The Addition of Transcriptomics to the Bead-Enabled Accelerated Monophasic Multi-Omics Method: A Step toward Universal Sample Preparation.

Omics-based measurements enable the study of biomolecules in a high-throughput fashion, leading to the characterization and quantification of biological systems. Multi-omics methods aim to incorporate several omics measurements for a more holistic approach, which is crucial for advancing our understanding of the diversity and redundancy of biological systems. Current multi-omics sample preparation methods have achieved proteomics, lipidomics, and metabolomics from individual samples; however, the bioinformatic tools currently available for interpreting data generated from these omics are limited. Alternately, transcriptomics has a wide arsenal of available bioinformatic tools offering intensive sample characterization but has yet to be incorporated into a unified, multi-omics sample preparation technique. Herein we describe the modified bead-enabled accelerated monophasic multi-omics (mBAMM) method, which incorporates RNA extraction for transcriptomics analysis. mBAMM was shown to enable RNA-seq without compromising the isolation of biomolecules for proteomics, lipidomics, and metabolomics. This methodology greatly improves sample characterization and represents a major innovation toward cohesive insights into biological systems.

Exposure to and Biomarker Responses From Legacy and Emerging Contaminants Along Three Drainages in the Milwaukee Estuary, Wisconsin, USA.

Legacy contaminants and contaminants of emerging concern (CECs) were assessed in tree swallow (Tachycineta bicolor) tissue and diet samples from three drainages in the Milwaukee estuary, Wisconsin, USA, to understand exposures and possible biomarker responses. Two remote Wisconsin lakes were assessed for comparative purposes. Bioaccumulative classes of contaminants, such as polybrominated diphenyl ethers and per- and polyfluoroalkyl substances, while at higher concentrations than the reference lakes, did not vary significantly among sites or among the three drainages. Polycyclic aromatic hydrocarbons were assessed in diet and sediment and were from primarily pyrogenic sources. Ten biomarkers were assessed relative to contaminant exposure. Polychlorinated biphenyls (PCBs) were elevated above reference conditions at all Milwaukee sites but did not correlate with any measured biomarker responses. Only one site, Cedarburg, just downstream from a Superfund site, had elevated PCBs compared to other sites in the Milwaukee estuary. Few non-organochlorine insecticides or herbicides were detected in tree swallow liver tissue, except for the atrazine metabolite desethylatrazine. Few pharmaceuticals and personal care products were detected in liver tissue except for N,N-diethyl-meta-toluamide, iopamidol, and two antibiotics. The present study is one of the most comprehensive assessments to date, along with the previously published Maumee River data, on the exposure and effects of a wide variety of CECs in birds. Environ Toxicol Chem 2024;43:856-877. © 2024 SETAC. This article has been contributed to by U.S. Government employees and their work is in the public domain in the USA.

Feature-agnostic metabolomics for determining effective subcytotoxic doses of common pesticides in human cells.

Although classical molecular biology assays can provide a measure of cellular response to chemical challenges, they rely on a single biological phenomenon to infer a broader measure of cellular metabolic response. These methods do not always afford the necessary sensitivity to answer questions of subcytotoxic effects, nor do they work for all cell types. Likewise, boutique assays such as cardiomyocyte beat rate may indirectly measure cellular metabolic response, but they too, are limited to measuring a specific biological phenomenon and are often limited to a single cell type. For these reasons, toxicological researchers need new approaches to determine metabolic changes across various doses in differing cell types, especially within the low-dose regime. The data collected herein demonstrate that LC-MS/MS-based untargeted metabolomics with a feature-agnostic view of the data, combined with a suite of statistical methods including an adapted environmental threshold analysis, provides a versatile, robust, and holistic approach to directly monitoring the overall cellular metabolomic response to pesticides. When employing this method in investigating two different cell types, human cardiomyocytes and neurons, this approach revealed separate subcytotoxic metabolomic responses at doses of 0.1 and 1 µM of chlorpyrifos and carbaryl. These findings suggest that this agnostic approach to untargeted metabolomics can provide a new tool for determining effective dose by metabolomics of chemical challenges, such as pesticides, in a direct measurement of metabolomic response that is not cell type-specific or observable using traditional assays.

Multi-omics responses in tree swallow (Tachycineta bicolor) nestlings from the Maumee Area of Concern, Maumee River, Ohio.

A multi-omics approach was utilized to identify altered biological responses and functions, and to prioritize contaminants to assess the risks of chemical mixtures in the Maumee Area of Concern (AOC), Maumee River, OH, USA. The Maumee AOC is designated by the United States Environmental Protection Agency as having significant beneficial use impairments, including degradation of fish and wildlife populations, bird or animal deformities or reproduction problems, and loss of fish and wildlife habitat. Tree swallow (Tachycineta bicolor) nestlings were collected at five sites along the Maumee River, which included wastewater treatment plants (WWTPs) and industrial land-use sites. Polychlorinated biphenyls (PCBs), polybrominated diphenyl ethers (PBDEs), polycyclic aromatic hydrocarbons (PAHs), polychlorinated dibenzo p dioxins and furans (PCDD/Fs), and chlorinated pesticide concentrations were elevated in Maumee tree swallows, relative to a remote reference site, Star Lake, WI, USA. Liver tissue was utilized for non-targeted transcriptome and targeted metabolome evaluation. A significantly differentially expressed gene cluster related to a downregulation in cell growth and cell cycle regulation was identified when comparing all Maumee River sites with the reference site. There was an upregulation of lipogenesis genes, such as PPAR signaling (HMGCS2, SLC22A5), biosynthesis of unsaturated fatty acids (FASN, SCD, ELOVL2, and FADS2), and higher lipogenesis related metabolites, such as docosapentaenoic acid (DPA), docosahexaenoic acid (DHA), eicosapentaenoic acid (EPA), and arachidonic acid (AA) at two industrial land-use sites, Ironhead and Maumee, relative to WWTP sites (Perrysburg and SideCut), and the reference site. Toledo Water, in the vicinity of the other two industrial sites and also adjacent to a WWTP, showed a mix of signals between industrial land-use and WWTP land-use. PAHs, oxychlordane, and PBDEs were determined to be the most likely causes of the differentiation in biological responses, including de novo lipogenesis and biosynthesis of unsaturated fatty acids.

Legacy and Contaminants of Emerging Concern in Tree Swallows Along an Agricultural to Industrial Gradient: Maumee River, Ohio.

Exposure to multiple classes of contaminants, both legacy and contaminants of emerging concern (CECs), were assessed in tree swallow (Tachycineta bicolor) tissue and diet samples from 6 sites along the Maumee River, Ohio, USA, to understand both exposure and possible effects of exposure to those CECs for which there are little avian data. The 6 sites represented a gradient from intensive agriculture upstream to highly urbanized and industrial landscapes downstream; 1 or 2 remote Wisconsin lakes were assessed for comparative purposes. Cytochrome P450 induction, DNA damage, and thyroid function were also assessed relative to contaminant exposure. Bioaccumulative CECs, such as polybrominated diphenyl ethers (PBDEs) and perfluorinated substances, did not follow any upstream to downstream gradient; but both had significantly greater concentrations along the Maumee River than at the remote lake sites. Greater exposure to PBDEs was apparent in swallows at or near wastewater-treatment facilities than at other sites. Total polychlorinated biphenyl and total polycyclic aromatic hydrocarbon concentrations were greater in swallows at downstream locations compared to upstream sites and were associated with higher ethoxyresorufin-O-dealkylase activity. Few herbicides or nonorganochlorine insecticides were detected in swallow tissues or their food, except for atrazine and its metabolite desethylatrazine. Few pharmaceuticals and personal care products were detected except for DEET and iopamidol. Both were detected in most liver samples but not in eggs, as well as detected at the remote lake sites. This is one of the most comprehensive assessments to date of exposure and effects of a wide variety of CECs in birds. Environ Toxicol Chem 2020;39:1936-1952. © 2020 SETAC. This article has been contributed to by US Government employees and their work is in the public domain in the USA.

Toxicity assessments of nanoscale zerovalent iron and its oxidation products in medaka (Oryzias latipes) fish.

Iron-based nanotechnologies are increasingly used for environmental remediation; however, toxicologic impacts of iron nanoparticles on the aquatic ecosystem remain poorly understood. We treated larvae of medaka fish (Oryzias latipes) with thoroughly characterized solutions containing carboxymethyl cellulose (CMC)-stabilized nanoscale zerovalent iron (nZVI), aged nanoscale iron oxides (nFe-oxides) or ferrous ion (Fe[II]) for 12-14 days' aqueous exposure to assess the causal toxic effect(s) of iron NPs on the fish. With the CMC-nZVI solution, the dissolved oxygen level decreased, and a burst of reactive oxygen species (ROS) was generated as Fe(II) oxidized to ferric ion (Fe[III]); with the other two iron solutions, these parameters did not significantly change. CMC-nZVI and Fe(II) solutions caused acute lethally and sublethally toxic effects in medaka larvae, with nFe-oxide-containing solutions causing the least toxic effects. We discuss modes of toxic action of iron NPs and chronic toxic effects in terms of hypoxia, Fe(II) toxicity and ROS-mediated oxidative damage.