Neonatal piglet diarrhoea associated with enteroadherent Enterococcus hirae

Neonatal porcine diarrhoea of uncertain aetiology is an increasing problem in several countries. The aim of the present study was to investigate the unexpected finding of enteroadherent cocci in the small intestine of piglets selected for necropsy examination from six herds (18 diarrhoeic piglets and 11 healthy controls). Gross and microscopical lesions were characterized and selected intestinal sections were further examined by immunohistochemistry for expression of active caspase-3. The enteroadherent bacterium was characterized in situ by Gram staining, ultrastructural imaging, fluorescence in-situ hybridization (FISH) and 16S rRNA gene analysis. Species identification of enterococci from intestinal cultures was performed by matrix-assisted laser desorption ionization time-of-flight mass spectrometry (MALDI-TOF MS) for one diarrhoeic and one control animal per herd. Gross changes were mild. Microscopically, small intestinal colonization by gram-positive cocci was observed in diarrhoeic animals only and was accompanied by villus atrophy (4/18) and mild epithelial lesions (10/18), including increased apoptosis of enterocytes. Transmission electron microscopy revealed coccoid bacteria adjacent to the epithelium, but without effacement of microvilli. 16S rRNA gene analysis yielded a sequence identical to Enterococcus hirae and FISH identified the enteroadherent bacteria as Enterococcus spp. in all colonized animals. The proportion of bacterial isolates identified as E. hirae by MALDI-TOF MS analysis was significantly higher (P = 0.0138) in diarrhoeic pigs. Species identification was confirmed by species-specific polymerase chain reaction for one E. hirae isolate per herd. These isolates were further tested for antimicrobial susceptibility, which indicated decreased susceptibility to ciprofloxacin for one isolate (minimum inhibitory concentration >4 mg/l). These findings suggested that neonatal porcine diarrhoea was associated with small intestinal colonization by E. hirae accompanied by mucosal lesions.