Comparing the Ability of Secretory Signal Peptides for Heterologous Expression of Anti-Lipopolysaccharide Factor 3 in Chlamydomonas reinhardtii.

Anti-lipopolysaccharide factor 3 (ALFPm3) possesses a wide antimicrobial spectrum and high antibacterial and viral activities for broad application prospects in the aquaculture industry. However, the application of ALFPm3 is limited by its low production in nature, as well as its low activity when expressed in Escherichia coli and yeast. Although it has been proven that its secretory expression can be used to produce antimicrobial peptides with strong antimicrobial activity, there is no study on the high-efficiency secretory expression of ALFPm3 in Chlamydomonas reinhardtii. In this study, signal peptides ARS1 and CAH1 were fused with ALFPm3 and inserted into the pESVH vector to construct pH-aALF and pH-cALF plasmids, respectively, that were transformed to C. reinhardtii JUV using the glass bead method. Subsequently, through antibiotic screening, DNA-PCR, and RT-PCR, transformants expressing ALFPm3 were confirmed and named T-JaA and T-JcA, respectively. The peptide ALFPm3 could be detected in algal cells and culture medium by immunoblot, meaning that ALFPm3 was successfully expressed in C. reinhardtii and secreted into the extracellular environment. Moreover, ALFPm3 extracts from the culture media of T-JaA and T-JcA showed significant inhibitory effects on the growth of V. harveyi, V. alginolyticus, V. anguillarum, and V. parahaemolyticus within 24 h. Interestingly, the inhibitory rate of c-ALFPm3 from T-JcA against four Vibrio was 2.77 to 6.23 times greater than that of a-ALFPm3 from T-JaA, indicating that the CAH1 signal peptide was more helpful in enhancing the secreted expression of the ALFPm3 peptide. Our results provided a new strategy for the secretory production of ALFPm3 with high antibacterial activity in C. reinhardtii, which could improve the application potentiality of ALFPm3 in the aquaculture industry.

Secretory Expression and Application of Antilipopolysaccharide Factor 3 in Chlamydomonas reinhardtii.

Anti-lipopolysaccharide factor is a class of antimicrobial peptides with lipopolysaccharide-binding structural domains, which has a broad antimicrobial spectrum, high antimicrobial activities, and broad application prospects in terms of the aquaculture industry. However, the low yield of natural antimicrobial peptides and their poor expression activity in bacteria and yeast have hindered their exploration and utilization. Therefore, in this study, the extracellular expression system of Chlamydomonas reinhardtii, by fusing the target gene with the signal peptide, was used to express anti-lipopolysaccharide factor 3 (ALFPm3) from Penaeus monodon in order to obtain highly active ALFPm3. Transgenic C. reinhardtii T-JiA2, T-JiA3, T-JiA5, and T-JiA6, were verified using DNA-PCR, RT-PCR, and immunoblot. Additionally, the IBP1-ALFPm3 fusion protein could be detected not only within the cells but also in the culture supernatant. Moreover, the extracellular secretion containing ALFPm3 was collected from algal cultures, and then its bacterial inhibitory activity was analyzed. The results showed that the extracts from T-JiA3 had an inhibition rate of 97% against four common aquaculture pathogenic bacteria, including Vibrio harveyi, Vibrio anguillarum, Vibrio alginolyticus, and Vibrio parahaemolyticus. The highest inhibition rate of 116.18% was observed in the test against V. anguillarum. Finally, the minimum inhibition concentration (MIC) of the extracts from T-JiA3 to V. harveyi, V. anguillarum, V. alginolyticus, and V. parahaemolyticus were 0.11 µg/µL, 0.088 µg/µL, 0.11 µg/µL, and 0.011 µg/µL, respectively. This study supports the foundation of the expression of highly active anti-lipopolysaccharide factors using the extracellular expression system in C. reinhardtii, providing new ideas for the expression of highly active antimicrobial peptides.

Exploring factors influencing the work-related morale for certified nursing assistants in hospice care: A structural equation modeling study.

BACKGROUND: Certified nursing assistants (CNAs) in palliative care units often encounter the death of patients and undergo emotional burdens because of their altruistic practices. However, the influences of altruism and perceptions of death on the work morale of CNAs remain unclear. The fact that morale can be improved by modifying related factors justifies the necessity of a theoretical model of morale for CNAs. However, such a model is currently absent. Based on relevant literature, factors including altruism, social support, perceptions of death, and job satisfaction are related to morale through the mediation of job stress. AIM: This study was aimed toward determining the influencing factors and developing a structural model of morale with regard to CNAs working in palliative care units. DESIGN: The hypothesized model was tested in a cross-sectional design with structural equation modeling using AMOS 23. SETTING/PARTICIPANTS: A total of 413 CNAs from five hospice units in Taiwan participated in the survey. RESULTS: Goodness-of-fit indexes, including χ2/DF = 2.187, PGFI = 0.461, and PNFI = 0.462, among others, confirmed the modified model fit the data well. Altruism, social support, perceptions of death, and job satisfaction were negatively related to job stress (-0.238, -0.196, -0.204, and -0.131, respectively). Altruism and social support positively affected job satisfaction (0.635 and 0.553). Factors directly affecting morale included altruism, social support, job stress, and job satisfaction (0.837, 0.711, -0.244, and 0.540, respectively). The relationship between altruism and morale was the strongest among all the direct paths toward morale. Job stress was the major mediator in these relationships. CONCLUSION: Altruism is imperative for morale. The confirmed model is called the Five Constructs to Morale (5CM) model. These findings indicate the need to assess the levels of altruism and perceptions of death during the recruitment of CNAs. From the perspective of human resource management and organizational behavior, CNAs with a high level of altruism and a positive attitude toward death assure the well-being of end of life patients because of work involvement and commitment to the challenging work of hospice care. The cultivation of a supportive environment and coping strategies are suggested for the management of CNAs facing patient death. Hospice care decision makers should carefully assess morale levels among health professionals by applying the 5CM model. Identifying modifiable factors that can improve morale among nursing staff in hospice care to ensure work fulfillment and quality care merits emphasis in clinical practice. The establishment of effective strategies to facilitate the work morale of nursing staff is suggested for palliative care units. Morale improvement related task should be a global public health priority.

[Clinical significance of serum S-100B protein in severe cerebral injury].

OBJECTIVE: To investigate the value of S-100B protein as a biology marker in diagnosis and prognostic after severe cerebral injury. METHODS: Sixty-six patients with severe head injury (Glasgow coma scale score (GCS)