Lethal Effects of Pulsed High-Voltage Discharge on Marine Plankton and Escherichia coli

Ballast seawater is considered globally as a major vector for invasions of non-indigenous organisms. Several technologies have been tested for their ability to remove organisms from ballast water. In the present study, we constructed a novel pulsed high-voltage discharge (PHVD) system that could operate in either high current mode with several hundred amperes or shockwave generating mode with relatively lower current in seawater. In laboratory-scale experiments, the PHVD system with shockwave-generating mode was found to be more effective in killing zooplankton (1.9- to 4.0-fold) and phytoplankton (3.3-fold) than high current mode at discharge with 300–500 pulses at 7.1 kV. Further experiments were carried out at different voltages and pulse-numbers to examine effects of the shockwave-generating PHVD system on viabilities of one zooplankton larva, two phytoplankton species, and an indicator bacterium suspended in seawater in a static chamber. For zooplankton, live cells were not detected at discharge with 400 pulses at 13 kV. For phytoplankton, the initial live cells of a dinoflagellate was decreased by 77?±?0.5%, and the initial chl a concentration of a diatom was decreased by 76?±?6% at discharge with 700 pulses at 13 kV. For an indicator bacterium Escherichia coli, live cells were not detected at discharges with 200 or 700 pulses at 13 kV. Measurements of ATP content of organisms showed congruent results with those obtained by the above methods, suggesting it may be a rapid method for evaluating treatment efficiency. Though further scale-up studies are necessary, these results suggest that the PHVD system have a high potential for applying to ballast seawater treatment.     

Animal Biowarfare Research: Historical Perspective and Potential Future Attacks

A biological attack on livestock or poultry could result in the loss of valuable animals, costs related to the containment of outbreaks and the disposal of carcasses, lost trade and other economic effects involving suppliers, transporters, distributors and restaurants; however, it is not possible to secure all livestock, and livestock are much less well guarded than human targets. Thus, the vulnerability of the livestock industry to the introduction of biological agents varies for the following reasons: (i) the majority of lethal and contagious biological agents are environmentally resilient, endemic in foreign countries and harmless to humans, making it easier for terrorists to acquire, handle and deploy these pathogens, (ii) with animals concentrated in fewer production facilities and frequently transported between these facilities, a single pathogen introduction could cause widespread infection and (iii) the extent of human travel around the globe makes it difficult to exclude exotic animal diseases as possible biological agents. Historically, many governments have developed and planned to use biological agents for direct attacks on livestock or poultry. In the past, developed nations have actively developed biological weapons to target animals. The potential spectrum of bioterrorism ranges from isolated acts against individuals by individuals to tactical and strategic military attacks and state‐sponsored international terrorism intended to cause mass casualties in animals, humans or both. This review provides an overview of the past development and use of biological weapons and describes potential future attacks.     

Sacrocaudal agenesis in a Korean native calf (Bos Taurus Coreanae)

A male deformed Korean native calf was examined macroscopically. The deformed calf had no caudal vertebral columns from 5th lumbar vertebra, sacrum and coccygeal vertebrae. The spinal cord was terminated in the vertebral foramen of the 3rd lumbar vertebra. The cervical vertebrae had scoliosis and the 3rd and 4th cervical vertebrae were fused. The 2nd and 3rd lumbar vertebrae were fused and the left and right transverse processes of the 4th lumbar vertebra articulated with ala of the ilium. The rectum was greatly expanded by the imperforate anus and a rectourethral fistula was formed between the rectum and urethra. The deformed calf was recorded as a first documentation of sacrocaudal agenesis confirmed in a Korean native calf.     

Serodiagnostic comparison of enzyme-linked immunosorbent assay and surface plasmon resonance for the detection of antibody to Porcine circovirus type 2

This paper describes the cloning and expression of the capsid protein of Porcine circovirus type 2 (PCV2) in an Escherichia coli expression system that was used to produce a fusion protein for subsequent immunologic studies: enzyme-linked immunosorbent assay (ELISA) and surface plasmon resonance (SPR). Polymerase chain reaction was used to amplify the gene encoding the capsid protein from the DNA of PCV2. The protein was then cloned into a pRSET prokaryotic expression vector. Western blot analysis revealed that the recombinant protein gave strong signals on a polyvinylidene difluoride membrane when exposed to the serum from a pig infected with PCV2. The expressed protein was purified and used as an antigen for the ELISA and SPR study. A protein chip based on SPR was developed, and the diagnostic potential of SPR was compared with that of ELISA with the use of 70 serum samples obtained from 6 pig farms. There was a strong positive correlation between the ELISA and SPR titers (r = 0.877, P < 0.01). Therefore, this recombinant capsid protein can be used as an antigen for serologic studies, and the SPR, a label-free method, appears to be a valuable and reproducible tool in the serodiagnosis of a PCV2 infection.     

A preliminary study of dietary protein requirement of juvenile marbled flounder (Pseudopleuronectes yokohamae)

An 8-week feeding trial was conducted to determine the optimal dietary protein level for juvenile marbled flounder. Five semi-purified test diets were formulated to contain different protein levels (CP) including 42.7%, 47.4%, 53.3%, 58.8%, and 64.5% (dry matter), named as CP42.7, CP47.4, CP53.3, CP58.8, and CP64.5, respectively. Five hundred and twenty-five juveniles (6.0 ± 0.1 g) were randomly distributed into 15 tanks (300 L tanks), resulting in 35 fish per tank (n = 3 tanks). Fish were fed the test diets 5 times per day until satiation. The CP58.8 resulted in the highest gain in weight and the best efficiency in feed utilization among the tested protein levels (P < 0.05). Fish fed the CP58.8 diet showed significantly higher whole-body protein and lipid contents than the fish that were fed the other diets (P < 0.05). Fish fed the CP53.3, CP58.8, and CP64.5 diets showed a significantly higher dorsal-muscle lipid content than the fish that were fed the CP42.7 and CP47.4 diets (P < 0.05). The one-slope straight broken-line regression analysis on the results of the thermal growth coefficient and feed conversion ratio indicated that the estimated optimum dietary protein level was 58.8%. Taken together, it is suggested that the dietary protein level of 58.8% is optimal for better growth and high efficiency in feed utilization for the juvenile marbled flounder.     

Serum immunoglobulin fused interferon-�� inhibited tumor growth in athymic mice bearing colon 26 adenocarcinoma cells

Interferon (IFN) has therapeutic potential for a wide range of infectious and proliferative disorders. However, the half-life of IFN is too short to have a stable therapeutic effect. To overcome this problem, serum immunoglobulin has been fused to IFN. In this study, the efficacy of serum immunoglobulin fused INFs (si-IFN1 and si-IFN2) was evaluated on athymic mice bearing colon 26 adenocarcinoma cells. Seven days after the implantation of tumor cells, each group of mice was injected once a week with si-IFN1 and si-IFN2 at two different concentrations (10 × : 30 µg/kg and 50 × : 150 µg/kg). A slight anti-tumoral effect was observed in all 10 × groups compared to the control. In the 50 × groups, however, si-IFN1 and si-IFN2 showed significant anti- tumoral effects compared to the control. To gain more information on the mechanisms associated with the decrease of tumor size, a Western blot assay of apoptosis-related molecules was performed. The protein expression of cytochrome c, caspase 9, 6, and 3 were increased by si-IFN1 and si-IFN2. These 2 IFNs also increased the expressions of p53, p21, Bax and Bad. Interestingly, si-IFN1 and si-IFN2 decreased the expression of VEGF-β. Taken together, serum immunoglobulin fused IFNs increased therapeutic efficacy under current experimental condition.     

Development of a multiplex loop-mediated isothermal amplification assay to detect shiga toxin-producing Escherichia coli in cattle

A multiplex loop-mediated isothermal amplification (mLAMP) assay was developed for simultaneous detection of the stx1 and stx2 genes and applied for detection of shiga toxin-producing Escherichia coli (STEC) in cattle farm samples. Two target genes were distinguished based on T_m values of 85.03 ± 0.54℃ for stx1 and 87.47 ± 0.35℃ for stx2. The mLAMP assay was specific (100% inclusivity and exclusivity), sensitive (with a detection limit as low as 10 fg/µL), and quantifiable (R² = 0.9313). The efficacy and sensitivity were measured to evaluate applicability of the mLAMP assay to cattle farm samples. A total of 12 (12/253; 4.7%) and 17 (17/253; 6.7%) STEC O157, and 11 (11/236; 4.7%) non-O157 STEC strains were isolated from cattle farm samples by conventional selective culture, immunomagnetic separation, and PCR-based culture methods, respectively. The coinciding multiplex PCR and mLAMP results for the types of shiga toxin revealed the value of the mLAMP assay in terms of accuracy and rapidity for characterizing shiga toxin genes. Furthermore, the high detection rate of specific genes from enrichment broth samples indicates the potential utility of this assay as a screening method for detecting STEC in cattle farm samples.     

Nitrite Toxicity and Methemoglobin Changes in Southern Flounder,Paralichthys lethostigma,in Brackish Water

This study was performed to estimate the nitrite toxicity to southern flounder, Paralichthys lethostigma, in brackish water (7.5 ppt of salinity). For a LC₅₀ test, 20 fingerlings (5.7 ± 0.4 cm) in each aquarium (15 L) were exposed to the concentrations of 0, 1, 5, 10, 15, 30, 60, 120, and 240 mg NO₂^?‐N/L in duplication for 10 d. Median lethal concentration at 96 h (96‐h LC₅₀) was calculated as 81.6 mg NO₂^?‐N/L. For a verification test, young flounder (164.2 ± 9.1 g) were exposed to a simulated culture condition in recirculating systems (1000 L). Sodium nitrite was not added to control system, whereas it was added to Treatment system 1 (TS 1) and Treatment system 2 (TS 2) to maintain nitrite concentrations of 20 and 30 mg NO₂^?‐N/L, respectively. The plasma nitrite concentrations of the young flounder in TS 1 and TS 2 were 4.5 and 6.6 mg NO₂^?‐N/L, respectively, after 2 wk. At this time, the methemoglobin percentages in TS 1 and TS 2 reached 85.8 and 89.7%, and survival rates were 37.5 and 25.0%, respectively. The results of these tests indicate that southern flounder do not concentrate nitrite in blood from the environment, but they seem to be more sensitive to nitrite compared with other species that do not concentrate nitrite.