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Figure 5: Detection of PorMs in M. fortuitum and M. smegmatis. 2D-Electrophoresis, Western Blot, ELISA and qRT-PCR experiments proved PorMs to be expressed in the analysed strains. Section A shows 2D-Electrophoresis of protein isolation from the strain M. fortuitum 10860/03 using the detergent nOPOE. The arrow indicates the porin spot proven by Western Blot analysis (see Additional file 2). Section B and C show comparative analysis of porin expression among RGM. Expression of porin was detected by means of ELISA (B) and qRT-PCR (C). Each value represents the mean (± SD) of at least three independent experiments. B: Quantification of porin by means of ELISA in cell extracts of different mycobacteria using the polyclonal antibody pAk MspA#813. C: RT-Real-time-PCR quantification of porin mRNA in various RGM using specific primers and probes for mspA or porM, respectively.

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Other Images from "Characterisation of porin genes from Mycobacterium fortuitum and their impact on growth":


Figure 6 Complementation of the porin-deficient m...

Figure 7 Effect of down-regulation and over-expre...

Figure 3 Occurrence of porin genes in M. fortuitu...

Figure 1 Growth rate of the M. fortuitum strains ...

Figure 4 Alignment of PorM1 and PorM2 from M. for...

Figure 2 Map of genomic regions containing porM1 ...

Figure 5 Detection of PorMs in M. fortuitum and M...

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Abstract

BackgroundHighly pathogenic mycobacteria like Mycobacterium tuberculosis are characterised by their slow growth and their ability to reside and multiply in the very hostile phagosomal environment and a correlation between the growth rate of mycobacteria and their pathogenicity has been hypothesised. Here, porin genes from M. fortuitum were cloned and characterised to address their impact on the growth rate of fast-growing and pathogenic mycobacteria.ResultsTwo genes encoding porins orthologous to MspA from M. smegmatis, porM1 and porM2, were cloned from M. fortuitum strains, which were originally isolated from human patients. Both porin genes were at least partially able to complement the mutations of a M. smegmatis mutant strain lacking the genes mspA and mspC with respect to the growth rate. PorM1 and porM2 were present in different strains of M. fortuitum including the type strain. Comparative expression analysis of porM genes revealed divergent porin expression among analysed M. fortuitum strains. Repression of the expression of porins by antisense technique decreased the growth rates of different M. fortuitum. The effects of over-expression of porM1 as well as porM2 varied depending on the strain and the concentration of antibiotic added to the medium and indicated that PorM1 and PorM2 enhance the growth of M. fortuitum strains, but also the diffusion of the antibiotic kanamycin into the cells.ConclusionThis study demonstrates the important role of porin expression in growth as well as antibiotic susceptibility of the opportunistic bacterium M. fortuitum.


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