TP Phytopathologie

Practice 6. SOIL AND AQUATIC FUNGI: OBSERVATION. ANAMORPHIC FUNGI. PITIACEOUS FUNGI.

Introduction: 

The laboratory session focused on the observation and identification of soil and aquatic fungi anamorphs, specifically pitaceae. The objective was to apply various isolation techniques to gain insights into fungal populations present in substrates. The examination aimed to identify and comprehend the diverse fungal species encountered in the provided samples using distinct methods, such as Serial Dilutions and the Leaves Method.

1. Enumeration (Serial Dilutions Method): 

During the experimentation utilizing the Serial Dilutions method, the first dilution was enumerated, resulting in the count of 20 colonies. The calculated population was determined using the formula provided in the protocol:

[pic 1]

For instance, considering the observation of 20 colonies per plate with 0.1ml plated from a 1:1000 dilution:

[pic 2]

[pic 3]

[pic 4]

1. Identification (Dilution Methods and Leaves Method): 

In the Dilution Methods, Cladosporium and Conidium were identified. Cladosporium displayed distinct characteristics, while Conidium was observed as round and elongated with septum, consistent with the details specified in the protocol. Concerning the Leaves Method, the identification included Oogonia with spines and Oomycetes with thicker hyphae, specifically Zoosporongus, characterized by a thicker hype.

[pic 5][pic 6]

Practices 7 Phyto parasitic bacteria (1) Observation of symptoms Isolation of necrosis

Introduction: This practice focuses on extracting Phyto parasitic bacteria from necrotic plant tissues through two primary methods: the Dilution Method and the Direct Method.

1. Dilution Method for Isolation:

• Step 1: Tissue Collection and Soaking:

1. Collection: Small pieces are gathered from the specific necrotic area of the leaf affected by bacterial action.
2. Soaking in Sterile Water: These tissue pieces are placed in tube 1 containing sterile water, previously washed with soap and water.

Purpose: This soaking process serves multiple purposes:

• Hydration: Rehydrates the dried-out necrotic tissues, potentially reviving dormant bacterial activity or growth.
• Surface Cleaning: Helps remove superficial contaminants or debris from the tissue surface, enhancing the isolation process.
• Conditioning: Facilitates the release of bacteria, if present, into the water, aiding in subsequent steps of analysis.

• Step 2: Serial Dilution and Observation:

1. Serial Dilution: Following the soaking period, perform the serial dilution method. Process: Transfer a drop of the liquid containing the soaked tissues onto a Petri dish using a seeding loop.
2. Cover and Store: Cover the Petri dish with parafilm and store it for subsequent observation.

Purpose: The half-hour soaking period prepares the liquid suspension for further processing, especially for the subsequent serial dilution step, by facilitating bacterial release and tissue hydration.

1. Direct Method of Isolation :

• Step 1: Tissue Collection and Washing:

1. Collection: Necrotic tissues from infected plants are gathered using sterile tools.
2. Washing: These tissues are cleansed in sterile distilled water to eliminate surface contaminants.

Purpose: Cleansing tissues ensures an accurate isolation process by removing potential contaminants.

...