Regional Distribution of Two Races of <Emphasis Type="Italic">Phialophora gregata </Emphasis>f. sp. <Emphasis Type="Italic">adzukicola, </Emphasis>Causal Agent of Brown Stem Rot of Adzuki Bean,and Their Genetic Diversity on Hokkaido,Northernmost Island of Japan

The distribution of two races (1 and 2) of Phialophora gregata f. sp. adzukicola, the causal agent of brown stem rot of adzuki bean, was examined using a total of 483 isolates obtained from 39 fields in 19 locations on Hokkaido, Japan between 1997 and 1999. Race 1 was predominant (416 isolates or 86.1%) in the commercial fields tested. Race 2 was found in 25 fields (64.1%), including two fields of cultivar Kita-no-otome (resistant to race 1, but susceptible to race 2), indicating that race 2 was widely distributed in most of the production areas in Hokkaido. Using amplified fragment length polymorphisms (AFLP), a total of 67 polymorphic AFLP markers was recorded among 72 representative isolates (37 and 35 isolates of races 1 and 2, respectively), and 57 distinct haplotypes were detected. Cluster analysis revealed no close correlation between races and AFLP groups. Thus, no difference was observed between values of gene diversity in each race (0.253 and 0.284 in races 1 and 2, respectively), and the coefficient of gene differentiation was very low (G _ST =0.015). Gene differentiation between both races by analysis of molecular variance was not significantly different from zero (Φ=−0.001; p=0.403). However, the results of gene differentiation among regional populations (G _ST =0.290, Φ=0.292; p<0.001) are not necessarily consistent with the result that isolates from the same district were generally not tightly clustered. Received 15 April 2002/ Accepted in revised form 6 September 2002     

Induction of ovine trophoblast cell fusion by fematrin‐1 in vitro

Endogenous retroviruses present in the genomes take a specific role in placental formation in various vertebrates, including bovine and sheep. Fematrin‐1, which is the envelope (Env) protein of bovine endogenous retrovirus found in bovine placenta, is involved in the formation of fetomaternal hybrid cells in cattle placenta. This study was conducted to clarify whether fematrin‐1 possesses fusogenic activity in trophoblast cells. Another question is whether Env proteins only have species‐specific activity or not. For this, fematrin‐1 gene was transfected in ovine trophoblast cells, and we examined fusogenic activity with Cos‐7 cells. Although fematrin‐1 fusogenic activity was detected in both neutral and acidic pH conditions, acidic condition significantly enhanced it. These activities were rather weaker than those of vesicular stomatitis virus G protein as a positive control. However, the ratio of fematrin‐1 and vesicular stomatitis virus G protein fusion index was confirmed similar to those in the previous reports. Some fusion cells showed multinucleate cells. These results imply that fematrin‐1 is involved in the formation of trophoblast hybrid cells even in different species trophoblastic cells.     

Isolation, characterization and quantitation of canine alpha-1-acid glycoprotein

The present study was designed to investigate the physicochemical properties of canine alpha-1-acid glycoprotein (AGP), and to establish a method for measuring serum AGP in canine. Fifty-three normal beagles were used in the study. AGP was purified from normal canine sera by successive ammonium sulfate precipitation, ion-exchange chromatography and gel filtration. The serum AGP concentration was measured by single radial immunodiffusion (SRID). Canine AGP had a molecular weight of 42,000 +/- 2,000 Da and contained 40.9% carbohydrate. Gel isoelectric focusing revealed microheterogeneity with 6-7 bands in a pI range (isoelectric points) of 3.2-3.8. AGP migrated to the alpha(1)-globulin region on immunoelectrophoresis. The serum AGP level of 35 normal beagles was 283.4 +/- 113.3 mug/ml. Canine AGP was isolated, and its physicochemical properties were clarified. SRID may be a useful method for quantification of serum AGP in canine.     

A field study of the associations between behaviors and reproductive performance in lactating sows

The present study examined factors associated with suckling, eating and postural behaviors (standing, lying and sitting) and the relationships between sow behaviors and reproductive performance. A commercial farrowing-to-finish farm with approximately 300 female pigs was visited eight times for 3 years to record sow behaviors using time-lapse video recorders for 24 hr. The lactational and postweaning reproductive performances of each sow were recorded using a recording software system. Statistical models were built to investigate the factors associated with each behavior. Regression analyses were performed to determine the associations between behavior measurements and adjusted 21-day litter weights (A21d-Wt), weaning-to-first-mating intervals (WMI) and occurrence of >or= 2 dead piglets during lactation. The means of suckling intervals and frequency of postural changes of the 145 recorded sows were 47.5 +/- 0.59 min and 86.8 +/- 2.97, respectively. As parity increased from 1 to >or= 4, the suckling interval and total standing duration increased (P<0.05). The number of nursing piglets and observation days 4 to 21 of lactation were not associated with the suckling intervals. A longer duration of lactation was associated with a longer total standing duration and a higher frequency of postural changes (P<0.05). Shorter suckling intervals were associated with heavier A21d-Wt (P<0.05). Each change in postural behavior increased the likelihood of occurrence of >or= 2 dead piglets during lactation (P<0.05). No postural or suckling behaviors were associated with WMI. Eating behavior was not associated with any reproductive performance. In conclusion, sow behaviors during lactation were related to lactational performance but were not related to postweaning reproductive performance.     

Intraoperative acridine orange photodynamic therapy and cribriform electron-beam irradiation for canine intranasal tumors: A pilot study

Untreated canine intranasal tumors carry a poor prognosis. We retrospectively evaluated the efficacy of marginal tumor resection in combination with intraoperative acridine orange (AO) photodynamic therapy (PDT) and 1 fraction of 5 Gy megavoltage irradiation for canine intranasal malignant tumors. When cribriform plate invasion or turbinate destruction around the cribriform plate was present, an additional fraction of 20 Gy was delivered with an electron beam during surgery. The study included 6 dogs, 2 of which were classified as stage I, 1 as stage II, and 3 as stage IV. The median local disease-free survival time and overall survival after the treatment were 8.5 and 13 months, respectively. Recurrence was noted in 2 of the 6 dogs after 4 and 7 months. Adverse events were mild (subcutaneous emphysema in 1 case, and rhinitis in 3 cases). Combination AO therapy may increase the tumor control time of dogs with marginally resectable intranasal malignant tumors.     

Isolation and characterization of canine tumor endothelial cells

The present study involved the isolation and characterization of canine tumor endothelial cells (TECs) from 2 malignancies. TECs were isolated using magnetic cell sorting following FITC labeling with UEA1 lectin, and they were characterized by measuring genetic and histopathological endothelial markers. Isolated TECs exhibited a cobblestone-like morphology and expressed both vascular endothelial growth factor receptor 2 (VEGFR2) and Von Willebrand factor (vWF). Further, both TECs and tumor cells derived from a seminoma exhibited increased C-X-C chemokine receptor type 7 (CXCR7) expression. However, CXCR7 expression was not detected in TECs and tumor cells derived from a hepatocellular carcinoma. Understanding TEC specific traits may be important in the development of more efficacious anti-angiogenic therapies that do not induce adverse effects.     

Establishing conditions for the storage and elution of rabies virus RNA using FTA? cards

The Flinders Technology Associates filter paper cards (FTA^® cards) can be used to store nucleic acid from various samples and are easily portable. However, RNA is physicochemically unstable compared with DNA, and appropriate methods have not been established for storage and extraction of RNA from FTA^® cards. The present study investigated the optimum conditions for storage and elution of viral RNA (vRNA) using rabies virus (RABV) applied to FTA^® cards. When TE buffer was used, the elution rates of vRNA increased with the length of the elution time. When the cards were stored at −80°C or −20°C, vRNA was stable over 3 months. Degradation of vRNAs occurred following storage at 4°C and room temperature, suggesting that RNA should be extracted from cards as soon as possible if no freezer is available. When we tried to amplify vRNA from RABV-infected animal brains applied to FTA^® cards and stored at −80°C for 6 months, we did not detect any amplified products with the primer set for 964 bp of RABV N gene. However, we were able to detect amplified products by increasing the elution time of vRNA from FTA^® cards from 30 min to 24 hr or by changing the primer sets to amplify 290 bp of N gene. Thus, we recommend extending the elution time for damaged or low concentration samples in FTA^® cards.     

A canine case of otitis media examined and cured using a video otoscope

Otitis media of the left ear was diagnosed by video otoscopic examination in a 7-year-old, intact male Shih-tzu dog (weight, 5.1 kg), that also had three complex ceruminous adenomas and a Pseudomonas aeruginosa infection in the left ear canal. In such cases, total ear canal ablation is usually required. However, a complete cure was achieved in the present case without total ear canal ablation. The complex ceruminous adenomas were excised using a diode laser, and repeated cleansing of the tympanic cavity and ear canal was implemented using a video otoscope. As a result, the ear canal was closed in a U-form, and the otitis media was cured.