Rapid Identification of Methicillin-Resistant Staphylococci Bacteremia

Abstract

Staphylococci represent the most commonly encountered blood culture isolates. With the spread of methicillin-resistant Staphylococcus aureus (MRSA) and methicillin-resistant coagulase-negative staphylococci (MRCoNS) in hospitals, rapid and reliable methods for their detection are warranted in order to provide choice of appropriate antimicrobial therapy. This study evaluated 4 rapid methods directly from positive blood cultures in parallel with each other (on the same day) for identification of methicillin-resistant staphylococcal isolates, in addition to antimicrobial susceptibility testing (AST), to compare the workflow for each test and to reduce the turnaround time (TAT) in order to be presented as practical applications in our microbiology laboratory. A total of 56 bacteremic patients' blood cultures with Gram stains showing gram-positive cocci (GPC) in clusters were included. The following direct assays were evaluated: direct tube coagulase (DTC) test, analytical profile index (API)-Staph kit for species identification coupled with antimicrobial susceptibility testing (AST), latex agglutination for detection of PBP2a (PBP2a LA Assay), and cefoxitin disk diffusion assay. The direct results were compared with results obtained with isolated colonies using standard methods as well as detection of the mecA gene by PCR. DTC and API-staph exhibited sensitivities of 96% and 96.8% and specificity of 100% for direct identification of staphylococcal isolates. Both PBP2a LA and cefoxitin DD assays exhibited sensitivity of 100% for detection of both MRSA and MRCoNS and specificities of 100% and 75% (PBP2a assay) and 90% and 100% (cefoxitin DD) for identification of methicillin-sensitive isolates, respectively. For direct antimicrobial susceptibility testing (DAST), the overall error rate was 1.11%. In conclusion, direct identification and susceptibility testing by any of these assays yielded acceptable performance and timely results -- 24 hours earlier than routine subculture -- and can be easily incorporated into routine processing of positive blood cultures to improve the outcomes for the patient and the costs to hospitals. Therefore, it is recommended to use the method with high sensitivity and the shortest TAT.

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Table 1. Characteristics of Patients With Staphylococcal Bacteremia
Characteristics	Staphylococcal Isolates
	No.	%
Sex:
Female	30	53.6
Male	26	46.4
Total	56	100
Age (years) (mean ± SD)^a	51.6 ±11.8
Underlying disease	32	57.1
Surgical intervention	34	60.7
Mechanical ventilation	4	7.1
Previous antibiotic treatment	56	100
Primary bacteremia	37	66.1
Intravascular catheter ^b	37	66.1
Secondary bacteremia	19	33.9
Skin and soft tissue	2	3.6
Lung	4	7.1
Urinary system	9	16.1
Surgical site	4	7.1

^aStudent t-test; ^bHickman-Broviac catheter for hemodialysis

Table 2. Incidence of Staphylococcal Species Isolated From Blood Cultures (N = 56)
Staphylococcal Species	No.	%
S aureus	25	44.6
CoNS species	31	55.4
S haemolyticus	10	32.3
S epidermidis	8	25.8
S hominis	8	25.8
S chromogenes	5	16.1

Table 3. Performance of Two Rapid Assays to Identify Staphylococci Isolates (N = 56) Directly From Blood Cultures
Assay	S aureus isolates (n=25)				CoNs isolates(n=31)
Assay	S (%)	SP (%)	PPV (%)	NPV (%)	S (%)	SP (%)	PPV (%)	NPV (%)
DTC API	96.0 100	100 100	100 100	96.9 100	NA 96.8	NA 100	NA 96.2	NA 100

S = sensitivity; SP = specificity; PPV = positive predictive value; NPV = negative predictive value; NA = not analyzed

Table 4. Performance of Two Rapid Assays to Detect Methicillin-Resistant Staphylococci Directly From Blood Cultures
Assay and result	No. of Isolates Yielding PCR^a Result
	Methicillin-resistant (n = 42)		Methicillin-sensitive (n = 14)		PPV (%)	NPV (%)
	S aureus N=15 N(S%)^b	CoNs N=27 N(S%) ^b	S aureus N=10 N(SP%) ^c	CoNs N=4 N(SP%) ^c	PPV (%)	NPV (%)
PBP -- 2a assay	15(100)	27(100)	10(100)	3(75)	97.7	100
Cefoxitin DD	15(100)	27(100)	9(90)	4(100)	97.7	100

^aDefinitive method; ^bsensitivity; ^cspecificity; PPV = positive predictive value; NPV = negative predictive value

Table 5. Antimicrobial Susceptibility Correlation for Methicillin-Resistant Staphylococcal Isolates^a
	MRSA (n = 15)			MRCoNS (n = 27)
Antimicrobial agent	No. very major^b error	No. major^cerror	No. minor^d error	No. very major^b error	No. major^cerror	No. minor^d error
Penicillin	0	0	0	0	0	1
Erythromycin	0	0	0	0	0	0
Amikacin	0	0	0	0	0	2
Cefoperazone	0	0	0	0	0	1
Ciprofloxacin	0	0	0	0	0	0
TMP/SMX	0	0	0	0	0	0
Vancomycin	0	0	0	0	0	0
Teicoplanin	0	0	0	0	1	3
Tetracycline	0	0	0	0	0	0
Clindamycin	0	0	0	0	2	0
Tobramycin	0	0	0	0	0	0
Chloramphenicol	2	0	0	0	0	0
Total (%)	2 (0.01)	0 (0.0)	0 (0.0)	0 (0.0)	3 (0.9)	7 (0.2)

^aAST was used for both direct and subculture isolates; TMP/SMX = trimethoprim-sulfamethoxazole; ^bFalse susceptibility; ^cFalse resistance; ^dIntermediate vs sensitive or resistant result

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Rapid Identification of Methicillin-Resistant Staphylococci Bacteremia Among Intensive Care Unit Patients

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