, 2011. (4) J. G. Tsay et al. Plant Dis. 95:1188, 2011. (5) T. J. White et al. Page 315 in: PCR Protocols: A Guide to Methods and Applications. Academic Press, San Diego, CA, 1990.

scholarly journals First report of leaf spot disease caused by Colletotrichum siamense on Cornus hongkongensis (Hemsl.) in China

Plant Disease ◽  
2021 ◽  
Author(s):  
Qing-Hai Wang ◽  
Yang Zhang ◽  
Yu-tong Zhang ◽  
Dong Li ◽  
Xiao-li Lin ◽  
...  
Keyword(s):  
Phylogenetic Analysis ◽  
Disease Incidence ◽  
Morphological Characteristics ◽  
Control Measures ◽  
Bootstrap Support ◽  
Leaf Spot Disease ◽  
Pathogenicity Test ◽  
Accurate Identification ◽  
First Report ◽  
Colletotrichum Siamense

Cornus hongkongensis (Hemsl.) is an excellent ornamental tree species in China and elsewhere. In 2019, C. hongkongensis anthracnose was firstly observed at the campus of Jiangxi Agricultural University (JXAU) (28°45′56″N, 115°50′21″E), then found in parks, Nanchang, China. In early August, the disease appeared and lasted until the leaves dropped (November). The disease incidence was above 60%, and the diseased leaf rate was above 70%. The lesions mostly appeared along the leaf edges. Some small round to irregular lesions also developed in other parts of the leaves. These diseased leaves had circular or irregularly shaped spots with gray-white color in the center and dark brown on the edge of the lesions. Later, the lesions became necrotic and shriveled. As the disease progressed, the spots coalesced so that affected leaves appeared blighted (Supplementary Figure 1 A-C). To identify the pathogen, leaves with typical symptoms from the campus of JXAU were collected and small pieces (5 × 5 mm) from the lesion borders were surfaced sterilized in 70% ethanol for 30 s, followed by 1 min in 3% NaOCl, and then rinsed with sterile distilled water three times. Leaf pieces were placed on potato dextrose agar (PDA) and incubated at 25 °C under a 12-h light/dark cycle (3000 lx). Pure cultures were obtained from individual conidia by single spore isolates. For studies of microscopic morphology, a representative isolate JX-S4 was subcultured on PDA. The colony of JX-S4 was white and turning gray and light gray on the reverse side, producing dark-green pigmentation near the center (Supplementary Figure 1 D). The conidia were one-celled, straight, hyaline, subcylindrical with rounded ends and 16.9 ± 1.6 × 6.0 ± 0.6 µm (n = 50) in size. Appressoria were one-celled, pale brown, thick-walled, ellipsoidal, and measured 8.7 ± 1.7 × 6.4 ± 0.8 µm (n = 50) (Supplementary Figure 1 E, F). The morphological characteristics of JX-S4 matched those of the Colletotrichum siamense species (Weir et al. 2012). For accurate identification, the internal transcribed spacer (ITS) and the genes encoding glyceraldehyde-3-phosphate dehydrogenase (GAPDH), chitin synthase (CHS-I), beta-tubulin 2 (TUB2), and calmodulin (CAL) were respectively amplified with primers ITS1/ITS4, GDF/GDR, CHS-79F/CHS-345R, βt2a/βt2b, and CL1/CL2. The sequences were deposited in GenBank (Accession nos. MT587807, MT628710, MT628709, MT628711, and MT628708). Phylogenetic analysis was calculated with concatenated sequences (ITS, GAPDH, CHS-I, CAL, and TUB2) using MEGA 7. In the maximum likelihood phylogenetic tree, Isolate JX-S4 was clustered with C. siamense with 93% bootstrap support (Supplementary Figure 2). Based on the morphological characteristics and phylogenetic analysis, JX-S4 was identified as C. siamense. Pathogenicity test of JX-S4 was verified on 45 attached healthy leaves from three C. hongkongensis plants (10-year-old) at the campus of JXAU inoculated with mycelial plugs (φ=5 mm) from the culture edge (6-day-old) on PDA. And an additional 45 healthy leaves were inoculated with PDA plugs as controls. The leaves were wounded with a red-hot needle (φ=0.5 mm). All treatment and control leaves were wrapped up with black plastic bags to keep them moist for 2 days. The pathogenicity tests were repeated twice. Within 7 days, all the inoculated leaves developed the lesions, which were similar to those observed in the field. Control leaves were asymptomatic (Supplementary Figure 1 G, H). The same fungus was re-isolated from the symptomatic tissues, fulfilling Koch’s postulates. To our knowledge, this is the first report of C. siamense causing C. hongkongensis anthracnose. This finding provides crucial information for managing this disease. For example, when diagnosing Cornus anthracnose, C. siamense needs to be looked out for and appropriate control measures implemented.

scholarly journals First Report of Leaf Spot Caused by Alternaria brassicae on Avena sativa in China

Plant Disease ◽  
2021 ◽  
Author(s):  
Jiangui Zhang ◽  
Xiumei Nie ◽  
Guiqin Zhao
Keyword(s):  
Avena Sativa ◽  
Leaf Spot ◽  
Unsaturated Fatty Acids ◽  
Disease Incidence ◽  
Morphological Characteristics ◽  
Northern China ◽  
Green Water ◽  
Bootstrap Support ◽  
Pathogenicity Test ◽  
First Report

Oat (Avena sativa) is an annual gramineous crop, which contains a source of soluble dietary fiber, β-glucan, unsaturated fatty acids, vitamins, minerals, phenolic acids and avenanthramides. It widely cultivated in cool and semi-arid areas in northern China (Li et al, 2017). In July 2018, a severe leaf spot infection was observed in the Forage Germplasm Nursery (31°17′22″N, 103°40′15″E, 2885 m elevation) in Tianzhu County, Wuwei City of Gansu Province in China. Disease incidence (total number of diseased leaves / total number of surveyed leaves X 100%) was 93% over 300 m2 planting area. Symptoms initially appeared as small circular to irregular, gray-green, water-soaked spots on the leaves in the middle or along the margin of leaves, that enlarged and coalesced. The center of the leaf spots turned brown to reddish-brown. Infected tissues from symptomatic leaves were cut into small pieces (5×5 mm), surface sterilized with 70% ethanol for 60 s, soaked in 5% commercial bleach (~0.275% NaClO) for 5 min (Xue et al, 2018), rinsed five times with distilled water, plated on potato dextrose agar (PDA) medium, and incubated for 3 days in the dark at 25°C (Zhang, 2003; Blagojević et al, 2020; Humpherson et al, 1989). Hyphae emerging from the tissue were subcultured on fresh PDA medium for purification. All colonies were light brown with intensive sporulation in rings that was grayish white, and later became grayish brown. The back of the colony was dark brown. Conidiophores were light brown, unbranched, grew vertically on hyphae, and each conidiophore produced 3 to 7 conidia (mostly 6). Conidia were light brown, septate, straight to slightly curved, single or in chains, oval or obclavate, measured 17 to 32 µm wide and 63 to 106 µm long with the conical beak cell, 7 to 12 transverse septa, 0 to 5 longitudinal septa. These morphological characteristics were similar to the descriptions of Alternaria spp. (Simmons, 2008). A single isolate, YMZZ1, was selected for molecular identification. The ITS region of rDNA, partial GAPDH and Tef1-α gene sequences were amplified by PCR with the primer pairs of ITS1/ITS4, gpd1/gpd2 and EF1-728F/EF1-986R, respectively (Woudenberg et al, 2013). Sequences were deposited in GeneBank under accessions MN446739 (ITS), MN481462 (GAPDH), and MN464104 (Tef1-α). A nucleotide BLAST search revealed ITS, GAPDH and Tef1-α sequences to be 99% similar to accessions numbers MN856410 (565/573 bp), MK026431 (575/575 bp), and MH754531 (211/211 bp), respectively) of A. brassicae. Neighbor-joining (NJ) and Maximum Likelihood (ML) phylogenetic analysis were conducted based on ITS, GAPDH and TEF-1α sequences using MEGA7.0 under Kimura 2-parameter model. The isolate YMZZ1 clustered with a representative strain A. brassicae LGBA22 with 100% bootstrap support. To test its pathogenicity, six healthy 3 week old plants were spray-inoculated with a suspension of 3×105 conidia/mL of YMZZ1. The same number of plants were sprayed with sterilized water as control. All plants were covered with transparent plastic bags for 48 h to maintain high relative humidity and incubated in a 25°C growth chamber (16/8 h light/dark) for observation. Ten days after inoculation, leaf spot symptoms were observed on leaves similar to those previously observed in nursery; no symptoms were observed on the control. The pathogenicity test was repeated twice under the same conditions and A. brassicae was re-isolated from inoculated plants each time fulfilling Koch’s postulates. A. brassicae has not been previously reported as a pathogen of A. sativa in the world, but has been mentioned as a pathogen of horse radish (Armoracia rusticana) in Serbia (Blagojevic et al, 2015). To our knowledge, this is the first report of leaf spot caused by A. brassicae on A. sativa in China. This study stresses an urgent need to identify appropriate management strategies of A. brassicae that help in preventing losses in quality and yield of oats in northern China.

scholarly journals First Report of Blackleg of Ajuga multiflora Caused by Phoma multirostrata in China

Plant Disease ◽  
2008 ◽  
Vol 92 (6) ◽  
pp. 981-981 ◽  
Author(s):  
B. G. Lou ◽  
X. Q. Hu ◽  
L. Wu
Keyword(s):  
Southern China ◽  
Disease Incidence ◽  
Morphological Characteristics ◽  
Continuous Cultivation ◽  
Multiple Infections ◽  
First Report ◽  
The Past ◽  
Landscape Plants ◽  
Ajuga Multiflora ◽  
Time Of Year

Ajuga multiflora Bunge (Labiatae) is an ornamental that is widely planted in gardens in southern China. Blackleg symptoms were observed on A. multiflora during the autumn of 2005 in the Hangzhou area, Zhejiang Province. A. multiflora had to be replaced by other landscape plants during the past 2 years because of a serious outbreak of the disease. When the plant is in continuous cultivation for more than 2 years, disease incidence is more than 80%. Brown water-soaked spots appeared initially on the base of the petioles. Blackleg lesions from multiple infections may coalesce. Lesions often expand rapidly along the petioles followed by wilting and drying of individual leaves, ultimately resulting in the death of plants. The disease can occur at any time of year, but is most severe during the flowering stage. A fungus was consistently isolated from symptomatic tissues on potato dextrose agar. After 14 days on SNA (synthetic low nutrient agar; 1.0 g of KH2PO4, 1.0 g of KNO3, 0.5 g of MgSO4·7H2O, 0.5 g of KCl, 0.2 g of glucose, 0.2 g of sucrose, and 20 g of agar per liter) medium, the fungus produced light olive-to-gray mycelium, and later, generated brown, globose to subglobose pycnidia with hyaline, unicellular, and ellipsoidal conidia. Diameter of pycnidia ranged from 163 to 260 μm (average 203 μm). Conidiophores measured 3.5 to 6.2 × 2.0 to 3.2 μm (average 5.4 × 2.3 μm). The fungus was identified as Phoma multirostrata (P.N. Mathur, S.K. Menon & Thirum) Dorenb. & Boerema based on its morphological characteristics (1). The fungus grew between 4 and 36°C and optimum growth was at 23°C. To fulfill Koch's postulates, 15 healthy 20-day-old A. multiflora seedlings were planted in sterile soil artificially infested with P. multirostrata, which was grown on a mixture (1:10 w/w) of corn meal and sand. Soil was amended with 20% (w/w) of inoculum mixture. Another 15 plants grown in noninfested soil served as controls. All plants were grown at 22 to 25°C in the greenhouse. After 5 to 7 days, inoculated plants showed symptoms closely resembling those seen on plants from the garden. P. multirostrata was isolated from the margins of necrotic tissues on diseased plants, but not from the control plants. To our knowledge, this is the first report of P. multirostrata causing blackleg of A. multiflora in China. Reference: (1) B. C. Sutton. The Coelomycetes. CMI, Kew, Surrey, UK, 1980.

scholarly journals First report of Coniella castaneicola causing leaf blight on blueberry (Vaccinium virgatum) in China

Plant Disease ◽  
2021 ◽  
Author(s):  
Jiahao Lai ◽  
Guihong Xiong ◽  
Bing Liu ◽  
Weigang Kuang ◽  
Shuilin Song
Keyword(s):  
Molecular Identification ◽  
Morphological Characteristics ◽  
Leaf Blight ◽  
Yield Potential ◽  
Effective Control ◽  
Economic Losses ◽  
Distilled Water ◽  
First Report ◽  
Fungal Isolates ◽  
Blight Disease

Blueberry (Vaccinium virgatum), an economically important small fruit crop, is characterized by its highly nutritive compounds and high content and wide diversity of bioactive compounds (Miller et al. 2019). In September 2020, an unknown leaf blight disease was observed on Rabbiteye blueberry at the Agricultural Science and Technology Park of Jiangxi Agricultural University in Nanchang, China (28°45'51"N, 115°50'52"E). Disease surveys were conducted at that time, the results showed that disease incidence was 90% from a sampled population of 100 plants in the field, and this disease had not been found at other cultivation fields in Nanchang. Leaf blight disease on blueberry caused the leaves to shrivel and curl, or even fall off, which hindered floral bud development and subsequent yield potential. Symptoms of the disease initially appeared as irregular brown spots (1 to 7 mm in diameter) on the leaves, subsequently coalescing to form large irregular taupe lesions (4 to 15 mm in diameter) which became curly. As the disease progressed, irregular grey-brown and blighted lesion ran throughout the leaf lamina from leaf tip to entire leaf sheath and finally caused dieback and even shoot blight. To identify the causal agent, 15 small pieces (5 mm2) of symptomatic leaves were excised from the junction of diseased and healthy tissue, surface-sterilized in 75% ethanol solution for 30 sec and 0.1% mercuric chloride solution for 2 min, rinsed three times with sterile distilled water, and then incubated on potato dextrose agar (PDA) at 28°C for 5-7 days in darkness. Five fungal isolates showing similar morphological characteristics were obtained as pure cultures by single-spore isolation. All fungal colonies on PDA were white with sparse creeping hyphae. Pycnidia were spherical, light brown, and produced numerous conidia. Conidia were 10.60 to 20.12 × 1.98 to 3.11 µm (average 15.27 × 2.52 µm, n = 100), fusiform, sickle-shaped, light brown, without septa. Based on morphological characteristics, the fungal isolates were suspected to be Coniella castaneicola (Cui 2015). To further confirm the identity of this putative pathogen, two representative isolates LGZ2 and LGZ3 were selected for molecular identification. The internal transcribed spacer region (ITS) and large subunit (LSU) were amplified and sequenced using primers ITS1/ITS4 (Peever et al. 2004) and LROR/LR7 (Castlebury and Rossman 2002). The sequences of ITS region (GenBank accession nos. MW672530 and MW856809) showed 100% identity with accessions numbers KF564280 (576/576 bp), MW208111 (544/544 bp), MW208112 (544/544 bp) of C. castaneicola. LSU gene sequences (GenBank accession nos. MW856810 to 11) was 99.85% (1324/1326 bp, 1329/1331 bp) identical to the sequences of C. castaneicola (KY473971, KR232683 to 84). Pathogenicity was tested on three blueberry varieties (‘Rabbiteye’, ‘Double Peak’ and ‘Pink Lemonade’), and four healthy young leaves of a potted blueberry of each variety with and without injury were inoculated with 20 μl suspension of prepared spores (106 conidia/mL) derived from 7-day-old cultures of LGZ2, respectively. In addition, four leaves of each variety with and without injury were sprayed with sterile distilled water as a control, respectively. The experiment was repeated three times, and all plants were incubated in a growth chamber (a 12h light and 12h dark period, 25°C, RH greater than 80%). After 4 days, all the inoculated leaves started showing disease symptoms (large irregular grey-brown lesions) as those observed in the field and there was no difference in severity recorded between the blueberry varieties, whereas the control leaves showed no symptoms. The fungus was reisolated from the inoculated leaves and confirmed as C. castaneicola by morphological and molecular identification, fulfilling Koch’s postulates. To our knowledge, this is the first report of C. castaneicola causing leaf blight on blueberries in China. The discovery of this new disease and the identification of the pathogen will provide useful information for developing effective control strategies, reducing economic losses in blueberry production, and promoting the development of the blueberry industry.

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