Fusobacterium Isolates Recovered From Colonic Biopsies of Inflammatory Bowel Disease Patients in Korea

Abstract

Dear Editor, Inflammatory bowel diseases (IBDs) such as Crohn’s disease (CD) and ulcerative colitis (UC) are chronic relapsing inflammatory disorders of the intestine that are characterized by abdominal pain and diarrhea [1, 2]. The incidence of IBD is increasing globally and has also increased in Korea recently [2, 3]. Although the etiology is unknown, the relationship between the colonic microbiota and IBD has been indicated as a possible contributor. Fusobacterium varium and F. nucleatum have emerged as compelling candidates responsible for IBD exacerbation [4, 5]. The colonization by these strains is possibly a useful biomarker for diagnosing gastrointestinal diseases [5]. We aimed to determine the prevalence of Fusobacterium spp. from colonic biopsies of IBD patients in Korea. All 54 patients fulfilling the diagnostic criteria for IBD including 26 of CD, 25 of UC, or 3 of Behcet’s disease (BD), at Hanyang University Guri Hospital between June 2014 and June 2015 were included in this study. Among them, 41 (76%) were male. The patient ages ranged from 14 to 75 yr (median: 31 yr). The Institutional Review Board of Hanyang University Guri Hospital approved this study. Biopsy material was obtained from patients undergoing colonoscopy for assessment of IBD status or for confirmation of gastrointestinal disease. Tissues were put into pre-reduced phosphate-buffered saline and immediately transported to the laboratory for bacterial culture. The specimens were ground, inoculated on crystal violet erythromycin (CVE) agar supplemented with tryptophan, and incubated at 37°C for 48 hr in an anaerobic jar [6]. All isolates showing a different colony morphology were identified by 16S rRNA gene sequencing by using PCR predicted to yield a product of 1,300 bp [7]. The PCR conditions were as follows: 95°C for 5 min; 30 cycles at 95°C for 30 sec, 55°C for 30 sec, and 72°C for 1 min; and a final 10-min step at 72°C. The sequencing of the amplification products were performed in a commercial facility (Macrogen, Seoul, Korea). The sequencing analysis was completed using the GenBank and EzTaxon databases. Some strains were identified using the MicroScan WalkAway automated system (Dade Behring, West Sacramento, CA, USA) and the Vitek MS system (bioMérieux, Marcy l’Etoile, France). Ten Fusobacterium isolates were recovered from nine of the 54 patients as follows: six F. mortiferum, two F. varium, and one F. nucleatum and F. mortiferum (Table 1). Clinically, the most important Fusobacterium species are F. nucleatum and F. necrophorum, but F. ulcerans and the F. mor-

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