Antibiotic amendment for suppression of indigenous microflora in feed sources for an Escherichia coli auxotroph lysine assay

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.

Development of a whole cell green fluorescent sensor for lysine quantification

“…In previous studies from our laboratory, bacterial growth response to lysine has been measured turbidimetrically (Zabala- Erickson et al 1999;Li and Ricke 2003) or by bioluminescence (Erickson et al 2000, Zabala-Díaz and. Both methods have drawbacks as discussed in the introduction.…”

“…Microbiological assays based on the growth response of an E. coli lysine auxotroph to external lysine, measured turbidimetrically, have been shown to be simple, inexpensive and specific (Payne et al 1977;Erickson et al 2002). The disadvantage of these methods is their relatively high level of time input ranging from 12 to 18 h (Li et al 2000;Prachaiyo and McLandsborough 2000;Erickson et al 1999). Even though some genetic alterations have been made to improve the lysine auxotrophic growth response (Li and Ricke 2003), and the OD-based growth assay can be completed in 8 h, the time required for the entire procedure still remains relatively inconvenient for a single day analysis.…”

Environmental Dissemination of FoodborneSalmonellain Preharvest Poultry Production: Reservoirs, Critical Factors, and Research Strategies

“…Such approaches have allowed for the selective growth, recovery, and quantitation of E. coli lysine auxotrophs in the presence of poultry feed backgrounds. 163,164 However, without maintenance of specific antibiotic-selective conditions, plasmids can be lost as bacterial cells undergo cell division. Insertion of antibiotic resistance genes into the chromosome can be accomplished by transposition with mobile transposons with selectable antibiotic genes.…”

Adaptation of Lactobacillus rhamnosus riboflavin assay to microtiter plates

“…In our study, L. rhamnosus antibiotic profile was determined by screening the bacterial drug susceptibility to a range of antibiotics including tetracycline, erythromycin, neomycin, nalidixic acid and kanamycin (Erickson et al, 1999;Froelich et al, 2002). Nalidixic acid was the only antibiotic from this group that L. rhamnosus exhibited resistance.…”