Bioaccumulation of ß-methylamino-L-alanine (BMAA) by mussels exposed to the cyanobacteria Microcystis aeruginosa.

Cyanobacterial blooms are increasingly common in aquatic environments, raising concerns about the health impacts associated with the toxins they produce. One of these toxins is ß-methylamino-L-alanine (BMAA), a neurotoxin linked to neurodegenerative diseases. Monitoring BMAA levels in the environment is challenging due to trace concentrations and complex matrices, and new approaches are needed for assessing exposure risk. In this laboratory study, Australian freshwater mussels, Velesunio ambiguus, were exposed to a BMAA-producing cyanobacterium, Microcystis aeruginosa, to assess its accumulation of the toxin over time. A sample preparation and analysis method was developed to allow accurate quantification of BMAA in the mussels at concentrations as low as 0.4 ng/g. Mussels exposed to M. aeruginosa accumulated BMAA, with concentrations increasing over the exposure period. Rapid depuration occurred after exposure to the cyanobacterium ended, with concentrations of BMAA quickly returning to pre-exposure levels. These results demonstrate the potential for mussels to be used as bioindicators in the field for monitoring BMAA levels over time, where rapid depuration is unlikely.

Microeukaryotes Associated with Freshwater Mussels in Rivers of the Southeastern United States.

Microeukaryotes are a diverse and often overlooked group of microbes that are important in food webs and other ecological linkages. Little is known about microeukaryotes associated with aquatic invertebrates, although filter feeders such as mussels are likely to take in and potentially retain microeukaryotes in their gut while feeding. Microeukaryotes such as apicomplexans have been reported in marine mussel species, but no studies have examined the presence of these microorganisms in freshwater mussels or how they relate to mussel host species or environmental conditions. In this study, microbial community DNA was extracted from the gut tissue of over 300 freshwater mussels, representing 22 species collected from rivers in the southeastern USA. Microeukaryote DNA was detected using PCR amplification, followed by the sequencing of positive amplicons. Microeukaryotes were found in 167 individual mussels (53%) of those tested. Amplicons included dinoflagellates/algae that differed between mussel species and are likely food sources that were distinct from those found in water and sediment samples analyzed concurrently. A total of 5% of the positive amplicons were non-photosynthetic alveolates that could represent parasitic microeukaryotes. Understanding the distribution of microeukaryotes in the freshwater mussel gut microbiome could further our understanding of the ongoing decline of mussel populations.

The complete female mitogenome of Potomida semirugata (Lamarck, 1819).

Freshwater mussels perform important ecological functions in ecosystems, such as water filtration and energy cycling. Unlike marine bivalves, freshwater mussels have unique characteristics including internal fertilization and parental care. Some freshwater mussels are facing a high risk of extinction due to several factors such as climate change and habitat loss. Potomida semirugata (Lamarck, 1819) is one of the freshwater mussel species with a high risk of extinction and listed as Endangered in the Red List of the International Union for Conservation of Nature. Here, we present the first F-type mitogenome sequence of P. semirugata. The genome was sequenced on an Illumina high-throughput platform from a P. semirugata specimen collected from the Tersakan River (Turkey). The 16,093 bp mitochondrial genome sequence contains 13 protein-coding genes, 22 transfer RNAs, and two ribosomal RNAs. Phylogenetic analysis placed P. semirugata in the Lamprotulini clade with Potomida littoralis, as expected. Potomida semirugata is a poorly studied species and the genomic resource provided here will contribute to a better understanding of its biological characterization.

Differential effects of antidepressant sertraline in glochidia-fish interactions involving drug transfer from parasite to host.

This study examined the impact of sertraline, an antidepressant common in treated wastewater, on the host-parasite dynamics between parasitic freshwater mussel (Unio tumidus, Unionidae) larvae (glochidia) and their host fish (Squalius cephalus, Cyprinidae). Employing a full-factorial design, both fish and glochidia were subjected to sertraline at the combinations of 0 µg L-1 (control), 0.2 µg L-1 (environmentally relevant concentration), and 4 µg L-1 (elevated concentration, short-term exposure of the parasite). The results showed that long-term host exposure (involving intensive sertraline accumulation in the fish brain) marginally increased subsequent glochidia attachment success by 2 %, while parasite exposure at the same environmentally relevant concentrations had no detectable effect. There was also no effect of exposure of glochidia to 0.2 µg L-1 of sertraline on their viability and encapsulation success during the initial parasitic stage. However, a significant alteration in attachment behavior, marked by a 3.3 % increase in attachment success and changes in the glochidia spatial distribution on the host body, was noted after 24 h of glochidia exposure to 4 µg L-1 of sertraline. Importantly, this study provides the first evidence of sertraline transfer from exposed glochidia to nonexposed host fish, as indicated by elevated levels of sertraline (12.8 ng g-1) in the brain tissue of nonexposed hosts. These findings highlight the subtle yet significant effects of pharmaceutical pollutants on freshwater ecosystems but also underscore the importance of understanding the unexpected dynamics of such contamination to predict and address future ecological changes.

Optimizing nutrient utilization, hydraulic loading rate, and feed conversion ratios through freshwater IMTA-aquaponic and hydroponic systems as an environmentally sustainable aquaculture concept.

Water quality in land-based fish production can be controlled through either instantaneous water exchange or costly wastewater treatment followed by recirculation. Agricultural-aquaculture integration is an excellent alternative technique for reducing nutrient discharge levels, boosting profitability, and converting fish culture wastewater into valuable products. The current study employed a solar energy system to power two separate IMTA-aquaponics systems (Nutrient Film Technique, NFT, and Floating Raft Systems, FRS) for the cultivation of Nile tilapia, African catfish, thin-lipped grey mullet, freshwater crayfish, freshwater mussels, and a variety of vegetables. Tilapia and catfish were fed exclusively on diets under the IMTA system. All wastewater from tilapia and catfish ponds, both dissolved and solid, flows sequentially to ponds containing other cultivated species. The water then flows through the IMTA system's terminal point to the NFT and FRS systems before returning to the tilapia and catfish ponds, allowing complete control of the nutrient flow throughout this entire circular system. Two 147-day production cycles were concluded. The results from the second production cycle are reported. Total biomass gain for aquatic species in the IMTA system was 736.46 kg, compared to 145.49 kg in the tilapia and 271.01 kg in the catfish monoculture systems. The current IMTA system had a cumulative feed conversion ratio (FCR) of 0.90, while the FCRs for tilapia and catfish were 1.28 and 1.42, respectively. Nile tilapia and catfish consumed 571.90 kg of feed containing 25.70 kg of nitrogen (N) and 9.70 kg of phosphorus (P), reflecting, and gaining 11.41 and 3.93 kg of dietary N and P, representing 44.40 and 40.46% dietary N and P retention, respectively. In the IMTA system, the addition of mullet and prawn as detrivores aquatic animals improves dietary N and P utilization efficiency to 59.06 and 51.19%, respectively, while the addition of mussels as herbivore animals improves dietary N and P utilization efficiency to 65.61 and 54.67%, respectively. Finally, using FRS and NFT as hydroponic systems increased dietary N and P efficiency to 83.51% N and 96.82% P, respectively. This study shows that the IMTA-Aquaponic system, as a bio-integrated food production system, can convert the majority of fish-fed residues into valuable products suitable for desert, rural, and urban areas in impoverished and developing countries.

The complete mitochondrial genome of Potomida acarnanica (Kobelt, 1879).

Freshwater mussels (Bivalvia, Unionida) play essential roles in the well-functioning of ecosystems, even providing essential services to humans. However, these bivalves face numerous threats (e.g. habitat loss and fragmentation, pollution, introduction of invasive species, and climate change) which have already led to the extinction of many populations. This underscores the need to fully characterize the biology of these species, particularly those, such as Potomida acarnanica, that are still poorly studied. This study presents the first mitogenome of P. acarnanica (Kobelt, 1879), an endemic species of Greece with a distribution limited to only two river basins. The mitochondrial genome of a P. acarnanica specimen, collected at Pamisos River (Peloponnese, Greece), was sequenced by Illumina high-throughput sequencing. This mitogenome (16,101 bp) is characterized by 13 protein-coding genes, 22 transfer RNA and 2 ribosomal RNA genes. The size of this mitogenome is within the range of another Potomida mitogenome already published for the species Potomida littoralis. In the phylogenetic inference, P. acarnanica was recovered as monophyletic with P. littoralis mitogenome in the Lamprotulini tribe, as expected. This genomic resource will assist in genetically characterizing the species, potentially benefiting future evolutionary studies and conservation efforts.

Plastic Analysis with a Plasmonic Nano-Gold Sensor Coated with Plastic-Binding Peptides.

Contamination with plastics of small dimensions (<1 µm) represents a health concern for many terrestrial and aquatic organisms. This study examined the use of plastic-binding peptides as a coating probe to detect various types of plastic using a plasmon nano-gold sensor. Plastic-binding peptides were selected for polyethylene (PE), polyethylene terephthalate (PET), polypropylene (PP), and polystyrene (PS) based on the reported literature. Using nAu with each of these peptides to test the target plastics revealed high signal, at 525/630 nm, suggesting that the target plastic limited HCl-induced nAu aggregation. Testing with other plastics revealed some lack of specificity but the signal was always lower than that of the target plastic. This suggests that these peptides, although reacting mainly with their target plastic, show partial reactivity with the other target plastics. By using a multiple regression model, the relative levels of a given plastic could be corrected by the presence of other plastics. This approach was tested in freshwater mussels caged for 3 months at sites suspected to release plastic materials: in rainfall overflow discharges, downstream a largely populated city, and in a municipal effluent dispersion plume. The data revealed that the digestive glands of the mussels contained higher levels of PP, PE, and PET plastic particles at the rainfall overflow and downstream city sites compared to the treated municipal effluent site. This corroborated earlier findings that wastewater treatment could remove nanoparticles, at least in part. A quick and inexpensive screening test for plastic nanoparticles in biological samples with plasmonic nAu-peptides is proposed.

Polymorphism in the aggressive mimicry lure of the parasitic freshwater mussel Lampsilis fasciola.

Unionoid freshwater mussels (Bivalvia: Unionidae) are free-living apart from a brief, obligately parasitic, larval stage that infects fish hosts, and gravid female mussels have evolved a spectrum of strategies to infect fish hosts with their larvae. In many North American species, this involves displaying a mantle lure: a pigmented fleshy extension that acts as an aggressive mimic of a host fish prey, thereby eliciting a feeding response that results in host infection. The mantle lure of Lampsilis fasciola is of particular interest because it is apparently polymorphic, with two distinct primary lure phenotypes. One, described as "darter-like", has "eyespots", a mottled body coloration, prominent marginal extensions, and a distinct "tail". The other, described as "worm-like", lacks those features and has an orange and black coloration. We investigated this phenomenon using genomics, captive rearing, biogeographic, and behavioral analyses. Within-brood lure variation and within-population phylogenomic (ddRAD-seq) analyses of individuals bearing different lures confirmed that this phenomenon is a true polymorphism. The relative abundance of the two morphs appears stable over ecological timeframes: the ratio of the two lure phenotypes in a River Raisin (MI) population in 2017 was consistent with that of museum samples collected at the same site six decades earlier. Within the River Raisin, four main "darter-like" lure motifs visually approximated four co-occurring darter species (Etheostoma blennioides, E. exile, E. microperca, and Percina maculata), and the "worm-like" lure resembled a widespread common leech, Macrobdella decora. Darters and leeches are typical prey of Micropterus dolomieui (smallmouth bass), the primary fish host of L. fasciola. In situ field recordings of the L. fasciola "darter" and "leech" lure display behaviors, and the lure display of co-occurring congener L. cardium, were captured. Despite having putative models in distinct phyla, both L. fasciola lure morphs have largely similar display behaviors that differ significantly from that of sympatric L. cardium individuals. Some minor differences in the behavior between the two L. fasciola morphs were observed, but we found no clear evidence for a behavioral component of the polymorphism given the criteria measured. Discovery of discrete within-brood inheritance of the lure polymorphism implies potential control by a single genetic locus and identifies L. fasciola as a promising study system to identify regulatory genes controlling a key adaptive trait of freshwater mussels.

Bisphenol a (BPA) aggravate the adverse effect on physiological and biochemical response in freshwater mussel potomida littoralis.

Bisphenol A (BPA) is a chemical compound extensively employed in plastic manufacturing, and this pollutant has been detected in diverse aquatic organisms, notably bivalves. In order to comprehend the ecological and toxicological consequences of BPA Bisphenol A in these organisms, it is essential to examine the physiological and biochemical effects and identify areas where our understanding is lacking. This knowledge is crucial for determining the environn ental threat posed by bisphenol A and assisting decision-makers in establishing the appropriate priorities. This investigation aimed to assess the impact of BPA on the biochemical and physiological parameters of the freshwater mussel Potomida littoralis. In a laboratory setting, mussels were subjected to two different levels of BPA (20 and 100 µg/L) for a duration of 21 days. Filtration rate was calculated from the clearance of neutral red, fed to mussels at different BPA concentrations. The mussel's filtration rate capacity declined as BPA exposure intensified, potentially due to the mussel's attempt to close its valves and minimize BPA absorption, thus preventing cellular damage. In the digestive gland tissue, key antioxidant and detoxification defenses, including catalase (CAT) activity, glutathione-S-transferase (GST) activity, and levels of H2O2 and glutathione (GSH), were activated, particularly at the 100 µg/L BPA concentration. This activation helped protect against lipid damage at higher BPA concentrations. This study underscores the significance of preventing and regulating BPA release into the environment to avert detrimental consequences for aquatic ecosystems.

Integrative phylogenetic, phylogeographic and morphological characterisation of the Unio crassus species complex reveals cryptic diversity with important conservation implications.

The global decline of freshwater mussels and their crucial ecological services highlight the need to understand their phylogeny, phylogeography and patterns of genetic diversity to guide conservation efforts. Such knowledge is urgently needed for Unio crassus, a highly imperilled species originally widespread throughout Europe and southwest Asia. Recent studies have resurrected several species from synonymy based on mitochondrial data, revealing U. crassus to be a complex of cryptic species. To address long-standing taxonomic uncertainties hindering effective conservation, we integrate morphometric, phylogenetic, and phylogeographic analyses to examine species diversity within the U. crassus complex across its entire range. Phylogenetic analyses were performed using cytochrome c oxidase subunit I (815 specimens from 182 populations) and, for selected specimens, whole mitogenome sequences and Anchored Hybrid Enrichment (AHE) data on âˆ¼ 600 nuclear loci. Mito-nuclear discordance was detected, consistent with mitochondrial DNA gene flow between some species during the Pliocene and Pleistocene. Fossil-calibrated phylogenies based on AHE data support a Mediterranean origin for the U. crassus complex in the Early Miocene. The results of our integrative approach support 12 species in the group: the previously recognised Unio bruguierianus, Unio carneus, Unio crassus, Unio damascensis, Unio ionicus, Unio sesirmensis, and Unio tumidiformis, and the reinstatement of five nominal taxa: Unio desectusstat. rev., Unio gontieriistat. rev., Unio mardinensisstat. rev., Unio nanusstat. rev., and Unio vicariusstat. rev. Morphometric analyses of shell contours reveal important morphospace overlaps among these species, highlighting cryptic, but geographically structured, diversity. The distribution, taxonomy, phylogeography, and conservation of each species are succinctly described.

The influence of rainfall events on the toxicity of urban wastewaters to freshwater mussels Elliptio complanata.

The cumulative impacts of rainfall frequency and intensity towards the ecotoxicity of urban pollution is gaining more and more attention in these times of climate change. The purpose of this study was to examine the ecotoxicological impacts of combined sewers overflows and municipal effluent discharge sites during 3 periods (years) of varying intensity precipitations to freshwater mussels Elliptio complanata. Mussels were placed in benthic cages for 3 months during the summer at 2 overflow discharge and 8 km downstream sites including an upstream site for three consecutive years with low (164 mm), medium (182 mm) and high (248 mm) amounts of rain. The results revealed that the effects were mainly influenced by suspended matter loadings and to the dissolved components to a lesser extent. Impacts at the downstream and overflow sites were noticeable at the reproduction (vitellogenin), genotoxicity, neurotoxicity (dopamine and serotonin changes) levels in addition to xenobiotic biotransformation revealed by glutathione S-transferase activity and metallothioneins for organic and heavy metals respectively. The site downstream the effluent produced most of the effects compared to the overflow sites in the Saint-Lawrence River. However, the impacts of combined sewers overflows could become problematic in low dilution systems such as small river and lakes.

The cerium oxide nanoparticles toxicity induced physiological, histological and biochemical alterations in freshwater mussels, Unio crassus.

INTRODUCTION: Releasing of cerium oxide nanoparticles (nano-CeO2) to the nature has increased due to the widespread use in many fields ranging from cosmetics to the food industry. Therefore, nano-CeO2 has been included in the Organization for Economic Co-operation and Development's (OECD) priority list for engineering nanomaterials. In this study, the effects of nano-CeO2 on the freshwater mussels were investigated to reveal the impact on the freshwater systems on model organism. METHODS: First, the chemical and structural properties of nano-CeO2 were characterized in details. Second, the freshwater mussels were exposed to environmentally relevant concentrations of nano-CeO2 as 10 mg, 25 mg and 50 mg/L during 48-h and 7-d. Third, after the exposure periods, hemolymph and tissue samples were taken to analyse the Total Hemocyte Counts (THCs) histology and oxidative stress parameters (total antioxidant status, glutathione, glutathione-S-transferase, and advanced oxidative protein products). RESULTS: Significant decrease of the THCs was observed in the nano-CeO2 exposed mussels compared to the control group (P < 0.05). The histological results showed a positive association between nano-CeO2 exposure concentration in the water and level of tissue damage and histopathological alterations were detected in the gill and the digestive gland tissues. Oxidative stress parameters were slightly affected after exposure to nano-CeO2 (P > 0.05). In conclusion, this study showed that acute exposure of freshwater mussels to nano-CeO2 did not pose significant biological risk. However, it has been proven that mussels are able to accumulate nano-CeO2 significantly in their bodies. CONCLUSION: This suggests that nano-CeO2 may be a potential risk to other organisms in the ecosystem through trophic transfer in the food-web based on their habitat and niche in the ecosystem.

Riverine flow rate drives widespread convergence in the shell morphology of imperiled freshwater mussels.

Frequent and strong morphological convergence suggests that determinism tends to supersede historical contingencies in evolutionary radiations. For many lineages living within the water column of rivers and streams, hydrodynamic forces drive widespread morphological convergence. Living below the sediment-water interface may release organisms from these hydrodynamic pressures, permitting a broad array of morphologies, and thus less convergence. However, we show here that the semi-infaunal freshwater mussels have environmentally determined convergence in shell morphology. Using 3D morphometric data from 715 individuals among 164 Nearctic species, we find that species occurring in rivers with high flow rates have evolved traits that resist dislodgement from their burrowed position in the streambed: thicker shells for their body size, with the thickest sector of the shell being the most deeply buried. Species occurring in low flow environments have evolved thinner and more uniformly thickened shells, corresponding to an alternative adaptation to dislodgement: increased burrowing efficiency. Within species, individuals also show increased shell thickness for their body size at higher flow rates, suggesting that ecophenotypy may, in part, be an important mechanism for establishing populations in new environments and thus evolutionary divergence in this highly imperiledinvertebrate group.

Characterizing the Gut Microbial Communities of Native and Invasive Freshwater Bivalves after Long-Term Sample Preservation.

Freshwater mussels are important indicators of the overall health of their environment but have suffered declines that have been attributed to factors such as habitat degradation, a loss of fish hosts, climate change, and excessive nutrient inputs. The loss of mussel biodiversity can negatively impact freshwater ecosystems such that understanding the mussel's gut microbiome has been identified as a priority topic for developing conservation strategies. In this study, we determine whether ethanol-stored specimens of freshwater mussels can yield representative information about their gut microbiomes such that changes in the microbiome through time could potentially be determined from museum mussel collections. A short-term preservation experiment using the invasive clam Corbicula fluminea was used to validate the use of ethanol as a method for storing the bivalve microbiome, and the gut microbiomes of nine native mussel species that had been preserved in ethanol for between 2 and 9 years were assessed. We show that ethanol preservation is a valid storage method for bivalve specimens in terms of maintaining an effective sequencing depth and the richness of their gut bacterial assemblages and provide further insight into the gut microbiomes of the invasive clam C. fluminea and nine species of native mussels. From this, we identify a "core" genus of bacteria (Romboutsia) that is potentially common to all freshwater bivalve species studied. These findings support the potential use of ethanol-preserved museum specimens to examine patterns in the gut microbiomes of freshwater mussels over long periods.

Mitochondrial sequence data reveal population structure within Pustulosa pustulosa.

Unionid mussels are among the most imperiled group of organisms in North America, and Pustulosa pustulosa is a freshwater species with a relatively wide latitudinal distribution that extends from southern Ontario, Canada, to Texas, USA. Considerable morphological and geographic variation in the genus Pustulosa (formerly Cyclonaias) has led to uncertainty over species boundaries, and recent studies have suggested revisions to species-level classifications by synonymizing C. aurea, C. houstonensis, C. mortoni, and C. refulgens with C. pustulosa (currently P. pustulosa). Owing to its wide range and shallow phylogenetic differentiation, we analyzed individuals of P. pustulosa using mitochondrial DNA sequence data under a population genetics framework. We included 496 individuals, which were comprised of 166 samples collected during this study and 330 additional sequences retrieved from GenBank. Pairwise ΦST measures based on ND1 data suggested there may be up to five major geographic groups present within P. pustulosa. Genetic differentiation between regions within Texas was higher compared to populations from the Mississippi and Great Lakes populations, which may reflect differences in historical connectivity. Mitochondrial sequence data also revealed varying demographic histories for each major group suggesting each geographic region has also experienced differential population dynamics in the past. Future surveys should consider exploring variation within species after phylogeographic delimitation has been performed. In this study, we begin to address this need for freshwater mussels via the P. pustulosa system.

A PacBio Hi-Fi Genome Assembly of the Painter's Mussel Unio pictorum (Linnaeus, 1758).

The highly diverse group of freshwater mussels from order Unionida is found in the world's freshwater systems due to several fascinating evolutionary adaptations, including "parental care," and most notably, an obligatory parasitic phase in their early life cycle, called glochidia, which infests and uses fish for nutrition and dispersal. Freshwater mussels play essential ecological roles in freshwater habitats, including water filtration, sediment bioturbation, and nutrient cycling. However, these species are also highly threatened, being one of the faunal groups with the highest recorded extinction rate in the wild. Genomics methods have an incredible potential to promote biodiversity conservation, allowing the characterization of population health, identification of adaptive genetic elements, delineation of conservation units, and providing a framework for predictive assessments of the impact of anthropogenic threats and climate change. Unfortunately, only six freshwater mussel species have had their whole genomes sequenced to date, and only two of these are European species. Here, we present the first genome assembly of the Painter's Mussel, Unio pictorum (Linnaeus, 1758), the type species representative of the order and the most widespread species of the genus in Europe. We used long-read PacBio Hi-Fi sequencing reads to produce a highly contiguous assembly that will pave the way for the study of European freshwater mussels in the Genome Era.

A novel assembly pipeline and functional annotations for targeted sequencing: A case study on the globally threatened Margaritiferidae (Bivalvia: Unionida).

The proliferation of genomic sequencing approaches has significantly impacted the field of phylogenetics. Target capture approaches provide a cost-effective, fast and easily applied strategy for phylogenetic inference of non-model organisms. However, several existing target capture processing pipelines are incapable of incorporating whole genome sequencing (WGS). Here, we develop a new pipeline for capture and de novo assembly of the targeted regions using whole genome re-sequencing reads. This new pipeline captured targeted loci accurately, and given its unbiased nature, can be used with any target capture probe set. Moreover, due to its low computational demand, this new pipeline may be ideal for users with limited resources and when high-coverage sequencing outputs are required. We demonstrate the utility of our approach by incorporating WGS data into the first comprehensive phylogenomic reconstruction of the freshwater mussel family Margaritiferidae. We also provide a catalogue of well-curated functional annotations of these previously uncharacterized freshwater mussel-specific target regions, representing a complementary tool for scrutinizing phylogenetic inferences while expanding future applications of the probe set.

Antimicrobial Susceptibility of Bacteria Isolated from Freshwater Mussels in the Wildcat Creek Watershed, Indiana, United States.

Antimicrobial resistance (AMR) is a global health crisis that threatens the health of humans and animals. The spread of resistance among species may occur through our shared environment. Prevention of AMR requires integrated monitoring systems, and these systems must account for the presence of AMR in the environment in order to be effective. The purpose of this study was to establish and pilot a set of procedures for utilizing freshwater mussels as a means of surveillance for microbes with AMR in Indiana waterways. One hundred and eighty freshwater mussels were sampled from three sites along the Wildcat Creek watershed in north-central Indiana. Specimens were evaluated for the presence of ESKAPE pathogens (Enterococcus faecium, Staphylococcus aureus, Klebsiella pneumoniae, Acinetobacter baumannii, Pseudomonas aeruginosa, Enterobacter species), Escherichia coli, Campylobacter, and Salmonella species, and the isolates were tested for antimicrobial resistance. A total of 24 bacterial isolates were obtained from tissue homogenates of freshwater mussels collected at a site directly downstream from Kokomo, Indiana. Of these, 17 were Enterobacter spp., five were Escherichia coli, one was Pseudomonas aeruginosa, and one was Klebsiella pneumoniae. All isolates were resistant to three or more antimicrobial drug classes. Further work is necessary to determine the source of the bacterial species found in the mussels.

In vitro assessment of the genotoxicity and immunotoxicity of treated and untreated municipal effluents and receiving waters in freshwater organisms.

Municipal wastewater effluent is one of the largest sources of pollution entering surface waters in the Laurentian Great Lakes. Exposure to wastewater effluent has been associated with impaired immune systems and induction of genotoxicity to aquatic animals. Due to habitat degradation and environmental pollution linked to industrial development and population growth, several regions of the Great Lakes have been designated Areas of Concern (AOCs). In this study, we assessed the effect of extracts of sewage influent, (treated) effluent and receiving surface waters from the Hamilton Harbour AOC and the Toronto and Region AOC (Ontario, Canada) on the phagocytic immune response of rainbow trout (Oncorhynchus mykiss) kidney leukocytes and the genotoxicity (DNA strand breaks) of these extracts on freshwater mussel (Eurynia dilatata) hemocytes. We identified and quantified numerous chemicals present in the various samples extracted for exposure. In freshwater mussels, extracts from Hamilton Harbour AOC induced DNA damage with the most frequency (12 out of 28 samples) regardless of sample type, reflecting past and present industrial activities. In contrast, extracts from Toronto and Region AOC induced DNA damage infrequently (2 out of 32 (summer) and 5 out of 32 (fall) samples, respectively) and from different WWTPs at different times. None of the extracts induced any significant effect on phagocytosis of rainbow trout kidney leukocytes. The present study indicates that despite overall improvements to effluent quality, treatment of influent by WWTPs may not result in a corresponding improvement of the genotoxicity of effluents. In vitro bioassays are useful and cost-effective rapid-screening tools for preliminary assessments of contamination of aquatic ecosystems.

Alterations to unionid transformation during agricultural and urban contaminants of emerging concern exposures.

Highly imperiled unionids have a complex life cycle including the metamorphosis of an obligate parasite life stage, larval glochidia, to the juvenile stage. Despite the known vulnerabilities of both glochidia and juveniles to pollutants, little is known on how metamorphosis success may be affected by chemical stress. Disruption of the transformation process in which glochidia encyst on the gills of a host fish, could lead to lowered recruitment and population declines. Transformation rates of Lampsilis cardium on host fish Micropterus salmoides were empirically derived from experimental exposures to low, medium, or high concentrations of an agricultural or urban mixture of contaminants of emerging concern (CECs) over two exposure durations. Transformation was characterized by: (1) a zero-inflated Poisson general linear mixed effects model to compare difference in transformation between exposure durations and (2) time response curves to describe the transformation curve using long-term exposure data. Lampsilis cardium transformation was similar between exposure durations. When compared to controls, CEC stress significantly reduced juvenile production (p « 0.05) except for the agricultural medium treatment and tended to increased encapsulation duration which while statistically insignificant (p = 0.16) may have ecological relevancy. Combining the empirically derived reduction of transformation rates with parameters values from the literature, a Lefkovich stage-based population model predicted strong declines in population size of L. cardium for all treatments if these results hold in nature. Management focus on urban CECs may lead to best conservation efforts though agricultural CECs may also have a concentration dependent impact on transformation and therefore overall recruitment and conservation success.