Morphological and molecular characterization for a Japanese isolate of tomato powdery mildew Oidium neolycopersici and its host range

 A single conidium of tomato powdery mildew was isolated from heavily infected leaves of tomato (cv. Moneymaker) grown in the greenhouse of Kinki University, Nara Prefecture, Japan. It was successively multiplied so the morphological and taxonomic characteristics of the pathogen and its host range under high humidity conditions could be analyzed. The isolate KTP-01 of the tomato powdery mildew optimally developed infection structures at 25°C under continuous illumination of 3500 lx. More than 90% of the conidia germinated and developed moderately lobed appressoria. After forming haustoria, the pathogen elongated secondary hyphae from both appressoria and conidia. The hyphae attached to leaf surfaces by several pairs of appressoria and produced conidiophores with noncatenated conidia. In addition to its morphological similarity to Oidium neolycopersici, the phylogenetic analysis (based on the sequence of internal transcribed spacer regions of rDNA) revealed that KTP-01 could be classified into the same cluster group as O. neolycopersici. In host range studies, KTP-01 produced abundant conidia on the foliage of all tomato cultivars tested and tobacco (Nicotiana tabacum), and it developed faint colonies accompanied by necrosis on leaves of potato (Solanum tuberosum), red pepper (Capsicum annuum), petunia (Petunia × hybrida), and eggplant (S. melongena). The pathogen did not infect other plant species including Cucurubitaceae plants, which have been reported to be susceptible to some foreign isolates. Thus, the present isolate of the tomato powdery mildew was assigned as O. neolycopersici, a pathotype different from foreign isolates of the pathogen. Received: December 5, 2002 / Accepted: December 26, 2002 Acknowledgments This work was supported in part by a Grant-in-Aid (12660050) from the Ministry of Education, Culture, Sports, Science, and Technology of Japan. We express our deepest thanks to professor Dr. Y. Sato, Toyama Prefectural University, for his kind and valuable suggestion on taxonomic analysis of the powdery mildew pathogen described in the present study.     

Rapid detection of chitosanase activity in chitosanase gene-transformed strains of <Emphasis Type="Italic">Enterobacter cloacae</Emphasis> by lytic infection of specific bacteriophages

 In combination with lytic infection by virulent phages, a simple method for monitoring transgenic strains of Enterobacter cloacae was developed in this study. First, 15 strains of E. cloacae were used as indicator bacteria to isolate virulent phages with different host ranges. Of the phages isolated, five isolates (EcP-22, -35, -45, -55, and -70) were used to construct a set of virulent phages corresponding to all strains of E. cloacae. Using this phage set, a rhizosphere strain (KRM-055E) of E. cloacae was effectively screened from field soil. KRM-055E was transformed with a prokaryotic chitosanase gene csnSM1 and infected with the phage EcP-03, which can lyse the strain most effectively. The lysis of KRM-055E/csn occurred 2 h after inoculation, and the chitosanase activity was simply detected by dropping the lysate onto an agar plate containing glycol chitosan. The positive signal for chitosanase activity was detected in the 2-h lysates, and the signal intensity reached a maximum in the 5-h lysate. The present assay was simple, rapid, inexpensive, easy to perform, and applicable to another strains. Received: August 2, 2002 / Accepted: October 31, 2002 Acknowledgments This work was supported in part by a grant (no. 99L01205) from the “Research for the Future” program of the Japan Society for the Promotion of Science. We are grateful to Dr. M. Sato, National Institute of Agrobiological Science, Dr. H. Okamoto, Fukui Agricultural Experiment Station, and Dr. K. Tsuda, Kyoto Prefectural Institute of Agricultural Biotechnology, for kindly providing E. cloacae strains. We thank Dr. P. Park, Kobe University, for technical support with the electron microscopic observations.     

Root Rot of Miniature Roses Caused by Pythium helicoides

Miniature roses growing in an ebb-and-flow watering system developed dieback during the summer growing season of 1996 in Gifu Prefecture. The main diagnostic symptoms were chlorosis of leaf followed by blight, and a brown, water-soaked root rot followed by dieback. Pythium isolates were recovered from the rotted root. The isolates form proliferous ellipsoidal papillate sporangia, spherical smooth oogonia, elongate antheridia, and aplerotic oospores. The optimum temperature for hyphal growth was 35°C with a growth rate of 34 mm/24 hr. Optimum temperature of zoospore formation (25-30°C) was lower than that of mycelial growth, and zoospores were produced even at 10°C. The isolates were identified as P. helicoides on the basis of these characteristics. In pathogenicity tests disease severity was highest at the highest tested temperature (35°C) at which the disease naturally occurred in summer. Four days after inoculation, the leaves turned yellow and the roots had a water-soaked rot, followed by leaf blight and root dieback after 7 days. The disease transmission test showed that diseased plants were found throughout the bench after 10 days. Received 4 July 2001/ Accepted in revised form 10 October 2001     

Partial Purification of Thai Isolate of Citrus Huanglongbing (Greening) Bacterium and Antiserum Production for Serological Diagnosis

We aimed to improve the purification of citrus Huanglongbing (greening) bacterium (HB), Candidatus Liberobacter asiaticum and to produce an antiserum against HB. Periwinkle plants Catharantus roseum L. graft-inoculated with HB were used to produce an antiserum. All young leaves of new shoots incubated at 20–25°C and 25–30°C, a few mature leaves incubated at 20–25°C, and all mature leaves incubated first at 25–30°C and later transferred to 20–25°C developed yellowing symptoms and were then used to prepare immunogen. The HB was partially purified from these leaves by an improved method that included a macerating enzyme treatment of the midribs of infected leaves and homogenization of infected phloem sieve tissues. An antiserum raised against partially purified HB reacted clearly at a dilution of 1/16 with HB-infected citrus extract prepared at a concentration of 40 times, but did not react with healthy or tristeza virus-infected citrus extract in microprecipitin tests. Received 23 August 2002/ Accepted in revised form 4 December 2002     

具有交联结构的丝素蛋白质凝胶的理化特性

用二缩水甘油基乙醚作为交联剂制备的丝素蛋白质凝胶（ Ｃ Ｆ Ｇ） 具有良好的强度和柔韧性。根据制作条件可达压缩强度＞ １００ｇ／ ｍ ｍ ２ ,压缩变形率＞ ６０ ％ 。 Ｃ Ｆ Ｇ 在中性溶液中很少有丝素蛋白质溶出,但能被胰蛋白酶分解。     

Production of lipid mediators in mastitic milk of cow

Bovine mastitis is one of the most prevalent and costly diseases in the dairy industry. Lipid mediators are signaling molecules which coordinately and intricately modulate inflammation. They are produced from polyunsaturated fatty acids (PUFAs) in the cellular membrane via several enzymes including cyclooxygenase (COX) and lipoxygenase (LOX). In the present study, we performed comprehensive analysis of lipid production in milk obtained from clinical or subclinical mastitic cows using liquid chromatography/mass spectrometry. We detected 26, 24, and 40 kinds of lipid constantly in healthy, subclinical, and clinical mastitic milk, respectively. In clinical mastitic milk, the amount of a major n‐6 PUFA, arachidonic acid (AA), tended to increase, whereas amounts of major n‐3 PUFAs, eicosapentaenoic acid and docosahexaenoic acid, tended to decrease. The amounts of several AA‐derived lipids including COX‐catalyzed prostaglandin (PG) D₂ and PGE₂, and LOX‐catalyzed leukotriene (LT) B₄ were increased in clinical mastitic milk. Although subclinical mastitic milk represented similar trend of lipid production to healthy milk, amounts of several lipids such as LTD₄, 14,15‐dihydroxyeicosatrienoic acid, and 14‐epoxyeicosatrienoic acid changed. These findings would be helpful for better understanding of mastitis pathology and give us some insights to develop a new diagnostic and therapeutic strategy.     

Effects of the butyric acid‐producing strain Clostridium butyricum MIYAIRI 588 on broiler and piglet zootechnical performance and prevention of necrotic enteritis

The objective of this study was to assess the effects of a probiotic strain Clostridium butyricumMIYAIRI 588 (CBM588) on broiler and weaned piglet health and zootechnical performance. Five field studies were carried out in broilers and five in weaned piglets under European feed additive guidelines. Each study followed a randomized blocked design with two treatments: Control (basal diet) and CBM588 supplemented groups. The zootechnical performance parameters selected were body weight, daily gain, feed intake and feed efficiency (feed:gain). Broilers fed diets with CBM588 gained significantly more weight (+2%, p < .001) and exhibited significantly better feed efficiency (?1.6%, p < .001) in comparison with Controls. Similarly, analysis of pooled data of weaned piglet trials showed that CBM588‐fed piglets were significantly heavier than Controls (+2.6%, p = .014), exhibited significantly higher mean daily gain (+4.7%; p = .004), and significantly improved feed efficiency (?4.2%, p = .001). In addition to the zootechnical efficacy studies, the preventive effect of CBM588 on necrotic enteritis (NE) was assessed in a natural challenge model in broilers where CBM588 reduced the incidence and severity of NE lesions. These data indicate the potential of CBM588 to improve broiler and weaned piglet zootechnical performance, and to make a positive contribution to animal health.     

Cardiac ganglioneuroma in a juvenile pig

A cardiac mass (3 × 5 × 3 cm) was detected at the base between the right auricular wall and right vena cava of a slaughtered 6-month-old female mixed-breed pig during a meat inspection. The tumor comprised infiltrative prominent interweaving fascicles of Schwann cells with Verocay bodies. Moreover, the ganglion cells were scattered or aggregated throughout the neoplastic tissue. The ganglion and Schwann cells had neither cellular atypism nor mitosis. On the basis of the bearing site as well as the morphological and immunohistochemical features, this is the first case of a cardiac ganglioneuroma in a pig.     

Identification of an antihypertensive peptide derived from chicken bone extract

A novel angiotensin‐converting enzyme (ACE) inhibitory peptide was isolated and purified from chicken bone extract by enzymatic digestion. The peptide was defined as an ACE inhibitor, and it demonstrated antihypertensive activity following oral administration to spontaneously hypertensive rats (SHRs). The results of this study suggest that peptides derived from an extract of chicken bones, administered orally, have the ability to reduce the blood pressure of SHRs significantly over a short period of time (3 h). Moreover, the blood pressure then remains low for 3 h. This peptide derived from chicken bones may therefore have great value as a short‐term remedy for chronic conditions such as high blood pressure. The amino acid sequence of the peptide was YYRA (Tyr‐Tyr‐Arg‐Ala), which was the origin of the Ig heavy chain V region (27–30 position). The IC₅₀ value of its synthetic peptide was 33.9 μg/mL. We suggest that the ACE inhibitory and antihypertensive peptides derived from chicken bone extract may contribute to develop physiologically functional foods or improve food functionality.