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The Mycobactericidal Efficacy of Orthophthalaldehyde and the Comparative Resistances of Mycobacterium Bovis, Mycobacterium Terrae, and Mycobacterium Chelonae

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The Mycobactericidal Efficacy of Orthophthalaldehyde and the Comparative Resistances of Mycobacterium Bovis, Mycobacterium Terrae, and Mycobacterium Chelonae Brigham Young University BYU ScholarsArchive Faculty Publications 1999-05-01 The Mycobactericidal Efficacy of Orthophthalaldehyde and the Comparative Resistances of Mycobacterium Bovis, Mycobacterium Terrae, and Mycobacterium Chelonae Richard A. Robison [email protected] Adam W. Gregory G. Bruce Schaalje Jonathan D. Smart Follow this and additional works at: https://scholarsarchive.byu.edu/facpub Part of the Microbiology Commons BYU ScholarsArchive Citation Robison, Richard A.; Gregory, Adam W.; Schaalje, G. Bruce; and Smart, Jonathan D., "The Mycobactericidal Efficacy of Orthophthalaldehyde and the Comparative Resistances of Mycobacterium Bovis, Mycobacterium Terrae, and Mycobacterium Chelonae" (1999). Faculty Publications. 620. https://scholarsarchive.byu.edu/facpub/620 This Peer-Reviewed Article is brought to you for free and open access by BYU ScholarsArchive. It has been accepted for inclusion in Faculty Publications by an authorized administrator of BYU ScholarsArchive. For more information, please contact [email protected], [email protected]. 324 INFECTION CONTROL AND HOSPITAL EPIDEMIOLOGY May 1999 THE MYCOBACTERICIDAL EFFICACY OF ORTHO- PHTHALALDEHYDE AND THE COMPARATIVE RESISTANCES OF MYCOBACTERIUM BOVIS, MYCOBACTERIUM TERRAE, AND MYCOBACTERIUM CHELONAE Adam W. Gregory, BS; G. Bruce Schaalje, PhD; Jonathan D. Smart, BS; Richard A. Robison, PhD ABSTRACT OBJECTIVES: To assess the mycobactericidal efficacy of against all mycobacterial test suspensions. Four replicates of an agent relatively new to disinfection, ortho-phthalaldehyde each organism-disinfectant combination were performed. (OPA) and to compare the resistances of three Mycobacterium RESULTS: Results were assessed by analysis of vari- species. Mycobacterium bovis (strain BCG) was compared with ance. M terrae was significantly more resistant to 0.05% OPA Mycobacterium chelonae and Mycobacterium terrae to investigate than either M bovis or M chelonae. At 0.21% OPA, M terrae was the feasibility of using either of the latter two species in tubercu- slightly more susceptible than one test suspension of M bovis, locidal testing. M chelonae (a rapid grower) and M terrae (an inter- but not significantly different from the other. M chelonae was sig- mediate grower) both grow faster and are less virulent than M nificantly less resistant than the other species at both OPA con- bovis (a slow grower). centrations. At their respective minimum effective concentra- DESIGN: The quantitative suspension protocol specified tion, OPA achieved a 6-log10 reduction of M bovis in nearly one by the Environmental Protection Agency (EPA), the sixth the time required by glutaraldehyde (5.5 minutes vs 32 Tuberculocidal Activity Test Method (EPA test), was used minutes). throughout this study. Standard suspensions of all three species CONCLUSIONS: These data, along with other recent were prepared in a similar manner. Two suspensions of M bovis, studies, lend support to the idea that M terrae may be a suitable created in different laboratories, were used. These were tested test organism for use in the tuberculocidal efficacy testing of dis- against two concentrations of alkaline glutaraldehyde to provide infectants. They also confirm the relatively rapid tuberculocidal reference data. Two concentrations of OPA were evaluated activity of OPA (Infect Control Hosp Epidemiol 1999;20:324-330). While airborne transmission of tuberculosis (TB) is One of the most critical variables in tuberculocidal the most common route of spread, cases due to indirect testing is the selection of the challenge organism (which contact with semicritical medical instruments (such as should possess resistance equivalent to that of M tubercu- bronchoscopes and endoscopes) continue to occur.1-4 losis). This has been a source of variability in past studies, Contributing to this problem are exaggerated tuberculoci- contributing to the difficulty of establishing a truly stan- dal label claims, which have been discovered even on high- dardized protocol. The quantitative suspension procedure level disinfectants.5-7 Worse yet, some disinfectants have currently approved by the Environmental Protection been found to support the presence of contaminating bac- Agency (EPA) was described first in 1987 by Ascenzi et al.5 teria for extended periods of time.8-10 Because mycobacte- Known as the EPA Tuberculocidal Activity Test Method ria generally are more resistant to chemical disinfection (EPA test), this quantitative procedure has attempted to than most other vegetative organisms,11 these data raise control many of the variables involved in tuberculocidal special concerns with respect to Mycobacterium tuberculo- testing. It specifies the use of Mycobacterium bovis (strain sis. One way to reduce the incidence of disease caused by BCG) as the challenge organism.12 M bovis also is specified inadequately disinfected medical devices is to assure that in the Association of Official Analytical Chemists tubercu- tuberculocidal assessment of disinfectants is accomplished locidal procedure.13 It is related closely to M tuberculosis with accurate and standardized test procedures. and has demonstrated similar levels of resistance to a vari- From the Department of Microbiology (Dr. Robison, Mr. Gregory, and Mr. Smart); the Department of Statistics (Dr. Schaalje), Brigham Young University, Provo, Utah. This work was supported by a grant from the Professional Development Committee of Brigham Young University and Johnson & Johnson Medical, Inc. The authors thank Heather Hall-Albert, Melissa Hardy, Jess Price, Jonathan Olivier, and Sara Maycock-Olivier for their many hours of generous technical assistance. Address reprint requests to Richard A. Robison, PhD, 791 WIDB, Department of Microbiology, Brigham Young University, Provo, UT 84602-5133. 98-OA-057. Gregory AW, Schaalje GB, Smart JD, Robison RA. The mycobactericidal efficacy of ortho-phthalaldehyde and the comparative resis- tances of Mycobacterium bovis, Mycobacterium terrae, and Mycobacterium chelonae. Infect Control Hosp Epidemiol 1999;20:324-330. Vol. 20 No. 5 MYCOBACTERICIDAL EFFICACY OF ORTHO-PHTHALALDEHYDE 325 ety of disinfecting agents.14 However, the use of M bovis has (2.5%), and one was stressed through simulated reuse, as drawbacks. In addition to being more virulent than several described below, and diluted to contain 1.5% glutaraldehyde. other known strains, it is slow-growing, requiring at least 3 This latter solution constitutes the manufacturer’s minimum weeks before final results can be obtained. effective concentration. The OPA samples originated from a Several researchers have experimented with faster- solution stressed through simulated use and diluted to con- growing, less pathogenic strains of mycobacteria in hopes tain 0.21% (the minimum effective concentration) and 0.05% of identifying a more suitable test species. This study OPA. The 0.05% OPA solution provided longer kill times, for involves a comparative analysis of a rapid-growing species more suitable comparison of the three Mycobacterium (Mycobacterium chelonae) and an intermediate-growing species. Simulated reuse of the disinfectant solutions consist- species (Mycobacterium terrae) with M bovis (a slow-growing ed of the addition of 5% horse serum and the processing of species). two sets of anesthesia equipment for 42 cycles over 14 days, The comparative resistances of these strains were according to a standard reuse protocol. A 0.8% phenol stan- evaluated using ortho-phthalaldehyde (OPA), a compound dard, the control disinfectant solution specified in the EPA relatively new to disinfection. The current standard for test method, also was assayed with each test suspension. high-level disinfection of semicritical instruments is 2% alka- line glutaraldehyde. Although this agent has well-documented Test Suspensions bactericidal, sporicidal, and mycobactericidal activity, it is Mycobacterial test suspensions were prepared not without deficiencies, including inactivation by sponta- according to the EPA test protocol. On each test day, the neous polymerization and loss of activity with dilution.15,16 It required number of frozen stock vials was allowed to thaw. also requires a relatively long exposure time to exact a sig- The contents of each vial were added to a prechilled 30-mL nificant tuberculosis kill.17 Teflon-on-glass tissue homogenizer with an equal volume Because OPA is a relatively novel disinfectant, few effi- of phosphate-buffered gelatin. The mixture was homoge- cacy studies have been performed. However, the limited nized on ice for 1 to 2 minutes. The homogenized suspen- available data suggest that it has much higher activity than sion then was diluted using ice-cold physiological saline 2% glutaraldehyde, especially with respect to its tuberculoci- solution containing 0.1% Tween 80 to approximately 1ϫ107 dal and virucidal action. In this study, we evaluated the CFU/mL. This suspension was kept on ice until used to mycobactericidal effects of two concentrations of OPA on the inoculate test disinfectants. three mycobacterial species. For comparison purposes, data also were collected on two concentrations of glutaraldehyde Disinfectant Test Procedure using the current tuberculocidal test organism, M bovis. All disinfectants were assayed according to the EPA test protocol. Each test consisted of inoculating the disin- METHODS fectant at time zero with a standardized test organism sus- Test Organisms pension and sampling this mixture at predetermined time Stock suspensions of M terrae
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