Identification of Defined Sequences in Domain V of E.coli 23S rRNA in the 50S Subunit Accessible for Hybridization with Complementary Oligodeoxyribonucleotides

The accessibility of specific sequences in domain V of E. coli 23s rRNA in the 50S subunit to complementary oligodeoxyribonucleotides (cDNA) has been investigated. The apparent percentage of subunits engaged in complex formation was determined by incubation of radiolabeled cDNA probe with 50S subunits, followed by nitrocellulose membrane filtration of the reaction mixtures and measurement of the bound radiolabeled cDNA probes by liquid scintillation counting of the filters. The site(s) of hybridization were determined by digestion of the RNA in the RNA/DNA heteroduplex by RNase H. The results of this study indicated that single-stranded sequences, 2058-2062, 2448-2454, 2467-2483, and 2497-2505 were available for hybridization to cDNA probes. Bases 2489-2496, which have been postulated to be base paired with 2455-2461 were also accessible for hybridization

Identification of a Protein in Several Borrelia Species which is Related to OspC of the Lyme Disease Spirochetes.

Using oligonucleotide probes which have previously been shown to be specific for the ospC gene found in the Lyme disease spirochete species Borrelia burgdorferi, B. garinii, and group VS461, we detected an ospC homolog in other Borrelia species including B. coriaceae, B. hermsii, B. anserina, B. turicatae, and B. parkeri. In contrast to the Lyme disease spirochetes, which carry the ospC gene on a 26-kb circular plasmid, we mapped the gene in other Borrelia species to linear plasmids which varied in size among the isolates tested. Some isolates carry multiple copies of the gene residing on linear plasmids of different sizes. The analyses conducted here also demonstrate that these Borrelia species contain a linear chromosome. Northern (RNA) blot analyses demonstrated that the gene is transcriptionally expressed in all species examined. High levels of transcriptional expression were observed in some B. hermsii isolates. Transcriptional start site analyses revealed that the length of the untranslated leader sequence was identical to that observed in the Lyme disease spirochete species. By Western blotting (immunoblotting) with antiserum (polyclonal) raised against the OspC protein of B. burgdorferi, we detected an immunoreactive protein of the same molecular weight as the OspC found in Lyme disease spirochete species. The results presented here demonstrate the presence of a protein that is genetically and antigenically related to OspC which is expressed in all species of the genus Borrelia tested

Analysis of the Distribution and Molecular Heterogeneity of the ospD Gene among the Lyme Disease Spirochetes: Evidence for Lateral Gene Exchange

Analysis of the ospD gene has revealed that this gene is not universal among Lyme disease spirochete isolates. The gene was found to be carried by 90, 50, and 24% of the Borrelia garinii, B. afzelii, and B. burgdorferi isolates tested. Size variability in the ospD-encoding plasmid was also observed. Sequence analysis has demonstrated the presence of various numbers of a 17-bp repeated sequence in the upstream control (promoter) region of the gene. In addition, a region within the coding sequence where various insertions, deletions, and direct repeats occur was identified. ospD gene sequences from 31 different isolates were determined and utilized in pairwise sequence comparisons and construction of a gene tree. These analyses suggest that the ospD gene was the target of several recombinational events and that the gene was recently acquired by Lyme disease spirochetes and laterally transferred between species

Coadministration of lopinavir/ritonavir and rifampicin in HIV and tuberculosis co-infected adults in South Africa

In HIV-infected patients receiving rifampicin-based treatment for tuberculosis (TB), the dosage of lopinavir/ritonavir (LPV/r) is adjusted to prevent sub-therapeutic lopinavir concentrations. In this setting, South African clinicians were advised to administer super-boosted LPV/r (400 mg/400 mg) twice daily, instead of standard dosed LPV/r (400 mg/100 mg) twice daily. We sought to determine--in routine practice--the tolerability and HIV treatment outcomes associated with super-boosted LPV/r compared to unadjusted LPV/r in combination with rifampicin-based TB treatment

Analysis of Mechanisms Associated with Loss of Infectivity of Clonal Populations of Borrelia burgdorferi B31MI

Numerous studies have provided suggestive evidence that the loss of plasmids correlates with the loss of infectivity of the Lyme disease spirochetes. In this study we have further investigated this correlation. Clonal populations were obtained from the skin of a mouse infected for 3 months with a clonal population of Borrelia burgdorferi B31MI. The complete plasmid compositions of these populations were determined using a combination of PCR and Southern hybridization. The infectivities of clones differing in plasmid composition were tested using the C3H-HeJ murine model for Lyme disease. While several clones were found to be noninfectious, a correlation between the loss of a specific plasmid and loss of infectivity in the clones analyzed in this report was not observed. While it is clear from recent studies that the loss of some specific plasmids results in attenuated virulence, this study demonstrates that additional mechanisms also contribute to the loss of infectivity

Comparison of effectiveness of cefovecin, doxycycline, and amoxicillin for the treatment of experimentally induced early Lyme borreliosis in dogs

Background While Koch’s postulates have been fulfilled for Lyme disease; causing transient fever, anorexia and arthritis in young dogs; treatment of sero-positive dogs, especially asymptomatic animals, remains a topic of debate. To complicate this matter the currently recommended antibiotic treatments of Lyme Disease in dogs caused by Borrelia burgdorferi require daily oral administrations for 31 days or longer, which makes non-compliance a concern. Additionally, there is no approved veterinary antimicrobial for the treatment of Lyme Disease in dogs in the USA and few recommended treatments have been robustly tested. In vitro testing of cefovecin, a novel extended-spectrum cephalosporin, demonstrated inhibition of spirochete growth. A small pilot study in dogs indicated that two cefovecin injections two weeks apart would be as efficacious against B. burgdorferi sensu stricto as the recommended treatments using doxycycline or amoxicillin daily for 31 days. This hypothesis was tested in 17–18 week old Beagle dogs, experimentally infected with B. burgdorferisensu stricto, using wild caught ticks, 75 days prior to antimicrobial administration. Results Clinical observations for lameness were performed daily but were inconclusive as this characteristic sign of Lyme Disease rarely develops in the standard laboratory models of experimentally induced infection. However, each antibiotic tested was efficacious against B. burgdorferi as measured by a rapid elimination of spirochetes from the skin and reduced levels of circulating antibodies to B. burgdorferi. In addition, significantly less cefovecin treated animals had Lyme Disease associated histopathological changes compared to untreated dogs. Conclusions Convenia was efficacious against B. burgdorferi sensu stricto infection in dogs as determined by serological testing, PCR and histopathology results. Convenia provides an additional and effective treatment option for Lyme Disease in dogs

Outcomes After Virologic Failure of First-Line ART in South Africa

Article approval pendingTo determine initial 24-week outcomes among prospectively enrolled patients with failure of initial antiretroviral therapy (ART)

The VMC survey - XI : Radial Stellar Population Gradients in the Galactic Globular Cluster 47 Tucanae

Copyright American Astronomical SocietyWe present a deep near-infrared color-magnitude diagram of the Galactic globular cluster 47 Tucanae, obtained with the Visible and Infrared Survey Telescope for Astronomy (VISTA) as part of the VISTA near-infrared Y, J, Ks survey of the Magellanic System (VMC). The cluster stars comprising both the subgiant and red giant branches exhibit apparent, continuous variations in color-magnitude space as a function of radius. Subgiant branch stars at larger radii are systematically brighter than their counterparts closer to the cluster core; similarly, red-giant-branch stars in the cluster's periphery are bluer than their more centrally located cousins. The observations can very well be described by adopting an age spread of ~0.5 Gyr as well as radial gradients in both the cluster's helium abundance (Y) and metallicity (Z), which change gradually from (Y = 0.28, Z = 0.005) in the cluster core to (Y = 0.25, Z = 0.003) in its periphery. We conclude that the cluster's inner regions host a significant fraction of second-generation stars, which decreases with increasing radius; the stellar population in the 47 Tuc periphery is well approximated by a simple stellar population.Peer reviewe

The VMC Survey -- XXXV. Model fitting of LMC Cepheid light curves

We present the results of the light curve model fitting technique applied to optical and near-infrared photometric data for a sample of 18 Classical Cepheids (11 fundamentals and 7 first overtones) in the Large Magellanic Cloud (LMC). We use optical photometry from the OGLE III database and near--infrared photometry obtained by the European Southern bservatory public survey "VISTA near--infrared survey of the Magellanic Clouds system". Iso--periodic nonlinear convective model sequences have been computed for each selected Cepheid in order to reproduce the multi--filter light curve amplitudes and shape details. The inferred individual distances provide an intrinsic weighted mean value for the LMC distance modulus of $\mu_0=18.56$ mag with a standard deviation of 0.13 mag. We derive also the Period--Radius, the Period--Luminosity and the Period--Wesenheit relations that are consistent with similar relations in the literature. The intrinsic masses and luminosities of the best--fitting models show that all the investigated pulsators are brighter than the redictions of the canonical evolutionary mass--luminosity relation, suggesting a significant efficiency of non--canonical phenomena, such as overshooting, mass loss and/or rotation.Comment: 14 pages, 10 figures, MNRAS accepte

Outer membrane protein a conservation among Orientia tsutsugamushi isolates suggests its potential as a protective antigen and diagnostic target

Scrub typhus threatens one billion people in the Asia-Pacific area and cases have emerged outside this region. It is caused by infection with any of the multitude of strains of the bacterium; Orientia tsutsugamushi; . A vaccine that affords heterologous protection and a commercially-available molecular diagnostic assay are lacking. Herein, we determined that the nucleotide and translated amino acid sequences of outer membrane protein A (OmpA) are highly conserved among 51; O. tsutsugamushi; isolates. Molecular modeling revealed the predicted tertiary structure of; O. tsutsugamushi; OmpA to be very similar to that of the phylogenetically-related pathogen,; Anaplasma phagocytophilum; , including the location of a helix that contains residues functionally essential for; A. phagocytophilum; infection. PCR primers were developed that amplified; ompA; DNA from all; O. tsutsugamushi; strains, but not from negative control bacteria. Using these primers in quantitative PCR enabled sensitive detection and quantitation of; O. tsutsugamushi ompA; DNA from organs and blood of mice that had been experimentally infected with the Karp or Gilliam strains. The high degree of OmpA conservation among; O. tsutsugamushi; strains evidences its potential to serve as a molecular diagnostic target and justifies its consideration as a candidate for developing a broadly-protective scrub typhus vaccine