Antimicrobial and Efflux Inhibitor Activity of Usnic Acid Against Mycobacterium abscessus

Abstract

New drugs are needed to treat infections with antimicrobial-resistant Mycobacterium abscessus; therefore, we evaluated usnic acid as an antimicrobial agent and efflux inhibitor (EI) against M. abscessus. Usnic acid showed antimicrobial activity, and synergistically, the EI verapamil increased this activity. In addition, when we evaluated the interaction of antimicrobials with usnic acid, the increase of their activity was observed. Finally, usnic acid showed an efflux inhibitory effect between the classical EIs verapamil and carbonyl cyanide m-chlorophenylhydrazine. In conclusion, usnic acid showed both antimicrobial and EI activity, indicating that this natural compound may be a promising scaffold for new drugs against this difficult-to-treat microorganism.

Key words

usnic acid - verapamil - Mycobacterium abscessus - efflux inhibitor - antimicrobial

• References

• 1 Stout JS. Treatment of Mycobacterium abscessus: all macrolides are equal, but perhaps some are more equal than others. Am J Respir Crit Care Med 2012; 186: 822-823
• 2 Tortoli E. Microbiological features and clinical relevance of new species of the genus Mycobacterium . Clin Microbiol Ver 2014; 27: 727-752
• 3 Mata R, Morales I, Perez O, Rivero-Cruz I, Acevedo L, Enriquez-Mendoza I, Bye R, Franzblau S, Timmermann B. Antimycobacterial compounds from Piper sanctum . J Nat Prod 2004; 67: 1961-1968
• 4 Lam KS. New aspects of natural products in drug discovery. Trends Microbiol 2007; 15: 279-289
• 5 Azevedo LG, Muccillo-Baisch AL, Filgueira DM, Boyle RT, Ramos DF, Soares AD, Lerner C, Silva PA, Trindade GS. Comparative cytotoxic and anti-tuberculosis activity of Aplysina caissara marine sponge crude extracts. Comp Biochem Physiol C Toxicol Pharmacol 2008; 147: 36-42
• 6 Shibata S, Ukita T, Tamura T, Miura Y. Relation between chemical constitutions and antibacterial effects of usnic acid and derivates. Jap Med J 1948; 1: 152-155
• 7 Ingolfsdottir K. Usnic acid. Phytochemistry 2002; 61: 729-736
• 8 Araújo AA, de Melo MG, Rabelo TK, Nunes PS, Santos SL, Serafini MR, Santos MR, Quintans-Júnior LJ, Gelain DP. Review of the biological properties and toxicity of usnic acid. Nat Prod Res 2015; 29: 2167-2180
• 9 Vijayakumara CS, Viswanathana S, Kannappa Reddya UM, Parvathavarthinia S, Kundub AB, Sukumar E. Anti-inflammatory activity of (+) -usnic acid. Fitoterapia 2000; 71: 564-566
• 10 Stoll A, Brack A, Renz J. The effect of lichenic substances on the tubercle bacillus and certain other microorganisms; seventh article on antibacterial substances. Schweiz Z Pathol Bakteriol 1950; 13: 729-751
• 11 Ingólfsdóttir K, Chung GA, Skulason VG, Gissurarson SR, Vilhelmsdottir M. Antimycobacterial activity of lichen metabolites in vitro . Eur J Pharm 1998; 6: 141-144
• 12 Copp BR, Pearce NA. Natural product growth inhibitors of Mycobacterium tuberculosis . Nat Prod Rep 2007; 24: 278-297
• 13 Ramos DF, Silva PEAS. Antimycobacterial activity of usnic acid against resistant and susceptible strains of Mycobacterium tuberculosis and non-tuberculous mycobacteria. Pharm Biol 2010; 48: 260-263
• 14 Carvalho RSF, Pereira MA, Linhares LA, Lira-Nogueira MCB, Cavalcanti IMF, Santos-Magalhães NS, Montenegro LML. Effects of the encapsulation of usnic acid into liposomes and interactions with antituberculous agents against multidrug-resistant tuberculosis clinical isolates. Mem Inst Oswaldo Cruz 2016; 111: 330-334
• 15 Nessar R, Reyrat EJM, Murray A, Gicquel B. Mycobacterium abscessus: a new antibiotic nightmare. J Antimicrob Chemother 2012; 67: 810-818
• 16 Carvalho NFG, Sato DN, Pavan FR, Ferrazoli L, Chimara E. Resazurin microtiter assay for clarithromycin susceptibility testing of clinical isolates of Mycobacterium abscessus group. J Clin Lab Anal 2016; 30: 751-755
• 17 Rodrigues L, Sampaio D, Couto I, Machado D, Kern WV, Amaral L, Viveiros M. The role of efflux pumps in macrolide resistance in Mycobacterium avium complex. Int J Antimicrob Agents 2009; 34: 529-533
• 18 Groblacher B, Kunert O, Bucar F. Compounds of Alpinia katsumadai as potential efflux inhibitors in Mycobacterium smegmatis . 2012; 20: 2701-2706
• 19 Gupta S, Tyagi S, Almeida DV, Maiga MC, Ammerman NC, Bishai WR. Acceleration of tuberculosis treatment by adjunctive therapy with verapamil as an efflux inhibitor. Am J Respir Crit Care Med 2013; 188: 600-607
• 20 Coelho T, Machado D, Couto I, Maschmann R, Ramos D, Von Groll A, Rossetti ML, Silva PA, Viveiros M. Enhancement of antibiotic activity by efflux inhibitors against multidrug resistant Mycobacterium tuberculosis clinical isolates from Brazil. Front Microbiol 2015; 6: 1-12
• 21 Maciazg-dorszynska M, Wezgrzyn G, Guzow-Krzeminska B. Antibacterial activity of lichen secondary metabolite usnic acid is primarily caused by inhibition of RNA and DNA synthesis. FEMS Microbiol Lett 2014; 353: 57-62
• 22 Clinical and Laboratory Standards Institute. Susceptibility Testing of Mycobacteria, Nocardiae, and other aerobic Actinomycetes; Approved Standard. 2nd ed.. ed. Wayne, PA: Clinical Standards Laboratory Institute; 2011: 1-76
• 23 Ramis IB, Cnockaert M, Von Groll A, Nogueira CL, Leão SC, Andre E, Simon A, Palomino JC, Da Silva PE, Vandamme P, Martin A. Antimicrobial susceptibility of rapidly growing mycobacteria using the rapid colorimetric method. Eur J Clin Microbiol Infect Dis 2015; 34: 1403-1413
• 24 Palomino JC, Martin A, Camacho M, Guerra H, Swings J, Portaels F. Resazurin microtiter assay plate: simple and inexpensive method for detection of drug resistance in Mycobacterium tuberculosis . 2002; 46: 2720-2722
• 25 Pillai S, Moellering R, Eliopoulos G. Antimicrobial Combinations. In: Lorian V. ed. Antibiotics in laboratory Medicine. 5th ed. Baltimore, MD: Williams and Wilkins; 2005: 365-440
• 26 Furuya R, Koga Y, Irie S, Ikeda F, Akiko K, Kobayashi I, Masatoshi T. In vitro activities of antimicrobial combinations against clinical isolates of Neisseria gonorrhoeae . Infect Chemother 2013; 19: 1218-1220
• 27 Ahmed SA, Gogal RM, Walsh jr. JE. A new rapid and simple non-radioactive assay to monitor and determine the proliferation of lymphocytes: an alternative to [3H] thymidine incorporation assay. J Immunol Methods 1994; 170: 211-224
• 28 Snewin VA, Gares MP, Gaora PO, Hasan Z, Brown IN, Young DB. Assessment of immunity to mycobacterial infection with luciferase reporter constructs. Infect Immun 1999; 67: 4586-4593
• 29 Orme I. Search for new drugs for treatment of tuberculosis. Antimicrob Agents Chemother 2001; 45: 1943-1946
• 30 Pavan FR, Sato DN, Leite CQF. An Approach to the Search for new Drugs against Tuberculosis. In: Cardona PJ. ed. Understanding Tuberculosis – New Approaches to Fighting against Drug Resistance. Rijek: In Tech; 2005: 137-146
• 31 Viveiros M, Rodrigues L, Martins M, Couto I, Spengler G, Martins A, Amaral L. Evaluation of efflux activity of bacteria by a semi-automated fluorometric system. Methods Mol Biol 2010; 642: 159-172
• 32 Paixão L, Rodrigues L, Couto I, Martins M, Fernandes P, Carvalho CCCR, Monteiro GA, Sansonetty F, Amaral L, Viveiros M. Fluorometric determination of ethidium bromide efflux kinetics in Escherichia coli . J Biol Eng 2009; 3: 1-18
• 33 Machado D, Coelho T, Perdigão J, Pereira C, Couto I, Portugal I, Maschmann R, Ramos D, Von Groll A, Rossetti MLR, Silva PA, Viveiros M. Interplay between mutations and efflux in drug resistant clinical isolates of Mycobacterium tuberculosis . Front Microbiol 2017; 8: 1-18