Molecular characterization of Swiss Ceratopogonidae (Diptera) and evaluation of real-time PCR assays for the identification of Culicoides biting midges

Biting midges of the genus Culicoides (Diptera, Ceratopogonidae) are vectors of several viruses of veterinary relevance, and they can cause insect bite hypersensitivity. As the morphological identification of these tiny insects is a difficult task in many cases, alternative approaches are expedient. With the aim to develop real-time PCRs, we determined partial mitochondrial cytochrome oxidase I gene (mt COI) sequences from 380 Culicoides midges representing three regions of Switzerland, namely the Alps, Midland north of the Alps (Atlantic climate), and South of the Alps (Mediterranean climate). The same region was also sequenced from non-biting midges of the genera Atrichopogon, Brachypogon, Dasyhelea, Forcipomyia and Serromyia. A total of 21 Culicoides species were identified by morphology. Sequence variability (haplotypes) was observed in all species. For each of C. grisescens and C. obsoletus, a novel cryptic species was identified. Whereas all individuals of C. grisescens and of the cryptic C. obsoletus species (O2) originated only from Alpine sites, the known C. obsoletus (O1) species was found in all three regions. Further, a sister taxon to C. pulicaris was identified based on the mt COI sequences and named Culicoides sp. Alignments of available mtCOI sequences from Ceratopogonidae (GenBank, this study) were used to design real-time PCR primers and probes to distinguish C. chiopterus, C. deltus, C. dewulfi, C. grisescens (including the cryptic species), C. imicola, C. lupicaris, C. obsoletus O1, C. obsoletus O2, C. pulicaris, C. scoticus and Culicoides sp. Specificities of primers and probes was tested with cloned targets representing 1 to 4 haplotypes of 18 Culicoides spp. and 1 haplotype each from 4 other Ceratopogonidae. No cross-reactivity was observed when plasmid template representing 5 × 10(6) gene copies was tested, but it was evident (Ct values ≤ 30) in few instances when plasmid template representing 5 × 10(9) gene copies was utilized, the latter corresponding to the total gene copy number (as determined in this study) in 20 insects. The sensitivities of two assays (C. imicola, C. grisescens) were tested by spiking single insects into pools of 99 or 999, randomly selected non-target Ceratopogonidae (with approx. 90% Culicoides specimens). In the pools of 100, Ct values were in the range of those obtained with single insects when employing 1% of the isolated DNA, whereas the sensitivity with the pools of 1000 was low, presumably due to the low DNA concentrations obtained with a protocol that seems inadequate for these larger pools. Thus, the assays as described are applicable for the specific identification of biting midges in small pools. Primers and probes of this study were devised to be suitable for multiplexed assays but these evaluations await to be performed.     

Die Entwicklung des Plazentastieles beim Meerschweinchen

As basis for detailed studies into the significance of the guinea pig subplacenta the formal development of the placental stalk which contains the subplacenta in its center was investigated in this species. Fifty conventional, domesticated, genetically undefined guinea pigs pregnant 8–63 days were used. The placentas were studied on semi-thin and paraffin sections on some of which morphometric observations were made. The findings provide the first three-dimensional picture of the subplacenta and give information of its development. The studies show that this region has neither a growth function nor a maternofetal transport function.     

Spatiotemporal variations of soil surface roughness from in-situ laser scanning

Microtopography and roughness are highly dynamic properties of the soil surface and important factors governing surface runoff and erosion processes. While various remote sensing technologies were successfully applied for topography measurements at different spatial scales, there is a lack of field studies that collected systematically microtopography data over long observation periods. In this paper an approach to measure and quantify surface roughness in the field based on laser scanning technologies is presented. Between June 2004 and November 2005 97 in-situ measurements were conducted in a test site with two different sandy substrates in vegetation-free conditions. Two-dimensional high-resolution (1 mm) datasets where generated for eight micro erosion plots of 0.25 to 2.9 m² in size. Dynamics and pattern formation were quantified for surface roughness and surface height changes. Roughness patterns at different scales were analyzed by local roughness indices using sliding windows of 3 to 55 mm in size. Results show strong spatial and temporal dynamics in surface roughness as well as substrate-specific variations. Temporal roughness variations could be detected and were linked to precipitation patterns. The methods presented in this paper are considered suitable to generate high-resolution datasets on spatiotemporal and multi-scale microtopography patterns and to advance the understanding of surface processes at small scales in natural environments.     

Genetic parameters for individual birth and weaning weight and for litter size of Large White pigs

Data from a French experimental herd recorded between 1990 and 1997 were used to estimate genetic parameters for individual birth and weaning weight, as well as litter size of Large White pigs using restricted maximum likelihood (REML) methodology applied to a multivariate animal model. In addition to fixed effects the model included random common environment of litter, direct and maternal additive genetic effects. The data consisted of 1928 litters including individual weight observations from 18 151 animals for birth weight and from 15 360 animals for weaning weight with 5% of animals transferred to a nurse. Estimates of direct and maternal heritability and proportion of the common environmental variance for birth weight were 0.02, 0.21 and 0.11, respectively. The corresponding values for weaning weight were 0.08, 0.16 and 0.23 and for litter size 0.22, 0.02 and 0.06, respectively. The direct and the maternal genetic correlations between birth and weaning weight were positive (0.59 and 0.76). Weak positive (negative) genetic correlations between direct effects on weight traits and maternal effects on birth weight (weaning weight) were found. Negative correlations were found between direct genetic effect for litter size and maternal genetic effects on all three traits. The negative relationship between litter size and individual weight requires a combined selection for litter size and weight.     

Population structure of short‐finned pilot whales in the oceanic archipelago of Madeira based on photo‐identification and genetic analyses: implications for conservation

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Response of osteoclasts to titanium surfaces with increasing surface roughness: an in vitro study

Osteoclasts are responsible for bone resorption and implant surface roughness promotes osseointegration. However, little is known about the effect of roughness on osteoclast activity. This study aims at the characterization of osteoclastic response to surface roughness. The number of osteoclasts, the tartrate-resistant acid phosphatase and matrix metalloproteinase (MMP) activities, the cell morphology and the actin-ring formation were examined on smooth (TS), acid-etched (TA) and sandblasted acid-etched (TLA) titanium and on native bone. Cell morphology was comparable on TA, TLA and bone, actin rings being similar in size on TLA and bone, but smaller on TA and virtually absent on TS. Gelatin zymography revealed increased proMMP-9 expression on TA, TLA, and bone compared to TS. In general, osteoclasts show similar characteristics on rough titanium surfaces and on bone, but reduced activity on smooth titanium surfaces. These results offer some insight into the involvement of osteoclasts in remodeling processes around implant surfaces.     

Increasing carbon sinks through forest management: a model-based comparison for Switzerland with its Eastern Plateau and Eastern Alps

The Kyoto Protocol brought a new forest function into focus: forests as carbon sinks. This new forest function may lead to new conflicts, because on the one hand, Switzerland has decided to account for forest management under Kyoto Protocol (Article 3.4), and on the other hand, Swiss Forestry statistics and the Swiss National Forest Inventory indicate that increasing amounts of wood are being harvested. This trend seems likely to continue. In this study, we used the empirical forest model MASSIMO and the soil model YASSO to analyse four different forest management scenarios. These scenarios basically feature different levels of harvesting frequencies and different rotation length, as well as their impact on regional potentials for carbon sequestration and harvesting amounts. Results were analysed both for the whole of Switzerland and for two very different regions: The Swiss Eastern Plateau and the Swiss Eastern Alps. The results indicate that Swiss forests can provide an increasing amount of harvested wood (+18% in relation to the base year 1996) for approximately 20 years and act as a carbon sink accountable under the Kyoto Protocol (0.5 million tons carbon per year). The corresponding forest management strategy aims for a sustainable and harvestable increment and may, therefore, avoid spurious carbon maximization in forests that can happen by accounting for only forest systems, and not for the effect of substitution of non-wood products and fossil fuels by forest products. The regional results indicate that (1) the carbon sink effect of Alpine forests in Switzerland might be limited, because generally, Alpine forests have low growth and yield and (2) a large increase in harvesting may lead to regional carbon sources and necessitate regional monitoring of increment to avoid overexploitation. As MASSIMO does not include the impacts of climate change, the conclusions of this study cannot be interpreted as actual predictions into the future but portray the impact of the applied management actions on the respective trends in carbon stocks and stock changes. They are, therefore, a contribution to support future management decisions. Further studies should focus on interactions with additional forest functions such as the preservation of biodiversity, increase the consideration of forest damage and account for the effect of climate change.     

Molecular cloning and characterization of markers and cytokines for equid myeloid cells

The myeloid cell system comprises of monocytes, macrophages (MPhi), dendritic cells (DC), Kupffer cells, osteoclasts or microglia and is also known as the mononuclear phagocytic system (MPS). Essential cytokines to differentiate or activate these cells include GM-CSF or IL-4. Important markers for characterization include CD1, CD14, CD68, CD163 and CD206. All these markers, however, were not cloned or further characterized in equids by use of monoclonal antibodies earlier. To overcome this problem with the present study, two approaches were used. First, we cloned equine cytokines and markers, and second we analyzed cross-reactivity of human homologues or anti-human monoclonal antibodies. For cloning of equine cytokines and markers, we used degenerate primers delineated from other species, or equine-specific primers based on previous information in Genbank. Flow cytometry was used to determine the expression of markers on myeloid cells. Cross-reactivity could be shown for anti-human CD14, CD163 and mannose receptor (CD206) mAbs. Surface markers such as CD1 and CD68 that distinguish MPhi and DC were cloned and sequenced. According to blast homology, equine CD1a and CD1b could be identified and distinguished. With the resulting information, dendritic cells and macrophages of horses may be characterized.     

Effect of strategic gastrointestinal nematode control on faecal egg count in traditional West African cattle

This paper reports on the effect of strategic anthelmintic treatments and other determinants on faecal egg counts (FEC) of Trichostrongyles in N'Dama cattle of a west African village. Initially, 527 animals from 13 private N'Dama cattle herds were monitored in a longitudinal study from October 1989 to December 1994. Each herd was stratified by age and animals were sequentially allocated to two groups with similar age distributions. One group received a single anthelmintic treatment of fenbendazole (7.5 mg/kg BW), in October 1989 (n = 250), whereas the other group remained untreated (n = 277) throughout the study. In the next rainy season (June to October), the treated animals were treated twice (in July and September). The same treatment schedule was used in the subsequent rainy seasons until December 1994. Biannual anthelmintic treatments decreased the level of FEC between 31% (late dry season) and 57% (rainy season), when compared to untreated controls. The highest levels of FEC were found during the rainy season from June to October. FEC levels decreased until 4 years of age, after which they remained on a constant low level. The variability of returns to anthelmintic treatments between herds did not seem to be influenced by FEC at the herd level. The financial evaluation of anthelmintic interventions cannot be predicted from FEC and must necessarily rely on the direct monitoring of livestock productivity parameters.