Glycopeptide derived from hen egg ovomucin has the ability to bind enterohemorrhagic Escherichia coli O157:H7

Ovomucin glycopeptide (OGP) was prepared by size exclusion chromatography after Pronase digestion of hen egg ovomucin, and the binding of OGP to foodborne pathogens (Bacillus cereus,Clostridium perfringens, Escherichia coli O157:H7, Listeria monocytogenes, Salmonella enteritidis, Salmonella typhimurium, and Staphylococcus aureus) was investigaed. Binding assays with biotinylated bacteria as probes in microtiter plates showed that OGP bound to only E. coli O157:H7 among these foodborne pathogens. Periodate treatment markedly reduced the binding ability, indicating that E. coli O157:H7 bound to carbohydrate moieties of OGP. Lectin blot analysis with Maackia amurensis (MAA) and Sambucus nigra (SNA), which are specific for oligosaccharides containing sialic acid, revealed their binding sites in OGP were similar to the E. coli O157:H7 binding sites that were probed with biotinylated E. coli O157:H7 after Western blotting of OGP. Sialydase treatment of OGP abolished its ability to bind E. coli O157:H7, demonstrating that sialic acid played an important role in the binding. These results suggest that OGP has E. coli O157:H7-specific binding sites that consist of sialic acid. On the basis of these properties, OGP has the potential to be an ingredient with a protective effect against E. coli O157:H7 infection and to be a novel probe for the detection of E. coli O157:H7 in the food hygiene field.     

Production and characterization of an alloplasmic and monosomic addition line of Brassica rapa carrying the cytoplasm and one chromosome of Moricandia arvensis

Intergeneric hybridization was performed between Moricandia arvensis and four inbred lines of Brassica rapa following embryo rescue. Three F₁ hybrid plants were developed from three cross combinations of M. arvensis × B. rapa, and amphidiploids were synthesized by colchicine treatment. Six BC₁ plants were generated from a single cross combination of amphidipolid × B. rapa ‘Ko1-303’ through embryo rescue. One BC₂ and three BC₃ plants were obtained from successive backcrossing with B. rapa ‘Ko1-303’ employing embryo rescue. Alloplasmic and monosomic addition lines of B. rapa (Allo-MALs, 2n = 21) were obtained from backcrossed progeny of three BC₃ plants (2n = 21, 22 and 23) without embryo rescue. An alloplasmic line of B. rapa (2n = 20) degenerated before floliation on 1/2 MS medium due to severe chlorosis. Allo-MALs of B. rapa (2n = 21) showed stable male sterility without any abnormal traits in vegetative growth and female fertility. Molecular analyses revealed that the same chromosome and cytoplasm of M. arvensis had been added to each Allo-MAL of B. rapa. This Allo-MAL of B. rapa may be useful material for producing cytoplasmic male sterile lines of B. rapa.     

Successive deaths of a captive snow leopard (Uncia uncia) and a serval (Leptailurus serval) by infection with feline panleukopenia virus at Sapporo Maruyama Zoo

Feline parvoviruses were isolated from frozen samples of intestines taken from a snow leopard (Uncia uncia) and a serval (Leptailurus serval) that died successively at Sapporo Maruyama Zoo in Hokkaido, Japan. Isolates possessed an antigenic epitope for both the feline panleukopenia virus (FPLV) and mink enteritis virus, identified with a hemagglutination inhibition test. Sequencing analyses of the VP2 region of the isolates revealed that the two isolates were identical and of the FPLV-type. These results suggested that FPLV was introduced from a feral cat which entered the zoo and transmitted the virus inside the zoo.     

Detachable Coils for Occlusion of Patent Ductus Arteriosus in 2 Dogs

OBJECTIVE: To describe the use of a detachable coil for transcatheter closure (TCC) of patent ductus arteriosus (PDA) in 2 dogs. STUDY DESIGN: Clinical study. ANIMALS: Two female Pembroke Welsh Corgi dogs with PDA. METHODS: Using fluoroscopic guidance, an 8-mm-diameter coil stent with 5 loops (detachable coils for PDA closure) was inserted via catheterization of the femoral artery. The catheter was passed through the PDA into the pulmonary artery. The coil was withdrawn so that 1.5 loops remained on the pulmonary side of the orifice of the ductus. The rest of the loops were pushed out from the catheter into the ductus. After confirming the correct placement of the coil and the effectiveness of the occlusion, the delivery wire was detached from the coil. RESULTS: Insertion of the coil was easily performed, even without previous experience. Immediate and marked decrease of the cardiac murmur was auscultated. Only slight residual flow was detected by angiography conducted 3 months' postoperatively. The dogs experienced quick and uneventful recovery after coil placement and required minimal postoperative care. Follow-up evaluation of the dogs showed no functional clinical signs of PDA, and no cardiac abnormalities were detected on electrocardiographic, phonocardiographic, and echocardiographic examination. In dog 1, the residual flow had disappeared on the color-flow Doppler echocardiographic examination at 18 months' postoperatively. CONCLUSION: TCC using a detachable coil was easy, safe, and effective in 2 dogs with PDA. The minimal residual shunting observed only by echocardiography seemed hemodynamically insignificant. CLINICAL RELEVANCE: This method can be used as an alternative to traditional surgical methods.     

Seroepidemiology of Toxoplasma gondii and Neospora caninum in seals around Hokkaido, Japan

Serological analysis was performed to detect Toxoplasma gondii and Neospora caninum infection in seals in Hokkaido. Serum samples were collected from 322 Kuril harbor seals (Phoca vitulina stejnegeri) at Nosappu, Akkeshi and Erimo, from 46 spotted seals (P. largha) at Nosappu, Erimo, Yagishiri Island, Hamamasu and Syakotan, and from 4 ribbon seals (P. fasciata) and a bearded seal (Erignathus barbatus) at Nosappu between 1998 and 2006. Recombinant surface antigen of T. gondii (SAG2t) and N. caninum (NcSAG1t) were used as antigens for ELISA to detect antibodies. Antibodies against SAG2t were detected from 4% of 77 Kuril harbor seals at Nosappu in 2005. Antibodies against NcSAG1t were detected from 2% (1/66) in 2003, 5% (4/79) in 2004 and 10% (8/77) in 2005 of Kuril harbor seals and 11% of 9 spotted seals in 2004 sampled at Nosappu. Eight percent of 12 Kuril harbor seals from Akkeshi and 25% of 4 spotted seals from Erimo in 2005 also contained antibodies against NcSAG1t. These suggest sporadic infection of T. gondii and N. caninum in Kuril harbor seals and spotted seals in Hokkaido. Of the ELISA-positive seals, 2 seals having anti-SAG2t antibodies and 3 seals having anti-NcSAG1t antibodies in 2005 were judged to be juveniles that have no maternal antibodies. These suggest that the protozoan infections have occurred in recent years. Infection of terrestrial protozoa such as T. gondii and N. caninum in seals indicates that the sea environment has been contaminated with protozoa.     

Effects of Salmonella enterica subsp. enterica serovar Enteritidis vaccination in layer hens subjected to S. Enteritidis challenge and various feed withdrawal regimens

Levels of Salmonella enterica subsp. enterica serovar Enteritidis infection and serum S. Enteritidis antibodies after experimental S. Enteritidis challenge and feed withdrawal were investigated in S. Enteritidis-vaccinated and unvaccinated hens. The results were used to determine whether formalin-inactivated S. Enteritidis vaccination can protect layer hens from S. Enteritidis challenge during feed withdrawal periods. S. Enteritidis infection rates were evaluated from cloacal swabs, eggs and organs. Serum antibody titers to deflagellated S. Enteritidis whole cells (DEWC) and S. Enteritidis FliC-specific 9-kDa polypeptide (SEp 9) were examined by commercial ELISA kits. Cloacal S. Enteritidis recovery rates were lower in the vaccinated than unvaccinated group. Recovery rates of S. Enteritidis from samples increased after feed withdrawal and decreased after re-introduction of feed. S. Enteritidis counts in cloacal swabs were lower in the vaccinated than in the unvaccinated group (P<0.05). More S. Enteritidis-positive eggs were detected from the unvaccinated group. Before S. Enteritidis challenge, the DEWC ELISA titer of the vaccinated group was higher (P<0.05) than the unvaccinated group; subsequently, the S. Enteritidis DEWC ELISA titers of both groups increased gradually. In contrast, only the vaccinated group elicited high SEp-9 antibody titer during post-challenge and feed withdrawal. Additionally, vaccinated hens yielded negative S. Enteritidis isolation rates from egg contents. There is a correlation between negative S. Enteritidis isolation rates and high SEp 9 titers in vaccinated layer hens challenged with S. Enteritidis and subjected to feed withdrawal regimens. These findings suggest the S. Enteritidis vaccination of pullets may protect against S. Enteritidis infection during forced molting and that SEp 9 titer could be a potential indicator of antibody protection against S. Enteritidis infection. The potential of the SEp 9 peptide as an antigen for S. Enteritidis vaccination in the future is worth noting.     

Molecular epidemiological characterization of poultry red mite, Dermanyssus gallinae,in Japan

Dermanyssus gallinae, the poultry red mite, is an obligatory blood-sucking ectoparasite. The genetic diversity of D. gallinae has been examined in some countries, but so far not in Asian countries. Here, we sequenced a part of the mitochondrial cytochrome oxidase subunit I (COI) and16S rRNA genes and nuclear internal transcribed spacers (ITS) region in 239 mite samples collected from 40 prefectures throughout Japan. The COI and 16S rRNA nucleotide sequences were classified into 28 and 26 haplotypes, respectively. In phylogenetic trees, the haplotypes clustered into 2 haplogroups corresponding to haplogroups A and B, which were previously reported. Haplogroups A and B were further subdivided into sub-haplogroups AJ1 and AJ2, and BJ1 and BJ2, respectively. In both trees, the sequences of haplotypes in AJ1 and BJ2 were relatively distant from those reported in other countries, while some sequences in AJ2 and BJ1 were identical to those in Europe. In addition, the ITS sequences were classified into two sequences, and both sequences were closely related to the sequences found in European countries. These findings indicate a possibility of international oversea transmission of D. gallinae.     

Isolation and characterization of a cDNA from Catharanthus roseus which is highly homologous with phosphate transporter

The PIT1 gene which is highly homologous with phosphate transporter was isolated from Catharanthus roseus and analyzed. The cBNA PIT1 contained an open reading frame of 542 amino acids and its sequence showed a 31, 30, and 34% identity with the phosphate transporter of Saccharomyces cerevisiae (PBO84), Neurospora crassa (PHO-5), and Glomus versiforme (GvPT), respectively. Furthermore, the cDNA PIT1 encoded a highly hydrophobic protein with 12 putative membrane-spanning regions and contained a conserved amino acid sequence reported in the human glucose transporter super-family* S. cerevisiae strain DpU (pho84 knockout strain) was unable to grow on low phosphate (55 μM) medium (LP medium). Expression of the PIT1 cDNA enabled DpU to grow on LP medium. Northern hybridization analysis revealed that the PIT1 gene was expressed in roots, stems, and young whole plant of C. roseus, but not in leaves.     

Accumulation of gamma-aminobutyrate (GABA) caused by heat-drying and expression of related genes in immature vegetable soybean (edamame)

We studied the effects of drying of immature seeds of vegetable soybean (Glycine max L. Merrill) on the accumulation of gamma-aminobutyrate (GABA) in the seeds. GABA accumulated after heat-drying, with the maximum at 40°C. The GABA content (447.5 mg/100 g DW) increased to more than 5 times the value in untreated seeds (79.6 mg/100 g DW). In contrast, the glutamate content decreased rapidly to 1/3 the level in the untreated seeds. The GABA content increased early in the heat-drying treatment: after 30 min, it had increased to 1.5 times the value in the untreated seeds. GABA did not accumulate in the vacuum-drying treatment. Among genes related to the GABA shunt, the gene for glutamate decarboxylase (EC 4.1.1.15), which catalyzes the decarboxylation of glutamate to produce GABA, showed relatively high expression, decreasing to only 70% of the value in untreated seeds even after 4 h of treatment. In contrast, expression of the genes for two catabolic mitochondrial enzymes, GABA transaminase (GABA-T; EC 2.6.1.19) and succinate semialdehyde dehydrogenase (SSADH; EC 1.2.1.16), decreased rapidly during heat-drying. These results suggest that the accumulated GABA was not metabolized rapidly by GABA-T and SSADH and therefore remained at high levels.