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1.
Twenty isolates ofMycobacterium tuberculosis resistant to rifampicin(RIF), isoniazid(INH) and streptomycin(STR) were analysed by Polymerase Chain Reaction (PCR) amplification of rpoB, katG and rrs genes to evaluate comparative diagnostic significance of genetic assays. Mutations were identified by single strand conformation polymorphism (SSCP) and cleavase fragment length polymorphism (CFLP) and were confirmed by DNA sequencing. SSCP of 4 RIF resistant and 14 INH resistant isolates showed an extra peak at the level of 75-bp and 85-bp respectively, while 2 STR resistant isolates showed 2 peaks with 9 bases difference. CFLP showed a different pattern among RIF, INH and STR sensitive and resistant isolates Thus SSCP and CFLP can be used as alternative diagnostic methods for identification of mutations in RIF, INH and STR resistant strains of M.tuberculosis.  相似文献   

2.
We have developed a simple, economical and reproducible method for processing blood samples from HIV infected patients for diagnosis of tuberculosis. The procedure was validated on 55 samples selected for tuberculosis based on clinical criteria. 52 patients had radiological changes indicative of pulmonary tuberculosis of which only 28 were positive for AFB in sputum (sensitivity 54%) and 27 for tuberculin (sensitivity 52%). 26 HIV positive patients who showed positive X-ray did not react to tuberculin. The genus PCR probe missed 3 samples (sensitivity 94%) compared to X-ray.M.tuberculosis was detected in the blood of all X-ray positive cases by PCR using TB400 probe (sensitivity 100%) and another probe forM. tuberculosis, IS6110, missed 6 of them (sensitivity 88% compared to X-ray and 89% compared to TB400). It is proposed that this simple sample processing method could be used to screen all blood samples quickly for mycobacteremia using the genus PCR and only those positive for mycobacteria need to be tested forM.tuberculosis. This would save the scarce resources and time by reducing significantly the number of samples to be screened for species confirmation.  相似文献   

3.
We have isolated and identified the biotype of environmental mycobacteria from the expectorate of leprosy patients, their contacts, their drinking water supply and also from the sputa samples of tuberculosis patients. 78% of the isolates from lepromatous leprosy patients and their contacts wereMycobacterium fortuitum- chelonae complex (MFC), 9%Mycobacterium avium complex (MAC), 9%Mycobacterium scrofulaceum and 4% wereMycobacterium smegmatis. Among the isolates from tuberculosis patients 63% belonged toM. fortuitum- chelonae complex, 19% toM. avium complex, 12% toMycobacterium Kansasii and 6% toM. smegmatis. All the isolates were multi-drug resistant when tested for sensitivity total of 21 drugs. TheMycobacterium fortuitum-chelonae complex organisms from leprosy contacts were more sensitive to rifampicin than those isolated from lepromatous leprosy and tuberculosis patients. Among 23 isolates from leprosy patients one isolate was resistant to 20 drugs, one isolate to 17 drugs and another isolate was resistant to 13 drugs. Among the 18 isolates from drinking water supply six showed resistance to more than 12 drugs. Polymerase Chain Reaction (PCR) and subsequent hybridisation with specific probes confirmed all the isolated strains as nontuberculous mycobacteria (Using genus primers and probe sensitivity 100%) and none asM. tuberculosis, suggesting that PCR could be used to rapidly identify mycobacteria at the genus level and to rule out tuberculosis in leprosy patients at an early stage to decide on appropriate course of therapy.  相似文献   

4.
A C-type mannose specific lectin (LRP) isolated from plasma of fresh water fishLabeo rohita showed 500 times higher specificity towards cell surface oligosaccharide (LAM) ofMycobacterium tuberculosis (H37Rv). Using biotinylated LRP, binding between lectin and LAM was demonstrated by ELISA and it was observed that even 3ng of oligosaccharides might be detected using only 1μg of biotinylated lectin.  相似文献   

5.
Allicin was prepared from the ethanol extract of garlic without the use of heat. It inhibited the growth ofMycobacterium tuberculosis H37Rv andM. tuberculosis TRC-C 1193 resistant to isoniazid completely when grown as surface cultures as well as shake cultures. The minimum inhibitory concentration (MIC) was 70 μg/ml for both organisms. Further purification of allicin over silica gel-G columns gave a chloroform elutable fraction called CEF-allicin which inhibited the growth of both the susceptible and the isoniazid resistant strains ofM. tuberculosis. Its MIC is 25 μg/ml and is the lowest reported so far when compared with very high inhibitory concentration of 1 to 3mg/ml reported by other workers. It completely inhibited the synthesis of lipids and DNA from14C-acetate and3H-thymidine respectively and almost completely that of proteins and glycine derived carbohydrates from14C-glycine ofM. tuberculosis within 6 hr of exposure to CEF-allicin.  相似文献   

6.
The antibody response to the 38kDa, 16kDa and Lipoarabinomannan (LAM) antigens ofMycobacterium tuberculosis was evaluated using three different ELISAs based on these antigens. The study group included tuberculosis patients (n=52), patients with HIV and TB co-infection (n=10), other chest symptomatics (n=5), HIV infected individuals (n=10), leprosy cases (n=7) and healthy controls (n=75). The results indicate that the 38kDa and LAM based ELISA for IgM/IgG has a low specificity (ranging from 69–85%) and sensitivity (ranging from 55–78%). When three ELISAs are carried out on a single patient the probability of detection of tuberculosis was significantly increased to 95.2% indicating that a single ELISA test is of low sensitivity and that a combination of ELISA’s may be needed to be of any value as a diagnostic test for tuberculosis. Additionally, a western blot assay of the serum antibody response to protein fraction ofM. tuberculosis was analysed in 15 tuberculosis patients and five healthy controls. A multiple antibody response to various M.tuberculosis proteins was observed which varied from patient to patient as compared to controls who showed a single 38–39 kDa protein band positivity. These finding suggest that a western blot assay which determines the antibody response to a set of antigenic components ofM. tuberculosis could be a better serological test for the diagnosis of tuberculosis in our population.  相似文献   

7.
Hemoglobin E (beta-26Glutamic acid→Lysine) is the second most prevalent hemoglobin variant in the world. 293 blood samples from cases referred from several hospitals in the region of Andhra Pradesh were screened for the detection of hemoglobinopathies. Four samples were found to be in heterozygous state for Hb E condition. Mutation in two of these heterozygotes was analysed using a 722 base pair (bp) amplified DNA fragment from beta-globin gene and restriction enzyme Mnl 1. A 232bp DNA fragment was found to be associated with the Hb E mutation.  相似文献   

8.
The effect of the antitubercular drugs isoniazid (10 μg/ml), ethambutol (10 μg/ml), rifampicin (0.5 μg/ml) and streptomycin (1 μg/ml) on the calmodulin like protein (CAMLP) content ofMycobacterium tuberculosis H37Rv andM. tuberculosis H37Ra was investigated. The drugs were added to actively growing cells at their mid log phase of growth (14 days) and after 12 more hours of incubation, CAMLP was estimated. In both the mycobacteria, all the four antitubercular drugs CAMLP.  相似文献   

9.
Based on our demonstration earlier that ethanol extract, water extract and a compound purified from garlic possessedin vitro antitubercular activity against drug resistant and susceptibleMycobacterium tuberculosis, we tried the effect of garlic extract in 30 patients of tubercular lymphadenitis. For ethical considerations, two groups of patients, 30 each, were given antitubercular therapy (ATT) consisting of isoniazid, rifampicin, ethambutol and pyrazinamide for 30 days. For the next 15 days (31 to 45 days) group 1 patients received 3–6 garlic pearls per day in addition to ATT while group 2 patients received ATT only. From 46th day onwards both the groups received ATT only for 6–8 months. Antitubercular activity of the serum samples collected on 45th day was assessed by its effect on the growth ofM. tuberculois. The serum of group 1 patients showed significantly much higher antitubercular activity than that of group 2 patients. Further, there was relief of dyspeptic symptoms caused by ATT therapy in patients of group 1 with garlic plus ATT therapy but no change in group 2 patients with ATT only. Liver function and hematological tests were normal in both the groups after 6 months of therapy. Garlic extracts or compounds have a good potential as antitubercular(s) drug if given as a supplement to ATT.  相似文献   

10.
Water extract of garlic (Allium sativum) inhibited the growth ofMycobacterium tuberculosis H37Rv andM. tuberculosis TRC-C1193 susceptible and resistant to isoniazid respectively. The minimum inhibitory concentration (MIC) was slightly above 80 but less than 160 μg/ml and slightly above 100 but less than 200 μg/ml for the susceptible and resistant strains respectively. Gel filtration in Sephadex G-100 columns showed that two protein fractions (43 & 38 kD) possessed antitubercular activity with much lower MICs of 20–40 μg/ml and 30–60 μg/ml for susceptible strain. Water extract, when added to actively growingM. tuberculosis in their mid log phase prevented their further growth. The water extract of garlic inhibited the incorporation of14C glycine into whole cells by 81% in 6 hrs. indicating that the primary mechanism of action is by inhibition of protein synthesis.  相似文献   

11.
The white spot syndrome virus (WSSV) is one of the deadly pathogens of penaeid shrimps and other crustaceans. The WSSV virion consists of an enveloped rod-shaped nucleocapsid enclosing a large circular double stranded DNA genome of 305 Kb with 181 open reading frames. The two major structural genes, VP19 and VP28 were amplified from the genomic DNA of Chinese isolate of WSSV and cloned in pUCm-T vector and sub cloned in pET-30a (+) vector. The expressions of genes inE. coli (BL21) were confirmed by SDS-PAGE analysis. The clones were sequenced, submitted to the gene bank and the Xiang Shan strain of WSSV were compared with the previous reported sequence of WSSV of various regions which revealed that VP19 and VP28 gene sequences had certain differences from the sequences of similar genes of the isolate already reported. The recombinant proteins expressed, purified and characterized.  相似文献   

12.
Tuberculosis remains a major public health problem globally, with India being one of the high burden countries. The common causative agent is Mycobacterium tuberculosis but in developing countries M. bovis is reported as a potential human pathogen. Almost 20% of all reported cases of tuberculosis are of extra pulmonary form of disease. Diagnosis of extra pulmonary tuberculosis (EPTB) is not always possible with conventional methods, due to the long time required and the paucibacillary nature of samples; hence the need of rapid molecular methods. A prospective study was conducted on 300 patients of EPTB over a period of 5 years. These patients were suspected cases of tubercular meningitis, tubercular ascites and tubercular lymphadenitis. Samples analyzed were cerebrospinal fluid, ascitic fluid and lymph node fine needle aspirate. A two step PCR targeting hup B gene was used. Clinical response to anti tubercular therapy (ATT) was taken as positive (gold standard). PCR for hup B gene was positive in 147 samples out of 155 ATT responders. Of these 85.71% were infected with M. tuberculosis, 9.52% with M. bovis alone and 4.76% showed co infection with both M.tb and M. bovis. The sensitivity and specificity of PCR was 90.32 and 94.48% respectively.  相似文献   

13.
Little is known about host-parasite inter-relationship in the lymphatic filarial parasites. There is no information available about the ability of these parasites to acquire cholesterol, though it is known that in general, nematodes lack the ability to synthesise cholesterolde novo. In this study, we have shown that the filarial parasites also lack the ability to incorporate labelled acetate into cholesterol, indicating the absence of the machinery for cholesterol biosynthesis. We have further shown that they elaborate a 43 kDa surface receptor for acquiring LDL-bound cholesterol. We have shown by polymerase chain reaction the presence of a 860 bp fragment indicating the presence of the gene for LDL-related protein (LRP) in the human filarial parasiteWuchereria bancrofti in the genomic DNA. We have also shown that it is expressed as seen in the cDNA clones identified from an expression library.  相似文献   

14.
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15.
施曼玲 《科技通报》2007,23(1):41-45
从浙江宁波雪菜上获得病毒分离物NBXC,病毒提纯后,电镜下可观察到大量长约740 nm的线形粒子。间接ELISA检测结果证实NBXC是芜菁花叶病毒(turnip mosaic virus,TuMV)的分离物。利用IC-RT-PCR对病毒分离物NBXC的CP和HC-Pro基因进行PCR扩增,分别得到约0.8 kb和1.4 kb的两条条带,与预期的TuMV CP和HC-Pro基因大小吻合。扩增产物克隆后进行序列测定,CP基因序列长度为864个核苷酸,编码288个氨基酸;HC-Pro基因序列长度为1374个核苷酸,编码458个氨基酸。NBXC的CP和HC-Pro基因与已报道的TuMV其他分离物的核苷酸序列同源率分别为90.0%～98.3%和82.2%～96.9%,氨基酸同源率分别为95.8%～99.3%和95.6%～99.3%。从CP和HC-Pro基因核苷酸序列同源性来看,NBXC与TuMV芸薹属分离物有更近的亲缘关系,而与萝卜属分离物亲缘关系较远。  相似文献   

16.
Background: Imbalance in cholesterol homeostasis may lead to gallstone disease. Apolipoprotein B is sole component of low-density lipoprotein and plays an important role in cholesterol metabolism. The present study was carried out to explore the association of APOB 3′ VNTR, exon 26 XbaI and signal peptide insertion/ deletion polymorphisms with gallstone disease. 214 ultrasonographically proven gallstone patients and 322 healthy, age and sex matched controls were taken for the study. Genotyping was done using PCR followed by polyacrylamide gel electrophoresis for VNTR and insertion/ deletion analysis. For APOB XbaI polymorphism PCR product was digested with XbaI restriction enzyme, followed by agarose gel electrophoresis. All statistical analyses were done using SPSS v11.5. Higher repeat alleles of APOB 3′ VNTR polymorphism were more frequent in gallstone patients than in controls. Alleles with more than 57 repeats were present only in patient group. Long (L) alleles with repeat higher than 49, were significantly higher (P=0.000; OR=3.705, 95% CI 2.577–5.326) and medium (M) alleles were lower (P=0.000; OR=0.406, 95% CI 0.304–0.542) in patients than in controls. To nullify the effect of gender, data was further stratified into male and female population. APOB 3′ VNTR, L alleles were imposing risk and M alleles were protective in both male and female population. APOB XbaI and insertion/deletion polymorphisms were not found to be associated with the gallstone disease. Longer alleles of APOB 3′ VNTR occur more frequently in gallstone patients, and may be an important risk factor for the development of gallstone disease. APOB XbaI and signal peptide insertion/deletion polymorphisms may not be contributing to the risk for gallstone disease.  相似文献   

17.
Trichloroacetic acid (TCA) solubilized and DEAE fractionatedMycobacterium tuberculosis H37Ra excretory-secretory (ES) antigen viz., Mtb EST DE1 and affinity purified goat antibodies to the TCA solubilized ES antigen (Mtb EST) were explored in detecting tubercular antibody and antigen respectively in sera of bone and joint tuberculosis by indirect and sandwich ELISA. Out of total 36 bone & joint tuberculosis cases, tubercular antibody was detected by indirect ELISA in 30 patients (sensitivity 83%), while circulating tubercular antigen was detected by sandwich ELISA in 27 patients (sensitivity 75%). Out of 34 non tubercular disease control cases, 10 patients showed positive reaction for antibody while only 4 patients showed positive reaction for antigen. In another group of 34 healthy subjects who were screened, 4 individuals showed positive reaction for tubercular antibody and 2 cases for antigen. This study shows that antigen detection assay using affinity purified anti Mtb EST antigen antibody is superior with overall specificity of 91% as compared to antibody detection assay with 75% specificity in bone & joint tuberculosis.  相似文献   

18.
BackgroundCellulose as a potential feed resource hinders its utilization because of its complex structure, and cellulase is the key to its biological effective utilization. Animal endogenous probiotics are more susceptible to colonization in the intestinal tract, and their digestive enzymes are more conducive to the digestion and absorption of feed in young animals. Min pigs are potential sources of cellulase probiotics because of the high proportion of dietary fiber in their feed. In this study, the cellulolytic bacteria in the feces of Min pigs were isolated and screened. The characteristics of enzymes and cellulase production were studied, which provided a theoretical basis for the rational utilization of cellulase and high-fiber food in animal production.ResultsIn our study, 10 strains of cellulase producing strains were isolated from Min pig manure, among which the M2 strain had the best enzyme producing ability and was identified as Bacillus velezensis. The optimum production conditions of cellulase from strain M2 were: 2% inoculum, the temperature of 35°C, the pH of 5.0, and the liquid loading volume of 50 mL. The optimum temperature, pH and time for the reaction of cellulase produced by strain M2 were 55°C, 4.5 and 5 min, respectively.ConclusionsMin pigs can be used as a source of cellulase producing strains. The M2 strain isolated from feces was identified as Bacillus velezensis. The cellulase from M2 strain had a good activity and the potential to be used as feed additive for piglets.How to cite: Li F, Xie Y, Gao X, et al. Screening of cellulose degradation bacteria from Min Pigs and optimization of its cellulase production. Electron J Biotechnol 2020;48. https://doi.org/10.1016/j.ejbt.2020.09.001  相似文献   

19.
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20.
Reactive oxygen species (ROS) are cytotoxic at higher concentration resulting in cell death, mutations, chromosomal aberrations or carcinogenesis. In this study DNA was modified by singlet oxygen and superoxide anion radicals generated by illumination of riboflavin under 365 nm UV-light. The modified DNA induced high titre antibodies in experimental animals. In enzyme immunoassay, serum antibodies from cancer patients (n = 34) showed a higher recognition of the modified DNA, as compared to the native form. This was further confirmed by the gel-shift assay. Immune IgG were used as a probe to detect oxidative lesions in the DNA of cancer patients. DNA isolated from lymphocytes of cancer patients proved to be an appreciable inhibitor of the experimentally induced antibodies against the ROS-DNA. This indicates the presence of oxidative lesions in the DNA obtained from cancer patients. The results show that ROS induced oxidative damage to DNA in cancer patients generate neo-epitopes that are alien for the immune system, resulting in autoantibody formation.  相似文献   

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