Genome Analysis of a Newly Discovered Yeast Species, Hanseniaspora menglaensis.

Annual surveys of Irish soil samples identified three isolates, CBS 16921 (UCD88), CBS 18246 (UCD443), and CBS 18247 (UCD483), of an apiculate yeast species within the Hanseniaspora genus. The internal transcribed spacer (ITS) and D1/D2 region of the large subunit (LSU) rRNA sequences showed that these are isolates of the recently described species Hanseniaspora menglaensis, first isolated from Southwest China. No genome sequence for H. menglaensis is currently available. The genome sequences of the three Irish isolates were determined using short-read (Illumina) sequencing, and the sequence of one isolate (CBS 16921) was assembled to chromosome level using long-read sequencing (Oxford Nanopore Technologies). Phylogenomic analysis shows that H. menglaensis belongs to the fast-evolving lineage (FEL) of Hanseniaspora. Only one MAT idiomorph (encoding MATα1) was identified in all three sequenced H. menglaensis isolates, consistent with one mating type of a heterothallic species. Genome comparisons showed that there has been a rearrangement near MATα of FEL species compared to isolates from the slowly evolving lineage (SEL).

Implementation of the CREED approach for environmental assessments.

Environmental exposure data are a key component of chemical and ecological assessments, supporting and guiding environmental management decisions and regulations. Measures taken to protect the environment based on exposure data can have social and economic implications. Flawed information may lead to measures being taken in the wrong place or to important action not being taken. Although the advantages of harmonizing evaluation methods have been demonstrated for hazard information, no comparable approach is established for exposure data evaluation. The goal of Criteria for Reporting and Evaluating Exposure Datasets (CREED) is to improve the transparency and consistency with which exposure data are evaluated regarding usability in environmental assessments. Here, we describe the synthesis of the CREED process, and propose methods and tools to summarize and interpret the outcomes of the data usability evaluation in support of decision-making and communication. The CREED outcome includes a summary that reports any key gaps or shortcomings in the reliability (data quality) and relevance (fitness for purpose) of the data being considered. The approach has been implemented in a workbook template (provided as Supporting Information), for assessors to readily follow the workflow and create a report card for any given dataset. The report card communicates the outcome of the CREED evaluation and summarizes important dataset attributes, providing a concise reference pertaining to the dataset usability for a specified purpose and documenting data limitations that may restrict data use or increase environmental assessment uncertainty. The application of CREED is demonstrated through three case studies, which also were used during beta testing of the methodology. As experience with the CREED approach application develops, further improvements may be identified and incorporated into the framework. Such development is to be encouraged in the interest of better science and decision-making, and to make environmental monitoring and assessment more cost-effective. Integr Environ Assess Manag 2024;00:1-16. © 2024 SETAC.

Draft genome sequence of the yeast Schwanniomyces capriottii strain UCD805, isolated from soil in Ireland.

Schwanniomyces capriottii is a member of the Debaryomycetaceae family in the order Saccharomycetales. Here, we present the genome sequence of S. capriottii UCD805, which was isolated from soil in Dublin, Ireland. This genome is 12.2 Mb and was assembled into 14 scaffolds plus a mitochondrial genome scaffold.

Evaluation of outbreak persistence caused by multidrug-resistant and echinocandin-resistant Candida parapsilosis using multidimensional experimental and epidemiological approaches.

Candida parapsilosis is known to cause severe and persistent outbreaks in clinical settings. Patients infected with multidrug-resistant C. parapsilosis (MDR Cp) isolates were identified in a large Turkish hospital from 2017-2020. We subsequently identified three additional patients infected with MDR Cp isolates in 2022 from the same hospital and two echinocandin-resistant (ECR) isolates from a single patient in another hospital. The increasing number of MDR and ECR isolates contradicts the general principle that the severe fitness cost associated with these phenotypes could prevent their dominance in clinical settings. Here, we employed a multidimensional approach to systematically assess the fitness costs of MDR and ECR C. parapsilosis isolates. Whole-genome sequencing revealed a novel MDR genotype infecting two patients in 2022. Despite severe in vitro defects, the levels and tolerances of the biofilms of our ECR and MDR isolates were generally comparable to those of susceptible wild-type isolates. Surprisingly, the MDR and ECR isolates showed major alterations in their cell wall components, and some of the MDR isolates consistently displayed increased tolerance to the fungicidal activities of primary human neutrophils and were more immunoevasive during exposure to primary human macrophages. Our systemic infection mouse model showed that MDR and ECR C. parapsilosis isolates had comparable fungal burden in most organs relative to susceptible isolates. Overall, we observed a notable increase in the genotypic diversity and frequency of MDR isolates and identified MDR and ECR isolates potentially capable of causing persistent outbreaks in the future.

Development and Validation of Multiple Linear Regression Models for Predicting Chronic Zinc Toxicity to Freshwater Microalgae.

Multiple linear regression (MLR) models were developed for predicting chronic zinc toxicity to a freshwater microalga, Chlorella sp., using three toxicity-modifying factors (TMFs): pH, hardness, and dissolved organic carbon (DOC). The interactive effects between pH and hardness and between pH and DOC were also included. Models were developed at three different effect concentration (EC) levels: EC10, EC20, and EC50. Models were independently validated using six different zinc-spiked Australian natural waters with a range of water chemistries. Stepwise regression found hardness to be an influential TMF in model scenarios and was retained in all final models, while pH, DOC, and interactive terms had variable influence and were only retained in some models. Autovalidation and residual analysis of all models indicated that models generally predicted toxicity and that there was little bias based on individual TMFs. The MLR models, at all effect levels, performed poorly when predicting toxicity in the zinc-spiked natural waters during independent validation, with models consistently overpredicting toxicity. This overprediction may be from another unaccounted for TMF that may be present across all natural waters. Alternatively, this consistent overprediction questions the underlying assumption that models developed from synthetic laboratory test waters can be directly applied to natural water samples. Further research into the suitability of applying synthetic laboratory water-based models to a greater range of natural waters is needed. Environ Toxicol Chem 2023;42:2630-2641. © 2023 The Authors. Environmental Toxicology and Chemistry published by Wiley Periodicals LLC on behalf of SETAC.

Applicability of Chronic Multiple Linear Regression Models for Predicting Zinc Toxicity in Australian and New Zealand Freshwaters.

Bioavailability models, for example, multiple linear regressions (MLRs) of water quality parameters, are increasingly being used to develop bioavailability-based water quality criteria for metals. However, models developed for the Northern Hemisphere cannot be adopted for Australia and New Zealand without first validating them against local species and local water chemistry characteristics. We investigated the applicability of zinc chronic bioavailability models to predict toxicity in a range of uncontaminated natural waters in Australia and New Zealand. Water chemistry data were compiled to guide a selection of waters with different zinc toxicity-modifying factors. Predicted toxicities using several bioavailability models were compared with observed chronic toxicities for the green alga Raphidocelis subcapitata and the native cladocerans Ceriodaphnia cf. dubia and Daphnia thomsoni. The most sensitive species to zinc in five New Zealand freshwaters was R. subcapitata (72-h growth rate), with toxicity ameliorated by high dissolved organic carbon (DOC) or low pH, and hardness having a minimal influence. Zinc toxicity to D. thomsoni (reproduction) was ameliorated by both high DOC and hardness in these same waters. No single trophic level-specific effect concentration, 10% (EC10) MLR was the best predictor of chronic toxicity to the cladocerans, and MLRs based on EC10 values both over- and under-predicted zinc toxicity. The EC50 MLRs better predicted toxicities to both the Australian and New Zealand cladocerans to within a factor of 2 of the observed toxicities in most waters. These findings suggest that existing MLRs may be useful for normalizing local ecotoxicity data to derive water quality criteria for Australia and New Zealand. The final choice of models will depend on their predictive ability, level of protection, and ease of use. Environ Toxicol Chem 2023;42:2614-2629. © 2023 The Authors. Environmental Toxicology and Chemistry published by Wiley Periodicals LLC on behalf of SETAC.

Evaluating the Protectiveness of a Bioavailability-Based Environmental Quality Standard for the Protection of Aquatic Communities from Zinc Toxicity Based on Field Evidence.

Environmental quality standards (EQS) are typically derived from the results of laboratory studies on single species. There is always uncertainty surrounding the protectiveness of an EQS when applied to real ecosystems containing a multitude of chemical and physical stressors. Quantile regression was used with field biological data on invertebrates in United Kingdom waters to identify taxa that are responsive to bioavailable zinc exposures. A threshold based on the total abundance of eight responsive taxa is used as an indicator of the overall ecosystem sensitivity. The inclusion of some responsive but insensitive taxa in this ecological metric could bias the results toward a higher threshold. The least responsive species were progressively removed from the collective ecological metric, basing the analysis on a progressively smaller number of the more responsive species. Quantile regression analysis at the 95th quantile for the three most responsive taxa resulted in a 10% effect concentration of 14.8 µg L-1 bioavailable zinc, suggesting that the EQS of 10.9 µg L-1 bioavailable zinc is sufficiently protective of sensitive members of the invertebrate community. There is a compromise between the robustness of the analysis and the sensitivity of the subcommunity that it is based on. Analyses based on fewer taxa provide a more sensitive result. This approach assessed real ecosystem data and evaluated the uncertainty associated with the protectiveness of the EQS for zinc. The zinc EQS is sufficiently protective of sensitive members of benthic macroinvertebrate communities under real environmental conditions, including a mix of multiple substances. Environ Toxicol Chem 2023;42:1010-1021. © 2023 The Authors. Environmental Toxicology and Chemistry published by Wiley Periodicals LLC on behalf of SETAC.

Comparison of Multiple Linear Regression and Biotic Ligand Models for Predicting Acute and Chronic Zinc Toxicity to Freshwater Organisms.

Multiple linear regression (MLR) models for predicting zinc (Zn) toxicity to freshwater organisms were developed based on three toxicity-modifying factors: dissolved organic carbon (DOC), hardness, and pH. Species-specific, stepwise MLR models were developed to predict acute Zn toxicity to four invertebrates and two fish, and chronic toxicity to three invertebrates, a fish, and a green alga. Stepwise regression analyses found that hardness had the most consistent influence on Zn toxicity among species, whereas DOC and pH had a variable influence. Pooled acute and chronic MLR models were also developed, and a k-fold cross-validation was used to evaluate the fit and predictive ability of the pooled MLR models. The pooled MLR models and an updated Zn biotic ligand model (BLM) performed similarly based on (1) R2 , (2) the percentage of effect concentration (ECx) predictions within a factor of 2.0 of observed ECx, and (3) residuals of observed/predicted ECx versus observed ECx, DOC, hardness, and pH. Although fit of the pooled models to species-specific toxicity data differed among species, species-specific differences were consistent between the BLM and MLR models. Consistency in the performance of the two models across species indicates that additional terms, beyond DOC, hardness, and pH, included in the BLM do not help explain the differences among species. The pooled acute and chronic MLR models and BLM both performed better than the US Environmental Protection Agency's existing hardness-based model. We therefore conclude that both MLR models and the BLM provide an improvement over the existing hardness-only models and that either could be used for deriving ambient water quality criteria. Environ Toxicol Chem 2023;42:393-413. © 2022 SETAC.

Natural organic matter source, concentration, and pH influences the toxicity of zinc to a freshwater microalga.

Zinc is a contaminant of concern in aquatic environments and is a known toxicant to many aquatic organisms. Dissolved organic matter (DOM) is a toxicity modifying factor for zinc and is an important water chemistry parameter. This study investigated the influence of DOM concentration, source, and water pH on the chronic toxicity of zinc to a freshwater microalga, Chlorella sp. The influence of DOM on zinc toxicity was dependent on both concentration and source. In the absence of DOM, the 72-h EC50 was 112 µg Zn.L-1. In the presence of a DOM high in fulvic-like components, zinc toxicity was either slightly decreased (<4-fold increase in EC10s across 15 mg C.L-1 range) or unchanged (minimal difference in EC50s). In the presence of a DOM high in humic-like (aromatic and high molecular weight) components, zinc toxicity was slightly decreased at the EC10 level and strongly increased at the EC50 level. The influence of pH on zinc toxicity was dependent on the source of DOM present in the water. In the presence of DOM high in humic-like components pH did not influence toxicity. In the presence of DOM high in fulvic-like components, pH had a significant effect on EC50 values. Labile zinc (measured by diffusive gradients in thin-films) followed linear relationships with dissolved zinc but could not explain the changes in observed toxicity, with similar DGT-labile zinc relationships shown for the two DOMs despite each DOM influencing toxicity differently. This indicates changes in toxicity may be unrelated to changes in zinc lability. The results suggest that increased toxicity of zinc in the presence of DOM may be due to direct uptake of Zn-DOM complexes. This study highlights the importance of considering DOM source and characteristics when incorporating DOM into water quality guidelines through bioavailability models.

Identification of European isolates of the lager yeast parent Saccharomyces eubayanus.

Lager brewing first occurred in Bavaria in the 15th century, associated with restrictions of brewing to colder months. The lager yeast, Saccharomyces pastorianus, is cold tolerant. It is a hybrid between Saccharomyces cerevisiae and Saccharomyces eubayanus, and has been found only in industrial settings. Natural isolates of S. eubayanus were first discovered in Patagonia 11 years ago. They have since been isolated from China, Tibet, New Zealand, and North America, but not from Europe. Here, we describe the first European strains UCD646 and UCD650, isolated from a wooded area on a university campus in Dublin, Ireland. We generated complete chromosome level assemblies of both genomes using long- and short-read sequencing. The UCD isolates belong to the Holarctic clade. Genome analysis shows that isolates similar to the Irish strains contributed to the S. eubayanus component of S. pastorianus, but isolates from Tibet made a larger contribution.

CRISPR-Cas9 Editing Induces Loss of Heterozygosity in the Pathogenic Yeast Candida parapsilosis.

Genetic manipulation is often used to study gene function. However, unplanned genome changes (including single nucleotide polymorphisms [SNPs], aneuploidy, and loss of heterozygosity [LOH]) can affect the phenotypic traits of the engineered strains. Here, we compared the effect of classical deletion methods (replacing target alleles with selectable markers by homologous recombination) with CRISPR-Cas9 editing in the diploid human-pathogenic yeast Candida parapsilosis. We sequenced the genomes of 9 isolates that were modified using classic recombination methods and 12 that were edited using CRISPR-Cas9. As a control, the genomes of eight isolates that were transformed with a Cas9-expressing plasmid in the absence of a guide RNA were also sequenced. Following gene manipulation using classic homologous recombination, only one strain exhibited extensive LOH near the targeted gene (8.9 kb), whereas another contained multiple LOH events not associated with the intended modification. In contrast, large regions of LOH (up to >1,100 kb) were observed in most CRISPR-Cas9-edited strains. LOH most commonly occurred adjacent to the Cas9 cut site and extended to the telomere in four isolates. In two isolates, we observed LOH on chromosomes that were not targeted by CRISPR-Cas9. Among the CRISPR-edited isolates, two exhibited cysteine and methionine auxotrophy caused by LOH at a heterozygous site in MET10, approximately 11 and 157 kb downstream from the Cas9 target site, respectively. C. parapsilosis isolates have relatively low levels of heterozygosity. However, our results show that mutation complementation to confirm observed phenotypes is required when using CRISPR-Cas9. IMPORTANCE CRISPR-Cas9 has greatly streamlined gene editing and is now the gold standard and first choice for genetic engineering. However, we show that in diploid species, extra care should be taken in confirming the cause of any phenotypic changes observed. We show that the Cas9-induced double-strand break is often associated with loss of heterozygosity in the asexual diploid human fungal pathogen Candida parapsilosis. This can result in deleterious heterozygous variants (e.g., stop gain in one allele) becoming homozygous, resulting in unplanned phenotypic changes. Our results stress the importance of mutation complementation even when using CRISPR-Cas9.

Systematic Analysis of Copy Number Variations in the Pathogenic Yeast Candida parapsilosis Identifies a Gene Amplification in RTA3 That is Associated with Drug Resistance.

We analyzed the genomes of 170 C. parapsilosis isolates and identified multiple copy number variations (CNVs). We identified two genes, RTA3 (CPAR2_104610) and ARR3 (CPAR2_601050), each of which was the target of multiple independent amplification events. Phylogenetic analysis shows that most of these amplifications originated only once. For ARR3, which encodes a putative arsenate transporter, 8 distinct CNVs were identified, ranging in size from 2.3 kb to 10.5 kb with 3 to 23 copies. For RTA3, 16 distinct CNVs were identified, ranging in size from 0.3 kb to 4.5 kb with 2 to ~50 copies. One unusual amplification resulted in a DUP-TRP/INV-DUP structure similar to some human CNVs. RTA3 encodes a putative phosphatidylcholine (PC) floppase which is known to regulate the inward translocation of PC in Candida albicans. We found that an increased copy number of RTA3 correlated with resistance to miltefosine, an alkylphosphocholine drug that affects PC metabolism. Additionally, we conducted an adaptive laboratory evolution experiment in which two C. parapsilosis isolates were cultured in increasing concentrations of miltefosine. Two genes, CPAR2_303950 and CPAR2_102700, coding for putative PC flippases homologous to S. cerevisiae DNF1 gained homozygous protein-disrupting mutations in the evolved strains. Overall, our results show that C. parapsilosis can gain resistance to miltefosine, a drug that has recently been granted orphan drug designation approval by the United States Food and Drug Administration for the treatment of invasive candidiasis, through both CNVs or loss-of-function alleles in one of the flippase genes. IMPORTANCE Copy number variations (CNVs) are an important source of genomic diversity that have been associated with drug resistance. We identify two unusual CNVs in the human fungal pathogen Candida parapsilosis. Both target a single gene (RTA3 or ARR3), and they have occurred multiple times in multiple isolates. The copy number of RTA3, a putative floppase that controls the inward translocation of lipids in the cell membrane, correlates with resistance to miltefosine, a derivative of phosphatidylcholine (PC) that was originally developed as an anticancer drug. In 2021, miltefosine was designated an orphan drug by the United States Food and Drug Administration for the treatment of invasive candidiasis. Importantly, we find that resistance to miltefosine is also caused by mutations in flippases, which control the outward movement of lipids, and that many C. parapsilosis isolates are prone to easily acquiring an increased resistance to miltefosine.

Clinical azole cross-resistance in Candida parapsilosis is related to a novel MRR1 gain-of-function mutation.

OBJECTIVES: Hereby, we describe the molecular mechanisms underlying the acquisition of azole resistance by a Candida parapsilosis isolate following fluconazole treatment due to candiduria. METHODS: A set of three consecutive C. parapsilosis isolates were recovered from the urine samples of a patient with candiduria. Whole-genome sequencing and antifungal susceptibility assays were performed. The expression of MRR1, MDR1, ERG11 and CDR1B (CPAR2_304370) was quantified by RT-qPCR. RESULTS: The initial isolate CPS-A was susceptible to all three azoles tested (fluconazole, voriconazole and posaconazole); isolate CPS-B, collected after the second cycle of treatment, exhibited a susceptible-dose-dependent phenotype to fluconazole and isolate CPS-C, recovered after the third cycle, exhibited a cross-resistance profile to fluconazole and voriconazole. Whole-genome sequencing revealed a putative resistance mechanism in isolate CPS-C, associated with a G1810A nucleotide substitution, leading to a G604R change in the Mrr1p transcription factor. Introducing this mutation into the susceptible CPS-A isolate (MRR1RI) resulted in resistance to fluconazole and voriconazole, as well as up-regulation of MRR1 and MDR1. Interestingly, the susceptible-dose-dependent phenotype exhibited by isolate CPS-B was associated with an increased copy number of the CDR1B gene. The expression of CDR1B was increased in both isolates CPS-B and CPS-C and in the MRR1RI strain, harbouring the gain-of-function mutation. CONCLUSIONS: Our results describe clinical azole cross-resistance acquisition in C. parapsilosis due to a G1810A (G604R) gain-of-function mutation, resulting in MRR1 hyperactivation and consequently, MDR1 efflux pump overexpression. We also associated amplification of the CDR1B gene with decreased fluconazole susceptibility and showed that it is a putative target of the MRR1 gain-of-function mutation.

The influence of hardness at varying pH on zinc toxicity and lability to a freshwater microalga, Chlorella sp.

Zinc is an essential element for aquatic organisms, however, activities such as mining and refining, as well as zinc's ubiquitous role in modern society can contribute to elevated environmental concentrations of zinc. Water hardness is widely accepted as an important toxicity modifying factor for metals in aquatic systems, though other factors such as pH are also important. This study investigated the influence of increasing water hardness, at three different pH values (6.7, 7.6 and 8.3), on the chronic toxicity of zinc to the growth rate of a microalgae, Chlorella sp. Zinc toxicity decreased with increasing hardness from 5 to 93 mg CaCO3 L-1 at all three pH values tested. The 72 h growth rate inhibition EC50 values ranged from 6.2 µg Zn L-1 (at 5 mg CaCO3 L-1, pH 8.3) to 184 µg Zn L-1 (at 92 mg CaCO3 L-1, pH 6.7). Increases in hardness from 93 to 402 mg CaCO3 L-1 generally resulted in no significant (p > 0.05) reduction in zinc toxicity. DGT-labile zinc measurements did not correspond with the observed changes in zinc toxicity as hardness was varied within a pH treatment. This suggests that cationic competition from increased hardness is decreasing zinc toxicity, rather than changes in metal lability. This study highlighted that current hardness algorithms used in water quality guidelines may not be sufficiently protective of sensitive species, such as Chlorella sp., in high hardness waters.

Draft Genome Sequence of a Diploid and Hybrid Candida Strain, Candida sanyaensis UCD423, Isolated from Compost in Ireland.

Candida sanyaensis is a CUG-Ser1 clade yeast that is associated with soil. Assembly of short-read and long-read data shows that C. sanyaensis has a diploid and hybrid genome, with approximately 97% identity between the haplotypes. The haploid genome size is approximately 15.4 Mb.

Modeling the Bioavailability of Nickel and Zinc to Ceriodaphnia dubia and Neocloeon triangulifer in Toxicity Tests with Natural Waters.

We studied biotic ligand model (BLM) predictions of the toxicity of nickel (Ni) and zinc (Zn) in natural waters from Illinois and Minnesota, USA, which had combinations of pH, hardness, and dissolved organic carbon (DOC) more extreme than 99.7% of waters in a nationwide database. We conducted 7-day chronic tests with Ceriodaphnia dubia and 96-hour acute and 14-day chronic tests with Neocloeon triangulifer and estimated median lethal concentrations and 20% effect concentrations for both species. Toxicity of Ni and Zn to both species differed among test waters by factors from 8 (Zn tests with C. dubia) to 35 (Zn tests with N. triangulifer). For both species and metals, tests with Minnesota waters (low pH and hardness, high DOC) showed lower toxicity than Illinois waters (high pH and high hardness, low DOC). Recalibration of the Ni BLM to be more responsive to pH-related changes improved predictions of Ni toxicity, especially for C. dubia. For the Zn BLM, we compared several input data scenarios, which generally had minor effects on model performance scores (MPS). A scenario that included inputs of modeled dissolved inorganic carbon and measured Al and Fe(III) produced the highest MPS values for tests with both C. dubia and N. triangulifer. Overall, the BLM framework successfully modeled variation in toxicity for both Zn and Ni across wide ranges of water chemistry in tests with both standard and novel test organisms. Environ Toxicol Chem 2021;40:3049-3062. © 2021 SETAC. This article has been contributed to by US Government employees and their work is in the public domain in the USA.

The Influence of pH on Zinc Lability and Toxicity to a Tropical Freshwater Microalga.

Increased focus on the development and application of bioavailability-based metal water quality guideline values requires increased understanding of the influence of water chemistry on metal bioavailability and toxicity. Development of empirical models, such as multiple linear regression models, requires the assessment of the influence of individual water quality parameters as toxicity-modifying factors. The present study investigated the effect of pH on the lability and toxicity of zinc (Zn) to a tropical green microalga (Chlorella sp.). Zinc speciation and lability were explored using the Windermere Humic Aqueous Model (WHAM7), ultrafiltration, and diffusive gradients in thin films (DGT). Zinc toxicity increased significantly with increasing pH from 6.7 to 8.3, with 50% growth inhibition effect concentrations decreasing from 185 to 53 µg l-1 across the pH range. Linear relationships between DGT-labile Zn and dissolved Zn did not vary across the tested pH range, nor did the linear relationship between dissolved (<0.45 µm) and ultrafiltered (<3 kDa) Zn. Our findings show that Zn toxicity to this freshwater alga is altered as a function of pH across environmentally realistic pH ranges and that these toxicity changes could not be explained by Zn speciation and lability as measured by DGT and WHAM7. Environ Toxicol Chem 2021;40:2836-2845. © 2021 SETAC.

The Cutting Edge: Surface Texture Analysis following Resection of Nerve Stumps Using Various Instruments.

BACKGROUND: Preparation of nerve ends is an essential part of nerve repair surgery. Multiple instruments have been described for this purpose; however, no consensus exists regarding which is the least traumatic for tissue handling. We believe that various instruments used for nerve-end excision will lead to different surface roughness. METHODS: Median and ulnar nerves from fresh frozen cadavers were dissected, and 1-2 cm lengths were excised using a No. 11 blade, a razor blade, or a pair of scissors. Using electron microscopy, 3-dimensional surface analysis of roughness (Sa) for each specimen was performed using ZeeScan optical hardware and GetPhase software (PhaseView, Buisson, France). An ANOVA or Kruskal-Wallis test compared roughness measures among cutting techniques. RESULTS: Forty nerves were included. Of these, 13 (32.5%) were cut using scissors, 15 (37.5%) using a razor blade, and 12 (30%) using a No. 11 blade. An ANOVA test showed statistical differences in Sa among the cutting techniques (P = 0.002), with the lowest mean Sa noted in the scissors group (7.2 µM, 95% CI: 5.34-9.06), followed by No. 11 blade (7.29 µM, 95% CI: 5.22-9.35), and razor blade (11.03 µM, 95% CI: 9.43-12.62). Median Ra (surface profile roughness) was 4.58 (IQR: 2.62-5.46). A Kruskal-Wallis test demonstrated statistical difference in Ra among techniques (P = 0.003), with the lowest by No. 11 blade (3 µM, IQR: 1.87-4.38), followed by scissors (3.29 µM, IQR: 1.56-4.96), and razor (5.41 µM, IQR: 4.95-6.21). CONCLUSION: This novel technique of 3-dimensional surface analysis found razor blade use demonstrated poor roughness, whereas a No. 11 blade or nerve-specific scissors led to equivocally smooth nerve ends.

A Review of Water Quality Factors that Affect Nickel Bioavailability to Aquatic Organisms: Refinement of the Biotic Ligand Model for Nickel in Acute and Chronic Exposures.

A review of nickel (Ni) toxicity to aquatic organisms was conducted to determine the primary water quality factors that affect Ni toxicity and to provide information for the development and testing of a biotic ligand model (BLM) for Ni. Acute and chronic data for 66 aquatic species were compiled for the present review. The present review found that dissolved organic carbon (DOC) and hardness act as toxicity-modifying factors (TMFs) because they reduced Ni toxicity to fish and aquatic invertebrates, and these effects were consistent in acute and chronic exposures. The effects of pH on Ni toxicity were inconsistent, and for most organisms there was either no effect of pH or, in some cases, a reduction in toxicity at low pH. There appears to be a unique pH effect on Ceriodaphnia dubia that results in increased toxicity at pHs above 8, but otherwise the effects of TMFs were consistent enough across all organisms and endpoints that a single set of parameters in the Ni BLM worked well with all acute and chronic toxicity data for fish, amphibians, aquatic invertebrates, and aquatic plants and algae. The unique effects of pH on C. dubia may be due to mixture toxicity involving both Ni and bicarbonate. The implications of this mixture effect on BLM modeling and a proposed set of BLM parameters for C. dubia are addressed in the review. Other than this exception, the Ni BLM with a single set of parameters could successfully predict toxicity to all acute and chronic data compiled in the present review. Environ Toxicol Chem 2021;40:2121-2134. © 2021 SETAC.