scholarly journals Comparison of Mycobacterium tuberculosis Genomes Reveals Frequent Deletions in a 20 kb Variable Region in Clinical Isolates

Yeast ◽  
2000 ◽  
Vol 1 (4) ◽  
pp. 272-282
Author(s):  
Timothy B. L. Ho ◽  
Brian D. Robertson ◽  
G. Michael Taylor ◽  
Rory J. Shaw ◽  
Douglas B. Young
Keyword(s):  
Mycobacterium Tuberculosis ◽  
Hot Spots ◽  
High Frequency ◽  
Biological Diversity ◽  
Variable Region ◽  
Clinical Isolates ◽  
Insertion Element ◽  
Genomic Deletions ◽  
Nucleotide Repeats ◽  
Strain H37rv

The Mycobacterium tuberculosis complex is associated with a remarkably low level of structural gene polymorphism. As part of a search for alternative forms of genetic variation that may act as a source of biological diversity in M. tuberculosis, we have identified a region of the genome that is highly variable amongst a panel of unrelated clinical isolates. Fifteen of 24 isolates examined contained one or more copies of the M. tuberculosis-specific IS6110 insertion element within this 20 kb variable region. In nine of the isolates, including the laboratory-passaged strain H37Rv, genomic deletions were identified, resulting in loss of between two and 13 genes. In each case, deletions were associated with the presence of a copy of the IS6110 element. Absence of flanking tri- or tetra-nucleotide repeats identified homologous recombination between adjacent IS6110 elements as the most likely mechanism of the deletion events. IS6110 insertion into hot-spots within the genome of M. tuberculosis provides a mechanism for generation of genetic diversity involving a high frequency of insertions and deletions.

scholarly journals Comparison ofMycobacterium tuberculosisGenomes Reveals Frequent Deletions in a 20 kb Variable Region in Clinical Isolates

Yeast ◽  
2000 ◽  
Vol 1 (4) ◽  
pp. 272-282 ◽  
Author(s):  
Timothy B. L. Ho ◽  
Brian D. Robertson ◽  
G. Michael Taylor ◽  
Rory J. Shaw ◽  
Douglas B. Young
Keyword(s):  
Genetic Diversity ◽  
Hot Spots ◽  
High Frequency ◽  
Biological Diversity ◽  
Variable Region ◽  
Clinical Isolates ◽  
Insertion Element ◽  
Genomic Deletions ◽  
Nucleotide Repeats ◽  
Strain H37rv

TheMycobacterium tuberculosiscomplex is associated with a remarkably low level of structural gene polymorphism. As part of a search for alternative forms of genetic variation that may act as a source of biological diversity inM. tuberculosis, we have identified a region of the genome that is highly variable amongst a panel of unrelated clinical isolates. Fifteen of 24 isolates examined contained one or more copies of theM. tuberculosis-specific IS6110insertion element within this 20 kb variable region. In nine of the isolates, including the laboratory-passaged strain H37Rv, genomic deletions were identified, resulting in loss of between two and 13 genes. In each case, deletions were associated with the presence of a copy of the IS6110element. Absence of flanking tri- or tetra-nucleotide repeats identified homologous recombination between adjacent IS6110elements as the most likely mechanism of the deletion events. IS6110insertion into hot-spots within the genome ofM. tuberculosisprovides a mechanism for generation of genetic diversity involving a high frequency of insertions and deletions.

Analysis of genetic polymorphisms affecting the four phospholipase C (plc) genes in Mycobacterium tuberculosis complex clinical isolates

Microbiology ◽  
2004 ◽  
Vol 150 (4) ◽  
pp. 967-978 ◽  
Author(s):  
C. Viana-Niero ◽  
P. E. de Haas ◽  
D. van Soolingen ◽  
S. C. Leão
Keyword(s):  
Mycobacterium Tuberculosis ◽  
Phospholipase C ◽  
Genetic Polymorphisms ◽  
Blot Hybridization ◽  
Southern Blot Hybridization ◽  
Clinical Isolates ◽  
Significant Similarity ◽  
Genomic Deletions ◽  
Genes Encoding ◽  
Mycobacterium Africanum

The Mycobacterium tuberculosis genome contains four highly related genes which present significant similarity to Pseudomonas aeruginosa genes encoding phospholipase C enzymes. Three of these genes, plcA, plcB and plcC, are organized in tandem (locus plcABC). The fourth gene, plcD, is located in a different region. This study investigates variations in plcABC and plcD genes in clinical isolates of M. tuberculosis, Mycobacterium africanum and ‘Mycobacterium canettii’. Genetic polymorphisms were examined by PCR, Southern blot hybridization, sequence analysis and RT-PCR. Seven M. tuberculosis isolates contain insertions of IS6110 elements within plcA, plcC or plcD. In 19 of 25 M. tuberculosis isolates examined, genomic deletions were identified, resulting in loss of parts of genes or complete genes from the plcABC and/or plcD loci. Partial plcD deletion was observed in one M. africanum isolate. In each case, deletions were associated with the presence of a copy of the IS6110 element and in all occurrences IS6110 was transposed in the same orientation. A mechanism of deletion resulting from homologous recombination of two copies of IS6110 was recognized in a group of genetically related M. tuberculosis isolates. Five M. tuberculosis isolates presented major polymorphisms in the plcABC and plcD regions, along with loss of expression competence that affected all four plc genes. Phospholipase C is a well-known bacterial virulence factor. The precise role of phospholipase C in the pathogenicity of M. tuberculosis is unknown, but considering the potential importance that the plc genes may have in the virulence of the tubercle bacillus, the study of isolates cultured from patients with active tuberculosis bearing genetic variations affecting these genes may provide insights into the significance of phospholipase C enzymes for tuberculosis pathogenicity.

scholarly journals New Multiplex PCR for Rapid Detection of Isoniazid-Resistant Mycobacterium tuberculosis Clinical Isolates

2005 ◽  
Vol 49 (1) ◽  
pp. 144-147 ◽  
Author(s):  
Laura Herrera-León ◽  
Tamara Molina ◽  
Pilar Saíz ◽  
Juan Antonio Sáez-Nieto ◽  
Maria Soledad Jiménez
Keyword(s):  
Mycobacterium Tuberculosis ◽  
Multiplex Pcr ◽  
High Frequency ◽  
Rapid Detection ◽  
Fragment Length Polymorphism ◽  
Regulatory Region ◽  
Clinical Isolates ◽  
Ribosome Binding ◽  
Frequent Mutations ◽  
Further Development

ABSTRACT In this study, we describe a multiplex PCR to detect a AGC→ACC (serine to threonine) mutation in the katG gene and a −15 C-to-T substitution (inhA C−15T) at the 5′ end of a presumed ribosome binding site in the promoter of the mabA-inhA operon. These mutations have been reported in the majority of previous studies as the most frequent mutations involved in the resistance to isoniazid (INH) of Mycobacterium tuberculosis clinical strains with high levels of resistance. The method was optimized and validated after an analysis of 30 M. tuberculosis clinical isolates with known sequences of the relevant part of the katG gene and the regulatory region of the mabA-inhA operon. We analyzed 297 INH-resistant M. tuberculosis isolates collected in Spain from 1996 to 2003 by PCR-restriction fragment length polymorphism (using the katG gene), DNA sequencing, and the newly developed multiplex PCR. The results were concordant for all 297 isolates tested. The analysis revealed that 204 (68.7%) of the isolates carried one or both of the mutations. This finding suggests that with further development this multiplex PCR will be able to detect the majority of the INH-resistant M. tuberculosis clinical isolates from Spain and other countries where a high frequency of similar mutations occur.

scholarly journals Emergence and Molecular Characterization of Extensively Drug-Resistant Mycobacterium tuberculosis Clinical Isolates from the Delhi Region in India

2010 ◽  
Vol 54 (11) ◽  
pp. 4789-4793 ◽  
Author(s):  
Alka Khanna ◽  
V. Samuel Raj ◽  
Bansidhar Tarai ◽  
Ruchi Sood ◽  
Pawan Kumar Pareek ◽  
...  
Keyword(s):  
Drug Resistance ◽  
Mycobacterium Tuberculosis ◽  
Hot Spots ◽  
Hot Spot ◽  
Amino Acid Position ◽  
Multidrug Resistant ◽  
Clinical Isolates ◽  
Common Mutation ◽  
Extensive Drug Resistance

ABSTRACT We screened 194 Mycobacterium tuberculosis strains isolated from tuberculosis (TB) patients in Delhi and neighboring regions in India to identify the prevalence of extensive drug resistance (XDR) in clinical isolates. Among these, 104 isolates were found to be multidrug resistant (MDR), and 6 were identified as XDR isolates, which was later confirmed by antimicrobial susceptibility testing against the respective drug screening panel. Genotyping was carried out by amplifying and sequencing the following genes: rpoB (rifampin), katG (isoniazid), gyrA (fluoroquinolones), and rrs (amikacin, kanamycin, and capreomycin). Our analyses indicated that mutations at the hot spots of these genes were positively correlated with drug resistance in clinical isolates. The key mutation observed for rpoB was in the codon for amino acid position 531 (S531L), and other mutations were seen in the hot spot, including those encoding Q510P, L511H, D516V, and H526Y mutations. We identified S315T and R463L substitutions encoded in the katG locus. An S95T substitution encoded in the gyrA locus was the most common mutation observed in fluoroquinolone-resistant isolates. In addition, we saw D94G and D94N mutations encoded in the QRDR region. The 16S rRNA (rrs) gene encoded mainly the A1401G mutation and an additional mutation, G1484T, resulting in ribosomal modifications. Taken together, the data in this report clearly establish the presence of phenotypically distinct XDR strains in India by molecular profiling and further identify specific mutational hot spots within key genes of XDR-TB strains.

scholarly journals Differences in Rate and Variability of Intracellular Growth of a Panel of Mycobacterium tuberculosis Clinical Isolates within a Human Monocyte Model

2002 ◽  
Vol 70 (11) ◽  
pp. 6489-6493 ◽  
Author(s):  
Qing Li ◽  
Christopher C. Whalen ◽  
Jeffrey M. Albert ◽  
Rhonda Larkin ◽  
Lynn Zukowski ◽  
...  
Keyword(s):  
Mycobacterium Tuberculosis ◽  
Human Monocyte ◽  
Clinical Isolates ◽  
Beijing Strain ◽  
Human Monocytes ◽  
Intracellular Growth ◽  
Strain Family ◽  
Strain H37rv

ABSTRACT Significant differences were observed in the capacities of Mycobacterium tuberculosis clinical isolates to grow within human monocytes. Genotyping indicated that the four most rapidly growing isolates were members of the Beijing strain family. M. tuberculosis strain H37Rv provided more reproducible infection than the clinical isolates or M. tuberculosis Erdman.

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