Pneumococcal Infection among Children before Introduction of 13-Valent Pneumococcal Conjugate Vaccine , Cambodia

I with Streptococcus pneumoniae remains a substantial cause of death among children (1). In high-income countries, introduction of pneumococcal conjugate vaccine (PCV) has substantially decreased incidence of invasive pneumococcal disease (IPD) (2). Data for PCV effect in low-income countries are less robust (2). We therefore studied the characteristics of pneumococci responsible for colonization and invasive disease among children in Cambodia before the early 2015 introduction of 13-valent PCV (PCV13).

I nfection with Streptococcus pneumoniae remains a substantial cause of death among children (1). In high-income countries, introduction of pneumococcal conjugate vaccine (PCV) has substantially decreased incidence of invasive pneumococcal disease (IPD) (2). Data for PCV effect in low-income countries are less robust (2). We therefore studied the characteristics of pneumococci responsible for colonization and invasive disease among children in Cambodia before the early 2015 introduction of 13-valent PCV (PCV13).

The Study
The study was conducted at Angkor Hospital for Children, Siem Reap, Cambodia. Before enrollment of a child, written consent was obtained from the parent/guardian. Ethical approval was granted by the hospital institutional review board and the Oxford Tropical Research Ethics Committee. For the colonization study, which was conducted in January (cool/dry season) and August (hot/wet season) 2014, colonization surveys were conducted in the outpatient department. Nasopharyngeal swab samples were collected from children 1 month to 15 years of age who had minor illnesses, excluding nonsevere pneumonia, not requiring hospital admission. Children were eligible for enrollment 1 time per survey. For the invasive disease study, which was conducted during August 1, 2013-July 31, 2014, samples were collected from hospitalized children 1 month to 15 years of age who met World Health Organization (WHO) clinical case definitions for pneumonia, meningitis, or sepsis (3). Children readmitted within 14 days were excluded from reenrollment. Samples were processed according to the WHO pneumococcal colonization detection protocol (4). Pneumococci were confirmed by optochin susceptibility and/or bile solubility and were serotyped by latex agglutination (5). Antimicrobial drug susceptibilities were determined according to Clinical and Laboratory Standards Institute guidelines (6). Serotype and antimicrobial drug susceptibilities were also determined for all invasive pneumococcal isolates cultured from patients during January 1, 2013-December 1, 2014. Pneumococci were grouped into vaccine serotypes (PCV13: 1, 3, 4, 5, 6A, 6B, 7F, 9V, 14, 18C, 19F, 19A, 23F), nonvaccine serotypes (all others), and nontypeable isolates. Multidrug resistance was defined as resistance to >3 agents (online Technical Appendix, http://wwwnc.cdc.gov/EID/article/21/11/15-0914-Techapp1.pdf) (7).
The outpatient colonization study included 974 children (Table 1; online Technical Appendix Figure 1). None were known to be HIV infected. Pneumococcal colonization was detected in 601 (61.7%) of children (online Technical Appendix Table 1). Colonization prevalence declined with age: 78.6% (206/262) in those 1-11 months, 61.9% (284/459) in those 12-59 months, and 43.9% (111/253) in those ≥5 years of age. The proportion colonized were 75.2% (342/455) in the cool/dry season and 49.9% (259/519) in the hot/wet season (p<0.001). The adjusted odds ratio for colonization in the hot/wet season was 0.38 (95% CI 0.28-0.51, p<0.001) after controlling for age, household size, cohabitation with other young children, current upper respiratory tract symptoms, and recent antimicrobial use. A total of 667 pneumococci were isolated (Figure 1 From August 1, 2013, through July 1, 2014, a total of 2,613 cases of medical admissions were screened; of these, 1,009 were included in the analysis (online Technical Appendix Figure 1). Median patient age at admission was 1.2 years (interquartile range 0.5-2.4), 56.5% (570/1,009) of patients were male, and 1.4% (14/1,006) were HIV positive. Most cases met the WHO category of severe pneumonia (online Technical Appendix Table 2). Pneumococcal colonization was identified in 29.1% (293/1,008) of children from whom a swab sample was obtained (online Technical Appendix  Table 4.

Conclusions
This study highlights the high potential for reduction of IPD among children after introduction of PCV13 in Cambodia; 88.4% (95% CI 74.9-96.1) of invasive isolates from this 1 surveillance site were serotypes covered by the vaccine. Vaccination should result in decreased drug-resistant pneumococcal infections, although the substantial reservoir of resistance in nonvaccine type and nontypeable pneumococci will probably erode any reduction over time (8)(9)(10). Colonization was high among outpatients and similar to that in other Southeast Asia locations (5,11). Multidrug resistance was common, probably the result of poor regulation of antimicrobial drug use in Cambodia (12); 72.1% of colonizing isolates and 55.8% of invasive isolates were multidrug resistant. For comparison, a recent study of children in Thailand found 31.6% of colonizing pneumococci to be multidrug resistant (13).
The range of serotypes detected in the colonization study was broad but slightly more restricted than that detected in other low-income country studies. In a longitudinal colonization study of refugee infants on the Thailand-Myanmar border, 67 serotypes were identified (5). This finding may reflect the high prevalence of antimicrobial drug use in the community, which would reduce the colonization prevalence of less resistant nonvaccine serotypes. However, the identification of several serotypes emerging as causes of IPD in South Africa, the United Kingdom, and the United States after introduction of PCV13 (e.g., serotypes 15A, 15B/C, 23B, 24F; which accounted for 7.8% of colonizing pneumococci in our study) is noteworthy, indicating the need for close monitoring for changes in colonization and IPD serotype distribution after PCV13 introduction (7,14,15).
The study has several limitations. The absolute number of IPD cases was small, and it was not possible to calculate disease incidence rates. The high prevalence of prehospitalization antimicrobial drug use hampered accurate IPD surveillance. Failure to detect more antimicrobial-drug susceptible nonvaccine type infections as a result of prehospitalization antimicrobial drug use may have falsely elevated the proportion of disease covered by PCV13. The low prevalence of colonization among hospitalized children highlights the need for swab sample collection before in-hospital antimicrobial drug administration for accurate evaluation of colonization in unwell children. Because the study was conducted at 1 site, caution is required when extrapolating the results to the general population of Cambodia. These data provide a baseline against which to monitor effectiveness of vaccinating children with PCV13 in Cambodia.     and 31 st December 2014 were retrieved from 80°C storage and sub-cultured onto 5% sheep blood agar for serotyping as described above. Antimicrobial susceptibilities for these isolates were extracted from the microbiology laboratory database.

Data analysis
Analyses were done using the R statistical package version 3.  (4). Resistance was defined as disk test "resistant" except for the β-lactam drugs: benzylpenicillin MICs of >0.06 µg/mL were classified as "resistant"; ceftriaxone MICs of >1 µg/mL were classified as "resistant." Multidrug resistance (MDR) was defined as resistance to three or more agents, with clindamycin/erythromycin and benzylpenicillin/ceftriaxone counting as a single agent (5).