Development of a nitrocellulose-enzyme immunoassay for the detection of yellow-head virus from penaeid shrimp

A nitrocellulose-enzyme immunoassay (NC-EIA) employing horseradish peroxidase (HRP) was developed for the detection of yellow-head virus in the tissues of infected penaeid shrimp. Two substrates, DAB (3,3¢-diaminobenzidine retrahydrochloride) and TMB (3,3¢5,5¢-tetramethylbenzidine) for HRP were evaluated. The TMB was found to produce a more distinct and easily recognizable colour product and provided enhanced sensitivity (0.4 ng for TMB versus 0.8 ng viral protein for DAB). Under the present format, the NC-EIA was capable of detecting as little as 100 TCID₅₀ of virus in infected tissues. Uninfected tissue homogenates yielded no significant background colouration.     

Natriuretic peptide inhibition of intestinal salt absorption in the Japanese eel: physiological significance

Electrophysiological studies in vitro demonstrated the significant inhibition by natriuretic peptides (NP) of short-circuit current across the eel intestine, an important osmoregulatory organ. Inhibitory potencies of several members of the NP family were assessed by voltage-clamp determination of net transepithelial salt absorption measured as the short-circuit current I_sc across the intestine of the freshwater-adapted (FW) and seawater-adapted (SW) Japanese eel (Anguilla japonica); the order of potency of synthetic eel peptides was: amidated atrial natriuretic peptide (ANP-NH₂) > ventricular natriuretic peptide (VNP) > atrial natriuretic peptide (ANP) >> C-type natriuretic peptide (CNP). Neither the order of potency nor the absolute potencies were effected by salinity adaptation. The observed potency sequence suggests that inhibition of intestinal absorption is mediated by A-type guanylyl cyclase-coupled NP receptors. The relatively low sensitivity of the intestinal response to NP compared with circulating NP concentrations suggests a role for intestinal regulation by NP which is independent of systemic delivery from cardiac sources. A novel model, incorporating the known immunohistochemical localization of NP-ergic cells and processes in the epithelial layer of the intestine and the dissipation of the Na⁺ electrochemical gradient along the alimentary tract, is developed in which local secretion of NP (in response to a bolus of food) inhibits salt absorption across the intestine regionally in favor of increased nutrient absorption.     

Effect of water temperature and chlorophyll abundance on shell growth of the Japanese pearl oyster, Pinctada fucata martensii, in suspended culture at different depths and sites

To understand the relationships between shell growth and some environmental factors, we examined the relationships between water temperature or chlorophyll abundance and the shell growth of the Japanese pearl oyster, Pinctada fucata martensii, suspended at three different depths at two sites. Growth in height, length and thickness of the shells were limited by water temperature during winter (< 20 °C), whereas growth in thickness correlated with food abundance, measured as chlorophyll, during early summer (> 20 °C). These results suggest that the shell of P. fucata martensii could grow well at locations with greater abundance of food and adequate water temperatures (20–26 °C), resulting in a longer growing season.     

Flumequine depletion from muscle plus skin tissue of gilthead seabream (Sparus aurata L.) fed flumequine medicated feed in seawater at 18 and 24 °C

We examined flumequine depletion from muscle plus skin of gilthead seabream held in seawater at 18 and 24 °C. Seven groups of 10 fish each were sampled at intervals ranging from 24 to 168 h after in-feed administration of flumequine at 35 mg/kg/day for 5 days. Muscle plus skin tissue samples were analyzed for flumequine by high-performance liquid chromatography and fluorescence detection (HPLC-SFD). Parent flumequine concentrations declined rapidly from muscle plus skin after dosing with elimination half-lives of t_1/2=22.14 and 21.43 h at 18 and 24 °C, respectively. Withdrawal periods for the maximum residue limit (MRL) of 600 μg/kg flumequine in muscle plus skin at 95% tolerance limit were 106.08 and 75.84 h at 18 and 24 °C, respectively, after treatment.     

The association between gall bladder mucoceles and hyperlipidaemia in dogs: A retrospective case control study

The diagnosis of gall bladder mucoceles (GM) in dogs has become increasingly frequent in veterinary medicine. Primary breed-specific hyperlipidaemia is reported in Shetland Sheepdogs and Miniature Schnauzers, breeds in which GM are known to occur more frequently than in other breeds. The objective of this study was to evaluate the association between GM and hyperlipidaemia in dogs. The study design was a retrospective case control study. Medical records of dogs diagnosed with GM at the Veterinary Medical Centre of The University of Tokyo between 1 April 2007 and 31 March 2012, were reviewed.Fifty-eight dogs with GM and a record of either serum cholesterol, triglyceride, or glucose concentrations were included in the study. Hypercholesterolaemia (15/37 cases; odds ratio [OR]: 2.92; 95% confidence interval [CI]: 1.02–8.36) and hypertriglyceridaemia (13/24 cases; OR: 3.55; 95% CI:1.12–15.91) showed significant association with GM. Pomeranians (OR: 10.69), American Cocker Spaniels (OR: 8.94), Shetland Sheepdogs (OR: 6.21), Miniature Schnauzers (OR: 5.23), and Chihuahuas (OR: 3.06) were significantly predisposed to GM. Thirty-nine out of 58 cases had at least one concurrent disease, including pancreatitis (five cases), hyperadrenocorticism (two cases), and hypothyroidism (two cases). A significant association between GM and hyperlipidaemia was confirmed, suggesting that hyperlipidaemia may play a role in the pathogenesis of GM.     

Effect of increasing levels of seven tree species extracts added to a high concentrate diet on in vitro rumen gas output

This study was conducted to investigate the effects of increasing levels of extracts of Byrsonima crassifolia, Celtis pallida, Enterolobium cyclocarpum, Fraxinus excelsior, Ficus trigonata, Phoradendrom brevifolium and Prunus domestica on in vitro gas production (GP) and ruminal fermentation of a high concentrate diet. Plant extracts were prepared at 1 g dry matter (DM)/8 mL of solvent mixture (methanol : ethanol : water, 1:1:8) and added at levels of 0, 0.6, 1.2 and 1.8 mL/g DM of a high concentrate diet. In vitro GP was recorded at 2, 4, 6, 8, 10, 12, 24, 48 and 72 h of incubation. Increasing addition of extracts linearly increased (P < 0.001), the GP₂₄, GP₄₈ and GP₇₂ (mL/g DM), and linearly decreased (P < 0.001), the discrete GP lag time. Moreover, increasing extract doses linearly increased (P < 0.001) the asymptotic GP and decreased (P < 0.001) the rate of GP. GP₆ was not impacted by treatments and GP₁₂ increased linearly (P = 0.01) with increasing addition of extracts. Rumen pH declined linearly (P < 0.05) with increasing doses of extracts added. As no interactions (P > 0.05) occurred between the extracts and doses, it could be conclude that all extracts positively modified rumen fermentation at doses of 1.2 to 1.8 mL extract/g diet DM.     

Development and characterization of monoclonal antibodies to subgroup A avian leukosis virus

Avian leukosis virus subgroup A (ALV‐A) is a retrovirus which infects egg‐type chickens and is the main pathogen of lymphoid leukosis (LL) and myeloid leukosis (ML). In order to greatly enhance the diagnosis and treatment of clinical avian leukemia, two monoclonal antibodies (MAbs) to ALV‐A were developed by fusion between SP2/0 and spleen cells from mice immunized with expressed ALV‐A env‐gp85 protein. Using immunofluorescence assay (IFA), two MAbs reacted with ALV‐A, but not with subgroups B and J of ALV. Western blot tests showed that molecular weight of ALV‐A envelope glycoprotein recognized by MAbs was about 53 kD. Isotyping test revealed that two MAbs (A5C1 and A4C8) were IgG1 isotypes. These MAbs can be used for diagnosis and epidemiology of ALV‐A.     

Extra small virus-like particles (XSV) and nodavirus associated with whitish muscle disease in the giant freshwater prawn, Macrobrachium rosenbergii

A disease of Macrobrachium rosenbergii, the giant freshwater prawn, farmed in China was recently recorded in Zhejiang, Jiangsu, Shanghai, Guangxi and Guangdong provinces. The clinical sign of the disease, which develops in post-larvae (PL), is a whitish appearance of the muscles, particularly noticeable in the abdomen. Mortalities may reach 100% in some hatcheries. Investigations by transmission electron microscopy after negative staining of diseased PL homogenates showed the presence of two types of viral particles: one, unenveloped, icosahedral in shape, 26-27 nm in diameter, the second, much smaller, about 14-16 nm in diameter, designated extra small virus particle (XSV). The large virus has a genome with two pieces of ssRNA (RNA-1 and RNA-2), of 3 and 1.2 kb, respectively. Hybridization tests confirmed that this large virus is closely related to M. rosenbergii nodavirus (MrNV) which was isolated from diseased prawns in a hatchery in the French West Indies. Its very small size and hypothesized biochemical and biological characteristics suggest XSV is a new type of crustacean virus. As XSV has always been found associated with the larger virus (nodavirus) and is located in muscle and connective cells of diseased animals, it could be an autonomous virus, a helper-type virus or a satellite-like virus.     

Immunohistochemical detection of gonadotropin-like material in the hagfish pituitary, correlated with their gonadal conditions

In the present study, immunoreactive gonadotropin (GTH)-like material was examined in the hagfish pituitary, in correlation with their gonadal conditions. For this purpose, three species of hagfish, Myxine glutinosa, Eptatretus burgeri and Paramyxine atami, were used. In all species, GTH-like cells were observed in the adenohypophysis. They were stained with several antibodies to LH-related GTHs, such as ovine LHβ, human LHβ, bullfrog LH, and salmon GTHIIβ. In the subsequent study, the relationship between the amount of GTH-like material and gonadal conditions was examined in Paramyxine atami. The adenohypophysis of adult animals exhibited well-developing features, and about half of the adenohypophysial cells were stained intensely with anti-ovine LHβ. On the other hand, the adenohypophysis of juveniles exhibited less developed features with few or no cells stained with anti-ovine LHβ. These results suggest that a GTH-gonadal system is truly present in the hagfish.     

Transgenic cell lines which stably express progestogen receptors (PRS) α and β and the PR-responsive reporter genes

Transgenic cell lines which stably express progestogen receptors (PRs) and the PR-responsive reporter genes were developed. They are a good system for rapid, sensitive and reproducible screening of various ligands.