Novel intestinal Helicobacter species isolated from cotton-top tamarins (Saguinus oedipus) with chronic colitis

Abstract

A disease similar to ulcerative colitis in humans has been identified in cotton-top tamarins (CTTs) in captivity. The clinical signs include weight loss, diarrhea, and rectal bleeding with the pathological features and biochemical abnormalities of ulcerative colitis. Approximately 25 to 40% of these animals develop colon cancer after 2 to 5 years of captivity. An infectious etiology has been proposed; however, no microbial agent to date has been identified. Helicobacter spp. have been associated with enterocolitis and inflammatory bowel disease (IBD) in humans and animals. Infection with Helicobacter pylori or Helicobacter mustelae is associated with an increased risk of gastric adenocarcinoma and lymphoma of the mucosa-associated lymphoid tissue. Helicobacter hepaticus causes hepatitis, hepatic adenomas, and hepatocellular carcinomas in susceptible strains of mice. The aim of this study was to assess a colony of CTTs with a high incidence of IBD and colon cancer for the presence of colonic Helicobacter spp. A fusiform, gram-negative bacterium with bipolar flagella and periplasmic fibers was isolated from the feces of CTTs. The bacterium grew under microaerobic conditions at 37 and 42 degrees C but not at 25 degrees C, did not hydrolyze urea, was positive for catalase and oxidase, did not reduce nitrate to nitrite, did not hydrolyze indoxyl acetate or alkaline phosphatase, and was resistant to nalidixic acid, cephalothin, and trimethoprim-sulfamethoxazole. On the basis of 16S rRNA gene sequence analysis, the organism was classified as a novel Helicobacter species. This is the first Helicobacter isolated from CTTs. Further studies are needed to elucidate the role of this novel Helicobacter sp. in the pathogenesis of ulcerative colitis and colonic adenocarcinoma in CTTs.

FIG. 1

Transmission electron micrograph of the novel Helicobacter sp. The typical bacterium is fusiform to slightly spiral and possesses periplasmic fibers and several sheathed flagella at each end. Bar, 0.5 μm.

FIG. 2

(A) Electrophoresis of DNA isolated from colonic biopsy samples, amplified by PCR with Helicobacter genus-specific primers, and run on a 1% agarose gel. Lane M, 100-bp DNA ladder; lane 1, MIT 97-6194-6; lane 2, MIT R97-6194-7; lane 3, MIT 97-6194-5; lane 4, MIT 97-6194-4; lane 5, MIT 97-6194-3; lane 6, MIT R97-6837; lane 7, MIT R97-6834; lane 8, MIT 97-6196-8; lane 9, MIT R97-6841; lane 10, MIT R97-6835; lane 11, MIT R97-6832; lane 12, MIT R97-6836; lane 13, blank; lane 14, positive control. (B) Electrophoresis of DNA isolated from fecal cultures, amplified by PCR with Helicobacter genus-specific primers, and run on a 1% agarose gel. The faint band in lane 2 at approximately 100 bases represents the PCR primers. Lane M, 100-bp DNA ladder; lane 1, positive control; lane 2, blank; lane 3, MIT 97-6194-4; lane 4, MIT 97-6194-3; lane 5, MIT 97-6194-5; lane 6, MIT R97-6834; lane 7, MIT R97-6841; lane 8, MIT R97-6837; lane 9, MIT R97-6840; lane 10, MIT R97-6842.

FIG. 3

Phylogenetic tree constructed on the basis of 16S rRNA sequence similarity values. The scale bar is equal to a 5% difference in nucleotide sequences as determined by measuring the lengths of the horizontal lines connecting two species.

FIG. 4

RFLP analysis of DNA isolated from pure fecal cultures, amplified by PCR with Helicobacter genus-specific primers, digested with AluI, and electrophoresed through a 3% agarose gel. Lane M, 100-bp DNA ladder; lane 1, MIT 97-6194-5; lane 2, MIT R97-6837; lane 3, MIT 97-6194-3; lane 4, MIT R97-6842; lane 5, MIT R97-6840.