Molecular Identification and Culture of Fungi Native to Heavy Metal Contaminated Kam Kotia Mine

Contaminated soil from the Kam Kotia mine site and comparable uncontaminated soil
(control treatment) was analyzed by the Queen’s Analytical Services Unit using
Inductively Coupled Plasma Atomic Emission Spectroscopy (ICP-AES). The copper,
zinc, arsenic and sulfur content of the soil was much higher in contaminated soil than in
the control soil analyzed (see Figure 1C). Soil was also collected from the root systems
of plants at the Kam Kotia mine in Timmins, ON for use in generating trap cultures to
propagate environmental samples of fungi (see Figure 1A). Trap cultures are used to
create a microcosm of the soil ecosystem in which environmental conditions are ideal
for the growth, and later for the sporulation, of plant-associated fungi (see Figure 1B).
Maize (Zea mays) was used as the host plant for our Kam Kotia trap cultures. Fungi
were cultivated for 3 months with minimal watering to keep the maize alive, but
stressed, so that fungal symbiosis would be encouraged. After 3 months of trap culture
cultivation, root sections were harvested, DNA was extracted and the polymerase chain
reaction (PCR) was performed using primers intended to detect AMF and the resulting
fragments were subjected to DNA sequencing. The sequences were then aligned with
the MaarjAM and NCBI genomics databases using a Standard Nucleotide BLAST
search. Of the ten fragments successfully sequenced, nine were aligned most closely
with the fungi Capnobotryella sp. MA 3612 with a sequence identity of 85% for the
Internal Transcribed Spacer region. One of the fragments sequenced aligned most
closely with the fungi Aureobasidium pullulans with a sequence identity of 91% for the
ITS region.