A retrospective study on bone metastasis in dogs with advanced-stage solid cancer.

OBJECTIVES: To review clinical characteristics, treatment, outcome and prognostic factors in dogs with solid cancer-bearing bone metastases. MATERIALS AND METHODS: Records were reviewed from dogs with histologically-proven solid cancer and bone metastases. Clinicopathologic variables, bone metastases characteristics and skeletal-related events were recorded. Endpoints were time to bone metastases and survival. RESULTS: Fifty dogs were included, 20 of them with synchronous and 30 of them with metachronous bone metastases. In the latter group, median time to diagnosis of bone metastases was 210 days (range, 30 to 1835). Most common primary cancer locations included mammary gland (n=6), spleen (n=5) and tonsil (n=5). Most common histotypes were carcinoma (n=32) and hemangiosarcoma (n=10). Nineteen dogs had multiple bones involvement, with humeri and vertebrae more commonly affected. Twenty-four dogs received antitumoural therapy, five symptomatic treatment and 21 were not treated. Overall median survival after bone metastases diagnosis was 30 days (range, 11 to 49); 83% of dogs died because of skeletal-related events. Lack of antitumoural therapy was significantly associated with shorter survival (hazard ratio: 2.7; 95% confidence interval: 1.3 to 5.6) and with increased risk of skeletal-related death (hazard ratio: 3.3; 95% confidence interval: 1.4 to 7.4). Dogs with endocrine/neuroendocrine tumours (odds ratio: 8.8; 95% confidence interval: 1.2 to 63.9), without appendicular metastases (odds ratio: 5.1; 95% confidence interval: 1.0 to 25.8), without extra-skeletal metastases (odds ratio: 5.2; 95% confidence interval: 1.1 to 24.5) and receiving antitumoural therapy (odds ratio: 14.8; 95% confidence interval: 1.7 to 131.4) had an increased chance of surviving more than 100 days. CLINICAL SIGNIFICANCE: Bone metastases in dogs with solid cancers are associated with poor prognosis and a high risk of skeletal-related events. Treatment appears to have an impact on survival.

Endodontic retreatment and bioceramics: A case report.

The success rate in endodontic retreatment can be significantly boosted by using materials that guarantee both intimate adhesion to the canal walls and long-term stability. Bioceramic cements used in root filling show interesting properties including extraordinary sealing capacities, antimicrobial activity that stimulates periapical healing and the continuous production of hydroxyopathitis for a long time. This case report deals with the application of bioceramic cement in endodontic retreatment. The Bioroot RCS (Septodont), thanks to its ability to firmly adhere both to the gutta-percha and to the walls of the canal and to definitively seal the apical third, can allow to obtain extraordinary healing in a relatively short time by exploiting its antimicrobial abilities. However, further studies are needed to assess the effectiveness of this technique over the long term.

Seasonal changes on microbial metabolism and biomass in the euphotic layer of Sicilian Channel.

As a part of a wider project on fisheries ecology, several biological and environmental parameters were monitored during two oceanographic cruises (BANSIC 2012 and NOVESAR 2013) in the Sicily Channel, which connects the Western and Eastern Mediterranean basins. The prokaryotic abundances and biomass as well as hydrolysis rates on organic matter were investigated in the euphotic layer of a retention area for fish larval stages including anchovy (Engraulis encrasicolus, Linnaeus, 1758) with the aim to investigate the different biogeochemical signatures in two seasonal conditions. The environmental parameters, particulate organic carbon and nitrogen together with heterotrophic production were also measured. Results showed significant increases for most of the studied parameters with increasing temperature during summer. This had effects on the Carbon cycle and recycling of nutrients; in fact total prokaryotic abundance and biomass, as well as carbon hydrolyzed by two enzymes (Leucine aminopeptidase and ß-glucosidase), increased significantly during summer. Conversely Alkaline phosphatase activity, Chlorophyll concentration and Oxygen increased during winter. The same environmental parameters affected also the presence of fish eggs. Moreover high percentages of free enzymes (i.e., enzymes not associated with cells) were measured, accounting for percentages variable from 12 to 95 % of the total enzymatic activity, with values generally higher in summer than in winter. In this oligotrophic environment, the prokaryotic biomass was supported by the C hydrolyzed by enzymatic activities. The ratio between the hydrolyzed C and prokaryotic biomass was higher in winter than in summer, indicating that alkaline phosphatase activity contribute to an efficient incorporation of C into biomass in winter.

Seasonal dynamics of prokaryotic abundance and activities in relation to environmental parameters in a transitional aquatic ecosystem (Cape Peloro, Italy).

This study examines the effects of temporal changes on microbial parameters in a brackish aquatic ecosystem. To this aim, the abundances of prokaryotes and vibrios together with the rates of enzymatic hydrolysis of proteins by leucine aminopeptidase (LAP), polysaccharides by ß-glucosidase (GLU) and organic phosphates by alkaline phosphatase (AP), heterotrophic prokaryotic production (HPP), respiration (R), were seasonally investigated, during a 2-year period in the coastal area of Cape Peloro (Messina, Italy), constituted by two brackish lakes (Faro and Ganzirri). In addition, physical and chemical parameters (temperature, salinity, nutrients) and particulate organic carbon and nitrogen (POC, PN) were measured. The influence of multiple factors on prokaryotic abundances and activities was analysed. The results showed that Cape Peloro area is characterised by high seasonal variability of the microbial parameters that is higher than the spatial one. Combined changes in particulate matter and temperature (T), could explain the variability in vibrios abundance, GLU and R activities in both lakes, indicating a direct stimulation of the warm season on the heterotrophic prokaryotic metabolism. Positive correlations between T (from 13.3 to 29.6 °C) and HPP, LAP, AP, POC, PN are also observed in Ganzirri Lake. Moreover, the trophic status index and most of the microbial parameters show significant seasonal differences. This study demonstrates that vibrios abundance and microbial activities are responsive to the spatial and seasonal changes of examined area. The combined effects of temperature and trophic conditions on the microbial parameters lead us to suggest their use as potential indicators of the prokaryotic response to climate changes in temperate brackish areas.

Enzymatic activities and prokaryotic abundance in relation to organic matter along a West-East Mediterranean transect (TRANSMED cruise).

The distribution of extracellular enzymatic activities (EEA) [leucine aminopeptidase (LAP), ß-glucosidase (GLU), alkaline phosphatase (AP)], as well as that of prokaryotic abundance (PA) and biomass (PB), dissolved organic carbon (DOC), particulate organic carbon and particulate total nitrogen (POC, PTN), was determined in the epi-, meso-, and bathypelagic waters of the Mediterranean Sea along a West-East transect and at one Atlantic station located outside the Strait of Gibraltar. This study represents a synoptical evaluation of the microbial metabolism during early summer. Decreasing trends with depth were observed for most of the parameters (PA, PB, AP, DOC, POC, PTN). Significant differences between the western and eastern basins of the Mediterranean Sea were found, displaying higher rates of LAP and GLU and lower C/N ratios more in the eastern than in the western areas. Conversely, in the epipelagic layer, PA and PB were found to be higher in the western than in the eastern basins. PB was significantly related to DOC concentration (all data, n = 145, r = 0.53, P < 0.01), while significant correlations of EEA with POC and PTN were found in the epipelagic layer, indicating an active response of microbial metabolism to organic substrates. Specific enzyme activities normalized to cell abundance pointed out high values of LAP and GLU in the bathypelagic layer, especially in the eastern basin, while cell-specific AP was high in the epi- and bathypelagic zone of the eastern basin indicating a rapid regeneration of inorganic P for both prokaryotes and phytoplankton needs. Low activity and abundance characterized the Atlantic station, while opposite trends of these parameters were observed along the Mediterranean transect, showing the uncoupling between abundance and activity data. In the east Mediterranean Sea, decomposition processes increased probably in response to mesoscale structures which lead to organic matter downwelling.

Characterization of chitinolytic bacteria and histological aspects of Shell Disease Syndrome in European spiny lobsters (Palinurus elephas) (Fabricius 1787).

The present research reports the first description of Shell Disease Syndrome in European spiny lobsters Palinurus elephas (Fabricius 1787), which occurred in an experimental aquaculture facility in Sicily (Italy). Both bacterial characterization and histopathological examination of the exoskeleton at site of lesions was carried out. Infected specimens showed tail fan erosions, and in one case uropod ulceration and complete loss of periods. Identified species included: Listonella anguillarum 50.5%, Vibrio parahaemolyticus 27.5% and Vibrio alginolyticus 22%. Microscopic evaluation of lesions indicate the presence of inflammatory responses, which include melanization and pseudomembrane formation, similar to those described for other crustaceans affected by SDS.

Development of a multiplex PCR assay for Photobacterium damselae subsp. piscicida identification in fish samples.

A multiplex polymerase chain reaction protocol for the detection of Photobacterium damselae and subspecies piscicida and damselae discrimination, with internal amplification control, was developed. Assay specificity was assessed by testing 19 target and 25 non-target pure cultures. The detection limit was 500 fg, corresponding to 100 genome equivalents. The optimized protocol was also prevalidated with spleen, kidney and blood samples from infected and uninfected sea bass, without any culture step, and it can be proposed as a valid alternative to culture standard methods for the rapid and specific diagnosis of photobacteriosis in fish.

Evaluation of different DNA-based fingerprinting methods for typing Photobacterium damselae ssp. piscicida.

This study evaluates the effectiveness of three different molecular techniques, repetitive extragenic palindromic PCR (REP-PCR), enterobacterial repetitive intergenic consensus sequence PCR (ERIC-PCR) and the random amplified polymorphic DNA (RAPD-PCR) for rapid typing of Photobacterium damselae ssp. piscicida strains isolated from different species of marine fish and geographic areas. The results obtained by the three methods showed that RAPD and ERIC-PCR were more discriminative for suitable rapid typing of Ph. damselae ssp. piscicida than REP-PCR. The analysis of DNA banding patterns generated by both molecular methods (RAPD and ERIC-PCR) clearly separated the strains into two main groups that strongly correlated with their geographic origin. Moreover, the REP-PCR analysis was less reproducible than the RAPD and ERIC-PCR methods and does not allow the establishment of genetic groups. RAPD and ERIC-PCR constitute valuable tools for molecular typing of Ph. damselae ssp. piscicida strains, which can be used in epidemiological studies of photobacteriosis infections.

Evaluation of different DNA-based fingerprinting methods for typing Photobacterium damselae ssp. piscicida

This study evaluates the effectiveness of three different molecular techniques, repetitive extragenic palindromic PCR (REP-PCR), enterobacterial repetitive intergenic consensus sequence PCR (ERIC-PCR) and the random amplified polymorphic DNA (RAPD-PCR) for rapid typing of Photobacterium damselae ssp. piscicida strains isolated from different species of marine fish and geographic areas. The results obtained by the three methods showed that RAPD and ERIC-PCR were more discriminative for suitable rapid typing of Ph. damselae ssp. piscicida than REP-PCR. The analysis of DNA banding patterns generated by both molecular methods (RAPD and ERIC-PCR) clearly separated the strains into two main groups that strongly correlated with their geographic origin. Moreover, the REP-PCR analysis was less reproducible than the RAPD and ERIC-PCR methods and does not allow the establishment of genetic groups. RAPD and ERIC-PCR constitute valuable tools for molecular typing of Ph. damselae ssp. piscicida strains, which can be used in epidemiological studies of photobacteriosis infections.

Heterotrophic bacteria in the northern Adriatic Sea: seasonal changes and ectoenzyme profile.

A seasonal study of the quantitative and qualitative distribution of heterotrophic bacterial community was carried out in the Adriatic Sea between April 1995 and January 1996, in order to evaluate its spatial and temporal variability and metabolic potential in the degradation processes of organic matter. The culturable bacteria (CFU) ranged between 0.1 and 22% of total bacterioplankton with a maximum percentage in surface samples of coastal zones. Their distribution was generally affected by the prevailing hydrological conditions. At the coastal stations about 44-75% of CFU variance could be explained by river runoff. The changes in the composition of heterotrophic bacterial community showed a seasonal succession of main bacterial groups, with a prevalence of Gram negative, non fermenting bacteria in the cold period (April-January) and an increase of Vibrionaccae and pigmented bacteria in summer. The seasonal variations were more important at the stations influenced by rivers than offshore. The bacterial community showed a greater versatility for organic polymers hydrolysis in the offshore station than in the coastal areas. Over 60% of all isolated heterotrophic bacteria expressed peptidase, lipase and phosphatase ectoenzymes activities, in all seasons and showed an increasing trend in warm period (in July October). The alpha- and beta-glucosidase potentials of bacteria were lower (20% on average) and showed different pattern during the year. These results suggest different role of the bacterial community in the decomposition of organic matter in the Adriatic Sea. Since only 20% of bacterial strains expressed glucosidase activity, carbohydrate-rich polymers such as mucilage might accumulate.

Estimates of leucine aminopeptidase activity in different marine and brackish environments.

AIM: Leucine aminopeptidase (LAP), an enzyme involved in the decomposition of natural peptides, was measured in different marine and brackish ecosystems, together with some environmental and microbiological parameters. METHODS AND RESULTS: The fluorogenic compound L-leucine-7-amido-4-methyl coumarin was specifically used for the determination of this in situ activity. The enzyme data obtained from this comparative study highlighted the strong spatial and temporal variability of the distribution of LAP in aquatic ecosystems, which was sometimes related to the course of environmental variables such as salinity and organic carbon content. CONCLUSIONS: LAP assay has proved to be a rapid method providing useful information on the microbial metabolic processes involved in the mineralization of organic matter. SIGNIFICANCE AND IMPACT OF THE STUDY: The determination of the potential rates of extracellular enzyme activity is of great ecological importance to extend knowledge on the role played by bacteria in aquatic biogeochemical cycles.

Use of the indirect immunofluorescence method for detection and enumeration of Escherichia coli in seawater samples.

The determination of Escherichia coli in marine waters through a rapid method, the microscopic indirect immunofluorescent technique, is evaluated in comparison with the conventional count on m-FC agar medium. The data obtained in seawater samples, collected monthly along the Messina coastline, show good sensitivity of the analysis and agreement between the microscopic and culture technique, with a detection limit of 10(2) cells 100 ml(-1) for immunofluorescence.

Bacterial pollution of Messina coastal waters: a one year study.

A year's monitoring of faecal pollution of marine coastal waters surrounding Messina was carried out in 1996/97. The distribution of faecal coliforms was evaluated in 15 stations located along the Sicilian coastline, sampled monthly in coincidence of the two opposing current phases ("montante" and "scendente" currents) which characterise the Straits of Messina. The data obtained provided a complete picture of hygienic-sanitary conditions of the area and highlighted the presence of heavily polluted sites in correspondence with river outflows. Higher bacterial counts were associated with lower salinity values and higher ammonia concentrations; over an annual study, they occurred during the coldest months, showing the negative impact of continental water inputs on the bacteriological quality of coastal waters.

Distribution of Synechococcus sp. and Synechococcus bacillaris in the waters of the Straits of Magellan (April 1995-early austral autumn).

During the last oceanographic cruise carried out in the Straits of Magellan (April 1995), a serological approach was used in order to determine the distribution and composition of the picophytoplankton community with respect to two cyanobacteria species, Synechococcus sp. and bacillaris, characterized respectively by phycoerythrin and phycocyanin as the main accessory photosynthetic pigment. In the period examined, the Straits were characterized by generally low concentrations of total picophytoplankton (10(5)-10(6) cells/l). The qualitative composition of the community showed the prevalence of the species Synechococcus sp. in the Pacific basin, whereas S. bacillaris appears to be predominant in the central area. The immunofluorescence method proved to be effective in the study of the diversity of these microorganisms in aquatic environments.

Microbiological monitoring of Castellammare Gulf (TP) waters for their suitability in marine aquaculture.

A three year investigation into the microbiological conditions of the waters of the Gulf of Castellammare (TP) was carried out in order to evaluate the possible utilization of this area as an aquaculture site. Analysis of a total of 168 samples was aimed at estimating the quantitative distribution of total and fecal coliforms, enterococci and halophilic vibrios. The qualitative distribution of Vibrio species was also studied with particular reference to potentially pathogenic species. Coliforms showed the highest densities (10(2)-10(3) CFU/100 ml) in November 1993 at the coastal stations and mostly negative values during the subsequent samplings; in contrast, halophilic vibrios prevailed in the warm months. V. alginolyticus and V. fluvialis prevailed respectively among the Vibrio population grown at 35 and 24 degrees C, grouped in 13 and in 4 clusters at similarity levels of 81-96%. Quantitative data demonstrate the presence of low levels of microbial contamination, which suggests the general suitability of the area for fish farming.

Detection of Nitrosococcus oceanus in a Mediterranean lagoon by immunofluorescence.

The autotrophic ammonium-oxidizing bacterium Nitrosococcus oceanus was studied in the brackish lake of Ganzirri by cultural and immunofluorescent methods. The preparation of the specific antiserum in rabbits is reported. The polyclonal antiserum for Nitrosococcus oceanus cross-reacted with other ammonia oxidizing strains, but not with other marine bacteria. Temporal changes were determined by taking monthly water samples from a station 6 m deep and the distribution in sediments was investigated in four stations. Isolation of a strain of Nitrosococcus sp. was obtained from a sediment sample collected in December. The abundance of Nitrosococcus spp. bacteria correlated positively with particulate organic carbon (POC), particulate organic nitrogen (PON), temperature and total bacteria, whereas there was a negative relationship with oxygen tension. No correlation was found between immunofluorescent and MPN counts of Nitrosococcus spp. bacteria.

Distribution and numerical taxonomy of Vibrionaceae in the waters of the Straits of Messina.

The results of a study carried out by numerical analysis on Vibrio strains isolated from the waters of the Straits of Messina are reported. The quantitative data showed the presence of low bacterial densities (ranging from 9 to 99 CFU/100 ml of water) due to the intense currents which characterize this area; also the highest bacterial counts were generally found during the "montante" current. With regard to the qualitative results, there was a predominance of vibrios belonging to V. mediterranei, V. splendidus II and V. pelagius II species, which represented respectively 25%, 19% and 13% of the total bacterial population. The species distribution did not seem related to sampling stations or depth. The taxonomic structure, obtained using the simple matching coefficient and unweighted average linkage clustering, revealed the presence of 7 main clusters (S = 80-95%), which included strains of various origin because of the particular turbulence of the waters of the Straits.