Scanning electron microscopy and fluorescent in situ hybridization of experimental Brachyspira (Serpulina) pilosicoli infection in growing pigs

Two groups of six 8-week-old pigs were challenged with 1X10(9) cfu Brachysp ira (Serpulina) pilosicoli or Serpulina intermedia daily for 3 consecutive days to study the pathology of porcine colonic spirochetosis by scanning el ectron microscopy (SEM) and fluorescent in situ hybridization (FISH) with o ligonucleotide probes targeting ribosomal RNA specific for B. pilosicoli an d the genus Brachyspira/Serpulina. Six pigs served as noninoculated control s. The animals were euthanatized successively between postinoculation days 14 and 24. B. pilosicoli was reisolated in feces from all of the inoculated pigs; however, only two pigs developed transient watery diarrhea. S. inter media was reisolated from four of the inoculated pigs, but clinical signs w ere not observed. Gross examination of the B. pilosicoli-infected pigs reve aled dilated large intestines with a hyperemic mucosa, whereas the large in testines of the S. intermedia-inoculated pigs and the control pigs appeared normal. SEM examination of B. pilosicoli-infected pigs revealed degenerate d epithelial cells and spirochetal colonization of the colonic mucosa in fo ur pigs. By FISH, B. pilosicoli cells were found colonizing and invading th e surface epithelium and the crypts in all the pigs. Spirochetal crypt colo nization markedly exceeded the occurrence of spirochetes on the mucosal sur face. SEM examination of S, intermedia-inoculated pigs revealed no abnormal ities, and Serpulina cells were detected only sporadically in the otherwise normal appearing mucosa of four pigs by FISH. The results provide further evidence that B. pilosicoli is associated with colitis in pigs, although th e gross lesions are mild. The spirochete is capable of colonizing the large intestine, inducing mucosal damage, invasion of the crypt and surface epit helium, and focal infiltration of the lamina propria. In addition, the stud y shows the applicability of FISH for specific identification of B. pilosic oli in formalin-fixed tissue.