Contrasting regulatory effects of organic acids on aerobic vinyl chloride biodegradation in etheneotrophs.

Vinyl chloride (VC) is a common groundwater pollutant generated during anaerobic biodegradation of chlorinated solvents (e.g., trichloroethene (TCE) or tetrachloroethene (PCE)). Aerobic VC biodegradation by etheneotrophs can support anaerobic PCE and TCE bioremediation to achieve complete removal in situ. However, anaerobic bioremediation strategies necessitate biostimulation with electron donors that are fermented in situ, generating organic acids that could influence aerobic VC biodegradation processes. We examined the effect of organic acids (lactate, acetate, propionate, and butyrate) on aerobic VC biodegradation by VC-assimilating etheneotrophs Mycobacterium strain JS60 and Nocardioides strain JS614. Strain JS60 grew on all organic acids tested, while strain JS614 did not respond to lactate. VC-grown strain JS60 fed VC and one or more organic acids showed carbon catabolite repression (CCR) behavior where VC biodegradation occurred only after organic acids were depleted. In contrast, CCR was not evident in VC-grown strain JS614, which degraded VC and organic acids simultaneously. Acetate-grown JS60 showed similar CCR behavior when fed VC and a single organic acid, except that extended lag periods (5-12 days) occurred before VC oxidation ensued. Acetate-grown JS614 fed VC and either acetate or butyrate displayed 5-8 day lag periods before simultaneous VC and organic acid biodegradation. In contrast, acetate-grown JS614 degraded VC and propionate without a significant lag, suggesting a regulatory link between propionate and VC oxidation in JS614. Different global regulatory mechanisms controlling VC biodegradation in the presence of organic acids in etheneotrophs have implications for developing combined anaerobic-aerobic bioremediation strategies at chlorinated ethene-contaminated sites. KEY POINTS: • With organic acids present, VC utilization was repressed in JS60, but not in JS614 • Strain JS60 grew readily on lactate, while strain JS614 did not • Propionate alleviated lag periods for VC utilization in acetate-grown JS614.

Natural Biodegradation of Vinyl Chloride and cis-Dichloroethene in Aerobic and Suboxic Conditions.

Chlorinated ethene (CE) groundwater contamination is commonly treated through anaerobic biodegradation (i.e., reductive dechlorination) either as part of an engineered system or through natural attenuation. Aerobic biodegradation has also been recognized as a potentially significant pathway for the removal of the lower CEs cis-1,2-dichloroethene (cDCE) and vinyl chloride (VC). However, the role of aerobic biodegradation under low oxygen conditions typical of contaminated groundwater is unclear. Bacteria capable of aerobic VC biodegradation appear to be common in the environment, while aerobic biodegradation of cDCE is less common and little is known regarding the organisms responsible. In this study, we investigate the role of aerobic cDCE and VC biodegradation in a mixed contaminant plume (including CEs, BTEX, and ketones) at Naval Air Station North Island, Installation Restoration Site 9. Sediment and groundwater collected from the plume source area, mid-plume, and shoreline were used to prepare microcosms under fully aerobic (8 mg/L dissolved oxygen (DO)) and suboxic (< 1 mg/L DO) conditions. In the shoreline microcosms, VC and cDCE were rapidly degraded under suboxic conditions (100% and 77% removal in < 62 days). In the suboxic VC microcosms, biodegradation was associated with a > 5 order of magnitude increase in the abundance of functional gene etnE, part of the aerobic VC utilization pathway. VC and cDCE were degraded more slowly under fully aerobic conditions (74% and 30% removal) in 110 days. High-throughput 16S rRNA and etnE sequencing suggest the presence of novel VC- and cDCE-degrading bacteria. These results suggest that natural aerobic biodegradation of cDCE and VC is occurring at the site and provide new evidence that low (< 1 mg/L) DO levels play a significant role in natural attenuation of cDCE and VC.

Graves' disease patients with iron deficiency anemia: serologic evidence of co-existent autoimmune gastritis.

BACKGROUND: Graves' disease (GD) has been associated with iron deficiency anemia (IDA). Atrophic gastritis leads to IDA and has been associated with autoimmune thyroid disease. This study prospectively determined the prevalence of atrophic gastritis markers and the relationship between these markers and markers of IDA in GD subjects. METHODS: Newly diagnosed GD patients (90) and controls (41) were studied. Of the newly diagnosed GD patients, 65 were consecutively enrolled and identified with GD irrespective of anemia, 25 had GD and IDA. Thyroid function, hematologic indices, and atrophic gastritis markers [parietal-cell antibodies (PCab), Helicobacter pylori antibodies (H. pylori ab), mean serum gastrin levels] were examined. RESULTS: GD patients presenting with IDA were twice as likely (64% vs. 32%, P=0.049) to harbor PCabs when compared to all other GD subjects. Unselected GD subjects (n=65) had significantly higher PCab (37% vs. 7%, P<0.001) compared to controls. Gastrin levels were significantly elevated in all GD subjects compared to controls (105 vs. 39 pg/ml, P<0.0001). This difference was magnified in PCab+ subjects (202 vs. 64 pg/ml, P=0.003). In all GD subjects, PCabs were associated with increased gastrin levels (202 vs. 75 pg/ml, P=0.0004) and lower ferritin levels (52 vs. 95, P=0.05). In GD anemic subjects, PCabs were associated with lower mean corpuscular volume (75 vs. 81, P=0.001). Gastrin levels correlated inversely with ferritin levels in all GD subjects and positively with TIBC in GD anemic subjects. CONCLUSIONS: A significant subset of patients presenting with GD may suffer from IDA due to concurrent autoimmune atrophic gastritis.

Detection of an alkene monooxygenase in vinyl chloride-oxidizing bacteria with GeneFISH.

Fluorescence in situ hybridization (FISH) can provide information on the morphology, spatial arrangement, and local environment of individual cells enabling the investigation of intact microbial communities. GeneFISH uses polynucleotide probes and enzymatic signal amplification to detect genes that are present in low copy numbers. Previously, this technique has only been applied in a small number of closely related organisms. However, many important functional genes, such as those involved in xenobiotic degradation or pathogenesis, are present in diverse microbial strains. Here, we present a geneFISH method for the detection of the functional gene etnC, which encodes the alpha subunit of an alkene monooxygenase used by aerobic ethene and vinyl chloride oxidizing bacteria (etheneotrophs). The probe concentration was optimized and found to be 100 pg/µl, similar to previous geneFISH reports. Permeabilization was necessary for successful geneFISH labeling of Mycobacteria; sequential treatment with lysozyme and achromopeptidase was the most effective treatment. This method was able to detect etnC in several organisms including Mycobacteria and Nocardioides, demonstrating for the first time that a single geneFISH probe can detect a variety of alleles (>80% sequence similarity) across multiple species. Detection of etnC with geneFISH has practical applications for bioremediation. This method can be readily adapted for other functional genes and has broad applications for investigating microbial communities in natural and engineered systems.

Cryogenic soil coring reveals coexistence of aerobic and anaerobic vinyl chloride degrading bacteria in a chlorinated ethene contaminated aquifer.

Vinyl chloride (VC) is a common groundwater contaminant and known human carcinogen. Three major bacterial guilds are known to participate in VC biodegradation: aerobic etheneotrophs and methanotrophs, and anaerobic organohalide-respiring VC-dechlorinators. We investigated the spatial relationships between functional genes representing these three groups of bacteria (as determined by qPCR) with chlorinated ethene concentrations in a surficial aquifer at a contaminated site. We used cryogenic soil coring to collect high-resolution aquifer sediment samples and to preserve sample geochemistry and nucleic acids under field conditions. All samples appeared to be anaerobic (i.e., contained little to no dissolved oxygen). VC biodegradation associated functional genes from etheneotrophs (etnC and/or etnE), methanotrophs (mmoX and/or pmoA), and anaerobic VC-dechlorinators (bvcA and/or vcrA) coexisted in 48% of the samples. Transcripts of etnC/etnE and bvcA/vcrA were quantified in contemporaneous groundwater samples, indicating co-located gene expression. Functional genes from etheneotrophs and anaerobic VC-dechlorinators were correlated to VC concentrations in the lower surficial aquifer (p < 0.05). Methanotroph functional genes were not correlated to VC concentrations. Cryogenic soil coring proved to be a powerful tool for capturing high-spatial resolution trends in geochemical and nucleic acid data in aquifer sediments. We conclude that both aerobic etheneotrophs and anaerobic VC-dechlorinators may play a significant role in VC biodegradation in aquifers that have little dissolved oxygen.

PCB dechlorination hotspots and reductive dehalogenase genes in sediments from a contaminated wastewater lagoon.

Polychlorinated biphenyls (PCBs) are a class of persistent organic pollutants that are distributed worldwide. Although industrial PCB production has stopped, legacy contamination can be traced to several different commercial mixtures (e.g., Aroclors in the USA). Despite their persistence, PCBs are subject to naturally occurring biodegradation processes, although the microbes and enzymes involved are poorly understood. The biodegradation potential of PCB-contaminated sediments in a wastewater lagoon located in Virginia (USA) was studied. Total PCB concentrations in sediments ranged from 6.34 to 12,700 mg/kg. PCB congener profiles in sediment sample were similar to Aroclor 1248; however, PCB congener profiles at several locations showed evidence of dechlorination. The sediment microbial community structure varied among samples but was dominated by Proteobacteria and Firmicutes. The relative abundance of putative dechlorinating Chloroflexi (including Dehalococcoides sp.) was 0.01-0.19% among the sediment samples, with Dehalococcoides sp. representing 0.6-14.8% of this group. Other possible PCB dechlorinators present included the Clostridia and the Geobacteraceae. A PCR survey for potential PCB reductive dehalogenase genes (RDases) yielded 11 sequences related to RDase genes in PCB-respiring Dehalococcoides mccartyi strain CG5 and PCB-dechlorinating D. mccartyi strain CBDB1. This is the first study to retrieve potential PCB RDase genes from unenriched PCB-contaminated sediments.

Continuous-Infusion Labetalol vs Nicardipine for Hypertension Management in Stroke Patients.

BACKGROUND: Labetalol and nicardipine are antihypertensives commonly used in the management of elevated blood pressure (BP) following an acute stroke, but there is limited evidence to suggest which agent as a continuous infusion should be used preferentially in this setting. OBJECTIVE: This study aimed to compare the safety, efficacy, and ease of administration of continuous-infusion labetalol with continuous-infusion nicardipine following an acute stroke. METHODS: This retrospective cohort study of patients with acute ischemic stroke or intracerebral hemorrhage included patients if they received either study agent within 24 hours of admission. The primary outcome was percent time spent at goal BP. Secondary outcomes included time to goal BP, the number of dose adjustments, and use of rescue antihypertensives. RESULTS: The analysis included 99 patients who received labetalol- (n = 34) or nicardipine- (n = 65) continuous infusions. Intracerebral hemorrhage was the most common stroke subset (n = 81) followed by acute ischemic stroke (n = 18). There was no statistical difference in time at goal BP (labetalol 68.0%, nicardipine 67.0%; P = .885), rescue antihypertensive use (labetalol 14.7%, nicardipine 24.6%; P = .2570), time spent 10% above or below mean systolic BP (labetalol 35.5%, nicardipine 33.5%; P = .885), time to goal BP (labetalol 81.4 minutes, nicardipine 56.3 minutes; P = .162), and mean number of dose adjustments (labetalol 5.9, nicardipine 6.9; P = .262). CONCLUSIONS: Labetalol- and nicardipine-continuous infusions were comparable in the studied safety and efficacy outcomes including time at goal and BP variability. Further prospective studies are needed to validate these safety and efficacy findings and to assess clinical outcomes.

Thymic Hyperplasia Associated with Graves' Disease: Pathophysiology and Proposed Management Algorithm.

BACKGROUND: The association between Graves' disease (GD) and thymic hyperplasia (TH) was first described in 1912 and has been reported numerous times thereafter. TH associated with GD presents as an incidental mediastinal mass on chest X-ray or computed tomography (CT). The pathogenesis of TH in the setting of GD is unclear but seems to involve a complex interplay of hormonal and immunological mechanisms. SUMMARY: Here, the effect that thyroid hormones and autoimmunity have on thymic growth and size is reviewed. The authors' experience, along with a review of published case reports, reveals that general physicians may be unfamiliar with this association. This lack of familiarity may result in an aggressive management course, including surgical intervention, along with its associated risks and costs. The differential diagnosis and diagnostic workup of thymic enlargement associated with GD is discussed in light of the available clinical evidence. CONCLUSION: Recent literature confirms the generally benign nature of TH associated with GD, and supports a conservative approach for the diagnostic workup and initial management. Practical management recommendations for thymic enlargement associated with GD have been formulated and are presented here.

Therapeutic drug monitoring and use of an adjusted body weight strategy for high-dose voriconazole therapy.

Objectives: A high-dose 12 mg/kg/day (6 mg/kg twice daily) voriconazole regimen was recommended by the CDC to treat patients injected with contaminated methylprednisolone acetate that caused a multi-state fungal outbreak in 2012-13. Therapeutic drug monitoring results of this unique regimen are unknown, as is the most appropriate dosing weight for obese patients. We evaluated voriconazole trough measurements for this dosing scheme, as well as the use of adjusted body weight dosing for obese patients. Methods: Voriconazole trough levels were analysed in obese (BMI ≥35 kg/m 2 ) and non-obese (BMI <35 kg/m 2 ) patients who were given initial therapy with 12 mg/kg/day. Results: Of 138 patients, the first steady-state voriconazole troughs were supratherapeutic (>5 mg/L) in 65 (47%) patients, therapeutic (2-5 mg/L) in 57 (41%) patients and subtherapeutic (<2 mg/L) in 16 (12%) patients. Twenty-three patients had pre-steady-state dose decreases due to supratherapeutic levels, with subsequent first steady-state troughs in the therapeutic ( n = 17) and subtherapeutic ( n = 6) categories. Voriconazole doses >11 and >8 mg/kg/day produced mainly first steady-state supratherapeutic troughs in 44 obese and 94 non-obese patients, respectively. An initial 12 mg/kg/day was progressively lowered to a median maintenance dose of 8.5 mg/kg/day in the obese and 8.6 mg/kg/day in the non-obese. Conclusions: A high-dose voriconazole regimen produced initial supratherapeutic troughs that required dose adjustment downward by nearly 30%. Adjusted body weight dosing in obese patients resulted in a similar maintenance dose to total body weight dosing in the non-obese, and appears to be a sensible dosing strategy for these patients.

Utilization of the clinical laboratory for the implementation of concussion biomarkers in collegiate football and the necessity of personalized and predictive athlete specific reference intervals.

BACKGROUND: A continued interest in concussion biomarkers makes the eventual implementation of identified biomarkers into routine concussion assessment an eventual reality. We sought to develop and test an interdisciplinary approach that could be used to integrate blood-based biomarkers into the established concussion management program for a collegiate football team. METHODS: We used a CLIA-certified laboratory for all testing and chose biomarkers where clinically validated testing was available as would be required for results used in clinical decision making. We summarized the existing methods and results for concussion assessment across an entire season to identify and demonstrate the challenges with the eventual integration of a parallel process using blood-based tests for concussion management. We analyzed the results of the biomarkers chosen for trends consistent with the outcome assessments provided from the current concussion management protocols. RESULTS: Baseline samples were collected with three additional post-concussion samples collected at three separate time points from players with a diagnosed concussion (n = 12). A summary of results from currently used concussion assessment tools were compared to the representative biomarkers S100B and NSE results. Nine sport-related concussions occurred during practice and three during play. For S100B, 50 % had follow-up testing results lower than the post-injury result. In contrast, 92 % of NSE follow-up results were lower than post-injury. One hundred percent of the results for S100B and NSE were within the athlete-derived reference intervals upon return-to-play and season end. CONCLUSIONS: The reported workflow provides a framework for the eventual implementation of biomarkers for concussion assessment into existing assessment protocols and strengthens the need for reliance on clinical laboratory testing. Athlete-specific reference intervals will be required to adequately interpret results.

Clinical and economic outcomes from a community hospital's antimicrobial stewardship program.

BACKGROUND: Data from community antimicrobial stewardship programs (ASPs) are limited. We describe clinical and economic outcomes from the first year of our hospital's ASP. METHODS: The ASP team comprised 2 infectious disease physicians and 3 intensive care unit pharmacists. The team prospectively audited the new starts and weekly use of 8 target antimicrobials: aztreonam, caspofungin, daptomycin, ertapenem, linezolid, meropenem, tigecycline, and voriconazole. Using administrative data, outcomes from the first year of the program, including death within 30 days of hospitalization, readmission within 30 days of discharge, and development of Clostridium difficile infection (CDI), were compared with outcomes from a similar period before institution of the program. RESULTS: A total of 510 antimicrobial orders were reviewed, of which 323 (63%) were appropriate, 94 (18%) prompted deescalation, 61 (12%) were denied, and 27 (5%) led to formal consultation with an infectious disease physician. On multivariate analysis, implementation of the ASP was associated with an approximate 50% reduction in the odds of developing CDI (odds ratio, 0.46; 95% confidence interval, 0.25-0.82). The ASP was not associated with decreased mortality at 30 days after discharge or readmission rate. The antimicrobial cost per patient-day decreased by 13.3%, from $10.16 to $8.81. The antimicrobial budget decreased by 15.2%, resulting in a total savings of $228,911. There was a 25.4% decrease in defined daily doses of the target antimicrobials. CONCLUSIONS: Implementation of the ASP was associated with significant reductions in CDI rate, antimicrobial use, and pharmacy costs.

Increased generation of fibrocytes in thyroid-associated ophthalmopathy.

CONTEXT: The pathogenic basis for Graves' disease (GD) continues to elude our understanding. Specifically why activating antibodies are generated against self-antigens remains uncertain as does the identity of the antigen(s) that provokes orbital involvement in GD, a process known as thyroid-associated ophthalmopathy (TAO). OBJECTIVE: The aim of the study was to determine whether CD34(+) fibrocytes are generated more frequently in GD, whether they infiltrate orbital connective tissues in TAO, and whether they express the thyrotropin receptor (TSHR). DESIGN/SETTING/PARTICIPANTS: Generation of fibrocytes from peripheral blood mononuclear cells was examined in samples from 70 patients with GD and 25 healthy control subjects. Fibrocytes were characterized by flow cytometry. Orbital tissues and fibroblast culture strains were examined for their presence. MAIN OUTCOME MEASURES: The frequency of CD34(+) fibrocyte generation from peripheral blood cells, characterization of their phenotype, cytokine production, and their presence in affected orbital tissues were analyzed. RESULTS: CD34(+)CXCR4(+)Col I(+) fibrocytes expressing IGF-I receptor are far more frequently generated from cultured peripheral blood mononuclear cells of donors with GD compared with healthy subjects. They express TSHR at high levels and TSH induces fibrocytes to produce IL-6 and TNF-alpha. Numerous CD34(+) fibrocytes were detected in orbital tissues in TAO but were absent in healthy orbits. Tissue-infiltrating fibrocytes express TSHR in situ and comprise a subpopulation of TAO-derived orbital fibroblasts. CONCLUSIONS: Our findings suggest that fibrocytes may participate in the pathogenesis of TAO because they express relevant autoantigens such as IGF-I receptor and functional TSHR and differentially accumulate in orbital tissue in TAO.

Characterization of the anaemia associated with Graves' disease.

BACKGROUND: Graves' disease (GD) is associated with hyperthyroidism. Thyrotoxicosis adversely affects multiple organ systems including haematopoiesis. Anaemia occurring specifically in GD has not been systematically studied previously. OBJECTIVE: To define the prevalence and characteristics of the anaemia associated with GD. DESIGN: Eighty-seven newly diagnosed patients with GD were recruited. Haematological indices, thyroid function and inflammatory parameters were examined at presentation and following successful treatment of hyperthyroidism. SETTING: Tertiary care academic referral centre. RESULTS: Thirty-three per cent of subjects presented with anaemia. The prevalence of anaemia not attributable to other causes (GD anaemia) was 22%. GD anaemia affected 41.6% (10/24) of men compared to 17.5% of women (11/63). Mean erythropoietin (EPO) levels (15.5 +/- 5.3 mIU/ml) were within normal reference limits but significantly higher (P = 0.004) than those of the non-anaemic controls. Hgb correlated inversely with EPO (P = 0.05) and CRP (P = 0.04) levels, a relationship that persisted after multivariate adjustment for TT3 or TT4. With antithyroid therapy for 16 +/- 6.3 weeks, Hgb levels normalized in 8 out of 9 subjects with GD anaemia (10.7 +/- 0.8 to 13.5 +/- 1.3 g/dl, P = 0.0001). After normalization of Hgb, mean MCV and TIBC were significantly increased, and median ferritin and mean EPO were significantly decreased. CONCLUSIONS: GD anaemia is common, resembles the anaemia of chronic disease, and is associated with markers of inflammation. It corrects promptly with return to the euthyroid state following treatment.