• BMB Reports
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• 제36권6호
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• pp.565-571
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• 2003

The locomotor responses of human peripheral blood neutrophils and lymphocytes were measured by the change from spherical to polarized shapes in the presence of endotoxins (lipopolysaccharide, LPS) of enteric pathogens: S. dysenteriae type 1, V. cholerae Inaba 569B, S. typhimurium, and K. pneumoniae. We reported earlier that these endotoxins are chemotactic factors for the neutrophils since they stimulated cell polarization within a few minutes of incubation. Endotoxins had an inhibitory effect upon neutrophil phagocytosis of opsonized yeast and the cells engulfed fewer yeasts. Interestingly, endotoxins increased neutrophil adhesion to clean glass surfaces, but stimulated the cells to exhibit increased random locomotion (chemokinesis) through cellulose nitrate filters and show an enhanced ability to reduce nitroblue tetrazolium (NBT) dye. Unlike neutrophils, lymphocytes direct from blood do not show polarized morphology towards chemotactic factors but the cells acquire locomotor capacity during 24-72 h culture with mitogens such as phytohemagglutinin (PHA), phorbol myristate acetate or concanavalin A. Stimulation of blood lymphocytes with endotoxins did not induce cell polarization in short-term but long-term culture resulted in an increase in the proportion of polarized cells that acquired locomotor morphologies. The majority of these cells were identified as esterase negative B-lymphocytes that migrated through filters. Despite the optimum time of incubation for each of these cell types being different, we found that lymphocytes respond to much lower concentrations of endotoxins than the neutrophils. These findings suggest that endotoxins of enteric pathogens modulate the functions of human blood neutrophils and lymphocytes.

• Journal of Microbiology and Biotechnology
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• 제6권3호
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• pp.189-193
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• 1996

Submicron-sized polymeric particles (SSPP) were used to remove nucleic acids and endotoxins from cell lysates. The positively charged SSPP selectively adsorb nucleic acids and endotoxins and form complexes with them. The complexes can be easily removed by sedimentation or centrifugation. The removal of nucleic acids and endotoxins using SSPP also can be accomplished in the presence of cell and cell debris. Therefore, nucleic acids and endotoxins can be removed in an initial step of the down-stream processes. In bakers yeast and E. coli lysate systems, the level of DNA could be reduced more than three orders of magnitudes and endotoxins more than seven orders of magnitudes concurrently willi the cell debris removal process using SSPP.

• BMB Reports
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• 제35권5호
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• pp.482-487
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• 2002

Early activation of human peripheral blood polymorphonuclear neutrophils is characterized by their morphological changes from spherical to polarized shapes. The endotoxins from enteric pathogens (S. dysenteriae type 1, V. cholerae Inaba 569B, S. typhimurium, and K. pneumoniae) were assessed by their ability to induce morphological polarization of the neutrophils as measures of early activation. Phagocytic activity, adhesion, chemokinetic locomotion, and nitroblue tetrazolium (NBT) dye-reduction ability measured the later activation of the cells. Neutrophils showed distinct morphological polarization in suspension over a wide range of concentrations of these endotoxins when were compared with those that were induced by the standard chemotactic factor, N-formyl-L-methionyl-L-leucyl-L-phenylalanine (FMLP). It was discovered that all of the endotoxins induced locomotor responses in neutrophils in suspension that were dose- and time-dependent. The optimum concentration for the endotoxins of S. dysenteriae, V. cholerae, and K. pneumoniae was 1 mg/ml in which 71, 69, and 66% of the neutrophils were polarized. However, the S. typhimurium dose was 2 mg/ml in which 50% of the cells responded. Neutrophils that were stimulated with endotoxins also showed increased random locomotion (p<0.005) through cellulose nitrate filters, but an enhanced adhesion of the cells to glass surfaces (p<0.03). These are important functions of these cells to reach and phagocytose damaged cells, as well as invading microorganisms. Interestingly, the endotoxins had a highly-significant inhibitory effect upon the proportions of neutrophils phagocytosing opsonized yeast (p<0.01) with a small number of yeast that were engulfed by the cells (p<0.02). Further, endotoxin-treated cells showed an enhanced ability to reduce NBT dye (p<0.03). Therefore, we concluded that endotoxins of enteric pathogens are neutrophil chemotactic factors.

• 한국환경보건학회지
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• 제36권3호
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• pp.191-198
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• 2010

The aim of this study was to determine the concentrations of air-borne microorganisms (bacteria, fungi, gram negative bacteria (GNB), endotoxins, and respirable suspended particles (RSPs), and their correlation with environmental parameters (temperature, humidity, and carbon dioxide levels) in duck houses. The mean concentrations of bacteria, fungi, and GNB were very high (> $10^6\;CFU/m^3$), and endotoxin levels exceeded $10^3\;EU/m^3$. Among the various work stages, in the task of receiving ducks, bacteria were $6.2{\times}10^6;CFU/m^3$, and GNB were $5.4{\times}10^6\;CFU/m^3$, while RSPs levels were $397.6\;{\mu}g/m^3$ and endotoxin levels were $47.2{\times}10^3\;EU/m^3$ in the task of dividing the ducks, and fungi were $4.9{\times}10^6\;CFU/m^3$ in the task of shipping the ducks. The concentrations of RSPs and endotoxins were significantly higher in tasks involving greater movement of ducks in the house, relative to tasks involving little movement of ducks (p<0.05). Further, with progression in growth of the ducks, the concentrations of bacteria, GNB, endotoxins, and RSPs were significantly increased. There was significant correlation between levels of GNB with bacteria (r=0.75) and fungi (r=0.86), endotoxins with RSPs (r=0.75), bacteria with fungi (r=0.39), and carbon dioxide with RSPs (r=0.38), bacteria (r=0.33), and endotoxins (r=0.31). These results suggest considerable respiratory hazard for farmers in these environments.

• Journal of Microbiology and Biotechnology
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• 제23권8호
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• pp.1099-1106
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• 2013

Tuta absoluta (Povolny, 1994) is a devastating moth to the Solanaceae plants. It is a challenging pest to control, especially on tomatoes. In this work, we studied the entomopathogenic activity of the Cry-forming ${\delta}$-endotoxins produced by Bacillus thuringiensis strain KS and B. thuringiensis kurstaki reference strain HD1 against T. absoluta. These strains carried the cry2, cry1Ab, cry1Aa/cry1Ac, and cry1I genes, and KS also carried a cry1C gene. The ${\delta}$-endotoxins of KS were approximately twofold more toxic against the third instar larvae than those of HD1, as they showed lower 50% and 90% lethal concentrations (0.80 and 2.70 ${\mu}g/cm^2$ (${\delta}$-endotoxins/tomato leaf)) compared with those of HD1 (1.70 and 4.50 ${\mu}g/cm^2$) (p < 0.05). Additionally, the larvae protease extract showed at least six caseinolytic activities, which activated the KS and HD1 ${\delta}$-endotoxins, yielding the active toxins of about 65 kDa and the protease-resistant core of about 58 kDa. Moreover, the histopathological effects of KS and HD1 ${\delta}$-endotoxins on the larvae midgut consisted of an apical columnar cell vacuolization, microvillus damage, and epithelial cell disruption. These results showed that the KS strain could be a candidate for T. absoluta control.

• 한국미생물·생명공학회지
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• 제41권2호
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• pp.242-248
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• 2013

Endotoxins are part of the outer membrane of the cell wall of gram-negative bacteria and are continuously released during bacterial growth. Endotoxins typically induce severe sepsis and septic shock, which cause more than 50% of mortalities. Endotoxins are easily measured in the serum by the limulus amebocyte lysate (LAL) test. However, a nonspecific result is obtained, because the high concentration of serum proteins disturbs the enzyme reaction of the LAL test. In order to solve this problem, the LAL test was performed in this study after the centrifugation of the boiled serum samples to remove the impurities. As a result, among the various conditions examined, endotoxin measurement with the LAL test was the most accurate and repeatable after centrifugation of the boiled serum at $100^{\circ}C$. Moreover, the endotoxin was accurately and repeatedly measured from the prepared sera of mice that had been administered an intraperitoneal injection of purified lipopolysaccharides (LPS) or E. coli. Therefore, the application of centrifugation to remove impurities from boiled serum gives an accurate measurement of endotoxins in the sera of normal subjects or patients, and this will lead to the improved diagnosis and prevention of diseases caused by endotoxins. In addition, the centrifugation of boiled serum samples should be considered and included in the development of endotoxin test kits.

• 한국산업보건학회지
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• 제31권4호
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• pp.331-341
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• 2021

Objectives: The aim of this study was to evaluate the concentrations and relationships of coarse particles with a diameter of 10 ㎛ (PM₁₀) with endotoxins according to the time of measurement in windowless poultry houses. Methods: In this study, measurement was performed on ten windowless poultry houses with a vertically integrated system from July to November. PM₁₀ was measured using personal environmental monitors and polytetrafluoroethylene (PTFE) filters with a 4 L/min-calibrated pump in selected sampling locations (two near the door and two near an exhaust fan). The endotoxin on PTFE filter was analyzed by the LAL turbidimetric method. Results: The range of geometric mean concentrations of PM₁₀ and endotoxins for each of the 38 samples were 0.12-3.30 mg/m³ and 11.9-3553.66 EU/m³, respectively. PM₁₀ and endotoxin concentrations varied by farm, increasing with the decrease in ventilation. The range of the coefficient of determination between PM₁₀ and endotoxin was 0.0009-0.9249. As the atmospheric temperature decreased, it was confirmed that the concentrations of PM₁₀ and endotoxin increased because the volume of ventilation was decreased. Conclusions: Endotoxins were more affected by time of measurement and ventilation than PM₁₀, which means that endotoxins could be an important indicator for intervention programs for improvement of indoor environments.

• Journal of Life Science
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• 제11권1호
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• pp.27-29
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• 2001

Vibrio vulnificus endotoxin was extracted, analyzed the chemical composition, tested its biological activity, and compared to those of Escherichia coli and Salmonella typhimurium. The major fatty acid of three endotoxins were different each other; V. vulnificus endotoxins were different each other; V. vulnificus endotoxin was myristric acid (C14:0), E. coli was lauric acid (C12:0), S. typhimurium was capric acid (C10:0). The biological activities of V. vulnificus endotoxin were similar to those of E. coli and S. typhimurium in terms of the gelation activity of the Limulus amebocyte lysate and the lethal toxicity. But the result of enzyme (AST, ALT, and LDH) analysis showed that the enzyme activity of V. vulnificus endotoxin was similar to that of E. coli, but smaller than that of S. typimurium.

• 한국미생물·생명공학회지
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• 제18권3호
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• pp.301-304
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• 1990

Bti와 Btd의 내독소에 대한 면역학적인 검토를 행한 결과 양균주의 내독소에는 동일한 기원의 단백질이 혼존하고 있음을 면역확산반응 및 면역전기영동법으로 확인되었다. 또한 동일한 기원의 단백질 함량은 Bti 내독소 보다 Btd 내독소에 많이 존재함을 효소항체법으로 확인하였다.

• Journal of Microbiology and Biotechnology
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• 제16권2호
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• pp.219-225
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• 2006

The rabbit pyrogen test and Limulus amoebocyte lysate (LAL) assay have been used to detect endotoxins present in vaccines. Currently, the rabbit pyrogen test is used to detect endotoxins in hepatitis B (HB) vaccines, even though the HB surface protein, which is the active ingredient, is overexpressed in and purified from eukaryotic cells that lack these endotoxins. Although the LAL clot assay is sensitive and reliable and can be used to replace the rabbit pyrogen test, its reaction is limited by the lack of responsiveness to the Gram-positive bacterial components. Furthermore, aluminum hydroxide in the HB vaccine can interfere with the LAL assay. In contrast, macrophages can detect the endotoxin as well as other pyrogens, and secrete $TNF-{\alpha}$. Therefore, this study was undertaken to examine the possibility of replacing the animal tests with a more efficient $TNF-{\alpha}$ secretion assay. With this in mind, we determined if aluminum hydroxide in the HB vaccines affects the $TNF-{\alpha}$ secretion assay. HB vaccines and the HB protein solutions spiked with lipopolysaccharide (LPS) produced the same level of dose-dependent $TNF{\alpha}$ secretion and temperature increase in rabbits, indicating that aluminum hydroxide in the HB vaccine does not interfere with the pyrogenic response in rabbits, nor does it interfere with $TNF-{\alpha}$ secretion. In addition, the $TNF-{\alpha}$ assay was found to be more sensitive than the LAL assay, and correlated well with the pyrogen test and the LAL assay. These results suggest that the $TNF-{\alpha}$ assay in RAW264.7 cells is a good substitute for the current pyrogen assays that are used for detecting LPS in HB vaccines as well as in other vaccines containing aluminum.