Silver nanomaterials (AgNMs) are broadly used and among the most studied nanomaterials. The underlying molecular mechanisms (e.g. protein and metabolite response) that precede phenotypical effects have been assessed to a much lesser extent. In this paper, we assess differentially expressed proteins (DEPs) and metabolites (DEMs) by high-throughput (HTP) techniques (HPLC-MS/MS with tandem mass tags, reversed-phase (RP) and hydrophilic interaction liquid chromatography (HILIC) with mass spectrometric detection). In a time series (0, 7, 14 days), the standard soil model Enchytraeus crypticus was exposed to AgNM300K and AgNO₃ at the reproduction EC20 and EC50. The impact on proteins/metabolites was clearly larger after 14 days. NM300K caused more upregulated DEPs/DEMs, more so at the EC20, whereas AgNO₃ caused a dose response increase of DEPs/DEMs. Similar pathways were activated, although often via opposite regulation (up vs down) of DEPs, hence, dissimilar mechanisms underlie the apical observed impact. Affected pathways included e.g. energy and lipid metabolism and oxidative stress. Uniquely affected by AgNO₃ was catalase, malate dehydrogenase and ATP-citrate synthase, and heat shock proteins (HSP70) and ferritin were affected by AgNM300K. The gene expression-based data in Adverse Outcome Pathway was confirmed and additional key events added, e.g. regulation of catalase and heat shock proteins were confirmed to be included. Finally, we observed (as we have seen before) that lower concentration of the NM caused higher biological impact. Data was deposited to ProteomeXchange, identifier PXD024444.

Monobutyl phthalate (MBP) is a primary metabolite of an environmental endocrine disruptor dibutyl phthalate (DBP), which poses a potential threat to living organisms. In this research, the acute toxicity of MBP on energy metabolism in zebrafish gills was studied. Transmission electron microscopy (TEM) results show that 10 mg L⁻¹ MBP can induce mitochondrial structural damage of chloride cells after 96 h of continuous exposure. The activity of ion ATPase and the expression level of oxidative phosphorylation-related genes suggest that MBP interferes with ATP synthesis and ion transport. Further leading to a decrease in mitochondrial membrane potential (MMP) and cell viability, thereby mediating early-stage cell apoptosis. Through a comprehensive analysis of principal component analysis (PCA) and integrated biomarker response (IBR) scores, atp5a1, a subunit of mitochondrial ATP synthase, is mainly inhibited by MBP, followed by genes encoding ion ATPase (atp1b2 and atp2b1). Importantly, MBP inhibits aerobic metabolism by inhibiting the key enzyme malate dehydrogenase (MDH) in the TCA cycle, forcing zebrafish to maintain ATP supply by enhancing anaerobic metabolism.

In the present study, the bioaccumulation of chromium, manganese, cobalt, copper, zinc, selenium, arsenic, strontium, cadmium, tin, antimony and lead in tissues of thirty marine fish species collected from New Ferry Whorf, Sassoon dock and Versova fishing harbour in Mumbai, India, were analysed. The bioaccumulation patterns of these twelve elements were determined to assess pollution biomarkers based on cellular and oxidative stresses. Catalase, superoxide dismutase and glutathione-s-transferase, glycolytic enzymes viz. lactate dehydrogenase and malate dehydrogenase, protein metabolism enzymes viz. aspartate transferase and alanine transferase, and lipid peroxidation were significantly higher in muscle and gill tissues. The activities of the neurotransmitter enzyme acetylcholine esterase in muscle and brain tissues was inhibited due to pollution. This study suggested that biochemical attributes such as oxidative stress enzymes, cellular biomarkers, neurotransmitter enzymes and metal and metalloid contamination could be successfully employed, even at low concentrations, as reliable biomarkers for biomonitoring of contaminated marine ecosystems.

The objective of this research is to investigate the enzymatic activities between protein disulfide isomerase (PDI) found in animals and plants and the properties found in a commonly used Chinese medicine called Sijunzi Tang. During the investigation, PDI, which is a monomer with a molecular mass of 57.0 kDa, was used to reactivate malate dehydrogenase (MDH). However, with the interference of polycyclic aromatic hydrocarbons (PAHs), evidence indicates that such chemicals are carcinogenic, mutagenic, and toxic to humans. The enzymatic activity of PDI found in animal’s liver and plant was 1657 folds of purification; 0.284 unit/mg of enzyme activity, and 5694.4 folds of purification; 1.00 unit/mg of enzyme activity, respectively. PDI extracted in treated animal and plant tissue revealed 2.40% and 80.44% of regaining MDH enzymatic activity, respectively. Although in its initial phase of investigation, it is assumed that the properties found in Sijunzi Tang can help regain enzymatic activity in those affected by xenobiotic substances, thus, making it a potential ingredient in assisting with PDI functions.

The main objective of this study was to characterize the Giardia duodenalis isolates from Iranian patients in Fars Province, south of Iran by biochemical and molecular methods. Fifteen mass cultivated of G. duodenalis isolates in modified TYI-S-33 medium were analyzed using isoenzyme electrophoresis and PCR genotyping. Polyacrylamide gel electrophoresis (PAGE) of five different enzyme systems was used to characterize isolates: (i) glucose-6-phosphate dehydrogenase, (ii) glucose phosphate isomerase, (iii) malate dehydrogenase, (iv) malic enzyme, and (v) phosphoglucomutase. As well, a fragment of the SSU-rDNA (292 bp) gene was amplified by PCR using the primers RH11 and RH4. The sequencing of the PCR products and phylogenetic tree were performed. The isoenzyme electrophoretic profiles divided fifteen G. duodenalis isolates into four zymodemes. G6PD, GPI, MDH, ME, and PGM enzyme systems showed 1, 2, 2, 3, and 3 enzyme pattern, respectively. G6PD isoenzyme pattern had the most homogeneity, while isoenzyme patterns of ME and PGM had the most heterogeneity in our study. Genotyping results indicated that the zymodemes 1–4 were categorized in assemblage A based on the SSU-rDNA gene. Phylogenetic analysis showed that all four zymodemes were distributed within the cluster of assemblage A. Our results indicated that both isoenzyme and DNA analyses were useful to characterize the isolates of Giardia and distinguishing various zymodemes and assemblages. It could be suggested that the genetic diversity among isoenzymes profiles of G. duodenalis may explain the variable clinical manifestations, pathogenicity, host response, drug susceptibility, and treatment efficacy of human giardiasis.

Polycyclic aromatic hydrocarbons (PAHs) are the most persistent organic pollutants in worldwide aquatic environments. The extensive isolation of genes responsive to PAH pollution in soft coral (Scleronephthya gracillimum) is described herein. Soft coral colonies were exposed to 100 μg/L of a standard mixture of PAHs. Gene candidates with transcript levels that changed in response to PAH exposure were identified by differential display polymerase chain reaction (DD-PCR). There were 37 types of candidate genes identified, of which 20 were upregulated in expression and 17 were downregulated. The functions of the genes identified included oxidative stress response, ribosomal structure maintenance, molecular chaperone activity, protein kinase activation and tumorigenesis, defense mechanisms, transcription, and other biological responses. mRNA quantification was carried out using real-time quantitative PCR in eight selected genes: cytosolic malate dehydrogenase, protein disulfide isomerase, ribosomal protein L6, ral guanine nucleotide dissociation stimulator-like 1, poly(ADP-ribose) polymerase 4, peptidylglycine α-hydroxylating monooxygenase, a disintegrin and metalloproteinase (ADAM) metallopeptidase protein, and eukaryotic initiation factor 4 gamma 3. Changes in transcript levels were consistent with DD-PCR results. The gene candidates isolated in this study were differentially expressed and therefore have potential as molecular biomarkers for understanding coral responses to environmental stressors.

A proteomic study of Cunninghamella echinulata recovery during exposure to tributyltin was conducted with 2-D SDS-PAGE protein separation and profiling, MALDI-TOF/TOF protein identification, and PCA analysis. The presence of TBT resulted in an upregulation of enzymes related to energy production via cellular respiration. The unique overexpression of NADH dehydrogenase and mitochondrial malate dehydrogenase, together with an increased level of cytochrome c oxidase, ATP synthase subunits, and inorganic pyrophosphatase, indicates a strong energy deficit in the cells, leading to an increase in the ATP production. The overexpression of Prohibitin-1, a multifunctional protein associated with the proper functioning of mitochondria, was observed as well. The data also revealed oxidative stress condition. Among reactive oxygen species (ROS)–scavenging enzymes, only superoxide dismutase (SOD) showed active response against oxidative stress induced by the xenobiotic. The induction of a series of ROS-scavenging enzymes was supported by a microscopic analysis revealing a considerably large concentration of ROS in the hyphae. The overexpression of cytoskeleton-related proteins in the TBT presence was also noticed. The obtained results allow explaining the recovery strategy of the fungus in response to the energy depletion caused by TBT.

The concentrations of 14 metals, namely, chromium (Cr), manganese (Mn), cobalt (Co), nickel (Ni), copper (Cu), zinc (Zn), selenium (Se), arsenic (As), strontium (Sr), cadmium (Cd), tin (Sn), antimony (Sb), mercury, (Hg), and lead (Pb), have been studied in freshwater mollusk Lamellidens marginalis collected from Dhimbe reservoir. In addition, also studied the cellular and antioxidative status to assess metal contamination as pollution biomarkers. The levels of all the 14 metals were elevated in different sampling sites of Dhimbe reservoir. The level of metals in the L. marginalis was followed as Mn > Zn > Ni > Sr > Cr > Cu > Hg > Se > Sn > Pb > As > Co > Cd > Sb. The oxidative biomarkers viz. catalase (CAT), superoxide dismutase (SOD), and glutathione-S-transferase (GST); glycolytic enzymes viz. lactate dehydrogenase (LDH) and malate dehydrogenase (MDH); and protein metabolic enzymes viz. aspartate amino transferase (AST), alanine amino transferase (ALT), and lipid peroxidation (LPO) were significantly (p < 0.01) higher, while neurotransmitter enzymes such as acetylcholine esterase (AChE) were significantly inhibited in muscle of L. marginalis (p < 0.01) collected from different sampling sites. Therefore, based on our results, we could recommend that the Dhimbe reservoir is moderately metal contaminated and investigation based on biochemical parameters such as antioxidative enzymes, cellular biomarkers, and AChE are strong indicators for metal contamination; hence, these parameters are successfully applied as reliable biomarkers for monitoring contaminated freshwater ecosystems.

The southwest coast of India along the Cochin region is seriously affected by metal contamination from increasing industrial activities. This contribution pertains to the chronic toxicity effects of lead and its biomarker responses on the black clam, Villorita cyprinoides, largely endemic to Southern India. The metal concentration (lead (Pb), copper (Cu), zinc (Zn), cadmium (Cd), nickel (Ni), and chromium (Cr)) and contamination indices such as geoaccumulation index and contamination factor suggests that the Cochin estuarine system (CES) is moderately to heavily polluted with metals. The 96 h effective median lethal concentration (LC₅₀) for Pb was 12.08 mg L⁻¹ at 95% confidence level, whereas the NOEC (no observed effect concentration) and LOEC (lowest observed effect concentration) were 0.46 mg L⁻¹ and 0.83 mg L⁻¹ respectively. The chronic toxicity value for Pb was 0.64 mg L⁻¹. Accumulation of Pb in V. cyprinoides after chronic toxicity test was several-fold higher than exposure concentration. On exposure to sublethal concentrations of lead, gills showed prominent pathological lesions such as damaged lateral cilia, congested hemolymph sinus, damaged abfrontal cilia, damaged ciliary filaments, fusion of gill lamellae, and reduced inter lamellar space. Filtration rate of clams exposed to Pb (86.69 ± 1.98 ml clam⁻¹ h⁻¹) was lower than that in control experiments (191.86 ± 6.48 ml clam⁻¹ h⁻¹). Antioxidant enzymes such as esterase (EST), superoxide dismutase (SOD), and malate dehydrogenase (MDH) exhibited different patterns in isoenzyme activity. Neutral red retention time (NRRT) showed a decrease from control to higher concentrations indicating membrane stability of hemocytic lysosomes decreased with the increasing metal exposure concentration. At LOEC and higher concentrations lysosomes showed enlargement and fragmentation. Lysosomal responses in V. cyprinoides can be used as a key cellular stress biomarker in assessing lead and other metal contamination.

The presence of nanoplastics (NPs) in various products and from the weathering of released plastic materials are of concern for the environment’s safety. The purpose of this study was to examine the biophysical effects of polystyrene NPs on freshwater mussels. Mussels were exposed to a range of concentrations of NPs (0.1, 0.5, 1, and 5 mg/L) for 24 h and allowed to depurate for 12 h in clean aquarium water. The digestive gland was isolated and analyzed for NPs, lipids, viscosity, protein aggregation, anisotropic changes (liquid crystals: LCs), and the oscillatory modulation in viscosity during the formation of self-organizing enzyme complex of fumarase, malate dehydrogenase, and citrate synthase. The results revealed that mussels accumulated NPs in the digestive gland and their levels were significantly correlated with lipids levels, LCs, the increase in the malate dehydrogenase/citrate synthase activity ratio, and oscillations in viscosity. Protein aggregation was also found to be correlated with lipid levels. The data suggests that the presence of NPs in the digestive gland involves changes in lipid content and LC formation and perturbs the normal oscillations in viscosity during sequential enzyme reactions of the above enzymes. It is concluded that the uptake of NPs in cells could disrupt the internal organization of cells which can interfere with the normal association of enzymes involved in energy metabolism.