Takayuki Kajiura

Changes of faecal microbiota in patients with Crohn's disease treated with an elemental diet and total parenteral nutrition.

BACKGROUND Intestinal microbiota contributes to the pathogenesis of Crohns disease. Elemental diet and total parenteral nutrition are effective therapies for Crohns disease; however, changes of microbiota as a result of both treatments have not been fully elucidated. AIM To elucidate changes of faecal microbiota in Crohns disease patients treated with elemental diet and total parenteral nutrition. METHODS Stool samples were collected from 33 active Crohns disease patients and 17 healthy subjects, and recollected after elemental diet (8 patients) and total parenteral nutrition (9 patients). Terminal restriction fragment length polymorphism analysis of bacterial 16srDNA was performed to evaluate the whole microbiota. Specific quantitative PCR was then used to determine populations of predominant bacterial groups. RESULTS In Crohns disease patients, the number of terminal restriction fragments, which reflects bacterial species, was significantly lower. Populations of total bacteria and Bifidobacterium were significantly lower and the ratio of Enterococcus was higher. The number of terminal restriction fragments was significantly decreased after total parenteral nutrition, but not after elemental diet. Population of Bacteroides fragilis significantly decreased after elemental diet, while population of Enterococcus significantly increased after total parenteral nutrition. CONCLUSION Faecal microbiota in Crohns disease patients was markedly different from healthy subjects. Species diversity was reduced by total parenteral nutrition, but not by elemental diet.

A new non-protein enediyne antibiotic N1999A2: Unique enediyne chromophore similar to neocarzinostatin and DNA cleavage feature

Abstract The present NMR and X-ray structural studies demonstrated that the new antibiotic N1999A2 isolated from the broth filtrate of Streptomyces sp. AJ9493, possesses a novel 9-membered ring enediyne chromophore similar to neocarzinostatin, but is not chromoprotein. Of special interest is the fact that stable N1999A2 exists as enediyne chromophore alone as well as dynemicin A, esperamicin A 1 and calicheamicin γ 1 . The major difference between N1999A2 and neocarzinostatin chromophore is lack of the amino sugar in N1999A2. The antibiotic N1999A2 revealed more random DNA cutting profile than neocarzinostatin chromophore.

Long-Term Alteration of Intestinal Microbiota in Patients with Ulcerative Colitis by Antibiotic Combination Therapy

Previous work has demonstrated that intestinal bacteria, such as Fusobacterium varium (F. varium), contribute to the clinical activity in ulcerative colitis (UC); thus, an antibiotic combination therapy (amoxicillin, tetracycline, and metronidazole (ATM)) against F. varium can induce and maintain UC remission. Therefore, we investigated whether ATM therapy induces a long-term alteration of intestinal microbiota in patients with UC. Patients with UC were enrolled in a multicenter, randomized, double-blind, placebo-controlled study. Biopsy samples at the beginning of the trial and again at 3 months after treatment completion were randomly obtained from 20 patients. The terminal restriction fragment length polymorphism (T-RFLP) in mucosa-associated bacterial components was examined to assess the alteration of the intestinal microbiota. Profile changes of T-RFLP in mucosa-associated bacterial components were found in 10 of 12 patients in the treatment group and in none of 8 in the placebo group. Dice similarity coefficients using the unweighted pair group method with arithmetic averages (Dice-UPGMA) confirmed that the similarity of mucosal microbiota from the descending colon was significantly decreased after the ATM therapy, and this change was maintained for at least 3 months. Moreover, at 3 months after treatment completion, the F. varium/β-actin ratio, examined by real-time PCR using nested PCR products from biopsy samples, was reduced less than 40% in 8 of 12 treated patients, which was higher, but not significantly, than in 4 of 8 patients in the placebo group. Together, these results suggest that ATM therapy induces long-term alterations in the intestinal microbiota of patients with UC, which may be associated, at least in part, with clinical effects of the therapy.

A newly discovered Anaerococcus strain responsible for axillary odor and a new axillary odor inhibitor, pentagalloyl glucose

Skin surface bacteria contribute to body odor, especially axillary odor. We aimed to investigate anaerobic bacteria that had not been previously studied for axillary odor formation. A new anaerobic Anaerococcus sp. A20, that releases 3-hydroxy-3-metyl-hexanoic acid (HMHA, main component of axillary odor) from its glutamyl conjugate, was discovered from axillary isolates. This strain showed strong resistance to the antimicrobial agents, triclosan and 4-isopropyl-3-methylphenol; therefore, we screened plant extracts that inhibit the A20 strain. We discovered that pentagalloyl glucose (PGG) extracted from the Chinese Gall plant exhibited both antibacterial and inhibitory activities against HMHA release by the A20 strain. As the excellent antibacterial activity and inhibitory effect of PGG against HMHA release were seen in vitro, we conducted an open study to evaluate the deodorant effects of PGG on axillary odor. The sensory tests on odor strength showed that application of the PGG solution could reduce axillary odors in vivo. Although there was a small change in axillary microbiota, the microbial count of A20 significantly reduced. These results strongly indicate PGG as a new innovative deodorant material that only affects odor-releasing bacteria in the axillary microbiota.

Screening of microorganisms producing a novel protein-asparaginase and characterization of the enzyme derived from Luteimicrobium album

A screening system using enrichment culture has been established with the aim of obtaining a novel enzyme for protein modification that has not been previously reported. This enzyme catalyzes deamidation of the side-chain amide group of asparagine in proteins. Enrichment culture of 390 soil samples was carried out with Z-Asn-Gly as the sole source of nitrogen, and the reaction product, Z-Asp-Gly, was detected in the culture supernatant of 102 strains. Strains with particularly high activity were Leifsonia sp., Luteimicrobium sp., Microbacterium sp., and Agromyces sp., all belonging to the class Actinobacteria. Of these, a protein-asparaginase (PA) was obtained from the culture supernatant of Luteimicrobium album 333B-h1, and its reactivity with different substrates and its basic enzymatic characteristics were investigated. Addition of the enzyme solution resulted in specific deamidation of only the asparagine residue in insulin chain B. The enzyme showed no reactivity with free asparagine or asparagine in low molecular weight peptides. It was demonstrated that the enzyme reacts with various protein substrates. In particular, proteins that have open structures, such as casein or gelatin, were good substrates. The activity and stability of PA at different temperatures and pH values were investigated. It was found that a temperature of 37°C and a roughly neutral pH are optimal conditions for the enzyme.