The effect of an experimentally induced acute mastitis on the test results of an Ostertagia ostertagi milk ELISA

Antibody levels against Ostertagia ostertagi in the milk are a promising parameter to identify dairy cows or herds with production losses due to gastrointestinal nematodes. However, milk antibody levels can be influenced by udder-related factors. In this study, the effect of an experimentally induced mastitis on the test results (ODR) of a milk O. ostertagi ELISA was investigated. Twenty-five cows that were naturally infected with gastrointestinal nematodes, were inoculated in their left udder quarters with Escherichia coli P4:O32 and quarter milk samples were collected at several intervals from 24 h before until 144 h after experimental infection. The effect of the contribution of milk from one or more infected quarters on the bulk tank milk ODR was estimated, based on a titration experiment. The mean O. ostertagi antibody levels of the infected udder quarters were significantly (P < 0.001) higher than those of the uninfected udder quarters at each sampling time post-infection. The largest difference was observed at 24 h post-infection with a mean difference of 0.251 ODR (95%CI: 0.172; 0.330). There was also a significant increase (P < 0.001) in total IgG levels with the largest difference being observed at 24 h post-infection. Highly significant (P < 0.005) correlation coefficients were observed between O. ostertagi ODR, total IgG ODR, Na+ and Cl- ion concentration and log transformed somatic cell counts at 24 h post-infection. The results demonstrate that an acute mastitis causes a flow of specific and non-specific antibodies from serum to milk with a subsequent increase in the O. ostertagi ODR values. The effect of the contribution of milk from infected quarters on the bulk tank milk O. ostertagi ODR was estimated to be minor if the relative number of infected quarters is small (< 3%).     

PIN proteins perform a rate-limiting function in cellular auxin efflux

Intercellular flow of the phytohormone auxin underpins multiple developmental processes in plants. Plant-specific pin-formed (PIN) proteins and several phosphoglycoprotein (PGP) transporters are crucial factors in auxin transport-related development, yet the molecular function of PINs remains unknown. Here, we show that PINs mediate auxin efflux from mammalian and yeast cells without needing additional plant-specific factors. Conditional gain-of-function alleles and quantitative measurements of auxin accumulation in Arabidopsis and tobacco cultured cells revealed that the action of PINs in auxin efflux is distinct from PGP, rate-limiting, specific to auxins, and sensitive to auxin transport inhibitors. This suggests a direct involvement of PINs in catalyzing cellular auxin efflux.     

Mental notes in tracking BVDV carriers

The detection and elimination of animals persistently infected with bovine viral diarrhoea virus (BVDV) is the key to BVD control. This method has proven to be very efficient in eradicating BVDV in a herd. Several pitfalls in the detection procedure can make that some persistently infected (PI) animals do not get identified or are removed too late, supporting the assumption that circulation of the virus could be possible in absence of PI animals. Furthermore the risk of reintroduction is high since the prevalence of BVD is high in the Netherlands and Belgium. Based on both practical experience and literature, here we review critical control points in order to minimise the risk of a false negative BVDV screening.     

Visual and quantitative electroencephalographic analysis of healthy young and adult cats under medetomidine sedation

A study was designed to investigate the effect of medetomidine sedation on quantitative electroencephalography (q-EEG) in healthy young and adult cats to determine objective guidelines for diagnostic EEG recordings and interpretation. Preliminary visual examination of EEG recordings revealed high-voltage low-frequency background activity. Spindles, k-complexes and vertex sharp transients characteristic of sleep or sedation were superimposed on a low background activity. Neither paroxysmal activity nor EEG burst-suppression were observed. The spectral analysis of q-EEG included four parameters, namely, relative power (%), and mean, median and peak frequency (Hz) of all four frequency bands (delta, theta, alpha and beta). The findings showed a prevalence of slow delta and theta rhythms as opposed to fast alpha and beta rhythms in both young (group A) and adult (group B) cats. A posterior gradient was reported for the theta band and an anterior gradient for the alpha and beta bands in both groups, respectively. The relative power value in group B compared to group A was significantly higher for theta, alpha and beta bands, and lower for the delta band. The mean and median frequency values in group B was significantly higher for delta, theta and beta bands and lower for the alpha band. The study has shown that a medetomidine sedation protocol for feline EEG may offer a method for investigating bio-electrical cortical activity. The use of q-EEG analysis showed a decrease in high frequency bands and increased activity of the low frequency band in healthy cats under medetomidine sedation.     

Long-Term Soil Reaction Changes in a Temperate Beech Forest Subject to Past Alkaline Pollution

Changes in deposition exert effects on forests. Some evidence for a slow recovery of strongly alkalised forest soils after a reduction in alkaline pollution exists, but the recovery rate is little known. The objective of this study was to estimate temporal soil reaction changes. For this purpose, measurements of soil reaction under a beech forest near magnesite works in Central Slovakia were taken in 1991 and 2006, following the alkaline pollution reduction in 1992. As a result, the mean active soil reaction decreased from pH 7.8 to 6.6. Beyond the range of beech stemflow, which amplifies water and acid pollution inputs into soils locally, soil reaction dropped from pH 7.8 to 6.8. The effect of stemflow on local exchangeable reaction minima was detected even before the pollution reduction. The logarithmic function fitted on the pH data indicates considerable differences between neutralisation rates within and outside the stemflow zone.     

Contour furrows for in situ soil and water conservation, Tigray, Northern Ethiopia

In Tigray (Northern Ethiopia), soil moisture has been identified as the most limiting factor in agricultural production; on the other hand, loss of rain water through runoff as well as the induced soil loss has been determined as a critical problem in the region in the last two to three decades. To alleviate the above paradox, the government has mobilized communities and resources for the construction of physical soil and water conservation structures (stone bunds, terraces) in almost all land uses. However, yield improvement was mainly concentrated within the vicinity of the structures and runoff continued to overtop the structures, as no measures for in situ soil conservation were taken. The terwah system, consisting of traditional ploughing followed by making every 1.5–2 m contour furrows, and permanent raised beds with contour furrows at 60–70 cm interval treatments, were considered and evaluated as practices that could increase the efficiency of in situ water utilization and soil conservation. An experiment was started in Gum Selasa, which is one of the drought prone areas in Tigray, whereby runoff volume and sediment load were measured after every rain event. Permanent raised beds with contour furrows at 60–70 cm interval significantly (P < 0.05) reduced runoff volume, runoff coefficient and soil loss as compared to traditional ploughing: 255, 381 and 653 m³ ha⁻¹ runoff was recorded from permanent bed, terwah and traditional ploughing, respectively during the whole cropping season. The above runoff induced 4.7 t ha⁻¹ soil loss from permanent bed, 7.6 t ha⁻¹ from terwah and 19.5 t ha⁻¹ from traditional ploughing. Overall, contour furrows and permanent raised beds can be part of the ongoing intensification process which includes physical soil and water conservation, slope reforestation, irrigation development and agro forestry in crop lands. Moreover, the use of permanent raised beds if combined with crop mulching and crop diversification is an important component for the development of sustainable conservation agriculture practices in the region.     

Generation of equine TSLP-specific antibodies and their use for detection of TSLP produced by equine keratinocytes and leukocytes

Allergic horses react to innocuous environmental substances by activation of Th2 cells and production of allergen-specific IgE antibodies. The mechanisms leading to Th2 differentiation are not well understood. In humans and mice, epithelial cell-derived thymic stromal lymphopoietin (TSLP) plays a central role in this process. Little is known about equine TSLP (eqTSLP) and its role in allergic diseases and our current knowledge is limited to the assessment of TSLP mRNA expression. In order to be able to study eqTSLP at the protein level, the aim of the present study was to produce recombinant eqTSLP protein and generate TSLP-specific antibodies. EqTSLP was cloned from a skin biopsy sample from a horse with chronic urticaria and eqTSLP protein was expressed in E.coli and in mammalian cells. Recombinant proteins were designed to include C-terminal Histag, which allowed subsequent purification and detection by Histag-specific Ab. Polyclonal and monoclonal eqTSLP-specific Ab were generated after immunization of mice with E.coli-expressed TSLP. Their specificity was tested by western blotting and ELISA. In addition, a commercially available polyclonal human TSLP-specific antibody was tested for cross-reactivity with eqTSLP. Expression of TSLP protein was confirmed by western blotting using Histag-specific Ab. E.coli-expressed TSLP appears as a band of ～13 kDa, whereas mammalian cell-expressed TSLP showed several bands at 20-25 kDa, probably representing several glycosylation forms. Polyclonal and monoclonal antibodies generated in this study, as well as commercially available human TSLP-specific Ab reacted with both E.coli- and mammalian cell-expressed TSLP in western blotting and ELISA. A capture ELISA was established to quantitate TSLP in cell supernatants and validated using supernatants from primary equine keratinocytes and peripheral blood leukocytes (PBL). Increased TSLP concentrations were found after stimulation of keratinocytes with PMA+ionomycine and with Culicoides extract. Similarly, increased TSLP concentrations were detected in PBL after stimulation with ConA, Culicoides extract, or IgE cross-linking. In conclusion, recombinant TSLP proteins and TSLP-specific antibodies produced in this study will allow further studies of the role of TSLP in equine allergic diseases.     

Characterization of the inflammatory infiltrate and cytokine expression in the skin of horses with recurrent urticaria

Background – Recurrent urticaria (RU) is a common skin disease of horses, but little is known about its pathogenesis. Hypothesis/Objective – The aim of this study was to characterize the inflammatory cell infiltrate and cytokine expression pattern in the skin of horses with RU. Animals – Biopsies of lesional and nonlesional skin of horses with RU (n = 8) and of skin from healthy control horses (n = 8) were evaluated. Methods – The inflammatory cell infiltrate was analysed by routine histology. Immunohistochemistry was used to identify T cells (CD3), B cells (CD79), macrophages (MAC387) and mast cells (tryptase). Expression of T‐helper 2 cytokines (interleukins IL‐4, IL‐5 and IL‐13), a T‐helper 1 cytokine (interferon‐γ), IL‐4 receptor α and thymic stromal lymphopoietin was assessed by quantitative RT‐PCR. Results – In subepidermal lesional skin of RU‐affected horses, increased numbers of eosinophils (P ≤ 0.01), CD79‐positive (P ≤ 0.01), MAC387‐positive (P ≤ 0.01) and tryptase‐positive cells (P ≤ 0.05) were found compared with healthy horses. Subepidermal lesional skin of RU‐affected horses contained more eosinophils (P ≤ 0.05) and tryptase‐positive cells (P ≤ 0.05) compared with nonlesional skin. There was no significant difference in infiltrating cells between nonlesional skin and skin of healthy horses. Expression of IL‐4 (P ≤ 0.01), IL‐13 (P ≤ 0.05), thymic stromal lymphopoietin (P ≤ 0.05) and IL‐4 receptor α (P ≤ 0.05) was increased in lesional skin of RU‐affected horses compared with control horses. Expression of IL‐4 was higher (P ≤ 0.05) in lesional compared with nonlesional RU skin. Conclusions and clinical importance – Analysis of cytokine expression and inflammatory infiltrate suggests that T‐helper 2 cytokines, eosinophils, mast cells and presumptive macrophages play a role in the pathogenesis of equine RU.     

ParaCalc®--a novel tool to evaluate the economic importance of worm infections on the dairy farm

Subclinical infections with gastrointestinal nematodes and liver fluke are important causes of production losses in grazing cattle. Although there is an extensive compilation of literature describing the effect of these infections on animal performance, only a few attempts have been made to convert these production losses to an economic cost. Here, we propose a novel tool (ParaCalc(?)), available as a web-application, to provide herd-specific estimates of the costs of these infections on dairy farms. ParaCalc(?) is a deterministic spread-sheet model where results from diagnostic methods to monitor the helminth infection status on a herd and anthelmintic usage are used as input parameters. Default values are provided to describe the effects of the infections on production and the cost of these production losses, but the latter can be adapted to improve the herd-specificity of the cost estimate. After development, ParaCalc(?) was applied on input parameters that were available for 93 Belgian dairy herds. In addition, the tool was provided to 6 veterinarians and their user experiences were evaluated. The estimated median [25th-75th percentile] cost per year per cow was € 46 [29-58] and € 6 [0-19] for gastrointestinal nematode and liver fluke infection, respectively. For both infections, the major components in the total costs were those associated with milk production losses in the adult cows. The veterinarians evaluated ParaCalc(?) as a useful tool to raise the farmers' awareness on the costs of worm infections, providing added value for their services. However, the score given for user-friendliness was diverse among users. Although the model behind ParaCalc(?) is a strong simplification of the real herd processes inducing economic losses, the tool may be used in the future to support economic decisions on helminth control.     

A survey to determine relationships between bulk tank milk antibodies against Ostertagia ostertagi and milk production parameters

In temperate climate regions, gastrointestinal nematodes are still widespread in adult dairy cows, but until now there exists no reliable diagnostic tool that can identify herds where the infection interferes with productivity. The objective of this study was to investigate the relationships between levels of antibodies against Ostertagia ostertagi in bulk tank milk and milk production. Bulk tank milk samples of 2553 dairy herds were obtained in spring and 2104 of these herds were sampled a second time in autumn. The antibody levels against O. ostertagi were determined with a milk ELISA and test results were expressed as an optical density ratio (ODR). The effect of bulk tank milk ODR on three different production parameters, kg milk, % and kg fat, % and kg protein was assessed by a multivariable linear regression model on the herds for which production data were available (n = 1063 and 867 in spring and autumn, respectively). The mean and standard deviation for ODRautumn (0.972+/-0.238) were higher than for ODRspring (0.825+/-0.201). Significant negative relationships were found between ODR and milk yield. An increase in ODRspring and ODRautumn from the 25th to the 75th percentile of the available ODR data was associated with a drop in the annual milk yield of 1.1 kg/cow/day, respectively 0.9 kg/cow/day. When a herd's ODR increased between spring and autumn with 0.142, it produced on average 0.4 kg/cow/day less in September than in April, in comparison with herds where the ODR did not increase. A significant negative association was found between ODRautumn and % protein averaged over the period of a year. No significant associations were found between ODR and % fat averaged over the year. When protein and fat production of September were expressed in kg an increase in ODRautumn from the 25th to the 75th percentile was associated with a decrease of 0.037 kg protein/cow/day and 0.042 kg fat/cow/day.