Data_Sheet_1_Heterologous prime-boost BCG with DNA vaccine expressing fusion antigens Rv2299c and Ag85A improves protective efficacy against Mycobacterium tuberculosis in mice.docx

The development of heterologous prime-boost regimens utilizing Bacille Calmette–Guerin (BCG) as the priming vaccine is a promising approach to improve the efficacy of vaccination against tuberculosis (TB). In this study, we examined the ability of a DNA vaccine that expressed a fusion of antigens Rv2299c and Ag85A to boost BCG immunity and protection against Mycobacterium tuberculosis (Mtb) in Balb/c mice. The fusion DNA vaccine was moderately immunogenic and afforded some protection when used on its own. After a priming BCG vaccination, the DNA boost significantly amplified Th1-type cell-mediated immunity compared to that resulting from either BCG or DNA immunization. In the DNA-boosted mice, Ag-specific CD4+ and CD8+ T cells that were mono-positive for IFN-γ alone were the most prominently expanded in infected lungs. The protective efficacy afforded by BCG against challenge infection was greatly improved by the DNA boost; bacterial loads were significantly reduced in both spleen and lung and histological damage in the lung was less. The use of a DNA vaccine containing the fusion antigens Rv2299c and Ag85A to boost BCG may be a good choice for the rational design of an efficient vaccination strategy against TB.</p

<i>Mtb</i> MazG is required for resistance to RNS shown <i>in vitro</i> and <i>ex vivo</i>.

<p>(<b>A</b>) <i>mazG</i>-null <i>Mtb</i> is susceptible to killing by acid nitrite. Exponential phase cultures (OD₆₀₀∼0.5) were suspended in 7H9-OADC pH4.5 with or without 2.5 mM NaNO₂ and treated for 20 hours. wt, wild-type <i>Mtb</i>; Δ<i>mazG</i>, <i>mazG</i>-null <i>Mtb</i>; compl, the complemented mutant. Data shown are mean ± S.E. of triplicates. * <i>p</i><0.05 <i>vs</i> wt. (<b>B</b>) Quantitative real-time PCR analyses of <i>mazG</i> and <i>dosR</i> from <i>Mtb</i> treated with DETA/NO or acid nitrite. All data were normalized to the levels of <i>sigA</i>. The <i>dosR</i> gene was used as a positive control <a href="http://www.plospathogens.org/article/info:doi/10.1371/journal.ppat.1003814#ppat.1003814-Voskuil1" target="_blank">[70]</a>. Results are expressed as the changes in expression levels compared to untreated samples. Mean ± S.E of three independent repeats. <b>C to E</b>, Survival of wild-type <i>Mtb</i>, the <i>mazG</i>-null mutant and the complemented mutant in resting (<b>C</b>), activated (<b>D</b>) or NMMA treated (<b>E</b>) cells of murine macrophage cell line RAW264.7. Shown is one representative result of two independent experiments.</p

Akupressur - Komplement till traditionell terapi vid postoperativt illamående och kräkningar

Postoperativt illamående och kräkningar, PONV, har sedan anestesins begynnelse vållat patienten stort lidande. Tillståndet kan förlänga den postoperativa vistelsen och är en av de vanligaste orsakerna till oplanerad inläggning av patienter i samband med dagkirurgi. Syftet med föreliggande arbete var att undersöka den förebyggande effekten av akupressur som ensam intervention eller i kombination med viss farmakologisk antiemetisk terapi vid postoperativt illamående och kräkningar hos vuxna patienter efter allmänkirurgi. En systematisk litteraturstudie genomfördes och tio vetenskapliga artiklar inkluderades och kvalitetsbedömdes. Sökning utfördes i databaserna PubMed, EBSCO HOST och Cochrane Library. Dessutom genomfördes manuell sökning. Resultatet visade att akupressur på en speciell triggerpunkt, P6, har en förebyggande effekt mot PONV. Ondansetron och akupressur är lika effektivt mot PONV, medan Droperidol verkar ha bättre effekt än akupressur. Slutsatsen var att akupressur har en plats som profylaktisk antiemetika för att förebygga PONV.Postoperative nausea and vomiting (PONV) has since the beginning of anesthesia caused the patient great suffering. It can prolong the post-operative stay and is one of the most common reasons for unplanned admittance of patients in connection to day surgery. The aim of this work was to study the preventive effect of acupressure as the lone intervention or in combination with certain pharmacological antiemetic therapies for post-operative nausea and vomiting in adult patients who have undergone general surgery. A systematic literature review was conducted and ten articles were included and each study subjected to a quality assessment. A PubMed, EBSCO HOST and Cochrane Library Database were conducted and a manual search of the literature references completed the search. The results showed that acupressure at the P6 meridian point has a preventive effect against PONV and that Ondansetron and acupressure are similarly effective against PONV, while Droperidal seems to have a better effect than acupressure. The conclusion was that acupressure can be used prophylactic to prevent PONV

The antimutator role of MazG in <i>Msm</i> (A) and <i>Mtb</i> (B).

<p>Both the bacterial culture conditions and the methods for determination of mutation frequencies were illustrated in <a href="http://www.plospathogens.org/article/info:doi/10.1371/journal.ppat.1003814#s4" target="_blank"><b><i>Materials and Methods</i></b></a> in detail. The frequencies conferring resistance to rifampicin in wild-type (wt), <i>mazG</i>-null (Δ<i>mazG</i>) and the complemented mutant (compl) strains were determined in exponential phase (OD₆₀₀∼0.5) with or without oxidative stress and in the stationary growth phase. Oxidative stress was induced by treating exponential phase cultures with 10 mM H₂O₂ for 5 h (<i>Msm</i>) or 24 h (<i>Mtb</i>). Stationary phase was at the 5^th-day or 28^th-day of culture for <i>Msm</i> or <i>Mtb</i>, respectively. (<b>C</b>) Survival rate of <i>Msm</i> and <i>Mtb</i> strains after exposure to H₂O₂. The numbers shown are mean ± S.E. of 3 independent experiments totaling 15 cultures of <i>Msm</i> and 6 of <i>Mtb</i>.</p

Kinetic constants of <i>Mtb</i> MazG.

<p>Kinetic constants of <i>Mtb</i> MazG.</p

MazG is required for <i>Mtb</i> survival <i>in vivo</i> and the corresponding lung pathogenesis.

<p><b>A to B</b>, Bacterial loads in spleen (<b>A</b>) and lung (<b>B</b>) of mice infected with wild-type <i>Mtb</i> (wt), the <i>mazG</i>-null mutant (Δ<i>mazG</i>) and the complemented mutant (compl). Data shown are mean ± S.E from 4 mice per group. (<b>C</b>) Lung sections taken from mice at 8-wk after infection and stained with hematoxylin and eosin. Inserted column shows mean ± S.E of lung inflammation of each group.</p

<i>mazG</i>-null <i>Msm</i> exhibited elevated CG to TA mutation under oxidative stress conditions and in the stationary phase of growth.

<p>Spontaneous rifampicin-resistant colonies were collected from 3 independent experiments (see Methods). Cluster I region of <i>rpoB</i> were PCR-amplified using <i>pfu</i> DNA polymerase and sequenced bi-directionally. All of the sequenced colonies contain single non-synonymous mutations (see also <b><a href="http://www.plospathogens.org/article/info:doi/10.1371/journal.ppat.1003814#ppat.1003814.s003" target="_blank">Table S1</a> and <a href="http://www.plospathogens.org/article/info:doi/10.1371/journal.ppat.1003814#ppat.1003814.s004" target="_blank">S2</a></b>). wt, wild-type <i>Msm</i>; Δ<i>mazG</i>, <i>mazG</i>-null <i>Msm</i>.</p

5-OH-dCTP is an <i>in vivo</i> substrate of mycobacterial MazG proved by chemical genetic analysis.

<p>The <i>Msm</i> competent cells were prepared as described in <a href="http://www.plospathogens.org/article/info:doi/10.1371/journal.ppat.1003814#s4" target="_blank"><b><i>Materials and Methods</i></b></a>, the nucleotides were incorporated by transformation. (<b>A</b>) <i>mazG</i>-null <i>Msm</i> (Δ<i>mazG</i>) exhibited a dose-dependent 5-OH-dCTP induced mutagenesis. wt, wild-type <i>Msm</i>. The data shown are mean ± S.E. of four repeats. (<b>B</b>) The rifampicin-resistance mutation frequencies of wild-type <i>Msm</i>, the <i>mazG</i>-null mutant and the complemented mutant treated with 5-OH-dCTP and dCTP. Nucleotides were added at 100 µM final concentration. Mean ± S.E of 12 independent transformations. **<i>p</i><0.01 <i>vs</i> wt, * <i>p</i><0.05 <i>vs</i> wt. (<b>C</b>) The rifampicin-resistance mutation frequencies of wild-type and <i>mazG</i>-null <i>Msm</i> treated with 100 µM 2-OH-dATP, 5-CHO-dUTP and normal dNTPs. Mean ± S.E of 8 independent transformations. (<b>D</b>) <i>mazG</i>-null <i>Msm</i> is susceptible to killing by 5-OH-dCTP. Strains were treated with 100 µM 5-OH-dCTP for 5 hours at 37°C. Shown is percent survival compared to an untreated control (100%). The data shown are mean ± S.E. of four repeats.</p