Pathogenicity of emerging Japanese type 1 porcine reproductive and respiratory syndrome virus in experimentally infected pigs

To clarify the pathogenicity of Japanese type 1 porcine reproductive and respiratory syndrome virus (PRRSV) isolate in experimentally infected pigs, we evaluated clinical signs and monitored viremia for 21 days post-inoculation (dpi). Lungs were mottled, tanned and reddish in appearance; had lesions predominantly in the cranial, middle and accessory lobes; and failed to collapse at 10 dpi. Although microscopic lesions of lungs were reproduced using the Japanese emerging type 1 PRRSV isolate under experimental conditions, no significant differences were noted between the challenge and control groups regarding mean rectal temperature and daily weight gain. These results provide useful insights into the limited pathogenicity of single infection with the Japanese type 1 PRRSV isolate in piglets, which differ from findings in reported field cases.     

Expression of amphiregulin during the pre- and post-implantation period in the mouse reproductive tract

In mammals, embryo implantation is an essential step in reproduction. Implantation is a phenomenon that involves crosstalk between the blastocyst and the maternal endometrium. However, the molecular basis of the connections between the blastocyst and endometrium is not yet fully understood. Amphiregulin is a member of the epidermal growth factor family and is known to be expressed in the luminal epithelium of the mouse uterus on 3.5 days post coitum (dpc). Thus, to clarify the mechanism of amphiregulin at fetomaternal interface, we analyzed the expression pattern of amphiregulin mRNA in the oviducts and uteri of pregnant and psuedopregnant mice by means of real-time PCR. Amphiregulin expression in the pregnant uterus dramatically increased on 2.5 dpc, peaked on 3.5 dpc, and declined by 5.5 dpc. Furthermore, to analyze the effect of the presence of an embryo on amphiregulin expression, we determined the expression pattern of amphiregulin mRNA in the uterus after embryo transfer on 0.5 and 1.5 dpc. A previous study showed that the expression of amphiregulin mRNA depends on the concentration of progesterone. However, our present results indicate that amphiregulin mRNA is upregulated by the presence of fertilized eggs in the lumen of the oviduct on 0.5 dpc.     

Preimplantation development of embryos in labrador retrievers

Preimplantation development of canine embryos is not well understood. To understand the timing of preattachment embryogenesis relative to the luteinizing hormone (LH) surge, early embryonic development was examined in Labrador Retrievers after artificial insemination. The embryos migrated from the oviduct to the uterus beginning on day 11 after the LH surge. This transport must be completed within 24 h. By day 13 after the LH surge, all of the embryos had moved and were localized in the uterus. The embryos developed to the morula stage within 11-13 days and to the blastocyst stage within 14 days after the LH surge, respectively. These findings add to the current understanding concerning the physiology of preimplantation development and should help further develop assisted reproductive techniques in canine species, such as cryopreservation and subsequent embryo transfer.     

A novel method for estimating the adaptive ability of guide dogs using salivary sIgA

Salivary secretory immunoglobulin A (sIgA) concentrations of prospective guide dogs for the blind were determined to clarify whether salivary sIgA is useful in evaluating the potential suitability of guide dogs for the blind. Saliva was collected from 73 prospective guide dogs in the kennel on day 1 (the day of separation from volunteer puppy-raisers), 2, 3, 7 and 14 during the estimation period (at about 1 year old). We selected particularly suitable dogs (superior dogs) and unsuitable dogs (inferior dogs) on the basis of the trainers' estimation. All dogs were divided into two groups, those were acceptable dogs would become the guide dogs and rejected dogs could not become guide dogs. The sIgA concentrations in superior dogs gradually increased from day 1 to 14 and those in inferior dogs remained at low levels. Moreover, the sIgA concentrations on day 14 in the acceptable dogs were significantly higher than those in rejected dogs. The cut-off point of sIgA concentrations on day 14 using an ROC curve was 90 EU/ml, and the specificity of the estimation at this point (70.4%) was higher than that of trainers' estimation (50%). Moreover, parallel testing using both trainers' estimation and sIgA estimation, showed that specificity was further improved (79.5%). The present study showed that sIgA concentration was extremely useful in estimating the adaptive ability for guide dogs for the blind.     

Screening of Neutrophil Activating Factors from a Metagenome Library of Sponge-Associated Bacteria

Marine sponge-associated bacteria are known as bio-active compound produce. We have constructed metagenome libraries of the bacteria and developed a metagenomic screening approach. Activity-based screening successfully identified novel genes and novel enzymes; however, the efficiency was only in 1 out of 10⁴ clones. Therefore, in this study, we thought that bioinformatics could help to reduce screening efforts, and combined activity-based screening with database search. Neutrophils play an important role for the immune system to recognize excreted bacterial by-products as chemotactic factors and are recruited to infection sites to kill pathogens via phagocytosis. These excreted by-products are considered critical triggers that engage the immune system to mount a defense against infection, and identifying these factors may guide developments in medicine and diagnostics. We focused on genes encoding amino acid ligase and peptide synthetase and selected from an in-house sponge metagenome database. Cell-free culture medium of each was used in a neutrophil chemiluminescence assay in luminol reaction. The clone showing maximum activity had a genomic sequence expected to produce a molecule like a phospho-N-acetylmuramyl pentapeptide by the metagenome fragment analysis.     

Immune response of largemouth bass, <Emphasis Type="Italic">Micropterus salmoides</Emphasis>, to whole cells of different <Emphasis Type="Italic">Nocardia seriolae</Emphasis> strains

Largemouth bass Micropterus salmoides were immunized with four different N. seriolae strains—two α-glucosidase-positive (961113, KU040801) and two α-glucosidase-negative (94260, OTTS) strains—along with Freund’s incomplete adjuvant. After primary immunization (week 0), a booster was administered at weeks 4 and 8. Nonspecific immune responses to multiple immunizations with the different N. seriolae strains were determined based on serum lysozyme activity and nitroblue tetrazolium (NBT)-positive cells in peripheral blood. The serum lysozyme activity and NBT-positive cells in peripheral blood were not significantly increased even after the two booster immunizations. Specific antibody responses against N. seriolae cells were investigated by enzyme-linked immunosorbent assay. At 4 weeks after immunization, all groups immunized with N. seriolae antigens showed significant increases in their specific antibody levels. The sera from fish immunized with different N. seriolae strains exhibited reactivity with N. seriolae sonicated antigens of 28, 30, 36 and 84 kDa by western blot analysis. After two boosters, fish were challenged with live N. seriolae to assess the vaccine’s efficacy; however, multiple injections of the N. seriolae strains did not reduce mortality, irrespective of the bacterin.