Bioconcentration and lethal effects of gas-condensate and crude oil on nearshore copepod assemblages.

The progressive establishment of gas platforms and increasing petroleum accidents pose a threat to zooplankton communities and thus to pelagic ecosystems. This study is the first to compare the impacts of gas-condensate and crude oil on copepod assemblages. We conducted microcosm experiments simulating slick scenarios at five different concentrations of gas-condensate and crude oil to determine and compare their lethal effects and the bioconcentration of low molecular weight polycyclic aromatic hydrocarbons (LMW-PAHs) in eastern Mediterranean coastal copepod assemblages. We found that gas-condensate had a two-times higher toxic effect than crude oil, significantly reducing copepod survival with increased exposure levels. The LMW-PAHs bioconcentration factor was 1-2 orders of magnitude higher in copepods exposed to gas-condensate than in those exposed to crude oil. The median lethal concentration (LC50) was significantly lower in calanoids vs. cyclopoid copepods, suggesting that calanoids are more susceptible to gas-condensate and crude oil pollution, with potential trophic implications.

Passive-Sampler-Derived PCB and OCP Concentrations in the Waters of the World─First Results from the AQUA-GAPS/MONET Network.

Persistent organic pollutants (POPs) are recognized as pollutants of global concern, but so far, information on the trends of legacy POPs in the waters of the world has been missing due to logistical, analytical, and financial reasons. Passive samplers have emerged as an attractive alternative to active water sampling methods as they accumulate POPs, represent time-weighted average concentrations, and can easily be shipped and deployed. As part of the AQUA-GAPS/MONET, passive samplers were deployed at 40 globally distributed sites between 2016 and 2020, for a total of 21 freshwater and 40 marine deployments. Results from silicone passive samplers showed α-hexachlorocyclohexane (HCH) and γ-HCH displaying the greatest concentrations in the northern latitudes/Arctic Ocean, in stark contrast to the more persistent penta (PeCB)- and hexachlorobenzene (HCB), which approached equilibrium across sampling sites. Geospatial patterns of polychlorinated biphenyl (PCB) aqueous concentrations closely matched original estimates of production and use, implying limited global transport. Positive correlations between log-transformed concentrations of Σ7PCB, ΣDDTs, Σendosulfan, and Σchlordane, but not ΣHCH, and the log of population density (p < 0.05) within 5 and 10 km of the sampling sites also supported limited transport from used sites. These results help to understand the extent of global distribution, and eventually time-trends, of organic pollutants in aquatic systems, such as across freshwaters and oceans. Future deployments will aim to establish time-trends at selected sites while adding to the geographical coverage.

Anthropogenic and natural disturbances along a river and its estuary alter the diversity of pathogens and antibiotic resistance mechanisms.

Antibiotic resistance (AMR) in pathogens threatens human health worldwide, and antibiotic-resistant bacteria (ARB) are widespread in the environment. In particular, anthropogenically-disturbed rivers became reservoirs of ARBs and hotspots of antibiotic resistance gene (ARG) transmission. However, the diversity and sources of ARB, and the mechanisms of ARG transmission are not fully known. Here, we used deep metagenomic sequencing to study the dynamics of pathogens and their antibiotic resistance mechanisms along the Alexander River (Israel), affected by sewage and animal farm runoffs. Putative pathogens such as Aeromicrobium marinum and Mycobacterium massilipolynesiensis were enriched in western stations, following the inputs of polluted Nablus River. Aeromonas veronii was dominant in eastern stations in Spring. Several AMR mechanisms showed distinct patterns in Summer-Spring (dry season) and Winter (rainy season). We found low abundance beta-lactamases conferring carbapenem resistance: e.g., OXA-912 was linked to A. veronii in Spring; OXA-119 and OXA-205 to Xanthomonadaceae in Winter. We classified 33 % of ARG-containing contigs as putative plasmid sequences, indicating the high potential for resistome transmission. A limited number of ARGs were linked to putative phages. Our results suggest that this model river is a hotspot for AMR activity and transmission, and highlight the merit of deep sequencing for AMR discovery.

BTEX and PAH contributions to Lake Kinneret water: a seasonal-based study of volatile and semi-volatile anthropogenic pollutants in freshwater sources.

Benzene , toluene, ethylbenzene, and xylenes (BTEX) BTEX molecules are toxic components, ubiquitous in the environment, often found in concentrations- a few orders of magnitude higher than the well-studied PAHs levels. This fact is demonstrated in either crude oil, fuels, water, and air samples. BTEX studies focus mainly on the airborne levels of these molecules, while their waterborne presence is understudied. In this study, BTEX levels were assessed at Lake Kinneret, Israel. As a result, 0-1.5 ppb of BTEX was recorded in five stations (2021-2022). Elevated BTEX levels (3-10 ppb) were recorded at the northern rivers nourishing this lake, implying the existence of remote polluting sources. Transect air samplings of BTEX conducted at the lake next to the bathing season of 2021 revealed airborne BTEX levels between 0.8 and 10 µg/m3, peaking up close to the bathing season, yet inconsistent with the BTEX water level trend. Lake water samples collected next to Tiberias city outfalls following the "Carmel" rainstorm showed elevated concentrations of BTEX up to 35 ppb and PAHs up to 0.47 ppb with an urban isotopic signal. The remote station's PAHs levels were less than one order of magnitude, with a distinct rural isotopic signal. Additionally, a human-specific microbial marker revealed increased sewer contributions at some of the urbansites. The results of this study show that a wide area dispersion of low atmospheric BTEX levels exists in the lake's perimeter. The dispersion rate is most likely influenced by season-based factors, e.g., motors and biomass fires. The unstudied waterborne BTEX levels in this lake are influenced by rivers, city runoff, and other yet unknown factors that may contribute to the sedimentation of these components. This process may result in a chronic pollution state. Despite the BTEX's medium-low solubility and high volatility, its under-evaluated waterborne transportation may lead to high toxic levels following bioaccumulation.

Seaweeds fast EDC bioremediation: Supporting evidence of EE2 and BPA degradation by the red seaweed Gracilaria sp., and a proposed model for the remedy of marine-borne phenol pollutants.

In the last few decades, Endocrine Disrupting Chemicals (EDCs) have taken significant roles in creating harmful effects to aquatic organisms. Many proposed treatment applications are time consuming, expensive and focus mainly on waste water treatment plants (WWTP), which are indeed a major aquatic polluting source. Nonetheless, the marine environment is the ultimate sink of many pollutants, e.g. EDCs, and has been largely neglected mainly due to the challenge in treating such salty and immense open natural ecosystems. In this study we describe the bromination and the yet unpresented degradation process of high concentrations (5 mg/L) of phenolic EDCs, by the marine red macroalgaeGracilaria sp. As shown, 17α-Ethinylestradiol (EE2), a well-known contraceptive drug, and one of the most persistent phenol EDCs in the environment, was eliminated from both the medium and tissues of the macroalga, in addition to the degradation of all metabolites as verified by the nil estrogenic activity recorded in the medium. Validation of the proposed bromination-degradation route was reinforced by identifying Bisphenol A (BPA) brominated degradation products only, following 168H of incubation in the presence of Gracilaria sp. As demonstrated in this assay for EE2, BPA and finally for paracetamol, it is likely that the phenol scavenging activity is nonspecific and, thus, possibly even a wider scope of various other phenol-based pollutants might be treated in coastal waters. As far as we know, Gracilaria sp. is the only marine sessile organism able of degrading various phenol based pollutants. The worldwide distribution of many Gracilaria species and their wide aquaculture knowhow, suggest that bioremediation based on these seaweeds is a possible cost effective progressive solution to the treatment of a wide scope of phenols at the marine environment.

Multidrug-resistant enterobacteriaceae in coastal water: an emerging threat.

BACKGROUND: The environmental role of carbapenemase-producing Enterobacteriaceae (CPE) acquisition and infection in human disease has been described but not thoroughly investigated. We aimed to assess the occurrence of CPE in nearshore aquatic bodies. METHODS: Enterobacteriaceae were cultured from coastal and estuary water near Netanya, Israel in June and July of 2018. Bacteria were identified by VITEK2® and their antimicrobial susceptibility was tested according to the CLSI guidelines. Enterobacteriaceae genomes were sequenced to elucidate their resistome and carbapenemase types. RESULTS: Among other clinically relevant bacteria, four CPE (three Enterobacter spp and one Escherichia coli isolate) were isolated from two river estuaries (Poleg and Alexander Rivers) and coastal water at a popular recreational beach (Beit Yanai). Molecular analysis and genome sequencing revealed the persistent presence of rare beta-lactamase resistance genes, including blaIMI-2 and a previously unknown blaIMI-20 allele, which were not found among the local epidemiological strains. Genome comparisons revealed the high identity of riverine and marine CPE that were cultivated one month apart. CONCLUSIONS: We show that CPE contamination was widespread in nearshore marine and riverine habitats. The high genome-level similarity of riverine and marine CPEs, isolated one month apart, hints at the common source of infection. We discuss the clinical implications of these findings and stress the urgent need to assess the role of the aquatic environment in CPE epidemiology.

Uptake and incorporation of PCBs by eastern Mediterranean rabbitfish that consumed microplastics.

Two experiments were executed to assess the feasibility of Polychlorinated Biphenyls (PCBs) transfer to fish tissues via MPs as a vector. PCBs that occur in the marine environment were tested for their adsorption to four different MP types. PCB congeners showed the highest adsorption levels to Polypropylene homo-polymer. The uptake of PCBs through MP ingestion was tested in an outdoor mesocosm using the herbivorous rabbitfish, Siganus rivulatus in the eastern Mediterranean Sea. Polypropylene homo-polymer particles (0.3-5.0 mm) pre saturated with 11 PCB congeners, in two concentrations (500 ng/g and 5000 ng/g), were mixed with dough and offered to the fish. PCBs were identified after two weeks in fish muscle tissues, but not in the liver. These results suggest that ingestion of contaminated MP by rabbitfish might harm them in the long run, and perhaps even those who consume them on a regular basis, e.g. rabbitfish predators and humans.

Phytoplankton response to N-rich well amelioration brines: A mesocosm study from the southeastern Mediterranean Sea.

Human-induced eutrophication of coastal water may be a major threat to aquatic life. Here, we investigated the effects of N-rich well amelioration brines (WAB) on coastal phytoplankton population's habitat in the surface oligotrophic waters of the southeastern Mediterranean Sea (SEM). To this end, we added WAB (2 concentrations) to mesocosms (1-m3 bags) to surface SEM water during summer and winter, where changes in phytoplankton biomass, activity and diversity was monitored daily for 8 days. Our results demonstrate that WAB addition triggered a phytoplankton bloom, resulting in elevated algal biomass (maximal +780%), increased primary production rates (maximal +675%) and a decrease in eukaryotic algal α-diversity (ca. -20%). Among the species that bloomed following WAB amendments, we found the potentially toxic dinoflagellate Karlodinium venificum. This study adds valuable perspective to the effect of nutrients discharged into nutrient limited SEM coastal waters, and in particular of N-derived WAB.

Computational and experimental analysis of drug binding to the Influenza M2 channel.

The Influenza Matrix 2 (M2) protein is the target of Amantadine and Rimantadine which block its H(+) channel activity. However, the potential of these aminoadamantyls to serve as anti-flu agents is marred by the rapid resistance that the virus develops against them. Herein, using a cell based assay that we developed, we identify two new aminoadamantyl derivatives that show increased activity against otherwise resistant M2 variants. In order to understand the distinguishing binding patterns of the different blockers, we computed the potential of mean force of the drug binding process. The results reveal that the new derivatives are less mobile and bind to a larger pocket in the channel. Finally, such analyses may prove useful in designing new, more effective M2 blockers as a means of curbing influenza. This article is part of a Special Issue entitled: Viral Membrane Proteins - Channels for Cellular Networking.

How do aminoadamantanes block the influenza M2 channel, and how does resistance develop?

The interactions between channels and their cognate blockers are at the heart of numerous biomedical phenomena. Herein, we unravel one particularly important example bearing direct pharmaceutical relevance: the blockage mechanism of the influenza M2 channel by the anti-flu amino-adamantyls (amantadine and rimantadine) and how the channel and, consequently, the virus develop resistance against them. Using both computational analyses and experimental verification, we find that amino-adamantyls inhibit M2's H(+) channel activity by electrostatic hindrance due to their positively charged amino group. In contrast, the hydrophobic adamantyl moiety on its own does not impact conductivity. Additionally, we were able to uncover how mutations in M2 are capable of retaining drug binding on the one hand yet rendering the protein and the mutated virus resistant to amino-adamantyls on the other hand. We show that the mutated, drug-resistant protein has a larger binding pocket for the drug. Hence, despite binding the channel, the drug remains sufficiently mobile so as not to exert a H(+)-blocking positive electrostatic hindrance. Such insight into the blocking mechanism of amino-adamantyls, and resistance thereof, may aid in the design of next-generation anti-flu agents.

Quantitative analysis of influenza M2 channel blockers.

The influenza M2 H(+) channel enables the concomitant acidification of the viral lumen upon endosomic internalization. This process is critical to the viral infectivity cycle, demonstrated by the fact that M2 is one of only two targets for anti-flu agents. However, aminoadamantyls that block the M2 channel are of limited therapeutic use due to the emergence of resistance mutations in the protein. Herein, using an assay that involves expression of the protein in Escherichia coli with resultant growth retardation, we present quantitative measurements of channel blocker interactions. Comparison of detailed K(s) measurements of different drugs for several influenza channels, shows that the swine flu M2 exhibits the highest resistance to aminoadamantyls of any channel known to date. From the perspective of the blocker, we show that rimantadine is consistently a better blocker of M2 than amantadine. Taken together, such detailed and quantitative analyses provide insight into the mechanism of this important and pharmaceutically relevant channel blocker system.

Resistance characteristics of influenza to amino-adamantyls.

The recent outbreaks of avian flu in Southeast Asia and swine flu in Mexico City painfully exemplify the ability of the influenza virus to rapidly mutate and develop resistance to modern medicines. This review seeks to detail the molecular mechanism by which the influenza virus has obtained resistance to amino-adamantyls, one of only two classes of drugs that combat the flu. Amino-adamantyls target the viral M2 H(+) channel and have become largely ineffective due to mutations in the transmembrane domain of the protein. Herein we describe these resistance rendering mutations and the compounded effects they have upon the protein's function and resulting virus viability.

Intracellular uptake and intracavitary targeting of folate-conjugated liposomes in a mouse lymphoma model with up-regulated folate receptors.

The folate receptor is overexpressed in a broad spectrum of malignant tumors and represents an attractive target for selective delivery of anticancer agents to folate receptor-expressing tumors. This study examines folate-lipid conjugates as a means of enhancing the tumor selectivity of liposome-encapsulated drugs in a mouse lymphoma model. Folate-derivatized polyethylene glycol (PEG3350)-distearoyl-phosphatidylethanolamine was post-loaded at various concentrations into the following preparations: radiolabeled PEGylated liposomes, PEGylated liposomes labeled in the aqueous compartment with dextran fluorescein, and PEGylated liposomal doxorubicin (PLD, Doxil). We incubated folate-targeted radiolabeled or fluorescent liposomes with mouse J6456 lymphoma cells up-regulated for their folate receptors (J6456-FR) to determine the optimal ligand concentration required in the lipid bilayer for liposomal cell association, and to examine whether folate-targeted liposomes are internalized by J6456-FR cells in suspension. Liposomal association with cells was quantified based on radioactivity and fluorescence-activated cell sorting analysis, and internalization was assessed by confocal fluorescence microscopy. We found an optimal ligand molar concentration of approximately 0.5% using our ligand. A substantial lipid dose-dependent increase in cell-associated fluorescence was found in folate-targeted liposomes compared with nontargeted liposomes. Confocal depth scanning showed that a substantial amount of the folate-targeted liposomes are internalized by J6456-FR cells. Binding and uptake of folate-targeted PLD by J6456-FR cells were also observed in vivo after i.p. injection of folate-targeted PLD in mice bearing ascitic J6456-FR tumors. The drug levels in ascitic tumor cells were increased by 17-fold, whereas those in plasma were decreased by 14-fold when folate-targeted PLD were compared with nontargeted PLD in the i.p. model. Folate-targeted liposomes represent an attractive approach for the intracellular delivery of drugs to folate receptor-expressing lymphoma cells and seem to be a promising tool for in vivo intracavitary drug targeting.

A novel method of resistance for influenza against a channel-blocking antiviral drug.

Effective antivirals are few and far between, and as such, the appearance of resistance toward such treatments is an obvious medical concern. In this article, we analyze the mechanism by which influenza attains resistance toward amantadine, a blocker of the viral M2 H(+) channel. Binding analyses of amantadine to M2 peptides from different viral strains showed that the virus has developed two alternate routes to avoid blockage of its channel: (1) a conventional route, in which the channel no longer binds the blocker and, hence, the blocker cannot exert its inhibitory function; and (2) a novel mechanism, in which binding of the blocker is retained, yet the function of the protein is unaffected. Pore diameter profiles revealed the molecular mechanism by which the virus may attain this novel type of resistance: an increase in the size of the channel. Thus, despite the drug binding the channel, it may not be able to block the pore, since the channel diameter has increased. Our findings may have broad ramifications in the design of new antivirals, and of novel blockers against malfunctioning human channels implicated in disease.