Genetic manipulation of Anopheles stephensi immunity to increase plasmodium falciparum salivary gland sporozoite infection levels

Human malaria is responsible for over 700,000 deaths a year. To stay abreast of the threat posed by the parasite, a constant stream of new drugs and vector control methods are required. This study focuses on a vaccine that has the potential to protect against parasite infection, but has been hindered by developmental challenges. In malaria prevention, live, attenuated, aseptic, Plasmodium falciparum sporozoites (PfSPZ) can be administered as a highly protective vaccine. PfSPZ are produced using adult female Anopheles stephensi mosquitoes as bioreactors. Production volume and cost of a PfSPZ vaccine for malaria are expected to be directly correlated with Plasmodium falciparum infection intensity in the salivary glands. The sporogonic development of Plasmodium falciparum in A. stephensi to fully infected salivary gland stage sporozoites is dictated by the activities of several known components of the mosquito’s innate immune system. Here I report on the use of genetic technologies that have been rarely, if ever, used in Anopheles stephensi Sda500 to increase the yield of sporozoites per mosquito and enhance vaccine production. By combining the Gal4/UAS bipartite system with in vivo expression of shRNA gene silencing, activity of the IMD signaling pathway downstream effector LRIM1, an antagonist to Plasmodium development, was reduced in the midgut, fat body, and salivary glands of A. stephensi. In infection studies using P. berghei and P. falciparum these transgenic mosquitoes consistently produced significantly more salivary gland stage sporozoites than wildtype controls, with increases in P. falciparum ranging from 2.5 to 10 fold. Using Plasmodium infection assays and qRT-PCR, two novel findings were identified. First, it was shown that 14 days post Plasmodium infection, transcript abundance of the IMD immune effector genes LRIM1, TEP1 and APL1c are elevated, in the salivary glands of A. stephensi, suggesting the salivary glands may play a role in post midgut defense against the parasite. Second, a non-pathogenic IMD signaling pathway response was observed which could suggest an alternative pathway for IMD activation. The information gained from these studies has significantly increased our knowledge of Plasmodium defense in A. stephensi and moreover could significantly improve vaccine production.

Relations among Topic Knowledge, Individual Interest, and Relational Reasoning, and Critical Thinking in Maternity Nursing

Critical thinking in learners is a goal of educators and professional organizations in nursing as well as other professions. However, few studies in nursing have examined the role of the important individual difference factors topic knowledge, individual interest, and general relational reasoning strategies in predicting critical thinking. In addition, most previous studies have used domain-general, standardized measures, with inconsistent results. Moreover, few studies have investigated critical thinking across multiple levels of experience. The major purpose of this study was to examine the degree to which topic knowledge, individual interest, and relational reasoning predict critical thinking in maternity nurses. For this study, 182 maternity nurses were recruited from national nursing listservs explicitly chosen to capture multiple levels of experience from prelicensure to very experienced nurses. The three independent measures included a domain-specific Topic Knowledge Assessment (TKA), consisting of 24 short-answer questions, a Professed and Engaged Interest Measure (PEIM), with 20 questions indicating level of interest and engagement in maternity nursing topics and activities, and the Test of Relational Reasoning (TORR), a graphical selected response measure with 32 items organized in scales corresponding to four forms of relational reasoning: analogy, anomaly, antithesis, and antinomy. The dependent measure was the Critical Thinking Task in Maternity Nursing (CT2MN), composed of a clinical case study providing cues with follow-up questions relating to nursing care. These questions align with the cognitive processes identified in a commonly-used definition of critical thinking in nursing. Reliable coding schemes for the measures were developed for this study. Key findings included a significant correlation between topic knowledge and individual interest. Further, the three individual difference factors explained a significant proportion of the variance in critical thinking with a large effect size. While topic knowledge was the strongest predictor of critical thinking performance, individual interest had a moderate significant effect, and relational reasoning had a small but significant effect. The findings suggest that these individual difference factors should be included in future studies of critical thinking in nursing. Implications for nursing education, research, and practice are discussed.