Team:SDU-Denmark/Proof
Proof of Concept
Bacteriocins - They work!
During the project we designed BioBricks, which contain the genes encoding a single bacteriocin or a hybrid bacteriocin. We purified the bacteriocins using the IMPACT method and determined the respective concentrations using a Bradford standard protein assay with Bovine Serum Albumin (BSA). The first key criteria in order to purify the bacteriocins were to correctly incorporate the bacteriocin into the IMPACT vector pTXB1. We identified successful ligation between our bacteriocins and the pTXB1 with colony PCR, thus the first key criteria were achieved. Check out demonstrate and results for specification. Let the purification begin!
Purified bacteriocins inhibit the growth of MRSA and P. aeruginosa
Our MIC test shows growth inhibition of the strains MRSA:USA300 (S.aureus), MRSA:CC398 (S.aureus), hetero-VISA (S.aureus) and Pseudomonas aeruginosa. The bacteriocins LacticinQ and the hybrid Laterosporulin-ThuricinS did not elicit their activity towards P. aeruginosa. However, the absence of inhibition could be due to a higher MIC value, i.e. the need of a higher concentration to inhibit growth than used in the MIC test, which still leaves the hybrid as a potential inhibitor of P. aeruginosa.
Synergistic effect of bacteriocins
Compared to traditional antibiotic, our results show that the bacteriocins has similar or better effect. The bacteriocins show inhibition of growth towards MRSA strains and P. aeruginosa at concentrations where the traditional antibiotics does not elicit an effect. The bacteriocins therefore show promising results to support the idea of bacteriocins being a supplement for traditional antibiotics of which multiresistant bacterias has become less sensitive towards.
The bacteriocins inhibit growth of the respective strains BUT the bacteriocins also showed synergistic effect. The synergistic effect is shown by a decrease in MIC when the bacteriocin Laterosporulin and ThuricinS are combined as a hybrid compared to their MIC value as single proteins. The hybrid LacticinQ-LacticinZ also shows a decreased MIC compared to LacticinQ as a single acting bacteriocin. The absence of inhibition towards P. aeruginosa by Laterosporulin-ThuricinS when in a hybrid compared to their single protein effect could be stated as a loss of effect. However, the MIC values overall decrease towards the S. aureus strains indicating a stronger effect. The observation leaves the possibility of a gain of effect towards strains not tested in the MIC test. The gain of effect could possible prove another characteristic of the designed hybrid bacteriocins. This leaves the opportunity to design an antimicrobial compound as needed.Our bacteriocins face AMR
The bacteriocins could serve as a supplement for traditional antibiotics, which bacterias are less sensitive towards, since our purified bacteriocins were capable of inhibiting the growth of methicillin resistant S. aureus strains. It could also be hypothesized that a combination of traditional antibiotics with bacteriocins could elicit a synergistic effect towards resistant bacterial strains. This leave us with the opportunity to shift the balance of resistance and prevent the evolution of antimicrobial resistance against traditional antibiotics (Dan, Y., M. P. Zacharof and R. W. Lovitt (2012)).
