Establishment and application of an in vitro methodology to study the effects of organic acids on coliform and lactic acid bacteria in the proximal part of the gastrointestinal tract of piglets

Abstract

In order to screen various feed additives for their anti-microbial effects in gastrointestinal content and thereby find candidates to replace antibiotic growth promoters in feed for piglets, a batch culture system was established to simulate the major environmental conditions in the stomach and proximal part of the small intestine of piglets. The applicability of the in vitro methodology was tested in typical screening experiments. The batch culture system was adjustable in regard to pH, temperature and incubation period. In addition, an anoxic atmosphere was created. The effect of potassium-diformate (PDF), a salt of formic acid, on the population changes of coliform and lactic acid bacteria was studied in stomach content at pH 3, 4 and 5, and in content from the proximal part of the small intestine at pH 5, 6 and 7. Furthermore, the anti-microbial effects of six different organic acids (formic-, propionic-, butyric-, lactic-, benzoic- and fumaric acid) were compared in stomach content (pH 4.5) and in small intestinal content (pH 5.5). Finally, a dose–response experiment involving stomach content (pH 4.5) with added benzoic acid was conducted. In contrast to lactic acid bacteria, coliform bacteria were unable to grow in stomach contents. A pH-dependent inhibition of bacterial growth was demonstrated in the two culture media. In particular, the growth rate of coliform bacteria was strongly influenced by pH, thus, showing generation times of 71 and 25 min at pH 5 and 7, respectively. Addition of PDF reduced the growth of coliform bacteria and lactic acid bacteria. Both type and dose-level of the acids employed affected the magnitude of response. In conclusion, the in vitro system reported offers a reliable method to investigate alterations in the microbiota in response to different environmental conditions and provides a useful approach to the screening of products to be tested in vivo.