Controlled Clinical Comparison of Plastic and Glass Bottles of BacT/ALERT FA Medium for Culturing Organisms from Blood of Adult Patients

Abstract

A new, clear-plastic nonvented aerobic FA bottle, designed to prevent breakage, has been developed for the BacT/ALERT blood culture system. We assessed the new plastic FA bottle by comparing its performance with that of the current glass FA bottle for recovery of microorganisms and time to detection of growth in blood samples obtained for culture from adult patients with suspected bloodstream infections. We conclude that the BacT/ALERT plastic and glass FA bottles are comparable for recovery of microorganisms and that the safety advantage of plastic bottles can be achieved without compromising performance.

The recovery of microorganisms from blood culture in order to diagnose bacteremia and fungemia remains one of the most important tests performed by a clinical microbiology laboratory. Prompt detection of microorganisms in blood culture bottles provides valuable information for clinicians in guiding therapeutic decisions. Glass bottles have been used routinely in blood culture systems; however, breakage of glass bottles, although a rare event, may compromise the integrity of a blood specimen, delay reporting, and expose health care workers to infectious blood-borne pathogens. To reduce these risks, clear plastic bottles have been developed for use in the BacT/ALERT (bioMérieux, Inc., Durham, N.C.) automated blood culture instrument. The new plastic FA (PFA) bottle contains a casein-soy-based medium with activated charcoal to counter the potential inhibitory activity of antibiotics, as does the glass BacT/ALERT FA bottle. We compared the new PFA bottle to the current FA bottle for the recovery of microorganisms, as well as for the time to detection of the growth of microorganisms in blood samples obtained for culture from adult patients with suspected bloodstream infections.

(This work was presented at the 12th European Congress of Clinical Microbiology and infectious Diseases, Milan, Italy, 25 April 2002 [abstract P713].)

From July 2000 to February 2002, blood was collected from adult patients at Duke University Medical Center with suspected bacteremia and fungemia. Institutional review board approval was obtained prior to the study, and all blood cultures were performed as part of standard patient care. Up to 30 ml of blood was collected from each adult patient and divided (10 ml each) among aerobic PFA and FA bottles and anaerobic FN bottles. Although not part of this comparative study, the FN bottle was included to enable recovery of anaerobic microorganisms. Upon receipt in the laboratory, each bottle was measured against known standards to determine the volume of blood contained therein. All bottles were processed regardless of volume received: however, only bottle pairs (one PFA and one FA bottle) containing 8 to 12 ml of blood per bottle were included in the data analysis. All bottles were incubated in the BacT/ALERT instrument for 5 days or until they signaled positive. Based on Gram stain results, aliquots of the blood-broth mixture were subcultured onto appropriate media and incubated per laboratory protocol. Subsequent identification of microorganisms was performed by standard laboratory methods (2). False positives were defined as bottles that were Gram stain and subculture negative after the instrument signaled positive. Gram stain-negative bottles were returned to the instrument for the remainder of the 5-day incubation period or until they were reflagged by the instrument. Specifically for this study, for bottle pairs with a negative bottle and a companion positive bottle, the contents of the negative bottle were subcultured at the end of the 5-day protocol. If microorganisms grew on subculture, these negative bottles were defined as false negative.

An infectious-disease physician reviewed each positive culture and coded it as clinically significant, a contaminant, or an isolate of unknown clinical significance based on previously published criteria (4). Episodes of bloodstream infection were defined by growth of a clinically significant isolate without recovery of a different microorganism during the succeeding 7-day period. If a different clinically significant microorganism was recovered within 72 h, the two isolates were considered evidence of a polymicrobial episode (not included in the analysis). If a different microorganism was recovered after 72 h, the second isolate was considered evidence of a new episode. Patients were considered to be on therapy at the time the blood was collected if they were receiving an agent with activity against the microorganism isolated. Testing for susceptibility of the blood isolate to antimicrobial agents was determined by NCCLS methods (3). In some cases (e.g., Candida spp.), resistance was inferred from known patterns of susceptibility. Statistical analysis of results was performed with the modified chi-square test described by McNemar (1).

We received a total of 5,862 blood culture sets that contained both PFA and FA bottles, 4,323 (74%) of which contained an adequate volume of blood. Three hundred twenty-two clinically significant isolates from 162 patients were detected in one or both study bottles (Table 1). Clinically significant isolates were detected with equal frequencies in both study bottles. The results of a subset of blood cultures from patients who were being treated with antimicrobial agents at the time the blood cultures were obtained are summarized in Table 2. There were no significant differences in recovery for any microorganism in this subgroup. In the subgroup of patients not receiving antimicrobial therapy (data not shown), Escherichia coli was detected more often in the FA (n = 8) than in the PFA (n = 1) bottles (P < 0.05); however, this difference was not present in the monomicrobial episode analysis (Table 3), which included 130 of 153 total episodes. When both bottles detected clinically significant microorganisms within 72 h the mean times to detection were similar (Table 4). Of the 4,323 paired blood culture bottles, the frequencies of false-positive bottles were similar for PFA (n = 10 [0.2%]) and FA (n = 11 [0.3%]) bottles. There was one false-negative result (Stenotrophomonas maltophilia) in a PFA bottle and one (Pseudomonas aeruginosa) in an FA bottle. Of all 185 contaminant isolates detected during this study (detailed data not shown), 53 were detected in both bottles, 52 were detected in PFA bottles only; and 80 were detected in FA bottles only (P < 0.05). Of the 131 coagulase-negative staphylococcal contaminants specifically, 38 were detected in both bottles, 36 were detected in PFA bottles only, and 57 were detected in FA bottles only (P < 0.05).

TABLE 1.

Comparative yields of clinically significant isolates in PFA versus FA aerobic blood culture bottles

Microorganism(s)	No. of isolates recovered from:			Pa
	Both bottles	PFA bottles only	FA bottles only
Staphylococcus aureus	50	9	8	NS
Coagulase-negative staphylococci	22	4	2	NS
Streptococcus sp.b	1	1	0	NS
Enterococcus sp.	12	6	6	NS
Gram-positive bacillic	6	0	1	NS
Enterobacteriaceaed	52	9	18	NS
Other gram-negative bacteriae	40	7	6	NS
Yeastsf	32	17	13	NS
All microorganisms	215	53	54	NS

^aNS, not significant.

^bIncludes viridans group streptococci (one isolate) and Streptococcus agalactiae (one isolate).

^cIncludes Corynebacterium jeikeium (four isolates), Listeria monocytogenes (two isolates), and diphtheroids (one isolate).

^dIncludes E. coli (26 isolates), Enterobacter cloacae (14 isolates), Klebsiella pneumoniae (13 isolates), Serratia marcescens (7 isolates), Enterobacter aerogenes (7 isolates), Klebsiella oxytoca (7 isolates), Citrobacter freundii (3 isolates), and Proteus mirabilis (2 isolates).

^eIncludes P. aeruginosa (16 isolates), Stenotrophomonas maltophilia (13 isolates), Burkholderia cepacia (9 isolates), Acinetobacter sp. (4 isolates), Chryseobacterium sp. (4 isolates), Pandoraea spp. (3 isolates), Neisseria sp. (1 isolate), Ochrobactrum anthropi (1 isolate), and unidentified gram-negative rods (2 isolates).

^fIncludes Candida albicans (27 isolates), Candida glabrata (11 isolates), Candida krusei (9 isolates), Candida parapsilosis (7 isolates), Candida lusitaniae (3 isolates), Candida tropicalis (2 isolates), and Cryptococcus neoformans (3 isolates).

TABLE 2.

Comparative yields of clinically significant isolates in PFA versus FA aerobic blood culture bottles from patients receiving antimicrobial therapy

Microorganism(s)	No. of isolates recovered from:			Pa
	Both bottles	PFA bottles only	FA bottles only
Staphylococcus aureus	14	3	3	NS
Coagulase-negative staphylococci	9	2	1	NS
Enterococcus sp.	5	0	1	NS
Corynebacterium jekeium	1	0	1	NS
Enterobacteriaceaeb	11	3	4	NS
Other gram-negative bacteriac	9	1	3	NS
Yeastsd	16	7	5	NS
All microorganisms	65	16	18	NS

^aNS, not significant.

^bIncludes E. coli (eight isolates), K. pneumoniae (four isolates), Enterobacter aerogenes (three isolates), Enterobacter cloacae (two isolates), and Proteus mirabilis (one isolate).

^cIncludes P. aeruginosa (four isolates), Stenotrophomonas maltophilia (three isolates), B. cepacia (three isolates), Acinetobacter sp. (one isolate), O. anthropi (one isolate), and an unidentified gram-negative rod (one isolate).

^dIncludes Candida albicans (10 isolates), Candida krusei (9 isolates), Candida parapsilosis (6 isolates), Cryptococcus neoformans (2 isolates), and Candida lusitaniae (1 isolate).

TABLE 3.

Comparative levels of detection of episodes of monomicrobial bacteremia or fungemia by PFA and FA aerobic culture bottles paired with an anaerobic FN bottle

Cause of episode	No. of episodes detected by:			Pa
	Both bottlesh	PFA bottles only	FA bottles only
Staphylococcus aureus	37	3	1	NS
Coagulase-negative staphylococci	14	0	0	NS
Streptococcus sp.b	1	1	0	NS
Enterococcus sp.	6	1	2	NS
Gram-positive bacillic	4	0	0	NS
Enterobacteriaceaed	36	3	5	NS
Other gram-negative bacteriae	20	0	2	NS
Yeastsf	10	5	1	NS
Anaerobesg	2	0	0	NS
All microorganisms	130	13	11	NS

^aNS, not significant.

^bIncludes viridans group streptococci (one isolate) and Streptococcus agalactiae (one isolate).

^cIncludes Listeria monocytogenes (two isolates), Cornyebacterium jeikeium (one isolate), and diphtheroids (one isolate).

^dIncludes E. coli (20 isolates),K. pneumoniae (9 isolates),Enterobacter cloacae (5 isolates), Enterobacter aerogenes (3 isolates), Serratia marcescens (3 isolates), K. oxytoca (2 isolates), Proteus mirabilis (1 isolate), and Pantoea agglomerans (1 isolate).

^eIncludes P. aeruginosa (six isolates), B. cepacia (five isolates), Stenotrophomonas maltophilia (four isolates), Acinetobacter sp. (one isolate), Chryseobacterium sp. (two isolates), Pandoraea sp. (one isolate), Neisseria sp. (one isolate), O. anthropi (one isolate), and an unidentified gram-negative rod (one isolate).

^fIncludes Candida albicans (eight isolates), Candida glabrata (four isolates), Candida krusei (one isolate), Candida parapsilosis (one isolate), Candida tropicalis (one isolate), and Cryptococcus neoformans (one isolate).

^gIncludes Clostridium perfringens (one isolate) and Bacteroides fragilis (one isolate).

^hWhen the anaerobic FN bottle was positive, the episode was considered to be positive in both systems.

TABLE 4.

Comparative times to positivity of clinically important bacteria and fungi when both the aerobic PFA and FA culture bottles were positive within 72 h

Microorganism(s)	No. of isolates	Mean time to positivity (h)
		PFA bottles	FA bottles
Staphylococcus aureus	50	17.3	17.7
Coagulase-negative staphylococci	21	24.4	23.0
Viridans group streptococcus	1	46.2	50.4
Enterococcus sp.	12	14.4	14.3
Gram-positive bacillia	6	26.1	27.5
Enterobacteriaceaeb	52	11.9	12.3
Other gram-negative bacteriac	35	20.2	19.9
Yeastsd	27	38.3	41.2
All microorganisms	204	20.2	20.6

^aIncludes Comyebacterium jeikeium (three isolates), L. monocytogenes (two isolates), and diphtheroids (one isolate).

^bIncludes E. coli (13 isolates), Enterobacter cloacae (13 isolates), K. pneumoniae (8 isolates), K. oxytoca (7 isolates), Serratia marcescens (6 isolates), Citrobacter freundii (2 isolates), Enterobacter aerogenes (1 isolate), Proteus mirabilis (1 isolate), and Pantoea agglomerans (1 isolate).

^cIncludes P. aeruginosa (12 isolates), Stenotrophomonas maltophilia (8 isolates), B. cepacia (7 isolates),Acinetobacter sp. (3 isolates), Chryseobacterium sp. (3 isolates), Neisseria sp. (1 isolate), and O. anthropi (1 isolate).

^dIncludes Candida albicans (14 isolates), Candida krusei (6 isolates), Candida parapsilosis (4 isolates), Candida lusitaniae (2 isolates), and Candida tropicalis (1 isolate).

The overall performance of the aerobic plastic (PFA) bottle was similar to that of the glass (FA) bottle for detection of clinically significant microorganisms from patients with suspected bacteremia or fungemia. Fewer contaminants were found in the PFA bottles. When both bottles were positive within 72 h, the times to detection were also similar. Because plastic bottles provide a performance comparable to that of glass bottles and have the potential for added safety, we recommend their use in the BacT/ALERT blood culture system.