Environmental exposure to active pharmaceutical ingredients (APIs) can have negative effects on the health of ecosystems and humans. While numerous studies have monitored APIs in rivers, these employ different analytical methods, measure different APIs, and have ignored many of the countries of the world. This makes it difficult to quantify the scale of the problem from a global perspective. Furthermore, comparison of the existing data, generated for different studies/regions/continents, is challenging due to the vast differences between the analytical methodologies employed. Here, we present a global-scale study of API pollution in 258 of the world’s rivers, representing the environmental influence of 471.4 million people across 137 geographic regions. Samples were obtained from 1,052 locations in 104 countries (representing all continents and 36 countries not previously studied for API contamination) and analyzed for 61 APIs. Highest cumulative API concentrations were observed in sub-Saharan Africa, south Asia, and South America. The most contaminated sites were in low- to middle-income countries and were associated with areas with poor wastewater and waste management infrastructure and pharmaceutical manufacturing. The most frequently detected APIs were carbamazepine, metformin, and caffeine (a compound also arising from lifestyle use), which were detected at over half of the sites monitored. Concentrations of at least one API at 25.7% of the sampling sites were greater than concentrations considered safe for aquatic organisms, or which are of concern in terms of selection for antimicrobial resistance. Therefore, pharmaceutical pollution poses a global threat to environmental and human health, as well as to delivery of the United Nations Sustainable Development Goals.

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In contrast to established zebrafish gene annotations, the question of sex determination has still not been conclusively clarified for developing zebrafish, Danio rerio, larvae, 28 dpf or earlier. Recent studies indicate polygenic sex determination (PSD), with the genes being distributed throughout the genome. Early genetic markers of sex in zebrafish help unravel co-founding sex-related differences to apply to human health and environmental toxicity studies. A qPCR-based method was developed for six genes: cytochrome P450, family 17, subfamily A, polypeptide 1 (cyp17a1); cytochrome P450, family 19, subfamily A, polypeptide 1a (cyp19a1a); cytochrome P450, family 19, subfamily A, polypeptides 1b (cyp19a1b); vitellogenin 1 (vtg1); nuclear receptor subfamily 0, group B, member 1 (nr0b1), sry (sex-determining region Y)-box 9b (sox9b) and actin, beta 1 (actb1), the reference gene. Sry-box 9a (Sox9a), insulin-like growth factor 3 (igf3) and double sex and mab-3 related transcription factor 1 (dmrt1), which are also known to be associated with sex determination, were used in gene expression tests. Additionally, Next-Generation-Sequencing (NGS) sequenced the genome of two adult female and male and two juveniles. PCR analysis of adult zebrafish revealed sex-specific expression of cyp17a1, cyp19a1a, vtg1, igf3 and dmrt1, the first four strongly expressed in female zebrafish and the last one highly expressed in male conspecifics. From NGS, nine female and four male-fated genes were selected as novel for assessing zebrafish sex, 28 dpf. Differences in transcriptomes allowed allocation of sex-specific genes also expressed in juvenile zebrafish.

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Invasive crayfish species were first documented in Switzerland in the 1970s. Today, North American crayfish species dominate in most major lakes and streams in Switzerland. In combination with the crayfish plague, they pose a substantial threat to our native crayfish. Over the past 20 years, various techniques have been applied to reduce negative impacts of these invasive crayfish in Switzerland: eradication (temporary drainage or destruction of a water system, biocides), suppression (intensive trapping, electricity introduction of predatory fish) and containment (construction of crayfish barriers). Temporary drainage or filling-in of isolated ponds, in combination with calcium hydroxide application has been successful in eradicating populations of invasive crayfish. However, trapping and introduction of predatory fish led to a reduction in population density but neither method has ever caused the extinction of a population. Invasive crayfish have not yet reached crayfish barriers, therefore, long-term functionality of these barriers still needs to be proven. Nevertheless, functional controls with native crayfish have shown that barriers prevent their upstream movement. Implementation of crayfish barriers is the most promising method to protect native crayfish from displacement by invasive crayfish species. Many measures are expensive, time consuming, and show little or no success in controlling invasive crayfish. Therefore, we recommend to focus on implementing drastic measures, such as filling-in or draining of isolated waters or a combination of various methods to maximise the reduction of population size.

Multiple parasites and pathogens cause disease in aquatic wildlife and in aquaculture species, generating a need for monitoring and management. Conventional disease monitoring methods involve laborious, costly, and invasive capture and examination of host species, and require specialized expertise for every host and pathogen of interest. Environmental DNA could provide simultaneous occurrence data for multiple pathogens across different host taxa, valuable for using parasite diversity as, for example, a bioindicator of ecosystem disturbance. Here, we tested the potential for simultaneous detection of four wildlife pathogens in water samples from 280, mainly riverine, sites across Switzerland. We targeted the crayfish pathogen Aphanomyces astaci, the amphibian pathogen Batrachochytrium dendrobatidis, and the fish pathogens Saprolegnia parasitica and Tetracapsuloides bryosalmonae. The eDNA detection showed a widespread distribution of A. astaci, S. parasitica, and T. bryosalmonae, although A. astaci and T. bryosalmonae were not detected in some alpine river catchments. B. dendrobatidis was detected only rarely, which was expected since the sampling did not target amphibian breeding sites. Co‐detection rates were higher in rivers than in lakes, likely reflecting the habitat preferences and distributions of the host species. We discuss the advantages and limitations of eDNA‐based pathogen monitoring and list a set of recommendations for managers. Our study illustrates how eDNA‐based techniques can monitor several pathogen species concurrently, thus facilitating more comprehensive disease monitoring schemes. Combined with metabarcoding approaches in the future, eDNA‐based sampling and detection can facilitate the incorporation of parasite and pathogen occurrence and diversity as an indicator for aquatic ecosystem health, and for revealing the hidden biodiversity and structure of parasite communities.

The sex of both humans and Danio rerio has previously been shown to affect the way individuals respond to drug exposure. Genes which allow identification of sex in juvenile zebrafish show potential to reveal these confounding variables between sex in toxicological and preclinical trials but the link between these is so far missing. These sex-specific, early expressed genes where expression is not altered by drug exposure must be carefully selected for this purpose. We aimed to discover genes which can be used in pharmaceutical trials and environmental toxicology studies to uncover sex-related variations in gene expression with drug application using the model organism Danio rerio. Previously published early sex determining genes from King et al. were evaluated as well as additional genes selected from our zebrafish Next-generation sequencing (NGS) data which are known from previously published works not to be susceptible to changes in expression with drug exposure. NGS revealed a further ten female-specific genes (vtg1, cyp17a1, cyp19a1a, igf3, ftz-f1, gdf9, foxl2a, Nr0b1, ipo4, lhcgr) and five male related candidate genes (FKBP5, apobb1, hbaa1, dmrt1, spata6) which are also expressed in juvenile zebrafish, 28 days post fertilisation (dpf). Following this, a literature review was performed to classify which of these early-expressed sex specific genes are already known to be affected by drug exposure in order to determine candidate genes to be used in pharmaceutical trials or environmental toxicology testing studies. Discovery of these early sex-determining genes in Danio rerio will allow identification of sex-related responses to drug testing to improve sex-specific healthcare and the medical treatment of human patients.