Characterization of Overwintering Sites of the Invasive Brown Marmorated Stink Bug in Natural Landscapes Using Human Surveyors and Detector Canines

<div><p><i>Halyomorpha halys</i> is an invasive species from Asia causing major economic losses in agricultural production in the mid-Atlantic region of the United States. Unlike other crop pests, <i>H. halys</i> is also well-known for nuisance problems in urban, suburban, and rural areas, as massive numbers of adults often invade human-made structures to overwinter inside protected environments. Research efforts have focused on populations in human-made structures while overwintering ecology of <i>H. halys</i> in natural landscapes is virtually unknown. We explored forested landscapes in the mid-Atlantic region to locate and characterize natural overwintering structures used by <i>H. halys.</i> We also evaluated the use of detector canines to locate overwintering <i>H. halys</i> to enhance the accuracy and efficiency of surveys. From these studies, we indentified shared characteristics of overwintering sites used by <i>H. halys</i> in natural landscapes. Overwintering <i>H. halys</i> were recovered from dry crevices in dead, standing trees with thick bark, particularly oak (<i>Quercus</i> spp.) and locust (<i>Robinia</i> spp.); these characteristics were shared by 11.8% of all dead trees in surveyed landscapes. For trees with favorable characteristics, we sampled ∼20% of the total above-ground tree area and recovered 5.9 adults per tree from the trees with <i>H. halys</i> present. Two detector canines were successfully trained to recognize and detect the odor of adult <i>H. halys</i> yielding >84% accuracy in laboratory and semi-field trials. Detector canines also found overwintering <i>H. halys</i> under field conditions. In particular, overwintering <i>H. halys</i> were recovered only from dead trees that yielded positive indications from the canines and shared key tree characteristics established by human surveyors. The identified characteristics of natural overwintering sites of <i>H. halys</i> will serve as baseline information to establish crop economic risk levels posed by overwintering populations, and accordingly develop sustainable management programs.</p></div

Buparlisib inhibits proliferation of pediatric sarcoma cell lines.

<p>(<b>A</b>) Cells were treated with media containing 0.1% DMSO or concentrations of buparlisib ranging from 10 nM to 10 μM for 72 hours. Cell viability was determined by MTT assay. Percent cell viability was plotted against log buparlisib concentration and IC50 values were calculated by fitting this data to a four-parameter, variable slope sigmoid dose-response model. Each point is the average of at least three independent experiments. (<b>B</b>) Cell viability IC50 values for pediatric sarcoma cell lines. Columns represent the average of at least three independent experiments, error bars represent 95% confidence intervals. Dashed line indicates median IC50. (<b>C</b>) Scatter plot showing the low correlation (R² = 0.07311) between cell viability and phospho-Akt IC50 values. The cell lines that possess the lowest (A204) and highest (A4573) phospho-Akt IC50s are indicated on the graph. Dot colors indicate sarcoma type: gray—ES, blue—OS, burgundy—RMS.</p

Buparlisib demonstrates increased synergy with rapamycin in PTEN deficient cell lines.

<p>(<b>A</b>) PTEN wild type (TC-32, A673, RD) or PTEN null (A4573) cell lines were exposed to a series of 1.5-fold dilutions of buparlisib and rapamycin alone or in combination at a constant ratio of 20:3 (TC-32, A673, RD) or 100:3 (A4573) for 72 hours, then cell viability was determined by MTT assay. Columns represent the average of three independent experiments, error bars represent standard deviation. Combination index values greater than 1, equal to 1, or less than one indicate antagonism, additivity, or synergy. (<b>B</b>) Isobologram plot of the effect of buparlisib combined with rapamycin. The effective doses of rapamycin and buparlisib are plotted on the x- and y-axis with lines of linear additivity connecting the ED₅₀, ED₇₅, and E₉₀ for individual treatments. Points for combination treatment above, on, or below the lines indicate antagonism, additivity, or synergy, respectively. (<b>C</b>) Immunoblot analysis of phospho-Akt (S473), total Akt, phospho-4E-BP1 (T37/46), total 4E-BP1, and actin in A4573 cells after 1 hour of treatment with buparlisib, rapamycin, or both. (<b>D</b>) Immunoblot analysis of phospho-Akt (S473), total Akt, phospho-4E-BP1 (T37/46), total 4E-BP1, cleaved PARP, and actin in A4573 cells after 24 hours of treatment with buparlisib, rapamycin, or both.</p

outgroup_alignment

The outgroup_alignment folder contains the alignment of the heterothallic species described in the paper in fasta format for each chromosome. The coordinates follow the 2509 N. tetrasperma mat a reference genome

Evaluation of <i>In Vitro</i> Activity of the Class I PI3K Inhibitor Buparlisib (BKM120) in Pediatric Bone and Soft Tissue Sarcomas

<div><p>Pediatric bone and soft tissue sarcomas often display increased Akt phosphorylation through up regulation of insulin-like growth factor (IGF1) signaling. Additionally, Akt signaling has been linked to resistance to IGF1 receptor (IGF1R) and mTOR (mammalian target of rapamycin) inhibitors in sarcoma, further demonstrating the role of Akt in tumor survival. This suggests targeting components of the PI3K/Akt pathway may be an effective therapeutic strategy. Here, we investigated the <i>in vitro</i> activity of the pan-class I PI3K inhibitor buparlisib (BKM120) in pediatric bone and soft tissue sarcomas. Buparlisib inhibited activation of Akt and signaling molecules downstream of mTORC1 (mTOR complex 1) in Ewing sarcoma, osteosarcoma, and rhabdomyosarcoma cell lines. Anti-proliferative effects were observed in both anchorage dependent and independent conditions and apoptosis was induced within 24 hours of drug treatment. Buparlisib demonstrated cytotoxicity as a single agent, but was found to be more effective when used in combination. Synergy was observed when buparlisib was combined with the IGF1R inhibitor NVP-AEW541 and the mTORC1 inhibitor rapamycin. The addition of NVP-AEW541 also further reduced phospho-Akt levels and more potently induced apoptosis compared to buparlisib treatment alone. Additionally, the combination of buparlisib with the MEK1/2 inhibitor trametinib resulted in synergy in sarcoma cell lines possessing MAPK pathway mutations. Taken together, these data indicate buparlisib could be a novel therapy for the treatment of pediatric bone and soft tissue sarcomas.</p></div

L9 VCF

The L9_vcf.tar.gz folder contains the compressed all-sites VCF file for L9 and the index file for that VCF file. The all-sites VCF file was generated using GATK UnifiedGenotyper, as described in the paper

L8 VCF - scaffold 3 to 7

The L8_vcf_scaffold_3_to_7.tar.gz folder contains the compressed all-sites VCF file for L8 and the index file for that VCF file. This file contains scaffold 3 to 7. The all-sites VCF file was generated using GATK UnifiedGenotyper, as described in the paper

L10 VCF - scaffold 3 to 7

The L10_vcf_scaffold_3_to_7.tar.gz folder contains the compressed all-sites VCF file for L10 and the index file for that VCF file. This file contains scaffold 3 to 7. The all-sites VCF file was generated using GATK UnifiedGenotyper, as described in the paper

Re-activation of Akt and induction of Erk occur within 24 hours of buparlisib treatment.

<p>(<b>A</b>) Immunoblot analysis of phospho-Akt (S473), total Akt, phospho-Erk1/2 (T202/T204 on Erk1, T185/T187 on Erk2), total Erk, and actin in ES (TC-174, A4573), OS (KHOS), and RMS (RD) cells after treatment with buparlisib for periods of 5 minutes to 24 hours. TC-174, KHOS, and RD cells were treated with 1 μM buparlisib. A4573 cells were treated with 3 μM buparlisib. (<b>B</b>) Immunoblot analysis of phospho-Akt (S473), total Akt, phospho-Erk1/2 (T202/T204 on Erk1, T185/T187 on Erk2), total Erk, and actin in KHOS cells after treatment with 3 μM buparlisib for periods of 5 minutes to 24 hours. (<b>C</b>) Immunoblot analysis of phospho-Akt (S473), total Akt, phospho-Erk1/2 (T202/T204 on Erk1, T185/T187 on Erk2), and total Erk in TC-32 and A4573 cells with increasing concentrations of buparlisib for 24 hours. (<b>D</b>) Immunoblot analysis of phospho-STAT3 (Y705), total STAT3, phospho-Akt (S473), total Akt, phospho-Erk1/2 (T202/T204 on Erk1, T185/T187 on Erk2), total Erk, and actin in TC-174 and A4573 parental cells and cells passaged in increasing concentrations of buparlisib. Phospho-Akt levels were quantitated based on Odyssey software integrated intensity values. Values are listed below each band.</p

Fig5 alignments

The Fig5_alignments folder contains a fasta format alignment gene alignments of the 6 SR and 2 PAR genes listed in Supplementary Table 12 and used in the reconstruction of the Fig5 phylogenetic trees