|LINE, ERIC - US Department Of Agriculture (USDA)|
Submitted to: Meeting Abstract
Publication Type: Abstract Only
Publication Acceptance Date: 5/21/2014
Publication Date: N/A
Technical Abstract: Antibiotic resistance is forcing us to revisit when, where, how and how much we use antibiotics. Bacteriophage endolysins show great promise as alternative antimicrobials with the added advantage that they are highly refractory to resistance development. As protein antimicrobials, there are hurdles to commercialization that have, in part, limited the acceptance of endolysins for therapeutic applications. However, there are numerous non- or sub-therapeutic uses of antibiotics where endolysins can have immediate application and fulfill a growing need while maintaining a low risk of resistant strain development. Ethanolic fermentations in the biofuel industry are contaminated with Lactobacillus contaminants (e.g. L. fermentum, L. reuteri) that drive down the pH, killing the yeast prior to optimial ethanol production. Endolysins ('Sa2 streptooccal prophage endolysin; L. casei phage endolysin, LysA) that are lytic for these bacteria can reduce the contamination up to 2.5 logs (CFU) when added to mock fermentation as purified protein. The cost of purified protein for large scale fermentations would be prohibitive. However, when expressed within the fermentative yeast, the lytic enzymes protect mock fermentations from contaminating Lactobacillus (presumably due to release of the lysin from yeast cells undergoing normal death/lysis). This avoids the need for secretion of the lysin, which can often result in unwanted side effects from abnormal glycosylation (improper folding, reduced activity). The known anti-biofilm activity of endolysins should also reduce the down-time required for decontamination to rid fermentation plants of bacterial contaminants. This strategy is expected to reduce the need to treat contaminated fermentations with antibiotics (or caustics to increase the pH) thus giving added value to the fermentation residue (mash) that is an important coproduct of the biofuel industry (sold as animal feed). As an extension of phage endolysin expression in yeast and the regulatory goal to reduce the use of antibiotics in animal feed, we have also expressed two phage endolysins (PlyCp26F and PlyCp39O) in yeast that are lytic for Clostridium perfringens, a major causative agent of Necrotic enteritis in chickens, and a disease that costs the poultry industry >$2 billion dollars annually. When purified, the yeast expressed endolysins maintain their anti-C. perfringens activity. The transgenic yeast are currently being tested in poultry feeding trials to determine if they can replace the use of sub-therapeutic antibiotic growth promotants in poultry feed. Currently the lysin transgenes are expressed on high copy vectors in laboratory yeast strains and will need to be integrated into the genome of industrial lignocellulosic fermenting yeast in order to make these approaches commercially viable.