HISA big data in biomedicine and healthcare 2013 conference

Additional file 5. Biofilm formation by the S. suis serotype 2 (S2) and serotype 9 (S9) wild-type and agI/II -deficient mutant strains in the absence of porcine fibrinogen. Biofilm formation capacity was quantified after 24 h of incubation at 37 °C in the absence of porcine fibrinogen. Data represent the mean ± SEM from at least three independent experiments

Comparison of CAMP factors and Uberis factor found in streptococci.

<p>A. multiple sequence alignment (<i>Sag</i>, <i>S. agalactiae</i>; <i>S. urinalis</i>; <i>S. canis</i>; <i>S. pyogenes</i>; <i>S. porcinus</i>; <i>S. pseudo porcinus</i> and <i>S. uberis</i>); B. phylogenetic tree showing the evolutionary relationships between the sequences of the alignment. Sequence alignment and construction of the phylogenetic tree were done using the AlignX module of VectorNTI advance 11 (InVitrogen). Conserved residues appear in light grey and identical amino acids appear in dark grey in the alignment. Position of residues in the sequence is indicated above the sequence. Sequence identities go from 56% (CAMP factor II of <i>S. agalactiae</i> and Uberis factor of <i>S. uberis</i>) to 100% (CAMP factor II of <i>S. agalactiae</i> and CAMP factor of <i>S. urinalis</i>). The CAMP factor of <i>P. acnes</i> is more distant (less than 30% of identity) and thus does not appear in this alignment. The phylogenetic tree has been constructed using the Neighbor Joining Method. Each branch of the tree has a length equal to the number of substitutions required to get from one nod to the next.</p

Impact of <i>MoPACC</i> deletion on the fungus biology.

<p>(a) Conidia production in the parental (P), control (C2 (ectopic)) and mutant (T2) strains grown for 14 days on solid rice medium buffered to pH 5 (grey) or to pH 8 (black). (b) Pathogenicity tests on barley leaves infected by conidia from the parent, mutant and complemented strains. Lesions on 10 separate leaves were counted after 5 days of incubation. All experiments were run in triplicates and standard deviations are shown.</p

Expression of CAMP factor II in pathogenic streptococcal strains.

<p>A. CAMP test using (1) <i>S. bovis</i> 1052 wt, (2) <i>S. bovis</i> (pOri23-camp⁵¹⁵), (3) <i>S. dysgalactiae</i> 593 wt, (4) <i>S. dysgalactiae</i> (pOri23-camp⁵¹⁵) and (5) GBS strain 515 as control; B. co-hemolytic activity was monitored in <i>S. bovis</i> 1052 (filled circle with pOri23-camp⁵¹⁵ and empty circle without) and in <i>S. dysgalactiae</i> (filled squares with pOri23-camp⁵¹⁵ and empty squares without treatment). Hemolytic activity was measured at OD₆₃₀ every 30 min using a microplate reader. The experiment was done in triplicate using three independent biological samples. Errors bars represent the standard deviation observed between the 9 values obtained for each strain. Controls without SMase treatment were carried out (data not shown).</p

Co-hemolytic activity of CAMP factor II expressed in <i>L. lactis</i> MG1363.

<p>A. CAMP test using (1) <i>L. lactis</i> MG1363 (pOri23), (2) <i>L. lactis</i> MG1363 (pOri23-camp⁵¹⁵) and (3) GBS strain 515 as control; B. <i>L. lactis</i> co-hemolytic activity was monitored in <i>L. lactis</i> MG1363 (pOri23) (filled circle with SMase treatment and empty circle without treatment) and in <i>L. lactis</i> MG1363 (pOri23-camp⁵¹⁵) (filled squares with SMase treatment and empty squares without treatment). Hemolytic activity was measured at OD₆₃₀ every 30 min using a microplate reader. The experiment was done in triplicate using three independent biological samples. Errors bars represent the standard deviation observed between the 9 values obtained for each strain. Controls without SMase treatment were done (data not shown).</p

Influence of pH on the biology of <i>M. oryzae</i>.

<p>(a) The wild type strain Guy11 was grown for 14 and 21 days on solid rice medium buffered to pH 5 (grey) or to pH 8 (black) and conidia from one 9 mm plate were collected and counted under a microscope; Student test analysis of the data revealed a Pvalue of 0,004 at 14 days and of 0.07 at 21 days, due to higher variance. (b) One hundred conidia collected from cultures on non-buffered solid rice medium were incubated for 8 hours in water adjusted to pH 3 to 8, and their germination was monitored by microscopic observation of the presence of a germ tube at least 3 µm in length. (c) Conidia collected from cultures on solid rice medium buffered to pH 5 (grey) or pH 8 (black) were sprayed onto rice or barley plants, and the lesions on 10 separate leaves were counted after 5 days of incubation; Student test analysis of the data revealed a Pvalue of 0.078 for the barley experiment and of 0.079 for the rice experiment. All experiments were run in triplicates and standard deviations are shown.</p

Schematic diagram of the ICE_<i>515_tRNA^Lys</i> mobile genetic element.

<p>ORFs appear as arrows. Genes encoding the putative bacteriocin system appear in light blue. The genes encoding the proteins that could be involved in oxidative stress response (NRAMP protein and thioredoxin-like) are colored in yellow and the gene encoding the putative new hemolytic CAMP factor (CAMP factor II) in purple. Genes encoding the proteins with LPxTG motif appear in pink. Genes of the conjugation module are indicated with blue arrows and regulation module with green arrows. Putative <i>oriT</i> is indicated by a star. Genes encoding a putative toxin-antitoxin system appear in orange and other genes encoding proteins with unknown function in white. The gene where the element is integrated (tRNA^Lys) is indicated in red. Recombination module is colored in red. Recombination sites are drawn as vertical rectangles. Black rectangles indicate identical sequences found in <i>attL</i>, <i>attR</i>, and <i>attI</i> sites; yellow rectangles indicate the arm of <i>attR</i> sites and the related arm of <i>attI</i> sites; and red rectangles indicate the arm of <i>attL sites</i> and the related arms of <i>attI</i> sites.</p

Analysis of the expression of the CAMP factor II (SAL_2074) gene.

<p>RT-PCR were performed using, as templates, RNA extracted from stationary-phase cultures of strain 515 (1, and negative control without RTase, 4); strain NEM316 (2 and negative control without RTase, 5); and transconjugant NEM316 (ICE_<i>515_tRNA^Lys</i>) (3 and negative control without RTase, 6). The same results were obtained in exponential phase. A positive control using genomic DNA of strain 515 was included (7) as well as a negative control with water (8). DNA molecular weight marker is marker number VI (Roche Applied Science).</p

Results of the complementation studies.

<p>Growth and conidiation <i>in vitro</i> were measured for the complemented strain Δ<i>PACC+PACC</i> under the same conditions used for the study of the Δ<i>PACC</i> mutant strain (<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0069236#pone-0069236-g005" target="_blank">Fig. 5</a>).</p

<i>MoPACC</i> and <i>MoPAL</i> genes.

<p>The <i>M. oryzae PAL</i> and <i>PACC</i> genes are presented. Introns were searched using the Softberry software and both their number (bold) and positions (start-end) are given. The length of the corresponding predicted proteins and the identity of these proteins to their <i>A. nidulans</i> counterparts is provided.</p