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Molecular profiling of multidrug-resistant river water isolates: insights into resistance mechanism and potential inhibitors

Abstract

Polluted waters are an important reservoir for antibiotic resistance genes and multidrug-resistant bacteria. This report describes the microbial community, antibiotic resistance genes, and the genetic profile of extended spectrum β-lactamase strains isolated from rivers at, Pune, India. ESBL-producing bacteria isolated from diverse river water catchments running through Pune City were characterized for their antibiotic resistance. The microbial community and types of genes which confer antibiotic resistance were identified followed by the isolation of antibiotic-resistant bacteria on selective media and their genome analysis. Four representative isolates were sequenced using next generation sequencing for genomic analysis. They were identified as Pseudomonas aeruginosa, Escherichia coli, and two isolates were Enterobacter cloacae. The genes associated with the multidrug efflux pumps, such as tolC, macA, macB, adeL, and rosB, were detected in the isolates. As MacAB-TolC is an ABC type efflux pump responsible for conferring resistance in bacteria to several antibiotics, potential efflux pump inhibitors were identified by molecular docking. The homology model of their MacB protein with that from Escherichia coli K12 demonstrated structural changes in different motifs of MacB. Molecular docking of reported efflux pump inhibitors revealed the highest binding affinity of compound MC207-110 against MacB. It also details the potential efflux pump inhibitors that can serve as possible drug targets in drug development and discovery.

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References

  1. Andrews S (2010) FastQC: a quality control tool for high throughput sequence data. Available online at: http://www.bioinformatics.babraham.ac.uk/projects/fastqc

  2. APHA (1998) Standard methods for the examination of water and waste water American Public Health Association. pp 874

  3. Bankevich A, Nurk S, Antipov D, Gurevich AA, Dvorkin M, Kulikov AS, Lesin VM, Nikolenko SI, Pham S, Prjibelski AD, Pyshkin AV, Sirotkin AV, Vyahhi N, Tesler G, Alekseyev MA, Pevzner PA (2012) SPAdes: a new genome assembly algorithm and its applications to single-cell sequencing. J Comput Biol 19(5):455–477

  4. Blaser MJ (2016) Antibiotic use and its consequences for the normal microbiome. Science 352(6285):544–545

  5. Bogomolnaya LM, Andrews KD, Talamantes M, Maple A, Ragoza Y, Vazquez-Torres A, Andrews-Polymenis H (2013) The ABC-type efflux pump MacAB protects Salmonella enteric serovar typhimurium from oxidative stress. M Bio 4(6):e00630–e00613

  6. Center for Disease Dynamics, Economics and Policy (2015) The State of the World’s Antibiotics 2015(13):70318–70319

  7. Coyne S, Rosenfeld N, Lambert T, Courvalin P, Périchon B (2010) Over expression of resistance-nodulation-cell division pump AdeFGH confers multidrug resistance in Acinetobacter baumannii. Antimicrob Agents Chemother 54(10):4389–4393

  8. Crow A, Greene NP, Kaplan E, Koronakis V (2017) Structure and mechanotransmission mechanism of the MacB ABC transporter superfamily. Proc Natl Acad Sci U S A 114:12572–12577

  9. Devarajan N, Laffite A, Mulaji CK, Otamonga JP, Mpiana PT, Mubedi JI, Prabakar K, Ibelings WB, Poté J (2016) Occurrence of antibiotic resistance genes and bacterial markers in a tropical river receiving hospital and urban wastewaters. PLoS One 11(2):e0149211

  10. Dhawde R, Macaden R, Ghadge A, Birdi T (2018) Seasonal prevalence of antibiotic-resistant bacteria in the river Mula-Mutha, India. Environ Monit Assess 190:533

  11. Environment Protection Authority June 2007 (2007) EPA guidelines-regulatory monitoring and testing water and waste water sampling. ISBN 978–1–921125-47-8

  12. Environmental Protection Agency (2001) Parameters of water quality interpretation and standards. ISBN 1-84096-015-3

  13. Fitzpatrick WP, Llabres S, Neuberger A, Blaza JN, Bai XC, Okada U, Murakami S, van Veen HW, Zachariae U, Scheres SHW, Luisi BF, Du D (2017) Structure of the MacAB–TolC ABC-type tripartite multidrug efflux pump. Nat Microbiol 2:17070–17070

  14. Guo J, Li J, Chen H, Bond P, Yuan Z (2017) Metagenomic analysis reveals wastewater treatment plants as hotspots of antibiotic resistance genes and mobile genetic elements. Water Res 123:468–478

  15. Gurevich A, Saveliev V, Vyahhi N, Tesler G (2013) QUAST: quality assessment tool for genome assemblies. Bioinformatics 29(8):1072–1075

  16. Jia B, Raphenya AR, Alcock B, Waglechner N, Guo P, Tsang KK, Lago BA, Dave BM, Pereira S, Sharma AN, Doshi S, Courtot M, Lo R, Williams LE, Frye JG, Elsayegh T, Sardar D, Westman EL, Pawlowski AC, Johnson TA, Brinkman FS, Wright GD, McArthur AG (2017) CARD 2017: expansion and model-centric curation of the comprehensive antibiotic resistance database. Nucleic Acids Res 45(D1):D566–D573

  17. Karkman A, Johnson TA, Lyra C, Stedtfeld RD, Tamminen M, Tiedje JM, Virta M (2016) High-throughput quantification of antibiotic resistance genes from an urban wastewater treatment plant. FEMS Microb Ecol 92(3) pii: fiw014). https://doi.org/10.1093/femsec/fiw014

  18. Kiratisin P, Apisarnthanarak A, Laesripa C, Saifon P (2008) Molecular characterization and epidemiology of extended-spectrum-β-lactamase-producing Escherichia coli and Klebsiella pneumoniae isolates causing health care-associated infection in Thailand, where the CTX-M family is endemic. Antimicrob Agents Chemother 52(8):2818–2824

  19. Klein E, Boeckel T, Martinez E, Pant S, Gandra S, Levin S, Goossens H, Laxminarayan R (2018) Global increase and geographic convergence in antibiotic consumption between 2000 and 2015. Proc Natl Acad Sci U S A 115:E3463–E3470

  20. Kohanski MA, Dwyer DJ, Collins JJ (2010) How antibiotics kill bacteria: from targets to networks. Nat Rev Microbiol 8(6):423–435

  21. Kramer B, Rarey M, Lengauer T (1999) Evaluation of the FLEXX incremental construction algorithm for protein-ligand docking. Proteins Struct Funct Genet 37(2):228–241

  22. Laffite A, Kilunga PI, Kayembe JM, Devarajan N, Mulaji CK, Giuliani G, Poté J (2016) Hospital effluents are one of several sources of metal, antibiotic resistance genes, and bacterial markers disseminated in Sub-Saharan urban Rivers. Front Microbiol 7:1128

  23. Larsen MV, Cosentino S, Rasmussen S, Friis C, Hasman H, Marvig RL, Jelsbak L, Sicheritz-Pontén T, Ussery DW, Aarestrup FM, Lund O (2012) Multilocus sequence typing of total genome sequenced bacteria. J Clin Micobiol 50(4):1355–1361

  24. Lemarchand K, Berthiaume F, Maynard C, Harel J, Payment P, Bayardelle P, Masson L, Brousseau R (2005) Optimization of microbial DNA extraction and purification from raw wastewater samples for downstream pathogen detection by microarrays. J Microbiol Methods 63:115–126

  25. Lomovskaya O, Warren MS, Lee A, Galazzo J, Fronko R, Lee M, Blais J, Cho D, Chamberland S, Renau T, Leger R, Hecker S, Watkins W, Hoshino K, Ishida H, Lee VJ (2001) Identification and characterization of inhibitors of multidrug resistance efflux pumps in pseudomonas aeruginosa: Novel agents for combination therapy. antimicrobial agents and chemotherapy 45(1):105–116

  26. Madec JY, Haenni M, Nordmann P, Poirel L (2017) Extended-spectrum β-lactamase/AmpC-and carbapenemase-producing Enterobacteriaceae in animals: a threat for humans? Clin Microbiol Infect 23(11):826–833

  27. Mahmood HY, Jamshidi S, Sutton JM, Rahman KM (2016) Current advances in developing inhibitors of bacterial multidrug efflux pumps. Curr Med Chem 23(10):1062–1081

  28. Marquez B, (2005) Bacterial efflux systems and efflux pumps inhibitors. Biochimie 87 (12):1137–1147

  29. Matsuo T, Chen J, Minato Y, Ogawa W, Mizushima T, Kuroda T, Tsuchiya T (2007) SmdAB, a heterodimeric ABC-type multidrug efflux pump, in Serratia marcescens. J Bacteriol 190(2):648–654

  30. Naas T, Poirel L, Nordmann P (2008) Minor extended-spectrum ß-lactamases. Clin Microbiol Infect 14:42–52

  31. Nakayama T, Hoa T, Harada K, Warisaya M, Asayama M, Hinenoya A, Lee J, Phu T, Ueda S, Sumimura Y, Hirata K, Phuong N, Yamamoto Y (2017) Water metagenomic analysis reveals low bacterial diversity and the presence of antimicrobial residues and resistance genes in a river containing wastewater from backyard aquacultures in the Mekong Delta, Vietnam. Environ Pollut 222:294–306

  32. Nawani N, Rahman A, Nahar N, Saha A, Kapadnis B, Mandal A (2016) Status of metal pollution in rivers flowing through urban settlements at Pune and its effect on resident microflora. Biologia 71(5):494–507

  33. Ni W, Li Y, Guan J, Zhao J, Cui J, Wang R, Liu Y (2016) Effects of efflux pump inhibitors on colistin resistance in multidrug-resistant Gram-negative bacteria. Antimicrob Agents Chemother 60(5):3215–3218

  34. Oliveira DV, Nunes LS, Barth AL, Van Der Sand ST (2017) Genetic background of β-lactamases in Enterobacteriaceae isolates from environmental samples. Microb Ecol:1–9

  35. Phan G, Picard M, Broutin I (2015) Focus on the outer membrane factor OprM, the forgotten player from efflux pumps assemblies. Antibio 4(4):544–566

  36. Rompré A, Servais P, Baudart J, De-Roubin R, Laurent P (2002) Detection and enumeration of coliforms in drinking water: current methods and emerging approaches. J Microbiol Methods 49(1):31–54

  37. Schrodinger Release 2017–4 (2017) Maestro Schrodinger. LLC, New York, NY

  38. Schwartz T, Kohnen W, Jansen B, Obst U (2003) Detection of antibiotic-resistant bacteria and their resistance genes in wastewater, surface water, and drinking water biofilms. FEMS Microbiol Ecol 43(3):325–335

  39. Shriram V, Khare T, Bhagwat R, Shukla R, Kumar V (2018) Inhibiting bacterial drug efflux pumps via phyto-therapeutics to combat threatening antimicrobial resistance. Front Microbiol 9:2990. https://doi.org/10.3389/fmicb.2018.02990

  40. Sun J, Deng Z, Yan A (2014) Bacterial multidrug efflux pumps: mechanism, physiology and pharmacological exploitations. Biochem Biophys Res Commun 453:254–267

  41. Sun W, Qian X, Gu J, Wang J, Duan L (2016) Mechanism and effect of temperature on variations in antibiotic resistance genes during anaerobic digestion of dairy manure. Sci Rep 6:30237

  42. Tikhonova B, Wang Q, Zgurskaya I (2002) Chimeric analysis of the multicomponent multidrug efflux transporters from gram-negative bacteria. J Bacteriol 184(23):6499–6507

  43. Wang Y, Chen Y, Zheng X, Gui C, Wei Y (2017) Spatio-temporal distribution of fecal indicators in three rivers of the Haihe River Basin, China. Environ Sci Pollut Res 24(10):9036–9047

  44. World Health Organization (2014) Antimicrobial resistance global report on surveillance. Accessed online at https://www.who.int/drugresistance/documents/surveillancereport/en

  45. Xu Y, Sim SH, Ki HN, Xiao LJ, Kim HM, Hwang KY, Lee K, Ha NC (2009) Crystal structure of the periplasmic region of MacB, a noncanonic ABC transporter. Biochem 48(23):5218–5225

  46. Zhao Z, Wang J, Han Y, Chen J, Liu G, Lu H, Chen S (2017) Nutrients, heavy metals and microbial communities co-driven distribution of antibiotic resistance genes in adjacent environment of mariculture. Environ Pollut 220:909–918

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Acknowledgements

Authors acknowledge the necessary infrastructure and support from University of Skӧvde, Sweden; Örebro University, Sweden and Dr. D.Y. Patil Vidyapeeth, Pune, India.

Funding

This study was supported by FORMAS, Swedish Research Council through the research grant (219-2014-837).

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Correspondence to Jana Jass or Neelu Nawani.

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Yewale, P.P., Lokhande, K.B., Sridhar, A. et al. Molecular profiling of multidrug-resistant river water isolates: insights into resistance mechanism and potential inhibitors. Environ Sci Pollut Res (2019). https://doi.org/10.1007/s11356-019-05738-2

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Keywords

  • Extended spectrum β-lactamase
  • Multidrug efflux pump
  • Efflux pump inhibitors
  • Antimicrobial resistance
  • MacB