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Objective To establish an inhalation infection pneumonia model of C57BL/6J mice with highly virulent and multi-drug resistant Pseudomonas aeruginosa(PA)strain F291007,and to study the microbiological,pathological and immunological characteristics of this model.Methods The strain F291007 was isolated and identified before the bacterial suspension was administered to the mice via aerosolized intratracheal inoculation to establish the pneumonia infection model.In the course of infection,the conditions and survival of the mice were observed,and the bacterial loads,the histopathological states and the cytokine expression levels in the major organs were detected.Finally,three key cytokines were blocked to observe the survival of mice.Results The strain F291007 was isolated and identified.After lethal dose infection,all the mice died within 24 h.After sub-lethal dose infection,a large number of immune cells in the body were capable of phagocytosis and killing of invading pathogens,which was manifested as rapid clearance of bacteria in lungs and the exponential decrease of bacterial load with the passage of time.The pathological changes in lungs were most severe at 1 to 3 days but gradually recovered.After infection,interleukin-6(IL-6),IL-17A and tumor necrosis factor-α(TNF-α)in alveolar lavage fluid and serum were significantly increased at 1 to 3 days.After blocking of these three cytokines with specific antibodies,the survival rates of infected mice decreased significantly.Conclusion A mouse model of gradually-recovered pneumonia infection caused by PA inhalation has been established,suggesting that the first one to three days are critical to immune response after infection through multiple indicators.This mouse model can be used for research on the pathogenesis,immunoregulation and treatment evaluation of highly virulent and multi-drug resistant PA inhalation pneumonia infection.
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Objective To construct a non-trace deletion mutant of exlA in Pseudomonas aeruginosa strain NY8755(NY8755ΔexlA)and investigate the basic characteristics of pore-forming toxin ExlA.Methods The NY8755ΔexlA was constructed using the secondary homologous recombination method.C57BL/6J female mice ages 6 to 8 weeks were infected with NY8755 and NY8755 ΔexlA via aerosolized intratraheal inoculation respectively.Within 7 days of infection,the survival and weight changes of the mice were observed and recorded before the proinflammatory cytokines in the bronchoal-veolar lavage fluid(BALF)of the infected mice in the two groups were detected.Results The sequencing results showed that NY8755 ΔexlA was constructed.After 1×107 CFU NY8755 and NY8755 ΔexlA were infected,all the mice in the wild-type strain group died within 48 hours,while those in the mutant strain group began to die after 48 hours,and 40%of them remained alive 7 days later.The weight of surviving mice in the mutant strain group decreased but recovered gradually.After 12 hours of infection,there were more bloody exudates(redder in color)in the BALF of the wild-type strain group than in the mutant strain group,and the contents of proinflammatory cytokines interleukin-1β(IL-1β)and interleukin-17A (IL-17A)were significantly different. Conclusion Pseudomonas aeruginosa pore-forming toxin ExlA is the key pathogenic virulence factor of the exlA-positive Pseudomonas aeruginosa,which can significantly affect the survival status of mice and cause obvious inflammation in mice. Very little information is available on the action mechanisms of ExlA. In this study, The NY8755ΔexlA and the C57BL/6J mouse models infected with NY8755 and NY8755ΔexlA have been constructed that may be used for the investigation of pathogenesis of exlA-positive Pseudomonas aeruginosa.
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Objective To develop a micro-circumstance airtight cabin for in the study of biological aerosols detection with such functions as airflow control and temperature and humidity detection .Methods Wind speed sensors , temperature and humidity sensors , electrical control valves , high efficiency filters and the vacuum pump formed the micro-circumstance regulating system .The techniques of airflow direction control , temperature compensation , air pressure control and aerosol uniformity distribution were used .Numerical simulation of aerosol concentration distribution in the airtight cabin was achieved using Fluent software .The bioaerosol concentration in different locations was tested by experiments .Results The micro-circumstance airtight cabin consisted of an airtight cabin and a control cabin .The control cabin used a single-chip microprocessor to provide air supply and exhaust air to the airtight cabin in a seaparate exhaust mode and cyclic ventilation mode.It worked under a negative pressure condition .Through numerical simulation,the aerosols were distributed through-out the cabin after five minutes of generation and the bottom airflow arrived at the top .The distribution of aerosol concentra-tion was approximately uniform .Conclusion The micro-circumstance airtight cabin is suited to various bioaerosols testing research thanks to its negative pressure working without bioaerosol leakage .
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Objective To investigate the immunogenicity of mycobacteriophage D29 (phage D 29) in guinea pig models with different delivery routes,and provide information for the application of phages in tuberculosis (TB) therapy.Methods Hartley guinea pigs were administrated with phage D29 through inhalation,intranasal drop or subcutaneous injection for 6 times within 35 days.7H9 broth aerosol inhalation and 0.85 ％ NaCl solution aerosol inhalation were set as solvent and negative controls,respectively.Anti-phage D29 neutralizing antibodies in sera collected weekly were measured by phage reduction neutralizing test (PRNT) and cytokine levels (interleukin-2,interleukin-4 and interferon-γ) were detected at day 35 by enzyme linked immunosorbent assay (ELISA).The data were analyzed by ANOVA and nonparametric test.ResultsNeutralizing antibodies were both negative in two control groups,while low-titer neutralizing antibodies (below 1 ∶ 100) appeared in inhalation and intranasal drop groups only at day 7 and day 14. Nevertheless, neutralizing antibodies were continuously detected in subcutaneous injection group,which increased rapidly and reached 1∶ 16 365.6 at day 35. After 35 days of experiments,serum concentrations of interleukin-2 (x2 =2.7605,P＞0.05),interleukin-4 (F=2.17,P＞0.05) and interferon-γ(F=0.75,P＞0.05) among three treatment groups and two control groups were all not significantly different.ConclusionsThe titer of anti-phage 29 neutralizing antibodies induced by inhalation or intranasal drop administration of phage D29 are both significantly lower than subcutaneous injection.Phage D29 administration doesn’t change the levels of cytokines,which indicates that it may not break the helper T cell (Th)1/Th2 balance.