From Antagonism to Enhancement: Triton X-100 Surfactant Affects Phenanthrene Interfacial Biodegradation by Mycobacteria through a Shift in Uptake Mechanisms.

Polycyclic aromatic hydrocarbons (PAHs), as persistent environmental pollutants, often reside in nonaqueous-phase liquids (NAPLs). Mycobacterium sp. WY10, boasting highly hydrophobic surfaces, can adsorb to the oil-water interface, stabilizing the Pickering emulsion and directly accessing PAHs for biodegradation. We investigated the impact of Triton X-100 (TX100) on this interfacial uptake of phenanthrene (PHE) by Mycobacteria, using n-tetradecane (TET) and bis-(2-ethylhexyl) phthalate (DEHP) as NAPLs. Interfacial tension, phase behavior, and emulsion stability studies, alongside confocal laser scanning microscopy and electron microscope observations, unveiled the intricate interplay. In surfactant-free systems, Mycobacteria formed stable W/O Pickering emulsions, directly degrading PHE within the NAPLs because of their intimate contact. Introducing low-dose TX100 disrupted this relationship. Preferentially binding to the cells, the surfactant drastically increased the cell hydrophobicity, triggering desorption from the interface and phase separation. Consequently, PAH degradation plummeted due to hindered NAPL access. Higher TX100 concentrations flipped the script, creating surfactant-stabilized O/W emulsions devoid of interfacial cells. Surprisingly, PAH degradation remained efficient. This paradox can be attributed to NAPL emulsification, driven by the surfactant, which enhanced mass transfer and brought the substrate closer to the cells, despite their absence at the interface. This study sheds light on the complex effect of surfactants on Mycobacteria and PAH uptake, revealing an antagonistic effect at low concentrations that ultimately leads to enhanced degradation through emulsification at higher doses. These findings offer valuable insights into optimizing bioremediation strategies in PAH-contaminated environments.

Heterotrophic bacteria isolated from a chloraminated system accelerate chloramine decay.

This work comprehensively demonstrates the ability of heterotrophic bacteria, isolated from a chloraminated system, to decay chloramine. This study non-selectively isolated 62 cultures of heterotrophic bacteria from a water sample (0.002 mg-N/L nitrite and 1.42 mg/L total chlorine) collected from a laboratory-scale reactor system; most of the isolates (93.3%) were Mycobacterium sp. Three species of Mycobacterium and one species of Micrococcus were inoculated to a basal inorganic medium with initial concentrations of acetate (from 0 to 24 mg-C/L) and 1.5 mg/L chloramine. Bacterial growth coincided with declines in the concentrations of chloramine, acetate, and ammonium. Detailed experiments with one of the Mycobacterium sp. isolates suggest that the common mechanism of chloramine loss is auto-decomposition likely mediated by chloramine-decaying proteins. The ability of the isolates to grow and decay chloramine underscores the important role of heterotrophic bacteria in the stability of chloramine in water-distribution systems. Existing strategies based on controlling nitrification should be augmented to include minimizing heterotrophic bacteria.

Effects of Graphene Oxide on Endophytic Bacteria Population Characteristics in Plants from Soils Contaminated by Polycyclic Aromatic Hydrocarbons.

Environmental pollution stands as one of the significant global challenges we face today. Polycyclic aromatic hydrocarbons (PAHs), a class of stubborn organic pollutants, have long been a focal point of bioremediation research. This study aims to explore the impact and mechanisms of graphene oxide (GO) on the phytoremediation effectiveness of PAHs. The results underscore the significant efficacy of GO in accelerating the degradation of PAHs. Additionally, the introduction of GO altered the diversity and community structure of endophytic bacteria within the roots, particularly those genera with potential for PAH degradation. Through LEfSe analysis and correlation studies, we identified specific symbiotic bacteria, such as Mycobacterium, Microbacterium, Flavobacterium, Sphingomonas, Devosia, Bacillus, and Streptomyces, which coexist and interact under the influence of GO, synergistically degrading PAHs. These bacteria may serve as key biological markers in the PAH degradation process. These findings provide new theoretical and practical foundations for the application of nanomaterials in plant-based remediation of polluted soils and showcase the immense potential of plant-microbe interactions in environmental restoration.

[A case of HIV combined with Mycobacterium celatum infection].

Here, we reported the diagnosis and treatment of a case of HIV infected person complicated by an extremely rare infection with Mycobacterium celatum. Due to the similarity of homologous sequence regions between Mycobacterium celatum and Mycobacterium tuberculosis complex, the identification of conventional Mycobacterium species was incorrect, which was corrected after first-generation 16S rRNA sequencing. This report aimed to improve the clinical understanding of Mycobacterium celatum infection and the level of differential diagnosis between non-tuberculous mycobacterial disease and tuberculosis.

Impact of the COVID-19 pandemic on the real-world diagnostic infrastructure for tuberculosis-An ESGMYC collaborative study.

We determined the impact of the COVID-19 pandemic on mycobacterial diagnostic services. 40 laboratories from 22 countries completed an online questionnaire covering the redeployment of the laboratory infrastructure and/or staff for SARS-CoV-2 testing, staff shortages and supply chain disruptions. 28 laboratories reported monthly numbers of samples processed for mycobacterial investigations and monthly numbers of M. tuberculosis complex (MTBC) PCRs performed between October 1st 2018 and October 31st 2020. More than half (23/40) of the participating TB laboratories reported having performed COVID-19 diagnostics in the early phase of the pandemic, in part with negative impact on the mycobacterial service activities. All participating laboratories reported shortages of consumables and laboratory equipment due to supply chain issues. Average monthly sample numbers decreased by 24% between January 2020 and October 2020 compared to pre-pandemic averages. At the end of the study period, most participating laboratories had not returned to pre-pandemic average MTBC PCR throughput.

Extracellular vesicles in mycobacteria: new findings in biogenesis, host-pathogen interactions, and diagnostics.

Since the discovery of extracellular vesicles (EVs) in mycobacterial species 15 years back, we have learned that this phenomenon is conserved in the Mycobacterium genus and has critical roles in bacterial physiology and host-pathogen interactions. Mycobacterium tuberculosis (Mtb), the tuberculosis (TB) causative agent, produces EVs both in vitro and in vivo including a diverse set of biomolecules with demonstrated immunomodulatory effects. Moreover, Mtb EVs (MEVs) have been shown to possess vaccine properties and carry biomarkers with diagnostic capacity. Although information on MEV biogenesis relative to other bacterial species is scarce, recent studies have shed light on how MEVs originate and are released to the extracellular space. In this minireview, we discuss past and new information about the vesiculogenesis phenomenon in Mtb, including biogenesis, MEV cargo, aspects in the context of host-pathogen interactions, and applications that could help to develop effective tools to tackle the disease.

Synchronization of Mycobacterium life cycle: A possible novel mechanism of antimycobacterial drug resistance evolution and its manipulation.

Mycobacterium Tuberculosis (Mtb) causing Tuberculosis (TB) is a widespread disease infecting millions of people worldwide. Additionally, emergence of drug resistant tuberculosis is a major challenge and concern in high TB burden countries. Most of the drug resistance in mycobacteria is attributed to developing acquired resistance due to spontaneous mutations or intrinsic resistance mechanisms. In this review, we emphasize on the role of bacterial cell cycle synchronization as one of the intrinsic mechanisms used by the bacteria to cope with stress response and perhaps involved in evolution of its drug resistance. The importance of cell cycle synchronization and its function in drug resistance in cancer cells, malarial and viral pathogens is well understood, but its role in bacterial pathogens has yet to be established. From the extensive literature survey, we could collect information regarding how mycobacteria use synchronization to overcome the stress response. Additionally, it has been observed that most of the microbial pathogens including mycobacteria are responsive to drugs predominantly in their logarithmic phase, while they show resistance to antibiotics when they are in the lag or stationary phase. Therefore, we speculate that Mtb might use this novel strategy wherein they regulate their cell cycle upon antibiotic pressure such that they either enter in their low metabolic phase i.e., either the lag or stationary phase to overcome the antibiotic pressure and function as persister cells. Thus, we propose that manipulating the mycobacterial drug resistance could be possible by fine-tuning its cell cycle.

Lipids Extracted from Mycobacterial Membrane and Enveloped PLGA Nanoparticles for Encapsulating Antibacterial Drugs Elicit Synergistic Antimicrobial Response against Mycobacteria.

Tuberculosis (TB) is a chronic disease caused byMycobacterium tuberculosis (Mtb), which shows a long treatment cycle often leads to drug resistance, making treatment more difficult. Immunogens present in the pathogen's cell membrane can stimulate endogenous immune responses. Therefore, an effective lipid-based vaccine or drug delivery vehicle formulated from the pathogen's cell membrane can improve treatment outcomes. Herein, we extracted and characterized lipids fromMycobacterium smegmatis, and the extracts contained lipids belonging to numerous lipid classes and compounds typically found associated with mycobacteria. The extracted lipids were used to formulate biomimetic lipid reconstituted nanoparticles (LrNs) and LrNs-coated poly(lactic-co-glycolic acid) nanoparticles (PLGA-LrNs). Physiochemical characterization and results of morphology suggested that PLGA-LrNs exhibited enhanced stability compared with LrNs. And both of these two types of nanoparticles inhibited the growth of M. smegmatis. After loading different drugs, PLGA-LrNs containing berberine or coptisine strongly and synergistically prevented the growth of M. smegmatis. Altogether, the bacterial membrane lipids we extracted with antibacterial activity can be used as nanocarrier coating for synergistic antibacterial treatment of M. smegmatis─an alternative model of Mtb, which is expected as a novel therapeutic system for TB treatment.

Disseminated mycobacterium genavense infection with central nervous system involvement in an HIV patient: a case report and literature review.

BACKGROUND: Immunodeficient patients, particularly HIV patients, are at risk of opportunistic infections. Nontuberculous mycobacteria can cause severe complications in immunodeficient patients. CASE PRESENTATION: We describe a 57-year-old HIV patient, primarily presented with coughs and constitutional symptoms, with a unique Mycobacterium genavense abdominal, pulmonary, and central nervous system infection, accompanied by intracranial masses. CONCLUSION: The diagnosis of NTM, including M. genavense, must always be considered by clinicians in immunodeficient patients, especially those with HIV, who have a compromised immune system.

Is the new tuberculous antigen-based skin test ready for use as an alternative to tuberculin skin test/interferon-gamma release assay for tuberculous diagnosis? A narrative review.

In recent years, novel specific Mycobacteria tuberculous (TB) antigen-based skin test (TBST) has become available for clinical use. The mechanism of TBST is similar to the interferon-gamma release assay (IGRA), making it a potential alternative for identifying latent tuberculous infection (LTBI), especially in subjects with history of bacille Calmette-Guérin vaccination. Three different commercial brands have been developed in Denmark, Russia, and China. Clinical studies in the respective countries have shown promising sensitivity, specificity, and safety profile. Some studies attempted to address the applicability of TBST in specific subject groups but the discrepancy in defining LTBI and problematic methodologies undermine the generalisation of the results to other communities across the world. Limited cost-effectiveness studies for TBST have been conducted without exploring the health economics for preventing development of LTBI into active TB. Unlike IGRA, no clinical studies have addressed the correlation of TBST results (magnitude of induration) with the likelihood of development of active TB. Moreover, the different TBSTs are not widely available for clinical use. While TBST is a promising test to overcome the shortcomings of tuberculin skin tests, more clinical data are needed to support its general application globally for the diagnosis of LTBI.

Beyond copper: examining the significance of His-mutations in mycobacterial GroEL1 HRCT for Ni(II) complex stability and formation.

Recently, we have studied the coordination chemistry of the Cu(II)-histidine-rich C-terminal tail (HRCT) complex of the mycobacterial GroEL1 protein. The structure of this domain differs significantly compared to the well-known methionine-glycine-rich GroEL chaperonin - it was predicted that mycobacterial GroEL1 could play a significant role in the metal homeostasis of Mycobacteria, especially copper. However, we found that this particular domain's pattern also repeats in a number of Ni(II)-binding proteins. Here, we present the studies concerning the properties of GroEL1 HRCT as a ligand for Ni(II) ions. For this purpose, we chose eight model peptides: L1 - Ac-DHDHHHGHAH, L2 - Ac-DKPAKAEDHDHHHGHAH, and 6 mutants of the latter in the pH range of 2-11. We examined the stoichiometry, stability, and spectroscopic features of copper complexes. We noticed that similar to the Cu(II)-complex, the presence of a Lys5 residue significantly increases the stability of the system. The impact of His mutations was also examined and carefully studied using NMR spectroscopy. His9 and His13 are the crucial residues for Ni(II) binding, whereas His12 has minimal relevance in complex formation.

Building size and disinfectant type influence the detection and concentration of Mycobacterium spp. in hot water plumbing.

The number of nontuberculous mycobacteria (NTM) lung disease cases is increasing in the United States (US). This respiratory disease is primarily caused by three NTM species: Mycobacterium avium, M. intracellulare, and M. abscessus. Since disease transmission could occur through water aerosolization, this study investigated these three species' occurrence (sporadic and persistent) in hot water samples collected from residences (n = 70) and office buildings (n = 30) across the US. A longitudinal survey design was used. Three quantitative Polymerase Chain Reaction (qPCR) assays were used to measure the mycobacterial species in the water samples. Additionally, the water's disinfectant residual was measured. A structure's age and square footage were evaluated to predict mycobacterial contamination. Also, the seasonal occurrence of each species was assessed by structure type. Residences had a 43 % (30/70), and office buildings had a 77 % (23/30) detection frequency of one or more Mycobacterium spp. in their hot water. The age of the structure influenced M. intracellulare detection frequency but not M. avium and M. abscessus. The structure's square footage affected M. avium and M. intracellulare detection frequency but not M. abscessus. In chlorinated water, M. intracellulare was detected 1.4× more often in office buildings' hot water than in chloraminated water. In chloraminated water, the Mycobacterium spp. were detected 2-2.5× more often in residences, while M. avium and M. abscessus were detected 1.5-2.3× more often in office buildings, compared to chlorinated water. Each Mycobacterium spp. had a different trend associated with the type of structure and disinfectant. Further research is needed to better understand NTM occurrence in the built environment to improve public health.

Evaluation of nucleotide MALDI-TOF-MS for the identification of Mycobacterium species.

Background: The accurate identification of the Mycobacterium tuberculosis complex (MTBC) and different nontuberculous mycobacteria (NTM) species is crucial for the timely diagnosis of NTM infections and for reducing poor prognoses. Nucleotide matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF-MS) has been extensively used for microbial identification with high accuracy and throughput. However, its efficacy for Mycobacterium species identification has been less studied. The objective of this study was to evaluate the performance of nucleotide MALDI-TOF-MS for Mycobacterium species identification. Methods: A total of 933 clinical Mycobacterium isolates were preliminarily identified as NTM by the MPB64 test. These isolates were identified by nucleotide MALDI-TOF-MS and Sanger sequencing. The performance of nucleotide MALDI-TOF MS for identifying various Mycobacterium species was analyzed based on Sanger sequencing as the gold standard. Results: The total correct detection rate of all 933 clinical Mycobacterium isolates using nucleotide MALDI-TOF-MS was 91.64% (855/933), and mixed infections were detected in 18.65% (174/933) of the samples. The correct detection rates for Mycobacterium intracellulare, Mycobacterium abscessus, Mycobacterium kansasii, Mycobacterium avium, MTBC, Mycobacterium gordonae, and Mycobacterium massiliense were 99.32% (585/589), 100% (86/86), 98.46% (64/65), 94.59% (35/37), 100.00% (34/34), 95.65% (22/23), and 100% (19/19), respectively. For the identification of the MTBC, M. intracellulare, M. abscessus, M. kansasii, M. avium, M. gordonae, and M. massiliense, nucleotide MALDI-TOF-MS and Sanger sequencing results were in good agreement (k > 0.7). Conclusion: In conclusion, nucleotide MALDI-TOF-MS is a promising approach for identifying MTBC and the most common clinical NTM species.

Newly Identified Mycobacterium africanum Lineage 10, Central Africa.

Analysis of genome sequencing data from >100,000 genomes of Mycobacterium tuberculosis complex using TB-Annotator software revealed a previously unknown lineage, proposed name L10, in central Africa. Phylogenetic reconstruction suggests L10 could represent a missing link in the evolutionary and geographic migration histories of M. africanum.

Soft tissue infection and osteomyelitis caused by Mycobacterium farcinogenes after heart surgery: Case report and literature review of human cases.

Mycobacterium farcinogenes (M. farcinogenes) is rapidly growing mycobacterium, belonging to non-tuberculous mycobacterial (NTM). M. farcinogenes is an exceedingly rare causative agent of human infection. Only seven cases with M. farcinogenes infections in humans were reported. This is a case of soft tissue infection and osteomyelitis caused by M. farcinogenes after heart surgery. Microbial identification was achieved by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF-MS). The clinical outcome was favorable after surgical debridement and 4-month antibiotics treatment. We also provide a comprehensive literature review on this disease.

Compartmentalization of galactan biosynthesis in mycobacteria.

Galactan polymer is a prominent component of the mycobacterial cell wall core. Its biogenesis starts at the cytoplasmic side of the plasma membrane by a build-up of the linker disaccharide [rhamnosyl (Rha) - N-acetyl-glucosaminyl (GlcNAc) phosphate] on the decaprenyl-phosphate carrier. This decaprenyl-P-P-GlcNAc-Rha intermediate is extended by two bifunctional galactosyl transferases, GlfT1 and GlfT2, and then it is translocated to the periplasmic space by an ABC transporter Wzm-Wzt. The cell wall core synthesis is finalized by the action of an array of arabinosyl transferases, mycolyl transferases, and ligases that catalyze an attachment of the arabinogalactan polymer to peptidoglycan through the linker region. Based on visualization of the GlfT2 enzyme fused with fluorescent tags it was proposed that galactan polymerization takes place in a specific compartment of the mycobacterial cell envelope, the intracellular membrane domain, representing pure plasma membrane free of cell wall components (previously denoted as the "PMf" domain), which localizes to the polar region of mycobacteria. In this work, we examined the activity of the galactan-producing cellular machine in the cell-wall containing cell envelope fraction and in the cell wall-free plasma membrane fraction prepared from Mycobacterium smegmatis by the enzyme assays using radioactively labeled substrate UDP-[14C]-galactose as a tracer. We found that despite a high abundance of GlfT2 in both of these fractions as confirmed by their thorough proteomic analyses, galactan is produced only in the reaction mixtures containing the cell wall components. Our findings open the discussion about the distribution of GlfT2 and the regulation of its activity in mycobacteria.

Identification, characterization and application of M16AT, a new organic solvent-tolerant (R)-enantio-selective type IV amine transaminase from Mycobacterium sp. ACS1612.

Biocatalysis has emerged as a powerful alternative to traditional chemical methods, especially for asymmetric synthesis. As biocatalysts usually exhibit excellent chemical, regio- and enantioselectivity, they facilitate and simplify many chemical processes for the production of a broad range of products. Here, a new biocatalyst called, R-selective amine transaminases (R-ATAs), was obtained from Mycobacterium sp. ACS1612 (M16AT) using in-silico prediction combined with a genome and protein database. A two-step simple purification process could yield a high concentration of pure enzyme, suggesting that industrial application would be inexpensive. Additionally, the newly identified and characterized R-ATAs displayed a broad substrate spectrum and strong tolerance to organic solvents. Moreover, the synthetic applicability of M16AT has been demonstrated by the asymmetric synthesis of (R)-fendiline from of (R)-1-phenylethan-1-amine.

Lsr2, a pleiotropic regulator at the core of the infectious strategy of Mycobacterium abscessus.

Mycobacterium abscessus is a non-tuberculous mycobacterium, causing lung infections in cystic fibrosis patients. During pulmonary infection, M. abscessus switches from smooth (Mabs-S) to rough (Mabs-R) morphotypes, the latter being hyper-virulent. Previously, we isolated the lsr2 gene as differentially expressed during S-to-R transition. lsr2 encodes a pleiotropic transcription factor that falls under the superfamily of nucleoid-associated proteins. Here, we used two functional genomic methods, RNA-seq and chromatin immunoprecipitation-sequencing (ChIP-seq), to elucidate the molecular role of Lsr2 in the pathobiology of M. abscessus. Transcriptomic analysis shows that Lsr2 differentially regulates gene expression across both morphotypes, most of which are involved in several key cellular processes of M. abscessus, including host adaptation and antibiotic resistance. These results were confirmed through quantitative real-time PCR, as well as by minimum inhibitory concentration tests and infection tests on macrophages in the presence of antibiotics. ChIP-seq analysis revealed that Lsr2 extensively binds the M. abscessus genome at AT-rich sequences and appears to form long domains that participate in the repression of its target genes. Unexpectedly, the genomic distribution of Lsr2 revealed no distinctions between Mabs-S and Mabs-R, implying more intricate mechanisms at play for achieving target selectivity.IMPORTANCELsr2 is a crucial transcription factor and chromosome organizer involved in intracellular growth and virulence in the smooth and rough morphotypes of Mycobacterium abscessus. Using RNA-seq and chromatin immunoprecipitation-sequencing (ChIP-seq), we investigated the molecular role of Lsr2 in gene expression regulation along with its distribution on M. abscessus genome. Our study demonstrates the pleiotropic regulatory role of Lsr2, regulating the expression of many genes coordinating essential cellular and molecular processes in both morphotypes. In addition, we have elucidated the role of Lsr2 in antibiotic resistance both in vitro and in vivo, where lsr2 mutant strains display heightened sensitivity to antibiotics. Through ChIP-seq, we reported the widespread distribution of Lsr2 on M. abscessus genome, revealing a direct repressive effect due to its extensive binding on promoters or coding sequences of its targets. This study unveils the significant regulatory role of Lsr2, intricately intertwined with its function in shaping the organization of the M. abscessus genome.