Additive Manufacturing of Devices Used for Collection and Application of Cereal Rust Urediniospores

Abstract

Optimized inoculation procedures are an important consideration in achieving repeatable plant infection when working with biotrophic rust fungi. Several plant pathology laboratories specializing in rust research employ a system where the collection and application of fungal spores are accomplished using an exchangeable gelatin capsule. Urediniospores are collected from erumpent pustules on plant surfaces into a capsule fitted to a cyclone collector controlled by a vacuum pump. By adding light mineral oil to the same capsule, the spore suspension is then sprayed onto plants by means of a dedicated atomizer (inoculator) connected to an air pressure source. Although devices are not commercially available, modern day technologies provide an opportunity to efficiently design and manufacture collectors and inoculators. Using a process called Additive Manufacturing (AM), also known as “3D printing,” the bodies of a collector and inoculator were digitally designed and then laser-sintered in nylon. Depending on availability, copper or aluminum tubes were fitted to the bodies of both devices afterward to either facilitate directed collection of spores from rust pustules on plant surfaces or act as a siphon tube to deliver the spore suspension contained in the capsule. No statistical differences were found between AM and metal inoculators for spray delivery time or spore deposition per unit area. In replicated collection and inoculation tests of wheat seedlings with urediniospore bulks or single pustule collections of Puccinia triticina and P. graminis f. sp. tritici , the causal organisms of leaf rust and stem rust, consistent and satisfactory infection levels were achieved. Immersing used devices in acetone for 60 s followed by a 2 h heat treatment at 75°C produced no contaminant infection in follow-up tests.

A new and effective method to induce infection of Phyllachora maydis into corn for tar spot studies in controlled environments

“…With a vacuum device and syringe tip, contamination of other fungal spores was minimized by targeting individual stromata for spore collection following a similar approach used in rust pathogenicity studies with Puccinia triticina and P. graminis f. sp. tritici, where spores are collected via vacuum from infected Triticum aestivum L. (wheat) and used for inoculation [20]. The strategy of using a vacuum collection device or syringe tip like the one used with our method for tar spot has not been used for Phyllachora species previously.…”

“…The strategy of using a vacuum collection device or syringe tip like the one used with our method for tar spot has not been used for Phyllachora species previously. This is likely because fruiting bodies of P. maydis are within stromata [9], and spore extrusion may require stimuli analogous to spores of Puccinia spp., which are collected from leaf or stem surfaces of wheat [20]. Our methodology allowed us to collect spores from various sources including green and dried corn leaves.…”

Fig rust caused by Phakopsora nishidana in South Africa

“…Procedures described by Boshoff et al (2020) were used to heat-shock the spores and for inoculation of plants and subsequent incubation. After 3 weeks, urediniospores were collected from the Kadota leaves into size 00 gelatine capsules, by connecting an air vacuum to a cyclone spore collector (Pretorius et al, 2019).…”

Accomplishments in wheat rust research in South Africa

“…and application of small amounts of urediniospores is essential. Because these specialised items are not commercially available, Pretorius et al 24 developed an additive manufacturing process to assemble spore collectors and atomisers through 3D printing. Using these devices, traditional race analysis is done by infecting seedlings of a predetermined (differential) set of wheat host lines with a rust isolate.…”