Usefulness of cefovecin disk‐diffusion test for predicting mecA gene‐containing strains of Staphylococcus pseudintermedius and clinical efficacy of cefovecin in dogs with superficial pyoderma

Background –  Cefovecin has been widely used to treat skin infections in dogs. The relationship of the cefovecin disk‐diffusion test results to the presence of the mecA gene and the clinical efficacy of cefovecin have not been fully evaluated. Hypothesis/Objectives –  To determine the usefulness of an in vitro cefovecin disk‐diffusion test in predicting the presence of the mecA gene in Staphylococcus pseudintermedius, as well as the in vivo efficacy of cefovecin therapy in dogs with superficial pyoderma. Methods –  Twenty‐six S. pseudintermedius strains isolated from 22 dogs with pyoderma were used. In vitro disk‐diffusion test results of cefovecin were compared with agar‐dilution test results, the presence of the mecA gene, and the improvement in clinical scores of dogs with superficial pyoderma at 14 days post treatment. Results –  There was a significant linear correlation (r = ?0.83) between the diameter of the obvious zone of inhibition by disk diffusion and the minimal inhibitory concentration for cefovecin (P < 0.0001). Receiver operating characteristic analysis revealed that zone diameters between 25 and 27 mm exhibited better sensitivity (92.9%) and specificity (100.0%) for detection of strains carrying the mecA gene. The mean improvement in clinical scores in dogs carrying cefovecin‐resistant strains was significantly lower than in dogs carrying cefovecin‐susceptible strains (P < 0.01). Conclusions and clinical importance –  The cefovecin disk‐diffusion test with a cut‐off value estimated in this study was valuable for predicting mecA gene carriage in S. pseudintermedius, as well as the in vivo efficacy of cefovecin therapy in dogs with superficial pyoderma caused by S. pseudintermedius.     

Oidium neolycopersici: intraspecific variability inferred from amplified fragment length polymorphism analysis and relationship with closely related powdery mildew fungi infecting various plant species

Previous works indicated a considerable variation in the pathogenicity, virulence, and host range of Oidium neolycopersici isolates causing tomato powdery mildew epidemics in many parts of the world. In this study, rDNA internal transcribed spacer (ITS) sequences, and amplified fragment length polymorphism (AFLP) patterns were analyzed in 17 O. neolycopersici samples collected in Europe, North America, and Japan, including those which overcame some of the tomato major resistance genes. The ITS sequences were identical in all 10 samples tested and were also identical to ITS sequences of eight previously studied O. neolycopersici specimens. The AFLP analysis revealed a high genetic diversity in O. neolycopersici and indicated that all 17 samples represented different genotypes. This might suggest the existence of either a yet unrevealed sexual reproduction or other genetic mechanisms that maintain a high genetic variability in O. neolycopersici. No clear correlation was found between the virulence and the AFLP patterns of the O. neolycopersici isolates studied. The relationship between O. neolycopersici and powdery mildew anamorphs infecting Aquilegia vulgaris, Chelidonium majus, Passiflora caerulea, and Sedum alboroseum was also investigated. These anamorphs are morphologically indistinguishable from and phylogenetically closely related to O. neolycopersici. The cross-inoculation tests and the analyses of ITS sequences and AFLP patterns jointly indicated that the powdery mildew anamorphs collected from the above mentioned plant species all represent distinct, but closely related species according to the phylogenetic species recognition. All these species were pathogenic only to their original host plant species, except O. neolycopersici which infected S. alboroseum, tobacco, petunia, and Arabidopsis thaliana, in addition to tomato, in cross-inoculation tests. This is the first genome-wide study that investigates the relationships among powdery mildews that are closely related based on ITS sequences and morphology. The results indicate that morphologically indistinguishable powdery mildews that differed in only one to five single nucleotide positions in their ITS region are to be considered as different taxa with distinct host ranges.     

Reducing eye fatigue through the use of wood

To clarify if eye fatigue is lessened when looking at wood, we carried out objective examinations using a near-point ruler and also performed sensory evaluations. Visual contact target materials were a white plastic panel, a black plastic panel or a wood panel which had Japanese ash flat grain surface. Each size of the materials was 140 × 280 mm. Test subjects were 30 undergraduate and graduate students. Each subject’s visual distance to a panel was 40–50 cm. The results showed that subjects suffered from more eye fatigue when looking at white and black plastic panels compared to looking at the wood panel in objective examinations. In sensory evaluations, almost all subjective symptom items for eyes and head progressed when subjects looked at white plastic panel or black plastic panel. However, almost no progression of subjective symptoms was noted when subjects looked at the wood panel. In both objective examinations and sensory evaluations, eye fatigue was most highly associated with the black plastic panel, followed by the white plastic panel and, finally, the wood panel.     

Cloning and characterization of pea apyrases: involvement of <Emphasis Type="Italic">PsAPY1 </Emphasis>in response to signal molecules from the pea pathogen <Emphasis Type="Italic">Mycosphaerella pinodes</Emphasis>

 Two nucleoside triphosphatase (NTPase) cDNA clones were isolated from a cDNA library of Pisum sativum L., cv. Midoriusui. The genes encoding the cDNAs were designated PsAPY1 and PsAPY2. PsAPY1 included the N-terminal amino acid sequence of an NTPase bound to pea cell wall. The phylogenic analysis indicated that PsAPY1 belongs to an NTPase subfamily responsive to environmental stimuli and that PsAPY2 belongs to a discrete subfamily, the physiological role of which is almost unknown. The adenosine triphosphatase activity of recombinant PsAPY1 was regulated by an elicitor and a suppressor from the pea pathogen Mycosphaerella pinodes. Based on these findings, we discuss the role of NTPases in response to biological stresses. Received: May 27, 2002 / Accepted: July 31, 2002     

Flagellin from Pseudomonas syringae pv. tabaci induced hrp-independent HR in tomato

We have previously shown that flagellin of Pseudomonas syringae pv. tabaci is an elicitor that induces a hypersensitive reaction (HR) in nonhost tomato cells. Flagellin is the major HR elicitor produced by this pathogen, as shown by the inability of a flagellin-defective mutant, ΔfliC, to induce HR. Also, a ΔfliD mutant that secretes large amounts of monomer flagellins induces a strong HR in tomato. In this study, the possible involvement of an Hrp type III secretion system (TTSS) in flagellin-induced HR was investigated using flagella-defective mutants or Hrp TTSS-defective mutants. The hrcC gene encodes HrcC protein, which is required for Hrp pilus formation in the outer membrane. An hrcC mutation, introduced into the wild-type, ΔfliC, and ΔfliD mutants of P. syringae pv. tabaci did not affect swimming motility or flagellin secretion, whereas all ΔhrcC, ΔfliC, and ΔfliD mutants lost the ability to cause disease on host tobacco leaves. However, the ΔhrcC mutant and the ΔfliD/ΔhrcC double mutant were still able to induce HR cell death, expression of one of the defense-related genes hsr203J, and the generation of hydrogen peroxide in nonhost tomato cells. Thus, flagellin is required for both pathogenicity in host tobacco and HR in nonhost tomato. On the other hand, hrp TTSS is necessary for pathogenicity on host tobacco but is not indispensable to induce HR in nonhost tomato. These results clearly show that flagellin-induced HR is hrp-independent in tomato.The nucleotide sequence data reported are available in the DDBJ/EMBL/GenBank databases under the accession number AB049570     

Defense responses of Arabidopsis thaliana inoculated with Pseudomonas syringae pv. tabaci wild type and defective mutants for flagellin (ΔfliC) and flagellin-glycosylation (Δorf1)

Flagellin, an essential component of the bacterial flagellar filament, is capable of inducing a hypersensitive response (HR), including cell death, in a nonhost plant. A flagellin-defective mutant (ΔfliC) of Pseudomonas syringae pv. tabaci lacks both the flagellar filament and motility, whereas a flagellin-glycosylation-defective mutant (Δorf1) retains the flagellar filament but lacks the glycosyl modification of flagellin protein. To investigate the role of flagellin protein and its glycosylation in the interaction with its nonhost Arabidopsis thaliana, we analyzed plant responses after inoculation with these bacteria. Inoculation with wild-type P. syringae pv. tabaci induced HR, with the generation of reactive oxygen species and cell death. In contrast, inoculation with either ΔfliC or Δorf1 mutant induced a low level of HR, and inoculated leaves developed a disease-like yellowing. These mutant bacteria multiplied better than the wild-type bacteria in A. thaliana. These results indicate that A. thaliana expresses a defense reaction in response to the bacterial flagellin with its glycosyl structure.     

Bacterial DNA activates immunity in <Emphasis Type="Italic">Arabidopsis thaliana</Emphasis>

To initiate defense responses against invasion of pathogenic organisms, animals and plants must recognize microbe-associated molecular patterns (MAMPs). In this study, the elicitor activity of bacterial DNA on the model plant Arabidopsis thaliana was examined. EcoRI-digested plasmid DNA induced defense responses such as generation of reactive oxygen species and deposition of callose, whereas SmaI- and HapII-digested plasmid DNA and EcoRI-digested herring DNA did not remarkably induce these responses. Further, methylation of the CpG sequence of plasmid DNA and Escherichia coli DNA reduced the level of the defense responses. The endocytosis inhibitors wortmannin and amantadine significantly inhibited DNA-induced defense responses. These results suggest that non-methylated CpG DNA, as a MAMP, induced defense responses in Arabidopsis and that non-methylated DNA seems to be translocated into the cytoplasm by endocytosis.     

Comparative study on effective and ineffective nodules of leguminous plants

It is well known that nitrogen fixation is mainly dependent upon the combination between rhizobial strain and host plant species, though it is also affected by the environmental condition. The site of association between the host and rhizobia is the root nodule. It is, therefore, quite natural that special attention has hitherto been focused on it. A typical symbiotic relationship is going on in effective nodule, while a parasitic relationship prevails in ineffective one. Between such types of nodules there must be some differences with respect to the inner structure as well as the metabolism in them. Experiments were made to compare the morphological change of rhizobia, the amount and distribution of starch, the kinds of free amino acids in effective nodule with those in ineffective one. To obtain materials, soybean, pea, red clover, and genge (Astragalus sinicus L.) Were used with their own effective and ineffective strains of rhizobia, respectively.