Determination of in vitro susceptibility to ABT-492 by BSAC standardized methodology

J. M. Andrews*, J. P. Ashby, G. Jevons and R. Wise

Department of Microbiology, City Hospital, Dudley Road, Birmingham B18 7QH, UK

Keywords: ABT-492, susceptibility, BSAC method

Sir,

ABT-492 is a new quinolone currently under development by Abbott Laboratories (Abbott Park, IL, USA) that has been shown to have broad spectrum in vitro activity, including anaerobes.1 In this study, we describe a standardized disc testing method to determine the susceptibility of organisms to ABT-492 based on the BSAC recommendations.2

A total of 885 organisms, comprising 100 Escherichia coli, 100 Klebsiella spp., 29 indole-positive Proteus, 50 Proteus mirabilis, 50 Enterobacter spp., 50 methicillin-susceptible Staphylococcus aureus (MSSA), 25 methicillin-resistant S. aureus (MRSA), 30 Staphylococcus saprophyticus, 20 Staphylococcus epidermidis, 44 Moraxella catarrhalis, 34 haemolytic streptococci, 87 Haemophilus influenzae (including one organism with reduced susceptibility to fluoroquinolones), 97 Streptococcus pneumoniae (including two mutants with ParC Ser-79 -> Tyr and either a GyrA or Glu-85 -> Tyr substitution; one recent fluoroquinolone-resistant clinical isolate), 29 Streptococcus milleri, 91 Enterococcus faecalis, 11 Neisseria meningitidis, and 38 Neisseria gonorrhoeae (including five isolates with reduced susceptibility to ciprofloxacin) were studied, together with appropriate ATCC and NCTC strains (shown in Table 1). MIC determinations and disc testing were undertaken by BSAC standardized methodology.2 Discs containing 1, 2 or 5 µg of ABT-492 were prepared as previously described.3 Acceptable limits for the control strains were determined by disc testing each strain 40 times on pre-poured plates from Oxoid and bioMérieux and media poured to a depth of 3.5, 4 and 4.5 mm. Zone diameters were combined and 95 percentiles calculated.2 Applying the BSAC formula4 to pharmacokinetic data (Cmax of ~3.2 mg/L following an oral dose of 250 mg with a terminal half-life of 7–8 h), an MIC susceptible breakpoint (BP) of 0.4 mg/L was calculated.


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Table 1.  MIC and zone diameter breakpoints for interpreting susceptibility to ABT-492 by BSAC methodology and expected MIC and zone diameter ranges for the control strains
 
MIC data were reviewed to ascertain the ABT-492 MIC ranges for the ‘wild sensitive’ populations. For Enterobacteriaceae, M. catarrhalis, haemolytic streptococci, S. milleri, N. meningitidis and E. faecalis, ABT-492 MICs for the population lacking a mechanism of resistance were 0.015–0.12 mg/L, 0.004–0.06 mg/L, 0.008–0.06 mg/L, 0.004–0.06 mg/L, 0.001 mg/L and 0.06–2 mg/L, respectively. In the case of staphylococci, the MIC range for the susceptible population was 0.001–0.015 mg/L. In contrast, for the 25 MRSA isolates that had reduced susceptibility to ciprofloxacin (ciprofloxacin MIC range 64–>128 mg/L), ABT-492 MICs ranged from 0.25 to 1 mg/L. For S. pneumoniae, ABT-492 MICs ranged between 0.002 and 0.015 mg/L except for the three isolates of S. pneumoniae with ciprofloxacin MICs of 128 mg/L. These organisms had corresponding ABT-492 MICs of 0.03 mg/L. In the case of H. influenzae, MICs for the susceptible population were between 0.001 and 0.004 mg/L. For the isolate with no zone of inhibition to a nalidixic acid 30 µg disc (ciprofloxacin MIC 0.06 mg/L), the ABT-492 MIC was 0.004 mg/L and therefore indistinguishable from the susceptible population. Five isolates of N. gonorrhoeae with reduced susceptibility to the quinolones (no zone of inhibition to a nalidixic acid 30 µg disc) had ABT-492 MICs between 0.03 and 0.25 mg/L. For the organisms lacking a mechanism of resistance, MICs ranged between 0.001 and 0.002 mg/L.

For all isolates except E. faecalis, zones for disc contents of 2 and 5 µg were unacceptably large, susceptible organisms having zones larger than 40 mm (data not shown). Zone diameter data for a 1 µg disc were therefore analysed for all genera except E. faecalis, looking at the populations lacking a mechanism of resistance. A summary of MIC and zone diameter BPs for interpreting susceptibility and expected MIC and zone diameter ranges for the control strains is shown in Table 1. Although not the drug of first choice, quinolones have been used to treat enterococcal urinary tract infections.5 In this case, an MIC BP of 0.4 mg/L would seem inappropriate as concentrations of ABT-492 in urine are significantly higher than those found in blood. Recommendations are therefore given based on microbiological BPs and a disc content of 5 µg (Table 1).

These data indicate that an ABT-492 disc content of 1 µg is the most appropriate concentration for determining susceptibility by BSAC methodology except for E. faecalis where a 5 µg disc is more suitable. For the detection of low-level quinolone resistance in H. influenzae and Neisseria spp., it would be prudent for a 30 µg nalidixic acid disc to be used (recommended by the BSAC).

Footnotes

* Corresponding author. Tel: +44-121-507-5693; Fax: +44-121-551-7763; E-mail: jenny.andrews{at}sbh.nhs.uk Back

References

1 . Nilius, A. M., Hensey-Rudloff, D., Almer, L. et al. (2002). Comparative in vitro activities of the new quinolone ABT-492, Trovafloxacin, levofloxacin and ciprofloxacin. In Program and Abstracts of the Forty-second Interscience Conference on Antimicrobial Agents and Chemotherapy, San Diego, CA, 2002. Abstract F-546, p. 183. American Society for Microbiology, Washington, DC, USA.

2 . Working Party of the British Society for Antimicrobial Chemotherapy. (2001). Antimicrobial Susceptibility Testing: BSAC Working Party Report. Journal of Antimicrobial Chemotherapy 48, Suppl. S1, 43–57.[Abstract/Free Full Text]

3 . Andrews, J. M., Ashby, J. P., Jevons, G. et al. (1999). Tentative minimum inhibitory concentration and zone diameter breakpoints for moxifloxacin using BSAC criteria. Journal of Antimicrobial Chemotherapy 44, 819–22.[Abstract/Free Full Text]

4 . Working Party of the British Society for Antimicrobial Chemotherapy. (1991). A Guide to Sensitivity Testing. Journal of Antimicrobial Chemotherapy 27, Suppl. D, 1–50.[ISI][Medline]

5 . Landman, D. J. & Quale, J. M. (1997). Management of infections due to resistant enterococci: a review of therapeutic options. Journal of Antimicrobial Chemotherapy 40, 161–70.[Abstract]