Chemiluminescence and chemotaxis assay of canine granulocytes: a methodological study

Chemiluminescence (CL) of isolated granulocytes and of whole blood from dogs was evaluated. Chemiluminescence of whole blood samples created an undesired quenching effect by the red blood cells which makes the assay difficult to apply in pathological cases with low formation of oxygen metabolites. This problem was avoided when chemiluminescence was determined, using isolated granulocytes. A cell concentration of 5 x 10(9)/l was needed to create optimal conditions. The Boyden chamber technique was used for study of random migration and chemotaxis. Casein (0.1%), zymosan activated serum with and without epsilon-amino-n-caproic-acid and homologous serum were effective chemoattractants for canine granulocytes, while FMLP (formyl-methionyl-leucyl-phenylalanin) did not attract canine granulocytes.     

畜牧科技论文中数据资料的检验判断方法初探

为便于判断科技论文中统计资料差异显著性标示的正误,基于论文中提供的样本大小、平均数和标准差3项信息,遵循统计检验的基本理论,提出了差比系数(均数差/标准差)的概念,并编制出最小显著差比系数表。利用该表可以快速准确地对论文中经t检验和方差分析多重检验的一些统计结果进行检验判断,纠正在科技论文资料统计过程中偶尔发生的错误。     

In vivo and in vitro effects of three glucocorticoids on blood leukocyte chemotaxis in the dog

The effects of three glucocorticoids on random migration (RM) and oriented migration (OM) of dog blood leukocytes either from dogs treated in vivo (at therapeutic dosage regimen) or leukocytes treated in vitro (at pharmacological concentrations), were investigated using an agarose gel technique. After in vivo treatment, methylprednisolone sodium succinate (MPSS) produced a stimulation of both RM and OM in the three treated dogs. Dexamethasone produced a stimulation of both RM and OM in the three treated dogs. Dexamethasone produced a stimulation of these parameters in all but one of the three treated dogs, but the difference was not statistically significant. After in vitro treatment of leukocytes from eight dogs, MPSS significantly stimulated OM. Dexamethasone was without significant effect except at a higher than therapeutic concentration (0.5 micrograms/ml), for which RM was stimulated. Hydrocortisone sodium succinate was without statistically significant effect. Under no conditions, except after suprapharmacological concentrations, did glucocorticoids inhibit RM or OM.     

Counting coccidial oocysts in chicken faeces: a comparative study of a standard McMaster technique and a new rapid method

For the assessment of coccidial oocyst production by chickens, some modified form of the McMaster counting method is commonly used. The objective of this study was to evaluate a standard method and to compare it to a new, faster method, in which all the preparative stages before counting are carried out in the same container into which the original faecal sample was collected. A stock suspension containing purified oocysts of all seven valid Eimeria species that parasitize chickens was prepared, from which seven concentrations of oocyst suspensions were made. Since the faecal material in a sample influences the ability of oocysts to float up in a McMaster chamber, the new method was tested to establish the optimal amount of faeces in the original sample. Control oocyst suspensions containing no faeces were also tested, and three series of counts using the new method were compared with the standard McMaster method. The results were statistically analysed by agreement analysis. Repeatability and between-operator variation of both methods were also tested by agreement analysis. Counting by the standard McMaster method underestimated the true number of oocysts. The new method gave counts in agreement with the true number of oocysts if using 1 g of faeces per sample. With 2 g of faeces, counts were obtained that agreed with counts by the standard McMaster method. Both methods showed agreement between repeated measurements. The new method used on a sample containing 2 g of faeces provides a convenient alternative to the standard modified McMaster method. A 1-g faecal sample increases agreement with the true numbers of oocysts. Processing of a sample with the new method is about nine times faster than with the standard method.     

Bile agar migration test: a novel method for testing the viability of Dictyocaulus viviparus larvae in lungworm vaccine

A bile agar migration (BAM) test was used to determine the viability of Dictyocaulus viviparus larvae in lungworm vaccines. Percentage migration values for 10 batches of vaccine varied from 8.8 to 33.2 per cent compared with 99 per cent of unirradiated larvae tested under similar conditions. On the other hand a conventional viability count (CVC), based on subjective assessment of larval shape, gave figures of 95.1 to 98.7 per cent for the 10 vaccine batches. The BAM test may therefore have potential for providing a more discriminatory assessment of vaccine quality than CVC.     

Grass awn migration in dogs and cats: a retrospective study of 182 cases

A retrospective study of 182 cases of grass awn migration in dogs and cats seen during a 1-year period was performed. The 182 cases comprised 61% of all foreign body-related cases during that year. Compared with the total hospital population, there was an increased prevalence of grass awn problems in the Springer Spaniel, Golden Retriever, Brittany Spaniel, and Airedale Terrier, but a decreased prevalence in German Shepherd Dogs, Miniature Poodles, and Dachshunds. The most common site of grass awn localization was the external ear canal, involving 51% of grass awn cases. Other common sites of grass awn localization included the interdigital webs, eye, nose, lumbar area, and thoracic cavity. Only 8 of the 182 animals were cats and 7 of the 8 had ocular involvement.     

Ex vivo viability of canine and feline sarcomas: a pilot study

Assay-based chemotherapeutic protocols are common in human gynecologic oncology, most notably for patients with ovarian or breast cancer. The current study examines ex vivo incubation conditions necessary for the assessment of sarcomatous tumor response to potential chemotherapeutic drugs. Slices of sarcomatous tumors were incubated in one of two culture media. Viability indices were measured and compared across time and between media. Neither medium was sufficient to support the growth of sarcomatous tumor tissue slices based on the indices studied. It is likely that sarcomatous tumors require a different approach for ex vivo assessment than their epithelial counterparts. Our long-term goal is to incubate tumor slices with chemotherapeutic agents to predict the in vivo tumor response based on the maintenance or loss of slice viability within this system.     

Challenges in estimating ancestral state reconstructions: the evolution of migration in Sylvia warblers as a study case

Our current understanding of how species have evolved is mainly based on comparative phylogenetic methods, which use phylogenies to infer the evolution of traits. The development of ancestral state reconstruction (ASR) methods has provided the tools to reconstruct trait evolution, which are widely used in fields like evolutionary biology, macroecology and paleontology. As there are different elements involved in those analyses, with different levels of uncertainty (i.e. relating to branch length estimation, trait coding, statistical framework, taxon sampling or software), the various combinations of these elements likely have a strong impact on the reconstruction of the evolution of traits, potentially leading to opposite conclusions. To assess the impact of these different elements in ASR, we performed a set of analyses, including all possible combinations of such elements and using the evolution of migratory behavior in Sylvia warblers as a case study, which was coded as a continuous or as a discrete character. Our results show that taxon sampling, character coding, tree shape, statistical framework and software all significantly affect ASR, both individually and in combination. Not all reconstructed tree nodes show discrepancies, but in the critical ones most pairwise comparisons between analyses lead to conflicting and unexpectedly antagonistic results (zero migration vs fully migratory), thus challenging interpretations of trait evolution. We propose some possible solutions to partly inform decisions, involving the method selection and the incorporation of biological or fossil evidence regarding how traits evolve, but our results demand serious rethinking about how the research community currently uses ASR.     

A new method for bovine embryo production: a potential alternative to superovulation

Eight cows were used to study the feasibility of transvaginal ultrasound-guided puncture of follicles as a method for the collection of immature oocytes for embryo production in vitro. In six trials at intervals of seven days, 104 oocytes were collected. After in vitro maturation and fertilisation the 104 oocytes were transferred to the oviducts of sheep. Six days later, 75 oocytes were recovered by flushing the oviducts. Twenty-four per cent of the recovered oocytes/embryos had developed into transferable and viable morulae and, or, blastocysts. The data show that this non-surgical and repeated collection of immature oocytes can be used successfully for the in vitro production of bovine embryos. The procedure may produce yields of embryos comparable to those obtainable by conventional superovulation procedures.     

Embryo transfer in pigs: a method for introducing genetic material into primary specific-pathogen-free herds.

Fifty-five embryos were transferred from 4 Duroc donor sows into 4 crossbred recipient gilts. The embryo survival rate was 66%; 53% of eggs ovulated by donors were represented by live pigs at term. Three of the litters were obtained by hysterectomy of the recipients and introduced into a primary specific-pathogen-free herd, thereby saving the donor sows for subsequent use. Embryo transfer provided a method for exploiting the genetic potential of superior females.     

Evaluation of a new in-clinic method for the detection of canine heartworm antigen

Canine heartworm is endemic in many parts of the world, and veterinarians rely on rapid in-clinic antigen tests to screen for this infection. Recently, an in-clinic, instrument-based rotor employing a colloidal gold agglutination immunoassay was launched in the marketplace (VetScan VS2(?) Canine Heartworm (HW) Antigen Test Kit; Abaxis, Inc.). Because of the widespread use of heartworm prevention and possible false negative test results in dogs with low heartworm burdens, the performance of the VetScan VS2(?) HW test and a commercially available in-clinic, membrane-based ELISA test (SNAP(?) Heartworm RT Test; IDEXX Laboratories) was compared using samples from dogs with low heartworm burdens and/or low levels of circulating antigen. Ninety serum samples were evaluated using the two methods. Testing was performed according to the manufacturer's product insert by personnel blinded to sample status. The samples were derived from two populations: dogs with necropsy-confirmed heartworm status (40 with 1-4 female worms, 30 with no worms), and field dogs (20) confirmed positive for antigen by microtiter plate ELISA (PetChek(?) Heartworm PF Antigen Test; IDEXX Laboratories). All 40 dogs with heartworms on necropsy were also confirmed to have circulating antigen by the PetChek HW ELISA. In necropsy-negative dogs (n=30), neither the VetScan VS2 HW nor SNAP HW tests detected heartworm antigen. Of the samples testing positive for antigen by PetChek HW (n=60), the VetScan VS2 HW and SNAP HW tests detected antigen in 15 and 56 samples, respectively. Percent agreement (plus 95% confidence interval) for each test relative to the PetChek HW qualitative result was 50% (40-60%) for VetScan VS2 HW and 96% (89-98%) for SNAP HW. Relative to the presence or absence of female worms at necropsy, agreement was 61% (50-72%) for VetScan VS2 HW and 99% (92-99.6%) for SNAP HW tests. It is clinically important that dogs with low heartworm burdens and/or low levels of circulating heartworm antigen be correctly identified by veterinarians in order to ensure prompt treatment, and the VetScan(?) VS2 HW test does not appear to be as accurate as the SNAP HW or PetChek HW tests when performed on this subset of patients.     

Negative staining method with nigrosin for the detection of cryptosporidial oocysts: a comparative study

A comparison was made between two methods for the detection of cryptosporidial oocysts in human faecal sediments of formalin-ether concentrates: a cover-slipped, wet method (containing Gram's iodine) and an air-dried, negative staining method using nigrosin solution. A modified Ziehl-Neelsen technique was used as a reference method. The negative staining method with nigrosin gave a positive diagnosis more often than the cover-slipped wet method. A comparison of the nigrosin method with the modified Ziehl-Neelsen method gave almost identical results. Restaining the nigrosin slides with the modified Ziehl-Neelsen technique showed that the round, refractile bodies were cryptosporidial occysts.     

In vitro and ex vivo inhibition of COX isoforms by robenacoxib in the cat: a comparative study

Schmid, V.B., Seewald, W., Lees, P., King, J.N. In vitro and ex vivo inhibition of COX isoforms by robenacoxib in the cat: a comparative study. J. vet. Pharmacol. Therap. 33 , 444–452. Robenacoxib is a novel nonsteroidal anti‐inflammatory drug (NSAID) developed for use in companion animals. Whole blood assays were used to characterize in the cat the pharmacodynamics of robenacoxib for inhibition of the cyclooxygenase (COX) isoforms, COX‐1 and COX‐2, in comparison with other NSAIDs. Based on in vitro IC₅₀COX‐1:IC₅₀COX‐2 ratios, robenacoxib was COX‐2 selective (ratio = 32.2), diclofenac (ratio = 3.9) and meloxicam (ratio = 2.7) were only weakly COX‐2 preferential, and ketoprofen (ratio = 0.049) was COX‐1 selective. In an in vivo pharmacokinetic ex vivo pharmacodynamic study, after both p.o. (1–2 mg/kg) and subcutaneous (2 mg/kg) dosing, robenacoxib achieved peak blood concentrations rapidly (T_max = 1 h for both administration routes) and was cleared from blood relatively rapidly (mean residence time was 1.70 h after p.o. and 1.79 h after subcutaneous dosing). In ex vivo COX isoform inhibition assays, orally (1–2 mg/kg) or subcutaneously (2 mg/kg) administered robenacoxib significantly inhibited COX‐2, with a relatively short duration of action in the central compartment, and had no effect on COX‐1. Therefore robenacoxib was COX‐2 selective and spared COX‐1 in vivo. In contrast, meloxicam (0.3 mg/kg via subcutaneous injection) inhibited both COX‐1 and COX‐2 isoforms significantly for at least 24 h, indicating nonselectivity in vivo.