Monthly Changes in the Rate of Recovery of Penicillin-Resistant Organisms From Children

The resistance to penicillin of oropharyngeal flora organisms has increased during the past decade. Two major mechanisms of resistance have developed: the production of betalactamase and changes in the penicillin protein-binding site.1-4 The aerobic beta-lactamase-producing bacteria (BLPB) consist of Staphylococcus aureus, Haemophilus influenzae and Moraxella catarrhalis, and the anaerobic bacteria are pigmented Prevotella species and Fusobacterium species.5 The emergence of BLPB and increase in Streptococcus pneumoniae resistance were shown to be associated with the administration of penicillin therapy.6-10 All previous studies that documented the recovery of BLPB and the isolation rate of penicillin-resistant bacteria reported the total isolation rate of these organisms during the study period. None described the dynamics of these changes with time.6-9 This study was designed to investigate the dynamics of penicillin resistance in the community by monitoring the monthly changes in the rate of recovery of penicillin-resistant bacteria in the oropharynx of children. Materials and methods. The study was conducted for 2 years between September 1, 1993, and August 31, 1995. The population studied was middle class, residing in suburban locations in the vicinity of Washington, DC. Included in the study were the first 30 patients seen each month by the authors for acute pharyngotonsillitis in an acute care clinic. Acute pharyngotonsillitis was defined as acute onset of sore throat plus at least one of the following: anterior cervical adenitis; temperature >38.3°C (101°F); pharyngeal or tonsillar exudate or pharyngeal injection. A total of 1891 visits for this diagnosis occurred in the clinic during the study period. The median age of the children was 7 years and 4 months (range, 2.5 to 14 years). Past medical history, including therapies with antimicrobial therapy and day-care attendance, was obtained by using a questionnaire and by checking medical records. Pharyngeal cultures were obtained from all children with two sterile cotton swabs. To obtain aerobic cultures one sterile cotton swab was placed in a transport system for aerobic bacteria (Culturette®; Marion Scientific Corp., Rockford, IL). For anaerobic bacteria the second swab was placed in an anaerobic transport medium (Port-A-Cul®; BBL, Becton Dickinson Co., Cockeysville, MD). The specimens were transported to the bacteriology laboratory and inoculated within 24 h of collection. Sheep blood (5%), chocolate and MacConkey's agar plates were inoculated for the isolation of aerobic organisms. The plates were incubated at 37°C aerobically (MacConkey's) and under 5% carbon dioxide (blood and chocolate), and examined at 24 and 48 h. To isolate anaerobic bacteria the specimens were plated onto prereduced vitamin K1-enriched Brucella blood agar, onto an anaerobic blood agar plate containing kanamycin and vancomycin, onto an anaerobic blood plate containing phenylethyl alcohol and into enriched thioglycolate broth.11 These media were incubated inside anaerobic jars (GasPak®; BBL) at 37°C and examined at 48 and 96 h. The thioglycolate broth was incubated for 14 days. Aerobic and anaerobic bacteria were identified by conventional methods.11, 12 Group A beta-hemolytic Streptococcus (GABHS) was identified by determining bacitracin sensitivity, and serologic grouping by Phadebact® coagglutination (Pharmacia Diagnostics, Inc., Piscataway, NJ).12 Five different colonies of each bacterial strain were picked from the primary agar plates and tested separately for production of beta-lactamase. Beta-lactamase activity was determined by chromogenic cephalosporin analog 87/312 methodology.13 All isolates of S. pneumoniae were screened for penicillin susceptibility with a 1-μg oxacillin disk by the Kirby-Bauer disk diffusion method. Strains that showed a zone of inhibition ≥20 mm were considered susceptible, and those that showed a zone of inhibition <20 mm were considered possibly resistant to penicillin. Resistance was confirmed by microdilution cation-supplemented Mueller-Hinton broth plus 5% lysed and centrifuged horse blood, as recommended by the National Committee for Clinical Laboratory Standards.14 Intermediate resistance to penicillin was defined as a MIC of 0.1 to 1.0 μg/ml and high resistance to penicillin was defined as MIC ≥ 2.0 μg/ml. Statistical analysis was done by the chi square test with continuity correction. Results. Three hundred sixty cultures were obtained each year. A total of 324 BLPB were recovered during the study period, 168 in the first and 156 in the second year. The BLPB in order of frequency were pigmented Prevotella species (87 isolates), H. influenzae (81, 75 of which were non-type b), M. catarrhalis (57 isolates), Fusobacterium species (50 isolates) and Staph. aureus (49 isolates). There were no differences in the recovery rate of these organisms each year. The recovery of BLPB varied for different months in each of the 2 years. The number of BLPB isolates and the number of patients with BLPB was maximum between January and April each year (P < 0.01). The lowest number of isolates was found between June and October each year. A gradual increase in the recovery rate of these organisms occurred between September and January, and a slow decline took place from April to June each year (Fig. 1).Fig. 1: Monthly recovery rate of beta-lactamase-producing bacteria from children (September, 1993, to August, 1995).BLPB were recovered from a total of 132 (37%) of the patients in the first year and 123 (34%) in the second. Beta-lactam antibiotics were administered within the past six weeks to 84 children in the first year and 81 in the second. Sixty-nine (52%) of these children in the first year were among the 132 who harbored BLPB, and 70 (57%) who received beta-lactam antibiotics in the second year were among the 123 who harbored BLPB. Fifteen beta-lactamtreated children (7%) were among the 228 children who did not harbor BLPB in the first year (P < 0.001), and 14 (6%) were among the 237 who did not carry BLPB in the second year (P < 0.001). Most beta-lactam antibiotics were administered during the peak time of recovery of BLPB in each year. A total of 88 isolates of S. pneumoniae were recovered during the study period, 39 in the first year and 49 in the second. Twelve were resistant (6 intermediate and 6 highly resistant) in the first year and 15 were resistant (12 intermediate and 3 highly resistant) in the second year. Most of the resistant strains were isolated between January and May each year. Beta-lactam antibiotics were given within the past 6 weeks to 10 of the 12 (83%) children who were colonized with penicillin-resistant S. pneumoniae in the first year and to 11 of the 15 in the second year (73%). Beta-lactam antibiotics were given to 74 (21%) of the 348 patients who were not colonized with penicillin-resistant S. pneumoniae in the first year and to 70 of the 345 (20%) in the second year (P < 0.001 in each year). No correlation was found between previous therapy with non-beta-lactam antimicrobials or day-care attendance (in 34 children in the first year and 42 in the second for the latter). However, no conclusion can be drawn from these findings because of the low prevalence of S. pneumoniae in this study. GABHS was identified in 35 children in the first year and 44 in the second. No correlation between recovery of penicillin-resistant bacteria and recovery of GABHS was noted. Discussion. This study demonstrates the monthly variations during a 2-year period in the recovery rate of aerobic and anaerobic BLPB and S. pneumoniae resistant to penicillin in children seen in an ambulatory clinic setting for acute pharyngotonsillitis. The maximal recovery rate of these organisms was in the winter months (January to May) where the rate of BLPB reached a peak of 63 to 67%. In contrast the lowest rate was recorded during the summer and fall (June to October) when it declined to a low of 10 to 13%. Similar seasonal changes in the prevalence of penicillin-resistant S. pneumoniae were previously reported.15 These seasonal variations may be a result of exposure to beta-lactam antibiotics that are given in greater frequency during the winter season. The administration of beta-lactam antibiotics has been previously shown to be associated with increased recovery rate of BLPB6-9 and S. pneumoniae resistant to penicillin.10 We also observed a correlation between recent intake of beta-lactam antibiotics and increase recovery of penicillinresistant bacteria. Beta-lactam antibiotics were given to more than one-half of the children colonized with BLPB and to 73 to 83% of those with penicillin-resistant S. pneumoniae. However, these agents were not taken by the remaining children, about one-half of those colonized with BLPB and 17 to 27% of those with penicillin-resistant S. pneumoniae. It is therefore plausible that factors other than direct exposure to antimicrobials contribute to the emergence of penicillin resistance. BLPB were also shown to spread and colonize family members of patients treated with beta-lactam antimicrobials.8 Another potential reason for the increased recovery of BLPB is the increased indoor crowding that occurs during the winter season and the possible spread of BLPB within schools and day-care centers that operate during the winter season. Infection with BLPB and penicillin-resistant bacteria may lead to failure of penicillins in the treatment of otitis media and tonsillitis.16 Furthermore the presence of BLPB can also interfere with the eradication of recurrent GABHS tonsillitis.17 Knowledge and monitoring of the local seasonal variations in the rate of recovery of penicillin-resistant organisms may have a direct effect on the physician's choice of antimicrobials therapy. If similar seasonal variations in the prevalence of BLPB occur in the respiratory flora of younger children, then this may affect treatment choices for acute otitis media. The decrease in recovery of penicillin-resistant organisms during the summer season was found to be associated with the decreased exposure of children to beta-lactam antimicrobials. This decrease underscores the importance of judicious use of antimicrobials. Avoidance of improper use of betalactam antibiotics may help control the increase in the rate of recovery of penicillin-resistant organisms. Acknowledgments. The assistance of Joanie Pietrafitta and Lee Gibbs is greatly appreciated. Itzhak Brook, M.D. Alan E. Gober, M.D.