First Report of Cirsium arvense (Canada thistle) as a New Host of Orobanche cumana Wallr. in Xinjiang, China

Cirsium arvense (Canada thistle) is a perennial herb native to Eurasia that has been introduced to temperate regions of the world where it is considered one of the serious weeds for arable and pastoral agriculture (Schröder et al. 1993). C. arvense reproduces both clonally and sexually. The weed is highly competitive, causes yield reductions in crops such as wheat, alfalfa, sugarbeet, and can reduce forage availability and production (Wilson 1981). Canada thistle is also a harbour for plant pathogens such as plant-parasitic nematodes (Tenuta et al. 2014). Sunflower broomrape (Orobanche cumana Wallr.) is a holoparasitic plant species with a restricted range of hosts both in the wild, where it mainly parasitizes a few species of the Asteraceae, and in agricultural fields, where it is exclusively found growing on sunflower (Fernández-Martínez et al. 2015). O. cumana infection can cause up to 80% of the yield loss in sunflower, which is a serious threat for sunflower production in Xinjiang and Inner Mongolia, China (Parker 2009). In July 2019, broomrape was observed parasitizing C. arvense in the greenhouse used for sunflower resistance identification (Shihezi, 86° 3' 36" E, 44° 18' 36" N, 500 m elevation) in Xinjiang, China. Fifty percent of the plants were parasitized by broomrape in the greenhouse and the host had an average of 1-2 broomrape shoots per plant. For molecular analysis, total genomic DNA was extracted from the flowers of broomrape and the rps2, rbcL, trnL-F genes, and ribosomal DNA internal transcribed spacer (ITS) region were amplified by PCR using the primer pairs rps2F/rps2R, rbcLF/rbcLR, C/F, ITS1/ITS4, respectively (Park et al. 2007; Manen et al. 2004; Taberlet et al. 1991; Anderson et al. 2004). The ITS (659bp), rps2 (451 bp), trnL-F (914 bp), and rbcL (961 bp) gene sequences of the broomrape were deposited in GenBank, the accession numbers are MT856745, MW809407, MW809408, and MW809409. The results of BLAST analysis showed that ITS sequence shared 100% similarity with O. cumana (659/659 nucleotide identity, MK567978), the rps2 sequence shared 99% similarity with O. cumana (449/451 nucleotide identity, KT387722), trnL-F sequence shared 99% similarity with O. cumana (907/911 nucleotide identity, MT027325), rbcL sequence shared 99% similarity with O. cumana (956/964 nucleotide identity, MK577840). The morphological characteristics such as stem, inflorescence, corolla, bracts, calyx, stamens, gynoecium are consistent with O. cumana described by Pujadas-Salvá and Velasco (2000). Morphological and molecular identification strongly support that the broomrape parasitic on C. arvense belonged to the O. cumana. Greenhouse pot experiments were carried out to assess the parasitic relationship between sunflower broomrape and C. arvense (Fernández-Martínez et al. 2000). In January 2020, C. arvense roots were harvested from an extant field of C. arvense in the greenhouse at Shihezi University (Supplementary Figure S1A). The soil was dug to 30-40 cm depth and C. arvense roots were removed and carefully washed in water. The healthy and living C. arvense roots were selected and cut into 10-11 cm pieces. Four C. arvense root pieces were grown (buried at a depth of 10-12 cm) in 8-L pots containing a mixture of sand-vermiculite-compost (1:1:1 v:v:v) and O. cumana seeds (50 mg of O. cumana seeds per 1 kg of the substrate) with 5 replicates. Three non-infected plants were grown and evaluated in parallel. Approximately 80 days after planting, at the flowering stage of the O. cumana, C. arvense plants were uprooted from the soil. Compared to non-infected plants, the hosts’ symptoms were slow growth, leaf wilting, and chlorosis, and similiar to the broomrape-infected C. arvense plants observed in the greenhouse field. The roots of C. arvense and broomrape were carefully washed in water and observed the parasitism of O. cumana. The infection was confirmed by observation of the attachment of the O. cumana to the C. arvense roots (Supplementary Figure S1D). To the best of our knowledge, this is the first report of O. cumana parasitizing C. arvense in Xinjiang, China. C. arvense as a new host of O. cumana indicates that sunflower broomrape can also propagate and survive in a host such as Canada thistle grown in sunflower fields. This finding suggests that it may be more difficult to control sunflower broomrape by rotation. In the next study, the contaminated area and the degree of parasitism of broomrape on C. arvense in the field will be investigated, and better-integrated control methods for controlling O. cumana will be designed. References: Schröder, D., et al. 1993. Weed. Res. 33:449-458. https://doi.org/10.1111/j.1365-3180.1993.tb01961.x Crossref, ISI, Google Scholar Wilson, R. G. 1981. Weed. Sci. 29:159-164. https://doi.org/10.1017/S0043174500061725 Crossref, ISI, Google Scholar Tenuta, M., et al. 2014. J. Nematol. 46(4):376–384. Fernández-Martínez, J. M., et al. 2015. Page 129 in: Sunflower Oilseed: Chemistry, Production, Processing and Utilization. AOCS Press, Champaign, IL. https://doi.org/10.1016/B978-1-893997-94-3.50011-8 Crossref, Google Scholar Parker, C. 2009. Pest Manag. Sci. 65:453-459. https://doi.org/10.1002/ps.1713 Crossref, ISI, Google Scholar Park, J. M., et al. 2007. Mol. Phylogenet. Evol. 43: 974. https://doi.org/10.1016/j.ympev.2006.10.011 Crossref, ISI, Google Scholar Manen, J. F., et al. 2004. Mol. Phylogenet. Evol. 33:482. https://doi.org/10.1016/j.ympev.2004.06.010 Crossref, ISI, Google Scholar Taberlet, P., et al. 1991. Plant Mol. Biol. 17:1105-1109. https://doi.org/10.1007/bf00037152 Crossref, ISI, Google Scholar Anderson, I.C., et al. 2004. Environ. Microbiol. 6: 769. https://doi.org/10.1111/j.1462-2920.2004.00675.x Crossref, ISI, Google Scholar Pujadas-Salvà, A. J., and Velasco, L. 2000. Bot. J. Linn. Soc. 134:513-527. https://doi.org/10.1006/bojl.2000.0346 Crossref, ISI, Google Scholar Fernández-Martínez, J. M., et al. 2000. Crop. Sci. 40:550-555. https://doi.org/10.2135/cropsci2000.402550x Crossref, ISI, Google Scholar

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First Report of Race Composition and Distribution of Sunflower Broomrape, Orobanche cumana, in China

Plant Disease ◽

10.1094/pdis-07-14-0721-pdn ◽

2015 ◽

Vol 99 (2) ◽

pp. 291-291 ◽

Cited By ~ 12

Author(s):

B. X. Shi ◽

G. H. Chen ◽

Z. J. Zhang ◽

J. J. Hao ◽

L. Jing ◽

...

Keyword(s):

Inner Mongolia ◽

Vascular System ◽

Tap Water ◽

Shanxi Province ◽

First Report ◽

Orobanche Cumana ◽

Sunflower Broomrape ◽

Differential Hosts ◽

Novi Sad ◽

Academy Of Sciences

Sunflower broomrape (Orobanche cumana Wallr.) is a holoparasitic plant that penetrates the vascular system of sunflower roots, absorbs plant nutrients and water, and thus causes stunting, reduced growth, and severe yield losses (3). To date, seven races of sunflower broomrape (O. cumana) have been identified by using international standard race differential hosts in Bulgaria, Serbia, Romania, Turkey, and Russia (4). However, the race types present in China are unknown. To identify the race composition of sunflower broomrape in China, race differential hosts of sunflower broomrape were received from Dr. Dragan Skoric (Serbian Academy of Sciences and Arts, Novi Sad, Serbia): Line AD66 has no resistant genes; Kruglik-41 contains resistant gene Or1; B-RO-02A has Or2; Record has Or3; LC1002B has Or4; LC1003B has Or5; LC-1093 has Or6, and Race-G-2 has Or7 (1). Eighteen sunflower broomrape samples were collected in August of 2011, 2012, and 2013 from different provinces/locations in China, including Xinjiang (Xinyuan, Shihezi, Tekesi, Beitun, Urumqi, and Yining), Inner Mongolia (Linhe, Xixiaozhao, Wuqianqi, Tuzuoqi, Keyouqianqi, and Aohanqi), Shanxi (Hunyuan, Shilou, Mizhi, and Dingbian), Jilin (Tongyu), and Hebei (Xuanhua). The differential hosts were each inoculated with the seeds of each broomrape isolate that was recovered, as described by Pancenko with minor modification (2). Briefly, two parts of field soil and one part of vermiculite were mixed together and used as potting mix. The mix was inoculated with broomrape seeds at 10 mg of seeds per 100 g of potting mix. The inoculated mix was placed in a 7-cm (diameter) × 11-cm (height) plastic pot to fill two-thirds of the pot volume. Three sunflower seeds were placed on the surface of the mix at an even distance from each other and covered with additional mix. The pots were kept in a greenhouse under a 16-h photoperiod at 10,000 lux of illumination intensity, temperature of 20–25°C, and 40% relative humidity. Forty days after incubation, sunflower seedlings were taken out from the pot and the roots washed with tap water. The number of tubercles was recorded on the root of each differential host. Race types were determined based on the reaction (tubercule formation on roots) of all the standard differential hosts to the test isolate. The results showed that races A, D, E, and G of O. cumana were present among the isolates. Race G was found in Wuqianqi, Xixiaozhao, and Linhe in the western part of Inner Mongolia. Race E was found only in Shihezi of Xinjiang. Race D was found in Aohanqi and Keyouqianqi (eastern part of Inner Mongolia); Xinyuan, Tekesi, Beitun, and Urumqi (northern part of Xinjiang); and Tongyu (northern part of Jilin). Race A was found in Mizhi, Shilou, and Hunyuan of Shanxi province and Xuanhua in Hebei province. Additionally, race A was also found in Tuzuoqi, the middle region of Inner Mongolia. Thus, races A, D, E, and G were the main race types of O. cumana in China. Race D was the predominant race type and had the widest distribution. Race G was the highest level race type in this study but was mainly limited to the western part of Inner Mongolia. This is the first report of race composition and distribution of sunflower broomrape (O. cumana) in China. References: (1) Y. Kaya et al. Helia 40:211, 2004. (2) A. N. Pancenko, Zbirnik VNIIMK. Page 107, 1973. (3) C. Parker. Page 17 in: Proc. 3rd Int. Workshop on Orobanche and Related Striga Research, 1994. (4) P. Shindrova et al. Helia 35:87, 2012.

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First Report of Stem Rot of Sunflower Broomrape (Orobanche cumana) Caused by Sclerotinia minor Jagger in Inner Mongolia, China

Plant Disease ◽

10.1094/pdis-08-17-1173-pdn ◽

2018 ◽

Vol 102 (3) ◽

pp. 683-683 ◽

Cited By ~ 1

Author(s):

J. Zhang ◽

R. Jia ◽

Y. Zhang ◽

M. Li ◽

H. Zhou ◽

...

Keyword(s):

Inner Mongolia ◽

Stem Rot ◽

Sclerotinia Minor ◽

First Report ◽

Orobanche Cumana ◽

Sunflower Broomrape

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A survey of subtropical nursery plants for fungal diseases in Northland

New Zealand Plant Protection ◽

10.30843/nzpp.2006.59.4449 ◽

2006 ◽

Vol 59 ◽

pp. 132-136 ◽

Cited By ~ 1

Author(s):

M. Braithwaite ◽

C.F. Hill ◽

S. Ganev ◽

J.M. Pay ◽

H.G. Pearson ◽

...

Keyword(s):

New Zealand ◽

Plant Pathogens ◽

Morphological Characteristics ◽

Molecular Techniques ◽

Plant Diseases ◽

New Host ◽

Disease Symptoms ◽

Plant Families ◽

First Time

During 2003 and 2004 fortyfive randomly selected wholesale and retail plant nurseries were surveyed for plant diseases The plant families Agavaceae Annonaceae Arecaceae Bromeliaceae Cycadaceae and Musaceae were targeted Plants were examined in situ for disease symptoms as well as samples being collected for laboratory analyses Fungi were identified using morphological characteristics and where necessary with molecular techniques The survey resulted in a range of fungi being identified from the target plants These fungi ranged from saprophytes to plant pathogens some of which may have undesirable effects on New Zealands biodiversity or economy Many new host/pathogen records were observed and several fungi were detected for the first time in New Zealand This paper presents and discusses the results of these findings

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Identification of Botryosphaeriaceae Species Causing Kiwifruit Rot in Sichuan Province, China

Plant Disease ◽

10.1094/pdis-07-14-0727-re ◽

2015 ◽

Vol 99 (5) ◽

pp. 699-708 ◽

Cited By ~ 16

Author(s):

You Zhou ◽

Guoshu Gong ◽

Yongliang Cui ◽

Daixi Zhang ◽

Xiaoli Chang ◽

...

Keyword(s):

Dna Sequences ◽

Plant Pathogens ◽

Elongation Factor ◽

Morphological Characteristics ◽

Sichuan Province ◽

Fruit Rot ◽

First Report ◽

Primary Inoculum ◽

Tubulin Genes ◽

Wide Range

Species of Botryosphaeriaceae fungi are important plant pathogens causing cankers, blight, and fruit rot in an extremely wide range of host. In recent years, kiwifruit rot has been a serious problem in Sichuan Province, one of the important kiwifruit production areas of China. Botryosphaeria dothidea has previously been associated with kiwifruit rot but little is known regarding whether other Botryosphaeriaceae genera also constitute kiwifruit rot pathogens in China. Accordingly, diseased fruit were collected from six different areas of Sichuan Province. Based on morphological characteristics, pathogenicity testing, and comparisons of DNA sequences of the internal transcribed spacer, transcription elongation factor 1-α, and β-tubulin genes, 135 isolates of Botryosphaeriaceae were identified as B. dothidea, Lasiodiplodia theobromae, and Neofusicoccum parvum. All of these species were found to cause kiwifruit rot. To understand the infection cycle of kiwifruit rot pathogens, these three species were used to inoculate leaves and shoots of kiwifruit. The results showed that these species could cause spots on leaves and lesions on shoots, producing abundant pycnidia on leaves and shoots surfaces. Moreover, B. dothidea conidia and ascospores from overwintered pycnidia and pseudothecia in kiwifruit orchards in April and August could cause fruit rot and spots on leaves of kiwifruit. Therefore, we concluded that overwintered pycnidia and pseudothecia of B. dothidea in kiwifruit orchards are the primary inoculum for kiwifruit rot, with new pycnidia that develop during the growing season serving as a secondary inoculum. This is the first report of N. parvum and L. theobromae causing kiwifruit rot in China and is also the first report that B. dothidea is able to overwinter as pycnidia and pseudothecia in kiwifruit orchards and serve as the primary inoculum for kiwifruit rot.

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First Report of Choanephora cucurbitarum Causing Leaf Wilt, Flower Rot, and Stem Necrosis on Crotalaria breviflora

Plant Disease ◽

10.1094/pdis-08-20-1719-pdn ◽

2021 ◽

Author(s):

Gislaine de Souza Oliveira ◽

Rildo Alexandre Fernandes ◽

Danilo Batista Pinho ◽

Solange Maria Bonaldo

Keyword(s):

Graduate Program ◽

Morphological Characteristics ◽

Google Scholar ◽

Environmental Sciences ◽

Mato Grosso ◽

First Report ◽

Initial Symptoms ◽

Wide Range ◽

Choanephora Cucurbitarum ◽

Stem Necrosis

Crotalaria breviflora (Fabaceae) is used as green manure crop because of its nitrogen fixation and nematode control (Nascimento et al. 2020). In April 2018, leaf wilting, flower rot, and stem necrosis symptoms were observed on C. breviflora with 100% incidence, in Sorriso (12° 33′ 31″ S, 55º 42′ 51″ W), Santa Carmem (11° 55′ 52″ S, 55º 16′ 47″ W), and Sapezal (12º 59′ 22″ S, 58º 45′ 52″ W) counties in the state of Mato Grosso, Brazil. Three monosporic isolates were isolated from symptomatic leaves, cultivated in potato dextrose agar (PDA) medium, and deposited at the Cultures Collection of the University of Brasilia (codes CCUB 1293, CCUB 1667, CCUB 1668). Colonies on PDA were white and cottony with presence of hyaline and coenocytic hyphae. The mycelia later became pale yellow with abundant reproductive structures. Sporangiophores were hyaline, aseptate, unbranched, and apically dilated to form a clavate vesicle, which produced secondary vesicles bearing sporangiola. Secondary vesicles were clavate, light brown, and 37 to 51 µm in diameter. Sporangia were brown to dark brown, globular to ellipsoid, 115 to 140 µm long, and 96 to 122 µm wide. Sporangiospores (n=30) were brown to reddish-brown, ellipsoid to ovoid, with longitudinal striae, 14 to 19 µm long, and 8 to 12 µm wide. Some with hyaline appendages at both ends. Their morphological characteristics were consistent with the descriptions of Choanephora cucurbitarum (Kirk 1984). To confirm the identity, the DNA of the three isolates was extracted and the sequences of Small Subunit (SSU), Large Subunit (LSU), and complete Internal Transcribed Spacer (ITS) of rDNA were amplified using V9G, ITS3, and LR5 primers (GenBank acc. no: MN897836, MN897837 and MN897838). The sequences were aligned with the MAFFT software. The alignment matrix was subjected to Maximum Likelihood (ML) analysis using RAxML v. 8 and Bayesian Inference performed in MrBayes v.3.1.2. The tree was edited in the FigTree software. The sequences showed 100% identity with the sequences from C. cucurbitarum found on the GenBank. To confirm pathogenicity, a suspension at 5.4 ×106 spores/ml was prepared from a 15-day-old culture grown at 25°C and sprayed on asymptomatic plants of C. breviflora. Sterilized water was sprayed as the control. Plants were kept in a humid chamber at 20°C for 48 h. Initial symptoms were visualized 16 days after inoculation. Complete necrosis of leaves and stems with spore mass on infected tissue was observed 19 days after inoculation. To satisfy the Koch’s postulates, the fungus was successfully reisolated from the infected tissues. No symptoms were observed on the control plants. In Brazil, this pathogen has been reported on Brassica oleracea var. capitata, Capsicum annuum, Crotalaria spectabilis, Cucurbita sp., and Vigna unguiculata (Alfenas et al. 2018; Mendes and Urben, 2019). C. cucurbitarum has been reported to have a wide range of hosts (Farr and Rossman, 2020). It can infect the crops grown in rotation or in succession, including common bean, corn, cotton, quinoa, soybean, and sunflower. Therefore, this pathogen is of epidemiological importance and poses a threat to the croplands where environmental conditions are conducive to the disease to develop and spread. To our knowledge, this is the first report of C. cucurbitarum causing leaf and flower wilt, and stem rot on C. breviflora in the world. Acknowledgment We thank the Environmental Sciences Graduate Program, Federal University of Mato Grosso, University of Brasilia, PROPeq/PROPG-UFMT, EMBRAPA, CODEX/UFMT, Institute of Agricultural and Environmental Sciences (ICAA)/UFMT and CAPES for providing the Master's scholarship. References Alfenas, R. F., et al. 2018. Plant Dis.102:1456. https://doi.org/10.1094/PDIS-10-17-1610-PDN, Google Scholar. Farr, D. F., and Rossman, A. Y. 2020. Fungal Databases, Syst. Mycol. Microbiol. Lab., ARS, USDA. Retrieved May 26, 2020 from https://nt.ars-grin.gov/fungaldatabases/, Google Scholar. Kirk, P. M. 1984. Mycol Paper. 152:1. Google Scholar. Mendes, M. A. S., and Urben, A. F. 2020. Fungos relatados em plantas no Brasil, Retrived May 26, 2020 from http://pragawall.cenargen.embrapa.br/aiqweb/michtml/fgbanco01.asp, Google Scholar. Nascimento, D. D. et al. 2020. Bioscience Journal. 36:713. https://doi.org/10.14393/BJ-v36n3a2020-42248, Google Scholar.

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First Report of a New Race of Sunflower Broomrape (Orobanche cumana) in Israel

Plant Disease ◽

10.1094/pdis.2004.88.11.1284c ◽

2004 ◽

Vol 88 (11) ◽

pp. 1284-1284 ◽

Cited By ~ 3

Author(s):

H. Eizenberg ◽

D. Plakhine ◽

T. Landa ◽

G. Achdari ◽

D. M. Joel ◽

...

Keyword(s):

Dna Analysis ◽

Local Development ◽

Random Amplified Polymorphic Dna ◽

Morphological Characteristics ◽

Intraspecific Diversity ◽

Plant Management ◽

Resistance Response ◽

Orobanche Cumana ◽

Sunflower Broomrape ◽

New Race

The genus Orobanche includes chlorophyll-lacking root parasites that parasitize many dicotyledonous species and causes severe damage to vegetable and field crops worldwide. Sunflower broomrape (Orobanche cumana Wallr.) is known in Eurasia as a specific parasite of sunflower, which differs from the nodding broomrape (O. cernua Loefl) in host specificity and morphological characteristics (3). Together with Egyptian broomrape (O. aegyptiaca Pers.), it seriously parasitizes sunflower (Helianthus annuus L.) in Israel (1). Prior to 2000, the local confectionary sunflower cvs. Ambar and Gitit proved to be resistant to the local O. cumana populations in Israel (2). A preliminary study, which we conducted in 1995 using the Vranceanu's differentials (4), indicated that O. cumana populations in Israel behave like the known race C. Using random amplified polymorphic DNA analysis, we also found a very low intraspecific diversity of this species in Israel at that time. However, in 2000, infection of the sunflower cvs. Ambar and Gitit was reported in two fields (Gadot and Afek) in northern Israel. In 2001 and 2002, O. cumana parasitized these cultivars in three more locations as much as 50 km apart (Tel-Adashim, Mevo-Hama, and Bet-Hilel). To determine the virulence of O. cumana populations on sunflower cultivars under controlled conditions, O. cumana seeds were collected in the above mentioned sunflower fields. In addition, we also used seeds from an O. cumana population collected in Alonim in 1997. This latter population did not infect the above mentioned ‘resistant’ sunflower cultivars in the field (2,); therefore, represented the previously known O. cumana populations in Israel. Resistant (Ambar) and susceptible (D.Y.3) sunflower cultivars were planted in separate pots that were differentially filled with soil that was inoculated with O. cumana seeds of the different populations. The experiment was performed in a full factorial arrangement with six replications. As expected, O. cumana from Alonim failed to attack the resistant sunflower. However, the O. cumana populations that were collected in the five other fields seriously attacked both sunflower cultivars, indicating higher virulence. O. cumana from all five new populations proved more virulent than the Alonim population on cvs. Ambar and D.Y.3. The occurrence of these new virulent populations could have several reasons including: (i) importation of virulent parasite seeds from abroad; or (ii) local development of virulence from previously avirulent populations. The latter could be favored by the continuous and repeated use of the available resistant varieties that are all based on a single resistance response (2). References: (1) H. Eizenberg and D. M. Joel. Orobanche in Israeli agriculture. Workshop of COST Action 849, Parasitic Plant Management in Sustainable Agriculture, 2001. (2) H. Eizenberg et al. Plant Dis. 88:479, 2003. (3) D. M. Joel. Phytoparasitica 16:375, 1988. (4) A. V. Vranceanu et al. Proc. 9th Sunflower Conf. 1:74–82, 1980.

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First Report of Postharvest Fruit Rot in Avocado (Persea americana) Caused by Lasiodiplodia theobromae in Italy

Plant Disease ◽

10.1094/pdis-10-11-0886 ◽

2012 ◽

Vol 96 (3) ◽

pp. 460-460 ◽

Cited By ~ 6

Author(s):

A. Garibaldi ◽

D. Bertetti ◽

M. T. Amatulli ◽

J. Cardinale ◽

M. L. Gullino

Keyword(s):

Plant Pathogens ◽

Morphological Characteristics ◽

The United States ◽

Disease Diagnosis ◽

Persea Americana ◽

Fruit Rot ◽

Pathogenicity Test ◽

Lasiodiplodia Theobromae ◽

First Report ◽

Pathogenicity Tests

Avocado (Persea americana Mill.) is grown in some areas of southern Italy. In spring 2011, a previously unknown rot was observed on fruit that was marketed in Torino (northern Italy). The decayed area started from the stalk, appeared irregular and soft, and was surrounded by a dark brown margin. The internal decayed area appeared rotten, brown, and surrounded by bleached tissue. Fragments (approximately 3 mm) were taken from the margin of the internal diseased tissues, cultured on potato dextrose agar (PDA), and incubated at temperatures between 21 and 25°C under alternating conditions of light and dark. Colonies of the fungus initially appeared whitish, later turning mouse gray to black. Mature mycelium was septate and produced a dark pigment. The fungus, grown on oat agar (2) and incubated at temperatures between 21 and 25°C under alternating light and darkness, produced grayish colonies with a fluffy aerial mycelium that became dark with age and produced black pigments. After 18 days of incubation, such colonies produced pycnidia aggregated into stromatic masses, emerging from decayed tissues, and up to 3 to 4 mm in diameter. Conidia produced in the pycnidia were initially unicellular, hyaline, granulose, ovoid to ellipsoidal, and measured 20.8 to 26.9 × 12.5 to 16.1 (average 24.4 × 13.5) μm. After 7 days, mature conidia became darker, uniseptate, and longitudinally striate. Paraphyses produced within the tissues of pycnidia were hyaline, cylindrical, nonseptate, and up to 63 μm long. Morphological characteristics of mycelia, pycnidia, and conidia observed with a light microscope permitted identify of the fungus as Lasiodiplodia theobromae (3). The internal transcribed spacer (ITS) region of rDNA was amplified using the primers ITS1/ITS4 and sequenced. BLAST analysis (1) of the 488-bp segment showed a 100% similarity with the corresponding sequence (GenBank Accession No. GQ502453) of L. theobromae Pat. Griffon & Maubl. The nucleotide sequence of the strain used for pathogenicity tests was submitted to GenBank (Accession No. JN849098). Pathogenicity tests were performed by inoculating 10 avocado fruits after surface disinfesting in 1% sodium hypochlorite and then wounding. Mycelial disks (8 mm in diameter) obtained from PDA cultures of one strain were placed on wounds. Ten control fruits were inoculated with plain PDA. Fruits were incubated at 15 ± 1°C. The first symptoms developed 4 days after the artificial inoculation. After 7 days, the rot was evident and L. theobromae was consistently reisolated. Noninoculated fruit remained healthy. The pathogenicity test was performed twice. To our knowledge, this is the first report of the presence of L. theobromae causing postharvest fruit rot on avocado in Italy, as well as in Europe. The occurrence of postharvest fruit rot on avocado caused by L. theobromae was described in many avocado-producing areas such as the United States (4), South Africa, and Israel. In Italy, the economic importance of avocado cultivation is currently limited. References: (1) S. F. Altschul et al. Nucleic Acids Res. 25:3389, 1997. (2). P. Narayanasamy. Microbial Plant Pathogens. Detection and Disease Diagnosis: Fungal Pathogens. Springer, Dordrecht, 2011. (3) E. Punithalingam. Sheet 519. CMI Description of Fungi and bacteria, 1976. (4) H. E. Stevens and R. B. Piper. Circular No. 582, USDA, 1941.

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First Report of white root rot of hemp (Cannabis sativa L.) caused by Dematophora necatrix in Campania region (Southern Italy)

Plant Disease ◽

10.1094/pdis-07-20-1521-pdn ◽

2021 ◽

Author(s):

Roberto Sorrentino ◽

Gian Maria Baldi ◽

Valerio Battaglia ◽

Francesco Raimo ◽

Giulio Piccirillo ◽

...

Keyword(s):

Root Rot ◽

Cannabis Sativa ◽

Elongation Factor ◽

Morphological Characteristics ◽

New Host ◽

Campania Region ◽

First Report ◽

History Of ◽

Industrial Hemp ◽

Dematophora Necatrix

Industrial hemp (Cannabis sativa L.) was cultivated in Italy until the end of the Second World War. Since then, it has been abandoned and substituted with other crops mainly due to legal restrictions and public concerns. Public legislation passed in 2016, has allowed for the production of hemp seeds, flowers and fibers (law n. 242/2016). During a 2019 survey on hemp sanitary status in the province of Naples (40°57'6"12 N, 14°22'37"56 E), hemp ‘Kompolty’ with symptoms of root rot were observed at a private farm and collected for further analysis at the phytosanitary laboratory of CREA in Caserta. Death generally occurred within 2-3 weeks after the appearance of the first symptoms, occurring on ca. 10% of plants, consisting of yellowing, canopy wilt and signs of roots covered with white mycelium and fan-like mycelium under the bark. The causal agent, was isolated from small root segments, excised from symptomatic plants, the surface was disinfected with 2% sodium hypochlorite, placed on potato dextrose agar (PDA) amended with streptomycin sulphate (100mg/L) and incubated in the dark at 25°C for 5 days. Small pieces (2-3 mm) at the edge of the resulting colonies were sub-cultured onto PDA and incubated at 25°C in the dark for one week. The mycelia from 15 isolates showed pear-shaped swellings adjacent to the septa. The conidia were aseptate, hyaline, ellipsoid to ovoid, and 3-5 × 2.5-3 µm (n=50). Based on the morphological characteristics, the fungus was identified as Rosellinia necatrix Berl. ex Prill. (Singleton et al., 1992) a fungus taxonomically revised to Dematophora necatrix R. Hartig (Wittstein et al., 2020). To confirm the identification, total DNA was extracted from five isolates using a DNeasy Plant Mini Kit (Qiagen, Hilden, Germany) and the ITS spacer was PCR-amplified with primers ITS1-ITS4 (White et al., 1990). The size-expected amplicons of 536 bp were purified and sequenced, the resulting sequence was trimmed and deposited in GenBank under the accession number MK937913. BLAST-n analysis revealed 98.83% nucleotide identity with some representative isolates of D. necatrix (MK888684.1; KT343972.1). To fulfill Koch’s postulates, the pathogenicity tests were carried out on fifteen 4-weeks-old potted hemp plants ‘Kompolty’. The inoculation was performed by adding 3 g of millet seeds inoculated with ten mycelial plugs, taken from the margins of a D. necatrix actively growing colony, per liter of sterile peat and perlite substrate in single pots. Moreover, ten hemp plants were inoculated with sterilized millet seed and served as negative controls. All plants were incubated at 25°C. After three weeks, inoculated plants exhibited foliar chlorosis, apical wilting, and death in two weeks, similar to what was observed in the field. Control plants did not show any symptoms. The fungus was isolated from the roots in all fifteen inoculated plants and confirmed to be D. necatrix based on morphological and molecular analysis, carried out with a second primer pair EF1-983F/ EF1-2218R targeting the transcription elongation factor 1- (Rehner and Buckley., 2005) (MW541068) that showed 99.67% nt in BLAST-n analysis. To our knowledge, this is the first report of D. necatrix infecting hemp in Europe. The farm where the problem arose has a history of cultivation for the production of apples for over 30 years. Therefore, an adaptation of D. necatrix to the new host is hypothesized. An in-depth knowledge on the diseases of hemp will be needed to relaunch hemp cultivation in this area.

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First Report of Foliar and Stem Blight on Sunflower Caused by Alternaria helianthiinficiens in Croatia

Plant Disease ◽

10.1094/pdis-05-12-0512-pdn ◽

2012 ◽

Vol 96 (11) ◽

pp. 1698-1698 ◽

Cited By ~ 1

Author(s):

K. Vrandečić ◽

D. Jurković ◽

J. Ćosić ◽

I. Stanković ◽

A. Vučurović ◽

...

Keyword(s):

Disease Incidence ◽

Pathogenic Fungus ◽

Morphological Characteristics ◽

Its Region ◽

Nucleotide Identity ◽

Oilseed Crop ◽

Conidial Suspension ◽

First Report ◽

Stem Blight ◽

Detached Leaves

Sunflower (Helianthus annus L.) is the most important oilseed crop in Croatia. In August 2009, in six localities of eastern Croatia, severe foliar and stem blight symptoms were observed on several genotypes with disease incidence ranging from 10 to 50%. At the initial stage of the infection, irregular to oval, brown spots different in size, surrounded by a chlorotic halo, appeared on the leaves that gradually became enlarged and coalesced, and whole leaves turned yellow and necrotic, followed by defoliation. Lesions on the stems were light to dark brown, randomly distributed, rounded and tapered on the ends; later becoming large and elongated causing stem breakage. Tissue within the lesion was reddish on the cross section. To determine the causal agent, small pieces of symptomatic leaves and stem tissue of sunflower were surface disinfested and placed on potato dextrose agar. A total of 17 isolates from leaves as well as six from stems were obtained and all formed cottony, dark olivaceous to black colonies under 12 h of fluorescent light per day. All isolates formed uniform solitary, pale brown to brown, long ovoid conidia with five to eight transverse and one to two longitudinal septa. The conidia of all isolates were slightly constricted at the transverse septa, measuring 55 to 90 × 14 to 20 μm. Based on the morphological characteristics, the pathogen was identified as Alternaria helianthiinficiens E.G. Simmons, Walcz & R.G. Roberts (4). The pathogenicity was tested with one representative isolate (Alt5) by injection of a conidial suspension (106 conidia/ml) into stems of 20 healthy sunflower seedlings and by spraying 20 non-wounded detached leaves with a suspension of spores. Small necrotic spots on all inoculated seedlings and leaves formed 5 and 9 days after inoculation, respectively. The control sunflower seedlings and detached leaves, inoculated with sterile water, showed no reactions. The identity of isolate Alt5 was futher confirmed by amplification and sequencing of the internal transcribed spacer (ITS) region of rDNA. Because there are no available corresponding ITS sequences of A. helianthiinficiens in the GenBank, reference type strain CBS 208.86 (publicly purchased, CBS, Utrecht, Netherlands) was also sequenced in this study. Total DNA was extracted directly from fungal mycelium and PCR amplification and sequencing were performed with primers ITS1F/ITS4. Sequence analysis of ITS region revealed 100% nucleotide identity between isolate Alt5 (GenBank Accession No. JX101648) and isolate CBS 208.86 (GenBank Accession No. JX101649). The nucleotide identity of both isolates compared with A. helianthi (HM449991), another sunflower pathogenic fungus, was only 80%. A. helianthiinficiens has previously been reported on sunflower in Hungary and the USA (3), Serbia (1), and Korea (2). However, to our knowledge, this is the first report of A. helianthiinficiens occurrence in Croatia as a new and harmful parasite of sunflower, illustrating an expansion of its geographical range and underscoring the need for phytosanitary control because it is a seedborne fungus. References: (3) M. Aćimović and N. Lačok. Helia 14:129, 1991. (4) H. S. Cho and S. H. Yu. Plant Pathol. J. 16:331, 2000. (2) E. G. Simmons. Mycotaxon 25:203, 1986. (1) E. G. Simmons. Alternaria: An Identification Manual. CBS Fungal Biodiversity Centre, Utrecht, the Netherlands, 2007.

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