Nucleotide sequence polymorphisms of beta1‐, beta2‐, and beta3‐adrenergic receptor genes on Jinhua,Meishan, Duroc and Landrace pigs

The full amino acid coding sequences of adrenergic receptor genes beta1, beta2, and beta3 (ADRB1, ADRB2, and ADRB3)were determined for Jinhua, Meishan, Duroc and Landrace pigs. Non‐synonymous substitution of Arg458Pro was found in the porcine ADRB1 gene, resulting in a 469 amino acid sequence. Continuous substitutions of Asn29Asp and Glu30Gln were found in the porcine ADRB2 gene, resulting in a 418 amino acid sequence. Additionally, a Lys30 polymorphism of the ADRB2 gene was found in the Jinhua pigs. There were three non‐synonymous substitutions of Asn24Thr, Arg264Gln and Asn398Asp on the porcine ADRB3 gene. A thymine insertion in the ADRB3 gene, resulting in a protein with two fewer amino acids, was found in the Jinhua and Meishan pigs. To assess the effect of ADRB polymorphisms on porcine subcutaneous fat layer thickness, we calculated the genetic frequency of the variants in fatty and lean groups, each consisting of 24 pigs that were crossbreds of Duroc and Jinhua pigs. The effect of the ADRB3 gene polymorphism was not evaluated, because there was insufficient variation on the ADRB3 gene in the examined groups. Although Fisher's exact test showed no significant difference in the frequency of ADRB1 and ADBR2 variants between the two groups, the Arg458 variant of ADRB1 was higher (P = 0.11) in the lean group, and pigs in that group had a thinner fat layer than did those with the Pro458 variant. These results imply a possibility of ADRB1 polymorphism as a minor factor in porcine fat layer thickness. The Asp29 variant of ADRB2 was higher in the lean group (P = 0.11), and the Glu30 variant was higher in the fatty group (P = 0.15), but the Asp29 variant was found only in the Chinese pigs. Thus, the effect of ADRB2 polymorphisms was not clear in this study.     

Tuberculosis seroprevalence and comparison of hematology and biochemistry parameters between seropositive and seronegative captive Asian elephants of Nepal

We conducted a tuberculosis (TB) serosurveillance program of captive elephants in Nepal and compared hematology and biochemistry parameters between seropositive and seronegative elephants. A total of 153 elephants (male=20, female=133) from four national parks were tested for TB using the ElephantTB STAT-PAK^® Assay (ChemBio Diagnostic Systems, Inc., Medford, NY, USA). The mean reported age for 138 elephants was 38.5 years (range 2–71 years). Seroprevalence for TB was 21.56% (33/153). The majority of seropositive elephants were female (n=30) and from Chitwan National Park (n=29). The occurrence of TB seropositive cases in other more remote national parks suggests TB may be widespread among the captive elephant population of Nepal. Hematology and biochemistry analyses were performed on 13 and 22 seropositive elephants, respectively and, nine elephants from a seronegative TB herd for comparison. Hematology parameters (hemoglobin, packed cell volume, platelet, white blood cells, and erythrocyte sedimentation rate) were comparable between the two groups. Total protein, globulin, and lactate dehydrogenase were significantly higher in seronegative elephants, and bilirubin was significantly higher in seropositive elephants whereas blood urea nitrogen, creatinine, glutamic oxaloacetic transaminase/aspartate aminotransferase (GOT/AST), glutamic pyruvic transaminase/alanine aminotransferase (GPT/ALT), gamma glutamyl transferase (GT), and albumin were not significantly different. The range of biochemical parameters that were significantly different between seropositive and seronegative elephants had narrow ranges. Thus, the potential of these parameters as a direct biomarker for TB diagnosis is limited based on the findings in this study. We recommend including blood parameters in future TB surveillance studies.     

Synergistic Antifungal Activity of Chitinolytic Enzymes and Prodigiosin Produced by Biocontrol Bacterium, Serratia marcescens Strain B2 against Gray Mold Pathogen, Botrytis cinerea

Serratia marcescens strain B2 is an effective biocontrol agent against gray mold of cyclamen, caused by Botrytis cinerea Persoon. Strain B2 has strong antifungal activity against B. cinerea in vitro. The culture filtrate was found to contain different types of chitinolytic enzyme activity, including endochitinase and chitobiase activity. Four chitinolytic enzymes were detected among the extracellular proteins of strain B2. Two major enzymes, a 58-kDa endochitinase and a 98-kDa chitobiase, were purified by chromatography and electrofocusing, respectively. Both enzymes inhibited spore germination of B. cinerea. And a red pigment, prodigiosin, extracted and purified from the bacterial cells, also inhibited spore germination. When prodigiosin and the chitinolytic enzymes were applied in concert, a synergistic inhibitory effect was observed. S. marcescens strain B2 has multiple modes of action against the pathogen. Received 14 February 2001/ Accepted in revised form 31 May 2001     

Functional Analysis of an ATP-Binding Cassette Transporter Gene in Botrytis cinerea by Gene Disruption

The BMR1 gene encoding an ABC transporter was cloned from Botrytis cinerea. To examine the function of BMR1 in B. cinerea, we isolated BMR1-deficient mutants after gene disruption. Disruption vector pBcDF4 was constructed by replacing the BMR1-coding region with a hygromycin B phosphotransferase gene (hph) cassette. The BMR1 disruptants had an increased sensitivity to polyoxin and iprobenfos. Polyoxin and iprobenfos, structurally unrelated compounds, may therefore be substrates of BMR1. Received 18 December 2000/ Accepted in revised form 18 April 2001     

Classification of the spermatogenic cycle,seasonal changes of seminiferous tubule morphology and estimation of the breeding season of the large Japanese field mouse (Apodemus speciosus) in Toyama and Aomori prefectures, Japan

The large Japanese field mouse, Apodemus speciosus, is a potential indicator of environmental stress, but this function has not been confirmed by histological studies. Since environmental stress affects the reproductive function of mice, we determined the reproductive characteristics of this species at two locations: Toyama (36°35ʹN, 137°24ʹE) and Aomori (40°35ʹN, 140°57ʹE). Mice were captured during May–November (n=119) and July–November (n=146) at these locations, respectively. We classified the breeding season from the numbers of pregnant females and young, in addition to the spermatogenic cycle and seasonal changes in seminiferous tubule morphology of males. Testicular weight was measured, and seminiferous tubule morphology was examined histologically. Fourteen stages were found in the seminiferous epithelium cycle based on acrosome formation and spermatid head morphology. At both locations, the breeding season peaked from late summer to early autumn and possibly in spring. Spermatogenic activity was classified into 4 periods from June to November: resting around June and October–November; resumptive around July; active around August; and degenerative around September. During the resting period, the seminiferous tubules consisted of Sertoli cells, spermatogonia and spermatocytes. Spermatogenesis began during the resumptive period, and spermatids were observed. During the active period, active spermatogenesis and a broad lumen were observed. During the degenerative period, spermatogenesis ended, and Sertoli cells, spermatogonia, spermatocytes and degenerating exfoliated round spermatids were observed. This study provides scientific information about the testicular histopathological evaluations of the large Japanese field mouse for its use as an index species of environmental pollution.     

Successful measures to prevent the spread of bovine papular stomatitis in a dairy farm

Nasal papules and oral ulcers were observed in calves that were group-housed at a dairy farm. The calves were diagnosed with bovine papular stomatitis (BPS) due to parapoxvirus (PPV) infection based on virologic examinations using polymerase chain reaction to detect PPV. To prevent the spread of BPS, we isolated the affected calves, made procedural changes so that the affected herd was managed after the healthy herd, disinfected the bedding with slaked lime, disinfected the stalls and fences with invert soap, and changed the animals’ feed to soft grass which does not damage the oral cavity. As a result, we succeeded in control the infection quickly.     

Improving the accuracy of estimating blood calcium concentration in Holstein cows using electrocardiographic variables

We previously reported the possibility of using the electrocardiogram variable to estimate blood calcium (Ca) concentration in dairy cows based on the strong positive correlation between the blood Ca concentration and the inverse of the corrected ST peak interval (STc⁻¹). To improve the accuracy of the estimation of blood Ca concentration, we investigated the relationship between blood Ca concentration and STc⁻¹ for each postpartum day and available variables other than STc⁻¹. We measured multiple variables (milk yield, calving number, age, body temperature, etc.), including serum total Ca concentration (tCa), blood ionized Ca concentration (iCa) and STc⁻¹ in 462 Holstein cows on days 0, 1, 2, 3, 5, and 7 postpartum. A very high correlation was observed between iCa and tCa. The association between tCa and STc⁻¹ for each postpartum day had a high coefficient of determination of 0.61–0.79 postpartum 0–2 days but decreased after the third day. In the investigation using the data from postpartum days 0–2, STc⁻¹, heart rate interval, calving number, and age were highly correlated with tCa. In addition, a multiple regression equation was obtained with tCa as the objective variable and STc⁻¹ and calving number as explanatory variables. The estimation accuracy was improved as compared with the simple regression equation using only STc⁻¹ as the explanatory variable. This multiple regression equation was used for 11 cows suspected of having hypocalcemia, and it was able to correctly detect cows requiring early treatment, except for one cow.