Effect of influenza A virus infection on nasopharyngeal colonization and otitis media induced by transparent or opaque phenotype variants of Streptococcus pneumoniae in the chinchilla model

Abstract

Phase variation in the colonial opacity of Streptococcus pneumoniae has been implicated as a factor in bacterial adherence, colonization, and invasion in the pathogenesis of pneumococcal disease. Additionally, the synergistic effects of influenza A virus and S. pneumoniae in the development of otitis media (OM) have been reported. This study examined the ability of opaque or transparent S. pneumoniae from the same strain in combination with an antecedent influenza A virus infection to colonize the nasopharynx and invade the middle ear in the chinchilla model. Our data indicated that there was no significant difference in the level of nasopharyngeal colonization and induction of OM between the opaque and transparent variants unless there was a prior challenge with influenza A virus. Subsequent to influenza A virus infection, there was a significant difference between the variants in the ability to colonize and persist in the nasopharynx and middle ear. The concentrations of the opaque variant in nasopharyngeal-lavage samples and middle-ear fluid remained consistently higher than those of the transparent variant for 10 days postinoculation. Data from this study indicate that the effects of influenza A virus on the pathogenesis of experimental S. pneumoniae-induced OM differ depending on the opacity phenotype involved.

FIG. 1

NP colonization dynamics in chinchillas inoculated i.n. with either opaque or transparent S. pneumoniae (Spn) type 6A 7 days after i.n. challenge with either diluent without influenza A virus (Influ A) or diluent with Influ A. Each data point represents the geometric mean number of CFU of S. pneumoniae bacteria (± the standard error of the mean) per milliliter of nasal-lavage fluid from a total of six animals combined from two separate experiments. ∗, P < 0.05 compared to the cohort inoculated with the transparent variant and Influ A.

FIG. 2

Survival of the opaque or transparent phenotype in the middle ears of chinchillas subsequent to i.n. inoculation with S. pneumoniae (Spn) type 6A 7 days after i.n. challenge with influenza A virus (Influ A). Each data point represents the geometric mean number of CFU of S. pneumoniae bacteria (± the standard error of the mean) per milliliter of MEF or middle-ear-lavage fluid from a total of two to four animals combined from two separate experiments. ∗, P < 0.05 compared to the cohort inoculated with the opaque variant and Influ A.

FIG. 3

Comparison of S. pneumoniae (Spn) concentrations in the middle ears for cohorts inoculated i.n. with the opaque or transparent variant of S. pneumoniae type 6A after a prior i.n. inoculation with influenza A virus (Influ A). Horizontal bars, median values; ∗, P = 0.02.

FIG. 4

Comparison of mean MEPs (± standard errors of the means) for cohorts inoculated with the S. pneumoniae (Spn) type 6A opaque or transparent phenotype either with or without prior i.n. inoculation with influenza A virus (Influ A). MEPs were determined by tympanometry over a 10-day observation period. Values below −60 daPa are considered abnormal for the chinchilla (17). ∗, P < 0.05 for the comparison of cohorts inoculated with influenza A virus and the opaque variant versus influenza A virus and the transparent variant.