Treatment of MDR, pre-XDR, XDR and rifampicin-resistant tuberculosis or in case of intolerance to at least rifampicin in Austria, Germany and Switzerland - Amendment to the 2022 consensus-based guideline: Tuberculosis in adulthood by the German Central Committee against Tuberculosis (DZK) on behalf of the German Respiratory Society (DGP) dated 19.09.2023.

Based on the assessment of new evidence, the World Health Organization (WHO) updated its guidelines for the treatment of drug-resistant tuberculosis (TB) in December 2022. The new recommendations and the latest study data made it necessary to update the existing guideline on the treatment of at least rifampicin- (RR-TB) for the German-speaking countries, replacing the respective chapters of the treatment guidelines published 2022. A shortened treatment of proven RR-TB and multidrug-resistant (MDR)-TB for at least 6 months using the fixed and non-modifiable drug combination of bedaquiline, pretomanid, linezolid, and moxifloxacin (BPaLM) is now also recommended for Austria, Germany, and Switzerland under certain conditions considering the existing barriers for the implementation of the new treatment regimen. For the treatment of pre-extensively drug-resistant (pre-XDR)-TB, an individualized treatment for 18 months continues to be the primary recommendation. The non-modifiable drug combination of bedaquiline, pretomanid, and linezolid (BPaL) may be used alternatively in selected pre-XDR-TB cases, provided that all prerequisites are met. The necessary requirements for using BPaLM and BPaL are presented in detail in this amendment to the consensus-based TB treatment guideline for adult patients.

Dual-centre evaluation of the FluoroType MTBDR version 2 assay for detection of Mycobacterium tuberculosis complex and resistance-conferring mutations in pulmonary and extrapulmonary samples from Denmark, Germany and Sierra Leone.

OBJECTIVES: We evaluated the ability of FluoroType MTBDR version 2 (FTv2; Hain Lifescience), a second-step real-time PCR assay, to simultaneously detect Mycobacterium tuberculosis complex (MTBC) DNA and mutations conferring resistance to rifampicin (RIF) and isoniazid (INH), in pulmonary and extrapulmonary samples from patients and compared them with corresponding cultures. METHODS: FTv2 MTBC was evaluated on 1815 and 432 samples from Denmark (DK) and Germany (DE), respectively. RIF and INH resistance mutations were assessed in the German samples and 110 samples from Sierra Leone and subsequently compared to phenotypic antimicrobial susceptibility testing and a composite reference DNA (CRD) based on the GenoType MTBDR line-probe assay and Sanger sequencing or whole-genome sequencing. RESULTS: Of the 584 (557 smear-negative) Danish and 277 (85 smear-negative) German sputum samples, 42 (16) and 246 (54) were culture positive, and 44 (18) and 222 (35) were FTv2 positive, providing an FTv2 sensitivity and specificity of 0.86 (0.63) and 0.98 (DK), 0.90 (0.65) and 1.00 (DE), respectively. The count, sensitivities, and specificities for all pulmonary samples were 1434, 0.79, and 0.99 (DK) and 347, 0.86, and 1.00 (DE), respectively; for extrapulmonary samples, 381, 0.33, 0.99 (DK) and 83, 0.50, and 1.00 (DE). The valid count, sensitivity, and specificity compared with CRD for detecting resistance mutations were RIF 355, 0.99, 0.96, and INH 340, 1.00, and 0.98, respectively. DISCUSSION: FTv2 reliably detects MTBC DNA in pulmonary and extrapulmonary samples and detects resistance mutations for INH and RIF resistance in inhA promoter, katG, and rpoB genes.

Genomic diversity and clinical relevance of Mycobacterium simiae.

Introduction: Mycobacterium simiae is a slow-growing non-tuberculous mycobacterium that can cause non-tuberculous mycobacterium (NTM) pulmonary disease and extrapulmonary infections. Until now, detailed genomic and clinical characteristics, as well as possible transmission routes of this rare pathogen remain largely unknown. Methods: We conducted whole genome sequencing of available M. simiae isolates collected at a tertiary care centre in Central Germany from 2006 to 2020 and set them into context with publicly available M. simiae complex sequences through phylogenetic analysis. Resistance, virulence and stress genes, as well as known Mycobacteriaceae plasmid sequences were detected in whole genome raw reads. Clinical data and course were retrieved and correlated with genomic data. Results: We included 33 M. simiae sensu stricto isolates from seven patients. M. simiae showed low clinical relevance with only two patients fulfilling American Thoracic Society (ATS) criteria in our cohort and three receiving NTM-effective therapy. The bacterial populations were highly stable over time periods of up to 14 years, and no instances of mixed or re-infections with other strains of M. simiae were observed. Clustering with <12 single nucleotide polymorphisms distance was evident among isolates from different patients; however, proof for human-to-human transmission could not be established from epidemiological data. Conclusion: Overall, the available sequence data for M. simiae complex was significantly extended and new insights into its pathogenomic traits were obtained. We demonstrate high longitudinal genomic stability within single patients. Although we cannot exclude human-to-human transmission, we consider it unlikely in the light of available epidemiological data.

[Treatment of MDR, pre-XDR, XDR and rifampicin resistant tuberculosis or in case of intolerance to at least rifampicin in Austria, Germany and Switzerland - Amendment dated 19.09.2023 to the Sk2-Guideline: Tuberculosis in adulthood of the German Central Committee against Tuberculosis (DZK) on behalf of the German Respiratory Society (DGP)]. / Therapie bei MDR-, prä-XDR-, XDR-Tuberkulose und Rifampicin-Resistenz oder bei Medikamentenunverträglichkeit gegenüber mindestens Rifampicin.

In December 2022, based on the assessment of new evidence, the World Health Organization (WHO) updated its guidelines for the treatment of drug-resistant tuberculosis (TB). The evaluation of both, these recommendations, and the latest study data, makes it necessary to update the existing guidelines on the treatment of at least rifampicin-resistant tuberculosis for the German-speaking region, hereby replacing the respective chapters. A shortened MDR-TB treatment of at least 6 month using the fixed and non-modifiable drug combination of bedaquiline, pretomanid, linezolid, and moxifloxacin (BPaLM) is now also recommended for Germany, Austria, and Switzerland under certain conditions. This recommendation applies to TB cases with proven rifampicin resistance, including rifampicin monoresistance. For treatment of pre-extensively drug resistant TB (pre-XDR-TB), an individualized treatment for 18 months adjusted to resistance data continues to be the primary recommendation. The non-modifiable drug combination of bedaquiline, pretomanid, and linezolid (BPaL) may be used alternatively in pre-XDR TB if all prerequisites are met. The necessary prerequisites for the use of BPaLM and BPaL are presented in this amendment to the S2k guideline for 'Tuberculosis in adulthood'.

[Tuberculosis Infection Control & Hygiene - Recommendations of the DZK]. / Infektionsprävention & Hygiene bei Tuberkulose ­ Empfehlungen des DZK.

Preventing the spread of the disease is an essential goal in the care and treatment of tuberculosis. In addition to early diagnosis and effective therapies, isolation of infectious patients and adequate hygiene measures are of particular importance for infection prevention. The present recommendations replace the previous recommendations "tuberculosis infection control" from 2012 and take into account the current national and international recommendations and as well as new scientific findings. After a description of the infection and the transmission pathways, the necessary prevention and hygiene measures in health care facilities are comprehensively presented. Since the last revision of the recommendations on infection prevention, international recommendations and the KRINKO recommendation on ending isolation have been changed. In accordance with this, under certain conditions in the case of sensitive tuberculosis, de-isolation in health care facilities can take place after 14 days without taking the sputum findings into account. The second part of the recommendations explains in detail the measures to be taken in special situations and areas, such as general practitioners, ambulance services and care facilities. Here, the recommendations on respiratory protection have been simplified; for staff, an FFP2 mask is now generally considered sufficient.

[Update: Diagnostics and treatment of pulmonary tuberculosis]. / Update: Diagnostik und Therapie der pulmonalen Tuberkulose.

Molecular diagnostic tools have changed the approach to the detection of Mycobacterium tuberculosis and associated drug-resistance substantially. PCR-based technologies allow a more rapid detection with higher diagnostic sensitivity in pulmonary and extrapulmonary specimens. However, a real point of care test, which needs minimal technical resources remains missing. Genome sequencing technologies are currently changing tuberculosis drug resistance testing, and for some questions are replacing phenotypic drug resistance testing, based on culture.New evidence on treatment for drug-sensitive tuberculosis allows shortening of treatment to 4 months, or in selected cases even to 2 months based on the use of fluoroquinolones, high dose rifamycins and newly developed TB medicines.Such developments will very likely simplify the management of tuberculosis, although prevention remains the most important pillar of any tuberculosis related public health strategy.

[Treatment of tuberculosis: what is new?] / Therapie der Tuberkulose: Was gibt es Neues?

Never before have so many people around the world been simultaneously affected by tuberculosis. Tuberculosis is the leading cause of death from a bacterial infectious disease worldwide. The World Health Organization's ambitious goal from 2014 of achieving global elimination of tuberculosis does not seem realistic, but on current trends, tuberculosis could be eliminated in the European Union by 2040. Since the beginning of 2022, there have been more innovations for the treatment of tuberculosis than in no other comparable time period before. One month of rifapentine and isoniazid is effective in treating latent tuberculosis infection. However, rifapentine is licensed in the USA but not in the EU and must be imported for individual cases. The duration of the standard treatment for tuberculosis can be shortened to four months but this treatment regimen is also based on rifapentine, in addition to isoniazid, pyrazinamide, and moxifloxacin. The approval of rifapentine in Europe is a much-needed step towards shortening the treatment of tuberculosis. With new drugs an even shorter standard treatment of only 2 months is possible. The treatment of multidrug-resistant/rifampicin-resistant tuberculosis (MDR-/RR-TB) has been shortened to six months, the same length as the standard treatment available in Germany. The combination of bedaquiline, pretomanid, linezolid ± moxifloxacin, cured around 90% of affected patients were cured in studies with a treatment duration of six months. With 19 drugs in clinical trials, the treatment of tuberculosis is expected to continue to improve rapidly in the coming years.

Rapid molecular diagnostics of tuberculosis resistance by targeted stool sequencing.

BACKGROUND: Stool is an important diagnostic specimen for tuberculosis in populations who struggle to provide sputum, such as children or people living with HIV. However, the culture of Mycobacterium tuberculosis (M. tuberculosis) complex strains from stool perform poorly. This limits the opportunity for phenotypic drug resistance testing with this specimen. Therefore, reliable molecular methods are urgently needed for comprehensive drug resistance testing on stool specimens. METHODS: We evaluated the performance of targeted next-generation sequencing (tNGS, Deeplex® Myc-TB) for the detection of mutations associated with M. tuberculosis complex drug resistance on DNA isolated from stool specimens provided by participants from a prospective cohort of patients treated for tuberculosis in Eswatini (n = 66; 56 with and 10 participants without M. tuberculosis complex DNA detected in stool by real-time quantitative PCR), and an independent German validation cohort of participants with culture-confirmed tuberculosis (n = 21). RESULTS: The tNGS assay detected M. tuberculosis complex DNA in 38 of 56 (68%) samples; for 28 of 38 (74%) samples, a full M. tuberculosis complex drug resistance prediction report was obtained. There was a high degree of concordance with sputum phenotypic drug susceptibility results (κ = 0.82). The ability to predict resistance was concentration-dependent and successful in 7/10 (70%), 18/25 (72%), and 3/21 (14%) of samples with stool PCR concentration thresholds of > 100 femtogram per microliter (fg/µl), 1 to 100 fg/µl, and < 1 fg/µl, respectively (p = 0.0004). The German cohort confirmed these results and demonstrated a similarly high concordance between stool tNGS and sputum phenotypic drug susceptibility results (κ = 0.84). CONCLUSIONS: tNGS can identify drug resistance from stool provided by tuberculosis patients. This affords the opportunity to obtain critical diagnostic information for tuberculosis patients who struggle to provide respiratory specimens.

Epidemiology and Prevalence of Oral Candidiasis in HIV Patients From Chad in the Post-HAART Era.

Oral candidiasis remains a common problem in HIV-infected individuals, especially in sub-Saharan Africa. Here, we performed the first study in Chad on the prevalence of oral yeasts carriage and oral candidiasis in HIV-positive subjects from southern Chad and analyzed the influence of HAART, CD4+ T-cell numbers, and antimycotics in 589 patients. These patients were recruited from a specialized medical center for HIV patients in Sarh and from a rural medical health dispensary in the vicinity, including a total of 384 HIV-positive and 205 HIV-negative individuals. Yeasts obtained from oral specimen were identified by MALDI-TOF MS and their antifungal susceptibility profiles determined. The overall prevalence of yeast colonization and symptomatic oral candidiasis in HIV-infected patients was 25.1%. The prevalence of oral candidiasis was higher in untreated than in HAART-treated HIV-positive patients (16% vs. 2%; p < 0.01). Oral candidiasis was furthermore associated with high fungal burdens of Candida albicans and a CD4+ T-cell number <200/µl. A shift toward non-albicans Candida species was observed under nucleoside-based HAART therapy. Azole antifungal drug resistance was only observed for the intrinsically resistant species Candida krusei and Candida glabrata. Prevalence of oral candidiasis in the studied area was very low. The species distribution was similar to other countries around the world, with C. albicans being dominant. Candida dubliniensis was not isolated. Nucleoside-based HAART therapy significantly reduced oral colonization as well as occurrence of oral candidiasis caused by C. albicans and led to a species shift toward non-albicans species. Antifungal resistance was not yet a concern in Chad.

Headspace analyses using multi-capillary column-ion mobility spectrometry allow rapid pathogen differentiation in hospital-acquired pneumonia relevant bacteria.

BACKGROUND: Hospital-acquired pneumonia (HAP) is a common problem in intensive care medicine and the patient outcome depends on the fast beginning of adequate antibiotic therapy. Until today pathogen identification is performed using conventional microbiological methods with turnaround times of at least 24 h for the first results. It was the aim of this study to investigate the potential of headspace analyses detecting bacterial species-specific patterns of volatile organic compounds (VOCs) for the rapid differentiation of HAP-relevant bacteria. METHODS: Eleven HAP-relevant bacteria (Acinetobacter baumanii, Acinetobacter pittii, Citrobacter freundii, Enterobacter cloacae, Escherichia coli, Klebsiella oxytoca, Klebsiella pneumoniae, Pseudomonas aeruginosa, Proteus mirabilis, Staphylococcus aureus, Serratia marcescens) were each grown for 6 hours in Lysogeny Broth and the headspace over the grown cultures was investigated using multi-capillary column-ion mobility spectrometry (MCC-IMS) to detect differences in the VOC composition between the bacteria in the panel. Peak areas with changing signal intensities were statistically analysed, including significance testing using one-way ANOVA or Kruskal-Wallis test (p < 0.05). RESULTS: 30 VOC signals (23 in the positive ion mode and 7 in the negative ion mode of the MCC-IMS) showed statistically significant differences in at least one of the investigated bacteria. The VOC patterns of the bacteria within the HAP panel differed substantially and allowed species differentiation. CONCLUSIONS: MCC-IMS headspace analyses allow differentiation of bacteria within HAP-relevant panel after 6 h of incubation in a complex fluid growth medium. The method has the potential to be developed towards a feasible point-of-care diagnostic tool for pathogen differentiation on HAP.

Tula Virus as Causative Agent of Hantavirus Disease in Immunocompetent Person, Germany.

We report molecular evidence of Tula virus infection in an immunocompetent patient from Germany who had typical signs of hantavirus disease. Accumulating evidence indicates that Tula virus infection, although often considered nonpathogenic, represents a threat to human health.

Two Pandemics, One Challenge-Leveraging Molecular Test Capacity of Tuberculosis Laboratories for Rapid COVID-19 Case-Finding.

In many settings, the ongoing coronavirus disease (COVID-19) pandemic coincides with other major public health threats, in particular tuberculosis. Using tuberculosis (TB) molecular diagnostic infrastructure, which has substantially expanded worldwide in recent years, for COVID-19 case-finding might be warranted. We analyze the potential of using TB diagnostic and research infrastructures for severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) testing. We focused on quality control by adapting the 12 Quality System Essentials framework to the COVID-19 and TB context. We conclude that diagnostic infrastructures for TB can in principle be leveraged to scale-up SARS-CoV-2 testing, in particular in resource-poor settings. TB research infrastructures also can support sequencing of SARS-CoV-2 to study virus evolution and diversity globally. However, fundamental principles of quality management must be followed for both TB and SARS-CoV-2 testing to ensure valid results and to minimize biosafety hazards, and the continuity of TB diagnostic services must be guaranteed at all times.

The Rnf complex is a Na+ coupled respiratory enzyme in a fermenting bacterium, Thermotoga maritima.

rnf genes are widespread in bacteria and biochemical and genetic data are in line with the hypothesis that they encode a membrane-bound enzyme that oxidizes reduced ferredoxin and reduces NAD and vice versa, coupled to ion transport across the cytoplasmic membrane. The Rnf complex is of critical importance in many bacteria for energy conservation but also for reverse electron transport to drive ferredoxin reduction. However, the enzyme has never been purified and thus, ion transport could not be demonstrated yet. Here, we have purified the Rnf complex from the anaerobic, fermenting thermophilic bacterium Thermotoga maritima and show that is a primary Na+ pump. These studies provide the proof that the Rnf complex is indeed an ion (Na+) translocating, respiratory enzyme. Together with a Na+-F1FO ATP synthase it builds a simple, two-limb respiratory chain in T. maritima. The physiological role of electron transport phosphorylation in a fermenting bacterium is discussed.

The Rnf complex from the acetogenic bacterium Acetobacterium woodii: Purification and characterization of RnfC and RnfB.

rnf genes are widespread in anaerobic bacteria and hypothesized to encode a respiratory enzyme that couples exergonic reduction of NAD with reduced ferredoxin as a reductant to vectorial ion (Na+, H+) translocation across the cytoplasmic membrane. However, despite its importance for the physiology of these bacteria, little is known about the subunit composition and the function of subunits. Here, we have purified the entire Rnf complex from the acetogen Acetobacterium woodii or after its production in Escherichia coli. These studies revealed covalently bound flavin in RnfB and RnfD. Unfortunately, the complex did not catalyze electron transfer from reduced ferredoxin to NAD. We, therefore, concentrated on the two cytosolic subunits RnfC and RnfB. RnfC was produced in E. coli, purified and shown to have 8.3 mol iron and 8.6 mol sulfur per mol of the subunit, consistent with the presence of two [4Fe-4S] centers, which were verified by EPR analysis. Flavins could not be detected, but RnfC catalyzed NADH-dependent FMN reduction. These data confirm RnfC as NADH-binding subunit and FMN as an intermediate in the electron transport chain. RnfB could only be produced as a fusion to the maltose-binding protein. It contained 25 mol iron and 26 mol sulfur, consistent with the predicted six [4Fe4S] centers. The FeS centers in RnfB were reduced with reduced ferredoxin as reductant. These data are consistent with RnfB as the ferredoxin-binding subunit of the complex.

Importation of Human Seoul Virus Infection to Germany from Indonesia.

Seoul hantavirus-associated hemorrhagic fever with renal syndrome cases are rare outside Asia and have not yet been found in Germany. We report clinical and molecular evidence for a Seoul virus infection in a patient in Germany. The infection was most likely acquired during a stay in Sulawesi, Indonesia.