Although the molecular mechanisms of carbapenem resistance of environmental isolates of Acinetobacter baumannii are well described, data on the mechanisms of colistin resistance are scarce. In this study, we report the molecular mechanisms of colistin resistance in environmental isolates of A. baumannii. Seven clinically relevant isolates of A. baumannii belonging to ST-2Pasteur were recovered from hospital wastewater and wastewater treatment plant. The phenotypic resistance to colistin was confirmed by broth microdilution with minimum inhibitory concentration values ranging from 20 to 160 mg/L. Colistin sulfate and colistimethate sodium showed bactericidal activity against two colistin-heteroresistant isolates in vitro, but substantially recovery of population was observed after prolonged incubation. In silico genome analysis revealed nucleotide variations resulting in amino acid changes in LpxC (N286D), LpxD (E117K), PmrB (A138T, R263S, L267W, Q309P, and A444V), and EptA (F166L, I228V, R348K, A370S, and K531T). According to reverse transcription quantitative PCR, all isolates had increased levels of eptA mRNA and decreased levels of lpxA and lpxD mRNA. Isolates expressed low hydrophobicity, biofilm, and pellicle formation, but showed excellent survival in river water during 50 days of monitoring. Colistin- and pandrug-resistant A. baumannii disseminated in the environment could represent the source for the occurrence of serious community-acquired infections.
Get full access to this article
View all access options for this article.
References
1.
AgodiA., VoulgariE., BarchittaM., QuattrocchiA., BellocchiP., PoulouA., and TsakrisA.. 2014. Spread of a carbapenem-and colistin-resistant Acinetobacter baumannii ST2 clonal strain causing outbreaks in two Sicilian hospitals. J. Hosp. Infect. 86:260–266.
2.
BakourS., OlaitanA.O, AmmariH., TouatiA., SaoudiS., SaoudiK., and RolainJ.M.. 2015. Emergence of colistin-and carbapenem-resistant Acinetobacter baumannii ST2 clinical isolate in Algeria: first case report. Microb. Drug Resist. 21:279–285.
3.
CafisoV., StracquadanioS., Lo VerdeF., GabrieleG., MezzatestaM.L, CaioC., PigolaG., FerroA., and StefaniS.. 2019. Colistin resistant A. baumannii: genomic and transcriptomic traits acquired under colistin therapy. Front. Microbiol. 9:3195.
ClausellA., Garcia-SubiratsM., PujolM., BusquetsM.A, RabanalF., and CajalY.. 2007. Gram-negative outer and inner membrane models: insertion of cyclic cationic lipopeptides. J. Phys. Chem. B. 111:551–563.
6.
MoffattJ.H., HarperM., HarrisonP., HaleJ.D, VinogradovE., SeemannT., HenryR., CraneB., St MichaelF., CoxA.D, AdlerB., NationR.L, LiJ., and BoyceJ.D.. 2010. Colistin resistance in Acinetobacter baumannii is mediated by complete loss of lipopolysaccharide production. Antimicrob. Agents Chemother. 54:4971–4977.
7.
OlaitanA.O., MorandS., and RolainJ.M.. 2014. Mechanisms of polymyxin resistance: acquired and intrinsic resistance in bacteria. Front. Microbiol. 5:643.
8.
ArroyoL.A., HerreraC.M, FernandezL., HankinsJ.V, TrentM.S, and HancockR.E.W. 2011. The pmrCAB operon mediates polymyxin resistance in Acinetobacter baumannii ATCC 17978 and clinical isolates through phosphoethanolamine modification of lipid A. Antimicrob. Agents Chemother. 55:3743–3751.
9.
LeeJ.Y., ChungE.S, and KoK.S.. 2017. Transition of colistin dependence into colistin resistance in Acinetobacter baumannii. Sci. Rep. 7:1–11.
10.
GersonS., BettsJ.W, LucaßeK., NodariC.S, WilleJ., JostenM., and VilaJ.. 2019. Investigation of novel pmrB and eptA mutations in isogenic Acinetobacter baumannii isolates associated with colistin resistance and increased virulence in vivo. Antimicrob. Agents Chemother. 63:e01586-18.
11.
NurtopE., BilmanF.B, MenekseS., AzapO.K, GönenM., ErgonulO., and CanF.. 2019. Promoters of colistin resistance in Acinetobacter baumannii infections. Microb. Drug Resist. 25:997–1002.
12.
LeshoE., YoonE.J, McGannP., SnesrudE., KwakY., MililloM., and WatermanP.E.. 2013. Emergence of colistin-resistance in extremely drug-resistant Acinetobacter baumannii containing a novel pmrCAB operon during colistin therapy of wound infections. J. Infec. Dis. 208:1142–1151.
13.
GharaibehM.H., and ShatnawiS.Q.. 2019. An overview of colistin resistance, mobilized colistin resistance genes dissemination, global responses, and the alternatives to colistin: a review. Vet. World, 12:1735–1746.
14.
HameedF., KhanM.A, MuhammadH., SarwarT., BilalH., and RehmanT.U. 2019. Plasmid-mediated mcr-1. gene in Acinetobacter baumannii and Pseudomonas aeruginosa: first report from Pakistan. Rev. Soc. Bras. Med. Trop. 52:e2019, 0237.
15.
MaF., ShenC., ZhengX., LiuY., ChenH., ZhongL., LiangY., LiaoK., XiaY., TianG.B, and YangY.. 2019. Identification of a novel plasmid carrying mcr-4.3 in an Acinetobacter baumannii strain in China. Antimicrob. Agents Chemother. 63:e00133-19.
16.
Seruga MusicM., HrenovicJ., Goic-BarisicI., HunjakB., SkoricD., and IvankovicT.. 2017. Emission of extensively-drug resistant Acinetobacter baumannii from hospital settings to the natural environment. J. Hosp. Infect. 96:232–237.
17.
HigginsP.G., HrenovicJ., SeifertH., and DekicS.. 2018. Characterization of Acinetobacter baumannii from water and sludge line of secondary wastewater treatment plant. Water Res. 140:261–267.
18.
EUCAST, European Committee on Antimicrobial Susceptibility Testing. Breakpoint tables for interpretation of MICs and zone diameters, Version 9.0. 2019.
19.
CLSI, Clinical and Laboratory Standards Institute. 2019. Performance Standards for Antimicrobial Susceptibility Testing, Document M100-ED29. Clinical and Laboratory Standards Institute, Wayne, NJ.
20.
LiJ., RaynerC.R, NationR.L, OwenR.J, SpelmanD., TanK.E, and LioliosL.. 2006. Htereroresistance to colistin in multidrug-resistant Acinetobacter baumannii. Antimicrob. Agents Chemother. 50:2946–2950.
21.
KojicM., StrahinicI., and TopisirovicL.. 2005. Proteinase PI and lactococcin A genes are located on the largest plasmid in Lactococcus lactis subsp. lactis bv. diacetylactis S50. Can. J. Microbiol. 51:305–314.
22.
PengY., LeungH.C, YiuS.M, and ChinF.Y.. 2012. IDBA-UD: a de novo assembler for single-cell and metagenomic sequencing data with highly uneven depth. Bioinformatics, 28:14208.
23.
LiH., and DurbinR.. 2010. Fast and accurate long-read alignment with Burrows-Wheeler transform. Bioinformatics, 26:58995.
24.
DiszT., AkhterS., CuevasD., OlsonR., OverbeekR., VonsteinV., StevensR., and EdwardsR.A.. 2010. Accessing the SEED genome databases via Web services API: tools for programmers. BMC Bioinf. 11:319.
25.
HoodM.I., JacobsA.C, SayoodK., DunmanP.M, and SkaarE.P.. 2010. Acinetobacter baumannii increases tolerance to antibiotics in response to monovalent cations. Antimicrob. Agents Chemother. 54:1029–1041.
26.
AdamsM.D., NickelG.C, BajaksouzianS., LavenderH., MurthyA.R, JacobsM.R, and BonomoR.A.. 2009. Resistance to colistin in Acinetobacter baumannii associated with mutations in the PmrAB two-component system. Antimicrob. Agents Chemother. 53:3628–3634.
27.
CoyneS., RosenfeldN., LambertT., CourvalinP., and PerichonB.. 2010. Overexpression of resistance-nodulation cell division pump AdeFGH confers multidrug resistance in Acinetobacter baumannii. Antimicrob. Agents Chemother. 54:4389–4393.
28.
LivakK.J., and SchmittgenT.D.. 2001. Analysis of relative gene expression data using real-time quantitative PCR and the 2−ΔΔCT method. Methods, 25:402–408.
29.
NovovicK., MihajlovicS., VasiljevicZ., FilipicB., BegovicJ., and JovcicB.. 2015. Carbapenem-resistant Acinetobacter baumannii from Serbia: Revision of CarO classification. PLoS One, 10:e0122793.
30.
RosenbergM., GutnickD., and RosenbergE.. 1980. Adherence of bacteria to hydrocarbons: a simple method for measuring cell-surface hydrophobicity. FEMS Microbiol. Lett. 9:29–33.
31.
KaliternaV., KaliternaM., HrenovicJ., BarisicI., TonkicM., and Goic-BarisicI.. 2015. Acinetobacter baumannii in the Southern Croatia: clonal lineages, biofilm formation and resistance patterns. Infect. Dis. 47:902–907.
32.
Nait ChabaneY., MartiS., RihoueyC., AlexandreS., HardouinJ., LesouhaitierO., VilaJ., KaplanJ.B, JouenneT., and DéE.. 2014. Characterisation of pellicles formed by Acinetobacter baumannii at the air-liquid interface. PLoS One, 9:e111660.
33.
HigginsP.G., PriorK., HarmsenD., and SeifertH.. 2017. Development and evaluation of a core genome multilocus typing scheme for whole- genome sequence-based typing of Acinetobacter baumannii. PLoS One, 12:e0179228.
34.
MustaphaM.M., LiB., PaceyM.P, MettusR.T, McElhenyC.L, MarshallC.W, ErnstR.K, CooperV.S, and DoiY.. 2018. Phylogenomics of colistin-susceptible and resistant XDR Acinetobacter baumannii. J. Antimicrob. Chemother. 73:2952–2959.
35.
MichalopoulosA., and FalagasM.E.. 2010. Treatment of Acinetobacter infections. Expert Opin. Pharmacother. 11:779–788.
36.
CAMS. 2016. Antibiotic Resistance in Croatia, 2015. Croatian Academy of Medical Sciences,, Zagreb, Croatia.
37.
D'OnofrioV., ConzemiusR., Varda-BrkicD., BogdanM., GrisoldA., GyssensI.C, BedenicB., and BarisicI.. 2020. Epidemiology of colistin-resistant, carbapenemase-producing Enterobacteriaceae and Acinetobacter baumannii in Croatia. Infect. Genet. Evol. 104263.
38.
VerlicchiP.2018. Hospital Wastewaters: Characteristics, Management, Treatment and Environmental Risks. Springer International Publishing,, Cham, Switzerland.
39.
IvankovicT., and HrenovicJ.. 2010. Surfactants in the environment. Arch. Ind. Hyg. Toxicol. 61:95–110.
40.
HrenovicJ., IvankovicT., IvekovicD., RepecS., StipanicevD., and GanjtoM.. 2017. The fate of carbapenem-resistant bacteria in a wastewater treatment plant. Water Res. 126:232–239.
41.
BiswasS., BrunelJ.M, DubusJ.C, Reynaud-GaubertM., and RolainJ.M.. 2012. Colistin: An Update on the Antibiotic of the 21st Century. Expert Rev. Anti. Infect. Ther. 10:917–934.
42.
ZhangC., QiuS., WangY., QiL., HaoR., LiuX., ShiY., HuX., AnD., LiZ., LiP., WangL., CuiJ., WangP., HuangL., KlenaJ.D, and SongH.. 2013. Higher isolation of NDM-1 producing Acinetobacter baumannii. from the sewage of the hospitals in Beijing. PLoS One 8: e6, 4857.
43.
HrenovicJ., Goic-BarisicI., KazazicS., KovacicA., GanjtoM., and TonkicM.. 2016. Carbapenem-resistant isolates of Acinetobacter baumannii in a municipal wastewater treatment plant, Croatia, 2014. Euro Surveill. 21:21–30.
44.
DexterC., MurrayG.L, PaulsenI.T, and PelegA.Y.. 2015. Community-acquired Acinetobacter baumannii: clinical characteristics, epidemiology and pathogenesis. Expert. Rev. Anti. Infect. Ther. 13:567–573.
45.
OikonomouO., SarrouS., PapagiannitsisC.C, GeorgiadouS., MantzarlisK., ZakynthinosE., DalekosG.N, and PetinakiE.. 2015. Rapid dissemination of colistin and carbapenem resistant Acinetobacter baumannii in Central Greece: mechanisms of resistance, molecular identification and epidemiological data. BMC Infect. Dis. 15:559.
46.
NhuN.T.K., RiordanD.W, NhuT.D.H, ThanhD.P, ThwaitesG., LanN.P.H, WrenB.W, BakerS., and StablerR.A.. 2016. The induction and identification of novel Colistin resistance mutations in Acinetobacter baumannii and their implications. Sci. Rep. 6:1–8.
Supplementary Material
Please find the following supplemental material available below.
For Open Access articles published under a Creative Commons License, all supplemental material carries the same license as the article it is associated with.
For non-Open Access articles published, all supplemental material carries a non-exclusive license, and permission requests for re-use of supplemental material or any part of supplemental material shall be sent directly to the copyright owner as specified in the copyright notice associated with the article.
