Functional Toxicogenomic Assessment of Triclosan in Human HepG2 Cells Using Genome-Wide CRISPR-Cas9 Screening

There are thousands of chemicals used by humans and detected in the environment for which limited or no toxicological data are available. Rapid and cost-effective approaches for assessing the toxicological properties of chemicals are needed. We used CRISPR-Cas9 functional genomic screening to identify the potential molecular mechanism of a widely used antimicrobial triclosan (TCS) in HepG2 cells. Resistant genes at IC50 (the concentration causing a 50% reduction in cell viability) were significantly enriched in the adherens junction pathway, MAPK signaling pathway, and PPAR signaling pathway, suggesting a potential role in the molecular mechanism of TCS-induced cytotoxicity. Evaluation of the top-ranked resistant genes, <i>FTO</i> (encoding an mRNA demethylase) and <i>MAP2K3</i> (a MAP kinase kinase family gene), revealed that their loss conferred resistance to TCS. In contrast, sensitive genes at IC10 and IC20 were specifically enriched in pathways involved with immune responses, which was concordant with transcriptomic profiling of TCS at concentrations o

Indigenous species barcode database improves the identification of zooplankton

<div><p>Incompleteness and inaccuracy of DNA barcode databases is considered an important hindrance to the use of metabarcoding in biodiversity analysis of zooplankton at the species-level. Species barcoding by Sanger sequencing is inefficient for organisms with small body sizes, such as zooplankton. Here mitochondrial <i>cytochrome c oxidase I</i> (<i>COI</i>) fragment barcodes from 910 freshwater zooplankton specimens (87 morphospecies) were recovered by a high-throughput sequencing platform, Ion Torrent PGM. Intraspecific divergence of most zooplanktons was < 5%, except <i>Branchionus leydign</i> (Rotifer, 14.3%), <i>Trichocerca elongate</i> (Rotifer, 11.5%), <i>Lecane bulla</i> (Rotifer, 15.9%), <i>Synchaeta oblonga</i> (Rotifer, 5.95%) and <i>Schmackeria forbesi</i> (Copepod, 6.5%). Metabarcoding data of 28 environmental samples from Lake Tai were annotated by both an indigenous database and NCBI Genbank database. The indigenous database improved the taxonomic assignment of metabarcoding of zooplankton. Most zooplankton (81%) with barcode sequences in the indigenous database were identified by metabarcoding monitoring. Furthermore, the frequency and distribution of zooplankton were also consistent between metabarcoding and morphology identification. Overall, the indigenous database improved the taxonomic assignment of zooplankton.</p></div

Species identified by metabarcoding analysis.

<p>The size of red dots indicated the frequency of each species that detected by morphology method (A) Reads number of each species in metabarcoding data. (B) The internal arcs indicate the species found in morphological analysis. The middle arcs indicate the species that have barcode sequences in indigenous species database. The external arcs indicate which species were detected by metabarcoding. Abundant (detected frequency > 1/2), moderate (detected frequency > 1/3) and rare (detected frequency < 1/3).</p

Schematic diagram of parallel barcode recovery using a high throughput sequencing protocol.

<p>Schematic diagram of parallel barcode recovery using a high throughput sequencing protocol.</p

Ecogenomics of Zooplankton Community Reveals Ecological Threshold of Ammonia Nitrogen

Communities of zooplankton can be adversely affected by contamination resulting from human activities. Yet understanding the influence of water quality on zooplankton under field-conditions is hindered by traditional labor-intensive approaches that are prone to incomplete or uncertain taxonomic determinations. Here, for the first time, an eco-genomic approach, based on genetic diversity in the mitochondrial cytochrome c oxidase I (COI) region of DNA of zooplankton was used to develop a site-specific, water quality criterion (WQC) for ammonia (NH₃). Ammonia has been recognized as a primary stressor in the catchment of the large, eutrophic Tai Lake, China. Nutrients, especially NH₃ and nitrite (NO₃^–) had more significant effects on structure of the zooplankton community than did other environmental factors. Abundances of rotifers increased along a gradient of increasing concentrations of total ammonia nitrogen (TAN), while abundances of copepods and cladocera decreased. A novel, rapid, species sensitivity distribution (SSD) approach based on operational taxonomic units (OTUs) was established to develop a WQC for NH₃. The WQC based on OTUs was consistent with the WQC based on the traditional morphology taxonomy approach. This genetics-based SSD approach could be a useful tool for monitoring for status and trends in species composition and deriving ecological criteria and an efficient biomonitoring tool to protect local aquatic ecosystems in virtually any aquatic ecosystem

Zooplankton species in the indigenous barcode database of Lake Tai.

<p>(A) A tree diagram of representative sequences for each species. Distance was measured as the number of base substitutions per site, based on the Kimura two-parameter (K2P) method. One thousand bootstrap trials were run using the neighbor-joining algorithm of the Mega 6.0 program. (B) Number of specimens of each species; red dot means that the species have barcode sequence in NCBI Genbank. (C) Intraspecific divergence based on the indigenous sequences. (D) <i>COI</i> sequences in NCBI Genbank. (E) Intraspecific divergence based on the NCBI Genbank sequences. (F) Similarity of indigenous DNA sequence against NCBI Genbank using Blastx. (G) Similarity of indigenous amino acid sequence against NCBI Genbank using Blastn. (H) Converge of indigenous DNA sequence against NCBI Genbank using Blastn.</p

Comparison of zooplankton identification in water samples between metabarcoding and morphology approaches.

<p>(A) Species number. (B) Frequency detected. The R2 and p-value are indicated for each regression axis.</p

Taxonomic assignment of NGS data.

<p>(A) Numbers of zooplankton OTUs and sequences in the NGS data. (B) Distribution of sequence similarity of OTUs against database (both indigenous and NCBI Genbank database). (C) Number of OTUs annotated by indigenous database and/or NCBI Genbank database. “Local” means the OTUs annotated by the indigenous database and “NCBI” means the OTUs annotated by NCBI Genbank. (D) Comparison of NGS data annotated by indigenous database and NCBI Genbank database. Only 24 species that have barcode sequence in NCBI Genbank were showed.</p

Proportion of bacterial sequences contributing to PAH degradation at ambient (370 ppm) or elevated (570 ppm) CO₂ levels.

<p>aCO₂, ambient CO₂; eCO₂, elevated CO₂. Data are means of three replicates ± standard error. Asterisks among columns indicate significant differences between aCO₂ and eCO₂ conditions (p < 0.05).</p

Elevated CO₂ accelerates polycyclic aromatic hydrocarbon accumulation in a paddy soil grown with rice

<div><p>The concentration of atmospheric carbon dioxide (CO₂) and polycyclic aromatic hydrocarbons (PAHs) contents in the environment have been rising due to human activities. Elevated CO₂ (eCO₂) levels have been shown to affect plant physiology and soil microbes, which may alter the degradation of organic pollutants. Here, we study the effect of eCO₂ on PAH accumulation in a paddy soil grown with rice. We collected soil and plant samples after rice harvest from a free-air CO₂ enrichment (FACE) system, which had already run for more than 15 years. Our results show that eCO₂ increased PAH concentrations in the soil, and we link this effect to a shift in soil microbial community structure and function. Elevated CO₂ changed the composition of soil microbial communities, especially by reducing the abundance of some microbial groups driving PAH degradation. Our study indicates that elevated CO₂ levels may weaken the self-cleaning ability of soils related to organic pollutants. Such changes in the function of soil microbial communities may threaten the quality of crops, with unknown implications for food safety and human health in future climate scenarios.</p></div