Multiplex real-time PCR SYBR Green for detection and typing of group III Clostridium botulinum

Clostridium botulinum type C and type D belonging to the group III organisms, are mainly responsible for animal botulism outbreaks. Clinical signs alone are often insufficient to make a diagnosis of botulism and a laboratory confirmation is required. Laboratory confirmation can be performed by demonstrating the presence of botulinum neurotoxins in serum, gastrointestinal contents, liver, wound of sick or dead animals, or by demonstrating the presence of C. botulinum in gastrointestinal contents, liver, and wound. Demonstration of spores in gastrointestinal contents or tissue of animals with clinical signs indicative of botulism reinforces the clinical diagnosis. With the aim of detecting and typing C. botulinum group III organisms, a multiplex real-time PCR SYBR Green was developed and in-house validated. Selectivity, limit of detection, relative accuracy, relative specificity, relative sensitivity, and repeatability of the method were investigated. The multiplex real-time PCR SYBR green used showed a 100% selectivity, 100% relative accuracy, 100% relative specificity, 100% relative sensitivity and a limit of detection of 277 and 580 DNA copies for C. botulinum type C and C. botulinum type D, respectively. The method reported here represents a suitable tool for laboratory diagnosis of type C and D botulism and for testing a large number of samples collected during the animal botulism surveillance and prevention activities.     

Novel Prunus rootstock somaclonal variants with divergent ability to tolerate waterlogging

Plants require access to free water for nutrient uptake, but excess water surrounding the roots can be injurious or even lethal because it blocks the transfer of free oxygen between the soil and the atmosphere. Genetic improvement efforts in this study were focused on the increased tolerance in roots to waterlogging. Among a pool of clones generated in vitro from leaf explants of rootstock Mr.S.2/5 of Prunus cerasifera L., the S.4 clone was flood tolerant whereas the S.1 clone was sensitive. The S.4 clone formed adventitious roots on exposure to flooding. Moreover, the chlorophyll content and mitochondrial activity in the leaf and root, soluble sugar content, alcohol dehydrogenase activity and ethylene content were different between the clones. The sorbitol transporter gene (SOT1) was up-regulated during hypoxia, the alcohol dehydrogenase genes (ADH1 and ADH3) were up-regulated in the leaves and down-regulated in the roots of the S.4 clone during hypoxia, and the 1-aminocyclopropane-1-oxidase gene (ACO1) was up-regulated in the leaves and roots of the S.4 clone during hypoxia and down-regulated in the wild-type roots. In addition, in the S.4 root, hypoxia induced significant down-regulation of a glycosyltransferase-like gene (GTL), which has a yet-undefined role. Although the relevant variation in the S.4 genome has yet to be determined, genetic alteration clearly conferred a flooding-tolerant phenotype. The isolation of novel somaclonals with the same genomic background but with divergent tolerance to flooding may offer new insights in the elucidation of the genetic machinery of resistance to flooding and aid in the selection of new Prunus rootstocks to be used in various adverse environments.     

Effects of fly attack (Bactrocera oleae) on the phenolic profile and selected chemical parameters of olive oil

The phenolic fraction of virgin olive oil influences both its quality and oxidative stability. One of the principal threats of the quality of olive fruit is the olive fly ( Bactrocera oleae) as it alters the chemical composition. The attack of this olive pest has been studied in order to evaluate its influence on the quality of virgin olive oil (free acidity, peroxide value, fatty acid composition, water content, oxidative stability, phenols, and antioxidant power of phenolic fraction). The study was performed using several virgin olive oils obtained from olives with different degrees of fly infestation. They were acquired in different Italian industrial mills from the Abruzzo region. Qualitative and quantitative analyses of phenolic profiles were performed by capillary electrophoresis-diode array detection, and electrochemical evaluation of the antioxidant power of the phenolic fraction was also carried out. These analyses demonstrated that the degree of fly attack was positively correlated with free acidity ( r = 0.77, p < 0.05) and oxidized products ( r = 0.58, p < 0.05), and negatively related to the oxidative stability index ( r = -0.54, p < 0.05) and phenolic content ( r = -0.50, p < 0.05), mainly with secoiridoid compounds. However, it has been confirmed that the phenolic fraction of olive oil depends on several parameters and that a clear correlation does not exist between the percentages of fly attack and phenolic content.     

Cystic liver disease related to high Platynosomum fastosum infection in a domestic cat

Platynosomum fastosum is a small fluke found in the biliary ducts and gallbladder of cats. Its lifecycle includes the snail Sublima octona as intermediate host, and lizards, toads and geckos as paratenic hosts. Affected cats are usually adult and acquire the parasite by feeding on infected lizards. This parasite occurs across the world but is more frequent in tropical areas. The clinical signs range from none to obstruction of the biliary tract, with hepatic failure and death, reinforcing the necessity of including the liver fluke Platynosomum fastosum in the differential diagnosis of hepatic diseases in cats. This report describes an unusual case of a cat with a polycystic hepatic disease and a severe infestation by Platynosomum fastosum and presents a review of the literature.     

Methacarn preserves mucus integrity and improves visualization of amoebae in gills of Atlantic salmon (Salmo salar L.)

Two aqueous fixation methods (modified Davidson's solution and modified Davidson's solution with 2% (w/v) Alcian blue) were compared against two non‐aqueous fixation methods (methacarn solution and methacarn solution with 2% (w/v) Alcian blue) along with the standard buffered formalin fixation method to (a) improve preservation of the mucous coat on Atlantic salmon, Salmo salar L., gills and (b) to examine the interaction between the amoebae and mucus on the gill during an infection with amoebic gill disease. Aqueous fixatives demonstrated excellent cytological preservation but failed to deliver the preservation of the mucus when compared to the non‐aqueous‐based fixatives; qualitative and semi‐quantitative analysis revealed a greater preservation of the gill mucus using the non‐aqueous methacarn solution. A combination of this fixation method and an Alcian blue/Periodic acid–Schiff staining was tested in gills of Atlantic salmon infected with amoebic gill disease; lectin labelling was also used to confirm the mucus preservation in the methacarn‐fixed tissue. Amoebae were observed closely associated with the mucus demonstrating that the techniques employed for preservation of the mucous coat can indeed avoid the loss of potential mucus‐embedded parasites, thus providing a better understanding of the relationship between the mucus and parasite.     

Multidisciplinary study of a Lateglacial-Holocene sedimentary sequence near Bologna (Italy): insights on natural and anthropogenic impacts on the landscape dynamics

Journal of Soils and Sediments - This study investigated a Lateglacial to Holocene sedimentary sequence derived from a small catchment located at San Lazzaro di Savena in the surroundings of...     

Soil organic matter dynamics and losses in manured maize-based forage systems

Soil organic matter (SOM) produces positive effects on multiple soil properties. Increasing its level also provides an opportunity to reduce atmospheric concentration of carbon dioxide (CO₂). Recycling animal manure and returning crop residue are among the main practices to enhance organic carbon (C) stock in arable croplands. This study analysed data of soil organic carbon (SOC) stocks from a medium-term field trial (established in 1992) through a Hénin–Dupuis-based equation to determine the proportion of different organic materials retained in the soil as SOM.The treatments included in the experiment simulated different typical management techniques and implied the application to the soil of various organic materials, i.e. cattle slurry, cattle farmyard manure, maize straw, root, maize and ley stubble. Soil organic carbon and total nitrogen (N) (Kjeldhal N) were measured in the 0–30 cm layer during spring 1999 and spring 2003. The relationship between SOC and N stocks measured in 1999 and 2003 and the annual additions of C and N was described by the Hénin–Dupuis-based equation, assuming the presence of two C and N pools (stable and fresh).Our results showed that the application of farmyard manure, slurry and maize straw induced higher C and N content compared to the application of urea without return of crop residue. The different levels of urea application did not produce any significant difference in C and N soil content.We found that approximately 2% of SOC is lost to the atmosphere annually. Furthermore, the amount of C and N retained in the soil each year varied by organic materials: 46% C and 44% N with farmyard manure, 26% C and 11% N with slurry, 28% C and 10% N with root and 6% C with maize straw and ley stubble.These results were used to inform an estimate of C lost or removed to/from atmosphere following manure application and crop residue return for the compilation of greenhouse gas inventories. We found that the most C conservative management is the production and spreading of farmyard manure and that increasing amounts of mineral N fertilizer did not affect the C sequestration extent.     

Comparison on accuracy of different nonlinear models in predicting growth of Podolica bulls

Animal growth does not follow a linear pattern, being explained mathematically by functions that have parameters with biological meaning. These parameters are used to estimate the expected weight of animals at specific ages. Several nonlinear models have been used to describe growth. This study was carried out to estimate the parameters of logistic, Gompertz, Richards and von Bertalanffy growth curve models in a sample of Podolica young bulls to determine the goodness of fit. Animals were weighed every 3 months from birth to 810 days of age. The results indicate that all the growth models used were easily fitted to the observed data with Gompertz and logistic functions presenting less computational difficulty in terms of number of iterations to achieve convergence. Moreover, logistic and Richards equations provided the best overall fit being useful to describe the growth of Podolica bulls. Considering that the literature lacks information on growth curves in Podolica breed, the study of a mathematical model for growth describing the developmental pattern of a specific population within a peculiar environment is a useful tool to improve Podolica breed production.     

Chitin and Chitosan as Multipurpose Natural Polymers for Groundwater Arsenic Removal and As2O3 Delivery in Tumor Therapy

Chitin and chitosan are natural polysaccharide polymers. These polymers have been used in several agricultural, food protection and nutraceutical applications. Moreover, chitin and chitosan have been also used in biomedical and biotechnological applications as drug delivery systems or in pharmaceutical formulations. So far, there are only few studies dealing with arsenic (As) removal from groundwater using chitin or chitosan and no evidence of the use of these natural polymers for arsenic trioxide (As₂O₃) delivery in tumor therapy. Here we suggest that chitin and/or chitosan might have the right properties to be employed as efficient polymers for such applications. Besides, nanotechnology offers suitable tools for the fabrication of novel nanostructured materials of natural origin. Since different nanostructured materials have already been employed successfully in various multidisciplinary fields, we expect that the integration of nanotechnology and natural polymer chemistry will further lead to innovative applications for environment and medicine.