Tuberculosis vaccine strain _Mycobacterium bovis_ BCG Russia is a natural _recA_ mutant

The current tuberculosis vaccine is a live vaccine derived from _Mycobacterium bovis_ and attenuated by serial _in vitro_ passaging. All vaccine substrains in use stem from one source, strain Bacille Calmette-Guérin. However, they differ in regions of genomic deletions, antigen expression levels, immunogenicity, and protective efficacy. As a RecA phenotype increases genetic stability and may contribute restricting the ongoing evolution of the various BCG substrains, we aimed to inactivate _recA_ by allelic replacement in BCG vaccine strains representing different phylogenetic lineages (Pasteur, Frappier, Denmark, Russia). Homologous gene replacement was successful in three out of four strains. However, only illegitimate recombination was observed in BCG substrain Russia. Sequence analyses of _recA_ revealed that a single nucleotide insertion in the 5' part of _recA_ led to a translational frameshift with an early stop codon making BCG Russia a natural _recA_ mutant. At the protein level BCG Russia failed to express RecA. According to phylogenetic analyses BCG Russia is an ancient vaccine strain most closely related to the parental _M. bovis_. Our data suggest that _recA_ inactivation in BCG Russia occurred early and is in part responsible for its high degree of genomic stability, resulting in a substrain that has less genetic alterations than other vaccine substrains with respect to _M. bovis_ AF2122/97 wild type

High-level resistance to isoniazid and ethionamide in multidrug-resistant Mycobacterium tuberculosis of the Lisboa family is associated with inhA double mutations

Objectives The purpose of this study was to determine the levels of isoniazid and ethionamide resistance and to identify associated mutations in endemic multidrug-resistant (MDR) strains of Mycobacterium tuberculosis from the Lisbon metropolitan area, Portugal. Methods Seventeen clinical MDR tuberculosis (TB) strains were characterized by standard and semi-quantitative drug susceptibility testing to assess the level of isoniazid and ethionamide resistance. The genes katG, inhA, ethA and ndh were screened for mutations. All strains were genotyped by 24 loci mycobacterial interspersed repetitive unit-variable number of tandem repeats (MIRU-VNTR) analysis. Results All strains showed high-level resistance to both isoniazid (>1 mg/L) and ethionamide (>25 mg/L). MIRU-VNTR typing revealed the presence of two main clusters, Lisboa3 and Q1, in 16/17 strains, all of which showed the C−15T mutation in the promoter region of the inhA gene. The 16 strains belong to the Latino-American-Mediterranean (LAM) genotype and the other strain belongs to the Beijing genotype. Sequencing of the inhA open reading frame revealed that the 16 strains also had mutations in the structural region of the gene, leading to the S94A substitution in 9 strains and the I194T substitution in 7 strains. Conclusions The results reveal that the presence of a mutation in the inhA regulatory region together with a mutation in the inhA coding region can lead to the development of high-level isoniazid resistance and cross-resistance to ethionamide among the MDR-TB strains circulating in Lisbon. This mutational pattern also hints to a possible involvement of strain-specific factors that could be a feature of the Portuguese MDR-TB strains where the LAM family is the major circulating genotyp

Conditioned Pain Modulation Is Associated with Common Polymorphisms in the Serotonin Transporter Gene

BACKGROUND: Variation in the serotonin transporter (5-HTT) gene (SLC6A4) has been shown to influence a wide range of affective processes. Low 5-HTT gene-expression has also been suggested to increase the risk of chronic pain. Conditioned pain modulation (CPM)--i.e. 'pain inhibits pain'--is impaired in chronic pain states and, reciprocally, aberrations of CPM may predict the development of chronic pain. Therefore we hypothesized that a common variation in the SLC6A4 is associated with inter-individual variation in CPM. Forty-five healthy subjects recruited on the basis of tri-allelic 5-HTTLPR genotype, with inferred high or low 5-HTT-expression, were included in a double-blind study. A submaximal-effort tourniquet test was used to provide a standardized degree of conditioning ischemic pain. Individualized noxious heat and pressure pain thresholds (PPTs) were used as subjective test-modalities and the nociceptive flexion reflex (NFR) was used to provide an objective neurophysiological window into spinal processing. RESULTS: The low, as compared to the high, 5-HTT-expressing group exhibited significantly reduced CPM-mediated pain inhibition for PPTs (p = 0.02) and heat-pain (p = 0.02). The CPM-mediated inhibition of the NFR, gauged by increases in NFR-threshold, did not differ significantly between groups (p = 0.75). Inhibition of PPTs and heat-pain were correlated (Spearman's rho = 0.35, p = 0.02), whereas the NFR-threshold increase was not significantly correlated with degree of inhibition of these subjectively reported modalities. CONCLUSIONS: Our results demonstrate the involvement of the tri-allelic 5-HTTLPR genotype in explaining clinically relevant inter-individual differences in pain perception and regulation. Our results also illustrate that shifts in NFR-thresholds do not necessarily correlate to the modulation of experienced pain. We discuss various possible mechanisms underlying these findings and suggest a role of regulation of 5-HT receptors along the neuraxis as a function of differential 5-HTT-expression

Mitochondrial Ribosome as the Target for the Macrolide Antibiotic Clarithromycin in the Helminth Echinococcus multilocularis

The mitochondrial rRNA of the tapeworm species Echinococcus multilocularis carries an adenine at sequence position 2058 (numbering according to that for Escherichia coli) of the large-subunit rRNA (lsrRNA), while the nucleus-encoded rRNA, as determined in this study, is characterized by 2058G. This indicates a dichotomy in the drug susceptibilities of ribosomes: cytoplasmic ribosomes are predicted to be resistant to macrolide antibiotics, while mitochondrial ribosomes lack the most common chromosomal resistance determinant, lsrRNA 2058G. Upon incubation with the macrolide clarithromycin, the formation of vesicles from metacestode tissue was reduced in a dose-dependent manner. Electron microscopy revealed distinct morphological alterations both of the mitochondria and of the vesicle wall (e.g., loss of microtriches) in drug-treated vesicles. Adult worms lost their motility and displayed morphological changes (shortening and constriction of proglottids and the presence of vacuoles) upon incubation with clarithromycin. Our findings demonstrate that macrolides have distinct in vitro effects on E. multilocularis, endorsing the use of sequence-based in silico approaches for exploitation of available ribosomal drugs as anthelmintic agents

High-level resistance to isoniazid and ethionamide in multidrug-resistant Mycobacterium tuberculosis of the Lisboa family is associated with inhA double mutations

OBJECTIVES: The purpose of this study was to determine the levels of isoniazid and ethionamide resistance and to identify associated mutations in endemic multidrug-resistant (MDR) strains of Mycobacterium tuberculosis from the Lisbon metropolitan area, Portugal. METHODS: Seventeen clinical MDR tuberculosis (TB) strains were characterized by standard and semi-quantitative drug susceptibility testing to assess the level of isoniazid and ethionamide resistance. The genes katG, inhA, ethA and ndh were screened for mutations. All strains were genotyped by 24 loci mycobacterial interspersed repetitive unit-variable number of tandem repeats (MIRU-VNTR) analysis. RESULTS: All strains showed high-level resistance to both isoniazid (>1 mg/L) and ethionamide (>25 mg/L). MIRU-VNTR typing revealed the presence of two main clusters, Lisboa3 and Q1, in 16/17 strains, all of which showed the C-15T mutation in the promoter region of the inhA gene. The 16 strains belong to the Latino-American-Mediterranean (LAM) genotype and the other strain belongs to the Beijing genotype. Sequencing of the inhA open reading frame revealed that the 16 strains also had mutations in the structural region of the gene, leading to the S94A substitution in 9 strains and the I194T substitution in 7 strains. CONCLUSIONS: The results reveal that the presence of a mutation in the inhA regulatory region together with a mutation in the inhA coding region can lead to the development of high-level isoniazid resistance and cross-resistance to ethionamide among the MDR-TB strains circulating in Lisbon. This mutational pattern also hints to a possible involvement of strain-specific factors that could be a feature of the Portuguese MDR-TB strains where the LAM family is the major circulating genotype

Erratum: ``A Measurement of the Cosmic Microwave Background B-Mode Polarization Power Spectrum at Sub-degree Scales with POLARBEAR'' (2014, ApJ, 794, 171)

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POLARBEAR constraints on cosmic birefringence and primordial magnetic fields

We constrain anisotropic cosmic birefringence using four-point correlations of even-parity E-mode and odd-parity B-mode polarization in the cosmic microwave background measurements made by the POLARization of the Background Radiation (POLARBEAR) experiment in its first season of observations. We find that the anisotropic cosmic birefringence signal from any parity-violating processes is consistent with zero. The Faraday rotation from anisotropic cosmic birefringence can be compared with the equivalent quantity generated by primordial magnetic fields if they existed. The POLARBEAR nondetection translates into a 95% confidence level (C.L.) upper limit of 93 nanogauss (nG) on the amplitude of an equivalent primordial magnetic field inclusive of systematic uncertainties. This four-point correlation constraint on Faraday rotation is about 15 times tighter than the upper limit of 1380 nG inferred from constraining the contribution of Faraday rotation to two-point correlations of B-modes measured by Planck in 2015. Metric perturbations sourced by primordial magnetic fields would also contribute to the B-mode power spectrum. Using the POLARBEAR measurements of the B-mode power spectrum (two-point correlation), we set a 95% C.L. upper limit of 3.9 nG on primordial magnetic fields assuming a flat prior on the field amplitude. This limit is comparable to what was found in the Planck 2015 two-point correlation analysis with both temperature and polarization. We perform a set of systematic error tests and find no evidence for contamination. This work marks the first time that anisotropic cosmic birefringence or primordial magnetic fields have been constrained from the ground at subdegree scales. \ua9 2015 American Physical Society