Melanin biosynthesis in Pyricularia oryzae: Site of tricyclazole inhibition and pathogenicity of melanin-deficient mutants

Tricyclazole (5-methyl-1,2,4-triazolo[3,4-b]benzothiazole) inhibits melanin synthesis in Pyricularia oryzae at concentrations less than 0.01 μg/ml. The primary site of inhibition in the biosynthetic pathway occurs between scytalone and vermelone. Accumulation of several metabolites derived from melanin precursors along branch pathways is associated with inhibition of melanin biosynthesis. At low tricyclazole concentrations (0.01–1 μg/ml), predominant accumulation of 2-hydroxyjuglone and 3,4-dihydro-3,4,8-trihydroxy-1-(2H)-naphthalenone (3,4,8-DTN) occurs as a result of the primary block between 1,3,8-trihydroxynaphthalene and vermelone. As the concentration of tricyclazole is increased from 1 to 10 μg/ml, flaviolin accumulation is markedly enhanced, whereas that of 3,4,8-DTN and 3,4-dihydro-4,8-dihydroxy-1-(2H)-naphthalenone is depressed, indicating possible secondary sites of inhibition in the main and branch pathways. Five melanin-deficient mutants of P. oryzae that phenotypically resemble the tricyclazole-treated wild-type strain were nonpathogenic or rarely infected two rice varieties. Three of the mutants studied were genetically defective in the melanin biosynthetic pathway at the site blocked by tricyclazole in the wild type. The wild-type strain converted both scytalone and vermelone to melanin; whereas the three mutants and the tricyclazole-treated wild type converted only vermelone to melanin. The data suggest a relationship between melanin biosynthesis and pathogenicity in P. oryzae.     

Endotoxin from Fusobacterium necrophorum of bovine hepatic abscess origin.

The endotoxic activity of Fusobacterium necrophorum bov 5 was investigated. The supernatant (S) fluid and cell wall (CW) preparation, obtained after differential centrifugation of the ruptured cell mass, were lethal for mice. The toxicity of the S fluid was stable during prolonged storage, treatment with formalin, and heating for 15 minutes at 80, 100, and 121 C, but was destroyed by alkaline hydrolysis with 0.25 N NaOH. The toxic factor was found in a high molecular weight (MW) fraction after gel filtration. The properties exhibited by the toxic S fluid resembled those of endotoxic lipopolysaccharide (LPS). Extracted and partially purified LPS (endotoxin) from F necrophorum bov 5 demonstrated a mouse median lethal dose (mouse LD50) of 16.8 mg/kg of body weight. The toxic LPS material, a high molecular weight moiety as estimated by gel filtration, was resistant to ribonuclease (RNase), deoxyribonuclease (DNase), and pronase treatment. A positive Shwartzman reaction (median skin lesion dose (SLD50) equal to 3.32 mug/kg of body weight) and biphasic fever response (minimal dose required to produce a fever index of 40 sq cm which falls on the linear portion of dose-response curve (FL40) equal to 0.41 mug/kg of body weight) further indicated the toxin was endotoxin in nature. The LPS from F necrophorum bov 5 was less toxic than Salmonella typhimurium LPS; but had considerable toxicity for experimental animals. The toxic activity of the partially purified F necrophorum bov 5 endotoxin was separated into 2 fraction regions by diethylaminoethyl (DEAE)-cellulose chromatography. The data provide evidence for the production of a potent endotoxin, possibly composed of more than one toxic component, which may be released upon cell disruption.     

Immunocytochemical localization of the Bacillus sphaericus binary toxin components in Culex quinquefasciatus (Diptera: Culicidae) larvae midgut

This study describes the immunocytochemical localization of the Bacillus sphaericus 2362 binary toxin components, BinA and BinB, in Culex quinquefasciatus larvae that had been intoxicated with this entomopathogen. Ultrathin sections of C. quinquefasciatus larvae midgut embedded in the hydrophilic resin L.R. White were incubated with the antibodies anti-BinA or anti-BinB and then revealed with goat anti-rabbit IgG coupled to gold particles. Immunocytochemical detection demonstrated the presence of specific labeling in ultra-thin sections that had been incubated with the BinA antiserum. Gold particles were detected on the apical areas of cell membranes and inside the epithelial cell cytoplasm, particularly the mitochondria, of cells from the gastric caeca and posterior stomach in larvae exposed during 2 or 24 h to the entomopathogen. A similar labeling pattern was observed in ultrathin sections from both midgut regions when incubated with BinB antiserum.     

A coat protein transgene from a Scottish isolate of potato mop-top virus mediates strong resistance against Scandinavian isolates which have similar coat protein genes

Resistance tests were made on seedlings of transformed lines of Nicotiana benthamiana which contain a transgene encoding the coat protein (CP) gene of a Scottish isolate of potato mop-top virus (PMTV). This transgene has been reported to confer strong resistance to the PMTV isolate from which the transgene sequence was derived and also to a second Scottish isolate. Plants of lines of the transgenic N. benthamiana were as resistant to two Swedish and two Danish PMTV isolates as to a Scottish isolate, and of five lines tested, greater than 93.5% of transgenic plants were immune. The coat protein gene sequences of these four Scandinavian isolates were very similar to those of the two Scottish isolates. The greatest divergence between the isolates was three amino acid changes and there was less than 2% change in CP gene nucleotide sequence. It is concluded that the PMTV CP transgene used in these experiments could confer resistance against isolates from different geographical areas because it is becoming apparent that the CP genes of PMTV isolates are highly conserved.     

The re-emergence of indigenous forest in an urban environment, Christchurch, New Zealand

Christchurch, the second largest city in New Zealand is a planned city on a coastal plain on the east coast of the South Island. The birth of the city and the subsequent century of development was characterised by colonial values and tree and garden planting with familiar European species along with those from Australia, North America, and eventually all other continents. The image of an “English garden city” with classical parks of oaks and willow-lined rivers became the accepted norm and the way in which the city has been promoted to potential tourists. Gardening is one of the top two recreational activities and exotic species greatly outnumber native species in the flora and in gardens. This has had serious consequences for the highly fragmented and degraded indigenous vegetation and its co-adapted wildlife. A few hardy indigenous species continued to regenerate through this period, but since the 1970s, there has been a progressive change of attitude and interest in reclaiming the natural heritage of the city, manifest in widespread private and public planting of indigenous species and active habitat restoration. In this article we examine the indigenous and exotic shrub and tree components of the Christchurch flora as planted street trees, in domestic gardens, and in parks. We also present data on shrub and tree regeneration in parks and domestic gardens in the city. Indications are that the more sensitive, less intrusive management of urban environments, combined with the greater density of indigenous seed sources, has allowed regeneration of a wide range of indigenous species across a broad spectrum of habitats – from neglected gardens to pavement cracks to exotic plantations. This is despite the competition from the prodigious seed banks and density of exotic trees, shrubs, and ground covers and albeit minimal impacts of introduced browsing and seed eating mammals. If the present trends continue through appropriate management and facilitation, these tentative signs of native forest regeneration should eventually proliferate into a sustainable mixed origin urban forest that resurrects and preserves the natural character of the region.     

Expression of the KIT protein (CD117) in primary cutaneous mast cell tumors of the dog.

Thirty-one canine cutaneous masses, diagnosed as mast cell tumors (MCT) by histopathologic analysis, were used to evaluate the immunohistochemical pattern of expression of KIT protein (CD117), a type III tyrosine kinase protein involved in mast cell growth and differentiation. Lesions were graded as I (well differentiated), II (intermediate differentiation), or III (poorly differentiated) according to the following morphologic features: invasiveness, cellularity and cellular morphology, mitotic index, and stromal reaction. Immunohistochemical KIT expression was compared with histologic grade and some histomorphologic features (cell differentiation and nuclear grade) evaluated separately. A possible predictive role of biologic behavior in MCTs for KIT expression was also investigated. Immunohistochemical analysis revealed three different patterns of KIT expression: a cytoplasmic diffuse pattern, a membranous pattern with immunostaining located on the cell surface, and a cytoplasmic perinuclear pattern, where KIT expression was detected in the cytoplasm of the neoplastic mast cells, close to the nucleus. Statistical analysis showed a close relationship between different KIT immunohistochemical patterns and histologic grade (P < 0.00000), cell differentiation (P < 0.00000), and nuclear grade (P < 0.0024). According to Kaplan-Meier-estimated survival curves compared by survival analysis, KIT expression was significantly associated with survival time (P = 0.037) but not cancer-free interval (P = 0.50). Similar to other well-known histomorphological features, KIT expression is a useful parameter of malignancy in cutaneous MCTs. KIT expression also predicted the biological behavior of the tumors in this study.     

An indirect enzyme linked immunosorbent assay for the detection of bovine antibodies to multiple Leptospira serovars

An indirect enzyme linked immunosorbent assay was developed for the detection of bovine antibodies to multiple pathogenic Leptospira serovars, including canicola, copenhageni (represents icterohaemorrhagiae), grippotyphosa, hardjobovis, pomona, and sejroe. The antigen utilized in this assay was a sonicated mixture of equal parts of killed whole cells of each of the 6 serovars named above. A mouse monoclonal antibody against bovine immunoglobulin (Ig)G₁ that was conjugated with horseradish peroxidase was used for detection of bound antibodies. This assay was evaluated with sera (n = 3107) that were microscopic agglutination test (MAT)-negative (at a 1:100 dilution) for each of the 6 serovars listed above and sera (n = 601) that were MAT-positive (at a 1:100 dilution) for 1, or any combination of the 6 listed serovars. In addition, sera from serial weekly bleedings of cows, which were individually experimentally infected with serovars hardjobovis, copenhageni, grippotyphosa, or canicola, were also tested in this assay.

At an optimal cut-off point determined by receiver operating characteristic (ROC) curve analysis, the relative sensitivity and specificity of the assay were 93.5% (95% confidence interval = 91.2% to 95.3%) and 94.7% (95% confidence interval = 93.9% to 95.5%), respectively. This assay was able to detect antibody in the sera of animals experimentally infected with serovar hardjobovis as early as 1 week postinoculation

     

Caprine toxoplasmosis in Southern Brazil: a comparative seroepidemiological study between the indirect immunofluorescence assay,the enzyme-linked immunosorbent assay,and the modified agglutination test

Tropical Animal Health and Production - This study investigated the prevalence of caprine toxoplasmosis in goat herds from Southern Brazil by the indirect immunofluorescence assay (IFA), and...     

Use of two doses of cloprostenol in different intervals for estrus synchronization in hair sheep under tropical conditions

Tropical Animal Health and Production - This study evaluated the effect of two doses of prostaglandin at different intervals on reproductive parameters of crossbred ewes. In Experiment 1, 30 ewes...