-
Microbiology (Reading, England) Oct 2023Bacteria use population heterogeneity, the presence of more than one phenotypic variant in a clonal population, to endure diverse environmental challenges - a...
Bacteria use population heterogeneity, the presence of more than one phenotypic variant in a clonal population, to endure diverse environmental challenges - a 'bet-hedging' strategy. Phenotypic variants have been described in many bacteria, but the phenomenon is not well-understood in mycobacteria, including the environmental factors that influence heterogeneity. Here, we describe three reproducible morphological variants in - smooth, rough, and an intermediate morphotype that predominated under typical laboratory conditions. has two recognized morphotypes, smooth and rough. Interestingly, exists in only a rough form. The shift from smooth to rough in both and was observed over time in extended static culture, however the frequency of the rough morphotype was high in pellicle preparations compared to planktonic culture, suggesting a role for an aggregated microenvironment in the shift to the rough form. Differences in growth rate, biofilm formation, cell wall composition, and drug tolerance were noted among and variants. Deletion of the global regulator shifted the intermediate morphotype to a smooth form but did not fully phenocopy the naturally generated smooth morphotype, indicating Lsr2 is likely downstream of the initiating regulatory cascade that controls these morphotypes. Rough forms typically correlate with higher invasiveness and worse outcomes during infection and our findings indicate the shift to this rough form is promoted by aggregation. Our findings suggest that mycobacterial population heterogeneity, reflected in colony morphotypes, is a reproducible, programmed phenomenon that plays a role in adaptation to unique environments and this heterogeneity may influence infection progression and response to treatment.
Topics: Humans; Mycobacterium abscessus; Mycobacterium smegmatis; Mycobacterium Infections, Nontuberculous; Mycobacterium
PubMed: 37862100
DOI: 10.1099/mic.0.001402 -
Nature Microbiology Sep 2023Mycobacteriophages show promise as therapeutic agents for non-tuberculous mycobacterium infections. However, little is known about phage recognition of Mycobacterium...
Mycobacteriophages show promise as therapeutic agents for non-tuberculous mycobacterium infections. However, little is known about phage recognition of Mycobacterium cell surfaces or mechanisms of phage resistance. We show here that trehalose polyphleates (TPPs)-high-molecular-weight, surface-exposed glycolipids found in some mycobacterial species-are required for infection of Mycobacterium abscessus and Mycobacterium smegmatis by clinically useful phages BPs and Muddy. TPP loss leads to defects in adsorption and infection and confers resistance. Transposon mutagenesis shows that TPP disruption is the primary mechanism for phage resistance. Spontaneous phage resistance occurs through TPP loss by mutation, and some M. abscessus clinical isolates are naturally phage-insensitive due to TPP synthesis gene mutations. Both BPs and Muddy become TPP-independent through single amino acid substitutions in their tail spike proteins, and M. abscessus mutants resistant to TPP-independent phages reveal additional resistance mechanisms. Clinical use of BPs and Muddy TPP-independent mutants should preempt phage resistance caused by TPP loss.
Topics: Mycobacteriophages; Trehalose; Bacteriophages; Amino Acid Substitution; Cell Membrane
PubMed: 37644325
DOI: 10.1038/s41564-023-01451-6 -
Human Vaccines & Immunotherapeutics Dec 2023The immune escape mutations of SARS-CoV-2 variants emerged frequently, posing a new challenge to weaken the protective efficacy of current vaccines. Thus, the...
The immune escape mutations of SARS-CoV-2 variants emerged frequently, posing a new challenge to weaken the protective efficacy of current vaccines. Thus, the development of novel SARS-CoV-2 vaccines is of great significance for future epidemic prevention and control. We herein reported constructing the attenuated () as a bacterial surface display system to carry the spike (S) and nucleocapsid (N) of SARS-CoV-2. To mimic the native localization on the surface of viral particles, the S or N antigen was fused with truncated PE_PGRS33 protein, which is a transportation component onto the cell wall of (). The sub-cellular fraction analysis demonstrated that S or N protein was exactly expressed onto the surface (cell wall) of the recombinant . After the immunization of the -based COVID-19 vaccine candidate in mice, S or N antigen-specific T cell immune responses were effectively elicited, and the subsets of central memory CD4+ T cells and CD8+ T cells were significantly induced. Further analysis showed that there were some potential cross-reactive CTL epitopes between SARS-CoV-2 and . Overall, our data provided insights that -based bacterial surface display system could be a suitable vector for developing T cell-based vaccines against SARS-CoV-2 and other infectious diseases.
Topics: Mice; Humans; Animals; Mycobacterium smegmatis; COVID-19 Vaccines; COVID-19; SARS-CoV-2; Spike Glycoprotein, Coronavirus
PubMed: 36785935
DOI: 10.1080/21645515.2023.2171233 -
ELife Aug 2023A DNA damage-inducible mutagenic gene cassette has been implicated in the emergence of drug resistance in during anti-tuberculosis (TB) chemotherapy. However, the...
A DNA damage-inducible mutagenic gene cassette has been implicated in the emergence of drug resistance in during anti-tuberculosis (TB) chemotherapy. However, the molecular composition and operation of the encoded 'mycobacterial mutasome' - minimally comprising DnaE2 polymerase and ImuA' and ImuB accessory proteins - remain elusive. Following exposure of mycobacteria to DNA damaging agents, we observe that DnaE2 and ImuB co-localize with the DNA polymerase III β subunit (β clamp) in distinct intracellular foci. Notably, genetic inactivation of the mutasome in an mutant containing a disrupted β clamp-binding motif abolishes ImuB-β clamp focus formation, a phenotype recapitulated pharmacologically by treating bacilli with griselimycin and in biochemical assays in which this β clamp-binding antibiotic collapses pre-formed ImuB-β clamp complexes. These observations establish the essentiality of the ImuB-β clamp interaction for mutagenic DNA repair in mycobacteria, identifying the mutasome as target for adjunctive therapeutics designed to protect anti-TB drugs against emerging resistance.
Topics: Bacterial Proteins; Mycobacterium tuberculosis; Mutagenesis; DNA Repair; Antitubercular Agents
PubMed: 37530405
DOI: 10.7554/eLife.75628 -
Frontiers in Cellular and Infection... 2023Mycobacteriophages are viruses that infect members of genus . Because of the rise in antibiotic resistance in mycobacterial diseases such as tuberculosis,...
Mycobacteriophages are viruses that infect members of genus . Because of the rise in antibiotic resistance in mycobacterial diseases such as tuberculosis, mycobacteriophages have received renewed attention as alternative therapeutic agents. Mycobacteriophages are highly diverse, and, on the basis of their genome sequences, they are grouped into 30 clusters and 10 singletons. In this article, we have described the isolation and characterization of a novel mycobacteriophage Kashi-VT1 (KVT1) infecting mc 155 () and isolated from Varanasi, India. KVT1 is a cluster K1 temperate phage that belongs to family as visualized in transmission electron microscopy. The phage genome is 61,010 base pairs with 66.5% Guanine/Cytosine (GC) content, encoding 101 putative open reading frames. The KVT1 genome encodes an immunity repressor, a tyrosine integrase, and an excise protein, which are the characteristics of temperate phages. It also contains genes encoding holin, lysin A, and lysin B involved in host cell lysis. The one-step growth curve demonstrated that KVT1 has a latency time of 90 min and an average burst size of 101 phage particles per infected cell. It can withstand a temperature of up to 45°C and has a maximum viability between pH 8 and 9. Some mycobacteriophages from cluster K are known to infect the pathogenic (); hence, KVT1 holds potential for the phage therapy against tuberculosis, and it can also be engineered to convert into an exclusively lytic phage.
Topics: Humans; Mycobacteriophages; Genome, Viral; Mycobacterium tuberculosis; Mycobacterium smegmatis; Tuberculosis; Bacteriophages
PubMed: 37545854
DOI: 10.3389/fcimb.2023.1173894 -
Current Opinion in Microbiology Apr 2024Members of the order Mycobacteriales are distinguished by a characteristic diderm cell envelope, setting them apart from other Actinobacteria species. In addition to the... (Review)
Review
Members of the order Mycobacteriales are distinguished by a characteristic diderm cell envelope, setting them apart from other Actinobacteria species. In addition to the conventional peptidoglycan cell wall, these organisms feature an extra polysaccharide polymer composed of arabinose and galactose, termed arabinogalactan. The nonreducing ends of arabinose are covalently linked to mycolic acids (MAs), forming the immobile inner leaflet of the highly hydrophobic MA membrane. The contiguous outer leaflet of the MA membrane comprises trehalose mycolates and various lipid species. Similar to all actinobacteria, Mycobacteriales exhibit apical growth, facilitated by a polar localized elongasome complex. A septal cell envelope synthesis machinery, the divisome, builds instead of the cell wall structures during cytokinesis. In recent years, a growing body of knowledge has emerged regarding the cell wall synthesizing complexes of Mycobacteriales., focusing particularly on three model species: Corynebacterium glutamicum, Mycobacterium smegmatis, and Mycobacterium tuberculosis.
PubMed: 38653035
DOI: 10.1016/j.mib.2024.102478 -
Vaccines Jul 2023() is frequently used as an alternative model organism in ) studies. While containing high sequence homology with it is considered non-pathogenic in humans. As such it... (Review)
Review
() is frequently used as an alternative model organism in ) studies. While containing high sequence homology with it is considered non-pathogenic in humans. As such it has been used to study and other infections in vivo and more recently been explored for potential therapeutic applications. A body of previous research has highlighted the potential of using genetically modified displaying rapid growth and unique immunostimulatory characteristics as an effective vaccine vector. Novel systems biology techniques can further serve to optimize these delivery constructs. In this article, we review recent advancements in vaccinomics tools that support the efficacy of a based vaccine vector. Moreover, the integration of systems biology and molecular omics techniques in these pioneering studies heralds a potential accelerated pipeline for the development of next-generation recombinant vaccines against rapidly developing diseases.
PubMed: 37631870
DOI: 10.3390/vaccines11081302 -
Nature Communications Sep 2023Tuberculosis continues to pose a serious threat to global health. Mycobacterium tuberculosis, the causative agent of tuberculosis, is an intracellular pathogen that...
Tuberculosis continues to pose a serious threat to global health. Mycobacterium tuberculosis, the causative agent of tuberculosis, is an intracellular pathogen that relies on various mechanisms to survive and persist within the host. Among their many virulence factors, mycobacteria encode Mce systems. Some of these systems are implicated in lipid uptake, but the molecular basis for Mce function(s) is poorly understood. To gain insights into the composition and architecture of Mce systems, we characterized the putative Mce1 complex involved in fatty acid transport. We show that the Mce1 system in Mycobacterium smegmatis comprises a canonical ATP-binding cassette transporter associated with distinct heterohexameric assemblies of substrate-binding proteins. Furthermore, we establish that the conserved membrane protein Mce1N negatively regulates Mce1 function via a unique mechanism involving blocking transporter assembly. Our work offers a molecular understanding of Mce complexes, sheds light on mycobacterial lipid metabolism and its regulation, and informs future anti-mycobacterial strategies.
Topics: Membrane Proteins; Mycobacterium smegmatis; Mycobacterium tuberculosis; Membrane Transport Proteins; ATP-Binding Cassette Transporters
PubMed: 37736771
DOI: 10.1038/s41467-023-41578-y