Queen Conch (Strombus gigas) Testis Regresses during the Reproductive Season at Nearshore Sites in the Florida Keys

BACKGROUND: Queen conch (Strombus gigas) reproduction is inhibited in nearshore areas of the Florida Keys, relative to the offshore environment where conchs reproduce successfully. Nearshore reproductive failure is possibly a result of exposure to environmental factors, including heavy metals, which are likely to accumulate close to shore. Metals such as Cu and Zn are detrimental to reproduction in many mollusks. METHODOLOGY/PRINCIPAL FINDINGS: Histology shows gonadal atrophy in nearshore conchs as compared to reproductively healthy offshore conchs. In order to determine molecular mechanisms leading to tissue changes and reproductive failure, a microarray was developed. A normalized cDNA library for queen conch was constructed and sequenced using the 454 Life Sciences GS-FLX pyrosequencer, producing 27,723 assembled contigs and 7,740 annotated transcript sequences. The resulting sequences were used to design the microarray. Microarray analysis of conch testis indicated differential regulation of 255 genes (p<0.01) in nearshore conch, relative to offshore. Changes in expression for three of four transcripts of interest were confirmed using real-time reverse transcription polymerase chain reaction. Gene Ontology enrichment analysis indicated changes in biological processes: respiratory chain (GO:0015992), spermatogenesis (GO:0007283), small GTPase-mediated signal transduction (GO:0007264), and others. Inductively coupled plasma-mass spectrometry analysis indicated that Zn and possibly Cu were elevated in some nearshore conch tissues. CONCLUSIONS/SIGNIFICANCE: Congruence between testis histology and microarray data suggests that nearshore conch testes regress during the reproductive season, while offshore conch testes develop normally. Possible mechanisms underlying the testis regression observed in queen conch in the nearshore Florida Keys include a disruption of small GTPase (Ras)-mediated signaling in testis development. Additionally, elevated tissue levels of Cu (34.77 ng/mg in testis) and Zn (831.85 ng/mg in digestive gland, 83.96 ng/mg in testis) nearshore are similar to reported levels resulting in reproductive inhibition in other gastropods, indicating that these metals possibly contribute to NS conch reproductive failure

Aquatic Ecotoxicity Testing of Nanoparticles—The Quest To Disclose Nanoparticle Effects

The number of products on the market containing engineered nanoparticles (ENPs) has increased significantly, and concerns have been raised regarding their ecotoxicological effects. Environmental safety assessments as well as relevant and reliable ecotoxicological data are required for the safe and sustainable use of ENPs. Although the number of publications on the ecotoxicological effects and uptake of ENPs is rapidly expanding, the applicability of the reported data for hazard assessment is questionable. A major knowledge gap is whether nanoparticle effects occur when test organisms are exposed to ENPs in aquatic test systems. Filling this gap is not straightforward, because of the broad range of ENPs and the different behavior of ENPs compared to “ordinary” (dissolved) chemicals in the ecotoxicity test systems. The risk of generating false negatives, and false positives, in the currently used tests is high, and in most cases difficult to assess. This Review outlines some of the pitfalls in the aquatic toxicity testing of ENPs which may lead to misinterpretation of test results. Response types are also proposed to reveal potential nanoparticle effects in the aquatic test organisms

The Gut Microbiome and Aquatic Toxicology: An Emerging Concept for Environmental Health

The microbiome plays an essential role in the health and onset of diseases in all animals, including humans. The microbiome has emerged as a central theme in environmental toxicology, as microbes interact with the host immune system in addition to its role in chemical detoxification. Pathophysiological changes in the gastrointestinal tissue caused by ingested chemicals, and metabolites generated from microbial biodegradation, can lead to systemic adverse effects. This critical review dissects what we know about the impacts of environmental contaminants on the microbiome of aquatic species, with special emphasis on the gut microbiome. We highlight some of the known major gut epithelium proteins in vertebrate hosts that are targets for chemical perturbation, proteins that also directly cross‐talk with the microbiome. These proteins may act as molecular initiators for altered gut function, and we propose a general framework for an adverse outcome pathway that considers gut dysbiosis as a major contributing factor to adverse apical endpoints. We present two case studies, nanomaterials and hydrocarbons with special emphasis on the Deepwater Horizon oil spill, to illustrate how investigations into the microbiome can improve understanding of adverse outcomes. Lastly, we present strategies to functionally relate chemical‐induced gut dysbiosis with adverse outcomes, as this is required to demonstrate cause‐effect relationships. Further investigations into the toxicant‐microbiome relationship may prove to be a major breakthrough for improving animal and human health. This article is protected by copyright. All rights reserve

Behavior and chronic toxicity of two differently stabilized silver nanoparticles to Daphnia magna

While differences in silver nanoparticle (AgNP) colloidal stability, surface potential, or acute aquatic toxicity for differently stabilized AgNP have often been reported, these have rarely been studied in long-term ecotoxicity tests. In the current study, we investigated the chronic toxicity of AgNP to Daphnia magna over a 21-day period with two different stabilizers (citrate and detergent), representative for charge and sterical stabilizers, respectively. This was coupled with a series of short-term experiments, such as mass balance and uptake/depuration testing, to investigate the behavior of both types of AgNP during a typical media exchange period in the D. magna test for chronic toxicity. As expected, the sterically stabilized AgNP were more stable in the test medium, also in the presence of food; however, a higher uptake of silver after 24 h exposure of the charge stabilized AgNP was found compared to the detergent-stabilized AgNP (0.046 ± 0.006 μg Ag μg DW−1 and 0.023 ± 0.005 μg Ag μg DW−1, respectively). In accordance with this, the higher reproductive effects and mortality were found for the charge-stabilized than for the sterically-stabilized silver nanoparticles in 21-d tests for chronic toxicity. LOEC was 19.2 μg Ag L−1 for both endpoints for citrate-coated AgNP and &gt;27.5 μg Ag L−1 (highest tested concentration for detergent-stabilized AgNP). This indicates a link between uptake and toxicity. The inclusion of additional short-term experiments on uptake and depuration is recommended when longer-term chronic experiments with nanoparticles are conducted

Uptake, Retention and Internalization of Quantum Dots in \u3ci\u3eDaphnia\u3c/i\u3e is Influenced by Particle Surface Functionalization

Nanomaterials are a diverse group of compounds whose inevitable release into the environment warrants study of the fundamental processes that govern the ingestion, uptake and accumulation in aquatic organisms. Nanomaterials have the ability to transfer to higher trophic levels in aquatic ecosystems, and recent evidence suggests that the surface chemistry of both the nanoparticle and biological membrane can influence uptake kinetics. Therefore, our study investigates the effect of surface functionalization on uptake, internalization and depuration in Daphnia spp. Uncharged (polyethylene glycol; PEG), positively charged (amino-terminated: NH2) and negatively charged (carboxyl-modified; COOH) cadmium selenide/zinc sulfide quantum dots were used to monitor ingestion, uptake and depuration of nanometals in Daphnia magna and Ceriodaphnia dubia over 24 h of exposure. These studies demonstrated that particles with higher negative charge (COOH quantum dots) were taken up to a greater extent by Daphnia (259.17 ± 17.70 RFU/20 Daphnia) than either the NH2 (150.01 ± 18.91) or PEG quantum dots (95.17 ± 9.78), however this is likely related to the functional groups attached to the nanoparticles as there were no real differences in zeta potential. Whole body fluorescence associates well with fluorescent microscopic images obtained at the 24 h timepoint. Confocal and electron microscopic analysis clearly demonstrated that all three types of quantum dots could cross the intestinal epithelial barrier and be translocated to other cells. Upon cessation of exposure, elimination of all three materials was biphasic with rapid initial clearance that likely represents elimination of material remaining in the GI tract followed by a much slower elimination phase that likely represents elimination of internalized material. These studies demonstrate that daphnids can take up intact nanomaterial from the water column and that this uptake is strongly influenced by particle surface functionalization. In addition, the usefulness of using quantum dots as a proxy for other nanometals (no acute toxicity, clear visualization in electron microscopy), in conjunction with several different imaging techniques in assessing uptake and accumulation of nanoparticles in daphnids was demonstrated

Gammaherpesvirus Readthrough Transcription Generates a Long Non-Coding RNA That Is Regulated by Antisense miRNAs and Correlates with Enhanced Lytic Replication In Vivo

Gammaherpesviruses, including the human pathogens Epstein–Barr virus (EBV) and Kaposi’s sarcoma-associated herpesvirus (KSHV) are oncogenic viruses that establish lifelong infections in hosts and are associated with the development of lymphoproliferative diseases and lymphomas. Recent studies have shown that the majority of the mammalian genome is transcribed and gives rise to numerous long non-coding RNAs (lncRNAs). Likewise, the large double-stranded DNA virus genomes of herpesviruses undergo pervasive transcription, including the expression of many as yet uncharacterized lncRNAs. Murine gammaperherpesvirus 68 (MHV68, MuHV-4, HV68) is a natural pathogen of rodents, and is genetically and pathogenically related to EBV and KSHV, providing a highly tractable model for studies of gammaherpesvirus biology and pathogenesis. Through the integrated use of parallel data sets from multiple sequencing platforms, we previously resolved transcripts throughout the MHV68 genome, including at least 144 novel transcript isoforms. Here, we sought to molecularly validate novel transcripts identified within the M3/M2 locus, which harbors genes that code for the chemokine binding protein M3, the latency B cell signaling protein M2, and 10 microRNAs (miRNAs). Using strand-specific northern blots, we validated the presence of M3-04, a 3.91 kb polyadenylated transcript that initiates at the M3 transcription start site and reads through the M3 open reading frame (ORF), the M3 poly(a) signal sequence, and the M2 ORF. This unexpected transcript was solely localized to the nucleus, strongly suggesting that it is not translated and instead may function as a lncRNA. Use of an MHV68 mutant lacking two M3-04-antisense pre-miRNA stem loops resulted in highly increased expression of M3-04 and increased virus replication in the lungs of infected mice, demonstrating a key role for these RNAs in regulation of lytic infection. Together these findings suggest the possibility of a tripartite regulatory relationship between the lncRNA M3-04, antisense miRNAs, and the latency gene M2