Brucella detection in blood: comparison of the BacT/Alert standard aerobic bottle, BacT/Alert FAN aerobic bottle and BacT/Alert enhanced FAN aerobic bottle in simulated blood culture

The objective of this study was to compare the performances of the standard aerobic bottle (StAe), FAN aerobic (FANAe) and enhanced FAN aerobic (E-FANAe) (the charcoal component of the FANAe was revised recently to improve the feasibility of Gram smear interpretation) blood culture bottles for BacT/Alert system for the detection of Brucella melitensis in simulated blood culture. Triplicate strains of eight clinical isolates of B. melitensis were studied. Each bottle was inoculated with 5 mL of freshly collected human blood at three different targeted bacterial inocula (10¹, 10² and 10³ CFU/bottle). All bottles were monitored for up to 21 days or until they became positive. The results of time to detection (TTD) on the eight B. melitensis samples were as follows: at 10¹ CFU/bottle, the E-FANAe had a mean TTD significantly shorter than the StAe (48 h vs. 56.2 h, P  < 0.05); and at 10³ CFU/bottle, the FANAe and E-FANAe had a mean TTD significantly shorter than the StAe (41.2 h and 40 h vs. 45.6 h, P  < 0.05). The reproducibilities (no.of positive signals/no.of all bottles) of three bottle systems were as follows: at 10¹ CFU/bottle, the reproducibilities of StAe, FANAe and E-FANAe were 96, 83 and 58%, respectively. At 10³ CFU/bottle, the reproducibilities of StAe, FANAe and E-FANAe were 95, 95 and 91%, respectively. Positive results for the presence of bacteria in Gram smears were confirmed in 68% of StAe, 54% of FANAe and 90% of E-FANAe. In case of suspected brucellosis, the combination of one StAe bottle and one E-FANAe bottle seems to provide the highest and fastest recovery of the organism.     

Evaluation of the BacT/Alert and VITAL blood culture systems for the diagnosis of bacteremia

Objective: To evaluate the detection of bacterial growth in the BacT/Alert (Organon Teknika) and VITAL (bioMérieux) automated blood culture systems.
Methods: In accordance with the protocol of study, 1021 blood sample pairs for culture were obtained from adult patients admitted to the Emergency Room and Intensive Care Unit.
Results: In total, 139 (13.6%) clinically significant blood cultures were detected, of which 79 (56.8%) were detected by both systems, 48 (34.5%) only by BacT/Alert and 12 (8.6%) only by VITAL (P <0.0001). The BacT/Alert system detected positive blood cultures more rapidly for all groups of microorganisms. The VITAL system showed six false-negative blood cultures, while the BacT/Alert system showed none ( P =0.03). There was no significant difference between the number of false-positive blood cultures detected by the two systems.
Conclusions: In our study, overall the BacT/Alert system achieved a better recovery of microorganisms than the VITAL system.     

Evaluation of the VITAL (bioMérieux) automated blood culture system using blind subculture

Objective   This study was performed to determine the ability of the VITAL system to detect and allow recovery of microorganisms that are difficult to grow, such as Brucella spp., yeasts, or anaerobes, as well as to determine the need for blind subcultures after the incubation period.
Methods   A prospective evaluation of the system was performed, and 8247 blood culture bottles were processed. The standard was blind subculture from all the bottles after 5 days of incubation.
Results   There were 3.2% false-positive and 0.6% false-negative results (72% of clinical importance). The system sensitivity for yeasts was 41%. The mean time for detection of Neisseria meningitidis was 31.9 ± 2.8 h, for Brucella spp. 119.7 ± 2 h, and for yeast 51.5 ± 27.8 h.
Conclusions   The VITAL system poses has serious difficulties in the detection of N. meningitidis , Brucella spp., yeast and methicillin- and aminoglycoside-resistant Staphylococcus aureus (MARSA). The low system sensitivity for yeast detection makes the blind subculture necessary after the incubation period.     

Detection of Campylobacter upsaliensis from a blood culture by using the BacT/Alert system.

Campylobacter upsaliensis was isolated from the blood of a 60-year-old female with hairy cell leukemia. This spiral-shaped organism was detected in the aerobic BacT/Alert bottle (Organon Teknika, Durham, N.C.) by acridine orange staining and was recovered only on chocolate agar in a microaerophilic atmosphere at 35 degrees C.     

Potential use of BacT/Alert automated blood culture system for antifungal susceptibility testing.

A simple, rapid susceptibility test is needed to determine the possible resistance of yeasts to commonly used antifungal agents. The BacT/Alert automated blood culture system was evaluated as one technology for development of such a test. Yeast nitrogen base was used as the growth medium, and amphotericin B was the test antifungal agent. Isolates of various Candida species, Torulopsis glabrata, Saccharomyces cerevisiae, and Cryptococcus neoformans were evaluated. The results suggest that detection of amphotericin B resistance of yeast isolates within 12 to 14 h after inoculation of test medium is possible.     

Comparison of BacT/Alert and BACTEC NR 860 blood culture systems in a laboratory not continuously staffed

Objective: To evaluate the performance of continuously working blood culture systems in a discontinuous laboratory system.
Method: The systems used were BacT/Alert (Organon Teknika Corp., Durham, NC) and BACTEC NR 860 (Becton Dickinson Diagnostic Instruments, Sparks, Md) in a comparison in a laboratory staffed 8 1/2 h on Mondays to Fridays and 4 1/2 h on Saturdays. Blood culture bottles (BacT/Alert aerobic and anaerobic, BACTEC NR 26 A and NR 27 A) were received thrice daily.
Results: From 1824 pairs of blood culture vials, 110 clinically significant microorganisms were recovered by both BACTEC and BacT/Alert, 43 by BACTEC alone, and 33 by BacT/Alert alone. The differences between the systems in total recovery and in recovery of individual species were not statistically significant. The average detection times were 13.36 h for BACTEC and 13.93 h for BacT/Alert ( P >0.1). These times represent only 35.6% (BACTEC) and 32.6% (BacT/Alert) of the total timespans from collection of blood to informing the ward of a positive result ( t _crd, clinically relevant detection time). If 24 h per day blood culture processing conditions and continuous transport of vials to the laboratory had been available, these percentages would have risen to 87% (BACTEC) and 87.5% (BacT/Alert). Under such 'ideal' conditions, t _trd could have been reduced by 22.16 h using BACTEC and by 26.81 h using BacT/Alert. The BacT/Alert system showed more false-positive results than the BACTEC system (80 (4.39%) versus 23 (1.26%), P <0.001).
Conclusions: No time benefit for detection of positive blood cultures is gained with continuously measuring systems, if loading and processing of vials is organized discontinuously, as in our laboratory.     

Detection of bacteraemia by the continuously monitoring BacT/Alert system.

AIMS--To analyse a continuously monitoring blood culture system with respect to the time to detection of various groups of organisms, their clinical importance, and the relative efficacy of the aerobic and anaerobic bottles. METHODS--Four thousand blood cultures were monitored and the information relating to the positive cultures was noted and analysed. RESULTS--Four hundred and seventy seven blood cultures were detected as positive, 81% (387/477) of which were detected within 48 hours. The most pathogenic organisms were detected in the shortest period, less pathogenic later and those generally regarded as contaminants last. Clinically important isolates were also detected earlier. Many positive blood cultures were detected in only one bottle of the set, even those regarded as clinically important. CONCLUSIONS--The management of continuously monitoring blood culture systems could be improved by considering time to detection trends. Clinicians should be aware of the relatively rapid detection of clinically important, positive blood cultures in relation to patient treatment.     

A comparison of two blood culture procedures for the isolation of staphylococci in a paediatric intensive care unit

Blood culture results obtained between January 2000 and July 2003 were reviewed for 1360 patients in a paediatric intensive care unit (PICU). The BacT/Alert FA aerobic medium was used with a blood volume of 1.5 mL for the first 23 months, and the BacT/Alert PF paediatric medium was used with a 0.5-mL volume for the remaining 18 months. The isolation rates were similar during both periods (13.4% vs. 13.1%), and staphylococci were the most common isolates (72.8%). There was a shorter time to detection of staphylococci with the smaller-volume (PF) procedure, which thus seems suitable for use in the diagnosis of staphylococcal bacteraemia in the PICU.     

Performance of VITEK-2 Compact and overnight MicroScan panels for direct identification and susceptibility testing of Gram-negative bacilli from positive FAN BacT/ALERT blood culture bottles

We describe the reliability of the VITEK-2 Compact and overnight MicroScan panels for direct identification and susceptibility testing from the BacT/ALERT blood culture system when using FAN (FA and FN) bottles. A simple procedure, in two centrifugation steps, was designed to remove the charcoal particles present in FA and FN bottles. A total of 113 positive blood cultures showing Gram-negative rods were investigated. Enterobacteriaceae were isolated in 104 cases, and Pseudomonas aeruginosa in nine. The MicroScan system correctly identified 106 (93.8%) of the 113 isolates. The seven identification errors included P. aeruginosa (three), Enterobacter cloacae (one), Escherichia coli (one), Klebsiella oxytoca (one), and Klebsiella pneumoniae (one). The VITEK-2 system correctly identified 109 (96.5%) of the 113 samples obtained directly from the blood culture bottles. The four unidentified isolates were Enterobacter cloacae (two), Escherichia coli (one), and P. aeruginosa (one). MicroScan yielded 4/779 (0.5%) very major errors and 28/2825 (0.9%) minor errors. VITEK-2 yielded 2/550 (0.36%) very major errors, 1/1718 (0.05%) major error, and 32/2373 (1.3%) minor errors. Both systems provided excellent identification (correlation of >90%) and susceptibility (correlation of >98%) results. The average times required to obtain identification and susceptibility results using the direct test applied to the VITEK-2 Compact system were 4.57 ± 1.37 h and 6.52 ± 1.64 h, respectively. The VITEK-2 compact system provided results on the same day that the blood culture was found to be positive.     

Detection of Brucella melitensis by BACTEC NR660 blood culture system.

Data on the performance of modern blood culture systems for the detection of Brucella spp. are insufficient. To evaluate the performance of the BACTEC NR660 blood culture system for the detection of Brucella melitensis within the routine 1-week blood culture protocol, a prospective 24-month study was conducted in an area endemic for B. melitensis in southern Israel. Blood samples obtained from patients with suspected brucellosis were monitored and blindly subcultured once per week for 4 weeks, and the fraction of blood cultures positive for B. melitensis detected by the BACTEC NR660 instrument within the first week was determined. During the study period, a total of 373 blood cultures were obtained from patients in whom brucellosis was suspected, and 27 (7.2%) of them, drawn from 21 different patients, were positive for B. melitensis. Twenty-one (78.8%) of these positive cultures were detected by the BACTEC instrument within 7 days, and six positive cultures were detected by subculture after 2 or 3 weeks of incubation. It is concluded that the BACTEC NR660 blood culture system detects the majority of B. melitensis isolates within the routine 1-week blood culture schedule. To maximize the recovery of the organism, however, prolonged incubation and periodic performance of blind subcultures are still required.     

Assessment of the BacT/Alert blood culture system: rapid bacterernia diagnosis with loading throughout the 24 h

Objective: To determine blood culture (BC) diagnostic speed when combining an automated BC system with rapid loading of inoculated bottles throughout the 24 h.
Methods: A total of 111 positive BCs representing bacteremia were investigated in retrospect. All bottles were loaded into the BacT/Alert BC system (Organon Teknika) as soon as possible after sampling and time from specimen collection to Gram stain result was recorded.
Results: The mean time from specimen collection to loading was 3.5 h (median 2.1 h). We found that 74% of all positive BCs collected during daytime (08.00–16.00) ere reported (as Gram stain) to the clinician before 17.00 the next day. For specimens collected between 16.00 and midnight the corresponding proportion was 67%. BCs drawn between midnight and 08.00 were reported before 17.00 the same day in 24% of the cases.
Conclusions: Rapid loading of an automated BC system throughout the 24 h results in fast diagnosis of bacteremia. The diagnostic speed in this study represents a fair estimation of the maximal diagnostic speed accomplishable in a clinical situation with the BacT/Alert system in conjunction with normal daytime laboratory working hours.     

Performance of the BacT/Alert Virtuo Microbial Detection System for the culture of sterile body fluids: prospective multicentre study

ObjectivesContinuous monitoring blood culture systems are commonly used for sterile body fluid cultures. In this multicentre study, we evaluated the performance of the new-generation BacT/Alert Virtuo system compared to the BacT/Alert 3D and conventional culture for the recovery of microorganisms from sterile body fluids.MethodsPeritoneal, cerebrospinal, pericardial, pleural and synovial fluids from adult patients submitted for culture were collected from three different centres. Specimens were inoculated into two bottles of the same bottle type (SA, SN, FA Plus or FN Plus) in equal volumes for simultaneous incubation in the Virtuo and 3D instruments. Each specimen was also Gram stained and seeded to solid media.ResultsA total of 811 specimens were inoculated to 1257 bottle pairs. The Virtuo and 3D showed equivalent recovery of clinically significant microorganisms (127/155, 81.9%, vs. 126/155, 81.3%, respectively). Solid media cultures recovered fewer pathogens than either continuous monitoring system (95/155, 61.3%, p <0.001), including significantly fewer Enterobacteriaceae and enterococci. The Virtuo was significantly faster than the 3D in median time to detection of isolates from the same specimen (12.5 (range, 2.8–101.5) hours vs. 15.5 (range, 4.3–78.5) hours, p <0.001). Direct specimen Gram stain detected the eventual pathogen in 30 (26.1%) of 115 significant positive specimens.ConclusionsThe BacT/Alert Virtuo system was equivalent to the 3D system in organism recovery from sterile body fluid culture but showed faster detection of growth as a result of design enhancements.     

Growth and detection of filamentous fungi in the BacT/Alert blood culture system.

Little is known about the behaviour of filamentous fungi in most blood culture systems, despite their increasingly recognised role in infections of immunocompromised hosts. The ability of the BacT/Alert system (Organon Teknika, Durham, North Carolina, USA) to detect the growth of 19 such fungi was examined. Eleven species grew and were detected rapidly; two species did not grow. Six species grew slowly, and were generally only recovered with terminal subculture after prolonged incubation. The CO2 production graph for some of these fungi showed a slow but steady rise, insufficient to cause the apparatus to signal positive. These results show that the BacT/Alert system may miss some fungi, either because of no growth in the medium or undetected slow growth. The latter problem could be overcome by prolonged incubation and terminal subculture when fungal infection is considered likely. Alteration of the signalling mechanism might permit earlier detection of some slow growing fungi.     

Comparison of the BacT/Alert pediatric blood culture system, Pedi-BacT, with conventional culture using the 20-milliliter Becton-Dickinson supplemented peptone broth tube.

The performance of the Pedi-BacT system, the BacT/Alert (Organon Teknika Corp., Durham, N.C.) pediatric blood culture bottle, was compared with that of a conventional 20-ml supplemented peptone broth tube (Becton-Dickinson Corp., Cockeysville, Md.) (BD system) in matched aerobic cultures. The tubes of the BD system were visually examined daily for 7 days and were subcultured during the first 24 h of incubation. Pedi-BacT cultures were mechanically agitated and continuously monitored for growth by the instrument. Of the 6,628 compliant pairs, 331 (5.0%) were positive in both systems, 220 (3.3%) were positive in the Pedi-BacT system only, and 170 (2.6%) were positive in the BD system only. One (0.02%) false-negative culture and 15 (0.2%) false-positive cultures occurred with the Pedi-BacT system while 20 (0.3%) false-negative cultures and 35 (0.5%) false-positive cultures occurred with the BD system. Of 288 clinically significant organisms detected in matched pairs from which a single isolate was recovered, 176 (61%) were recovered from both systems, 83 (29%) were recovered from the Pedi-BacT system only (P < 0.0001), and 29 (10%) were recovered from the BD system only. Members of the family Enterobacteriaceae (P < 0.01), miscellaneous nonfermenters (P < 0.05), and Candida spp. (P < 0.01) were isolated more frequently in the Pedi-BacT system than in the BD system. No significant difference in recovery of other organisms was found between the systems. The average time to detection for the Pedi-BacT system ranged from 11.5 h for streptococci to 29.7 h for enterococci, while that for the BD system ranged from 20.3 h for streptococci to 66.4 h for some nonfermenters. The BacT/Alert system is a reliable, labor-saving alternative to conventional blood culture methods.     

Prevalence of mycobacteremia in Indian HIV-infected patients detected by the MB/BacT automated culture system

The use of automated blood cultures system, such as MB/BacT, has provided a novel opportunity for laboratories to diagnose mycobacteremia in HIV-infected patients. However, no such study has been carried out in India so far. This prospective study was conducted on 52 HIV-positive patients with suspected tuberculosis who were referred to our tertiary care hospital in New Delhi. In these patients, the prevalence of mycobacteremia was evaluated using the MB/BacT automated culture system (bioMérieux, France). Twenty-seven HIV-negative but suspected tuberculosis patients were also included for comparison. Mycobacteria could be isolated from sputa or fecal samples of 20 HIV-positive patients (38.4%), and in nine (45%) of these 20 cases, mycobacteria could also be isolated simultaneously from their blood specimens. In the remaining 32 patients, all relevant non-hematological clinical samples remained negative for mycobacteria, but the pathogen could be detected from the blood samples of seven (21.87%) of these 32 patients. Therefore, only 25 (48%) clinically suspected patients remained negative in both Löwenstein-Jensen (L-J) and MB/BacT culture methods, and 12 of these responded to anti-tubercular treatment, while in the rest either non-tubercular diagnosis was established or they were lost to follow-up. The study revealed that low \( {\text{CD}}^{ + }_{4} \) counts and poor or no reactivity to purified protein derivative (PPD) were the best clinical predictors for the occurrence of mycobacteremia in HIV-positive patients. Of the 16 isolates from blood, 13 were diagnosed as Mycobacterium tuberculosis and one each were identified as M. avium, M. kansasii, and a mixed infection of M. tuberculosis and M. avium complex. The prevalence rate of mycobacteremia was significantly low (11.1%) in HIV-negative patients. In conclusion, this study showed that blood culture could be an important adjunct investigation for confirming the clinical diagnosis of tuberculosis in HIV-positive patients.     

Evaluation of the MB/BacT System for recovery of mycobacteria from clinical specimens in comparison to Lowenstein–Jensen medium

Objective   To evaluate the performance of the MB/BacT system (Organon Teknika) in comparison to Lowenstein–Jensen (LJ) solid medium for recovery of mycobacteria from clinical specimens.
Methods   Two thousand three hundred and ten specimens (1626 respiratory, 593 urine, 60 body fluids, five tissue and 26 others) were inoculated in MB/BacT (0.5 mL) and on two LJ slants (0.25 mL each). N -acetyl- l -cysteine-NaOH (final concentration 2%) was used for decontamination.
Results   Two hundred and fifty-one (10.8%) mycobacterial isolates [190 Mycobacterium tuberculosis complex (MTBC) and 61 non-tuberculous mycobacteria (NTM)] were detected. Of these 251 isolates, 234 (93.2%; 181 MTBC and 53 NTM) were detected in MB/BacT and 169 (67.3%; 154 MTBC and 15 NTM) on LJ. The mean (median) times to detection of MTBC by MB/BacT and LJ were 13.8 (13) and 22.1 (20) days, respectively, while overall contamination rates were 7.7% and 8.1%, respectively.
Conclusions   Sensitivity and time to detection were significantly better with MB/BacT than with solid LJ medium.     

Clinical comparison of the isolator and BacT/Alert aerobic blood culture systems.

The performance characteristics of the Isolator (Wampole Laboratories, Cranbury, N.J.) and the BacT/Alert (Organon Teknika Corporation, Durham, N.C.) aerobic blood culture systems were compared for 6,009 blood culture sets obtained from patients with suspected bloodstream infections. The BacT/Alert aerobic bottle [BTA(O2)] was continuously agitated while it was incubated in 5% CO2 at 36 degrees C; culture plates prepared from the Isolator tube [I(O2)] were incubated in 5% CO2 at 37 degrees C. From 394 blood cultures, 416 clinically significant isolates of bacteria and yeasts were recovered. The overall yields for BTA(O2) and I(O2) were not significantly different (319 versus 336; P = 0.20). I(O2) recovered significantly more staphylococcus (P < 0.05) and yeast isolates (P < 0.01). BTA(O2) recovered significantly more aerobic and facultatively anaerobic gram-negative bacilli (P < 0.05). In blood culture sets which produced growth of the same organisms in both the BTA(O2) and I(O2) systems, the BTA(O2) system detected growth sooner, but more rapid identification was possible with the I(O2) system by virtue of earlier isolation of colonies on solid media.