Concentration and retention of Toxoplasma gondii oocysts by marine snails demonstrate a novel mechanism for transmission of terrestrial zoonotic pathogens in coastal ecosystems.

The parasite Toxoplasma gondii is an environmentally persistent pathogen that can cause fatal disease in humans, terrestrial warm-blooded animals and aquatic mammals. Although an association between T. gondii exposure and prey specialization on marine snails was identified in threatened California sea otters, the ability of kelp-dwelling snails to transmit terrestrially derived pathogens has not been previously investigated. The objective of this study was to measure concentration and retention of T. gondii by marine snails in laboratory aquaria, and to test for natural T. gondii contamination in field-collected snails. Following exposure to T. gondii-containing seawater, oocysts were detected by microscopy in snail faeces and tissues for 10 and 3 days respectively. Nested polymerase chain reaction was also applied as a method for confirming putative T. gondii oocysts detected in snail faeces and tissues by microscopy. Toxoplasma gondii was not detected in field-collected snails. Results suggest that turban snails are competent transport hosts for T. gondii. By concentrating oocysts in faecal pellets, snails may facilitate entry of T. gondii into the nearshore marine food web. This novel mechanism also represents a general pathway by which marine transmission of terrestrially derived microorganisms can be mediated via pathogen concentration and retention by benthic invertebrates.

Aquatic polymers can drive pathogen transmission in coastal ecosystems.

Gelatinous polymers including extracellular polymeric substances (EPSs) are fundamental to biophysical processes in aquatic habitats, including mediating aggregation processes and functioning as the matrix of biofilms. Yet insight into the impact of these sticky molecules on the environmental transmission of pathogens in the ocean is limited. We used the zoonotic parasite Toxoplasma gondii as a model to evaluate polymer-mediated mechanisms that promote transmission of terrestrially derived pathogens to marine fauna and humans. We show that transparent exopolymer particles, a particulate form of EPS, enhance T. gondii association with marine aggregates, material consumed by organisms otherwise unable to access micrometre-sized particles. Adhesion to EPS biofilms on macroalgae also captures T. gondii from the water, enabling uptake of pathogens by invertebrates that feed on kelp surfaces. We demonstrate the acquisition, concentration and retention of T. gondii by kelp-grazing snails, which can transmit T. gondii to threatened California sea otters. Results highlight novel mechanisms whereby aquatic polymers facilitate incorporation of pathogens into food webs via association with particle aggregates and biofilms. Identifying the critical role of invisible polymers in transmission of pathogens in the ocean represents a fundamental advance in understanding and mitigating the health impacts of coastal habitat pollution with contaminated runoff.

Research commentary: Association of zoonotic pathogens with fresh, estuarine, and marine macroaggregates.

Aquatic macroaggregates (flocs ≥ 0.5 mm) provide an important mechanism for vertical flux of nutrients and organic matter in aquatic ecosystems, yet their role in the transport and fate of zoonotic pathogens is largely unknown. Terrestrial pathogens that enter coastal waters through contaminated freshwater runoff may be especially prone to flocculation due to fluid dynamics and electrochemical changes that occur where fresh and marine waters mix. In this study, laboratory experiments were conducted to evaluate whether zoonotic pathogens (Cryptosporidium, Giardia, Salmonella) and a virus surrogate (PP7) are associated with aquatic macroaggregates and whether pathogen aggregation is enhanced in saline waters. Targeted microorganisms showed increased association with macroaggregates in estuarine and marine waters, as compared with an ultrapure water control and natural freshwater. Enrichment factor estimations demonstrated that pathogens are 2-4 orders of magnitude more concentrated in aggregates than in the estuarine and marine water surrounding the aggregates. Pathogen incorporation into aquatic macroaggregates may influence their transmission to susceptible hosts through settling and subsequent accumulation in zones where aggregation is greatest, as well as via enhanced uptake by invertebrates that serve as prey for marine animals or as seafood for humans.

Toxic diatoms and domoic acid in natural and iron enriched waters of the oceanic Pacific.

Near-surface waters ranging from the Pacific subarctic (58°N) to the Southern Ocean (66°S) contain the neurotoxin domoic acid (DA), associated with the diatom Pseudo-nitzschia. Of the 35 stations sampled, including ones from historic iron fertilization experiments (SOFeX, IronEx II), we found Pseudo-nitzschia at 34 stations and DA measurable at 14 of the 26 stations analyzed for DA. Toxin ranged from 0.3 fg·cell(-1) to 2 pg·cell(-1), comparable with levels found in similar-sized cells from coastal waters. In the western subarctic, descent of intact Pseudo-nitzschia likely delivered significant amounts of toxin (up to 4 µg of DA·m(-2)·d(-1)) to underlying mesopelagic waters (150-500 m). By reexamining phytoplankton samples from SOFeX and IronEx II, we found substantial amounts of DA associated with Pseudo-nitzschia. Indeed, at SOFeX in the Antarctic Pacific, DA reached 220 ng·L(-1), levels at which animal mortalities have occurred on continental shelves. Iron ocean fertilization also occurs naturally and may have promoted blooms of these ubiquitous algae over previous glacial cycles during deposition of iron-rich aerosols. Thus, the neurotoxin DA occurs both in coastal and oceanic waters, and its concentration, associated with changes in Pseudo-nitzschia abundance, likely varies naturally with climate cycles, as well as with artificial iron fertilization. Given that iron fertilization in iron-depleted regions of the sea has been proposed to enhance phytoplankton growth and, thereby, both reduce atmospheric CO(2) and moderate ocean acidification in surface waters, consideration of the potentially serious ecosystem impacts associated with DA is prudent.

The role of domoic acid in abortion and premature parturition of California sea lions (Zalophus californianus) on San Miguel Island, California.

Domoic acid is a glutaminergic neurotoxin produced by marine algae such as Pseudo-nitzschia australis. California sea lions (Zalophus californianus) ingest the toxin when foraging on planktivorous fish. Adult females comprise 60% of stranded animals admitted for rehabilitation due to acute domoic acid toxicosis and commonly suffer from reproductive failure, including abortions and premature live births. Domoic acid has been shown to cross the placenta exposing the fetus to the toxin. To determine whether domoic acid was playing a role in reproductive failure in sea lion rookeries, 67 aborted and live-born premature pups were sampled on San Miguel Island in 2005 and 2006 to investigate the causes for reproductive failure. Analyses included domoic acid, contaminant and infectious disease testing, and histologic examination. Pseudo-nitzschia spp. were present both in the environment and in sea lion feces, and domoic acid was detected in the sea lion feces and in 17% of pup samples tested. Histopathologic findings included systemic and localized inflammation and bacterial infections of amniotic origin, placental abruption, and brain edema. The primary lesion in five animals with measurable domoic acid concentrations was brain edema, a common finding and, in some cases, the only lesion observed in aborted premature pups born to domoic acid-intoxicated females in rehabilitation. Blubber organochlorine concentrations were lower than those measured previously in premature sea lion pups collected in the 1970s. While the etiology of abortion and premature parturition was varied in this study, these results suggest that domoic acid contributes to reproductive failure on California sea lion rookeries.

Fecal indicator bacteria and zoonotic pathogens in marine snow and California mussels (Mytilus californianus).

Pollution of nearshore waters with disease-causing microorganisms impacts ecosystems health through illness and deaths in people and wildlife, as well as negative socioeconomic consequences of impaired marine resources. Insight on pathogen ecology in coastal habitats is crucial for accurately mitigating inputs and impacts of microbial pollution. Three objectives were addressed to (i) compare fecal pollution in proximity to (a) freshwater runoff, and (b) endemic marine wildlife; (ii) evaluate presence and magnitude of fecal microorganisms in marine snow and mussels and (iii) determine if pathogens in mussels and FIB levels in seawater or mussels are correlated. Sampling during the wet season, proximity to freshwater, and FIB levels in mussel homogenates (but not seawater) were associated with pathogen presence in mussels. Pathogens and FIB were enriched in aggregate-rich fractions, further supporting an important role of marine snow in pathogen transmission. The lack of association between FIB in surrounding waters and presence of pathogens in mussels calls into question current regulations for insuring safe seafood to consumers in the United States, and alternative monitoring approaches such as direct testing for select pathogens should be further evaluated.

Respiratory Problems Associated with Surfing in Coastal Waters.

A pilot project was conducted to examine the health status and possible adverse health effects associated with seawater exposure (microbial water-quality indicators and phytoplankton abundance and their toxins) of surfers in Monterey Bay, Central California coastal waters. Forty-eight surfers enrolled in the study and completed an initial health background survey and weekly health surveys online using Survey Monkey. Descriptive statistics and generalized estimating equation, a regression technique, were used to identify longitudinal and correlated results. The surfers were predominately Caucasian, male, and physically active. They surfed approximately 4 h a week. Their average age was 34 years. The data indicated that the surfers were generally "healthy," with a low prevalence of diabetes, high cholesterol, and hypertension. Their most common health problems were allergies and asthma. During the study, 10% of the surfers reported gastrointestinal symptoms and 29% reported upper respiratory symptoms. This study suggests surfers were significantly more likely to report upper respiratory symptoms when they had a history of allergies, housemates with upper respiratory symptoms, and/or a history of previous adverse health symptoms while surfing during a "red tide" (an event often associated with the presence of phytoplankton toxins). Additionally, female surfers reported upper respiratory symptoms more than males.

Comparison of N-acetylcysteine and fenoldopam for preventing contrast-induced nephropathy (CAFCIN).

BACKGROUND: N-acetylcysteine and fenoldopam are commonly prescribed for prevention of contrast mediated nephropathy, however, comparative superiority of either agent is unknown. METHODS: In a prospective, randomized, parallel-group trial, adult cardiac catheterization patients at the university and veterans' hospitals with pre-existing stable renal insufficiency were randomized to N-acetylcysteine 600 mg orally twice daily for 4 doses or fenoldopam 0.1 mcg/kg/min intravenously for a minimum of 8 h. All patients received intravenous hydration with normal saline (5% dextrose in normal saline for diabetics on insulin). Randomization was stratified for diabetes. The primary endpoint was mean change in Scr at 72 h. Secondary endpoint was the incidence of contrast-induced nephropathy (25% increase above baseline Scr or absolute increase of 0.5 mg/dL). RESULTS: Study termination occurred after ninety-five patients (mean age 68+/-10 years, female 25%, diabetic 42%, mean baseline Scr 1.5+/-0.4 mg/dL) were randomized, with 84 completing follow-up (44 N-acetylcysteine, 40 fenoldopam). Overall, there were no significant differences in mean change in Scr at 72 h (N-acetylcysteine 0.20+/-0.72 vs. fenoldopam 0.08+/-0.48 mg/dL, p=0.4) or incidence of contrast-induced nephropathy (N-acetylcysteine 5 vs fenoldopam 8, p=0.4). No differences were detected in subgroup analyses for diabetes, baseline Scr >1.7 or 2.0 mg/dL, gender, age >70 years, or contrast volume >150 mL. Results were similar after multivariate adjustment for diabetes, contrast volume, heart failure and gender. CONCLUSIONS: Our randomized comparison failed to demonstrate a significant difference in the abilities of N-acetylcysteine and fenoldopam to prevent the decline in renal function or the incidence of contrast-induced nephropathy during cardiac catheterization.

From sanddabs to blue whales: the pervasiveness of domoic acid.

Domoic acid (DA) is a potent food web transferred algal toxin that has caused dramatic mortality events involving sea birds and sea lions. Although no confirmed DA toxicity events have been reported in whales, here we present data demonstrating that humpback and blue whales are exposed to the toxin and consume DA contaminated prey. Whale fecal samples were found to contain DA at levels ranging from 10 to 207microg DA g(-1) feces via HPLC-UV methods. SEM analysis of whale feces containing DA, collected from krill-feeding whales, revealed the presence of diatom frustules identified as Pseudo-nitzschia australis, a known DA producer. Humpback whales were observed feeding on anchovies and sardines that contained DA at levels ranging from 75 to 444microg DA g(-1) viscera. DA contamination of whale feces and fish occurred only during blooms of toxic Pseudo-nitzschia. Additionally, several novel fish species collected during a toxic diatom bloom were tested for DA. Fish as diverse as benthic sanddabs and pelagic albacore were found to contain the neurotoxin, suggesting that DA permeates benthic as well as pelagic communities.

A new pathogen transmission mechanism in the ocean: the case of sea otter exposure to the land-parasite Toxoplasma gondii.

Toxoplasma gondii is a land-derived parasite that infects humans and marine mammals. Infections are a significant cause of mortality for endangered southern sea otters (Enhydra lutris nereis), but the transmission mechanism is poorly understood. Otter exposure to T. gondii has been linked to the consumption of marine turban snails in kelp (Macrocystis pyrifera) forests. It is unknown how turban snails acquire oocysts, as snails scrape food particles attached to surfaces, whereas T. gondii oocysts enter kelp beds as suspended particles via runoff. We hypothesized that waterborne T. gondii oocysts attach to kelp surfaces when encountering exopolymer substances (EPS) forming the sticky matrix of biofilms on kelp, and thus become available to snails. Results of a dietary composition analysis of field-collected snails and of kelp biofilm indicate that snails graze the dense kelp-biofilm assemblage composed of pennate diatoms and bacteria inserted within the EPS gel-like matrix. To test whether oocysts attach to kelp blades via EPS, we designed a laboratory experiment simulating the kelp forest canopy in tanks spiked with T. gondii surrogate microspheres and controlled for EPS and transparent exopolymer particles (TEP - the particulate form of EPS). On average, 19% and 31% of surrogates were detected attached to kelp surfaces covered with EPS in unfiltered and filtered seawater treatments, respectively. The presence of TEP in the seawater did not increase surrogate attachment. These findings support a novel transport mechanism of T. gondii oocysts: as oocysts enter the kelp forest canopy, a portion adheres to the sticky kelp biofilms. Snails grazing this biofilm encounter oocysts as 'bycatch' and thereby deliver the parasite to sea otters that prey upon snails. This novel mechanism can have health implications beyond T. gondii and otters, as a similar route of pathogen transmission may be implicated with other waterborne pathogens to marine wildlife and humans consuming biofilm-feeding invertebrates.

Revisiting carbon flux through the ocean's twilight zone.

The oceanic biological pump drives sequestration of carbon dioxide in the deep sea via sinking particles. Rapid biological consumption and remineralization of carbon in the "twilight zone" (depths between the euphotic zone and 1000 meters) reduce the efficiency of sequestration. By using neutrally buoyant sediment traps to sample this chronically understudied realm, we measured a transfer efficiency of sinking particulate organic carbon between 150 and 500 meters of 20 and 50% at two contrasting sites. This large variability in transfer efficiency is poorly represented in biogeochemical models. If applied globally, this is equivalent to a difference in carbon sequestration of more than 3 petagrams of carbon per year.

A method of detecting and quantifying severity of myocardial perfusion defects with intravenous ultrasound contrast and breath holding during stress echocardiography.

Although breath holding is commonly used to improve and maintain image quality during stress echocardiography, its effects on the qualitative and quantitative analysis of myocardial contrast enhancement (MCE) following intravenously injected microbubbles is unknown. The purpose of this study was to determine how breath holding affects MCE following either an intravenous bolus or continuous infusion of perfluorocarbon containing microbubbles. In 48 patients, intravenous Optison was given at peak dobutamine stress to assess myocardial perfusion. The degree of myocardial opacification was assessed immediately following a breath hold in inspiration (BH(ini)), at the end of a breath hold (BH(term)), and following expiration and a subsequent second breath hold (BH(reinsp)). Pulmonary venous time velocity integrals were recorded during these different phases as well. Eleven patients had quantitative coronary angiography. Mean duration of the breath hold was 7 +/- 1 seconds. Pulmonary venous return fell by 29% +/- 18% at BH(term) (P < 0.001). There was complete disappearance of MCE at BH(term) in 27 of 35 bolus injection patients at peak stress, and no return of MCE following flash destruction during breath holding in 11 of 13 patients receiving continuous infusions. BH(reinsp) resulted in a boluslike return of contrast, with a transient, bright MCE in 44 of 48 patients, and a time intensity plot that resembled a gamma variate function. Perfusion defects were visualized in 25 patients during BH(ini) and 28 patients during BH(reinsp). Coronary artery territory agreement between perfusion assessed during BH(ini) and BH(reinsp) and quantitative coronary angiography was 76% and 81%, respectively.