Mycobacterium tuberculosis drug-resistance testing: challenges, recent developments and perspectives

Drug-resistance testing, or antimicrobial susceptibility testing (AST), is mandatory for Mycobacterium tuberculosis in cases of failure on standard therapy. We reviewed the different methods and techniques of phenotypic and genotypic approaches. Although multiresistant and extensively drug-resistant (MDR/XDR) tuberculosis is present worldwide, AST for M. tuberculosis (AST-MTB) is still mainly performed according to the resources available rather than the drug-resistance rates. Phenotypic methods, i.e. culture-based AST, are commonly used in high-income countries to confirm susceptibility of new cases of tuberculosis. They are also used to detect resistance in tuberculosis cases with risk factors, in combination with genotypic tests. In low-income countries, genotypic methods screening hot-spot mutations known to confer resistance were found to be easier to perform because they avoid the culture and biosafety constraint. Given that genotypic tests can rapidly detect the prominent mechanisms of resistance, such as the rpoB mutation for rifampicin resistance, we are facing new challenges with the observation of false-resistance (mutations not conferring resistance) and false-susceptibility (mutations different from the common mechanism) results. Phenotypic and genotypic approaches are therefore complementary for obtaining a high sensitivity and specificity for detecting drug resistances and susceptibilities to accurately predict MDR/XDR cure and to gather relevant data for resistance surveillance. Although AST-MTB was established in the 1960s, there is no consensus reference method for MIC determination against which the numerous AST-MTB techniques can be compared. This information is necessary for assessing in vitro activity and setting breakpoints for future anti-tuberculosis agents

On dispersive energy transport and relaxation in the hopping regime

A new method for investigating relaxation phenomena for charge carriers hopping between localized tail states has been developed. It allows us to consider both charge and energy {\it dispersive} transport. The method is based on the idea of quasi-elasticity: the typical energy loss during a hop is much less than all other characteristic energies. We have investigated two models with different density of states energy dependencies with our method. In general, we have found that the motion of a packet in energy space is affected by two competing tendencies. First, there is a packet broadening, i.e. the dispersive energy transport. Second, there is a narrowing of the packet, if the density of states is depleting with decreasing energy. It is the interplay of these two tendencies that determines the overall evolution. If the density of states is constant, only broadening exists. In this case a packet in energy space evolves into Gaussian one, moving with constant drift velocity and mean square deviation increasing linearly in time. If the density of states depletes exponentially with decreasing energy, the motion of the packet tremendously slows down with time. For large times the mean square deviation of the packet becomes constant, so that the motion of the packet is ``soliton-like''.Comment: 26 pages, RevTeX, 10 EPS figures, submitted to Phys. Rev.

Characterisation of the mycobacterial NER system reveals novel functions of uvrD1 helicase

In this study, we investigated the role of the nucleotide excision repair (NER) pathway in mycobacterial DNA repair. Mycobacterium smegmatis lacking the NER excinuclease component uvrB, the helicase uvrD1 and a double knock-out lacking both proteins were constructed and their sensitivity to a series of DNA damaging agents wa analysed. As anticipated, the mycobacterial NER system was shown to be involved in the processing of bulky DNA adducts and inter-strand cross-links. In addition, it could be shown to exert a protective effect against oxidising and nitrosating agents. Interestingly, inactivation of uvrB and uvrD1 significantly increased marker integration frequencies in gene conversion assays. This implies that in mycobacteria, which lack the postreplicative mismatch repair system, NER, and particularly the UvrD1 helicase, is involved in the processing of a subset of recombination-associated mismatches

Revisiting susceptibility testing in MDR-TB by a standardized quantitative phenotypic assessment in a European multicentre study

Objectives Treatment outcome of MDR-TB is critically dependent on the proper use of second-line drugs as per the result of in vitro drug susceptibility testing (DST). We aimed to establish a standardized DST procedure based on quantitative determination of drug resistance and compared the results with those of genotypes associated with drug resistance. Methods The protocol, based on MGIT 960 and the TB eXiST software, was evaluated in nine European reference laboratories. Resistance detection at a screening drug concentration was followed by determination of resistance levels and estimation of the resistance proportion. Mutations in 14 gene regions were investigated using established techniques. Results A total of 139 Mycobacterium tuberculosis isolates from patients with MDR-TB and resistance beyond MDR-TB were tested for 13 antituberculous drugs: isoniazid, rifampicin, rifabutin, ethambutol, pyrazinamide, streptomycin, para-aminosalicylic acid, ethionamide, amikacin, capreomycin, ofloxacin, moxifloxacin and linezolid. Concordance between phenotypic and genotypic resistance was >80%, except for ethambutol. Time to results was short (median 10 days). High-level resistance, which precludes the therapeutic use of an antituberculous drug, was observed in 49% of the isolates. The finding of a low or intermediate resistance level in 16% and 35% of the isolates, respectively, may help in designing an efficient personalized regimen for the treatment of MDR-TB patients. Conclusions The automated DST procedure permits accurate and rapid quantitative resistance profiling of first- and second-line antituberculous drugs. Prospective validation is warranted to determine the impact on patient car

Ac hopping conduction at extreme disorder takes place on the percolating cluster

Simulations of the random barrier model show that ac currents at extreme disorder are carried almost entirely by the percolating cluster slightly above threshold; thus contradicting traditional theories contributions from isolated low-activation-energy clusters are negligible. The effective medium approximation in conjunction with the Alexander-Orbach conjecture leads to an excellent analytical fit to the universal ac conductivity with no nontrivial fitting parameters

Hopping Conductivity of a Nearly-1d Fractal: a Model for Conducting Polymers

We suggest treating a conducting network of oriented polymer chains as an anisotropic fractal whose dimensionality D=1+\epsilon is close to one. Percolation on such a fractal is studied within the real space renormalization group of Migdal and Kadanoff. We find that the threshold value and all the critical exponents are strongly nonanalytic functions of \epsilon as \epsilon tends to zero, e.g., the critical exponent of conductivity is \epsilon^{-2}\exp (-1-1/\epsilon). The distribution function for conductivity of finite samples at the percolation threshold is established. It is shown that the central body of the distribution is given by a universal scaling function and only the low-conductivity tail of distribution remains $\epsilon$-dependent. Variable range hopping conductivity in the polymer network is studied: both DC conductivity and AC conductivity in the multiple hopping regime are found to obey a quasi-1d Mott law. The present results are consistent with electrical properties of poorly conducting polymers.Comment: 27 pages, RevTeX, epsf, 5 .eps figures, to be published in Phys. Rev.

Quantum creep and variable range hopping of one-dimensional interacting electrons

The variable range hopping results for noninteracting electrons of Mott and Shklovskii are generalized to 1D disordered charge density waves and Luttinger liquids using an instanton approach. Following a recent paper by Nattermann, Giamarchi and Le Doussal [Phys. Rev. Lett. {\bf 91}, 56603 (2003)] we calculate the quantum creep of charges at zero temperature and the linear conductivity at finite temperatures for these systems. The hopping conductivity for the short range interacting electrons acquires the same form as for noninteracting particles if the one-particle density of states is replaced by the compressibility. In the present paper we extend the calculation to dissipative systems and give a discussion of the physics after the particles materialize behind the tunneling barrier. It turns out that dissipation is crucial for tunneling to happen. Contrary to pure systems the new metastable state does not propagate through the system but is restricted to a region of the size of the tunneling region. This corresponds to the hopping of an integer number of charges over a finite distance. A global current results only if tunneling events fill the whole sample. We argue that rare events of extra low tunneling probability are not relevant for realistic systems of finite length. Finally we show that an additional Coulomb interaction only leads to small logarithmic corrections.Comment: 15 pages, 3 figures; references adde

Dielectric quantification of conductivity limitations due to nanofiller size in conductive powders and nanocomposites

Conducting submicron particles are well-suited as filler particles in non-conducting polymer matrices to obtain a conducting composite with a low percolation threshold. Going to nanometer-sized filler particles imposes a restriction to the conductivity of the composite, due to the reduction of the density of states involved in the hopping process between the particles, compared to its value within the crystallites. We show how those microscopic parameters that govern the charge-transport processes across many decades of length scales, can accurately and consistently be determined by a range of dielectric-spectroscopy techniques from a few Hz to infrared frequencies. The method, which is suited for a variety of systems with restricted geometries, is applied to densely packed 7-nm-sized tin-oxide crystalline particles with various degree of antimony doping and the quantitative results unambiguously show the role of the nanocrystal charging energy in limiting the hopping process.Comment: 6 pages, 4 figure