Mycobacterium Tuberculosis In Cerebrospinal Fluid Research Articles

Diagnostic performance of real time PCR for the detection of Mycobacterium tuberculosis in cerebrospinal fluid samples.

We aimed this study to standardize real time - polymerase chain reaction (RT-PCR) for the detection of Mycobacterium tuberculosis (Mtb) in cerebrospinal fluid (CSF) samples and compare its diagnostic performance with GeneXpert (Xpert), Mycobacteria Growth Indicator Tube (MGIT) and Multiplex PCR (MPCR) for tuberculous meningitis (TBM). A total of 217 CSF samples were obtained from patients with suspected TBM during the study period between January 2019 and December 2021. The optimal cycle threshold (CT) of RT-PCR was determined by comparing different gene targets of Mtb (IS6110, 16SrRNA, HSP65 and Ag85B). Sensitivity, specificity, positive predictive value (PPV) and negative predictive value (NPV) was determined for RT-PCR, Xpert, MGIT960 and MPCR. Diagnostic accuracy of these assays was compared by using clinical diagnosis as reference standard. IS6110RT-PCR was found to be highly sensitive as compared to other gene targets. Sensitivities of IS6110RT-PCR, MPCR, Xpert and MGIT against a reference standard of definite, probable and possible TBM were 36.7%, 21.1%, 16.7% and 6.7%, respectively; specificities were 97.6%, 100%, 100% and 100%, respectively. Xpert, RT-PCR, MPCR and MGIT960 detected 6.91% (n = 15), 5.99% (n = 13), 5.99% (n = 13) and 2.76% (n = 6) of definite TBM, respectively. RT-PCR detected 6.45% (n = 14) and 2.76% (n = 6) of possible TBM and probable TBM, respectively and MPCR detected 1.38% (n = 3) of possible and probable TBM each. IS6110RT-PCR is highly sensitive for primary screening of suspected TB cases, which may help clinicians to start appropriate patient's treatment with clinical suspicion of TBM.

Microbiological diagnosis and mortality of tuberculosis meningitis: Systematic review and meta-analysis.

Tuberculosis (TB) which is caused by Mycobacterium tuberculosis poses a significant public health global treat. Tuberculosis meningitis (TBM) accounts for approximately 1% of all active TB cases. The diagnosis of Tuberculosis meningitis is notably difficult due to its rapid onset, nonspecific symptoms, and the difficulty of detecting Mycobacterium tuberculosis in cerebrospinal fluid (CSF). In 2019, 78,200 adults died of TB meningitis. This study aimed to assess the microbiological diagnosis TB meningitis using CSF and estimated the risk of death from TBM. Relevant electronic databases and gray literature sources were searched for studies that reported presumed TBM patients. The quality of included studies was assessed using the Joanna Briggs Institute Critical Appraisal tools designed for prevalence studies. Data were summarized using Microsoft excel ver 16. The proportion of culture confirmed TBM, prevalence of drug resistance and risk of death were calculated using the random-effect model. Stata version 16.0 was used perform the statistical analysis. Moreover, subgroup analysis was conducted. After systematic searching and quality assessment, 31 studies were included in the final analysis. Ninety percent of the included studies were retrospective studies in design. The overall pooled estimates of CSF culture positive TBM was 29.72% (95% CI; 21.42-38.02). The pooled prevalence of MDR-TB among culture positive TBM cases was 5.19% (95% CI; 3.12-7.25). While, the proportion of INH mono-resistance was 9.37% (95% CI; 7.03-11.71). The pooled estimate of case fatality rate among confirmed TBM cases was 20.42% (95%CI; 14.81-26.03). Based on sub group analysis, the pooled case fatality rate among HIV positive and HIV negative TBM individuals was 53.39% (95%CI; 40.55-66.24) and 21.65% (95%CI;4.27-39.03) respectively. Definite diagnosis of TBM still remains global treat. Microbiological confirmation of TBM is not always achievable. Early microbiological confirmation of TBM has great importance to reduce mortality. There was high rate of MDR-TB among confirmed TBM patients. All TB meningitis isolates should be cultured and drug susceptibility tested using standard techniques.

Evaluation of the molecular bacterial load assay for detecting viable Mycobacterium tuberculosis in cerebrospinal fluid before and during tuberculous meningitis treatment

New tools to monitor treatment response and predict outcome from tuberculous meningitis (TBM) are urgently required. We retrospectively evaluated the 16S rRNA-based molecular bacterial load assay (MBLA) to quantify viable Mycobacterium tuberculosis in serial cerebrospinal fluid (CSF) from adults with TBM. 187 CSF samples were collected before and during the first two months of treatment from 99 adults TBM, comprising 56 definite, 43 probable or possible TBM, and 18 non-TBM and preserved at −80°C prior to MBLA. We compared MBLA against MGIT culture, GeneXpert MTB/RIF (Xpert) and Ziehl-Neelsen (ZN) smear. Before treatment, MBLA was positive in 34/99 (34.3%), significantly lower than MGIT 47/99 (47.5%), Xpert 51/99 (51.5%) and ZN smear 55/99 (55.5%). After one month of treatment, MBLA and MGIT were positive in 3/38 (7.9%) and 4/38 (10.5%), respectively, whereas Xpert and ZN smear remained positive in 19/38 (50.0%) and 18/38 (47.4%). In summary, MBLA was less likely to detect CSF bacteria before the start of treatment compared with MGIT culture, Xpert and ZN smear. MBLA and MGIT positivity fell during treatment because of detecting only viable bacteria, whereas Xpert and ZN smear remained positive for longer because of detecting both live and dead bacteria. Sample storage and processing may have reduced MBLA-detectable viable bacteria; and sampling earlier in treatment may yield more useful results. Prospective studies with CSF sampling after 1–2 weeks are warranted.

Xpert MTB/RIF Ultra for the Diagnosis of Tuberculous Meningitis: A Small Step Forward.

The delayed diagnosis of tuberculous meningitis (TBM) leads to poor outcomes, yet the current diagnostic methods for identifying Mycobacterium tuberculosis in cerebrospinal fluid (CSF) are inadequate. The first comparative study of the new GeneXpert MTB/RIF Ultra (Xpert Ultra) for TBM diagnosis suggested increased sensitivity of Xpert Ultra. Two subsequent studies have shown Xpert Ultra has improved sensitivity, but has insufficient negative predictive value to exclude TBM. Collecting and processing large volumes of CSF for mycobacterial testing are important for optimal diagnostic test performance. But clinical, radiological, and laboratory parameters remain essential for TBM diagnosis and empiric therapy is often needed. We therefore caution against the use of Xpert Ultra as a single diagnostic test for TBM; it cannot be used to “rule out” TBM.

Evaluation of multiplex PCR using MPB64 and IS6110 primers for rapid diagnosis of tuberculous meningitis

Tuberculous meningitis (TBM) is one of those most serious manifestations of extra-pulmonary tuberculosis and prompt diagnosis and treatment is required for better clinical outcome. It is difficult to diagnose due to lack of rapid, sensitive, and specific tests. Newer methods, which are easy and reliable, are required to diagnose TBM at an early stage. Thus our aim was to evaluate the Multiplex polymerase chain reaction (PCR) technique, using primers directed against the insertion sequence IS6110 and MPB64 gene for the detection of Mycobacterium tuberculosis in Cerebrospinal fluid (CSF), for rapid diagnosis of TBM patients. 102 CSF samples were analyzed from patients suspected with TBM along with a control group of 10 patients having other neurological disorders. CSF sediments were analyzed individually for M.tuberculosis DNA by Multiplex PCR using two set of primers targeting insertion sequence IS6110 and gene MBp64, which is very specific for MTBC. Out of 37 patients diagnosed with TBM clinically, MPB64 PCR was positive in 22, IS6110 PCR was positive in 28, both PCR using Multiplex were positive in 34 and Microscopy was positive in one. Thus Sensitivity of MPB64 PCR, IS6110 PCR, Multiplex PCR and Microscopy were found to be 62.3%, 75.4%, 91.8% and 2.7% respectively. In non TBM group PCR was negative in all cases hence, the specificity was 100%. Multiplex PCR system using primers targeting IS6110 and MPB64, for the detection of M.tuberculosis DNA in CSF samples, has high sensitivity than any one of them alone, and could be used for the early detection of TBM in CSF samples.

GeneXpert for Diagnosis of Tubercular Meningitis.

Diagnosis of tubercular meningitis (TBM) is difficult in children. The GeneXpert MTB/RIF assay has been recommended by WHO in 2013 to be used in children and in extra pulmonary clinical specimens. The present study was designed to assess the diagnostic utility of GeneXpert in detecting Mycobacterium tuberculosis in cerebrospinal fluid (CSF) in TBM cases and to compare the results with liquid culture BACTEC 460. Thirty four subjects <15y were diagnosed as TBM based on clinical, CSF and imaging details. Sensitivity of GeneXpert in CSF was 38.24% as compared to Bactec culture which was only 14.71% (p 0.0279). To conclude, GeneXpert MTB/RIF test is able to rapidly confirm diagnosis of TBM with higher sensitivity as compared to conventional methods and liquid culture.

Have we made any headway in the diagnosis of neurotuberculosis? Place of real time PCR or real time PCR

The diagnosis of Central Nervous System tuberculosis (CNS TB) had been always an enigma as yield of Mycobacterium tuberculosis in cerebrospinal fluid (CSF) is very low and diagnosis mainly rested on combination of clinical, CSF analysis and radiological findings. We attempted to find whether 16s rRNA primer based real time Polymerase chain reaction (PCR) aids in diagnosis of tubercular meningitis (TBM) and tuberculoma. Literature is flooded with newer techniques for the diagnosis of neuro-tuberculosis by real time PCR. But no clear cut guidelines are available about their role. 40 cases of tubercular meningitis, diagnosed on basis of Modified Ahuja Criteria and 40 controls were included in this prospective study. All patients were evaluated clinically and investigated in detail including cerebrospinal fluid analysis, Acid Fast Bacilli staining, culture for TB by Bac T Alert 3-D system,16s rRNA real time PCR assay, chest X ray, computed tomography scans of head, contact survey using chest X-ray, Mantoux test and hemogram. Comparative analysis of real time PCR were done in control group vs. cases of TBM diagnosed on the basis of Modified Ahuja criteria. Results: CSF for tuberculosis by real time PCR was positive in 26/40 cases (65%) of TBM, as against 2/40 controls. Keeping the CSF culture as the gold standard its sensitivity is 62.50 (95% confidence intervals- 54.7, 85.2), specificity is 77.40 (95% confidence intervals- 54.7, 85.2), positive predictive value is 46.15 and negative predictive value is 88.89. Modified Ahuja criteria can serve as useful tool in diagnosis. CSF analysis by real time PCR has shown better sensitivity and quicker results than culture. However, it cannot be recommended alone as it was negative in about a third of cases. But its significance is underlined in early diagnosis and as an adjunct to culture.

Psychosis Secondary to Tuberculous Meningitis: A Case Report

Tuberculous meningitis (TB meningitis) is a subacute meningitis known for its various form of initial manifestations, which often make early diagnosis difficult. Psychosis is a rare manifestation of this disease. We reported a case of 19-year-old woman who presented with worsening psychotic disorder of one year duration. She presented initially with social isolation with subsequent mutism and stupor. Initial brain imaging & Electroencephalography (EEG) was unremarkable. Cerebrospinal fluid (CSF) investigations revealed positive cerebrospinal fluid Mycobacterium tuberculosis polymerase chain reaction (MTB PCR). She was treated with empirical antituberculosis drugs and steroids. Subsequently her psychotic symptoms resolved. As a conclusions, the psychotic disorder was most likely caused by TB meningitis. TB meningitis should be considered in patients with no background history of psychiatric illness presenting with psychotic disorder especially in countries with high TB burden.

Bilateral brachial neuritis secondary to varicella reactivation in an HIV-positive man

We present the case of a 48-year-old HIV-positive man, who developed acute onset of pain in both upper limbs associated with proximal weakness and distal paraesthesia. Eight weeks prior to this presentation he had had varicella zoster affecting his right S1 dermatome. CD4 count was 355 cells/mm(3) and he was antiretroviral therapy (ART) naive. Power was 0/5 proximally and 4/5 distally in the upper limbs. Reflexes were absent and there was sensory loss in the C5, C6 and T1 dermatomes. Cerebrospinal fluid (CSF) examination showed a lymphocytosis with low glucose; however, CSF Mycobacterium tuberculosis (TB), and herpes simplex virus polymerase chain reaction (HSV PCR) were negative as was syphilis serology. Electromyography showed marked motor axonal loss. Magnetic resonance imaging (MRI) did not show any cervical spinal lesion. Varicella zoster virus (VZV) PCR was positive in the CSF. He was treated with high-dose intravenous aciclovir with good resolution of his syndrome over time and was commenced on ART. We believe this to be the first case report of varicella reactivation causing bilateral neuralgic amyotrophy in an HIV-positive patient.

Tuberculous Meningoencephalitis in a Patient with Hairy Cell Leukemia in Complete Remission

Tuberculous meningoencephalitis is a rare disease associated with high morbidity and mortality. We report a patient with hairy cell leukemia in complete remission who, after a single cycle of chemotherapy with cladribine, presented fever and neurological deficits. Laboratory diagnosis of tuberculous meningoencephalitis was made by polymerase chain reaction testing for Mycobacterium tuberculosis in cerebrospinal fluid. Despite the prompt institution of antitubercular-therapy, patient's general condition did not improve and he died. Mycobacterial infection should be considered in patients with intra-cranial lesions, affected by hematological malignancies and persistent immunosuppression.

Improved diagnosis of central nervous system tuberculosis by MPB64-Target PCR.

Central nervous system (CNS) tuberculosis is a serious clinical problem, the treatment of which is sometimes hampered by delayed diagnosis. Clearly, prompt laboratory diagnosis is of vital importance as the spectrum of disease is wide and abnormalities of the cerebrospinal fluid (CSF) are incredibly variable. Since delayed hypersensitivity is the underlying immune response, bacterial load is very low. The conventional bacteriological methods rarely detect Mycobacterium tuberculosis in CSF and are of limited use in diagnosis of tuberculous meningitis (TBM). This double blind study was, therefore, directed to the molecular analysis of CNS tuberculosis by an in-house-developed PCR targeted for amplification of a 240bp nucleotide sequence coding for MPB64 protein specific for Mycobacterium tuberculosis. Based on the clinical criteria, 47 patients with CNS tuberculosis and a control group of 10 patients having non-tubercular lesions of the CNS were included in the study. Analyses were done in three groups; one group consisting of 27 patients of TBM, a second group of 20 patients with intracranial tuberculomas and a third group of 10 patients having nontubercular lesions of the CNS acted as control. There were no false positive results by PCR and the specificity worked out to be 100%. In the three study groups, routine CSF analysis (cells and chemistry), CSF for AFB smear and culture were negative in all cases. PCR was positive for 21/27 patients (77.7% sensitivity) of the first group of TBM patients, 6/20 patients (30% sensitivity) of the second group with intracranial tuberculomas were positive by PCR and none was PCR-positive (100% specificity) in the third group. Thus, PCR was found to be more sensitive than any other conventional method in the diagnosis of clinically suspected tubercular meningitis.

Meningitis tuberculosa: revisión de 27 años

To analyze the features and outcome of children with tuberculous meningitis (TM) in a tertiary hospital in Madrid, with special emphasis on the possible influence of HIV infection, immigration and withdrawal of the Bacille-Calmette-Guérin (BCG) vaccine in Madrid in 1987. We reviewed the medical records of patients with TM diagnosed over a 27-year period (1977-2003). TM was diagnosed on the basis of compatible cytochemical findings in cerebrospinal fluid (CSF), plus a) isolation of Mycobacterium tuberculosis in CSF or gastric fluid; b) positive Mantoux test, or c) previous contact with active tuberculosis. Twenty-eight cases of TM were diagnosed. There were 24 cases (85 %) in the first 14 years versus only four cases (15 %) in the last 13 years. None of the children was co-infected with HIV. The median age was 3 years and 4 months. Only two of these had received the BCG vaccine. The source of infection was identified in 64 %. At admission, 57 % had nuchal rigidity, 46 % had neurological focality, and 54 % had an abnormal chest roentgenogram. The Mantoux purified protein derivative (PPD) test was positive in 89 % at diagnosis. Zhiel-Nielsen smear in CSF was positive in 3.5 %, and a positive culture from CSF was obtained in 32 %. Computed tomography scan was performed in 22 children and showed hydrocephalus in 90 %. All patients were treated with four drugs, with no significant adverse events. Corticoids (89 %), ventricular-peritoneal shunt (43 %) and antiepileptic drugs (39 %) were widely used as complementary treatments. Mortality was 12 %. Half of the patients had sequels, which were mostly permanent. The most frequent and severe complication was neurodevelopmental delay (21 %). Severe sequels occurred in children with more advanced stages of illness. The frequency of TM has decreased in the last 15 years, despite the emergence of HIV and immigration from countries where tuberculosis is endemic. These data are in agreement with the general reduction of the incidence of TM in our environment. Withdrawal of the BCG vaccine in Madrid in 1987 did not increase cases of TM in our hospital; on the contrary, these have decreased. A considerable percentage of children with advanced stages of TM show severe sequels.

Genomic variation in the PLCD gene region of clinical strains of Mycobacterium Tuberculosis

Tuberculous meningitis (TBM) is one of those most serious manifestations of extra-pulmonary tuberculosis and prompt diagnosis and treatment is required for better clinical outcome. It is difficult to diagnose due to lack of rapid, sensitive, and specific tests. Newer methods, which are easy and reliable, are required to diagnose TBM at an early stage. Thus our aim was to evaluate the Multiplex polymerase chain reaction (PCR) technique, using primers directed against the insertion sequence IS6110 and MPB64 gene for the detection of Mycobacterium tuberculosis in Cerebrospinal fluid (CSF), for rapid diagnosis of TBM patients.102 CSF samples were analyzed from patients suspected with TBM along with a control group of 10 patients having other neurological disorders. CSF sediments were analyzed individually for M. tuberculosis DNA by Multiplex PCR using two set of primers targeting insertion sequence IS6110 and gene MBp64, which is very specific for MTBC.Out of 37 patients diagnosed with TBM clinically, MPB64 PCR was positive in 22, IS6110 PCR was positive in 28, both PCR using Multiplex were positive in 34 and Microscopy was positive in one. Thus Sensitivity of MPB64 PCR, IS6110 PCR, Multiplex PCR and Microscopy were found to be 62.3%, 75.4%, 91.8% and 2.7% respectively. In non TBM group PCR was negative in all cases hence, the specificity was 100%.Multiplex PCR system using primers targeting IS6110 and MPB64, for the detection of M. tuberculosis DNA in CSF samples, has high sensitivity than any one of them alone, and could be used for the early detection of TBM in CSF samples.

Evaluation of polymerase chain reaction for rapid diagnosis of clinically suspected tuberculous meningitis

Setting: Since conventional bacteriological methods rarely detect Mycobacterium tuberculosis in cerebrospinal fluid (CSF) and are of limited use in the diagnosis of tuberculous meningitis (TBM), clinical features suggestive of TBM supported by indirect evidence such as CSF examination and computerized tomography (CT) of the head have been used for the early diagnosis of TBM. Objective: We evaluated the efficacy of polymerase chain reaction (PCR) in the diagnosis of TBM. Methods: Coded CSF samples from 40 patients with TBM and from 49 patients with other neurological disorders were processed. In the absence of a reliable sensitive and specific test for M. tuberculosis in CSF, we used a set of established clinical criteria as the gold standard. Accordingly, the patients were divided into definite, highly probable, probable and possible TBM. The samples were decoded only after completion of the laboratory tests. Results: PCR was positive in 2 4 , 19 20 , 13 16 patients with highly probable, probable and possible TBM respectively. None of the samples were positive by conventional bacteriological methods. However, 3 49 CSF samples from non-TBM patients were also found positive by PCR. PCR detected M. tuberculosis genomic DNA in the CSF of 85% of clinically suspected TBM cases and 6.1% of non-tuberculous controls. Conclusion: PCR, along with the suggested clinical criteria, offers a rapid and fairly accurate diagnosis of TBM.

DETECTION OF MYCOBACTERIAL ANTIGENS IN CEREBROSPINAL FLUID OF PATIENTS WITH TUBERCULOUS MENINGITIS BY ENZYME-LINKED IMMUNOSORBENT ASSAY

Culture and identification of Mycobacterium tuberculosis in cerebrospinal fluid (CSF) of patients with tuberculous meningitis is slow, and prognosis depends on early diagnosis and tratament. An enzyme-linked immunosorbent assay with commercial BCG and anti-BCG was designed to detect mycobacterial antigens in human body fluids. Absorbance at 405 nm was measured and readings ≽0·20 were considered positive. The readings were above 0·20 for four patients with confirmed tuberculous meningitis and nine of twelve patients with probable tuberculous meningitis. In all but one of ten patients with meningitis of other causes and in eleven control subjects the readings were below 0·20. The specificity of the test was 95% and its sensitivity 81·25%