Guidance for Residents Addressing Copper Problems in Drinking Water: Opportunities and Challenges.

Residents and their pets may experience aesthetic or health concerns resulting from elevated copper in their drinking water. The United States Environmental Protection Agency Lead and Copper Rule focuses on addressing systemwide corrosion issues, but gaps in the rule leave some municipal water consumers and residents with private well water vulnerable to high cuprosolvency. We developed guidance to aid residents in understanding, detecting, and addressing cuprosolvency issues in their drinking water. Three types of at-home test kits for copper and one for pH were determined to be accurate enough (R 2 > 0.9 (lab, based on average values from n = 5 replicates each) and >0.7 (field)) to detect concerns related to high cuprosolvency and inform selection of intervention options. Case study results indicate that, although water treatments such as increasing pH on-site may be effective, long-term treatment (>36 weeks or permanently) may be needed to maintain reductions in cuprosolvency. A decision tree is provided to help residents and citizen scientists navigate these concerns for both public water systems and private wells.

A Possible Nocebo Effect in Children Following the Flint Water Crisis: Evidence From Schoolteacher Perceptions and Neuropsychological Evaluations.

OBJECTIVE: Special education enrollment increased in Flint following the 2014-2015 Flint Water Crisis, but lead exposure is not plausibly responsible. Labeling Flint children as lead poisoned and/or brain damaged may have contributed to rising special education needs (ie, nocebo effect). To better document this possibility, we surveyed schoolteachers and reviewed neuropsychological assessments of children for indications of negative labeling. METHODS: A survey of Flint and Detroit (control) public schoolteachers using a modified Illness Perception Questionnaire was conducted 5 years post-crisis. We also examined neuropsychological assessments from a recently settled class lawsuit. RESULTS: Relative to Detroit (n = 24), Flint teachers (n = 11) believed that a higher proportion of their students had harmful lead exposure (91.8% Flint vs 46% Detroit; P = 0.00034), were lead poisoned (51.3% vs 24.3%; P = 0.018), or brain damaged (28.8% vs 12.9%; P = 0.1), even though blood lead of Flint children was always less than half of that of Detroit children. Neuropsychological assessments diagnosed lead poisoning and/or brain damage from water lead exposure in all tested children (n = 8), even though none had evidence of elevated blood lead and a majority had prior learning disability diagnoses. CONCLUSION: Teachers' responses and neuropsychological assessments suggest Flint children were harmed by a nocebo effect.

Zinc Orthophosphate Can Reduce Nitrate-Induced Corrosion of Lead Solder.

Nitrate-induced spallation of lead-bearing solder particles into drinking water is not sufficiently controlled by phosphate-based inhibitors, although adding zinc can improve their performance. Studies using copper coupons coated with new lead-tin solder in water with up to 12 mg/L nitrate demonstrated that zinc orthophosphate reduced lead release by more than 90% and outperformed orthophosphate alone. Lead release and spallation from harvested pipes with decades-old lead-tin solder in a high nitrate water were improved but not eliminated with zinc orthophosphate over a period of months. When applied at a water utility with high source water nitrate, monthly in-home field sampling showed that 90th percentile lead levels dropped below the action level after dosing zinc orthophosphate at full scale for 6 months. Scanning electron microscopy (SEM) analysis of pipe scales revealed that zinc and orthophosphate codeposit at the copper-solder interface and may act as a mixed inhibitor, with zinc inhibiting the cathodic reaction on the copper pipe, phosphate limiting the anodic reaction, and an added benefit of zinc orthophosphate preferentially precipitating at the galvanic interface between the anode and the cathode. Updates to corrosion control guidance for waters with higher nitrate due to seasonal runoff or source water changes are needed.

Effects of Copper on Legionella pneumophila Revealed via Viability Assays and Proteomics.

Cu is an antimicrobial that is commonly applied to premise (i.e., building) plumbing systems for Legionella control, but the precise mechanisms of inactivation are not well defined. Here, we applied a suite of viability assays and mass spectrometry-based proteomics to assess the mechanistic effects of Cu on L. pneumophila. Although a five- to six-log reduction in culturability was observed with 5 mg/L Cu2+ exposure, cell membrane integrity only indicated a <50% reduction. Whole-cell proteomic analysis revealed that AhpD, a protein related to oxidative stress, was elevated in Cu-exposed Legionella relative to culturable cells. Other proteins related to cell membrane synthesis and motility were also higher for the Cu-exposed cells relative to controls without Cu. While the proteins related to primary metabolism decreased for the Cu-exposed cells, no significant differences in the abundance of proteins related to virulence or infectivity were found, which was consistent with the ability of VBNC cells to cause infections. Whereas the cell-membrane integrity assay provided an upper-bound measurement of viability, an amoebae co-culture assay provided a lower-bound limit. The findings have important implications for assessing Legionella risk following its exposure to copper in engineered water systems.

Drinking water sources, quality, and associated health outcomes in Appalachian Virginia: A risk characterization study in two counties.

OBJECTIVES: In the US, violations of drinking water regulations are highest in lower-income rural areas overall, and particularly in Central Appalachia. However, data on drinking water use, quality, and associated health outcomes in rural Appalachia are limited. We sought to assess public and private drinking water sources and associated risk factors for waterborne pathogen exposures for individuals living in rural regions of Appalachian Virginia. METHODS: We administered surveys and collected tap water, bottled water, and saliva samples in lower-income households in two adjacent rural counties in southwest Virginia (bordering Kentucky and Tennessee). Water samples were tested for pH, temperature, conductivity, total coliforms, E. coli, free chlorine, nitrate, fluoride, heavy metals, and specific pathogen targets. Saliva samples were analyzed for antibody responses to potentially waterborne infections. We also shared water analysis results with households. RESULTS: We enrolled 33 households (83 individuals), 82% (n = 27) with utility-supplied water and 18% with private wells (n = 3) or springs (n = 3). 58% (n = 19) reported household incomes of <$20,000/year. Total coliforms were detected in water samples from 33% (n = 11) of homes, E. coli in 12%, all with wells or springs (n = 4), and Aeromonas, Campylobacter, and Enterobacter in 9%, all spring water (n = 3). Diarrhea was reported for 10% of individuals (n = 8), but was not associated with E. coli detection. 34% (n = 15) of saliva samples had detectable antibody responses for Cryptosporidium spp., C. jejuni, and Hepatitis E. After controlling for covariates and clustering, individuals in households with septic systems and straight pipes had significantly higher likelihoods of antibody detection (risk ratios = 3.28, 95%CI = 1.01-10.65). CONCLUSIONS: To our knowledge, this is the first study to collect and analyze drinking water samples, saliva samples, and reported health outcome data from low-income households in Central Appalachia. Our findings indicate that utility-supplied water in this region was generally safe, and individuals in low-income households without utility-supplied water or sewerage have higher exposures to waterborne pathogens.

Competition for engineering tenure-track faculty positions in the United States.

How likely are engineering PhD graduates to get a tenure-track faculty position in the United States? To answer this question, we analyzed aggregated yearly data on PhD graduates and tenure-track/tenured faculty members across all engineering disciplines from 2006 to 2021, obtained from the American Society of Engineering Education. The average likelihood for securing a tenure-track faculty position for engineering overall during this 16-year period was 12.4% (range = 10.9-18.5%), implying that roughly 1 in 8 PhD graduates attain such positions. After a significant decline from 18.5 to 10.9% between 2006 and 2014 (R2 = 0.62; P < 0.05), a trend consistent with a period of rising competition, the outlook has since stabilized between 11.3 and 12% (R2 = 0.04; P > 0.05). Given that most engineering PhD graduates will never secure a tenure-track faculty position, emphasizing alternative career tracks during doctoral training could align expectations better with reality.

Poly-ß-(1→6)-N-acetyl-D-glucosamine mediates surface attachment, biofilm formation, and biocide resistance in Cutibacterium acnes.

Background: The commensal skin bacterium Cutibacterium acnes plays a role in the pathogenesis of acne vulgaris and also causes opportunistic infections of implanted medical devices due to its ability to form biofilms on biomaterial surfaces. Poly-ß-(1→6)-N-acetyl-D-glucosamine (PNAG) is an extracellular polysaccharide that mediates biofilm formation and biocide resistance in a wide range of bacterial pathogens. The objective of this study was to determine whether C. acnes produces PNAG, and whether PNAG contributes to C. acnes biofilm formation and biocide resistance in vitro. Methods: PNAG was detected on the surface of C. acnes cells by fluorescence confocal microscopy using the antigen-specific human IgG1 monoclonal antibody F598. PNAG was detected in C. acnes biofilms by measuring the ability of the PNAG-specific glycosidase dispersin B to inhibit biofilm formation and sensitize biofilms to biocide killing. Results: Monoclonal antibody F598 bound to the surface of C. acnes cells. Dispersin B inhibited attachment of C. acnes cells to polystyrene rods, inhibited biofilm formation by C. acnes in glass and polypropylene tubes, and sensitized C. acnes biofilms to killing by benzoyl peroxide and tetracycline. Conclusion: C. acnes produces PNAG, and PNAG contributes to C. acnes biofilm formation and biocide resistance in vitro. PNAG may play a role in C. acnes skin colonization, biocide resistance, and virulence in vivo.

Considering a Utility-Centric Framework Based on "Minimum Orthophosphate" Criteria for Mitigation of Elevated Cuprosolvency in Drinking Water.

Gaps in the United States Environmental Protection Agency (US EPA) Lead and Copper Rule (LCR) leave some consumers and their pets vulnerable to high cuprosolvency in drinking water. This study seeks to help proactive utilities who wish to mitigate cuprosolvency problems through the addition of orthophosphate corrosion inhibitors. The minimum doses of orthophosphate necessary to achieve acceptable cuprosolvency in relatively new copper pipe were estimated as a function of alkalinity via linear regressions for the 90th, 95th, and 100th percentile copper tube segments (R2 > 0.98, n = 4). Orthophosphate was very effective at reducing cuprosolvency in the short term but, in some cases, resulted in higher long-term copper concentrations than the corresponding condition without orthophosphate. Alternatives to predicting "long-term" results for copper tubes using simpler bench tests starting with fresh Cu(OH)2 solids showed promise but would require further vetting to overcome limitations such as maintaining water chemistry and orthophosphate residuals and to ensure comparability to results using copper tube.

Mercury poisoning in a bodybuilder.

A 64-year-old man had progressive unsteadiness over several years, with tingling in his feet. He was a longstanding bodybuilding enthusiast. Clinical assessment and neurophysiology confirmed a cerebellar ataxia and axonal peripheral neuropathy. His serum mercury concentration was significantly raised. We diagnosed chronic mercury toxicity secondary to excessive tuna consumption. We advised him to stop eating tuna and prescribed dimercaptosuccinic acid, after which his serum mercury concentrations subsequently fell. This case report highlights the importance of considering dietary and nutritional causes of neurological disease. We also discuss the mechanisms, diagnosis and treatment of mercury toxicity.

Evaluation of pre-hospital cannabis exposure and hospital opioid utilization in a trauma population: A retrospective cohort.

PURPOSE: Cannabis utilization has increased over time for recreational and medical purposes due to its legalization or decriminalization. The effects of cannabis use on opioid utilization are not well understood. The primary objective was to evaluate the total opioid utilization, measured in morphine milligram equivalents (MME), in hospitalized trauma patients that tested positive for tetrahydrocannabinol (THC) on a urine drug screen (UDS). METHODS: This was a retrospective, cohort study in a level 1 trauma center between 10/17/17 and 12/31/19. Adult trauma patients (aged 15 years and older) who had a UDS completed within 48 h of hospital arrival were eligible for inclusion. Patients were excluded for a hospitalization >14 days, death within 24 h, severe alcohol withdrawal, prescribed cannabinoids, high daily opioid use prior-to-arrival, or transitioned to hospice or palliative care. Group assignments were determined based on the presence or absence of THC on the UDS. RESULTS: The analysis included 750 patients with 160 (21 %) THC positive patients. The population were primarily male (64.6 %), median age of 56 years [IQR 35-72], with blunt (93 %) injuries from motor vehicle crash or falls (79 %). The THC group was more likely to have other substances present, including amphetamines, benzodiazepines, opiates, and cocaine. The THC group had a higher median injury severity score (10 [IQR 5-17] vs. 9 [5-14], p = 0.0056), and maximum abbreviated injury score (3 [IQR 2-3] vs. 2 [IQR 2-3], p = 0.0009). The THC group had a total higher median opioid utilization during the hospitalization (155 [IQR 68-367] vs. 62 [IQR 13-175] MME; p < 0.0001), which included higher opioid use in the emergency department, floor, and intensive care unit. There were no significant differences in secondary outcomes except the THC group was more likely to receive an opioid prescription at discharge and more likely to require mechanical ventilation. Based on multivariable regression analyses, other variables were associated with increased opioid utilization. CONCLUSION: Pre-existing THC exposure may be associated with an increased hospital opioid utilization in a trauma population. However, other variables may also play a role in opioid utilization.

Premise Plumbing Pipe Materials and In-Building Disinfectants Shape the Potential for Proliferation of Pathogens and Antibiotic Resistance Genes.

In-building disinfectants are commonly applied to control the growth of pathogens in plumbing, particularly in facilities such as hospitals that house vulnerable populations. However, their application has not been well optimized, especially with respect to interactive effects with pipe materials and potential unintended effects, such as enrichment of antibiotic resistance genes (ARGs) across the microbial community. Here, we used triplicate convectively mixed pipe reactors consisting of three pipe materials (PVC, copper, and iron) for replicated simulation of the distal reaches of premise plumbing and evaluated the effects of incrementally increased doses of chlorine, chloramine, chlorine dioxide, and copper-silver disinfectants. We used shotgun metagenomic sequencing to characterize the resulting succession of the corresponding microbiomes over the course of 37 weeks. We found that both disinfectants and pipe material affected ARG and microbial community taxonomic composition both independently and interactively. Water quality and total bacterial numbers were not found to be predictive of pathogenic species markers. One result of particular concern was the tendency of disinfectants, especially monochloramine, to enrich ARGs. Metagenome assembly indicated that many ARGs were enriched specifically among the pathogenic species. Functional gene analysis was indicative of a response of the microbes to oxidative stress, which is known to co/cross-select for antibiotic resistance. These findings emphasize the need for a holistic evaluation of pathogen control strategies for plumbing.

Influence of pipe materials on in-building disinfection of P. aeruginosa and A. baumannii in simulated hot water plumbing.

A framework is needed to account for interactive effects of plumbing materials and disinfectants on opportunistic pathogens (OPs) in building water systems. Here we evaluated free chlorine, monochloramine, chlorine dioxide, and copper-silver ionization (CSI) for controlling Pseudomonas aeruginosa and Acinetobacter baumannii as two representative OPs that colonize hot water plumbing, in tests using polyvinylchloride (PVC), copper-PVC, and iron-PVC convectively-mixed pipe reactors (CMPRs). Pipe materials vulnerable to corrosion (i.e., iron and copper) altered the pH, dissolved oxygen, and disinfectant levels in a manner that influenced growth trends of the two OPs and total bacteria. P. aeruginosa grew well in PVC CMPRs, poorly in iron-PVC CMPRs, and was best controlled by CSI disinfection, whereas A. baumannii showed the opposite trend for pipe material and was better controlled by chlorine and chlorine dioxide. Various scenarios were identified in which pipe material and disinfectant can interact to either hinder or accelerate growth of OPs, illustrating the difficulties of controlling OPs in portions of plumbing systems experiencing warm, stagnant water.

A Comprehensive Examination of the Contaminants in Drinking Water in Public Schools in California, 2017-2022.

OBJECTIVES: Reports of unsafe school drinking water in the United States highlight the importance of ensuring school water is safe for consumption. Our objectives were to describe (1) results from our recent school drinking water sampling of 5 common contaminants, (2) school-level factors associated with exceedances of various water quality standards, and (3) recommendations. METHODS: We collected and analyzed drinking water samples from at least 3 sources in 83 schools from a representative sample of California public schools from 2017 through 2022. We used multivariate logistic regression to examine school-level factors associated with lead in drinking water exceedances at the American Academy of Pediatrics (AAP) recommendation level (1 part per billion [ppb]) and state action-level exceedances of other contaminants (lead, copper, arsenic, nitrate, and hexavalent chromium). RESULTS: No schools had state action-level violations for arsenic or nitrate; however, 4% had ≥1 tap that exceeded either the proposed 10 ppb action level for hexavalent chromium or the 1300 ppb action level for copper. Of first-draw lead samples, 4% of schools had ≥1 tap that exceeded the California action level of 15 ppb, 18% exceeded the US Food and Drug Administration (FDA) bottled water standard of 5 ppb, and 75% exceeded the AAP 1 ppb recommendation. After turning on the tap and flushing water for 45 seconds, 2%, 10%, and 33% of schools exceeded the same standards, respectively. We found no significant differences in demographic characteristics between schools with and without FDA or AAP exceedances. CONCLUSIONS: Enforcing stricter lead action levels (<5 ppb) will markedly increase remediation costs. Continued sampling, testing, and remediation efforts are necessary to ensure drinking water meets safety standards in US schools.

Shotgun Metagenomics Reveals Impacts of Copper and Water Heater Anodes on Pathogens and Microbiomes in Hot Water Plumbing Systems.

Hot water building plumbing systems are vulnerable to the proliferation of opportunistic pathogens (OPs), including Legionella pneumophila and Mycobacterium avium. Implementation of copper as a disinfectant could help reduce OPs, but a mechanistic understanding of the effects on the microbial community under real-world plumbing conditions is lacking. Here, we carried out a controlled pilot-scale study of hot water systems and applied shotgun metagenomic sequencing to examine the effects of copper dose (0-2 mg/L), orthophosphate corrosion control agent, and water heater anode materials (aluminum vs magnesium vs powered anode) on the bulk water and biofilm microbiome composition. Metagenomic analysis revealed that, even though a copper dose of 1.2 mg/L was required to reduce Legionella and Mycobacterium numbers, lower doses (e.g., ≤0.6 mg/L) measurably impacted the broader microbial community, indicating that the OP strains colonizing these systems were highly copper tolerant. Orthophosphate addition reduced bioavailability of copper, both to OPs and to the broader microbiome. Functional gene analysis indicated that both membrane damage and interruption of nucleic acid replication are likely at play in copper inactivation mechanisms. This study identifies key factors (e.g., orthophosphate, copper resistance, and anode materials) that can confound the efficacy of copper for controlling OPs in hot water plumbing.

A conserved pressure-driven mechanism for regulating cytosolic osmolarity.

Controlling intracellular osmolarity is essential to all cellular life. Cells that live in hypo-osmotic environments, such as freshwater, must constantly battle water influx to avoid swelling until they burst. Many eukaryotic cells use contractile vacuoles to collect excess water from the cytosol and pump it out of the cell. Although contractile vacuoles are essential to many species, including important pathogens, the mechanisms that control their dynamics remain unclear. To identify the basic principles governing contractile vacuole function, we investigate here the molecular mechanisms of two species with distinct vacuolar morphologies from different eukaryotic lineages: the discoban Naegleria gruberi and the amoebozoan slime mold Dictyostelium discoideum. Using quantitative cell biology, we find that although these species respond differently to osmotic challenges, they both use vacuolar-type proton pumps for filling contractile vacuoles and actin for osmoregulation, but not to power water expulsion. We also use analytical modeling to show that cytoplasmic pressure is sufficient to drive water out of contractile vacuoles in these species, similar to findings from the alveolate Paramecium multimicronucleatum. These analyses show that cytoplasmic pressure is sufficient to drive contractile vacuole emptying for a wide range of cellular pressures and vacuolar geometries. Because vacuolar-type proton-pump-dependent contractile vacuole filling and pressure-dependent emptying have now been validated in three eukaryotic lineages that diverged well over a billion years ago, we propose that this represents an ancient eukaryotic mechanism of osmoregulation.

Pilot-scale assessment reveals effects of anode type and orthophosphate in governing antimicrobial capacity of copper for Legionella pneumophila control.

Copper (Cu) is sometimes applied as an antimicrobial for controlling Legionella in hot water plumbing systems, but its efficacy is inconsistent. Here we examined the effects of Cu (0 - 2 mg/L), orthophosphate corrosion inhibitor (0 or 3 mg/L as phosphate), and water heater anodes (aluminum, magnesium, and powered anodes) on both bulk water and biofilm-associated L. pneumophila in pilot-scale water heater systems. Soluble, but not total, Cu was a good predictor of antimicrobial capacity of Cu. Even after months of exposure to very high Cu levels (>1.2 mg/L) and low pH (<7), which increases solubility and enhances bioavailability of Cu, culturable L. pneumophila was only reduced by ∼1-log. Cu antimicrobial capacity was shown to be limited by various factors, including binding of Cu ions by aluminum hydroxide precipitates released from corrosion of aluminum anodes, higher pH due to magnesium anode corrosion, and high Cu tolerance of the outbreak-associated L. pneumophila strain that was inoculated into the systems. L. pneumophila numbers were also higher in several instances when Cu was dosed together with orthophosphate (e.g., with an Al anode), revealing at least one scenario where high levels of total Cu appeared to stimulate Legionella. The controlled, pilot-scale nature of this study provides new understanding of the limitations of Cu as an antimicrobial in real-world plumbing systems.

Did a Nocebo Effect Contribute to the Rise in Special Education Enrollment Following the Flint, Michigan Water Crisis?

Background: Exposure to waterborne lead during the Flint Water Crisis during April 2014-October 2015 is believed to have caused increased special education enrollment in Flint children. Method: This retrospective population-based cohort study utilized de-identified data for children under six years of age who had their blood lead tested during 2011 to 2019, and special education outcomes data for children enrolled in public schools for corresponding academic years (2011-12 to 2019-20) in Flint, Detroit (control city) and the State of Michigan. Trends in the following crisis-related covariates were also evaluated: waterborne contaminants, poverty, nutrition, city governance, school district policies, negative community expectations, media coverage and social media interactions. Results: Between 2011 and 2019, including the 2014-15 crisis period, the incidence of elevated blood lead in Flint children (≥ 5µg/dL) was always at least 47% lower than in the control city of Detroit (p < .0001) and was also never significantly higher than that for all children tested in Michigan (p = 0.33). Nonetheless, special education enrollment in Flint spiked relative to Detroit and Michigan (p < .0001). There is actually an inverse relationship between childhood blood lead and special education enrollment in Flint. Conclusion: This study failed to confirm any positive association between actual childhood blood lead levels and special education enrollment in Flint. Negative psychological effects associated with media predictions of brain damage could have created a self-fulfilling prophecy via a nocebo effect. The findings demonstrate a need for improved media coverage of complex events like the Flint Water Crisis.

NSF Fellows' perceptions about incentives, research misconduct, and scientific integrity in STEM academia.

There is increased concern about perverse incentives, quantitative performance metrics, and hyper-competition for funding and faculty positions in US academia. Recipients of the prestigious National Science Foundation Graduate Research Fellowships (n = 244) from Civil and Environmental Engineering (45.5%) and Computer Science and Engineering (54.5%) were anonymously surveyed to create a baseline snapshot of their perceptions, behaviors and experiences. NSF Fellows ranked scientific advancement as the top metric for evaluating academics followed by publishing in high-impact journals, social impact of research, and publication/citation counts. The self-reported rate of academic cheating was 16.7% and of research misconduct was 3.7%. Thirty-one percent of fellows reported direct knowledge of graduate peers cheating, and 11.9% had knowledge of research misconduct by colleagues. Only 30.7% said they would report suspected misconduct. A majority of fellows (55.3%) felt that mandatory ethics trainings left them unprepared for dealing with ethical issues. Fellows stated academic freedom, flexible schedules and opportunity to mentor students were the most positive aspects of academia, whereas pressures for funding, publication, and tenure were cited as the most negative aspects. These data may be useful in considering how to better prepare STEM graduate trainees for academic careers.

A conserved pressure-driven mechanism for regulating cytosolic osmolarity.

Controlling intracellular osmolarity is essential to all cellular life. Cells that live in hypo-osmotic environments like freshwater must constantly battle water influx to avoid swelling until they burst. Many eukaryotic cells use contractile vacuoles to collect excess water from the cytosol and pump it out of the cell. Although contractile vacuoles are essential to many species, including important pathogens, the mechanisms that control their dynamics remain unclear. To identify basic principles governing contractile vacuole function, we here investigate the molecular mechanisms of two species with distinct vacuolar morphologies from different eukaryotic lineagesâ€"the discoban Naegleria gruberi , and the amoebozoan slime mold Dictyostelium discoideum . Using quantitative cell biology we find that, although these species respond differently to osmotic challenges, they both use actin for osmoregulation, as well as vacuolar-type proton pumps for filling contractile vacuoles. We also use analytical modeling to show that cytoplasmic pressure is sufficient to drive water out of contractile vacuoles in these species, similar to findings from the alveolate Paramecium multimicronucleatum . Because these three lineages diverged well over a billion years ago, we propose that this represents an ancient eukaryotic mechanism of osmoregulation.