A case study on pharmaceutical residues and antimicrobial resistance genes in Costa Rican rivers: A possible route of contamination for feline and other species.

In this investigation, the presence of antibiotics and pharmaceuticals in Costa Rican surface waters, specifically in regions near feline habitats, was examined. The study revealed that 47% of the water samples contained detectable traces of at least one antibiotic. Ciprofloxacin and norfloxacin were the most frequently detected compounds, each with a detection rate of 27%. Other antibiotics, such as erythromycin, roxithromycin, and trimethoprim, were also found but at lower frequencies, around 14%. Notably, all antibiotic concentrations remained below 10 ng/L, with ciprofloxacin, norfloxacin, and erythromycin showing the highest concentrations. Furthermore, the investigation revealed the presence of non-antibiotic pharmaceutical residues in the water samples, typically at concentrations below 64 ng/L. Tramadol was the most frequently detected compound, present in 18% of the samples. The highest concentrations were observed for acetaminophen and tramadol, measuring 64 and 10 ng/L, respectively. Comparing these findings with studies conducted in treated wastewater and urban rivers, it became evident that the concentrations of antibiotics and pharmaceuticals were notably lower in this study. While previous research reported higher values, the limited number of studies conducted in protected areas raises concerns about the potential environmental impact on biodiversity. In summary, these results emphasize the importance of monitoring pharmaceutical residues and antimicrobial resistance genes ARGs in vulnerable ecosystems, especially those in close proximity to feline habitats in Costa Rica. Additionally, the study delved into the detection of (ARGs). All tested water samples were positive for at least one ARG, with the blaTEM gene being the most prevalent at 82%, followed by tetS at 64% and qnrB at 23%. Moreover, this research shed light on the complexity of evaluating ARGs in environmental samples, as their presence does not necessarily indicate their expression. It also highlighted the potential for co-selection and co-regulation of ARGs, showcasing the intricate behaviors of these genes in aquatic environments.

Passive monitoring techniques to evaluate environmental pesticide exposure: Results from the Infant's Environmental Health study (ISA).

BACKGROUND: Pesticides used in agriculture may expose populations living nearby. Costa Rica is a major banana-exporting country, its production depends on extensive pesticide use. OBJECTIVES: To evaluate environmental pesticide exposure, we measured levels of current-use pesticides in air and dust from 12 schools in Matina County, Costa Rica, with passive sampling methods. METHODS: We selected ten proximal and two non-proximal schools and placed polyurethane foam passive air samplers outdoors at each school, during four consecutive periods. At three of these schools, we also placed an active air sampler during the first 24 h of each sampling period. We collected passive dust samples by placing a glass Petri Dish at the inside of each school. We subsequently performed a chemical analysis of 18 pesticides, using gas chromatography with mass detector. RESULTS: With passive air samplers we detected ten different pesticides: two insecticides, two nematicides, and six fungicides, of which nine reported to be used on banana plantations. More than half of the samples contained at least five pesticides. Chlorpyrifos was detected most-frequently, in 98% of samples, followed by the nematicides etoprophos and the fungicide pyrimethanil that were both detected in 81% of samples. Chlorpyrifos concentrations were five times higher in proximal as compared to non-proximal schools: mean = 18.2 ng/m3 (range = 6.1-36.1) and mean = 3.5 ng/m3 (range= <0.5-11.4) and varied more between schools than in time (intra-class correlation coefficient = 0.80). In general, results from passive and active samplers showed similar exposure patterns; yet median concentrations tended to be higher in passive samplers. In dust samples, mostly fungicides were detected; chlorothalonil was detected most frequently, in 50% of samples. DISCUSSION: Passive air sampling is a promising technique to characterize environmental exposure to current-use pesticides; more studies are needed to characterize the sampling rates, reproducibility and optimum sampling times for passive samplers. School environments near banana plantations are contaminated with multiple pesticides that include insecticides, nematicides, and fungicides, which is of concern.

Effect of pesticides used in banana and pineapple plantations on aquatic ecosystems in Costa Rica.

Current knowledge on fate and effect of agricultural pesticides comes is mainly from temperate ecosystems. More studies are needed in tropical systems in order to assess contamination risks to nontarget endemic tropical species from the extensive use of pesticides e.g. in banana and pineapple plantations. In this study, acute laboratory toxicity tests with organophosphate pesticides ethoprophos and chlorpyrifos were conducted on two Costa Rican species, cladoceran Daphnia ambigua and fish Parachromis dovii. Tests showed that chlorpyrifos was more toxic than ethoprophos to D. ambigua and P. dovii and that D. ambigua was also more sensitive than P. dovii to both pesticides. Additionally, bioassays were performed by exposing D. magna and P. dovii to contaminated water collected from the field. Chemical analyses of field water revealed that fungicides were generally the most frequent pesticide group found, followed by insecticides/nematicides and herbicides. The bioassays and values obtained from the literature confirmed that D. magna was more sensitive to pesticide contamination than P. dovii and that D. ambigua was more sensitive than D. magna, suggesting that the native cladoceran is a more suitable test species than its temperate counterpart. Species sensitivity distributions showed no significant difference in sensitivity between tropical and temperate fish and the arthropod species exposed to chlorpyrifos in this study. Choline esterase activity (ChE) was measured in P. dovii in laboratory tests in order to assess the applicability of this biomarker. ChE inhibition in P. dovii was observed in the laboratory at levels below the LC10 of both ethoprophos and chlorpyrifos, confirming that ChE is an efficient biomarker of exposure. Both indigenous Costa Rican species used in this study were found to be suitable standard tropical test species. Further studies are needed to investigate how protective the safe environmental concentrations, derived from LC50 of native tropical species, are for protecting tropical aquatic natural communities.

Immunological and physiological responses to predation risk and sublethal concentrations of chlorothalonil and ß-endosulfan in Lithobates taylori (Anura: Ranidae) tadpoles.

Pesticide exposure and its interaction with other natural stressors can play a role in amphibian population declines because disruptions in stress hormone regulatory mechanisms may inhibit immune responses during metamorphosis. Here, we determined the interactive effects of predation risk and sublethal concentration of two pesticides on immunological and physiological responses in tadpoles of the tropical frog Lithobates taylori. Using mesocosms, we used chronic exposure to three levels of chlorothalonil and ß-endosulfan in the presence or absence of Odonate larvae. Our results show that ß-endosulfan in high concentrations reduced the weight of the tadpoles and increased the neutrophil count and corticosterone (CORT) levels. Larval development was accelerated by high concentrations of chlorothalonil. Also, this pesticide in low and high concentrations increases the absolute values of lymphocytes. Tadpoles exposed to chlorothalonil increased the numbers of monocytes (in low concentrations), and lymphocytes (in high and low concentrations). The interactions of the low concentrations of both pesticides with and without the predator's presence also increased the number of lymphocytes. A combination of pesticides increases the number of lymphocytes in the blood due to synergistic cytotoxicity. This study proves that ß- endosulfan elevates circulating CORT and thus generates physiological stress in tadpoles. Given that both pesticides are widely used within the distribution of L. taylori in Costa Rica, it is likely that tadpoles' development and immune function are altered by pesticide use. In combination with stressors such as emerging diseases and altered precipitation regimes, widespread agrochemical uses likely caused this species enigmatic decline in recent decades.

Biotransformation and oxidative stress responses in fish (Astyanax aeneus) inhabiting a Caribbean estuary contaminated with pesticide residues from agricultural runoff.

The estuarine ecosystem of Madre de Dios Lagoon (MDL), in the Caribbean Coast of Costa Rica, is exposed to contamination with pesticide residues coming from the upstream agricultural areas. Biomarkers can provide a better indication of the fitness of biota in real mixture exposure scenarios than traditional lethal dose toxicity measurements. Here, we measured biomarkers of biotransformation, oxidative stress, and neurotoxicity on Astyanax aeneus, an abundant fish species in MDL. Glutathione S-transferase activity (GST), catalase activity (CAT), lipid peroxidation (LPO), and cholinesterase activity (ChE) were measured in fish collected during seven sampling campaigns, carried out between 2016 and 2018. Pesticide residues were analyzed in surface water samples collected every time fish were sampled. Residues of 25 pesticides, including fungicides, insecticides, and herbicides, were detected. The biomarkers measured in A. aeneus varied along the sampling moments, with biotransformation and oxidative stress signals showing a coupled response throughout the assessment. Furthermore, significant correlations were established between three biomarkers (GST, LPO, and CAT) and individual pesticides, as well as between GST and LPO with groups of pesticides with shared biocide action. Among pesticides, insecticide residues had a major influence on the responses observed in fish. This work demonstrates the chronic exposure to pesticide residues in MDL and how such exposure is related to physiological responses in fish that can affect their health and potentially, the trophic networks. This early warning information should be considered to improve the protection of estuarine ecosystems in the tropics.

Pesticide use in banana plantations in Costa Rica - A review of environmental and human exposure, effects and potential risks.

Biodiversity is declining on a global scale. Especially tropical ecosystems, containing most of the planetary biodiversity, are at risk. Agricultural monocrop systems contribute to this decline as they replace original habitats and depend on extensive use of synthetic pesticides that impact ecosystems. In this review we use large-scale banana production for export purposes in Costa Rica as an example for pesticide impacts, as it is in production for over a century and uses pesticides extensively for more than fifty years. We summarise the research on pesticide exposure, effects and risks for aquatic and terrestrial environment, as well as for human health. We show that exposure to pesticides is high and relatively well-studied for aquatic systems and humans, but hardly any data are available for the terrestrial compartment including adjacent non target ecosystems such as rainforest fragments. Ecological effects are demonstrated on an organismic level for various aquatic species and processes but are not available at the population and community level. For human health studies exposure evaluation is crucial and recognised effects include various types of cancer and neurobiological dysfunctions particularly in children. With the many synthetic pesticides involved in banana production, the focus on insecticides, revealing highest aquatic risks, and partly herbicides should be extended to fungicides, which are applied aerially over larger areas. The risk assessment and regulation of pesticides so far relies on temperate models and test species and is therefore likely underestimating the risk of pesticide use in tropical ecosystems, with crops such as banana. We highlight further research approaches to improve risk assessment and, in parallel, urge to follow other strategies to reduce pesticides use and especially hazardous substances.

Indigenous children living nearby plantations with chlorpyrifos-treated bags have elevated 3,5,6-trichloro-2-pyridinol (TCPy) urinary concentrations.

BACKGROUND: The US Environmental Protection Agency voluntary phased-out residential use of chlorpyrifos in 2001. In contrast, in Costa Rica, chlorpyrifos-treated bags are increasingly used to protect banana and plantain fruits from insects and to fulfill product standards, even in populated areas. OBJECTIVES: To evaluate children's exposure to chlorpyrifos in villages situated nearby banana plantations and plantain farms in Costa Rica. METHODS: The study targeted two villages with use of chlorpyrifos-treated bags in nearby banana plantations and plantain farms and one village with mainly organic production. For 140 children from these villages, mostly indigenous Ngäbe and Bribri, parent-interviews and urine samples (n=207) were obtained. Urinary 3,5,6-trichloro-2-pyridinol (TCPy) levels were measured as a biomarker for chlorpyrifos exposure. In the banana and plantain village also environmental contamination to chlorpyrifos was explored. RESULTS: Children from the banana and plantain villages had statistically significant higher urinary TCPy concentrations than children from the referent village; 2.6 and 2.2 versus 1.3µg/g creatinine, respectively. Chlorpyrifos was detected in 30% of the environmental samples as well as in 92% of the hand/foot wash samples. For more than half of the children their estimated intake exceeded the US EPA chronic population adjusted dose. For some, the acute population adjusted dose and the chronic reference dose were also exceeded. CONCLUSIONS: Our results suggest that children living nearby plantations with chlorpyrifos-treated bags are exposed to chlorpyrifos levels that may affect their health. Interventions to reduce chlorpyrifos exposure are likely to improve children's health and environment in banana and plantain growing regions.

Ecotoxicological characterization of a tropical soil after diazinon spraying.

The impact of diazinon spraying in an agricultural tropical soil through the evaluation of both the habitat and retention functions of the soil system was never reported. To fill this gap, five times the recommended dose of a commercial diazinon formulation was sprayed in an agricultural area of Costa Rica, and dilution gradients of the sprayed soil were prepared in the laboratory. Avoidance and reproduction tests with soil organisms (Eisenia andrei, Enchytraeus crypticus and Folsomia candida) to evaluate losses in terrestrial habitat function, and growth and reproduction tests with aquatic organisms (Chlorella vulgaris and Daphnia magna, respectively) to evaluate the retention function of soil were performed. Results demonstrated that regarding habitat function, F. candida reproduction was the most sensitive endpoint (EC(50) = 0.288 mg a.i./kg), followed by avoidance behaviour of E. andrei (EC(20) = 1.75 mg a.i./kg). F. candida avoidance and the reproduction of E. andrei and E. crypticus were not affected by diazinon. The toxicity tests with aquatic organisms showed that the soil retention function was insufficient to prevent effects of diazinon either on microalgae growth (EC(50) ≤ 0.742 mg/L and EC(20) ≤ 0.223 mg/L) and on the reproduction of the cladoceran (EC(50) ≤ 0.00771 mg/L and EC(20) ≤ 0.00646 mg/L). Results suggested that diazinon exerted toxic effects even at the dilution corresponding to the recommended dose, fact which makes its misuse an issue of environmental concern. Furthermore, the present study highlighted the importance and complementary nature of the assessment of both habitat and retention functions to an ecological risk assessment in tropical systems.

Ecological Integrity Impairment and Habitat Fragmentation for Neotropical Macroinvertebrate Communities in an Agricultural Stream.

The Volcán River watershed in the south Pacific of Costa Rica comprises forests, small urban settlements, cattle fields, and intensive agriculture (mostly pineapple and sugarcane). The ecological integrity and quality of its waters was assessed from 2011-2013 and 2018-2019 by means of physical-chemical parameters (pH, conductivity, temperature, DO, DBO, nitrate, total phosphorus, and pesticide residues) and benthic macroinvertebrate (MI) sampling in eight sites (Volcán, Cañas, and Ángel Rivers, and Peje and Maura streams), resulting in high ecological integrity in all sites except the Peje stream, which is polluted with nitrates and pesticides. Only in this stream was there a marked seasonal variation in the abundance of 16 MI families including Leptohyphidae, Leptophlebiidae, Philopotamidae, Glossossomatidae, and Corydalidae, among others, whose presence was limited exclusively to the dry season (December to April), disappearing from the stream in the rainy season, with corresponding peaks in nitrate (max 20.3 mg/L) and pesticides (mainly herbicides and organophosphate insecticides). The characteristics of the watershed, with large areas of forest and excellent water quality, allow for the re-colonization of organisms into the Peje stream; however, those organisms are incapable of development and growth, providing evidence of a contaminant-driven habitat fragmentation in this stream during the rainy season.

Effects of the insecticide ß-endosulfan on tadpoles of Isthmohyla pseudopuma (Anura: Hylidae).

Conventional agriculture uses pesticides intensively. Once pesticides are released into the environment, they can be toxic to non-target organisms. Exposure of amphibians to pesticides can be lethal and affect their growth, development and behavior. ß-endosulfan is a persistent organochlorine that has been detected in environmental samples within protected sites in Costa Rica, far from agricultural areas. The aim of this study was to evaluate the lethal and sublethal effects, as well as changes in three biomarkers (Cholinesterase activity [ChE], glutathione S-transferase activity [GST] and lipid peroxidation [LPO]) in tadpoles of Isthmohyla pseudopuma exposed to ß-endosulfan. A 96-h acute test (20, 40, 60, 80, 100 and 200 µg/L) was performed in order to calculate the median lethal concentration (LC50), while effects on growth and development were assessed during a 4-weeks chronic test (10, 20, 30 and 50 µg/L). In addition, we measured the aforementioned biomarkers in tadpoles exposed to concentrations below the LC50. The 96-h LC50 for this species was 123.6 µg/L. We found no evidence of ß-endosulfan influencing any of the three biomarkers evaluated. At 50 µg/L, both length and total weight of tadpoles decreased with respect to the control. Also, at 30 and 50 µg/L we observed that individuals showed a slower development. Therefore, we demonstrated that at sublethal concentrations, ß-endosulfan negatively affects I. pseudopuma at early stages causing tadpoles to develop slower and smaller than normal.

Pyrimethanil and chlorpyrifos air concentrations and pregnant women's urinary metabolites in the Infants' Environmental Health Study (ISA), Costa Rica.

BACKGROUND: Only few studies have compared environmental pesticide air concentrations with specific urinary metabolites to evaluate pathways of exposure. Therefore, we compared pyrimethanil and chlorpyrifos concentrations in air with urinary 4-hydroxypyrimethanil (OHP, metabolite of pyrimethanil) and 3,5,6-trichloro-2-pyridinol (TCPy, metabolite of chlorpyrifos) among pregnant women from the Infant's Environmental Health Study (ISA) in Matina County, Costa Rica. METHODS: During pregnancy, we obtained repeat urinary samples from 448 women enrolled in the ISA study. We extrapolated pyrimethanil and chlorpyrifos concentrations measured with passive air samplers (PAS) (n = 48, from 12 schools), across space and time using a Bayesian spatiotemporal model. We subsequently compared these concentrationswith urinary OHP and TCPy in 915 samples from 448 women, usingseparatemixed models andconsidering several covariables. RESULTS: A 10% increase in air pyrimethanil (ng/m3) was associated with a 5.7% (95% confidence interval (CI 4.6, 6.8) increase in OHP (µg/L). Women living further from banana plantations had lower OHP: -0.7% (95% CI -1.2, -0.3) for each 10% increase in distance (meters) as well as women who ate rice and beans ≥15 times a week -23% (95% CI -38, -4). In addition, each 1 ng/m3 increase in chlorpyrifos in air was associated with a 1.5% (95% CI 0.2, 2.8) increase in TCPy (µg/L), and women working in agriculture tended to have increased TCPy (21%, 95% CI -2, 49). CONCLUSION: The Bayesian spatiotemporal models were useful to estimate pyrimethanil and chlorpyrifos air concentrations across space and time. Our results suggest inhalation of pyrimethanil and chlorpyrifos is a pathway of environmental exposure. PAS seems a useful technique to monitor environmental current-use pesticide exposures. For future studies, we recommend increasing the number of locations of environmental air measurements, obtaining all air and urine measurements during the same month, and, ideally, including dermal exposure estimates as well.

Accumulation of heavy metals (Cd, Cr, Cu, Mn, Pb, Ni, Zn) in sediments, macroalgae (Cryptonemia crenulata) and sponge (Cinachyrella kuekenthali) of a coral reef in Moín, Limón, Costa Rica: An ecotoxicological approach.

Moín, on the Caribbean coast of Costa Rica, is a multi-use coastal zone with a variety of human activities that can cause metal pollution. With the purpose of assessing the current environmental burden due to heavy metal presence in the marine environment of Moín, and their bioaccumulation in organisms of the nearby coral reef, we determined seven metals in samples of bottom sediments, macroalgae (Cryptonemia crenulata) and sponge (Cinachyrella kuekenthali). The results were compared with samples from the southern Caribbean, an area with little human activity. Using ICP-MS, results showed a concentration range for sediments Mn > Cu > Zn > Cr > Ni > Pb > Cd, algae Mn > Cu > Zn > Ni > Cr > Pb > Cd and sponge Mn > Cu > Zn > Ni > Cr > Cd > Pb, relatively low concentrations overall and no differences observed between sites. Bioconcentration factor > 1 was determined for Cd, Cu, Ni and Zn, while concentrations in sediments were below the SQG thresholds. Our study provides the first data on metal concentrations in a macroalgae and a sponge from the Costa Rican Caribbean.

Pesticide sequestration in passive samplers (SPMDs): considerations for deployment time, biofouling, and stream flow in a tropical watershed.

Semi-permeable membrane devices (SPMDs) provide an informative and cost-effective approach for monitoring contaminants in remote tropical streams. Estimation and interpretation of contaminant concentrations in streams derived from SPMDs can vary based on a number of environmental factors, including stream flow, biofouling, and deployment time. In three one-month long trials, SPMDs were concurrently deployed for 4, 15, and 28 days at three stream sites in an extensive agricultural area of southeastern Costa Rica. Water, bottom sediment, and suspended solids grab samples were also collected and several environmental variables were monitored at corresponding time intervals during each month-long study period. At all three sites, SPMD concentrations of the widely used insecticide chlorpyrifos increased with deployment time, with no relationship between SPMD biofouling and pesticide sequestration. Differences in SPMD chlorpyrifos sequestration among sites are likely due to differences in stream chlorpyrifos concentration rather than differences in SPMD sampling rates. The longer exposure period of SPMDs allowed for the detection of lower concentrations of chlorpyrifos, terbufos, and difenoconazole compared to water grab samples. In addition to the use of appropriate performance reference compounds (PRCs), other environmental variables such as stream turbidity, flow regime, stream morphology, and knowledge of pesticide application methods are important considerations for optimizing SPMD deployment and data interpretation in tropical regions.

Tracking pesticide fate in conventional banana cultivation in Costa Rica: A disconnect between protecting ecosystems and consumer health.

Conventional banana cultivation in Costa Rica relies on heavy pesticide use. While pesticide residues in exported bananas do not generally represent a safety concern for consumers abroad, ecosystem and human health in producing regions are not likewise protected. In Costa Rica, most studies on pesticide residues in the environment are snapshots, limiting our ability to identify temporal dynamics that can inform risk mitigation strategies. To help bridge this gap, we created a dynamic multimedia model for the Caño Azul River drainage area, which is heavily influenced by banana and pineapple plantations. This model estimates chemical concentrations in water, air, soil, sediments, and banana plants through time, based on pesticide properties and emission patterns and on variable environmental conditions. Case studies for three representative chemicals-the herbicide diuron, the nematicide ethoprofos, and the fungicide epoxiconazole-show that concentrations in fruit remain below EU and US maximum residue limits set to ensure consumer health, while those in the environment are highly variable, reaching peak concentrations in water that can exceed thresholds for ecosystem health. Critical research needs, including incorporating sediment dynamics and the effects of adjuvants on the properties and transport of active ingredients into multimedia models, were identified.

Environmental risk assessment of pesticides in the River Madre de Dios, Costa Rica using PERPEST, SSD, and msPAF models.

This study assesses the ecological risks (ERA) of pesticides to aquatic organisms in the River Madre de Dios (RMD), which receives surface runoff water from banana, pineapple, and rice plantations on the Caribbean coast of Costa Rica. Water samples collected over 2 years at five sites in the RMD revealed a total of 26 pesticides. Their toxicity risk to aquatic organisms was assessed using three recent ERA models. (1) The PERPEST model showed a high probability (>50 %) of clear toxic effects of pesticide mixtures on algae, macrophytes, zooplankton, macroinvertebrates, and community metabolism and a low probability (<50 %) of clear effects on fish. (2) Species sensitivity distributions (SSD) showed a moderate to high risk of three herbicides: ametryn, bromacil, diuron and four insecticides: carbaryl, diazinon, ethoprophos, terbufos. (3) The multi-substance potentially affected fraction (msPAF) model showed results consistent with PERPEST: high risk to algae (maximum msPAF: 73 %), aquatic plants (61 %), and arthropods (25 %) and low risk to fish (0.2 %) from pesticide mixtures. The pesticides posing the highest risks according to msPAF and that should be substituted with less toxic substances were the herbicides ametryn, diuron, the insecticides carbaryl, chlorpyrifos, diazinon, ethoprophos, and the fungicide difenoconazole. Ecological risks were highest near the plantations and decreased progressively further downstream. The risk to fish was found to be relatively low in these models, but water samples were not collected during fish kill events and some highly toxic pesticides known to be used were not analyzed for in this study. Further sampling and analysis of water samples is needed to determine toxicity risks to fish during peaks of pesticide mixture concentrations. The msPAF model, which estimates the ecological risks of mixtures based on their toxic modes of action, was found to be the most suitable model to assess toxicity risks to aquatic organisms in the RMD. The PERPEST model was found to be a strong tool for screening risk assessments. The SSD approach is useful in deriving water quality criteria for specific pesticides. This study, through the application of three ERA models, clearly shows that pesticides used in plantations within the RMD watershed are expected to have severe adverse effects on most groups of aquatic organisms and that actions are urgently needed to reduce pesticide pollution in this high biodiversity ecosystem.

Risk assessment of agriculture impact on the Frío River watershed and Caño Negro Ramsar wetland, Costa Rica.

The Caño Negro Ramsar wetland is a conservation area of great natural and societal value, located in the lower part of the Frío River watershed in the north of Costa Rica. Its aquatic ecosystems may be considered vulnerable to pollution due to recent changes in land use toward agriculture. In 2011 and 2012, quarterly sampling was done at ten sites located in the middle and lower sections of the Frío River Basin that pass through crop areas and later drain into Caño Negro wetland. Pesticide residues, nitrates, sediment concentrations, and diversity of benthic macroinvertebrates and fish biomarkers were studied in the selected sites. Additionally, risk of toxicity was calculated in two different ways: (1) by using a ratio of MEC to hazard concentrations threshold for 5% of species (HC5) to calculate a risk quotient (RQ), and (2) by using a ratio of MEC to available ecotoxicity data of native fish and cladocera for diazinon and ethoprophos, to obtain a risk quotient for native species (RQns). Results indicated that three out of the ten sites (rivers Thiales, Mónico, and Sabogal) showed variable levels of pollution including six different active ingredients (a.i.) of pesticide formulations (herbicides ametryn, bromacil, and diuron; insecticides cypermethrin, diazinon, and ethoprophos). Moreover, potential adverse effects on fishes in Thiales and Mónico rivers were indicated by cholinesterase (ChE) inhibition and glutathione S-transferase (GST) enhancement. Risk evaluations indicated pesticide residues of ametryn, bromacil, and ethoprophos to be exceeding the limits set by MTR, also RQ was high (>1) in 70% of the positive samples for diuron (most frequently found pesticide in water samples), cypermethrin, diazinon, and ethoprophos, and RQns was high for diazinon. Therefore, these substances might be of major concern for the ecological health of aquatic ecosystems in the middle basin of the Frío River. The most critical site was Mónico River, which had the highest pollution (75% detection samples with 3-5 a.i.) and highest calculated risk (RQ > 1 in 75% of the samples). This is also the river that most directly drains into the protected wetland. Even though pesticide pollution in this area is not as severe as in other parts of Costa Rica, it is imperative that measures are taken, particularly in the surroundings of Mónico River, in order to diminish and mitigate possible detrimental effects to biota in Caño Negro Ramsar Site.

In situ toxicity and ecological risk assessment of agro-pesticide runoff in the Madre de Dios River in Costa Rica.

The River Madre de Dios (RMD) and its lagoon is a biodiversity rich watershed formed by a system of streams, rivers, channels, and a coastal lagoon communicating with the Caribbean Sea. This basin sustains a large area of agricultural activity (mostly banana, rice, and pineapple) with intensive use of pesticides, continually detected in water samples. We investigated in situ the toxicological effects caused by pesticide runoff from agriculture and the relation of pesticide concentrations with different biological organization levels: early responses in fish biomarkers (sub-organismal), acute toxicity to Daphnia magna (organismal), and aquatic macroinvertebrate community structure. The evaluation was carried out between October 2011 and November 2012 at five sites along the RMD influenced by agricultural discharges and a reference site in a stream outside the RMD that receives less pesticides. Acute toxicity to D. magna was observed only once in a sample from the RMD (Caño Azul); the index of biomarker responses in fish exposed in situ was higher than controls at the same site and at the RMD-Freeman. However, only macroinvertebrates were statistically related to the presence of pesticides, combined with both physical-chemical parameters and habitat degradation. All three groups of variables determined the distribution of macroinvertebrate taxa through the study sites.

Occurrence and fate of illicit drugs and pharmaceuticals in wastewater from two wastewater treatment plants in Costa Rica.

Chemical analysis of raw wastewater in order to assess the presence of biological markers entering a wastewater treatment plant can provide objective information about the health and lifestyle of the population connected to the sewer system. This work was performed in a tropical country of Central America, Costa Rica, with the aim of extending this knowledge to new world regions. This work is the first to report wastewater-based epidemiological data on the use of illicit drugs in this region of the world. Composite wastewater samples from the influents of two different wastewater treatment facilities and surface water samples from surrounding areas were collected applying the best practice protocol and analysed to investigate the occurrence and fate of selected illicit drugs of abuse and pharmaceuticals. Results showed the presence of chemical indicators of the classic drugs cocaine and cannabis at high concentration levels, besides the moderate presence of the opiates codeine and morphine. Neither the worldwide commonly used psychoactive substances of abuse such as synthetic phenethylamines, nor pharmaceuticals from the family of benzodiazepines were detected, demonstrating the spatial differences in drug use among different world regions. In addition, effluent wastewater samples were analysed and compared to influent concentrations in order to evaluate the decrease in concentration of the targeted analytes through two treatment technologies. As a final step, a wide-scope qualitative screening, including hundreds of suspect compounds, was applied in order to have a better knowledge on the presence of pharmaceuticals in waters and to assess the potential impact of the treated wastewater into the receiving aquatic ecosystems.

Water quality and macroinvertebrate community response following pesticide applications in a banana plantation, Limon, Costa Rica.

Pesticides used in banana production may enter watercourses and pose ecological risks for aquatic ecosystems. The occurrence and effects of pesticides in a stream draining a banana plantation was evaluated using chemical characterization, toxicity testing and macrobenthic community composition. All nematicides studied were detected in the surface waters of the banana plantation during application periods, with peak concentrations following applications. Toxicity tests were limited to the carbofuran application and no toxicity was observed with the acute tests used. However, since pesticide concentrations were generally below the lowest LC50 value for crustaceans but above calculated aquatic quality criteria, there remains a risk of chronic toxicity. Accurate ecological assessments of pesticide use in banana plantations are currently limited by the lack of local short-term chronic toxicity tests and tests using sensitive native species. Relatively constant levels of four pesticides (imazalil, thiabendazole, chlorpyrifos and propiconazole), which had toxic effects according to the 96h hydra and 21d daphnia chronic test, were recorded in the effluent of the packing plant throughout the study, indicating that the solid waste trap used in this facility was not effective in eliminating toxic chemicals. Certain taxa, such as Heterelmis sp. (Elmidae), Heteragrion sp. (Megapodagrionidae, Odonata), Caenis sp. (Caenidae, Ephemerotera), and Smicridea sp. (Hidropsychidae, Trichoptera), were more abundant at reference sites than in the banana farm waters, and may be good candidates for toxicity testing. Multivariate analyses of the macroinvertebrate communities clearly showed that the banana plantation sites were significantly different from the reference sites. Moreover, following the pesticide applications, all the banana plantation sites showed significant changes in community composition, with the same genera being affected at all sites and for all pesticides (terbufos, cadusafos and carbofuran). Consequently, the results presented here show that multivariate analysis of community composition was more sensitive in distinguishing pesticide effects than the toxicity tests and richness and composition measures used. We conclude that monitoring macroinvertebrate communities can be a powerful tool in the assessment of ecological effects of banana production.

Acute, chronic and biochemical effects of chlorothalonil on Agalychnis callidryas, Isthmohyla pseudopuma and Smilisca baudinii tadpoles.

Declines of amphibian populations have been a worldwide issue of concern for the scientific community during the last several decades. Efforts are being carried out to elucidate factors related to this phenomenon. Among these factors, pathogens, climate change, and environmental pollution have been suggested as possible causes. Regarding environmental pollutants, some pesticides are persistent in the environment and capable of being transported long distances from their release point. In Costa Rica, some pesticides have been detected in protected areas, at locations where amphibian populations have declined. Information about toxicity of pesticides used in Costa Rican agriculture to amphibians is still scarce, particularly for native species.Toxicity tests with chlorothalonil, a fungicide intensively used in Costa Rica, were carried out exposing tadpoles of three Costa Rican native species: Agalychnis callidryas, Isthmohyla pseudopuma, and Smilisca baudinii in order to evaluate acute and chronic toxicity as well as the biomarkers cholinesterase activity (ChE), glutathione-S transferase activity (GST), and lipid peroxidation (LPO).96-h LC50: 26.6 (18.9-35.8) µg/L to A. callidryas, 25.5 (21.3-29.7) µg/L to I pseudopuma and 32.3 (26.3-39.7) µg/L to S. baudinii were determined for chlorothalonil. These three species of anurans are among the most sensitive to chlorothalonil according to the literature. Besides, GST was induced in S. baudinii after exposure to sub-lethal concentrations of chlorothalonil while evisceration occurred in S. baudinii and A. callidryas tadpoles exposed to lethal concentrations of the fungicide. Chronic exposure to sub-lethal concentrations accelerated development in S. baudinii and caused lesions in tail of S. baudinii and I. pseudopuma tadpoles. Our results demonstrate that chlorothalonil is highly toxic to native amphibian species and that low concentrations can cause biochemical responses related to phase II of biotransformation and effects on development.