Drug-resistant Mycobacterium tuberculosis among Nepalese patients at a tuberculosis referral center.

BACKGROUND: Multidrug-resistant tuberculosis (MDR-TB), characterized by isoniazid and rifampicin resistance, is caused by chromosomal mutations that restrict treatment options and complicate tuberculosis management. This study sought to investigate the prevalence of pre-extensively drug-resistant (pre-XDR) and extensively drug-resistant (XDR) tuberculosis, as well as mutation pattern, in Nepalese patients with MDR/rifampicin-resistant (RR)-TB strains. METHODS: A cross-sectional study was conducted on MDR/RR-TB patients at the German Nepal Tuberculosis Project from June 2017 to June 2018. The MTBDRsl line probe assay identified pre-XDR-TB and XDR-TB. Pre-XDR-TB included MDR/RR-TB with resistance to any fluoroquinolone (FLQ), while XDR-TB included MDR/RR-TB with resistance to any FLQ and at least one additional group A drug. Mutation status was determined by comparing bands on reaction zones [gyrA and gyrB for FLQ resistance, rrs for SILD resistance, and eis for low-level kanamycin resistance, according to the GenoType MTBDRsl VER 2.0, Hain Lifescience GmbH, Nehren, Germany definition of pre-XDR and XDR] to the evaluation sheet. SPSS version 17.0 was used for data analysis. RESULTS: Out of a total of 171 patients with MDR/RR-TB, 160 had (93.57%) had MTBC, of whom 57 (35.63%) had pre-XDR-TB and 10 (6.25%) had XDR-TB. Among the pre-XDR-TB strains, 56 (98.25%) were FLQ resistant, while 1 (1.75%) was SLID resistant. The most frequent mutations were found at codons MUT3C (57.14%, 32/56) and MUT1 (23.21%, 13/56) of the gyrA gene. One patient had SLID resistant genotype at the MUT1 codon of the rrs gene (100%, 1/1). XDR-TB mutation bands were mostly detected on MUT1 (30%, 3/10) of the gyrA and rrs, MUT3C (30%, 3/10) of the gyrA, and MUT1 (30%, 3/10) of the rrs. CONCLUSIONS: Pre-XDR-TB had a significantly higher likelihood than XDR-TB, with different specific mutation bands present in gyrA and rrs genes.

Determinants of multidrug-resistant tuberculosis among adults undergoing treatment for tuberculosis in Tigray Region, Ethiopia: a case-control study.

BACKGROUND: Multidrug-resistant tuberculosis is a type of tuberculosis that is resistant to at least the first-line antituberculosis drugs namely, rifampicin and isoniazid. However, most of these studies were limited only to a single hospital. Therefore, this study aimed to identify the determinants of multidrug-resistant tuberculosis among adults undergoing treatment for tuberculosis in the Tigray region of Ethiopia. METHODS: Hospital-based unmatched case-control study was conducted from 1 April 2019 to 30 June 2019. A simple random sampling method was used to select the required sample size. Variables at a p value less than 0.25 in bivariate analysis were entered into a multivariable analysis to identify the determinant factors of multidrug-resistant tuberculosis. Finally, the level of significance was declared at p<0.05. RESULTS: Rural residence (adjusted OR (AOR) 2.54; 95% CI 1.34 to 4.83), HIV (AOR 4.5; 95% CI 1.4 to 14.2), relapse (AOR 3.86; 95% CI 1.98 to 7.5), return after lost follow-up (AOR 6.29; 95% CI 1.64 to 24.2), treatment failure (AOR 5.87; 95% CI 1.39 to 24.8) were among the determinants of multidrug-resistant tuberculosis. CONCLUSION: Rural residence, HIV, relapses, return after lost follow-up and treatment failure were the identified determinant factors of multidrug-resistance tuberculosis.

Comparative evaluation of intensified short course regimen and standard regimen for adults TB meningitis: a protocol for an open label, multi-center, parallel arms, randomized controlled superiority trial (INSHORT trial).

BACKGROUND: Despite several incremental improvements in the management of tuberculous meningitis (TBM), the mortality rates remain high. In spite of national and international guidelines, variation in the choice, dose, and duration of drugs exist between countries and clinicians. We propose to evaluate a shorter and more effective regimen containing agents with augmented intracerebral drug exposure and anti-inflammatory approaches to improve disability-free survival among patients with TBM. Our strategy incorporates the various developments in the field of TBM over the last two decades and only few trials have evaluated a composite of these strategies in the overall outcomes of TBM. METHODS: An open label, parallel arms, randomized controlled superiority trial will be conducted among 372 participants across 6 sites in India. Eligible participants will be randomly allocated in 1:1:1 ratio into one of the three arms. The intervention arm consists of 2 months of high-dose rifampicin (25 mg/kg), moxifloxacin (400 mg), pyrazinamide, isoniazid, aspirin (150 mg), and steroids followed by rifampicin, isoniazid, and pyrazinamide for 4 months. The second intervention arm includes all the drugs as per the first arm except aspirin and the patients in the control arm will receive treatment according to the National TB Elimination Program guidelines. All participants will be followed up for 1 year after the treatment.  DISCUSSION: Current WHO regimens have agents with poor central nervous system drug exposure and is too long. It does not reflect the accumulating evidence in the field. We propose a comprehensive clinical trial incorporating the emerging evidence accrued over the last two decades to shorten the duration and improve the treatment outcomes. This multi-centric trial may generate crucial evidence with policy and practice implications in the treatment of TBM. TRIAL REGISTRATION: Clinical Trial Registry India CTRI/2023/05/053314. Registered on 31 May 2023 ( https://ctri.nic.in/Clinicaltrials/pmaindet2.php?EncHid=ODYzMzg=&Enc=&userName=CTRI/2023/05/053314 ). CLINICALTRIALS: gov NCT05917340. Registered on 6 August 2023 ( https://classic. CLINICALTRIALS: gov/ct2/show/NCT05917340 ). PROTOCOL VERSION: Version 1.3 dated 12 July 2023.

Genotypic and phenotypic drug resistance patterns of Mycobacterium tuberculosis isolated from presumptive pulmonary tuberculosis patients in Ethiopia: A multicenter study.

BACKGROUND: The emergence of drug-resistant tuberculosis (DR-TB) has been a major obstacle to global tuberculosis control programs, especially in developing countries, including Ethiopia. This study investigated drug resistance patterns and associated mutations of Mycobacterium tuberculosis Complex (MTBC) isolates from the Amhara, Gambella, and Benishangul-Gumuz regions of Ethiopia. METHODS: A cross-sectional study was conducted using 128 MTBC isolates obtained from patients with presumptive tuberculosis (TB). Phenotypic (BACTEC MGIT 960) and genotypic (MTBDRplus and MTBDRsl assays) methods were used for drug susceptibility testing. Data were entered into Epi-info and analyzed using SPSS version 25. Frequencies and proportions were determined to describe drug resistance levels and associated mutations. RESULTS: Of the 127 isolates recovered, 100 (78.7%) were susceptible to four first-line anti-TB drugs. Any drug resistance, polydrug resistance, and multi-drug resistance (MDR) were detected in 21.3% (27), 15.7% (20), and 15% (19) of the isolates, respectively, by phenotypic and/or genotypic methods. Mono-resistance was observed for Isoniazid (INH) (2, 1.6%) and Streptomycin (STR) (2, 1.6%). There were two genotypically discordant RIF-resistant cases and one INH-resistant case. One case of pre-extensively drug-resistant TB (pre-XDR-TB) and one case of extensively drug-resistant TB (XDR-TB) were identified. The most frequent gene mutations associated with INH and rifampicin (RIF) resistance were observed in the katG MUT1 (S315T1) (20, 76.9%) and rpoB (S531L) (10, 52.6%) genes, respectively. Two MDR-TB isolates were resistant to second-line drugs; one had a mutation in the gyrA MUT1 gene, and the other had missing gyrA WT1, gyrA WT3, and rrs WT1 genes without any mutation. CONCLUSIONS: The detection of a significant proportion of DR-TB cases in this study suggests that DR-TB is a major public health problem in Ethiopia. Thus, we recommend the early detection and treatment of DR-TB and universal full first-line drug-susceptibility testing in routine system.

Diagnostic Performance of STANDARD™ M10 Multidrug-resistant Tuberculosis Assay for Detection of Mycobacterium tuberculosis and Rifampicin and Isoniazid Resistance in Zimbabwe.

BACKGROUND: Although Zimbabwe has transitioned out of the 30 high-burden countries, it still remained in the 30 high multidrug-resistant (MDR)/rifampicin-resistant tuberculosis (TB) burden. Rapid detection of rifampicin (RIF) and isoniazid (INH) is essential for the diagnosis of MDR-TB. The World Health Organization has recommended the use of molecular WHO-recommended rapid diagnostic (mWRD) for TB and DR-TB. STANDARD™ M10 MDR-TB assay is a new molecular rapid diagnostic assay developed by SD Biosensor for the detection of Mycobacterium tuberculosis (MTB) and RIF and INF resistance. This study aims to determine the diagnostic accuracy of STANDARD™ M10 MDR-TB assay. METHODS: The study was conducted on 214 samples with different MTB and RIF and INH resistance status. The STANDARD™ M10 MDR-TB assay was performed according to the manufacturer's instructions. Xpert MTB/RIF Ultra, MGIT culture, and phenotypic drug susceptibility testing are used as comparative methods. RESULTS: The sensitivity and specificity of STANDARD™ M10 MDR-TB assay for the detection of MTB are 99% and 97.9%, respectively. The sensitivity and specificity of the assay for detection of MDR-TB were 97.8% and 100%, respectively. CONCLUSION: The STANDARD™ M10 MDR-TB assay demonstrated high diagnostic accuracy in the detection of MTB and RIF and INH resistance. This molecular assay can also be used as an alternative to other mWRD assays.

Rapid Detection of M. tuberculosis and Its Resistance to Rifampicin and Isoniazid with the mfloDx™ MDR-TB test.

BACKGROUND: Rapid detection of tuberculosis (TB) and its resistance are essential for the prompt initiation of correct drug therapy and for stopping the spread of drug-resistant TB. There is an urgent need for increased use of rapid diagnostic tests to control the threat of increased TB and multidrug-resistant TB (MDR-TB). METHODS: EMPE Diagnostics has developed a multiplex molecular diagnostic platform called mfloDx™ by combining nucleotide-specific padlock probe-dependent rolling circle amplification with sensitive lateral flow biosensors, providing visual signals, similar to a COVID-19 test. The first test kit of this platform, mfloDx™ MDR-TB can identify Mycobacterium tuberculosis (MTB) complex and its clinically significant mutations in the rpoB and katG genes and in the inhA promotor contributing resistance to rifampicin (RIF) and isoniazid (INH), causing MDR-TB. RESULTS: We have evaluated the performance of the mfloDx™ MDR-TB test on 210 sputum samples (110 from suspected TB cases and 100 from TB-negative controls) received from a tertiary care center in India. The clinical sensitivity for detecting MTB compared to acid-fast microscopy and mycobacteria growth indicator tube (MGIT) cultures was 86.4% and 84.9%, respectively. All the 100 control samples were negative indicating excellent specificity. In smear-positive sputum samples, the mfloDx™ MDR-TB test showed a sensitivity of 92.5% and 86.4% against MGIT culture and Xpert MTB/RIF, respectively. The clinical sensitivity for the detection of RIF and INH resistance in comparison with MGIT drug susceptibility testing was 100% and 84.6%, respectively, while the clinical specificity was 100%. CONCLUSION: From the above evaluation, we find mfloDx™ MDR-TB to be a rapid and efficient test to detect TB and its multidrug resistance in 3 h at a low cost making it suitable for resource-limited laboratories.

Disseminated Tuberculosis: A 6-year Case Series Experience in a Tertiary Care Center.

BACKGROUND: Disseminated tuberculosis (dTB) disease is associated with a significant burden of morbidity and mortality and it requires improved awareness among clinicians. Case reports revealing the clinical and microbiological characteristics of dTB patients will help us to extend our knowledge of dTB. In our study, we have documented dTB cases followed for 6 years and revealed patients' clinical characteristics. METHODS: Patients followed between 2017 and 2023 who were diagnosed with dTB in a tertiary referral hospital in Istanbul have been evaluated. Data regarding patients' characteristics, methods used in establishing the definitive diagnosis, radiological patterns in chest X-rays, extrapulmonary sites involved, antituberculosis (TB) treatment regimens received, medication side effects, and drug resistance have been examined. Descriptive statistics were performed. RESULTS: Clinical characteristics of 55 patients with a median age of 41 (range 20-85, 52.7% male) were examined. The most common extrapulmonary involvements in our study were the skeletal system (n = 24), central nervous system (n = 7), and genitourinary tract (n = 7). Isoniazid (INH) resistance was detected in four patients. Mono resistance was reported for pyrazinamide in one patient. Multidrug resistance was detected in two patients and one of them was also resistant to ethambutol. Preextensively, drug resistance was reported in three patients. Another three patients were evaluated as resistant to both INH and streptomycin. CONCLUSION: Migrating from a high TB burden country and comorbidities such as diabetes mellitus, human immunodeficiency virus, and rheumatoid arthritis that are related to immunocompromisation are thought to be risk factors for dTB.

Tuberculosis preventive treatment uptake among people living with HIV during COVID-19 period in Addis Ababa, Ethiopia: a retrospective data review.

BACKGROUND: Screening for tuberculosis (TB) and providing TB preventive treatment (TPT) along with antiretroviral therapy is key components of human immune deficiency virus (HIV) care. The uptake of TPT during the coronavirus disease 2019 (COVID-19) period has not been adequately assessed in Addis Ababa City Administration. This study aimed at assessing TPT uptake status among People living with HIV (PLHIV) newly initiated on antiretroviral therapy during the COVID-19 period at all public hospitals of Addis Ababa City Administration, Ethiopia. METHODS: A retrospective data review was conducted from April-July 2022. Routine District Health Information System 2 database was reviewed for the period from April 2020-March 2022. Proportion and mean with standard deviation were computed. Logistic regression analysis was conducted to assess factors associated with TPT completion. A p-value of < 0.05 was considered statistically significant. RESULTS: A total of 1,069 PLHIV, aged 18 years and above were newly initiated on antiretroviral therapy, and of these 1,059 (99.1%) underwent screening for TB symptoms. Nine hundred twelve (86.1%) were negative for TB symptoms. Overall, 78.8% (719) of cases who were negative for TB symptoms were initiated on TPT, and of these 70.5% and 22.8% were completed and discontinued TPT, respectively. Of 719 cases who were initiated on TPT, 334 (46.5%) and 385 (53.5%) were initiated on isoniazid plus rifapentine weekly for three months and Isoniazid preventive therapy daily for six months, respectively. PLHIV who were initiated on isoniazid plus rifapentine weekly for three months were more likely to complete TPT (adjusted odds ratio [AOR],1.68; 95% confidence interval [CI], 1.01, 2.79) compared to those who were initiated on Isoniazid preventive therapy daily for six months. CONCLUSION: While the proportion of PLHIV screened for TB was high, TPT uptake was low and far below the national target of achieving 90% TPT coverage. Overall a considerable proportion of cases discontinued TPT in this study. Further strengthening of the programmatic management of latent TB infection among PLHIV is needed. Therefore, efforts should be made by the Addis Ababa City Administration Health Bureau authorities and program managers to strengthen the initiation and completion of TPT among PLHIV in public hospitals.

Peroxynitrite imaging in ferroptosis-mediated drug-induced liver injury with a near-infrared fluorescence probe.

BACKGROUND: Over-consumption of drugs can result in drug-induced liver damage (DILI), which can worsen liver failure. Numerous studies have shown the significant role ferroptosis plays in the pathophysiology of DILI, which is typified by a marked imbalance between the generation and breakdown of lipid reactive oxygen species (ROS). The content of peroxynitrite (ONOO-) rapidly increased during this process and was thought to be a significant marker of early liver injury. Therefore, the construction of fluorescence probe for the detection and imaging of ONOO- holds immense importance in the early diagnosis and treatment of ferroptosis-mediated DILI. RESULTS: We designed a probe DILI-ONOO based on the ICT mechanism for the purpose of measuring and visualizing ONOO- in ferroptosis-mediated DILI processes and associated studies. This probe exhibited significant fluorescence changes with good sensitivity, selectivity, and can image exogenous and endogenous ONOO- in cells with low cytotoxicity. Using this probe, we were able to show changes in ONOO- content in ferroptosis-mediated DILI cells and mice models induced by the intervention of acetaminophen (APAP) and isoniazid (INH). By measuring the concentration of ferroptosis-related indicators in mice liver tissue, we were able to validate the role of ferroptosis in DILI. It is worth mentioning that compared to existing alanine transaminase (ALT) and aspartate aminotransferase (AST) detection methods, this probe can achieve early identification of DILI prior to serious liver injury. SIGNIFICANCE: This work has significant reference value in researching the relationship between ferroptosis and DILI and visualizing research. The results indicate a strong correlation between the progression of DILI and ferroptosis. Additionally, the use of DILI-ONOO shows promise in investigating the DILI process and assessing the effectiveness of medications in treating DILI.

Risk factors of adult isoniazid-resistant and rifampicin-susceptible tuberculosis in Nanjing, 2019-2021.

INTRODUCTION: This study aimed to analyze the risk factors associated with isoniazid-resistant and rifampicin-susceptible tuberculosis (Hr-TB) in adults. METHOD: The clinical data of 1,844 adult inpatients diagnosed with culture-positive pulmonary tuberculosis (PTB) in Nanjing Second Hospital from January 2019 and December 2021 were collected. All culture positive strain from the patient specimens underwent drug susceptibility testing (DST). Among them, 166 patients with Hr-TB were categorized as the Hr-TB group, while the remaining 1,678 patients were classified as having drug-susceptible tuberculosis (DS-TB). Hierarchical logistic regression was employed for multivariate analysis to identify variables associated with Hr-TB. RESULTS: Multivariate logistic regression analysis revealed that individuals with diabetes mellitus (DM) (OR 1.472, 95% CI 1.037-2.088, p = 0.030) and a history of previous tuberculosis treatment (OR 2.913, 95% CI 1.971-4.306, p = 0.000) were at higher risk of developing adult Hr-TB, with this risk being more pronounced in male patients. Within the cohort, 1,640 patients were newly treated, and among them, DM (OR 1.662, 95% CI 1.123-2.461, p = 0.011) was identified as risk factors for Hr-TB. CONCLUSIONS: Diabetes mellitus is a risk factor for Hr-TB in adults, and the contribution of diabetes as a risk factor was more pronounced in the newly treatment or male subgroup. And previous TB treatment history is also a risk factor for Hr-TB in adults.

Large-scale statistical analysis of Mycobacterium tuberculosis genome sequences identifies compensatory mutations associated with multi-drug resistance.

Tuberculosis (TB), caused by Mycobacterium tuberculosis, has a significant impact on global health worldwide. The development of multi-drug resistant strains that are resistant to the first-line drugs isoniazid and rifampicin threatens public health security. Rifampicin and isoniazid resistance are largely underpinned by mutations in rpoB and katG respectively and are associated with fitness costs. Compensatory mutations are considered to alleviate these fitness costs and have been observed in rpoC/rpoA (rifampicin) and oxyR'-ahpC (isoniazid). We developed a framework (CompMut-TB) to detect compensatory mutations from whole genome sequences from a large dataset comprised of 18,396 M. tuberculosis samples. We performed association analysis (Fisher's exact tests) to identify pairs of mutations that are associated with drug-resistance, followed by mediation analysis to identify complementary or full mediators of drug-resistance. The analyses revealed several potential mutations in rpoC (N = 47), rpoA (N = 4), and oxyR'-ahpC (N = 7) that were considered either 'highly likely' or 'likely' to confer compensatory effects on drug-resistance, including mutations that have previously been reported and validated. Overall, we have developed the CompMut-TB framework which can assist with identifying compensatory mutations which is important for more precise genome-based profiling of drug-resistant TB strains and to further understanding of the evolutionary mechanisms that underpin drug-resistance.

Experimental and Computational Studies on the Interaction of DNA with Hesperetin Schiff Base CuII Complexes.

The interactions with calf thymus DNA (CT-DNA) of three Schiff bases formed by the condensation of hesperetin with benzohydrazide (HHSB or L1H3), isoniazid (HIN or L2H3), or thiosemicarbazide (HTSC or L3H3) and their CuII complexes (CuHHSB, CuHIN, and CuHTSC with the general formula [CuLnH2(AcO)]) were evaluated in aqueous solution both experimentally and theoretically. UV-Vis studies indicate that the ligands and complexes exhibit hypochromism, which suggests helical ordering in the DNA helix. The intrinsic binding constants (Kb) of the Cu compounds with CT-DNA, in the range (2.3-9.2) × 106, from CuHTSC to CuHHSB, were higher than other copper-based potential drugs, suggesting that π-π stacking interaction due to the presence of the aromatic rings favors the binding. Thiazole orange (TO) assays confirmed that ligands and Cu complexes displace TO from the DNA binding site, quenching the fluorescence emission. DFT calculations allow for an assessment of the equilibrium between [Cu(LnH2)(AcO)] and [Cu(LnH2)(H2O)]+, the tautomer that binds CuII, amido (am) and not imido (im), and the coordination mode of HTSC (O-, N, S), instead of (O-, N, NH2). The docking studies indicate that the intercalative is preferred over the minor groove binding to CT-DNA with the order [Cu(L1H2am)(AcO)] > [Cu(L2H2am)(AcO)] ≈ TO ≈ L1H3 > [Cu(L3H2am)(AcO)], in line with the experimental Kb constants, obtained from the UV-Vis spectroscopy. Moreover, dockings predict that the binding strength of [Cu(L1H2am)(AcO)] is larger than [Cu(L1H2am)(H2O)]+. Overall, the results suggest that when different enantiomers, tautomers, and donor sets are possible for a metal complex, a computational approach should be recommended to predict the type and strength of binding to DNA and, in general, to macromolecules.

Screening for Latent Tuberculosis Infection in People Living with HIV: TUBHIVIT Project, a Multicenter Italian Study.

BACKGROUND: The coexistence of HIV infection and latent tuberculosis infection (LTBI) presents a significant public health concern due to the increased risk of tuberculosis (TB) reactivation and progression to active disease. The multicenter observational cohort study, TUBHIVIT, conducted in Italy from 2017 to 2023, aimed to assess the prevalence of LTBI among people living with HIV (PLHIV) and their outcomes following LTBI screening and therapy initiation. METHODS: We performed a prospective study in five referral centers for HIV care in Italy. PLHIV who consented Tto participate underwent QuantiFERON-TB Gold Plus and clinical, microbiological, and radiological assessments to exclude subclinical tuberculosis, as opportune. PLHIV diagnosed with LTBI who started chemoprophylaxis were followed until the end of therapy. RESULTS: A total of 1105 PLHIV were screened for LTBI using the QuantiFERON-TB Gold Plus test, revealing a prevalence of 3.4% of positive results (38/1105). Non-Italy-born individuals exhibited a significantly higher likelihood of testing positive. Thirty-one were diagnosed with LTBI, 1 showed active subclinical TB, and 6 were lost to follow-up before discriminating between latent and active TB. Among the PLHIV diagnosed with LTBI, 83.9% (26/31) started chemoprophylaxis. Most individuals received 6-9 months of isoniazid-based therapy. Of the 26 PLHIV commencing chemoprophylaxis, 18 (69.2%) completed the therapy, while 3 discontinued it and 5 were still on treatment at the time of the analysis. Adverse events were observed in two cases, while in one case the patient refused to continue the treatment.

Correlation between serum isoniazid concentration and therapeutic response in patients of pulmonary tuberculosis in Central India: A prospective observational study.

BACKGROUND: Tuberculosis (TB), an infectious disease caused by Mycobacterium tuberculosis is one of the top ten causes of death worldwide. Isoniazid (INH) is an important component of anti-tuberculosis therapy (ATT). Low isoniazid levels can serve as a risk factor for the development of treatment failure, relapse of disease and acquired secondary resistance. Hence, serum level of isoniazid becomes a critical factor in determining the treatment outcome of patients on ATT. This study aimed to evaluate the correlation between serum isoniazid concentration and therapeutic response in patients of pulmonary tuberculosis in Central India. METHODS: This was a prospective single cohort observational study conducted at a tertiary care hospital. Therapeutic response in newly diagnosed patients of pulmonary TB was determined based the microbiological, clinical and radiological parameters. Serum INH levels were estimated based on a spectrophotometric method using nano-spectrophotometer. RESULTS: In this study, patients had a significant improvement in treatment outcome as evident by a significant decrease in the TB score I at end of IP (p = 0.001) and a significant decline in the Timika score at end of CP (p = 0.001). Although all patients converted to sputum negative at end of CP, 20% remained positive at end of IP. Lower INH levels were seen in 13.3% of the study population. Higher INH levels were observed in sputum converters, patients with low TB score I and low Timika score, although no statistically significant difference was noted (p > 0.05). CONCLUSION: In this study, we could not find any statistically significant association between serum INH levels and therapeutic outcome of the patients. Further studies on a larger population could provide better understanding about the prevalence of low serum isoniazid levels among the Indian population and establish its relationship with therapeutic outcome. Also, the usage of a comparatively less expensive spectrophotometric method of analysis makes this feasible in almost every district hospital without the need of high-performance liquid chromatography which is costlier and needs more expertise.

Application of the Hollow-Fiber Infection Model to Personalized Precision Dosing of Isoniazid in a Clinical Setting.

BACKGROUND: The hollow-fiber infection model (HFIM) is a valuable tool for evaluating pharmacokinetics/pharmacodynamics relationships and determining the optimal antibiotic dose in monotherapy or combination therapy, but the application for personalized precision medicine in tuberculosis treatment remains limited. This study aimed to evaluate the efficacy of adjusted antibiotic doses for a tuberculosis patient using HFIM. METHODS: Model-based Bayesian forecasting was utilized to assess the proposed reduction of the isoniazid dose from 300 mg daily to 150 mg daily in a patient with an ultra-slow-acetylation phenotype. The efficacy of the adjusted 150-mg dose was evaluated in a time-to-kill assay performed using the bacterial isolate Mycobacterium tuberculosis (Mtb) H37Ra in a HFIM that mimicked the individual pharmacokinetic profile of the patient. RESULTS: The isoniazid concentration observed in the HFIM adequately reflected the target drug exposures simulated by the model. After 7 days of repeated dose administration, isoniazid killed 4 log10 Mtb CFU/mL in the treatment arm, while the control arm without isoniazid increased 1.6 log10 CFU/mL. CONCLUSION: Our results provide an example of the utility of the HFIM for predicting the efficacy of specific recommended doses of anti-tuberculosis drugs in real clinical setting.

Development of a multiplex droplet digital PCR method for detection and monitoring of Mycobacterium tuberculosis and drug-resistant tuberculosis.

BACKGROUND: The prevalence of multidrug-resistant tuberculosis (MDR-TB) among Korean tuberculosis patients is about 4.1%, which is higher than the OECD average of 2.6%. Inadequate drug use and poor patient compliance increase MDR-TB prevalence through selective pressure. Therefore, prompt detection of drug resistance in tuberculosis patients at the time of diagnosis and quantitative monitoring of these resistant strains during treatment are crucial. METHODS: A multiplex droplet digital PCR (ddPCR) assay was developed and assessed using DNA material of nine Mycobacterium tuberculosis strains with known mutation status that were purchased from the Korean National Tuberculosis Association. We collected a total of 18 MDR-TB residual samples referred for PCR analysis. Total DNA was extracted from the samples and subjected to the quadruplex ddPCR assay. Their results were compared to those of known resistance phenotypes. RESULTS: The analytical sensitivity and specificity of the multiplex ddPCR assay for detecting INH, RIF, EMB, FQ, and SM resistance-causing mutations ranged from 71.43 to 100% and 94.12-100%, respectively. Follow-up sample results showed that the quadruplex ddPCR assay was sensitive enough to detect IS6110 and other mutations even after onset of treatment. CONCLUSIONS: We developed a sensitive and accurate multiplex ddPCR assay that can detect the presence of tuberculosis quantitatively and resistance-conveying mutations concurrently. This tool could aid clinicians in the diagnosis and treatment monitoring of tuberculosis.

AI-driven design of customized 3D-printed multi-layer capsules with controlled drug release profiles for personalized medicine.

Personalized medicine aims to effectively and efficiently provide customized drugs that cater to diverse populations, which is a significant yet challenging task. Recently, the integration of artificial intelligence (AI) and three-dimensional (3D) printing technology has transformed the medical field, and was expected to facilitate the efficient design and development of customized drugs through the synergy of their respective advantages. In this study, we present an innovative method that combines AI and 3D printing technology to design and fabricate customized capsules. Initially, we discretized and encoded the geometry of the capsule, simulated the dissolution process of the capsule with classical drug dissolution model, and verified it by experiments. Subsequently, we employed a genetic algorithm to explore the capsule geometric structure space and generate a complex multi-layer structure that satisfies the target drug release profiles, including stepwise release and zero-order release. Finally, Two model drugs, isoniazid and acetaminophen, were selected and fused deposition modeling (FDM) 3D printing technology was utilized to precisely print the AI-designed capsule. The reliability of the method was verified by comparing the in vitro release curve of the printed capsules with the target curve, and the f2 value was more than 50. Notably, accurate and autonomous design of the drug release curve was achieved mainly by changing the geometry of the capsule. This approach is expected to be applied to different drug needs and facilitate the development of customized oral dosage forms.

Pharmacogenetic variability of tuberculosis biomarkers in native and mestizo Peruvian populations.

In Peru, 29 292 people were diagnosed with tuberculosis in 2022. Although tuberculosis treatments are effective, 3.4%-13% are associated with significant adverse drug reactions, with drug-induced liver injury (DILI) considered the most predominant. Among the first-line antituberculosis drugs, isoniazid is the main drug responsible for the appearance of DILI. In liver, isoniazid (INH) is metabolized by N-acetyltransferase-2 (NAT2) and cytochrome P450 2E1 (CYP2E1). Limited information exists on genetic risk factors associated with the presence of DILI to antituberculosis drugs in Latin America, and even less is known about these factors in the native and mestizo Peruvian population. The aim of this study was to determine the prevalence of NAT2 and CYP2E1 genotypes in native and mestizo population. An analytical cross-sectional analysis was performed using genetic data from mestizo population in Lima and native participants from south of Peru. NAT2 metabolizer was determined as fast, intermediate and slow, and CYP2E1 genotypes were classified as c1/c1, c1/c2 and c2/c2, from molecular tests and bioinformatic analyses. Of the 472 participants, 36 and 6 NAT2 haplotypes were identified in the mestizo and native population, respectively. In mestizo population, the most frequent NAT2*5B and NAT2*7B haplotypes were associated with DILI risk; while in natives, NAT2*5G and NAT2*13A haplotypes were associated with decreased risk of DILI. For CYP2E1, c1/c1 and c1/c2 genotypes are the most frequent in natives and mestizos, respectively. The linkage disequilibrium of NAT2 single nucleotide polymorphisms (SNPs) was estimated, detecting a block between all SNPs natives. In addition, a block between rs1801280 and rs1799929 for NAT2 was detected in mestizos. Despite the limitations of a secondary study, it was possible to report associations between NAT2 and CYP2E alleles with Peruvian native and mestizo by prevalence ratios. The results of this study will help the development of new therapeutic strategies for a Tuberculosis efficient control between populations.

Efficacy, safety, and pharmacokinetics of isoniazid affected by NAT2 polymorphisms in patients with tuberculosis: A systematic review.

N-acetyltransferase 2 (NAT2) genetic polymorphisms might alter isoniazid metabolism leading to toxicity. We reviewed the impact of NAT2 genotype status on the pharmacokinetics, efficacy, and safety of isoniazid, a treatment for tuberculosis (TB). A systematic search for research articles published in Scopus, PubMed, and Embase until August 31, 2023, was conducted without filters or limits on the following search terms and Boolean operators: "isoniazid" AND "NAT2." Studies were selected if NAT2 phenotypes with pharmacokinetics or efficacy or safety of isoniazid in patients with TB were reported. Patient characteristics, NAT2 status, isoniazid pharmacokinetic parameters, early treatment failure, and the prevalence of drug-induced liver injury were extracted. If the data were given as a median, these values were standardized to the mean. Forty-one pharmacokinetics and 53 safety studies were included, but only one efficacy study was identified. The average maximum concentrations of isoniazid were expressed as supratherapeutic concentrations in adults (7.16 ± 4.85 µg/mL) and children (6.43 ± 3.87 µg/mL) in slow acetylators. The mean prevalence of drug-induced liver injury was 36.23 ± 19.84 in slow acetylators, which was significantly different from the intermediate (19.49 ± 18.20) and rapid (20.47 ± 20.68) acetylators. Subgroup analysis by continent showed that the highest mean drug-induced liver injury prevalence was in Asian slow acetylators (42.83 ± 27.61). The incidence of early treatment failure was decreased by genotype-guided isoniazid dosing in one study. Traditional weight-based dosing of isoniazid in most children and adults yielded therapeutic isoniazid levels (except for slow acetylators). Drug-induced liver injury was more commonly observed in slow acetylators. Genotype-guided dosing may prevent early treatment failure.

Guidelines for tuberculosis screening and preventive treatment among pregnant and breastfeeding women living with HIV in PEPFAR-supported countries.

BACKGROUND: Tuberculosis (TB) preventive treatment (TPT) is recommended by the World Health Organization (WHO) for persons living with HIV, including pregnant and breastfeeding women. Given the President's Emergency Plan for AIDS Relief (PEPFAR)'s investment in TPT services for persons living with HIV as a strategy to prevent TB as well as uncertainty in guidelines and policy regarding use of TPT during pregnancy and the postpartum period, we conducted a review of current relevant national guidelines among PEPFAR-supported countries. METHODS: Our review included 44/49 PEPFAR-supported countries to determine if TB screening and TPT are recommended specifically for pregnant and breastfeeding women living with HIV (WLHIV). National guidelines reviewed and abstracted included TB, HIV, prevention of vertical HIV transmission, TPT, and any other relevant guidelines. We abstracted information regarding TB screening, including screening tools and frequency; and TPT, including timing, regimen, frequency, and laboratory monitoring. RESULTS: Of 44 PEPFAR-supported countries for which guidelines were reviewed, 66% were high TB incidence countries; 41% were classified by WHO as high TB burden countries, and 43% as high HIV-associated TB burden countries. We found that 64% (n = 28) of countries included TB screening recommendations for pregnant WLHIV in their national guidelines, and most (n = 35, 80%) countries recommend TPT for pregnant WLHIV. Fewer countries included recommendations for breastfeeding as compared to pregnant WLHIV, with only 32% (n = 14) mentioning TB screening and 45% (n = 20) specifically recommending TPT for this population; most of these recommend isoniazid-based TPT regimens for pregnant and breastfeeding WLHIV. However, several countries also recommend isoniazid combined with rifampicin (3RH) or rifapentine (3HP). CONCLUSIONS: Despite progress in the number of PEPFAR-supported countries that specifically include TB screening and TPT recommendations for pregnant and breastfeeding WLHIV in their national guidelines, many PEPFAR-supported countries still do not include specific screening and TPT recommendations for pregnant and breastfeeding WLHIV.