Additional file 2: of Development of a bead-based Luminex assay using lipopolysaccharide specific monoclonal antibodies to detect biological threats from Brucella species

Specificity of the developed bead-based Luminex immunoassay for B. anthracis, F. tularensis and Y. pestis. (PDF 170 kb

Influence of Chlorinating Agents on the Formation of Stable Biomarkers in Hair for the Retrospective Verification of Exposure

Chlorine, as a dual-use chemical, is an essential industrial chemical which has been used as a chemical weapon in the past due to its toxicity and availability. The retrospective verification of chlorine intoxication is often especially challenging, and unambiguous markers are still missing. In this study, the effects of different chlorinating and oxidizing agents on human hair were investigated. Samples were exposed to a variety of chlorinating chemicals for a short time and then completely hydrolyzed by a HBr solution to break down their keratin proteins into individual amino acids. After derivatization and targeted liquid chromatography-mass spectrometry analysis, 3-chlorotyrosine and 3,5-dichlorotyrosine were unambiguously identified from human hair exposed to chlorine, hypochlorite, and sulfuryl chloride. Our results show long-term stability of these markers in the biological matrix, as the chlorotyrosines can still be found 10 months post-exposure at the same levels. Finally, an untargeted analysis was able to discriminate between some of the different intoxicants

Laboratory colonies included to build the shrinkage discriminant analysis (SDA) models.

1<p>Model 1: SDA model to discriminate between members of the <i>A. gambiae</i> species complex.</p>2<p>Model 2: SDA model to discriminate between M and S molecular forms within <i>A. gambiae</i> s.s.</p>3<p>Model 3: SDA model to classify specimens to their colony of a single species, <i>i.e. A. arabiensis</i>.</p

Model selection and cross-validation for colony authentication in <i>Anopheles arabiensis</i> (Model 3).

<p>(<b>A</b>) Error rate as a function of the number of peaks included in the SDA model for five <i>A. arabiensis</i> colonies and the total error rate over all colonies. The peaks were ranked according to the correlation-adjusted <i>t</i>-scores (CAT scores). The vertical, red line shows the 23 peaks chosen for the SDA model (<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0057486#pone.0057486.s004" target="_blank">Table S4</a>). (<b>B</b>) Top 23 peaks included in SDA model after they were ranked according to CAT scores (<i>i.e.</i> peak with highest CAT score appears at the top of the list). The length and direction of the horizontal blue bars represents the CAT scores of the centroid versus the pooled mean and shows the influence of a particular peak in differentiating between the colonies.</p

Dendrogram of hierarchical, unsupervised clustering of binary peaks (presence/absence).

<p>While the <i>Anopheles</i> species (complexes) are well separated by the cluster algorithm, the sibling species of the <i>A. gambiae</i> complex (coloured lines) do not segregate into well defined clusters. Specimens, both from the same species and colony, are split into different groups. The external branches represent each measured specimen. For each colony spectra from 10 specimens were recorded and included in the cluster analysis. The labels give the names of the colonies (<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0057486#pone-0057486-t001" target="_blank">Table 1</a>). The length of the branches corresponds to the size of the Dice similarity coefficient.</p

Examples of MALDI-TOF MS spectra for <i>Anopheles gambiae</i> sensu stricto and <i>A. arabiensis</i>.

<p>Examples of representative MALDI-TOF MS spectra measured from 3 <i>A. arabiensis</i> (blue) and 3 <i>A. gambiae</i> s.s. (red) colonies. The spectra were taken from crude suspensions of heads and thoraces in SA solution. The vertical, dashed lines indicate peaks that are characteristic (but not exclusive) for one or the other species. The left panels show the whole spectra between 2 and 14 kDa, while the right panels zoom into two peaks. The two peaks are separated by only a few Daltons. While the left peak is more common in <i>A. arabiensis</i>, the right peak is more common in <i>A. gambiae</i> s.s. In this representation the peak intensities were normalised against the highest intensity measured in each spectrum.</p

Model selection and cross-validation to discriminate between species of the <i>Anopheles gambiae</i> complex (Model 1).

<p>(<b>A</b>) The graph shows the error rate from the cross-validation plotted as a function of the number of the ranked peaks included in the SDA model that discriminates between members of the <i>A. gambiae</i> species complex. The peaks were ranked (left to right) according to the correlation-adjusted <i>t</i>-scores (CAT scores). The vertical, red line shows the 68 peaks chosen for the SDA model. (<b>B</b>) List with the 68 ranked peaks (top equals highest rank) their corresponding CAT scores. The length and direction of the horizontal blue bars represents the CAT scores of the centroid versus the pooled mean and show the influence of a particular peak in differentiating between the groups (<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0057486#pone.0057486.s002" target="_blank">Table S2</a>). For example the top peak, M12369.6 has a strong influence in separating <i>A. merus</i> from all the other species, emphasised by the length of the bar and the opposite direction from the bars of the other species. In contrast, the tenth peak, M12527.3 has a stronger influence in separating <i>A. gambiae</i> s.s. from <i>A. arabiensis</i>. AR: <i>A. arabiensis</i>; GA: <i>A. gambiae</i> s.s.; ME: <i>A. merus</i>; QD: <i>A. quadriannulatus</i>.</p

Total number of peaks versus number of diagnostic peaks present in average spectra from the <i>Anopheles gambiae</i> species complex.

<p>The number of diagnostic peaks present is associated with the number of total peaks present in an average peak list. The diagnostic peaks refer to the 68 selected peaks to distinguish within the <i>A. gambiae</i> species complex (Model 1). The plot suggests that the field specimens collected by aspiration (green crosses) were generally of lower quality (<i>i.e.</i> showing fewer peaks) than the specimens that were raised from the larvae; regardless whether from laboratory (black circles) or field caught larvae (red triangles).</p

Laboratory colonies included in the MALDI-TOF MS analysis.

1<p>MR4: Malaria Research and Reference Reagent Center, VA, USA. Numbers in brackets are MR4 reference numbers; IRD: Institut de Recherche pour le Développement, Montpellier, France; Swiss TPH: Swiss Tropical and Public Health Institute, Basel, Switzerland. Numbers in brackets indicate the MR4 catalogue number.</p

Additional file 1: of Complete genome sequence of “Thiodictyon syntrophicum” sp. nov. strain Cad16T, a photolithoautotrophic purple sulfur bacterium isolated from the alpine meromictic Lake Cadagno

Figure S1. Phylogenetic placement of “T. syntrophicum” strain Cad16T within the other 12 Chromatiaceae species with a publicly available whole genome sequences. Additionally, the closely related phylogenetic lineages Nitrosococcus, Rheinheimera and Arsukibacterium are also included. Strain Cad16T is most closely related to L. purpurea DSM 4197. The maximum likelihood tree was inferred from 100 concatenated single-copy orthologues sequences [61] and a total of 1000 bootstrap replicates were performed. Numbers at the nodes indicate the SH-aLRT support (%) and ultrafast bootstrap support (%). OrthoMCL [64], was used to define at set orthologues proteins between these 23 species. Hundred single-copy orthologues were randomly chosen and aligned with MUSCLE [66] . The best-fit phylogenetic model and subsequent consensus tree computation, based on maximum-likelihood and 1000 bootstrap iterations, was performed with the IQ-TREE software [62]. Nodes with both, 100% SH-aLRT and ultrafast bootstrap support, are indicated with filled black circle symbols for convenience. (TIF 57220 kb