Effect of Adenovirus Type 1 and Influenza a Virus onStreptococcus PneumoniaeNasopharyngeal Colonization and Otitis Media in the Chinchilla

Abstract

Considerable evidence has implicated respiratory tract virus potentiation of bacterial adherence, colonization, and superinfection as a significant factor contributing to the pathogenesis of otitis media (OM). Influenza A and B viruses, adenovirus, and respiratory syncytial virus are the primary respiratory tract viruses associated with this disease. Investigations have established a dramatic increase in the development of experimental OM in chinchillas co-inoculated with influenza A virus and Streptococcus pneumoniae (Spn). The mechanism underlying this phenomenon was suggested to involve, in part, viral compromise of eustachian tube mucosal integrity and function. This study was designed to assess and compare the effect of adenovirus and influenza A virus infection on adherence, the kinetics of colonization, and invasion of the middle ear by Spn in the chinchilla model of OM. Cohorts were inoculated intranasally with adenovirus type 1 or influenza A virus, and then inoculated intranasally 7 days later with Spn 6A. All cohorts were observed over a 14-day period after challenge with Spn, and the incidence and severity of OM were assessed by several methods, including culture of the nasopharynx and middle ear effusions. The data indicated that influenza A virus promotes a significant increase in nasopharyngeal colonization by Spn, an increased incidence and severity of OM, and a sustained presence of Spn in the effusions. Adenovirus infection, however, did not enhance colonization by Spn or result in an increased incidence or severity of OM.

Expression of Cytokine and Chemokine Genes by Human Middle Ear Epithelial Cells Induced by Influenza A Virus andStreptococcus pneumoniaeOpacity Variants

“…Influenza virus A/Alaskan (6/77) (H 3 N 2 ) was propagated and its titers were determined by a plaque assay as was previously described in detail and previously used by our laboratory (9,31,39,41). Our primary cultures of HMEE cells were infected with influenza A virus at the multiplicities of infection (MOIs) of 0.01 and 0.1 in serum-free alpha minimal essential medium (MEM␣) (Gibco BRL, Gaithersburg, Md.)…”

“…S. pneumoniae type 6A (EF 3114), kindly provided by B. Andersson, Department of Clinical Immunology, University of Gröteborg, Gröteborg, Sweden, has been described in detail previously (3). This strain has been used in previous reports from our laboratory (39,40). The isogenic opaque and transparent variants of S. pneumoniae type 6A were also used in a previous report from our laboratory (41) and originally isolated by Jeffery Weiser, Children's Hospital of Philadelphia.…”

Immunization with Native or Recombinant Streptococcus pneumoniae Neuraminidase Affords Protection in the Chinchilla Otitis Media Model

“…Neuraminidase was purified from whole-cell lysates as previously described by Lock et al (12). S. pneumoniae 6A (EF3114) was used for purification of native neuraminidase as well as intranasal inoculation in the chinchilla model of OM and has been described in detail previously (1,22). Fractions from a DEAESepharose column containing neuraminidase (identified by use of a neuraminidase assay described below) were pooled, washed, and concentrated at 4°C with buffer D (50 mM Tris base, 0.15 M NaCl, 0.1 mM Na-EDTA, 200 mM phenylmethylsulfonyl fluoride, 0.01% sodium azide) by using an Amicon stirred ultrafiltration cell model 8200 fitted with a polyethersulfone membrane with a 50,000 molecular weight cutoff (Millipore Corporation, Bedford, Mass.).…”

“…Influenza virus A/Alaska/6/77 (H3N2) has previously been used in combination with S. pneumoniae by our laboratory and others to induce experimental OM in the chinchilla model (8,25), and it has been described in detail previously (6,16). Fourteen days following the final immunization with neuraminidase, the chinchillas were anesthetized and inoculated intranasally with approximately 6 ϫ 10 6 PFU of influenza A virus per our standard protocol (22).…”

“…For each chinchilla, tympanic membrane inflammation was assessed in both ears by means of an otoscope by using the rating scale previously described (22). Animals with significant to severe erythema, retraction, and middle ear fluid (obtained by tympanocentesis) were considered to have developed OME.…”