First confirmed report of Pythium aphanidermatum causing Phaseolus vulgaris root rot in Sinaloa, Mexico
Snap bean (Phaseolus vulgaris L.) is an important crop in Sinaloa, Mexico where 2,076 ha were cultivated in 2024, representing 21.88% of the national production area (DGSIAP 2026). In December 2024, root rot was observed in ~ 40% 4‑month‑old P. vulgaris plants grown in a 5‑ha outdoor commercial field in Culiacán, Sinaloa (24°14'44.7"N 107°10'59.6"W). Plants also exhibited crown rot, stunted growth, wilting, leaf yellowing, and partial foliage necrosis and defoliation, and plant death. Thirty pieces (1‑cm²) of rotted tissue were collected from the roots of 15 randomly selected diseased plants. The fragments were surface‑sterilized in 1% NaClO, rinsed in sterile distilled water and dried on paper. Five pieces were placed equidistantly per Petri dish containing PDA supplemented with streptomycin sulfate (0.3 g L⁻¹) and incubated at 30 °C for 5 days. Out of the 6 isolates generated, 3 were selected for identification. The isolates were plated on V8 medium (800 mL distilled water, 200 mL V8 juice, 2 g CaCO₃, and 15 g agar) and incubated for 7 days at 30 °C. Cultures developed white, dense, cottony, aerial, and well‑branched mycelia; the hyphae were hyaline and coenocytic, and the sporangia were filamentous, lobulate, and irregular in shape. These traits were consistent with Pythium aphanidermatum (Watanabe 2002; Castro-Diego et al. 2025). PCR analysis was performed with the internal transcribed spacers ITS1 and ITS4 (White et al. 1990) and the cytochrome oxidase c subunit 1 (COI) (Robideau et al. 2011). The sequences were deposited in GenBank under PX671483, PX671484, and PX671485 for ITS, PX677389, PX677390, and PX677391 for COI, corresponding to isolates PYTHFRI3CULSIN, PYTHFRI1CULSIN, and PYTHFRI8CULSIN, respectively. BLAST analysis showed that PX671483, PX671484, and PX671485 shared 100% identity with P. aphanidermatum sequence HQ643439 (555/555 bp, 664/664 bp, and 739/739 bp, respectively), and that PX677389, PX677390, and PX677391 also exhibited 100% identity with sequence HQ708486 (596/596 bp, 628/628 bp, and 626/626 bp, respectively) (Robideau et al. 2011). Phylogenetic analyses using the Neighbor‑Joining method with COI sequences and 1,000 bootstrap replications in MEGA 11 placed the three isolates within the P. aphanidermatum clade. To verify the pathogenicity of the isolates, 2 pathogenicity assays were conducted. In each assay, 10 P. vulgaris seedlings were individually inoculated at the stem base with 5 mL of inoculum (0.25 g of macerated mycelium in 100 mL of sterile water) 15 days after emergence. An additional 10 seedlings were inoculated with sterile water to serve as controls. All plants were maintained in a growth chamber at 28 ± 2 °C, 70 ± 5% RH, and a 12:12 h photoperiod for 30 days. Inoculated plants developed symptoms similar to those observed in the field, whereas control plants remained asymptomatic. Pythium aphanidermatum was successfully reisolated and identified from symptomatic root tissues, fulfilling Koch’s postulates, while no isolates were obtained from control plants. Although P. aphanidermatum was previously reported on P. vulgaris in Mexico (McGuire and Crandall 1967), details on methodology and isolation site were not provided. This study represents the first confirmed occurrence of P. aphanidermatum causing root rot in P. vulgaris in Mexico supported by both morphological and molecular evidence and the first documented case in the state of Sinaloa.