In vitro markers for virulence in Yersinia ruckeri

In this study, different traits that have been associated with bacterial virulence were studied in Yersinia ruckeri. Two isolates that had been shown to cause disease and mortality in experimentally infected rainbow trout were compared with five avirulent isolates. Both virulent isolates showed high adhesion to gill and intestinal mucus of rainbow trout, whereas the majority of non‐virulent strains demonstrated significantly lower adhesion. A decrease in adherence capability following bacterial treatment with sodium metaperiodate and proteolytic enzymes suggested the involvement of carbohydrates and proteins. All strains were able to adhere to and invade chinook salmon embryo cell line (CHSE‐214), fathead minnow epithelial cell line (FHM) and rainbow trout liver cell line (R1). One non‐virulent strain was highly adhesive and invasive in the three cell lines, whereas the virulent strains showed moderate adhesive and invasive capacity. The internalization of several isolates was inhibited by colchicine and cytochalasin‐D, suggesting that microtubules and microfilaments play a role. For all strains, intracellular survival assays showed a decrease of viable bacteria in the cells 6 h after inoculation, suggesting that Y. ruckeri is not able to multiply or survive inside cultured cells. Analysis of the susceptibility to the bactericidal effect of rainbow trout serum demonstrated that virulent Y. ruckeri strains were serum resistant, whereas non‐virulent strains were generally serum sensitive.     

Conjunctival dacryops in a domestic shorthair cat

A 1-year-old spayed female domestic shorthair cat with a 1-week history of a conjunctival mass located in the dorsolateral quadrant of the right eye was presented to a referral hospital. Ophthalmic examination revealed a pink, fluctuant, and non-painful cystic swelling of the upper right palpebral conjunctiva. An ocular ultrasound confirmed the presence of a fluid-filled cystic structure that extended into the temporal orbit. The histopathology of the excisional biopsy revealed multiple cysts lined by ductal epithelium along with dilated lacrimal gland ducts and lymphocytic nodules adjacent to the dorsal lacrimal gland; consistent with a diagnosis of dacryops.     

Influence of amount and frequency of protein supplementation to ruminants consuming low-quality cool-season forages: efficiency of nitrogen utilization in lambs and performance of gestating beef cows

We evaluated the influence of amount and crude protein (CP) supplementation frequency (SF) on nitrogen (N) use by wethers and the performance of late-gestation beef cows. In exp. 1, seven Western whiteface wethers (31.8 ± 1.4 kg) were used in an incomplete 7 × 4 Latin square to evaluate intake and N use. Wethers received one of the seven treatments in a 2 × 3 factorial design containing two levels of supplemental soybean meal offered at a rate of 100% (F) or 50% (H; 50% of F) of the estimated CP requirement daily, once every 5, or once every 10 d, plus a non-supplemented control (CON). Low-quality cool-season forage (4.9 % CP; dry matter [DM] basis) was provided daily for ad libitum intake. Experimental periods lasted 30 d. In exp. 2, 84 Angus × Hereford cows (560 ± 35 kg) were stratified by age, body condition score (BCS), and expected calving date and allocated to 1 of the 21 feedlot pens (three pens per treatment). Pens were randomly assigned to receive the same treatments as in exp. 1 and cows had free access to low-quality cool-season forage (2.9% CP; DM basis). Cow body weight (BW) and BCS were measured every 14 d until calving and within 24 h after calving. In exp. 1, supplementation did not alter total DM and organic matter (OM) intake (P ≥ 0.26), but both parameters linearly decreased as SF decreased (P = 0.02). Supplementation increased DM, OM, and neutral detergent fiber (NDF) digestibility (P ≤ 0.02). Additionally, F feeding linearly increased DM, OM, and NDF digestibility as SF decreased (P ≤ 0.04). Digestibility of N, N balance, and digested N retained were greater with supplementation (P < 0.01), and N digestibility linearly increased as SF decreased (P = 0.01). Mean plasma urea-N concentration was not only greater (P < 0.01) for supplemented vs. CON wethers but also greater (P = 0.03) for F vs. H. In exp. 2, pre-calving BCS change was greater (P = 0.03) for supplemented cows. A linear effect of SF × supplementation rate for pre-calving BCS change was noted (P = 0.05), as F-supplemented cows lost more BCS compared with H as SF decreased. When considering supplementation intervals greater than 5 d, reducing the quantity of supplement provided, compared with daily supplementation, may be a feasible management strategy to maintain acceptable nutrient use and animal performance while reducing supplement and labor costs.     

Diffuse idiopathic skeletal hyperostosis of the spine in a nine‐year‐old cat

A nine‐year‐old intact female domestic shorthair cat was evaluated for paraparesis, ataxia and severe spinal hyperaesthesia. Neurological examination indicated a T3‐L3 spinal cord segment lesion. Computed tomography of the thoracolumbar and lumbosacral vertebral column was performed. This showed contiguous smooth new bone formation ventral and lateral to the vertebrae extending from the cranial thoracic area to the lumbosacral junction and appearing similar to canine diffuse idiopathic skeletal hyperostosis. There was also marked dorsolateral stenosis of the vertebral canal at the level of T4‐T5 because of degenerative changes of the facet joints. To the authors’ knowledge, this is the first published report of feline diffuse idiopathic skeletal hyperostosis.     

Tenogenic induction of equine mesenchymal stem cells by means of growth factors and low-level laser technology

Tendons regenerate poorly due to a dense extracellular matrix and low cellularity. Cellular therapies aim to improve tendon repair using mesenchymal stem cells and tenocytes; however, a current limitation is the low proliferative potential of tenocytes in cases of severe trauma. The purpose of this study was to develop a method useful in veterinary medicine to improve the differentiation of Peripheral Blood equine mesenchymal stem cells (PB-MSCs) into tenocytes. PB-MSCs were used to study the effects of the addition of some growth factors (GFs) as TGFβ3 (transforming growth factor), EGF2 (Epidermal growth factor), bFGF2 (Fibroblast growth factor) and IGF-1 (insulin-like growth factor) in presence or without Low Level Laser Technology (LLLT) on the mRNA expression levels of genes important in the tenogenic induction as Early Growth Response Protein-1 (EGR1), Tenascin (TNC) and Decorin (DCN). The singular addition of GFs did not show any influence on the mRNA expression of tenogenic genes whereas the specific combinations that arrested cell proliferation in favour of differentiation were the following: bFGF2 + TGFβ3 and bFGF2 + TGFβ3 + LLLT. Indeed, the supplement of bFGF2 and TGFβ3 significantly upregulated the expression of Early Growth Response Protein-1 and Decorin, while the use of LLLT induced a significant increase of Tenascin C levels. In conclusion, the present study might furnish significant suggestions for developing an efficient approach for tenocyte induction since the external administration of bFGF2 and TGFβ3, along with LLLT, influences the differentiation of PB-MSCs towards the tenogenic fate.     

Central nervous system signs in chickens caused by a new avian reovirus strain: a pathogenesis study

The present study describes the pathogenesis of infection of chicks with a new avian reovirus strain, belonging to the so-called enteric reovirus strains (ERS) that is capable of causing central nervous system signs in SPF white leghorns. After intramuscular (IM) or oral inoculation birds were either observed for clinical signs or sacrificed for macroscopic, histological and virological examination for 21 days. Virus isolation was performed on the brain, leg muscle, hock joint, liver and spleen. For the detection of viral antigen the immunohistochemistry (IHC) technique was performed on the caudal part of the cerebrum, spinal cord including spinal ganglia and right N. Ischiadicus. High mortality (79% in 7 days) was seen in birds that were inoculated IM. Survivors were depressed and stayed small until the end of the experiment. One bird had tremor and showed torticollis at 9 days after IM inoculation. Birds that were inoculated orally were depressed from day 4 and stayed small until the end of the experiment. One bird showed a torticollis at 10 days after inoculation. After both IM and oral inoculation ERS was isolated from the brain between 3 and 10 days after inoculation. Other examined organs were positive for virus isolation from day 1 or 5 until day 21. IHC revealed viral antigen positive cells in the Plexus chorioideus (plexus epithelial cells or cells within the underlying connective tissue) and in a spinal ganglion. The results indicate that the pathogenesis of ERS infection in chickens bears some resemblance with that of the mammalian reoviruses serotype 1 in mice.     

The Exploitation of Nematode-Responsive Plant Genes in Novel Nematode Control Methods

The molecular interactions between plants and sedentary nematodes are undergoing intense study, not only for reasons of fundamental research but also for the potential benefits to agriculture. The present technology allows the transformation of an increasing number of crop plants, providing new ways to introduce resistance against plant-parasitic nematodes. The ability of sedentary nematodes to induce specialized feeding sites in plant roots is one of the most fascinating aspects of this host–parasite interaction. Molecular approaches have been initiated to identify and characterize plant genes altered in expression after infection by sedentary nematodes. The results obtained indicate that many genes indeed become up-regulated upon nematode infection. Surprisingly, several so-called constitutive promoters that are normally used to achieve high expression in plant cells are completely ‘silenced’ in the feeding sites within days after nematode infection. Generally, there are two options available for the genetic engineering of nematode resistance: the synthesis of anti-nematode proteins or the localized production of a cytotoxic protein that interferes with the development of feeding cells. Nematode-induced promoters are very useful for the production by plants of sufficiently high levels of anti-nematode proteins at feeding sites. Alternatively, interfering with feeding-cell development is somewhat similar to the hypersensitive response evoked by nematodes in a naturally resistant plant. Here, destruction of specific plant cells can be achieved by the localized expression of a cytotoxin such as barnase, a potent ribonuclease. This approach, however, calls for a highly specific ‘non-leaky’ promoter, which is active only in the feeding cells. Another possibility is to use a two-component system, where the leakiness of the promoter in other tissues is counterbalanced by the constitutive expression of a neutralizing gene.     

Agrobacterium-mediated transformation of the CrDAT gene and selection of transgenic periwinkle lines with a high vincristine accumulation

Catharanthus roseus contains vincristine and vinblastine, which are outstanding drugs for cancer. In the biosynthetic pathways of terpenoid indole alkaloids (TIAs) in C. roseus, deacetylvindoline 4-O-acetyltransferase (DAT) is a key enzyme that catalyses the last reaction of vindoline biosynthesis to form vinblastine and vincristine. In this study, the CrDAT transgene was transferred into the periwinkle by Agrobacterium-mediated transformation and generated transgenic periwinkle lines with an increase in vincristine accumulation. The C. roseus DAT gene was introduced into C. roseus plants and it was confirmed that CrDAT was successfully transferred into the genome of periwinkle plants and efficiently translated to synthesise recombinant DAT protein. Four transgenic periwinkle lines in T1 generation, T1-1, T1-3, T1-6, and T1-7, expressed recombinant DAT protein with the total protein content in the range of 2.86 μg.mg^?1 to 5.12 μg.mg^?1. Moreover, the vincristine contents of four transgenic lines increased by 1.63?2.48-fold compared to non-transgenic plants, ranging from 6.91 µg.g^?1 (fresh weight) to 10.53 µg.g^?1 (fresh weight). The T1-1 line had the highest vincristine content. Hence, the overexpression of the recombinant DAT protein can improve the vincristine accumulation of transgenic C. roseus plants.

Abbreviation: CrDAT - Catharanthus roseus Deacetylvindoline-4-O-Acetyl Transferase; D4H - Deacetoxyvindoline 4-hydroxylase; ELISA - Enzyme-Linked Immunosorbent Assay Monoterpene indole alkaloid; T0, T1 - Generations of transgenic plants; TIAs - Terpenoid indole alkaloids; WT- The wild-type tobacco plants (non transgenic plant); 35S - Cauliflower mosaic virus 35S promoter     

Ketoprofen and phenylbutazone attenuation of PAF-induced lung inflammation in calves

The purposes of this study were: (1) to investigate which arachidonic acid metabolites contributed to platelet-activating factor (PAF) induced pulmonary dysfunction; and (2) to compare the effect of two non-steroidal anti-inflammatory drugs, phenylbutazone and ketoprofen in a model of PAF-induced reversible lung inflammation in six calves. In placebo and phenylbutazone groups, PAF infusion induced significant dysfunctions in the pattern of breathing, mechanics of breathing and gas exchange. These dysfunctions were prevented by ketoprofen pretreatment, except for the mechanics of breathing which was moderately but significantly altered by the PAF challenge. In all calves, leukotriene (LT) B4 plasma concentrations did not significantly increase above baseline values at any time. Prostaglandin (PG) E2 plasma concentrations showed a minor significant increase in phenylbutazone pretreated calves (55.8 +/- 25.8 pg/mL from 36.7 +/- 16.13 pg/mL). Thromboxane (TX) B2 plasma concentration was significantly increased during PAF challenge in placebo- and phenylbutazone-pretreated groups, but not in ketoprofen-pretreated calves (1580.0 +/- 1370 from 42.7 +/- 10.7 pg/mL; 2340 +/- 477 from 63 +/- 32 pg/mL; and 36.5 +/- 4.12 from 39.3 +/- 12.0 pg/mL, respectively). These data suggest that TXA2 is an important cyclooxygenase metabolite of arachidonic acid produced in response to PAF and that ketoprofen (intramuscular injection, 3 mg/kg) is more effective than phenylbutazone (intramuscular injection, 10 mg/kg) in preventing respiratory dysfunctions induced by the PAF challenge 30 min after drug administration. Ketoprofen did not suppress totally the PAF-induced changes in mechanics of breathing, which suggests that PAF or a secondary release of mediators could have a direct action on airway smooth muscle.     

Assessment of a bovine viral diarrhea virus antigen capture ELISA and a microtiter virus isolation ELISA using pooled ear notch and serum samples.

To reduce the cost of whole herd screening for bovine viral diarrhea virus persistently infected animals, the sensitivity and specificity of an antigen-capture enzyme-linked immunosorbent assay (AC-ELISA) and a microtiter virus isolation ELISA using saline from ear notch samples or pooled serum was determined. Pooled saline from ear notch samples, assayed by AC-ELISA, gave a sensitivity and specificity of 98% and 94%, respectively, for pools containing 2 samples and 72% and 100%, respectively, for pools of 5. The sensitivity of pooled ear notch or serum samples for bovine viral diarrhea virus detection by microtiter virus isolation (sensitivity < 5%) or serum samples for detection by AC-ELISA (sensitivity < 15%) is too low to be used for whole herd screening. Pooling saline from ear notch samples from 2 animals tested by AC-ELISA, however, could provide a less expensive, reliable method for whole herd screening for bovine viral diarrhea virus.