Combinatorial efficacy of Manuka honey and antibiotics in the in vitro control of staphylococci and their small colony variants

Abstract

IntroductionStaphylococci are among the list of problematic bacteria contributing to the global antibiotic resistance (ABR) crisis. Their ability to adopt the small colony variant (SCV) phenotype, induced by prolonged antibiotic chemotherapy, complicates staphylococcal infection control options. Novel and alternative approaches are needed to tackle staphylococcal infections and curb ABR. Manuka honey (MH), a non-antibiotic alternative is recognized for its unique antibacterial activity based on its methylglyoxal (MGO) component.MethodsIn this study, MH (MGO 830+) was tested in combination with gentamicin (GEN), rifampicin (RIF), or vancomycin (VA) against staphylococcal wildtype (WT) and SCVs. To our knowledge, there are no current studies in the literature documenting the effects of MH on staphylococcal SCVs. While Staphylococcus aureus is well-studied for its international ABR burden, limited data exists demonstrating the effects of MH on S. epidermidis and S. lugdunensis whose pathogenic relevance and contribution to ABR is also rising.Results & discussionThe three staphylococci were most susceptible to RIF (0.06-0.24 μg/ml), then GEN (0.12-0.49 μg/ml), and lastly VA (0.49-0.96 μg/ml). The MICs of MH were 7%, 7-8%, and 6-7% (w/v), respectively. Fractional inhibitory concentration (FIC) evaluations showed that the combined MH + antibiotic effect was either additive (FICI 1-2), or partially synergistic (FICI >0.5-1). While all three antibiotics induced SCVs in vitro, stable SCVs were observed in GEN treatments only. The addition of MH to these GEN-SCV-induction analyses resulted in complete suppression of SCVs (p<0.001) in all three staphylococci, suggesting that MH’s antibacterial properties interfered with GEN’s SCV induction mechanisms. Moreover, the addition of MH to growth cultures of recovered stable SCVs resulted in the inhibition of SCV growth by at least 99%, indicating MH’s ability to prevent subsequent SCV growth. These in vitro analyses demonstrated MH’s broad-spectrum capabilities not only in improving WT staphylococci susceptibility to the three antibiotics, but also mitigated the development and subsequent growth of their SCV phenotypes. MH in combination with antibiotics has the potential to not only resensitize staphylococci to antibiotics and consequently require less antibiotic usage, but in instances where prolonged chemotherapy is employed, the development and growth of SCVs would be hampered, providing a better clinical outcome, all of which mitigate ABR.

Manuka honey as a non-antibiotic alternative against Staphylococcus spp. and their small colony variant (SCVs) phenotypes

“…CNS isolates featured in 7 of the articles ( S. capitis , S. epidermidis (MRSE), S. lugdunensis , S. haemolyticus , S. simulans , S. saprophyticus , and S. warneri ( Supplementary Table S1 )). Among the 48 studies, only one covered the application of MH on Staphylococcal SCVs ( Liang et al., 2023 ). Other than MH (unprocessed or medical grade), the research articles concurrently tested 12 other types of honey from various sources and geographical origins for their efficacy against Staphylococci.…”

“…It was not indicated, however, whether the SCVs were detectable prior to MH treatment (since SCVs are a naturally occurring sub-population), or if they were induced following MH treatment. A recent study demonstrated both the growth inhibition and clearance of stable gentamicin-induced staphylococcal SCVs in the presence of MH, suggesting MH’s current efficacy in staphylococcal-mediated infections ( Liang et al., 2023 ). Knowing the relevance of SCV phenotypes in exacerbating the ABR cycle, additional investigations evaluating the influence of MH directly on SCV phenotypes are needed, including the possibility of SCV induction from this product.…”

“…However, strain-dependent effects were evident, with minimal to no synergism observed in the growth of other MRSA or non-MRSA isolates (both planktonic and biofilm; FICI 0.87 - 2) ( Liu et al., 2014 ), and even instances of antagonism. Against CNS, the combination of MH with gentamicin demonstrated partial synergy (FICI 0.93) against S. lugdunensis , while an additive effect (FICI 1.5) was observed against S. epidermidis isolates ( Liang et al., 2023 ).…”

“…Interestingly, repeated exposure of both planktonic and biofilm S. epidermidis to MH increased the susceptibility to subsequent vancomycin challenges, suggesting that the physiological modification by MH was not transient. In addition, the simultaneous application of MH and vancomycin resulted in additive effects against planktonic S. epidermidis and S. lugdunensis isolates ( Liang et al, 2023 ).…”

“…Despite variations in outcomes of other MH-antibiotic combinations, the MH-rifampicin combination exhibited relatively consistent synergism against the growth of different S. aureus strains, both in the state of planktonic cell and biofilm, irrespective of their resistance nature. However, this efficacy did not extend to the case of CNS, with only additive effects shown in the growth of S. epidermidis and S. lugdunensis isolates (FICI 1.14 and 1.50) ( Liang et al, 2023 ). Nonetheless, it is crucial to acknowledge that the current data is insufficient to draw statistical conclusions, highlighting the need for extensive investigations before clinical applications.…”