Ultrastructural findings in feline corneal sequestra

OBJECTIVES: (1) To describe the ultrastructural features of corneal sequestra in cats; and (2) to enhance our understanding regarding the pathogenesis of feline corneal sequestration. METHODS: Nine corneal sequestra were harvested via keratectomy from globes of nine cats. The sequestra were routinely fixed then postfixed for high resolution light and transmission electron microscopy (HR-LM and TEM, respectively). The tissues were embedded in Epon/Araldite. Sections of 0.5-microm thickness were cut and stained with 1% toluidine blue in 1% sodium tetraborate solution for HR-LM. Ultrathin sections were collected on copper grids and stained with uranyl acetate and Sato's lead stain for TEM. Ultrathin sections were examined and the images were captured on an Advantage HR CCD camera using a Hitachi 7500 electron microscope operated at 80 kV. Two healthy corneas from two cats were harvested immediately following euthanasia. These corneal tissues (control samples) were processed in the same manner as the corneal sequestra for HR-LM and TEM. A portion of each sequestrum was also submitted for polymerase chain reaction (PCR) testing for infectious agents including feline herpesvirus-1 (FHV-1), Toxoplasma gondii, Chlamydophila felis and Mycoplasma spp. RESULTS: Ultrastructure of healthy corneal tissues revealed basal corneal epithelial cells aligned adjacent to a thin acellular layer similar to Bowman's layer with underlying tightly packed, regularly arranged, collagen fibrils oriented in different planes. Keratocytes were elongated and had long and irregularly shaped nuclei, and cytoplasm contained rough endoplasmic reticulum and abundant membrane-bound vesicles. In contrast, corneal sequestra contained varying amounts of an amorphous, electron-dense substance, continuous with intact basal epithelial basement membranes peripherally, and overlying corneal ulceration and loosely packed collagen fibrils. Remnants of necrotic keratocytes were seen in spaces between disarranged collagen layers. In all samples, occasional keratocytes exhibited morphology indicative of apoptosis including clumping and margination of chromatin, and shrunken cytoplasm. Varying degrees of inflammation were noted on HR-LM and TEM of affected corneas including peri- and intralesional neutrophils, lymphocytes, plasma cells, and macrophages. Corneal sequestra were FHV-1-positive (n = 3), FHV-1- and T. gondii-positive (n = 1), T. gondii-positive (n = 3), or negative for DNA of these infectious agents (n = 2) using PCR. All corneal sequestra were negative for DNA of Chlamydophila felis and Mycoplasma spp. using PCR. CONCLUSIONS: Apoptosis may play a role in the pathogenesis of feline corneal sequestration independent of the presence of DNA of these infectious organisms. Prospective clinical studies are warranted to further understand the significance of T. gondii in relation to feline corneal sequestration.     

Possibility of vancomycin-resistant enterococci transmission from human to broilers, and possibility of using the vancomycin-resistant gram-positive cocci as a model in a screening study of vancomycin-resistant enterococci infection in the broiler chick

In this study, vancomycin‐resistant enterococci (VRE) from humans and vancomycin‐resistant gram‐positive cocci (VRPC) from pigs were examined for their ability to transmit in the chick intestine (Experiment 1). A model study on the spread speed of VRPC was also estimated from chick to chick under semi‐production conditions with different administration routes (not inoculated, oral administration to a chick, sprayed on the floor) (Experiment 2). Furthermore, the disappearance of VRPC from their litter with composting processes was examined (Experiment 3). Each of six chicks was inoculated with VRPC or VRE at 1 day old in Experiment 1. All the chicks had VRPC or VRE in their glandular stomach at 22 days of age. In Experiment 2, 6 floor pens covered with sawdust were prepared and 20 chicks were allotted to each pen. The chicks were inoculated with VRPC at 1 day of age. The VRPC were detected in each group in cloacal swabs at 2 days of age (detection rate; 20–80%). And they were also detected in the not‐inoculated group. The VRPC detection rate gradually decreased, and detection was rare (0–10%) in the packing chicks (50 days old). VRPC were detected in the litter of each pen in Experiment 2. A composting process was effectively used to eliminate VRPC by the 6th week (Experiment 3).     

Genetic and antigenic characterization of bovine viral diarrhea viruses isolated from cattle in Hokkaido,Japan

In our previous study, we genetically analyzed bovine viral diarrhea viruses (BVDVs) isolated from 2000 to 2006 in Japan and reported that subgenotype 1b viruses were predominant. In the present study, 766 BVDVs isolated from 2006 to 2014 in Hokkaido, Japan, were genetically analyzed to understand recent epidemics. Phylogenetic analysis based on nucleotide sequences of the 5′-untranslated region of viral genome revealed that 766 isolates were classified as genotype 1 (BVDV-1; 544 isolates) and genotype 2 (BVDV-2; 222). BVDV-1 isolates were further divided into BVDV-1a (93), 1b (371) and 1c (80) subgenotypes, and all BVDV-2 isolates were grouped into BVDV-2a subgenotype (222). Further comparative analysis was performed with BVDV-1a, 1b and 2a viruses isolated from 2001 to 2014. Phylogenetic analysis based on nucleotide sequences of the viral glycoprotein E2 gene, a major target of neutralizing antibodies, revealed that BVDV-1a, 1b and 2a isolates were further classified into several clusters. Cross-neutralization tests showed that BVDV-1b isolates were antigenically different from BVDV-1a isolates, and almost BVDV-1a, 1b and 2a isolates were antigenically similar among each subgenotype and each E2 cluster. Taken together, BVDV-1b viruses are still predominant, and BVDV-2a viruses have increased recently in Hokkaido, Japan. Field isolates of BVDV-1a, 1b and 2a show genetic diversity on the E2 gene with antigenic conservation among each subgenotype during the last 14 years.     

Inhibitory effects of SKF96365 on the activities of K+ channels in mouse small intestinal smooth muscle cells

In order to investigate the effects of {"type":"entrez-protein","attrs":{"text":"SKF96365","term_id":"1156357400","term_text":"SKF96365"}}SKF96365 (SKF), which is a non-selective cationic channel blocker, on K⁺ channel currents, we recorded currents through ATP sensitive K⁺ (I_KATP), voltage-gated K⁺ (I_Kv) and Ca²⁺ activated K⁺ channels (I_BK) in the absence and presence of SKF in single small intestinal myocytes of mice with patch-clamp techniques. SKF (10 µM) reversibly abolished I_KATP that was induced by cromakalim (10 µM), which is a selective ATP sensitive K⁺ channel opener. These inhibitory effects were induced in a concentration-dependent and voltage-independent manner. The 50% inhibitory concentration (IC₅₀) was 0.85 µM, which was obviously lower than that reported for the muscarinic cationic current. In addition, SKF (1 µM ≈ the IC₅₀ value in I_KATP suppression) reversibly inhibited the I_Kv that was induced by repetitive depolarizing pulses from −80 to 20 mV. However, the extent of the inhibitory effects was only ~30%. In contrast, SKF (1 µM) had no significant effects on spontaneous transient I_BK and caffeine-induced I_BK. These results indicated that SKF inhibited ATP sensitive K⁺ channels and voltage-gated K⁺ channels, with the ATP sensitive K⁺ channels being more sensitive than the voltage-gated K⁺ channels. These inhibitory effects on K⁺ channels should be considered when SKF is used as a cationic channel blocker.     

Effect of volume of non-surgical embryo transfer medium on ability of porcine embryos to survive to term

Non-surgical embryo transfer is a promising method for improving efficiency in the pork industry and also for biotechnology applications, such as in vitro embryo production, transgenesis and cloning. Several groups have reported successful piglet production using an artificial insemination catheter or flexible catheter designed for this procedure; however, the efficiency of the technique is still low. The critical points that need to be addressed in order to improve this procedure are (1) the embryo deposition site and (2) volume of transfer medium associated with the embryos; however, the latter has not yet been examined systematically. In the present study, we evaluated the effect of the volume of non-surgical embryo transfer medium on the ability of porcine embryos to survive to term by using a recently produced flexible catheter. The catheter consists of a guide and an injector. Blastocysts 200-230 mum in diameter were collected from donor gilts and transferred to recipient gilts. The time required for the completion of embryo transfer using this catheter was 14.6 +/- 3.9 min. The tip of the injector was determined by laparotomy to be located in a uterine horn 20-30 cm anterior from the branching point of the uterus body. We transferred 17.0-17.3 embryos with different volumes of medium (1.6, 3.2 and 10 ml) into each of 5, 4 and 4 recipients, respectively, and pregnancy was confirmed in 4, 3 and 1 of these recipients, respectively. Three recipients in the 1.6 ml group farrowed a total of 19 piglets (4, 5 and 10 piglets, respectively). These results suggest that successful non-surgical embryo transfer is affected by the volume of transfer medium.     

Rheological properties and structural changes in steamed and boiled abalone meat

ABSTRACT: Changes in tissue structures, rheological properties, and water content of abalone meat were studied in relation to boiling and steaming time. The adductor muscle of abalone Haliotis discus, which was removed directly from the shell, was boiled or steamed for 1 h, 2 h, and 3 h. When observed under a light microscope and by scanning electron microscopy, structural changes in the myofibrils were greatest in the boiled abalone meat compared with the steamed meat. When heating time was increased from 1 h to 3 h, the instantaneous modulus E ₀ of boiled abalone meat decreased gradually with increased heating time, whereas the E ₀ of steamed abalone meat was reduced when heated for 2 h. When heated for 1 h, the relaxation time of steamed abalone meat was much longer than that of boiled meat. There were no significant changes in the relaxation time of abalone meat among the different boiling times, but the relaxation time of steamed abalone meat was reduced gradually with increasing heating times. The study's results confirmed that the difference in rheological properties between the boiled and steamed meats was due mainly to the denaturation level of myofibrils when heated for 1 h, as well as due to the changes in water and solid content and the manner in which the inner water was exchanged after heating time was increased from 1 h to 3 h.     

Productivity of single-grip harvesters in clear-cutting operations in the northern European part of Russia

A field-based study was carried out to broaden our knowledge of fully mechanized cut-to-length harvesting productivity in naturally grown forests in the northern European part of Russia (NEPR). The recorded data comprised 38 midsized single-grip harvesters (JD 1270D) in clear-cutting operations in the Karelia, Komi, Vologda, Leningrad, Tver, and Kirov regions in NEPR, 4.3 million felled trees, and 1.4 million m³ u.b. (under bark) of processed timber. Harvesting operations were conducted in forest stands composed of spruce (48% on average), pine (19%), birch (22%), and aspen (11%), with an average stem volume 0.31 m³ u.b. The cut-to-length harvesters produced from 4.3 to 14.9 m³ u.b./productive machine hour (PMH) and 16.0–49.5 m³ u.b./stem processing machine hour (S _proc MH). A machine evaluation analysis and a regression analysis were used to formulate models for predicting cutting productivity of modern single-grip harvester. The regression models were developed to estimate the productivity of the harvesters in the regions taking into account two significant factors influencing the productivity: the stem volume and tree species of the felled trees. Productivity/cubic meter u.b. of processed timber/PMH was calculated according to stem volume and tree species distributions in most forest-covered NEPR regions. Further research is suggested to improve the developed productivity models and to allow prediction of system performance over a broad range of stand and site conditions.     

Relationships between internal ethylene and optical reflectance in ripening ‘Antonovka’ apples grown under sunlit and shaded conditions

The feasibility of non-destructive estimation of internal ethylene concentration (IEC) in apple fruit via fruit reflectance using recently developed approaches and a fiber-optics reflectometer was investigated. The relationships between IEC and fruit reflectance in the 400–800 nm range were studied in stored apple (Malus × domestica Borkh., cv. Antonovka) fruit. A strong correlation between IEC and optical reflectance spectra taken from sunlit surfaces of the fruit was detected whereas reflectance of the shaded fruit surface showed a weak correlation with IEC. The increase of the reflectance in the red occurred along with IEC build-up during ripening resulting a strong (r² > 0.80) correlation. By contrast, reflectance in the blue-green part of the spectrum remained low and was negatively (r² ≈ 0.65) correlated with IEC. These observations are consistent with the phenomenon of degradation of chlorophylls which often occurs in parallel with the retention of carotenoids in ripening apple skin. As a result, IEC showed a significant correlation (r² > 0.69; P < 0.001) with the index based on reflectances in the red and blue-green regions of the spectrum (R₆₇₈ ? R₄₈₀)/R₈₀₀. The effects of strong solar light on the relationships between IEC and fruit reflectance are considered. The possibilities and limitations of a non-destructive reflectance-based assay of IEC in apple fruit are discussed.     

Diverse antimicrobial killing by Enterococcus faecium E 50-52 bacteriocin

An effective bacteriocin was identified and characterized. Lactic acid bacteria were screened against Campylobacter jejuni. One bacteriocin producer, Enterococcus faecium (NRRL B-30746), was studied. The isolate was grown, and the bacteriocin was purified to single-band homogeneity. Biochemical traits indicated that the peptide was a Class IIa bacteriocin, and it was named E 50-52. The bacteriocin had a molecular weight of 3339.7 and an isoelectric point of 8.0. The minimal inhibitory concentrations of E 50-52 against C. jejuni, Yersinia spp., Salmonella spp., Escherichia coli O157:H7, Shigella dysenteriae, Morganella morganii, Staphylococcus spp., and Listeria spp. ranged from 0.025 to 32 microg/mL. In therapeutic broiler trials, oral treatment with E 50-52 reduced both C. jejuni and Salmonella enteritidis by more than 100,000-fold in the ceca, and systemic S. enteritidis was reduced in the liver and spleen. The wide range of antibacterial activity of bacteriocin E 50-52 against pathogens provides a promising alternative to antibiotics.     

Carbon and nutrient budgets of the Chagga home garden system in the Kilimanjaro highlands,Tanzania

Sustainable land management of smallholder farms is crucial for ensuring food security in sub-Saharan Africa. However, little is known about the nutrient dynamics of smallholder farming systems at the farm level based on primary data. In this study, carbon (C) and nutrient budgets of the home garden system in the Kilimanjaro highlands, where Andosols predominate, were quantified at the farmer's field. We evaluated (1) the soil C and nutrient flow in the main three land-use blocks (banana (Musa spp.) garden, maize (Zea mays L.) field and grassland) in one representative home garden and (2) the internal flow between farmland and livestock and the external nutrient flow across the inside and outside of the six home gardens. Intensive applications of livestock dung to the banana trees resulted in a positive C budget (7.2 Mg C ha⁻¹ year⁻¹) in the banana garden. Nitrogen loss through the harvesting of feed and crops was almost balanced with the livestock dung application, while nitrogen loss through leaching only accounted for 3% of that applied. Banana productivity has been maintained despite a negative potassium budget (−241 kg K ha⁻¹ year⁻¹), probably owing to the replenished exchangeable potassium from Andosols. In the maize field, the C budget was negative (−1.7 Mg C ha⁻¹ year⁻¹) owing to high organic matter decomposition. Carbon and nutrient budgets in the grassland were all negative. Our results revealed that the village average livestock density (4.4 TLU ha⁻¹: TLU means tropical livestock unit) was sufficient for P, Ca and Mg balance in the home garden, whereas it was not sufficient for N and K balance. Increasing livestock density improved the nutrient balance of the system. However, it is noteworthy that 33%–47% of the feed supplied as C and nutrients was collected from outside the home gardens, suggesting a high reliance on external inflow to fulfil feed demands. In conclusion, intensive livestock dung application to banana cultivation was fundamental for maintaining agricultural productivity to replenish the nutrients lost from the system. At the same time, this system was sustained not only by C and nutrient cycling within the system, but also by transporting resources from the external environment into the system.