Antimicrobial properties and death-inducing mechanisms of saccharomycin, a biocide secreted by Saccharomyces cerevisiae.

We recently found that Saccharomyces cerevisiae (strain CCMI 885) secretes antimicrobial peptides (AMPs) derived from the glycolytic enzyme glyceraldehyde 3-phosphate dehydrogenase (GAPDH) that are active against various wine-related yeast and bacteria. Here, we show that several other S. cerevisiae strains also secrete natural biocide fractions during alcoholic fermentation, although at different levels, which correlates with the antagonistic effect exerted against non-Saccharomyces yeasts. We, therefore, term this biocide saccharomycin. The native AMPs were purified by gel-filtration chromatography and its antimicrobial activity was compared to that exhibited by chemically synthesized analogues (AMP1 and AMP2/3). Results show that the antimicrobial activity of the native AMPs is significantly higher than that of the synthetic analogues (AMP1 and AMP2/3), but a conjugated action of the two synthetic peptides is observed. Moreover, while the natural AMPs are active at pH 3.5, the synthetic peptides are not, since they are anionic and cannot dissolve at this acidic pH. These findings suggest that the molecular structure of the native biocide probably involves the formation of aggregates of several peptides that render them soluble under acidic conditions. The death mechanisms induced by the AMPs were also evaluated by means of epifluorescence microscopy-based methods. Sensitive yeast cells treated with the synthetic AMPs show cell membrane disruption, apoptotic molecular markers, and internalization of the AMPs. In conclusion, our work shows that saccharomycin is a natural biocide secreted by S. cerevisiae whose activity depends on the conjugated action of GAPDH-derived peptides. This study also reveals that S. cerevisiae secretes GAPDH-derived peptides as a strategy to combat other microbial species during alcoholic fermentations.

Male Breast Metastasis: A Case of Treatment-Emergent Neuroendocrine Prostate Cancer.

Treatment-emergent transformed neuroendocrine prostate cancer (NEPC) is a highly aggressive type of prostate cancer that may arise from typical adenocarcinoma of the prostate, which is associated with rapidly progressive disease involving visceral sites and refractoriness to hormonal therapy. We present the case of a 74-year old male with a known history of stable prostate adenocarcinoma treated with transurethral prostate resection, local radiotherapy (RT), and androgen deprivation therapy (ADT) in 2020 who presented to the emergency room with complaints of shoulder and anterior chest pain, dyspnoea, and fatigue. Upon examination, a solid, adherent breast mass and infra-clavicular adenopathy were palpable. Thoracic computed tomography (CT) scan showed adenopathies in multiple thoracic chains, bilateral pulmonary nodular opacities, multiple osteolytic lesions, and bilateral enlargement of retro areolar tissue. A staging CT scan revealed further hepatic and penile lesions. Breast mass biopsy was compatible with small cell neuroendocrine cancer. Biopsies of the prostate, penis, lymph nodes, and bronchus were also performed. Histology of the prostate showed focal infiltration by the known adenocarcinoma while all others documented extensive infiltration by neuroendocrine carcinoma, whose morphology and immunohistochemical profile were identical to that of the breast. This case highlights the challenges a diagnosis of neuroendocrine prostate cancer might pose, and the aggressiveness of this type of cancer, which frequently presents with advanced disease and is associated with poor outcomes.

Analysis and direct quantification of Saccharomyces cerevisiae and Hanseniaspora guilliermondii populations during alcoholic fermentation by fluorescence in situ hybridization, flow cytometry and quantitative PCR.

Traditionally, it was assumed that non-Saccharomyces (NS) yeasts could only survive in the early stages of alcoholic fermentations. However, recent studies applying culture-independent methods have shown that NS populations persist throughout the fermentation process. The aim of the present work was to analyze and quantify Saccharomyces cerevisiae (Sc) and Hanseniaspora guilliermondii (Hg) populations during alcoholic fermentations by plating and culture-independent methods, such as fluorescence in situ hybridization (FISH) and quantitative PCR (QPCR). Species-specific FISH probes labeled with fluorescein (FITC) were used to directly hybridize Sc and Hg cells from single and mixed cultures that were enumerated by epifluorescence microscopy and flow cytometry. Static and agitated fermentations were performed in synthetic grape juice and cell density as well as sugar consumption and ethanol production were determined throughout fermentations. Cell density values obtained by FISH and QPCR revealed the presence of high populations (107-108 cells/ml) of Sc and Hg throughout fermentations. Plate counts of both species did not show significant differences with culture-independent results in pure cultures. However, during mixed fermentations Hg lost its culturability after 4-6 days, while Sc remained culturable (about 108 cells/ml) throughout the entire fermentation (up to 10 days). The rRNA content of cells during mixed fermentations was also analyzed by flow cytometry in combination with FISH probes. The fluorescence intensity conferred by the species-specific FISH probes was considerably lower for Hg than for Sc. Moreover, the rRNA content of Hg cells, conversely to Sc cells, remained almost unchanged after boiling, which showed that rRNA stability is species-dependent.

Fungal Endocarditis: A Case Report.

Fungal endocarditis is a rare disease, and it is associated with severe complications and poor prognosis despite combined clinical and surgical treatment. Although Candida albicans (C. albicans) is the most common etiological agent of this severe form of endocarditis, Candida parapsilosis (C. parapsilosis) is the most common non-albicans causative species. It occurs mostly in patients with predisposing risk factors, and the rarity of this disease demands a high index of suspicion; the diagnosis must be vigilantly pursued by echocardiography and multiple blood cultures. The past few decades have witnessed a rise in the incidence of this disease, mainly due to improvements in the diagnostic approach. We report the case of a 63-year-old man with no previous medical history of cardiac disease and no risk factors who was diagnosed with fungal endocarditis due to C. parapsilosis without fungemia. This report illustrates a rare case of fungal endocarditis in a patient with no risk factors and highlights the challenges encountered in the diagnosis, along with complications and predictors of poor prognosis.

When a Late Metastasis Is Hard to Swallow.

Pseudoachalasia is an uncommon disorder characterised by aperistalsis in the tubular oesophagus and impaired relaxation of the lower oesophageal sphincter (LES). It presents with symptoms and radiologic, endoscopic and manometric findings that mimic idiopathic achalasia. There is a huge spectrum of underlying causes for pseudoachalasia, although malignancy is the most common aetiology. We report the case of a 70-year-old Portuguese female with a history of breast cancer, submitted to tumourectomy, radiotherapy and hormonotherapy, in complete remission for 16 years, who presented in the emergency department with a two-month history of dysphagia, weight loss, heartburn and nausea. Blood work, body computed tomography (CT) scan, mammography, upper endoscopy, colonoscopy and skeletal scintigraphy did not show any alterations, but barium swallow scan and oesophageal manometry suggested achalasia. She was submitted to oesophageal dilatation with partial symptomatic improvement. Six months later, new onset of dysphonia and worsening of initial symptoms was noticed. A new CT scan revealed unilateral pleural effusion, large mediastinal adenopathy and multiple pulmonary nodules highly suggestive of a metastatic malignancy. Endobronchial ultrasound-guided transbronchial needle aspiration (EBUS-TBNA) from mediastinal adenopathies confirmed the tumoural invasion by a carcinoma, and immunohistochemistry suggested a breast origin. She underwent a nasoendoscopy that revealed bilateral vocal cord paralysis. After chemotherapy was started, symptoms of achalasia completely resolved, and tumour markers, which were increased, have normalised. The presented case highlights a pseudoachalasia as the first manifestation of a late breast metastasis. It is essential to always have in mind patients' past history as a key that can help resolve clinical doubts.

Epidural Abscess: A Cause of Back Pain That Must Not Be Missed.

Low back and cervical pain are common emergency department complaints. Most cases represent low-risk pathologies but in rare cases, it can be a symptom of a serious underlying disease and the distinction can be challenging. A case is reported of a 58-year-old male with recurrent cervical pain requiring several emergency department visits. He eventually presented with intense cervical and low back pain, tetraparesis, and dyspnea. His condition deteriorated in less than 24 hours, evolving with fever, shock, and respiratory insufficiency and was admitted to the Intensive Care Unit for aminergic support, invasive mechanical ventilation and started on empiric antibiotics. Investigations revealed raised inflammatory markers and a cervical pre-vertebral purulent collection associated with an epidural collection at the level of C2 to C5 with spinal cord compression. The patient underwent transoral drainage and removal of the posterior C1 arch along with C2-C3-C4-C5 laminectomy. Staphylococcus aureus was isolated in the blood, pus, and bone. Re-intervention was required due to persistent pre-vertebral abscesses with clinical improvement thereafter. Eight weeks of antibiotic treatment were completed in total. The patient was discharged after 72 days of hospitalization with significant improvement in force and autonomy. Epidural abscess is a rare finding that can cause rapid irreversible neurological deterioration. Clinical suspicion must be high for an early diagnosis and treatment, essential for a better prognosis.

Leptospirosis, a Re-emerging Threat.

Leptospirosis is a zoonosis transmitted by an animal vector and caused by the spirochete bacteria Leptospira. Human infection is rare and acquired by exposure to environmental sources (animal urine, contaminated water, soil, or infected animal tissue). It can have an extremely broad presentation, ranging from asymptomatic to serious illness. We report the case of a 59-year-old man admitted to the hospital with myalgia, fever, and abdominal discomfort. Routine laboratory tests revealed raised inflammatory markers, thrombocytopenia, kidney dysfunction, and hepatic cytolysis and cholestasis. The hypothesis of zoonosis was raised based on symptoms and analytical changes despite the weak epidemiological history to support it. Although leptospira serology tests were negative on admission, a polymerase chain reaction test was requested due to a high degree of suspicion which came back positive. The patient completed eight days of doxycycline with favorable clinical and analytical progression. This case highlights the changing epidemiology of leptospirosis and the importance of having a high degree of suspicion even outside endemic zones. It also highlights the importance of a wise choice of diagnostic tests according to the disease stage.

Impact of sterile leukocyturia on outcome of critically ill patients with severe acute kidney injury.

INTRODUCTION: The role of immunological mechanisms on renal regeneration and functional recovery after an episode of Acute Kidney Injury (AKI) is still understudied. We aim to evaluate the impact of sterile leukocyturia on outcomes of critically-ill AKI patients. METHODS: A retrospective analysis of critically-ill patients with stage ≥2 AKI by KDIGO was performed. Patients with urinary tract infection, previous renal replacement therapy, chronic kidney disease stage >3 and kidney, urinary tract or prostatic cancer were excluded. Sterile leukocyturia was defined as a positive leukocyte esterase value. RESULTS: 108 patients with stage ≥2 AKI were included, 39.8% of which had sterile leukocyturia. AKI patients with sterile leukocyturia were older, had more cardiovascular disease and a lower baseline renal function (p < 0.05). They had a higher serum creatinine and leukocytosis at admission, were more frequently septic (p < 0.05) and had more persistent AKI by both KDIGO criteria at multivariable analysis (OR 6.130, 95% CI 2.007-18.747). CONCLUSION: Sterile leukocyturia was associated with different patient baseline and AKI characteristics and more persistent AKI by both KDIGO criteria. Sterile leukocyturia may represent a surrogate marker of renal inflammation during AKI.

Large-vessel vasculitis induced by pegfilgrastim.

Granulocyte colony-stimulating factor (G-CSF) is increasingly being used to prevent febrile neutropenia associated with chemotherapy. Large-vessel vasculitis (LVV) has been recognized as a rare side effect of G-CSF treatment. We report a case of G-CSF associated LVV in a patient with breast cancer. While clear pathogenic mechanisms remain unknown, G-CSF may cause vasculitis due to inflammatory cytokines production. This adverse reaction should be recognized in patients with suggestive symptoms following the administration of pegfilgrastim. A 56-year-old woman with luminal B breast cancer who had undergone surgery and adjuvant chemotherapy, initially with paclitaxel, was started on a doxorubicin plus cyclophosphamide protocol, followed by supportive use of long-acting G-CSF pegfilgrastim. Following the administration of pegfilgrastim, the patient developed intermittent fever and was given empiric antibiotics in the outpatient setting with no improvement. There were no signs of cancer progression, and the contrast-enhanced CT scan highlighted wall thickening of the aortic arch and the proximal segment of the subclavian artery, which was not present in previous imaging studies. The patient was diagnosed with LVV, and a differential diagnosis was performed to rule out paraneoplastic setting, immune-mediated diseases, infection or other drug-induced vasculitis. Treatment with steroids was initiated and tapered with significant improvement and resolution of the radiological signs of aortitis.

Seasonal variations in lipid composition of the hydrothermal vent mussel Bathymodiolus azoricus from the Menez Gwen vent field.

Specimens of the hydrothermal mussel Bathymodiolus azoricus collected in Menez Gwen hydrothermal vent field (NE Atlantic) during 2002-2003 were examined for feeding patterns variations through three seasons. The fatty acid profile and lipid classes of the mussels were studied, together with the MODIS/AQUA-derived near-surface chlorophyll a to test the hypothesis that surface productivity might be related to the feeding patterns of this species. The lipid levels showed pronounced seasonal fluctuations with the highest values occurring in January and August. Seasonal variations in lipid classes and fatty acid composition of neutral and polar lipids in the mussels are presented. Differences in the fatty acid profile of lipid classes in different seasons suggest that the higher energy requirements in summer and winter were supplied by bacterial biomarkers omega7 MUFA (monounsaturated fatty acids), whereas omega6 PUFA (polyunsaturated fatty acids) and NMI (non-methylene-interrupted) fatty acids predominated during the spring. The MODIS/AQUA data show marked seasonal variability and an anomalous peak during January of 2003, although this cannot be directly linked to lipid composition variation.

Survival rate of wine-related yeasts during alcoholic fermentation assessed by direct live/dead staining combined with fluorescence in situ hybridization.

Real-time detection of microorganisms involved in complex microbial process, such as wine fermentations, and evaluation of their physiological state is crucial to predict whether or not those microbial species will be able to impact the final product. In the present work we used a direct live/dead staining (LDS) procedure combined with fluorescence in situ hybridization (FISH) to simultaneously assess the identity and viability of Saccharomyces cerevisiae (Sc) and Hanseniaspora guilliermondii (Hg) during fermentations performed with single and mixed cultures. The population evolution of both yeasts was determined by plating and by LDS combined with species-specific FISH-probes labeled with Fluorescein. Since the FISH method involves the permeabilization of the cell membrane prior to hybridization and that it may influence the free diffusion of PI in and out of the cells, we optimized the concentration of this dye (0.5 µg of PI per 10(6) cells) for minimal diffusion (less than 2%). Fluorescent cells were enumerated by hemocytometry and flow cytometry. Results showed that the survival rate of Sc during mixed cultures was high throughout the entire process (60% of viable cells at the 9th day), while Hg began to die off at the 2nd day, exhibited 98% of dead cells at the 3rd day (45 g/l of ethanol) and became completely unculturable at the 4th day. However, under single culture fermentation the survival rate and culturability of Hg decreased at a much slower pace, exhibiting at the 7th day (67 g/l of ethanol) 8.7×10(4) CFU/ml and 85% of dead cells. Thus, our work demonstrated that the LDS-FISH method is able to simultaneously assess the viability and identity of these wine-related yeast species during alcoholic fermentation in a fast and reliable way. In order to validate PI-staining as a viability marker during alcoholic fermentation, we evaluated the effect of ethanol on the membrane permeability of Sc and Hg cells, as well as their capacity to recover membrane integrity after being exposed to different levels of ethanol (1%, 6%, 10%, 12% v/v). Results showed that while Sc cells were able to recover membrane integrity after ethanol exposure, Hg cells were not. However, under alcoholic fermentation Sc cells didn't recover membrane integrity after the mid-term (4-5 days) of alcoholic fermentation.

Concentration of sterols of Porphyridium cruentum biomass at stationary phase.

The objective of this study was to investigate sterols content of Porphyridium cruentum batch cultured in laboratory at 18 degrees C and harvested in the stationary. The sterol distribution of this species is characterized by a predominance of cholesterol, with values as 199.0 mg 100 g(-1) freeze dry weight (92.2%). The second most important sterol was stigmasterol (4.9%) followed by beta-sitosterol (2.2%). Studied sterols give to this species a special importance in for being used in food as supplements/nutraceuticals (including aquaculture).

Prevenção da violência no namoro: Avaliação do workshop ver, pensar e agir: Projeto (o)usar & ser laço branco / Prevenção da violência no namoro: Avaliação do workshop ver, pensar e agir: Projeto (o)usar & ser laço branco

A violência no namoro (VN) constitui uma preocupação crescente enquanto grave problema de saúde mundial, destacando-se pela elevada prevalência e por se assumir um forte preditor da violência nas relações de intimidade futuras. Segundo a OMS (2012), a implementação de programas de prevenção primária deve ser uma prioridade. Com este estudo pretendemos conhecer a eficácia dos workshops VPA: Ver-Pensar-Agir, do projecto ?(O)Usar ? Ser Laço Branco? (OSLB) que procura prevenir a VN. Tivemos como objetivos: avaliar o efeito dos workshops VPA do projeto OSLB, sobre os conhecimentos das/os adolescentes sobre VN; descrever as práticas das/os adolescentes, perante o conhecimento de situações de VN; identificar os comportamentos de vitimização e perpetração de VN, nas/os adolescentes; e avaliar a importância atribuída pelas/os adolescentes às estratégias utilizadas nos workshops. Foi realizado um estudo quase-experimental. Participaram 89 estudantes do ensino secundário que pertenciam a 4 turmas. A seleção da amostra foi por conveniência. Foram constituídos dois grupos com duas turmas cada: grupo intervenção com 48 estudantes (31 rapazes e 17 raparigas) e grupo controlo com 41 estudantes (8 rapazes e 33 raparigas).Os dados foram recolhidos através de questionários concebidos no âmbito do projeto OSLB. Foi efetuado um workshop no grupo intervenção, a quem foi aplicado um questionário antes, imediatamente após e passados 5 meses (follow-up). Ao grupo controlo foi aplicado o mesmo questionário no dia do workshop e passados 5 meses. Foram salvaguardados todos os procedimentos éticos e legais, com parecer favorável da Comissão de Ética da UICISA-E da ESEnfC. Os resultados permitem-nos verificar que as/os adolescentes referem ter conhecimentos sobre a quem recorrer numa situação de VN. O workshop do projeto OSLB foi eficaz para aumentar os conhecimentos das/os participantes após a intervenção, verificando-se a sua manutenção após 5 meses, sendo que as raparigas possuem mais conhecimentos sobre a VN, no pré-teste. Os principais comportamentos quer de vitimização quer de perpetuação são de violência psicológica. A utilização do teatro fórum e a proximidade de idades dos educadores de pares foi considerado pelas/os adolescentes como uma estratégia facilitadora para a sensibilização, comunicação e participação na sessão. Os resultados obtidos convergem com a literatura consultada. A VN é uma realidade para muitos adolescentes, mas é sensível a programas de prevenção primária, de que é exemplo o projeto OSLB. Consideramos que a principal limitação do estudo foi a não randomização da amostra. Sugerimos que os enfermeiros ? em especial os especialistas em saúde materna e obstetrícia - devem procurar desenvolver ou replicar programas de prevenção primária, cuja eficácia seja comprovada, com o objetivo de fomentarem relações de intimidade saudáveis e sobretudo prevenir a VN.