In-vitro anti-Mycobacterium tuberculosis effect of essential oil of Ocimum sanctum L. (Tulsi/Basil) leaves

Background/ObjectivesMycobacterium tuberculosis, the causative agent of tuberculosis has developed resistance to most of the available antimicrobials. Consequently, it is difficult to cure all the patients with tuberculosis and in future, the incidence of tuberculosis by drug resistant M. tuberculosis is likely to increase, worldwide. Therefore detection and development of new antimicrobials against M. tuberculosis is needed urgently.MethodsEssential oil from the leaves of Ocimum sanctum L (Tulsi/Basil) was obtained by hydro distillation. The anti-mycobacterial effect of essential oil was evaluated against H37Rv and nine clinical isolates of M. tuberculosis in the BD BACTEC MGIT instrument using different volumes of essential oil.ResultsThe essential oil inhibited the growth of H37Rv and all the nine clinical isolates of M. tuberculosis. The minimal inhibitory concentration of H37Rv was 3 μl (2.931 μg) and those of the clinical isolates of M. tuberculosis ranged from 1.5 μl (1.4655 μg) to 6 μl (5.862 μg).ConclusionThe Essential oil from the leaves of O. sanctum L.(Tulsi/Basil) has anti-M. tuberculosis effect in the in-vitro BD BACTEC MGIT method.     

A case of brucellar septic arthritis of the knee with a prolonged clinical course

Brucellosis is a systemic infectious disease with a broad spectrum of clinical manifestations. Arthritis is frequently observed in its course and may be one of the main presenting clinical features of the disease. We report a case of brucellar monoarthritis of the knee with a prolonged clinical course despite efficient antibiotic treatment.     

Rapid recovery of Mycobacterium tuberculosis complex from clinical specimens using the BACTEC 9000 MB system, a new automated fluorimetric technique

Objective: To evaluate the new non-radioactive automated method BACTEC 9000 MB system for the rapid detection of mycobacteria in clinical specimens.
Method: Ninety clinical specimens from 90 patients with a clinical diagnosis of tuberculosis were tested by both BACTEC 9000 and standard microbiological methods, and the results compared.
Results: The BACTEC 9000, in comparison with the standard method, showed significantly higher detection rates (45 of 90 positive versus 34), shorter time to culture positivity (mean time 18.8 versus 27.4 days) and lower contamination rate (2.2% versus 5.5%).
Conclusions: These results encourage the use of this new system and suggest its use in microbiological laboratories involved in mycobacteriology.     

Determination of the activity of standard anti-tuberculosis drugs against intramacrophage Mycobacterium tuberculosis,in vitro: MGIT 960 as a viable alternative for BACTEC 460

BACTEC 460 has now been phased out, so the search for an alternative is imperative. We have determined the activity of standard anti-tuberculosis drugs against intramacrophage Mycobacterium tuberculosis, in vitro, by using BACTEC 460 and MGIT 960 methods. The minimum inhibitory concentrations of isoniazid, rifampicin, ethambutol and streptomycin against intracellular M. tuberculosis H37Rv were found to be 0.2, 0.8, 8.0, and 5.0 μg/mL, respectively, by both methods. These results show a significant (p < 0.001) concordance between minimum inhibitory concentrations obtained by these two different methods. MGIT 960 system uses a robust florescence quenching-based oxygen sensor, requires no radioisotope, is safe, and relatively easy to operate. Apparently, this is the first report wherein MGIT 960 has been validated for anti-tubercular susceptibility testing against intracellular M. tuberculosis H37Rv. Our preliminary data thus clearly demonstrate that the MGIT 960 method can be considered as a promising alternative to BACTEC 460 method.     

实时荧光PCR与BACTEC MGIT 960快速培养在初治痰涂片阴性未受抗结核治疗的肺结核诊断中的应用

目的:探讨实时荧光PCR与BACTEC MGIT 960分枝杆菌快速培养(快速培养法)准确诊断初治痰涂片阴性(涂阴)且未经受抗结核治疗的肺结核可能性。方法:实验组:368份临床诊断活动性肺结核患者的初治涂片阴性痰标本;对照组:55份非结核性肺部疾病患者的痰液标本。用FQ-PCR技术、快速培养法及改良罗氏培养参考法对两组标本进行分析,观察指标为TB-DNA和结核菌阳性率。结果:实验组和对照标本中FQPCR技术、快速培养法和改良罗氏培养参考法检测的TB-DNA或结核菌阳性率分别为:16.85%、23.37%、22.01%和1%、0%、0%。三种方法分别平均耗时3h、9.6d和28d。以改良罗氏培养参考法为参照,FQ-PCR法和快速培养法的敏感性分别为69.1%和100%;特异性分别为91.2%和98.6%。结论:FQ-PCR和快速培养法均能提高初治涂阴肺结核患者的早期确诊率,但FQ-PCR的敏感性尚需进一步提高。     

BACTEC MGIT 960用于结核分枝杆菌快速培养的初步评价

目的评价BACTEC MGIT 960系统快速分离结核分枝杆菌及药敏试验的效果。方法对临床拟诊结核病患者痰样本,采用直接涂片萋-尼法抗酸染色显微镜检、BACTEC MGIT 960和改良罗氏(L-J)培养法进行检测,并对分离到的结核分枝杆菌进行药敏试验。比较其分离的阳性率、培养及药敏所需的时间。结果共对634例拟诊结核病患者痰样本进行了检测,MGIT960培养的阳性率(51.7%)明显高于L-J法(37.7%),直接涂片法阳性率最低(14.7%)。MGIT960和L-J法的污染率分别为4.9%和4.4%。MGIT960和传统的L-J法的检出时间分别为12.0 d和32.0 d。MGIT960和L-J法对异烟肼、利福平、链霉素和乙胺丁醇的药物敏感性检出时间分别为9.0 d和36.5 d。结论 BACTEC MGIT 960可以明显提高分离培养阳性率、缩短培养和药敏试验的时间。     

实时荧光聚合酶链反应和结核培养仪法对非痰液标本结核杆菌检测的比较

目的评估比较实时荧光聚合酶链反应(FQ-PCR)和结核培养仪(BACTEC960)法在检测非痰液标本中结核分支杆菌的特异性和灵敏度。方法对34例临床可疑患者用BACTEC960法和FQ-PCR技术2种方法作阳性检出率比较。结果FQ-PCR诊断肺外结核病的特异度为95%,灵敏度为14·3%。BACTEC960法诊断肺外结核病的特异度为100%,灵敏度为21·4%。结论BACTEC960法诊断结核病的特异性和灵敏度与国内外文献报道较一致,而实时荧光PCR技术诊断结核病的特异性和灵敏度与国内外文献报告有较大差异,明显低于平均阳性检出率。     

Diagnostic appraisal of simultaneous application of two nested PCRs targeting MPB64 gene and IS6110 region for rapid detection of M. tuberculosis genome in culture proven clinical specimens

Background: Early diagnosis of tuberculosis is critical for its effective management and prevention. Several gene amplification methods are used in the detection of tubercle bacilli from clinical specimens. MPB64 gene and IS6110 region have been identified as potential gene targets for the specific detection of Mycobacterium tuberculosis from direct clinical specimens. Objective: The present study was conducted to evaluate the diagnostic utility of simultaneous application of two nested polymerase chain reaction (nPCRs) targeting MPB64 and IS6110 region for the detection of M. tuberculosis genome. Materials and Methods: A total of 100 and 354 clinical specimens from the control group and clinically suspected tuberculosis patients, respectively, were included in the study. nPCRs targeting MPB64 and IS6110 region were performed. Results and Conclusion: All of the 100 clinical specimens from the control group were negative for both nPCRs. Out of the 354 clinical specimens, 339 were positive for both culture and nPCRs, 10 and 5 were positive for culture, and nPCR targeting IS6110 and MPB64 regions, respectively. To conclude, nPCRs targeting MPB64 and IS6110 region are reliable and specific targets when applied simultaneously on clinical specimens to attain 100% sensitivity for the detection of M. tuberculosis genome.     

Direct inoculation on Phoenix panels for identification and antimicrobial susceptibility from positive BACTEC cultures: first study from India

Purpose: This was a prospective study planned in a super-specialty hospital in Delhi to reduce turnaround times of identification–susceptibility results of positive blood cultures. Materials and Methods: One hundred consecutive single morphology non-duplicate cultures were inoculated on Becton Dickinson Phoenix™ panels by growth recovered directly from liquid BACTEC™ media and after pure growth on solid media. Results: Complete concordance was observed in 72.4% of gram-negative and 45.8% of gram-positive isolates. For gram-negative isolates, categorical agreement (CA) was >83% and essential agreement (EA) was >96% among all antibiotics tested, very major errors (VME) were 0.13%, major errors (ME) 0.54%, and minor errors (MiE) were 3.01%. For gram-positive isolates, VME was 0.73%, 1.10% MiE and no ME. It was observed that average time from receipt of specimen to release of reports was 30:34 h and 32 h for gram-negative and gram-positive isolates if reports of “Direct” panels were to be released. Conclusions: By direct panel inoculation, a decrease of at least 18–20 h in turnaround time was observed compared with the standard method. This helps early change to effective antibiotic therapy and also reduces the expenditure incurred for a patient’s hospital stay by average Rs 20,000 ($443) per day.     

血培养联合PCR对HIV感染者和AIDS患者分枝杆菌血症诊断的分析

目的评价应用BACTEC Myco/F Lytic（MFL）培养基血培养联合聚合酶链反应（PCR）对人类免疫缺陷病毒（HIV）感染者和获得性免疫缺陷综合征（AIDS）患者合并分枝杆菌（MB）感染诊断的效果。方法用MFL配合BACTEC 9120血培养仪对55例可疑合并MB感染的HIV/AIDS患者血标本进行培养,联合抗酸染色和PCR对阳性标本进行菌种鉴定。结果 55例患者血培养分析显示,MFL血培养的总报阳率为32.7%（18/55）,其中7例（12.7%）报阳标本经证实为MB生长,8例（14.5%）报阳标本中未检出任何微生物,暂定为假报警,3例（5.4%）检出污染菌。其中MB的平均培养周期为746.79 h,约31.1 d。7例MB培养阳性标本经PCR鉴定,5例（71.4%）为结核分枝杆菌（TB）,2例（28.6%）为鸟分枝杆菌复合体（MAC）。结论 MFL血培养能协助临床诊断MB血症,该法操作简单、安全性高,但培养周期较长,且存在假报警及某些需氧菌污染的情况,故宜联合其他检测手段对MB血症进行检测。