BackgroundNontuberculous mycobacteria (NTM) are rare but emerging pathogens in pediatric populations, particularly in countries where BCG vaccination has been discontinued. Their diagnosis is often delayed due to nonspecific symptoms and limited microbiological sensitivity.MethodsWe conducted a retrospective cohort study of all pediatric patients with confirmed or clinically probable NTM disease, diagnosed at the National Institute for Pediatric Tuberculosis and Respiratory Diseases in Slovakia between 2017 and 2024. Medical records from 2012 to 2016 were screened, but no cases fulfilling inclusion criteria were identified. Clinical characteristics, diagnostic approaches, therapeutic strategies, and patient outcomes were systematically evaluated.ResultsIn total, 30 patients were diagnosed. The majority of cases (27/30) involved cervical lymphadenitis (including two with concurrent pulmonary involvement), most commonly affecting children between 1 and 2 years. In addition, one patient was diagnosed with a pulmonary form of NTM disease, one with NTM-associated osteomyelitis, and one with a skin and soft tissue infection. Mycobacterium avium complex was the most frequently identified species, detected in 8 patients. Bacteriological confirmation was achieved in 53.3% of cases, while histological evidence of granulomatous inflammation was found in 86.7%. The median time to diagnosis was 45 days (IQR: 31–109.5), with longer delays in culture-negative patients (p = 0.0648). Surgical excision was performed in 25 of 27 patients with lymphadenitis, 60% received adjunctive antibiotic therapy. Follow-up data were available for 19 patients: 15 had full recovery, 3 experienced recurrent upper respiratory tract infections, and 1 immunocompromised patient died from miliary tuberculosis.ConclusionThis is the first national study on pediatric NTM disease in a post-BCG vaccination era in Slovakia. Despite centralization of care, diagnostic delays were common, particularly in bacteriologically negative cases. These findings underscore the need for early tissue sampling, comprehensive microbiological evaluation, and interdisciplinary collaboration to improve diagnostic efficiency.

The war in Ukraine triggered unprecedented migration into Central Europe, raising concerns regarding tuberculosis (TB) spread. We conducted a molecular-epidemiological study including all Ukrainian migrants with culture-confirmed TB in Slovakia and the Czech Republic between September 2021 and December 2024 (n = 229), together with all other multidrug-resistant (MDR) TB cases reported in these countries between 2023 and 2024 (n = 28). Whole-genome sequencing (WGS) was performed on cultured Mycobacterium tuberculosis complex (MTBC) isolates, and the resulting data were used for phylogenetic reconstruction, lineage classification, genomic resistance prediction, and cluster analysis. Since the war began in 2022, the share of TB cases from Ukrainian migrants in the Czech Republic rose significantly (P < 0.001), along with an evident increase in total MDR-TB cases. Out of 199 MTBC strains from Ukrainian migrants with good-quality WGS data, 129 (64.8

Introduction: Tuberculosis (TB) in the pediatric population remains a major diagnostic challenge. Alterations in iron metabolism, including changes in ferritin and transferrin levels, may reflect not only the host inflammatory response but also iron availability for Mycobacterium tuberculosis (Mtb). The aim of this study was to compare plasma ferritin and transferrin concentrations in pediatric patients with different forms of TB and in controls. Methods: A total of 145 pediatric patients with TB or suspected TB hospitalized at pediatric departments in Dolný Smokovec, Slovakia, and Prague between 2024 and 2025 were included in the study. Plasma ferritin and transferrin concentrations were measured in these patients. The patients were divided into four groups: bacteriologically confirmed TB (bact. TB; n = 22), clinically confirmed TB (clin. TB; n = 36), tuberculosis infection (TBI; n = 29), and controls (n = 58). Results: Transferrin levels were significantly decreased in patients with bact. TB compared with the other groups. In contrast, ferritin concentrations did not differ significantly among the groups, although higher levels were observed in the bact. TB group. These findings reflect differing patterns of ferritin and transferrin in association with active disease. Conclusion: Transferrin and ferritin may be useful biomarkers of TB in pediatric patients, with transferrin appearing to have greater diagnostic potential in our cohort. Since both parameters may reflect not only TB-related processes but also alterations in iron metabolism and the inflammatory response, their diagnostic value may be enhanced when evaluated in combination with other biomarkers.

Paediatric tuberculosis (TB) diagnosis remains a challenge due to the paucibacillary nature of the disease, resulting in 51.00

OBJECTIVES:While the reported incidence of non-tuberculous mycobacterial (NTM) infections is increasing, the true prevalence remains uncertain due to limitations in diagnostics and surveillance. The emergence of rare and novel species underscores the need for characterization to improve surveillance, detection, and management. METHODS:We performed whole-genome sequencing (WGS) and/or targeted deep-sequencing using the Deeplex Myc-TB assay on all NTM isolates collected in Slovakia and the Czech Republic between the years 2019 to 2023 that were unidentifiable at the species level by the routine diagnostic line probe assays (LPA) GenoType CM/AS and NTM-DR. Minimal inhibitory concentrations against amikacin, ciprofloxacin, moxifloxacin, clarithromycin, and linezolid were determined, and clinical data were collected. RESULTS:Twenty-eight cultures from different patients were included, of which 9 (32.1%) met the clinically relevant NTM disease criteria. The majority of those had pulmonary involvement, while two children presented with lymphadenitis. Antimycobacterial resistance rates were low. In total, 15 different NTM species were identified, predominantly rare NTM like M. neoaurum, M. kumamotonense and M. arupense. Notably, clinically relevant M. chimaera variants were also identified with WGS and Deeplex-Myc TB, which, unlike other M. chimaera strains, appeared to be undetectable by LPA assays. Deeplex detected four mixed infections that were missed by WGS analysis. In contrast, WGS identified two novel species, M. celatum and M. branderi, which were not detected by Deeplex-Myc TB. Importantly, one of these novel species strains was associated with clinically relevant pulmonary disease. DISCUSSION:Our study demonstrates the clinical relevance of uncommon NTM and the effectiveness of targeted deep-sequencing combined with WGS in identifying rare and novel NTM species.

Anecdotal information suggests that clinical practice regarding the use of putative hepatoprotective agents in TB treatment varies across countries in the WHO European Region.Between November 2023 and May 2024, we conducted a standardised questionnaire survey on the use of putative hepatoprotective agents in patients receiving TB treatment among Tuberculosis Network European Trials Group (TBnet) representatives in countries in the WHO European Region.We received valid responses from 37 of 53 countries (69.8%), with 16 (43.2%) reporting regular use of putative hepatoprotective agents during anti-TB treatment. Half of these countries ( n = 8) are part of the former Soviet Union. In five countries, these agents are recommended by national guidelines. The most commonly used hepatoprotective agents were silibin/silymarin ( n = 9, 56.3%), ursodeoxycholic acid ( n = 5, 31.3%), and soy phospholipids ( n = 4, 25.0%). Treatment duration varied, with 56.3% ( n = 9) using them for less than 1 month, 18.8% ( n = 3) for 1–3 months, and 18.8% ( n = 3) for 4–6 months.Putative hepatoprotective agents are widely used as an adjunct to TB treatment in the WHO European Region, particularly in the countries of the former Soviet Union, some of which have included them in their national guidelines.

ObjectivesPoint out the social specifics of patients with tuberculosis, which persist despite the socioeconomic development of Slovak society.MethodsA questionnaire survey was conducted on a sample of consecutive patients with newly diagnosed tuberculosis during the first half of 2023. The collected data were aggregated and compared with the results of European Health Interview Survey (EHIS) in 2019.ResultsWe found significant differences in the distribution of the level of education, labour status, nutritional and marital status, financial poverty, smoking habits, alcohol consumption and number of rooms in the dwelling between the general population and TB patients in Slovakia. Moreover, a significant trend in the proportions was observed across contingency table categories for all ordinal variables with more than two levels.ConclusionThere are several significant social differences between patients with tuberculosis and the general Slovak population.

BACKGROUND:Tuberculosis remains a global health concern, with rising pediatric and adolescent cases. The advancement of diagnostic strategies is crucial for effective control, with whole-genome sequencing emerging as a promising tool. This study explores using whole-genome sequencing in pediatric Tuberculosis. METHODS:Mycobacterium tuberculosis isolates from pediatric patients and their contacts were collected between January 2023 and June 2024 in Slovakia and the Czech Republic. The isolates were subjected to WGS to characterize the resistance patterns and transmission. RESULTS:The study included 37 patients in total-30 pediatric cases and 7 adult index cases-with a single M. tuberculosis isolate collected per patient. The phylogenetic analysis results revealed that 32 out of 37 (86.5%) isolates belonged to the Euro-American lineage. Five isolates (13.5%) belonged to the East-Asian lineage. Genotypic resistance to at least one drug was confirmed in 6 patients (16%). 24 patients were divided into 9 clusters (65%), leaving 13 unclustered (35%). Moreover, the concordance between the identification of source case by WGS and epidemiological anamnesis was confirmed in 60% of patients. CONCLUSIONS:Epidemiological data may not always provide accurate insights into the transmission of TB. Consequently, integrating molecular methods, such as WGS, is essential to enhance the reliability and precision of epidemiological analyses.

Despite developing new diagnostics, drugs, and vaccines, treating tuberculosis (TB) remains challenging. Monitoring inflammatory markers can contribute to more precise diagnostics of TB, identifying its active and latent forms, or monitoring its treatment success. We assessed alterations in plasma levels of 48 cytokines in 20 patients (17 males) with active pulmonary TB compared to age-matched healthy controls (n = 18). Blood samples were collected from individuals hospitalised with TB prior to commencing antibiotic therapy, after the first week, and following the third week. The majority of patients received treatment with a combination of four first-line antituberculosis drugs: rifampicin, isoniazid, ethambutol, and pyrazinamide. Plasmatic cytokine levels from patients three times and controls were analyzed using a Bio-Plex Pro Human Cytokine Screening Panel. The results showed significantly higher levels of 31 cytokines (p<0.05) than healthy controls. Three-week therapy duration showed significantly decreased levels of nine cytokines: interferon alpha-2 (IFN-α2), interleukin (IL) 1 alpha (IL-1α), IL-1 receptor antagonist (IL-1ra), IL-6, IL-10, IL-12 p40, IL-17, leukemia inhibitory factor (LIF), and tumor necrosis factor alpha (TNF-α). Out of these, only levels of IL-1α and IL-6 remained significantly elevated compared to controls. Moreover, we have found a negative correlation of 18 cytokine levels with BMI of the patients but no correlation with age. Our results showed a clinical potential for monitoring the levels of specific inflammatory markers after a short treatment duration. The reduction in cytokine levels throughout the course of therapy could indicate treatment success but should be confirmed in studies with more individuals involved and a longer observation period.

Background Inhaled corticosteroids have been widely reported as a preventive measure against the development of severe forms of COVID-19 not only in patients with asthma.Methods In 654 Czech and Slovak patients with asthma who developed COVID-19, we investigated whether the correct use of inhaler containing corticosteroids was associated with a less severe course of COVID-19 and whether this had an impact on the need for hospitalisation, measurable lung functions and quality of life (QoL).Results Of the studied cohort 51.4% had moderate persistent, 29.9% mild persistent and 7.2% severe persistent asthma. We found a significant adverse effect of poor inhaler adherence on COVID-19 severity (p=0.049). We also observed a lower hospitalisation rate in patients adequately taking the inhaler with OR of 0.83. Vital capacity and forced expiratory lung volume deterioration caused by COVID-19 were significantly reversed, by approximately twofold to threefold, in individuals who inhaled correctly.Conclusion Higher quality of inhalation technique of corticosteroids measured by adherence to an inhaled medication application technique (A-AppIT) score had a significant positive effect on reversal of the vital capacity and forced expiratory lung volume in 1 s worsening (p=0.027 and p<0.0001, respectively) due to COVID-19. Scoring higher in the A-AppIT was also associated with significantly improved QoL. All measured variables concordantly and without exception showed a positive improvement in response to better adherence. We suggest that corticosteroids provide protection against the worsening of lungs in patients with COVID-19 and that correct and easily assessable adherence to corticosteroids with appropriate inhalation technique play an important role in preventing severe form of COVID-19.

Mycobacterium abscessus is an emerging opportunistic pathogen affecting patients with chronic lung diseases, primarily cystic fibrosis (CF), or those under immunosuppression. Hence, investigations into the epidemiology and transmission of M. abscessus and accurate antibiotic susceptibility data are essential for the effective treatment of infections caused by this pathogen. This retrospective nationwide study included all clinical M. abscessus isolates (n = 59) from 29 patients diagnosed in the Czech Republic and Slovakia between 2018 and 2023. Whole genome sequencing (WGS) was performed to identify clusters and classify isolates into predominant circulating clones (DCC). Subspecies identification of unique isolates showed subspecies abscessus as the most prevalent (69.0%). The results of drug-susceptibility testing showed that 65.5% of all isolates were resistant to at least three antibiotics tested. CF patients under 24 years of age were the most at-risk group for M. abscessus infection. WGS identified seven clusters (including two cross-border) comprising CF and non-CF patients with a total clustering rate of 48.3%. One cluster involved patients infected with subspecies massiliense strains differing by 0 single nucleotide polymorphisms hospitalized in the same center. Furthermore, we identified representatives of all major DCCs. This study revealed predominant Mycobacterium abscessus complex clones circulating in the Czech Republic and Slovakia. The results show the high discriminatory power of WGS in the molecular epidemiology of M. abscessus and provide supporting evidence of direct or indirect cross-transmission of subspecies massiliense among both CF and non-CF patients. IMPORTANCEThis study highlights the importance of understanding Mycobacterium abscessus transmission because it poses a growing threat to vulnerable populations, especially young cystic fibrosis patients. Investigating how it spreads and which antibiotics work best is crucial for effective treatment. This research used whole genome sequencing to track M. abscessus and found evidence of potential transmission between patients, including across borders. The findings suggest that dominant strains are circulating and some patients may be infected through direct or indirect contact. This knowledge can inform infection control and treatment strategies.

INTRODUCTION AND OBJECTIVES:Post-tuberculosis lung disease (PTLD), as other chronic respiratory disorders, may have infectious complications; some of them can be prevented with vaccinations. So far, no document has discussed the potential role of vaccination in PTLD. Therefore, the objective of this review was to describe vaccination recommendations to prevent infections potentially capable of complicating PTLD. MATERIALS AND METHODS:A non-systematic review of the literature was conducted. The following keywords were used: tuberculosis, vaccination, vaccines and PTLD. PubMed/MEDLINE and Embase were used as the search engine, focusing on English-language literature only. RESULTS:We identified 9 vaccines potentially useful in PTLD. Influenza, pneumococcal and anti-COVID-19 vaccinations should be recommended. Patients with PTLD can also benefit from vaccination against shingles. Vaccination against pertussis is mainly relevant during childhood. Diphtheria, tetanus and measles vaccination are recommended for general population and should be considered in patients with PTLD not previously vaccinated. Tdap (Tetanus, diphtheria, and pertussis) booster should be repeated in every adult every ten years. Vaccination against BCG retains its importance during early childhood in countries where TB is endemic. CONCLUSIONS:Vaccination deserves to be considered among the strategies to prevent and/or mitigate PTLD complications. Further evidence is necessary to better understand which vaccines have the greatest impact and cost-benefit.

Background: The war in Ukraine has led to significant migration to neighboring countries, raising public health concerns. Notable tuberculosis (TB) incidence rates in Ukraine emphasize the immediate requirement to prioritize approaches that interrupt the spread and prevent new infections.Methods: We conducted a prospective genomic surveillance study to assess migration's impact on TB epidemiology in the Czech Republic and Slovakia. Mycobacterium tuberculosis isolates from Ukrainian war refugees and migrants, collected from September 2021 to December 2022 were analyzed alongside 1574 isolates obtained from Ukraine, the Czech Republic, and Slovakia.Results: Our study revealed alarming results, with historically the highest number of Ukrainian tuberculosis patients detected in the host countries. The increasing number of cases of multidrug-resistant TB, significantly linked with Beijing lineage 2.2.1 (p < 0.0001), also presents substantial obstacles to control endeavors. The genomic analysis identified the three highly related genomic clusters, indicating the recent TB transmission among migrant populations. The largest clusters comprised war refugees diagnosed in the Czech Republic, TB patients from various regions of Ukraine, and incarcerated individuals diagnosed with pulmonary TB specialized facility in the Kharkiv region, Ukraine, pointing to a national transmission sequence that has persisted for over 14 years.Conclusions: The data showed that most infections were likely the result of reactivation of latent disease or exposure to TB before migration rather than recent transmission occurring within the host country. However, close monitoring, appropriate treatment, careful surveillance, and social support are crucial in mitigating future risks, though there is currently no evidence of local transmission in EU countries.

Intro: Drug-resistant tuberculosis (DR-TB) has been declared to be a major contributor to antimicrobial resistance worldwide and remains a global public health concern. The genetic basis of antibiotic resistance in clinical DR Mycobactetium tuberculosis isolates has been widely studied and is predominantly attributed to mutations in specific genes. However, the role of some specific mutations in these genes has not yet been fully elucidated. This study aimed to characterize the mutations associated with resistance to first- and second-line anti-TB drugs classified in Group 3: Uncertain significance in the WHO catalogue and to investigate their relation with the phenotypic resistance profile. Methods: We performed whole genome sequencing of 50 strains of Mycobacterium tuberculosis phenotypically resistant to at least one anti-TB drug. All mutations in genes associated with resistance to first- and second-line anti-TB drugs were called and compared with the catalogue of mutations published by WHO. Findings: Variable sensitivities and specificities for genotype-phenotype correlation of resistance predictions ranging from 15% (pyrazinamide) to 92.7% (isoniazid) and 92.3% (izoniazid) to 100% (streptomycin) were obtained. Resistance conferring mutation was found in 22/50 (40%) isolates. Genotypic resistance to streptomycin was confirmed only in 11/30 phenotypically resistant isolates. For 6/30 isolates without a known mutation, we characterized a unique variants in the gidB gene (V77A; V202A; L35P; A119D; a-60c; c11-t) with uncertain significance in STM resistance. In 3 phenotypic PZA-monoresistant isolates we detected the mutation with uncertain significance in pncA (G162R) and kefB (A107V). All these mutations were present in the genome of phenotypically resistant isolates. Conclusion: In this study we characterized several mutations with uncertain significance in resistance to first- and second-line anti-TB drugs. Comparison of the phenotypic and genotypic resistance profile may contribute in the reclassification of some of these mutations with into the group of mutations associated with resistance.

Tuberculosis is a major global health issue, with approximately 10 million people falling ill and 1.4 million dying yearly. One of the most significant challenges to public health is the emergence of drug-resistant tuberculosis. For the last half-century, treating tuberculosis has adhered to a uniform management strategy in most patients. However, treatment ineffectiveness in some individuals with pulmonary tuberculosis presents a major challenge to the global tuberculosis control initiative. Unfavorable outcomes of tuberculosis treatment (including mortality, treatment failure, loss of follow-up, and unevaluated cases) may result in increased transmission of tuberculosis and the emergence of drug-resistant strains. Treatment failure may occur due to drug-resistant strains, non-adherence to medication, inadequate absorption of drugs, or low-quality healthcare. Identifying the underlying cause and adjusting the treatment accordingly to address treatment failure is important. This is where approaches such as artificial intelligence, genetic screening, and whole genome sequencing can play a critical role. In this review, we suggest a set of particular clinical applications of these approaches, which might have the potential to influence decisions regarding the clinical management of tuberculosis patients.

Rationale Tuberculosis (TB) remains a challenging global infectious disease, mainly affecting the lungs. First‐line anti‐TB drugs play a crucial role in slowing down the rapid spread of TB. In addition, the patient might benefit from therapeutic drug monitoring since it has become an accepted clinical tool for optimizing TB treatment. Methods A simple and sensitive liquid chromatography/tandem mass spectrometry method was developed to monitor the plasma level of isoniazid, ethambutol and pyrazinamide in plasma samples. A one‐step extraction procedure using an Ostro™ plate was applied, and extracts were analyzed by gradient elution followed by detection on a mass spectrometer by multiple reaction monitoring mode. Results The analytes were separated within 4.2 min and over the concentration range of 0.2–10 μg/ml for isoniazid and ethambutol and 1–65 μg/ml for pyrazinamide. The method was successfully validated according to the European Medicine Agency guideline for the selectivity, linearity and lower limit of detection, precision and accuracy, matrix effect, extraction recovery, carryover, dilution integrity and stability, and applied for quantification of analytes in clinical samples from TB patients. Conclusions The presented method allows sensitive and reproducible determination of selected anti‐TB drugs with advantages such as low sample volume requirement, short run time of analysis, one‐step sample preparation procedure with capabilities for phospholipids removal, and a low quantification limit as well as a high degree of selectivity.