Does pre-enrichment of anodes with acetate to select for <em>Geobacter</em> spp. enhance performance of microbial fuel cells when switched to more complex substrates?

Copyright \ua9 2023 Christgen, Spurr, Milner, Izadi, McCann, Yu, Curtis, Scott and Head. Many factors affect the performance of microbial fuel cells (MFCs). Considerable attention has been given to the impact of cell configuration and materials on MFC performance. Much less work has been done on the impact of the anode microbiota, particularly in the context of using complex substrates as fuel. One strategy to improve MFC performance on complex substrates such as wastewater, is to pre-enrich the anode with known, efficient electrogens, such as Geobacter spp. The implication of this strategy is that the electrogens are the limiting factor in MFCs fed complex substrates and the organisms feeding the electrogens through hydrolysis and fermentation are not limiting. We conducted a systematic test of this strategy and the assumptions associated with it. Microbial fuel cells were enriched using three different substrates (acetate, synthetic wastewater and real domestic wastewater) and three different inocula (Activated Sludge, Tyne River sediment, effluent from an MFC). Reactors were either enriched on complex substrates from the start or were initially fed acetate to enrich for Geobacter spp. before switching to synthetic or real wastewater. Pre-enrichment on acetate increased the relative abundance of Geobacter spp. in MFCs that were switched to complex substrates compared to MFCs that had been fed the complex substrates from the beginning of the experiment (wastewater-fed MFCs - 21.9 \ub1 1.7% Geobacter spp.; acetate-enriched MFCs, fed wastewater - 34.9 \ub1 6.7% Geobacter spp.; Synthetic wastewater fed MFCs – 42.5 \ub1 3.7% Geobacter spp.; acetate-enriched synthetic wastewater-fed MFCs - 47.3 \ub1 3.9% Geobacter spp.). However, acetate pre-enrichment did not translate into significant improvements in cell voltage, maximum current density, maximum power density or substrate removal efficiency. Nevertheless, coulombic efficiency (CE) was higher in MFCs pre-enriched on acetate when complex substrates were fed following acetate enrichment (wastewater-fed MFCs – CE = 22.0 \ub1 6.2%; acetate-enriched MFCs, fed wastewater – CE =58.5 \ub1 3.5%; Synthetic wastewater fed MFCs – CE = 22.0 \ub1 3.2%; acetate-enriched synthetic wastewater-fed MFCs – 28.7 \ub1 4.2%.) The relative abundance of Geobacter ssp. and CE represents the average of the nine replicate reactors inoculated with three different inocula for each substrate. Efforts to improve the performance of anodic microbial communities in MFCs utilizing complex organic substrates should therefore focus on enhancing the activity of organisms driving hydrolysis and fermentation rather the terminal-oxidizing electrogens

Understanding drivers of antibiotic resistance genes in High Arctic soil ecosystems

This work is licensed under a Creative Commons Attribution 4.0 International License.Soils in tropical and temperate locations are known to be a sink for the genetic potential of anthropogenic-driven acquired antibiotic resistance (AR). In contrast, accumulation of acquired AR is less probable in most Polar soils, providing a platform for characterizing background resistance and establishing a benchmark for assessing AR spread. Here, high-throughput qPCR and geochemistry were used to quantify the abundance and diversity of both antibiotic resistance genes (ARGs) and selected mobile genetic elements (MGEs) across eight soil clusters in the Kongsfjorden region of Svalbard in the High Arctic. Relative ARG levels ranged by over two orders of magnitude (10−6 to 10−4 copies/16S rRNA gene copy), and showed a gradient of potential human and wildlife impacts across clusters as evidenced by altered geochemical conditions and increased “foreign” ARG abundances (i.e., allochthonous), including blaNDM-1. Impacted clusters exhibited 100× higher total ARGs and MGEs in tandem with elevated secondary nutrients, especially available P that is typically low and limiting in Arctic soils. In contrast, ARGs in less-impacted clusters correlated strongly to local soil lithology. The most plausible source of exogenous P and allochthonous ARGs in this region is bird and other wildlife guano, disseminated either by local human wastes or via direct carriage and deposition. Regardless of pathway, accumulation of apparent allochthonous ARGs and MGEs in High Arctic soils is concerning, highlighting the importance of characterizing Arctic sites now to establish benchmarks for tracking AR spread around the world

In vitro culture with gemcitabine augments death receptor and NKG2D ligand expression on tumour cells

Much effort has been made to try to understand the relationship between chemotherapeutic treatment of cancer and the immune system. Whereas much of that focus has been on the direct effect of chemotherapy drugs on immune cells and the release of antigens and danger signals by malignant cells killed by chemotherapy, the effect of chemotherapy on cells surviving treatment has often been overlooked. In the present study, tumour cell lines: A549 (lung), HCT116 (colon) and MCF-7 (breast), were treated with various concentrations of the chemotherapeutic drugs cyclophosphamide, gemcitabine (GEM) and oxaliplatin (OXP) for 24 hours in vitro. In line with other reports, GEM and OXP upregulated expression of the death receptor CD95 (fas) on live cells even at sub-cytotoxic concentrations. Further investigation revealed that the increase in CD95 in response to GEM sensitised the cells to fas ligand treatment, was associated with increased phosphorylation of stress activated protein kinase/c-Jun N-terminal kinase and that other death receptors and activatory immune receptors were co-ordinately upregulated with CD95 in certain cell lines. The upregulation of death receptors and NKG2D ligands together on cells after chemotherapy suggest that although the cells have survived preliminary treatment with chemotherapy they may now be more susceptible to immune cell-mediated challenge. This re-enforces the idea that chemotherapy-immunotherapy combinations may be useful clinically and has implications for the make-up and scheduling of such treatments

Effects of copper mineralogy and methanobactin on cell growth and sMMO activity in <i>Methylosinus trichosporium</i> OB3b

Controls on in situ methanotroph activity are not well understood. One potentially important parameter is copper (Cu) because it is the metal-centre of particulate methane monooxygenase (pMMO), the most active enzyme for oxidizing methane to methanol. Further, Cu-to-cell ratios influence the relative expression of pMMO versus the alternate soluble MMO (sMMO) in some species. However, most methanotroph studies only have assessed readily soluble forms of Cu (e.g. CuCl₂) and there is a dearth of Cu-related activity data for Cu sources more common in the environment. Here we quantified sMMO activity (as a practical indicator of Cu availability) and growth kinetics in <i>Methylosinus trichosporium</i> OB3b, an organism that expresses both pMMO and sMMO, when grown on Cu-minerals with differing dissolution equilibria to assess how mineral type and methanobactin (mb) might influence in situ methanotroph activity. Mb is a molecule produced by <i>M. trichosporium</i> OB3b that has a high affinity for Cu, reduces Cu toxicity, and may influence Cu availability in terrestrial systems. CuCO₃.Cu(OH)₂ and CuO were chosen for study based on modelling data, reflecting more and less soluble minerals, respectively, and were found to affect <i>M. trichosporium</i> OB3b activity differently. Cells grew without growth lag and with active pMMO on CuCO₃.Cu(OH)₂, regardless of the amount of mineral supplied (<500 μmoles Cu-total l^&minus;1). The organism also grew well on CuO; however, significant sMMO activity was retained up to 50 μmoles Cu-total l^&minus;1, although sMMO activity was suppressed by supplemental mb and-or direct cell-mineral contact. Mb addition increased growth rates (<i>p</i> < 0.05) with both minerals. Results show mb broadly stimulates growth, but Cu mineralogy and mb dictate whether sMMO or pMMO is active in the cells. This explains why sMMO activity has been seen in soils with high Cu and also has implications for predicting dominant MMO activity in terrestrial bioremediation applications

Microenvironment‐induced restoration of cohesive growth associated with focal activation of P ‐cadherin expression in lobular breast carcinoma metastatic to the colon

Invasive lobular carcinoma (ILC) is a special breast cancer type characterized by noncohesive growth and E‐cadherin loss. Focal activation of P‐cadherin expression in tumor cells that are deficient for E‐cadherin occurs in a subset of ILCs. Switching from an E‐cadherin deficient to P‐cadherin proficient status (EPS) partially restores cell–cell adhesion leading to the formation of cohesive tubular elements. It is unknown what conditions control EPS. Here, we report on EPS in ILC metastases in the large bowel. We reviewed endoscopic colon biopsies and colectomy specimens from a 52‐year‐old female (index patient) and of 18 additional patients (reference series) diagnosed with metastatic ILC in the colon. EPS was assessed by immunohistochemistry for E‐cadherin and P‐cadherin. CDH1 /E‐cadherin mutations were determined by next‐generation sequencing. The index patient's colectomy showed transmural metastatic ILC harboring a CDH1 /E‐cadherin p.Q610* mutation. ILC cells displayed different growth patterns in different anatomic layers of the colon wall. In the tunica muscularis propria and the tela submucosa, ILC cells featured noncohesive growth and were E‐cadherin‐negative and P‐cadherin‐negative. However, ILC cells invading the mucosa formed cohesive tubular elements in the intercryptal stroma of the lamina propria mucosae. Inter‐cryptal ILC cells switched to a P‐cadherin‐positive phenotype in this microenvironmental niche. In the reference series, colon mucosa infiltration was evident in 13 of 18 patients, one of which showed intercryptal EPS and conversion to cohesive growth as described in the index patient. The large bowel is a common metastatic site in ILC. In endoscopic colon biopsies, the typical noncohesive growth of ILC may be concealed by microenvironment‐induced EPS and conversion to cohesive growth

Lack of correlation of stem cell markers in breast cancer stem cells

BACKGROUND: Various markers are used to identify the unique sub-population of breast cancer cells with stem cell properties. Whether these markers are expressed in all breast cancers, identify the same population of cells, or equate to therapeutic response is controversial. METHODS: We investigated the expression of multiple cancer stem cell markers in human breast cancer samples and cell lines in vitro and in vivo, comparing across and within samples and relating expression with growth and therapeutic response to doxorubicin, docetaxol and radiotherapy. RESULTS: CD24, CD44, ALDH and SOX2 expression, the ability to form mammospheres and side-population cells are variably present in human cancers and cell lines. Each marker identifies a unique rather than common population of cancer cells. In vivo, cells expressing these markers are not specifically localized to the presumptive stem cell niche at the tumour/stroma interface. Repeated therapy does not consistently enrich cells expressing these markers, although ER-negative cells accumulate. CONCLUSIONS: Commonly employed methods identify different cancer cell sub-populations with no consistent therapeutic implications, rather than a single population of cells. The relationships of breast cancer stem cells to clinical parameters will require identification of specific markers or panels for the individual cancer

Identification of differentially expressed microRNAs in human male breast cancer

<p>Abstract</p> <p>Background</p> <p>The discovery of small non-coding RNAs and the subsequent analysis of microRNA expression patterns in human cancer specimens have provided completely new insights into cancer biology. Genetic and epigenetic data indicate oncogenic or tumor suppressor function of these pleiotropic regulators. Therefore, many studies analyzed the expression and function of microRNA in human breast cancer, the most frequent malignancy in females. However, nothing is known so far about microRNA expression in male breast cancer, accounting for approximately 1% of all breast cancer cases.</p> <p>Methods</p> <p>The expression of 319 microRNAs was analyzed in 9 primary human male breast tumors and in epithelial cells from 15 male gynecomastia specimens using fluorescence-labeled bead technology. For identification of differentially expressed microRNAs data were analyzed by cluster analysis and selected statistical methods.</p> <p>Expression levels were validated for the most up- or down-regulated microRNAs in this training cohort using real-time PCR methodology as well as in an independent test cohort comprising 12 cases of human male breast cancer.</p> <p>Results</p> <p>Unsupervised cluster analysis separated very well male breast cancer samples and control specimens according to their microRNA expression pattern indicating cancer-specific alterations of microRNA expression in human male breast cancer. miR-21, miR519d, miR-183, miR-197, and miR-493-5p were identified as most prominently up-regulated, miR-145 and miR-497 as most prominently down-regulated in male breast cancer.</p> <p>Conclusions</p> <p>Male breast cancer displays several differentially expressed microRNAs. Not all of them are shared with breast cancer biopsies from female patients indicating male breast cancer specific alterations of microRNA expression.</p

Does pre-enrichment of anodes with acetate to select for Geobacter spp. enhance performance of microbial fuel cells when switched to more complex substrates?

Many factors affect the performance of microbial fuel cells (MFCs). Considerable attention has been given to the impact of cell configuration and materials on MFC performance. Much less work has been done on the impact of the anode microbiota, particularly in the context of using complex substrates as fuel. One strategy to improve MFC performance on complex substrates such as wastewater, is to pre-enrich the anode with known, efficient electrogens, such as Geobacter spp. The implication of this strategy is that the electrogens are the limiting factor in MFCs fed complex substrates and the organisms feeding the electrogens through hydrolysis and fermentation are not limiting. We conducted a systematic test of this strategy and the assumptions associated with it. Microbial fuel cells were enriched using three different substrates (acetate, synthetic wastewater and real domestic wastewater) and three different inocula (Activated Sludge, Tyne River sediment, effluent from an MFC). Reactors were either enriched on complex substrates from the start or were initially fed acetate to enrich for Geobacter spp. before switching to synthetic or real wastewater. Pre-enrichment on acetate increased the relative abundance of Geobacter spp. in MFCs that were switched to complex substrates compared to MFCs that had been fed the complex substrates from the beginning of the experiment (wastewater-fed MFCs - 21.9 ± 1.7% Geobacter spp.; acetate-enriched MFCs, fed wastewater - 34.9 ± 6.7% Geobacter spp.; Synthetic wastewater fed MFCs – 42.5 ± 3.7% Geobacter spp.; acetate-enriched synthetic wastewater-fed MFCs - 47.3 ± 3.9% Geobacter spp.). However, acetate pre-enrichment did not translate into significant improvements in cell voltage, maximum current density, maximum power density or substrate removal efficiency. Nevertheless, coulombic efficiency (CE) was higher in MFCs pre-enriched on acetate when complex substrates were fed following acetate enrichment (wastewater-fed MFCs – CE = 22.0 ± 6.2%; acetate-enriched MFCs, fed wastewater – CE =58.5 ± 3.5%; Synthetic wastewater fed MFCs – CE = 22.0 ± 3.2%; acetate-enriched synthetic wastewater-fed MFCs – 28.7 ± 4.2%.) The relative abundance of Geobacter ssp. and CE represents the average of the nine replicate reactors inoculated with three different inocula for each substrate. Efforts to improve the performance of anodic microbial communities in MFCs utilizing complex organic substrates should therefore focus on enhancing the activity of organisms driving hydrolysis and fermentation rather the terminal-oxidizing electrogens

Evidence-based guidelines for managing patients with primary ER+ HER2− breast cancer deferred from surgery due to the COVID-19 pandemic

Many patients with ER+ HER2- primary breast cancer are being deferred from surgery to neoadjuvant endocrine therapy (NeoET) during the COVID-19 pandemic. We have collated data from multiple international trials of presurgical endocrine therapy in order to provide guidance on the identification of patients who may have insufficiently endocrine-sensitive tumors and should be prioritised for early surgery or neoadjuvant chemotherapy rather than NeoET during or in the aftermath of the COVID-19 pandemic for safety or when surgical activity needs to be prioritized. For postmenopausal patients, our data provide strong support for the use of ER and PgR status at diagnosis for triaging of patients into three groups in which (taking into account clinical factors): (i) NeoET is likely to be inappropriate (Allred ER 10%) indicates a higher priority for early surgery. Too few data were available for premenopausal patients to provide a similar treatment algorithm. These guidelines should be helpful for managing patients with early ER+ HER2- breast cancer during and in the aftermath of the COVID-19 crisis

Understanding drivers of antibiotic resistance genes in High Arctic soil ecosystems

Soils in tropical and temperate locations are known to be a sink for the genetic potential of anthropogenic-driven acquired antibiotic resistance (AR). In contrast, accumulation of acquired AR is less probable in most Polar soils, providing a platform for characterizing background resistance and establishing a benchmark for assessing AR spread. Here, high-throughput qPCR and geochemistry were used to quantify the abundance and diversity of both antibiotic resistance genes (ARGs) and selected mobile genetic elements (MGEs) across eight soil clusters in the Kongsfjorden region of Svalbard in the High Arctic. Relative ARG levels ranged by over two orders of magnitude (10-6 to 10-4 copies/16S rRNA gene copy), and showed a gradient of potential human and wildlife impacts across clusters as evidenced by altered geochemical conditions and increased "foreign" ARG abundances (i.e., allochthonous), including blaNDM-1. Impacted clusters exhibited 100× higher total ARGs and MGEs in tandem with elevated secondary nutrients, especially available P that is typically low and limiting in Arctic soils. In contrast, ARGs in less-impacted clusters correlated strongly to local soil lithology. The most plausible source of exogenous P and allochthonous ARGs in this region is bird and other wildlife guano, disseminated either by local human wastes or via direct carriage and deposition. Regardless of pathway, accumulation of apparent allochthonous ARGs and MGEs in High Arctic soils is concerning, highlighting the importance of characterizing Arctic sites now to establish benchmarks for tracking AR spread around the world