| 1. |
- Behra, Phani Rama Krishna, et al.
(author)
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Comparative genome analysis of Mycobacteria focusing on tRNA and non-coding RNA
- 2022
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In: BMC Genomics. - : BioMed Central (BMC). - 1471-2164. ; 23
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Journal article (peer-reviewed)abstract
- Background: The Mycobacterium genus encompasses at least 192 named species, many of which cause severe diseases such as tuberculosis. Non-tuberculosis mycobacteria (NTM) can also infect humans and animals. Some are of emerging concern because they show high resistance to commonly used antibiotics while others are used and evaluated in bioremediation or included in anticancer vaccines.Results: We provide the genome sequences for 114 mycobacterial type strains and together with 130 available mycobacterial genomes we generated a phylogenetic tree based on 387 core genes and supported by average nucleotide identity (ANI) data. The 244 genome sequences cover most of the species constituting the Mycobacterium genus. The genome sizes ranged from 3.2 to 8.1 Mb with an average of 5.7 Mb, and we identified 14 new plasmids. Moreover, mycobacterial genomes consisted of phage-like sequences ranging between 0 and 4.64% dependent on mycobacteria while the number of IS elements varied between 1 and 290. Our data also revealed that, depending on the mycobacteria, the number of tRNA and non-coding (nc) RNA genes differ and that their positions on the chromosome varied. We identified a conserved core set of 12 ncRNAs, 43 tRNAs and 18 aminoacyl-tRNA synthetases among mycobacteria.Conclusions; Phages, IS elements, tRNA and ncRNAs appear to have contributed to the evolution of the Mycobacterium genus where several tRNA and ncRNA genes have been horizontally transferred. On the basis of our phylogenetic analysis, we identified several isolates of unnamed species as new mycobacterial species or strains of known mycobacteria. The predicted number of coding sequences correlates with genome size while the number of tRNA, rRNA and ncRNA genes does not. Together these findings expand our insight into the evolution of the Mycobacterium genus and as such they establish a platform to understand mycobacterial pathogenicity, their evolution, antibiotic resistance/tolerance as well as the function and evolution of ncRNA among mycobacteria.
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| 2. |
- Behra, Phani Rama Krishna, et al.
(author)
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Insight into the biology of Mycobacterium mucogenicum and Mycobacterium neoaurum Glade members
- 2019
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In: Scientific Reports. - : NATURE PUBLISHING GROUP. - 2045-2322. ; 9
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Journal article (peer-reviewed)abstract
- Nontuberculous mycobacteria, NTM, are of growing concern and among these members of the Mycobacterium mucogenicum (Mmuc) and Mycobacterium neoaurum (Mneo) clades can cause infections in humans and they are resistant to first-line anti-tuberculosis drugs. They can be isolated from different ecological niches such as soil, tap water and ground water. Mycobacteria, such as Mmuc and Mneo, are classified as rapid growing mycobacteria, RGM, while the most familiar, Mycobacterium tuberculosis, belongs to the slow growing mycobacteria, SGM. Modern "omics" approaches have provided new insights into our understanding of the biology and evolution of this group of bacteria. Here we present comparative genomics data for seventeen NTM of which sixteen belong to the Mmuc- and Mneo-clades. Focusing on virulence genes, including genes encoding sigma/anti-sigma factors, serine threonine protein kinases (STPK), type VII (ESX genes) secretion systems and mammalian cell entry (Mce) factors we provide insight into their presence as well as phylogenetic relationship in the case of the sigma/anti-sigma factors and STPKs. Our data further suggest that these NTM lack ESX-5 and Mce2 genes, which are known to affect virulence. In this context, Mmuc- and Mneo-clade members lack several of the genes in the glycopeptidolipid (GLP) locus, which have roles in colony morphotype appearance and virulence. For the M. mucogenicum type strain, Mmuc(T), we provide RNASeq data focusing on mRNA levels for sigma factors, STPK, ESX proteins and Mce proteins. These data are discussed and compared to in particular the SGM and fish pathogen Mycobacterium marinum. Finally, we provide insight into as to why members of the Mmuc- and Mneo-clades show resistance to rifampin and isoniazid, and why Mmuc(T) forms a rough colony morphotype.
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| 3. |
- Das, Sarbashis, et al.
(author)
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Characterization of three Mycobacterium spp. with potential use in bioremediation by genome sequencing and comparative genomics
- 2015
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In: Genome Biology and Evolution. - : Oxford University Press (OUP). - 1759-6653. ; 7:7, s. 1871-1886
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Journal article (peer-reviewed)abstract
- We provide the genome sequences of the type strains of the polychlorophenol-degrading Mycobacterium chlorophenolicum (DSM43826), the degrader of chlorinated aliphatics Mycobacterium chubuense (DSM44219) and Mycobacterium obuense (DSM44075) that has been tested for use in cancer immunotherapy. The genome sizes of M. chlorophenolicum, M. chubuense and M. obuense are 6.93, 5.95 and 5.58 Mbps with GC-contents of 68.4, 69.2 and 67.9%, respectively. Comparative genomic analysis revealed that 3254 genes are common and we predicted approximately 250 genes acquired through horizontal gene transfer from different sources including proteobacteria. The data also showed that the biodegrading Mycobacterium spp. NBB4, also referred to as M. chubuense NBB4, is distantly related to the M. chubuense type strain and should be considered as a separate species, we suggest it to be named M. ethylenense NBB4. Among different categories we identified genes with potential roles in: biodegradation of aromatic compounds, and copper homeostasis. These are the first non-pathogenic Mycobacterium spp. found harboring genes involved in copper homeostasis. These findings would therefore provide insight into the role of this group of Mycobacterium spp. in bioremediation as well as the evolution of copper homeostasis within the Mycobacterium genus.
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| 4. |
- Das, Sarbashis, et al.
(author)
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Extensive genomic diversity among Mycobacterium marinum strains revealed by whole genome sequencing
- 2018
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In: Scientific Reports. - : Springer Science and Business Media LLC. - 2045-2322. ; 8
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Journal article (peer-reviewed)abstract
- Mycobacterium marinum is the causative agent for the tuberculosis-like disease mycobacteriosis in fish and skin lesions in humans. Ubiquitous in its geographical distribution, M. marinum is known to occupy diverse fish as hosts. However, information about its genomic diversity is limited. Here, we provide the genome sequences for 15 M. marinum strains isolated from infected humans and fish. Comparative genomic analysis of these and four available genomes of the M. marinum strains M, E11, MB2 and Europe reveal high genomic diversity among the strains, leading to the conclusion that M. marinum should be divided into two different clusters, the "M"- and the "Aronson"-type. We suggest that these two clusters should be considered to represent two M. marinum subspecies. Our data also show that the M. marinum pan-genome for both groups is open and expanding and we provide data showing high number of mutational hotspots in M. marinum relative to other mycobacteria such as Mycobacterium tuberculosis. This high genomic diversity might be related to the ability of M. marinum to occupy different ecological niches.
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| 5. |
- Das, Sarbashis, et al.
(author)
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The Mycobacterium phlei Genome : Expectations and Surprises
- 2016
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In: Genome Biology and Evolution. - : Oxford University Press (OUP). - 1759-6653. ; 8:4, s. 975-985
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Journal article (peer-reviewed)abstract
- Mycobacterium phlei, a nontuberculosis mycobacterial species, was first described in 1898-1899. We present the complete genome sequence for the IV, phlei CCUG21000(T) type strain and the draft genomes for four additional strains. The genome size for all five is 5.3 Mb with 69.4% Guanine-Cytosine content. This is approximate to 0.35 Mbp smaller than the previously reported M. phlei RIVM draft genome. The size difference is attributed partly to large bacteriophage sequence fragments in the M. phlei RIVM genome. Comparative analysis revealed the following: 1) A CRISPR system similar to Type 1E (cas3) in M. phiei RIVM; 2) genes involved in polyamine metabolism and transport (potAD, potT) that are absent in other mycobacteria, and 3) strain specific variations in the number of sigma-factor genes. Moreover, M. phlei has as many as 82 mce (mammalian cell entry) homologs and many of the horizontally acquired genes in M. phlei are present in other environmental bacteria including mycobacteria that share similar habitat. Phylogenetic analysis based on 693 Mycobacterium core genes present in all complete mycobacterial genomes suggested that its closest neighbor is Mycobacterium smegmatis JS623 and Mycobacterium rhodesiae NBB3, while it is more distant to M. smegmatis mc2 155.
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| 6. |
- Pettersson, B. M. Fredrik, et al.
(author)
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Comparative Sigma Factor-mRNA Levels in Mycobacterium marinum under Stress Conditions and during Host Infection
- 2015
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In: PLOS ONE. - : Public Library of Science (PLoS). - 1932-6203. ; 10:10
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Journal article (peer-reviewed)abstract
- We have used RNASeq and qRT-PCR to study mRNA levels for all s-factors in different Mycobacterium marinum strains under various growth and stress conditions. We also studied their levels in M. marinum from infected fish and mosquito larvae. The annotated s-factors were expressed and transcripts varied in relation to growth and stress conditions. Some were highly abundant such as sigA, sigB, sigC, sigD, sigE and sigH while others were not. The s-factor mRNA profiles were similar after heat stress, during infection of fish and mosquito larvae. The similarity also applies to some of the known heat shock genes such as the a-crystallin gene. Therefore, it seems probable that the physiological state of M. marinum is similar when exposed to these different conditions. Moreover, the mosquito larvae data suggest that this is the state that the fish encounter when infected, at least with respect to s-factor mRNA levels. Comparative genomic analysis of s-factor gene localizations in three M. marinum strains and Mycobacterium tuberculosis H37Rv revealed chromosomal rearrangements that changed the localization of especially sigA, sigB, sigD, sigE, sigF and sigJ after the divergence of these two species. This may explain the variation in species-specific expression upon exposure to different growth conditions.
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| 7. |
- Ramesh, Malavika, et al.
(author)
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Age-dependent pleomorphism in Mycobacterium monacense cultures
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Other publication (pop. science, debate, etc.)abstract
- Changes in cell shape and pleomorphism have been shown to be an integral part of the mycobacterial life cycle, however, systematic investigations into its patterns of pleomorphic behaviour in connection with stages or conditions of growth is scarce. We have studied the complete growth-cycle of Mycobacterium monacense cultures, a Non-Tuberculous Mycobacterium (NTM), in solid as well as liquid media. We provide data showing changes in cell shape from rod to coccoid and occurrence of refractive cells ranging from Phase Grey to phase Bright (PGB) in appearance upon ageing. Data further showed that changes in cell shape observed under the microscope could be correlated to the biphasic nature of the growth curves for M. monacense (as well as the NTM Mycobacterium boenickei) as measured by the absorbance of liquid cultures. Using the complete M. monacense genome we identified genes involved in cell morphology, and transcriptome analyses at different stages of growth revealed changes in their mRNA levels. One gene of interest, dnaK_3, that showed strong upregulation during stationary phase, was identified as an MreB-like homolog based on the protein domain architecture. Exogenous overexpression of M. monacense dnaK_3 in Mycobacterium marinum resulted in morphological changes with an impact on the frequency of occurrence of PGB cells. However, the introduction of an anti-sense "gene" targeting M. marinum dnaK_3 did not show such effects. Using dnaK_3-lacZ reporter constructs we provide data that these differences could be attributed to differences in the regulation of dnaK_3 in the two species. Together, this suggests that although its regulation may vary between mycobacterial species, dnaK_3 might be involved in the mechanism influencing mycobacterial cell shape.
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| 8. |
- Ramesh, Malavika, et al.
(author)
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Age-dependent pleomorphism in Mycobacterium monacense cultures
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Other publication (pop. science, debate, etc.)abstract
- Changes in cell shape and pleomorphism have been shown to be an integral part of the mycobacterial life cycle, however, systematic investigations into its patterns of pleomorphic behaviour in connection with stages or conditions of growth is scarce. We have studied the complete growth-cycle of Mycobacterium monacense cultures, a Non-Tuberculous Mycobacterium (NTM), in solid as well as liquid media. We provide data showing changes in cell shape from rod to coccoid and occurrence of refractive cells ranging from Phase Grey to phase Bright (PGB) in appearance upon ageing. Data further showed that changes in cell shape observed under the microscope could be correlated to the biphasic nature of the growth curves for M. monacense (as well as the NTM Mycobacterium boenickei) as measured by the absorbance of liquid cultures. Using the complete M. monacense genome we identified genes involved in cell morphology, and transcriptome analyses at different stages of growth revealed changes in their mRNA levels. One gene of interest, dnaK_3, that showed strong upregulation during stationary phase, was identified as an MreB-like homolog based on the protein domain architecture. Exogenous overexpression of M. monacense dnaK_3 in Mycobacterium marinum resulted in morphological changes with an impact on the frequency of occurrence of PGB cells. However, the introduction of an anti-sense "gene" targeting M. marinum dnaK_3 did not show such effects. Using dnaK_3-lacZ reporter constructs we provide data that these differences could be attributed to differences in the regulation of dnaK_3 in the two species. Together, this suggests that although its regulation may vary between mycobacterial species, dnaK_3 might be involved in the mechanism influencing mycobacterial cell shape.
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| 9. |
- Ramesh, Malavika, et al.
(author)
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Branching morphology in non-tuberculous mycobacteria Mycobacterium senegalense and Mycobacterium malmoense
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Other publication (pop. science, debate, etc.)abstract
- Understanding mycobacterial growth cycle has been a challenging topic ever since the identification of Mycobacterium tuberculosis (Mtb) as the causal agent of Tuberculosis. Diverse metabolic pathways, changes in cell shape and/or size under various growth and environmental conditions, dormancy and virulence are some of the major factors that have made mycobacterial studies complex and challenging. Over the years, many mycobacterial species classified as Non-Tuberculous Mycobacteria (NTM) have been isolated from humans, animals and identified as pathogens. In this study, we have observed and described the cell morphology of two NTMs that showed branching/filamentous growth. Mycobacterium senegalense (Msen), causative agent of bovine farcy showed extensive branching with spore-like phase grey and phase bright (PGB) structures similar to the ones observed in a slow-growing NTM Mycobacterium malmoense (Mmal), known to cause TB-like lung infections in humans. Global transcriptome analyses of various genes is suggestive of pathways and regulations that may be responsible for the pleiomorphsim in general and specifically for branching in Msen and Mmal. One such gene that was analysed was the wag31, which has been shown to localise preferably at branch points.
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| 10. |
- Ramesh, Malavika, et al.
(author)
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Branching morphology in the non-tuberculous mycobacteria Mycobacterium senegalense and Mycobacterium malmoense
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Other publication (pop. science, debate, etc.)abstract
- Understanding mycobacterial growth cycle has been a challenging topic ever since the identification of Mycobacterium tuberculosis (Mtb) as the causal agent of Tuberculosis. Diverse metabolism, changes in cell shape and/or size under various growth and environmental conditions, dormancy and virulence are some of the major factors that has posed mycobacterial studies as very challenging. Over the years, many mycobacterial species classified as Non-Tuberculous Mycobacteria (NTM) have been isolated from humans, animals and identified as pathogens. In this study, we have observed and described the cell morphology of two NTMs that showed branching/filamentous growth. Mycobacterium senegalense (Msen), causative agent of bovine farcy showed extensive branching with spore-like phase grey and phase bright (PGB) structures similar to the ones observed in a slow-growing NTM Mycobacterium malmoense (Mmal), known to cause TB-like lung infections in humans. Global transcriptome analyses of various genes is suggestive of pathways and regulations that may be responsible for the pleiomorphsim in general and specifically for branching in Msen and Mmal. One such gene that was analysed was the wag31, which has been shown to localise preferably at branch points.
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| 11. |
- Ramesh, Malavika, et al.
(author)
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Intracellular localization of the mycobacterial stressosome complex
- 2021
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In: Scientific Reports. - : Springer Nature. - 2045-2322. ; 11:1
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Journal article (peer-reviewed)abstract
- Microorganisms survive stresses by alternating the expression of genes suitable for surviving the immediate and present danger and eventually adapt to new conditions. Many bacteria have evolved a multiprotein "molecular machinery" designated the "Stressosome" that integrates different stress signals and activates alternative sigma factors for appropriate downstream responses. We and others have identified orthologs of some of the Bacillus subtilis stressosome components, RsbR, RsbS, RsbT and RsbUVW in several mycobacteria and we have previously reported mutual interactions among the stressosome components RsbR, RsbS, RsbT and RsbUVW from Mycobacterium marinum. Here we provide evidence that "STAS" domains of both RsbR and RsbS are important for establishing the interaction and thus critical for stressosome assembly. Fluorescence microscopy further suggested co-localization of RsbR and RsbS in multiprotein complexes visible as co-localized fluorescent foci distributed at scattered locations in the M. marinum cytoplasm; the number, intensity and distribution of such foci changed in cells under stressed conditions. Finally, we provide bioinformatics data that 17 (of 244) mycobacteria, which lack the RsbRST genes, carry homologs of Bacillus cereus genes rsbK and rsbM indicating the existence of alternative σF activation pathways among mycobacteria.
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| 12. |
- Ramesh, Malavika
(author)
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Morphological changes in mycobacteria
- 2021
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Doctoral thesis (other academic/artistic)abstract
- Morphological variations at different stages and conditions of growth has been observed frequently in various mycobacteria. Despite years of research, the molecular basis of such variations is still not clear and requires further elucidation. Owing to the pathogenic significance of mycobacteria, currently, mycobacterial research emphasizes on understanding responses to oxygen deprivation, starvation, various antibiotic treatments, latency and dormancy.In this thesis, change in cell shape from rod to coccoid, occurrence of refractive spore-like phase grey, phase bright bodies and branching are some of the morphological variations discussed. Further, this thesis also discusses microaerobic condition (oxygen deprivation) and the localisation pattern of the stressosome complex.
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| 13. |
- Ramesh, Malavika, et al.
(author)
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Pleiomorphism and gene expression profile of SGM Mycobacterium marinum and RGM Mycobacterium monacense under microaerobic condition
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Other publication (pop. science, debate, etc.)abstract
- Microaerobic condition is often studied in relation to pathogenesis of mycobacteria as it mimics the environment inside the lungs of an infected TB patient. Adaptation to a dormant state by Mycobacterium tuberculosis (Mtb) help the pathogen to persist within the host system and cause latent infections. Hypoxia, also referred to as microaerobic condition, is one of the major factors inducing dormancy in Mtb. Studies have characterised dormancy and latency by certain features such as inability to replicate, low metabolic activity, and expression of various genetic markers such as expression genes belonging to the Dos regulon (dormancy survival regulon). In addition to the mRNA profiles of various dormancy associated genes, this study also shows the morphological changes observed in the slow growing (SGM) Mycobacterium marinum and the rapid growing (RGM) Mycobacterium monacense. The observed morphological changes refers to increasing frequencies of refractive spore-like Phase Grey and phase Bright (PGB) cells in mycobacterial cultures when subjected to microaerobic condition. Similar observations with respect to PGB cells has also been detected in Mtb cultures. Together this suggests that changes in cell morphology under microaerobic condition occurs in mycobacteria irrespective of the species. Our microscopic and RNASeq data further suggested that sigB and the mreB-like homolog dnaK_3 both influence cell shape upon depletion of oxygen.
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| 14. |
- Ramesh, Malavika, et al.
(author)
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Pleomorphism and gene expression profile of SGM Mycobacterium marinum and RGM Mycobacterium monacense under microaerobic condition
-
Other publication (pop. science, debate, etc.)abstract
- Microaerobic condition is often studied in relation to pathogenesis of mycobacteria as it mimics the environment inside the lungs of an infected TB patient. Adaptation to a dormant state by Mycobacterium tuberculosis (Mtb) help the pathogen to persist within the host system and cause latent infections. Hypoxia, also referred to as microaerobic condition, is one of the major factors inducing dormancy in Mtb. Studies have characterised dormancy and latency by certain features such as inability to replicate, low metabolic activity, and expression of various genetic markers such as expression genes belonging to the Dos regulon (dormancy survival regulon). In addition to the mRNA profiles of various dormancy associated genes, this study also shows the morphological changes observed in the slow growing (SGM) Mycobacterium marinum and the rapid growing (RGM) Mycobacterium monacense. The observed morphological changes refers to increasing frequencies of refractive spore-like Phase Grey and phase Bright (PGB) cells in mycobacterial cultures when subjected to microaerobic condition. Similar observations with respect to PGB cells has also been detected in Mtb cultures. Together this suggests that changes in cell morphology under microaerobic condition occurs in mycobacteria irrespective of the species. Our microscopic and RNASeq data further suggested that sigB and the mreB-like homolog dnaK_3 both influence cell shape upon depletion of oxygen.
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| 15. |
- Singh, Bhupender, et al.
(author)
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Asymmetric growth and division in Mycobacterium spp. : compensatory mechanisms for non-medial septa
- 2013
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In: Molecular Microbiology. - : Wiley. - 0950-382X .- 1365-2958. ; 88:1, s. 64-76
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Journal article (peer-reviewed)abstract
- Mycobacterium spp., rod-shaped cells belonging to the phylum Actinomycetes, lack the Min- and Noc/Slm systems responsible for preventing the placement of division sites at the poles or over the nucleoids to ensure septal assembly at mid-cell. We show that the position for establishment of the FtsZ-ring in exponentially growing Mycobacterium marinum and Mycobacterium smegmatis cells is nearly random, and that the cells often divide non-medially, producing two unequal but viable daughters. Septal sites and cellular growth disclosed by staining with the membrane-specific dye FM4-64 and fluorescent antibiotic vancomycin (FL-Vanco), respectively, showed that many division sites were off-centre, often over the nucleoids, and that apical cell growth was frequently unequal at the two poles. DNA transfer through the division septum was detected, and translocation activity was supported by the presence of a putative mycobacterial DNA translocase (MSMEG2690) at the majority of the division sites. Time-lapse imaging of single live cells through several generations confirmed both acentric division site placement and unequal polar growth in mycobacteria. Our evidence suggests that post-septal DNA transport and unequal polar growth may compensate for the non-medial division site placement in Mycobacterium spp.
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