Masahiko Kuroki

Oral passive immunization against experimental salmonellosis in mice using chicken egg yolk antibodies specific for Salmonella enteritidis and S. typhimurium

The efficacy of chicken egg yolk homotypic antibodies specific for outer membrane proteins (OMP), lipopolysaccharide (LPS) or flagella (Fla) in controlling experimental salmonellosis in mice was investigated. Mice challenged orally with 2 x 10(9) c.f.u. of Salmonella enteritidis or 2 x 10(7) c.f.u. of S. typhimurium were orally treated with 0.2 ml anti-OMP, -LPS or -Fla yolk antibody three times a day for three consecutive days. In mice challenged with S. enteritidis, antibody treatment resulted in a survival rate of 80%, 47% and 60% using OMP, LPS or Fla specific antibodies respectively, in contrast to only 20% in control mice. In the S. typhimurium trial, survival rate was 40%, 30% and 20% using OMP, LPS or Fla specific antibodies respectively in contrast to 0% in control mice. In vitro adhesion of S. enteritidis and S. typhimurium to HeLa cells was significantly reduced by anti-OMP, -LPS, and -Fla homotypic antibodies. Results suggest that egg yolk antibodies specific for Salmonella OMP, LPS, and Fla may protect mice from experimental salmonellosis when passively administered orally. Of these antibodies, anti-OMP exhibited the highest level of protection in vivo and in vitro.

Passive protection of neonatal calves against bovine coronavirus-induced diarrhea by administration of egg yolk or colostrum antibody powder

Abstract The protective effect of egg yolk and colostrum powders prepared from hens and cows vaccinated with inactivated bovine coronavirus (BCV) antigen was evaluated in a challenge model with a virulent BCV strain. Twenty three calves from BCV-free herds were randomly divided into control and several treatment groups. All calves were orally challenged with 1 × 109 TCID50 of the virulent Kakegawa strain of BCV at 24 to 36 h after birth. Calves in treatment groups received either egg yolk powder or cow colostrum containing BCV specific antibodies. Daily treatment with these antibody preparations started 6 h until 7 days post-challenge. Control calves which received no antibody had severe diarrhea and all died within 6 days after infection. In contrast, calves fed milk containing egg yolk or colostrum with neutralization titers of 1:2560 or 1:10 240 respectively all survived and had positive weight gain unlike the other treatment groups. These results indicate that the orally administered egg yolk and colostrum powders protected against BCV-induced diarrhea in neonatal calves and that the egg yolk used provided a higher degree of protection compared to colostrum powder on a titer basis. Treatment with whole egg yolk from immunized hens therefore provides a more efficacious alternative to the existing methods of specific passive protection against BCV.

Passive protection against bovine rotavirus-induced diarrhea in murine model by specific immunoglobulins from chicken egg yolk

Chicken egg yolk immunoglobulins (yIg) specific against bovine rotavirus (BRV) serotypes 6 (strain Shimane) and 10 (strain KK-3) were used for oral passive immunization of suckling mice against experimental BRV challenge. The protective capacity of the antibody preparation was tested using different concentrations of yIg against a challenge dose of 10(7.5) TCID50 for Shimane and 10(7.0) TCID50 for KK-3 strain. There was a significant homotypic (P < 0.05) and heterotypic (P < 0.01) protection using 160 anti-Shimane or 160 anti-KK-3 neutralizing antibody titer (NAT) compared to control mice given yIg derived from eggs of mock-immunized (control) hens. The titer of infectious BRV recovered from intestinal tissue or luminal chyme decreased with increasing homotypic yIg NAT. A decrease in degree and duration of BRV antigen localization in the villus epithelial lining was observed in mice treated with homotypic yIg at optimum dose for prevention of diarrhea. The NAT in sera of challenged mice increased with decreasing NAT in the yIg given before challenge suggesting that protection was dose-dependent. The present findings indicate that a passive protection could be achieved by the use of yIg against BRV-induced diarrhea in this murine model.

Affinity Purification of Helicobacter pyloriUrease RELEVANCE TO GASTRIC MUCIN ADHERENCE BY UREASE PROTEIN

A simple, reproducible and high yield method ofHelicobacter pylori urease enzyme purification was developed using a heparinoid (Cellufine sulfate) affinity gel. The purification method involved two sequential steps using the same gel that takes advantage of the differential affinity of urease to the heparinoid at two levels of hydrogen ion concentration. SDS-polyacrylamide gel electrophoresis analysis of affinity-purified urease revealed two major protein bands with about 62- and 30-kDa molecular mass. When whole cell lysates of clinical and laboratory strains of H. pylori were probed by Western blot, anti-urease hyperimmune serum produced by affinity-purified urease in rabbit recognized only the two bands corresponding to the urease A and B subunits. To probe the molecular relevance of affinity gel adherence to mucin adherence, the purified urease was derivatized withN-hydroxysuccinimidobiotin and used in adherence assays. Competitive inhibition tests revealed commonality of urease receptors among gastric mucin, heparin, and heparinoid. Composite data on adherence kinetics modulated by pH, salt, incubation time, and concentration of urease or mucin were indicative of conformation-dependent ligand-receptor interaction.

Enhancement of Chicken Resistance Against Escherichia coli Infection by Oral Administration of Bifidobacterium thermophilum Preparations

SUMMARY. Three types of Bifidobacterium thermophilum extract were prepared and fed to 2-wk-old chickens to evaluate their usefulness in enhancing the defense activity of the chickens against pathogenic Escherichia coli. All three preparations resulted in significant reduction (P < 0.05) of E. coli numbers in the lungs of the treated chicken groups compared with the control nontreated group. Besides, improvement in the survival rate was observed in the treated chicken groups, especially the one administered the enzyme-digested B. thermophilum extract sample. Concanavalin A–stimulated lymphocytes from the latter group demonstrated significantly higher proliferation activity compared with those from the control group. These results suggest that oral administration of B. thermophilum preparations may be used to enhance the resistance of chickens against E. coli infection.

Adherence Protects the Binding Sites of Helicobacter pylori Urease from Acid-Induced Damage

Colonization by Helicobacter pylori partly depends on acid‐dependent adherence by urease to gastric mucin. To further verify the relevance of urease adherence to colonization, the influence of acidity on the binding sites of H. pylori urease was investigated. When enzyme‐based in vitro ligand capture assays were used, the effect of acidity on the binding site of H. pylori urease was determined against a backdrop medium consisting of acidic buffers simulating the luminal side of gastric mucus. A high degree of stability was exhibited by adherent urease, suggesting a pivotal role by the denatured enzyme in the persistence of the bacterium within the acidified compartment of gastric mucus.