First Report of Diaporthe amygdali Associated with Twig Canker and Shoot Blight of Nectarine in Spain

Nectarine (Prunus persica (L.) Batsch var. nucipersica (Suckow) C. K. Schneid.) is a fruit crop widely cultivated throughout the Mediterranean basin. In Spain, it is mainly grown in eastern regions of the country. In March 2018, 5-year-old nectarine trees showing twig canker symptoms were observed after a rainy spring period in a 0.5 ha orchard located at Alaior, Menorca island (Spain). Cankers were frequent on affected trees (approximately, 80% of the total trees), thus leading to shoot blight. Ten twig segments of one-year old wood with cankers were cut, washed under running tap water, surface disinfected for 1 min in a 1.5% sodium hypochlorite solution and rinsed twice in sterile distilled water. Small pieces (2 mm) of affected tissues were taken from the margin of the cankers and plated on potato dextrose agar (PDA) supplemented with 0.5 g/L of streptomycin sulphate (PDAS). The plates were then incubated at 25 ºC in the dark for 7 to 10 d. Actively growing colonies were first hyphal-tipped and then transferred to PDA and 2% water agar supplemented with sterile pine needles and incubated at 21-22ºC under a 12h/12h near UV / darkness cycle during 21 d (León et al. 2020). Colonies were white at first, becoming light cream, with visible solitary and aggregate pycnidia at maturity. Alpha conidia were aseptate, fusiform, hyaline, multi-guttulated (mean ± SD = 7.4 ± 0.7 × 2.8 ± 0.4 µm, n = 100). Beta and gamma conidia were not observed. The morphological and cultural characteristics of the isolates were congruent with those of Diaporthe spp. (Gomes et al. 2013). The ITS1-5.8S-ITS2 (ITS) region and fragments of β-tubulin (tub2), the translation elongation factor 1-alpha (tef1-α) gene regions, histone H3 (his3) and calmodulin (cal) genes of representative isolate DAL-59 were amplified and sequenced (Santos et al. 2017). The BLASTn analysis revealed 100% similarity with sequences of D. mediterranea (Synonym D. amygdali) (Hilário et al. 2021) isolate DAL-34 from almond (ITS: MT007489, tub2: MT006686, tef1-α: MT006989, his3: MT007095 and cal: MT006761). Sequences of isolate DAL-59 were deposited in GenBank Database (ITS: MT007491, tub2: MT006688, tef1-α: MT006991, his3: MT007097 and cal: MT006763). Pathogenicity tests were conducted using one-year-old potted plants of nectarine cv. Boreal, which were inoculated with isolate DAL-59. In each plant, a 3 mm wound was made in the center of the main branch (about 30 cm length) with a scalpel. Colonized agar plugs with 3 mm diameter, which were obtained from active 10-day-old colonies growing on PDA, were inserted underneath the epidermis and the wounds sealed with Parafilm. Inoculated plants were incubated in a growth chamber at 23 ºC with 12 h of light per day. Controls were inoculated with uncolonized PDA plugs. There were twelve plants per treatment, which were arranged in a completely randomized design. Five days after inoculation necrosis development was observed in the area of inoculation. Wilting and twig blight symptoms over the lesion occurred 3-wk after inoculation and pycnidia were detected, while the controls remained asymptomatic. Diaporthe amygdali was re-isolated from symptomatic tissues and identified as described above to satisfy Koch’s postulates. To our knowledge, this is the first report of D. amygdali causing twig canker and shoot blight disease on nectarine in Spain.

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First report of soybean stem blight caused by Diaporthe phaseolorum in Sichuan province, China

Plant Disease ◽

10.1094/pdis-04-21-0697-pdn ◽

2021 ◽

Author(s):

Xiaolin Zhao ◽

Sujiao Zheng ◽

Wenwu Ye ◽

Xiaobo Zheng ◽

Yuanchao Wang

Keyword(s):

Phylogenetic Trees ◽

Elongation Factor ◽

Its Region ◽

Stem Canker ◽

Sichuan Province ◽

Nuclear Ribosomal Dna ◽

Sodium Hypochlorite Solution ◽

First Report ◽

Stem Blight ◽

Soybean Seedlings

In September 2020, a disease resembling stem blight on soybean was found in Chengdu city, Sichuan province, southwestern of China. Symptoms began as a brown lesion on the stems, usually at the nodes, then lesions expanded, darkened, and even girdled the stems, causing wilt of the above stems (Figure 1A). In three 0.33-ha fields, a total of 300 soybean plants (20 plants/site × 5 sites/filed × 3 fields) were investigated, and 3% of the plants showed the disease symptoms. The symptoms were consistent with those previously reported for stem canker and stem blight on soybean caused by Diaporthe complex (Cui et al. 2009; Mena et al. 2019; Santos et al. 2011). The tissues of symptomatic soybean stems were rinsed by water, disinfected by submerging them in 75% ethanol for 30 s and in 2% sodium hypochlorite solution for 2 min, then followed by washing with sterile distilled water. Small diseased tissue fragments were placed on selective potato dextrose agar (PDA) containing rifampicin and ampicillin (both 50 mg/μl). Plates were sealed and incubated at 26°C for two days. Developing mycelia of isolates were transferred to fresh PDA and purified by single hyphal tip. For the five obtained isolates (Figure 1B), five markers, including the internal transcribed spacer (ITS) region of the nuclear ribosomal DNA, parts of the translation elongation factor 1-α (TEF1), part of the histone H3 (HIS) gene, the calmodulin gene (CAL), and the beta-tubulin gene (TUB), were amplified using the established primers ITS4/ITS5 (White et al. 1990), EF1-728F/EF1-986R (Carbone and Kohn 1999), CYLH3F (Crous et al. 2006) and H3-1b (Glass and Donaldson 1995), CAL228F/CAL737R (Carbone and Kohn 1999), and Bt-2a/Bt-2b (Glass and Donaldson 1995), respectively, and sequenced (GenBank IDs: MW595761-MW595780 and MW624472-MW624476). Phylogenetic trees were constructed based on concatenated sequences of the five markers using the maximum-likelihood (ML) method in MEGA-5.2.2. Based the results of morphological (Figure 1C-E) and phylogenetic analysis (Figure 2), the five isolates were all identified as D. phaseolorum. Pathogenicity tests for the isolates were conducted on 7-day-old soybean seedlings (cv. Shangdou 1310) using a hypocotyl slit inoculation method. At the stem 2-cm below cotyledon, a 6-mm long slit was cut with a sterile scalpel, and placed inside with a 3 mm × 3 mm PDA plug with or without mycelia of pathogen. Ten plants were assayed for each treatment, and the plants were maintained in greenhouse at 26°C, with humidity higher than 90% for the first two days. The assay was repeated at least three times. Typical brown lesions on the stems were observed four days after inoculation (Figure 1F), even 20% treated plants died. The D. phaseolorum was reisolated from these stem lesions. No disease symptom was observed on control plants (Figure 1G). Thus D. phaseolorum was the pathogen causing the soybean stem blight in field. To our knowledge, this is the first report of D. phaseolorum causing this disease in Sichuan province, China. The result may provide useful information for soybean disease control in this region of China. The authors declare no conflict of interest. Funding: China Agriculture Research System (CARS-004-PS14).

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First Report of Xylella fastidiosa in Peach in New Mexico

Plant Disease ◽

10.1094/pdis-10-10-0719 ◽

2011 ◽

Vol 95 (7) ◽

pp. 871-871 ◽

Cited By ~ 1

Author(s):

J. J. Randall ◽

J. French ◽

S. Yao ◽

S. F. Hanson ◽

N. P. Goldberg

Keyword(s):

New Mexico ◽

Prunus Persica ◽

Xylella Fastidiosa ◽

Its Region ◽

23S Rrna ◽

Pcr Analysis ◽

Sequencing Analysis ◽

First Report ◽

Dark Green ◽

Branch Dieback

Xylella fastidiosa is a gram-negative bacterium that causes disease in a wide variety of plants such as grapes, citrus trees, oleanders, and elm and coffee trees. This bacterium is xylem limited and causes disease symptoms such as leaf scorch, stunting of plant growth, branch dieback, and fruit loss. The presence of X. fastidiosa was previously reported in New Mexico where it was found to be infecting chitalpa plants and grapevines (3). In the summer of 2010, peach (Prunus persica (L.) Batsch) trees from two locations in northern New Mexico exhibited leaf deformity and stunting, dark green venation, slight mottling, and branch dieback. Preliminary viral diagnostic screening was performed by Agdia (Elkhart, IN) on one symptomatic tree and it was negative for all viruses tested. Three trees from two different orchards tested positive for X. fastidiosa by ELISA and PCR analysis using X. fastidiosa-specific primer sets HL (1) and RST (2). Bacterial colonies were also cultured from these samples onto periwinkle wilt media. Eight colonies obtained from these three plants tested PCR positive using the X. fastidiosa-specific primers. The 16S ribosomal and 16S-23S rRNA internal transcribed spacer (ITS) region (557 nucleotides) (GenBank Accession No. HQ292776) along with the gyrase region (400 nucleotides) (GenBank Accession No. HQ292777) was amplified from the peach total DNA samples and the bacterial colonies. Sequencing analysis of these regions indicate that the X. fastidiosa found in peach is 100% similar to other X. fastidiosa multiplex isolates including isolates from peach, pecan, sycamore, and plum trees and 99% similar to the X. fastidiosa isolates previously found in New Mexico. Further analysis of the 16S ribosomal and 16S-23S rRNA ITS sequences with maximum likelihood phylogenetic analysis using Paup also groups the peach isolates into the X. fastidiosa multiplex subspecies. The gyrase sequence could not be used to differentiate the peach isolates into a subspecies grouping because of the lack of variability within the sequence. This X. fastidiosa multiplex subspecies could possibly be a threat to the New Mexico pecan industry since pecan infecting X. fastidiosa isolates belong to the same bacterial subspecies. It is not known if X. fastidiosa subspecies multiplex isolates from peach are capable of infecting pecans but they are closely genetically related. It is interesting to note that the isolates from peach are different than previously described X. fastidiosa isolates in New Mexico that were infecting chitalpa and grapes (3). X. fastidiosa has previously been described in peach; the disease is called “phony peach”. The peach trees exhibited stunting and shortened internodes as reported for “phony peach”. They also exhibited slight mottling and branch dieback that may be due to the environment in New Mexico or perhaps they are also exhibiting mineral deficiency symptoms in association with the X. fastidiosa disease. To our knowledge, this is the first report of X. fastidiosa in peach in New Mexico. References: (1) M. H. Francis et al. Eur. J. Plant Pathol. 115:203, 2006. (2) G. V. Minsavage et al. Phytopathology 84:456, 1994. (3) J. J. Randall et al. Appl. Environ. Microbiol. 75:5631, 2009.

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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 Euphorbia larica Dieback Caused by Fusarium brachygibbosum in Oman

Plant Disease ◽

10.1094/pdis-09-12-0828-pdn ◽

2013 ◽

Vol 97 (5) ◽

pp. 687-687 ◽

Cited By ~ 9

Author(s):

I. H. Al-Mahmooli ◽

Y. S. Al-Bahri ◽

A. M. Al-Sadi ◽

M. L. Deadman

Keyword(s):

Elongation Factor ◽

Aerial Mycelium ◽

Its Region ◽

Common Component ◽

First Report ◽

Ethnobotanical Uses ◽

Artificial Inoculations ◽

Stem Lesions ◽

Desert Flora

Euphorbia larica Boiss. (Arabic = Isbaq) is a dominant and common component of the native desert flora of northern Oman. Traditional ethnobotanical uses have included use of the latex for treating camels with parasites. In February 2011, E. larica plants showing stem lesions up to several cm long and in many cases with stem dieback were collected from Al-Khoudh 50 km west of Muscat. The disease appeared widespread within the location where several dead specimens were also recorded, although the cause was unclear. Sections (5 mm) of five diseased branches taken from different plants and placed on potato dextrose agar (PDA) in all cases yielded Fusarium-like colonies. Colonies recovered were initially white becoming rose to medium red in color with abundant aerial mycelium. Macroconidia were scarce and scattered (mean of 20 spores: 26.83 × 4.73 μm) with three to four septa per spore; microconidia were slightly curved, ovoid, and fusiform (mean of 20 spores: 11.64 × 4.03 μm) with zero to two septa per spore. Spherical chlamydospores (mean of 20 spores: 11.05 μm) were terminal and intercalary, single, and in chains. In vitro characters and spores measurements conformed to previously described features of Fusarium brachygibbosum Padwick (1). Mycelial plugs (5 mm) were taken from 7-day-old cultures of the fungus grown on 2.5% PDA and applied to a small incision (3 mm) on the stems of healthy E. larica grown in situ and protected with wet cotton and Parafilm. The residual agar, mycelium, cotton, and Parafilm were removed after 7 days and symptoms were recorded. Control stems were inoculated using PDA (5 mm) plugs alone and inoculations were repeated twice. Artificial inoculations resulted in dieback of all stems within 11 days and fungal colonies identical to initial isolations were recovered from artificially infected surface-sterilized stem pieces. Identification of F. brachygibbosum was confirmed by comparing sequences generated from the internal transcribed spacer (ITS) region of the ribosomal DNA (ITS1 and ITS4 primers) and the intron region of translation elongation factor alpha (EF1-α) (EF-1-986 and EF-728 primers). The ITS and EF1-α sequences were found to share 100% and 99% nucleotide similarity to previously published sequences of the ITS (HQ443206) and EF1-α (JQ429370) regions of F. brachygibbosum in GenBank. The accession number of ITS sequence of one isolate assigned to EMBL-Bank was HF562936. The EF sequence was assigned to EMBL-Bank accession (submission number Hx2000027017; number will be sent later). This pathogen has previously been reported on date palm (2) in Oman but, to our knowledge, this is the first report of this pathogen on E. larica. References: (1) A. M. Al-Sadi et al. Crop Prot. 37:1, 2012. (2) G. W. Padwick. Mycol. Pap. 12:11, 1945.

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First Report of Fruit Rot Disease on Pomelo Caused by Lasiodiplodia theobromae in China

Plant Disease ◽

10.1094/pdis-03-11-0214 ◽

2011 ◽

Vol 95 (9) ◽

pp. 1190-1190 ◽

Cited By ~ 5

Author(s):

M. Luo ◽

Z. Y. Dong ◽

S. Y. Bin ◽

J. T. Lin

Keyword(s):

Its Region ◽

Fruit Rot ◽

Economic Losses ◽

Single Spore ◽

Lasiodiplodia Theobromae ◽

First Report ◽

Potted Plants ◽

Diseased Fruit ◽

Long Time ◽

Water Plants

Pomelo (Citrus grandis) is widely cultivated in MeiZhou Guangdong Province of China. In 2008, a disease on pomelo fruit caused significant economic losses by affecting fruit quality. Diseased fruit was collected in December 2008 from MeiZhou Guangdong, surface sterilized in 75% ethanol for 1 min and internal necrotic tissue was transferred to potato dextrose agar (PDA) and incubated at 28°C for 5 days. Three single-spore isolates were obtained from different fruit and identified as Lasiodiplodia theobromae (Pat.) Griffon & Maubl. (synonyms Diplodia natalensis Pole-Evans and Botryodiplodia theobromae Pat.; teleomorph Botryosphaeria rhodina (Cooke) Arx) on the basis of morphological and physiological features. The fungus produced dark brown colonies (initially grayish) on PDA. Young hyphae were hyaline and aseptate, whereas mature hyphae were septate with irregular branches. Cultures of L. theobromae produced globular or irregular pycnidia abundantly on PDA (pH 3.5) at 28°C after 1 month. Mature conidia of L. theobromae were 20 to 26 × 12 to 15.5 μm, subovoid to ellipsoid-ovoid, initially hyaline and nonseptate, remaining hyaline for a long time, and finally becoming dark brown and one septate with melanin deposits on the inner surface of the wall arranged longitudinally giving a striate appearance to the conidia. The internal transcribed spacer (ITS) region of the rDNA was amplified from gDNA using primers ITS1 (5′-TCCGATGGTGAACCTGCGG-3′) and ITS4 (5′-TCCTCCGCTTATTGATATGC-3′) (1). Amplicons were 542 bp long (GenBank Accession No. JF693024) and had 100% nucleotide identity with the corresponding sequence (GenBank Accession No. EU860391) of L. theobromae isolated from a Pinus sp. (2). To satisfy Koch's postulates, six asymptomatic fruit on potted plants were sprayed until runoff with a spore suspension (1 × 106 spores/ml) prepared from 30-day-old cultures of one isolate. Control fruit received water. Plants were covered with sterile wet gauze to maintain high humidity. Fruit spot symptoms similar to those on diseased field fruit appeared after 15 days on all inoculated fruits. L. theobromae was reisolated from all inoculated test fruit. No symptoms were observed on the fruit of control plants. To our knowledge, this is the first report of L. theobromae causing disease on pomelo fruit in China. This pathogen has also been previously reported to be economically important on a number of other hosts by mostly affecting the leaves. References: (1) J. C. Batzer et al. Mycologia 97:1268, 2005. (2) C. A. Pérez et al. Fungal Divers. 41:53,2010.

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First Report of Powdery Mildew Caused by Erysiphe macleayae on Macleaya microcarpa in Poland

Plant Disease ◽

10.1094/pdis-03-12-0244-pdn ◽

2012 ◽

Vol 96 (9) ◽

pp. 1376-1376 ◽

Cited By ~ 3

Author(s):

M. J. Park ◽

S. E. Cho ◽

M. Piątek ◽

H. D. Shin

Keyword(s):

Powdery Mildew ◽

Sequence Similarity ◽

Signs And Symptoms ◽

Its Region ◽

Botanical Garden ◽

Central China ◽

Perennial Herb ◽

Width Ratio ◽

First Report ◽

Potted Plants

Macleaya microcarpa (Maxim.) Fedde, also known as smallfruit plume poppy, is a perennial herb belonging to the family Papaveraceae. The plant, together with the better-known species M. cordata (Willd.) R. Br., is native to central China and is now planted worldwide for medicinal purposes. In October 2008 and August 2009, dozens of smallfruit plume poppy planted in the Kraków Botanical Garden, Poland, were found to be severely infected with a powdery mildew. White colonies with abundant sporulation developed on both sides of leaves and young stems, forming circular to irregular patches. Infections caused leaf yellowing and premature defoliation. The damage has been observed every year since 2009. Representative voucher specimens were deposited in the fungal herbarium of the W. Szafer Institute of Botany of the Polish Academy of Sciences (KRAM) and the Korea University herbarium (KUS). Appressoria on the mycelia were lobed, often in pairs. Conidiophores composed of three to four cells arose from the upper part of creeping hyphae, 65 to 120 × 7 to 10 μm, attenuated toward the base, sub-straight or slightly flexuous in foot-cells, and produced conidia singly. Conidia were hyaline, oblong-elliptical to doliiform, 25 to 38 × 12 to 18 μm with a length/width ratio of 1.8 to 2.6; lacked fibrosin bodies; and produced germ tubes on the subterminal position with club-shaped or lobed appressoria. The conidial surface was wrinkled to irregularly reticulate. No chasmothecia were found. The structures described above match well with the anamorph of Erysiphe macleayae R.Y. Zheng & G.Q. Chen (3). To confirm the identity of the causal fungus, the internal transcribed spacer (ITS) region of rDNA from KUS-F24459 was amplified using primers ITS5 and P3 (4) and directly sequenced. The resulting sequence of 553 bp was deposited in GenBank (Accession No. JQ681217). A GenBank BLAST search using the present data revealed >99% sequence similarity of the isolate with E. macleayae on M. cordata from Japan (AB016048). Pathogenicity was confirmed through inoculation by gently pressing diseased leaves onto leaves of three healthy potted plants. Three noninoculated plants served as controls. Plants were maintained in a greenhouse at 25°C. Inoculated plants developed signs and symptoms after 7 days, whereas the control plants remained healthy. The fungus present on the inoculated plants was morphologically identical to that originally observed on diseased plants. The powdery mildew infections of M. cordata associated with E. macleayae have been recorded in China and Japan (2), and more recently in Germany (1,3). To our knowledge, this is the first report of E. macleayae on M. microcarpa globally as well as in Poland. This mildew species was described in China and is endemic to Asia, where chasmothecia of the fungus were found. Only recently have powdery mildews been found on M. cordata in Germany (1,3) and now on M. microcarpa in Poland, indicating the fungus is spreading in Europe. References: (1) N. Ale-Agha et al. Schlechtendalia 17:39, 2008. (2) D. F. Farr and A. Y. Rossman. Fungal Databases, Systematic Mycology and Microbiology Laboratory, ARS, USDA. Retrieved from http://nt.ars-grin.gov/fungaldatabases/ , February 7, 2012. (3) A. Schmidt and M. Scholler. Mycotaxon 115:287, 2011. (4) S. Takamatsu et al. Mycol. Res. 113:117, 2009.

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First Report of Powdery Mildew Caused by Podosphaera xanthii on Calendula officinalis in Italy

Plant Disease ◽

10.1094/pdis-92-1-0174c ◽

2008 ◽

Vol 92 (1) ◽

pp. 174-174 ◽

Cited By ~ 1

Author(s):

A. Garibaldi ◽

G. Gilardi ◽

M. L. Gullino

Keyword(s):

Powdery Mildew ◽

Its Region ◽

Pathogenicity Test ◽

Podosphaera Xanthii ◽

Calendula Officinalis ◽

First Report ◽

Potted Plants ◽

Blastn Analysis ◽

Voucher Specimens ◽

Pot Marigold

Calendula officinalis L. (Asteraceae) (pot marigold or English marigold) is an ornamental species grown in gardens and as potted plants for the production of cut flower. It was also used in ancient Greek, Roman, Arabic, and Indian cultures as a medicinal herb as well as a dye for fabrics, foods, and cosmetics. During the summer of 2007, severe outbreaks of a previously unknown powdery mildew were observed on plants in several gardens near Biella (northern Italy). Both surfaces of leaves of infected plants were covered with dense, white mycelia and conidia. As the disease progressed, infected leaves turned yellow and died. Mycelia and conidia also were observed on stems and flower calyxes. Conidia were hyaline, ellipsoid, born in short chains (four to six conidia per chain), and measured 27.0 to 32.1 (31.4) × 12.9 to 18.4 (18.2) μm. Conidiophores measured 49 to 77.3 (67.2) × 8 to 13.3 (10.8) μm and showed a foot cell measuring 44 to 59 (51.9) × 9.3 to 12.6 (11.3) μm followed by one shorter cell measuring 15.6 to 18.9 (17.6) × 10.4 to 13.6 (12.2) μm. Fibrosin bodies were present. Chasmothecia were spherical, amber colored, with a diameter of 89 to 100 (94.5) μm. Each chasmothecium contained one ascus with eight ascospores. On the basis of its morphology, the causal agent was determined to be a Podosphaera sp. (2). The internal transcribed spacer (ITS) region of rDNA was amplified using the primers ITS4/ITS6 and sequenced. BLASTn analysis (1) of the 588 bp showed a 100% homology with the sequence of Podosphaera xanthii (2). The nucleotide sequence has been assigned GenBank Accession No. EU100973. Pathogenicity was confirmed through inoculations by gently pressing diseased leaves onto leaves of healthy C. officinalis plants. Five plants were inoculated. Five noninoculated plants served as control. Plants were maintained in a greenhouse at temperatures ranging from 20 to 26°C. Eleven days after inoculation, typical symptoms of powdery mildew developed on inoculated plants. Noninoculated plants did not show symptoms. The pathogenicity test was carried out twice. To our knowledge, this is the first report of powdery mildew on C. officinalis in Italy. C. officinalis was previously described as a host to Sphaerotheca fuliginea (synonym S. fusca) in Great Britain (4) as well as in Romania (3). Voucher specimens are available at the AGROINNOVA Collection, University of Torino. References: (1) S. F. Altschul et al. Nucleic Acids Res. 25:3389, 1997. (2) U. Braun and S. Takamatsu. Schlechtendalia 4:1, 2000. (3) E. Eliade. Rev. Appl. Mycol. 39:710, 1960. (4) F. J. Moore. Rev. Appl. Mycol. 32:380, 1953.

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First Report of Diaporthe australafricana Associated with Stem Canker on Blueberry in Chile

Plant Disease ◽

10.1094/pdis-12-11-1025-pdn ◽

2012 ◽

Vol 96 (5) ◽

pp. 768-768 ◽

Cited By ~ 3

Author(s):

B. A. Latorre ◽

K. Elfar ◽

J. G. Espinoza ◽

R. Torres ◽

G. A. Díaz

Keyword(s):

Vitis Vinifera ◽

Vascular Tissue ◽

Its Region ◽

Pathogenic Fungi ◽

Stem Canker ◽

Vaccinium Corymbosum ◽

Academic Press ◽

First Report ◽

Potted Plants ◽

Blastn Analysis

Stem cankers of blueberry (Vaccinium corymbosum L.) have been observed on as much as 15% of the plants in plantations in central and southern Chile since 2006. Symptoms consisted of apical necrosis of the shoots and brown-to-reddish necrotic lesions on the stems. Internally, a brown-to-reddish discoloration of the vascular tissue can be observed. Twenty, single-plant samples were collected in 12 blueberry plantings (approximately 33°27′ to 40°53′S). Isolations from the margins of the necrotic lesions on the stems were made by plating small pieces (5 mm) on potato dextrose agar acidified with 0.5 μl/ml of 92% lactic acid (APDA). The plates were incubated at 20°C for 5 to 7 days, and hyphal tips of white colonies with septate and hyaline mycelium were transferred to APDA. Colonies were then transferred to autoclaved Pinus radiata needles on 2% water agar and incubated for 20 days at 20°C. Twelve isolates producing black pycnidia and alpha conidia were tentatively identified as a Phomopsis sp. (teleomoph Diaporthe Nitschke). Other fungi, including Botryosphaeriaceae spp. and Pestalotiopsis spp., were also isolated. Alpha conidia were smooth, unicellular, hyaline, fusoid, biguttulate, and 6.4 to 7.9 × 2.3 to 3.3 μm (n = 20). Beta conidia were not observed. The internal transcribed spacer (ITS) region of the rDNA was amplified using primers ITS1 and ITS2 (4) and sequenced. BLASTn analysis of the 473-bp fragment (GenBank Accession No. JQ045712) showed 100% identity to Diaporthe australafricana Crous & J.M. van Niekerk from Vitis vinifera (3). The pathogenicity of D. australafricana was studied on blueberry cv. O'Neal using detached stems (n = 4) in the laboratory, on 2-year-old potted plants (n = 4) in a greenhouse, and on attached stems of mature plants (n = 4) established in the ground. Inoculations were done by placing mycelial plugs taken from 7-day-old APDA cultures in a 7-mm long incision made on the stems. Inoculations with sterile mycelium plugs served as negative controls. Inoculation sites were wrapped with Parafilm to avoid rapid dehydration. Dark brown, necrotic lesions on the internal tissues were obtained on all inoculated stems 15 days after inoculation. Mean lesion lengths were 18.0 ± 7.4 mm on detached stems, 7.8 ± 6.9 mm on stems of 2-year-old plants, and 7.3 ± 2.5 mm on mature plants in the field. No symptoms developed on control stems. Reisolations were successful in 100% of the inoculated stems and D. australafricana was confirmed by the presence of pycnidia and alpha conidia. To our knowledge, this is the first report of D. australafricana causing stem canker in V. corymbosum. Previously, this pathogen has been reported to be affecting Vitis vinifera in Australia and South Africa (3). These results do not exclude that other plant-pathogenic fungi may be involved in this syndrome (1,2). References: (1) J. G. Espinoza et al. Plant Dis 92:1407, 2008. (2) J. G. Espinoza et al. Plant Dis. 93:1187, 2009. (3) J. M. van Niekerk et al. Australas. Plant Pathol. 34:27, 2005. (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, NY, 1990.

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First report of Nigrospora sphaerica causing fruit dried-shrink disease in Akebia trifoliata from China

Plant Disease ◽

10.1094/pdis-11-20-2471-pdn ◽

2021 ◽

Author(s):

Xiujing Hong ◽

Shijia Chen ◽

linchao Wang ◽

Bo Liu ◽

Yuruo Yang ◽

...

Keyword(s):

Elongation Factor ◽

Morphological Characteristics ◽

Its Region ◽

Average Diameter ◽

Ethanol Solution ◽

Likelihood Method ◽

Conidial Suspension ◽

Multiple Sequence ◽

First Report ◽

Pure Cultures

Akebia trifoliata, a recently domesticated horticultural crop, produces delicious fruits containing multiple nutritional metabolites and has been widely used as medicinal herb in China. In June 2020, symptoms of dried-shrink disease were first observed on fruits of A. trifoliata grown in Zhangjiajie, China (110.2°E, 29.4°N) with an incidence about 10%. The infected fruits were shrunken, colored in dark brown, and withered to death (Figure S1A, B). The symptomatic fruits tissues (6 × 6 mm) were excised from three individual plants, surface-disinfested in 1% NaOCl for 30s and 70% ethanol solution for 45s, washed, dried, and plated on potato dextrose agar (PDA) containing 50 mg/L streptomycin sulfate in the dark, and incubated at 25℃ for 3 days. Subsequently, hyphal tips were transferred to PDA to obtain pure cultures. After 7 days, five pure cultures were obtained, including two identical to previously reported Colletotrichum gloeosporioides causing leaf anthracnose in A. trifoliata (Pan et al. 2020) and three unknown isolates (ZJJ-C1-1, ZJJ-C1-2, and ZJJ-C1-3). The mycelia of ZJJ-C1-1, ZJJ-C1-2 and ZJJ-C1-3 were white, and formed colonies of approximate 70 mm (diameter) in size at 25℃ after 7 days on potato sucrose agar (PSA) plates (Figure S1C). After 25 days, conidia were formed, solitary, globose, black, shiny, smooth, and 16-21 μm in size (average diameter = 18.22 ± 1.00 μm, n = 20) (Figure S1D). These morphological characteristics were similar to those of N. sphaerica previously reported (Li et al. 2018). To identify species of ZJJ-C1-1, ZJJ-C1-2 and ZJJ-C1-3, the internal transcribed spacer (ITS) region, β-tubulin (TUB2), and the translation elongation factor 1-alpha (TEF1-α) were amplified using primer pairs including ITS1/ITS4 (Vilgalys and Hester 1990), Bt-2a/Bt-2b (Glass and Donaldson 1995), and EF1-728F/EF-2 (Zhou et al. 2015), respectively. Multiple sequence analyses showed no nucleotide difference was detected among genes tested except ITS that placed three isolates into two groups (Figure S2). BLAST analyses determined that ZJJ-C1-1, ZJJ-C1-2 and ZJJ-C1-3 had 99.73% to N. sphaerica strains LC2705 (KY019479), 100% to LC7294 (KY019397), and 99.79-100% to LC7294 (KX985932) or LC7294 (KX985932) based on sequences of TUB2 (MW252168, MW269660, MW269661), TEF-1α (MW252169, MW269662, MW269663), and ITS (MW250235, MW250236, MW192897), respectively. These indicated three isolates belong to the same species of N. sphaerica. Based on a combined dataset of ITS, TUB2 and TEF-1α sequences, a phylogenetic tree was constructed using Maximum likelihood method through IQ-TREE (Minh et al. 2020) and confirmed that three isolates were N. sphaerica (Figure S2). Further, pathogenicity tests were performed. Briefly, healthy unwounded fruits were surface-disinfected in 0.1% NaOCl for 30s, washed, dried and needling-wounded. Then, three fruits were inoculated with 10 μl of conidial suspension (1 × 106 conidia/ml) derived from three individual isolates, with another three fruits sprayed with 10 μl sterilized water as control. The treated fruits were incubated at 25℃ in 90% humidity. After 15 days, all the three fruits inoculated with conidia displayed typical dried-shrink symptoms as those observed in the farm field (Figure S1E). The decayed tissues with mycelium and spores could be observed on the skin or vertical split of the infected fruits after 15 days’ inoculation (Figure S1F-H). Comparably, in the three control fruits, there were no dried-shrink-related symptoms displayed. The experiment was repeated twice. The re-isolated pathogens were identical to N. sphaerica determined by sequencing the ITS, TUB2 and TEF-1α. Previous reports showed N. sphaerica could cause postharvest rot disease in kiwifruits (Li et al. 2018). To our knowledge, this is the first report of N. sphaerica causing fruits dried-shrink disease in A. trifoliata in China.

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First Report of Lasiodiplodia theobromae Associated with Decline of Grapevine Rootstock Mother Plants in Spain

Plant Disease ◽

10.1094/pdis-92-5-0832b ◽

2008 ◽

Vol 92 (5) ◽

pp. 832-832 ◽

Cited By ~ 13

Author(s):

A. Aroca ◽

R. Raposo ◽

D. Gramaje ◽

J. Armengol ◽

S. Martos ◽

...

Keyword(s):

Elongation Factor ◽

Aerial Mycelium ◽

Its Region ◽

Pathogenic Fungi ◽

Vascular Lesions ◽

Malt Extract ◽

Pathogenicity Test ◽

Lasiodiplodia Theobromae ◽

First Report ◽

Mother Plants

A field of Richter 110 rootstock mother plants in Valencia Province (eastern Spain) was surveyed during November 2006 to study the mycoflora of declining plants. Two canes with stunted leaves were collected from a plant with a reduced number of shoots. No cankers or vascular lesions were observed in the collected canes. Six wood chips (1 to 2 mm thick) were taken from one basal fragment (3 to 4 cm long) of each cane, surface sterilized in 70% ethanol for 1 min, and plated on malt extract agar supplemented with 0.5 g L–1 of streptomycin sulfate. Petri dishes were incubated for 7 days at 25°C. A fungus was consistently isolated from all samples that showed the following characteristics: colonies grown on potato dextrose agar (PDA) at 25°C developed a white, aerial mycelium that turned gray after 4 to 6 days and produced pycnidia after 1 month on sterile grapevine slivers of twigs placed on the PDA surface; conidia from culture were ellipsoidal, thick walled, initially hyaline, nonseptate, and measuring 20 to 25 (22.5) × 12 to 14 (13) μm; aged conidia were brown, 1-septate with longitudinal striations in the wall; and pseudoparaphyses variable in form and length were interspersed within the fertile tissue. The fungus was identified as Lasiodiplodia theobromae (Pat.) Griffon & Maubl. from the above characteristics (2). Identity was confirmed by analysis of the nucleotide sequences of the internal transcribed spacer (ITS) region from the rRNA repeat and part of the translation elongation factor 1-alpha (EF1-α) and the β-tubulin (B-tub) genes, as done elsewhere (1,3). BLAST searches at GenBank showed a high identity with reference sequences (ITS: 100%, EF1-α: 97%; B-tub: 99%). Representative sequences of the studied DNA regions were deposited at GenBank (Accession Nos.: ITS: EU254718; EF1-α: EU254719; and B-tub: EU254720). A pathogenicity test was conducted on 1-year-old grapevine plants cv. Macabeo grafted onto Richter 110 rootstocks maintained in a greenhouse. A superficial wound was made on the bark of 10 plants with a sterilized scalpel, ≈10 cm above the graft union. A mycelial plug obtained from the margin of an actively growing fungal colony (isolate JL664) was placed in the wound and the wound was wrapped with Parafilm. Ten additional control plants were inoculated with sterile PDA plugs. All control plants grew normally, and the inoculation wound healed 3 months after inoculation. Plants inoculated with L. theobromae showed no foliar symptoms in the same period, but developed cankers variable in size surrounding the inoculation sites. Vascular necroses measuring 8.4 ± 1.5 cm (mean ± standard error) developed in the inoculated plants that were significantly longer than the controls (0.3 ± 0.2 cm). The pathogen was reisolated from all inoculated plants and no fungus was reisolated from the controls. These results confirmed the pathogenicity of L. theobromae to grapevine and points to a possible involvement of L. theobromae in the aetiology of grapevine decline as previously reported (3,4). To our knowledge, this is the first report of L. theobromae isolated from grapevine in Spain. References: (1) J. Luque et al. Mycologia 97:1111, 2005. (2) E. Punithalingam. No. 519 in: Descriptions of Pathogenic Fungi and Bacteria. CMI, Kew, Surrey, UK, 1976. (3) J. R. Úrbez-Torres et al. Plant Dis. 90:1490, 2006. (4) J. M. van Niekerk et al. Phytopathol. Mediterr. 45(suppl.):S43, 2006.

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