Synergistic activity between Triphala and selected antibiotics against drug resistant clinical isolates

Triphala is an indigenous medicinal product used in diseases in nose and skin wounds, tumours, cough and upper respiratory diseases, peptic ulcer, mouth ulcers, anemia, abdominal pain, obesity, liver diseases and toxic conditions [1‐3]. It is composed of dried pericarp of the fruits of three plants, Terminalia chebula Retz. (Combretaceae), Terminalia bellirica (Gaertn.) Roxb. (Combretaceae) and Emblica officinalis L. (Euphorbiaceae) mixed in equal proportions (1:1:1) [1‐4].

The biological activity of Triphala has been studied in some detail, including in animal experiments. Triphala is known to have antimicrobial activity, anti-viral activity, anti-oxidant activity including the ability to scavenge free radicals, anti-inflammatory activity, anti-pyretic activity, wound healing properties, anti-mutagenic and anti-carcinogenic activity as well as hypoglycaemic effects [4‐6]. Extracts of Triphala and its individual plants have demonstrated varying degree of activity against a range of Gram negative and Gram positive bacteria [7‐10].

Furthermore, Triphala has also shown antibacterial activity against Enterococcus faecalis in biofilms formed on dental substrates [11]. A mouthwash prepared from Triphala has been effective in reducing dental plaque formation [12]. Triphala has been incorporated in collagen sponges to reduce wound infection and facilitate healing [13].

Antimicrobial resistance (AMR) is a major issue faced by health care professionals globally. In addition to the identification of novel antibiotics, enhancing the activity of existing antibiotics by combining with novel compounds can also be considered as a strategy to overcome the problem of AMR.

Gentamicin is an aminoglycoside with activity against both Gram negative and positive organisms. Gentamicin is used routinely in combination with other classes of antibiotics in the treatment of different types of bacterial infections. Oxacillin is an indicator drug used to assess sensitivity of Staphylococcus aureus to β-lactam antibiotics, the main group of antibiotics used to treat staphylococcal infections unless the isolate is a methicillin-resistant Staphylococcus aureus (MRSA). Although synergistic activity of component plants of Triphala with some antibiotics have been studied [14], the synergy between Triphala and gentamicin and oxacillin has not been studied previously to the best of our knowledge. The current study was therefore conducted with the objective of characterizing the antimicrobial activity of Triphala against multi-drug resistant (MDR) organisms and to evaluate synergistic activity with gentamicin and oxacillin against appropriate organisms.

All isolates tested were confirmed to be MDR organisms being resistant to three or more classes of antibiotics (Table 1).

In the current study, we demonstrated that Triphala interacts in a synergistic manner with gentamicin against the selected MDR Gram negative bacilli and with oxacillin against MRSA. Aminoglycosides including gentamicin act by interfering with bacterial protein synthesis. Gentamicin has been used widely in combination with other β-lactam antibiotics [19]. Non-lethal damage by theβ-lactam antibiotics to the cell wall of bacteria is considered to facilitate the entry of the aminoglycoside antibiotic to the bacterial cell, enhancing its killing ability [20]. Aminoglycoside resistance, including resistance to gentamicin occurs through enzymatic deactivation of the drug, active efflux of the drug or cell wall changes leading to reduced uptake of the drug and decreased binding of the drug to the 16S ribosome due to mutations or methylation [19]. Both ethanolic and aqueous extracts of Triphala contain phenolics, flavonoids and carotenoids [21]. Antibacterial activity of Triphala has been shown to correlate to its phenolic content [22]. Phenolic compounds are known to disrupt the cell membranes of bacteria [23]. Therefore, the mechanism of synergy between gentamicin and Triphala may be similar to that between gentamicin andβ-lactams.

Oxacillin acts by interfering with cell wall synthesis. The main mechanism of resistance to all β-lactams in MRSA is the presence of PBP2a that has a lower affinity to this group of drugs [24]. Synergy between phenolic compounds and oxacillin against MRSA, similar to our findings, has been documented previously [25, 26].

The chemical structures of active phenolic compounds determine their antibacterial activity [23]. Therefore, it is important that the active phytochemicals of Triphala be identified and their structures elucidated in furthering this study.

While synergism or additive activity was noted across all antimicrobial-plant compounds tested in the study, not all gentamicin MIC drops resulted in achieving clinical sensitive breakpoints. Furthermore, no consistency has been observed in the reversion of gentamicin MIC to sensitive levels across the organisms tested. Presence of varying mechanisms of resistance among the test isolates may have contributed to this. As the mechanisms of resistance for the test isolates were not established, we are unable to further validate this claim.

While there was a drop in the MIC of Triphala against all organisms tested when used in combination with the antibiotics, the values remained relatively high (original MIC values not shown). However, the reported MIC values for Triphala and its constitutive plant products when tested alone has ranges similar to that obtained in our study [22, 23]. Similar levels have been reported for other plant products as well [27, 28].

Aminoglycoside use had increased in the recent past for the treatment of infections caused by MDR organisms as the rate of emergence of resistance to aminoglycosides has been low. However, the reason for discontinuation of the aminoglycoside in many instances is nephrotoxicity [29]. Interestingly, Triphala has renal protective effects both in animal studies [30] and limited clinical studies [31], presumably due to its antioxidant properties. In vitro and in-vivo studies of Triphala and gentamicin together to explore the possible benefits are warranted in future.

In conclusion, Triphala demonstrated synergistic activity with gentamicin against selected MDR Gram negative bacilli and with oxacillin against MRSA, warranting further studies to identify the possibility of using such combinations in clinical practice.