Detection of Streptococcus pneumoniae Strain Cocolonization in the Nasopharynx

Abstract

Colonization with more than one distinct strain of the same species, also termed cocolonization, is a prerequisite for horizontal gene transfer between pneumococcal strains that may lead to change of the capsular serotype. Capsule switch has become an important issue since the introduction of conjugated pneumococcal polysaccharide vaccines. There is, however, a lack of techniques to detect multiple colonization by S. pneumoniae strains directly in nasopharyngeal samples. Two hundred eighty-seven nasopharyngeal swabs collected during the prevaccine era within a nationwide surveillance program were analyzed by a novel technique for the detection of cocolonization, based on PCR amplification of a noncoding region adjacent to the pneumolysin gene (plyNCR) and restriction fragment length polymorphism (RFLP) analysis. The numbers of strains and their relative abundance in cocolonized samples were determined by terminal RFLP. The pneumococcal carriage rate found by PCR was 51.6%, compared to 40.0% found by culture. Cocolonization was present in 9.5% (10/105) of samples, most (9/10) of which contained two strains in a ratio of between 1:1 and 17:1. Five of the 10 cocolonized samples showed combinations of vaccine types only (n ‫؍‬ 2) or combinations of nonvaccine types only (n ‫؍‬ 3). Carriers of multiple pneumococcal strains had received recent antibiotic treatment more often than those colonized with a single strain (33% versus 9%, P ‫؍‬ 0.025). This new technique allows for the rapid and economical study of pneumococcal cocolonization in nasopharyngeal swabs. It will be valuable for the surveillance of S. pneumoniae epidemiology under vaccine selection pressure.Streptococcus pneumoniae is among the most important of human pathogens and is responsible for bacterial meningitis, sepsis, pneumonia, and acute otitis media. The habitat of the pneumococcus is the mucosa of the human nasopharynx. Colonization of the nasopharynx occurs early in life, with a prevalence of about 40% in infants and 15% in adults (depending on the local epidemiology). A single strain can persist in the nasopharynx for weeks or months, to then be replaced by other strains. Colonization is the starting point for all relevant aspects of this pathogen, such as invasive disease, exchange of genetic material, genetic recombination, and transmission. By the age of two, more than 95% of children have been colonized with up to six different serotypes (8). Sometimes more than one pneumococcal strain colonizes the nasopharynx at the same time. This phenomenon is known as cocolonization, or multiple colonization with more than one distinct strain of the same species. Cocolonization is probably required for horizontal gene transfer between different pneumococcal strains. Such genetic exchange has been shown to occur for the capsule gene locus and has been observed in the past for dominant international multiresistant pneumococcal clones (5). There are few data on rates of multiple colonization, but existing estimates range from 1.3% (13) up to 20% (9, 26). No...

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