Direct Susceptibility Testing of Positive Blood Cultures by Using Sensititre Broth Microdilution Plates

Abstract

Traditional susceptibility testing of blood cultures requires overnight incubation in order to obtain isolated colonies. Susceptibility results can be reported up to 24 h sooner by using a bacterial pellet from the blood culture broth. This study evaluated the accuracy of direct susceptibility testing from positive ESP blood culture broths by using Sensititre broth microdilution plates compared to testing with isolated colonies. Practical inclusion criteria were applied to gram-positive organisms to avoid reporting susceptibilities for probable contaminants. All gram-negative organisms were tested directly. An aliquot of the blood culture was centrifuged, and the resulting pellet was used to make a 0.5 McFarland suspension. Microdilution plates were inoculated and interpreted according to the manufacturer's instructions. Colony counts were performed to ensure proper colony density was achieved. A total of 199 patient and seeded blood cultures were evaluated for both essential (within ؎1 twofold dilution) and categorical (sensitive, intermediate, or resistant) agreement. Testing of 93 gram-positive isolates (1,214 antimicrobial agent-organism combinations) yielded 98% essential agreement and categorical error rates of 0.3% minor, no major (false resistance), and 1.7% very major (false susceptibility) errors. For 106 gram-negative isolates (1,828 antimicrobial agent-organism combinations), the essential agreement was 99%. Categorical error rates were 0.5, 0, and 2.0% for minor, major, and very major errors, respectively. Performance was comparable for both gram-positive and gram-negative isolates, as well as for both aerobic and anaerobic media. Using this direct testing methodology, reliable susceptibility results can be reported to physicians 24 h sooner, allowing earlier appropriate modification of antimicrobial therapy.Timely intervention in the treatment of bloodstream infection is of paramount importance to increase the chances of a favorable outcome, since empirical therapy can be modified upon receipt of in vitro antimicrobial susceptibility testing (AST) results. AST results can assist in modifying antimicrobial therapy (13), and investigators have demonstrated decreased mortality with early treatment (10). Further data from outcome-based studies assessing the effect of rapid reporting of susceptibility results have shown a decrease in the number of laboratory tests and procedures ordered (4), decreased length of stay (1), and decreased health care costs and quicker modification of antimicrobial therapy (1,4,23).The standard protocol for identification and susceptibility testing of positive blood cultures involves Gram stain and subculture of blood culture broth onto solid medium and overnight incubation to obtain isolated colonies. These colonies are then used to make a standardized inoculum used for AST. A direct susceptibility testing algorithm from positive ESP (TREK Diagnostic Systems, Cleveland, Ohio) blood culture bottles was designed that circumvents the isolation process and expedites the reporti...

Evaluation of the Merlin MICRONAUT System for Rapid Direct Susceptibility Testing of Gram-Positive Cocci and Gram-Negative Bacilli from Positive Blood Cultures

“…In general, good agreement between direct and standard susceptibility testing results was observed when gram-negative bacilli were tested, including both members of the Enterobacteriaceae and Pseudomonas species (4-6, 12, 14, 18, 21, 24, 26, 28). For direct testing of gram-positive cocci from blood cultures, only limited data from small studies are available for the VITEK, the VITEK 2, the Sensititre, and the MicroScan systems (5,6,8,18,26,29). A significantly higher rate of disagreement between direct and standard testing results for gram-positive cocci than for gramnegative bacilli was found.…”

Evaluation of Penicillin Binding Protein 2a Latex Agglutination Assay for Identification of Methicillin-Resistant Staphylococcus aureus Directly from Blood Cultures

“…The supernatant was discarded, and the bacterial pellet was used as the inoculum for the PBP2a test. Direct testing specifically for susceptibility results by using this preparation has been described previously (3). From this point, the PBP2a assay was performed according to manufacturer instructions.…”

Early diagnosis of resistant pathogens

“…For example, Chapin and Musgnug reported 98% essential agreement and categorical error rates of 0.3% minor, no major (false resistance) and 1.7% very major (false susceptibility) errors in Gram-positive isolates, and 99% essential agreement, 0.5, and 0 and 2.0% respectively for minor, major and very major errors, respectively 28 . The reporting time for the direct testing of susceptibility for blood culture isolates by the VITEK 2 system ranged from 3.3 to 17.5 h. Compared with conventional methods that require 1 or 2 d, this method can make same-day reporting possible and thus permit better patient management 29 .…”