Skip to main page content

• HOME
• Current Issue
• Archive
• Alerts
• About ASM
• Contact us
• Tech Support
• Journals.ASM.org

This Article Abstract Full Text (PDF) Alert me when this article is cited Alert me if a correction is posted Citation Map Services E-mail this article to a friend Similar articles in this journal Similar articles in PubMed Alert me to new issues of the journal Download to citation manager Reprints and Permissions Copyright Information Books from ASM Press MicrobeWorld Citing Articles Citing Articles via Google Scholar Google Scholar Articles by Kosowska-Shick, K. Articles by Appelbaum, P. C. Search for Related Content PubMed PubMed Citation Articles by Kosowska-Shick, K. Articles by Appelbaum, P. C.

 Previous Article  |  Next Article 

Antimicrobial Agents and Chemotherapy, June 2009, p. 2239-2247, Vol. 53, No. 6
0066-4804/09/$08.00+0     doi:10.1128/AAC.01531-08
Copyright © 2009, American Society for Microbiology. All Rights Reserved.

Comparative Antipneumococcal Activities of Sulopenem and Other Drugs

Klaudia Kosowska-Shick, Lois M. Ednie, Pamela McGhee, and Peter C. Appelbaum^*

Hershey Medical Center, Hershey, Pennsylvania 17033

Received 17 November 2008/ Returned for modification 9 March 2009/ Accepted 13 March 2009

Top ABSTRACT INTRODUCTION MATERIALS AND METHODS RESULTS DISCUSSION REFERENCES

 
For 297 penicillin-susceptible, -intermediate, and -resistant pneumococcal strains, the sulopenem MIC₅₀s were 0.008, 0.06, and 0.25, respectively, and the sulopenem MIC₉₀s were 0.016, 0.25, and 0.5 µg/ml, respectively. The MIC₅₀s of amoxicillin for the corresponding strains were 0.03, 0.25, and 2.0 µg/ml, respectively, and the MIC₉₀s were 0.03, 1.0, and 8.0 µg/ml, respectively. The combination of amoxicillin and clavulanate gave MICs similar to those obtained with amoxicillin alone. The sulopenem MICs were similar to those of imipenem and meropenem. The MICs of ß-lactams increased with those of penicillin G, and among the quinolones tested, moxifloxacin had the lowest MICs. Additionally, 45 strains of drug-resistant type 19A pneumococci were tested by agar dilution and gave sulopenem MIC₅₀s and MIC₉₀s of 1.0 and 2.0 µg/ml, respectively. Both sulopenem and amoxicillin (with and without clavulanate) were bactericidal against all 12 strains tested at 2x MIC after 24 h. Thirty-one strains from 10 countries with various penicillin, amoxicillin, and carbapenems MICs, including those with the highest sulopenem MICs, were selected for sequencing analysis of the pbp1a, pbp2x, and pbp2b regions encoding the transpeptidase active site and MurM. We did not find any correlations between specific penicillin-binding protein-MurM patterns and changes in the MICs.

Top ABSTRACT INTRODUCTION MATERIALS AND METHODS RESULTS DISCUSSION REFERENCES

 
The incidence of pneumococci resistant to penicillin G and other ß-lactam and non-ß-lactam compounds has increased at an alarming rate worldwide, including the United States. The major foci of infections currently include South Africa, Spain, and Central and Eastern Europe (26, 27). A 1997 survey showed that 50.4% of 1,476 clinically significant pneumococcal isolates in the United States were not susceptible to penicillin (26). Of all strains tested, approximately 33% were macrolide resistant, with the highest rate of macrolide resistance being seen among penicillin-resistant strains, for which the MICs were 2.0 µg/ml (26). A later survey, conducted as part of the Alexander Project (27), reported that the worldwide prevalence of pneumococci isolated between 1998 and 2000 for which penicillin G MICs were 2 µg/ml was 18.2%, with the overall macrolide resistance rate being 24.6%. It is also important to note the higher rates of isolation of penicillin-intermediate and -resistant pneumococci (approximately 30%) in middle ear fluids from patients with refractory otitis media than from other isolation sites (4). Quinolone-resistant pneumococci have also been described (6, 22, 31). The problem of drug-resistant pneumococci is compounded by the ability of resistant clones to spread from country to country and from continent to continent (32, 34). In a recent paper, Pichichero and Casey (40) reported on the emergence in the United States of an otopathogenic strain of pneumococcus type 19A which is not covered by the current pediatric conjugate vaccine and which is resistant to all currently FDA-approved antibiotics for the treatment of acute otitis media in children. Another very recent study has shown that of 393 isolates of pneumococci collected from children in the United States between 2005 and 2006, 30.5% of all the isolates were serotype 19A; >50% of the serotype 19A strains showed multiresistant susceptibility patterns (9).

ß-Lactam resistance in Streptococcus pneumoniae is mediated by stepwise alterations of penicillin-binding proteins (PBPs) that result in decreased antibiotic affinities (21). Pneumococci contain a set of six PBPs. The decreased affinities of PBP 1A, 2X, and 2B for β-lactams have been reported to play an important part in resistance and to cause high-level penicillin G resistance (1, 2, 30, 41, 42, 51). Alterations in the conserved motifs in PBP 2B are associated with penicillin G resistance, and alterations in PBP 2X mediate low-level resistance to cephalosporins (1, 8, 11, 18). The region of PBP 2B from amino acids 538 to 642 also appears to be relevant for resistance, especially among strains with high amoxicillin MICs (5, 29). Additional alterations in PBP 1A raise the penicillin G and cefotaxime MICs (1, 35, 42, 47). These specific PBPs are thought to be the key to resistance to each antibacterial agent. Non-PBP-related resistance determinants are also essential for the development of high-level penicillin G and cephalosporin resistance in pneumococcal isolates. The mechanism used by non-PBP-related resistance determinants involves alterations in MurM, an enzyme involved in the biosynthesis of branched-stem cell wall muropeptides (14).

The stated need for other ß-lactam and non-ß-lactam compounds active against resistant pneumococcal strains isolated from patients with community-acquired respiratory tract infections is as current as ever (17). Amoxicillin is currently the ß-lactam with the most potent activity against penicillin-susceptible and -resistant pneumococci. In combination with clavulanate, amoxicillin also provides excellent coverage against ß-lactamase-positive Haemophilus influenzae and Moraxella catarrhalis strains (26, 27).

PF-03709270 is a novel oral prodrug of sulopenem, a parenteral penem antibiotic (Fig. 1) (16). The current study tested (i) the activities of penicillin, amoxicillin, amoxicillin-clavulanate, imipenem, meropenem, sulopenem, ceftriaxone, cefuroxime, cefpodoxime, cefdinir, ciprofloxacin, levofloxacin, gatifloxacin, moxifloxacin, azithromycin, and clarithromycin against 297 pneumococci and 45 serotype 19A pneumococci resistant to all available oral agents (9, 40) by the agar dilution MIC determination method; (ii) the activities of amoxicillin, amoxicillin-clavulanate, imipenem, meropenem, sulopenem, ertapenem, ceftriaxone, cefuroxime, cefpodoxime, cefdinir, ciprofloxacin, levofloxacin, gatifloxacin, moxifloxacin, azithromycin, and telithromycin against 12 selected pneumococci by time-kill analysis; (iii) whether alterations in PBPs 2B, 2X, and 1A in combination with mutations in the murM gene and specific patterns of the PBP 2B region (amino acids 557 to 647) correlate with resistance to sulopenem and other ß-lactam antibiotics among selected pneumococcal strains; and (iv) the identities of the pneumococcal clones with the highest sulopenem MICs (1 µg/ml) by multilocus sequence typing (MLST).

View larger version (8K): [in this window] [in a new window]

FIG. 1. Sulopenem structure.

Top ABSTRACT INTRODUCTION MATERIALS AND METHODS RESULTS DISCUSSION REFERENCES

 
Two hundred ninety-seven selected pneumococcal strains were tested by the agar dilution MIC method. These comprised 80 penicillin-susceptible strains (MICs, 0.06 µg/ml), 88 penicillin-intermediate strains (MICs, 0.125 to 1.0 µg/ml), and 129 penicillin-resistant strains (MICs, 2.0 to 16.0 µg/ml). Of these strains, 152 were macrolide resistant (azithromycin MICs, 1.0 µg/ml) and had defined macrolide resistance mechanisms. Additionally, 30 strains with levofloxacin MICs of 4.0 µg/ml and defined mutations in type II topoisomerase were tested. Strains were selected to include a majority which had penicillin G MICs of 2.0 µg/ml. Some strains were macrolide susceptible but penicillin resistant; this does not reflect the current situation in the United States. We also tested an additional 45 strains of drug-resistant type 19A pneumococci (provided by S. Brown and R. Jones) by the agar dilution MIC method. Among the 12 strains tested by time-kill analysis, 2 were penicillin susceptible, 5 were penicillin intermediate resistant, and 5 were penicillin resistant; 6 strains were macrolide resistant: 3 because of erm(B) mutations, 2 because of mef(A) mutations, and 1 because of an L4 ribosomal protein mutation. Two strains were also quinolone resistant and had defined changes in their quinolone resistance-determining region. Thirty-one isolates were selected from among the 297 strains on the basis of the following criteria: all strains with the highest sulopenem MIC of 1 µg/ml (n = 9), 8 strains with amoxicillin MICs equal to their penicillin G MICs, 8 strains with amoxicillin MICs greater than their penicillin G MICs, and 6 strains with penicillin G MICs greater than their amoxicillin MICs. This collection comprised 28 penicillin-resistant S. pneumoniae strains (MICs, 2.0 µg/ml) and 3 penicillin-intermediate resistant S. pneumoniae strains (MICs, 1.0 µg/ml) (see Table 5). Identification of the strains as S. pneumoniae was based on optochin susceptibility, bile solubility, the presence of the lytA gene, and sequencing of the 16S rRNA gene (36). All 31 strains were analyzed by pulsed-field gel electrophoresis (PFGE) to determine clonal relatedness. MLST of all strains (n = 9) with sulopenem MICs of 1 µg/ml and one with an MIC of 0.25 µg/ml was performed as described previously (13).

View this table: [in this window] [in a new window]

TABLE 5. MICs, PFGE types, and MLST types for strains tested for β-lactam resistancea

Fragments of the pbp1A gene (the region from nucleotides 870 to 1950) encoding 350 amino acids, the pbp2B gene (the region from nucleotides 655 to 2028) encoding 458 amino acids, and the pbp2X gene (the region from nucleotides 301 to 2034) encoding 578 amino acids were amplified from chromosomal DNA by PCR with the primers and under the conditions described previously and were directly sequenced (CEQ8000 genetic analysis system; Beckman Coulter, Fullerton, CA) (29). The sequences obtained were analyzed with the BLAST and ClustalW programs (46). The PBP alleles were identified by comparison with the sequences of the analyzed gene from strain R6 and named with a letter of the alphabet on the basis of the degree of homology to the strain R6 sequence; e.g., allele A has the highest degree of homology to the allele in the R6 sequence, and allele L has the lowest degree of homology to the allele in the R6 sequence. The murM gene was amplified from chromosomal DNA by PCR with the primers and under the conditions described previously and was directly sequenced (14). Subsequently, additional primers were designed for amplification of the murein protein variant 149193/50012, as follows: primers mur75up (5'-ATTACAAAGTAGTGCTTGGG) and mur608dn (5'-CGCTTCTCAGTTTTTTTCATCAA) or primers 1491mur135up (5'-CTATGAAGAGGGGAAGTTACTGGCTGTGGCT) and 1491mur549dn (5'-ACCAAATTGAATCTCTACACCCTTA) (43).

Sulopenem susceptibility powder was obtained from Pfizer Central Research, Groton, CT. The other antimicrobials were obtained from their respective manufacturers. The agar dilution method was performed with Mueller-Hinton agar (Becton Dickinson, Sparks, MD) supplemented with 5% sheep blood and inocula of 10⁴ CFU/spot, as previously described by our group (23, 37-39, 44, 45, 48, 49). Clavulanate was combined with amoxicillin in a 1:2 ratio. Standard quality control strains, including S. pneumoniae ATCC 49619, were included in each run of the agar dilution MIC determinations (7).

For the time-kill studies, methods previously described by our group were used (23, 48, 49). The medium was cation-adjusted Mueller-Hinton broth containing 5% lysed horse blood, and the inocula were 5 x 10⁵ to 5 x 10⁶ CFU/ml. The suspensions were incubated at 35°C in a shaking water bath; and viability counts were done after 0, 3, 6, 12, and 24 h. Drugs were considered bactericidal if the original inoculum was reduced by 3 log₁₀ CFU/ml (99.9%) at the lowest concentration tested during each of the time periods and bacteriostatic if the inoculum was reduced by 0 to <3 log₁₀ CFU/ml. Drug carryover was addressed by dilution, as reported previously (23, 48, 49).

Nucleotide sequence accession numbers. All sequences of pbp1a, pbp2X, and pbp2B obtained were deposited in the GenBank database and had the following accession numbers: EU863659 to EU863689 for pbp2B, EU863690 to EU863720 for pbp2X, and EU863721 to EU863751 for pbp1A.

Top ABSTRACT INTRODUCTION MATERIALS AND METHODS RESULTS DISCUSSION REFERENCES

 
The results of the MIC testing of 297 strains can be seen in Table 1. For the 297 penicillin-susceptible, -intermediate, and -resistant pneumococcal strains, sulopenem gave MIC₅₀s of 0.008, 0.06, and 0.25, respectively, and MIC₉₀s of 0.016, 0.25, and 0.5 µg/ml, respectively. The MIC₅₀s of amoxicillin for the corresponding strains were 0.03, 0.25, and 2.0 µg/ml, respectively, and the MIC₉₀s were 0.03, 1.0, and 8.0 µg/ml, respectively. The combination of amoxicillin and clavulanate gave MICs similar to those obtained with amoxicillin alone. The sulopenem MICs were similar to those of imipenem and meropenem. The sulopenem MICs were similar to those of imipenem and meropenem and lower than those of ceftriaxone. All oral cephalosporins tested had MICs higher than those of amoxicillin. The results for the 45 strains of drug-resistant type 19A pneumococci are presented in Table 2. The sulopenem MIC₅₀s and MIC₉₀s for these 45 strains were 1.0 and 2.0 µg/ml, respectively, and the amoxicillin MIC₅₀s and MIC₉₀s for these 45 strains were 8.0 and 8.0 µg/ml, respectively. As can be seen, these strains were highly resistant to all available oral antipneumococcal compounds approved for pediatric use.

View this table: [in this window] [in a new window]

TABLE 1. Agar dilution MICs for 297 strains

View this table: [in this window] [in a new window]

TABLE 2. Agar dilution MICs for 45 drug-resistant type 19A pneumococci

The MICs of the 12 strains tested by time-kill analysis are presented in Table 3, and the results of the time-kill analyses are presented in Table 4. As can be seen, all ß-lactams were bactericidal (99.9% killing) against all 12 strains tested at 2x MIC after 24 h and against 8 to 11 strains (the range for the different ß-lactams tested) at 2x MIC after 12 h. All four quinolones were bactericidal at 2x MIC against all 10 strains tested after 24 h; azithromycin was bactericidal against all 8 strains tested, and telithromycin at 2x MIC was bactericidal against 8 of 12 strains after 24 h.

View this table: [in this window] [in a new window]

TABLE 3. MICs for 12 strains tested by time-kill analysis

View this table: [in this window] [in a new window]

TABLE 4. Results of killing kinetics

Nine PFGE types (five types represented by 2 closely related strains, three types represented by 3 closely related strains, and one type represented by 6 closely related strains) were found among the 31 isolates characterized by PFGE, and 6 strains had unique patterns (Table 5). Eight sequence types (STs) were found among 10 selected isolates (9 with the highest sulopenem MIC [1 µg/ml] and 1 with the lowest sulopenem MIC [0.125 µg/ml] in the collection of 10 strains) (Table 5).

The amino acid alterations of the three conserved motifs of PBP 1A, 2X, and 2B are presented in Table 6. During sequence comparisons, we noticed that the sequence numbering for PBP 2B was off by 6 amino acids compared to the original sequence of S. pneumoniae R6 (GenBank accession number AE008520.1) (24).

View this table: [in this window] [in a new window]

TABLE 6. Amino acid alterations of the three conserved motifs of PBPs 1A, 2X, and 2B in 31 tested strains compared to S. pneumoniae R6 sequence and MurM and PBP allele attributions

Previous reports have described PBP 2B motifs which were off by the 6 amino acids described above, and because of that we have retained the old amino acid numbering to be able to correlate mutations (10, 12, 29, 41).

Analysis of PBP 1A showed that 7 strains (penicillin G MICs, 1 to 4 µg/ml; amoxicillin MICs, 1 to 8 µg/ml) had T₃₇₁A substitutions in the ₃₇₀STMK₃₇₃ motif, 7 strains (penicillin G MICs, 2 to 8 µg/ml; amoxicillin MICs, 0.25 to 4 µg/ml) had T₃₇₁S substitutions in the ₃₇₀STMK₃₇₃ motif, and 14 strains had P₄₃₂T substitutions (accompanied by T₃₇₁A in 2 strains and with a T₃₇₁S mutation in ₃₇₀STMK₃₇₃ in all 14 strains) close to the ₄₂₈SRN₄₃₀ motif of PBP 1A. PBP 1A was present in 10 different variants (variants A1 to J1) (Table 6).

Ten isolates showed a single PBP 2X mutation, T₃₃₈A, in the ₃₃₇STMK₃₄₀ motif and had penicillin G MICs of 1 to 16 µg/ml and amoxicillin of MICs 1 to 8 µg/ml, and 10 strains showed the T₃₃₈P substitution and had penicillin G MICs of 1 to 8 µg/ml and amoxicillin MICs of 0.25 to 8 µg/ml. Eight strains had ₃₃₈TM_{339 338}AF₃₃₉ mutations and had penicillin G MICs of 2 to 8 µg/ml and amoxicillin MICs of 2 to 16 µg/ml. Fourteen of the 31 strains had an L₅₄₆V substitution close to the ₅₄₇KSG₅₄₉ motif of PBP 2X (penicillin G MICs, 1 to 16 µg/ml; amoxicillin MICs, 1 to 8 µg/ml). No change in the ₃₈₅SVVK₃₈₈ motif of PBP 2X was noted. PBP 2X showed the presence of 16 different variants (variants A3 to P3) (Table 6).

All strains showed the same substitution T₄₄₅A in the ₄₄₂SSNT₄₄₅ motif in PBP 2B and no change in the ₃₈₅SVVK₃₈₈ motif. Sequence analysis of PBP 2B yielded 12 different alleles (alleles A2 to L2) (Table 6). One strain (strain 3374) had an insertion ₄₂₆YTQ₄₂₇ (₄₃₂YTQ₄₃₃) in PBP 2B. The insertion of three amino acids (₄₂₅WYT₄₂₆) has been described previously, and authors have speculated that such alterations may have been responsible for the altered PBP 2B and may have been involved as one of the determinants of penicillin resistance (50).

Six groups were distinguished on the basis of the specific mutation patterns in 90 amino acid fragments from amino acid positions 557 to 647 (Fig. 2).

View larger version (9K): [in this window] [in a new window]

FIG. 2. Comparison of the sequence of PBP 2B (region from amino acid positions 557 to 647 of the deduced amino acid sequence) in all strains tested compared to the PBP 2B sequence from strain R6 of S. pneumoniae.

The murM gene was sequenced from 31 isolates and yielded six different alleles. The deduced amino acid sequences represented six previously published MurM variants: MurMA (14 strains), MurMB6 (5 strains), MurMB1 (5 strains), 149193/5002 (which has 98% homology to allele MurMB1; 3 strains), and MurMB7 and MurMB4 (2 strains each) (Table 6).

Top ABSTRACT INTRODUCTION MATERIALS AND METHODS RESULTS DISCUSSION REFERENCES

 
PF-03709270 is an oral prodrug of sulopenem, an experimental penem (16). In a study of the comparative activities of sulopenem and other drugs against a collection of recently isolated gram-positive and -negative organisms, the sulopenem MIC₉₀s ranged from 0.03 to 1 µg/ml against all clinically significant bacterial species tested. This high in vitro potency was also confirmed by in vitro time-kill studies (25). Our studies confirm the activity of sulopenem against 114 pneumococci of various resistance phenotypes described previously (25) and expand on the information on the activity of sulopenem available.

The results of this study indicate that the combination of amoxicillin and clavulanate did not influence the amoxicillin MICs against all pneumococcal groups, and among all oral ß-lactams tested, amoxicillin had the lowest MIC. Sulopenem, imipenem, meropenem, and ceftriaxone all had low MICs against the pneumococci tested, with the oral cephalosporins having higher MICs. Sulopenem in particular was very active, with the MICs ranging from 0.004 to 2.0 µg/ml, even against drug-resistant pneumococci with very high penicillin G and amoxicillin MICs. The MICs of the ß-lactams rose with those of penicillin G, and moxifloxacin was the most active quinolone tested (23, 37-39, 44, 45, 48, 49) Against the drug-resistant type 19A pneumococci, sulopenem had MICs lower than those of amoxicillin whether amoxicillin was tested with or without clavulanate. The MICs of sulopenem for these 45 strains were similar to those of imipenem and meropenem. Amoxicillin, with and without clavulanate, and sulopenem showed excellent killing kinetics relative to their MICs, with bactericidal activity against all 12 strains at 2x MIC being achieved after 24 h. Although azithromycin and telithromycin are commonly described to be bacteriostatic against pneumococci, they are, in reality, slowly bactericidal (as it is defined) at multiples of the MIC after 24 h (23, 48, 49). The clinical significance of this phenomenon awaits clarification.

Among the PBPs produced by S. pneumoniae, PBPs 1A, 2B, and 2X are the most important in the development of resistance. Other mechanisms, such as altered protein kinase CiaH and glycosyltransferase CpoA, have been described as potential mechanisms of resistance; but these aspects were not investigated in the current study (19, 20, 28).

Elevated sulopenem MICs have not previously been associated with any MLST pattern. We found that MLST analysis (10 strains) showed a correlation only between the ST and the country of origin. All defined STs, STs 41, 1094, 663 and 610, and 603 (isolated in the United States and South Africa, Russia, and Poland, respectively), have already been described in the MLST database and have previously been isolated in the countries mentioned above. ST81 represents global S. pneumoniae clone 23F-Spain, which has spread in many countries (33).

Among the PBP alleles analyzed, the most variable was PBP 2X, followed by PBP 2B and PBP 1A, with homologies at the protein level with the sequence of strain R6 of 95 to 87%, 96 to 90%, and 93 to 84%, respectively. A similar correlation has been described by Biçmen et al. (3) and del Campo et al. (10).

Altered stem peptide (mur) genes can also affect the activities of β-lactams (15, 43). Six types of MurM were detected in our study, but no correlation to the increased MICs of any antibiotic tested was detected. Each individual clonal group had the same MurM allele (the exceptions being strains 3455 and 3458). Our studies confirm that the role of murM in resistance cannot be explained at this time as a simple correlation between the presence of an altered MurM protein and resistance to ß-lactams: strains with high-level resistance to penicillin G had unaltered MurM proteins, and also, strains with identical PBP patterns but different MurM alleles had no changes in MICs. In general, our data support the hypothesis by du Plessis et al. that the involvement of MurM in penicillin resistance appears to be dependent on specific mutations in PBPs, especially in PBP 2B (12).

In our study, the region of PBP 2B from amino acids 557 to 647 appeared to be relevant for resistance, especially among strains with high amoxicillin MICs, which confirms previous findings (5, 12, 29).

All mutations previously reported to affect β-lactam resistance (e.g., PBP 1A with a T₃₇₁A or S substitution within the ₃₇₀STMK₃₇₃ motif, a T₃₃₈S or A substitution in PBP 2X, and a T₄₄₅A substitution in the ₄₄₂SSNT₄₄₅ motif of PBP 2B) were found in all isolates tested in the current study, but we were unable to find a clear correlation between those alterations and specific patterns of resistance, including susceptibility to sulopenem (1, 8, 41, 42). This may suggest that at least one change in crucial PBP motifs may be required to develop β-lactam resistance.

The results of the present study indicate that sulopenem (administered either parenterally or as the oral prodrug) has a potential place in the treatment of infections caused by drug-resistant pneumococci, including the drug-resistant nonvaccine type 19A phenotype (40), which is probably spreading throughout all countries in which the vaccine is available. The only current therapeutic options for the latter strains are linezolid or the respiratory quinolones. The clinical use of sulopenem must await the results of pharmacokinetic/pharmacodynamic analyses, as well as data from toxicological, safety, experimental animal, and clinical efficacy studies.

 
This study was supported by a grant from Pfizer Central Research, Groton, CT.

We thank S. Brown (Clinical Microbiology Institute, Wilsonville, OR) and R. Jones (JMI Laboratories, North Liberty, IA) for the provision of drug-resistant type 19A strains.

 
* Corresponding author. Mailing address: Department of Pathology, Hershey Medical Center, P.O. Box 850, Hershey, PA 17033. Phone: (717) 531-5113. Fax: (717) 531-7953. E-mail: pappelbaum{at}psu.edu

Published ahead of print on 23 March 2009.

Top ABSTRACT INTRODUCTION MATERIALS AND METHODS RESULTS DISCUSSION REFERENCES

 

1
1. Asahi, Y., Y. Takeuchi, and K. Ubukata. 1999. Diversity of substitutions within or adjacent to conserved amino acid motifs of penicillin-binding protein 2X in cephalosporin-resistant Streptococcus pneumoniae isolates. Antimicrob. Agents Chemother. 43:1252-1255.[Abstract/Free Full Text]
2
2. Barcus, V. A., K. Ghanekar, M. Yeo, T. J. Coffey, and C. G. Dowson. 1995. Genetics of high level penicillin resistance in clinical isolates of Streptococcus pneumoniae. FEMS Microbiol. Lett. 126:299-303.[CrossRef][Medline]
3
3. Biçmen, M., Z. Gülay, D. Ramaswamy, M. Musher, and M. Gur. 2006. Analysis of mutations in the pbp genes of penicillin-non-susceptible pneumococci from Turkey. Clin. Microbiol. Infect. 12:150-155.[CrossRef][Medline]
4
4. Block, S., C. J. Harrison, J. A. Hedrick, R. D. Tyler, R. A. Smith, E. Keegan, and S. A. Chartrand. 1995. Penicillin-resistant Streptococcus pneumoniae in acute otitis media: risk factors, susceptibility patterns and antimicrobial management. Pediatr. Infect. Dis. J. 14:751-759.[Medline]
5
5. Cafini, F., R. del Campo, L. Alou, D. Sevillano, M. I. Morosini, F. Baquero, and J. Prieto. 2006. Alterations of the penicillin-binding proteins and murM alleles of clinical Streptococcus pneumoniae isolates with high-level resistance to amoxicillin in Spain. J. Antimicrob. Chemother. 57:224-229.[Abstract/Free Full Text]
6
6. Chen, D. K., A. McGeer, J. C. de Azavedo, and D. E. Low. 1999. Decreased susceptibility of Streptococcus pneumoniae to fluoroquinolones in Canada. N. Engl. J. Med. 22:233-239.
7
7. Clinical and Laboratory Standards Institute. 2006. Methods for dilution antimicrobial susceptibility tests for bacteria that grow aerobically. Approved standard M7-A7, 7th ed. Clinical and Laboratory Standards Institute, Wayne, PA.
8
8. Coffey, T. J., M. Daniels, L. K. McDougal, C. G. Dowson, F. C. Tenover, and B. G. Spratt. 1995. Genetic analysis of clinical isolates of Streptococcus pneumoniae with high-level resistance to expanded-spectrum cephalosporins. Antimicrob. Agents Chemother. 39:1306-1313.[Abstract/Free Full Text]
9
9. Critchley, I. A., M. R. Jacobs, S. D. Brown, M. M. Traczewski, G. S. Tillotson, and N. Janjic. 2008. Prevalence of serotype 19A Streptococcus pneumoniae among isolates from U.S. children in 2005-2006 and activity of faropenem. Antimicrob. Agents Chemother. 52:2639-2643.[Abstract/Free Full Text]
10
10. del Campo, R., F. Cafini, M. I. Morosini, A. Fenoll, J. Linares, L. Alou, D. Sevillano, R. Canton, J. Prieto, and F. Baquero on behalf of the Spanish Pneumococcal Network. 2006. Combinations of PBPs and MurM protein variants in early and contemporary high-level penicillin-resistant Streptococcus pneumoniae isolates in Spain. J. Antimicrob. Chemother. 57:983-986.[Abstract/Free Full Text]
11
11. Dowson, C. G., A. P. Johnson, E. Cercenado, and R. C. George. 1994. Genetics of oxacillin resistance in clinical isolates of Streptococcus pneumoniae that are oxacillin resistant and penicillin susceptible. Antimicrob. Agents Chemother. 38:49-53.[Abstract/Free Full Text]
12
12. du Plessis, M., E. Bingen, and K. P. Klugman. 2002. Analysis of penicillin-binding protein genes of clinical isolates of Streptococcus pneumoniae with reduced susceptibility to amoxicillin. Antimicrob. Agents Chemother. 46:2349-2357.[Abstract/Free Full Text]
13
13. Enright, M., and B. G. Spratt. 1998. A multilocus sequence typing scheme for Streptococcus pneumoniae: identification of clones associated with serious invasive disease. Microbiology 144:3049-3060.[Abstract/Free Full Text]
14
14. Filipe, S. R., E. Severina, and A. Tomasz. 2000. Distribution of the mosaic structured murM genes among natural populations of Streptococcus pneumoniae. J. Bacteriol. 182:6798-6805.[Abstract/Free Full Text]
15
15. Filipe, S. R., and A. Tomasz. 2000. Inhibition of the expression of penicillin resistance in Streptococcus pneumoniae by inactivation of cell wall muropeptide branching genes. Proc. Natl. Acad. Sci. USA 97:4891-4896.[Abstract/Free Full Text]
16
16. Forrest, A., A. Hazra, D. Girard, S. Finegan, J. O'Donnell, and K. Soma. 2008. Use of PK/PD analysis to select doses for PF-03709270 with and without probenecid (P), to be studied in phase 2b trials, abstr. A-034. Abstr. 48th Intersci. Conf. Antimicrob. Agents Chemother. American Society for Microbiology, Washington, DC.
17
17. Friedland, I. R., and G. H. McCracken, Jr. 1994. Management of infections caused by antibiotic-resistant Streptococcus pneumoniae. N. Engl. J. Med. 331:377-382.[Free Full Text]
18
18. Grebe, T., and R. Hakenbeck. 1996. Penicillin-binding proteins 2b and 2x of Streptococcus pneumoniae are primary resistant determinants for different classes of β-lactam antibiotics. Antimicrob. Agents Chemother. 40:829-834.[Abstract/Free Full Text]
19
19. Guenzi, E., A. M. Gasc, M. A. Sicard, and R. Hakenbeck. 1994. A two-component signal-transducing system is involved in competence and penicillin susceptibility in laboratory mutants of Streptococcus pneumoniae. Mol. Microbiol. 12:505-515.[Medline]
20
20. Hakenbeck, R., A. Konig, I. Kern, M. van der Linden, W. Keck, D. Billot-Klein, R. Legrand, B. Schoot, and L. Gutmann. 1998. Acquisition of five high-M_r penicillin-binding protein variants during transfer of high-level beta-lactam resistance from Streptococcus mitis to Streptococcus pneumoniae. J. Bacteriol. 180:1831-1840.[Abstract/Free Full Text]
21
21. Hakenbeck, R. 1999. β-Lactam-resistant Streptococcus pneumoniae: epidemiology and evolutionary mechanism. Chemotherapy 45:83-94.[CrossRef][Medline]
22
22. Ho, P.-L., T.-L. Que, D . N. -C. Tsang, T. -K. Ng, K. -H. Chow, and W.-H. Seto. 1999. Emergence of fluoroquinolone resistance among multiply resistant strains of Streptococcus pneumoniae in Hong Kong. Antimicrob. Agents Chemother. 43:1310-1313.[Abstract/Free Full Text]
23
23. Hoellman, D. B., G. Lin, M. R. Jacobs, and P. C. Appelbaum. 1999. Anti-pneumococcal activity of gatifloxacin compared with other quinolone and non-quinolone agents. J. Antimicrob. Chemother. 43:645-649.[Abstract/Free Full Text]
24
24. Hoskins, J., W. E. Alborn, Jr., J. Arnold, L. C. Blaszczak, S. Burgett, B. S. DeHoff, S. T. Estrem, L. Fritz, D. J. Fu, W. Fuller, C. Geringer, R. Gilmour, J. S. Glass, H. Khoja, A. R. Kraft, R. E. Lagace, D. J. LeBlanc, L. N. Lee, E. J. Lefkowitz, J. Lu, P. Matsushima, S. M. McAhren, M. McHenney, K. McLeaster, C. W. Mundy, T. I. Nicas, F. H. Norris, M. O'Gara, R. B. Peery, G. T. Robertson, P. Rockey, P. M. Sun, M. E. Winkler, Y. Yang, M. Young-Bellido, G. Zhao, C. A. Zook, R. H. Baltz, S. R. Jaskunas, P. R. Rosteck, Jr., P. L. Skatrud, and J. I. Glass. 2001. Genome of the bacterium Streptococcus pneumoniae strain R6. J. Bacteriol. 183:5709-5717.[Abstract/Free Full Text]
25
25. Huband, M. D., T. D. Gootz, M. A. Cohen, L. M. Mullins, S. P. McCurdy, L. A. Brennan, J. M. Duignan, P. J. Pagano, and R. W. Murray. 2008. In vitro antibacterial activity of sulopenem: a new oral penem antimicrobial versus recent bacterial isolates, abstr. F1-344. Abstr. 48th Intersci. Conf. Antimicrob. Agents Chemother. American Society for Microbiology, Washington, DC.
26
26. Jacobs, M. R., S. Bajaksouzian, A. Zilles, G. Lin, G. A. Pankuch, and P. C. Appelbaum. 1999. Susceptibilities of Streptococcus pneumoniae and Haemophilus influenzae to 10 oral antimicrobial agents based on pharmacodynamic parameters: 1997 U.S. surveillance study. Antimicrob. Agents Chemother. 43:1901-1908.[Abstract/Free Full Text]
27
27. Jacobs, M. R., D. Felmingham, P. C. Appelbaum, R. N. Grueneberg, and the Alexander Project Group. 2003. The Alexander Project 1998-2000: susceptibility of pathogens isolated from community-acquired respiratory tract infections to commonly used antimicrobial agents. J. Antimicrob. Chemother. 52:229-246.[Abstract/Free Full Text]
28
28. Klimek, J. J., C. Nightingale, W. B. Lehmann, and R. Quintiliani. 1977. Comparison of concentrations of amoxicillin and ampicillin in serum and middle ear fluid of children with chronic otitis media. J. Infect. Dis. 135:999-1002.[Medline]
29
29. Kosowska, K., M. R. Jacobs, S. Bajaksouzian, L. Koeth, and P. C. Appelbaum. 2004. Alterations of penicillin-binding proteins 1A, 2X, and 2B in Streptococcus pneumoniae isolates for which amoxicillin MICs are higher than penicillin MICs. Antimicrob. Agents Chemother. 48:4020-4022.[Abstract/Free Full Text]
30
30. Laible, G., B. G. Spratt, and R. Hakenbeck. 1991. Interspecies recombinational events during the evolution of altered PBP2X genes in penicillin-resistant clinical isolates of Streptococcus pneumoniae. Mol. Microbiol. 5:1993-2002.[Medline]
31
31. Liñares, J., A. G. Campa, and R. Pallares. 1999. Fluoroquinolone-resistance in Streptococcus pneumoniae. N. Engl. J. Med. 20:1546-1548.
32
32. McDougal, L. K., R. Facklam, M. Reeves, S. Hunter, J. M. Swenson, B. C. Hill, and F. C. Tenover. 1992. Analysis of multiply antimicrobial-resistant isolates of Streptococcus pneumoniae from the United States. Antimicrob. Agents Chemother. 36:2176-2184.[Abstract/Free Full Text]
33
33. McGee, L., L. McDougal, J. Zhou, B. G. Spratt, F. C. Tenover, R. George, R. Hakenbeck, W. Hryniewicz, J. C. Lefevre, A. Tomasz, and K. P. Klugman. 2001. Nomenclature of major antimicrobial-resistant clones of Streptococcus pneumoniae defined by the Pneumococcal Molecular Epidemiology Network. J. Clin. Microbiol. 39:2565-2571.[Abstract/Free Full Text]
34
34. Munoz, R., J. M. Musser, M. Crain, D. E. Briles, A. Marton, A. J. Parkinson, U. Sorensen, and A. Tomasz. 1992. Geographic distribution of penicillin-resistant clones of Streptococcus pneumoniae: characterization by penicillin-binding protein profile, surface protein A typing, and multilocus enzyme analysis. Clin. Infect. Dis. 15:112-118.[Medline]
35
35. Munoz, R., C. G. Dowson, M. Daniels, T. J. Coffey, C. Martin, R. Hakenbeck, and B. G. Spratt. 1992. Genetics of resistance to third-generation cephalosporins in clinical isolates of Streptococcus pneumoniae. Mol. Microbiol. 6:2461-2465.[Medline]
36
36. Neeleman, C., C. H. W. Klaassen, D. M. Klomberg, H. A. de Valk, and J. W. Mouton. 2004. Pneumolysin is a key factor in misidentification of macrolide-resistant Streptococcus pneumoniae and is a putative virulence factor of S. mitis and other streptococci. J. Clin. Microbiol. 42:4355-4357.[Abstract/Free Full Text]
37
37. Pankuch, G. A., M. R. Jacobs, and P. C. Appelbaum. 1994. Study of comparative antipneumococcal activities of penicillin G, RP 59500, erythromycin, sparfloxacin, ciprofloxacin, and vancomycin by using time-kill methodology. Antimicrob. Agents Chemother. 38:2065-2072.[Abstract/Free Full Text]
38
38. Pankuch, G. A., M. R. Jacobs, and P. C. Appelbaum. 1995. Activity of CP99,219 compared with DU-6859a, ciprofloxacin, ofloxacin, levofloxacin, lomefloxacin, tosufloxacin, sparfloxacin and grepafloxacin against penicillin-susceptible and -resistant pneumococci. J. Antimicrob. Chemother. 35:230-232.[Free Full Text]
39
39. Pankuch, G. A., M. R. Jacobs, and P. C. Appelbaum. 1995. Comparative activity of ampicillin, amoxicillin, amoxicillin/clavulanate and cefotaxime against 189 penicillin-susceptible and -resistant pneumococci. J. Antimicrob. Chemother. 35:883-888.[Abstract/Free Full Text]
40
40. Pichichero, M., and J. R. Casey. 2007. Emergence of a multiresistant serotype 19A pneumococcal strain not included in the 7-valent conjugate vaccine as an otopathogen in children. JAMA 298:1772-1778.[Abstract/Free Full Text]
41
41. Smith, A. M., and K. P. Klugman. 1995. Alterations in penicillin-binding protein 2B from penicillin-resistant wild-type strains of Streptococcus pneumoniae. Antimicrob. Agents Chemother. 39:859-867.[Abstract/Free Full Text]
42
42. Smith, A. M., and K. P. Klugman. 1998. Alterations in PBP 1A essential for high-level penicillin resistance in Streptococcus pneumoniae. Antimicrob. Agents Chemother. 42:1329-1333.[Abstract/Free Full Text]
43
43. Smith, A. M., and K. P. Klugman. 2001. Alterations in MurM, a cell wall muropeptide branching enzyme, increase high-level penicillin and cephalosporin resistance in Streptococcus pneumoniae. Antimicrob. Agents Chemother. 45:2393-2396.[Abstract/Free Full Text]
44
44. Spangler, S. K., M. R. Jacobs, and P. C. Appelbaum. 1992. Susceptibilities of penicillin-susceptible and -resistant strains of Streptococcus pneumoniae to RP 59500, vancomycin, erythromycin, PD 131628, sparfloxacin, temafloxacin, Win 57273, ofloxacin, and ciprofloxacin. Antimicrob. Agents Chemother. 36:856-859.[Abstract/Free Full Text]
45
45. Spangler, S. K., M. R. Jacobs, G. A. Pankuch, and P. C. Appelbaum. 1993. Susceptibility of 170 penicillin-susceptible and -resistant pneumococci to six oral cephalosporins, four quinolones, desacetylcefotaxime, Ro 23-9424 and RP 67829. J. Antimicrob. Chemother. 31:273-280.[Abstract/Free Full Text]
46
46. Thompson, J. D., D. G. Higgins, and T. J. Gibson. 1994. CLUSTAL W: improving the sensitivity of progressive multiple sequence alignment through sequence weighting, position-specific gap penalties and weight matrix choice. Nucleic Acids Res. 22:4673-4680.[Abstract/Free Full Text]
47
47. Ubukata, K., T. Muraki, A. Igarashi, Y. Asahi, and M. Konno. 1997. Identification of penicillin and other beta-lactam resistance in Streptococcus pneumoniae by polymerase chain reaction. J. Infect. Chemother. 3:190-197.[CrossRef]
48
48. Visalli, M. A., M. R. Jacobs, and P. C. Appelbaum. 1996. MIC and time-kill study of DU-6859a, ciprofloxacin, levofloxacin, sparfloxacin, cefotaxime, imipenem, and vancomycin against nine penicillin-susceptible and -resistant pneumococci. Antimicrob. Agents Chemother. 40:362-366.[Abstract/Free Full Text]
49
49. Visalli, M. A., M. R. Jacobs, and P. C. Appelbaum. 1997. Antipneumococcal activity of BAY 12-8039, a new quinolone, compared with activities of three other quinolones and four ß-lactams. Antimicrob. Agents Chemother. 41:2786-2789.[Abstract/Free Full Text]
50
50. Yamane, A., H. Nakano, Y. Asahi, K. Ubukata, and M. Konno. 1996. Directly repeated insertion of 9-nucleotide sequence detected in penicillin-binding protein 2B gene of penicillin-resistant Streptococcus pneumoniae. Antimicrob. Agents Chemother. 40:1257-1259.[Abstract/Free Full Text]
51
51. Zhao, G., T. I. Meier, J. Hoskins, and K. A. McAllister. 2000. Identification and characterization of the penicillin-binding protein 2a of Streptococcus pneumoniae and its possible role in resistance to beta-lactam antibiotics. Antimicrob. Agents Chemother. 44:1745-1748.[Abstract/Free Full Text]

---

Antimicrobial Agents and Chemotherapy, June 2009, p. 2239-2247, Vol. 53, No. 6
0066-4804/09/$08.00+0     doi:10.1128/AAC.01531-08
Copyright © 2009, American Society for Microbiology. All Rights Reserved.

This Article Abstract Full Text (PDF) Alert me when this article is cited Alert me if a correction is posted Citation Map Services E-mail this article to a friend Similar articles in this journal Similar articles in PubMed Alert me to new issues of the journal Download to citation manager Reprints and Permissions Copyright Information Books from ASM Press MicrobeWorld Citing Articles Citing Articles via Google Scholar Google Scholar Articles by Kosowska-Shick, K. Articles by Appelbaum, P. C. Search for Related Content PubMed PubMed Citation Articles by Kosowska-Shick, K. Articles by Appelbaum, P. C.