In vitro infection efficiency of nervous necrosis virus alters depending on amount of viral particles adsorbed onto cells.

Nervous necrosis virus (NNV) in the family Nodaviridae is one of the simplest spherical RNA viruses and is pathogenic to many fish species. We investigated the effect of purified NNV on striped snakehead cells (SSN-1) in terms of adsorption ratio and infection efficiency using the 96-well titration system. The proportion of cytopathic effect (CPE)-positive wells among total number of wells inoculated with the virus (CPE appearance ratio) reduced by 17% each time the NNV infectivity dose was halved (y = 55.7x + 50.6). Thus, subtle differences in NNV infectivity could be accurately detected using this system. Experiments performed to observe alteration of CPE appearance ratio with changing viral doses and adsorption times showed that NNV particles introduced into microplate wells as suspensions in ≤ 100 µl inoculum were adsorbed almost completely onto cells seeded on the wells within 4 days of incubation. Density profile analysis of NNV coat proteins revealed that the NNV suspension at 1 50% tissue culture infectious dose (TCID50) contained 60 particles. Infection efficiency/NNV peaked at 20 particles (1.20%/particle) and then declined gradually with increasing NNV doses. Therefore, in vitro infection efficiency of NNV may alter depending on the quantity of viral particles adsorbed onto cells.

Sites responsible for infectivity and antigenicity on nervous necrosis virus (NNV) appear to be distinct.

Nervous necrosis virus (NNV) is a pathogenic fish-virus belonging to the genus Betanodavirus (Nodaviridae). Surface protrusions on NNV particles play a crucial role in both antigenicity and infectivity. We exposed purified NNV particles to different physicochemical conditions to investigate the effects on antigenicity and infectivity, in order to reveal information regarding the conformational stability and spatial relationships of NNV neutralizing-antibody binding sites and cell receptor binding sites. Treatment with PBS at 37 °C, drastically reduced NNV antigenicity by 66-79% on day one, whereas its infectivity declined gradually from 107.6 to 105.8 TCID50/ml over 10 days. When NNV was treated with carbonate/bicarbonate buffers at different pHs, both antigenicity and infectivity of NNV declined due to higher pH. However, the rate of decline with respect to antigenicity was more moderate than for infectivity. NNV antigenicity declined 75-84% after treatment with 2.0 M urea, however, there was no reduction observed in infectivity. The antibodies used in antigenicity experiments have high NNV-neutralizing titers and recognize conformational epitopes on surface protrusions. The maintenance of NNV infectivity means that receptor binding sites are functionally preserved. Therefore, it seems highly likely that NNV neutralizing-antibody binding sites and receptor binding sites are independently located on surface protrusions.

Reproducibility of antigen-immobilized enzyme-linked immunosorbent assay (ELISA) and sandwich ELISA for quantitative detection of NNV particles.

Nervous necrosis virus (NNV) is a fish virus belonging to family Nodaviridae. In this study, we prepared partially aggregated and monometric NNV particles to determine reproducibility of two different enzyme-linked immunosorbent assays (ELISAs): antigen-immobilized ELISA and sandwich ELISA. Passing ratios of purified NNV particles through ultrafilters with molecular weight cut off (MWCO) of 105, 3 × 105 and 106 were 0%, 35.2% and 80.3%, respectively, suggesting that purified NNV particles were partially aggregated whereas those in filtrates with MWCO of 3 × 105 could be monometric. Both NNV particles were subjected to ELISAs. Reduction ratios of ELISA values by 2-fold dilution of antigens were 50% in sandwich ELISA regardless of aggregation state of NNV particles. In contrast, those in antigen-immobilized ELISA were 42% (partially aggregated NNV) to 43% (monometric NNV), which were lower than the theoretical value (50%). This could be due to changes in aggregation state of NNV particles during dry-immobilization. Sandwich ELISA has excellent reproducibility from five times of experiments, in comparison with antigen-immobilized ELISA. Furthermore, available range of regression lines (R2 > 0.99) in sandwich ELISA was wider than that in antigen-immobilized ELISA. These results revealed that sandwich ELISA had better quantitativeness, reproducibility and available range of ELISA values than antigen-immobilized ELISA.

Altered conformational structures of nervous necrosis virus surface protrusions and free coat proteins after incubation at moderate-low temperatures.

Nervous necrosis virus (NNV) is a pathogenic fish virus belonging to family Nodaviridae. The objective of this study was to analyze stabilities of NNV surface protrusion and free coat protein (CP) conformational structures by analyzing changes of NNV infectivity and antigenicity after incubation at moderate-low temperatures. When cultured NNV suspension was incubated at 45 °C, its infectivity declined gradually but its antigenicity maintained. In contrast, both infectivity and antigenicity of purified NNV declined after incubation at 45 °C. After heat-treatment, surface protrusions of NNV particles disappeared completely, although viral particle structures maintained. Therefore, the reduction in NNV infectivity appeared to specifically occur as a result of heat-denaturation of virus surface protrusions. The loss of NNV infectivity in the presence of fetal bovine serum (FBS) was delayed compared to virus heated in the absence of FBS, demonstrating that FBS could function as a stabilizer for conformational structures of NNV surface protrusions. Moreover, the stabilizing function of FBS changed depending on salt concentration. Continued maintenance of antigenicity for heated cultured NNV suspension containing free-CPs may suggest that conformational structures corresponding to protrusion-domain of free-CP are more heat-stable than those of surface protrusions on NNV particles.

The Moderating Effect of Interpersonal Skills on the Relationship between Childhood Emotional Trauma and Depression in Nursing Students / 한국간호교육학회지

PURPOSE: The purpose of this study was to identify the moderating effect of interpersonal skills on the relationship between childhood emotional trauma and depression. METHODS: From June to July, 2017, a convenience sample of 226 nursing students was recruited. Research data were collected through self-report questionnaires and analyzed using SPSS version 22.0. RESULTS: 68.1% (154) respondents experienced emotional abuse, and 48.1% (110) emotional neglect in childhood emotional trauma. The average depression score was 10.76. There were 54.4% (123), 34.5% (78), 12.8% (29), and 7.1% (16) of respondents with mild, moderate, and severe depression, respectively. The average interpersonal skills score was 3.59. There was a significant correlation between childhood emotional trauma (emotional abuse, emotional neglect), interpersonal skills and depression. And the moderating effect of interpersonal skills on the relationship between childhood emotional neglect and depression was significant. CONCLUSION: Interpersonal skills play a role as a moderating variable influencing the relationship between emotional neglect of childhood emotional trauma and depression, and also reduce the effects of childhood emotional trauma on depression. When developing a depression prevention program for nursing college students, such programs should consider strategies to reduce the negative effects of childhood emotional trauma and to improve interpersonal skills.

Lack of nervous necrosis virus (NNV) neutralizing antibodies in convalescent sevenband grouper Hyporthodus septemfasciatus after NNV infection.

Viral nervous necrosis (VNN) is caused by nervous necrosis viruses (NNVs) belonging to genus Betanodavirus (Nodaviridae). It is one of the most serious diseases in aquaculture industry worldwide. In the present study, the kinetics of NNV-infectivity and NNV-specific antibodies in convalescent sevenband grouper Hyporthodus septemfasciatus after NNV infection was determined. When fish were infected with NNV at 17.5 °C, and reared for 84 days at natural seawater temperature (increasing rate: approximately 0.1 °C/day), NNV infectivity peaked on day 14 with 107.80 TCID50/g at the highest, and declined to below the detection limit. When convalescent fish were reared at 27 °C, and re-infected with NNV at 104.3 or 106.3 TCID50/fish, no mortality was observed although NNV multiplied up to 108.80 and 107.80 TCID50/g at the highest, respectively, suggesting NNV-specific immune response. It also revealed that convalescent fish were re-infected by NNV although NNV multiplication was strongly regulated. Interestingly, NNV-specific antibodies were detectable in 20% and ≥80% of convalescent fish before and after re-infection with NNV, respectively. However, no NNV-neutralizing activity was detected before and after re-infection in almost all of the convalescent fish. Therefore, NNV-neutralizing antibodies might not be necessary for the protection of convalescent fish against NNV re-infection after previous NNV infection.

Purification of nervous necrosis virus (NNV) particles by anion-exchange chromatography.

Nervous necrosis virus (NNV) belongs to genus Betanodavirus (family Nodaviridae). It is highly pathogenic to various marine fishes. In the present study, cultured NNV suspension was placed in dialysis tube at molecular weight cut off (MWCO) of 106 and dialyzed against Dulbecco's phosphate buffered saline (D-PBS), 15mM Tris-HCl (pH 8.0), or deionized water (DIW) for 14days followed by anion-exchange chromatography. Infectivity titers of NNV suspensions were stable during dialyses. However, the antigenicity of NNV suspension was decreased to 2.5% by D-PBS dialysis, 11.8% by Tris-HCl dialysis, and 56.2% by DIW dialysis. Anion-exchange chromatograms revealed a total of four peaks (P300, P400, P600 and P700) for NNV suspension after D-PBS dialysis. Additional two peaks (P800 and P-OH) were detected in the NNV suspension after Tris-HCl or DIW dialysis. The substance from the P700 peak had the highest NNV-infectivity. Peak P700 commonly shared by the NNV suspensions after dialysis against the three different buffers. After Tris-HCl dialysis, no other protein except NNV coat protein (CP) at Mr 41,000 was detected from P700. However, after D-PBS dialysis, the P700 peak also contained P600 antigens. Therefore, the P700 peak after Tris-HCl dialysis represented the peak of highly purified NNV particles.

Dialysis buffer with different ionic strength affects the antigenicity of cultured nervous necrosis virus (NNV) suspensions.

Nervous necrosis virus (NNV) belongs to the genus Betanodavirus (Nodaviridae). It is highly pathogenic to various marine fishes. Here, we investigated the antigenicity changes of cultured NNV suspensions during 14days of dialyses using a dialysis tube at 1.4×10(4) molecular weight cut off (MWCO) in three different buffers (Dulbecco's phosphate buffered saline (D-PBS), 15mM Tris-HCl (pH 8.0), and deionized water (DIW)). Total NNV antigen titers of cultured NNV suspension varied depending on different dialysis buffers. For example, total NNV antigen titer during D-PBS dialysis was increased once but then decreased. During Tris-HCl dialysis, it was relatively stable. During dialysis in DIW, total NNV antigen titer was increased gradually. These antigenicity changes in NNV suspension might be due to changes in the aggregation state of NNV particles and/or coat proteins (CPs). ELISA values of NNV suspension changed due to changing aggregates state of NNV antigens. NNV particles in suspension were aggregated at a certain level. These aggregates were progressive after D-PBS dialysis, but regressive after Tris-HCl dialysis. The purified NNV particles self-aggregated after dialysis in D-PBS or in Tris-HCl containing 600mM NaCl, but not after dialysis in Tris-HCl or DIW. Quantitative analysis is merited to determine NNV antigens in the highly purified NNV particles suspended in buffer at low salt condition.

Cell Culture Medium Inhibits Antigen Binding Used in an ELISA for Detection of Antibodies against Nervous Necrosis Virus.

Abstract We investigated the optimum dilution of nervous necrosis virus (NNV) for use as antigens to detect antibodies by an enzyme-linked immunosorbent assay (ELISA) in the Sevenband Grouper Epinephelus septemfasciatus. The ELISA values for a standardized suspension of antigens diluted with L-15 medium containing 1% fetal bovine serum decreased gradually with the dilution of the antigens, whereas those for the antigens diluted with distilled water (DW) initially increased with the dilution of the antigens, peaked at a 320-fold dilution, and then decreased thereafter. Additional studies revealed that binding of NNV antigens to ELISA wells was inhibited by fetal bovine serum and other substances in the L-15 medium. Sera obtained from Sevenband Grouper vaccinated with live NNV vaccine and survivors from natural NNV-infection were subjected to antibody detection by ELISA. All of the sera were positive by ELISA when the standardized suspension was diluted 320-fold, whereas sera from five out of the six survivors and two out of the six vaccinated fish were negative or weakly positive by ELISA using NNV antigens diluted 10-fold. We therefore concluded that cultured NNV solutions prepared in cell culture media may need to be diluted with distilled water for use in ELISA. Received January 28, 2014; accepted April 16, 2014.

Concept Analysis on the Organizational Commitment: Focused on the Nursing Organizations / 한국간호교육학회지

PURPOSE: The purpose of this study was to explore the clear concept of organizational commitment for effective nursing personnel management. METHOD: This study was conducted using Walker & Avant's conceptual analysis framework(2005). RESULTS: Organizational commitment was identified with six attributes: acknowledgment, acceptance, trust, sacrifice, attachment, regulation. Antecedents of organizational commitment were personal characteristics, perceived organizational support, empowerment, fair compensation, job satisfaction, job autonomy, organizational culture, and leadership. Consequences of organizational commitment were turnover intention, job performance and organizational citizenship behavior. CONCLUSION: Organization commitment is a core concept for understanding the management of nursing personnel. Appropriate instruments to operationalize the concept of organizational commitment including six attributes need to be developed.

Assessment of the sevenband grouper Epinephelus septemfasciatus with a live nervous necrosis virus (NNV) vaccine at natural seawater temperature.

We investigated the long-term kinetics of nervous necrosis virus (NNV) infectivity in sevenband grouper, Epinephelus septemfasciatus, injected with a live NNV vaccine at 17.3°C and reared at natural seawater temperature. We also evaluated horizontal infection of NNV from fish vaccinated with live NNV to naïve fish in a cohabitation experiment. Although 10.5% mortality was observed in the vaccinated fish, they were strongly protected from the challenge with homologous NNV. The NNV infectivity titer was detected from day 5 after vaccination, peaked on day 10 at 10(7.43±0.35) TCID50/g, but NNV was under the detection limit (≤10(2.8) TCID50/g) between days 42 and 128. No mortalities or NNV were detected in any of the vaccinated and cohabitated naïve fish, suggesting that NNV spread from fish vaccinated with live NNV should be low if it is limited to fish in the late stage of vaccination (≥42 days from NNV inoculation). The present results demonstrate that a protective immune response to NNV was mounted in sevenband grouper by the live NNV vaccine without controlling fish rearing temperature.

Analysis of Intervention Studies for Married Immigrant Women

PURPOSE: The purpose of this study is to analyze the intervention studies for married immigrant women. METHODS: Based on inclusion criteria, 45 articles published from 2006 to 2011 were reviewed and analyzed. RESULTS: Among the total amount of studies, 77.8% have been conducted since 2009; and 46.7% of them were for a master's thesis, 11.1% were for a doctoral thesis, and 42.2% were journal articles. Most of them were quantitative research (77.8%) and 71.1% were conducted in urban areas. The most frequently conducted intervention was art therapy (35.6%), whereas 6.7% was health education. In total, 104 outcome indicators were used with 57 (54.8%) in the mental health domain, 29 (27.9%) in the social health domain, and 18 (17.3%) in the physical health domain. The most commonly used outcome indicator was self-esteem (23.2%), and the next, in the order of frequency, included self-efficacy (23.2%), acculturative stress (21.4%), and depression (10.7%). CONCLUSION: Most intervention studies were conducted to support the sociocultural adaptation of married immigrant women, while few intervention studies were conducted to support the health of married immigrant women. To promote the health and quality of life of married immigrant women, many health related intervention programs should be undertaken.

Potentiality of a live vaccine with nervous necrosis virus (NNV) for sevenband grouper Epinephelus septemfasciatus at a low rearing temperature.

Nervous necrosis virus (NNV) is the causative agent of viral nervous necrosis (VNN), one of the most serious diseases in over 30 species of cultured marine fishes worldwide. Although several kinds of NNV vaccines have been developed, none of these vaccines have been yet marketed. Here, we demonstrate the potentiality of a live NNV vaccine for sevenband grouper Epinephelus septemfasciatus at a low rearing temperature (17°C). Moreover, we investigated the kinetics of NNV infectivity titer in fish reared at low and optimum temperatures (17°C and 26°C) for VNN onset to determine why sevenband grouper reared at 17°C survive NNV infection. In pathogenicity tests of NNV, fish mortality was reduced by decreasing the fish rearing temperature, and no mortality was observed in fish reared at 17°C regardless of the infection method. During fish acclimation to the optimum temperature of VNN onset (26°C), increased mortalities were observed in the survivors from the 1st NNV-infection. Little or no mortality was observed in the 2nd NNV-infection. Thus, it was demonstrated that the survivors from the 1st NNV-infection mounted a specific protective immune response against NNV. Especially, in the fish infected with NNV by immersion at 17°C, only two out of 30 fish died until the end of the 2nd infection (total survival rate: 93.3%), suggesting a positive potentiality for a live NNV vaccine. In the analysis of NNV kinetics in the fish reared at 26°C, NNV rapidly multiplied up to ≥ 10(9)TCID(50)g(-1) before fish began to die, and the critical level of NNV was around 10(10)TCID(50)g(-1). Probability of NNV multiplication reduced by decreasing the inoculated NNV dose, but NNV multiplication rate was independent of the NNV dose. The threshold of NNV for fish mounting a protective immune response was around >10(4)TCID(50)g(-1). Against this, in the fish reared at 17°C, NNV slowly multiplied in comparison with that in fish at 26°C. NNV titer in the peak was at 10(7.1 ± 1.4)TCID(50)g(-1), which was far behind the critical level of NNV but still greatly above the threshold level (10(4)TCID(50)g(-1)). Thus, it was demonstrated that the multiplication rate of NNV in vivo was strongly correlated to NNV virulence and fish mortality, and down-regulation of NNV multiplication in fish reared at 17°C enabled control of VNN onset for development of a live NNV vaccine.

Contraceptive Knowledge and Practice among Married Immigrant Women

PURPOSE: The purpose of this study was to investigate the contraceptive knowledge and practice among married immigrant women. METHODS: A cross-sectional survey design with a convenient sampling was used. 170 married immigrant women were recruited in Daejeon city and Chungcheong area. A structured questionnaire was self-administered from November 1, 2011 to January 13, 2012. RESULTS: 48.2% of subjects have used contraceptive practice. The widely used contraception methods were IUD, condom, oral pill and withdrawal. 73.2% of subjects got information about contraception from family and friends, and 18.3% of subjects had no source of information. The subjects' employment state (p = .006), duration of marriage (p = .019), number of baby (p = .021), family type (p = .046) have a significant influence on their contraceptive practice. The subjects' mean score of contraceptive knowledge was very low (5.72 points out of 15 points). There were significant differences in subjects' contraceptive knowledge depending on their education level (p = .002), employment state (p < .001), country of origin (p = .010), and family type (p = .003). CONCLUSION: To improve contraceptive knowledge and practice for married immigrant women, it is necessary to develop a proper contraceptive education program to enhance contraceptive knowledge and practice. Adjusted education program by this results will contribute to increase sexual health for married immigrant women.