ABSTRACT
Objective To evaluate the therapeutic effect of antibiotics combined with recombinant tuberculosis vaccine AEC/BC02 on Mycobacterium tuberculosis infection in guinea pigs. Methods Two weeks after guinea pigs were challenged subcutaneously with a high dose of Mycobacterium tuberculosis,the guinea pigs with the positive skin test responses to the recombinant ESAT6-CFP10 allergen were randomly di-vided into four groups:normal saline (NS),AEC/BC02,antibiotics and antibiotics+AEC/BC02. In antibiotics+AEC/BC02 group,guinea pigs firstly received isoniazid ( INH) and rifapentine ( RFT) treatment once a week for a total of three times,and then were immunized with a single dose of AEC/BC02 vaccine six times at 10-day intervals. Guinea pigs in AEC/BC02 and antibiotics groups were respectively vaccinated with AEC/BC02 vaccine and given INH and RFT treatment at the same dose and frequency as given to antibiotics+ AEC/BC02 group. Thirteen weeks after challenge,all guinea pigs were sacrificed for necropsy. Results The gross pathological scores of NS,AEC/BC02,antibiotics and antibiotics+AEC/BC02 groups were 83±8,77± 22,45±28 and 19±14,respectively. Antibiotics+AEC/BC02 group had a significantly lower gross pathological score than antibiotics,AEC/BC02 and NS groups (P<0. 05,P<0. 01,P<0. 01,respectively). Moreover,the gross pathological scores of antibiotics and AEC/BC02 groups were significantly decreased as compared with that of NS group (both P<0. 01). However,there was no significant difference between AEC/BC02 and NS groups. The spleen bacterial load of antibiotics+AEC/BC02 group was (2. 50±1. 26) lg CFU,which was sig-nificantly lower than those of NS [(4. 92+0. 52) lg CFU],AEC/BC02 [(4. 78+0. 84) lg CFU] and antibi-otics [(4. 39+0. 50) lg CFU] groups (P<0. 01). Compared with NS group,antibiotic and AEC/BC02 groups showed no significant difference in spleen bacterial load. Histopathological changes indicated different levels of granulomatous lesions appeared in lung tissues of all groups and the most severe change was ob-served in AEC/BC02 group,followed by that in NS,antibiotics and antibiotics+AEC/BC02 groups. Conclu-sion INH and RFT treatment in combination with AEC/BC02 vaccine in the treatment of guinea pigs with Mycobacterium tuberculosis infection was superior to either treatment alone as it significantly alleviated organ lesions and lowered the bacterial loads in spleen and lung.
ABSTRACT
Objective To identify the cross-reactive antigens shared by Mycobacteria smegmatis(MS) and Mycobacteria tuberculosis(MTB) and to analyze their antigenicity.Methods Bacterial antigens were extracted from strains of MS and MTB by ultrasonication.Western blot assay was performed to analyze common antigens that reacted with both of the antiserum samples against MS and MTB.The extracted bacterial antigens were mixed with incomplete Freund′s adjuvant and then were injected into muscles of mice.Cytokines secreted by murine spleen lymphocytes following stimulation with various antigens of MS and MTB were determined by ELISPOT and flow cytometry on the 7th day.IgG levels in serum samples were detected by ELISA 7 days after injection.Results There were cross-reactive antigens shared by MS and MTB.Potent humoral immune responses and cellular immunity against both MS and MTB could be induced by those cross-reactive antigens after sensitization the mice by either MS or MTB antigens.Cytokines of IL-2 and IFN-γ in CD4+ and CD8+T cells of mice stimulated with MS or MTB antigens were significantly increased as compared with those of non-sensitization group and those of Brucella antigens stimulation group.ConclusionCross-reactive antigens shared by MS and MTS can effectively promote specific immune reactions to the infection of MTB, which provides a scientific basis for the development of tuberculosis vaccines.
ABSTRACT
More than 60% of active tuberculosis(TB) patients are smear- and culture-negative, constituting a prime group in the prevention and control of TB in China. In the existing laboratory testing technologies, immunological diagnosis is more advantageous than etiological diagnosis in the detection of smear-and culture-negative TB. Serum antibody detection reagents are cheap,easy to operate and time-sav-ing,and have been widely used in China. However,these agents are not stable in sensitivity and specificity, and because of that their accuracy in the diagnosis of smear-and culture-negative TB is doubtful. In this re-view,we summarize some problems in the use of serum antibody detection among smear- and culture-nega-tive pulmonary TB patients and discuss possible methods to solve these problems expecting to provide some ideas for promoting its development,application and policy formulation.
ABSTRACT
Objective To investigate the effect of risperidone on S100B protein and its intervention on the relapse of heroin addicts. Methods From January 2014 to March 2014, 70 cases heroin addicts in the second detection center in Xining were selected as objects, three patients dropped out, and the left were randomly divided into control group(n=33) and experimental group(n=34) , which were given detoxification for 10 days, rehabilitation for 5 days, while the experimental group were given risperidone intervention.The condition of craving degree for heroin, relapse and incidence of adverse reactions between two groups were compared.Serum S100B protein levels were detected by ELISA.The efficacy of risperidone was analyzed.Results The craving degree for heroin between two groups was of no significant differences, after drug treatment, the craving degree for heroin of experimental group were significantly lower than those of control group at the time-point of hospital discharge, 1 months after discharging and 3 months after discharging(P=0.013, P =0.008, P =0.005).The differences of relapse condition between two groups in 3 months after discharging were statistically significant (P=0.003), the relapse incidence of experimental group(11.8%) was significant lower than that of control group(54.5%). Besides, the differences of relapse condition among different craving degree groups were statistically significant in two groups respectively(P=0.005, P=0.008).Serum S100B protein levels of experimental group , in different periods were lower than those of control group, with statistical significance ( P<0.05); There were 5 cases of insomnia, 3 cases of headache, 3 cases of thirsty, 2 cases of mild extrapyramidal symptoms in test group, but all above were mild and relieved on their own after withdrawal, no similar symptoms were observed in control group.Conclusion Risperidone intervention is a safe and effective method to prevent heroin addicts from relapsing, it maybe with the mechanism of reducing the levels of serum S100B protein, but it needs further research and efforts to put it in clinical applications of herion addiction treatment.
ABSTRACT
ABSTRACT:The guinea pigs were immunized by the Brucella vaccine through intradermal injections and the skin scratch respectively ,and then the immune effects of the two ways were evaluated .Serum samples were collected one month after the last injection and detected for the total IgG titer by interval ELISA .Cell‐mediated immune was evaluated by late‐onset hyper‐sensitivity .The guinea pigs were challenged with Brucella melitensis M5 ,and then were killed to isolated M5 from spleen of each guinea pig to compare the protective effects of two methods of immunization .The ELISA results showed that both of the two methods of immunization could induce strong humoral immune response ,and DTH response to Br‐PPD antigen were 100%in both methods .No significant difference in the immune protective effect of two methods was detected .Results of humoral im‐munity ,cellular immunity and protective effect showed the same effect by intradermal injections and skin scratches .
ABSTRACT
Objective To establish a suitable guinea pig model for evaluating the protective effects of new TB vaccines in BCG prime-boost regimen .Methods Two different immunization strategies by using the recombinant TB vaccine were employed to boost BCG primed guinea pigs in this study .One was for short-term evaluation with 14 weeks interval between prime and boost immunization and another was for long -term evaluation with 54 weeks interval .In the short-term evaluation group , guinea pigs were boosted twice with the recombinant TB vaccine ( AEC/BC02 ) in every two weeks , while guinea pigs in the long-term evaluation group were boosted for three times with two weeks interval between each injection .A negative con-trol group ( NS→NS) and a BCG control group ( BCG→NS) were both set up in two evaluation groups .One week after the last immunization , all guinea pigs were challenged with M.tuberculosis.Six to seven weeks after bacteria challenge , all animals were euthanized and dissected to evaluate lesion scores of liver , spleen and lung, as well as the viable bacterial load in spleen .Results In the short-term evaluation group , the le-sion scores in those boosted with vaccine (3.33±5.00) was lower than that of BCG control group (5.56± 7.27) (P>0.05) and negative control group (47.00±28.11) (P=0.0001).The difference between BCG control group and negative control group in lesion score was also significant .The animals in vaccine boosted group had lower bacterial loads (0.78±1.55 log10 ) in spleen than that in BCG control group (1.06±1.87) (P>0.05) and negative control group (5.47±0.61) (P=0.0003).In the long-term evaluation group, the lesion score in those boosted with vaccine was lower (5.0±7.6) than that in BCG control group (14.4± 13.5) (P=0.0394) and negative control group (56.9±14.1) (P0.05) and negative control group (5.43±0.56) (P=0.01).There was a significant difference in bacterial load between BCG control group and negative group (P=0.0089).Conclusion The results suggest that the interval time between BCG-prime and boost immunization should be properly prolonged in the guinea pig model used for evaluating the protective effects of new TB vaccines in BCG prime -boost regimen .
ABSTRACT
Objective To establish a guinea pig model of latent Mycobacterium tuberculosis infec-tion for evaluating the effects of therapeutic vaccines .Methods Guinea pigs were subcutaneously inocula-ted with 5.0×103 CFU Mtb.The skin test was performed with 0.5μg recombinant ESAT6-CFP10 protein to detect positive conversion rates at different time points .Two weeks after Mtb inoculation , guinea pigs in model group received 5 mg isoniazid treatment ( three times a week for four weeks ) by oral gavage , while those in control group received normal saline .At the sixth week after Mtb infection , guinea pigs with and without isoniazid treatment were dissected for pathology examination .The pathological scores of liver , spleen and lung, as well as bacteria loads in spleen were compared between two groups .The established guinea pig model of latent infection was then validated by testing two reference vaccines ( AEC/BC02 and AEC/BC03 ) . Results Two weeks after Mtb inoculation , all guinea pigs showed positive EC skin test with induration area of (19.9±3.0) mm.Upon four weeks of isoniazid treatment , the guinea pigs in model group showed no pathological changes with zero scores in the examined organs .No bacterium was detected in spleen of ani-mals from model group.However, the total pathological score was 38.8±16.5 and bacteria load in spleen was (5.1±0.3) Log10 CFU with the guinea pigs from control group .Natural recurrence of tuberculosis in model group was observed after drug withdrawal .The total pathological scores were 48.5±23.9 and 51.3± 23.41.The bacterial loads in spleen were (4.5±1.3) and (4.2±1.1) Log10 CFU and bacterial loads in lung were (4.1±1.2) and (3.4±1.3) Log10 CFU respectively as verified with reference vaccines of AEC /BC02 and AEC/BC03.Conclusion Isoniazid treatment inhibited the proliferation of inoculated Mtb in guinea pigs.A guinea pig model of latent Mycobacterium tuberculosis infection is successfully established with an advantage of good repeatability .Therefore, it can be used to evaluate the effects of therapeutic vaccines on latent Mycobacterium tuberculosis infection.
ABSTRACT
Objective To establish a suitable guinea pig model for evaluating the protective effects of new therapeutic TB vaccines in preclinical study .Methods The guinea pigs were subcutaneously injected with single-cell suspension of multi-drug resistant M.tuberculosis at a dose of 1000 CFU, 0.2 ml per animal.The study was divided into experimentⅠand experimentⅡ.In experimentⅠ, the guinea pigs were given immuno-therapy and/or chemical treatment on day 3 after infection .Four guinea pigs in each group were dissected at weeks 5, 7 and 9 after infection for evaluating lesion scores and histopathology changes of liver , spleen and lung, as well as bacterial load in spleen .In experimentⅡ, the guinea pigs were given immunotherapy and/or chemical treatment with different doses on day 14 after infection .All guinea pigs were dissected at week 5 after infection for evaluating lesion scores of liver , spleen and lung , as well as bacterial load in spleen .Results In experimentⅠ, all of the three treatments including immunotherapy combined with chemotherapy , immunotherapy alone and chemotherapy alone could effectively prevent organ lesion and reduce bacterial load in spleen , which were significantly different from negative control group .The immunotherapy combined with chemotherapy showed better treatment effects than other two treatments .Along with a prolonged period after drug withdrawal , the de-gree of organ lesion in immunotherapy group and chemotherapy group rebounded sharply , but only slightly in im-munotherapy combined with chemotherapy group .In experimentⅡ, animals in all treatment groups showed alle -viated organ lesion and reduced bacterial load in spleen .A relatively better treatment effect was observed in im-munotherapy combined with chemotherapy group .Conclusion The established guinea pig model of M.tubercu-losis infection showed an advantage of good repeatability .It might be used to evaluate the protective effects of new therapeutic vaccines against tuberculosis in preclinical study .
ABSTRACT
Objective To comparatively evaluate the effects of a recombinant Mtb protein ESAT 6-CFP10 ( rESAT6-CFP10 ) and a purified protein derivative ( PPD ) as skin test reagents in guinea pigs . Methods Guinea pigs were sensitized with different Mycobacteria species .After sensitization , all guinea pigs were intradermally injected with rESAT6-CFP10 and PPD.At 48 h after the injection, the size of ery-thema at injection sites was measured by using a double-blind method .For guinea pigs sensitized with viable Mtb, the size of erythema at injection sites were measured at 24 h after the injection .The positive conversion rates of skin test with rESAT 6-CFP10 and PPD were calculated .Results The results of PPD skin test were positive in all guinea pigs sensitized with viable Mtb , killed Mtb and BCG with erythema diameters of (11.4 ±0.9) mm, (11.8±1.1) mm and (13.2±0.8) mm, respectively.Positive skin test with rESAT6-CFP10 was only observed in guinea pigs infected by viable Mtb-showing erythema diameters of (13.7±5.7) mm. The skin test with rESAT6-CFP10 was negative in guinea pigs sensitized by killed Mtb-and vaccinated by BCG.The skin tests by using rESAT6-CFP10 and PPD were performed on randomly selected guinea pigs at ninth day after infection by Mycobacterium tuberculosis H37Rv.At the 2nd week, totally 24 selected guinea pigs showed positive skin test results with rESAT6-CFP10 (24/24) with erythema diameters of (19.9± 3.0) mm, while only 15 out of 24 had positive PPD skin test with erythema diameters of (6.1±5.5) mm. At the 4th week, all guinea pigs showed positive PPD skin test (3/3) with erythema diameters of (12.7± 2.5) mm.Conclusion The skin test by using recombinant ESAT 6-CFP10 protein can effectively distin-guish viable Mtb infection from BCG vaccination and killed Mtb sensitization , which is a more suitable anti-gen than PPD for the early diagnosis of Mtb infection .
ABSTRACT
Objective To establish guinea pig model of type Ⅰ anaphylaxis, isolate the anaphy-lactic antibody(IgE and IgG) preliminarily from sera of sensitized guinea pigs, and investigate its functional characteristics. Methods Animal model was established by sensitizing guinea pigs with OVA and Al(OH)_3, level of antibody was determined by ELISA, IgE and IgG in sera were preliminarily isolated through saturated ammonium sulphate precipitate and affinity chromatograph of Protein A. A continuous passive cutaneous ana-phylaxis test (PCA) was performed by sensitizing guinea pigs individually with IgE and IgG and then challenging at different time , and the variation of blue spots in skin were observed after challenge . Results Concentration of IgE in model group and control group were 719.3750 ng/ml and 2.5250 ng/ml, the optical density of IgG in model group and control group were 0.9921 and 0.0174, the level of two antibodies in model group were significantly higher than that of control group (P<0.05). In 9 d continuous PCA test, the blue spot induced by IgE in skin lasted for 9 days and appeared the largest when challenged at day 5. The diameter of blue spot induced by IgG was the largest when challenged at day 2 and then decreased fast. Con-clusion Anaphylactic antibodies were successfully preliminarily isolated from sera of sensitizing guinea pig, both IgE and IgG play roles in type Ⅰ anaphylaxis of guinea pig, the hypersensitive reaction induced by IgG is fast and short than that induced by IgE, and IgG may become an important surrogate marker in immunotoxic-ity evaluation(type Ⅰ anaphylaxis)of vaccine.
ABSTRACT
OBJECTIVE To isolate Acinetobacter bacteriophages and analyze their lysis spectrum.METHODS A total of 78 Acinetobacter strains were isolated from the sewage of different sources whose lysis spectrum was determined by method of single layer plates.RESULTS From them only two kinds of Acinetobacter bacteriophages AB46 and AB54 with different lysis spectrum were isolated from 2 strains.The former had a lysis spectrum of 46(1.28%)and the latter had a lysis spectrum of 54,57,59,60,62,69,70 and 78(10.27%).CONCLUSIONS Acinetobacter bacteriophages are isolated and their lysis spectrums are obtained.The lysis spectrums are narrow which need to improve their lysis ranges but they can infect their hosts virulently and form transparent plaques.
ABSTRACT
Objective To study a new and rapid method for detection of anti-histoplasma antibody by dot immunogold filtration assay (DIGFA). Methods DIGFA was developed by coating purified protein derivative of histoplasmin (P-HTPM) on nitrocellulose membrane as membrane antigen and labeling SPA with colloidal gold. Anti-histoplasma antibodies in sera from mice immunized with Histoplasma were detected by DIGFA. Results All immunizedsera with Histoplasma were positive by DIGFA as well as the results detected by ELISA. The sera immunized respectively with Monilia and Penicillium marneffei were negtive by DIGFA while 1 of 8 immunizedsera with Blastomyces dermatitidis and 2 of 9 immunizedsera with Paracoccidioides brasiliensis were found to be positive. Conclusion DIGFA was a valuable and rapid method to detect histoplasmaantibody in serum.
ABSTRACT
Objective To realize the simultaneous control of two laboratory instruments by a computer. Methods On the basis of analysing the mode of communication between computer and laboratory instrument, the author expanded the computer's COM interface through the USB / COM converter. Results The author realized a computer controlling MP280 CLIA chemiluminescence analyzer and anthos 2010 enzyme tester at the same time. Conclusion User can use computer's USB interface and USB / COM converter to achieve controlling multiple instruments and equipments at the same time, as long as the communication interface of equipment is open. This method has preferable potential application.
ABSTRACT
BCG Purified Protein Derivative (BCG-PPD)was isolated and purified from BCG Culture filtrate by trichloroacetic acid and ammonium sulfate methods. The purity of BCG-PPD was Similar to PPD-S(international standard) and PPD-C(China), but more than that of PPD-CT68 (Canada)and PPD-RT23(Danish). The Delayed-Type Hyperseusitivity(DTH) to BCG-PPD was more sensitivity than other PPD on BCG vaccinated guinea pigs, but less sensitivity than other PPD on Mycobacterium tuberculosis infected guinea pigs. The conversion rate and induration diameter to BCG-PPD was higher than PPD in 333 of 12 weeks after BCG vaccination newborns, but lower than that of other PPD in 97 tuberculosis patients. It was shown that DTH reaction to PPD was more sensitivity in Mycobacteria homogeneous strain vaccinated individual than Mycobacteria heterogeneous strain vaccinated individual. It was demonstrated that BCG-PPD was better than other PPD on observation conversion rates and induration diameter of BCG vaccinated individual. It maybe help to identification BCG vaccinated or tuberculosis infected with DTH of BCG-PPD and PPD in same individual.
ABSTRACT
Objective To evaluate the quality of immunological diagnosis kits of schistosomiasis. Methods The national reference IgG serum of schistosomiasis was applied for evaluating eight immunological diagnosis kits of schistosomiasis. Results All eight diagnosis kits had no National Production Certificate issued by State Food and Drug Administration. Six kits had good quality with high precision. After the physical examination, two kits did not meet the quality standards. Conclusions The good and bad quality of immunological diagnosis kits of schistosomiasis used in market is intermingled. The quality of these kits should be monitored.