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In vitro antibacterial and antibiotic-potentiation activities of the methanol extracts from Beilschmiedia acuta, Clausena anisata, Newbouldia laevis and Polyscias fulva against multidrug-resistant Gram-negative bacteria
BMC Complementary and Alternative Medicinevolume 15, Article number: 412 (2015)
The present study was designed to investigate the antibacterial activities of the methanol extracts from different parts of Beilschmedia acuta Kosterm (Lauraceae), Clausena anisata (Willd) Hook (Rutaceae), Newbouldia laevis Seem (Bignoniaceae) and Polyscias fulva (Hiern) Harms (Araliaceae) as well as their synergistic effects with antibiotics against a panel of Gram-negative bacteria, including multi-drug resistant (MDR) phenotypes expressing active efflux pumps.
Broth microdilution method was used to determine the minimum inhibitory concentrations (MICs) and the minimum bactericidal concentrations (MBCs) of the extracts, as well as those of antibiotics in association with the most active ones, B. acuta, N. laevis and P. fulva.
MIC values obtained indicate that extracts from the bark of B. acuta were active on all the 26 tested Gram-negative bacteria, with MICs ranging from values below 8 to 256 μg/mL. Other samples displayed selective activities, their inhibitory effects being observed on 9 (34.62 %) of the 26 bacterial strains for N. laevis leaves extract, 6 (23.10 %) for both C. anisata leaves and roots extracts, 7 (26.9 %) and 4 (15.4 %) for leaves and roots extracts of P. fulva respectively. Extract from B. actua bark displayed the best antibacterial activity with MIC values below 100 μg/mL against 16 (61.5 %) of the 26 tested microorganisms. The lowest MIC values (below 8 μg/mL) were obtained with this extract against Escherichia coli W3110 and Klebsiella pneumoniae ATCC11296. The MIC values of this extract were lower than those of ciprofloxacin against E. coli W3110, Enterobacter aerogenes ATCC13048, CM64 and Providencia stuartii NAE16. At MIC/2, the best percentages of synergistic effects (100 %), were obtained with B. acuta bark extract and tetracycline (TET) as well as with P. fulva leaves extract and TET and kanamycin (KAN).
The overall results of the present study provide information for the possible use of the studied plants and mostly Beilschmedia acuta in the control of bacterial infections including MDR phenotypes.
Fighting multi-drug resistant (MDR) Gram-negative (MDRGN) bacteria remains a challenging issue worldwide. Microbial infections involving MDRGN bacteria constitute a major public health problem in developing countries  where the high cost of antibiotics makes them unaffordable to the majority of the population. Clinically, the continuous emergence of MDRGN bacteria drastically reduced the efficacy of antibiotic arsenal and, consequently, increased the frequency of therapeutic failure . Therefore, the discovery of new antimicrobial agents is still relevant nowadays. Also, the shortcomings of drugs available today and scarcity of novel antibiotics propel the discovery of new chemotherapeutic agents from medicinal plants . Approximately 60 % of the world population still relies on medicinal plants for their primary healthcare . Medicinal plants have been used as a source of remedies since ancient times in Africa. In addition, promising new concepts such as the efflux pump inhibitors [5, 6], and synergy between antibiotics and phytochemicals are now being developed. The ability of several African medicinal plants to inhibit the growth of MDRGN bacteria, as well as their ability to potentiate the activity of commonly used antibiotics was previously reported. Some of these plants include Dorstenia psilurus, Dichrostachys glomerata and Beilschmiedia cinnamomea [7–9].
In our continuous search of plant extracts with antibiotic-potentiating activity to combat MDR bacteria, the present work was designed to investigate the antibacterial activity of four Cameroonian medicinal plants used traditionally in the treatment of bacterial infections, namely Beilschmiedia acuta Kosterm (Lauraceae), Clausena anisata (Willd) Hook (Rutaceae), Newbouldia laevis Seem (Bignoniaceae) and Polyscias fulva (Hiern) Harms (Araliaceae), against MDRGN expressing active efflux via the Resistance-Nodulation Cell Division (RND)-type pumps. In the treatment of infectious diseases, Beilschmiedia acuta is traditionally used for gastrointestinal infections , Clausena anisata for fungal, bacterial and viral infections, Newbouldia laevis for bacterial and fungal infections [11–14], dysentery, worms, malaria, dental caries and diarrhea  and Polyscias fulva for venereal infections [16, 17].
Plant material and extraction
All medicinal plants used in the present work were collected in different areas of Cameroon between January and April 2012. The plants were identified at the National Herbarium (Yaounde, Cameroon), where voucher specimens were deposited under the reference numbers (Table 1). Air-dried and powdered plant material was weighed (300 g) and soaked in 1 L of methanol (MeOH) for 48 h at room temperature. The filtrate obtained through Whatman filter paper No.1 was concentrated under reduced pressure in a vacuum to obtain the crude extract. All crude extracts were kept at 4 °C until further use.
Chemicals for antimicrobial assays
Tetracycline (TET), ciprofloxacine (CIP), chloramphenicol (CHL), ampicillin (AMP) and kanamycin (KAN) (Sigma-Aldrich, St Quentin Fallavier, France) were used as reference antibiotics (RA). p-Iodonitrotetrazolium chloride (INT, Sigma-Aldrich) was used as a microbial growth indicator [18, 19].
Microbial strains and culture media
The studied microorganisms included sensitive and resistant strains of Pseudomonas aeruginosa, Klebsiella pneumoniae, Enterobacter aerogenes, Escherichia coli obtained from the American Type Culture Collection (ATCC). Their bacterial features are summarized in Table 2. Nutrient agar was used to activate the tested Gram-negative bacteria .
INT colorimetric assay for MIC and MBC determinations
The MIC determination on the tested bacteria was conducted using rapid p-iodonitrotetrazolium chloride (INT) colorimetric assay according to described methods  with some modifications [21, 22]. The test samples and RA were first dissolved in DMSO/Mueller Hinton Broth (MHB). The final concentration of DMSO was lower than 2.5 % and does not affect the microbial growth [23, 24]. The solution obtained was then added to Mueller Hinton Broth, and serially diluted two fold (in a 96- wells microplate). One hundred microlitre (100 μL) of inoculum 1.5 x 106 CFU/mL prepared in appropriate broth was then added [21, 22]. The plates were covered with a sterile plate sealer, then agitated to mix the contents of the wells using a plate shaker and incubated at 37 °C for 18 h. The assay was repeated thrice. Wells containing adequate broth, 100 μL of inoculum and DMSO to a final concentration of 2.5 % served as negative control. The MIC of samples was detected after 18 h incubation at 37 °C, following addition (40 μL) of 0.2 mg/mL of INT and incubation at 37 °C for 30 min. Viable bacteria reduced the yellow dye to pink. The MIC was defined as the sample concentration that prevented the color change of the medium and exhibited complete inhibition of microbial growth . The MBC was determined by adding 50 μL aliquots of the preparations, which did not show any growth after incubation during MIC assays, to 150 μL of adequate broth. These preparations were incubated at 37 °C for 48 h. The MBC was regarded as the lowest concentration of extracts, which did not produce a color change after addition of INT as mentioned above [21, 22].
Samples were tested alone and the best four extracts (those from the leaves and bark of Beilschmedia acuta, and from the leaves of Newbouldia laevis and Polyscias fulva) were also selected and tested in association with antibiotics at the sub-inhibitory concentrations (MIC/2 and MIC/5) [7–9] against nine MDR bacteria. Fractional inhibitory concentration (FIC) was calculated as the ratio of MICAntibiotic in combination/MICAntibiotic alone and the results were discussed as follows: synergy (≤0.5), indifferent (0.5 to 4), or antagonism (>4) [25, 26]. All assays were performed in triplicate.
The antibacterial activities of methanol extracts from various parts of Beilschmedia acuta, Clausena anisata, Newbouldia laevis and Polyscias fulva are summarized in Table 3 (MIC values up to 1024 μg/mL are provided as supporting information; Additional file 1: Table S1). It can be observed that extracts from the bark of B. acuta were active on all 26 tested Gram-negative bacteria, with MICs ranging from values below 8 to 256 μg/mL. Other samples displayed selective activities, their inhibitory effects being observed against nine (34.62 %) of the 26 bacterial strains for N. laevis leaves extract, six (23.10 %) for both C. anisata leaves and roots extracts, seven (26.9 %) and four (15.4 %) for leaves and roots extracts of P. fulva respectively. Extract from the bark of B. actua showed the best antibacterial activity with MIC values below 100 μg/mL against 16/26 (61.5 %) of the tested microorganisms. The lowest MIC values below 8 μg/mL were obtained with this extract against Escherichia coli W3110 and Klebsiella pneumoniae ATCC11296. MIC values of this extract were lower than those of ciprofloxacin against E. coli W3110, Enterobacter aerogenes ATCC13048 and CM64 and Providencia stuartii NAE16 (Table 3). The bactericidal activities of studied samples were mostly noted with the extract from B. acuta, with MBC values observed against 23/26 (88.5 %) tested bacteria (see Additional file 1: Table S2, supporting information).
Five commonly used antibiotics (CIP, TET, KAN, AMP and CHL) were combined with extracts from B. acuta leaves and bark and those from the leaves of N. laevis and P. fulva at their MIC/2 and MIC/5, as obtained on each of nine tested bacterial strains (Tables 4 and 5). Synergistic effects were observed with all tested extracts and all studied antibiotics on at least one of the nine selected bacteria. The best percentages of synergistic effect (100 %) were obtained at MIC/2 with B. acuta bark extract in combination with TET (Table 5) as well as with P. fulva leaves extract in association with TET and KAN (Table 5).
Phytochemicals are routinely classified as antimicrobials on the basis of susceptibility tests that produce MICs in the range of 100 to 1000 μg/mL . Moreover, for crude extracts, the antimicrobial activity is considered to be significant if MIC values are below 100 μg/mL and moderate when 100< MIC <625 μg/mL [28, 29]. Therefore, the activity recorded with B. acuta bark extract against the 26 tested bacterial strains can be considered as very important. If we consider the alternative criteria described by Fabry et al. , where extracts having MIC values less than 8000 μg/mL have noteworthy antimicrobial activity, the overall activity recorded with the leaves and fruit extracts of B. acuta, P. fulva and N. laevis leaves extracts can also be considered promising. A keen look of the results of MIC and MBC determinations (Table 3, Additional file 1: Tables S1 and S2) indicates that MBC/MIC ratios were mostly above four, suggesting that studied extracts, including the most active ones, generally displayed bacteriostatic effects (MBC/MIC > 4) [31–33]. Various classes of phytochemicals (Table 2) were previously detected in the extracts of the four tested plants  and this may explain their antibacterial activity.
The results obtained in this study, and mostly those obtained with the bark of B. acuta are very important when taking in consideration the fact that most of the bacterial strains used were MDR phenotypes expressing active efflux pumps [7–9, 34, 35]. In fact, the activity of antibiotics against the studied MDR bacteria was previously found to increase in the presence of phenylalanine arginine β-naphthylamide (PAßN), a potent inhibitor of RND efflux systems, particularly AcrAB–TolC (of Enterobaceriaceae) and MexAB–OprM (of Pseudomonas species) [7–9, 34, 35]. In the present study, we demonstrated that beneficial effects when combining four of the tested plant extracts [namely those from B. acuta (leaves and bark), N. leavis (leaves) and P. fulva (leaves)] with the first line antibiotics could be achieved. High percentages of synergistic effects (100 %) obtained with B. acuta bark extract and TET as well as P. fulva leaves extract in combination with TET and KAN, clearly suggest that such associations could improve the fight against MDR bacterial infections. This also suggests that some of the constituents of the corresponding plants can act as efflux pump inhibitors, as more than 70 % synergistic cases were observed with many combinations .
The antimicrobial potential of the genus Beilschmiedia has previously been documented. Chouna et al.  demonstrated that compounds such as beilschmiedic acid C isolated from B. anacardioides were significantly active against Bacillus subtilis, Micrococcus luteus and Streptococcus faecalis. Beilschmiedia cinnamomea was previously reported to have significant to moderate activities (64–1024 μg/mL) against the MDRGN tested in this work . Beilschmedia obscura was also found to show a good and large spectrum of antibacterial activity against MDRGN . Some compounds previously isolated from the genus Beilschmiedia and belonging to alkaloids, phenols, saponines, sterols and triterpenoids [36, 38] were shown to possess antimicrobial activities . The genus Beilschmiedia is also known traditionally to possess antimicrobial activities . Beilschmedia acuta tested in this study is also used in Cameroon to treat gastrointestinal infections . The obtained data highlight the importance of this plant in the control of microbial infections and mostly those involving MDR phenotypes. The antimicrobial activities of extracts and compounds from Newbouldia laevis towards sensitive bacteria and fungi were also reported [39, 40], and the present study provides additional data on the potential of this plant to fight MDR bacteria. Also, the antimicrobial activity of essential oil from Clausena anisata was reported against Staphylococcus aureus, Streptococcus pneumoniae, Enterococcus species, Salmonella typhimurium and Pseudomonas aeruginosa [41, 42]. The present report provides more evidence of the antimicrobial potential of this plant.
The results of this study are very interesting, in regards to the medical importance of the studied microorganisms. These data provided evidence that crude extracts from the studied plants and mostly that from the bark of Beilschmedia acuta are potential sources of antimicrobial drugs to fight MDR bacterial infections. The purification of this plant will be carried out to isolate its active constituents. The cytotoxicity assays on normal cell lines constitute the limitation of the present work and will further be performed to ensure the safety of the tested extracts.
Adwan G, Abu-Shanab B, Adwan K. Antibacterial activities of some plant extracts alone and in combination with different antimicrobials against multidrug-resistant Pseudomonas aeruginosa strains. Asian Pac J Trop Med. 2010;3(4):266–9.
Rice LB. Unmet medical needs in antibacterial therapy. Biochem Pharmacol. 2006;71(7):991–5.
Ates DA, Erdogrul OT. Antimicrobial activity of various medicinal and commercial plants extracts. Turkian J Biol. 2003;27:157–62.
WHO WHO. Tuberculosis, Fact sheet No. 104. World Health Organization: Geneva, Switzerland 2010, http://www.kff.org/globalhealth/upload/7883-02.pdf. Accessed on February 02, 2012.
Newman DJ, Cragg GM. Natural Products as Sources of New Drugs over the Last 25 Years. J Nat Prod. 2007;70(3):461–77.
Coutinho HD, Lima JG, Siqueira-Júnior JP. Additive effects of Hyptis martiusii Benth with aminoglycosides against Escherichia coli. Indian J Med Res. 2010;131:106–8.
Fankam AG, Kuete V, Voukeng IK, Kuiate JR, Pages JM. Antibacterial activities of selected Cameroonian spices and their synergistic effects with antibiotics against multidrug-resistant phenotypes. BMC Complement Altern Med. 2011;11:104.
Voukeng IK, Kuete V, Dzoyem JP, Fankam AG, Noumedem JA, Kuiate JR, et al. Antibacterial and antibiotic-potentiation activities of the methanol extract of some cameroonian spices against Gram-negative multi-drug resistant phenotypes. BMC Res Notes. 2012;5:299.
Noumedem JA, Mihasan M, Kuiate JR, Stefan M, Cojocaru D, Dzoyem JP, et al. In vitro antibacterial and antibiotic-potentiation activities of four edible plants against multidrug-resistant gram-negative species. BMC Complement Altern Med. 2013;13:190.
Kuete V, Tankeo SB, Saeed ME, Wiench B, Tane P, Efferth T. Cytotoxicity and modes of action of five Cameroonian medicinal plants against multi-factorial drug resistance of tumor cells. J Ethnopharmacol. 2014;153(1):207–19.
Kupchan SM, Moniot JL, Sigel CW, Hemingway RJ. Tumor inhibitors. LXV. Bersenogenin, berscillogenin, and 3-epiberscillogenin, three new cytotoxic bufadienolides from Bersama abyssinica. J Org Chem. 1971;36(18):2611–6.
Bowen IH, Jackson BP, Motawe HM. An Investigation of the Stem Bark of Bersama abyssinica. Planta Med. 1985;51(6):483–7.
Gafner S, Wolfender J, Nianga M, Stoeckli-Evans H, Hostettmann K. Antifungal and antibacterial naphtoquinones from Newbouldia laevis roots. Phytochemistry. 1996;42(5):1315–20.
Dandjesso C, Klotoé J, Dougnon T, Sègbo J, Atègbo J, Gbaguidi F, et al. Phytochemistry and hemostatic properties of some medicinal plants sold as anti-hemorrhagic in Cotonou markets (Benin). Indian J Sci Technol. 2012;5(8):3105–9.
Eyong KO, Krohn K, Hussain H, Folefoc GN, Nkengfack AE, Schulz B, et al. Newbouldiaquinone and newbouldiamide: a new naphthoquinone-anthraquinone coupled pigment and a new ceramide from Newbouldia laevis. Chem Pharm Bull (Tokyo). 2005;53(6):616–9.
Jeruto P, Lukhoba C, Ouma G, Otieno D, Mutai C. Herbal treatments in Aldai and Kaptumo divisions in Nandi district, Rift valley province, Kenya. Afr J Tradit Complement Altern Med. 2007;5(1):103–5.
Focho D, WT N, Fonge B. Medicinal plants of Aguambu - Bamumbu in the Lebialem highlands, southwest province of Cameroon. African J Pharm Pharmacol. 2009;3(1):1–13.
Eloff JN. A sensitive and quick microplate method to determine the minimal inhibitory concentration of plant extracts for bacteria. Planta Med. 1998;64(8):711–3.
Mativandlela SPN, Lall N, Meyer JJM. Antibacterial, antifungal and antitubercular activity of (the roots of) Pelargonium reniforme (CURT) and Pelargonium sidoides (DC) (Geraniaceae) root extracts. S Afr J Bot. 2006;72(2):232–7.
Kuete V, Kamga J, Sandjo LP, Ngameni B, Poumale HM, Ambassa P, et al. Antimicrobial activities of the methanol extract, fractions and compounds from Ficus polita Vahl. (Moraceae). BMC Complement Altern Med. 2011;11:6.
Kuete V, Nana F, Ngameni B, Mbaveng AT, Keumedjio F, Ngadjui BT. Antimicrobial activity of the crude extract, fractions and compounds from stem bark of Ficus ovata (Moraceae). J Ethnopharmacol. 2009;124(3):556–61.
Kuete V, Wansi JD, Mbaveng AT, Kana Sop MM, Tadjong AT, Beng VP, et al. Antimicrobial activity of the methanolic extract and compounds from Teclea afzelii (Rutaceae). S Afr J Bot. 2008;74(4):572–6.
Kuete V, Ngameni B, Simo CC, Tankeu RK, Ngadjui BT, Meyer JJ, et al. Antimicrobial activity of the crude extracts and compounds from Ficus chlamydocarpa and Ficus cordata (Moraceae). J Ethnopharmacol. 2008;120(1):17–24.
Kuete V, Wabo GF, Ngameni B, Mbaveng AT, Metuno R, Etoa FX, et al. Antimicrobial activity of the methanolic extract, fractions and compounds from the stem bark of Irvingia gabonensis (Ixonanthaceae). J Ethnopharmacol. 2007;114(1):54–60.
Shahverdi AR, Monsef-Esfahani HR, Tavasoli F, Zaheri A, Mirjani R. Trans-Cinnamaldehyde from Cinnamomum zeylanicum bark essential oil reduces the clindamycin resistance of Clostridium difficile in vitro. J Food Sci. 2007;72(1):S055–8.
Braga LC, Leite AAM, Xavier KGS, Takahashi JA, Bemquerer MP, Chartone-Souza E, et al. Synergic interaction between pomegranate extract and antibiotics against Staphylococcus aureus. Can J Microbiol. 2005;51(7):541–7.
Simões M, Bennett R, Rosa E. Understanding antimicrobial activities of phytochemicals against multidrug resistant bacteria and biofilms. Nat Prod Rep. 2009;26:746–57.
Kuete V. Potential of Cameroonian plants and derived products against microbial infections: a review. Planta Med. 2010;76(14):1479–91.
Kuete V, Efferth T. Cameroonian medicinal plants: pharmacology and derived natural products. Front Pharmacol. 2010;1:123.
Fabry W, Okemo P, Ansorg R. Antibacterial activity of East African medicinal plants. J Ethnopharmacol. 1998;60:79–84.
Mims C, Playfair J, Roitt I, Wakelin D, Williams R. Antimicrobials and chemotherapy. In: Mims CA, et al. Eds, Med Microbiol Rev. 1993; 35:1–34.
Mbaveng AT, Kuete V, Mapunya BM, Beng VP, Nkengfack AE, Meyer JJ, et al. Evaluation of four Cameroonian medicinal plants for anticancer, antigonorrheal and antireverse transcriptase activities. Environ Toxicol Pharmacol. 2011;32(2):162–7.
Mbaveng AT, Ngameni B, Kuete V, Simo IK, Ambassa P, Roy R, et al. Antimicrobial activity of the crude extracts and five flavonoids from the twigs of Dorstenia barteri (Moraceae). J Ethnopharmacol. 2008;116(3):483–9.
Kuete V, Ngameni B, Tangmouo JG, Bolla JM, Alibert-Franco S, Ngadjui BT, et al. Efflux pumps are involved in the defense of Gram-negative bacteria against the natural products isobavachalcone and diospyrone. Antimicrob Agents Chemother. 2010;54(5):1749–52.
Kuete V, Tangmouo JG, Meyer JJ, Lall N. Diospyrone, crassiflorone and plumbagin: three antimycobacterial and antigonorrhoeal naphthoquinones from two Diospyros spp. Int J Antimicrob Agents. 2009;34(4):322–5.
Chouna JR, Nkeng-Efouet PA, Lenta BN, Devkota KP, Neumann B, Stammler HG, et al. Antibacterial endiandric acid derivatives from Beilschmiedia anacardioides. Phytochemistry. 2009;70(5):684–8.
Fankam AG, Kuiate JR, Kuete V. Antibacterial activities of Beilschmiedia obscura and six other Cameroonian medicinal plants against multi-drug resistant Gram-negative phenotypes. BMC Complement Altern Med. 2014;14:241.
Chen JJ, Chou ET, Peng CF, Chen IS, Yang SZ, Huang HY. Novel epoxyfuranoid lignans and antitubercular constituents from the leaves of Beilschmiedia tsangii. Planta Med. 2007;73(6):567–71.
Kuete V, Eyong KO, Folefoc GN, Beng VP, Hussain H, Krohn K, et al. Antimicrobial activity of the methanolic extract and of the chemical constituents isolated from Newbouldia laevis. Pharmazie. 2007;62(7):552–6.
Eyong KO, Folefoc GN, Kuete V, Beng VP, Krohn K, Hussain H, et al. Newbouldiaquinone A: A naphthoquinone-anthraquinone ether coupled pigment, as a potential antimicrobial and antimalarial agent from Newbouldia laevis. Phytochemistry. 2006;67(6):605–9.
Nelson RR. In-vitro activities of five plant essential oils against methicillin-resistant Staphylococcus aureus and vancomycin-resistant Enterococcus faecium. J Antimicrob Chemother. 1997;40(2):305–6.
Senthilkumar A, Venkatesalu V. Phytochemical analysis and antibacterial activity of the essential oil of Clausena anisata (Willd.) Hook. F. ex Benth. Int J Integr Biol. 2009;5(2):116–20.
Adesinan SK, Ette EI. The isolation and identification of anticonvulsant agents from Clausena anisata and Afraegle paniculata. Fitoterapia. 1982;53:63–6.
Hutchings A, Scott A, Lewis G, Cunningham A. Zulu medicinal plants. Pietermaritzburg: Natal University Press; 1996.
Ojewole JA. Hypoglycaemic effect of Clausena anisata (Willd) Hook methanolic root extract in rats. J Ethnopharmacol. 2002;81(2):231–7.
Ito C, Itoigawa M, Aizawa K, Yoshida K, Ruangrungsi N, Furukawa H. Gamma-lactone carbazoles from Clausena anisata. J Nat Prod. 2009;72(6):1202–4.
Ngadjui B, Mouncherou S, Ayafor J, Sondengam B, Tillequin F. Geranyl coumarins from Clausena anisata. Phytochemistry. 1991;30:2809–11.
Usman H, Osuji JC. Phytochemical and in vitro antimicrobial assay of the leaf extract of Newbouldia laevis. Afr J Tradit Complement Altern Med. 2007;4(4):476–80.
Kuete V, Mbaveng AT, Tsaffack M, Beng VP, Etoa FX, Nkengfack AE, et al. Antitumor, antioxidant and antimicrobial activities of Bersama engleriana (Melianthaceae). J Ethnopharmacol. 2008;115(3):494–501.
Tshibangu JN, Chifundera K, Kaminsky R, Wright AD, Konig GM. Screening of African medicinal plants for antimicrobial and enzyme inhibitory activity. J Ethnopharmacol. 2002;80(1):25–35.
Bedir E, Toyang NJ, Khan IA, Walker LA, Clark AM. A new dammarane-type triterpene glycoside from Polyscias fulva. J Nat Prod. 2001;64(1):95–7.
Kuete V, Efferth T. Pharmacogenomics of Cameroonian traditional herbal medicine for cancer therapy. J Ethnopharmacol. 2011;137(1):752–66.
Njayou FN, Moundipa PF, Tchana AN, Ngadjui BT, Tchouanguep FM. Inhibition of microsomal lipid peroxidation and protein oxidation by extracts from plants used in Bamun folk medicine (Cameroon) against hepatitis. Afr J Tradit Complement Altern Med. 2008;5(3):278–89.
Viveiros M, Jesus A, Brito M, Leandro C, Martins M, Ordway D, et al. Inducement and reversal of tetracycline resistance in Escherichia coli K-12 and expression of proton gradient-dependent multidrug efflux pump genes. Antimicrob Agents Chemother. 2005;49(8):3578–82.
Okusu H, Ma D, Nikaido H. AcrAB efflux pump plays a major role in the antibiotic resistance phenotype of Escherichia coli multiple-antibiotic-resistance (Mar) mutants. J Bacteriol. 1996;178(1):306–8.
Elkins CA, Mullis LB. Substrate competition studies using whole-cell accumulation assays with the major tripartite multidrug efflux pumps of Escherichia coli. Antimicrob Agents Chemother. 2007;51(3):923–9.
Kuete V, Alibert-Franco S, Eyong KO, Ngameni B, Folefoc GN, Nguemeving JR, et al. Antibacterial activity of some natural products against bacteria expressing a multidrug-resistant phenotype. Int J Antimicrob Agents. 2011;37(2):156–61.
Baglioni P, Bini L, Liberatori S, Pallini V, Marri L. Proteome analysis of Escherichia coli W3110 expressing an heterologous sigma factor. Proteomics. 2003;3(6):1060–5.
Sar C, Mwenya B, Santoso B, Takaura K, Morikawa R, Isogai N, et al. Effect of Escherichia coli wild type or its derivative with high nitrite reductase activity on in vitro ruminal methanogenesis and nitrate/nitrite reduction. J Anim Sci. 2005;83(3):644–52.
Ghisalberti D, Masi M, Pages JM, Chevalier J. Chloramphenicol and expression of multidrug efflux pump in Enterobacter aerogenes. Biochem Biophys Res Commun. 2005;328(4):1113–8.
Mallea M, Chevalier J, Bornet C, Eyraud A, Davin-Regli A, Bollet C, et al. Porin alteration and active efflux: two in vivo drug resistance strategies used by Enterobacter aerogenes. Microbiology. 1998;144(Pt 11):3003–9.
Mallea M, Mahamoud A, Chevalier J, Alibert-Franco S, Brouant P, Barbe J, et al. Alkylaminoquinolines inhibit the bacterial antibiotic efflux pump in multidrug-resistant clinical isolates. Biochem J. 2003;376(Pt 3):801–5.
Pradel E, Pages JM. The AcrAB-TolC efflux pump contributes to multidrug resistance in the nosocomial pathogen Enterobacter aerogenes. Antimicrob Agents Chemother. 2002;46(8):2640–3.
Chevalier J, Pages JM, Eyraud A, Mallea M. Membrane permeability modifications are involved in antibiotic resistance in Klebsiella pneumoniae. Biochem Biophys Res Commun. 2000;274(2):496–9.
Tran QT, Mahendran KR, Hajjar E, Ceccarelli M, Davin-Regli A, Winterhalter M, et al. Implication of porins in beta-lactam resistance of Providencia stuartii. J Biol Chem. 2010;285(42):32273–81.
Lorenzi V, Muselli A, Bernardini AF, Berti L, Pages JM, Amaral L, et al. Geraniol restores antibiotic activities against multidrug-resistant isolates from Gram-negative species. Antimicrob Agents Chemother. 2009;53(5):2209–11.
Authors are thankful to the Cameroon National Herbarium (Yaounde) for the plant identification. Authors are also thankful to UMR-MD1 (Mediterranean University, Marseille, France) for providing some clinical bacteria.
The authors declare that there are no conflict of interest.
SBT carried out the study; VK designed the experiments and wrote the manuscript; VK and PT supervised the work; VK provided the bacterial strains; all authors read and approved the final manuscript.
MICs up to 1024 μg/mL of the crude extracts and ciprofloxacin on the panel of tested bacteria. Table S2. MBCs up to 1024 μg/mL of the crude extracts and ciprofloxacin on the panel of tested bacteria. (DOCX 29 kb)