Expression of the C-terminal family 22 carbohydratebinding module of xylanase 10B of Clostridium themocellum in tobacco plant

Carbohydrate-binding modules have been shown to alter plant cell wall structural architecture. Hence, they have the potential application of being used to engineer the plant to produce tailor-made natural fibers in the cell wall. The Clostridium thermocellum xylanase, Xyn10B, contains two CBMs that belong to family 22 (CBM22). The C-terminal CBM22-2 of the glycoside hydrolase (GH) 10 had been characterized to interact with xylan, a major hemicellulosic component in the secondary cell wall of plants. In this work, the expression of the CBM22-2 in transgenic tobacco plants was evaluated.Histological examinations of the transgenic stems did not reveal marked cell wall phenotype. In addition, there were no observable changes in the height or the appearance of the transgenic plants expressing the CBM22-2 module. The results indicate that the family 22 carbohydrate binding module isnot a potential candidate for use in in planta modification of the cell wall

The plant biotechnology flight: Is Africa on board?

The development of plant biotechnologies has been very rapid in recent times, especially in the developed countries. The technologies have created a new branch of biotechnology known as molecular farming, where plants are engineered to produce pharmaceutical and technical proteins in large quantities. An evaluation of the status of plant biotechnology development in Africa revealed that majority of the countries that are involved in biotech activities are still at the level of tissue culture applications. This calls for urgent and sincere commitments on the part of the various stakeholders inAfrica, especially the governments, to the development of biotechnology capacity

Gastric Acid Secretion, Mucus Concentration and Ulceration Following Orogastic Feeding of Cannabis Sativa to Albino Wistar Rats

This study was carried out to evaluate the effect of consumption (ingestion) of Cannabis sativa on the gastrointestinal tract using mucus concentration, acid secretion and ulceration in animal (rats) model as indices. Three groups of six (6) rats each were used. The control group were fed on rat chow only while another group were fed on low dose of Cannabis sativa 0.5gm/100g body weight and another group were fed on high dose of Cannibis sativa 0.8mg/100g body weight for 28 days respectively and allowed free access to food and water. At the end of 28 days, the results showed that the acid secretion in the high dose group was significantly (P<0.001) higher than in low dose group while the mucus concentration in low dose and high dose group respectively were significantly lower (P<0.01) than control. The mucus concentration in high dose was significantly lower (P<0.001) than low dose group when the test groups were compared. Similarly the ulcer scores were higher in low dose group (P<0.05) and in high dose group (P<0.001) respectively when compared with control group. The ulceration in high dose group was significantly higher (P<0.001) than the low dose group. In conclusion, ingestion of Cannabis sativa causes decrease in adherent gastric mucus, increase acid secretion and increase in gastric ulceration in a dose dependent fashion. The liberal use of Cannabis sativa should therefore be discouraged

Short-duration exposure to 2.45 GHz microwave radiation induces DNA damage in Sprague Dawley rat’s reproductive systems

The genotoxic effects of 2.45 GHz microwave (MW) radiation on the testis and ovary of Sprague Dawley rats was investigated. The animals were exposed to varying levels of specific absorption rate (SAR) of 0 (control), 0.48, 0.95, 1.43, 1.91, 2.39, 2.90, 3.40, 3.80 and 4.30 Wkg-1, for 10 min. The induction of DNA damages was assessed using DNA direct amplification of length polymorphisms (DALP) and validated with single cell gel electrophoresis (SCGE) comet assay for same cells at SAR 2.39 Wkg-1. Potential damage at the organ level was assessed by histopathological study. The results show significant differences in the Olive moment and % DNA in the blood of the exposed animals when compared with the control (p < 0.05). Hyperchromasia was observed in the ovary of the animals exposed to MW radiation. Also, there was reduction in the number of germ cells and cell disorganization in the testis of exposed group with increasing SARs. These results suggest that MW radiation has the potential to affect both male and female fertility adversely.Keywords: 2.45 GHz microwave radiation, histopathology, DNA single strand break, ovary, testisAfrican Journal of Biotechnology Vol. 12(2), pp. 115-12

Expression of an expansin carbohydrate-binding module affects xylem and phloem formation

Expansins are believed to be involved in disrupting the non-covalent adhesion of cellulose to matrix polysaccharides, thereby promoting wall creep. We have targeted a putative potato expansin (EXPA)carbohydrate-binding module (CBM) to the cell walls of tobacco plants. Histological examinations and electron microscopy indicated that 30% of the xylem cells of the transgenic stems with high expressionof the expansin CBM are wider (radial surface area) than those of the controls. Similarly, 37% of the xylem cells of the stems of the high expressers have thinner cell walls than those of the controls. Therewere no such phenotypes in the low and none expressers, as well as in the control plants. The transgenic tobacco plants expressing the potato expansin CBM did not exhibit marked change in plant morphology. Analysis of cellulose content in the stem cell walls was similar between the high expresser of the transgene and the control plants. Nonetheless, our results taken together demonstrate that expansin CBM alone can bring about changes in the plant cell walls

Promiscuous, non-catalytic, tandem carbohydrate-binding modules modulate the cell-wall structure and development of transgenic tobacco (Nicotiana tabacum) plants

We have compared heterologous expression of two types of carbohydrate binding module (CBM) in tobacco cell walls. These are the promiscuous CBM29 modules (a tandem CBM29-1-2 and its single derivative CBM29-2), derived from a non-catalytic protein1, NCP1, of the Piromyces equi cellulase/hemicellulase complex, and the less promiscuous tandem CBM2b-1-2 from the Cellulomonas fimi xylanase 11A. CBM-labelling studies revealed that CBM29-1-2 binds indiscriminately to every tissue of the wild-type tobacco stem whereas binding of CBM2b-1-2 was restricted to vascular tissue. The promiscuous CBM29-1-2 had much more pronounced effects on transgenic tobacco plants than the less promiscuous CBM2b-1-2. Reduced stem elongation and prolonged juvenility, resulting in delayed flower development, were observed in transformants expressing CBM29-1-2 whereas such growth phenotypes were not observed for CBM2b-1-2 plants. Histological examination and electron microscopy revealed layers of collapsed cortical cells in the stems of CBM29-1-2 plants whereas cellular deformation in the stem cortical cells of CBM2b-1-2 transformants was less severe. Altered cell expansion was also observed in most parts of the CBM29-1-2 stem whereas for the CBM2b-1-2 stem this was observed in the xylem cells only. The cellulose content of the transgenic plants was not altered. These results support the hypothesis that CBMs can modify cell wall structure leading to modulation of wall loosening and plant growth

Functional ectodomain of the hemagglutinin-neuraminidase protein is expressed in transgenic tobacco cells as a candidate vaccine against Newcastle disease virus.

Recently, the use of plants for the production of recombinant proteins has been well demonstrated with promising outcomes. In this study, an efficient Nicotiana tabacum L. cv. Bright Yellow 2 (BY-2) cells system expressing the ectodomain of hemagglutinin-neuraminidase (eHN) protein from Newcastle disease virus (NDV) strain AF2240 was established. Transgenic tobacco BY-2 cell cultures expressing the immunogenic eHN protein were generated and the translation efficiency of eHN protein was enhanced using the 5′-untranslated region of Nicotiana tabacum alcohol dehydrogenase gene (NtADH 5′-UTR) under the control of strong cauliflower mosaic virus (CaMV 35S) promoter. Transgenic lines verified by real-time PCR showed high level of eHN mRNA transcripts and immunoblotting confirmed the presence of 66 kD eHN protein. The eHN protein was stably produced in an average of 0.2–0.4 % total soluble protein. Green fluorescent protein-tagged eHN protein was expressed and localized at the cytosol of BY-2 cell. All mice receiving purified eHN protein from transgenic tobacco BY-2 cells produced specific anti-NDV antibodies. We concluded that plant made eHN elicit immune response and can serve as candidate vaccine against NDV

Low Cost Tuberculosis Vaccine Antigens in Capsules: Expression in Chloroplasts, Bio-Encapsulation, Stability and Functional Evaluation In Vitro

Tuberculosis (TB) caused by Mycobacterium tuberculosis is one of the leading fatal infectious diseases. The development of TB vaccines has been recognized as a major public health priority by the World Health Organization. In this study, three candidate antigens, ESAT-6 (6kDa early secretory antigenic target) and Mtb72F (a fusion polyprotein from two TB antigens, Mtb32 and Mtb39) fused with cholera toxin B-subunit (CTB) and LipY (a cell wall protein) were expressed in tobacco and/or lettuce chloroplasts to facilitate bioencapsulation/oral delivery. Site-specific transgene integration into the chloroplast genome was confirmed by Southern blot analysis. In transplastomic leaves, CTB fusion proteins existed in soluble monomeric or multimeric forms of expected sizes and their expression levels varied depending upon the developmental stage and time of leaf harvest, with the highest-level of accumulation in mature leaves harvested at 6PM. The CTB-ESAT6 and CTB-Mtb72F expression levels reached up to 7.5% and 1.2% of total soluble protein respectively in mature tobacco leaves. Transplastomic CTB-ESAT6 lettuce plants accumulated up to 0.75% of total leaf protein. Western blot analysis of lyophilized lettuce leaves stored at room temperature for up to six months showed that the CTB-ESAT6 fusion protein was stable and preserved proper folding, disulfide bonds and assembly into pentamers for prolonged periods. Also, antigen concentration per gram of leaf tissue was increased 22 fold after lyophilization. Hemolysis assay with purified CTB-ESAT6 protein showed partial hemolysis of red blood cells and confirmed functionality of the ESAT-6 antigen. GM1-binding assay demonstrated that the CTB-ESAT6 fusion protein formed pentamers to bind with the GM1-ganglioside receptor. The expression of functional Mycobacterium tuberculosis antigens in transplastomic plants should facilitate development of a cost-effective and orally deliverable TB booster vaccine with potential for long-term storage at room temperature. To our knowledge, this is the first report of expression of TB vaccine antigens in chloroplasts

Global patient outcomes after elective surgery: prospective cohort study in 27 low-, middle- and high-income countries.

BACKGROUND: As global initiatives increase patient access to surgical treatments, there remains a need to understand the adverse effects of surgery and define appropriate levels of perioperative care. METHODS: We designed a prospective international 7-day cohort study of outcomes following elective adult inpatient surgery in 27 countries. The primary outcome was in-hospital complications. Secondary outcomes were death following a complication (failure to rescue) and death in hospital. Process measures were admission to critical care immediately after surgery or to treat a complication and duration of hospital stay. A single definition of critical care was used for all countries. RESULTS: A total of 474 hospitals in 19 high-, 7 middle- and 1 low-income country were included in the primary analysis. Data included 44 814 patients with a median hospital stay of 4 (range 2-7) days. A total of 7508 patients (16.8%) developed one or more postoperative complication and 207 died (0.5%). The overall mortality among patients who developed complications was 2.8%. Mortality following complications ranged from 2.4% for pulmonary embolism to 43.9% for cardiac arrest. A total of 4360 (9.7%) patients were admitted to a critical care unit as routine immediately after surgery, of whom 2198 (50.4%) developed a complication, with 105 (2.4%) deaths. A total of 1233 patients (16.4%) were admitted to a critical care unit to treat complications, with 119 (9.7%) deaths. Despite lower baseline risk, outcomes were similar in low- and middle-income compared with high-income countries. CONCLUSIONS: Poor patient outcomes are common after inpatient surgery. Global initiatives to increase access to surgical treatments should also address the need for safe perioperative care. STUDY REGISTRATION: ISRCTN5181700