October 3 Lab

The goals for this lab were to:

• Familiarize ourselves with some of the Hyphomycetes
• Check in on our Neurospora crosses from last week
• Check in on our Ustilago infected corn from last week and stain for the fungus

The Hyphomycetes

The hyphomycetes are a broad term for conidiating fungi. These are very common and it would be a good idea to become familiar with the group in general. We looked at 12 different species throughout the lab to get a better idea of their spore shapes, hyphal forms, conidiophores and other various structures. I drew pictures of each of these fungi, and took a few photographs of some. The following images were observed from either a tape or squash mount slide. 

Trichoderma viride showing a conidiophore on the right, and after looking at the Illustrated Genera of Imperfect Fungi (IGIF, from here on out), what I can only assume is hyphae layered with conidia on the left and bottom. 

The coolest conidia we looked at today. Pestalotia conidia that bear appendages on both sides.  Hyphae threaded together on the right.

Fusarium graminearum conidia. They remind me of compartmentalized bananas. 

Monilinia fructicola. This looks like a section of the branched conidiophore on top, loose conidia in the center and an incompletely drawn hypha on bottom. IGIF shows a much better image.

Rhizoctonia solani does not have conidia or asesexual fruit bodies. My sample appears to be a bit dried out, but similar in appearance to IGIF. 

Thielaviopsis brasicola showing its aleuriospores. This also produces phialospores, but they are not shown here.

Aleuriospores of Thielaviopsis brasicola. 

Epicoccum spp. The conidia did not remain with the conidiophore in my images.

Microscopic image of Epicoccum conidia.

This Curvularia spp. reminds me of alternaria, except these are a more grey conidia. The conidia can be straight or bent under the scope.

Not the best Aspergillus niger mount that I have done. You can see conidia in chains or part of the conidiophore here. 

Botrytis cinera. The only useful part of this image is my attempt to demonstrate the slender nature of this fungus. A better image can be seen in IGIF.

Alternaria brassicola. We have seen this before, but here again is a simple drawing. The conidia are brown.

We were also going to look at Colletotrichum coccodes and Nigrospora, but they were contaminated with an unknown fungus. 

Neurospora crosses from last week

We each checked in on our Neurospora crosses from last week to see if any perithecia containing ascospores had formed. You can see growth on my plate, and you can see were the two fungi have met in the center of the plate. Interestingly enough, my plate did not bear perithecia in the center of the plate, but instead on the far ends of the plate. They were not completely developed this week, but I will try to get a better image next week.

My Neurospora cross one week after plating. 

Ustilago infected corn 

Last week we infected one-week-old corn plants with Ustilago maydis. This fungus causes corn smut and leads to large, bulbous growth on the plant after affecting plant cell growth and division. After just one week there were already visible signs of the disease.  

Damage present on the corn plant after just one week.

A better view of a small gall forming on one of my infected plants from last week. 

To confirm that a fungus was causing these galls, I removed a portion of the plant and began a staining protocol that binds a blue dye to the chitin in the fungal cell wall. To begin, I placed the excised leaf material in a plastic petri dish and submerged this leaf in a 2:1 Acetic acid: ethanol solution to clear the leaf. The leaf remained in this solution for 24 hours, was then removed from the solution, rinsed with ethanol, and placed in a microcentrifuge tube with lactophenol blue. This solution has three purposes. It kills any living specimens, preserves fungal components, and stains chitin blue. I kept the leaf in this solution for 4 hours, and then removed it into a jar of ethanol to remove any excess blue dye. I kept the leaf in this solution for several hours, then placed the leaf onto a slide, covered with a long cover slip, and then observed the locations of fungus in the plant.

The excised leaf material freshly placed in acetic acid/ethanol mix.

My stained corn leaf showing pockets of blue where the fungus is present.

The color is a bit off on my camera, but you can see blue color has remained in the veins of the plant. 

The location of a gall. You can see a congregation of blue mycelia.

A small blue lesion, likely the start of a gall.

Closed stomata on the underside of the leaf. You can see the cells dotted with blue spots,  spores perhaps?