First Report of Vascular Wilt on Peanut in Mexico Caused by Fusarium incarnatum

Abstract

During the fall of 2020, wilt symptoms were presented in a commercial peanut field (Arachis hypogaea L.), variety 'CHAMPS' in Buenavista de Benito Juárez, México (18.460501 N, 98.627100 W). A peanut field was scout 80 days after planting, and plants presenting symptoms of root and crown rot, vascular chlorosis, and tissue death, were sampled. Disease incidence was estimated at 55% of the field. Isolations of the pathogen were made from stem and root tissues. These samples were disinfected by immersion in 1% sodium hypochlorite (NaClO) for 3 min and immersion in 70% ethanol for 1 min with 3 rinses with sterile distilled water. Subsequently, 0.5 cm fragments were removed and placed on media of potato dextrose agar (PDA) and Rose Bengal. Petri dishes were incubated in complete darkness at 26 °C for 7 days. Abundant aerial white mycelium was observed, which turned tan to brown and showed a slightly orange color on the back of the plate. Finally, pure cultures were obtained by single sporing (Aslam et al. 2020). Colonies identified as Fusarium spp. (Leslie and Summerell, 2006) were sub cultured on PDA agar media and Spezieller Nährstoffarmer (Pérez-Vázquez et al. 2022) to observe microscopic characteristics of ten isolates. Colonies of a representative strain (MA-PET-03) produced hyaline septate hyphae, macroconidia dorsoventrally curved, tapering towards both ends of 51–57 × 4.6–5.4 μm (n = 80) with most having 7 septa. Microconidia were unicellular, nonseptate, hyaline, and ovoid, 12.4–20.6 × 3.6–4.1 μm (n = 80). Chlamydospores were abundant and globose 5–11 μm diam (n = 80), intercalary, and solitary in short chains (Figure 1). The observed microscopic characteristics correspond to the description of Fusarium incarnatum (Khoa et al. 2006; Leslie and Summerell, 2006; Xia et al. 2019). The molecular analyses were done with genomic DNA extracted as previously reported by Pérez et al. (2022). A region from the translation elongation factor gene was PCR amplified using EF688/EF1251 primers (Alves et al. 2008) and from the calmodulin (CMDA) gene, using CALDF1 and CALDR1 primers (Noel et al. 2022). The corresponding PCR products were purified with the Gen Elute™ PCR Clean-Up Kit from Sigma-Aldrich Co. (St. Louis Mo. USA) and sequenced at Macrogen Inc. (Seoul, South Korea). The phylogenetic analysis was inferred using the Bayesian Inference method with 1 million generations, final standard deviation was 0.008516. The nucleotide substitution model for Calmodulin (CMDA) was GTR + G and for TEF1 GTR + I + G. This analysis showed that strain MA-PET-03 shared 100% identity (Figure 2) with F. incarnatum ex-type strain CBS 132.73 (CMDA: MN170342; TEF1: JMN1704761) from Pointed gourd (Trichosanthes dioica) in Malawi Africa. The sequences of strain MA-PET-03 were deposited in GenBank (CMDA: OQ679820; TEF1: OQ679821). The pathogenicity tests were carried out with a total of 20 peanut plants, variety 'CHAMPS', 18 days after having been sown in groups of five seeds in 250 g plastic pots, containing a sterilized mixture of Peatmoss and Agrellite (1:1 v./v), with four repetitions. The seeds were inoculated by immersion in 20 mL of spore suspension (106 conidia/mL) isolated from F. incarnatum for 10 min. The plants were maintained in a greenhouse (70% relative humidity and 28 °C) until the appearance of disease symptoms of. Likewise, 10 control plants were inoculated with sterile water. The experiment was repeated twice. The symptoms developed 15 days after inoculation, the plants presented symptoms of chlorosis, wilting of leaves, stems, and roots, a manifestation similar to that observed in the field, while the control plants remained healthy. F. incarnatum was consistently reisolated from inoculated stems and roots and identified by the microscopic characteristics described above. Peanut leaf blight and wilt disease caused by F. incarnatum has been reported in India (Thirumalaisamy et al. 2019). This first report emphasizes that this phytopathogen is a new threat for peanut producers in Mexico, which is why our finding suggests the need to seek new strategies for its control.