Holarrhena antidysenterica Extract and Its Steroidal Alkaloid, Conessine, as Resistance-Modifying Agents Against Extensively Drug-Resistant Acinetobacter baumannii

Abstract

Emergence and spread of antibiotic-resistant Acinetobacter baumannii have become a major public health concern. This study was designed to investigate the efficacy of Holarrhena antidysenterica extract and its major steroidal alkaloid conessine as resistance-modifying agents (RMAs) on the susceptibility of A. baumannii to novobiocin and rifampicin. A significant synergistic activity of both the extract and conessine in combination with either novobiocin or rifampicin with fractional inhibitory concentration index ≤0.5 was demonstrated. Fluorescent dyes and different efflux pump inhibitors were used to further investigate the synergism. Increase in the uptake of 1-N-phenylnaphthylamine in the bacterial cells treated with the extract and conessine was not observed indicating that both substances did not act as permeabilizers. With regard to efflux pump inhibition, no accumulation in ethidium bromide (EtBr) was noticed suggesting that the AdeABC pump was not involved. In contrast, accumulation in Pyronin Y was significantly increased (p < 0.05) demonstrating that the synergism was due to interference with the AdeIJK pump. Study on frequencies of the spontaneous mutational resistance to the extract in combination with antibiotics demonstrated attenuation in drug-resistant organisms. Thus, H. antidysenterica extract and conessine as RMAs may offer a combinatory therapy to restore antibiotic susceptibility in the extensively drug-resistant A. baumannii.

Steroidal alkaloids and conessine from the medicinal plant Holarrhena antidysenterica restore antibiotic efficacy in a Galleria mellonella model of multidrug-resistant Pseudomonas aeruginosa infection

“…The susceptibility of the P. aeruginosa strains to a group of anti-pseudomonal antibiotics alone or in the presence of steroidal alkaloids (128 or 1024 mg/L) or conessine (32 mg/L) is shown in Table 1 . In this study, steroidal alkaloids extract at 1024 mg/L contained approximately 37 mg/L conessine content [ 19 ]. The strain overexpressing the MexAB-OprM efflux pump (PAM1032) was more resistant to ceftazidime, piperacillin, meropenem and levofloxacin in comparison to the parent strain (PAM1020).…”

“…Steroidal alkaloids present within the extracts possess antibacterial, anti-diarrhoeal and astringent properties [ 15 , 16 ]. Previous studies indicated that H. antidysenterica extract, and the principal active ingredient alone (the steroidal alkaloid conessine [ 16 ]), were able to restore the inhibitory activity of novobiocin and rifampicin against extensively drug-resistant Acinetobacter baumannii in vitro [ 17 – 19 ]. Exposure to either H. antidysenterica extract, or conessine alone, at concentrations that did not result in increased membrane disruption, resulted in accumulation of the fluorescent dye Pyronin Y suggesting that both treatments may have EPI-like properties [ 19 ].…”

“…Previous studies indicated that H. antidysenterica extract, and the principal active ingredient alone (the steroidal alkaloid conessine [ 16 ]), were able to restore the inhibitory activity of novobiocin and rifampicin against extensively drug-resistant Acinetobacter baumannii in vitro [ 17 – 19 ]. Exposure to either H. antidysenterica extract, or conessine alone, at concentrations that did not result in increased membrane disruption, resulted in accumulation of the fluorescent dye Pyronin Y suggesting that both treatments may have EPI-like properties [ 19 ]. Moreover, exposure to conessine restored the MICs of antibiotics against a P. aeruginosa strain over-expressing MexAB-OprM to values comparable with the wild-type parent and conessine induced the accumulation of Hoechst 33342 in cells over-expressing the same efflux pump [ 20 ].…”

Mechanisms Efflux Pumps of &lt;i&gt;Acinetobacter baumannii&lt;/i&gt; (MDR): Increasing Resistance to Antibiotics

“…In other studies, PAβN at 10 μg/ml decreased predominantly MIC concentrations of trimethoprim, clindamycin and chloramphenicol [97] [102]. Twice a time on clinical isolates, whereas PAβN at 20 μg/ml reduced nicidixic acid MIC to 16-fold but showed little effect on sensitiveness to ciprofloxacin [106]. At 100 μg/ml PAβN are also sensible minocycline activity by decreasing ≥ 04-fold MIC values [107].…”

Conessine as a novel inhibitor of multidrug efflux pump systems in Pseudomonas aeruginosa

“…It has been demonstrated that the extract potentiated the effect of antibiotics by acting as a permeabilizer [ 13 ]. Moreover, a recent study indicated that both Holarrhena antidysenterica extract and conessine, a steroidal alkaloid compound, could restore antibiotic activity due to interference with AdeIJK pump in A. baumannii [ 14 ]. Previous study documented that AdeIJK pump and MexAB-OprM pump are functionally equivalent pumps in both organisms [ 15 ].…”