Journal: BMC Infectious Diseases

Article Title: Development and evaluation of an in-house single step loop-mediated isothermal amplification (SS-LAMP) assay for the detection of Mycobacterium tuberculosis complex in sputum samples from Moroccan patients

doi: 10.1186/s12879-016-1864-9

Figure Lengend Snippet: Nucleotide sequences of the SS-LAMP primers targeting the IS6110 sequence of MTBC strains

Article Snippet: SS-LAMP primers have been also tested on some MTBC strains DNA ( M. tuberculosis H37Rv, ATCC 25618D-2; M. tuberculosis H37Ra, ATCC 25177D-5 ; M. bovis, ATCC-BAA- 935D-2 ) and non-MTBC strains DNA ( M. avium, ATCC-BAA-968D-5 ; M. kansasii, ATCC 12478; M. intracellulare, ATCC 35769; M. abscessus, ATCC 19977D-5; M. gordonae, ATCC 14470D-5; M. smegmatis, ATCC 700084D-5; Escherichia coli, ATCC 8739; Pseudomonas aeruginosa, ATCC 9027; Staphylococcus aureus, ATCC 6538; Candida albicans, ATCC 10231; Aspergillus niger, ATCC 16888 ).

Techniques: Sequencing

Journal: PLoS ONE

Article Title: Comparative Whole-Genome Analysis of Clinical Isolates Reveals Characteristic Architecture of Mycobacterium tuberculosis Pangenome

doi: 10.1371/journal.pone.0122979

Figure Lengend Snippet: Arrow heads indicate the variably sized products in ATCC H37Ra and ATCC H37Rv genomes. Gene 4 and Gene 8 shares partial homology with laboratory strains and showed the presence of a differently sized product in ATCC H37Rv and ATCC H37Ra. (B) The alignment of Gene 4 against reference genomes showed an insertion of 175bp sequence in Gene 4 of OSDD strains with respect to reference strains. (C) The alignment of Gene 8 against reference genomes showed a deletion of 1,352bp sequence in Gene 8 of OSDD strains.

Article Snippet: Detailed insights into the genomic loci of Gene 4 alignment with reference strains showed partial overlapping of Gene 4 with hypothetical proteins in Mtb ATCC H37Rv and Mtb ATCC H37Ra thereby resulting in the formation of differentially sized gene products (Figures A and B in ).

Techniques: Sequencing

Journal: PLoS ONE

Article Title: Comparative Whole-Genome Analysis of Clinical Isolates Reveals Characteristic Architecture of Mycobacterium tuberculosis Pangenome

doi: 10.1371/journal.pone.0122979

Figure Lengend Snippet: Summary of gene clusters absent from Mtb ATCC H37Ra and Mtb ATCC H37Rv.

Article Snippet: Detailed insights into the genomic loci of Gene 4 alignment with reference strains showed partial overlapping of Gene 4 with hypothetical proteins in Mtb ATCC H37Rv and Mtb ATCC H37Ra thereby resulting in the formation of differentially sized gene products (Figures A and B in ).

Techniques:

Journal: PLoS ONE

Article Title: Comparative Whole-Genome Analysis of Clinical Isolates Reveals Characteristic Architecture of Mycobacterium tuberculosis Pangenome

doi: 10.1371/journal.pone.0122979

Figure Lengend Snippet: Arrow heads indicate the variably sized products in ATCC H37Ra and ATCC H37Rv genomes. Gene 4 and Gene 8 shares partial homology with laboratory strains and showed the presence of a differently sized product in ATCC H37Rv and ATCC H37Ra. (B) The alignment of Gene 4 against reference genomes showed an insertion of 175bp sequence in Gene 4 of OSDD strains with respect to reference strains. (C) The alignment of Gene 8 against reference genomes showed a deletion of 1,352bp sequence in Gene 8 of OSDD strains.

Article Snippet: Detailed insights into the genomic loci of Gene 4 alignment with reference strains showed partial overlapping of Gene 4 with hypothetical proteins in Mtb ATCC H37Rv and Mtb ATCC H37Ra thereby resulting in the formation of differentially sized gene products (Figures A and B in ).

Techniques: Sequencing

Journal: PLoS ONE

Article Title: Comparative Whole-Genome Analysis of Clinical Isolates Reveals Characteristic Architecture of Mycobacterium tuberculosis Pangenome

doi: 10.1371/journal.pone.0122979

Figure Lengend Snippet: Summary of gene clusters absent from Mtb ATCC H37Ra and Mtb ATCC H37Rv.

Article Snippet: Detailed insights into the genomic loci of Gene 4 alignment with reference strains showed partial overlapping of Gene 4 with hypothetical proteins in Mtb ATCC H37Rv and Mtb ATCC H37Ra thereby resulting in the formation of differentially sized gene products (Figures A and B in ).

Techniques:

Journal: Scientific Reports

Article Title: A modular paper-and-plastic device for tuberculosis nucleic acid amplification testing in limited-resource settings

doi: 10.1038/s41598-019-51873-8

Figure Lengend Snippet: LAMP in tube. ( A ) Gel electrophoresis analysis of products of LAMP reactions conducted in tube with varying starting copy numbers of Mtb gDNA (10 6 copies up to 10 copies). The numbers on top of the lanes indicate the starting copy number of the template. N- No template control, M- DNA marker and D- digested LAMP amplicons. Reaction time was 40 minutes. ( B ) Specificity of the LAMP assay tested against non- Mtb targets. +: Positive control ( Mtb gDNA), 1: Human gDNA, 2: E. coli gDNA, 3: Mycobacterium smegmatis gDNA, 4: plasmid with hepatitis C insert, 5: plasmid with dengue insert. ( C ) Real-time amplification curves for LAMP in tube. Reaction time was 80 minutes. (i–iv) denote lanes from different gels. Full-length gels are presented in Supplementary Fig. S9 (i), (ii), (iii) and (iv) .

Article Snippet: Mtb gDNA (strain H37Rv) was obtained from BEI resources (Manassas, VA, USA) and quantified using a Nanodrop 2000 (ThermoFisher Scinetific).

Techniques: Nucleic Acid Electrophoresis, Control, Marker, Lamp Assay, Positive Control, Plasmid Preparation, Amplification

Journal: Scientific Reports

Article Title: A modular paper-and-plastic device for tuberculosis nucleic acid amplification testing in limited-resource settings

doi: 10.1038/s41598-019-51873-8

Figure Lengend Snippet: LAMP amplification in paper. ( A ) Schematic of the method used for retrieving the LAMP amplicons from the paper pads. ( B ) Gel electrophoresis analysis of products of LAMP reactions conducted in paper starting from varying number of starting Mtb gDNA copies (10 4 up to 10). The numbers on top of the wells indicate the starting copy number of the template. H- 1000 copies of human gDNA were added to the reaction mix along with 1000 copies of Mtb gDNA; N- No template control; M- DNA marker; and D- digested LAMP amplicons. The reaction time was 40 minutes. Larger spaces between sections of lanes in a gel image indicate lanes from different gels and smaller spaces represent lanes from the same gel. Full-length gels are presented in Supplementary Fig. and (v).

Article Snippet: Mtb gDNA (strain H37Rv) was obtained from BEI resources (Manassas, VA, USA) and quantified using a Nanodrop 2000 (ThermoFisher Scinetific).

Techniques: Amplification, Nucleic Acid Electrophoresis, Control, Marker

Journal: Scientific Reports

Article Title: A modular paper-and-plastic device for tuberculosis nucleic acid amplification testing in limited-resource settings

doi: 10.1038/s41598-019-51873-8

Figure Lengend Snippet: LAMP in FLIPP-NAAT and end-point fluorescence detection. ( A ) Testing uniformity of field in FLIPP-NAAT. Numbers on each reaction zone mark the 12 replicates. The mean intensity for signals from the 12 reaction zones was 160 with a coefficient of variation of only 5.4%. ( B ) Cell phone image of FLIPP-NAAT after 5 minutes of addition of SYBR Green I dye for fluorescence detection. Numbers on top of each reaction zone indicate log 10 of Mtb gDNA starting copy number. N – no template control. ( C ) Mean pixel intensity of green color for the different starting copy numbers. The mean intensity of each starting copy number was statistically different from that of others (*P < 0.05; N = 3). ( D ) LAMP reactions in FLIPP-NAAT in presence of 3.1 * 10 3 copies of human gDNA. Numbers on top of each reaction zone indicate log 10 of Mtb gDNA starting copy number. N – no template control containing only human gDNA. ( E ) Mean pixel intensity of green color for the different starting copy numbers. The mean intensity of 100 and 10 copy number was statistically different from that of the negative controls (*P < 0.05; N = 3). The one drop out for 10 copies (second reaction zone from the right in the bottom row, Panel D) was not included for the intensity analysis as it was a false negative. The dashed line represents the threshold value of (µ N + 3σ N ). Samples with intensity below the threshold were considered as TB negative while samples with green color intensity above the threshold were considered as TB positive.

Article Snippet: Mtb gDNA (strain H37Rv) was obtained from BEI resources (Manassas, VA, USA) and quantified using a Nanodrop 2000 (ThermoFisher Scinetific).

Techniques: Fluorescence, SYBR Green Assay, Control

Journal: Scientific Reports

Article Title: A modular paper-and-plastic device for tuberculosis nucleic acid amplification testing in limited-resource settings

doi: 10.1038/s41598-019-51873-8

Figure Lengend Snippet: LAMP in FLIPP-NAAT from dry-stored reagents. ( A ) End-point imaging of the FLIPP-NAAT device after 5 minutes of addition of SYBR green I dye. Numbers on top of each reaction zone indicate log 10 of starting copy number. N – no template control. ( B ) Mean pixel intensity of the green color from all starting copy numbers was statistically higher (*P < 0.05; N = 3) from that of no template control. The dashed line represents the threshold value of (µ N + 3σ N ). Samples with intensity below the threshold were considered as TB negative while samples with green color intensity above the threshold were considered as TB positive. ( C ) LAMP in FLIPP-NAAT from dry-stored reagents and rehydration with mock sputum. Each reaction zone had 100 copies of Mtb gDNA as the starting template. The markings on top of each of the reaction zone indicate the dilution of mock sputum used for rehydration. 3 W, 2 W, 1 W – reaction set-up with template suspended in water containing 1000, 100 and 10 copies of Mtb gDNA respectively, as positive controls. All reactions worked despite the presence of the complex sputum matrix.

Article Snippet: Mtb gDNA (strain H37Rv) was obtained from BEI resources (Manassas, VA, USA) and quantified using a Nanodrop 2000 (ThermoFisher Scinetific).

Techniques: Imaging, SYBR Green Assay, Control