Cloning and characterization of two glutathione S-transferases from pyrethroid-resistant Culex pipiens

BACKGROUND: The Marin strain of Culex pipiens Say is a pyrethroid‐resistant population that was collected in Marin County, California, in 2001 and subsequently maintained in the laboratory under regular permethrin exposure. RESULTS: In this study, two cDNAs, CpGSTd1 and CpGSTd2, encoding glutathione S‐transferase (GST) were cloned from Cx. pipiens Marin. Phylogenetic analysis of the deduced amino acid sequences, CpGSTD1 and CpGSTD2, of these genes indicated that they belong to the Delta class of insect GSTs. The nucleotide and deduced amino acid sequences of CpGSTd1 and CpGSTd2 were 59 and 48% identical respectively. CpGSTD1 and CpGSTD2 were expressed in Escherichia coli and purified by affinity chromatography. The recombinant GSTs exhibited unique selectivity towards the general GST substrates 1‐chloro‐2,4‐dinitrobenzene (CDNB) and 1,2‐dichloro‐4‐nitrobenzene (DCNB), and also differed in their sensitivity to known inhibitors of GSTs. CpGSTD1 exhibited peroxidase activity with cumene hydroperoxide, while CpGSTD2 appeared to lack this activity. CpGSTD1 was able to metabolize 1,1,1‐trichloro‐2,2‐bis(4‐chlorophenyl)ethane (DDT), while DDT metabolism by CpGSTD2 was not detectable. CpGSTD1 and CpGSTD2 showed no detectable metabolism of permethrin. Gene expression of CpGSTd1 and CpGSTd2 in Marin mosquitoes was elevated about twofold in comparison with that found in a pyrethroid‐sensitive mosquito strain. CONCLUSION: The results indicate that CpGSTD1 and CpGSTD2 have unique biochemical characteristics, but they do not appear to play major roles in permethrin resistance in Marin mosquitoes. Copyright © 2012 Society of Chemical Industry     

Oral vaccination of foxes against rabies in Turkey between 2008 and 2010

Following a sustained spill-over event from dogs to foxes, fox rabies spread rapidly in the Aegean region, Turkey. In order to control the outbreak a program of oral vaccination of foxes against rabies was introduced. In the selected vaccination area three annual campaigns between 2008 and 2010 were undertaken during the winter months whereby the vaccine baits were distributed exclusively by plane using a density of 18 baits per km2. Subsequently, fox rabies cases were reported only from locations bordering the non-vaccinated areas. Hence, it was shown that fox rabies control by means of oral rabies vaccination is feasible in Turkey. However, for the progress towards the elimination of fox-mediated rabies in Turkey to be maintained, it is necessary that political and financial support is secured to extend oral vaccination where infected foxes remain.     

Retrospective study and review of ocular radiation side effects following external-beam Cobalt-60 radiation therapy in 37 dogs and 12 cats

This retrospective study evaluated the ocular side effects of cancer-bearing dogs and cats treated with external–beam Cobalt-60 (Co-60) radiation in which one or both orbit(s) were included in the radiation field. A total of 37 dogs and 12 cats presented to the Ontario Veterinary College during the 10-year study period (1999–2009) were evaluated. The radiation protocols ranged from a maximum of 60 Gray (Gy) in 24 fractions for curative intent to a minimum of 8 Gy in 1 fraction for palliative treatment. The main ocular side effect reported in both dogs and cats was conjunctivitis (79% and 55%, respectively). Other common ocular side effects included eyelid lesions in dogs (44%), ulcerative keratitis in cats (36%), and keratoconjunctivitis sicca in both dogs and cats (44% and 27%, respectively). The high incidence of ocular side effects in both patient populations indicates a need for regular ophthalmic examinations as a component of routine follow-up for radiation therapy involving the orbit. Radiation damage to ocular tissues is also reviewed.     

Effectiveness of vaccines and vaccination programs for the control of foot-and-mouth disease in Uganda, 2001–2010

Foot-and-mouth disease (FMD) is a highly contagious disease of cloven-hoofed animals. In Uganda, FMD outbreaks are mainly controlled by ring vaccination and restriction of animal movements. Vaccination stimulates immunity and prevents animals from developing clinical signs which include lameness, inappetence, and decreased production. Ring vaccination and restriction of animal movements have, however, not successfully controlled FMD in Uganda and outbreaks reoccur annually. The objective of this study was to review the use of FMD virus (FMDV) vaccines and assess the effectiveness of vaccination programs for controlling FMD in Uganda (2001–2010), using retrospective data. FMD vaccine distribution patterns in Uganda (2001–2010) matched occurrence of outbreaks with districts reporting the highest number of outbreaks also receiving the largest quantity of vaccines. This was possibly due to “fire brigade” response of vaccinating animals after outbreaks have been reported. On average, only 10.3 % of cattle within districts that reported outbreaks during the study period were vaccinated. The average minimum time between onset of outbreaks and vaccination was 7.5 weeks, while the annual cost of FMDV vaccines used ranged from US $58,000 to 1,088,820. Between 2001 and 2010, serotyping of FMD virus was done in only 9/121 FMD outbreaks, and there is no evidence that vaccine matching or vaccine potency tests have been done in Uganda. The probability of FMDV vaccine and outbreak mismatch, the delayed response to outbreaks through vaccination, and the high costs associated with importation of FMDV vaccines could be reduced if virus serotyping and subtyping as well as vaccine matching were regularly done, and the results were considered for vaccine manufacture.     

Characterization of turkey inducible nitric oxide synthase and identification of its expression in the intestinal epithelium following astrovirus infection

The inducible nitric oxide synthase (iNOS) enzyme has long been recognized as a key mediator of innate immune responses to infectious diseases across the phyla. Its role in killing or inactivating bacterial, parasitic, and viral pathogens has been documented in numerous host systems. iNOS, and its innate immune mediator NO has also been described to have negative consequence on host tissues as well; therefore understanding the pathogenesis of any infectious agent which induces iNOS expression requires a better understanding of the role iNOS and NO play in that disease. Previous studies in our laboratory and others have demonstrated evidence for increased levels of iNOS and activity of its innate immune mediator NO in the intestine of turkeys infected with astrovirus. To begin to characterize the role iNOS plays in the innate immune response to astrovirus infection, we identified, characterized, developed tkiNOS specific reagents, and demonstrated that the intestinal epithelial cells induce expression of iNOS following astrovirus infection. These data are the first to our knowledge to describe the tkiNOS gene, and demonstrate that astrovirus infection induces intestinal epithelial cells to express iNOS, suggesting these cells play a key role in the antiviral response to enteric infections.     

Marked, biased, filter (MBF): use of digital X-radiography and mark-recapture to partition seed lots based on sampled individual seed quality attributes

We report results of preliminary evaluations of marked, biased, filter (MBF), a method for both estimating and improving the effectiveness of separating conifer seeds. MBF integrates elements of X-radiographic seed quality assessment and mark-recapture methods from wildlife population sampling. Given a sample drawn from a population, individual seeds with attributes targeted for separation are labelled with a mark which biases the subsequent removal probability of other, non-marked seeds in the lot during grading and processing, after returning the sample to the population. Bias in the marking filter, in the form of extra mass or density added to seeds, minimizes the likelihood that uncertainty due to sampling error in the prior estimate will result in empty seeds remaining in the population during processing. In two experiments, a ponderosa pine seed lot was partitioned into empty and filled fractions with an air column separator. Powder dye talc was used for marking in the first trial, with low bias (<1%) and a small sample size (n = 150) with dry (2% moisture content) and soaked (26% moisture content) seed lots. A second trial conducted with two coats of spray paint used for marking and a larger sample size (n = 400) resulted in removal of 100% of non-filled seeds in the population. A simulation was conducted to evaluate sample statistics upon which to base the value of threshold bias, τ, below which seeds in an X-rayed sample should be marked to achieve consistent separation. Estimating the marking threshold based on quantiles of filled seeds most consistently resulted in removal of the non-filled fraction. MBF is a useful method for statistically linking one or more individual seed quality attributes extracted from X-ray image analysis with population-level processing of seed lots.     

Sulfometuron methyl influences seedling growth and leaf function of three conifer species

Seedling growth and gas exchange responses were measured for two potted seedling trials testing herbicide phytotoxicity to three important tree species of the Inland Northwest, USA. Media-filled pots were treated with sulfometuron methyl (Oust^?) in varying concentrations and planted with seedlings of Larix occidentalis Nutt., Pseudotsuga menziesii var. glauca (Beissn.) Franco, and Pinus monticola Dougl. ex D. Don. Seedlings were grown in two trials to determine the effects of two important residue breakdown factors, substrate moisture and pH, relative to that of herbicide application rate on seedling health. Changes in seedling height, root-collar diameter, and root volume were morphologic variables of interest, and physiological variables measured were net photosynthetic assimilation, stomatal conductance, and transpiration rate. While three levels of media moisture and four levels of media pH had no effect on seedling performance, most growth and leaf function variables were hindered across application rate treatments of all three species. Label-recommended dosages resulted in growth suppression levels potentially detrimental to seedling establishment for all three species. This was most pronounced for Pinus monticola, in particular for root growth, where untreated control seedlings showed 109% more root volume growth than treated seedlings. We conclude that when possible, a species-specific application rate might be found that balances the benefits of vegetation control with the phytotoxicity to promote optimal growth gains.     

In vitro evaluation of topical biocide and antimicrobial susceptibility of Staphylococcus pseudintermedius from dogs

Background – Staphylococcus pseudintermedius is an important canine pathogen, and the emergence and widespread dissemination of meticillin‐resistant strains (MRSP) is of significant concern. Multidrug‐resistant infections may require alternative approaches, such as the use of topical therapy. There is minimal information about the in vitro susceptibility of meticillin‐susceptible S. pseudintermedius (MSSP) and MRSP to biocides and topical antimicrobials. Hypothesis/Objectives – The hypothesis was that clinical isolates of MSSP and MRSP would not have universal susceptibility to topical biocides and antimicrobials. The goal of this study was to assess the susceptibility of a collection of S. pseudintermedius isolates to selected antimicrobials and biocides. Animals – The study was performed on clinical isolates of MSSP and MRSP from dogs with skin and soft tissue infections collected throughout North America between 2006 and 2008. Methods – The minimal inhibitory concentrations (MICs) of chlorhexidine digluconate, benzalkonium chloride, triclosan, accelerated hydrogen peroxide, geranium oil, tea tree oil and grapefruit seed extract were tested for 25 MRSP and 25 MSSP isolates from dogs using the agar dilution method. The MICs of fusidic acid, bacitracin and mupirocin were determined using Etests. Results – Triclosan demonstrated excellent activity against all bacterial isolates, with no growth at the lowest concentration evaluated (MIC ≤ 0.5 μg/mL). Conversely, grapefruit seed extract did not inhibit growth at the highest concentration tested (MIC > 3.84 μg/mL). All isolates were susceptible to mupirocin, fusidic acid and bacitracin. There were no significant differences noted in the range, MIC₅₀ or MIC₉₀ between MSSP and MRSP isolates. Conclusions and clinical importance – While isolates were susceptible to most of the tested compounds, universal susceptibility to all compounds with potential antimicrobial activity cannot be assumed, and specific testing is required.     

Stomatal response of an anisohydric grapevine cultivar to evaporative demand, available soil moisture and abscisic acid

Stomatal responsiveness to evaporative demand (air vapour pressure deficit (VPD)) ranges widely between species and cultivars, and mechanisms for stomatal control in response to VPD remain obscure. The interaction of irrigation and soil moisture with VPD on stomatal conductance is particularly difficult to predict, but nevertheless is critical to instantaneous transpiration and vulnerability to desiccation. Stomatal sensitivity to VPD and soil moisture was investigated in Semillon, an anisohydric Vitis vinifera L. variety whose leaf water potential (Ψ(l)) is frequently lower than that of other grapevine varieties grown under similar conditions in the warm grape-growing regions of Australia. A survey of Semillon vines across seven vineyards revealed that, regardless of irrigation treatment, midday Ψ(l) was dependent on not only soil moisture but VPD at the time of measurement. Predawn Ψ(l) was more closely correlated to not only soil moisture in dry vineyards but to night-time VPD in drip-irrigated vineyards, with incomplete rehydration during high night-time VPD. Daytime stomatal conductance was low only under severe plant water deficits, induced by extremes in dry soil. Stomatal response to VPD was inconsistent across irrigation regime; however, in an unirrigated vineyard, stomatal sensitivity to VPD-the magnitude of stomatal response to VPD-was heightened under dry soils. It was also found that stomatal sensitivity was proportional to the magnitude of stomatal conductance at a reference VPD of 1kPa. Exogenous abscisic acid (ABA) applied to roots of Semillon vines growing in a hydroponic system induced stomatal closure and, in field vines, petiole xylem sap ABA concentrations rose throughout the morning and were higher in vines with low Ψ(l). These data indicate that despite high stomatal conductance of this anisohydric variety when grown in medium to high soil moisture, increased concentrations of ABA as a result of very limited soil moisture may augment stomatal responsiveness to low VPD.