Macrolide and fluoroquinolone resistance associated mutations in Mycoplasma genitalium in men who have sex with men attending STI clinic: A pilot study from India.

Background Increasing rates of macrolide and fluroquinolone resistance in Mycoplasma genitalium (MG) are being reported worldwide with resultant treatment failure. Aims and objectives We aimed to determine the level of antibiotic resistance of MG in men who have sex with men (MSM) attending a sexually transmitted infections (STIs) clinic in New Delhi, India. Methods Real-time polymerase chain reaction (PCR) assays targeting MgPa and pdhD genes were performed to detect MG rectal, urogenital or oropharyngeal infections in 180 MSM between January 2022 and June 2023. Macrolide resistance-associated mutations (MRM) and quinolone resistance-associated mutations (QRM) were detected by specific amplification of domain V of 23SrRNA gene and appropriate regions of parC and gyrA genes respectively followed by sequencing. PCR-based screening for Chlamydia trachomatis (CT) infection was also performed. Results A total of 13 (7.2%) MSM were positive for MG infection. The most common site of infection was anorectum (8/13; 61.5%) followed by the urethra (5/13; 38.5%). None of the patients had infection at both the sites, and no oropharyngeal MG infection was detected. CT infection was detected in 37 (20.6%) MSM. Of the 13 MG-infected MSM, 6 (46.2%) were co-infected with CT. MRM and QRM were found in five (46.2%) and two (15.4%) strains, respectively. Both Quinolone resistance mutation (QRM)-harbouring strains also harboured MRM. All the five MG isolates carried the MRM A2071G. Both the QRM isolates co-harboured the parC and gyrA single-nucleotide polymorphisms. There was no correlation between the presence of antibiotic resistance and co-infection with CT (P = 0.52). Limitation Because all patients in the study were MSM, the high rate of resistance to macrolides and fluoroquinolones could not be extrapolated for non-MSM patients. Conclusion This is a report of an initial survey of antibiotic resistance to MG in a country where its diagnosis and treatment are not routinely available. We found a high prevalence of MG-carrying MRM, QRM and dual-class resistance in MSM in the absence of antibiotic exposure. This study mandates the need for both screening and detection of antimicrobial resistance against MG.

Performance of Cephid CT/NG Xpert rapid PCR test for detection of Chlamydia trachomatis and Neisseria gonorrhoeae at a tertiary care centre in North India.

INTRODUCTION: Chlamydia trachomatis (CT) and Neisseria gonorrhoeae (NG) are prevalent causes of sexually transmitted infections (STIs) globally, leading to substantial morbidity and transmission risks. METHODS: This study evaluates the diagnostic efficacy of Xpert CT/NG compared to conventional PCR and culture methods in 121 patients at a tertiary care centre in North India. RESULTS: Xpert CT/NG demonstrated high sensitivity (85.8%) and specificity (96.3%) outperforming conventional PCR. Xpert CT/NG's rapidity and accuracy underscore its utility in timely diagnosis and control of STIs. CONCLUSION: As sexually transmitted infections pose a serious health concern implementation of such rapid diagnostic methods/point of care testing methods are to be implemented for early diagnosis.

Brucella Seropositivity at a Tertiary Care Hospital in Chandigarh.

There are reports that the neglected zoonotic tropical disease brucellosis is reemerging today. Serological tests are being widely used in the diagnosis of brucellosis. In the present study, we performed a standard agglutination test (SAT) on 1348 suspected cases of brucellosis during the period of four years from April 2018 to March 2022. We noticed an increase in seropositivity from 2.6% in the year 2018-19 to 7.4% in the year 2021-22. We also noticed a spike in seropositivity in the years 2019-20 (12.5%). Our study shows the recent trend in seropositivity of the disease in Chandigarh and, hence, can be a meaningful addition to the existing serological diagnostic data related to brucellosis.

Xpert MTB/XDR assay: rapid TB drug resistance detection.

PURPOSE: To assess the Xpert MTB/XDR assay's efficiency in promptly detecting resistance to isoniazid, fluoroquinolones, ethionamide, and second-line injectable drugs among tuberculosis (TB) patients. METHODS: From August 2020 to July 2021, TB suspected patient samples were enrolled at a tertiary care center for our study. We conducted mycobacterial culture, phenotypic DST using proportion method in liquid culture at WHO-recommended concentrations, and the line probe assay (LPA). Simultaneously, the Index test, Xpert MTB/XDR, was performed following the manufacturer's instructions. RESULTS: Among 360 samples, 107 were excluded due to incomplete information. Resistance to isoniazid, levofloxacin and moxifloxacin was found in 45/251, 21/251 and 20/251 samples, respectively by phenotypic DST. The diagnostic accuracy of Index test, taking phenotypic DST as a reference standard, was 95.8%, 99.04%, and 99.05% for isoniazid, levofloxacin, and moxifloxacin, respectively. The Index test assay demonstrated a specificity of 99.1% for detecting SLID resistance, yielding a diagnostic accuracy of 99.2. Comparing the Index test with LPA revealed a significant enhancement in sensitivity for detecting isoniazid resistance (86.7% vs. 82.2%). CONCLUSIONS: The Index test exhibited promising outcomes in identifying resistance to isoniazid and fluoroquinolones, surpassing the performance of the LPA. This could be valuable for promptly initiating treatment in cases of drug-resistant tuberculosis.

Performance evaluation of loop-mediated isothermal amplification, polymerase chain reaction and real-time polymerase chain reaction methods to detect Neisseria gonorrhoeae among symptomatic patients from India.

Background: Neisseria gonorrhoeae is one of the most important causative organisms in causing sexually transmitted infections. The clinical presentation of gonorrhoea mimics the symptoms of other sexually transmitted infections, and a proper diagnosis of the same is therefore crucial in patient management. The current study intended to compare different in-house molecular methods: that is, conventional PCR, real-time PCR, and LAMP assay for detection of N. gonorrhoeae. Methods: A total of 163 samples were collected from 145 patients who presented with urethral and vaginal discharge. Collected samples were processed for culture on GC agar base, and three different molecular diagnostic tests (conventional PCR, real-time PCR, and LAMP assay) were performed simultaneously on all the samples. Results: Culture of N. gonorrhoeae was positive in 17 out of 21 (80.9%) swab samples. With culture as the gold standard method, conventional and real-time PCR had a sensitivity of 94.1%, whereas the sensitivity of the LAMP assay was found to be 88.2%. All three methods had a specificity of 100%. In addition to swab samples, evaluation of urine samples by different molecular methods yielded a good concordance with a kappa value of 0.85 by conventional PCR and real-time PCR showing a perfect level of agreement, while the LAMP assay was found to have a substantial level of agreement. Conclusion: LAMP assay had a comparable diagnostic accuracy to other molecular methods for the detection of N. gonorrhoeae and can be used as a point-of-care test in resource-limited settings.

Unmasking the Hidden Invader: A case report of chest wall tuberculosis presenting as an abscess.

Chest wall tuberculosis is a very rare clinical entity that accounts for 1-5% of musculoskeletal tuberculosis (TB). Here we present a case of chest wall TB. A 16-year-old boy presented with complaints of progressive increasing swelling over the right hemithorax following a history of trauma. Radiologically it was a localized liquid collection with no connection to the thoracic cavity. Pus sample was sent to the mycobacteriology laboratory. The Gene Xpert result came as positive, and sensitive to rifampicin while the acid-fast bacilli (AFB) smear test was negative. Diagnosing a case of chest wall TB is always challenging for clinicians.

Whole genome sequencing for the prediction of resistant tuberculosis strains from northern India.

PURPOSE: Tuberculosis is an important public health problem among infectious diseases. The problem becomes more concerning with the emergence of MDR-TB and pre-XDR-TB. Whole genome sequencing (WGS) detection of resistance has recently gained popularity as it has advantages over other commercial techniques. METHODS: We performed in-house WGS followed by detailed analysis by an in-house pipeline to identify the resistance markers. This was accompanied by Phenotypic DST, and Sanger sequencing on all the 12 XDR, 06 pre-XDR, and 06 susceptible M. tb isolates. These results were collated with online M. tb WGS pipelines (TB profiler, PhyResSE, Mykrobe predictor) for comparative analysis. RESULTS: Following our in-house analysis, we observed 64 non-synonymous SNPs, fifteen synonymous SNPs, and five INDELs in 25 drug resistance-associated genes/intergenic regions (IGRs) in M. tb isolates. Sensitivity for detecting XDR is 33%, 58%, 83%, and 83%, respectively, using Mykrobe predictor, PhyResSE, TB-profiler, and in-house pipeline for WGS analysis, respectively. TB-profiler detected a rare mutation H70R in the gyrA gene in one pre-XDR isolate. Lineage 2.2.1 East-Asian (Beijing sublineage type) predominated (60%) in WGS data analysis of the XDR isolates. CONCLUSIONS: Our findings suggest that in-house analysis of WGS data and TB-profiler sensitivity was better for the detection of second-line resistance as compared to other automated tested tools. Frequent upgradation of newer mutations associated with resistance needs to be updated, as it potentiates tailored treatment for patients.

In-vivo expressed mycobacterial transcripts as diagnostic targets for pulmonary tuberculosis.

The nucleic acid amplification tests (NAATs) such as Xpert MTB/RIF have transformed the TB diagnostic field by significantly increasing the case detection. However, newer improved diagnostic assays are still needed to meet the WHO targets to end TB. Present study is based on a novel approach of utilizing the in-vivo expressed specific mycobacterial transcriptomic biomarkers for the diagnosis of pulmonary tuberculosis (PTB). Total 61 subjects were recruited including smear positive (smear+; n = 15), smear negative (smear-; n = 30) PTB patients and disease controls (n = 16). Transcripts of three mycobacterial genes Rv0986, Rv0971c and Rv3121 were analyzed using real time PCR (qRT-PCR) in sputum samples. qRT-PCR with Rv0986, Rv0971c and Rv3121 identified smear + PTB patients with 100 %, 78.6 % and 86.7 % sensitivity respectively. In smear- PTB patients, both Rv0986 and Rv0971c based qRT-PCR resulted in 63 %, sensitivity whereas Rv3121 identified these patients with ∼40 % sensitivity only. The sensitivity of the assay for smear-patients increased to 85 % when combinatorial analysis of qRT-PCR data for all the three genes was used. Thus, in-vivo expressed mycobacterial transcripts have promising potential as biomarkers for PTB diagnosis.

Delta checks.

Delta check is an electronic error detection tool. It compares the difference in sequential results within a patient against a predefined limit, and when exceeded, the delta check rule is considered triggered. The patient results should be withheld for review and troubleshooting before releasing to the clinical team for patient management. Delta check was initially developed as a tool to detect wrong-blood-in-tube (sample misidentification) errors. It is now applied to detect errors more broadly within the total testing process. Recent advancements in the theoretical understanding of delta check has allowed for more precise application of this tool to achieve the desired clinical performance and operational set up. In this Chapter, we review the different pre-implementation considerations, the foundation concepts of delta check, the process of setting up key delta check parameters, performance verification and troubleshooting of a delta check flag.

Tuberculosis and Bacillus Calmette-Guérin Disease in Patients with Chronic Granulomatous Disease: an Experience from a Tertiary Care Center in North India.

Chronic granulomatous disease (CGD) is a phagocytic defect characterized by recurrent bacterial and fungal infections. We report clinical profile of patients with CGD and mycobacterial infections in a cohort from North India. A review of clinical and laboratory records was carried out for patients with CGD registered at our center between 1990 and 2021. Of the 99 patients with CGD, 22 had mycobacterial infections-Mycobacterium tuberculosis and M. bovis-BCG in 11 each. Among the children with M. bovis-BCG infection, 6 had localized and 5 had disseminated BCG disease. Median age at onset of symptoms and diagnosis of BCG disease was 5 months and 15 months, respectively. While disseminated forms of BCG were noted only in CYBB defect, none of the patients with NCF1 defect developed complications due to BCG vaccine. A recurring radiological feature was left axillary lymph node calcification, which was present in around 50% of CGD patients with BCG infections. Of 11 patients with tuberculosis, pulmonary, pleuro-pulmonary, abdominal, and disseminated forms were present in 6, 1, 2, and 2, respectively. Median age at onset of symptoms and diagnosis of tuberculosis was 129 months and 130 months, respectively. Molecular defects were identified in CYBB (5), NCF1 (4), and CYBA (1). Incidence of tuberculosis and BCG-related complications in patients with CGD is higher than the normal population. Screening for CGD is warranted in any patient with adverse reactions to BCG vaccination, calcification of left axillary lymph node, and persistent, recurrent or disseminated forms of tuberculosis.

Association of intermediate Nugent Score and bacterial vaginosis with sexually transmitted infections and vulvovaginal candidiasis.

Background Bacterial vaginosis is a common vaginal syndrome among females, which leads to significant morbidity and complications, if left untreated. The association of bacterial vaginosis with various sexually transmitted infections has been mentioned in previous literature. However, studies on the intermediate Nugent Score are lacking. This study was planned to examine the association of sexually transmitted infections with the intermediate Nugent Score. Materials and Methods The study included was conducted to include females presenting with vaginal discharge, burning micturition, itching, lower abdominal pain and infertility. The Nugent scoring was used to categorize patients into those having normal flora, intermediate or bacterial vaginosis. Conventional and molecular techniques targeting Trichomonas vaginalis, Chlamydia trachomatis, Ureaplasma urealyticum, Mycoplasma hominis, Syphilis, Neisseria gonorrhoeae and vulvovaginal candidiasis were performed. Results A total of 3,531 clinical samples were collected from females with a median age of 28.0 years. The number of patients with bacterial vaginosis and intermediate Nugent Score and positive cases were significantly higher in the 21-35 years age group (P < 0.0001). We observed that the likelihood of test results being positive for Trichomonas vaginalis was higher (P < 0.05), as the abnormality of the vaginal flora increased. Mycoplasma hominis was observed to be significantly higher in the intermediate Nugent Score group than the BV-positive patients (0.6 vs 0.2, P = 0.002). The number of vulvovaginal candidiasis cases in both the bacterial vaginosis-negative and bacterial vaginosis-positive groups were nearly the same (9.3 vs 9.8%). Limitation Individual follow-up couldn't be performed on the patients. Conclusion We observed that the dysbiosis in vaginal microbiota, with an increase in Nugent scoring, was significantly associated with an increased risk for the acquisition of sexually transmitted infections and vulvovaginal candidiasis.

Evaluation of Mycobacterium tuberculosis derived cell-free DNA using pleural fluid and paired plasma samples for the diagnosis of pleural tuberculosis.

Pleural tuberculosis (pTB) is a grave clinical challenge. A novel cell-free M. tuberculosis DNA (cfM.tb-DNA) probe-based-qPCR assay was developed for the diagnosis of pTB. Total cell-free DNA was extracted from pleural fluid (PF) and paired plasma samples and cfM.tb-DNA was quantified by probe-based qPCR targeting devR (109-bp) gene of M. tuberculosis in patients with pleural effusion. Patient categorization was done using 'Composite-Reference-Standard' formulated for the study. Assay cut-offs were determined from samples in the 'Development set' (n = 17; 'Definite & Probable' pTB; n = 9 and 'Non-TB'; n = 8) by ROC-curve analysis and applied to 'Validation set' (n = 112; 'Definite' pTB; n = 8, 'Probable' pTB; n = 34, 'Possible' pTB; n = 28 and 'Non-TB'; n = 42). cfM.tb-DNA qPCR had a sensitivity of 62.5% (95%CI; 24.4,91.4) in 'Definite' pTB category and 59.5% (95%CI; 43.2,74.3) in 'Definite & Probable' pTB category with 95.2% (95%CI; 83.8,99.4) specificity using PF. In plasma (n = 85), the assay had a sub-optimal sensitivity of 7.6% (95%CI; 0.95,25.1) with 88.2% (95%CI; 72.5,96.7) specificity in 'Definite & Probable' pTB group. Xpert MTB/RIF assay detected only six-samples in the 'Validation set'. Logistic regression analysis indicated that PF-cfM.tb-DNA qPCR provided incremental advantage over existing pTB diagnostic algorithms. To the best of our knowledge, this is the first report describing the utility of cfM.tb-DNA for pTB diagnosis in India.

Prevalence of Trichomonas vaginalis by polymerase chain reaction-based molecular method among symptomatic women from Northern India.

Introduction: Trichomoniasis remains one of the most common sexually transmitted infections, which is curable. To prevent complications and transmission, prompt and correct diagnosis is essential to treat Trichomonas vaginalis. The present study was done to evaluate polymerase chain reaction (PCR) with other conventional techniques for the diagnosis of T. vaginalis infection and determine the prevalence of T. vaginalis in women with vaginal discharge based on PCR assay. Methods: Vaginal swabs were collected by the trained health-care professional using FLOQSwabs™ (Copan, Italy) during routine pelvic examinations among 1974 symptomatic females. The wet microscopy, culture, and PCR were performed. Results: The sensitivity of wet mount and culture in comparison to PCR was 60.87% and 56.52%, respectively. The kappa inter-rater agreement of T. vaginalis PCR showed substantial agreement with wet mount microscopy (κ = 0.742) and culture (κ = 0.707). The PCR detected an additional 17 cases that were missed by conventional techniques. Discussion: The study highlights the importance of PCR for T. vaginalis screening among symptomatic females.

Stable isotope labeling as a promising tool for rapid drug susceptibility testing in Neisseria gonorrhoeae.

The world is heading towards an era of intractable and impending untreatable N. gonorrhoeae, thereby underlining the significance of rapid and accurate prediction of drug resistance as an indispensable need of the hour. In the present study, we optimized and evaluated a stable isotope labeling-based approach using the MALDI-TOF MS (Matrix-Assisted Laser Desorption/Ionization-Time of Flight Mass Spectrometry) for rapid and reliable detection of ciprofloxacin and azithromycin resistance in N. gonorrhoeae. All the isolates were cultured under three varied condition setups viz. medium supplemented with normal lysine, heavy lysine (isotope), and heavy lysine along with the antibiotics (ciprofloxacin/azithromycin), respectively. After incubation, spectra were acquired using the MALDI-TOF MS which were further screened for unique patterns (media-specific spectra) to differentiate drug-susceptible and resistant isolates. The results of the stable isotope labeling assay were comparable to the results of phenotypic methods used for susceptibility testing.

Advances in internal quality control.

Quality control practices in the modern laboratory are the result of significant advances over the many years of the profession. Major advance in conventional internal quality control has undergone a philosophical shift from a focus solely on the statistical assessment of the probability of error identification to more recent thinking on the capability of the measurement procedure (e.g. sigma metrics), and most recently, the risk of harm to the patient (the probability of patient results being affected by an error or the number of patient results with unacceptable analytical quality). Nonetheless, conventional internal quality control strategies still face significant limitations, such as the lack of (proven) commutability of the material with patient samples, the frequency of episodic testing, and the impact of operational and financial costs, that cannot be overcome by statistical advances. In contrast, patient-based quality control has seen significant developments including algorithms that improve the detection of specific errors, parameter optimization approaches, systematic validation protocols, and advanced algorithms that require very low numbers of patient results while retaining sensitive error detection. Patient-based quality control will continue to improve with the development of new algorithms that reduce biological noise and improve analytical error detection. Patient-based quality control provides continuous and commutable information about the measurement procedure that cannot be easily replicated by conventional internal quality control. Most importantly, the use of patient-based quality control helps laboratories to improve their appreciation of the clinical impact of the laboratory results produced, bringing them closer to the patients.Laboratories are encouraged to implement patient-based quality control processes to overcome the limitations of conventional internal quality control practices. Regulatory changes to recognize the capability of patient-based quality approaches, as well as laboratory informatics advances, are required for this tool to be adopted more widely.

Evaluation of immunodominant peptides of in vivo expressed mycobacterial antigens in an ELISA-based diagnostic assay for pulmonary tuberculosis.

Non-sputum-based biomarker assay is urgently required as per WHO's target product pipeline for diagnosis of tuberculosis. Therefore, the current study was designed to evaluate the utility of previously identified proteins, encoded by in vivo expressed mycobacterial transcripts in pulmonary tuberculosis, as diagnostic targets for a serodiagnostic assay. A total of 300 subjects were recruited including smear+, smear- pulmonary tuberculosis (PTB) patients, sarcoidosis patients, lung cancer patients and healthy controls. Proteins encoded by eight in vivo expressed transcripts selected from previous study including those encoded by two topmost expressed and six RD transcripts (Rv0986, Rv0971, Rv1965, Rv1971, Rv2351c, Rv2657c, Rv2674, Rv3121) were analyzed for B-cell epitopes by peptide arrays/bioinformatics. Enzyme-linked immunosorbent assay was used to evaluate the antibody response against the selected peptides in sera from PTB and controls. Overall 12 peptides were selected for serodiagnosis. All the peptides were initially screened for their antibody response. The peptide with highest sensitivity and specificity was further assessed for its serodiagnostic ability in all the study subjects. The mean absorbance values for antibody response to selected peptide were significantly higher (p<0.001) in PTB patients as compared to healthy controls; however, the sensitivity for diagnosis of PTB was 31% for smear+ and 20% for smear- PTB patients. Thus, the peptides encoded by in vivo expressed transcripts elicited a significant antibody response, but are not suitable candidates for serodiagnosis of PTB.

Evaluation of low-density lipoprotein cholesterol equations by cross-platform assessment of accuracy-based EQA data against SI-traceable reference value.

OBJECTIVES: Low-density lipoprotein cholesterol (LDLC) is the primary cholesterol target for the diagnosis and treatment of cardiovascular disease (CVD). Although beta-quantitation (BQ) is the gold standard to determine LDLC levels accurately, many clinical laboratories apply the Friedewald equation to calculate LDLC. As LDLC is an important risk factor for CVD, we evaluated the accuracy of Friedewald and alternative equations (Martin/Hopkins and Sampson) for LDLC. METHODS: We calculated LDLC based on three equations (Friedewald, Martin/Hopkins and Sampson) using the total cholesterol (TC), triglycerides (TG), and high-density lipoprotein cholesterol (HDLC) in commutable serum samples measured by clinical laboratories participating in the Health Sciences Authority (HSA) external quality assessment (EQA) programme over a 5 years period (number of datasets, n=345). LDLC calculated from the equations were comparatively evaluated against the reference values, determined from BQ-isotope dilution mass spectrometry (IDMS) with traceability to the International System of Units (SI). RESULTS: Among the three equations, Martin/Hopkins equation derived LDLC had the best linearity against direct measured (y=1.141x - 14.403; R2=0.8626) and traceable LDLC (y=1.1692x - 22.137; R2=0.9638). Martin/Hopkins equation (R2=0.9638) had the strongest R2 in association with traceable LDLC compared with the Friedewald (R2=0.9262) and Sampson (R2=0.9447) equation. The discordance with traceable LDLC was the lowest in Martin/Hopkins (median=-0.725%, IQR=6.914%) as compared to Friedewald (median=-4.094%, IQR=10.305%) and Sampson equation (median=-1.389%, IQR=9.972%). Martin/Hopkins was found to result in the lowest number of misclassifications, whereas Friedewald had the most numbers of misclassification. Samples with high TG, low HDLC and high LDLC had no misclassification by Martin/Hopkins equation, but Friedewald equation resulted in ∼50% misclassification in these samples. CONCLUSIONS: The Martin/Hopkins equation was found to achieve better agreement with the LDLC reference values as compared to Friedewald and Sampson equations, especially in samples with high TG and low HDLC. Martin/Hopkins derived LDLC also enabled a more accurate classification of LDLC levels.

Cerebrospinal fluid adenosine deaminase for the diagnosis of tuberculous meningitis.

Background: A consensus on the diagnostic utility of cerebrospinal fluid adenosine deaminase (ADA) for tuberculous meningitis (TBM) is lacking. Methods: Patients aged ≥12 years admitted with CNS infections were enrolled prospectively. ADA was measured with spectrophotometry. Results: We enrolled 251 TBM and 131 other CNS infections. The optimal cutoff of ADA was calculated at 5.5 U/l against microbiological reference standard with area under curve 0.743, sensitivity 80.7%, specificity 60.3%, positive likelihood ratio 2.03 and negative likelihood ratio 3.12. The widely used cutoff value 10 U/l had specificity 82% and sensitivity 50%. The discriminating power was higher for TBM versus viral meningoencephalitis than bacterial or cryptococcal meningitis. Conclusion: Cerebrospinal fluid ADA has a low-to-modest diagnostic utility.

The diagnosis of tuberculosis (TB) of the brain is mainly made by testing cerebrospinal fluid, a clear liquid that flows in and around the brain and spinal cord. Adenosine deaminase (ADA) is a protein whose production and activity are increased in many diseases, such as TB. ADA testing in cerebrospinal fluid is widely used for the diagnosis of brain TB. However, the experts have split opinions regarding its confirmatory role. This study explores ADA measurement in cerebrospinal fluid for differentiating TB from other brain infections. The report says that this simple and inexpensive test can be helpful, but it cannot make or refute the diagnosis of brain TB and should only be considered along with other tests.