E D I T O R I A L C O M M E N TA R Y

Reduction of Carriage and Transmission of Streptococcus pneumoniae: The Beneficial “Side Effect” of Pneumococcal Conjugate Vaccine Chiara Azzari and Massimo Resti Department of Pediatrics, Anna Meyer Children’s Hospital, University of Florence, Florence, Italy

(See the article by Millar et al. on pages 989–96)

In 2000, the 7-valent polysaccharide pneumococcal conjugate vaccine (PCV7) was recommended in the United States for routine use in all infants and children aged !2 years and in high-risk children aged ⭐4 years [1]. Several controlled clinical trials have shown that the vaccine is highly efficacious in preventing invasive pneumococcal disease (IPD) in vaccinated children. The incidence of vaccine-type IPD among children aged !5 years decreased 94% from 1998 to 2003 [2] and plateaued in 2002–2005 [3]. Use of the vaccine also reduced the incidence of IPD among unvaccinated populations. In fact, from 1998 to 2003, the incidence of vaccine-type IPD decreased up to 60% among persons aged 15 years [2–4], and a statistically significant decrease of ∼40% among infants too young to be vaccinated was also found [5]. These data suggest a key role for indirect effects of PCV7, and it has been suggested that both reduction in nasopharyngeal carriage and reduction in transmission of Received 30 June 2008; accepted 1 July 2008; electronically published 9 September 2008. Reprints or correspondence: Prof. Chiara Azzari, Dept. of Pediatrics, Anna Meyer Children’s Hospital, Viale Pieraccini 24, 50139 Firenze, Italy ([email protected]). Clinical Infectious Diseases 2008; 47:997–9  2008 by the Infectious Diseases Society of America. All rights reserved. 1058-4838/2008/4708-0002$15.00 DOI: 10.1086/591967

vaccine-type pneumococci could be possible mechanisms of herd immunity to Streptococcus pneumoniae. Nasopharyngeal carriage at every age is well documented, but it is particularly frequent among children aged !2 years, children attending child day care centers [6], children living with tobacco-smoking parents, and children whose families have a higher number of preschool-attending children. However, the true role of nasopharyngeal carriage in herd immunity still needs to be completely clarified. In this issue of Clinical Infectious Diseases, Millar et al. [7] report data on the indirect effect of PCV7 on carriage among unvaccinated adults and children living with children who have been vaccinated. In their study, conducted at the Navajo and White Mountain Apache reservations from 1997 through 2000, they evaluated both vaccinated and unvaccinated children and their household members (children aged !18 years and adults aged ⭓18 years) for a study of carriage. They collected single nasopharyngeal swab specimens from the children and both nasopharyngeal and oropharyngeal swab specimens from the adults. Isolates, obtained on sheep-blood agar plates, were serotyped by the Quellung reaction. Pneumococcal serotypes were divided into vaccine-type serotypes (4, 6B, 9V, 14,

18C, 19F, and 23F), vaccine-associated serotypes (serotypes in serogroups 6, 9, 18, 19 or 23 that are not vaccine types), and non–vaccine type/non–vaccine associated (NVT/NVA) serotypes (all others). The results by Millar et al. [7] demonstrate that adults and unvaccinated children aged !5 years who were living in households with a PCV7 vaccinee were less likely to be colonized with vaccinetype pneumococci than were those living in a household with unvaccinated children. These results are consistent with those obtained by Hammitt et al. [8] in a community-based, observational study of adult carriage, and a similar conclusion was reached, that the protective effect in unvaccinated populations is attributable to reduced exposure and reduced transmission when exposure occurs. Millar et al. [7] also observed that there was no difference in carriage for children aged 5– 17 years and comment that the degree of indirect protection may be smaller for this age group because the older children attend school and thus have much less exposure to the youngest children than do the adults. Which mechanisms are involved in this indirect effect? The hypotheses suggest first the involvement of reduced acquisition of vaccine-type serotype pneumo-

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cocci among vaccinated individuals and, therefore, the absence of transmission to households. Second, there is a reduced transmissibility from vaccinated individuals, even when they are colonized with vaccine-type serotypes, because of a low density of carriage. Third, the authors speculate a reduction in carriage duration. Millar et al. [7] acknowledge some limitations of their study. First, some carriers could have been missed because of the use of a low-sensitivity sampling procedure and detection method. Moreover, the duration of carriage and density of carriage were not evaluated. One point in the study by Millar et al. [7] that deserves discussion is the increase in NVT/NVA serotypes among children aged !5 years who were vaccinated with PCV7. The evaluation of serotype replacement was not one of the aims of the study; although this result needs to be researched further, some comments should be made because it is a key point in epidemiological evaluation and in prediction of vaccine efficacy. In the study by Millar et al. [7] and in other studies of carriage, the diagnostic procedure used might cause a bias that could affect results of an evaluation of serotype replacement. The authors report that a single colony was selected from each culture plate to determine serotype by the Quellung reaction, and when colonies of multiple morphologies were present, each morphological type was serotyped. But, as the title of an article by Charalambous et al. [9] asks, “How valid is single colony isolation for surveillance of Streptococcus pneumoniae carriage?” By the single-colony culture method, a single serotype per patient can be found, and multiple colonization, which is a very frequent condition, cannot be revealed. Thus, the singlecolony method should be used, as by Millar et al. [7], only to seek average prevalence of common serotypes. On the other hand, if we are interested in finding all circulating serotypes or if we want to draw conclusions about limited phenomena, such as initial serotype replacement, in-

dividuation of multiple colonization is essential. Selection by morphology adds some information but, nevertheless, underestimates the true rate of multiple carriage [10]. This point has important implications for any carriage study. In parallel with a decrease in vaccine-type serotypes, Millar et al. [7] describe an increase in NVT/NVA serotypes. The crucial point is that a serotype replacement may be the first step toward a loss in vaccine efficacy. It has been proposed [11] that what appears to be a replacement of nasopharyngeal pneumococcal serotypes is nothing other that an “unmasking” phenomenon. When both vaccine-type and NVT/NVA serotypes are present at the same time in the nasopharynx, a single-colony method can individuate only the serotype with the higher density. After elimination of vaccine-type serotypes through vaccination, only NVT/NVA serotypes will be detected, and the result could be interpreted erroneously as a replacement by a different serotype. It is not easy to individuate multiple colonization by culture methods. First, culture has been demonstrated to have little sensitivity in evaluating multiple infections in respiratory samples, and, second, it is extremely expensive. How can the problem be solved? Many authors have demonstrated that molecular methods, such as PCR and real-time PCR, are useful tools and can be used to increase diagnostic sensitivity for pneumococcal disease or colonization, because timely and accurate detection does not depend on the microbiological viability of the organism [12–16]. Use of multiplex PCR directly on clinical samples [13, 15] could be the way to overcome the diagnostic limitation of culture methods in carriage studies and to obtain a complete epidemiological picture of pneumococcal serotypes. The lower cost of molecular methods creates an opportunity for a large number of researchers in all areas of the world, such that the use of these methods has been considered to be a complement to traditional meth-

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ods for surveillance of infectious diseases. Moreover, quantitative molecular methods could provide information on the density of different serotypes in the same sample. However, as with any new test, criteria of quality must be carefully observed [17] before the methods are transferred from a research setting to wider use. Doubts have been raised about whether bacterial DNA found in biological samples could derive from nonviable and therefore irrelevant bacteria. Experimental data suggest that DNA from nonviable bacteria is removed from respiratory specimens in a few days, so that the presence of bacterial DNA should be considered evidence of very recent or ongoing colonization and/ or infection [18]. In conclusion, investigation of multiple colonization is essential for all studies aimed at evaluating pneumococcal serotype replacement, either spontaneous [19, 20] or vaccine-modulated [3] replacement. The study by Millar et al. [7] makes an important contribution to furthering our understanding of the mechanisms involved in the indirect protection given by PCV7. Carriage studies are important because colonization of the nasopharynx is the first step in the pathogenesis of IPD. IPD results from a complex network of events that is still incompletely clarified. It is known that the ability to cause IPD is different among different pneumococcal clones and serotypes [21, 22]. However, acquisition of a new strain is a sine qua non for development of subsequent IPD. Moreover, recent studies have demonstrated that a high bacterial load detected by real-time PCR at nonsterile sites can help in predicting an etiological role for S. pneumoniae in invasive bacterial diseases [23, 24]. If these preliminary results can be confirmed, they will increase the importance of molecular methods in pneumococcal diagnosis. Carriage studies provide information on serotype distribution and allow monitoring of biological pressure derived from antibiotic therapy or vaccinations. The study by Millar et al. [7] has shed light on the mechanisms that

allow PCV7 to reach a high protective efficacy in unvaccinated populations, demonstrating the role of both reduced transmission from vaccinated carriers and reduced frequency of carriage. Other aspects remain to be studied: the duration of carriage for single serotypes, the relationship between carriage density and the risk of IPD, and antibiotic resistance, for which culture methods remain irreplaceable. In conclusion, in accord with Millar et al. [7], we think that continued surveillance of pneumococcal carriage and IPD for serotype changes and possible serotype replacement is essential, and if performed with specific and sensitive methods— among which PCR and real-time PCR should absolutely be considered—it will help us predict the impact of PCV7 and new pneumococcal conjugate vaccines.

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Acknowledgments Potential conflicts of interest. C.A. and M.R.: no conflicts.

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