Gram-negative pathogen Klebsiella oxytoca is associated with spontaneous chronic otitis media in Toll-like receptor 4-deficient C3H/HeJ mice

CONCLUSION: This report confirms the presence of gram-negative Klebsiella bacteria in the middle ear of the C3H/HeJ mouse by culture, polymerase chain reaction (PCR), and electron microscopy. Identification of the bacterial pathogen supports the C3H/HeJ mouse as an excellent model for spontaneous chronic otitis media and its effects on the middle and inner ear. OBJECTIVES: The C3H/HeJ mouse has a single amino acid substitution in its Toll-like receptor 4, making it insensitive to endotoxin and suppressing initiation of the innate immune system. This study explored the bacteriology of the resultant middle ear infection by culture, PCR, histology, and electron microscopy. MATERIALS AND METHODS: Twelve-month-old C3H/ HeJ mice were screened positive for spontaneous otitis media. Tympanocentesis and blood cultures of mice were carried out under sedation. Middle ear aspirate material and blood samples were then sent for culture and PCR. Mice were then sacrificed for bright-field and electron microscopy analysis. RESULTS: All tympanocentesis and blood specimens grew gram-negative Klebsiella oxytoca, which was confirmed by PCR. Histopathology confirmed an intense inflammatory reaction and gram-negative bacteria in the middle and inner ears. Electron microscopy of the middle ears revealed abundant rod-shaped Klebsiella bacteria, both free and being engulfed by neutrophils.