Introduction

Because of their potent antimicrobial activity, wine and numerous wine-based mixtures with other plant-derived antimicrobials have been investigated as a part of non-antibiotic strategies for tackling foodborne and medical pathogens (Friedman, 2014). Although the exact mechanisms and relative contribution of wine components to the antimicrobial activity of wine are still a matter of debate, it is possible that they are different to those of standard medicinal antibiotics (Pojer et al., 2013; Cushnie and Lamb, 2005; Cisowska et al., 2011). Hence, wine might be less susceptible to the development of bacterial resistance.

In contrast to numerous studies demonstrating the effectiveness of using different phytochemicals and plant extracts against resistant bacteria (Friedman, 2015), the antimicrobial effects of wine in relation to bacterial resistance to antibiotics is largely unexplored.

Therefore, the present study was undertaken to examine whether resistance to medicinal antibiotics of standard strains and clinical isolates of E. coli and S. enteritidis is related to their resistance to wine as an unspecific antimicrobial medium.

Materials and methods

Results and discussion

The minimal incubation time of the bacterial suspension with wine, necessary for prevention of bacterial growth, was 3 and 20 min for E. coli ATCC and ESBL E. coli respectively (Figure 1A). This notable difference in the time-kill curves between the two E. coli strains can be clearly linked to a clinically relevant difference in their susceptibility to a spectrum of antimicrobial drugs. Table 1 shows minimal inhibitory concentrations (MIC) in μg/mL for antimicrobial drugs to which ESBL E. coli was intermediately or fully resistant relative to ATCC E. coli, which proved sensitive to all test antibiotics.

In the case of S. enteritidis strains, they were both susceptible with practically the same MIC values for all tested antibiotics (clinical isolate S. enteritidis KK 962 showed slightly higher MIC values for piperacillin, tigecycline and chloramphenicol; data not shown). This was reflected in the similar time-kill curves (4 and 5 min for S. enteritidis ATCC 13076 and S. enteritidis KK 962 respectively (Figure 1B)).

Figure 1. Time-kill curves for Escherichia coli ATCC 25922 and multi-resistant, extended-spectrum beta-lactamase - producing clinical isolate Escherichia coli UR 3612 (A) and Salmonella enteritidis ATCC 13076 and clinical isolate Salmonella enteritidis KK 926 (B) following incubation with red wine.

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Data are averages of at least three independent experiments.

The fact that wine affected the same bacterial species of differing origin in different ways does not support the notion that wine may not kill bacteria, but just cause their unspecific aggregation due to the polyphenolic compounds contained in wine (Cushnie and Lamb, 2005). On the other hand, the inhibition of growth in the nutrient agar of the bacteria which were previously inoculated in wine, has been described to be just a transient phenomenon; namely, after exposure to a harsh antimicrobial medium such as wine, bacteria may turn into a viable, but non-cultivable, state, thereby preserving potential to restore colony-forming activity when incubated in a favourable growing environment (Millet and Lonvaud-Funel, 2000). Whatever the mechanism of the loss of colony-forming activity observed in this study, it does not alter the key finding that bacterial sensitivity to wine is associated with the susceptibility of bacteria to medicinal antibiotics.

Moreover, we believe that the method in which test strains are exposed to whole wine, in which physical-chemical properties and the matrix are not disturbed, provide the most realistic insight into the “true” effects of wine. ESBL producing E. coli as the most resistant bacteria to wine was also resistant to a wide range of antimicrobial drugs which belong to different classes depending on their principal mechanism of action. Among known basic mechanisms of bacterial resistance, multidrug efflux systems capable of pumping a broad spectrum of unrelated antibiotics and biocides (Poole, 2005) are likely to be involved.

However, the antimicrobial activity of wine and its constituents involves many modes of action (Cisowska et al., 2011; Cho et al., 2014; Cushnie and Lamb, 2005; Daglia et al., 2007). The relationship between bacterial multidrug resistance and various mechanisms of antibacterial action of wine and its constituents is practically unexplored.

In an attempt to distinguish the roles of polyphenols, pH, ethanol and other wine components, in our previous work we examined the antimicrobial effects of intact wine in comparison to those of phenols-stripped wine, dealcoholized wine, ethanol and low pH applied separately and in combination (Boban et al., 2010). It was not possible to relate the antibacterial activity of the samples to their total phenolic and resveratrol content, ethanol content, or pH. It was concluded that the antimicrobial activity of complex solutions, such as an intact wine, cannot be exclusively attributed to its phenolic or nonphenolic constituents, nor can the antimicrobial activity of wine be predicted on the basis of its particular components.

We hope that this simple yet straightforward study may be inspiring for further studies, thus deepening insight into this complex matter.

Conclusion

The key finding of this study is that bacterial resistance to wine appears to be closely associated with bacterial resistance to antimicrobial drugs. This indicates that bacterial resistance to wine and medicinal antibiotics share, at least in part, the same mechanisms.

Acknowledgements

This study was supported by the Croatian Science Foundation (Project no. 8652)

References

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