Antibacterial and antibiotic-modulation activity of six Cameroonian medicinal plants against Gram-negative multi-drug resistant phenotypes

Infectious diseases still represent one of the major health concern worldwide [1]. According to the National Institute of Health, infectious diseases are the second cause of death and the leading cause of loss of productive life years worldwide. Bacterial infections are responsible of about 70 % of cases of death related to microorganisms [1]. The use of antibiotics and hygiene rules helped to fight infectious diseases in the past. However, they are becoming increasingly difficult to control as results of the spread of resistant phenotypes. The resistance to antibiotics has increased in recent decades, mainly due of their inappropriate use [2]. Bacteria have developed several mechanisms of resistance including active efflux which plays an important role in multi-drug resistance (MDR), mainly in Gram-negative bacteria [3]. There is a need for the discovery of new active antimicrobials to combat MDR microorganisms. Amongst the new areas explored to overcome infectious diseases caused by MDR bacteria, medicinal plants seem to offer an ideal alternative since they are readily available source of bioactive agents and are well accepted by about 80 % of the world population. Many African medicinal plants and their metabolites were previously found active against MDR Gram-negative bacteria [4, 5]. Also the synergistic activities of some African medicinal plants with antibiotics against MDR Gram-negative bacteria were reported [5, 6]. It was demonstrated that several naturally occurring efflux pump inhibitors can restore the activity of antibiotics against MDR bacteria [7, 8]. The present study was therefore designed to investigate the antibacterial potential against MDR Gram-negative phenotypes expressing active efflux pumps of six Cameroonian medicinal plants used traditionally in the treatment of bacterial infections, namely Anthocleista schweinfurthii Gilg.(Loganiaceae), Boehmeria platyphylla D. Don (Urticaceae), Caucalis melanantha (Hochst/Hien) (Urticaceae), Erigeron floribundus (H.BK) (Asteraceae), Nauclea latifolia Smith (Rubiaceae) and Zehneria scobra (cf) Sondev (Cucurbitaceae).

Medicinal plants are potential source of antimicrobial agents used in the treatment of infectious diseases [16]. According to Rios and Recio [17], and Kuete et al. [17], the antibacterial activity of a plant extract is considered significant when the MICs are below 100 μg/mL. The activity is considered moderate when 100 ≤ MIC ≤ 625 μg/mL and weak when MIC are above 625 μg/mL [17]. Therefore, the antibacterial activities reported in the present study can mostly be regarded as moderate or low. This could be explained by the fact that the tested bacteria are mostly MDR phenotypes. In fact, P. aeruginosa and MDR Enterobacteriaceae ( K. pneumoniae, E. aerogenes, E.cloacae and P. stuartii and E. coli) tested in the present study have been classified as antimicrobial-resistant organisms of concern in healthcare facilities [18‐20]. The previously reported activities of A. schweinfurthii include antibacterial inhibitory effects of n-hexane, dichloromethane, ethyl acetate and methanol extracts from leaves and stem bark against Staphylococcus aureus ATCC 33591 and E. coli ATCC 27195 [21]. The MIC values obtained in the present study were respectively 62.5 and 125 μg/ml against S. aureus and E. coli. Such values were higher than those previously documented, highlighting the MDR feature of the studied bacteria. MBC values were obtained in few cases (Additional file 1: Table S2). A keen look of data (Additional file 1: Table S2) indicates that, in most of the cases, the tested extract exerted bacteriostatic effects with a ratio MBC/MIC above 4. The overall antibacterial activity of the tested extracts could be due their phytochemical composition. However, the presence of a specific class of second metabolite could not guarantee the antibacterial activity of the plant, as this will depend on nature of the compounds, its concentration as well as the possible interactions with other constituents of the extract. It is also surprising that saponins, known to possess antibacterial activities were not detected in the tested extracts; However, this does means that the extract were completely devoid of this class of secondary metabolite; One of the most understandable explanation should that saponins could be present in very little amounts in the tested extract, and therefore could not be detected using the qualitative phytochemical methods. Some cardiac glycosides such as bufalin, oubain, digoxin are toxic meanwhile many of them have therapeutic uses and these primarily involve the treatment of cardiac failure [22‐24]. Their utility results from an increased cardiac outpout by increasing the force of contraction. By increasing intracellular calcium, cardiac glycosides increase calcium-induced calcium release and thus contraction [23, 24]. The traditional use of the studied plants could suggest that their cardiac glycoside could be not toxic and have very low toxic effects.

To the best of our knowledge, the present work describes for the first time the antibacterial activity of B. platyphylla. This activity could be due to the presence of the detected phytochemicals. In fact, antibacterial compounds such as acetophenone [25] and cryptopleurine [26‐28] were previously isolated from B. Platyphylla. The antibacterial activity of C. melanantha and E. floribundus is also reported here for the first time. However these plants were previously reported for their antifungal activities [29‐32]. The antibacterial activities of extracts from Zehneria scobra and Nauclea latifolia [33, 34] were reported on some bacteria: The present study therefore provides additional information on the activity of these plants against MDR Gram-negative phenotypes.

The synergistic effects between antibiotics and the tested plants are also reported here for the first time. The observed synergistic effects could be due to possible interaction between plant constituents and the tested antibiotics. As the strains used in this study are known to actively expressed efflux pumps, one of the possible explanations for the observed synergistic effects could be the ability of the constituents of the extracts to act as efflux pumps inhibitor. This can explain why the effect of antibiotics with intracellular targets such as STR, CHL and KAN increased contrary to that of beta-lactamine (AMP) acting in the cell wall (Tables 3, 4, 5 and 6).

The overall results of the present study provides baseline information for the possible use of the tested plants, especially A. schweinfurthii, N. Latifolia, B. platyphylla and E. floribundus in the control of infections due to Gram-negative bacteria. The present study indicates that the tested plant extracts alone could not be used efficiently to tackle MDR bacterial infections. However, it was demonstrated that extracts from A. schweinfurthii fruits and Z. scobra could be used in combination with some antibiotics to fight bacterial multi-drug resistance.