Morphological and molecular characterization of Fusarium spp pathogenic to pecan tree in Brazil.

The occurrence of Fusarium spp associated with pecan tree (Carya illinoinensis) diseases in Brazil has been observed in recent laboratory analyses in Rio Grande do Sul State. Thus, in this study, we i) obtained Fusarium isolates from plants with disease symptoms; ii) tested the pathogenicity of these Fusarium isolates to pecan; iii) characterized and grouped Fusarium isolates that were pathogenic to the pecan tree based on morphological characteristics; iv) identified Fusarium spp to the species complex level through TEF-1α sequencing; and v) compared the identification methods used in the study. Fifteen isolates collected from the inflorescences, roots, and seeds of symptomatic plants (leaf necrosis or root rot) were used for pathogenicity tests. Morphological characterization was conducted using only pathogenic isolates, for a total of 11 isolates, based on the mycelial growth rate, sporulation, colony pigmentation, and conidial length and width variables. Pathogenic isolates were grouped based on morphological characteristics, and molecular characterization was performed by sequencing TEF-1α genes. Pathogenic isolates belonging to the Fusarium chlamydosporum species complex, Fusarium graminearum species complex, Fusarium proliferatum, and Fusarium oxysporum were identified based on the TEF-1α region. Morphological characteristics were used to effectively differentiate isolates and group the isolates according to genetic similarity, particularly conidial width, which emerged as a key morphological descriptor in this study.

First Report of Fusarium sambucinum Associated on Pinus elliottii Seeds in Brazil.

An elevated incidence of the fungal genus Fusarium was ascertained during a health quality analysis of a batch of Pinus elliottii Englm. seeds obtained from the Florestas Institute for Agricultural and Forest Research (Fundação Estadual de Pesquisa Agropecuária [FEPAGRO] Florestas) in Santa Maria (29° 39' 55″ S and 53° 54' 45″ W), state of Rio Grande do Sul, Brazil. This genus comprised about 75% of all fungal genera observed in a blotter test. The fungus was then isolated and purified to perform pathogenicity tests. Healthy seeds of P. elliottii were inoculated by contact with fungal mycelium for 48 h (3). Forty-two days after inoculation, a reduction was observed in the germination potential of the seeds; however, those seeds that germinated developed normally until, as seedlings, they suffered damping-off. Fusarium was isolated from the affected vegetal material by transferring mycelium tips to potato dextrose agar (PDA) medium in petri dishes in order to morphologically identify the species. After 72 h, a tan mycelial pad 5.5 cm in diameter had formed. After transfer to carnation leaf agar (CLA), pale orange sporodochia that formed macroconidia could be observed. The macronidia were relatively short and narrow (40.2 × 4.7 µm), each containing a mean of 5 septa; the apical cell was pointed, while the basal one was foot-shaped (2,4). The chlamydospores formed in clusters, while the conidiogenous cells could be seen on top of monophialides. Primer pairs ITS1 and ITS4, EF1-T and EF1-567R, and ßtub-F and ßtub were employed to amplify the three regions ITS1.8S ITS2, elongation factor - 1α (TEF 1-α), and ß-tubulin, respectively. The sequences of these three regions showed 97, 95, and 99% of similarity with Fusarium sambucinum Fückel, respectively. The pathogen was reinoculated on P. elliottii seeds in order to complete Koch's postulates. The pathogenicity test was repeated with the same conditions described before and the results were confirmed. No occurrence of damping-off was observed in the control seedlings. The inoculated seedlings showed, besides damping-off, a visible reduction in root system expansion as well as reductions in fresh and dry tissue weight. F. sambucinum has already been reported on P. radiata D. Don in New Zealand, causing root rot and dieback (1); however, in Brazil, the present study is, to the best of our knowledge, the first to report the association of this pathogen with P. elliottii. References: (1) M. A. Dick and K. Dobbie. N. Z. Plant Prot. 55:58, 2002. (2) W. Gerlach and H. Nirenberg. The Genus Fusarium - A Pictorial Atlas. Biologische Bundesanstalt für Land - und. Forstwirtschaft, Berlin, 1982. (3) M. Lazarotto et al. Summa Phytopathol. 36:134, 2010. (4) J. F. Leslie and B. A. Summerell. The Fusarium Laboratory Manual, 1st ed. Wiley-Blackwell, Hoboken, NJ, 2006.