First Report of Bacterial Bark Canker of Walnut Caused by Brenneria nigrifluens in Hungary.

During August 2012, vertical oozing cankers were sporadically observed on trunks and branches of walnut trees (Juglans regia) in the city of Zánka, near Lake Balaton and other parts of Hungary including Budapest, Gyor, and Tatabánya cities. Cankers were observed on trunks and branches where brownish-black exudates staining the bark appeared mainly in the summer. Isolations were performed primarily from exudates but also from infected tissues using King's medium B (KB) (3) and EMB medium (2). Colonies similar in appearance to Brenneria nigrifluens (syn.: Erwinia nigrifluens) (1,5) were isolated. The bacterium, first reported in California, was also recorded in Iran, Spain, France, and several Italian locations, on walnut trees. The bacterial strain was gram negative and did not induce a hypersensitive response on tobacco (Nicotiana tabacum L. 'White Burley') leaves. The bacterium grew at 26°C. Colonies on KB were white and non-fluorescent, but on EMB medium were a typical dark purple with metallic green sheen. The results of substrate utilization profiling using the API 20E kit (Biomérieux, Marcy l'Etoile, France) showed that the bacterium belonged to the Enterobacteriaceae. The strain was positive for citrate utilization, H2S, and acetoin production and urease, glucose, inositol, saccharose, and arabinose reactions. Pathogenicity was tested by injecting five young healthy walnut branches on two separate 2-year-old grafted potted plants with a bacterial suspension containing 107 CFU/ml. Negative controls were walnut branches injected with sterile distilled water. Branches were enclosed in plastic bags and incubated in a greenhouse under 80% shade at 26°C day and 17°C night temperatures. Three months after inoculation, necrotic lesions were observed in the inner bark and dark lines were observed in internal wood, but no external cankers were observed on inoculated branches. The negative control appeared normal. B. nigrifluens was re-isolated from lesions on inoculated branches and identified as described above; thus, Koch's postulates were fulfilled. For molecular identification of the pathogen, 16S rDNA amplification was performed using genomic DNA from strain Bn-WalnutZa-Hun1 with a universal bacterial primer set (63f and 1389r) (4). The PCR products were cloned into a pGEM T-Easy vector (Promega, Madison, WI) and transformed into Escherichia coli DH5α cells. A recombinant plasmid (2A2.5) was sequenced using M13 forward and reverse primers. The sequence was deposited in NCBI GenBank (Accession No. HF936707) and showed 99% sequence identity with a number of B. nigrifluens strains, including type strains Z96095.1, AJ233415.1, JX484740.1, JX484739.1, JX484738.1, and FJ611884.1. On the basis of the symptoms, colony morphology, biochemical tests, and 16S rDNA sequence identity, the pathogen was identified as Brenneria nigrifluens. To our knowledge, this is the first report of a natural outbreak of bacterial bark canker on walnut in Hungary and the presence of the pathogen may seriously influence in local orchards and garden production in the future. References: (1) L. Hauben et al. Appl Microbiol 21:384, 1998. (2) J. E. Holt-Harris and O. Teague. J. Infect. Dis. 18:596, 1916. (3) E. O. King et al. J. Lab. Clin. Med. 44:301, 1954. (4) A. M. Osborn et al. Environ. Microbiol. 2:39, 2000. (5) E. E. Wilson et al. Phytopathology 47:669, 1957.

Comparative animal study on hard tissue integration and bone formation of different Nobel Biocare implants.

Dental implantation aims at optimal and long-term hard tissue integration. Beside primary stability, loading time and other factors, e.g. the surface of the endosteal part of the implant, is a matter of special importance. In this animal trial, hard tissue integration of two different implant types was studied using radiological, histological and histomorphometric analysis. Two different implants with an oxidized surface (TiUnite; Nobel Biocare AB, Goteborg, Sweden, NobelReplace Tapered Groovy 4.3 x 10 mm and Replace Select Tapered 4.3 x 10 mm) were inserted into the right and left mandibles of 10 German domestic pigs between canine and premolar and immediately provided with a ceramic crown. The primary implant stability was determined using resonance frequency analysis. After 70 days, the test animals were killed and specimens were collected for histological and histomorphometric examination. All implants showed good primary stability after surgery. Histological and histomorphometrical analysis revealed no significant differences in the bone apposition. The immediate loading of the different implant types don't have any negative effects on the bone apposition in the period of 70 days. The long-term effects of immediate loading of these types of implant requires further studies.

Evaluation of shape and size changes of bone and remodelled bone substitute after different fixation methods.

Suitable tissue fixation is indispensable to histological analysis. This investigation, therefore, sought to evaluate changes of shape and size of bone specimens and remodelled bone substitute material following different fixation methods. Mandibular bones of 9 pigs (Sus scrofa domesticus) served as specimens. Two mandibular premolars were extracted respectively and the extraction alveoli were filled with synthetic bone substitute material. The samples were collected after 70 days. Fixation of 6 specimens respectively was done for 7 days in 4% formalin (formaldehyde), 70% ethanol and glycerol at 18 degrees C room temperature. The samples were radiographically examined before and after fixation using a reference specimen and subsequently underwent histological analysis. After fixation in formalin, the samples showed no size changes. After fixation in glycerol, morphological analysis revealed minor shape changes. Fixation in ethanol causes shrinking of the tissue specimens. Histological inspection of the tissues shows no morphological changes except slight shrinking. In conclusion there is no universal fixative that could met all requirements and permited proper examination without affecting tissues or bone specimens.

Different effects of naproxen on the organ blood flows in normo- and hypervolemic anaesthetized rats.

The effects of naproxen, an inhibitor of the enzyme cyclooxigenase (10 mg/kg i.v.) on the distribution of the cardiac output (CO) and on the intrarenal hemodynamics were investigated in normovolemic (free salt and water uptake till the beginning of experiment) and hypervolemic (with i.v. infusion of 50 ml 0.9% NaCl solution/kg/10 min) narcotized rats. The cardiac output was measured on the basis of he Stewart-Hamilton principle, the blood flow of the organs by the Sapirstein method. 86Rb was used as indicator. In hypervolemia, the blood pressure is the same, the cardiac output is higher (CO-normovolemia: 23.1 +/- 7.04, CO-hypervolemia: 29.0 +/- 6.43 ml/min/100 g; p < 0.05) the total peripheral resistance (TPR) is lower (TPR-normovolemia: 40.0 +/- 9.39 R, TPR-hypervolemia: 31.2 +/- 8.34 R, p < 0.05) than in normovolemic animals. In hypervolemia the vascular resistance of the investigated organs (heart, lungs, kidney, skin, muscle, liver, spleen, intestine) is also lower and the intrarenal blood flow shifts toward the medulla. One hour following the naproxen administration a) in normovolemia joining to a slightly decreased cardiac output and increased TPR, the vascular resistance of the skin (R-control: 85.1 +/- 32.7, R-naproxen: 161 +/- 57.4; p < 0.001) and of the skeletal muscle (R-control: 114 +/- 35.1, R-naproxen: 190 +/- 81.9; p < 0.01) increases. The blood flow of the other organs and the intrarenal hemodynamics does not change under the effect of naproxen. b) in hypervolemia the general circulatory parameters (blood pressure, cardiac output, TPR) and the parameters of the organ circulation and intrarenal hemodynamics remain unchanged. The results suggest that in rats the prostanoid compounds (PGE2, PGI2, TXA2) a) can modify the blood flow of the skin and muscle in normovolemic animals, but they do not have any role in determining the blood flow of the other organs or the intrarenal distribution of blood flow. b) in hypervolemia they play no role in determining organ-, or intrarenal blood flow. The consequences of cyclooxygenaze enzyme inhibition--at least in the case of the organ blood flow--depend on the magnitude of sodium and water load in the organism.

Dibromodulcitol plus bleomycin compared with bleomycin alone in head and neck cancer.

Advanced recurrent squamous cell head and neck cancer patients were prospectively randomized to receive or not receive dibromodulcitol 10 mg/kg PO weekly for 8 consecutive weeks in addition to bleomycin chemotherapy. Patients initially entered in the study received bleomycin 15 mu/m2 three times weekly for 8 weeks. This was later changed to 15 mu/m2 twice weekly for 8 weeks because of unacceptable stomatitis. Most patients had relapsed following surgery and/or radiotherapy, but none had received prior chemotherapy. A2 : 1 randomization in favor of the dibromodulcitol-containing therapy was used. There were 12 partial responses in the 44 evaluable patients receiving the combination (27%), and 4 partial responses in the 18 patients receiving single-agent bleomycin chemotherapy (22%). This difference was not statistically significant. Response durations were also relatively short for both therapies. Within the limitations of this study, we were unable to demonstrate that patient benefit resulted from the addition of dibromodulcitol to bleomycin chemotherapy for this patient population.