Inoculation of Lupinus luteus with White Root Rot Fungus, Rosellinia necatrix, to Estimate Virulence

An inoculation method using Lupinus luteus was developed for estimating virulence of isolates of the white root rot fungus, Rosellinia necatrix Prillieux. Fungal cultures grown on pieces of mulberry twigs were placed in contact with the hypocotyl of 3-week-old seedlings growing in pots of soil. Disease development was uniform and reproducible in repeated experiments. Of 24 isolates with double-stranded RNA, eight were weakly virulent. This method is useful throughout the year for estimating virulence of many isolates of the fungus and for screening for hypovirulent isolates. Received 2 March 2001/ Accepted in revised form 25 May 2001     

D-Alanine-D-Alanine Ligase Gene (ddl) of Erwinia chrysanthemi Strain EC16 I. Isolation and Gene Dosage Effect on Pectate Lyase Synthesis

New regulatory gene for pectate lyase (Pel) production of Erwinia chrysanthemi strain EC16 was searched by observing the gene dosage effect of each cosmid library in Pel production. From this survey, a cosmid clone, p5A, had reduced in Pel production under both inducing and non-inducing conditions and caused less tissue maceration of potato tubers. The 2.64-kb HindIII fragment from p5A was found to be responsible for this phenotype and to contain one major ORF consisting of 1107 nucleotides, which had homology with ddlA (49.6%) and ddlB (49.6%) of Escherichia coli and with ddlA (44.6%) of Salmonella typhimurium. These genes had been shown to encode D-alanine-D-alanine ligase (Ddl), an enzyme which is involved in the synthesis of peptidoglycan. This ORF encodes 368 amino acid residues and its molecular weight is estimated to be 43 kDa. This ORF of EC16 could complement ddl deficient mutant of Escherichia coli. Received 15 May 2002/ Accepted in revised form 20 June 2002     

稻曲病菌离体培养体系的构建与优化

稻曲病是水稻穗期的一种重要病害,严重影响稻米的产量和品质。稻曲病菌Ustilaginoidea virens 在离体培养条件下生长缓慢,严重制约了杀菌剂室内生测试验的观察和高效筛选。本研究以稻曲病菌培养基为研究对象,通过筛选、组合和优化固体培养体系的氮源、碳源、凝固剂以及阳离子浓度,以期构建适合稻曲病菌生长的室内培养体系。结果发现:在5种常见的商品化培养基中,稻曲病菌的生长速率仅为0.7~2.3 mm/d;而在18种配制培养基中,有77%的培养基对稻曲病菌表现出不同程度的生长促进效应,其中蛋白胨蔗糖结冷胶 (PSGG) 培养基的促进效果最显著 (P < 0.05),生长速率达3.4 mm/d,且其对不同地理分布的稻曲病菌菌株均有生长促进效应。此外,在以结冷胶作为凝固剂的培养基中添加质量分数为0.02%的硫酸镁 (Mg2+),可有效提高培养基的凝固程度,且对菌丝体的生长速率无影响。在适合稻曲病菌生长的PSGG培养基中,蛋白胨和蔗糖是合适的氮源和碳源组合,结冷胶是合适的凝固剂。此研究结果可提高室内培养稻曲病菌的生长速率,为进一步理解稻曲病菌在离体条件下的生长机制及提高室内杀菌剂生测试验的效率提供了重要参考。     

气相色谱-串联质谱检测蔬菜中氟吡菌酰胺及其代谢物残留

采用气相色谱-串联质谱（GC-MS/MS）技术建立了同时检测7种类型蔬菜中氟吡菌酰胺及其代谢物残留的分析方法。样品采用乙腈提取,无水硫酸镁和氯化钠盐析后,经分散固相萃取（PSA）净化,GC-MS/MS检测。结果表明:在0.01、0.1和1 mg/kg 3个添加水平下,氟吡菌酰胺及其代谢物2-（三氟甲基）苯甲酰胺（BZM）的平均回收率在80%~108%之间,相对标准偏差（RSD）在1.5%~8.4%之间。氟吡菌酰胺及其代谢物在7种蔬菜中的检出限（LOD）（S/N=3）分别为2.17~3.13 μg/kg和1.22~2.04 μg/kg,两者的定量限（LOQ）均为0.01 mg/kg。该方法操作简便、稳定和快速,可以满足大部分蔬菜中氟吡菌酰胺及其代谢物的定性与定量分析要求。     

Phytoremediation of the herbicides atrazine and metolachlor by transgenic rice plants expressing human CYP1A1, CYP2B6, and CYP2C19

This study evaluated the expression of human cytochrome P450 genes CYP1A1, CYP2B6, and CYP2C19 in rice plants (Oryza sativa cv. Nipponbare) introduced using the plasmid pIKBACH. The transgenic rice plants (pIKBACH rice plants) became more tolerant toward various herbicides than nontransgenic Nipponbare rice plants. Rice plants expressing pIKBACH grown in soil showed tolerance to the herbicides atrazine, metolachlor, and norflurazon and to a mixture of the three herbicides. The degradation of atrazine and metolachlor by pIKBACH rice plants was evaluated to confirm the metabolic activity of the introduced P450s. Although both pIKBACH and nontransgenic Nipponbare rice plants could decrease the amounts of the herbicides in plant tissue and culture medium, pIKBACH rice plants removed greater amounts in greenhouse experiments. The ability of pIKBACH rice plants to remove atrazine and metolachlor from soil was confirmed in large-scale experiments. The metabolism of herbicides by pIKBACH rice plants was enhanced by the introduced P450 species. Assuming that public and commercial acceptance is forthcoming, pIKBACH rice plants may become useful tools for the breeding of herbicide-tolerant crops and for phytoremediation of environmental pollution by organic chemicals.     

Evaluation of the air sac volume of penguins with respiratory diseases using computed tomography

Captive penguins with respiratory diseases exhibit advanced pathological conditions upon the appearance of clinical signs. Therefore, the successful treatment of respiratory diseases remains difficult after the onset of clinical signs, leading to high mortality rates. In this study, we measured air sac volume using computed tomography (CT) to evaluate the respiratory condition of penguins. In a regular quarterly health checkup, blood samples were collected from 45 penguins housed at an aquarium in Hokkaido, Japan. A total of 12 penguins with abnormal blood parameters underwent CT. The air sac volumes were calculated in three-dimensional CT, and the scatter plots of the air sac volumes and body weights were analyzed. No correlation was found between the air sac volume and body weight in both the gentoo and king penguins. Two gentoo penguins with infiltration and one king penguin with multiple nodules on CT were tentatively diagnosed with aspergillosis and treated with oral administration of itraconazole. Follow-up CT examination was performed until the outcome: healed or died. The mean air sac volumes of the two gentoo penguins, which recovered after treatment, increased from 273.9 and 329.0 cm³ before healing to 449.0 and 424.6 cm³ after healing, respectively. Meanwhile, the air sac volume of the king penguin, which subsequently died, decreased from 1,556.9 to 920.6 cm³ despite treatment. Changes of the air sac volume in the same individual could be useful for evaluating the respiratory condition of penguins.     

Detection and molecular characterization of bovine leukemia virus in beef cattle presented for slaughter in Egypt

Bovine leukemia virus (BLV) is the etiological agent of enzootic bovine leukosis, the most common neoplastic disease of cattle worldwide and a serious problem for the cattle industry. Previous studies have shown the molecular prevalence of BLV and the coexistence of BLV genotype-1 and -4 in Egyptian dairy cattle; however, the molecular characteristics of BLV in Egyptian beef cattle are unknown. Therefore, we collected blood samples of 168 beef cattle from slaughterhouses in three governorates in Egypt. Based on BLV-CoCoMo-qPCR-2 targeting long terminal repeats and nested PCR targeting the env-gp51 gene, the BLV provirus infection rates were found to be 47/168 (28.0%) and 42/168 (25.0%), respectively. Phylogenetic analysis based on 501 bp of the BLV env-gp51 gene from 42 BLV isolates revealed that at least six distinctive strains (b, e, f, g, x, and z) were prevalent in cattle across the examined regions. Furthermore, phylogenetic analysis of the 420 bp sequence of the BLV env-gp51 region of the six strains against 11 known genotypes showed that the strains b, e, f, and g were clustered into genotype-1, and strains x and z were clustered into genotype-4. Our results also indicated that strains b and x exist in both dairy and beef cattle in Egypt. The present study is the first to detect and genotype BLV among beef cattle in Egypt.     

Non‐surgical transfer of vitrified porcine embryos using a catheter designed for a proximal site of the uterus

This study aimed to compare the efficiency of non‐surgical embryo transfer (ET) using a newly developed catheter, which enables transferring embryos into a proximal site of the uterus (mostly uterine body), and surgical ET of vitrified porcine embryos. In Experiment 1, the catheter was inserted into 12 gilts, with each half of the group allocated to skilled or novice operators. The time required for insertion into the uterus did not differ between skilled and novice operators (4 min 9 s and 4 min 6 s, respectively). In Experiment 2, 12 gilts were used as recipients for non‐surgical and surgical ET with vitrified embryos (n = 6, each). There was no significant difference in the rate of piglet production based on the number of transferred embryos between surgical and non‐surgical ET (25.8% vs. 15.4%, p = .098). The results suggest that non‐surgical ET catheter allowed for easy insertion and transfer of embryos without special training. Although the catheter is effective for deposition of embryos into the proximal site of uterus, the efficiency of piglet production is not enhanced compared with surgical ET. The ET method using this catheter, being labor‐saving and less‐invasive, may contribute to the improvement of ET in pigs.