Real-time PCR for Quantification of Toxigenic Fusarium Species in Barley and Malt

A real-time PCR technique was applied for the quantification of trichothecene-producing Fusarium species (TMTRI assay) as well as the highly toxigenic Fusarium graminearum (TMFg12 assay) present in barley grain and malt. PCR results were compared to the amounts of trichothecenes detected in the samples to find out if the PCR assays can be used for trichothecene screening instead of expensive and laborious chemical analyses. DNA was extracted from ground kernels using a commercial DNA extraction kit and analysed in a LightCycler® system using specific primers and fluorogenic TaqMan probes. Both naturally and artificially contaminated grains were analysed. The TMTRI assay and the TMFg12 assay enabled the quantification of trichothecene-producing Fusarium DNA and F. graminearum DNA present in barley grain and malt samples, respectively. Both TaqMan assays were considered to be sensitive and reproducible. Linearity of the assays was 4–5 log units when pure Fusarium DNAs were tested. The amount of Fusarium DNA analysed with the TMTRI-trichothecene assay could be used for estimation of the deoxynivalenol (DON) content in barley grain. Furthermore, the TMFg12 assay for F. graminearum gave a good estimation of the DON content in north American barley and malt samples, whilst the correlation was poor among Finnish samples. DON content and the level of F. graminearum DNA were found to be naturally low in Finnish barleys.     

Development of Strain-specific Primers for a Strain of Gliocladium catenulatum Used in Biological Control

The randomly amplified polymorphic DNA (RAPD) technique was used to develop strain-specific primers for Gliocladium catenulatum strain J1446, which is promising in biological control. One of the primer pairs developed proved to be strain-specific; strain J1446 was differentiated from 16 G. catenulatum strains and six other strains of two Gliocladium species, as well as from Trichoderma virens, and isolates of Nectria spp. and Fusarium spp. Specific primers were also tested with DNA isolated from cucumber leaves, treated or untreated with a solution made from Gliocladium powder. The expected amplification product was produced only from treated leaves. DNA isolated from Gliocladium-treated potato tubers and fungi grown in peat was also used in amplification reactions. Strain-specific primers detected strain J1446 when the amount of DNA was 5pg or more. Some variation between the Gliocladium strains was found by the random amplified microsatellites method (RAMS) and the universally primed polymerase chain reaction method (UP-PCR), but no clear fragments specific to strain J1446 were produced. Cross-blot hybridisation of UP-PCR products differentiated strain J1446 from T. virens, but not from the Gliocladium isolates. The 28S rDNA sequences and -tubulin sequences were identical or very similar in all Gliocladium strains. Thus, it is possible that the Gliocladium strains of the present study are conspecific, which means that a revision in the taxonomy of Gliocladium species may be necessary.     

Etiology of respiratory disease in non-vaccinated, non-medicated calves in rearing herds

The aim of this study was to examine the occurrence of bacterial, mycoplasmal and viral pathogens in the lower respiratory tract of calves in all-in all-out calf-rearing units. According to clinical status, non-medicated calves with and without respiratory disease signs were selected of the 40 herds investigated to analyse the micro-organisms present in healthy and diseased calves. Tracheobronchial lavage (TBL) and paired serum samples were analysed for bacteria, mycoplasmas, respiratory syncytial virus (RSV), parainfluenza virus 3 (PIV3), bovine corona virus (BCV) and bovine adenovirus (BAV). Pasteurella multocida was the most common bacterial pathogen. It was isolated from 34% of the TBL samples in 28 herds and was associated with clinical respiratory disease (p < 0.05) when other pathogenic bacteria or mycoplasma were present in the sample. Mannheimia spp. and Histophilus somni were rarely found. Mycoplasma bovis was not detected at all. Ureaplasma diversum was associated with clinical respiratory disease (p < 0.05). TBL samples from healthy or suspect calves were more often negative in bacterial culture than samples from diseased calves (p < 0.05). No viral infections were detected in six herds, while 16-21 herds had RSV, BCV, BAV or PIV3. In the herds that had calves seroconverted to BCV, respiratory shedding of BCV was more frequently observed than faecal shedding. This study showed that the microbial combinations behind BRD were diverse between herds. M. bovis, an emerging pathogen in many countries, was not detected.     

Modelling the predisposition of Norway spruce to <Emphasis Type="Italic">Ips typographus</Emphasis> L. infestation by means of environmental factors in southern Finland

New measures for effective monitoring and controlling of bark beetle infestations are needed as a response to intensified outbreaks caused by the climate change. Various environmental factors affect tree health and susceptibility, as well as stand predisposition to bark beetles. European spruce bark beetle Ips typographus L. abundance and outbreak frequency in Finland has significantly increased during the last decade. The ability to identify sites under a high risk of infestation would facilitate adaptation to this new situation and help target limited forest health management resources. Accordingly, our goal was to investigate the importance of various stand, soil and topographic characteristics in the assessing predisposition of Norway spruce dominated urban forest in southern Finland to I. typographus infestations. Information on the environmental factors was assessed in the field in 2014 and derived from a digital elevation model. Ips typographus infestation intensity was classified into three infestation index classes based on tree-wise symptoms of resin flow, discoloration and defoliation. Cumulative logit link models were utilized for investigating stand-level infestation probability. The best explanatory factors were aspect, slope, site type and soil texture. Models with the highest cumulative probabilities for severe infestation were linked with eastern aspect, moderate steep slope and rich site type fertility (0.72) and eastern aspect, shallow soil and rich site type fertility (0.71). Higher soil C/N ratios with east aspect and rich site type fertility was associated with an increased risk of severe infestation in a third model. The lowest risk was associated with southern and southwestern aspects, fine soil texture, moderate site fertility and gentle slopes.     

Agro-morphological traits of <Emphasis Type="Italic">Cicer reticulatum</Emphasis> Ladizinsky in comparison to <Emphasis Type="Italic">C. echinospermum</Emphasis> P.H. Davis in terms of potential to improve cultivated chickpea (<Emphasis Type="Italic">C. arietinum</Emphasis> L.)

Germplasm collections of C. reticulatum and C. echinospermum are limited, while both species face threats from over-grazing and habitat change in their natural environments. Recently many new accessions of C. reticulatum and C. echinospermum were collected in east and south-east Anatolia (Turkey) but they have not yet been evaluated for agro-morphological traits. Therefore, the current study investigated agro-morphological traits of new germplasm sources of C. reticulatum and C. echinospermum and evaluated resistance to biotic and abiotic stresses for chickpea improvement. The most attractive agro-morphological traits were canopy width, number of stems and pods per plant and biological yield. The most productive accessions of C. reticulatum and C. echinospermum had 712 and 625 pods per plant, respectively. Two distinct seed, flower and leaf shapes were found in accessions of C. echinospermum. Path analyses indicated that biological yield and harvest index had the most direct influence on seed yield in both species. Factor analyses showed that high seed yield in C. reticulatum depended on high biological yield and number of pods per plant, whereas high seed yield in C. echinospermum depended on harvest index. It was concluded that most accessions of C. reticulatum and C. echinospermum were not only resistant to some biotic and abiotic stresses but also had hidden alleles that could produce transgressive segregation in crosses to cultivated material.     

Estimation of light interception properties of conifer shoots by an improved photographic method and a 3D model of shoot structure

The spherical mean of the shoot silhouette-to-total leaf area ratio (STAR) and the shoot transmission coefficient (c) are two key structural parameters in radiative transfer models for calculating canopy photosynthesis and leaf area index. The standard optical method for estimating these parameters might introduce errors in the estimates for species with flexible shoots and needles by changing shoot inclination relative to its inclination in situ. We devised and tested two methods to address this problem. First, we modified the standard optical method by designing an apparatus that allows shoots to be photographed in their original orientation. Second, we developed a faster, model-based approach to replace photography and tested the results against the established approach. We used shoots of three pine species, Pinus echinata Mill. (needle length ~50 mm), P. taeda L. (~150 mm) and P. palustris Mill. (~300 mm). Values of the parameters simulated by the model were similar to those measured from the photographs. In our data, STAR varied about twofold among the pine species and was ~40% higher in shade shoots than in sun shoots of P. taeda. The transmission coefficient for P. taeda shade shoots was also ~40% higher than that of sun shoots of all three species. We tested the versatility of the model by employing it on shoots of two other pine species (P. strobus L. and P. thumbergiana Parl.) as well as on shoots of Tsuga canadensis L. Carr. and Picea pungens Engelm. Regardless of shoot characteristics, the model generated values of shoot structural parameters similar to those estimated with the optical method. Although species-specific and vertical gradients in parameter values are best for modeling radiative transfer in conifer canopies, our results suggest that, in the absence of adequate data, STAR can be approximated as 0.16 for a wide range of shoot structures. For applications requiring angle-dependent parameterization, our new model facilitates rapid generation of these radiative transfer parameters.     

Whole‐community facilitation by beaver: ecosystem engineer increases waterbird diversity

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