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Infection and Immunity Jul 2023Macrophage (MΦ) infection models are important tools for studying host-mycobacterial interactions. Although the multiplicity of infection (MOI) is an important...
Macrophage (MΦ) infection models are important tools for studying host-mycobacterial interactions. Although the multiplicity of infection (MOI) is an important experimental variable, the selection of MOI in mycobacterial infection experiments is largely empirical, without reference to solid experimental data. To provide relevant data, we used RNA-seq to analyze the gene expression profiles of MΦs 4 or 24 h after infection with Mycobacterium marinum () at MOIs ranging from 0.1 to 50. Analysis of differentially expressed genes (DEGs) showed that different MOIs are linked to distinct transcriptomic changes and only 10% of DEGs were shared by MΦ infected at all MOIs. KEGG pathway enrichment analysis revealed that type I interferon (IFN)-related pathways were inoculant dose-dependent and enriched only at high MOIs, whereas TNF pathways were inoculant dose-independent and enriched at all MOIs. Protein-protein interaction (PPI) network alignment showed that different MOIs had distinct key node genes. By fluorescence-activated cell sorting and follow-up RT-PCR analysis, we could separate infected MΦs from uninfected MΦs and found phagocytosis of mycobacteria to be the determinant factor for type I IFN production. The distinct transcriptional regulation of RAW264.7 MΦ genes at different MOIs was also seen with Mycobacterium tuberculosis () infections and primary MΦ infection models. In summary, transcriptional profiling of mycobacterial infected MΦs revealed that different MOIs activate distinct immune pathways and the type I IFN pathway is activated only at high MOIs. This study should provide guidance for selecting the MOI most appropriate for different research questions.
Topics: Transcriptome; Signal Transduction; Macrophages; Mycobacterium tuberculosis; Interferon Type I
PubMed: 37338365
DOI: 10.1128/iai.00155-23 -
Clinical, Cosmetic and Investigational... 2023is an atypical bacterium, and skin infections caused by it are relatively rare, usually occurring in workers engaged in seafood processing and housewives who clean and...
is an atypical bacterium, and skin infections caused by it are relatively rare, usually occurring in workers engaged in seafood processing and housewives who clean and prepare fish for consumption. The infection often occurs after the skin is punctured by fish scales, spines, etc. The JAK/STAT signaling pathway is closely related to the human immune response to infections. Therefore, JAK inhibitors may induce and exacerbate various infections in clinical practice. This article reports a case of skin infection in the left upper limb of a female patient with chronic idiopathic myelofibrosis during treatment with ruxolitinib. The patient denied being punctured or scratched by fish scales or spines. Clinical manifestations included multiple infiltrative erythemas and subcutaneous nodules in the thumb and forearm. Histopathological examination showed infiltration of mixed acute and chronic inflammatory cells in the subcutaneous tissue. The diagnosis was ultimately confirmed by NGS sequencing. The patient was cured after taking moxifloxacin and clarithromycin for 10 months. Infection is a common adverse reaction of JAK inhibitors, but no literature has reported on skin infections occurring during JAK inhibitor treatment, which is relatively rare. As the clinical application of JAK inhibitors becomes more widespread, the skin infections they cause may present in various forms and require the attention of clinicians.
PubMed: 37333514
DOI: 10.2147/CCID.S413592 -
Microbiology Spectrum Aug 2023Among the numerous pathogenic nontuberculous mycobacteria (NTM), which may cause disease in both poikilothermic and homoeothermic organisms, members of the unique clade...
Among the numerous pathogenic nontuberculous mycobacteria (NTM), which may cause disease in both poikilothermic and homoeothermic organisms, members of the unique clade Mycobacterium ulcerans/Mycobacterium marinum (MuMC) may cause disease in both fish and humans. Here, we describe the emergence of Mycobacterium pseudoshottsii, one of the four MuMC members, in Israel. For many years, M. marinum was the dominant NTM that was diagnosed in Israel as a fish pathogen. To the best of our knowledge, this is the first isolation and genomic characterization of infecting edible fish from two different fish species farmed in offshore sea cages in the eastern Mediterranean as well as in a recirculating aquaculture system in Israel. We compared the whole-genome sequences to all available genomic sequences of MuMC in free, publicly accessible databases. Mycobacterium pseudoshottsii was first detected in 1997 in the USA, infecting wild striped bass (Morone saxatilis). Since then, several reports from different countries worldwide have shown its capacity to become established in new regions as well as its pathogenicity to saltwater and euryhaline finfish of different genera. Our phylogenetic analysis revealed that the Mycobacterium ulcerans/Mycobacterium marinum clade (MuMC) is divided into two main branches: one that includes M. marinum and , and the second, which includes other M. marinum isolates as well as two isolates of M. shottsii. Our results reinforce the proposition that the geographical distribution of is much more extensive than is commonly believed. The emergence of in different parts of the world and its pathogenic traits that affect finfish of different genera may be a cause for concern among fish farmers, researchers, and environmental organizations.
Topics: Humans; Animals; Phylogeny; Mycobacterium; Bass; Phenotype; Mycobacterium marinum; Mycobacterium Infections, Nontuberculous; Fish Diseases
PubMed: 37272844
DOI: 10.1128/spectrum.00856-23 -
Pathogens (Basel, Switzerland) Apr 2023Caused by the intracellular pathogen (Mtb), tuberculosis (TB) remains a massive global public health issue. A well-known and key TB trait is caseous necrotic granuloma,...
Caused by the intracellular pathogen (Mtb), tuberculosis (TB) remains a massive global public health issue. A well-known and key TB trait is caseous necrotic granuloma, which allows mycobacteria to reactivate and disseminate, thus confounding TB eradication programs. Amino acid (AA) metabolism is key to regulating immune responses in Mtb infections; however, it is currently unclear if AAs can be used to treat tuberculous granulomas. Here, we screened 20 proteinogenic AAs using a -infected zebrafish granuloma model. Only L-tyrosine simultaneously reduced (. ) levels in zebrafish larvae and adults and inhibited intracellular pathogen survival levels. Mechanistically, L-tyrosine significantly upregulated interferon-γ (IFN-γ) expression in . -infected zebrafish adults but not in larvae. Using N-acetylcysteine (NAC) to inhibit reactive oxygen species (ROS), L-tyrosine appeared to inhibit Mtb intracellular survival by promoting ROS production. Thus, L-tyrosine as a non-essential AA may reduce mycobacterial survival in both macrophages and tuberculous granulomas. Our research provides a platform for the clinical development of AAs for active or latent TB patients infected with drug-sensitive or drug-resistant Mtb.
PubMed: 37242324
DOI: 10.3390/pathogens12050654 -
Microbiology Spectrum Jun 2023Mycobacterium ulcerans, an environmental opportunistic pathogen, causes necrotic cutaneous and subcutaneous lesions, named Buruli ulcers, in tropical countries....
Mycobacterium ulcerans, an environmental opportunistic pathogen, causes necrotic cutaneous and subcutaneous lesions, named Buruli ulcers, in tropical countries. PCR-derived tests used to detect M. ulcerans in environmental and clinical samples do not allow one-shot detection, identification, and typing of M. ulcerans among closely related Mycobacterium marinum complex mycobacteria. We established a 385-member M. marinum/M. ulcerans complex whole-genome sequence database by assembling and annotating 341 M. marinum/M. ulcerans complex genomes and added 44 M. marinum/M. ulcerans complex whole-genome sequences already deposited in the NCBI database. Pangenome, core genome, and single-nucleotide polymorphism (SNP) distance-based comparisons sorted the 385 strains into 10 M. ulcerans taxa and 13 M. marinum taxa, correlating with the geographic origin of strains. Aligning conserved genes identified one (proline-proline-glutamate) gene sequence to be species and intraspecies specific, thereby genotyping the 23 M. marinum/M. ulcerans complex taxa. PCR sequencing of the gene correctly genotyped nine M. marinum/M. ulcerans complex isolates among one M. marinum taxon and three M. ulcerans taxa in the African taxon (T2.4). Further, successful gene PCR sequencing in 15/21 (71.4%) swabs collected from suspected Buruli ulcer lesions in Côte d'Ivoire exhibited positive M. ulcerans 2404 real-time PCR and identified the M. ulcerans T2.4.1 genotype in eight swabs and M. ulcerans T2.4.1/T2.4.2 mixed genotypes in seven swabs. gene sequencing could be used as a proxy for whole-genome sequencing for the one-shot detection, identification, and typing of clinical M. ulcerans strains, offering an unprecedented tool for identifying M. ulcerans mixed infections. We describe a new targeted sequencing approach that characterizes the gene to disclose the simultaneous presence of different variants of a single pathogenic microorganism. This approach has direct implications on the understanding of pathogen diversity and natural history and potential therapeutic implications when dealing with obligate and opportunistic pathogens, such as Mycobacterium ulcerans presented here as a prototype.
Topics: Humans; Buruli Ulcer; Mycobacterium ulcerans; Cote d'Ivoire; Real-Time Polymerase Chain Reaction; Personal Protective Equipment
PubMed: 37222600
DOI: 10.1128/spectrum.00342-23 -
Emerging Infectious Diseases Jun 2023Infections after reptile bites are uncommon, and microbial etiologies are not well defined. We describe a case of Mycobacterium marinum soft-tissue infection after an...
Infections after reptile bites are uncommon, and microbial etiologies are not well defined. We describe a case of Mycobacterium marinum soft-tissue infection after an iguana bite in Costa Rica that was diagnosed through 16S rRNA sequencing and mycobacterial culture. This case informs providers of potential etiologies of infection after iguana bites.
Topics: Animals; Humans; Costa Rica; Iguanas; Mycobacterium Infections, Nontuberculous; RNA, Ribosomal, 16S; Bites and Stings
PubMed: 37209698
DOI: 10.3201/eid2906.230062 -
Molecular epidemiology of nontuberculous mycobacteria isolated from tuberculosis-suspected patients.AMB Express May 2023It is a growing problem around the world to deal with nontuberculous mycobacteria infection (NTM), but its clinical significance is still largely unknown. This study...
It is a growing problem around the world to deal with nontuberculous mycobacteria infection (NTM), but its clinical significance is still largely unknown. This study aims to investigate the epidemiology of NTM infections from various clinical samples and determine their clinical significance. From December 2020 to December 2021, 6125 clinical samples were collected. In addition to phenotypic detection, genotypic detection through multilocus sequence typing (hsp65, rpoB, and 16S rDNA genes) and sequencing was also conducted. Records of patients were consulted for clinical information, such as symptoms and radiological findings. Of the 6,125 patients, 351 (5.7%) were positive for acid-fast bacteria (AFB). Out of 351 AFB, 289 (82.3%) and 62 (17.7%) subjects were identified as M. tuberculosis complex (MTC) and NTM strains, respectively. Isolates of Mycobacterium simiae and M. fortuitum were the most frequent, followed by isolates of M. kansasii and M. marinum. We also isolated M. chelonae, M. canariasense, and M. jacuzzii, which are rarely reported. Symptoms (P = 0.048), radiographic findings (P = 0.013), and gender (P = 0.039) were associated with NTM isolates. M. Fortuitum, M. simiae, and M. kansasii presented with bronchiectasis, infiltration, and cavitary lesions most frequently, while cough was the most common symptom. In conclusion, Mycobacterium simiae and M. fortuitum were presented in seventeen and twelve NTM isolates from the collected samples. There is evidence that NTM infections in endemic settings may contribute to the dissemination of various diseases and the control of tuberculosis. In spite of this, further research is needed to evaluate the clinical significance of NTM isolates.
PubMed: 37202495
DOI: 10.1186/s13568-023-01557-4 -
PLoS Pathogens May 2023Infections caused by members of the mycobacterium tuberculosis complex [MTC] and nontuberculous mycobacteria [NTM] can induce widespread morbidity and mortality in...
Infections caused by members of the mycobacterium tuberculosis complex [MTC] and nontuberculous mycobacteria [NTM] can induce widespread morbidity and mortality in people. Mycobacterial infections cause both a delayed immune response, which limits rate of bacterial clearance, and formation of granulomas, which contain bacterial spread, but also contribute to lung damage, fibrosis, and morbidity. Granulomas also limit access of antibiotics to bacteria, which may facilitate development of resistance. Bacteria resistant to some or all antibiotics cause significant morbidity and mortality, and newly developed antibiotics readily engender resistance, highlighting the need for new therapeutic approaches. Imatinib mesylate, a cancer drug used to treat chronic myelogenous leukemia [CML] that targets Abl and related tyrosine kinases, is a possible host-directed therapeutic [HDT] for mycobacterial infections, including those causing TB. Here, we use the murine Mycobacterium marinum [Mm] infection model, which induces granulomatous tail lesions. Based on histological measurements, imatinib reduces both lesion size and inflammation of surrounding tissue. Transcriptomic analysis of tail lesions indicates that imatinib induces gene signatures indicative of immune activation and regulation at early time points post infection that resemble those seen at later ones, suggesting that imatinib accelerates but does not substantially alter anti-mycobacterial immune responses. Imatinib likewise induces signatures associated with cell death and promotes survival of bone marrow-derived macrophages [BMDMs] in culture following infection with Mm. Notably, the capacity of imatinib to limit formation and growth of granulomas in vivo and to promote survival of BMDMs in vitro depends upon caspase 8, a key regulator of cell survival and death. These data provide evidence for the utility of imatinib as an HDT for mycobacterial infections in accelerating and regulating immune responses, and limiting pathology associated with granulomas, which may mitigate post-treatment morbidity.
Topics: Humans; Animals; Mice; Imatinib Mesylate; Pyrimidines; Piperazines; Benzamides; Anti-Bacterial Agents; Granuloma
PubMed: 37200402
DOI: 10.1371/journal.ppat.1011387 -
MSystems Jun 2023The antibiotic-tolerant biofilms present in tuberculous granulomas add an additional layer of complexity when treating mycobacterial infections, including tuberculosis...
The antibiotic-tolerant biofilms present in tuberculous granulomas add an additional layer of complexity when treating mycobacterial infections, including tuberculosis (TB). For a more efficient treatment of TB, the biofilm forms of mycobacteria warrant specific attention. Here, we used (Mmr) as a biofilm-forming model to identify the abundant proteins covering the biofilm surface. We used biotinylation/streptavidin-based proteomics on the proteins exposed at the Mmr biofilm matrices to identify 448 proteins and proteomics to detect 91 Mmr proteins from the mycobacterial granulomas isolated from adult zebrafish. and proteomics data are available via ProteomeXchange with identifiers PXD033425 and PXD039416, respectively. Data comparisons pinpointed the molecular chaperone GroEL2 as the most abundant Mmr protein within the and proteomes, while its paralog, GroEL1, with a known role in biofilm formation, was detected with slightly lower intensity values. To validate the surface exposure of these targets, we created in-house synthetic nanobodies (sybodies) against the two chaperones and identified sybodies that bind the mycobacterial biofilms and those present in granulomas. Taken together, the present study reports a proof-of-concept showing that surface proteomics and proteomics combined is a valuable strategy to identify surface-exposed proteins on the mycobacterial biofilm. Biofilm surface-binding nanobodies could be eventually used as homing agents to deliver biofilm-targeting treatments to the sites of persistent biofilm infection. IMPORTANCE With the currently available antibiotics, the treatment of TB takes months. The slow response to treatment is caused by antibiotic tolerance, which is especially common among bacteria that form biofilms. Such biofilms are composed of bacterial cells surrounded by the extracellular matrix. Both the matrix and the dormant lifestyle of the bacterial cells are thought to hinder the efficacy of antibiotics. To be able to develop faster-acting treatments against TB, the biofilm forms of mycobacteria deserve specific attention. In this work, we characterize the protein composition of Mmr biofilms in bacterial cultures and in mycobacteria extracted from infected adult zebrafish. We identify abundant surface-exposed targets and develop the first sybodies that bind to mycobacterial biofilms. As nanobodies can be linked to other therapeutic compounds, in the future, they can provide means to target therapies to biofilms.
Topics: Animals; Proteomics; Mycobacterium marinum; Zebrafish; Single-Domain Antibodies; Anti-Bacterial Agents; Tuberculosis; Biofilms
PubMed: 37184670
DOI: 10.1128/msystems.01073-22 -
Microbiology Resource Announcements Jun 2023We report the complete genome sequence of a Mycobacterium marinum strain, which was isolated from skin tissue of a wound infection. This strain was subjected to short-...
We report the complete genome sequence of a Mycobacterium marinum strain, which was isolated from skin tissue of a wound infection. This strain was subjected to short- and long-read sequencing. Its complete genome contains a single 6,393,703-bp circular chromosome. Phylogenomic analysis of all M. marinum genomes assigned this strain to cluster I.
PubMed: 37125927
DOI: 10.1128/mra.00174-23