No abstract

Growth responses of lysine auxotrophic mutants of Escherichia coli have been used as a measurement of bioavailable lysine in protein sources and animal feeds. Sterilizing feed samples by autoclaving to eliminate non‐specific background growth of indigenous feed micro‐organisms prior to conducting the bacterial assay may introduce chemical and physical alterations to the feeds, influencing the estimation of available feed lysine. In this study, an antibiotic‐ and antifungal‐supplemented medium was constructed to support growth of an E. coli lysine auxotroph assay organism, and was tested for its ability to repress indigenous bacterial and fungal growth in feed samples. To determine which antibiotics to include, an ampicillin‐sensitive E. coli lysine mutant strain (ATCC no. 23812) was screened for antibiotic resistance and transformed with a plasmid carrying an ampicillin resistance gene. Maximum optical density quantitative response of the E. coli auxotroph to lysine was not altered by the antibiotic medium amendments (ampicillin, novobiocin and cycloheximide). Indigenous microfloral growth in a variety of typical animal feeds was suppressed in the presence of the antistatic agents. The estimated lysine recovery was 91·6% and 98·1% when the medium was used in an assay of available lysine in a lysine‐supplemented feed. This indicates that the antibiotic‐amended basal medium can be used for the E. coli‐determined lysine availability of a variety of animal feeds without prior sterilization of the feed sources.

Microbiological assays involving Escherichia coli lysine auxotrophs must be optimized to facilitate routine use. Our objectives in this study were to characterize growth of an auxotrophic E. coli lysine mutant (American Type Culture Collection strain #23812) and examine the effect of agitation on E. coli mutant growth. A defined minimal salts basal medium was used and supplemented with various lysine concentrations. The E. coli lysine auxotroph responded to increasing lysine concentration with increasing optical density. When maximum optical density (MOD) was determined for the auxotroph, a linear increase was obtained as lysine concentrations were increased (R2± 0.96) for both agitation and static cultures. Growth rates were not significantly (p > 0.05) affected by lysine concentrations, cultural conditions or their combined effect. However, growth with agitation significantly (p < 0.05) reduced the assay time by shortening the lag phase and causing stationary phase to occur earlier. The values of R2 (± 0.96) relatively remained constant over the range while the bacterial population were in the stationary phase. In conclusion, the lysine growth assay using the E. coli lysine auxotroph can be made more rapid by agitating the culture during incubation.

No abstract

Optimizing economic utilization of feed protein sources for poultry nutritional requiremens is difficult to achieve given the varied protein quality of the respective sources. Although there are several limiting amino acids in feeds that would benefit from development of rapid and more reliable bioavailability assays, lysine is of key importance since this amino acid is usually the first or second limiting amino acid in poultry feeds and is susceptible to processing treatments. However, to incorporate incoming sources in the most cost‐effective manner, accurate and timely prediction of lysine bioavailability prior to use is desired to achieve a consistent nutritional value. Animal bioassays involving chicks are one of the standard accepted practices for evaluating protein quality and amino acid bioavailability, but such assays have several limitations. Alternative in vitro tests that accurately predict lysine bioavailability for feed proteins would solve many of the problems associated with currently used animal bioassays. The expected focus should be on the development of more rapid in vitro lysine bioavailability assays that could be easily used for evaluation of poultry feed protein sources.