Expression of an Escherichia coli antigenic fusion protein comprising the heat labile toxin B subunit and the heat stable toxin, and its assembly as a functional oligomer in transplastomic tobacco plants

Enterotoxigenic Escherichia coli (ETEC) strains are important pathogens in developing countries. Some vaccine formulations containing the heat labile toxin B subunit (LTB) have been used in clinical trials; however, the induction of neutralizing antibodies against the heat-stable toxin (ST), a poor immunogenic peptide, is necessary, as most ETEC strains can produce both toxins. In this study, a plant optimized synthetic gene encoding for the LTB-ST fusion protein has been introduced into plastids of tobacco leaf tissues, using biolistic microprojectile bombardment, in an effort to develop a single plant-based candidate vaccine against both toxins. Transplastomic tobacco plants carrying the LTB-ST transgene have been recovered. Transgene insertion into the plastid was confirmed by both PCR and Southern blot analysis. GM1-ELISA revealed that the LTB-ST fusion protein retained its oligomeric structure, and displayed antigenic determinants for both LTB and ST. Western blot analysis, using LTB antisera, confirmed the presence of a 17-KDa protein in transplastomic lines, with the correct antigenicity of the fusion protein. Expression levels of this fusion protein in different lines reached up to 2.3% total soluble protein. Oral immunization of mice with freeze-dried transplastomic tobacco leaves led to the induction of both serum and mucosal LTB-ST specific antibodies. Following cholera toxin challenge, a decrease of intestinal fluid accumulation was observed in mice immunized with LTB-ST-containing tobacco. These findings suggest that tobacco plants expressing LTB-ST could serve as a plant-based candidate vaccine model providing broad-spectrum protection against ETEC-induced diarrhoeal disease.     

A protein complex bearing an oxidase with napthalene dihydrodiol dehydrogenase activity is induced in <Emphasis Type="Italic">Mucor circinelloides</Emphasis> strain YR-1 during growth on polycyclic aromatic compounds

Fungi are organisms capable of growing in a myriad of conditions and respond to counteract environmental cues. Several locations in the world are polluted with oil and its derivatives, and some microorganisms tolerant to these compounds have been isolated. Some fungi can grow in the presence of molecules such as polycyclic aromatic hydrocarbons as sole carbon sources. In this report, we further characterized the induced enzymes with phenanthrene from Mucor circinelloides YR-1 strain, isolated from a polluted field near a petrochemical facility in México. We identified a putative oxidase that is induced when growth with phenanthrene as sole carbon source at a pH of 8.5 and is NADP⁺ dependent. We show that this enzyme bears naphthalene dihydrodiol dehydrogenase activity with substrate preference for the cis-naphthalene over the trans-naphthalene, with an optimal pH in the range of 8–10. Mass spectrometry analysis revealed that the induced enzyme belongs to the NADP⁺ oxidase family enzymes with the typical Rossmann-fold for NADP⁺ binding. This enzyme seems to form a high molecular weight structure (~ 541 kDa) and with a monomer of 57 kDa, suggesting that the multimer is constituted of 10 subunits. Our findings contribute to understanding of the roles that dihydrodiol dehydrogenases have in organisms exposed to toxic compounds in the environment and can regulate their expression.     

Complex segregation analysis of diabetes mellitus

Complex segregation analysis was applied to a sample of 12,293 nuclear families each with at least 1 diabetic patient. The families were divided into two groups depending on the proband's treatment: insulin-dependent (IDG) and insulin-independent (IIG). Heterogeneity analysis has revealed a highly significant difference in the IIG group when families were divided into different mating types. The higher recurrence risk was found in the group with affected mothers. Also evidence for a major recessive gene was found in the IGG group, while it was not possible to distinguish between the hypothesis for absence of a major locus and absence of polygenic inheritance in the IDG group. Risks to develop the disease were calculated for a few typical situations.     

Comparison of Carotid Intima-Media Thickness in Children and Adults With and Without Obesity: A Hysteresis Model

ObjectiveTo describe a third-degree polynomial function (hysteresis) of the effect size of age, obesity, and insulin sensitivity over the carotid intima-media thickness (c-IMT), in the pediatric and adult groups.MethodsA quasi-experimental study with fixed factor analysis of age (children aged 8-12 years, n = 73; adults aged 21-45 years, n = 82) and obesity (yes, n = 76; no, n = 79) was conducted to analyze the effect on the c-IMT and Matsuda insulin sensitivity index values. This quasi-experimental design was analyzed with robust regression modeling.ResultsThe additive effect of obesity, independent of age, was evident in the case of the c-IMT values. There was no interaction effect, but a significant difference between participants with normal weight and those with obesity was found (P < .0001). The difference between adults and children was also significant, but the effect size was smaller. A model was created based on age, Tanner stage, and obesity using the c-IMT and Matsuda insulin sensitivity index values. A linear function fit as R², and the cubic function estimated parameters like a polynomial model.ConclusionThis practical study design showed that children with obesity displayed the same levels of carotid intima-media abnormalities as adults with obesity. The polynomial shape of the model suggests potentially poor outcomes that resemble the hysteresis process and may predict chronic cardiometabolic events during early adulthood.