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1.
Viruses ; 13(7)2021 07 15.
Artigo em Inglês | MEDLINE | ID: mdl-34372584

RESUMO

Double-stranded DNA bacteriophages end their lytic cycle by disrupting the host cell envelope, which allows the release of the virion progeny. Each phage must synthesize lysis proteins that target each cell barrier to phage release. In addition to holins, which permeabilize the cytoplasmic membrane, and endolysins, which disrupt the peptidoglycan (PG), mycobacteriophages synthesize a specific lysis protein, LysB, capable of detaching the outer membrane from the complex cell wall of mycobacteria. The family of LysB proteins is highly diverse, with many members presenting an extended N-terminus. The N-terminal region of mycobacteriophage Ms6 LysB shows structural similarity to the PG-binding domain (PGBD) of the φKZ endolysin. A fusion of this region with enhanced green fluorescent protein (Ms6LysBPGBD-EGFP) was shown to bind to Mycobacterium smegmatis, Mycobacterium vaccae, Mycobacterium bovis BGC and Mycobacterium tuberculosis H37Ra cells pretreated with SDS or Ms6 LysB. In pulldown assays, we demonstrate that Ms6 LysB and Ms6LysBPGBD-EGFP bind to purified peptidoglycan of M. smegmatis, Escherichia coli, Pseudomonas aeruginosa and Bacillus subtilis, demonstrating affinity to PG of the A1γ chemotype. An infection assay with an Ms6 mutant producing a truncated version of LysB lacking the first 90 amino acids resulted in an abrupt lysis. These results clearly demonstrate that the N-terminus of Ms6 LysB binds to the PG.


Assuntos
Bacteriólise/fisiologia , Micobacteriófagos/metabolismo , Proteínas Virais/genética , Membrana Celular/metabolismo , Parede Celular/metabolismo , Endopeptidases , Hidrólise , Mycobacterium/metabolismo , Mycobacterium/virologia , Peptidoglicano/metabolismo , Ligação Proteica
2.
Methods Mol Biol ; 2131: 329-347, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32162265

RESUMO

Mycobacterium sp. is exhibiting complex evolution of antimicrobial resistance (AMR) and can therefore be considered as a serious human pathogen. Many strategies were employed earlier to evade the pathogenesis but AMR became threatened. Molecular tools employing bacteriophage can be an alternative to effective treatment against Mycobacterium. Phage treatment using phage-encoded products, such as lysins, causes lysis of cells; particularly bacteria could be used instead of direct use of these bacteriophages. Modern technologies along with bacteriophage strategies such as in silico immunoinformatics approach, machine learning, and artificial intelligence have been described thoroughly to escape the pathogenesis. Therefore, understanding the molecular mechanisms could be a possible alternative to evade the pathogenesis.


Assuntos
Micobacteriófagos/fisiologia , Infecções por Mycobacterium/prevenção & controle , Mycobacterium/crescimento & desenvolvimento , Animais , Biologia Computacional , Enzimas/farmacologia , Interações Hospedeiro-Patógeno , Humanos , Aprendizado de Máquina , Mycobacterium/efeitos dos fármacos , Mycobacterium/virologia , Infecções por Mycobacterium/tratamento farmacológico , Terapia por Fagos
3.
Int J Mycobacteriol ; 8(2): 170-174, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31210161

RESUMO

Background: Mycobacteriophages are viruses that infect Mycobacterium spp. Till date, 10427 mycobacteriophages have been isolated and 1670 mycobacteriophage genomes have been sequenced https://phagesdb.org/hosts/genera/1/ (cited on 30th December,2018). In the previous study, 10 different mycobacteriophages from 14 soil samples were isolated, by qualitative plaque formation method using Mycobacterium smegmatis as host. Among these, three phages were found to infect four different species of Mycobacterium, i.e., Mycobacterium fortuitum subsp. fortuitum MTCC993, Mycobacterium kansasii MTCC3058, Mycobacterium avium subsp. avium MTCC1723, and Mycobacterium tuberculosis MTCC300, besides the host M. smegmatis. The phage lysates were concentrated by polyethylene glycol (PEG) precipitation. One of the three phages showing host diversity was selected for further study. The various phage growth parameters such as incubation temperature, time of adsorption, host cell density and effect of cations were standardised. Methods: The studies were done by qualitative and quantitative plaque assay method. Results: The phage selected for further study showed an optimum adsorption time of 15 min. The optimum temperature for propagation was found to be 37°C. The phage was found to be stable at 42°C. In the presence of calcium, the phage showed a higher rate of infectivity. Conclusion: Understanding the biology of mycobacteriophages and their host diversity is the key to understanding mycobacterial systems. This could be the first step toward exploiting the potential of phages as therapeutic agents.


Assuntos
Micobacteriófagos/fisiologia , Mycobacterium/virologia , Cátions/química , Especificidade de Hospedeiro , Infecções por Mycobacterium/terapia , Mycobacterium smegmatis/virologia , Mycobacterium tuberculosis/virologia , Terapia por Fagos , Temperatura , Ensaio de Placa Viral
4.
Viruses ; 9(11)2017 11 17.
Artigo em Inglês | MEDLINE | ID: mdl-29149017

RESUMO

All dsDNA phages encode two proteins involved in host lysis, an endolysin and a holin that target the peptidoglycan and cytoplasmic membrane, respectively. Bacteriophages that infect Gram-negative bacteria encode additional proteins, the spanins, involved in disruption of the outer membrane. Recently, a gene located in the lytic cassette was identified in the genomes of mycobacteriophages, which encodes a protein (LysB) with mycolyl-arabinogalactan esterase activity. Taking in consideration the complex mycobacterial cell envelope that mycobacteriophages encounter during their life cycle, it is valuable to evaluate the role of these proteins in lysis. In the present work, we constructed an Ms6 mutant defective on lysB and showed that Ms6 LysB has an important role in lysis. In the absence of LysB, lysis still occurs but the newly synthesized phage particles are deficiently released to the environment. Using cryo-electron microscopy and tomography to register the changes in the lysis phenotype, we show that at 150 min post-adsorption, mycobacteria cells are incompletely lysed and phage particles are retained inside the cell, while cells infected with Ms6wt are completely lysed. Our results confirm that Ms6 LysB is necessary for an efficient lysis of Mycobacterium smegmatis, acting, similarly to spanins, in the third step of the lysis process.


Assuntos
Esterases/metabolismo , Micobacteriófagos/genética , Micobacteriófagos/fisiologia , Mycobacterium/virologia , Microscopia Crioeletrônica , Endopeptidases , Esterases/genética , Galactanos , Hidrólise , Micobacteriófagos/enzimologia , Micobacteriófagos/ultraestrutura , Mycobacterium/metabolismo , Mycobacterium/ultraestrutura , Tomografia , Proteínas Virais/genética
5.
Sci Rep ; 7(1): 16514, 2017 11 28.
Artigo em Inglês | MEDLINE | ID: mdl-29184079

RESUMO

Mycobacteriophage are viruses that infect mycobacteria. More than 1,400 mycobacteriophage genomes have been sequenced, coding for over one hundred thousand proteins of unknown functions. Here we investigate mycobacteriophage Giles-host protein-protein interactions (PPIs) using yeast two-hybrid screening (Y2H). A total of 25 reproducible PPIs were found for a selected set of 10 Giles proteins, including a putative virion assembly protein (gp17), the phage integrase (gp29), the endolysin (gp31), the phage repressor (gp47), and six proteins of unknown function (gp34, gp35, gp54, gp56, gp64, and gp65). We note that overexpression of the proteins is toxic to M. smegmatis, although whether this toxicity and the associated changes in cellular morphology are related to the putative interactions revealed in the Y2H screen is unclear.


Assuntos
Proteínas de Bactérias/metabolismo , Interações Hospedeiro-Patógeno , Micobacteriófagos/fisiologia , Mycobacterium/metabolismo , Mycobacterium/virologia , Mapeamento de Interação de Proteínas , Proteínas Virais/metabolismo , Regulação Viral da Expressão Gênica , Fenótipo , Mapas de Interação de Proteínas , Técnicas do Sistema de Duplo-Híbrido , Proteínas Virais/genética
6.
BMC Microbiol ; 16(1): 111, 2016 06 17.
Artigo em Inglês | MEDLINE | ID: mdl-27316672

RESUMO

BACKGROUND: A large collection of sequenced mycobacteriophages capable of infecting a single host strain of Mycobacterium smegmatis shows considerable genomic diversity with dozens of distinctive types (clusters) and extensive variation within those sharing evident nucleotide sequence similarity. Here we profiled the mycobacterial components of a large composting system at the São Paulo zoo. RESULTS: We isolated and sequenced eight mycobacteriophages using Mycobacterium smegmatis mc(2)155 as a host. None of these eight phages infected any of mycobacterial strains isolated from the same materials. The phage isolates span considerable genomic diversity, including two phages (Barriga, Nhonho) related to Subcluster A1 phages, two Cluster B phages (Pops, Subcluster B1; Godines, Subcluster B2), three Subcluster F1 phages (Florinda, Girafales, and Quico), and Madruga, a relative of phage Patience with which it constitutes the new Cluster U. Interestingly, the two Subcluster A1 phages and the three Subcluster F1 phages have genomic relationships indicating relatively recent evolution within a geographically isolated niche in the composting system. CONCLUSIONS: We predict that composting systems such as those used to obtain these mycobacteriophages will be a rich source for the isolation of additional phages that will expand our view of bacteriophage diversity and evolution.


Assuntos
Micobacteriófagos/genética , Micobacteriófagos/isolamento & purificação , Mycobacterium/genética , Mycobacterium/virologia , Microbiologia do Solo , Solo , Bacteriófagos/genética , Sequência de Bases , Brasil , DNA Bacteriano/genética , DNA Viral/genética , Evolução Molecular , Genes Bacterianos , Variação Genética , Genoma Viral , Família Multigênica , Micobacteriófagos/classificação , Mycobacterium/classificação , Mycobacterium/isolamento & purificação , Mycobacterium smegmatis/classificação , Mycobacterium smegmatis/genética , Mycobacterium smegmatis/isolamento & purificação , Mycobacterium smegmatis/virologia , Filogenia
7.
Indian J Med Microbiol ; 34(2): 186-92, 2016.
Artigo em Inglês | MEDLINE | ID: mdl-27080770

RESUMO

PURPOSE: The aim of this study was to isolate a novel mycobacteriophage and then explore its anti-tuberculosis (TB) potential. MATERIALS AND METHODS: Phage was isolated from enriched soil sample. A total of 36 mycobacterial strains obtained from clinical specimens were subjected to investigate the host range of phage by the spot lysis assay. Biological characteristics were investigated through growth curve, host range and phage antimicrobial activity in vitro. Then, genome sequencing and further analysis were accomplished by using an ABI3730XL DNA sequencer and comparative genome, respectively. RESULTS: A lytic mycobacteriophage (Chy1) was isolated and the plaque morphology was similar to D29. The genome of Chy1 was estimated to be about 47,198 base pair (bp) and strong similarity (97.4% identity) to D29, especially, the Chy1 gene 7 encoding holin which is considered as a clock controlling growth cycle of the corresponding phage, was identical (100% identity) to phage D29 gene 11, thus classifying Chy1 as a member of the cluster A2 family. However, to our surprise, Chy1 can infect a narrower range of host-mycobacterial strains than that of D29. The latent period of Chy1 was quite longer compared to D29. Moreover, Chy1 has a weaker ability to lyse Mycobacterium smegmatis compared to D29. CONCLUSIONS: The sequence of Chy1 showed 97.4% homology with the genome sequence of D29, but there was a large difference in their biological characteristics. Overall, the results of this investigation indicate that Chy1 is not an ideal candidate for developing mycobacteriophage-based anti-TB therapies but for future researches to investigate the reason why biological characteristics of Chy1 and D29 were remarkably different.


Assuntos
Genótipo , Especificidade de Hospedeiro , Micobacteriófagos/isolamento & purificação , Micobacteriófagos/fisiologia , Mycobacterium/virologia , Microbiologia do Solo , Genoma Viral , Humanos , Micobacteriófagos/classificação , Micobacteriófagos/genética , Mycobacterium/isolamento & purificação , Infecções por Mycobacterium/microbiologia , Mycobacterium smegmatis , Análise de Sequência de DNA , Replicação Viral
8.
BMC Bioinformatics ; 17: 30, 2016 Jan 13.
Artigo em Inglês | MEDLINE | ID: mdl-26757899

RESUMO

BACKGROUND: In recent years, many studies focused on the description and comparison of large sets of related bacteriophage genomes. Due to the peculiar mosaic structure of these genomes, few informative approaches for comparing whole genomes exist: dot plots diagrams give a mostly qualitative assessment of the similarity/dissimilarity between two or more genomes, and clustering techniques are used to classify genomes. Multiple alignments are conspicuously absent from this scene. Indeed, whole genome aligners interpret lack of similarity between sequences as an indication of rearrangements, insertions, or losses. This behavior makes them ill-prepared to align bacteriophage genomes, where even closely related strains can accomplish the same biological function with highly dissimilar sequences. RESULTS: In this paper, we propose a multiple alignment strategy that exploits functional collinearity shared by related strains of bacteriophages, and uses partial orders to capture mosaicism of sets of genomes. As classical alignments do, the computed alignments can be used to predict that genes have the same biological function, even in the absence of detectable similarity. The Alpha aligner implements these ideas in visual interactive displays, and is used to compute several examples of alignments of Staphylococcus aureus and Mycobacterium bacteriophages, involving up to 29 genomes. Using these datasets, we prove that Alpha alignments are at least as good as those computed by standard aligners. Comparison with the progressive Mauve aligner - which implements a partial order strategy, but whose alignments are linearized - shows a greatly improved interactive graphic display, while avoiding misalignments. CONCLUSIONS: Multiple alignments of whole bacteriophage genomes work, and will become an important conceptual and visual tool in comparative genomics of sets of related strains. A python implementation of Alpha, along with installation instructions for Ubuntu and OSX, is available on bitbucket (https://bitbucket.org/thekswenson/alpha).


Assuntos
Bacteriófagos/genética , Genoma Viral , Mycobacterium/virologia , Alinhamento de Sequência/métodos , Staphylococcus aureus/virologia , Algoritmos , Biologia Computacional/métodos , Genômica/métodos
9.
Microb Genom ; 2(10): e000079, 2016 10.
Artigo em Inglês | MEDLINE | ID: mdl-28348827

RESUMO

In an genomics course sponsored by the Howard Hughes Medical Institute (HHMI), undergraduate students have isolated and sequenced the genomes of more than 1,150 mycobacteriophages, creating the largest database of sequenced bacteriophages able to infect a single host, Mycobacterium smegmatis, a soil bacterium. Genomic analysis indicates that these mycobacteriophages can be grouped into 26 clusters based on genetic similarity. These clusters span a continuum of genetic diversity, with extensive genomic mosaicism among phages in different clusters. However, little is known regarding the primary hosts of these mycobacteriophages in their natural habitats, nor of their broader host ranges. As such, it is possible that the primary host of many newly isolated mycobacteriophages is not M. smegmatis, but instead a range of closely related bacterial species. However, determining mycobacteriophage host range presents difficulties associated with mycobacterial cultivability, pathogenicity and growth. Another way to gain insight into mycobacteriophage host range and ecology is through bioinformatic analysis of their genomic sequences. To this end, we examined the correlations between the codon usage biases of 199 different mycobacteriophages and those of several fully sequenced mycobacterial species in order to gain insight into the natural host range of these mycobacteriophages. We find that UPGMA clustering tends to match, but not consistently, clustering by shared nucleotide sequence identify. In addition, analysis of GC content, tRNA usage and correlations between mycobacteriophage and mycobacterial codon usage bias suggests that the preferred host of many clustered mycobacteriophages is not M. smegmatis but other, as yet unknown, members of the mycobacteria complex or closely allied bacterial species.


Assuntos
Biodiversidade , Genoma Viral/genética , Especificidade de Hospedeiro , Micobacteriófagos/genética , Mycobacterium/virologia , Códon/genética , Micobacteriófagos/classificação , Micobacteriófagos/fisiologia
10.
Biol Direct ; 9: 19, 2014 Sep 15.
Artigo em Inglês | MEDLINE | ID: mdl-25224692

RESUMO

BACKGROUND: Mycobacterium abscessus is an emerging opportunistic pathogen which diversity was acknowledged by the recent description of two subspecies accommodating M. abscessus, Mycobacterium bolletii and Mycobacterium massiliense isolates. RESULTS: Here, genome analysis found 1-8 prophage regions in 47/48 M. abscessus genomes ranging from small prophage-like elements to complete prophages. A total of 20,304 viral and phage proteins clustered into 853 orthologous groups. Phylogenomic and phylogenetic analyses based on prophage region homology found three main clusters corresponding to M. abscessus, M. bolletii and M. massiliense. Analysing 135 annotated Tape Measure Proteins found thirteen clusters and four singletons, suggesting that at least 17 mycobacteriophages had infected M. abscessus during its evolution. The evolutionary history of phages differed from that of their mycobacterial hosts. In particular, 33 phage-related proteins have been horizontally transferred within M. abscessus genomes. They comprise of an integrase, specific mycobacteriophage proteins, hypothetical proteins and DNA replication and metabolism proteins. Gene exchanges, loss and gains which occurred in M. abscessus genomes have been driven by several mycobacteriophages. CONCLUSIONS: This analysis of phage-mycobacterium co-evolution suggests that mycobacteriophages are playing a key-role in the on-going diversification of M. abscessus. REVIEWERS: This article was reviewed by Eric Bapteste, Patrick Forterre and Eugene Koonin.


Assuntos
Variação Genética , Micobacteriófagos/fisiologia , Mycobacterium/genética , Mycobacterium/virologia , Proteínas de Bactérias/genética , Mapeamento Cromossômico , Análise por Conglomerados , Transferência Genética Horizontal/genética , Genoma Bacteriano/genética , Anotação de Sequência Molecular , Filogenia , Prófagos/genética
11.
BMC Genomics ; 15: 243, 2014 Mar 27.
Artigo em Inglês | MEDLINE | ID: mdl-24673856

RESUMO

BACKGROUND: Prophages, integral components of many bacterial genomes, play significant roles in cognate host bacteria, such as virulence, toxin biosynthesis and secretion, fitness cost, genomic variations, and evolution. Many prophages and prophage-like elements present in sequenced bacterial genomes, such as Bifidobacteria, Lactococcus and Streptococcus, have been described. However, information for the prophage of Mycobacterium remains poorly defined. RESULTS: In this study, based on the search of the complete genome database from GenBank, the Whole Genome Shotgun (WGS) databases, and some published literatures, thirty-three prophages were described in detail. Eleven of them were full-length prophages, and others were prophage-like elements. Eleven prophages were firstly revealed. They were phiMAV_1, phiMAV_2, phiMmcs_1, phiMmcs_2, phiMkms_1, phiMkms_2, phiBN42_1, phiBN44_1, phiMCAN_1, phiMycsm_1, and phiW7S_1. Their genomes and gene contents were firstly analyzed. Furthermore, comparative genomics analyses among mycobacterioprophages showed that full-length prophage phi172_2 belonged to mycobacteriophage Cluster A and the phiMmcs_1, phiMkms_1, phiBN44_1, and phiMCAN_1 shared high homology and could be classified into one group. CONCLUSIONS: To our knowledge, this is the first systematic characterization of mycobacterioprophages, their genomic organization and phylogeny. This information will afford more understanding of the biology of Mycobacterium.


Assuntos
Genoma Bacteriano , Genoma Viral , Mycobacterium/genética , Prófagos/genética , Análise por Conglomerados , Ordem dos Genes , Genes Virais , Genômica , Mycobacterium/virologia , Filogenia , Prófagos/classificação , Integração Viral
13.
Arch Microbiol ; 196(3): 209-18, 2014 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-24504137

RESUMO

Mycobacteriophage therapy is a potential alternative treatment for Mycobacterium tuberculosis infection. Here, we further characterized a mycobacteriophage, Bo4, and evaluated its ability to infect and kill M. tuberculosis. We first found that Bo4 can infect M. tuberculosis and Mycobacterium smegmatis. The observed clear plaques created by Bo4 infection indicated that Bo4 might be a lytic phage able to lyse mycobacterial strains, which was confirmed by phage antimicrobial activity. Bo4 formed clear zones in a medium with pH values of 7.4 or 5.0, suggesting the possibility that Bo4 could lyse mycobacteria, such as M. tuberculosis, in blood as well as in lysosomal macrophages. Further investigation into the Bo4 genome revealed that Bo4 had a dsDNA genome. Moreover, Bo4 contained ~39,318 bp comprised of 66.76 % G+C content. Complete genome sequencing showed high nucleotide identity with cluster G mycobacteriophages, thus classifying Bo4 as a member of the cluster G family. Additionally, annotation of the Bo4 genome indicated that it was a lytic bacteriophage and did not contain any harmful genes that increased mycobacterial virulence or decreased human immunity. Overall, the results of investigation indicate that the Bo4 possesses the potential to destroy M. tuberculosis, making it a potentially useful tool for diagnosing and treating tuberculosis.


Assuntos
Micobacteriófagos/classificação , Mycobacterium tuberculosis/virologia , Tipagem de Bacteriófagos , Composição de Bases , Genoma Viral/genética , Especificidade de Hospedeiro , Humanos , Concentração de Íons de Hidrogênio , Micobacteriófagos/genética , Mycobacterium/virologia , Filogenia , Sintenia , Tuberculose/microbiologia , Tuberculose/terapia , Replicação Viral
14.
Microbiol Spectr ; 2(2)2014 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-26105821

RESUMO

Mycobacteriophages have provided numerous essential tools for mycobacterial genetics, including delivery systems for transposons, reporter genes, and allelic exchange substrates, and components for plasmid vectors and mutagenesis. Their genetically diverse genomes also reveal insights into the broader nature of the phage population and the evolutionary mechanisms that give rise to it. The substantial advances in our understanding of the biology of mycobacteriophages including a large collection of completely sequenced genomes indicates a rich potential for further contributions in tuberculosis genetics and beyond.


Assuntos
Micobacteriófagos/genética , Micobacteriófagos/fisiologia , Mycobacterium/virologia , Evolução Molecular , Genoma Viral
15.
Appl Environ Microbiol ; 79(18): 5608-15, 2013 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-23851082

RESUMO

Addition of affinity tags to bacteriophage particles facilitates a variety of applications, including vaccine construction and diagnosis of bacterial infections. Addition of tags to phage capsids is desirable, as modification of the tails can lead to poor adsorption and loss of infectivity. Although tags can readily be included as fusions to head decoration proteins, many phages do not have decoration proteins as virion components. The addition of a small (10-amino-acid) Strep-tag II (STAG II) to the mycobacteriophage TM4 capsid subunit, gp9, was not tolerated as a genetically homogenous recombinant phage but could be incorporated into the head by growth of wild-type phage on a host expressing the capsid-STAG fusion. Particles with capsids composed of wild-type and STAG-tagged subunit mixtures could be grown to high titers, showed good infectivities, and could be used to isolate phage-bacterium complexes. Preparation of a STAG-labeled fluoromycobacteriophage enabled capture of bacterial complexes and identification of infected bacteria by fluorescence.


Assuntos
Técnicas Bacteriológicas/métodos , Proteínas do Capsídeo/metabolismo , Capsídeo/metabolismo , Micobacteriófagos/fisiologia , Mycobacterium/isolamento & purificação , Coloração e Rotulagem/métodos , Montagem de Vírus , Proteínas do Capsídeo/genética , Fluorescência , Mycobacterium/virologia , Proteínas Recombinantes de Fusão/genética , Proteínas Recombinantes de Fusão/metabolismo
16.
J Virol ; 87(14): 8099-109, 2013 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-23678183

RESUMO

The unique characteristics of the waxy mycobacterial cell wall raise questions about specific structural features of their bacteriophages. No structure of any mycobacteriophage is available, although ∼3,500 have been described to date. To fill this gap, we embarked in a genomic and structural study of a bacteriophage from Mycobacterium abscessus subsp. bolletii, a member of the Mycobacterium abscessus group. This opportunistic pathogen is responsible for respiratory tract infections in patients with lung disorders, particularly cystic fibrosis. M. abscessus subsp. bolletii was isolated from respiratory tract specimens, and bacteriophages were observed in the cultures. We report here the genome annotation and characterization of the M. abscessus subsp. bolletii prophage Araucaria, as well as the first single-particle electron microscopy reconstruction of the whole virion. Araucaria belongs to Siphoviridae and possesses a 64-kb genome containing 89 open reading frames (ORFs), among which 27 could be annotated with certainty. Although its capsid and connector share close similarity with those of several phages from Gram-negative (Gram(-)) or Gram(+) bacteria, its most distinctive characteristic is the helical tail decorated by radial spikes, possibly host adhesion devices, according to which the phage name was chosen. Its host adsorption device, at the tail tip, assembles features observed in phages binding to protein receptors, such as phage SPP1. All together, these results suggest that Araucaria may infect its mycobacterial host using a mechanism involving adhesion to cell wall saccharides and protein, a feature that remains to be further explored.


Assuntos
Genoma Viral/genética , Micobacteriófagos/genética , Mycobacterium/virologia , Siphoviridae/genética , Componentes Genômicos , Processamento de Imagem Assistida por Computador , Funções Verossimilhança , Microscopia Eletrônica , Anotação de Sequência Molecular , Micobacteriófagos/ultraestrutura , Siphoviridae/ultraestrutura , Vírion/ultraestrutura , Ligação Viral
17.
PLoS One ; 8(2): e56384, 2013.
Artigo em Inglês | MEDLINE | ID: mdl-23468864

RESUMO

Mycobacteriophages have been essential in the development of mycobacterial genetics through their use in the construction of tools for genetic manipulation. Due to the simplicity of their isolation and variety of exploitable molecular features, we searched for and isolated 18 novel mycobacteriophages from environmental samples collected from several geographic locations. Characterization of these phages did not differ from most of the previously described ones in the predominant physical features (virion size in the 100-400 nm, genome size in the 50-70 kbp, morphological features compatible with those corresponding to the Siphoviridae family), however novel characteristics for propagation were noticed. Although all the mycobacteriophages propagated at 30°C, eight of them failed to propagate at 37°C. Since some of our phages yielded pinpoint plaques, we improved plaque detection by including sub-inhibitory concentrations of isoniazid or ampicillin-sulbactam in the culture medium. Thus, searches for novel mycobacteriophages at low temperature and in the presence of these drugs would allow for the isolation of novel members that would otherwise not be detected. Importantly, while eight phages lysogenized Mycobacterium smegmatis, four of them were also capable of lysogenizing Mycobacterium tuberculosis. Analysis of the complete genome sequence obtained for twelve mycobacteriophages (the remaining six rendered partial genomic sequences) allowed for the identification of a new singleton. Surprisingly, sequence analysis revealed the presence of parA or parA/parB genes in 7/18 phages including four that behaved as temperate in M. tuberculosis. In summary, we report here the isolation and preliminary characterization of mycobacteriophages that bring new information to the field.


Assuntos
Micobacteriófagos/genética , Mycobacterium/virologia , Motivos de Aminoácidos , Sequência de Aminoácidos , Antituberculosos/farmacologia , Cátions/metabolismo , Biologia Computacional , DNA Viral , Ordem dos Genes , Genoma Viral , Isoniazida/farmacologia , Dados de Sequência Molecular , Micobacteriófagos/classificação , Micobacteriófagos/isolamento & purificação , Micobacteriófagos/fisiologia , Filogenia , Alinhamento de Sequência , Ensaio de Placa Viral , Proteínas Virais/química , Proteínas Virais/genética , Tropismo Viral , Replicação Viral/efeitos dos fármacos
19.
20.
J Bacteriol ; 194(18): 5128, 2012 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-22933758

RESUMO

The genome of Mycobacterium massiliense M172, isolated from a human sputum sample, was sequenced using Illumina GA IIX technology and found to contain 5,204,460 bp, including putative genes for virulence and antibiotic resistance as well as a 92-kb genomic region most likely to correspond to a mycobacteriophage.


Assuntos
DNA Bacteriano/química , DNA Bacteriano/genética , Genoma Bacteriano , Mycobacterium/genética , Análise de Sequência de DNA , Farmacorresistência Bacteriana , Humanos , Dados de Sequência Molecular , Micobacteriófagos/genética , Mycobacterium/isolamento & purificação , Mycobacterium/patogenicidade , Mycobacterium/virologia , Infecções por Mycobacterium/microbiologia , Escarro/microbiologia , Fatores de Virulência/genética
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