Nocardia cyriacigeorgica Elicits Gut Disturbances in a Leaky Gut Model of Colitis, but Not the Harmful Cascade Leading to Gut-First Parkinson's Disease.

Parkinson's disease (PD) is a progressive neurodegenerative disorder with an unknown cause. Recent research has highlighted the importance of the gut in neuronal and immune maturation through the exchange of nutrients and cellular signals. This has led to the "gut-first PD" hypothesis, which aims to explain many of the sporadic cases and their prodromal intestinal symptoms, such as constipation and intestinal α-synuclein (aSyn) aggregation. The link between mitochondrial dysfunction and aSyn deposition is central to PD pathophysiology, since they can also trigger pro-inflammatory signals associated with aSyn deposition, potentially contributing to the onset of PD. As mitochondria are derived from ancestral alpha-proteobacteria, other bacteria may specifically target this organelle. We sought to use Nocardia cyriacigeorgica, a bacterium previously associated with parkinsonism, and dextran sulfate sodium (DSS) as pro-inflammatory modulators to gain further insight into the onset of PD. This study indicates that aSyn aggregation plus mitochondrial dysfunction without intestinal barrier leakage are not sufficient to trigger gut-first PD.

Influence of COVID-19 pandemic on molar pregnancy and postmolar gestational trophoblastic neoplasia: An observational study.

OBJECTIVE: To assess whether the incidence and aggressiveness of molar pregnancy (MP) and postmolar gestational trophoblastic neoplasia (GTN) changed during the COVID-19 pandemic. DESIGN: Observational study with two separate designs: retrospective multicentre cohort of patients with MP/postmolar GTN and a cross-sectional analysis, with application of a questionnaire. SETTING: Six Brazilian Reference Centres on gestational trophoblastic disease. POPULATION: 2662 patients with MP/postmolar GTN treated from March-December/2015-2020 were retrospectively evaluated and 528 of these patients answered a questionnaire. METHODS: Longitudinal retrospective multicentre study of patients diagnosed with MP/ postmolar GTN at presentation and a cross-sectional analysis, with application of a questionnaire, exclusive to patients treated during the period of study, to assess living and health conditions during the COVID-19 pandemic compared with previous years. MAIN OUTCOME MEASURES: The incidence of MP/postmolar GTN. RESULTS: Compared with the last 5 pre-pandemic years, MP/postmolar GTN incidence remained stable during 2020 (COVID-19 pandemic). Multivariable logistic regression, adjusted for the patient age, showed that during 2020, presentation with MP was more likely to be >10 weeks of gestation (adjusted odds ratio [aOR] 2.50, 95% confidence interval [CI] 1.90-3.29, P < 0.001), have a pre-evacuation hCG level ≥100 000 iu/l (aOR 1.77, 95% CI 1.38-2.28, P < 0.001) and time to the initiation of chemotherapy ≥7 months (aOR 1.86, 95% CI 1.01-3.43, P = 0.047) when compared with 2015-2019. CONCLUSIONS: Although the incidence of MP/postmolar GTN remained stable during the COVID-19 pandemic in Brazil, the pandemic was associated with greater gestational age at MP diagnosis and more protracted delays in initiation of chemotherapy for postmolar GTN.

Microalgal cultures for the remediation of wastewaters with different nitrogen to phosphorus ratios: Process modelling using artificial neural networks.

Microalgae have remarkable potential for wastewater bioremediation since they can efficiently uptake nitrogen and phosphorus in a sustainable and environmentally friendly treatment system. However, wastewater composition greatly depends on its source and has a significant seasonal variability. This study aimed to evaluate the impact of different N:P molar ratios on the growth of Chlorella vulgaris and nutrient removal from synthetic wastewater. Furthermore, artificial neural network (ANN) threshold models, optimised by genetic algorithms (GAs), were used to model biomass productivity (BP) and nitrogen/phosphorus removal rates (RRN/RRP). The impact of various inputs culture variables on these parameters was evaluated. Microalgal growth was not nutrient limited since the average biomass productivities and specific growth rates were similar between the experiments. Nutrient removal efficiencies/rates reached 92.0 ± 0.6%/6.15 ± 0.01 mgN L-1 d-1 for nitrogen and 98.2 ± 0.2%/0.92 ± 0.03 mgP L-1 d-1 for phosphorus. Low nitrogen concentration limited phosphorus uptake for low N:P ratios (e.g., 2 and 3, yielding 36 ± 2 mgDW mgP-1 and 39 ± 3 mgDW mgP-1, respectively), while low phosphorus concentration limited nitrogen uptake with high ratios (e.g., 66 and 67, yielding 9.0 ± 0.4 mgDW mgN-1 and 8.8 ± 0.3 mgDW mgN-1, respectively). ANN models showed a high fitting performance, with coefficients of determination of 0.951, 0.800, and 0.793 for BP, RRN, and RRP, respectively. In summary, this study demonstrated that microalgae could successfully grow and adapt to N:P molar ratios between 2 and 67, but the nutrient uptake was impacted by these variations, especially for the lowest and highest N:P molar ratios. Furthermore, GA-ANN models demonstrated to be relevant tools for microalgal growth modelling and control. Their high fitting performance in characterising this biological system can contribute to reducing the experimental effort for culture monitoring (human resources and consumables), thus decreasing the costs of microalgae production.

The Role of Bacteria-Mitochondria Communication in the Activation of Neuronal Innate Immunity: Implications to Parkinson's Disease.

Mitochondria play a key role in regulating host metabolism, immunity and cellular homeostasis. Remarkably, these organelles are proposed to have evolved from an endosymbiotic association between an alphaproteobacterium and a primitive eukaryotic host cell or an archaeon. This crucial event determined that human cell mitochondria share some features with bacteria, namely cardiolipin, N-formyl peptides, mtDNA and transcription factor A, that can act as mitochondrial-derived damage-associated molecular patterns (DAMPs). The impact of extracellular bacteria on the host act largely through the modulation of mitochondrial activities, and often mitochondria are themselves immunogenic organelles that can trigger protective mechanisms through DAMPs mobilization. In this work, we demonstrate that mesencephalic neurons exposed to an environmental alphaproteobacterium activate innate immunity through toll-like receptor 4 and Nod-like receptor 3. Moreover, we show that mesencephalic neurons increase the expression and aggregation of alpha-synuclein that interacts with mitochondria, leading to their dysfunction. Mitochondrial dynamic alterations also affect mitophagy which favors a positive feedback loop on innate immunity signaling. Our results help to elucidate how bacteria and neuronal mitochondria interact and trigger neuronal damage and neuroinflammation and allow us to discuss the role of bacterial-derived pathogen-associated molecular patterns (PAMPs) in Parkinson's disease etiology.

Microalgae systems - environmental agents for wastewater treatment and further potential biomass valorisation.

Water is the most valuable resource on the planet. However, massive anthropogenic activities generate threatening levels of biological, organic, and inorganic pollutants that are not efficiently removed in conventional wastewater treatment systems. High levels of conventional pollutants (carbon, nitrogen, and phosphorus), emerging chemical contaminants such as antibiotics, and pathogens (namely antibiotic-resistant ones and related genes) jeopardize ecosystems and human health. Conventional wastewater treatment systems entail several environmental issues: (i) high energy consumption; (ii) high CO2 emissions; and (iii) the use of chemicals or the generation of harmful by-products. Hence, the use of microalgal systems (entailing one or several microalgae species, and in consortium with bacteria) as environmental agents towards wastewater treatment has been seen as an environmentally friendly solution to remove conventional pollutants, antibiotics, coliforms and antibiotic resistance genes. In recent years, several authors have evaluated the use of microalgal systems for the treatment of different types of wastewater, such as agricultural, municipal, and industrial. Generally, microalgal systems can provide high removal efficiencies of: (i) conventional pollutants, up to 99%, 99%, and 90% of total nitrogen, total phosphorus, and/or organic carbon, respectively, through uptake mechanisms, and (ii) antibiotics frequently found in wastewaters, such as sulfamethoxazole, ciprofloxacin, trimethoprim and azithromycin at 86%, 65%, 42% and 93%, respectively, through the most desirable microalgal mechanism, biodegradation. Although pathogens removal by microalgal species is complex and very strain-specific, it is also possible to attain total coliform and Escherichia coli removal of 99.4% and 98.6%, respectively. However, microalgal systems' effectiveness strongly relies on biotic and abiotic conditions, thus the selection of operational conditions is critical. While the combination of selected species (microalgae and bacteria), ratios and inoculum concentration allow the efficient removal of conventional pollutants and generation of high amounts of biomass (that can be further converted into valuable products such as biofuels and biofertilisers), abiotic factors such as pH, hydraulic retention time, light intensity and CO2/O2 supply also have a crucial role in conventional pollutants and antibiotics removal, and wastewater disinfection. However, some rationale must be considered according to the purpose. While alkaline pH induces the hydrolysis of some antibiotics and the removal of faecal coliforms, it also decreases phosphates solubility and induces the formation of ammonium from ammonia. Also, while CO2 supply increases the removal of E. coli and Pseudomonas aeruginosa, as well as the microalgal growth (and thus the conventional pollutants uptake), it decreases Enterococcus faecalis removal. Therefore, this review aims to provide a critical review of recent studies towards the application of microalgal systems for the efficient removal of conventional pollutants, antibiotics, and pathogens; discussing the feasibility, highlighting the advantages and challenges of the implementation of such process, and presenting current case-studies of different applications of microalgal systems.

Predictors for single-agent resistance in FIGO score 5 or 6 gestational trophoblastic neoplasia: a multicentre, retrospective, cohort study.

BACKGROUND: Patients with gestational trophoblastic neoplasia who have an International Federation of Gynaecology and Obstetrics (FIGO) risk score of 5 or 6 usually receive non-toxic single-agent chemotherapy as a first-line treatment. Previous studies suggest that only a third of patients have complete remission, with the remaining patients requiring toxic multiagent chemotherapy to attain remission. As stratification factors are unknown, some centres offer multiagent therapy upfront, resulting in overtreatment of many patients. We aimed to identify predictive factors for resistance to single-agent therapy to inform clinicians on which patients presenting with a FIGO score of 5 or 6 are likely to benefit from upfront multiagent chemotherapy. METHODS: We did a multicentre, retrospective, cohort study of patients with gestational trophoblastic neoplasia presenting with a FIGO score of 5 or 6, who received treatment at three gestational trophoblastic neoplasia reference centres in the UK, Brazil, and the USA between Jan 1, 1964, and Dec 31, 2018. All patients who had been followed up for at least 12 months after remission were included. Patients were excluded if they had received a non-standard single-agent treatment (eg, etoposide); had been given a previously established first-line multiagent chemotherapy regimen; or had incomplete data for our analyses. Patient data were retrieved from medical records. The primary outcome was the incidence of chemoresistance after first-line or second-line single-agent chemotherapy. Variables associated with chemoresistance to single-agent therapies were identified by logistic regression analysis. In patient subgroups defined by choriocarcinoma histology and metastatic disease status, we did bootstrap modelling to define thresholds of pretreatment human chorionic gonadotropin concentrations and identify groups of patients with a greater than 80% risk (ie, a positive predictive value [PPV] of 0·8) of resistance to single-agent chemotherapy. FINDINGS: Of 5025 patients with low-risk gestational trophoblastic neoplasia, we identified 431 patients with gestational trophoblastic neoplasia presenting with a FIGO risk score of 5 or 6. All patients were followed up for a minimum of 2 years. 141 (40%) of 351 patients developed resistance to single-agent treatments and required multiagent chemotherapy to achieve remission. Univariable and multivariable logistic regression revealed metastatic disease status (multivariable logistic regression analysis, odds ratio [OR] 1·9 [95% CI 1·1-3·2], p=0·018), choriocarcinoma histology (3·7 [1·9-7·4], p=0·0002), and pretreatment human chorionic gonadotropin concentration (2·8 [1·9-4·1], p<0·0001) as significant predictors of resistance to single-agent therapies. In patients with no metastatic disease and without choriocarcinoma, a pretreatment human chorionic gonadotropin concentration of 411 000 IU/L or higher yielded a PPV of 0·8, whereas in patients with either metastases or choriocarcinoma, a pretreatment human chorionic gonadotropin concentration of 149 000 IU/L or higher yielded the same PPV for resistance to single-agent therapy. INTERPRETATION: Approximately 60% of women with gestational trophoblastic neoplasia presenting with a FIGO risk score of 5 or 6 achieve remission with single-agent therapy; almost all remaining patients have complete remission with subsequent multiagent chemotherapy. Primary multiagent chemotherapy should only be given to patients with metastatic disease and choriocarcinoma, regardless of pretreatment human chorionic gonadotropin concentration, or to those defined by our new predictors. FUNDING: None. TRANSLATION: For the Portuguese translation of the abstract see Supplementary Materials section.

Right bundle brunch block in patients with acute myocardial infarction is associated with a higher in-hospital arrhythmic risk and mortality, and a worse prognosis after discharge.

INTRODUCTION: Recently, the presence of right bundle brunch block (RBBB) in patients with persistent ischaemic symptoms has been suggested as an indication for emergent coronary angiography. OBJECTIVE: The aim of this study was to assess the prognostic impact of RBBB in patients with acute myocardial infarction (AMI) before the implementation of the recent recommendations. METHODS: We retrospectively studied consecutive patients admitted with AMI between 2011 and 2013. Patients with left bundle brunch block, pacemaker, or nonspecific intraventricular conduction delay were excluded. Patients with RBBB were compared with those without RBBB. Clinical characteristics, in-hospital evolution, and major adverse cardiovascular events (MACE) during follow-up, defined as cardiovascular death, sustained ventricular arrhythmias, acute heart failure syndromes, recurrent myocardial infarction, or acute stroke, were analysed. RESULTS: The analysis included 481 patients. Thirty two patients (6.7%) had RBBB. Patients with RBBB were older. During hospital admission, RBBB patients had a higher rate of sustained ventricular tachycardia and death. Survival curve analysis showed that patients with RBBB had a lower in-hospital survival rate (Log-rank, p = 0.004). After discharge, during a mean follow-up time of 24.3 ± 11.6 months, 53 patients (12%) died. Survival curve analysis showed a lower survival rate free of MACE for those patients with RBBB (Log-rank, p = 0.011). RBBB was independently associated with MACE occurrence (HR 2.17, 95% CI 1.07-4.43; p = 0.033), after adjusting for demographic data, coronary angiography findings, treatment performed, echocardiographic evaluation, and medical therapy. CONCLUSION: Patients with RBBB had a higher rate of in-hospital mortality and arrhythmic events, and an increased risk of MACE during follow-up.

Premature ventricular contractions of the right ventricular outflow tract: is there an incipient underlying disease? New insights from a speckle tracking echocardiography study.

CONTEXT: Premature ventricular contractions (PVCs) originating in the right ventricular outflow tract (RVOT) are traditionally considered idiopathic and benign. Echocardiographic conventional measurements are typically normal. AIMS: To assess whether right ventricle longitudinal strain, determined by two-dimensional speckle tracking echocardiography, differ between RVOT PVCs patients (treated with catheter ablation) and healthy controls. METHODS: We retrospectively selected patients with PVCs from the RVOT who underwent electrophysiological study and catheter ablation between 2016 and 2019. Patients with documented structural heart disease were excluded. Transthoracic echocardiography was performed and right ventricle global longitudinal strain (RV-GLS), free wall longitudinal strain (RVFW-LS) and left ventricle global longitudinal strain (LV-GLS) were determined as well as conventional ultrasound measurements of RV and LV function. RESULTS: We studied 21 patients with RVOT PVCs and 13 controls. Patients with PVCs from the RVOT had lower values of RV-GLS and RVFW-LS compared with the control group (-19.4% versus -22.5%, P = 0.015 and -22.1% versus -25.5, P = 0.041, respectively). They also had lower values of LV-GLS, although still within the normal range (-19.1% versus -20.9%, P = 0.047). Regarding RVOT PVCs patients only, RV-GLS and RVFW-LS had no correlation with the PVCs burden prior to catheter ablation and they did not differ between the patients in whom the catheter ablation was successful and those in whom it was not. RV-GLS also had a positive correlation with RVOT proximal diameter (r = 0.487, P = 0.025). CONCLUSIONS: In this group of RVOT PVCs patients, we found worse RV longitudinal strain values (and therefore sub-clinical myocardial dysfunction) when compared to healthy controls.

Gestational trophoblastic neoplasia lethality among Brazilian women: A retrospective national cohort study.

OBJECTIVE: To investigate GTN lethality among Brazilian women comparing cases of death by GTN with those who survived, thereby identifying factors associated with GTN lethality. METHODS: We retrospectively reviewed medical records of women with GTN treated at ten Brazilian GTN Reference Centers, from January 1960 to December 2017. We evaluated factors associated with death from GTN and used Cox proportional hazards regression models to identify independent variables with significant influence on the risk of death. RESULTS: From 2186 patients with GTN included in this study, 2092 (95.7%) survived and 89 (4%) died due to GTN. When analyzing the relative risk (RR), adjusted for WHO/FIGO score, patients with low risk disease had a significantly higher risk of death if they had choriocarcinoma (RR: 12.40), metastatic disease (RR: 12.57), chemoresistance (RR: 3.18) or initial treatment outside the Reference Center (RR: 12.22). In relation to patients with high-risk GTN, these factors were significantly associated with death due to GTN: the time between the end of antecedent pregnancy and the initiation of chemotherapy (RR: 4.10), metastatic disease (RR: 14.66), especially in brain (RR: 8.73) and liver (RR: 5.76); absence of chemotherapy or initial treatment with single agent chemotherapy (RR: 10.58 and RR: 1.81, respectively), chemoresistance (RR: 3.20) and the initial treatment outside the Reference Center (RR: 28.30). CONCLUSION: The risk of mortality from low and high-risk GTN can be reduced by referral of these patients to a Reference Center or, if not possible, to involve clinicians in a Reference Center with consultation regarding management.

Comparison of treatment for low-risk GTN with standard 8-day MTX/FA regimen versus modified MTX/FA regimen without chemotherapy on the weekend.

OBJECTIVE: To compare the outcomes of patients with low-risk gestational trophoblastic neoplasia (GTN) treated with standard 8-day methotrexate/folinic acid (MTX/FA) versus modified regimen. METHODS: Retrospective cohort study of patients with low-risk GTN followed at Rio de Janeiro Federal University, from January/1990-December/2017 with standard 8-day MTX/FA or modified regimen (MTX administered on the 8th day rather than 7th) to avoid treatment on the weekend. RESULTS: From 937 patients with low-risk GTN, 538 were treated with standard MTX/FA and 98 patients received modified regimen. Both groups were comparable in age (p = .749), antecedent pregnancy (p = .221), time to initiate chemotherapy (p = .926), hCG pretreatment level (p = .112) and WHO/FIGO prognostic risk score (p = .723). Patients treated with modified MTX/FA had twice of cases of metastatic lung disease compared with the standard regimen (22.5% vs 10.6%; p = .002). The rate of remission (p = .999), number of cycles to remission in the first-line (p = .966), chemoresistance (p = .500), time to switch to second-line therapy (p = .176), need for multiagent chemotherapy (p = .084), relapse (p = .122) or death (p = .475) was the same for both MTX/FA regimen. However, although patients receiving modified MTX/FA required a higher total number of remission cycles (6 vs 5 cycles; p = .004) and longer time to remission (19 vs 16 weeks; p < .001) when compared with the standard regimen, these variables showed no significant differences after multivariate logistic regression adjusted for lung metastasis. CONCLUSION: The modified 8-day MTX/FA regimen didn't compromise oncologic outcomes for women with low-risk GTN. This regimen appears to be an acceptable alternative to standard 8-day MTX/FA when treatment on weekend isn't an option.

Prostate cancer cells enhance interleukin-15-mediated expansion of NK cells.

OBJECTIVES: To identify cytokines that can activate and expand NK cells in the presence of prostate cancer cells in order to determine whether these agents may be useful in future intra-tumoural administration in pre-clinical and clinical prostate cancer trials. MATERIALS AND METHODS: Lymphocytes isolated from normal donor blood were set up in co-cultures with either cancer or non-cancerous prostate cell lines, together with each of the cytokines interleukin (IL)-2, IL-12, IL-15, interferon (IFN)-γ or IL-21 for a period of 7 days. Then, expansion of NK cells, NKT cells and CD8 T cells was measured by flow cytometry and compared with the expansion of the same cells in the absence of prostate cells. The cytotoxic activity of NK cells, as measured by perforin and tumour cell killing, was also assessed. NK cell receptors and their corresponding ligands on prostate tumour cells were analysed to determine whether any of these were modulated by co-culture. The role of the tumour-secreted heat shock proteins HSP90 and HSP70 in the expansion of NK cells in the co-cultures was also investigated because of their effects on NK and CD8 T-cell activation. RESULTS: We showed that, among a panel of cytokines known to cause NK cell activation and expansion, only IL-15 could actively induce expansion of NK, NKT and CD8 T cells in the presence of prostate cancer cell lines. Furthermore, the expansion of NK cells was far greater (up to 50% greater) in the presence of the cancer cells (LNCaP, PC3) than when lymphocytes were incubated alone. In contrast, non-cancerous cell lines (PNT2 and WPMY-1) did not exert any expansion of NK cells. The cytolytic activity of the NK cells, as measured by perforin, CD107a and killing of tumour cells, was also greatest in co-cultures with IL-15. Examination of NK cell receptors shows that NKG2D is upregulated to a greater degree in the presence of prostate cancer cells, compared with the upregulation with IL-15 in lymphocytes alone. However, blocking of NKG2D does not inhibit the enhanced expansion of NK cells in the presence of tumour cells. CONCLUSIONS: Among a panel of NK cell-activating cytokines, IL-15 was the only cytokine that could stimulate expansion of NK cells in the presence of prostate cancer cells; therefore IL-15 may be a good candidate for novel future intra-tumoural therapy of the disease.

Revealing the hidden diversity of marine fungi in Portugal with the description of two novel species, Neoascochyta fuci sp. nov. and Paraconiothyrium salinum sp. nov.

Fungi are ubiquitous organisms with a wide distribution in almost all ecosystems, including marine environments. Coastal and estuarine ecosystems remain poorly unexplored as fungal habitats, potentially harbouring a hidden diversity with important ecological roles. During an extensive survey of marine fungi in coastal and estuarine Portuguese environments, a collection of 612 isolates was obtained from water, algae, sponges and driftwood. From these, 282 representative isolates were selected through microsatellite-primed PCR (MSP-PCR) fingerprinting analysis, which were identified based on DNA sequence data. The collection yielded 117 taxa from 38 distinct genera, which were identified using DNA sequence analysis. Overall, fungal community composition varied with host/substrate, but the most abundant taxa in the collection were Cladosporium cladosporioides, Penicillium terrigenum, Penicillium brevicompactum and Fusarium equiseti/incarnatum complex. The occurrence of a high fungal diversity harbouring novel species was disclosed. Through a multilocus phylogeny based on ITS, tub2 and tef1-α sequences, in conjunction with morphological and physiological data, we propose Neoascochyta fuci sp. nov. and Paraconiothyrium salinum sp. nov.

Effect of γ-Aminobutyric Acid (GABA) on the Metabolome of Two Strains of Lasiodiplodia theobromae Isolated from Grapevine.

The effect of γ-aminobutyric acid (GABA) on the metabolome of two strains of Lasiodiplodia theobromae isolated from grapevine that hold a different degree of virulence to the host plant (LA-SOL3 (more virulent), LA-SV1 (less virulent)) was investigated. The culture filtrates and crude extracts from the two strains grown in the presence and absence of 10 mM of GABA were tested for phytotoxicity on tomato plant cuttings and leaves, respectively. Considering the opportunistic nature of this fungus for humans, crude extracts were also tested for cytotoxicity on mammalian cell lines. We found that culture filtrates and crude extracts have a decreased toxicity in the presence of GABA. Metabolomic analysis, conducted on both strains at both growth conditions, revealed the production of several compounds, such as indole-3-carboxylic acid (ICA, which is the main compound produced by L. theobromae), 3-indolecarboxyaldehyde, (3R,4S)-botryodiplodin, (R)-mellein. Finally, data demonstrate that GABA both induces a decrease in the amount of ICA, and a diversification of the metabolites produced by L. theobromae.

Can carboplatin or etoposide replace actinomycin-d for second-line treatment of methotrexate resistant low-risk gestational trophoblastic neoplasia?

OBJECTIVE: To evaluate the impact of periodic shortage of actinomycin-d (Act-d) in the treatment of Brazilian patients with low-risk gestational trophoblastic neoplasia (GTN) after methotrexate and folinic acid rescue (MTX/FA) resistance, treated alternately with carboplatin or etoposide as a second-line regimen. METHODS: Retrospective cohort that included patients with failure of first-line MTX/FA regimen for low-risk GTN treated at Rio de Janeiro Federal University, Universidade Federal de São Paulo and Irmandade da Santa Casa de Misericórdia de Porto Alegre, from January/2010- December/2017. RESULTS: From 356 patients with low-risk GTN treated with MTX/FA, 75 (21.1%) developed resistance, of which 40 (53.3%) received Act-d, 23 (30.7%) carboplatin and 7 (9.3%) etoposide. Although patients treated with single-agent chemotherapy as a second-line regimen had comparable clinical and primary treatment characteristics, those treated with Act-d (80%, p = 0.033) or etoposide (71.4%, p = 0.025) had higher remission rates when compared with carboplatin (47.8%). Only 29% of patients treated with carboplatin received the chemotherapy cycles without delay compared to Act-d (98%, p < 0.001) or etoposide (85%, p = 0.009). Patients treated with carboplatin had significantly more hematological toxicity, notably anemia (30.4%, p = 0.008), lymphopenia (47.7%, p < 0.001) and thrombocytopenia (43.4%, p < 0.001), as well as a higher occurrence of febrile neutropenia (14.4%, p = 0.044) and vomiting (60%, p < 0.001) than those receiving Act-d (5%, none, 2.5%, none, 10%, respectively). CONCLUSION: Carboplatin did not have a satisfactory clinical response rate, likely due to severe hematological toxicity, which postponed chemotherapy. Our results reinforce the preference for Act-d as a second-line agent in patients with low-risk GTN after MTX/FA resistance.

Verrucoconiothyrium ambiguum sp. nov., a novel species isolated from sea water, and affiliation of the genus Verrucoconiothyrium to the family Didymellaceae.

The genus Verrucoconiothyrium was erected to accommodate Coniothyrium-like species with verruculose conidia. So far, it includes only four species, which have been found in association with plants, and very little is known about their distribution and host preferences. In this study, a Coniothyrium-like fungus isolated from sea water from the north of Portugal was characterised. Phylogenetic analysis, based on sequence data of the internal transcribed spacer and beta-tubulin loci, placed this fungus within the genus Verrucoconiothyrium but clearly distinct from the other known species. A novel species Verrucoconiothyrium ambiguumsp. nov. is described and illustrated. The taxonomic affiliation of the genus Verrucoconiothyrium at the family level was addressed through individual and combined gene genealogies. Our results show that the genus Verrucoconiothyrium is a member of the family Didymellaceae.

Biodiversity of Penicillium species from marine environments in Portugal and description of Penicillium lusitanum sp. nov., a novel species isolated from sea water.

During an extensive survey of marine fungi in coastal marine environments from Portugal, a collection of Penicillium isolates were obtained from sea water, macroalgae and driftwood. Sixteen distinct Penicillium species were identified with Penicillium terrigenum and Penicillium brevicompactum being the most frequent. A Penicillium species isolated from sea water could not be affiliated to any known species. Phylogenetic analyses based on the ITS region of the rDNA and the beta-tubulin (benA) gene placed it into Penicillium section Ramosa, distinct from all currently known species and with Penicillium tunisiense as its closest relative. Although having similar morphological characteristics, these species differ in micromorphological and molecular characters. Thus, Penicillium lusitanum sp. nov. is proposed as a novel species.

Combined transcriptomics-metabolomics profiling of the heat shock response in the hyperthermophilic archaeon Pyrococcus furiosus.

Pyrococcus furiosus is a remarkable archaeon able to grow at temperatures around 100 °C. To gain insight into how this model hyperthermophile copes with heat stress, we compared transcriptomic and metabolomic data of cells subjected to a temperature shift from 90 °C to 97 °C. In this study, we used RNA-sequencing to characterize the global variation in gene expression levels, while nuclear magnetic resonance (NMR) and targeted ion exchange liquid chromatography-mass spectrometry (LC-MS) were used to determine changes in metabolite levels. Of the 552 differentially expressed genes in response to heat shock conditions, 257 were upregulated and 295 were downregulated. In particular, there was a significant downregulation of genes for synthesis and transport of amino acids. At the metabolite level, 37 compounds were quantified. The level of di-myo-inositol phosphate, a canonical heat stress solute among marine hyperthermophiles, increased considerably (5.4-fold) at elevated temperature. Also, the levels of mannosylglycerate, UDP-N-acetylglucosamine (UDPGlcNac) and UDP-N-acetylgalactosamine were enhanced. The increase in the pool of UDPGlcNac was concurrent with an increase in the transcript levels of the respective biosynthetic genes. This work provides the first metabolomic analysis of the heat shock response of a hyperthermophile.

Dual RNA Sequencing of Vitis vinifera during Lasiodiplodia theobromae Infection Unveils Host-Pathogen Interactions.

Lasiodiplodia theobromae is one of the most aggressive agents of the grapevine trunk disease Botryosphaeria dieback. Through a dual RNA-sequencing approach, this study aimed to give a broader perspective on the infection strategy deployed by L. theobromae, while understanding grapevine response. Approximately 0.05% and 90% of the reads were mapped to the genomes of L. theobromae and Vitis vinifera, respectively. Over 2500 genes were significantly differentially expressed in infected plants after 10 dpi, many of which are involved in the inducible defense mechanisms of grapevines. Gene expression analysis showed changes in the fungal metabolism of phenolic compounds, carbohydrate metabolism, transmembrane transport, and toxin synthesis. These functions are related to the pathogenicity mechanisms involved in plant cell wall degradation and fungal defense against antimicrobial substances produced by the host. Genes encoding for the degradation of plant phenylpropanoid precursors were up-regulated, suggesting that the fungus could evade the host defense response using the phenylpropanoid pathway. The up-regulation of many distinct components of the phenylpropanoid pathway in plants supports this hypothesis. Moreover, genes related to phytoalexin biosynthesis, hormone metabolism, cell wall modification enzymes, and pathogenesis-related proteins seem to be involved in the host responses observed. This study provides additional insights into the molecular mechanisms of L. theobromae and V. vinifera interactions.

Models for the bystander effect in gradient radiation fields: Range and signalling type.

Bystander responses to radiation are responsible for a significant fraction of cell death, but are not included in the conventional linear-quadratic (LQ) radiobiological model. Strong dose gradients in radiation fields affect the distribution of bystander signals and can be used to decrease the survival of cancer cells. Predictive models incorporating bystander effects are needed to design the dose gradients in modulated fields to improve cancer treatments. Fundamental questions concern the nature and range of bystander signalling. Some authors propose bystander signals are carried by diffusing molecular factors expressed into the extracellular medium and that strong dose gradients drive their diffusion. Others propose bystander effects occur between neighbouring cells through gap-junctions, leaving no universal agreement. Here we test three assumptions concerning the effective range of bystander signals using both average and local measures of survival. Model 1 assumes short range signalling (e.g. gap-junction mediated) proportional to the local dose gradient, without relying on diffusion across the extracellular medium; Model 2 assumes metabolite diffusion governed by Fick's second law with either negative or both signs of bystander effect; Model 3 assumes that the extent of signal production is dependent on the average of the dose gradient over the field and that the signals have long range distribution. A single bystander parameter for each model was fitted to observed average survival of cancer cells in uniform and modulated fields. All models gave better fits than the classical LQ model. Model 2 fitted best with one sign of bystander effect on survival. Model 3 gave the best overall fit of average survival. The models were then used to predict local survival and survival as a function of dose in modulated fields, using independent datasets, without changing the bystander parameter. Model 3 gave the best overall prediction. This study demonstrates that the bystander effect can be controlled by design of the radiation field modulation.

Lasiodiplodia theobromae as a Producer of Biotechnologically Relevant Enzymes.

Phytopathogenic fungi are known to produce several types of enzymes usually involved in plant cell wall degradation and pathogenesis. The increasing of global temperature may induce fungi, such as Lasiodiplodia theobromae (L. theobromae), to alter its behavior. Nonetheless, there is only limited information regarding the effect of temperature on L. theobromae production of enzymes. The need for new, thermostable enzymes, that are biotechnologically relevant, led us to investigate the effect of temperature on the production of several extracellular enzymatic activities by different L. theobromae strains. Fungi were grown at 25 °C, 30 °C and 37 °C and the enzymatic activities were detected by plate assays, quantified by spectrophotometric methods and characterized by zymography. The thermostability (25-80 °C) of the enzymes produced was also tested. Strains CAA019, CBS339.90, LA-SOL3, LA-SV1 and LA-MA-1 produced amylases, gelatinases, caseinases, cellulases, lipases, laccases, xylanases, pectinases and pectin liases. Temperature modulated the expression of the enzymes, and this effect was more visible when fungi were grown at 37 °C than at lower temperatures. Contrary to proteolytic and endoglucanolytic activities, whose highest activities were detected when fungi were grown at 30 °C, lipolytic activity was not detected at this growth temperature. Profiles of proteases and endoglucanases of fungi grown at different temperatures were characterized by zymography. Enzymes were shown to be more thermostable when fungi were grown at 30 °C. Proteases were active up to 50 °C and endoglucanases up to 70 °C. Lipases were the least stable, with activities detected up to 45 °C. The enzymatic profiles detected for L. theobromae strains tested showed to be temperature and strain-dependent, making this species a good target for biotechnological applications.