Haemophilus influenzae ATCC 49766, an alternative quality control strain for monitoring broth microdilution susceptibility tests with selected beta-lactams

Abstract

A P-lactamase-negative, ampicillin-resistant strain of Haemophilus influenzae is currently used for quality control of broth microdilution tests performed with Haemophilus test medium (HTM). Studies with eight lots of HTM broth documented the fact that MIC limits for some antimicrobial agents are unrealistically stringent; i.e., only three of eight lots of HTM broth were satisfactory for testing cefaclor. An alternative, ampicillinsusceptible strain of H. influenzae (ATCC 49766) was found to provide much more reproducible results with five problematic drugs (cefaclor, cefuroxime, cefamandole, loracarbef, and cefonicid). Multilaboratory studies defined MIC control limits for both control strains tested against 12 antimicrobial agents.For testing the susceptibility ofHaemophilus influenzae to antimicrobial agents, supplemented Mueller-Hinton medium has been recommended (2-4, 6). However, Mendelman et al. (5) have described unsatisfactory results with the agar version of Haemophilus test medium (HTM). A P-lactamasenegative, ampicillin-resistant strain of H. influenzae (ATCC 49247) was selected for quality control of susceptibility tests that use HTM agar or broth (1, 6). It was selected because its relative resistance to ampicillin and other P-lactam compounds maximized the number of drugs with MIC endpoints that are likely to be within the range of concentrations normally tested in microdilution trays. However, in vitro tests with some cephalosporins are extremely inoculum dependent and are also influenced by a wide range of other technical variables that make it very difficult to obtain reproducible results with the control strain of H. influenzae (ATCC 49247). Doern et al. (1) proposed MIC limits for broth microdilution tests with 18 antimicrobial agents. They were unable to obtain useful control limits with cefaclor and also noted marked variability in cefuroxime and cefamandole control test results. The current report extends those observations by documenting the amount of variability with eight lots of HTM broth. Such variability emphasized the need for an alternative control strain that provides more reproducible results with cefaclor, cefuroxime, and related f-lactams.After careful evaluation of several clinical isolates that might serve that function, H. influenzae ATCC 49766 was selected by one of us (J.H.J.). A five-laboratory collaborative effort was subsequently initiated to define MIC control limits for 12 antimicrobial agents tested against two strains of H. influenzae (ATCC 49247 and ATCC 49766). The results of those studies are described in this report. * Corresponding author. MATERIALS AND METHODSInitial screening tests. Two lots of Mueller-Hinton broth were purchased from each of the following manufacturers: Difco Laboratories, Detroit, Mich.; Becton Dickinson Microbiology Systems, Cockeysville, Md.; Acumedia, Baltimore, Md.; and Unipath Oxoid, Ogdensburg, N.Y. Twofold dilutions of each antimicrobial agent were prepared in each of the eight broth media and dispensed into wells in microdilution tr...

In vitro susceptibility testing of Haemophilus influenzae: review of new National Committee for Clinical Laboratory Standards recommendations

“…A further difficulty pertains to inter-and intramanufacturer variability with batches of HTM (2,7,11,12). Such variability is perhaps not surprising, as the individual ingredients used to make HTM have profound effects on media performance.…”

“…In addition to lot-to-lot variability, within-lot variability also exists with HTM-based susceptibility tests (2,7,12). The magnitude of this problem is unknown.…”

Quality control parameters for susceptibility tests of Haemophilus influenzae against three carbapenems

“…However, tests of that control strain against some cephem drugs have proven to be too variable to be useful for quality control purposes. A second ampicillin‐susceptible strain of H. influenzae (ATCC 49766) has been subsequently selected for monitoring tests of those problematic drugs [1,2]. The latter control strain is very susceptible to many β‐lactam drugs, and MICs are likely to be below the range of concentrations normally tested.…”

“…In 1991, multi‐laboratory studies were carried out in order to evaluate both control strains of H. influenzae against a variety of antimicrobial agents. Broth microdilution tests with two carbapenems (imipenem and meropenem) were included [1]. For testing meropenem, H. influenzae ATCC 49766 was preferred, because MICs were much more reproducible.…”

Comparison of agar dilution, broth dilution, and disk diffusion testing of ampicillin against Haemophilus species by using in-house and commercially prepared media

“…Nevertheless, problems with HTM have been reported, including its inability to support the growth of H. infiuenzae and provide consistency in susceptibility testing results (11). Interand intramanufacturer variabilities with batches of HTM (1, 3, 6, 7) as well as lot-to-lot and within-lot variabilities of HTM have also been reported (1,3,7). In June 1992, NCCLS realized that the HTM DD method could falsely categorize ,3-lactamase-negative, ampicillin-susceptible (BLNAS) H. influenzae strains as 3-lactamase-negative, ampicillin-resistant (BLNAR) strains.…”