Evaluation of a phytogenic product from two western herbal medicines to replace an antimicrobial growth promoter in poultry production

Abstract

Antimicrobial growth promoters (AGPs) are substances that are added to feed in sub-therapeutic levels in intensive animal production to improve weight gain and conversion of feed (FCR) into body mass. AGPs have been used widely as growth promoters in broiler and pig production under high-density growth conditions. Despite the observed efficacy, the use of AGPs has been criticized due to its possible role in the development of antibiotic resistance in human pathogens. Directive 183/2003 of the European Parliament, issued in 2003, banned of the use of all antibiotic agents as growth promoters in the European Union from 2006. The new context caused an increase in the search for alternative growth promoters. The aim of this study was to produce a commercially viable prophylactic antibacterial phytogenic product from Ginkgo biloba and Hypericum perforatum with a low potential to develop resistance, as an alternative to AGPs in poultry production. The first objective of this study based on earlier results of the Phytomedicine Programme, was to evaluate the activity and potentize extracts from Ginkgo biloba and Hypericum perforatum for optimal activity against relevant bacterial pathogens. Extracts of ethyl acetate (EA), hexane, dichloromethane (DCM) and acetone (in order of activity) from a direct extraction procedure of powdered G. biloba leaves were active against Enterococcus faecalis, Staphylococcus aureus and Clostridium perfringens. The EA, hexane and DCM extracts were 2 to 3 times more active than the acetone extract (average total activity 1728 ml/g dry extract for the 3 pathogens). The DCM-, EA-, acetone- and hexane extracts (in order of activity) from the direct extraction procedure from H. perforatum were only active against C. perfringens with the first three extracts having a total activity of between 1026 and 1333 ml/g dry material and the hexane extract a total activity of 333 ml/g dry material. The spectrum of activity of G. biloba corresponds to that of Zn-bacitracin, which is commonly used an antibiotic growth promoter in the poultry industry. The second objective in this study was to combine extracts or fractions of extracts of G. biloba and H. perforatum to optimise activity against selected bacterial pathogens. A synergistic effect could be observed when combining a ratio of 1:5 of G. biloba: H. perforatum (hexane extracts) or 1:15 (acetone extracts) against E. faecalis while only an indifferent (neutral) effect was observed against C. perfringens. After elucidation of the quantitative and qualitative aspects involved in the antimicrobial activity, the major antibacterial compound from G. biloba was isolated and characterized as ginkgolic acid (C17:1). It was also determined whether activity against E. faecalis and C. perfringens in an extract or fraction of and extract of G. biloba can be attributed only to ginkgolic acid or whether synergism or other interactions also play a role in the antibacterial activity. It was shown that synergistic interactions are at play between constituents in the hexane and EA fraction, with the last mentioned fraction not containing any ginkgolic acid. These results support the use of the whole extract as opposed to isolated compounds as antimicrobial agents against pathogenic organisms. Two important pharmacodynamic parameters were investigated i.e. resistance development to a hexane extract and the isolated ginkgolic acid from G. biloba against E. faecalis and secondly the time-kill dynamics of this hexane extract over 24 h against E. faecalis. The bactericidal nature of the hexane extract from G. biloba as well the absence of decreased susceptibility to this extract (and the isolated ginkgolic acid) in the resistance studies against E. faecalis indicate that this extract has potential to be exploited as a alternative to AGPs in the poultry industry. The final objective was to determine the effect of extracts of G. biloba alone or in combination with H. perforatum extracts on the performance of broiler chickens over a 35 day period. The effect of these extracts on C. perfringens in the intestine of broilers was also investigated. No significant differences were found with relation to any of the production parameters studied (FCR, live weight or % survival) although a trend towards more favourable European Performance Efficiency Factor index values were observed for treatments containing G. biloba (5% improvement) or a combination of G. biloba and H. perforatum (2.1% improvement) compared to the untreated control. Similarly, Zn-Bacitracin resulted in a 5.5% improvement compared to the untreated control. There was a general trend (not statistically significant, P=0.05) towards a reduction in C. perfringens scores in the feed supplemented with G. biloba- in combination with H. perforatum extract which can probably be ascribed to the direct antimicrobial effect. The rate of colonization was however too low to cause infection probably due to lack of virulence of the C. perfringens challenge and the absence of predisposing factors due to the hygienic growth conditions used. It is necessary for an effective disease model to be developed in order for the efficacy of any new treatment method to be properly evaluated. Such a model will require a much higher incidence of disease and reproducibility than was achieved in this project. The safety of using extracts of G. biloba with ginkgolic acid as the prime antibacterial compound was considered. The active dose was at least 42 times lower than safe dosage recommended in the literature. The combination of extracts of G. biloba and H. perforatum holds promise as a potential growth promoter in poultry production. Better results may be achieved if potentized extracts are used and compared with Zn-Bacitracin and a negative control under industrial growth conditions where the birds are stressed and natural infections would take place. Copyright