The bacterial causes of diarrhea can be frustrating to identify, and it is likely that many remain undiagnosed. The pathogenic potential of certain bacteria becomes less ambiguous when they are observed to intimately associate with intestinal epithelial cells. In the present study we sought to retrospectively characterize the clinical, in situ molecular, and histopathological features of enteroadherent bacteria in seven unrelated kittens that were presumptively diagnosed with enteropathogenic Escherichia coli (EPEC) on the basis of postmortem light microscopic and, in some cases, microbiological examination. Characterization of the enteroadherent bacteria in each case was performed by Gram staining, in situ hybridization using fluorescence-labeled oligonucleotide probes, PCR amplification of species-specific gene sequences, and ultrastructural imaging applied to formalin-fixed paraffin-embedded sections of intestinal tissue. In only two kittens was EPEC infection confirmed. In the remaining five kittens, enteroadherent bacteria were identified as Enterococcus spp. The enterococci were further identified as Enterococcus hirae on the basis of PCR amplification of DNA extracted from the formalin-fixed, paraffin-embedded tissue and amplified by using species-specific primers. Transmission electron microscopy of representative lesions from E. coli-and Enterococcus spp.-infected kittens revealed coccobacilli adherent to intestinal epithelial cells without effacement of microvilli or cup-and-pedestal formation. Enterococci were not observed, nor were DNA sequences amplified from intestinal tissue obtained from age-matched kittens euthanized for reasons unrelated to intestinal disease. These studies suggest that E. hirae may be a common cause of enteroadherent bacterial infection in pre-weaning-age kittens and should be considered in the differential diagnosis of bacterial disease in this population.Diarrhea is a frequent clinical sign at the time of death or euthanasia of kittens, particularly those residing in shelters or rescue facilities. Enteritis is second only to the specific diagnosis of feline parvovirus infection as the most common cause of kitten mortality identified by histopathology-based studies (2). Aside from viral, protozoal, and helminthic causes of enteritis, the bacterial culprits of diarrhea are particularly problematic to identify. The intestinal tract harbors a diverse population of bacteria that play critical roles in nutrient assimilation, mucosal immunity, and colonization resistance. Recognition of pathogenic bacteria within this population is hampered by our limited knowledge of normal bacterial diversity, the challenge of distinguishing commensal from pathogenic bacteria, the frequent presence of pathogenic bacteria in clinically normal animals, and the ability of commensal bacteria to become pathogenic in genetically susceptible individuals or under abnormal environmental conditions. The pathogenic potential of certain bacteria becomes less ambiguous when they are observed to intimately adh...