Diversity of aquatic parasites in pristine spring waters in Tehsil Babuzai, Swat, Pakistan.

Global access to clean and safe drinking water remains a formidable challenge, contributing to a myriad of health issues. This research exposes the existence of waterborne parasites in seemingly pristine spring waters, indicating potential contamination. Daily extensive sampling of Seventeen water sources was conducted in the untarnished freshwater streams of Tehsil Babuzai, District Swat, Khyber Pakhtunkhwa, Pakistan, from February to September 2021. Employing a stringent filtration process, the collected samples were effectively concentrated to detect any waterborne parasites. Subsequent application of the wet mount technique, combined with the capabilities of a compound microscope, revealed a disconcerting reality: all examined samples tested positive for various parasites. Identified parasites included Schistosoma species, Ascaris lumbricoides, Trichiuria trichiuria, Taenia saginata, Entamoeba histolytica, Amoeba, Lacrymana olor, Tintinnids, Paramecium, Dileptus, Euglena, Loxodes striatus, Acanthocyclops lynceus, Spondylosium, Oscillatoria, Cyanobacteria, Cilindros, Cilindros cerro, Commensal amoeba mature cysts,, Filliform larva of Strongyloides, Cercaria larva, Larva of Taenia solium, Egg of Enterobius vermiculais, Egg of Isospora belli, Egg of Tapeworm, Egg of Schistosoma species, Egg of Toxocara, and Egg of Diphyllobothrium latum. These findings clearly demonstrate the presence of a diverse array of parasites in the freshwater springs of Tehsil Babuzai, Swat, Pakistan. Implementing robust water treatment protocols, conducting regular monitoring and testing, and raising awareness about the risks of waterborne parasites are crucial steps to safeguard public health in the region.

Extensive sampling and citizen science expand the distribution of the threatened freshwater turtle Ranacephala hogei (Mertens, 1967).

Ranacephala hogei is a South American freshwater turtle considered one of the 25 most endangered chelonian species in the world. Endemic to the Atlantic Forest hotspot of southeastern Brazil, the conservation status of R. hogei is subject to continuous assessment at various levels. However, the scarcity of data regarding this species, particularly its geographic range, challenges these evaluations. In an effort to address these gaps, we conducted a comprehensive long-term inventory using different methods to study this species. Our efforts resulted in a 144% increase in documented occurrence points, including a new hydrographic basin and protected areas. By combining historical and current records, we have observed the persistence of R. hogei in rural areas, even in locations where its extinction was previously predicted. Consequently, our data significantly contribute to supporting future conservation assessments.

Hazardous effects of nickel ferrite nanoparticles and nickel chloride in early life stages of the freshwater snail Biomphalaria glabrata (Say, 1818).

Nickel ferrite nanoparticles (NiF NPs) have growing applications in biomedical and nanomedicine fields. However, knowledge concerning their ecotoxicity during the early developmental stages of invertebrates, such as gastropods, remains scarce. Thus, the current study aimed to evaluate whether NiF NPs and nickel chloride (NiCl2) induce toxic effects on embryos and newly hatched snails of freshwater species Biomphalaria glabrata (Say, 1818). NiF NPs were synthesized and characterized by multiple techniques, and their ecotoxicity was assessed by Biomphalaria embryotoxicity test (BET) during 144 h of exposure and an acute toxicity test (96 h) using newly hatched snails. NiF NPs induced mortality, developmental delay, reduced hatching rate, and promoted morphological changes in B. glabrata. Also, NiF NPs induced higher toxicity in embryos than in newly hatched B. glabrata. Overall, results showed that the early developmental stages of gastropods are a target group for nanoparticle toxicity in freshwater ecosystems.

Reconstructing 90 years of anthropogenic activities in a subtropical reservoir: a chemometric and paleolimnological perspective.

Environmental reference conditions (RC) and historical trends are crucial for assessing the degree of freshwater impact and formulating restoration measures. This is particularly relevant for freshwater systems used as potable water sources. Using sediments from the Carlos Maggiolo reservoir (Minas, Uruguay), located in a watershed with a mining history, this study aimed to (1) establish metal (As, Cr, Cu, Ni, Pb, and Zn) RC using a predicted interval (PI) chemometric technique; (2) evaluate metal enrichment and toxicity over time and space; and (3) assess environmental changes examining geochemical proxies. Surface sediments from 29 stations were used to establish RC and a core from the dam area was sampled to infer past conditions. The sediments were submitted to partial digestion and analyzed by inductively coupled plasma optical emission spectrometry- ICP-OES. Enrichment factor (EF) calculated using both PI and bottom core values did not show significant differences over time. Over space and time, most metals primarily originated from natural sources (EF < 2). The PI in mg/kg was Cr: 23.74-37.32; Cu: 25.75-48.99; Ni: 16.29-25.55; Pb: 7.63-13.75; and Zn: 94.34-174.80. A stratigraphically constrained cluster analysis corroborated by a permutational multivariate analysis of variance categorized the reservoir into two zones: Zone I, before reservoir operation to ~ 1996, and Zone II from ~ 1997 to 2017. The average concentrations of the main metals of toxicological interest in zones I and II, respectively, were as follows: Cr 37.60 ± 1.59, 34.54 ± 1.49 mg/kg; Cu 49.76 ± 2.84, 44.55 ± 2.70 mg/kg; Ni 24.11 ± 0.67, 22.53 ± 1.22 mg/kg; Pb 12.40 ± 0.63, 13.52 ± 0.82 mg/kg; Zn 99.25 ± 3.12, 93.86 ± 4.42 mg/kg; Mn 1160.56 ± 68.88, 1441.61 ± 83.55 mg/kg; and P 1243.21 ± 271.56, 1128.42 ± 183.10 mg/kg. According to a principal component analysis, the period preceding reservoir operation until ~ 1985 was linked to mining activities and application of Cu2SO4, and the period from 1985s to 2000 was influenced by C and P concentrations due to increasing agricultural and afforestation activities in the watershed that consume P fertilizers. The most recent period, from 2000 to ~ 2017, was characterized by an increase in sedimentation rates mainly associated with erosion, particularly in the agricultural areas. This subsequent soil loss in the watershed could compromise the reservoir's useful life. This study contributes to a better understanding of metal geochemistry in subtropical reservoirs and aids in formulating effective recovery and restoration measures.

Remodeling of Uterine Tissues During Gestation of Potamotrygon wallacei (Elasmobranchii), a Neotropical Freshwater Stingray Endemic to the Negro River, Central Amazonia.

Neotropical freshwater stingrays of the subfamily Potamotrygoninae exhibit aplacental viviparity with uterine trophonemata. In this reproductive mode, females nourish and provide oxygenation to the embryo via the mucosa of the uterine wall. The aim of this study was to describe and histologically quantify the tissue components of the gravid uterus in an Amazonian freshwater stingray. Adult females of Potamotrygon wallacei were studied in different reproductive periods: resting stage, pregnant, and postpartum. During reproductive rest, the left ovary has numerous follicles compared to the right side. Therefore, uterine fertility is usually higher on the left side. The presence of an embryo in the right uterus suggests that the right ovary is also functional, although this only occurs in larger females. In females at reproductive rest, the wall of the uterus is formed by a mucosal layer (without the trophonemata) that contributes 16.7% to the thickness, while the myometrium accounts for 83.3% of the thickness. The mass-specific volume of the mucosal layer, inner circular, and outer longitudinal smooth muscle sheets tend to increase in the gravid uterus, indicating hypertrophy and hyperplasia of these components. During pregnancy, the trophonemata undergo marked tissue remodeling. Epithelial cells are organized into glandular acini and have apical secretory vesicles; furthermore, peripheral blood vessels proliferate and become dilated. These characteristics demonstrate that the gravid uterus of P. wallacei presents intense uterolactation activity and provides oxygenation to the fetus. Tissue remodeling occurs only in the uterus with the presence of an embryo. During postpartum, females have low body condition factor indicating a high reproductive cost. This study contributes to the knowledge of the reproductive biology of this species and will help us understand the impacts of climate change on the breeding areas of potamotrygonids.

Novel members of the order Picornavirales identified in freshwater from Guarapiranga reservoir in São Paulo.

The global challenge of water resource availability is exacerbated by anthropogenic influences that promote the emergence of pollutants. Among these pollutants are microbiological agents, including viruses, which are ubiquitous in the biosphere and play a pivotal role in both ecological balance and the occurrence of diseases in animals and plants. Consequently, monitoring viruses in water sources becomes indispensable for the establishment of effective prevention, promotion, and control strategies. Within this context, the study focuses on the identification of novel viruses belonging to the Picornavirales order in freshwater from the Guarapiranga Reservoir in the state of São Paulo, Brazil. The samples were subjected to viral metagenomics. Our analysis led to the characterization of four distinct sequences (GinkV-05, AquaV_10, MarV_14, and MarV_64), which exhibited significant divergence compared to other members of the Picornavirales order. This remarkable diversity prompted the identification of a potential new genus within the Marnaviridae family, tentatively named Ginkgonavirus. Additionally, we characterized four sequences in a very distinct clade and propose the recognition of a novel family (named Aquaviridae) within the Picornavirales order. Our findings contribute valuable insights into the previously uncharted diversity of Picornavirales present in water sources, shedding light on an important facet of viral ecology and evolution in aquatic environments.

Realistic nitrate concentrations diminish reproductive indicators in Skiffia lermae, an endemic species in critical endangered status.

Goodeinae is a subfamily of critically endangered fish native to central Mexico. Populations of Skiffia lermae, a species belonging to this subfamily, have significantly decreased in the past two decades. A previous study showed that S. lermae is sensitive to acute nitrate-nitrogen (NO3-N) exposure, leading to noticeable changes in both behavioral and histopathological bioindicators. The aim herein was to determine the vulnerability of S. lermae to NO3-N exposure at realistic concentrations registered in freshwater ecosystems in central Mexico where the species was historically reported. Offspring of S. lermae were chronically exposed during 60 days to concentrations of 5, 10 and 20 mg NO3-N/L, with 2 mg NO3-N/L used as the reference value (control). Survival rate, feeding behavior, aquatic surface respiration, body growth, scaled mass index, immature red blood cells, as well as histopathological changes in branchial, hepatic and gonadal tissues were evaluated. Additionally, this study analyzed water quality in freshwater ecosystems where S. lermae presently persists. The results showed decreased survival as NO3-N concentration increased, as well as increased feeding latency, aquatic surface respiration and histological damage in the gills and liver. These organs showed differential sex-dependent responses to NO3-N exposure; females were more sensitive than males. In the ovaries, a decreased density of stage III oocytes was associated with increased NO3-N concentrations. No changes were observed in body growth and number of immature red blood cells. Concentrations recorded in the three freshwater ecosystems that S. lermae inhabit were below 2 mg NO3-N/L. Together, the results could explain why the species has disappeared from more contaminated freshwater ecosystems where NO3-N levels exceed 5 mg/L. Moreover, the study warns about the risks of increasing NO3-N concentrations in the current sites where the species lives.

Ballast water transport of alien phytoplankton species to the brazilian amazon coast.

The aim of this study was to investigate the presence of alien phytoplankton species transported through ballast water of ships that docked on the Amazon coast. Phytoplankton samples were collected from 25 ships between 2012 and 2014, revealing 215 identified species, mostly comprising oceanic planktonic marine species. However, several coastal and freshwater species not yet documented on the Maranhão coast were also observed. The identification of several coastal and freshwater species not yet recorded for Amazonian environments in the ballast water of the Ponta da Madeira Maritime Terminal (TMPM), as well as toxic microalgae genera such as the dinoflagellates Alexandrium and Gymnodinium and of some diatom species from the genus Pseudo-nitzchia, raises concerns regarding the possibility of introducing species. This indicates that ballast water can be responsible for the introduction of alien species in Amazonian aquatic environments, thereby highlighting the TMPM as a critical hotspot in the Amazonian region.

Morphology of the Digestive Tube of the Amazonian Freshwater Stingray Potamotrygon wallacei (Elasmobranchii: Potamotrygonidae): A Stereological Approach.

This work aimed to describe and quantify the tissue components of the digestive tube of the neotropical freshwater stingray, Potamotrygon wallacei. For this, conventional histology and stereological methods were used to estimate tissue volume. The volumes of the four fundamental layers and the tissue components in the stomach (cardiac and pyloric) and spiral intestine were also estimated. In the cardiac stomach, the mucosa layer occupies 44.7% of the total volume of the organ wall. The gastric glands are the main components, and these structures alone represent 49.7% of this layer. This large number of gastric glands suggests a high potential for processing food items with a high protein content. The stereological methods were sensitive enough to show a reduction in the volume of the gastric glands from the cardiac region toward the pyloric region. Gastric glands are absent in the pyloric region of the stomach. However, the muscularis becomes thicker towards the pyloric region. The increase in smooth muscle thickness is due to the thickening of the inner muscular layer. This suggests that the role of the pyloric stomach may be related to the mixing of the chyme and assisting its passage to the spiral intestine. In the spiral intestine, data on the volume of the mucosa layer (and epithelial lining) suggest that the spiral valve has a large absorptive area. In several respects, the morphology of the digestive tube of P. wallacei is similar to that of other batoids. However, its slight morphological variations may be related to the habitat specificity of this species.

Critical swimming speed at different temperatures for small-bodied freshwater native riverine fish species.

This study evaluated the effect of fish total length (LT) and three water temperatures (10, 15 and 20 °C) on the critical swimming speed (Ucrit) of the species Percilia irwini (2.9-6.3 cm LT), Cheirodon galusdae (3.4-5.5 cm LT), and Trichomycterus areolatus (4.0-6.3 cm LT). An Ucrit estimation model was constructed for each species as a function of temperature and size. The results showed mean Ucrit for P. irwini of 44.56, 53.83 and 63.2 cm s-1 at 10, 15 and 20 °C, respectively: 55.34, 61.74 and 70.05 cm s-1 for C. galusdae and 56.18, 63.01 and 71.09 cm s-1 for T. areolatus. Critical velocity depended on the interaction between species, body length and water. The swimming performance increased significantly with rising temperature in all three species. The velocity also increased with greater fish total length. After controlling for fish total length, velocity also increased with higher temperature in the three species. This research is relevant to small fish species that require conservation measures.

Taylor's power law for freshwater fishes: Functional traits beyond statistical inevitability.

Taylor's power law (TPL) describes the expected range of parameters of the mean-variance scaling relationship and has been extensively used in studies examining temporal variations in abundance. Few studies though have focused on biological and ecological covariates of TPL, while its statistical inherences have been extensively debated. In the present study, we focused on species-specific features (i.e. functional traits) that could be influential to temporal TPL. We combined field surveys of 180 fish species from 972 sites varying from small streams to large rivers with data on 31 ecological traits describing species-specific characteristics related to three main niche dimensions (trophic ecology, life history, and habitat use). For each species, the parameters of temporal TPL (intercept and slope) were estimated from the log-log mean-variance relationships while controlling for spatial dependencies and biological covariates (species richness and evenness). Then, we investigated whether functional traits explained variations in TPL parameters. Differences in TPL parameters among species were explained mostly by life history and environmental determinants, especially TPL slope. Life history was the main determinant of differences in TPL parameters and thereby aggregation patterns, with traits related to body size being the most influential, thus showing a high contrast between small-sized species with short lifespans and large-bodied migratory fishes, even after controlling for phylogenetic resemblances. We found that life history traits, especially those related to body size, mostly affect TPL and, as such, can be determinants of temporal variability of fish populations. We also found that statistical effects and phylogenetic resemblances are embedded in mean-variance relationships for fish, and that environmental drivers can interact with ecological characteristics of species in determining temporal fluctuations in abundance.

Behavioral biomarkers in fishes: A non-lethal approach to assess the effects of chemical pollution on freshwater ecosystems.

The expansion of the human population and the escalating use of chemical products pose a considerable threat to aquatic biodiversity. Consequently, there is an imperative need for the implementation of non-lethal, cost-effective, and easily deployable biomonitoring tools. In this context, fish and their behavior as biomarkers have gained prominence in monitoring of freshwater ecosystems. The aim of this study was to assess the state of art in the use of behavioral biomarkers in ecotoxicology, emphasizing their role as informative tools for global environmental monitoring. Through a systematic literature search, ninety-two articles focusing on the evaluation of behavioral changes in freshwater fish in response to pollution were identified. The most prevalent keywords were "behavior" (7%) and "zebrafish" (6%). Experiments were conducted in countries with expansive territories, such as the United States (18%) and Brazil (17%). Exotic species were primarily employed (58%), with Danio rerio (26%) being the most frequently studied species. Among pollutants, pesticides (32%) and medicines (25%) were the most frequently studied, while locomotion (38%) and social behaviors (18%) were the most frequently evaluated behaviors. Across these studies, authors consistently reported significant changes in the behavior of fish exposed to contaminants, including decreased swimming speed and compromised feeding efficiency. The review findings affirm that evaluating behavioral biomarkers in freshwater fish offers an informative, non-lethal, cost-effective, and easily implementable approach to understanding pollution impacts on freshwater ecosystems. Although few studies on behavioral biomarkers were available to date, the number has rapidly increased in recent years. Furthermore, a variety of novel approaches and study models are being included. Research into behavioral biomarkers is crucial for understanding and managing environmental risks in freshwater ecosystems. Nevertheless, further studies are needed to enhance our understanding of behavioral toxicity indicators, considering factors such as life stage, sex, and breeding season in the tested species.

Negative effects of cigarette butt leachate on freshwater phytoplankton communities.

We experimentally tested the effects of different concentrations of cigarette butt leachate on freshwater phytoplankton chlorophyll-a, species richness, cell density, and community composition. For this, we sampled the phytoplankton from a eutrophic lake and acclimated it for 24 h in microcosms. We then conducted the experiment in microcosms maintained for 96 h. The experiment consisted of four treatments: control and leachate from 1 butt L-1 (T1), 5 butts L-1 (T5), and 10 butts L-1 (T10), which were prepared by diluting a stock solution of leachate from 50 butts L-1. We found that algal chlorophyll-a content was not affected by different leachate concentrations. In contrast, phytoplankton cell density decreased in a dose-dependent manner as concentrations of the leachate increased. Similarly, the number of species was highest in the control group relative to all other treatments, with T1 and T5 showing higher species richness than T10. Additionally, the exposition to different concentrations of the leachate impacted community composition across all treatments in comparison to the control group. Our results suggest that cigarette butt leachate alters the number of cells and species, as well as the distribution of abundance, without necessarily reducing chlorophyll-a concentrations. Our findings indicate that to gain a comprehensive understanding of the effects of cigarette butt leachate on freshwater ecosystems, it is essential to evaluate more realistic scenarios that incorporate aquatic communities, rather than isolated species.

Helminths collected from some freshwater fishes and amphibians in Ecuador and Venezuela.

The present paper comprises a systematic survey of helminths (trematodes, an acanthocephalan and nematodes) found in nine species of freshwater fishes in Ecuador collected in March 1999 and those (a trematode and acanthocephalans) collected from an amphibian and two species of freshwater fishes in Venezuela in 1992, 1996 and 2001. The following 17 helminth species were recorded: Trematoda: Prosthenhystera ornamentosa sp. n., P. obesa (Diesing, 1850), Crassicutis intermedius (Szidat, 1954), C. cichlasomae Manter, 1936 and Glypthelmins eleutherodactyli sp. n. Acanthocephala: Quadrigyrus torquatus Van Cleave, 1920, Gracilisentis variabilis (Diesing, 1851) and Neoechinorhynchus (Neoechinorhynchus) ecuadoris sp. n. Nematoda: Cosmoxynema vianai Travassos, 1949, Travnema travnema Pereira, 1938, Touzeta ecuadoris Petter, 1987, Sprentascaris hypostomi Petter et Cassone, 1984, Sprentascaris sp., Contracaecum sp. Type 1 larvae, Contracaecum sp. Type 2 larvae, Procamallanus (Procamallanus) peraccuratus Pinto, Noronha et Rolas, 1976 and Procamallanus (Spirocamallanus) sp. juv. Nearly all of these parasites are reported from Ecuador or Venezuela for the first time and many of these findings represent new host records. The new species P. ornamentosa sp. n. was collected from the gall-bladder of an unidentified anostomid (Anostomidae, Characiformes) in Ecuador, G. eleutherodactyli sp. n. from the digestive tract of the frog Eleutherodactylus sp. (Eleutherodactylidae, Anura) in Venezuela and N. (N.) ecuadoris sp. n. from the intestine of Lebiasina sp. (Lebiasinidae, Characiformes) in Ecuador. Most parasites are briefly described and illustrated and problems concerning their morphology, taxonomy, hosts and geographical distribution are discussed.

Effect of emerging pollutants on the gut microbiota of freshwater animals: Focusing on microplastics and pesticides.

In recent years, emerging environmental pollutants have increasingly endangered the health of freshwater organisms. The gut microbiota exhibits sensitivity to medications, dietary factors and environmental pollutants, rendering it a novel target for toxicological studies. The gut microbiota can be a potential exposure route affecting the host's health. Herein, we review the current knowledge on two different but concurrent pollutants, microplastics and pesticides, regarding their impact on the gut microbiota, which includes alterations in microbial composition, gene expression, function, and health effects in the hosts. Moreover, synergetic interactions between microplastics and pesticides can exacerbate dysbiosis and health risks. We discuss health-related implications of gut microbial changes based on the consequences in metabolism, immunity, and physiology function. Further research is needed to discover the mechanisms underlying these effects and develop strategies for mitigating their harmful impacts on freshwater animals.

Climate change can disproportionately reduce habitats of stream fishes with restricted ranges in southern South America.

Freshwater fishes are among the most threatened taxa worldwide owing to changes in land use, species introductions, and climate change. Although more than half of the freshwater fishes in the Chilean Mediterranean ecoregion are considered vulnerable or endangered, still little is known about their biogeography. Fishes of the family Perciliidae are endemic of this region and ideal cases to study potential implications of global warming given their endangered conservation status, small size, restricted range, and limited dispersal capacity in fragmented habitats. Here, we model the spatial distribution of habitats for Percilia irwini and P. gillissi under current (1970-2000) and future (2050-2080) climatic scenarios (SSP245, SSP585). We implement maximum entropy (MaxEnt) models adapted for stream networks using high-resolution datasets of selected geophysical and climatic variables. At present, both species inhabit relatively low-quality habitats. In the future (SSP585), suitable habitats for P. irwini are predicted to be reduced drastically (99%) with potential local extirpations in its northern range. Similarly, up to 62% of suitable habitats for P. gillissi would also be reduced in the future. Our study provides insights about assessing future threats and vulnerability of endemic, endangered, range-restricted, and small-bodied freshwater species in this region and elsewhere.

Unoculubranchiobdella sp. (Hirudinea: Ozobranchidae) as a vector for Haemogregarina spp. in freshwater turtles from Brazil.

Species of Haemogregarina are blood parasites known to parasitise vertebrate hosts, including fishes (Haemogregarina sensu lato) and freshwater turtles (Haemogregarina sensu stricto). Their vectors, include gnathiid isopods and leeches, respectively. In turtles, Haemogregarina balli has the best-characterized life cycle in the genus. However, no studies in Brazil have suggested a possible vector for any species of Haemogregarina from freshwater turtles. Therefore, in the present study, we provide insights into a leech vector based on specimens found feeding on two species of freshwater turtles, Podocnemis unifilis and Podocnemis expansa, using morphological and molecular data. In 2017 and 2019, freshwater turtles were collected in Goiás State, Brazil. Hosts were inspected for ectoparasites and leeches were collected from two specimens of P. expansa and nine specimens of P. unifilis. Leeches were subsequently identified as members of the genus Unoculubranchiobdella. Leech histological slides revealed haemogregarine-like structures, similar to post-sporogonic merogony, found near the gills and within the posterior sucker. Molecular analysis of the haemeogregarines resulted in the identification of three species of Haemogregarina: Haemogregarina embaubali, Haemogregarina goianensis, and Haemogregarina brasiliana. Therefore, our findings, based on morphology and DNA data suggest leeches of the genus Unoculubranchiondella as vectors for at least three species of Haemogregarina from Brazilian turtles.

Lethal and behavioral effects of semi-purified microcystins, Micropeptin and apolar compounds from cyanobacteria on freshwater microcrustacean Thamnocephalus platyurus.

The mass proliferation of cyanobacteria, episodes known as blooms, is a concern worldwide. One of the most critical aspects during these blooms is the production of toxic secondary metabolites that are not limited to the four cyanotoxins recognized by the World Health Organization. These metabolites comprise a wide range of structurally diverse compounds that possess bioactive functions. Potential human and ecosystem health risks posed by these metabolites and co-produced mixtures remain largely unknown. We studied acute lethal and sublethal effects measured as impaired mobility on the freshwater microcrustaceans Thamnocephalus platyurus for metabolite mixtures from two cyanobacterial strains, a microcystin (MC) producer and a non-MC producer. Both cyanobacterial extracts, from the MC-producer and non-MC-producer, caused acute toxicity with LC50 (24 h) values of 0.50 and 2.55 mgdw_biomass/mL, respectively, and decreased locomotor activity. Evaluating the contribution of different cyanopeptides revealed that the Micropeptin-K139-dominated fraction from the MC-producer extract contributed significantly to mortality and locomotor impairment of the microcrustaceans, with potential mixture effect with other cyanopeptolins present in this fraction. In the non-MC-producer extract, compounds present in the apolar fraction contributed mainly to mortality, locomotor impairment, and morphological changes in the antennae of the microcrustacean. No lethal or sublethal effects were observed in the fractions dominated by other cyanopetides (Cyanopeptolin 959, Nostoginin BN741). Our findings contribute to the growing body of research indicating that cyanobacterial metabolites beyond traditional cyanotoxins cause detrimental effects. This underscores the importance of toxicological assessments of such compounds, also at sublethal levels.

Investigating the etiology of Haff disease: Optimization and validation of a sensitive LC-MS/MS method for palytoxins analysis in directly associated freshwater and marine food samples from Brazil.

Haff disease typically develops after eating contaminated marine or freshwater species, especially fish. Despite still having an unknown etiology, recent reports have suggested its possible correlation with palytoxins. Therefore, the present work aimed to optimize and perform a validation of a sensitive method using liquid chromatography coupled with tandem mass spectrometry (LC-MS/MS) for the analysis of palytoxin and some of its analogs, with the main purpose of investigating their presence in marine and freshwater food samples associated with Haff disease in Brazil. The method optimization was performed using a central composite rotatable design and fish samples fortified with the palytoxin standard. Then, the optimized method was validated for different food matrices, including freshwater and marine fish, mollusks, and crustaceans. The sample preparation involved a solid-liquid extraction using methanol and water, solid-phase extraction using Strata-X cartridges, and on-column palytoxin oxidation. The detection of the main oxidized fragments (amino and amide aldehydes) was achieved by LC-MS/MS with electrospray ionization in positive mode, using a C18 column, as well as acetonitrile and water as mobile phases, both acidified with 0.1 % of formic acid. After optimization and validation, the etiological investigation involved the analysis of 16 Brazilian Haff disease-related food samples (in natura and leftover meals) from 2022. The method was demonstrated to be appropriate for quantitative analysis of freshwater and marine species. So far, it has proven to be one of the most sensitive methods related to palytoxin detection (LOD 10 µg/kg), being able to work in a range that includes the provisional ingestion limit (30 µg/kg). Regarding the Haff disease-related samples analysis, there is a strong indication of palytoxin contamination since the amino aldehyde (common fragment for all palytoxins) was detected in 15 of the 16 samples. Selected results were confirmed using liquid chromatography coupled with high-resolution mass spectrometry (LC-HRMS).

Hematological and genotoxic biomarkers in a natural population of freshwater turtles Trachemys dorbigni (Duméril & Bibron, 1835) (Testudines: Emydidae) living in an urban area in Southern Brazil.

The present study aimed to report the morphometric and hematological indices and genotoxicity of a free-life population of D'Orbigny's slider turtles (Trachemys dorbigni) living in an urban area in Southern Brazil. For that, 16 specimens were randomly captured in an urban canal that receives irregular releases of wastewater. Biometrics and external visual changes were analyzed, such as turtle shell deformities, and the presence of parasites. Blood samples were collected to evaluate the hematological profile and the presence of micronuclei and other erythrocyte nuclear abnormalities as potential mutagenic and genotoxic effects. Water physicochemical parameters were also measured. Organisms with ectoparasites (31.25%) and small carapace deformations (56.25%) were observed, but maximum carapace length and weight were considered normal for the species according to the literature. The blood profile indicated low hemoglobin and hematocrit and a high number of total leukocytes, particularly eosinophils which characterize parasitic infections. A frequency of 0.12% for the micronucleus was considered basal, but the frequency of other erythrocyte abnormalities was evident, mainly of blebbed nuclei (63.79%), indicating chromosomal damage in the early stage. The results of this study suggest that natural populations of chelonian inhabiting urbanized areas are impacted by anthropogenic activities in the surrounding environment. Furthermore, it provides comprehensive data which can serve as a comparative model for environmental monitoring studies involving turtles.