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First Report of Neonectria neomacrospora Causing European Silver Fir (Abies alba) Dieback in France

Saurat, C., Schenck, N., Fourrier-Jeandel, C., Bialais, L., Daubrée, J.-B., Ioos, R.
Plant disease 2019 v.103 no.2 pp. 365
Abies alba, Christmas trees, DNA, Neonectria, agar, branches, chloramphenicol, conidia, culture media, death, dieback, economic impact, ethanol, fungi, internal transcribed spacers, malt, malting, mountains, mycelium, necrosis, oligodeoxyribonucleotides, plantations, polymerase chain reaction, shoots, tissues, tree mortality, Belgium, Denmark, France, Norway, Sweden, United Kingdom
Neonectria neomacrospora is responsible for a severe canker disease on fir (Abies spp.) in Norway, Denmark (EPPO 2013), Sweden (Pettersson et al. 2016), the United Kingdom (Pérez-Sierra et al. 2016), and Belgium (Schmitz et al. 2017). The disease is characterized by the death of shoots and branches, cankers, and resin flow, and it may lead to tree mortality. Severe incidences have been observed, notably in Christmas tree plantations (Abies spp.), and N. neomacrospora was added to the EPPO alert list in 2017. In December 2017, extensive dieback symptoms, reddening to browning of the crown with shoot necroses especially on the distal part, and twig bulges were observed in a natural Abies alba stand (state forest) located in the Pyrenean mountains (Cauterets, 0°10′12.5″ W; 42°50′24.5″ N). At least 10% of the 50-ha stand of A. alba was affected. Symptomatic tissues from shoots were collected and sent to the lab for analysis. After removal of the needles, symptomatic sections of necrotic tissues were surface disinfected with 70% ethanol. Pieces of tissue including necrosis edges were plated onto malt agar supplemented with 100 mg/ml of chloramphenicol and incubated for 5 to 8 days at 22°C. A homogeneous beige to yellow-ochre mycelium consistently grew out from the tissue pieces and was transferred to potato dextrose agar (PDA) and malt agar media and was incubated at 22°C under light conditions (12 h/12 h light alternation). After 6 to 8 days, a dense and felted mycelium grew quite slowly and appeared whitish to cream with a yellowish tint on PDA or malt agar. Observed on PDA from the reverse side, the colony looked first beige then turned to tan or brown. The colony produced two types of conidia that could be observed and characterized under the microscope: (i) typical multiseptate (mostly three septa), hyaline, straight or sometimes slightly curved macroconidia, 36.3 to 53.8 (44.9) × 5 to 5.6 (5.4) µm and (ii) hyaline ovoid to cylindrical microconidia 7.5 to 12.5 (9.4) × 2.7 to 3.9 (3.2) µm. These conidial features were consistent with Neonectria conidia. To further assess the identity of the species, DNA was extracted from the pure culture obtained, and the internal transcribed spacer (ITS) region was amplified by PCR using the ITS5/ITS4 primer pair. The nucleotide sequence was deposited in GenBank (accession no. MH396677). BLAST analysis of the sequence showed 100% identity with recently available N. neomacrospora reference ITS sequences (Schmitz et al. [2017], MG049669; Pettersson et al. [2016], KT383061). These molecular results were in line with the morphological features observed. To our knowledge, this is the first official report of N. neomacrospora causing European silver fir dieback in France. Because this fungus was shown to be pathogenic on A. alba (Nielsen et al. 2017) and is already causing severe dieback in Scandinavian countries, the spread and establishment of N. neomacrospora, especially in Christmas trees production areas, could have severe economic consequences.