Morphological and Molecular Identification of Fungi Causing Canker Disease on Melia Azidarch Trees in Some Regions of Mosul Provinces, Iraq

The aim of this study was to identify the fungi associated with canker disease on Melia azidarch trees inside Mosul University campus and the presidential palaces regions in Mosul Province, Iraq. Results of isolation showed the presenting of the fungi (Nattrassia mangiferae, Neoscylitidium dimidiatum Penz., Fusarium graminearium Schw., Alternaria brassicicola Schw., Aspergillus sp. and Penecillium sp.), which accompanied with the samples displayed cankering symptoms during the period from April to December/2020, the maximum of dominance was 85% for the fungus Neoscylitidium dimidiatum in August, while the lowest was 49% in April for the same year, followed by Fusarium graminearium with 38% in December, while the lowest percentage was 4% in October, then Alternaria brassicicola Schw. was 25% in April and the lowest value was 0% in August, followed by Aspergillus sp. and Penecillium sp. with low isolation percentages the maximum of which 25% and the lowest is 0% in August. When studying the pathogenicity of the isolated fungi, the results showed a high pathogenic effect in terms the length, diameter and the area of cankers symptom. Based on the results of the molecular diagnosis, the morphological identification was confirmed and it was clear that Fusarium austroamericanum, detection is considered the first record of this fungus in Iraq Melia azidarch trees.

LC-MS/MS Tandem Mass Spectrometry for Analysis of Phenolic Compounds and Pentacyclic Triterpenes in Antifungal Extracts of Terminalia brownii (Fresen)

Decoctions, macerations and fumigations of the stem bark and wood of Terminalia brownii Fresen. are used in traditional medicine for fungal infections and as pesticides on field crops and in traditional granaries in Sudan. In addition, T. brownii is commonly used for protecting wooden houses and furniture. Therefore, using agar disc diffusion and macrodilution methods, eight extracts of various polarities from the stem wood and bark were screened for their growth inhibitory effects against filamentous fungi commonly causing fruit, vegetable and grain decay, as well as infections in the immunocompromised host. Ethyl acetate extracts of the stem wood and bark gave the best antifungal activities, with MIC values of 250 µg/ml against Nattrassia mangiferae and Fusarium verticillioides, and 500 µg/ml against Aspergillus niger and Aspergillus flavus. Aqueous extracts gave almost as potent effects as the ethyl acetate extracts against the Aspergillus and Fusarium strains, and were slightly more active than the ethyl acetate extracts against Nattrassia mangiferae. Thin layer chromatography, RP-HPLC-DAD and tandem mass spectrometry (LC-MS/MS), were employed to identify the chemical constituents in the ethyl acetate fractions of the stem bark and wood. The stem bark and wood were found to have a similar qualitative composition of polyphenols and triterpenoids, but differed quantitatively from each other. The stilbene derivatives, cis- (3) and trans- (4) resveratrol-3-O-β-galloylglucoside, were identified for the first time in T. brownii. Moreover, methyl-(S)-flavogallonate (5), quercetin-7-β-O-di-glucoside (8), quercetin-7-O-galloyl-glucoside (10), naringenin-4`-methoxy-7-pyranoside (7), 5,6-dihydroxy-3`,4`,7-tri-methoxy flavone (12), gallagic acid dilactone (terminalin) (6), a corilagin derivative (9) and two oleanane type triterpenoids (1) and (2) were characterized. Our results justify the traditional uses of macerations and decoctions of T. brownii stem wood and bark for crop and wood protection and demonstrate that standardized extracts could have uses for the eco-friendly control of plant pathogenic fungi in African agroforestry systems. Likewise, our results justify the traditional uses of these preparations for the treatment of skin infections caused by filamentous fungi.

First Report of Multiple Species of the Botryosphaeriaceae Causing Bot Canker Disease of Indian Laurel-Leaf Fig in California

Indian laurel-leaf fig (Ficus microcarpa L.) is a commonly used indoor and outdoor ornamental tree. F. microcarpa is most frequently encountered as lining city streets, especially in warmer southern California climates. A disease known as ‘Sooty Canker,' caused by the fungus Nattrassia mangiferae (Syd. & P. Syd) B. Sutton & Dyko, is particularly devastating on F. microcarpa. Disease symptoms are characterized by branch dieback, crown thinning, and if the disease progresses to the trunk, eventual tree death (2). Recent taxonomic revisions have renamed Nattrassia mangiferae as Neofusicoccum mangiferae (Syd. & P. Syd.) Crous, Slippers & A. J. L. Phillips (1). An initial survey conducted during the spring of 2011 across four cities in Los Angeles County included, Culver City, Lakewood, Santa Monica, and Whittier. Five symptomatic branches per city were collected from trees showing branch cankers and dieback. Pieces of symptomatic tissue (2 mm2) were plated onto one-half-strength potato dextrose agar. Most isolates initially identified by morphological characteristics, such as growth pattern, speed of growth, and colony color, resembled those in the Botryosphaeriaceae (4). Two representative isolates from each site location were sequenced. Sequences obtained from amplification of the internal transcribed spacer region (ITS1-5.8rDNA-ITS2) and the β-tubulin gene were compared in a BLAST search in GenBank. Results identified isolates as Botryosphaeria dothidea (identity of 99% to EF638767 and 100% to JN183856.1 for ITS and β-tubulin, respectively); Neofusicoccum luteum (100% to EU650669 and 100% to HQ392752); N. mediterraneum (100% to HM443605 and 99% to GU251836); and N. parvum (100% to GU188010 and 100% to HQ392766) and have been deposited in GenBank with the following accession numbers: JN543668 to JN543671 (ITS) and JQ080549 to JQ080552 (β-tubulin). Pathogenicity tests were conducted in the greenhouse on 6-month-old F. microcarpa with one isolate from each previously listed fungal species. Five plants per isolate were stem-wound inoculated with mycelial plugs and wrapped with Parafilm. Uncolonized agar plugs were used as a control. Inoculations were later repeated a second time in the same manner for a total of 10 plants per isolate. Plants were observed for 6 weeks and destructively sampled to measure vascular lesion lengths. Mean vascular lesion lengths were 26, 22, 54, and 46 mm for B. dothidea, N. luteum, N. mediterraneum, and N. parvum, respectively. The mean lesion lengths for all isolates were significantly different (P = 0.05) from the control. Each species was consistently recovered from inoculated plants, except the control, thus fulfilling Koch's postulates. To our knowledge, this is the first report on the pathogenicity of multiple Botryosphaeriaceae species causing branch canker and dieback on F. microcarpa in California. These results are significant since trees along sidewalks in southern California are often crowded and undergo extensive root and branch pruning and some Botryosphaeriaceae spp. are known to enter its host through wounds caused by pruning or mechanical injury (2,3). Further sampling is imperative to better assess the distribution of these canker-causing fungal pathogens on F. microcarpa. References: (1) P. W. Crous et al. Stud. Mycol. 55:235, 2006. (2) D. R. Hodel et al. West. Arborist 35:28, 2009. (3) V. McDonald et al. Plant Dis. 93:967, 2009. (4) B. Slippers et al. Fungal Biol. Rev. 21:90, 2007.

Prevalence and Severity of Nattrassia mangiferae Root and Stem Rot Pathogen of Cassava in Bénin

Root rot pathogens were found through diagnostic surveys in all departments (regions) of Bénin, West Africa, to affect 86 to 100% and 96 to 100% of cassava fields during the dry and rainy seasons, respectively. Disease incidence in individual fields ranged between 0 and 53%, and averaged 16 to 27% per department. Nattrassia mangiferae was consistently the most frequently isolated root rot pathogen (56% in the dry season and 22 to 52% in the rainy season). Pathogenicity of N. mangiferae was confirmed on four cultivars of cassava using stem cuttings and storage roots. For all four cultivars, N. mangiferae significantly reduced the number of roots. Lesions (3 to 15 cm long) formed on the lower stem portion of all inoculated plants, whereas control plants remained symptom free. On storage roots, the disease profile was similar to that formed on stem cuttings. Other root rot pathogens detected during the dry season were Macrophomina phaseolina (14.2%), Fusarium spp. (11.8%), Botryodiplodia theobromae (7.7%), and Pythium spp. (2.9%). During the rainy season, Fusarium spp. were the second most commonly isolated root rot pathogens in three departments (Atlantique, Borgou, and Mono). In Oueme and Zou, B. theobromae was the second most isolated root rot pathogen (ranging between 24 and 28%) during the rainy season. During the same season, Pythium spp. were pronounced in Borgou (18%), followed by Mono (11%), Atlantique (9%), Atacora (8%), Oueme (5%), and Zou (6%). Results of the study are discussed with a view to creating awareness of the destructive power of N. mangiferae, a hitherto poorly recognized root rot pathogen of cassava in Bénin and West Africa in general.