3D Printing in alloy design to improve biocompatibility in metallic implants.

3D Printing (3DP) or additive manufacturing (AM) enables parts with complex shapes, design flexibility, and customization opportunities for defect specific patient-matched implants. 3DP or AM also offers a design platform that can be used to innovate novel alloys for application-specific compositional modifications. In medical applications, the biological response from a host tissue depends on a biomaterial's structural and compositional properties in the physiological environment. Application of 3DP can pave the way towards manufacturing innovative metallic implants, combining structural variations at different length scales and tailored compositions designed for specific biological responses. This study shows how 3DP can be used to design metallic alloys for orthopedic and dental applications with improved biocompatibility using in vitro and in vivo studies. Titanium (Ti) and its alloys are used extensively in biomedical devices due to excellent fatigue and corrosion resistance and good strength to weight ratio. However, Ti alloys' in vivo biological response is poor due to its bioinert surface. Different coatings and surface modification techniques are currently being used to improve the biocompatibility of Ti implants. We focused our efforts on improving Ti's biocompatibility via a combination of tantalum (Ta) chemistry in Ti, the addition of designed micro-porosity, and nanoscale surface modification to enhance both in vitro cytocompatibility and early stage in vivo osseointegration, which was studied in rat and rabbit distal femur models.

Plasma metabolome of healthy and Rhodococcus equi-infected foals over time.

BACKGROUND: Foals that develop pulmonary ultrasonographic lesions on Rhodococcus equi (R. equi) endemic farms are treated with antibiotics because those at risk of developing clinical pneumonia (~20%) cannot be recognised early. Candidate biomarkers identified using metabolomics may aid targeted treatment strategies against R. equi. OBJECTIVES: (1) To describe how foal ageing affects their plasma metabolome (birth to 8 weeks) and (2) to establish the effects that experimental infection with Rhodococcus equi (R. equi) has on foal metabolome. STUDY DESIGN: Experimental study. METHODS: Nine healthy newborn foals were experimentally infected with R. equi as described in a previous study. Foals were treated with oral antibiotics if they developed clinical pneumonia (n = 4, clinical group) or remained untreated if they showed no signs of disease (n = 5, subclinical group). A group of unchallenged foals (n = 4) was also included in the study. By the end of the study period (8 weeks), all foals were free of disease. This status was confirmed with transtracheal wash fluid evaluation and culture as well as thoracic ultrasonography. Plasma metabolomics was determined by GC-MS weekly for the study duration (8 weeks). RESULTS: Foals' plasma metabolome was altered by ageing (birth to 8 weeks) and experimental infection with R. equi as demonstrated using multivariate statistical analysis. The intensities of 25 and 28 metabolites were altered by ageing and infection (p < 0.05) respectively. Furthermore, 20 metabolites changed by more than 2-fold between clinical and subclinical groups. MAIN LIMITATIONS: The number of foals is limited. Foals were experimentally infected with R. equi. CONCLUSIONS: Ageing and R. equi infection induced changes in the plasma metabolome of foals. These results provide an initial description of foal's plasma metabolome and serve as background for future identification of R. equi pneumonia biomarkers.

INTRODUCTION/CONTEXTE: Les poulains qui développent des lésions pulmonaires échographiques dans les fermes d'élevage où Rhodococcus equi (R. equi) est endémique sont traités avec antibiotiques car ceux à risque de développer des lésions cliniques (~20%) ne peuvent être identifiés précocement. Certains biomarqueurs identifiés par le biais de la métabolomique pourraient aider à orienter les stratégies de traitement pour R. equi. OBJECTIFS: (1) Décrire les changements de métabolome plasmatique qui surviennent chez les poulains en lien avec l'âge (naissance jusqu'à 8 semaines d'âge) et (2) Établir les effets d'une infection expérimentale à Rhodococcus Equi sur le métabolome des poulains. TYPE D'ÉTUDE: Étude expérimentale. MÉTHODES: Neufs poulains nouveaux-nés en santé ont été infectés de façon expérimentale par R. equi tel que décrit précédemment. Ils ont été traités avec des antibiotiques s'ils ont développé une pneumonie clinique (n = 4, groupe clinique) ou ont simplement été suivi dans le temps s'ils n'ont pas montré de signes de la maladie (n = 5, groupe sous-clinique). Un groupe de poulains sains (n = 4) était aussi inclus dans l'étude. À la fin de l'étude (8 semaines), tous les poulains étaient sains tel que confirmé par l'évaluation et la culture de leur fluide de lavage transtrachéal de même qu'à l'échographie thoracique. Les métabolomiques plasmastiques ont été déterminées par GC-MS de façon hebdomadaire pour la durée de l'étude (8 semaines). RÉSULTATS: À la fois l'âge et l'infection expérimentale ont altéré le métabolome plasmatique des poulains tel que démontré par l'analyse statistique multivariée. L'âge a altéré l'intensité de 25 métabolites et l'infection a modifié l'intensité de 28 métabolites (p < 0.05). De plus, 20 métabolites ont changé de plus de 2 fois leur valeur initiale, entre les groupes cliniques et sous-cliniques. LIMITES PRINCIPALES: Le nombre de poulains reste limité. Les poulains ont été infecté par R. equi de façon expérimentale. CONCLUSIONS: Le vieillissement et l'infection par R. equi induisent des changements dans le métabolome plasmatique des poulains. Ces résultats représentent une description initiale du métabolome plasmatique chez le poulain et peuvent servir de base pour l'identification future de biomarqueurs pour la détection de pneumonie à Rhodococcus equi.

Dietary intake of c9,t11-conjugated linoleic acid correlates with its concentration in plasma lipid fractions of men but not women.

The c9,t11-18:2 isomer of conjugated linoleic acid (c9,t11-CLA) represents the main dietary CLA form with putative health benefits. Whereas CLA intake influences the tissue CLA concentration, little is known about the association between dietary CLA and the CLA content of plasma lipid fractions. This study was designed to document fasting and nonfasting plasma c9,t11-CLA concentrations in a population of free-living adults (n = 94) and relate these concentrations to c9,t11-CLA intake. We also determined the c9,t11-CLA content of the primary plasma lipid fractions in a subset (n = 50) of our participants, related these to c9,t11-CLA intake, and determined whether c9,t11-CLA intake or plasma c9,t11-CLA was correlated with plasma cholesterol. Mean fasting plasma c9,t11-CLA concentrations were 0.46 ± 0.01 and 0.54 ± 0.01% (wt:wt) of total fatty acids for men and women, respectively (P < 0.05); nonfasting concentrations were 0.28 ± 0.01 and 0.38 ± 0.01% of total fatty acids, respectively (P < 0.001). All major esterified plasma lipid fractions contained c9,t11-CLA; TG had the highest percentages. In men, c9,t11-CLA intake correlated (r = 0.47; P < 0.05) with TG c9,t11-CLA content, suggesting that TG c9,t11-CLA may serve as a biomarker for c9,t11-CLA intake. In females, there were no correlations between c9,t11-CLA intake and the c9,t11-CLA content of any esterified plasma lipid fraction. In neither sex was there a relation between dietary c9,t11-CLA or plasma c9,t11-CLA concentration and circulating lipoprotein cholesterol concentration. The influence of sex on circulating c9,t11-CLA content and further validation of biomarkers of c9,t11-CLA intake warrant further investigation.

Binary Dynamic Logit for Correlated Ordinal: estimation, application and simulation.

We evaluate the estimation performance of the Binary Dynamic Logit model for correlated ordinal variables (BDLCO model), and compare it to GEE and Ordinal Logistic Regression performance in terms of bias and Mean Absolute Percentage Error (MAPE) via Monte Carlo simulation. Our results indicate that when the proportional-odds assumption does not hold, the proposed BDLCO method is superior to existing models in estimating correlated ordinal data. Moreover, this method is flexible in terms of modeling dependence and allows unequal slopes for each category, and can be used to estimate an apple bloom data set where the proportional-odds assumption is violated. We also provide a function in R to implement BDLCO.

Plasma lipidome of healthy and Rhodococcus equi-infected foals over time.

BACKGROUND: Many foals that develop thoracic ultrasonographic lesions as a result of Rhodococcus equi infection heal on their own. However, most of these foals receive antimicrobials because foals at risk of developing clinical pneumonia cannot be identified. Untargeted lipidomics is useful to identify candidate biomarkers. OBJECTIVES: (a) To describe the changes that occur in foal lipidomics as a result of ageing (birth to 8 weeks) and (b) To compare these results with those observed in foals after experimental infection with R. equi. STUDY DESIGN: Experimental study. METHODS: Healthy newborn foals (n = 9) were challenged with R. equi intratracheally the first week of life. Foals were treated with antimicrobials if they developed clinical pneumonia (n = 4, "clinical group") or were closely monitored if they showed no signs of disease (n = 5 "subclinical group"). An unchallenged group (n = 4) was also included. All foals were free of disease (transtracheal wash fluid evaluation and culture as well as thoracic ultrasonography) by 8 weeks of life. Plasma lipidomics was determined by LC-MS weekly for the study duration (8 weeks). RESULTS: Both ageing and experimental infection altered the foal's plasma lipidome as demonstrated by multivariate statistical analysis. The intensities of 31 lipids were altered by ageing and 12 by infection (P < .05). Furthermore, nine lipids changed by more than twofold between clinical and subclinical groups. MAIN LIMITATIONS: The number of foals is limited. Foals were experimentally challenged with R. equi. CONCLUSIONS: Ageing and R. equi infection induced changes in the plasma lipidome of foals. These experimental results provide the background for future work in the discovery of earlier biomarkers of R. equi pneumonia. Early identification of foals at risk of developing clinical pneumonia is key in order to decrease antimicrobial use and development of antimicrobial resistance.

Direct comparison of additively manufactured porous titanium and tantalum implants towards in vivo osseointegration.

Material properties of implants such as volume porosity and nanoscale surface modification have been shown to enhance cell-material interactions in vitro and osseointegration in vivo. Porous tantalum (Ta) and titanium (Ti) coatings are widely used for non-cemented implants, which are fabricated using different processing routes. In recent years, some of those implants are being manufactured using additive manufacturing. However, limited knowledge is available on direct comparison of additively manufactured porous Ta and Ti structures towards early stage osseointegration. In this study, we have fabricated porous Ta and Ti6Al4V (Ti64) implants using laser engineered net shaping (LENS™) with similar volume fraction porosity to compare the influence of surface characteristics and material chemistry on in vivo response using a rat distal femur model for 5 and 12 weeks. We have also assessed whether surface modification on Ti64 can elicit similar in vivo response as porous Ta in a rat distal femur model for 5 and 12 weeks. The harvested implants were histologically analyzed for osteoid surface per bone surface. Field emission scanning electron microscopy (FESEM) was done to assess the bone-implant interface. The results presented here indicate comparable performance of porous Ta and surface modified porous Ti64 implants towards early stage osseointegration at 5 weeks post implantation through seamless bone-material interlocking. However, a continued and extended efficacy of porous Ta is found in terms of higher osteoid formation at 12 weeks post-surgery.

An optimal set of features for predicting type IV secretion system effector proteins for a subset of species based on a multi-level feature selection approach.

Type IV secretion systems (T4SS) are multi-protein complexes in a number of bacterial pathogens that can translocate proteins and DNA to the host. Most T4SSs function in conjugation and translocate DNA; however, approximately 13% function to secrete proteins, delivering effector proteins into the cytosol of eukaryotic host cells. Upon entry, these effectors manipulate the host cell's machinery for their own benefit, which can result in serious illness or death of the host. For this reason recognition of T4SS effectors has become an important subject. Much previous work has focused on verifying effectors experimentally, a costly endeavor in terms of money, time, and effort. Having good predictions for effectors will help to focus experimental validations and decrease testing costs. In recent years, several scoring and machine learning-based methods have been suggested for the purpose of predicting T4SS effector proteins. These methods have used different sets of features for prediction, and their predictions have been inconsistent. In this paper, an optimal set of features is presented for predicting T4SS effector proteins using a statistical approach. A thorough literature search was performed to find features that have been proposed. Feature values were calculated for datasets of known effectors and non-effectors for T4SS-containing pathogens for four genera with a sufficient number of known effectors, Legionella pneumophila, Coxiella burnetii, Brucella spp, and Bartonella spp. The features were ranked, and less important features were filtered out. Correlations between remaining features were removed, and dimensional reduction was accomplished using principal component analysis and factor analysis. Finally, the optimal features for each pathogen were chosen by building logistic regression models and evaluating each model. The results based on evaluation of our logistic regression models confirm the effectiveness of our four optimal sets of features, and based on these an optimal set of features is proposed for all T4SS effector proteins.

Genomic variants link to hepatitis C racial disparities.

Chronic liver diseases are one of the major public health issues in United States, and there are substantial racial disparities in liver cancer-related mortality. We previously identified racially distinct alterations in the expression of transcripts and proteins of hepatitis C (HCV)-induced hepatocellular carcinoma (HCC) between Caucasian (CA) and African American (AA) subgroups. Here, we performed a comparative genome-wide analysis of normal vs. HCV+ (cirrhotic state), and normal adjacent tissues (HCCN) vs. HCV+HCC (tumor state) of CA at the gene and alternative splicing levels using Affymetrix Human Transcriptome Array (HTA2.0). Many genes and splice variants were abnormally expressed in HCV+ more than in HCV+HCC state compared with normal tissues. Known biological pathways related to cell cycle regulations were altered in HCV+HCC, whereas acute phase reactants were deregulated in HCV+ state. We confirmed by quantitative RT-PCR that SAA1, PCNA-AS1, DAB2, and IFI30 are differentially deregulated, especially in AA compared with CA samples. Likewise, IHC staining analysis revealed altered expression patterns of SAA1 and HNF4α isoforms in HCV+ liver samples of AA compared with CA. These results demonstrate that several splice variants are primarily deregulated in normal vs. HCV+ stage, which is certainly in line with the recent observations showing that the pre-mRNA splicing machinery may be profoundly remodeled during disease progression, and may, therefore, play a major role in HCV racial disparity. The confirmation that certain genes are deregulated in AA compared to CA tissues also suggests that there is a biological basis for the observed racial disparities.

Loss of body fat and associated decrease in leptin in early lactation are related to shorter duration of postpartum anovulation in healthy US women.

BACKGROUND: The mechanisms regulating postpartum anovulation (PPA) are not well understood, although poor nutritional status and "intense" breastfeeding behaviors are associated with longer durations. OBJECTIVES: This study was designed primarily to investigate the hypothesis that higher body fat and leptin concentrations are related to shorter PPA in healthy, lactating US women. Nutrient intake, breastfeeding behaviors, prolactin, and insulin levels were also evaluated. METHODS: Fasting serum was collected during the third trimester of pregnancy, at 2 and 4 weeks postpartum, and every 4 weeks until the occurrence of the first postpartum menses. Body composition and breastfeeding behaviors were assessed at the same times postpartum. Women (n = 13) were categorized based on initial percentage body fat and timing of first postpartum menses. RESULTS: There were no consistent relationships between body composition, leptin, or breastfeeding behaviors and duration of PPA. However, there was an interaction (P = .005) between ovulation group and time on adiposity; earlier ovulating women had less percentage body fat in early lactation than did later ovulating women. There was also an interaction (P = .1) between ovulation group and time postpartum on leptin, such that leptin concentrations tended to be greater in later than earlier ovulating groups beginning at 12 weeks postpartum. CONCLUSION: In summary, although baseline percentage body fat and leptin were not related to duration of PPA, fertility returned earlier in women who lost greater percentage body fat during the early postpartum period.

Maternal supplementation with CLA decreases milk fat in humans.

CLA refers to isomers of octadecadienoic acid with conjugated double bonds. The most abundant form of CLA (rumenic acid (RA): c9,t11-18:2) is found in milk and beef fat. Further, CLA supplements containing RA and t10,c12-18:2 are now available. Consumption of commercially produced CLA has been shown to decrease adipose accretion in growing laboratory and production animals and cause milk fat depression in cows. We tested the hypothesis that CLA supplementation would increase milk CLA concentration and decrease milk fat content in humans. Breastfeeding women (n = 9) participated in this double-blind, placebo-controlled, crossover study divided into three periods: intervention I (5 d), washout (7 d), and intervention II (5 d). Women were randomized to treatment order. During each intervention period, women consumed 1.5 g of CLA supplement or placebo (olive oil) daily; during the washout period, no supplements were consumed. Milk was collected by complete breast expression on the final day of each period; milk output was estimated by 24-h weighing on the penultimate day of each intervention period. Milk RA and t10,c12-18:2 concentrations were greater (P < 0.05) during the CLA treatment period as compared to the placebo period. Milk fat content was significantly lower during the CLA treatment, as compared to the placebo treatment (P < 0.05). Data indicate no effect of treatment on milk output. Therefore, it would be prudent that lactating women not consume commercially available CLA supplements at this time.

Effects of exposure to a combination of zinc- and lead-spiked sediments on mouthpart development and growth in Chironomus tentans.

Exposures to either zinc or lead in contaminated sediments have been shown to induce characteristic deformities in larval chironomids. This study examined the effects of exposure to lead and zinc in combination on Chironomus tentans larvae. Proportions of mouthpart deformities in populations of larvae reared in sediments containing nominal combinations of lead and zinc were tested for additive, synergistic, and antagonistic interactions using logistic regression. Metal body burdens, body size measurements, and survival were used to evaluate toxicity and developmental impacts. Results demonstrate zinc and lead mixtures produce fewer deformities than the individual metal, so their interaction may be characterized as antagonistic. However, exposure to the metal mixtures also caused delayed development and failure to hatch. The apparent decline in deformities may be an artifact of higher mortalities or developmental effects. This research provides better understanding of some of the problems and considerations for use of chironomid population deformity proportions in bioassessments for sediment metal contamination.

Optimizing experimental design using the house mouse (Mus musculus L.) as a model for determining grain feeding preferences.

There is little research evaluating flavor preferences among wheat varieties. We previously demonstrated that mice exert very strong preferences when given binary mixtures of wheat varieties. We plan to utilize mice to identify wheat genes associated with flavor, and then relate this back to human preferences. Here we explore the effects of experimental design including the number of days (from 1 to 4) and number of mice (from 2 to 15) in order to identify designs that provide significant statistical inferences while minimizing requirements for labor and animals. When mice expressed a significant preference between 2 wheat varieties, increasing the number of days (for a given number of mice) increased the significance level (decreased P-values) for their preference, as expected, but with diminishing benefit as more days were added. However, increasing the number of mice (for a given number of days) provided a more dramatic log-linear decrease in P-values and thus increased statistical power. In conclusion, when evaluating mouse feeding preferences in binary mixtures of grain, an efficient experimental design would emphasize fewer days rather than fewer animals thus shortening the experiment duration and reducing the overall requirement for labor and animals.

A yeast's eye view of mammalian reproduction: cross-species gene co-expression in meiotic prophase.

BACKGROUND: Meiotic prophase is a critical stage in sexual reproduction. Aberrant chromosome recombination during this stage is a leading cause of human miscarriages and birth defects. However, due to the experimental intractability of mammalian gonads, only a very limited number of meiotic genes have been characterized. Here we aim to identify novel meiotic genes important in human reproduction through computational mining of cross-species and cross-sex time-series expression data from budding yeast, mouse postnatal testis, mouse embryonic ovary, and human fetal ovary. RESULTS: Orthologous gene pairs were ranked by order statistics according to their co-expression profiles across species, allowing us to infer conserved meiotic genes despite obvious differences in cellular synchronicity and composition in organisms. We demonstrated that conserved co-expression networks could successfully recover known meiotic genes, including homologous recombination genes, chromatin cohesion genes, and genes regulating meiotic entry. We also showed that conserved co-expression pairs exhibit functional connections, as evidenced by the annotation similarity in Gene Ontology and overlap with physical interactions. More importantly, we predicted six new meiotic genes through their co-expression linkages with known meiotic genes, and subsequently used the genetically more amenable yeast system for experimental validation. The deletion mutants of all six genes showed sporulation defects, equivalent to a 100% validation rate. CONCLUSIONS: We identified evolutionarily conserved gene modules in meiotic prophase by integrating cross-species and cross-sex expression profiles from budding yeast, mouse, and human. Our co-expression linkage analyses confirmed known meiotic genes and identified several novel genes that might be critical players in meiosis in multiple species. These results demonstrate that our approach is highly efficient to discover evolutionarily conserved novel meiotic genes, and yeast can serve as a valuable model system for investigating mammalian meiotic prophase.

Dietary fat type influences total milk fat content in lean women.

Trans fatty acids (TFA) are found naturally in some foods (e.g., dairy products) as well as many processed foods made with partially hydrogenated vegetable oils (PHVO). Data from a growing literature suggest that some TFA decrease milk fat in lactating animals. Because the physiologic effects of TFA in lactating women are unknown, this study was designed to investigate the effects of TFA consumption on human milk fat. A randomized, crossover design (n = 12) was used to study the effect of 3 dietary treatments: high PHVO (regular margarine), low PHVO (low TFA margarine), or low PHVO but high in naturally occurring TFA (butter) on milk fat. Treatments were administered for 5 d, with 7-d washout periods. Maternal adiposity was estimated by dual-energy X-ray absorptiometry. Milk and blood were collected on d 5 of each intervention period. In general, milk and serum fatty acid concentrations mirrored those of the dietary treatments. There were significant interactions between treatment and maternal adiposity on milk fat and infant milk consumption, as well as on serum glucose and nonesterified fatty acid (NEFA) concentrations. Consumption of regular margarine, compared with low TFA margarine, resulted in lower milk fat in leaner, but not in more obese women. Consumption of either regular or low TFA margarine, compared with butter, elevated serum NEFA concentrations in the more obese women. In summary, consumption of regular margarine, compared with low TFA margarine, decreased milk fat in lean women. Further studies are required to determine whether infant milk consumption might compensate for this potentially important change in milk composition.

Consumption of conjugated linoleic acid (CLA) from CLA-enriched cheese does not alter milk fat or immunity in lactating women.

Isomers of conjugated linoleic acid (CLA) decreased milk fat, altered immunity, and reduced the risk for cardiovascular disease (CVD) in some animals. The major form of CLA in the human diet is c9,t11-18:2 (rumenic acid; RA). We studied the effects of high RA consumption on plasma and milk RA concentration, milk composition, immunity, and CVD risk factors in lactating women (n = 36) assigned to 1 of 3 treatments: control, low CLA cheese (LCLA; 160 mg RA/d), or high CLA cheese (HCLA; 346 mg RA/d). The increase in plasma RA concentration between baseline and 8 wk in women consuming HCLA cheese was significantly greater than that of controls. At study completion (8 wk), milk RA concentration among women consuming HCLA cheese was greater (P < 0.05) than that of controls (0.37 vs. 0.26% of fatty acids). Treatment did not affect milk fat, protein, or lactose concentrations, immune indices (e.g., plasma T-helper cells and interleukin-2), or measured risk factors for CVD (e.g., plasma triacylglyceride and cholesterol). In summary, consumption of a RA-enriched cheese modestly increased plasma and milk RA concentrations without affecting total milk fat, plasma and milk indices of immunity, or selected risk factors for CVD.

Morphological abnormalities in Chironomus tentans exposed to cadmium-and copper-spiked sediments.

The induction of mouthpart deformities and the developmental response with exposure to sediments spiked with three concentrations (9, 39, and 61 microgg(-1) Cd dry wt.) of cadmium (Cd) and three concentrations (30, 125, and 215 microgg(-1) Cu dry wt.) of copper (Cu) were investigated. Mouthpart deformity proportions in Chironomus tentans larvae were compared between metal-spiked and control populations and between parent and offspring (F1) populations. Cd- and Cu-treated sediments induced deformities (low Cd=13%, medium Cd=7%, high Cd=4%, low Cu=6%, medium Cu=9%, high Cu=6%) at significantly higher proportions than control (3%) sediments. No negative developmental response was determined. Larval sizes in metal-treated aquaria and control aquaria were not significantly different. F1 larvae from parents reared in medium and high Cu had significantly lower deformity rates than their parents. Our research adds to the growing evidence implicating heavy metals in general, and Cd and Cu specifically, as teratogenic agents.

Teratogenic versus mutagenic abnormalities in chironomid larvae exposed to zinc and lead.

Before chironomid mouthpart deformities can be utilized as indicators of aquatic metal pollution with certainty, it must first be established that deformities are teratogenic and not mutagenic. A laboratory experiment was conducted to assess this question using Zn and Pb as causative agents. Parent populations were reared in sediments spiked with zinc (Zn) or lead (Pb) and their resulting offspring (F1 generation) were reared in clean sediments. The proportions of mouthpart deformities in C. tentans larvae were compared via logistic regression, accounting for time of exposure, between parent and offspring populations. Results indicate that 14% of chironomids from Zn-spiked sediment contained deformed menta and/or mandibles. However, the F1-Zn generation displayed a deformity of 1.7%. Larvae reared in Pb-spiked sediments displayed a deformity frequency of 9% and the F1 generations (F1-Pb a and F1-Pb b) had deformity proportion of 7 and 6%, respectively. We concluded that the deformities caused by Zn stress were morphological because the resulting F1 deformity frequencies declined to control levels. However, deformities caused by Pb appear to be genetic since F1 deformity percentages did not differ from the parent deformity frequency. Because larvae reared in Zn- and Pb-spiked sediments were larger than larvae reared in uncontaminated sediments, we could not conclude that Zn and Pb in the sediments stunted the development of C. tentans.