Towards a rapid molecular identification of the common phlebotomine sand flies in the Mediterranean region     

Latrofa MS  Annoscia G  Dantas-Torres F  Traversa D  Otranto D 《Veterinary parasitology》2012,184(2-4):267-270

The present study reports two simple molecular approaches allowing a rapid identification of the most prevalent species of phlebotomine sand flies in the Mediterranean region. A PCR protocol for the amplification of ITS2 ribosomal region and a PCR-RFLP on a mitochondrial DNA fragment (cytb-nd1) were settled in order to identify and discriminate among Phlebotomus perniciosus, Phlebotomus neglectus, Phlebotomus perfiliewi, Phlebotomus papatasi and Sergentomyia minuta. The ITS2 regions showed a certain degree of interspecific variability, which led to PCR amplicons of different sizes, i.e., 450, 490, 460, 480 and 530 bp for P. perniciosus, P. neglectus, P. perfiliewi, P. papatasi, and S. minuta, respectively. Analogously, the digestion of a mitochondrial DNA amplicon with Ase I enzyme showed five different restriction profiles, which allowed the unequivocal differentiation of the sand fly species examined. These methods might represent useful tools for a molecular large scale screening of phlebotomine sand fly species caught in areas where leishmaniasis is endemic, in order to plan appropriate epidemiological surveillance programs for both Leishmania spp. and their vectors.  相似文献   

A duplex real-time polymerase chain reaction assay for the detection of and differentiation between Dirofilaria immitis and Dirofilaria repens in dogs and mosquitoes     

Latrofa MS  Dantas-Torres F  Annoscia G  Genchi M  Traversa D  Otranto D 《Veterinary parasitology》2012,185(2-4):181-185

This study describes a duplex real-time polymerase chain reaction (PCR) assay for the detection and differentiation between Dirofilaria immitis and Dirofilaria repens in dog blood and mosquitoes. Regions of a cytochrome oxidase 1 (cox1) mitochondrial DNA fragment and the second internal transcribed spacer (ITS-2) of nuclear ribosomal DNA were amplified from microfilariae and adult worm samples, using a sensitive SsoFast? EvaGreen(?) based real-time PCR method coupled with melting-curve analysis. The limit of the real-time PCR in detecting microfilaria and adult worm DNA was also tested both in dog blood and in artificially infected microfilarial. Two peaks at different melting temperatures (T(m)) for D. immitis (mean ± SD=75.7 ± 0.3°C) and D. repens (mean ± SD=70 ± 0.7°C), respectively, were obtained for microfilarial and adult positive controls of both species when examined separately and together. The real-time PCR protocol was also efficient in detecting microfilarial and adult DNA of both species when tested in samples spiked with DNA from Aedes albopictus, in Aedes aegypti experimentally infected by D. repens and in Culex pipiens naturally infected by D. repens and D. immitis. The high sensitivity of real-time PCR confirmed its reliability in detecting small amounts of genomic DNA either in dog blood or mosquitoes (2.5 pg/μl and 3 × 10(-1)pg/μl for D. immitis and D. repens, respectively). This assay is proposed as a tool for the epidemiological surveillance of the two most important Dirofilaria species in areas where they are endemic and sympatric.  相似文献   

Towards assessing the sustainability of European logging operations     

Staffan Berg  Janine Fischbach  Franka Brüchert  Mikael Poissonnet  Stefania Pizzirani  Anne Varet  Udo H. Sauter 《European Journal of Forest Research》2012,131(1):81-94

The forest-based sector has been at the forefront in operationally implementing the sustainability concept, its associated principles and indicators for sustainable forest management. Several methods have been developed to study environmental impacts of forestry activities, but none of the existing tools address all the dimensions of sustainability along the whole forest wood chain (FWC) in a balanced way. Consequently, the decision was made to develop a tool for sustainability impact assessment (ToSIA), the modelling framework for sustainability impact assessment of FWCs. The objective of the EU Project Eforwood was to develop ToSIA, a decision support tool. Within ToSIA, a FWC is modelled as a number of interconnected processes. For each process, a range of economic, environmental and social indicators and their respective values are calculated, thus representing the three pillars of sustainability. By this method, the multifunctionality of forests can be assessed and supply chains can be compared with respect to sustainability. Sensitivity analysis and scenario techniques can be applied to learn about the effect of expected changes to the structure of the chain, the material flows and the indicator values. In order to provide the tool with information about forest and logging operations, data were collected at two fundamental levels: (1) a regional level with case studies in Scandinavia, Iberia and Baden-Württemberg and (2) a European level with a case study that reflects conditions in the 27 countries of the European Union. This paper describes and details the harvesting and logging processes for the European countries. The results are displayed for each of the three regional case studies as well as aggregated to five principal areas in Europe: Eastern, Northern, Western, Central and Southwest Europe.  相似文献   

Detection of Mycobacterium avium subsp. paratuberculosis by buoyant density centrifugation, sequence capture PCR and dot blot hybridisation     

Halldórsdóttir S  Englund S  Nilsen SF  Olsaker I 《Veterinary microbiology》2002,87(4):327-340

Detection of Mycobacterium avium subsp. paratuberculosis (M. paratuberculosis) by polymerase chain reaction (PCR) is often hampered by the lack of efficient methods for sample treatment. We report a protocol for analysis of faecal samples based on buoyant density centrifugation in Percoll and IS900 sequence capture PCR combined with a dot blot assay for detection of low-grade infection of M. paratuberculosis. Serial dilutions of M. paratuberculosis genomic DNA and M. paratuberculosis bacteria were used to assess the sensitivity of the method. The final evaluation was performed with spiked faecal samples, which also were analysed by culture. The presence of PCR inhibitory substances in processed faecal samples was evaluated by including a PCR internal control. By using buoyant density centrifugation, sequence capture PCR, and dot blot hybridisation, we achieved a sensitivity of 10(3)CFU (colony forming units)/g of faeces. The detection limit by culture was assessed to 10(2)CFU/g of faeces. We conclude that the described protocol is a fast and sensitive alternative to bacterial culture of faecal samples.  相似文献   

Genetic analysis and phenotypic associations for drought tolerance in Hordeum spontaneum introgression lines using SSR and SNP markers     

Berhane Lakew  Robert J. Henry  Salvatore Ceccarelli  Stefania Grando  Jason Eglinton  Michael Baum 《Euphytica》2013,189(1):9-29

Associations between markers and drought related traits were investigated on a set of 57 advanced barley breeding lines, carrying various levels of introgression from Hordeum spontaneum lines 41-1 and 41-5, the best sources of drought tolerance in the ICARDA barley breeding program, using 74 simple sequences repeats (SSR) and 20 single nucleotide polymorphism markers. The 57 lines were evaluated for grain yield and drought related traits for three years (2003/04, 2004/05, 2005/06) in nine Mediterranean low rainfall environments. A high level of polymorphism was found with SSR markers, and the mean polymorphism information content and gene diversity values were 0.67 and 0.71, respectively. The number of alleles per locus varied from 2 to 11, with an average of 5.8 alleles per marker. Considering all the 57 lines, the linkage disequilibrium (LD) analysis was significant at a comparison-wise P?<?0.01 level in nearly 9 % of the SSR marker pairs used and a decay of LD was observed to a value of r ²?<?0.2 at a genetic distance of 40?cM. The association analysis revealed a total of 147 significant marker?Ctrait associations for grain yield and drought related traits. A total of 72 (49 %) marker?Ctrait associations showed favorable effects of the exotic germplasm where the H. spontaneum lines contributed to an improvement of the trait under drought stress conditions. The number of significant marker?Ctrait associations per trait were: 12 for growth habit; 2 for growth vigor; 11 for peduncle extrusion; 5 for number of grains per spike; 20 for peduncle length; 16 for days to heading; 20 for plant height; 8 for spike length; 17 for thousand kernel weight; 30 for grain yield; 4 for harvest index and 2 for biological yield. The phenotypic variation explained by individual marker?Ctrait associations ranged from 7.6 % to 36.2 %. The identification of genomic regions associated with grain yield and drought related traits is useful for the genetic improvement of cultivars better adapted to drought stress environments. Thus, the present study is encouraging in identifying significant marker?Ctrait associations through LD based association mapping analysis, which could complement and augment previous quantitative trait loci information for the potential use of Marker Assisted Selection for drought.  相似文献   

TRAP molecular markers as a system for saturation of the genetic map of durum wheat     

Virginia Menzo  Angelica Giancaspro  Stefania Giove  Domenica Nigro  Silvana Zacheo  Pasqualina Colasuonno  Ilaria Marcotuli  Ornella Incerti  Antonio Blanco  Agata Gadaleta 《Euphytica》2013,194(2):151-160

Target region amplification polymorphism (TRAP) is a relatively new PCR-based technique that detects large numbers of loci in a single reaction without extensive pre-PCR processing of samples. The aim of this study was to integrate TRAP markers in an EST-derived SSR linkage map of a RIL mapping population from the cross of the durum wheat cultivars Ciccio and Svevo, for a more general purpose of establishing a high-throughput system for genetic map saturation. Primer combinations producing PCR products with at least 4–5 polymorphic bands were selected and analyzed across the mapping population. The PCR reactions produced a total of 2,881 fragments with an average of 52 peaks per reaction. A total of 142 new TRAP markers were mapped and found to be randomly distributed in the genome. The total length of the map was 2,043.0 cM, with an average chromosome length of 145.9 cM. Homoeologous group one had the highest number of TRAP markers (38 loci) and the longest map length (407.9 cM) for a total of 87 markers, while the homoeologous group five had the lowest TRAP marker number (5 loci) and the shortest map length (232.5 cM). The distribution of markers among the seven homoeologous groups was random. The results indicate that TRAP is highly efficient in genetic mapping, generating a large number of markers scattered across the genome. This closes many existing gaps in marker coverage and may join otherwise separate linkage groups.  相似文献   

Effect of storage time and temperature on the total protein concentration and electrophoretic fractions in equine serum     

Daniela Alberghina  Stefania Casella  Claudia Giannetto  Simona Marafioti  Giuseppe Piccione 《Canadian journal of veterinary research》2013,77(4):293-296

Serum protein electrophoresis (SPE) is a technique that could be considered one of the most useful diagnostic aids available to the clinician. The effect of storage time and temperature on the total proteins and electrophoretic fractions (albumin, α₁-, α₂-, β₁-, β₂-, and γ-globulins) was assessed in 24 healthy horses. All samples, collected by jugular vein puncture, were centrifuged and divided into 4 aliquots. The 1st aliquot was analyzed within 3 h from collection (time 0), the 2nd was refrigerated at +4°C for 24 h, the 3rd was refrigerated at +4°C for 48 h, and the last was frozen at −20°C for 48 h. One-way repeated-measures analysis of variance (ANOVA) showed a significant effect (P < 0.05) of the different storage conditions on the concentrations of all the parameters studied and significant variations in the percentages of albumin, α₁-globulins, α₂-globulins, and γ-globulins. Compared with time 0 the total protein concentration increased significantly after 48 h at −20°C, the albumin percentage decreased after 48 h at −20°C, the α₁-globulin percentage increased after 24 h at +4°C, the α₂-globulin percentage increased after 48 h at +4°C and at −20°C, and the γ-globulin percentage increased after 48 h at −20°C. The results should help veterinary practitioners handle and store equine serum samples appropriately. Further investigations at different storage times and temperatures could be useful.  相似文献   

The Bio-Functional Properties of Pigmented Cereals may Involve Synergies among Different Bioactive Species     

Parizad  Parisa Abbasi  Capraro  Jessica  Scarafoni  Alessio  Bonomi  Francesco  Blandino  Massimo  Marengo  Mauro  Giordano  Debora  Carpen  Aristodemo  Iametti  Stefania 《Plant foods for human nutrition (Dordrecht, Netherlands)》2019,74(1):128-134

This study was aimed at characterizing the anthocyanins and phenolics profile in different varieties of pigmented corn and wheat and in some of their milling fractions. Acid/ethanol extracts were used to assess total anthocyanins, overall antioxidant activity, the overall polyphenol profile, and for evaluating the inhibition of pancreatic α-amylase and of intestinal α-glucosidase. Both enzymes were inhibited in a dose-dependent manner by all extracts, but individual extracts had specific effects on each enzyme. Anti-inflammatory response was evaluated by using acid-free extracts and Caco-2 cells transiently transfected with a luciferase reporter gene responding to cytokine stimulation. The immune response of interleukin-stimulated cells decreased significantly in a dose-dependent manner in the presence of 20–50 μM/l anthocyanins from all grains extracts, again with a different efficiency. The inhibitory ability and the anti-inflammatory capability of these extracts are in most cases higher than in similar extracts from other sources, suggesting that activities in each extract may imply specific synergies between anthocyanins and other phenolics.

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Genetic diversity of ICARDA’s worldwide barley landrace collection     

Abderrazek Jilal  Stefania Grando  Robert J. Henry  L. Slade Lee  Nicole Rice  Helen Hill  Michael Baum  Salvatore Ceccarelli 《Genetic Resources and Crop Evolution》2008,55(8):1221-1230

Twenty genic- and genomic SSR markers were used to study genetic diversity and geographical differentiation of barley from 29 countries through analysis of a worldwide collection of 304 ICARDA’s barley landraces. Of these, 19 loci were highly polymorphic in the material studied. Based on Nei-distance matrix, Principal Component Analysis (PCoA) and cluster analysis using UPGMA associated with AMOVA the data revealed countries’ grouping within regions. Three distinct germplasm pools were identified in the landraces. The first of these was from Eastern Africa (Eritrea and Ethiopia) and South America (Ecuador, Peru and Chile) suggesting that barley introduced to South America might have originated specifically from East Africa or that they share a common genetic basis for adaptation. The second was the Caucasus (Armenia and Georgia) and the third included the remaining regions of Central Asia, Near East, Northern Africa and Eastern Asia. Genetic diversity of barley subspecies (Six-rowed barley, Two-rowed barley, H. spontaneum C. Koch and H. agriocrithon Åberg) also discriminates them into three groups: cultivated barleys (Six-rowed barley and Two-rowed barley), wild barley H. spontaneum and subspecies H. agriocrithon. These results associated with parsimony analysis demonstrate that H. agriocrithon and H. spontaneum might be distinct and do not support a hybrid origin for H. agriocrithon suggesting further investigation of the basis of more intense sampling of the two subspecies H. spontaneum and H. agriocrithon.  相似文献   

Cat and dog ownership and management patterns in central Italy     

Slater MR  Di Nardo A  Pediconi O  Villa PD  Candeloro L  Alessandrini B  Del Papa S 《Preventive veterinary medicine》2008,85(3-4):267-294

Three hundred and ninety-seven randomly selected households were interviewed by telephone to determine the numbers and management of owned cats and dogs in the Teramo Province of Italy. The households were selected using stratified random sampling for each municipality; municipalities were combined into coastal, central hills and mountain regions for analysis. The interviews were completed during May and June of 2004 with a response rate of 74% (397/536). Forty-six percent of households (n=181) owned pets; 15% of all households (n=60) owned cats and 33% (n=130) owned dogs. Twenty-seven of these households (7%) owned both cats and dogs. Data were provided on 91 cats evenly divided between males and females. The median age was 3 years (range 0.2-10 years). Forty-one percent of cats (36/87) entered the household as strays. Nearly half lived entirely outside. Seventy percent (62/88) had visited a veterinarian at least once; 43% (39/91) were sterilized. About 1/3 had had a litter and all litters were considered accidental rather than planned. Age, indoor/outdoor status, veterinarian visit and region were all associated with sterilization. Age, confined to a yard, veterinary visit and region were associated with allowing the cat to roam freely. Data were provided on 182 dogs. Sixty-two percent (113/181) were male, with a median age of 4 years (range newborn to 17 years) and 40% (72/181) were purebred. Almost half were acquired as a gift. Sixty-two percent (112/180) were kept entirely outside despite the fact that 82% (147/180) were considered companions rather than working dogs. Almost all of the dogs had been to a veterinarian at least once; only 20% (n=29) were sterilized. Male dogs were significantly less likely to be sterilized than females. Almost half the dogs had had at least one litter. Seventy-six percent (137/180) of dogs knew some basic commands. Sex, source and training to sit/stay/come were significantly associated with whether the dog was sterilized. Dog size, confinement to a yard, trained to leash walk, veterinary visit and region were predictors of roaming freely. Source and trained to leash walk were associated with dog registration. For cats and dogs, education about sterilization seemed to be critically important. For dogs, factors relating to training, which may reflect the strength of the human-animal bond, were also important.  相似文献