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Studies of interactions among cultured ericoid mycorrhizal fungi of salal (gaultheria shallon pursh)

University of British Columbia

This thesis compares molecular and cultural methods for the detection and identification of the fungal partners of mixed in vitro mycorrhizae of salal. Mixed in vitro mycorrhizae were synthesized by inoculating salal with pairs of three different fungi in all possible combinations. The fungi were sterile isolates S246 and S9, and Oidiodendron griseum. Mycorrhizae formed after three to four months and the fungi in the mycorrhizal roots were detected by (1) re-isolation and morphological identification of the cultured fungi and (2) direct polymerase chain reaction (PCR) amplification of the ribosomal internal transcribed spacer regions using primers ITS1-F and ITS4, followed by restriction fragment length polymorphism (RFLP) identification. The detection results from the synthesized mycorrhizae were compared with PCR/RFLP detection from mixtures of fungal DNAs at known concentrations. The O. griseum. DNA could not be detected at any concentration when it was combined with DNA from either S9 or S246, indicating a PCR bias against O. griseum. This bias explained why O. griseum never appeared in the RFLP patterns from amplified DNA from synthesized mycorrhizae despite being re-isolated from 71% of the root segments. The RFLPs proved useful in clearly indicating mixtures of the two sterile isolates, especially because mixtures of sterile isolates were difficult to recognize based on morphology. This thesis also compared culture and PCR/RFLP methods to identify ericoid mycorrhizal fungi colonizing mapped roots. Two mycorrhizal fungi often occupied the same 5 mm segment of a salal root. From the roots located under both inocula with the same chance for colonization, 70% of the cultures recovered were mixed cultures when a sporulating and sterile fungi were used as inoculum. The three ericoid mycorrhizal fungi mentioned above were paired in culture to look at their competitive abilities. All pairings showed deadlock, an inability of the paired species to invade the territory occupied by the other one. For better observation of the colonized roots within the agar medium, phytagel was substituted by granulated agar. Phytagel improved the clarity of the gel and significantly improved the shoot growth of salal; the percent root colonization did not show any significant difference.

Thesis/Dissertation

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2009-05-19

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http://hdl.handle.net/2429/7958

Botany

University of British Columbia

UBCV

DSpace