The effects of <i>BTB1</i> and <i>BTB2</i> dsRNA deliveries on the mRNA levels of <i>BTB1</i> and <i>BTB2</i>, respectively.

<p><i>BTB1</i><b>(A)</b> and <i>BTB2</i> <b>(B)</b> dsRNA deliveries resulted in significant knockdown in the mRNA levels of <i>BTB1</i> and <i>BTB2</i>, respectively, in either F10A or AO females. Student’s <i>t</i> test results for the comparison of the relative <i>BTB1</i> (or <i>BTB2</i>) mRNA levels in F10A (or AO) females that received control and <i>BTB1</i> dsRNA are <i>P</i> < 0.000001, 2-tailed <i>t</i> test. The <i>BTB1</i> (or <i>BTB2</i>) mRNA levels in control females were scaled to 1. Different letters denote significant differences.</p

The effects of <i>clathrin heavy chain</i> gene knockdown on a subsequent RNAi response as measured by <i>cathepsin L</i> knockdown in <i>M. occidentalis</i> females.

<p>(<b>A</b>) The effects of different treatment combinations on the relative mRNA levels of <i>clathrin heavy chain</i> (mean + SEM). Ingestion of <i>clathrin heavy chain</i> dsRNA resulted in a reduction of <i>clathrin heavy chain</i> mRNA levels in <i>Clathrin + Control</i> and <i>Clathrin + Cathepsin</i> mites. (<b>B</b>) The effect of different treatment combinations on the relative mRNA levels of <i>cathepsin L</i> (mean + SEM). <i>Clathrin heavy chain</i> dsRNA pretreatment reduced subsequent RNAi responses as evaluated by gene knockdown of <i>cathepsin L</i>. For <i>clathrin heavy chain</i> and <i>cathepsin L</i> mRNAs comparisons: One-way ANOVA, <i>F_3,19</i> = 470.10, <i>P</i><0.0001 and <i>F_3,19</i> = 49.16, P<0.0001, respectively, Tukey-Kramer HSD lettering for all comparisons. The mRNA levels of <i>clathrin heavy chain</i> and <i>cathepsin L</i> in <i>Control + Control</i> group are scaled to 1. Different letters denote significant differences.</p

The Glutathione-S-Transferase, Cytochrome P450 and Carboxyl/Cholinesterase Gene Superfamilies in Predatory Mite <i>Metaseiulus occidentalis</i>

<div><p>Pesticide-resistant populations of the predatory mite <i>Metaseiulus</i> (= <i>Typhlodromus</i> or <i>Galendromus) occidentalis</i> (Arthropoda: Chelicerata: Acari: Phytoseiidae) have been used in the biological control of pest mites such as phytophagous <i>Tetranychus urticae</i>. However, the pesticide resistance mechanisms in <i>M</i>. <i>occidentalis</i> remain largely unknown. In other arthropods, members of the glutathione-S-transferase (GST), cytochrome P450 (CYP) and carboxyl/cholinesterase (CCE) gene superfamilies are involved in the diverse biological pathways such as the metabolism of xenobiotics (e.g. pesticides) in addition to hormonal and chemosensory processes. In the current study, we report the identification and initial characterization of 123 genes in the GST, CYP and CCE superfamilies in the recently sequenced <i>M</i>. <i>occidentalis</i> genome. The gene count represents a reduction of 35% compared to <i>T</i>. <i>urticae</i>. The distribution of genes in the GST and CCE superfamilies in <i>M</i>. <i>occidentalis</i> differs significantly from those of insects and resembles that of <i>T</i>. <i>urticae</i>. Specifically, we report the presence of the Mu class GSTs, and the J’ and J” clade CCEs that, within the Arthropoda, appear unique to Acari. Interestingly, the majority of CCEs in the J’ and J” clades contain a catalytic triad, suggesting that they are catalytically active. They likely represent two Acari-specific CCE clades that may participate in detoxification of xenobiotics. The current study of genes in these superfamilies provides preliminary insights into the potential molecular components that may be involved in pesticide metabolism as well as hormonal/chemosensory processes in the agriculturally important <i>M</i>. <i>occidentalis</i>.</p></div

A comparison of cytosolic GST gene numbers in the genomes of nine arthropods.

<p>Data are derived from Hayes et al. [<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0160009#pone.0160009.ref073" target="_blank">73</a>], Oakeshott et al. [<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0160009#pone.0160009.ref084" target="_blank">84</a>], Grbic et al. [<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0160009#pone.0160009.ref058" target="_blank">58</a>], Reddy et al. [<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0160009#pone.0160009.ref059" target="_blank">59</a>], and the current study.</p

Domain structure and phylogenetic tree of clathrin heavy chain proteins of <i>M. occidentalis</i> and selected species.

<p>(<b>A</b>) Schematic structures of predicted clathrin heavy chain (CHC) proteins from the mites <i>M. occidentalis</i> (Mo) and <i>T. urticae</i> (Tu), the insect <i>D. melanogaster</i> (Dm), and the mammal <i>Homo sapiens</i> (Hs). Names and pfam ID of conserved domains are shown in brackets. (<b>B</b>) Phylogenetic analysis of predicted clathrin heavy chain proteins from selected species (Mo = <i>M. occidentalis</i>, Tu = <i>T. urticae</i>, Is = <i>I. scapularis</i>, Am = <i>A. mellifera</i>, Ag = <i>An. gambiae</i>, Tc = <i>T. castaneum</i>, Dm = <i>D. melanogaster</i>, Hs = <i>H. sapiens</i>, Ms = <i>M. musculus</i>, Ce = <i>C. elegans</i>). The tree was generated using a maximum likelihood approach <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0110874#pone.0110874-Stamatakis1" target="_blank">[28]</a> with bootstrap support values shown at the nodes. The tree was rooted using the clathrin heavy chain of the nematode <i>C. elegans</i> (Ce_CHC). The scale bar represents the numbers of substitutions per site.</p

A comparison of CCE gene numbers in the genomes of eight arthropods.

<p>Data are derived from Yu et al.[<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0160009#pone.0160009.ref106" target="_blank">106</a>], Oakeshott et al [<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0160009#pone.0160009.ref084" target="_blank">84</a>]. Grbic et al [<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0160009#pone.0160009.ref058" target="_blank">58</a>]. and the current study.</p

Phylogenetic relationships of the different CYP clans.

<p>The deduced amino acid sequences of 63 <i>M</i>. <i>occidentalis</i> CYP genes were aligned with those of selected CYPs from <i>D</i>. <i>melanogaster</i> (Dm), <i>A</i>. <i>mellifera</i> (Am) and <i>T</i>. <i>urticae</i> (Tu) (<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0160009#pone.0160009.s003" target="_blank">S3 Fig</a>). The midpoint-rooted tree was generated using MrBayes. The <i>M</i>. <i>occidentalis</i> CYP genes are shown in colors. Posterior probabilities are shown at the nodes. Details of the gene names for the CYPs from <i>M</i>. <i>occidentalis</i> and other arthropods are shown in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0160009#pone.0160009.s005" target="_blank">S1</a> and <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0160009#pone.0160009.s007" target="_blank">S3</a> Tables, respectively.</p

A list of clathrin heavy chain proteins in selected species.

<p>*Identifier for BOGAS database (<a href="http://bioinformatics.psb.ugent.be/orcae/overview/Tetur" target="_blank">http://bioinformatics.psb.ugent.be/orcae/overview/Tetur</a>).</p><p>Percentage identity was determined by BLASTP searches.</p><p>A list of clathrin heavy chain proteins in selected species.</p

Domain structure and phylogenetic tree of BTB1 and BTB2 proteins of <i>M</i>. <i>occidentalis</i> and their closest homologs from selected species.

<p><b>(A)</b> Schematic structures of BTB proteins from the chelicerates <i>M</i>. <i>occidentalis</i>, <i>T</i>. <i>urticae</i> (Tu), <i>I</i>. <i>scapularis</i> (Is) and <i>S</i>. <i>mimosarum</i> (Sm), the insects <i>D</i>. <i>melanogaster</i> (Dm), <i>Nasonia vitripennis</i> (Nv) and <i>A</i>. <i>aegypti</i> (Aa), and the mammal <i>H</i>. <i>sapiens</i> (Hs). Names and pfam IDs of conserved domains are shown in brackets. <b>(B)</b> A phylogenetic analysis of BTB proteins from selected species. The tree was generated using a maximum likelihood approach [<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0144291#pone.0144291.ref049" target="_blank">49</a>] with bootstrap support values shown at the nodes. The tree was rooted using BTB18 of the mammal <i>H</i>. <i>sapiens</i> (Hs_BTB18). The scale bar represents the numbers of substitutions per site.</p

A schematic diagram showing the point mutation at a conserved position in the AChEs of <i>M</i>. <i>occidentalis</i> and <i>T</i>. <i>urticae</i>.

<p>The deduced amino acid sequences of the AChEs from several species were aligned using the same method as described for other multiple sequence alignments (e.g. GSTs). A partial alignment is shown with the G-to-S point mutation in the <i>M</i>. <i>occidentalis</i> and <i>T</i>. <i>urticae</i> AChEs highlighted in bold. Numbers on the top and bottom of the alignment denote the positions of corresponding amino acid residues in the AChEs of <i>M</i>. <i>occidentalis</i> and <i>T</i>. <i>californica</i>, respectively. Species include <i>M</i>. <i>occidentalis</i> (Mo; CCE16 in this study), <i>T</i>. <i>urticae</i> (Tu; OrcAE ID: tetur19g00850), <i>I</i>. <i>scapularis</i> (Is; GenBank accession no.: XP_002413212.1), <i>Culex pipiens</i> (Cp; GenBank accession no.: AAV28503.1), <i>A</i>. <i>mellifera</i> (Am; BeeBase ID: GB18414), <i>D</i>. <i>melanogaster</i> (Dm; FlyBase ID: CG17907) and <i>T</i>. <i>californica</i> (Tc; UniProtKB ID: P04058.2).</p