Characterization of porcine circovirus type 2 (PCV2) infection in swine lymphocytes using mitogen-stimulated peripheral blood lymphocytes from healthy PCV2-carrier pigs

Information regarding the susceptibility of swine lymphocytes to PCV2 is rather limited. To further explore and characterize the PCV2 infection in swine lymphocytes, an in vitro model using concanavalin A (Con A)-stimulated peripheral blood lymphocytes (PBLs) obtained from clinically healthy PCV2-carrier pigs was introduced. It was found that the PCV2 antigen-containing rate was below 2% in PBLs from healthy PCV2-free pigs following treated simultaneously with Con A and PCV2. However, significantly higher PCV2 antigen- and nucleic acid-containing rates could be seen in Con A-stimulated PBLs from clinically healthy PCV2-carrier pigs. Prior to Con A treatment, both of the PCV2 antigen- and nucleic acid-containing rates in PBLs from healthy PCV2-carrier pigs were less than 1%; however, they reached 22.1+/-5.7% by flow cytometry and 27.1+/-6.5% by in situ hybridization, respectively, at 4-day post-incubation with Con A. Phenotyping of PCV2 antigen-containing cells revealed that PCV2-positive cells could be detected in both T and B lymphocyte populations within which IgM-positive B lymphocytes appeared to have a relatively higher positive rate. The Con A-stimulated PBLs also displayed a significantly higher viral load by the measurement of either PCV2 DNA copy number or viral titer when compared with the non-treated PBLs from healthy PCV2-carrier pigs. The results indicate that PBLs, especially IgM-bearing B lymphocytes, are indeed susceptible to PCV2 infection and PCV2 is capable of replicating in dividing lymphocytes. This activation-induced replication may explain in part the pathogenesis of lymphoid depletion in PMWS-affected pigs.     

Evaluation of dietary inclusion of housefly maggot (Musca domestica) meal on growth,fillet composition and physiological responses for barramundi,Lates calcarifer

The study was to evaluate the effects of dietary fish meal (FM) partially replaced by housefly maggot meal (HMM) on growth, fillet composition and physiological responses of juvenile barramundi, Lates calcarifera. HMM at 100, 150, 200 and 300 g kg^?1 was supplemented in the basal diet to replace dietary FM protein. Basal diet without HMM supplementation was used as control. Total of five experimental diets were fed to triplicate groups of juvenile barramundi (initial weight: 9.66 ± 0.22 g) in a flow‐through rearing system for 8 weeks. Fish fed all experimental diets showed no effects (P > 0.05) on weight gain and whole body protein, lipid and moisture content. Fish fed control diet and 100 g kg^?1 HMM diet had the highest (P < 0.05) hepatic superoxide dismutase (SOD) activity, followed by 150 g kg^?1 HMM group, the lowest in 200 and 200 g kg^?1 HMM groups. Hepatic thiobarbituric acid reactive substance (TBARS) value was the highest in fish fed 150–300 g kg^?1 HMM diets, followed by 100 g kg^?1 HMM group and the lowest in fish fed the control diet. Fish fed the 300 g kg^?1 HMM diet had lower plasma lysozyme activity than fish fed other diets. The results indicated that up to 300 g kg^?1 HMM can be used to substitute dietary FM protein without negative effect on growth. Although physiological responses were also considered, up to 100 g kg^?1 HMM in barramundi diet was recommended.     

Immune response of largemouth bass, <Emphasis Type="Italic">Micropterus salmoides</Emphasis>, to whole cells of different <Emphasis Type="Italic">Nocardia seriolae</Emphasis> strains

Largemouth bass Micropterus salmoides were immunized with four different N. seriolae strains—two α-glucosidase-positive (961113, KU040801) and two α-glucosidase-negative (94260, OTTS) strains—along with Freund’s incomplete adjuvant. After primary immunization (week 0), a booster was administered at weeks 4 and 8. Nonspecific immune responses to multiple immunizations with the different N. seriolae strains were determined based on serum lysozyme activity and nitroblue tetrazolium (NBT)-positive cells in peripheral blood. The serum lysozyme activity and NBT-positive cells in peripheral blood were not significantly increased even after the two booster immunizations. Specific antibody responses against N. seriolae cells were investigated by enzyme-linked immunosorbent assay. At 4 weeks after immunization, all groups immunized with N. seriolae antigens showed significant increases in their specific antibody levels. The sera from fish immunized with different N. seriolae strains exhibited reactivity with N. seriolae sonicated antigens of 28, 30, 36 and 84 kDa by western blot analysis. After two boosters, fish were challenged with live N. seriolae to assess the vaccine’s efficacy; however, multiple injections of the N. seriolae strains did not reduce mortality, irrespective of the bacterin.     

Averting the baiji syndrome: conserving habitat for critically endangered dolphins in Eastern Taiwan Strait

• 1. Numbering no more than 100 individuals and facing many threats, the geographically isolated Eastern Taiwan Strait population of Indo‐Pacific humpback dolphins (Sousa chinensis) is in peril. The estuarine and coastal waters of central‐western Taiwan have historically provided prime habitat for these dolphins, but environmental conditions today bear little resemblance to what they were in the past.
• 2. The humpback dolphins must share their habitat with thousands of fishing vessels and numerous factories built upon thousands of hectares of reclaimed land.
• 3. They are exposed to chemicals and sewage released from adjacent terrestrial activities. Noise and disturbance associated with construction, vessel traffic and military activities are features of everyday life for these animals.
• 4. Measures to slow the pace of habitat deterioration and reduce the many risks to the dolphins are urgently needed. As one practical step in this direction, this paper describes the habitat needs of these small cetaceans so that decision makers will be better equipped to define ‘priority habitat’ and implement much needed protection measures under the terms of local legislation.
• 5. The preferred habitat of these dolphins in Taiwan consists of shallow (<30 m), near‐shore marine waters with regular freshwater inputs.
• 6. For such a small, isolated and threatened population, ‘priority habitat’ should not be limited to areas of particularly intensive dolphin use or high dolphin density, but rather it should encompass the entire area where the animals have been observed (their current ‘habitat’), as well as additional coastal areas with similar bio‐physical features (‘suitable habitat’). Such a precautionary approach is warranted because the loss of only a few individuals could have serious population‐level consequences.
• 7. While conventional socio‐economic analysis might suggest that implementing protection measures over an area stretching ～350 km north–south along Taiwan's west coast and ～3 km out to sea would be too ‘costly’, the loss of this charismatic species from Taiwan's waters would send a troubling message regarding our collective ability to reconcile human activities with environmental sustainability. Copyright © 2010 John Wiley & Sons, Ltd.

     

Induced ovulation of yellow catfish (Pelteobagrus fulvidraco) using a combination of a gonadotrop‐releasing hormone analogue and domperidone

The effects of an intraperitoneal hormone injection of gonadotropin‐releasing hormone agonist (D‐Ala⁶, Pro⁹‐NEt GnRHa) alone or in combination with a dopamine antagonist, domperidone (DOM), on ovulation induction in yellow catfish Pelteobagrus fulvidraco were tested. The hormone treatments were as follows: 6 mg kg⁻¹ body weight (BW) of carp pituitary extract as a positive control, GnRHa 10, 20, 40 and 80 μg kg⁻¹ BW and a combination of GnRHa and DOM as follows: 10 μg+5 mg, 20 μg+10 mg, 40 μg+20 mg and 80 μg+40 mg kg⁻¹ BW. Physiological saline (0.7% NaCl) was used as a negative control. Significant differences in the ovulation ratio, latency period and ovulation index (OI) were observed among treatments (P<0.05). The combination of GnRHa and DOM at doses of 40 μg+20 mg kg⁻¹ BW had higher values of the ovulation ratio and OI, and a shorter latency period compared with other treatments. The highest OI in GnRHa treatments was only 56.67%, suggesting a dopaminergic tone on gonadotropin secretion in this fish at the pre‐ovulatory stage. Therefore, ovulation can be successfully induced in yellow catfish with 40 μg kg⁻¹ GnRHa+20 mg kg⁻¹ DOM without affecting the egg quality.     

Development and testing of a Bayesian population model for the bigeye thresher shark,Alopias superciliosus,in an area subset of the western North Pacific

A Bayesian population modelling tool integrating separable virtual population analysis, per‐recruit models and age‐structured demographic analysis was developed for the bigeye thresher Alopias superciliosus (Lowe) population in an area subset of the western North Pacific. The mortality rates for years 1989–2016 were estimated, various biological reference points and associated risks of decline were also estimated, and alternative harvest strategies for the stock were evaluated. Estimates of the posterior mean of fishing mortality for bigeye thresher shark suggest fishing pressure has been high in recent years (2011–2016). The estimated population growth rate (λ) (without fishing) obtained from age‐structured demographic model was relatively low (λ = 1.01 per year; 95% confidence intervals of 1.00 and 1.03 per year). Risk analyses revealed that only low levels of fishing pressure (10% of the current fishing pressure) over a wide range of ages could maintain a relatively low risk of population decline for bigeye threshers. Sensitivity testing indicated that the model is robust to prior specification. The developed framework could be used as an assessment tool to evaluate the risk of decline for other widely distributed pelagic shark species where insufficient catch and effort data are available.     

Chemical Characteristics of Different Tissues of Spectacled Caiman (Caiman crocodilus)

Proximate compositions of dorsal, ventral, leg, and tail cuts of farmed spectacled caiman (Caiman crocodilus) were determined. The water, protein, fat, and ash contents of muscle were 77–78%, 19–23%, 0.3–1.2%, and 1.0%, respectively. The dominant nucleotide-related compound in the tissues was inosine monophosphate (IMP) (68.6–84.9%). The predominant free amino acids (FAAs) in the muscle of spectacled caiman were taurine (Tau), glycine (Gly), alanine (Ala), and histidine (His), along with a dipeptide of anserine (Ans) (80–183 mg/100 g) and carnosine (Car) (35–103 mg/100 g) being detected in each sample. The major fatty acid composition included palmitic (16:0), oleic (18:1 cis-9), linoleic (18:2n-6), and stearic (C18:0) acids. The amount of docosahexaenoic acid (4.27–8.76%) was higher than that of eicosapentaenoic acid (0.99–2.24%). The ratio of polyunsaturated fatty acid/saturated fatty acid in spectacled caiman (0.83–1.19) was higher than that in other commonly consumed meat. Meat from the spectacled caiman can be characterized as a valuable food for human consumption in terms of these nutritional features.     

Dihydroaustrasulfone Alcohol Inhibits PDGF-Induced Proliferation and Migration of Human Aortic Smooth Muscle Cells through Inhibition of the Cell Cycle

Dihydroaustrasulfone alcohol is the synthetic precursor of austrasulfone, which is a marine natural product, isolated from the Taiwanese soft coral Cladiella australis. Dihydroaustrasulfone alcohol has anti-inflammatory, neuroprotective, antitumor and anti-atherogenic properties. Although dihydroaustrasulfone alcohol has been shown to inhibit neointima formation, its effect on human vascular smooth muscle cells (VSMCs) has not been elucidated. We examined the effects and the mechanisms of action of dihydroaustrasulfone alcohol on proliferation, migration and phenotypic modulation of human aortic smooth muscle cells (HASMCs). Dihydroaustrasulfone alcohol significantly inhibited proliferation, DNA synthesis and migration of HASMCs, without inducing cell death. Dihydroaustrasulfone alcohol also inhibited platelet-derived growth factor (PDGF)-induced expression of cyclin-dependent kinases (CDK) 2, CDK4, cyclin D1 and cyclin E. In addition, dihydroaustrasulfone alcohol inhibited PDGF-induced phosphorylation of extracellular signal-regulated kinase 1/2 (ERK1/2), whereas it had no effect on the phosphorylation of phosphatidylinositol 3-kinase (PI3K)/(Akt). Moreover, treatment with PD98059, a highly selective ERK inhibitor, blocked PDGF-induced upregulation of cyclin D1 and cyclin E and downregulation of p27^kip1. Furthermore, dihydroaustrasulfone alcohol also inhibits VSMC synthetic phenotype formation induced by PDGF. For in vivo studies, dihydroaustrasulfone alcohol decreased smooth muscle cell proliferation in a rat model of restenosis induced by balloon injury. Immunohistochemical staining showed that dihydroaustrasulfone alcohol noticeably decreased the expression of proliferating cell nuclear antigen (PCNA) and altered VSMC phenotype from a synthetic to contractile state. Our findings provide important insights into the mechanisms underlying the vasoprotective actions of dihydroaustrasulfone alcohol and suggest that it may be a useful therapeutic agent for the treatment of vascular occlusive disease.     

Genetic characteristics of Streptococcus dysgalactiae isolated from cage cultured cobia,Rachycentron canadum (L.)

Disease outbreaks occurred during 2007–2013 in Taiwan with 2.5–10% mortality among the cage cultured cobia, Rachycentron canadum (L.), characterized by the presence of polyserositis, pericarditis and peritonitis. The micro‐organisms isolated from internal organs were Gram‐positive cocci. The isolates were confirmed as Streptococcus dysgalactiae by a polymerase chain reaction assay that yielded the expected specific 259 bp amplicon. Additionally, partial sequence of the 16S–23S rDNA intergenic spacer region of the GCS strain isolates from fish was also compared and produced 100% sequence identity with S. dysgalactiae (GenBank accession number AB252398 ). The genetic characterization was then determined by pulsed‐field gel electrophoresis (PFGE) analysis. Based on PFGE, the Apa I or Sma I digestion patterns of chromosomal DNA of these isolates were grouped into three main clusters. Taiwanese strains were divided into two clusters, and the tet(M) gene was detected in cluster 1 (pulsotypes: A1–A2 and S1–S3), but not in cluster 2 strains (pulsotypes: A3–A4 and S4–S5). Three Japanese strains from amberjack, Seriola dumerili (Risso), were grouped into cluster 3 (pulsotypes: A5–A7 and S6–S8) and displayed no mortality to cobia in the challenge experiment. Conversely, Taiwanese strains from cobia and snubnose pompano, Trachinotus blochii (L.), displayed a mortality rate of 50–87.5% in cobia.     

Induction of Apoptosis by Sinulariolide from Soft Coral through Mitochondrial-Related and p38MAPK Pathways on Human Bladder Carcinoma Cells

Sinulariolide, an isolated compound from the soft coral Sinularia flexibilis, possesses the anti-proliferative, anti-migratory and apoptosis-inducing activities against the TSGH bladder carcinoma cell. The anti-tumor effects of sinulariolide were determined by 3-(4,5-cimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide assay, cell migration assay and flow cytometry, respectively. Sinulariolide inhibited the growth and migration of bladder carcinoma cells in a dose-dependent manner, as well as induced both early and late apoptosis as determined by the flow cytometer. Also, the sinulariolide-induced apoptosis is related to the mitochondrial-mediated apoptosis via caspase-dependent pathways, elucidated by the loss of mitochondrial membrane potential, release of cytochrome C, activation of caspase-3/-9, Bax and Bad, as well as suppression of Bcl-2/Bcl-xL/Mcl-1. Detection of the PARP-1 cleaved product suggested the partial involvement of caspase-independent pathways. Moreover, inhibition of p38MAPK activity leads to the rescue of the cell cytotoxicity of sinulariolide-treated TSGH cells, indicating that the p38MAPK pathway is also involved in the sinulariolide-induced cell apoptosis. Altogether, these results suggest that sinulariolide induces apoptosis against bladder cancer cells through mitochondrial-related and p38MAPK pathways.