Yoshikazu Hirai

Antibacterial Activity of Hydrolyzable Tannins Derived from Medicinal Plants against Helicobacter pylori

Helicobacter pylori is a major etiological agent in gastroduodenal disorders. In this study, we isolated 36 polyphenols and 4 terpenoids from medicinal plants, and investigated their antibacterial activity against H. pylori in vitro. All hydrolyzable tannins tested demonstrated promising antibacterial activity against H. pylori. Monomeric hydrolyzable tannins revealed especially strong activity. Other compounds demonstrated minimal antibacterial activity with a few exceptions. A monomeric hydrolyzable tannin, Tellimagrandin I demonstrated time‐ and dose‐dependent bactericidal activity against H. pylori in vitro. On the other hand, hydrolyzable tannins did not affect the viability of MKN‐28 cells derived from human gastric epithelium. Hydrolyzable tannins, therefore, have potential as new and safe therapeutic regimens against H. pylori infection. Furthermore, we investigated effects of hydrolyzable tannins on lipid bilayer membranes. All the hydrolyzable tannins tested demonstrated dose‐dependent membrane‐damaging activity. However, it remains to be elucidated whether their membrane‐damaging activity directly contributes to their antibacterial action.

The haemagglutinin of Clostridium botulinum type C progenitor toxin plays an essential role in binding of toxin to the epithelial cells of guinea pig small intestine, leading to the efficient absorption of the toxin

Binding of the purified type C 7S (neurotoxin), 12S and 16S botulinum toxins to epithelial cells of ligated small intestine or colon of the guinea pig (in vivo test) and to pre-fixed gastrointestinal tissue sections (in vitro test) was analysed. The 16S toxin bound intensely to the microvilli of epithelial cells of the small intestine in both in vivo and in vitro tests, but did not bind to cells of the stomach or colon. The neurotoxin and 12S toxin did not bind to epithelial cells of the small intestine or to cells of the stomach or colon. Absorption of the toxins was assessed by determining the toxin titre in the sera of guinea pigs 6-8 h after the intra-intestinal administration of the toxins. When the 16S toxin [1 x 10(5) minimum lethal dose (MLD)] was injected, 200-660 MLD ml-1 was detected in the sera, whereas when the 12S toxin (2 x 10(5) MLD) or 7S toxin (2 x 10(5) MLD) was injected, little toxin activity was detected in the sera. Therefore, the haemagglutinin of type C 16S toxin is apparently very important in the binding and absorption of botulinum toxin in the small intestine.

Antibacterial Properties of Antimicrobial-Finished Textile Products

The antibacterial properties of five kinds of antimicrobial‐finished textile products (AFTPs) were examined against Staphylococcus aureus, including methicillin‐resistant (MRSA) strains and Pseudomonas aeruginosa, under wet and dry conditions. Textile products containing Ag. Zn. ammonium Zeolite and chitosan were found to be effective against methicillin‐sensitive S. aureus (MSSA) for up to 6 hr of incubation under wet and dry conditions, and effective against MRSA for up to 24 hr of incubation only under wet conditions. Under dry conditions, however, all AFTPs were ineffective against one MRSA strain. When organic matter was added to the incubation mixture, textile products containing Ag. Zn. ammonium Zeolite and chitosan still showed antibacterial activities, but not as strongly. The results of this study suggested the following: (1) There are differences in antibacterial properties among commercially available AFTPs; (2) Determining effectiveness requires several hours of incubation; (3) Water content as an environmental factor can affect effectiveness; and (4) Some bacterial species and strains are not affected by AFTPs. The antibacterial properties of AFTPs in the clinical setting may be of limited value.

Bacillus cereus bacteremia outbreak due to contaminated hospital linens

We describe an outbreak of Bacillus cereus bacteremia that occurred at Jichi Medical University Hospital in 2006. This study aimed to identify the source of this outbreak and to implement appropriate control measures. We reviewed the charts of patients with blood cultures positive for B. cereus, and investigated B. cereus contamination within the hospital environment. Genetic relationships among B. cereus isolates were analyzed. Eleven patients developed B. cereus bacteremia between January and August 2006. The hospital linens and the washing machine were highly contaminated with B. cereus, which was also isolated from the intravenous fluid. All of the contaminated linens were autoclaved, the washing machine was cleaned with a detergent, and hand hygiene was promoted among the hospital staff. The number of patients per month that developed new B. cereus bacteremia rapidly decreased after implementing these measures. The source of this outbreak was B. cereus contamination of hospital linens, and B. cereus was transmitted from the linens to patients via catheter infection. Our findings demonstrated that bacterial contamination of hospital linens can cause nosocomial bacteremia. Thus, blood cultures that are positive for B. cereus should not be regarded as false positives in the clinical setting.

Antibodies to human gastric epithelial cells and heat shock protein 60 in Helicobacter pylori positive mucosa associated lymphoid tissue lymphoma.

BACKGROUND Development of gastric mucosa associated lymphoid tissue (MALT) lymphoma is thought to be closely associated with host immune reactions toHelicobacter pylori. AIM To investigate humoral immune responses in patients with MALT lymphoma to antigens shared by H pylori and human gastric epithelial cells. METHODS Sera were obtained from H pylori positive patients with MALT lymphoma (n = 11) or other gastroduodenal diseases (peptic ulcer, n = 40; non-ulcer dyspepsia, n = 20) and fromH pylori negative healthy control subjects (n = 10). Antibodies to HGC-27 human gastric epithelial cells and human recombinant heat shock protein (Hsp) 60 were examined using an enzyme linked immunosorbent assay (ELISA) and immunoblotting. RESULTS Antibody titres to HGC-27 cells were significantly elevated inH pylori positive patients with MALT lymphoma when compared with titres in patients with other gastroduodenal diseases and in healthy subjects. Immunoblotting of sera from patients with MALT lymphoma often detected a band with a molecular mass corresponding to Hsp60, and both ELISA and immunoblotting showed elevated antibody titres to the recombinant human Hsp60. Antigenic similarity between Hsp60 and H pylori HspB was documented by immunoblotting experiments. CONCLUSIONS Autoantibodies reactive with host gastric epithelial cells are often increased in MALT lymphoma, and Hsp60 is a major target antigen. Immune responses induced by immunological cross reactivity between H pylori HspB and human Hsp60 in gastric epithelium may be involved in the development of MALT lymphoma.

Sensitive Detection of Bacillus anthracis Using a Binding Protein Originating from γ-Phage

Detection of biological weapons is a primary concern in force protection, treaty verification, and safeguarding civilian populations against domestic terrorism. One great concern is the detection of Bacillus anthracis, the causative agent of anthrax. Therefore, there is a pressing need to develop novel methods for rapid, simple, and precise detection of B. anthracis. Here, we report that the C‐terminal region of γ‐phage lysin protein (PlyG) binds specifically to the cell wall of B. anthracis and the recombinant protein corresponding to this region (positions, 156–233), PlyGB, is available as a bioprobe for detection of B. anthracis. Our detection method, based on a membrane direct blot assay using recombinant PlyGB, was more rapid and sensitive than the γ‐phage test and was simpler and more inexpensive than genetic methods such as PCR, or immunological methods using specific antibodies. Furthermore, its specificity was comparable to the γ‐phage test. PlyGB is applicable in conventional methods instead of antibodies and could be a potent tool for detection of B. anthracis.

Immunoglobulin G1 antibody response to Helicobacter pylori heat shock protein 60 is closely associated with low-grade gastric mucosa-associated lymphoid tissue lymphoma.

ABSTRACT Gastric mucosa-associated lymphoid tissue (MALT) lymphoma is related to Helicobacter pylori infection. Specifically, it has been pointed out that pathogenesis of MALT lymphoma involves the 60-kDa heat shock protein (hsp60). To investigate humoral immune responses to the H. pylori hsp60 in patients with gastroduodenal diseases and patients with MALT lymphoma, the hsp60 ofH. pylori was expressed with a glutathioneS-transferase fusion protein and was purified (recombinant hsp60). Sera were obtained from H. pylori-positive patients with gastroduodenal diseases (MALT lymphoma, n = 13; gastric ulcer, n = 20; duodenal ulcer, n = 20; gastritis,n = 20) and from H. pylori-negative healthy volunteers (n = 9). Sera from patients with MALT lymphoma were also obtained at two times: before and after eradication therapy. Antibodies to hsp60 and H. pylori were assessed by enzyme-linked immunosorbent assay. The levels of immunoglobulin G (IgG) antibodies to the hsp60 of H. pylori-positive patients with gastroduodenal diseases were significantly elevated compared to those in the controls. The levels of IgG1 antibodies to hsp60 were elevated and correlated with the levels of anti-H. pylori antibodies in patients with MALT lymphoma. Humarol immunity against hsp60 may be important and relevant to gastroduodenal diseases induced by H. pylori infection.

Characterization of Streptococcus sanguis isolated from patients with Behçet's disease.

The DNA homology and cell wall sugar constituents of eight Streptococcus sanguis(‐like) strains, three isolated from the patients with Behçets disease (BD114‐23, BD113‐20, BD118‐1), two from patients with Kawasaki disease (MCLS‐1, MCLS‐2), and three type and reference strains of ATCC (ATCC10556T: S. sanguis, ATCC10557: S. oralis, and ATCC10558T: S. gordonii) were analyzed. Strains BD114‐23 and BD118‐1 showed high DNA homology to ATCC10556T, and their cell wall constituents were identical. Conversely, BD113‐20, MCLS‐1, MCLS‐2, and ATCC10557 showed little DNA homology to ATCC10556T and ATCC10558T, but showed approximately 50 to 60% homology to each other. The cell wall constituents of BD113‐20, MCLS‐1, MCLS‐2, and ATCC10557, however, were somewhat different, indicating that some of the clinical isolates have different characters from those of the three ATCC strains.

Detection of Helicobacter pylori associated antigen and heat shock protein 60 on follicular dendritic cells in the germinal centres of low grade B cell lymphoma of gastric mucosa associated lymphoid tissue (MALT)

AIMS: To investigate the localisation of Helicobacter pylori antigens and the expression of human heat shock proteins (HSP) in stomachs affected by MALT lymphoma. METHODS: Surgically resected stomachs from 24 patients with MALT lymphoma were immunostained with anti-H pylori rabbit antibodies (ORP-1 and ORP-2) and anti-human HSP60 mouse monoclonal antibodies (mAb) (LK-1 and LK-2). RESULTS: Follicular dendritic cells of germinal centres in the stomachs affected by MALT lymphoma were immunostained with anti-H pylori polyclonal antibodies and with anti-human HSP60 mAb, as were the epithelial cells. None of the lymph node samples reacted. CONCLUSIONS: Human HSP60, which cross reacts with anti-H pylori polyclonal antibodies, is often expressed on follicular dendritic cells in gastric MALT lymphoma tissues and may be aetiologically relevant to lymphomagenesis of MALT lymphoma.

Alteration in phospholipid composition of Staphylococcus aureus during formation of autoplast

The phospholipid composition of autoplasts (protoplasts made by autolysis of Staphylococcus aureus 209P was examined. The autoplasts were prepared by incubation of 209P cells in 1.2 M sucrose--0.33 M acetate buffer (pH 5.8). Cardiolipin comprised nearly half the total phospholipid in these autoplasts. Autoplasts had a lower phosphatidylglycerol content than intact cells but similar lysylphosphatidylglycerol content. The increase in cardiolipin content during release of autoplasts was not affected by pH or temperature. The result indicates that removal of the cell wall caused the increase in cardiolipin content. The total amount of phospholipids increased slightly during autoplast formation, but there was no significant increase in fatty acids or diglycerides. The changes of phospholipid composition during formation of the autoplast was due to de novo synthesis of cardiolipin from phosphatidylglycerol.