A reforma do pensamento: por uma educação que descubra as relações ocultas do saber

Reforming education involves reforming thought, and a reform of thought implies a paradigm shift in the current knowledge organization model. Complex thinking, interdisciplinary and transdisciplinary research methods and recognition of the relationship between knowledge, the knower and the surrounding medium are elements that can contribute to the construction of educational models that address the problems faced by the world nowadays. A holistic view in education contributes and commits the educator and the learner with society and its problems.Reformar la educación implica reformar el pensamiento y una reforma del pensamiento implica un cambio de paradigma en el modelo actual de organización del conocimiento. El pensamiento complejo, los métodos de investigación inter y transdisciplinar y el reconocimiento de la relación entre el saber, el sujeto cognoscente y el medio que lo rodea son elementos que pueden contribuir a la construcción de modelos educativos que respondan a los problemas que enfrenta el mundo actual. Una visión holística dentro de la educación contribuye y compromete al educador y al educando con la sociedad y sus problemáticas.Reformar a educação implica reformar o pensamento e uma reforma do pensamento implica uma mudança de paradigma no modelo atual de organização do conhecimento. O pensamento complexo, os métodos de pesquisa inter e transdisciplinar e o reconhecimento da relação entre o saber, o sujeito cognoscente e o meio que o rodeia são elementos que podem contribuir para a construção de modelos educativos que respondam aos problemas enfrentados pelo mundo atual. Uma visão holística dentro da educação contribui e compromete o educador e o educando com a sociedade e suas problemáticas

Expression of ζY83C toxin enhances the efficacy of different antimicrobials during exponential growth.

<p>BG689 cells were grown in MMS7 at 37°C up to ∼5×10⁷ cells/ml, then 0.5% Xyl and/or an antimicrobial (AM) were added and the cultures were incubated for 120 min A) or 240 min B) and then plated onto LB agar plates. The antimicrobials used were Amp, 3 µg/ml; Ery, 20 µg/ml; Cip, 4 µg/ml or Tri, 3 µg/ml. The number of CFUs relative to the non-induced/non-AM treated control is shown. + and − denote the presence or the absence of the indicated compound. Error bars show 95% confidence intervals of more than three independent experiments.</p

Expression of ζY83C toxin enhances the efficacy of different antimicrobials in high- and low-density non-growing cells.

<p>BG689 cells were grown in MMS7 at 37°C up to early stationary phase and diluted into fresh pre-warmed MMS7 to ∼1×10⁹ cells/ml (A and B) or to ∼1×10⁶ cells/ml (C and D). Then 0.5% Xyl and/or an AM were added and the cultures were incubated for 120 min (A and C) or 240 min (B and D). The symbols, the plating conditions, and the antimicrobial concentrations are as described in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0086615#pone-0086615-g001" target="_blank">Figure 1</a>. Error bars show 95% confidence intervals of more than three independent experiments.</p

RelA is required for ζY83C toxin enhanced efficacy to different antimicrobials.

<p>A) BG1145 cells (Δ<i>relA</i>) were grown to ∼5×10⁷ cells/ml. Then 0.5% Xyl and/or an AM were added and the cultures were incubated for 120 min. B) BG1145 cells were pre-treated with limiting relacin (1 mM) concentrations (+ Rel) or not (− Rel), and then 0.5% Xyl, Amp or both were added and the cultures were incubated for 120 min. The symbols, the plating conditions, and the antimicrobial concentrations were those indicated in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0086615#pone-0086615-g001" target="_blank">Figure 1</a>. Error bars show 95% confidence intervals of more than three independent experiments.</p

Effect of a chromosomal-encoded TA locus on the formation of antimicrobial persisters.

<p>BG689 (ζY83C <i>mazF</i>⁺) and BG1243 (ζY83C Δ<i>mazF</i>) cells were grown in MMS7 to ∼5×10⁷ cell ml⁻¹. Then 0.5% Xyl^a or 2x-MIC Amp^b or both was added and the culture was incubated for 120 min.</p>c<p>Induction or not of the ζY83C toxin is indicated by + and − superscript symbols, respectively.</p>d<p>The CFUs were measured after 120 min of toxin induction and/or antimicrobial addition by plating appropriate dilutions on LB plates. The results are the average of at least three independent experiments and are within a 10% standard error.</p

Expression of the ε₂ antitoxin reverses the effect of ζ toxin, but does not increase MDT.

<p>(A and B) BG1125-borne ζ gene was induced by the addition of 1 mM IPTG, and pCB799-borne ε gene was induced by addition of Xyl. BG1125 cells were grown to ∼5×10⁷ cells/ml in MMS7 containing traces of Xyl (0.005%). Then, expression of ζ was induced and/or cells treated with an antimicrobial, and the cultures incubated for 120 min (A) or 240 min (B). At those times, samples were taken and plated on LB agar or on LB-0.5% Xyl plates to induce the expression of the ε₂ antitoxin. Expression of the ε₂ antitoxin reverses the effect of ζ toxin in non-growing cells. (C and D) BG1125 cells were grown in MMS7, containing traces of Xyl (0.005%), up to early stationary phase, and diluted to ∼1×10⁹ cells/ml. Then, expression of <b>ζ</b> was induced and/or cells treated with an antimicrobial, and the cultures incubated for 120 min (C) or 240 min (D) with agitation at 37°C. At these times samples were taken and plated on LB agar or LB-0.5% Xyl to induce the expression of the ε₂ antitoxin. The symbols, the plating conditions, and the antimicrobial concentrations were those indicated in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0086615#pone-0086615-g001" target="_blank">Figure 1</a>. Error bars show 95% confidence intervals of more than three independent experiments.</p

G<i>44</i>P-mediated cleavage of replicated D-loops.

<p>Different D-loop variants resembling several recombination intermediates were end-labeled at the 5′end of oligonucleotide 19-M (the invading strand, in A) or of oligonucleotide 17-M (the displaced strand, in B) and were incubated with 10 nM G<i>44</i>P in buffer B containing 10 mM MgCl₂ for 30 min at 37°C. Reaction products were analyzed using 20% denaturing PAGE and revealed by autoradiography. Drawings indicate the different substrates analyzed (D-loops A to F, and control HJ and ssDNA). Asterisks indicate the [γ³²P]-ATP labeling of oligonucleotides at the 5′end. In the HJ substrate, an arrow indicates the major cleavage site. As markers, the G+A sequencing ladder obtained for the corresponding labeled oligonucleotide and the 41-nt and 21-nt primers for the corresponding sequence were loaded.</p

Determination of the cleavage ability of G<i>44</i>P on static and mobile HJs.

<p>(A and C) A fixed HJ (HJ-23M, in A) or a mobile HJ containing a 13-bp homologous core (HJ-jbm6, in C) [γ³²P]-labeled at the indicated strand was incubated with 10 nM G<i>44</i>P in buffer B containing 10 mM MgCl₂ for 30 min at 37°C. Reaction products were analyzed using 15% denaturing PAGE in the presence (+) or absence (-) of protein. “m” indicates the G+A sequencing ladder obtained for the corresponding labeled oligonucleotide. To serve as additional molecular weight markers, and denoted by C, 41-nt, 21-nt, 23-nt and 18-nt primers were loaded. In lane 13 of panel A, a degraded 17-M oligonucleotide was loaded. (B and D) The cleavage sites detected are indicated by arrows in the core of the two HJ sequences.</p

Characterization of the Holliday Junction Resolving Enzyme Encoded by the <em>Bacillus subtilis</em> Bacteriophage SPP1

<div><p>Recombination-dependent DNA replication, which is a central component of viral replication restart, is poorly understood in Firmicutes bacteriophages. Phage SPP1 initiates unidirectional theta DNA replication from a discrete replication origin (<em>ori</em>L), and when replication progresses, the fork might stall by the binding of the origin binding protein G<em>38</em>P to the late replication origin (<em>ori</em>R<em>).</em> Replication restart is dependent on viral recombination proteins to synthesize a linear head-to-tail concatemer, which is the substrate for viral DNA packaging. To identify new functions involved in this process, uncharacterized genes from phage SPP1 were analyzed. Immediately after infection, SPP1 transcribes a number of genes involved in recombination and replication from <em>P</em>_E2 and <em>P</em>_E3 promoters. Resequencing the region corresponding to the last two hypothetical genes transcribed from the <em>P</em>_E2 operon (genes <em>44</em> and <em>45</em>) showed that they are in fact a single gene, re-annotated here as gene <em>44</em>, that encodes a single polypeptide, named gene <em>44</em> product (G<em>44</em>P, 27.5 kDa). G<em>44</em>P shares a low but significant degree of identity in its C-terminal region with virus-encoded RusA-like resolvases. The data presented here demonstrate that G<em>44</em>P, which is a dimer in solution, binds with high affinity but without sequence specificity to several double-stranded DNA recombination intermediates. G<em>44</em>P preferentially cleaves Holliday junctions, but also, with lower efficiency, replicated D-loops. It also partially complemented the loss of RecU resolvase activity in <em>B. subtilis</em> cells. These <em>in vitro</em> and <em>in vivo</em> data suggest a role for G<em>44</em>P in replication restart during the transition to concatemeric viral replication.</p> </div

G<i>44</i>P binding to different DNA substrates.

<p>EMSAs showing binding of G<i>44</i>P to the indicated [γ³²P]-labeled DNA substrates: (A) HJ-J3, (B) D-loop DL-D, (C) 80-bp dsDNA, and (D) 80-nt ssDNA. DNA (0.2 nM) was incubated with increasing amounts of G<i>44</i>P as indicated in buffer B containing 1 mM EDTA for 20 min at 37°C. The three types of complexes formed are denoted by I, II, and III.</p