Post-PCV7 Changes in Colonizing Pneumococcal Serotypes in 16 Massachusetts Communities, 2001 and 2004
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《小儿科》
Channing Laboratory, Brigham and Women's Hospital, Boston, Massachusetts
Department of Ambulatory Care and Prevention, Harvard Medical School and Harvard Pilgrim Health Care, Boston, Massachusetts
Department of Pediatrics, Boston University School of Medicine, Boston, Massachusetts
Divisions of Infectious DiseasesGeneral Pediatrics, Children's Hospital Boston, Boston, Massachusetts
Department of Pediatrics, Harvard Medical School, Boston, Massachusetts
ABSTRACT
Objective. The introduction of heptavalent conjugate pneumococcal vaccine (PCV7) has raised concerns for replacement with nonvaccine serotypes in both invasive disease and asymptomatic carriage. Analysis of colonizing serotypes among healthy children in the community provides critical data on such changes.
Methods. Nasopharyngeal specimens were obtained from children who were younger than 7 years during well-child or sick visits in primary care practices in 16 Massachusetts communities during 2001 and 2004. Susceptibility testing and serotyping were performed on isolated Streptococcus pneumoniae strains. Vaccination history with PCV7 was abstracted from the medical record.
Results. Among colonizing pneumococcal isolates, PCV7 serotypes decreased from 36% to 14%, and non-PCV7 serotypes increased from 34% to 55%. Overall carriage did not change (26% to 23%); neither did carriage of potentially cross-reactive serotypes (30% to 31%). The most common non-PCV7 serotypes were serotypes 11, 15, and 29. There was a substantial increase in penicillin nonsusceptibility from 8% to 25% in non-PCV7 serotypes; 35% were highly resistant to penicillin. Penicillin nonsusceptibility increased from 45% to 56% among PCV7 serotypes while remaining stable among PCV7 potentially cross-reactive strains (51% vs 54%).
Conclusions. Pneumococcal colonization has changed after the introduction of PCV7, both in serotype distribution and in patterns of antibiotic resistance. The frequency of nonvaccine strains has increased, and the proportion of nonvaccine isolates that are not susceptible to penicillin has tripled. This shift toward increased carriage of nonvaccine serotypes warrants vigilance for changes in the epidemiology of invasive pneumococcal disease.
Key Words: Streptococcus pneumoniae pneumococcal conjugate vaccine antibiotic resistance colonization
Abbreviations: PCV7, heptavalent pneumococcal conjugate vaccine PNSP, penicillin nonsusceptible Streptococcus pneumoniae MDR, multidrug resistance AOM, acute otitis media
Before the release of a conjugate vaccine for US children, Streptococcus pneumoniae caused invasive disease in 1 in 500 to 600 children who were younger than 2 years.1 In addition, rising antibiotic resistance among pneumococcal strains was a major concern among public health officials.2,3 By 1999, 38% of laboratory isolates in children who were younger than 2 years were nonsusceptible to penicillin,4 with higher rates reported in local areas.5–7
In February 2000, heptavalent pneumococcal conjugate vaccine (PCV7) was released to provide immunity to the 7 serotypes responsible for 85% of pediatric invasive pneumococcal disease8 and 78% of penicillin-nonsusceptible S pneumoniae (PNSP) isolates in children.9 Prelicensure trials showed a 90% reduction in invasive disease10 and a 23% reduction in all-cause pneumonia in the first 2 years of life.11 Postlicensure studies have also shown declines in invasive pneumococcal disease from 188 per 100000 to 59 cases per 100000 in US children who are younger than 2 years.4,12
Despite the success of PCV7 in reducing invasive disease, critical questions remain about the changing pneumococcal reservoir. Some trials of immunization with PCV7 have shown a 50% to 65% decrease in carriage of vaccine serotypes13–15 and have raised concerns about replacement of vaccine serotypes with nonvaccine serotypes that may ultimately result in invasive disease.16,17 Although, currently, the majority of invasive disease is caused by vaccine-included serotypes, the relationship between carriage of and invasive disease is complex and varies by serotype.18,19 Shifts in colonizing serotypes after PCV7 may serve as early indicators of changes in invasive disease.
We assessed secular trends in community-level S pneumoniae carriage and resistance after the licensure of PCV7. Community-level analyses assess the full impact of both direct and indirect (herd immunity) effects of vaccines. In 2001, we conducted surveillance for pneumococcal carriage among generally healthy children in 16 distinct Massachusetts communities.20 At that time, just after introduction of PCV7, we found that 26% of children carried S pneumoniae overall and that PCV7 serotypes accounted for 36% of isolates.20 Resistance to penicillin (and other antimicrobials) was concentrated among PCV7 serotypes and potentially cross-reactive serotypes, with penicillin nonsusceptibility of 45% and 51%, respectively. By November 2003, all children through 3.5 years of age were born into the era of routine PCV7 immunization in infancy. In this context, we resampled the same 16 communities in the winter and spring of 2003–2004 to determine (1) changes in carriage of PCV serotypes, PCV7 cross-reactive serotypes, and non-PCV7 S pneumoniae serotypes; (2) changes in overall antimicrobial resistance; and (3) serotype-specific changes in antimicrobial resistance.
METHODS
Data Collection
Nasopharyngeal specimens were collected from young children in pediatric and family practice physician offices in 16 Massachusetts communities between November 17, 2003, and April 16, 2004. We had performed surveillance in the same 16 communities from March 13 to May 11, 2001, as part of the baseline microbiologic data collection for a randomized trial of community-level intervention to promote judicious antibiotic prescribing for children.20 These 16 communities were selected on the basis of physical geographic separation, evidence that few children crossed community boundaries to seek pediatric care, and diversity of size and demographic characteristics using available US Census data. In 2001, 31 private practice offices participated, and in 2004, 23 offices participated, all of which had participated in 2001.
In 2001, children were eligible for inclusion if they were younger than 7 years, resided in a study community, and presented for either a routine well-child or "sick visit" at a participating practice.20 Parental consent and nasopharyngeal swabs were obtained by either trained study personnel or trained office practice nurses. PCV7 vaccination history was recorded from the medical record. In 2004, to increase sample size, we modified criteria to include children who received care in the study practices and lived beyond the zip code boundaries of the community. Recruitment in 2004 was independent of participation in 2001. All study procedures were approved by the Harvard Pilgrim Health Care institutional review board.
Microbiologic Processing
Nasopharyngeal samples were processed for S pneumoniae growth, antibiotic susceptibility, and serotype as previously described.20 Standard National Committee for Clinical Laboratory Standards susceptibility breakpoints were used to classify organisms as susceptible, intermediate, and resistant.21 Two definitions of multidrug resistance (MDR) were used: resistance to the -lactam class plus 1 other antibiotic class (MDR1) and resistance to at least 3 antibiotic classes (MDR2). Penicillins, cephalosporins, and carbapenems were considered a single class. Comparisons of third-generation cephalosporin resistance were performed using 2004 breakpoint criteria.21,22 We had previously reported carriage data from 2001 using the breakpoint criteria at that time. These 2001 data were reanalyzed with 2004 criteria for this analysis.
Serotypes included in the PCV7 were considered vaccine serotypes, and serotypes from serogroups that were not in PCV7 were considered nonvaccine serotypes. Serotypes in the same serogroup as vaccine serotypes (eg, 6A and 6B both are serogroup 6 strains) but not specifically included in the vaccine were classified as potentially cross-reactive serotypes. The extent to which the vaccine provides protection in vivo varies by serotype, but there is likely to be clinically important cross-protection against some of these strains. For quality control, we reserotyped a 36% random sample (82 of 226) of pneumococcal isolates from 2004 in a blinded manner and calculated percentage concordance.
Data Analysis
Microbiologic data were analyzed using SAS software version 8.2 (SAS Institute, Cary, NC). Proportions of children who carried S pneumoniae and proportions of isolates that were nonsusceptible to antibiotics were calculated and compared between the 2004 and previously published 2001 sample20 using Fisher exact tests. Carriage proportions were calculated by sampling period and by age group (<6 months, 6–<12 months, 12–<24 months, 24–<36 months, and 36 months). Resistance patterns were analyzed for the 2004 sample, by vaccine group (PCV7, PCV7 cross-reactive, and non-PCV7 serotypes) and by individual serotype and compared with the 2001 sample.
RESULTS
We collected 996 nasopharyngeal samples from children in 23 practices from November 17, 2003, through April 16, 2004 (Table 1), 2 of which were lost in transit. Of the 994 remaining samples, 73% (726) came from children who lived within the specified 16 Massachusetts communities described in Table 2; the remainder were patients of these practices who lived outside the designated community zip codes. Because pneumococcal carriage and descriptive characteristics of these children were similar to those who lived within the designated zip codes, data were combined for analysis. Fifteen percent (153) of participants were siblings of other participants, compared with 12% (87) in 2001.
Pneumococcal carriage was similar between 2001 and 2004 (Table 2). Age-specific pneumococcal carriage increased in children who were younger than 6 months (8% in 2001 vs 22% in 2004; P = .01) and decreased in those who were older than 36 months (30% in 2001 vs 19% in 2004; P = .002). Overall carriage of Haemophilus influenzae was 15% in 2004 compared with 17% in 2001 (P = .4), and overall carriage of Moraxella catarrhalis was 26% in 2004 compared with 16% in 2001 (P < .0001).
Of the 232 S pneumoniae isolates, 226 were serotyped (6 were unable to be grown after transfer). The proportion of PCV7 serotypes declined from 36% (51 of 143) in 2001 to 14% (32 of 226) in 2004 (P < .0001), whereas non-PCV7 serotypes increased from 34% (49 of 143) in 2001 to 55% (125 of 226) in 2004 (P < .0001; Fig 1). The proportion of PCV7 cross-reactive isolates remained stable. In all age groups, the proportion of PCV7 strains carried decreased, whereas the proportion of non-PCV7 serotypes either remained the same (<6-month-olds) or increased (all other age groups). There was no difference in the proportion of serotypes included in the 23-valent adult pneumococcal polysaccharide vaccine between 2001 (57%; 82 of 143) and 2004 (50%; 114 of 226). Serotype concordance during repeat serotyping was 96.3%. Previous studies that compared concordance of pneumococcal serotyping between laboratories showed a similar serotype discordance level.23
Every PCV7-included serotype decreased as a proportion of isolates, except for 9V, which had no representation in 2001 and 1 isolate in 2004 (Fig 2). Serotype 19F showed minimal decline and thus increased proportionally among PCV7 serotypes from 31% (16 of 51) in 2001 to 63% (20 of 32; P = .007) in 2004. Among PCV7 cross-reactive serotypes, there was a significant decrease in the proportion of serotype 6A (53% [23 of 43] to 32% [22 of 69]; P = .03) but significant increases in 19A (14% [6 of 43] to 38% [26 of 69]; P = .009) and 23A (2% [1 of 43] to 20% [14 to 69]; P = .008).
The most common non-PCV7 serotypes (N = 125) in 2004 were 15 (N = 34), 29 (N = 19), 11A (N = 18), 22F (N = 9), 10A (N = 7), and 35F (N = 7). Between 2001 and 2004, there was a significant increase in serotype 29 (2% in 2001 vs 8% in 2004; P = .04) and a trend toward an increase in serotype 15 (8% in 2001 vs 15% in 2004; P = .07). There were insufficient numbers to detect a significant change in the remaining serotypes.
Vaccine penetration (receipt of at least 1 PCV7 dose) increased from 60% in 2001 to 96% in 2004 among children who were younger than 12 months and from 38% to 79% among children who were at least 12 months of age. Among children who were at least 12 months of age and received 1 or more doses of PCV7, non-PCV7 strains increased from 8% (16 of 191) in 2001 to 14% (87 of 623) in 2004 (P = .05). Children who were at least 12 months of age and had not received any doses of PCV7 (primarily older children who were born before the introduction of PCV7) had similar percentages of non-PCV7 carriage in 2001 (7%; 20 of 306) as compared with 2004 (10%; 16 of 164; P = .27). In 2001, overall carriage of PCV7 serotypes among those who were at least 12 months of age was low among both vaccine recipients (5%; 10 of 191) and nonrecipients (3%; 8 of 306). No reduction in overall carriage of vaccine serotypes was seen in 2004, although the proportion of PCV7 serotypes among carriers declined.
Overall carriage of a PNSP isolate was similar in 2001 (7%; 49 of 695) and 2004 (9%; 86 of 988). However, penicillin nonsusceptibility declined among PCV7 isolates and increased among non-PCV7 isolates (Fig 1). The former reflected a decrease in high-level penicillin-resistant isolates, whereas the latter reflected an increase in intermediate resistance to penicillin (Table 3).
Penicillin nonsusceptibility varied among commonly carried serotypes. In 2004, the proportion of PNSP was 64% in 6A, 0% in 11A, 38% in 15, 65% in 19A, 65% in 19F, 36% in 23A, and 74% in 29 (see Fig 2).
Changes in susceptibility to additional antibiotics is shown in Table 3. Ceftriaxone nonsusceptibility declined significantly between 2001 and 2004 isolates, largely as a result of declines in intermediate resistance (14% in 2001 vs 0.4% in 2004; P < .0001). This decline was seen across PCV7, PCV7 cross-reactive, and non-PCV7 serogroups, even when accounting for the fact that National Committee for Clinical Laboratory Standards breakpoints had changed between 2001 and 2004.22 Trimethoprim-sulfamethoxazole nonsusceptibility also declined significantly. On the contrary, clindamycin nonsusceptibility increased, particularly among non-PCV7 serotypes. Clindamycin nonsusceptibility was highly correlated with macrolide nonsusceptibility (6 of 6 in 2001; 20 of 21 in 2004). Fluoroquinolone nonsusceptibility was minimal in both 2001 (3%; 4 of 143) and 2004 (0%; 0 of 226). Notably, among non-PCV7 serotypes, nonsusceptibility significantly increased for penicillin, clindamycin, and MDR (Table 3). The proportion of isolates that were resistant to 3 or more antibiotic classes increased from 9% (4 of 43) to 20% (14 of 69) among PCV7 cross-reactive serotypes and from 2% (1 of 49) to 10% (13 of 125) among non-PCV7 serotypes.
DISCUSSION
The licensure of PCV7 was accompanied by projections of decreased S pneumoniae carriage, invasive disease, and antibiotic resistance.13,15,17 When evaluating young healthy children in the same 16 Massachusetts communities in 2001 and 2004, we found significantly increased carriage of nonvaccine strains and increased antibiotic resistance among nonvaccine serotypes.
Despite the significant decline in PCV7 serotypes, overall pneumococcal carriage did not change between 2001 and 2004 because of significant increases in the carriage of non-PCV7 serotypes. This serotype replacement was seen in all pediatric age groups studied and resulted in similar proportions of pneumococcal carriage in age groups between 6 and 36 months for both sampling periods. Among children who were 36 months of age, overall carriage declined between 2001 and 2004. This is not surprising given differential vaccination of children of this age in 2004 compared with 2001 (unlikely to be vaccinated unless at high risk). By contrast, we have no clear explanation for the observed increased rate of carriage among young infants (<6 months).
These changes suggest a rapid replacement of vaccine serotypes by nonvaccine serotypes, either by the unmasking of previous minority strains24 or by replacement of previous PCV7 carriage by serotypes unaffected by vaccine immunity. Another possibility is serotype switch,25 whereby pneumococcal strains express a new capsular serotype, possibly to evade host immunity. This phenomenon has been described between 15B and 15C,26 19F and 23F,27 9V and 11A,28 and others.25
Although serotype replacement has been a concern after the release of the conjugate pneumococcal vaccine, experts hoped that antibiotic resistance would decrease because resistance had been highly concentrated in PCV7 and PCV7 cross-reactive serotypes.1,20,29 As hoped, our serial community-based sampling showed that high-level penicillin resistance declined 38% and nonsusceptibility to third-generation cephalosporins decreased 88% across all isolates. However, this reduction in high-level penicillin resistance, produced by the reduction in carriage of certain PCV7 and PCV7 cross-reactive serotypes, was offset by increased carriage of nonvaccine serotypes, in which intermediate penicillin resistance has risen significantly. Antibiotic resistance among non-PCV7 pneumococcal serotypes (which had been negligible in 2001) increased to 25%, and resistance to 3 or more antibiotic classes increased to 10%.
In addition to increasing intermediate penicillin resistance, non-PCV7 strains showed an increase in clindamycin nonsusceptibility that was highly associated with macrolide resistance. Until now, most S pneumoniae isolates that are resistant to macrolides in the United States have displayed the M phenotype, related to a macrolide-specific efflux mechanism.30 The mechanism of resistance associated with dual macrolide and clindamycin resistance is related to the MLS phenotype, which is a result of target modification at the 50S ribosomal subunit.30 As the prevalence of non-PCV7 carriage increases, antibiotic exposure may be providing a selective pressure toward the emergence of additional resistance in these serotypes.
These data must be interpreted with the following caveats. First, we assume that carriage among children who seek care from primary care providers reflects carriage in their community. Although this assumption depends on the distribution of patients in a given practice, we believe it to be better than sampling schemes in child care settings that likely overrepresent shared colonizing strains. Second, the surveillance period occurred later in the season in 2001 compared with 2004 and may have affected carriage prevalence slightly, although we expect the proportion of serotypes among carriers to be unaffected. Third, because we sampled only twice, it is possible that we have observed natural fluctuations in the pneumococcal carriage state, rather than a trend related to the introduction of pneumococcal conjugate vaccine. We believe this unlikely given the magnitude and consistent direction of the shifts.
These data provide insight into the impact of PCV7 on pneumococcal carriage in young children. All communities included both immunized and unimmunized children, but, by 2004, all children who were younger than 42 months were born after routine PCV7 immunization began. As reflected in our study subjects, Massachusetts has one of the highest documented immunization rates in the United States.31 In this sample, we find evidence for increased carriage of non-PCV7 serotypes among vaccinated children who were older than 12 months.
It remains to be seen whether invasive infections caused by non-PCV serotypes will become more common. Recent data32 suggest that nonvaccine serotypes, including novel ones,33,34 may become important pathogens and that increased antibiotic resistance among these strains33,34 may have adverse consequences for children over time. Continued evaluation of the pneumococcal reservoir may presage changes in strains that are responsible for localized and invasive disease as a result of the widespread use of PCV7. Such information may inform clinicians' expectations of antibiotic resistance in pneumococcal infections as well as development of the next generation of conjugate pneumococcal vaccines.
ACKNOWLEDGMENTS
This study was funded by the US Agency for Healthcare Research and Quality (grant HS10247).
We thank the nurses and research assistants who made the nasopharyngeal sampling possible in our 16 Massachusetts communities: Andrea Gayle-Bennet, PA; Judy LaChance, RN; Patricia McCrann, RN; Verna Senna, RN; Martha Tata, RN; Susan DeAmario; Bobbi Fish; Sheila Lambi; Sara Lehroff; Ann Marie Rezendes; and Dan Steinberg. Also, we extend special thanks to each of the practices that participated in this study.
FOOTNOTES
Accepted Feb 8, 2005.
Conflict of interest: Dr Pelton receives research support from Wyeth Lederle.
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Black S, Shinefield H, Fireman B, et al. Efficacy, safety and immunogenicity of heptavalent pneumococcal conjugate vaccine in children. Northern California Kaiser Permanente Vaccine Study Center Group. Pediatr Infect Dis J. 2000;19 :187 –195
Black SB, Shinefield HR, Ling S, et al. Effectiveness of heptavalent pneumococcal conjugate vaccine in children younger than five years of age for prevention of pneumonia. Pediatr Infect Dis J. 2002;21 :810 –815
Black S, Shinefield H, Baxter R, et al. Postlicensure surveillance for pneumococcal invasive disease after use of heptavalent pneumococcal conjugate vaccine in Northern California Kaiser Permanente. Pediatr Infect Dis J. 2004;23 :485 –489
Dagan R, Melamed R, Muallem M, et al. Reduction of nasopharyngeal carriage of pneumococci during the second year of life by a heptavalent conjugate pneumococcal vaccine. J Infect Dis. 1996;174 :1271 –1278
Mbelle N, Huebner RE, Wasas AD, Kimura A, Chang I, Klugman KP. Immunogenicity and impact on nasopharyngeal carriage of a nonavalent pneumococcal conjugate vaccine. J Infect Dis. 1999;180 :1171 –1176
Dagan R, Givon-Lavi N, Zamir O, Fraser D. Effect of a nonavalent conjugate vaccine on carriage of antibiotic-resistant Streptococcus pneumoniae in day-care centers. Pediatr Infect Dis J. 2003;22 :532 –540
Spratt BG, Greenwood BM. Prevention of pneumococcal disease by vaccination: does serotype replacement matter Lancet. 2000;356 :1210 –1211
Pelton SI, Klein JO. The future of pneumococcal conjugate vaccines for prevention of pneumococcal diseases in infants and children. Pediatrics. 2002;110 :805 –814
Brueggemann AB, Griffiths DT, Meats E, Peto T, Crook DW, Spratt BG. Clonal relationships between invasive and carriage Streptococcus pneumoniae and serotype- and clone-specific differences in invasive disease potential. J Infect Dis. 2003;187 :1424 –1432
Hanage WP, Kaijalainen TH, Syrjanen RK, et al. Invasiveness of serotypes and clones of Streptococcus pneumoniae among children in Finland. Infect Immun. 2005;73 :431 –435
Finkelstein JA, Huang SS, Daniel J, et al. Antibiotic-resistant Streptococcus pneumoniae in the heptavalent pneumococcal conjugate vaccine era: predictors of carriage in a multicommunity sample. Pediatrics. 2003;112 :862 –869
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Porat N, Barkai G, Jacobs MR, Trefler R, Dagan R. Four antibiotic-resistant Streptococcus pneumoniae clones unrelated to the pneumococcal conjugate vaccine serotypes, including 2 new serotypes, causing acute otitis media in southern Israel. J Infect Dis. 2004;189 :385 –392(Susan S. Huang, MD, MPH, )
Department of Ambulatory Care and Prevention, Harvard Medical School and Harvard Pilgrim Health Care, Boston, Massachusetts
Department of Pediatrics, Boston University School of Medicine, Boston, Massachusetts
Divisions of Infectious DiseasesGeneral Pediatrics, Children's Hospital Boston, Boston, Massachusetts
Department of Pediatrics, Harvard Medical School, Boston, Massachusetts
ABSTRACT
Objective. The introduction of heptavalent conjugate pneumococcal vaccine (PCV7) has raised concerns for replacement with nonvaccine serotypes in both invasive disease and asymptomatic carriage. Analysis of colonizing serotypes among healthy children in the community provides critical data on such changes.
Methods. Nasopharyngeal specimens were obtained from children who were younger than 7 years during well-child or sick visits in primary care practices in 16 Massachusetts communities during 2001 and 2004. Susceptibility testing and serotyping were performed on isolated Streptococcus pneumoniae strains. Vaccination history with PCV7 was abstracted from the medical record.
Results. Among colonizing pneumococcal isolates, PCV7 serotypes decreased from 36% to 14%, and non-PCV7 serotypes increased from 34% to 55%. Overall carriage did not change (26% to 23%); neither did carriage of potentially cross-reactive serotypes (30% to 31%). The most common non-PCV7 serotypes were serotypes 11, 15, and 29. There was a substantial increase in penicillin nonsusceptibility from 8% to 25% in non-PCV7 serotypes; 35% were highly resistant to penicillin. Penicillin nonsusceptibility increased from 45% to 56% among PCV7 serotypes while remaining stable among PCV7 potentially cross-reactive strains (51% vs 54%).
Conclusions. Pneumococcal colonization has changed after the introduction of PCV7, both in serotype distribution and in patterns of antibiotic resistance. The frequency of nonvaccine strains has increased, and the proportion of nonvaccine isolates that are not susceptible to penicillin has tripled. This shift toward increased carriage of nonvaccine serotypes warrants vigilance for changes in the epidemiology of invasive pneumococcal disease.
Key Words: Streptococcus pneumoniae pneumococcal conjugate vaccine antibiotic resistance colonization
Abbreviations: PCV7, heptavalent pneumococcal conjugate vaccine PNSP, penicillin nonsusceptible Streptococcus pneumoniae MDR, multidrug resistance AOM, acute otitis media
Before the release of a conjugate vaccine for US children, Streptococcus pneumoniae caused invasive disease in 1 in 500 to 600 children who were younger than 2 years.1 In addition, rising antibiotic resistance among pneumococcal strains was a major concern among public health officials.2,3 By 1999, 38% of laboratory isolates in children who were younger than 2 years were nonsusceptible to penicillin,4 with higher rates reported in local areas.5–7
In February 2000, heptavalent pneumococcal conjugate vaccine (PCV7) was released to provide immunity to the 7 serotypes responsible for 85% of pediatric invasive pneumococcal disease8 and 78% of penicillin-nonsusceptible S pneumoniae (PNSP) isolates in children.9 Prelicensure trials showed a 90% reduction in invasive disease10 and a 23% reduction in all-cause pneumonia in the first 2 years of life.11 Postlicensure studies have also shown declines in invasive pneumococcal disease from 188 per 100000 to 59 cases per 100000 in US children who are younger than 2 years.4,12
Despite the success of PCV7 in reducing invasive disease, critical questions remain about the changing pneumococcal reservoir. Some trials of immunization with PCV7 have shown a 50% to 65% decrease in carriage of vaccine serotypes13–15 and have raised concerns about replacement of vaccine serotypes with nonvaccine serotypes that may ultimately result in invasive disease.16,17 Although, currently, the majority of invasive disease is caused by vaccine-included serotypes, the relationship between carriage of and invasive disease is complex and varies by serotype.18,19 Shifts in colonizing serotypes after PCV7 may serve as early indicators of changes in invasive disease.
We assessed secular trends in community-level S pneumoniae carriage and resistance after the licensure of PCV7. Community-level analyses assess the full impact of both direct and indirect (herd immunity) effects of vaccines. In 2001, we conducted surveillance for pneumococcal carriage among generally healthy children in 16 distinct Massachusetts communities.20 At that time, just after introduction of PCV7, we found that 26% of children carried S pneumoniae overall and that PCV7 serotypes accounted for 36% of isolates.20 Resistance to penicillin (and other antimicrobials) was concentrated among PCV7 serotypes and potentially cross-reactive serotypes, with penicillin nonsusceptibility of 45% and 51%, respectively. By November 2003, all children through 3.5 years of age were born into the era of routine PCV7 immunization in infancy. In this context, we resampled the same 16 communities in the winter and spring of 2003–2004 to determine (1) changes in carriage of PCV serotypes, PCV7 cross-reactive serotypes, and non-PCV7 S pneumoniae serotypes; (2) changes in overall antimicrobial resistance; and (3) serotype-specific changes in antimicrobial resistance.
METHODS
Data Collection
Nasopharyngeal specimens were collected from young children in pediatric and family practice physician offices in 16 Massachusetts communities between November 17, 2003, and April 16, 2004. We had performed surveillance in the same 16 communities from March 13 to May 11, 2001, as part of the baseline microbiologic data collection for a randomized trial of community-level intervention to promote judicious antibiotic prescribing for children.20 These 16 communities were selected on the basis of physical geographic separation, evidence that few children crossed community boundaries to seek pediatric care, and diversity of size and demographic characteristics using available US Census data. In 2001, 31 private practice offices participated, and in 2004, 23 offices participated, all of which had participated in 2001.
In 2001, children were eligible for inclusion if they were younger than 7 years, resided in a study community, and presented for either a routine well-child or "sick visit" at a participating practice.20 Parental consent and nasopharyngeal swabs were obtained by either trained study personnel or trained office practice nurses. PCV7 vaccination history was recorded from the medical record. In 2004, to increase sample size, we modified criteria to include children who received care in the study practices and lived beyond the zip code boundaries of the community. Recruitment in 2004 was independent of participation in 2001. All study procedures were approved by the Harvard Pilgrim Health Care institutional review board.
Microbiologic Processing
Nasopharyngeal samples were processed for S pneumoniae growth, antibiotic susceptibility, and serotype as previously described.20 Standard National Committee for Clinical Laboratory Standards susceptibility breakpoints were used to classify organisms as susceptible, intermediate, and resistant.21 Two definitions of multidrug resistance (MDR) were used: resistance to the -lactam class plus 1 other antibiotic class (MDR1) and resistance to at least 3 antibiotic classes (MDR2). Penicillins, cephalosporins, and carbapenems were considered a single class. Comparisons of third-generation cephalosporin resistance were performed using 2004 breakpoint criteria.21,22 We had previously reported carriage data from 2001 using the breakpoint criteria at that time. These 2001 data were reanalyzed with 2004 criteria for this analysis.
Serotypes included in the PCV7 were considered vaccine serotypes, and serotypes from serogroups that were not in PCV7 were considered nonvaccine serotypes. Serotypes in the same serogroup as vaccine serotypes (eg, 6A and 6B both are serogroup 6 strains) but not specifically included in the vaccine were classified as potentially cross-reactive serotypes. The extent to which the vaccine provides protection in vivo varies by serotype, but there is likely to be clinically important cross-protection against some of these strains. For quality control, we reserotyped a 36% random sample (82 of 226) of pneumococcal isolates from 2004 in a blinded manner and calculated percentage concordance.
Data Analysis
Microbiologic data were analyzed using SAS software version 8.2 (SAS Institute, Cary, NC). Proportions of children who carried S pneumoniae and proportions of isolates that were nonsusceptible to antibiotics were calculated and compared between the 2004 and previously published 2001 sample20 using Fisher exact tests. Carriage proportions were calculated by sampling period and by age group (<6 months, 6–<12 months, 12–<24 months, 24–<36 months, and 36 months). Resistance patterns were analyzed for the 2004 sample, by vaccine group (PCV7, PCV7 cross-reactive, and non-PCV7 serotypes) and by individual serotype and compared with the 2001 sample.
RESULTS
We collected 996 nasopharyngeal samples from children in 23 practices from November 17, 2003, through April 16, 2004 (Table 1), 2 of which were lost in transit. Of the 994 remaining samples, 73% (726) came from children who lived within the specified 16 Massachusetts communities described in Table 2; the remainder were patients of these practices who lived outside the designated community zip codes. Because pneumococcal carriage and descriptive characteristics of these children were similar to those who lived within the designated zip codes, data were combined for analysis. Fifteen percent (153) of participants were siblings of other participants, compared with 12% (87) in 2001.
Pneumococcal carriage was similar between 2001 and 2004 (Table 2). Age-specific pneumococcal carriage increased in children who were younger than 6 months (8% in 2001 vs 22% in 2004; P = .01) and decreased in those who were older than 36 months (30% in 2001 vs 19% in 2004; P = .002). Overall carriage of Haemophilus influenzae was 15% in 2004 compared with 17% in 2001 (P = .4), and overall carriage of Moraxella catarrhalis was 26% in 2004 compared with 16% in 2001 (P < .0001).
Of the 232 S pneumoniae isolates, 226 were serotyped (6 were unable to be grown after transfer). The proportion of PCV7 serotypes declined from 36% (51 of 143) in 2001 to 14% (32 of 226) in 2004 (P < .0001), whereas non-PCV7 serotypes increased from 34% (49 of 143) in 2001 to 55% (125 of 226) in 2004 (P < .0001; Fig 1). The proportion of PCV7 cross-reactive isolates remained stable. In all age groups, the proportion of PCV7 strains carried decreased, whereas the proportion of non-PCV7 serotypes either remained the same (<6-month-olds) or increased (all other age groups). There was no difference in the proportion of serotypes included in the 23-valent adult pneumococcal polysaccharide vaccine between 2001 (57%; 82 of 143) and 2004 (50%; 114 of 226). Serotype concordance during repeat serotyping was 96.3%. Previous studies that compared concordance of pneumococcal serotyping between laboratories showed a similar serotype discordance level.23
Every PCV7-included serotype decreased as a proportion of isolates, except for 9V, which had no representation in 2001 and 1 isolate in 2004 (Fig 2). Serotype 19F showed minimal decline and thus increased proportionally among PCV7 serotypes from 31% (16 of 51) in 2001 to 63% (20 of 32; P = .007) in 2004. Among PCV7 cross-reactive serotypes, there was a significant decrease in the proportion of serotype 6A (53% [23 of 43] to 32% [22 of 69]; P = .03) but significant increases in 19A (14% [6 of 43] to 38% [26 of 69]; P = .009) and 23A (2% [1 of 43] to 20% [14 to 69]; P = .008).
The most common non-PCV7 serotypes (N = 125) in 2004 were 15 (N = 34), 29 (N = 19), 11A (N = 18), 22F (N = 9), 10A (N = 7), and 35F (N = 7). Between 2001 and 2004, there was a significant increase in serotype 29 (2% in 2001 vs 8% in 2004; P = .04) and a trend toward an increase in serotype 15 (8% in 2001 vs 15% in 2004; P = .07). There were insufficient numbers to detect a significant change in the remaining serotypes.
Vaccine penetration (receipt of at least 1 PCV7 dose) increased from 60% in 2001 to 96% in 2004 among children who were younger than 12 months and from 38% to 79% among children who were at least 12 months of age. Among children who were at least 12 months of age and received 1 or more doses of PCV7, non-PCV7 strains increased from 8% (16 of 191) in 2001 to 14% (87 of 623) in 2004 (P = .05). Children who were at least 12 months of age and had not received any doses of PCV7 (primarily older children who were born before the introduction of PCV7) had similar percentages of non-PCV7 carriage in 2001 (7%; 20 of 306) as compared with 2004 (10%; 16 of 164; P = .27). In 2001, overall carriage of PCV7 serotypes among those who were at least 12 months of age was low among both vaccine recipients (5%; 10 of 191) and nonrecipients (3%; 8 of 306). No reduction in overall carriage of vaccine serotypes was seen in 2004, although the proportion of PCV7 serotypes among carriers declined.
Overall carriage of a PNSP isolate was similar in 2001 (7%; 49 of 695) and 2004 (9%; 86 of 988). However, penicillin nonsusceptibility declined among PCV7 isolates and increased among non-PCV7 isolates (Fig 1). The former reflected a decrease in high-level penicillin-resistant isolates, whereas the latter reflected an increase in intermediate resistance to penicillin (Table 3).
Penicillin nonsusceptibility varied among commonly carried serotypes. In 2004, the proportion of PNSP was 64% in 6A, 0% in 11A, 38% in 15, 65% in 19A, 65% in 19F, 36% in 23A, and 74% in 29 (see Fig 2).
Changes in susceptibility to additional antibiotics is shown in Table 3. Ceftriaxone nonsusceptibility declined significantly between 2001 and 2004 isolates, largely as a result of declines in intermediate resistance (14% in 2001 vs 0.4% in 2004; P < .0001). This decline was seen across PCV7, PCV7 cross-reactive, and non-PCV7 serogroups, even when accounting for the fact that National Committee for Clinical Laboratory Standards breakpoints had changed between 2001 and 2004.22 Trimethoprim-sulfamethoxazole nonsusceptibility also declined significantly. On the contrary, clindamycin nonsusceptibility increased, particularly among non-PCV7 serotypes. Clindamycin nonsusceptibility was highly correlated with macrolide nonsusceptibility (6 of 6 in 2001; 20 of 21 in 2004). Fluoroquinolone nonsusceptibility was minimal in both 2001 (3%; 4 of 143) and 2004 (0%; 0 of 226). Notably, among non-PCV7 serotypes, nonsusceptibility significantly increased for penicillin, clindamycin, and MDR (Table 3). The proportion of isolates that were resistant to 3 or more antibiotic classes increased from 9% (4 of 43) to 20% (14 of 69) among PCV7 cross-reactive serotypes and from 2% (1 of 49) to 10% (13 of 125) among non-PCV7 serotypes.
DISCUSSION
The licensure of PCV7 was accompanied by projections of decreased S pneumoniae carriage, invasive disease, and antibiotic resistance.13,15,17 When evaluating young healthy children in the same 16 Massachusetts communities in 2001 and 2004, we found significantly increased carriage of nonvaccine strains and increased antibiotic resistance among nonvaccine serotypes.
Despite the significant decline in PCV7 serotypes, overall pneumococcal carriage did not change between 2001 and 2004 because of significant increases in the carriage of non-PCV7 serotypes. This serotype replacement was seen in all pediatric age groups studied and resulted in similar proportions of pneumococcal carriage in age groups between 6 and 36 months for both sampling periods. Among children who were 36 months of age, overall carriage declined between 2001 and 2004. This is not surprising given differential vaccination of children of this age in 2004 compared with 2001 (unlikely to be vaccinated unless at high risk). By contrast, we have no clear explanation for the observed increased rate of carriage among young infants (<6 months).
These changes suggest a rapid replacement of vaccine serotypes by nonvaccine serotypes, either by the unmasking of previous minority strains24 or by replacement of previous PCV7 carriage by serotypes unaffected by vaccine immunity. Another possibility is serotype switch,25 whereby pneumococcal strains express a new capsular serotype, possibly to evade host immunity. This phenomenon has been described between 15B and 15C,26 19F and 23F,27 9V and 11A,28 and others.25
Although serotype replacement has been a concern after the release of the conjugate pneumococcal vaccine, experts hoped that antibiotic resistance would decrease because resistance had been highly concentrated in PCV7 and PCV7 cross-reactive serotypes.1,20,29 As hoped, our serial community-based sampling showed that high-level penicillin resistance declined 38% and nonsusceptibility to third-generation cephalosporins decreased 88% across all isolates. However, this reduction in high-level penicillin resistance, produced by the reduction in carriage of certain PCV7 and PCV7 cross-reactive serotypes, was offset by increased carriage of nonvaccine serotypes, in which intermediate penicillin resistance has risen significantly. Antibiotic resistance among non-PCV7 pneumococcal serotypes (which had been negligible in 2001) increased to 25%, and resistance to 3 or more antibiotic classes increased to 10%.
In addition to increasing intermediate penicillin resistance, non-PCV7 strains showed an increase in clindamycin nonsusceptibility that was highly associated with macrolide resistance. Until now, most S pneumoniae isolates that are resistant to macrolides in the United States have displayed the M phenotype, related to a macrolide-specific efflux mechanism.30 The mechanism of resistance associated with dual macrolide and clindamycin resistance is related to the MLS phenotype, which is a result of target modification at the 50S ribosomal subunit.30 As the prevalence of non-PCV7 carriage increases, antibiotic exposure may be providing a selective pressure toward the emergence of additional resistance in these serotypes.
These data must be interpreted with the following caveats. First, we assume that carriage among children who seek care from primary care providers reflects carriage in their community. Although this assumption depends on the distribution of patients in a given practice, we believe it to be better than sampling schemes in child care settings that likely overrepresent shared colonizing strains. Second, the surveillance period occurred later in the season in 2001 compared with 2004 and may have affected carriage prevalence slightly, although we expect the proportion of serotypes among carriers to be unaffected. Third, because we sampled only twice, it is possible that we have observed natural fluctuations in the pneumococcal carriage state, rather than a trend related to the introduction of pneumococcal conjugate vaccine. We believe this unlikely given the magnitude and consistent direction of the shifts.
These data provide insight into the impact of PCV7 on pneumococcal carriage in young children. All communities included both immunized and unimmunized children, but, by 2004, all children who were younger than 42 months were born after routine PCV7 immunization began. As reflected in our study subjects, Massachusetts has one of the highest documented immunization rates in the United States.31 In this sample, we find evidence for increased carriage of non-PCV7 serotypes among vaccinated children who were older than 12 months.
It remains to be seen whether invasive infections caused by non-PCV serotypes will become more common. Recent data32 suggest that nonvaccine serotypes, including novel ones,33,34 may become important pathogens and that increased antibiotic resistance among these strains33,34 may have adverse consequences for children over time. Continued evaluation of the pneumococcal reservoir may presage changes in strains that are responsible for localized and invasive disease as a result of the widespread use of PCV7. Such information may inform clinicians' expectations of antibiotic resistance in pneumococcal infections as well as development of the next generation of conjugate pneumococcal vaccines.
ACKNOWLEDGMENTS
This study was funded by the US Agency for Healthcare Research and Quality (grant HS10247).
We thank the nurses and research assistants who made the nasopharyngeal sampling possible in our 16 Massachusetts communities: Andrea Gayle-Bennet, PA; Judy LaChance, RN; Patricia McCrann, RN; Verna Senna, RN; Martha Tata, RN; Susan DeAmario; Bobbi Fish; Sheila Lambi; Sara Lehroff; Ann Marie Rezendes; and Dan Steinberg. Also, we extend special thanks to each of the practices that participated in this study.
FOOTNOTES
Accepted Feb 8, 2005.
Conflict of interest: Dr Pelton receives research support from Wyeth Lederle.
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