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Evaluation and Comparison of Clostridium Epsilon-Alpha Fusion Gene Expression Using Different Commercial Expression Vector.

ABSTRACT: Clostridium perfringens and Clostridium septicum are gram-positive, anaerobic, spore-forming rods and pathogens for humans and livestock, which are widespread in nature as well as human and animal digestive systems. C. perfringens produces numerous different exoproteins, which are various systems of action. The major C. perfringens toxins include alpha, beta, epsilon, and iota. C. perfringens are classified into five groups (A-E) on the basis of the production of these lethal toxins. Furthermore, toxins secreted from C. septicum include alpha, beta, delta, and gamma. Epsilon and alpha toxins of C. perfringens and C. septicum are the major causes of enterotoxemia and braxy in sheep and goats, respectively. The production of recombinant immunogenic proteins of these bacteria using suitable expression vectors and expression prokaryotic hosts can be a convenient method for the reduction of the costs and production time of clostridial anaerobic vaccines. In the present study, recombinant Escherichia coli strain TOP10 containing pJETεα was used for the evaluation of C. perfringens type D and C. septicum epsilon-alpha fusion protein using different commercial vectors. After the extraction of pJETεα from the recombinant cell, it was digested by NdeI and XhoI restriction enzymes and subcloned into pET22b (+), pET26b (+), and pGEM-B1 expression vectors in E. coli/Rosetta and E. coli/BL21 (DE3). The expression of recombinant fusion toxin was evaluated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and western blotting in three different temperatures, various isopropyl β-D-1-thiogalactopyranoside (IPTG) gradients, and different times using pGEMεα, pET22εα, and pET26εα vectors in E. coli/Rosetta and E. coli/BL21 (DE3). According to the obtained results, recombinant E. coli/Rosetta/pET22εα showed better expression at a temperature of 37°C after 6 h of induction by IPTG.

SUBMITTER: Sepehrifar HR

PROVIDER: S-EPMC8410204 | biostudies-literature |

REPOSITORIES: biostudies-literature

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Project description:Sampling for adult mosquito populations is a means of evaluating the efficacy of vector control operations. The goal of this study was to evaluate and identify the most efficacious mosquito traps and combinations of attractants for malaria vector surveillance along the Thai-Myanmar border.In the first part of the study, the BG-Sentinel™ Trap (BGS Trap) and Centers for Disease Control and Prevention miniature light trap (CDC LT) baited with different attractants (BG-lure® and CO2) were evaluated using a Latin square experimental design. The six configurations were BGS Trap with BG-lure, BGS Trap with BG-lure plus CO2, BGS Trap with CO2, CDC LT with BG-lure, CDC LT with BG lure plus CO2, and CDC LT with CO2. The second half of the study evaluated the impact of light color on malaria vector collections. Colors included the incandescent bulb, ultraviolet (UV) light-emitting diode (LED), green light stick, red light stick, green LED, and red LED.A total of 8638 mosquitoes consisting of 42 species were captured over 708 trap-nights. The trap types, attractants, and colored lights affected numbers of female anopheline and Anopheles minimus collected (GLM, P < 0.01). Results revealed that BGS Trap captured many anophelines but was significantly less than the CDC LT. The CDC LT, when baited with BG-lure plus CO2 captured the greatest number of anopheline females with a catch rate significantly higher than the CDC LT baited with BG-lure or CO2 alone (P < 0.05). The number of anopheline females collected from the CDC LT baited with CO2 was greater than the CDC LT baited with BG-lure (646 vs 409 females). None of the alternative lights evaluated exceeded the performance of the incandescent light bulb in terms of the numbers of anopheline and An. minimus collected.We conclude that the CDC LT augmented with an incandescent light shows high potential for malaria vector surveillance when baited with CO2 and the BG-lure in combination and can be effectively used as the new gold standard technique for collecting malaria vectors in Thailand.

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Evaluation and Comparison of Clostridium Epsilon-Alpha Fusion Gene Expression Using Different Commercial Expression Vector.

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