Polygalacturonase-inhibiting protein (PGIP) from Japanese pear: possible involvement in resistance against scab

Venturia nashicola is the causal agent of scab, a fungal disease affecting Asian pears. The Japanese pear cv. ‘Kousui’ is highly susceptible to the race 1 of this fungus whereas the cv. ‘Kinchaku’ and the non-host European pear cv. ‘Flemish Beauty’ are resistant. The aim of this work is to investigate the role of polygalacturonase-inhibiting proteins (PGIPs) of pear during the interactions with V. nashicola leading to susceptibility or resistance. PGIP protein was detected from immature fruit of Kousui and Kinchaku. It showed a molecular mass of 42 kDa that shifted to 35 kDa after chemical deglycosylation. The gene pgip was amplified by PCR using genomic DNA and/or cDNA from young leaves of Kousui, Kinchaku, and European pear cvs. Flemish Beauty, ‘Bartlett’, and an Asian wild pear strain ‘Mamenashi 12’, then sequenced after sub-cloning. Some conserved variations were identified in the sequence indicating that gene family also exists in pgip of Japanese pear and confirmed by Southern blot analysis. The expression of PGIP was studied in scab-inoculated leaves of the susceptible Kousui and the resistant Kinchaku and Flemish Beauty. pgip Gene and its encoding protein were highly and rapidly activated in these resistant plants. In addition, PGIP extracts derived from Kinchaku and Flemish Beauty partially inhibited the activity of polygalacturonase (PG) from V. nashicola suggesting a possible role of PGIP in limiting fungal growth frequently observed in these resistant cultivars.     

Morphological and genetic analysis of Vietnamese Sus scrofa bones for evidence of pig domestication

In the present study, we used morphological and genetic analyzes to distinguish bones of domestic boars from those of wild boars. We analyzed 65 Sus bones (cranium, mandible and teeth) stored in three research institutes in Vietnam and in a village in Vietnam. Based on comparison of bucco‐lingual measurements of mandibular parts, the 58 specimens were morphologically classified into two size groups: a large bone group and a small bone group. Analysis of 572‐bp mitochondrial DNA (mtDNA) sequences indicated that the large bones had genetic links to wild boar lineage including Ryukyu, Taiwan and Korean wild boars, and that the small bone group was closely related to East Asian domestic pigs. The phylogenetic analysis and parsimonious networks constructed among mtDNA haplotypes belonging to Ryukyu wild boar lineage showed that the Ryukyu wild boar is closely related to the Vietnamese wild boars, and uniquely miniaturized on their islands after the Ryukyu archipelago became isolated from the Asian continent.     

The inhibitory effect of a combination of imidacloprid and permethrin on blood feeding by mosquitoes in dogs raised under outdoor conditions

Combinations of imidacloprid and permethrin were frequently used to control harmful arthropod of companion animals. The inhibitory effects on blood-feeding activity of mosquitoes in dogs raised under outdoor conditions were evaluated by using combination of 10% (w/v) of imidacloprid and 50% (w/v) of permethrin as spot-on form. Dogs in the treated group received the combination imidacloprid/permethrin spot-on. After treatment, dogs in the control and treated groups were kept separately from the evening (17:00) to the morning of the following day (09:00) in two different kennels installed outdoors to mimic realistic dog-raising conditions. Mosquitoes in the kennels were collected by light traps placed in the kennels and a sweep net to determine evidence of blood feeding, and for species identification. Mosquitoes were collected at Days 5, 3 and 1 before agent treatment, and the Day of treatment, and Days 3, 7, 14, 21, 28, 35 and 42 after treatment. The percentages of blood-fed mosquitoes measured at Days 0, 3, 21, 28 and 42 after treatment were statistically significantly lower (p<0.01) in the treated group than in the control group. The most commonly collected mosquito, Culex tritaeniorhynchus, revealed statistically significant lower percentages (p<0.01) of blood-fed mosquitoes in the treated group than in the control group at the Day of treatment, and Days 3, 7, 21, 28 and 42 after treatment. It appeared that the test agent was effective in inhibiting blood feeding by adult female mosquitoes, and the efficacy lasts for 42 days after treatment under outdoor conditions.     

Genomic analyses of bovine viral diarrhea viruses isolated from cattle imported into Japan between 1991 and 2005

Thirty-one isolates of bovine viral diarrhea virus (BVDV) isolated within the past 15 years from imported cattle by the Japanese Animal Quarantine Service (AQS) were used in this study in which a 5'-untranslated region of each isolate was genetically analyzed. Twenty-six of the 31 isolates were classified as BVDV1 and the remainder as BVDV2. Phylogenetic analysis of the RT-PCR fragments amplified from the isolates showed the presence of viruses belonging to the BVDV1a, BVDV1b, BVDV1c, unclassified BVDV1 genotypes, and BVDV2. From the cattle of Australian origin, 16 of 17 isolates were classified as BVDV1c. This result was in agreement with a report showing that BVDV1c was a predominant subgenotype in Australia. From the cattle of North American origin, BVDV1 and BVDV2 species were both found. BVDV2 from the North American cattle was identified as the same cluster as the BVDV 890 strain, which is the prototype of BVDV2. These results suggest that the BVDVs isolated from exported cattle at the AQS reflect the predominant genotypes of BVDVs found in the exporting countries. The unclassified BVDV1 genotype of Chinese origin was in the same cluster as the ZM-95 strain, which was isolated from pigs in China. In this study, the genomic properties of 31 isolates of BVDV collected in the AQS were investigated. We concluded that isolates are genetically heterogeneous but geographically restricted. The information obtained from this report will be useful when carrying out epidemiological surveys of BVDV isolated in Japan.     

Anti-anxiety effect of ovary lipid extracted from Skipjack tuna (Katsuwonus pelamis) in rats

Using an elevated plus-maze test, we evaluated anxiety level in rats given ovary lipid extracted from Skipjack tuna (Katsuwonus pelamis; OLS). The percentage of open time was significantly higher in rats given OLS than in rats in the control group, but lower than in rats given diazepam (1.0 mg/kg body weight). Based on this fact and findings about other indicators, this study showed that OLS does not have as fast-acting and strong an anti-anxiety effect as diazepam but that continuous ingestion of OLS causes an anti-anxiety effect in animals.     

Acrosome reaction of mouse epididymal sperm on oocyte zona pellucida

To improve assessment of the acrosome reaction of mouse epididymal sperm, we employed anti-Izumo1 antibody instead of antibodies against acrosomal proteins. The acrosomal states among acrosome-intact, spontaneously acrosome-reacted, truly acrosome-reacted, and probably dead and/or membrane-damaged sperm were clearly distinguished by combined application of anti-Izumo1 antibody, DNA dye Hoechst 33342, and monoclonal antibody MN7 to paraformaldehyde-fixed sperm. When the acrosome reaction of capacitated epididymal sperm on the oocyte zona pellucida was examined using anti-Izumo1 antibody, approximately 20% of sperm bound onto the zona pellucida were acrosome-reacted 30 min after insemination. We also observed the moment of the acrosome reaction of live sperm on the zona pellucida by time-lapse monitoring using fluorescein isothiocyanate-conjugated anti-Izumo1 antibody.     

Optimization of cryoprotectant treatment for the vitrification of immature cumulus-enclosed porcine oocytes: comparison of sugars,combinations of permeating cryoprotectants and equilibration regimens

Our aim was to optimize the cryoprotectant treatment for the preservation of immature porcine cumulus-oocyte complexes (COCs) by solid surface vitrification. In each experiment, the vitrification solution consisted of 50 mg/ml polyvinyl pyrrolidone, 0.3 M of the actual sugar and in total 35% (v/v) of the actual permeating cryoprotectant (pCPA) combination. After warming, the COCs were subjected to in vitro maturation, fertilization and embryo culture. In Experiment 1, trehalose and sucrose were equally effective during vitrification and warming in terms of facilitating oocyte survival and subsequent embryo development. In Experiment 2, when equilibration was performed at 38.5 C in a total of 4% (v/v) pCPA for 15 min, the combination of ethylene glycol and propylene glycol (EG + PG = 1:1) was superior to EG and dimethyl sulfoxide (EG + DMSO = 1:1) in terms of oocyte survival after vitrification and the quality of resultant blastocysts. In Experiment 3, equilibration in 4% (v/v) pCPA for 15 min before vitrification was superior to that in 15% (v/v) CPA for 5 min for achievement of high survival rates irrespective of the pCPA combination used. In Experiment 4, when equilibration was performed in 4% EG + PG for 5 min, 15 min or 25 min, there was no difference in oocyte survival and subsequent embryo development after vitrification and warming; however, the developmental competence of cleaved embryos was tendentiously reduced when equilibration was performed for 25 min. In conclusion, trehalose and sucrose were equally effective in facilitating vitrification, and the optimum pCPA treatment was 5–15 min equilibration in 4% (v/v) of EG + PG followed by vitrification in 35% (v/v) EG + PG.     

Screening for ketosis using multiple logistic regression based on milk yield and composition

Multiple logistic regression was applied to milk yield and composition data for 632 records of healthy cows and 61 records of ketotic cows in Hokkaido, Japan. The purpose was to diagnose ketosis based on milk yield and composition, simultaneously. The cows were divided into two groups: (1) multiparous, including 314 healthy cows and 45 ketotic cows and (2) primiparous, including 318 healthy cows and 16 ketotic cows, since nutritional status, milk yield and composition are affected by parity. Multiple logistic regression was applied to these groups separately. For multiparous cows, milk yield (kg/day/cow) and protein-to-fat (P/F) ratio in milk were significant factors (P<0.05) for the diagnosis of ketosis. For primiparous cows, lactose content (%), solid not fat (SNF) content (%) and milk urea nitrogen (MUN) content (mg/dl) were significantly associated with ketosis (P<0.01). A diagnostic rule was constructed for each group of cows: (1) 9.978 × P/F ratio + 0.085 × milk yield <10 and (2) 2.327 × SNF − 2.703 × lactose + 0.225 × MUN <10. The sensitivity, specificity and the area under the curve (AUC) of the diagnostic rules were (1) 0.800, 0.729 and 0.811; (2) 0.813, 0.730 and 0.787, respectively. The P/F ratio, which is a widely used measure of ketosis, provided the sensitivity, specificity and AUC values of (1) 0.711, 0.726 and 0.781; and (2) 0.678, 0.767 and 0.738, respectively.