PKS and NRPS Amplification

Amplicons acquired from the PCR were observed in 1% (w/v) agarose gel electrophoresis. Fragments amplified with K1F/MR6 primers have 1,200 – 1,400 bp (Figure 1). Fragments amplified with A3F/A7R primers have 700 – 800 bp (Not shown), encoding for conserved motifs in multidimeric peptides associated NRPS. Nonspecific amplifications were observed in both reactions.

Testing Isolated Actinomycetes Against Microbial Targets

Antimicrobial activity was observed from both actinomycetes towards distinct targets. Antifungal activity was observed from AR against fungal contaminants H1 and H2. The red pigment secreted by AR in solid media was not observed when it was cultured in liquid media. Antifungal activity was observed when solid media discs containing the red pigment were tested against fungal contaminants, as opposed to having the supernatants from the liquid cultures that were absent of the red pigment (Figure 2). No antifungal activity was observed from the second actinomycete, AV, given that it presented dearly identical fungal growth in both antibiograms. Antibacterial activity was observed from AV against Staphylococcus aureus, but not towards Pseudomonas aeruginosa, while AR had no apparent effect over either bacterium. As with AR’s pigment, AV secreted a yellow pigment in solid media only. Media disks containing the yellow pigment were effective against S. aureus presenting an inhibition zone of approximately 0.5 cm.

 

In the Near Future

We expect to use additional whole-cell techniques in order to detect antimicrobial activity from cultivable microbes. Also, we intend to verify whether the antimicrobial substances produced by AR and AV are novel.