Métodos físicos para reducir inóculo de Sclerotium cepivorum en semilla de ajo morado (Allium sativum L.)

Introducción. En Colombia, la producción de ajo se ve afectada por patógenos, principalmente Sclerotium cepivorum, el cual se disemina por el material de siembra. Es necesario establecer tecnologías de bajo costo que permitan controlar este patógeno. Objetivo. Evaluar dos métodos de termoterapia, termoterapia por calor seco y termoterapia por inmersión en agua, para disminuir la presencia de S. cepivorum en semilla de ajo y su efecto sobre características agronómicas de plantas emergidas a partir de bulbillos tratados. Materiales y métodos. Este trabajo se llevó a cabo en el Centro de Investigación Tibaitatá de la Corporación Colombiana de Investigación Agropecuaria (AGROSAVIA), ubicado en Cundinamarca, Colombia, durante el primer semestre del 2019. Se evaluaron nueve tratamientos; cinco en termoterapia por calor seco: 35 ºC por 12 h, 35 ºC por 129,6 h, 50 ºC por 48 min, 5 ºC por 20 días, 38 °C por 1 h; dos en termoterapia por inmersión en agua caliente: 48 °C por 30 min, 50 °C por 20 min y dos controles: desinfección química (tebuconazol) y temperatura ambiente (20 ºC por 20 días), en un diseño en bloques completos al azar. Se evaluó brotación de bulbillos y crecimiento de las plantas de ajo después de aplicar termoterapia a la semilla y viabilidad de esclerocios de S. cepivorum. Resultados. El almacenamiento de la semilla a 5 ºC por 20 días aceleró su brotación, alcanzó una germinación de 80 % a los cinco días, mayor número de hojas y acumulación de biomasa. La germinación de esclerocios estuvo entre 33 y 67 %; el tratamiento que presentó la menor incidencia fue 35 ºC por 129,6 h (33 %). Conclusión. El tratamiento de termoterapia por calor seco a 35 °C por 129,6 h mostró potencial en la reducción del inóculo de S. cepivorum en semillas de ajo.

Establishment of a Cell Suspension Culture of Eysenhardtia platycarpa: Phytochemical Screening of Extracts and Evaluation of Antifungal Activity

Eysenhardtia platycarpa (Fabaceae) is a medicinal plant used in Mexico. Biotechnological studies of its use are lacking. The objective of this work was to establish a cell suspension culture (CSC) of E. platycarpa, determine the phytochemical constituents by spectrophotometric and gas chromatography‒mass spectrometry (GC‒MS) methods, evaluate its antifungal activity, and compare them with the intact plant. Friable callus and CSC were established with 2 mg/L 1-naphthaleneacetic acid plus 0.1 mg/L kinetin. The highest total phenolics of CSC was 15.6 mg gallic acid equivalents (GAE)/g dry weight and the total flavonoids content ranged from 56.2 to 104.1 µg quercetin equivalents (QE)/g dry weight. The GC‒MS analysis showed that the dichloromethane extracts of CSC, sapwood, and heartwood have a high amount of hexadecanoic acid (22.3–35.3%) and steroids (13.5–14.7%). Heartwood and sapwood defatted hexane extracts have the highest amount of stigmasterol (~23.4%) and β-sitosterol (~43%), and leaf extracts presented β-amyrin (16.3%). Methanolic leaf extracts showed mostly sugars and some polyols, mainly D-pinitol (74.3%). Compared with the intact plant, dichloromethane and fatty hexane extracts of CSC exhibited percentages of inhibition higher for Sclerotium cepivorum: 71.5% and 62.0%, respectively. The maximum inhibition for Rhizoctonia solani was with fatty hexane extracts of the sapwood (51.4%). Our study suggests that CSC extracts could be used as a possible complementary alternative to synthetic fungicides.

Visualization of Three Sclerotiniaceae Species Pathogenic on Onion Reveals Distinct Biology and Infection Strategies

Botrytis squamosa, Botrytis aclada, and Sclerotium cepivorum are three fungal species of the family Sclerotiniaceae that are pathogenic on onion. Despite their close relatedness, these fungi cause very distinct diseases, respectively called leaf blight, neck rot, and white rot, which pose serious threats to onion cultivation. The infection biology of neck rot and white rot in particular is poorly understood. In this study, we used GFP-expressing transformants of all three fungi to visualize the early phases of infection. B. squamosa entered onion leaves by growing either through stomata or into anticlinal walls of onion epidermal cells. B. aclada, known to cause post-harvest rot and spoilage of onion bulbs, did not penetrate the leaf surface but instead formed superficial colonies which produced new conidia. S. cepivorum entered onion roots via infection cushions and appressorium-like structures. In the non-host tomato, S. cepivorum also produced appressorium-like structures and infection cushions, but upon prolonged contact with the non-host the infection structures died. With this study, we have gained understanding in the infection biology and strategy of each of these onion pathogens. Moreover, by comparing the infection mechanisms we were able to increase insight into how these closely related fungi can cause such different diseases.

Establishment of a Cell Suspension Culture of Eysenhardtia platycarpa: Phytochemical Screening of extracts and Evaluation of Antifungal Activity.

Eysenhardtia platycarpa (Fabaceae) is a medicinal plant used in México and it lacks biotechnological studies for its use. The aim of this work was to establish a cell suspension cultures (CSC) of E. platycarpa, determine the phytochemical profile, and evaluate its antifungal activity. Friable callus and CSC were established with 2 mg/L 1-naphthaleneacetic acid plus 0.1 mg/L kinetin. The highest total phenolics of CSC was 15.6 mg GAE/g dry weight and the total flavonoids content ranged from 56.2 to 104.1 µg QE/g dry weight. CG‒MS analysis showed that the dichloromethane extracts of CSC, sapwood and heartwood have a high amount of hexadecanoic acid (22.3 ‒ 35.3 %) and steroids (13.5 ‒ 14.7%). Heartwood and sapwood defatted hexane extracts have the highest amount of stigmasterol (≈ 23.4%) and β-sitosterol (≈ 43%), and leaf extracts presented β-amyrin (16.3%). Methanolic leaves extracts showed mostly sugars and some polyols, mainly D-pinitol (74.3%). Dichloromethane and fatty hexane extracts of CSC exhibited the percentages inhibition higher for Sclerotium cepivorum with 71.5 and 62.0%, respectively. The maximum inhibition for Rhizoctonia solani was with fatty hexane extracts of the sapwood (51.4%). Our study suggest that CSC extracts could be used as a possible complementary alternative to synthetic fungicides.

Dynamics in Secondary Metabolite Gene Clusters in Otherwise Highly Syntenic and Stable Genomes in the Fungal Genus Botrytis

Fungi of the genus Botrytis infect >1,400 plant species and cause losses in many crops. Besides the broad host range pathogen Botrytis cinerea, most other species are restricted to a single host. Long-read technology was used to sequence genomes of eight Botrytis species, mostly pathogenic on Allium species, and the related onion white rot fungus, Sclerotium cepivorum. Most assemblies contained <100 contigs, with the Botrytis aclada genome assembled in 16 gapless chromosomes. The core genome and pan-genome of 16 Botrytis species were defined and the secretome, effector, and secondary metabolite repertoires analyzed. Among those genes, none is shared among all Allium pathogens and absent from non-Allium pathogens. The genome of each of the Allium pathogens contains 8–39 predicted effector genes that are unique for that single species, none stood out as potential determinant for host specificity. Chromosome configurations of common ancestors of the genus Botrytis and family Sclerotiniaceae were reconstructed. The genomes of B. cinerea and B. aclada were highly syntenic with only 19 rearrangements between them. Genomes of Allium pathogens were compared with ten other Botrytis species (nonpathogenic on Allium) and with 25 Leotiomycetes for their repertoire of secondary metabolite gene clusters. The pattern was complex, with several clusters displaying patchy distribution. Two clusters involved in the synthesis of phytotoxic metabolites are at distinct genomic locations in different Botrytis species. We provide evidence that the clusters for botcinic acid production in B. cinerea and Botrytis sinoallii were acquired by horizontal transfer from taxa within the same genus.