Prevalence and Some Possible Mechanisms of Colistin Resistance Among Multidrug-Resistant and Extensively Drug-Resistant Pseudomonas aeruginosa

Figures & data

Figure 1 Antibiotic resistance pattern of all isolated P. aeruginosa isolates.

Figure 2 Phenotypic detection for mcr positive isolates by combined disc diffusion test (CDT). (A): mcr-1 positive strain showed an increase in the zone diameter of discs with colistin and EDTA ≥ 3mm in comparison to colistin alone. (B): mcr-1 negative isolate showed slight change (1 mm) in the inhibition zone diameter of colistin and EDTA disc in comparison to colistin alone.

Figure 3 Antibiotic resistance pattern of colistin-resistant isolates.

Table 1 Colistin-Resistant Isolates, Some Possible Mechanisms of Resistance to Colistin and Their Susceptibility to Other Antibiotics

Isolates*	Colistin MIC µg/mL	mcr1/mcr-2	CDT^a	RZP^b	MIC of Colistin in Presence of 0.5 MIC CCCP (µg/mL) (Fold Change)^c	MDR, XDR or PDR	Antibiotics Showing Activity on the Tested Isolates
P1	8	+/−	+	1.07	8 (no change)	MDR	MEM, TGC, CIP, AK, CAR, PRL
P2	16	+/−	+	0.8	8 (2 folds)	MDR	MEM, TGC, CAR, CN, PRL
P3	256	−/−	-	0.7	128 (2 folds)	XDR	MEM, IMP, AK
P4	256	+/−	+	0.57	128 (2 folds)	MDR	TGC, CIP, MEM, PRL, AK
P5	256	+/−	+	1.08	256 (no change)	MDR	MEM, TGC, CAR, CN, PRL, CIP
P6	128	−/−	-	1.035	16 (8 folds)	MDR	MEM, TGC, CAR, AK, PRL
P7	128	−/−	-	1.28	64 (2 fold)	MDR	MEM, TGC, CAR, CN, PRL, ATM
P8	256	−/−	-	0.99	16 (16 folds)	XDR	MEM, TGC
P9	256	+/−	+	1.07	64 (4 folds)	MDR	TGC, CIP, MEM, PRL, AK
P10	32	−/−	-	1.08	16 (2 folds)	MDR	TGC, CIP, MEM
P11	128	−/−	-	1.055	64 (2 folds)	MDR	TGC, CIP, MEM, IMP, AK, CN
P12	256	−/−	-	1.48	256 (no change)	MDR	TGC, CIP, AK, LEV
P13	64	+/−	+	2.7	64 (no change)	MDR	TGC, CIP, MEM, PRL, AK
P14	256	−/−	-	0.935	128 (2 folds)	MDR	TGC, CIP, MEM, PRL, AK
P15	128	+/−	-	1.57	32 (4 folds)	XDR	MEM, PB
P16	64	+/−	-	2.9	8 (8 folds)	MDR	TGC, CIP, MEM, PRL, AK, CN

Notes: *All isolates were positive for toxA gene; ^aCombined disc diffusion test, ^bZeta potential Ratio= ZP_+EDTA/ZP_−EDTA, ^cstrains were considered as positive for efflux if fold change ≥ 8 folds. MDR: multi-drug resistant: nonsusceptible to ≥ 1 agent in ≥ 3 antimicrobial categories. XDR: nonsusceptible to ≥ 1 agent in all but ≤ 2 antimicrobial categories.

Abbreviations: CN, Gentamicin; AK, Amikacin; PB, ploymxin B; MEM, meropenem; TGC, Tigecycline; CEP, cefeperazone; CIP, ciprofloxacin; IPM, imipenem; PRL, piperacillin; LEV, levofloxacin; CAR, Carbenicillin.

Table 2 Molecular Weights and Amount % of Extracted Outer Membrane Proteins of Colistin Resistant and Colistin Sensitive P. Aeruginosa

Lanes:	M		Colistin Resistant Isolates				Colistin Sensitive Isolates
			R1 (P1)		R2 (P12)		S1		S2		S3
Rows	Mol.w.	Amount	Mol.w.	Amount	Mol.w.	Amount	Mol.w.	Amount	Mol.w.	Amount	Mol.w.	Amount
r1	170	5.326371
r2	125	9.451697
r3	81	10.44386	105	27.13755			105.75	7.895928	105.5	61.66282	105.55	37.32484
r4	62	9.112272	66.75	3.475836	66.071	3.128621	66.071	5.723982	66.07	3.903002	66.714	41.65605
r5			54.607	5.390335	54.686	36.3847	54.643	11.04072	54.693	6.30485	56.857	5.070064
r6	53	7.650131	47.874	3.289963	47.848	10.39397	47.839	11.04072	47.806	10.46189	47.774	9.312102
r7	43	8.381201	40.186	5.130112	40.174	10.16686	40.195	14.47964	40.163	7.459584	40.186	2.267516
r8	32	11.69713							31.364	5.034642
r9	25	10.86162	23.684	7.843866	23.673	6.31924	23.177	5.904977	23.684	5.17321	23.886	4.280255
r10			21.456	6.003717	21.21	7.31205
r11	17	11.93211
r12	14	15.22193					14.482	43.8914

Figure 4 Outer membrane SDS-PAGE of colistin resistant and sensitive strains. Lane 1: Protein Marker, Lane 2 and Lane 3: colistin-resistant strains (P1 & P12), Lanes 4–6: colistin sensitive strains.

Figure 5 LPS bands pattern. Lanes 1, 2 & 3: colistin-resistant mcr-1 negative strains (P3, P6 & P10, respectively), Lane 4: Colistin sensitive strains and Lane 5: Colistin-resistant mcr-1 positive strain (P1). O-antigen repeats are boxed and arrow refers to LPS core.