Molecular epidemiology of coagulase-negative bloodstream isolates: detection of <i>Staphylococcus epidermidis</i> ST2, ST7 and linezolid-resistant ST23

Martínez-Meléndez, Adrián; Morfín-Otero, Rayo; Villarreal-Treviño, Licet; Camacho-Ortíz, Adrián; González-González, Gloria; Llaca-Díaz, Jorge; Rodríguez-Noriega, Eduardo; Garza-González, Elvira

doi:10.1016/j.bjid.2016.05.007

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Original Articles • Braz J Infect Dis 20 (5) • Sep-Oct 2016 • https://doi.org/10.1016/j.bjid.2016.05.007 copy

Molecular epidemiology of coagulase-negative bloodstream isolates: detection of Staphylococcus epidermidis ST2, ST7 and linezolid-resistant ST23

Authorship SCIMAGO INSTITUTIONS RANKINGS

Abstract

The mechanisms contributing to persistence of coagulase-negative staphylococci are diverse; to better understanding of their dynamics, the characterization of nosocomial isolates is needed. Our aim was to characterize phenotypic and molecular characteristics of Staphylococcus epidermidis and Staphylococcus haemolyticus human blood isolates from two tertiary care hospitals in Mexico, the Hospital Universitario in Monterrey and the Hospital Civil in Guadalajara.

Antimicrobial susceptibility was determined. Biofilm formation was assessed by crystal violet staining. Detection of the ica operon and Staphylococcal Cassette Chromosome mec typing were performed by PCR. Clonal relatedness was determined by Pulsed-fiel gel electrophoresis and Multi locus sequence typing.

Methicillin-resistance was 85.5% and 93.2% for S. epidermidis and S. haemolyticus, respectively. Both species showed resistance >70% to norfloxacin, clindamycin, levofloxacin, trimethoprim/sulfamethoxazole, and erythromycin. Three S. epidermidis and two S. haemolyticus isolates were linezolid-resistant (one isolate of each species was cfr+). Most isolates of both species were strong biofilm producers (92.8% of S. epidermidis and 72.9% of S. haemolyticus). The ica operon was amplified in 36 (43.4%) S. epidermidis isolates. SCCmec type IV was found in 47.2% of the S. epidermidis isolates and SCCmec type V in 14.5% of S. haemolyticus isolates. No clonal relatedness was found in either species. Resistance to clindamycin, levofloxacin, erythromycin, oxacillin, and cefoxitin was associated with biofilm production for both species (p < 0.05). A G2576T mutation in 23S rRNA gene was detected in an S. haemolyticus linezolid-resistant isolate. All linezolid-resistant S. epidermidis isolates belonged to ST23; isolate with SCCmec type IV belonged to ST7, and isolate with SCCmec type III belonged to ST2. This is the first report of ST7 in Mexico.

There was a high genetic diversity in both species, though both species shared characteristics that may contibute to virulence.

Keywords
Linezolid resistance; Staphylococcal Cassette; Chromosome mec; Coagulase-negative staphylococci; Multilocus Sequence Typing

Antimicrobial	S. epidermidis				S. haemolyticus
	MIC₅₀^a a MIC50, minimal inhibitory concentration for 50% of the isolates. (μg/mL)	MIC₉₀^b b MIC90, minimum inhibitory concentration for 90% of the isolates. (μg/mL)	Resistantn (%)	Susceptiblen (%)	MIC₅₀^a a MIC50, minimal inhibitory concentration for 50% of the isolates. (μg/mL)	MIC₉₀^b b MIC90, minimum inhibitory concentration for 90% of the isolates. (μg/mL)	Resistantn (%)	Susceptiblen (%)
Oxacillin	>0.5	>0.5	72 (86.7)	11 (13.3)	>0.5	>0.5	55 (93.2)	4 (6.8)
Vancomycin	2	4	0 (0)	83 (100)	2	2	0 (0)	59 (100)
Erythromycin	>4	>4	66 (79.5)	16 (19.3)	>4	>4	53 (89.8)	6 (10.2)
Tetracycline	≤2	>8	10 (12)	71 (85.6)	≤2	>8	13 (22.0)	44 (74.6)
Levofloxacin	>2	>2	62 (74.7)	18 (21.7)	>2	>2	54 (91.5)	4 (6.8)
Norfloxacin	>8	>8	64 (77.1)	18 (21.7)	>8	>8	53 (89.8)	6 (10.2)
Clindamycin	>2	>2	68 (81.9)	12 (14.5)	>2	>2	53 (89.8)	5 (8.5)
Trimethoprim/sulfamethoxazole	>2/38	>2/38	60 (72.3)	23 (27.7)	>2/38	>2/38	45 (76.3)	14 (23.7)
Linezolid	≤2	4	3 (3.6)	80 (96.4)	≤2	≤2	2 (3.4)	57 (96.6)

Isolate	Species	SCCmec typing			Susceptibility profile^a a All isolates were resistant to norfloxacin, clindamycin, levofloxacin, trimethoprim/sulfamethoxazole, and erythromycin. All isolates were susceptible to vancomycin.			Linezolid resistance^b b Mutation G2576T was found in isolate 2975.		Biofilm production
		mecA	mec class	ccr type	FOX	OX	TET	MIC (μg/mL)	cfr	Glucose	NaCl	ica operon
14583	S. epidermidis	+	A	3 + 5	R	R	S	8	Negative	Strong	Strong	Negative
14565	S. epidermidis	+	A	3 + 5	R	R	I	>32	Positive	Strong	Strong	Positive
12701	S. epidermidis	+	A	3	R	R	S	8	Negative	Strong	Strong	Positive
9976	S. haemolyticus	−	NA	NA	S	S	S	8	Positive	NP	NP	Negative
2975	S. haemolyticus	+	NT	NT	R	R	R	32	Negative	Strong	NP	Negative

No. of isolates	SCCmec typing			Resistance profile^a a All isolates were resistant to cefoxitin and oxacillin. All isolates were susceptible to vancomycin. n (%)							Biofilm production^b b Strong producers. n (%)
	mec class	ccr type	SCCmec	NOR	CLI	TET	LEV	LZD	SXT	ERY	Glucose	NaCl
34	B	2	IV	30 (88)	30 (88)	3 (8.8)	30 (88)	0 (0)	25 (74)	30 (88)	33 (97)	30 (88)
5	A	3	III	5 (100)	4 (80)	0 (0)	5 (100)	1 (20)	4 (80)	4 (80)	5 (100)	4 (80)
3	A	1	New	3 (100)	3 (100)	2 (67)	3 (100)	0 (0)	2 (67)	3 (100)	3 (100)	2 (67)
1	A	5	New	1 (100)	1 (100)	0 (0)	1 (100)	0 (0)	1 (100)	1 (100)	1 (100)	1 (100)
1	A	1 + 2	Variant	0 (0)	1 (100)	0 (0)	0 (0)	0 (0)	0 (0)	1 (100)	1 (100)	0 (0)
5	B	1 + 2	Variant	5 (100)	5 (100)	1 (20)	5 (100)	0 (0)	4 (80)	5 (100)	5 (100)	4 (80)
1	A	2 + 3	Variant	1 (100)	1 (100)	0 (0)	1 (100)	0 (0)	1 (100)	1 (100)	1 (100)	1 (100)
7	A	3 + 5	Variant	7 (100)	7 (100)	1 (14)	7 (100)	2 (29)	7 (100)	7 (100)	7 (100)	7 (100)
5	B	2 + 5	Variant	3 (60)	5 (100)	1 (20)	2 (40)	0 (0)	3 (60)	4 (80)	5 (100)	5 (100)
2	A	2 + 4	Variant	2 (100)	2 (100)	0 (0)	2 (100)	0 (0)	2 (100)	2 (100)	2 (100)	2 (100)
1	A	3 + 4	Variant	1 (100)	1 (100)	0 (0)	1 (100)	0 (0)	1 (100)	1 (100)	0 (0)	1 (100)
1	B	2 + 4	Variant	1 (100)	1 (100)	0 (0)	1 (100)	0 (0)	1 (100)	1 (100)	1 (100)	1 (100)
2	A	1 + 4 + 5	Variant	0 (0)	2 (100)	0 (0)	0 (0)	0 (0)	2 (100)	2 (100)	2 (100)	1 (50)
1	NT	2 + 5	NT	1 (100)	0 (0)	0 (0)	1 (100)	0 (0)	1 (100)	0 (0)	1 (100)	1 (100)
1	NT	2	NT	1 (100)	1 (100)	0 (0)	1 (100)	0 (0)	1 (100)	1 (100)	1 (100)	1 (100)
2	A	NT	NT	2 (100)	2 (100)	1 (50)	1 (50)	0 (0)	2 (100)	2 (100)	2 (100)	2 (100)

No. of isolates	SCCmec typing			Resistance profile^a a All isolates were resistant to cefoxitin, oxacillin, norfloxacin, levofloxacin and trimethoprim/sulfamethoxazole. All isolates were susceptible to vancomycin.				Biofilm production^b b Strong producers.
	mec class	ccr type	SCCmec	CLI	TET	LZD	ERY	Glucose	NaCl
8	C2	5	V	7 (88)	3 (38)	0 (0)	7 (88)	7 (88)	4 (50)
1	C2	1 + 2 + 5	Variant	0 (0)	0 (0)	0 (0)	0 (0)	1 (100)	0 (0)
3	C2	1 + 4 + 5	Variant	3 (100)	3 (100)	0 (0)	3 (100)	3 (100)	2 (67)
12	NT	4	NT	10 (83)	3 (25)	0 (0)	11 (92)	8 (67)	5 (42)
1	NT	5	NT	1 (100)	1 (100)	0 (0)	1 (100)	1 (100)	0 (0)
30	NT	NT	NT	30 (100)	3 (10)	1 (3)	30 (100)	23 (77)	18 (60)

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[1] Corresponding author. elvira_garza_gzz@yahoo.com