KEY TAKEAWAYS
- Synbiotics and direct fed microbials can be used as an alternative to reduce the use of antibiotics in beef cattle and enhance the effectiveness of probiotics or direct fed microbials used in the industry.
- Prebiotics that enhance the abundance of beneficial microbes in the gut and additionally provide an opportunity to increase the potential of a direct fed microbial to grow and colonize in the cattle gut can be used to manipulate the rumen microbiome.
- Substrates that enhanced the antimicrobial activity against pathogens in Generally Regarded As Safe (GRAS) approved strains and closely related strains that are capable of inhibiting opportunistic pathogens in the rumen were identified.
BACKGROUND
Livestock agriculture, and especially the beef industry, has come under great scrutiny for the prophylactic use of antibiotics (especially antibiotics used as growth promotants). Thus, to ensure antibiotic stewardship while improving health, well-being and performance of the animal to meet future demands, novel alternatives for antibiotics are greatly needed. One of the largest animal health related issues in the current beef industry is liver abscesses. Liver abscesses cause an economic loss to the producer and the packer and are the leading cause of beef condemnation in the United States. While the liver is not a significant loss, abscesses formed results in reduced animal performance and carcass yield from reduced feed intake, reduced weight gain, decreased feed efficiency, and decreased carcass dressing percentage. Currently, the most effective method for controlling liver abscesses is the use of antibiotics belonging to the macrolide family, named Tylosin. Even with the use of antibiotics, 10- 20% of fed cattle are affected and form liver abscesses. However, apart from prophylactic use of antibiotics, a proven strategy for reducing liver abscesses has not been developed. Direct fed microbials (DFMs) are potential promising alternatives to reduce antibiotic use in beef cattle production and to improve animal health, well-being and productivity. Over the last 2 years, novel generally regarded as safe (GRAS) approved DFMs have been developed and have demonstrated that such microbials can be used to decrease pathogens such as
Fusobacterium necrophorum and
Streptococcus bovis using in-vitro and in-vivo studies. To further enhance the activity of the DFMs identified, pre-biotics (substrates that probiotic or DFMs use for growth) in combination with the DFM to develop a symbiotic that would have enhanced capability to reduce antibiotic use were proposed. Here, to improve the applicability of the GRAS approved isolates identified to use as an alternative to antibiotics and to increase the naturally occurring “beneficial microbes” in the gut, experiments were conducted to identify substrates that enhance the growth of the GRAS approved isolates to develop novel feed additives that increase the growth and inhibitory ability against pathogens.
Freshly grown overnight cultures of
Bacillus spp. and
Anerobacillus spp. were each inoculated on 95 different substrates to identify carbon sources that increased growth. Additionally, each microbe was also grown in yeast media, and in the presence of 1% fructo-oligosaccharides, galacto-oligosaccharides, and 2-2-Fucosyllactose. Following growth, each organism grown in each substrate was screened against
Streptococcus bovis to identify substrates that enhanced inhibition. Each culture was grown in BHI overnight. 10 ml of each culture was centrifuged and resuspended in 10ml of minimal media and 100 ml of each isolate was used for inoculation of each substrate. The substrates were organized on Biolog anaerobic plates with a water control and 95 different carbon sources. Following incubation of the cultures in biology plates, growth was read at 24 and 48 hr intervals and functional screening of each bacterial strain was conducted using
S. bovis ATCC 70041 on low profile Bioassay plates using BHI agar medium. A sterilized 96 well plate replicator was utilized to replicate each isolate on all substrates on a lawn of
S. bovis ATCC 70041. Between each replication, the plate replicator was sterilized. The inoculated plates were incubated at 37°C overnight and were inspected for the presence of clearing zones at 24 and 48 hour intervals. The inhibition assays were performed by screening on 0.5 McFarland
Streptococcus bovis cultures.
Substrates that enhance the antimicrobial activity against pathogens in GRAS approved strains and closely related strains that are capable of inhibiting opportunistic pathogens in the rumen (
F. necrophorum and
S. bovis) were identified. Among these organisms and substrate combinations,
B. subtilis,
B. licheniformis and
B. pumilis were seen to be the most effective GRAS-approved bacteria species that can inhibit
F. necrophorum and
S. bovis in the rumen. Fructo-oligosaccharides and galacto-oligosaccharides were identified to be the most beneficial prebiotics to increase inhibitory features in the
Bacillus genera. While limited studies have looked at developing synbiotics to improve bacterial strains, this strategy provides a unique opportunity to increase naturally occurring beneficial microbes of similar metabolic capabilities as the GRAS approved strains in the host animal to increase. Thus, addition of prebiotics helps increase the abundance of beneficial microbes in the gut and additionally provides an opportunity to increase the potential effect of the direct fed microbial on the host.
Implications
This project develops novel symbiotic combinations (prebiotic + direct fed microbial) that can be used as alternatives to antibiotic use. Additionally, this innovative approach to identify novel symbiotics as alternatives to antibiotic use will help ensure “antibiotic stewardship” and safer beef production practices. This research will help provide new methods for reducing antibiotic use while maintaining animal health and performance.