Journal of Antimicrobial Chemotherapy (2004) 53, 1039 1044 DOI: 10.1093/jac/dkh248 Advance Access publication 5 May 2004 Surveillance of susceptibility patterns in 1297 European and US anaerobic and capnophilic isolates to co-amoxiclav and five other antimicrobial agents Laura M. Koeth 1 *, Caryn E. Good 1, Peter C. Appelbaum 2, Ellie J. C. Goldstein 3, Arne C. Rodloff 4, Marina Claros 4 and Luc J. Dubreuil 5 1 Laboratory Specialists, Inc., Cleveland, OH; 2 M. S. Hershey Medical Center, Hershey, PA; 3 R. M. Alden Research Laboratory, Santa Monica, CA, USA; 4 Institute für Medizinische Mikrobiologie und Infektionsepidemiologie der Universität Leipzig, Leipzig, Germany; 5 ADRINORD, Lille, France Received 27 October 2003; returned 14 January 2004; revised 12 March 2004; accepted 22 March 2004 In vitro susceptibility data were collected for co-amoxiclav and other antimicrobial agents against 1297 recent anaerobe isolates collected in Europe and the USA. The co-amoxiclav (amoxicillin/clavulanic acid) MIC 50/90 s (amoxicillin/clavulanic acid concentration in a ratio of 2:1, expressed in terms of amoxicillin concentration in mg/l) were 0.5/4 for Bacteroides fragilis, 0.125/1 for Prevotella species, 0.125/0.25 for Fusobacterium nucleatum, 0.5/1 for Eikenella corrodens, 0.25/8 for Peptostreptococcus anaerobius, 0.125/0.5 for Micromonas (Peptostreptococcus) micros, 0.25/0.5 for Fingoldia (Peptostreptococcus) magna, and 0.125/0.125 for Porphyromonas species. The co-amoxiclav susceptibility rate for B. fragilis was 94.6%, for P. anaerobius 84.3% and for all other species tested 100%. These data indicate that co-amoxiclav remains an effective drug for the antimicrobial treatment and prophylaxis of many anaerobic infections. Among the comparator drugs, metronidazole was very active against all bacterial species (>96% susceptible) except E. corrodens (MIC 50/90 of >32/>64 mg/l), which is a capnophilic organism. Imipenem was also highly active against all species (>98% susceptible). Levofloxacin and clindamycin were the least potent agents tested, particularly against Bacteroides, Prevotella and Peptostreptococcus (levofloxacin susceptibility rates: Bacteroides 72.7%, Prevotella 71.5%, F. magna 72.4%; clindamycin susceptibility rates: Bacteroides 79.5%, Prevotella 92.1%, F. magna 84.7%). Keywords: amoxicillin/clavulanic acid, anaerobes, susceptibility data Introduction As part of a global surveillance study, this study was carried out to collect susceptibility data for co-amoxiclav and five other antimicrobial agents against key anaerobic organisms. Gram-negative anaerobic bacilli are the most commonly encountered anaerobes in clinical infections, found in more than half of the specimens yielding anaerobes. Bacteroides, Fusobacterium, Porphyromonas and Prevotella, which are all members of this group, were included in this study. Peptostreptococcus, an important genus among the Gram-positive nonspore-forming anaerobic bacteria, was also examined. Two of the peptostreptococci studied have recently been reclassified from Peptostreptococcus magnus to Fingoldia magna and Peptostreptococcus micros to Micromonas micros. Eikenella corrodens, a capnophilic organism that is part of the normal flora of mucous membranes especially of the mouth, was included because of its significance in cutaneous infections and wounds. Anaerobic bacteria are commonly found in polymicrobial infections with reported incidence ranging from 4% to 50%, 1,2 however, anaerobic cultures are not routinely carried out. Therefore, it is important to include an antimicrobial agent with known efficacy against anaerobes when empirically treating an infection with a high probability of anaerobic presence. 2 4 Like the increasing antimicrobial resistance in common aerobic bacteria, anaerobes have also showed trends towards some increasing resistance in recent years. 5 7 Since susceptibility testing for anaerobes is often not done, except for sterilesite isolates (blood, CSF, etc.), it is especially important to know the background susceptibility patterns in the community. 8 This is commonly and efficiently done through regular surveillance studies. These studies, when carried out repeatedly over time, may additionally provide researchers and clinicians with advance notice of emerging patterns of resistance. At present, empirical treatment of an infection with a high probability of anaerobe presence involves either the use of a very broad-... *Corresponding author. Tel: +1-440-835-4458; Fax: +1-440-835-5786; E-mail: amdlmk@aol.com... 1039 JAC vol.53 no.6 The British Society for Antimicrobial Chemotherapy 2004; all rights reserved.
L. M. Koeth et al. Table 1. Collecting centres and numbers of isolates collected at each Number of each bacterial species c Collecting centre City State/Country Total no. isolates collected BF PR FN EC PS PO VAMC West Los Angeles a Los Angeles California 191 24 47 34 0 81 5 Mount Sinai Medical Center a New York New York 12 10 2 0 0 0 0 M. S. Hershey Medical Center Hershey Pennsylvania 105 61 16 6 0 21 1 Creighton University a Omaha Nebraska 9 6 1 0 0 2 0 Cleveland Clinic Foundation a Cleveland Ohio 30 0 19 0 4 7 0 R. M. Alden Research Foundation Santa Monica California 249 0 0 71 58 67 53 University of Lille Lille France 131 34 44 13 13 21 6 University of Leipzig Leipzig Germany 236 60 46 6 0 79 45 Eijkman-Winkler Institute b Utrecht The Netherlands 11 0 1 2 8 0 0 Humbold University b Berlin Germany 25 0 22 0 0 3 0 University of Halle b Halle Germany 139 33 53 11 0 27 15 University of Bonn b Bonn Germany 38 20 13 1 0 4 0 Sera & Vaccines Research Laboratory b Warsaw Poland 31 0 28 0 0 3 0 University Hospital of Wales b Cardiff UK 90 45 11 13 0 21 0 Isolate number totals 1297 293 303 157 83 336 125 a Tested at M. S. Hershey Medical Center. b Tested at University of Leipzig. c BF, B. fragilis; PR, Prevotella spp.; FN, F. nucleatum; EC, E. corrodens; PS, Peptostreptococcus spp. (includes P. anaerobius, M. micros and F. magna); PO, Porphyromonas spp. spectrum antimicrobial agent or combined therapy with a drug effective against the known or suspected facultative anaerobes and one known to be effective against strict anaerobes. 9 Some of the drugs known to target strict anaerobes include clindamycin and metronidazole. Both drugs have good efficacy against many anaerobes but clindamycin has no effect on enteric Gram-negative facultative anaerobic bacilli and metronidazole is essentially ineffective against most other classes of bacteria. Either drug is generally given in combination with aminoglycosides or cephalosporins. Penicillin and related β-lactam drugs are often effective against both strict anaerobes and facultative anaerobes, but are often inactivated by bacteria that produce β-lactamases. Carbapenems, a class of β-lactams that includes imipenem and meropenem, are also very effective against both classes of bacteria and are generally not inactivated by β-lactamases. Since many of the strict anaerobes are β-lactamase producers, especially Bacteroides, Prevotella and some Fusobacterium species, 10 it is often preferable to use a non-β-lactam antimicrobial agent if that genus is suspected or include a β-lactamase inhibitor such as clavulanic acid, sulbactam, or tazobactam, in combination with a broadly effective β-lactam antimicrobial agent. 11 Co-amoxiclav (amoxicillin/clavulanic acid) was developed to enhance the activity of amoxicillin against β-lactamase-producing bacteria by the addition of clavulanic acid, an active β-lactamase inhibitor. In over 20 years of use as an orally administered antimicrobial agent, co-amoxiclav has maintained exceptional in vitro activity against respiratory tract pathogens. This study was conducted to provide susceptibility data for co-amoxiclav and five other antimicrobial agents against several important pathogenic anaerobic bacteria collected from 14 institutions in the USA and Europe. The antimicrobial agents tested included amoxicillin alone, to provide background comparison for amoxicillin without the β-lactamase inhibitor clavulanic acid. Any bacterial isolates that do not produce β-lactamase would be expected to have similar MIC data with amoxicillin and co-amoxiclav. Both metronidazole and clindamycin were chosen as comparators because of their historical efficacy against anaerobes, but also because of reports of increasing incidence of resistance. 12,13 Since fluoroquinolones and related drugs are often used in polymicrobial infections, levofloxacin was included to represent the group. 14,15 Finally, imipenem, a broad-spectrum carbapenem that is often used to treat anaerobic infections, was also included. Many of the drugs used as comparators in this study have also been used in several previously published surveillance and comparison reports. 16 19 Materials and methods We studied 1297 isolates that included 293 Bacteroides fragilis, 303 Prevotella species, 157 Fusobacterium species, 83 Eikenella corrodens, 92 Peptostreptococcus anaerobius, 146 Micromonas micros, 98 Finegoldia magna and 125 Porphyromonas species obtained from 14 institutions (eight centres throughout Europe and six in the USA, Table 1). Isolates were collected, from May 1999 to May 2000, primarily from blood cultures, bronchoalveolar lavage cultures, brush specimens, sinus aspirate or drainage, and pleural specimens. Wound, abscess, or tissue cultures were also used to ensure an adequate range of species. The Prevotella isolates tested included representatives from nearly all species of human clinical interest. Most of the Prevotella species tested were those found in the oral cavity but included representatives most commonly found in the female genital tract (Prevotella bivia and Prevotella disiens), which accounted for about 10% of the Prevotella isolates tested. The Porphyromonas isolates were mostly Porphyromonas asaccharolytica (46%) and Porphyromonas gingivalis (25%), but also included Porphyromonas levii, Porphyromonas endodontalis, and unspecified Porphyromonas spp. The following antimicrobial agents were tested: co-amoxiclav at a fixed clavulanic acid concentration of 2 mg/l (A/C 2), co-amoxiclav at a 2:1 amoxicillin/clavulanic acid concentration ratio (A/C 2:1), amoxicillin, 1040
Antimicrobial susceptibility in anaerobic and capnophilic isolates clindamycin, imipenem, levofloxacin, and metronidazole. Co-amoxiclav was tested in two formulations to account for the differences in testing methods in the USA and Europe. In the USA, the concentration of amoxicillin to clavulanic acid is tested at a ratio of 2:1, whereas in parts of Europe, the drug is tested with a fixed concentration of 2 mg/l clavulanic acid. A/C 2:1 was tested by the US centres and the German centre and A/C 2 was tested by the German and French centres. Susceptibility testing of the isolates was carried out at Université de Lille Laboratoire de Microbiologie Clinique (Lille, France), Institut für Medizinische Mikrobiologie und Infektionsepidemiologie der Universität Leipzig (Leipzig, Germany), M. S. Hershey Medical Center (Hershey, PA, USA), and R. M. Alden Research Laboratory (Santa Monica, CA, USA) by agar dilution. Agar dilutions were carried out according to current NCCLS guidelines 20 unless otherwise indicated. Each test concentration of antimicrobial agent was added to molten agar, which was mixed, poured into a Petri dish, and allowed to solidify. Standardized suspensions of isolates to be tested were inoculated onto the surface of each plate in the concentration series with the use of a replicating device. The inocula were allowed to absorb into the medium and placed in an anaerobic atmosphere within 30 min of inoculation. After plates were incubated anaerobically with an anaerobic indicator for approximately 48 h, the plates were examined visually. The lowest concentration of each antimicrobial agent that inhibited growth of an organism was reported as the MIC of the antimicrobial agent. Quality control strains as specified by the NCCLS guidelines 20 (Bacteroides fragilis ATCC 25285 and Bacteroides thetaiotaomicron ATCC 29741) were tested in each batch. Results for the study strains were accepted only if quality control results were within established NCCLS ranges. MIC 50 s, MIC 90 s and the percentages of isolates susceptible, intermediate and resistant were determined. Susceptibility rates were calculated using NCCLS interpretive criteria 20 and pharmacokinetic/pharmacodynamic (PK/PD) breakpoints for amoxicillin and levofloxacin. 21,22 The same NCCLS breakpoints were used for interpretation of A/C 2:1 and A/C 2 based on prior studies that reported no differences in MICs. 11,23,24 Results Results are summarized in Table 2. In vitro activities (based on percentage susceptibility) of all antimicrobial agents tested against the B. fragilis isolates, ranging from most active to least active, were: metronidazole (99.7%), imipenem (98.3%), A/C 2:1 (94.6%), clindamycin (79.5%), levofloxacin (72.7%) and amoxicillin (5.1%). The prevalence of B. fragilis resistance to amoxicillin/clavulanic acid (2:1) was 4.2%. Sixteen B. fragilis were resistant by either one or both co-amoxiclav methodologies (ratio of 2:1 or fixed clavulanic acid concentration of 2 mg/l). Of these 16 isolates, five were also resistant to imipenem, three were resistant to clindamycin, three had levofloxacin MICs of 32 mg/l, two had metronidazole MICs of 8 mg/l and included one isolate from France that was multi-resistant to imipenem, levofloxacin and clindamycin. The majority of these isolates (nine of 16) were isolated from one collection centre in Cardiff, UK. Clindamycin resistance rates were 23.5% in France, 17.7% in Germany, 12.6% in the USA and 2.2% in Cardiff, UK. Of the 42 clindamycin-resistant isolates, 11 had levofloxacin MICs 8 mg/l, two isolates were resistant to imipenem and three were resistant to co-amoxiclav. There were a total of five B. fragilis resistant to imipenem. All five of these isolates were also resistant to coamoxiclav, two were also resistant to clindamycin and two had levofloxacin MICs 8 mg/l. Two of the imipenem-resistant isolates were from France, two were from Hershey, PA, USA and one was from Cardiff, UK. Metronidazole resistance was rare. There were two isolates of B. fragilis from Cardiff, UK that were resistant (>256 mg/l) and both of these isolates were also resistant to coamoxiclav. One isolate from France had a metronidazole MIC of 8 mg/l. The Prevotella data were analysed to determine whether any species-specific or collecting centre-specific patterns of resistance were present that should preclude analysis of the genus as a whole. No significant pattern was discernable in the data, so the MIC 50, MIC 90 and susceptibility rates were not determined for each species. Of the isolates tested, there was one Prevotella oralis from France with an A/C 2 MIC of 32 mg/l and three Prevotella loeschii from France with an A/C 2 MIC of 8 mg/l. There were 20 isolates resistant to clindamycin (including six isolates from France and six from Cleveland, OH, USA). In addition, four isolates showed intermediate susceptibility to clindamycin. There were no other isolates with resistance to any of the other drugs tested. The only multidrug resistance found in Prevotella was in one isolate of P. loeschii, which had a clindamycin MIC of >256 mg/l and an A/C 2 MIC of 8 mg/l. All Prevotella isolates were susceptible to A/C 2:1, metronidazole and imipenem and 92.1% were susceptible to clindamycin. Amoxicillin activity (57.9% susceptible) was considerably less than co-amoxiclav activity, presumably due to β-lactamase production. Levofloxacin MICs for Prevotella were higher, although 71.5% of isolates were in the susceptible range. All agents were very active against Fusobacterium nucleatum. Amoxicillin was similar (±1 doubling dilution) in activity for most isolates to A/C 2 and A/C 2:1, but the MIC range was significantly wider (0.03 >128 versus 0.06 2 and 0.03 1). Only one isolate (from Santa Monica, CA, USA) was resistant to clindamycin, six isolates were resistant to amoxicillin and four isolates were resistant to levofloxacin. Clindamycin and metronidazole were not active against the E. corrodens isolates tested; the other agents tested showed good activity. All amoxicillin and co-amoxiclav MICs were 2 mg/l. Levofloxacin was also very active, with all isolates at 0.125 mg/l. Only one isolate from the Netherlands had an elevated imipenem MIC of 8 mg/l. Peptostreptococci, which have previously been examined in the aggregate as Peptostreptococcus species, were identified and analysed in this study as P. anaerobius, M. micros and F. magna. P. anaerobius was significantly different in susceptibility to coamoxiclav compared with the other two genera. Whereas M. micros and F. magna were all 100% susceptible to co-amoxiclav, 84.3% of P. anaerobius were susceptible. Amoxicillin was similar (±1 doubling dilution) in activity to co-amoxiclav for all three species. However, there were two isolates of M. micros from Leipzig with amoxicillin MICs of 128 mg/l and co-amoxiclav MICs of 0.125 mg/l. Differences in susceptibility were also present with clindamycin. Both P. anaerobius and M. micros were highly susceptible at 98.9% and 99.3%, respectively. F. magna had a relatively lower susceptibility rate of 84.7%. All peptostreptococci were susceptible to imipenem, with the exception of one P. anaerobius isolate from France (MIC of 8 mg/l). It is noteworthy, however, that the imipenem MIC 90 of 1 mg/l for P. anaerobius is higher than the MIC 90 s for M. micros and F. magna (0.06 and 0.125 mg/l, respectively). All P. anaerobius were susceptible to metronidazole. One M. micros and three F. magna from France were resistant to metronidazole. Levofloxacin was less active against F. magna. Levofloxacin MICs for 22.4% of F. magna were >4 mg/l, compared to 3.4% and 5.4% for M. micros and P. anaerobius, respectively. 1041
L. M. Koeth et al. Table 2. Summary of MIC results (MIC 50 s, MIC 90 s, MIC ranges and susceptibility rates) MIC (mg/l) Percentage Organism Antimicrobial agent n MIC 50 MIC 90 range susceptible intermediate resistant B. fragilis A/C 2 a 192 0.25 4 0.06 >128 91.1 1.6 7.3 A/C 2:1 b 259 0.5 4 0.125 128 94.6 1.2 4.2 amoxicillin 293 32 >128 0.125 >128 5.1 94.9 clindamycin 293 1 64 0.016 >256 79.5 6.8 13.7 imipenem 293 0.25 1 0.016 128 98.3 0.0 1.7 levofloxacin 293 2 8 0.125 >16 72.7 27.3 metronidazole 293 1 1 0.06 16 99.7 0.3 0.0 Prevotella spp. A/C 2 a 218 0.125 0.5 0.016 32 98.2 1.4 0.5 A/C 2:1 b 259 0.125 1 0.125 4 100.0 0.0 0.0 amoxicillin 303 1 64 0.016 >128 57.9 42.1 clindamycin 303 0.016 2 0.016 >256 92.1 1.3 6.6 imipenem 303 0.03 0.25 0.016 2 100.0 0.0 0.0 levofloxacin 303 1 8 0.06 64 71.5 28.5 metronidazole 303 0.5 2 0.06 >4 100.0 0.0 0.0 F. nucleatum A/C 2 a 46 0.125 0.5 0.06 2 100.0 0.0 A/C 2:1 b 144 0.125 0.25 0.03 1 100.0 0.0 0.0 amoxicillin 157 0.125 1 0.03 >128 95.5 3.2 clindamycin 157 0.06 0.125 0.016 8 99.4 0.0 0.6 imipenem 157 0.016 0.25 0.016 2 100.0 0.0 0.0 levofloxacin 157 0.5 2 0.06 >16 97.5 2.5 metronidazole 157 0.125 1 0.06 2 100.0 0.0 0.0 E. corrodens A/C 2 a 21 0.25 2 0.016 2 NA NA NA A/C 2:1 b 70 0.5 1 0.06 2 NA NA NA amoxicillin 83 1 2 0.06 32 NA NA NA clindamycin 83 >32 128 0.016 256 NA NA NA imipenem 83 0.125 0.25 0.016 8 NA NA NA levofloxacin 83 0.06 0.125 0.06 0.125 NA NA NA metronidazole 83 >32 >64 16 >64 NA NA NA P. anaerobius A/C 2 a 40 0.125 8 0.06 32 87.5 2.5 10.0 A/C 2:1 b 83 0.25 8 0.03 32 84.3 7.2 8.4 amoxicillin 92 0.25 16 0.06 32 74.2 25.8 clindamycin 92 0.06 0.25 0.016 32 98.9 0.0 1.1 imipenem 92 0.06 1 0.016 8 98.9 1.1 0.0 levofloxacin 92 0.5 1 0.125 32 92.5 7.5 metronidazole 92 0.5 1 0.06 2 100.0 0.0 0.0 F. magna A/C 2 a 40 0.125 0.5 0.06 1 100.0 0.0 A/C 2:1 b 90 0.25 0.5 0.125 1 100.0 0.0 0.0 amoxicillin 98 0.25 0.5 0.06 1 100.0 0.0 clindamycin 98 0.5 32 0.016 256 84.7 2.0 13.3 imipenem 98 0.125 0.125 0.016 0.5 100.0 0.0 0.0 levofloxacin 98 1 16 0.125 64 72.4 27.6 metronidazole 98 0.5 1 0.06 >64 96.9 0.0 3.1 M. micros A/C 2 a 78 0.125 0.125 0.06 1 100.0 0.0 A/C 2:1 b 142 0.125 0.5 0.03 2 100.0 0.0 0.0 amoxicillin 146 0.125 0.5 0.03 >128 97.3 2.1 clindamycin 146 0.125 0.5 0.016 8 99.3 0.0 0.7 imipenem 146 0.016 0.06 0.016 0.5 100.0 0.0 0.0 levofloxacin 146 0.5 2 0.125 >16 91.1 8.9 metronidazole 146 0.25 0.5 0.06 >64 99.3 0.0 0.7 Porphyromonas spp. A/C 2 a 66 0.125 0.125 0.06 0.5 100.0 0.0 A/C 2:1 b 119 0.125 0.125 0.03 0.5 100.0 0.0 0.0 amoxicillin 125 0.125 0.25 0.03 16 95.2 4.8 clindamycin 125 0.016 0.06 0.016 >32 96.8 0.0 3.2 imipenem 125 0.016 0.5 0.016 0.5 100.0 0.0 0.0 levofloxacin 125 0.5 2 0.125 8 92.0 8.0 metronidazole 125 0.125 0.5 0.06 2 100.0 0.0 0.0 A/C 2, co-amoxiclav at a fixed clavulanic acid concentration of 2 mg/l; A/C 2:1, co-amoxiclav at a 2:1 amoxicillin/clavulanic acid concentration ratio; NA, not available. Co-amoxiclav MICs are expressed in terms of amoxicillin concentration (mg/l). a Tested at German and French centres. b Tested at US and German centres. 1042
Antimicrobial susceptibility in anaerobic and capnophilic isolates The Porphyromonas isolates included P. asaccharolytica, P. endodontalis, P. levii, P. gingivalis, and Porphyromonas spp. All isolates were susceptible to co-amoxiclav, imipenem and metronidazole. Amoxicillin was similar (±1 doubling dilution) in activity to co-amoxiclav, with the exception of five isolates that had amoxicillin MICs of 8 16 mg/l and co-amoxiclav MICs of 0.125 0.5 mg/l. There were two P. asaccharolytica and two P. levii from Santa Monica, CA that were resistant to clindamycin (>32 mg/l). Although all isolates were susceptible to imipenem, it is noteworthy that onethird of MICs for isolates from Germany are >0.03 mg/l and all MICs for the other isolates are at or below 0.03 mg/l. Levofloxacin MICs for Porphyromonas showed moderate activity and 92.0% were within the susceptible range. There were seven isolates with levofloxacin MICs of 4 mg/l and three with MICs of 8 mg/l. With the exception of B. fragilis and Prevotella spp., the A/C 2 MICs correlated with the A/C 2:1 MICs. A total of 77.2% of B. fragilis with A/C 2 MICs 2 mg/l had lower MICs (by 1 4 dilutions) with A/C 2 compared to A/C 2:1. However, 56.5% of B. fragilis with A/C 2 MICs 4 mg/l had higher MICs (by 1 2 dilutions) with A/C 2 compared to A/C 2:1; 64.6% of Prevotella spp. with amoxicillin MICs 0.5 mg/l had lower MICs with A/C 2 compared to A/C 2:1. Discussion Most of the large surveillance studies in the past decade have examined Bacteroides species. Many of the studies have been of only B. fragilis, but the B. fragilis group is often also examined in aggregate. Dubreuil et al. 6 examined B. fragilis group isolates in France over the period 1977 1992 and found little or no change in rates of resistance to metronidazole, co-amoxiclav and imipenem. Resistance to clindamycin increased significantly over that period of time, stabilizing at 8 12%. Amoxicillin resistance was high in 1977 and continued to increase over time. Other studies 18,19,25 27 have reported B. fragilis and B. fragilis group isolates with increasing resistance to clindamycin and amoxicillin. Imipenem resistance has been reported at a range of 0 3% across several of these studies in the past decade. Metronidazole is reported as consistently active over time across all studies examined. In 1990, Appelbaum et al. 11 studied 320 non- B. fragilis B. fragilis group isolates from 28 US centres, and found that 9.2% of the 207 β-lactamase positive isolates were resistant to co-amoxiclav. Other studies 17 19,25 reported diminishing activity with clindamycin, but consistent high activity with metronidazole, regardless of β-lactamase status. The susceptibility data for B. fragilis in this study is consistent with prior published studies. Two previous papers 16,26 reported slightly increased imipenem MIC 90 s, but this was not found to be the case in this study. Co-amoxiclav has been a consistently active antimicrobial agent against Prevotella species, as have been metronidazole and imipenem. 16,18,19,26,28 Clindamycin resistance has been variable, with one study showing resistance of greater than 10%, 26 one with a low range of MIC 90 s ( 0.015 0.03 mg/l) across species but with an MIC range maximum of >128 mg/l, 16 and others showing full susceptibility. 19,28 Prevotella have been consistently susceptible to imipenem and metronidazole in the literature and this study. The genus has showed very little or no resistance to co-amoxiclav; however, some resistance to clindamycin exists, but was not found to be over 10% in this study. Fusobacterium are analysed and reported in the literature in aggregate as Fusobacterium species and as F. nucleatum. In a 1990 study by Appelbaum et al., 11 11.3% of β-lactamase-positive Fusobacterium species were resistant to co-amoxiclav and none of the isolates was resistant to metronidazole. Subsequent studies 16,17,29 continue to report no resistance to either metronidazole or imipenem. Studies specifically describing F. nucleatum 16 18,26 report high activity with co-amoxiclav, imipenem, metronidazole and clindamycin. The one study that included levofloxacin reported an MIC 90 of 0.5 mg/l (range 0.06 1 mg/l). 16 F. nucleatum susceptibility data in this study were largely consistent with that found in the literature. Two 1998 studies combined Porphyromonas isolates with other bacteria to report susceptibilities. Ednie et al. 17 examined a pool of 103 Porphyromonas and Prevotella species, finding no resistance to clindamycin and metronidazole (MIC 90 = 0.03 and 4 mg/l, respectively), but reporting maximum MICs at the intermediate breakpoint for both antimicrobials (4 and 16 mg/l, respectively). Edlund et al. 29 pooled Porphyromonas, Prevotella and Bacteroides species for a sample of 100 isolates. No resistance was reported to clindamycin or imipenem, and the MIC 90 of metronidazole was 1.0 mg/l, well below the susceptible breakpoint of 8 mg/l. In 1999, Goldstein et al. 16 reported no Porphyromonas resistant to co-amoxiclav, imipenem, or metronidazole, and high levofloxacin activity (MIC 90 = 0.5 mg/l). The MIC 90 reported for clindamycin was 128 mg/l, for a rate of resistance well over 10%. However, with a sample size of only 11, this could represent as few as two resistant isolates. The susceptibility data for Porphyromonas in this study were largely consistent with the 1999 Goldstein study, showing no resistance to co-amoxiclav or metronidazole and 3.2% resistance to clindamycin. There is a shift in imipenem MIC 90 and MIC max from 0.015 and 0.015 mg/l in the earlier study to 0.5 and 0.5 mg/l in this study. Peptostreptococci have often been examined in the aggregate as Peptostreptococcus species. 17,18,28,29 In these prior studies, F. magna and M. micros were still considered part of the Peptostreptococcus genus. The four studies consulted that examined the genus as a whole consistently reported no resistance to metronidazole. Clindamycin was also tested and reported by Aldridge & Johnson 19 to be fully active against the 17 isolates tested. In 1992, a study by Wexler et al. 18 included co-amoxiclav and reported 12% resistance. That study and the 1998 study by Edlund et al. 29 reported no resistance to imipenem. In 1998, Ednie et al. 17 reported resistance to clindamycin. The paper reported MIC 50/90 and range, with the MIC 90 at the intermediate breakpoint of 4 mg/l and MIC max at >32 mg/l. When the genus is evaluated by species, the sample sizes are often small, making comparisons statistically questionable. The 1999 Goldstein et al. 16 study reported on four species of Peptostreptococcus with sample sizes of 12 14. All species were susceptible to imipenem; F. magna, M. micros and P. anaerobius were all susceptible to clindamycin as well; and F. magna and M. micros were also susceptible to metronidazole and co-amoxiclav. Levofloxacin had high activity against M. micros. An earlier publication by Sheikh et al. 26 reported MICs for F. magna, M. micros and other Peptostreptococcus species. Imipenem was consistently active against all species tested. Metronidazole was active against F. magna, but M. micros and the other species showed some resistance. Clindamycin was the least active of the three antimicrobials tested, especially against M. micros. The major changes noted in the Peptostreptococcus/Finegoldia/Micros group in this study, compared with prior publications, are the prevalence of clindamycin resistance and the appearance of metronidazole resistance in F. magna. Also of note in this study were the significant differences seen among these three genera. In the clinical setting, it is recommended that these isolates are identified (as they are no longer grouped as peptostreptococci) in order to assure appropriate therapy for good outcomes. Clindamycin, for example, is one drug that could be used with some confidence to treat P. anaerobius or M. micros infections, but which should be used 1043
L. M. Koeth et al. more carefully in treating F. magna. And while F. magna and M. micros were 100% susceptible to co-amoxiclav, P. anaerobius had a susceptibility rate of 84.3%. There were some collecting centre differences in susceptibility that raise some questions about emerging resistance. Among B. fragilis, co-amoxiclav resistance was most prevalent in Cardiff, UK and the only metronidazole resistance was also observed in Cardiff. In contrast, the clindamycin resistance among B. fragilis was relatively low in Cardiff and much higher in France. The highest prevalence of elevated co-amoxiclav MICs for Prevotella was observed in France. Among Porphyromonas spp., the only clindamycin resistance was observed in Santa Monica, CA, USA and a slight shift in imipenem MICs was observed in Germany. Nine metronidazole-resistant peptostreptococci (including isolates identified as Peptostreptococcus spp., F. magna and M. micros) were also isolated from France. It would be instructive to attempt to determine whether the rates of clinical treatment failure are likewise increasing, a project perhaps to be undertaken through national registries and beyond the scope of this paper. The apparent shift in the metronidazole MIC curve against Porphyromonas also bears watching, as this, too, might represent emergence of resistance. The comparatively large sample sizes used in this study allow some statistical confidence in the results. The lack of similarly large numbers in many of the older published studies makes longitudinal comparisons difficult, and underscores the importance of large surveillance studies. The appearance of possible shifts in MIC curves may provide advance notice of emerging resistance and should be followed closely and correlated with clinical outcomes. Acknowledgements This work was supported by a project grant from SmithKline Beecham. References 1. Goldstein, E. J. C. (1996). Anaerobic bacteremia. Clinical Infectious Diseases 23, Suppl. 1, S97 101. 2. Feleke, G. & Forlenza, S. (1991). Anaerobic infections: the basics for primary care physicians. Postgraduate Medicine 89, 221 34. 3. Finegold, S. M. & Wexler, H. M. (1996). Present studies of therapy for anaerobic infections. Clinical Infectious Diseases 23, Suppl. 1, S9 14. 4. Sanders, C. V. & Aldridge, K. E. (1992). Current antimicrobial therapy of anaerobic infections. European Journal of Clinical Microbiology and Infectious Diseases 11, 999 1011. 5. Lee, K., Chong, Y., Jeong, S. H. et al. (1996). Emerging resistance of anaerobic bacteria to antimicrobial agents in South Korea. Clinical Infectious Diseases 23, Suppl. 1, S73 7. 6. Dubreuil, L., Breuil, J., Dublanchet, A. et al. (1992). Survey of the susceptibility patterns of Bacteroides fragilis group strains in France from 1977 to 1992. European Journal of Clinical Microbiology and Infectious Diseases 11, 1094 9. 7. Baquero, F. & Reig, M. (1992). Resistance of anaerobic bacteria to antimicrobial agents in Spain. European Journal of Clinical Microbiology and Infectious Diseases 11, 1016 20. 8. Finegold, S. M. (1997). Perspective on susceptibility testing of anaerobic bacteria. Clinical Infectious Diseases 25, Suppl. 2, S251 3. 9. Hardman, J. G. & Limbird, L. E. (1996). Goodman & Gilman s The Pharmacological Basis of Therapeutics, 9th edn. McGraw-Hill Professional, New York, NY, USA. 10. Nord, C. E. (1986). Mechanisms of β-lactam resistance in anaerobic bacteria. Review of Infectious Diseases 8, Suppl. 5, S543 8. 11. Appelbaum, P. C., Spangler, S. K. & Jacobs, M. R. (1990). β-lactamase production and susceptibilities to amoxicillin, amoxicillinclavulanate, ticarcillin, ticarcillin-clavulanate, cefoxitin, imipenem, and metronidazole of 320 non-bacteroides fragilis Bacteroides isolates and 129 fusobacteria from 28 U.S. centers. Antimicrobial Agents and Chemotherapy 34, 1546 50. 12. Rasmussen, B. A., Bush, K. & Tally, F. P. (1997). Antimicrobial resistance in anaerobes. Clinical Infectious Diseases 24, Suppl. 1, S110 20. 13. Jones, R. N. & the MYSTIC Advisory Board. (2000). Detection of emerging resistance patterns within longitudinal surveillance systems: data sensitivity and microbial susceptibility. Journal of Antimicrobial Chemotherapy 46, Topic T2, 1 8. 14. Stein, G. E., Schooley, S., Tyrrell, K. L. et al. (2003). Bactericidal activities of methoxyfluoroquinolones gatifloxacin and moxifloxacin against aerobic and anaerobic respiratory pathogens in serum. Antimicrobial Agents and Chemotherapy 47, 1308 12. 15. Stein, G. E. & Goldstein, E. J. C. (2003). Review of the in vitro activity and potential clinical efficacy of levofloxacin in the treatment of anaerobic infections. Anaerobe 9, 75 81. 16. Goldstein, E. J. C., Citron, D. M., Warren, Y. et al. (1999). In vitro activity of gemifloxacin (SB 265805) against anaerobes. Antimicrobial Agents and Chemotherapy 43, 2231 5. 17. Ednie, L. M., Jacobs, M. R. & Appelbaum, P. C. (1998). Activities of gatifloxacin compared to those of seven other agents against anaerobic organisms. Antimicrobial Agents and Chemotherapy 42, 2459 62. 18. Wexler, H. M., Molitoris, E. & Finegold, S. M. (1992). In vitro activities of three of the newer quinolones against anaerobic bacteria. Antimicrobial Agents and Chemotherapy 36, 239 43. 19. Aldridge, K. E. & Johnson, W. D. (1997). A comparison of susceptibility results of the Bacteroides fragilis group and other anaerobes by traditional MIC results and statistical methods. Journal of Antimicrobial Chemotherapy 39, 319 24. 20. National Committee for Clinical Laboratory Standards. (2001). Methods for Antimicrobial Susceptibility Testing of Anaerobic Bacteria Fifth Edition: Approved Standard M11-A5. NCCLS, Villanova, PA, USA. 21. Craig, W. A. (1998). Pharmacokinetic/pharmacodynamic parameters: rationale for antibacterial dosing of mice and men. Clinical Infectious Diseases 26, 1 10. 22. Jacobs, M. R. (2001). Optimisation of antimicrobial therapy using pharmacokinetic and pharmacodynamic parameters. Clinical Microbiology and Infection 7, 589 96. 23. Jacobs, M. R., Spangler, S. K. & Appelbaum, P. C. (1990). Susceptibility of Bacteroides non-fragilis and fusobacteria to amoxicillin, amoxicillin/clavulanate, ticarcillin, ticarcillin/clavulanate, cefoxitin, imipenem and metronidazole. European Journal of Clinical Microbiology and Infectious Diseases 9, 417 21. 24. Jacobs, M. R., Spangler, S. K. & Appelbaum, P. C. (1990). β-lactamase production, β-lactam sensitivity and resistance to synergy with clavulanate of 737 Bacteroides fragilis group organisms from thirtythree US centres. Journal of Antimicrobial Chemotherapy 26, 361 70. 25. Cuchural, G. J., Tally, F. P., Jacobus, N. V. et al. (1990). Comparative activities of newer β-lactam agents against members of the Bacteroides fragilis group. Antimicrobial Agents and Chemotherapy 34, 479 80. 26. Sheikh, W., Pitkin, D. H. & Nadler, H. (1993). Antibacterial activity of meropenem and selected comparative agents against anaerobic bacteria at seven North American centers. Clinical Infectious Diseases 16, Suppl. 4, S361 6. 27. Betriu, C., Sanchez, A., Gomez, M. et al. (1999). In-vitro susceptibilities of species of the Bacteroides fragilis group to newer β-lactam agents. Journal of Antimicrobial Chemotherapy 43, 133 6. 28. Dubreuil, L., Singer, E., Odou, M. F. et al. (1999). Antianaerobic activity of grepafloxacin compared with ofloxacin, ciprofloxacin, clindamycin, metronidazole and 5 β-lactams. Drugs 58, Suppl. 2, 110 2. 29. Edlund, C., Sabouri, S. & Nord, C. E. (1998). Comparative in vitro activity of BAY 12-8039 and five other antimicrobial agents against anaerobic bacteria. European Journal of Clinical Microbiology and Infectious Diseases 17, 193 5. 1044